Noninvasive Tests for Hepatic Fibrosis

Number: 0690

Table Of Contents

Policy Applicable CPT / HCPCS / ICD-10 Codes Background References

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Policy

Scope of Policy

This Clinical Policy Bulletin addresses non-invasive tests for hepatic fibrosis.

1. Medical Necessity

   1. Aetna considers transient elastography (e.g., FibroScan) medically necessary for follow-up of primary sclerosing cholangitis, monitoring of liver function in Wilson’s disease, and for distinguishing hepatic cirrhosis from non-cirrhosis in persons with hepatitis B, hepatitis C or other chronic liver diseases (e.g., hereditary hemochromatosis, non-alcoholic fatty liver disease (NAFLD) and non-alcoholic steatohepatitis (NASH); now known as metabolic dysfunction-associated steatohepatitis (MASH)). Performance of transient elastography more than twice per year is considered not medically necessary. Performance of transient elastography within 6 months following a liver biopsy, or FibroTest-ActiTest/HCV-Fibrosure is considered not medically necessary.
   2. Aetna considers FibroTest-ActiTest/HCV-FibroSure medically necessary for distinguishing hepatic cirrhosis from non-cirrhosis in persons with hepatitis C and other chronic liver diseases (e.g., hereditary hemochromatosis, NAFLD and NASH (now known as MASH)). Performance of this test more than twice per year is considered not medically necessary. Performance of this test within 6 months following a liver biopsy or transient elastography is considered not medically necessary.
   3. Aetna considers magnetic resonance elastography medically necessary for non-alcoholic steatohepatitis (NASH; now known as MASH) for the detection and prognosis of liver fibrosis. Performance of magnetic resonance elastography more than twice per year is considered not medically necessary. Performance of this test within 6 months following a liver biopsy (or other test for liver fibrosis) is considered not medically necessary.
   4. Aetna considers the Enhanced Liver Fibrosis (ELF) test medically necessary for the detection and prognosis of liver fibrosis in persons with chronic liver diseases. Performance of the Enhanced Liver Fibrosis (ELF) test more than twice per year is considered not medically necessary. Performance of this test within 6 months following a liver biopsy (or other test for liver fibrosis) is considered not medically necessary.
   5. AST, ALT, and platelets (used with age to calculate the FIB-4 index) for assessing risk of progression of liver disease in persons with risk factors for nonalcoholic fatty liver disease (NAFLD).

2. Experimental, Investigational, or Unproven

   The following procedures are considered experimental, investigational, or unproven because the effectiveness of these approaches has not been established:

   1. Acoustic radiation forced impulse (ARFI) for distinguishing hepatic cirrhosis from non-cirrhosis in persons with hepatitis C and other chronic liver diseases, and for all other indications;
   2. Artificial intelligence in distinguishing healthy versus NAFLD/non-alcoholic steatohepatitis (NASH; now known as MASH), fibrosis versus non-fibrosis in the evaluation of NAFLD progression, as well as screening and early diagnosis of fatty liver disease;
   3. FibroTest-ActiTest/HCV-FibroSure for all other indications (e.g., diagnosis and/or monitoring of primary biliary cholangitis);
   4. Hepatic artery resistive index for evaluation of fibrosis progression in individuals with non-alcoholic fatty liver disease (NAFLD);
   5. Magnetic resonance elastography for distinguishing hepatic cirrhosis from non-cirrhosis in persons with hepatitis C or other chronic liver diseases, and for all other indications (e.g., diagnosis of hepatic fibrosis, prediction of ascites in persons with chronic liver disease);
   6. NASHnext for evaluation of NASH activity and fibrosis;
   7. OWLiver Panel for detecting or monitoring hepatic fibrosis in persons with hepatitis C or other chronic liver diseases (e.g., NAFLD, now known as metabolic dysfunction-associated fatty liver disease (MAFLD));
   8. Quantitative magnetic resonance (e.g., LiverMultiScan) for analysis of liver tissue composition;
   9. Serum ferritin as a biomarker for detecting or monitoring hepatic fibrosis in persons with hepatitis C or other chronic liver diseases (e.g., NAFLD);
   10. Transient elastography for detection of esophageal varices in individuals with cirrhosis, diagnosis of acute cellular rejection following liver transplantation, diagnosis of glycogenic hepatopathy, diagnosis of portal hypertension, and evaluation of alpha-1 antitrypsin deficiency, Alagille syndrome, and Gilbert syndrome;
   11. Transient elastography for routine follow-up of liver transplant recipients;
   12. The following serum marker tests for detecting or monitoring hepatic fibrosis in persons with hepatitis C or other chronic liver diseases (e.g., NAFLD) (not an all-inclusive list):
       • Angiotensin converting enzyme
       • FibroMAX
       • FibroSpect
       • HepaScore
       • LIVERFAST
       • Micro-fibrillar associated glycoprotein 4 (MFAP4)
       • MicroRNA-21
       • miR-29a and miR-122
       • miRNA-221 and miRNA-222
       • NASH FibroSure
       • Plasma cytokeratin-18
       • Signal-induced proliferation-associated 1 like 1 (SIPA1L1).

Table:

CPT Codes / HCPCS Codes / ICD-10 Codes

Code Code Description

Transient elastography (e.g., FibroScan):

CPT codes covered if selection criteria are met:

76981 Ultrasound, elastography; parenchyma (eg, organ) 91200 Liver elastography, mechanically induced shear wave (eg, vibration), without imaging, with interpretation and report

CPT codes not covered for indications listed in the CPB:

76982 – 76983 Ultrasound, elastography

ICD-10 codes covered if selection criteria are met:

B18.0 – B18.1 Chronic viral hepatitis B B18.2 Chronic viral hepatitis C E83.01 Wilson’s disease E83.110 Hereditary hemochromatosis K74.60 – K74.69 Other and unspecified cirrhosis of liver K75.81 Nonalcoholic steatohepatitis (NASH) K76.0 Fatty (change of) liver, not elsewhere classified K83.01 Primary sclerosing cholangitis

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

E74.00 – E74.09 Glycogen storage disease [glycogenic hepatopathy] E80.4 Gilbert syndrome E88.01 Alpha-1-antitrypsin deficiency I85.00 – I85.01 Esophageal varices K76.6 Portal hypertension Q44.70, Q44.71, Q44.79 Other congenital malformations of liver [Alagille syndrome] T86.41 Liver transplant rejection Z94.4 Liver transplant status

FibroTest-ActiTest, HCV-FibroSure and NASH FibroSure:

CPT codes covered if selection criteria are met:

81596 Infectious disease, chronic hepatitis C virus (HCV) infection, six biochemical assays (ALT, A2-macroglobulin, apolipoprotein A-1, total bilirubin, GGT, and haptoglobin) utilizing serum, prognostic algorithm reported as scores for fibrosis and necroinflammatory activity in liver

CPT codes not covered for indications listed in the CPB:

0003M Liver disease, ten biochemical assays (ALT, A2-macroglobulin, apolipoprotein A-1, total bilirubin, GGT, haptoglobin, AST, glucose, total cholesterol and triglycerides) utilizing serum, prognostic algorithm reported as quantitative scores for fibrosis, steatosis and nonalcoholic steatohepatitis (NASH) [NASH FibroSURE]

Other CPT codes related to the CPB:

47000 Biopsy of liver, needle; percutaneous 47001 Biopsy of liver, needle; when done for indicated purpose at time of other major procedure (List separately in addition to code for primary procedure) 47100 Biopsy of liver, wedge 91200 Liver elastography, mechanically induced shear wave (eg, vibration), without imaging, with interpretation and report

ICD-10 codes covered if selection criteria are met:

B18.0 – B18.1 Chronic viral hepatitis B B18.2 Chronic viral hepatitis C E83.110 Hereditary hemochromatosis K70.0 – K77 Diseases of liver [chronic] Z94.4 Liver transplant status

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

K83.01 Primary sclerosing cholangitis K83.09 Other cholangitis [primary biliary cholangitis]

Magnetic Resonance Elastography:

CPT codes covered if selection criteria are met:

76391 Magnetic resonance (eg, vibration) elastography

Other CPT codes related to the CPB:

47000 Biopsy of liver, needle; percutaneous 47001 when done for indicated purpose at time of other major procedure (List separately in addition to code for primary procedure) 47100 Biopsy of liver, wedge

ICD-10 codes covered if selection criteria are met:

K75.81 Nonalcoholic steatohepatitis (NASH)

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

K74.00 – K74.02 Hepatic fibrosis

Enhanced Liver Fibrosis (ELF) test:

CPT codes covered if selection criteria are met:

81517 Liver disease, analysis of 3 biomarkers (hyaluronic acid [HA], procollagen III amino terminal peptide [PIIINP], tissue inhibitor of metalloproteinase 1 [TIMP-1]), using immunoassays, utilizing serum, prognostic algorithm reported as a risk score and risk of liver fibrosis and liver-related clinical events within 5 years

Other CPT codes related to the CPB:

47000 Biopsy of liver, needle; percutaneous 47001 when done for indicated purpose at time of other major procedure (List separately in addition to code for primary procedure) 47100 Biopsy of liver, wedge

ICD-10 codes covered if selection criteria are met:

K70.0 – K77 Diseases of liver [chronic]

Artificial intelligence:

CPT codes not covered for indications listed in the CPB:

Artificial intelligence –no specific code

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

K70.0 – K77 Diseases of liver Z13.818 Encounter for screening for other digestive system disorders [fatty liver disease]

NASHnext:

CPT codes not covered for indications listed in the CPB:

0468U Hepatology (nonalcoholic steatohepatitis [NASH]), miR-34a- 5p, alpha 2-macroglobulin, YKL40, HbA1c, serum and whole blood, algorithm reported as a single score for NASH activity and fibrosis

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

K74.00 – K74.02 Hepatic fibrosis K75.81 Nonalcoholic steatohepatitis (NASH)

OWLiver Panel:

CPT codes not covered for indications listed in the CPB:

0344U Hepatology (nonalcoholic fatty liver disease [NAFLD]), semiquantitative evaluation of 28 lipid markers by liquid chromatography with tandem mass spectrometry (LC-MS/MS), serum, reported as at-risk for nonalcoholic steatohepatitis (NASH) or not NASH

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

B18.2 Chronic viral hepatitis C K74.00 – K74.02 Hepatic fibrosis K76.0 Fatty (change of) liver, not elsewhere classified

Quantitative magnetic resonance (e.g., LiverMultiScan):

CPT codes not covered for indications listed in the CPB:

0648T Quantitative magnetic resonance for analysis of tissue composition (eg, fat, iron, water content), including multiparametric data acquisition, data preparation and transmission, interpretation and report, obtained without diagnostic MRI examination of the same anatomy (eg, organ, gland, tissue, target structure) during the same session 0649T Quantitative magnetic resonance for analysis of tissue composition (eg, fat, iron, water content), including multiparametric data acquisition, data preparation and transmission, interpretation and report, obtained with diagnostic MRI examination of the same anatomy (eg, organ, gland, tissue, target structure) (List separately in addition to code for primary procedure) 0697T Quantitative magnetic resonance for analysis of tissue composition (eg, fat, iron, water content), including multiparametric data acquisition, data preparation and transmission, interpretation and report, obtained without diagnostic MRI examination of the same anatomy (eg, organ, gland, tissue, target structure) during the same session; multiple organs 0698T Quantitative magnetic resonance for analysis of tissue composition (eg, fat, iron, water content), including multiparametric data acquisition, data preparation and transmission, interpretation and report, obtained with diagnostic MRI examination of the same anatomy (eg, organ, gland, tissue, target structure); multiple organs (List separately in addition to code for primary procedure) 0723T Quantitative magnetic resonance cholangiopancreatography (QMRCP) including data preparation and transmission, interpretation and report, obtained without diagnostic magnetic resonance imaging (MRI) examination of the same anatomy (eg, organ, gland, tissue, target structure) during the same session 0724T Quantitative magnetic resonance cholangiopancreatography (QMRCP) including data preparation and transmission, interpretation and report, obtained with diagnostic magnetic resonance imaging (MRI) examination of the same anatomy (eg, organ, gland, tissue, target structure) (List separately in addition to code for primary procedure)

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

K70.0 – K77 Diseases of liver

Serum ferritin:

CPT codes not covered for indications listed in the CPB:

82728 Ferritin

ICD-10 codes not covered for indications listed in the CPB (not all-inclusive):

K70.0 – K77 Diseases of liver

Serum marker tests:

CPT codes not covered for indications listed in the CPB:

Hepatic artery resistive index, serum markers tests angiotensin converting enzyme, serum miR-29a and miR-122 and serum miRNA-221 and miRNA-222 – no specific code [not covered for liver Fibrosis]:

0166U Liver disease, 10 biochemical assays (α2-macroglobulin, haptoglobin, apolipoprotein A1, bilirubin, GGT, ALT, AST, triglycerides, cholesterol, fasting glucose) and biometric and demographic data, utilizing serum, algorithm reported as scores for fibrosis, necroinflammatory activity, and steatosis with a summary interpretation 83520 Immunoassay, analyte, quantitative; not otherwise specified [If billed for FIBROspect or HCV-FIBROSUREFibroMAX, FibroTest-ActiTest, HepaScore] 83883 Nephelometry, each analyte not elsewhere specified [If billed for FIBROspect or HCV-FIBROSURE FibroMAX, HepaScore] 88342 Immunohistochemistry or immunocytochemistry, per specimen; initial single antibody stain procedure [for the evaluation of non-alcoholic fatty liver disease and other liver disease]

Other CPT codes related to the CPB:

47000 Biopsy of liver, needle; percutaneous 47001 Biopsy of liver, needle; when done for indicated purpose at time of other major procedure (List separately in addition to code for primary procedure) 47100 Biopsy of liver, wedge 82977 Glutamyltransferase, gamma (GGT)

ICD-10 codes covered if selection criteria are met:

B18.0 – B18.1 Chronic viral hepatitis B B18.2 Chronic viral hepatitis C K70.0 – K77 Diseases of liver [chronic] Z94.4 Liver transplant status

Background

Hepatic fibrosis is the excessive accumulation of fibrotic connective tissue resulting from prolonged inflammation and progressive scarring of the liver due to a sustained wound-healing response to alcohol or nonalcohol-induced liver injury (nonalcoholic liver disease includes, but not limited to, hepatitis B and hepatitis C infections). The increased fibrosis and liver stiffness reduces blood flow through the liver, which leads to hardening and death of liver cells. Other chronic liver diseases include alcoholic liver disease, chronic hepatitis B, non-alcoholic steatosis, and chronic viral hepatitis B.

Liver biopsy is considered the gold standard for diagnosis and management of chronic liver disease. However, it is an invasive procedure that may result in complications. For that reason, non-invasive hepatic fibrosis tests are being introduced. Examples of these tests include, but may not be limited to, the following.

Serum Markers of Hepatic Fibrosis

Liver fibrosis serum panels are blood serum laboratory tests that have been developed as an alternative to liver biopsy to purportedly determine the extent of liver damage that has occurred in individuals with liver disease, such as hepatitis C virus (HCV).

Biochemical marker combinations are being developed as alternatives to liver biopsy in patients with chronic hepatitis C and other chronic liver diseases, including chronic hepatitis B, alcoholic liver disease, or non-alcoholic steatosis. Non-invasive tests are being developed to replace liver biopsy, and thus avoid the risk of biopsy-related adverse events. Non-invasive tests also have the potential to avoid limitations of liver biopsy, including the risk of sampling errors and inter- and intra-pathologist variability.

An assessment by the Belgian Healthcare Knowledge Center (KCE, 2016) reported that until recently, the usual examination to establish the degree of hepatic fibrosis was a histopathological examination of a liver sample taken by means of a liver biopsy. The result of a biopsy is expressed on the basis of the METAVIR scoring system which qualifies the degree of liver fibrosis in function of the histologic characteristics (F score) and the degree of inflammatory activity (A score) observed in the sample. The assessment noted, however, that liver biopsy is an invasive test which carries a risk of bleeding, especially in patients in an advanced stage of the illness. The assessment noted that there are now a number of non-invasive tests for hepatic fibrosis. However, there are no studies that measure the long-term outcomes of patients managed with these non-invasive tests (the longest study ran over a 9-year period but was conducted on a relatively small sample). The only studies available are cross-sectional studies which evaluate the sensitivity and specificity of these non-invasive tests in comparison with biopsies.

The FibroSpect II (Prometheus Laboratories, San Diego, CA) is a non-invasive diagnostic panel to assist in the detection of liver fibrosis. The FibroSpect II uses a combination of components in the fibro-genic cascade, such as hyaluronic acid, TIMP-1 (tissue inhibitor of metalloproteinase), and alpha-2-macroglobulin. Using an algorithm, the results of the measurements are converted into a score to determine an individual’s fibrosis score. The test is intended to differentiate mild fibrosis from more severe disease.

HepaScore measures four markers for liver fibrosis – bilirubin, gamma glutamyl transpeptidase, hyaluronic acid, and alpha-2 macroglobulin – and applies the results to an algorithm, combined with an individual’s age and sex, to determine a liver fibrosis score.

Non-alcoholic fatty liver disease (NAFLD) fibrosis score is based on analytes that are supposedly individually useful for evaluating patients with liver disease. The test includes ALT, albumin, aspartate aminotransferase (AST) glucose and platelet count. Age and body mass index (BMI) are also used to calculate the fibrosis score.

The FibroTest (Biopredictive, Houilles, France) and the ActiTest (Biopredictive, Houilles, France), marketed in the United States as the HCV-FibroSure Test (LabCorp, Burlington, NC) (also known as HCV-FibroSure, ASH FibroSure, NASH FibroSure) are serum biochemical tests for the assessment of fibrosis and necro-inflammatory activity, respectively. The HCV-FibroSure Test includes the following five markers, as well as age and gender: alpha2-macroglobulin, haptoglobin, gamma-glutamyl transpeptidase (GGT), total bilirubin, apolipoprotein A1, plus alanine aminotransferase (ALT). These measurements are applied to an algorithm, combined with an individual’s age and sex, to determine liver fibrosis severity. In addition, ASH FibroSure and NASH FibroSure purportedly provide markers for steatosis, alcoholic steatohepatitis (ASH) and non-alcoholic steatohepatitis (NASH) as well as for liver fibrosis.

Rossi et al (2003) reported on the results of FibroTest scores of 125 patients with hepatitis C. Of these, 57 had FibroTest scores either less than 0.1 (indicating no fibrosis) or greater than 0.6 (indicating significant fibrosis). Although 33 of the 125 patients had FibroTest scores of less than 0.1 and were therefore deemed unlikely to have fibrosis, 6 (18 %) had significant fibrosis. Conversely, of the 24 patients with scores of greater than 0.6 who were likely to have significant fibrosis, 5 (21 %) had mild fibrosis. The investigators concluded that, “[o]f the 125 patients in the cohort, 57 (46 %) could have avoided liver biopsy”; but discrepant results were recorded in 11 of those 57 (19 %). In other words, discrepancies with the biopsy gold standard were found in one-fifth of patients. There are no prospective clinical outcome studies of the HCV-FibroSure in the management of patients with hepatitis C or other chronic liver diseases.

An National Institutes of Health Consensus Statement on Management of Hepatitis C (NIH, 2002) concluded that liver biopsy is useful in defining baseline abnormalities of liver disease and in enabling patients and healthcare providers to reach a decision regarding antiviral therapy. The NIH Consensus Statement concludes that noninvasive tests are not adequate substitutes for liver biopsy.

Various noninvasive tests of hepatic fibrosis have been examined for monitoring patients with chronic hepatitis C virus (HCV) infection. These include routinely available laboratory tests, such as liver- associated chemistries, platelet count, and prothrombin time, as well as specific serum markers of fibrosis and inflammation not currently widely available or well validated. No single test or panel of serologic markers can provide an accurate assessment of intermediate stages of hepatic fibrosis. Similarly, quantitative tests of liver function and radiologic imaging of the liver are sensitive for diagnosing advanced cirrhosis but are not useful in assessing hepatic fibrosis and early cirrhosis.

In a review on newer markers for hepatocellular carcinoma, Marrero and Lok (2004) stated that there is a scarcity of longitudinal studies evaluating the ability of biomarkers to detect pre-clinical disease. There is an urgent need for novel biomarkers for the detection of early hepatocellular carcinoma.

Suzuki et al (2005) stated that “use of an accurate serum marker for severe hepatic fibrosis may also improve accuracy of non-invasive diagnostic models. Hyaluronic acid, a serum marker for severe hepatic fibrosis, has been reported to have a high diagnostic performance in assessing the severity of hepatic fibrosis in patients with alcoholic liver disease. In this issue, a non-invasive diagnostic model including hyaluronic acid was shown to have excellent performance in excluding the presence of medium to large esophageal varices in severe alcohol abusers. Based on current evidence, the strategy of using a non-invasive diagnostic model together with a serum marker for severe hepatic fibrosis may improve cost-benefit in the prevention of variceal hemorrhage among patients with alcoholic liver disease. The independent verification of such diagnostic models is needed”.

Evidence based guidelines on the management of hepatitis C from the American Association for the Study of Liver Diseases (Strader et al, 2004) stated: “Although liver fibrosis markers are commercially available, they are currently insufficiently accurate to support their routine use. Until sensitive serum markers can be developed that will define all stages of fibrosis and mirror the information derived from liver biopsy, the procedure remains the only means of defining the severity of damage from HCV infection in many patients”.

Serum gamma glutamyl-transferase (GGT) is elevated in individuals with acute and chronic alcohol toxicity. Serum GGT assay may be useful in evaluating patients when heavy drinking is suspected but the patient denies it (NIAAA, 2005).

Wilson et al (2006) stated that although most HCV infections are acquired by injection drug use, prospective data on the progression of liver fibrosis are sparse. In this study, baseline liver biopsies were obtained on a random sample of 210 out of 1,667 HCV-positive injection drug users (IDUs). Subjects were followed biannually, with a second biopsy offered to those eligible. Paired biopsies were scored 0 to 6 (modified Ishak score), significant fibrosis was defined as score 3 or greater, and progression of fibrosis was defined as an increase 2 or more units or clinical evidence of end-stage liver disease. Predictive values of blood markers (FibroSure, aspartate aminotransferase-to-platelet-ratio index (APRI) and alanine aminotransferase (ALT)) were assessed for detection of contemporaneous and future liver fibrosis. Among 119 prospectively followed IDUs, 96 % were African American; 97 % HCV genotype 1a/b; 27 % HIV-infected, and median age was 42 years. Most (90.7 %) did not have significant liver fibrosis at first biopsy. Although predictive value for detecting insignificant fibrosis at first biopsy was greater than 95 % for FibroSure, APRI, and ALT, specificities were 88.9 %, 72.7 %, and 72.7 %, respectively. After 4.2 years median follow-up, 21 % had progression of fibrosis, which was significantly associated with serum level of HCV RNA and ALT. No serological test had predictive value greater than 40 % for contemporaneous or future significant fibrosis. Even initial biopsy result had only a 30.4 % value for predicting future significant fibrosis. The authors concluded that significant liver fibrosis and progression were detected in some, but not most, IDUs in this cohort. In this setting with low fibrosis prevalence, FibroSure, ALT, and APRI tests predict insignificant fibrosis; however, further work is needed to find non-invasive markers of significant liver fibrosis.

Nourani and Pockros (2007) noted that biochemical markers are a potentially useful alternative to liver biopsy in patients with chronic hepatitis C aged 65 years and older. Furthermore, Rossi et al (2007) stated that an obstacle to widespread adoption of serum marker models (e.g., FibroSpect) for assessing liver fibrosis has been the lack of uniform performance indicators, such as diagnostic odds ratios and likelihood ratios. At present, serum marker models are not considered sufficiently reliable to replace liver biopsy in patients with chronic liver disease.

Shaheen and Myers (2008) performed a systematic review and meta-analysis of the diagnostic accuracy of fibrosis marker panels in patients with HIV/hepatitis C coinfection. Random effects meta-analyses and areas under summary receiver operating characteristics curves (AUC) examined test accuracy for detecting significant fibrosis (F2 to F4) and cirrhosis. Heterogeneity was explored using meta-regression. Five studies (n = 574) including 4 fibrosis measures (APRI [n = 4 studies], Forns’ [n = 2], FibroTest [n = 1], SHASTA [n = 1]) met the inclusion criteria. The prevalence of significant fibrosis and cirrhosis were 51 % and 16 %, respectively. For the prediction of significant fibrosis, the summary AUC was 0.82 (95 % confidence interval [CI]: 0.78 to 86) and diagnostic odds ratio was 7.8 (5.1 to 11.9). For cirrhosis, these figures were 0.83 (0.69 to 0.97) and 11.0 (4.6 to 26.2), respectively. Meta-regression including study factors (methodological quality and biopsy adequacy), patient characteristics (age, gender, CD4 count), and fibrosis measure failed to identify important predictors of accuracy. The authors concluded that available fibrosis marker panels have acceptable performance for identifying significant fibrosis and cirrhosis in HIV/HCV-coinfected patients but are not yet adequate to replace liver biopsy. They noted that additional studies are needed to identify the optimal measure.

Smith and Sterling (2009) reviewed non-invasive measures and their ability to replace biopsy for assessing hepatic fibrosis in patients with chronic HCV. A systematic review of PUBMED and EMBASE was carried out through 2008 using the following search terms: HCV, liver, elastography, hepatitis, Fibroscan, SPECT, non-invasive liver fibrosis, ultrasonography, Doppler, MRI, Fibrotest, Fibrosure, Actitest, APRI, Forns and breath tests, alone or in combination. These investigators identified 151 studies: 87 using biochemical, 57 imaging and 7 breath tests either alone or in combination. The authors concluded that great strides are being made in the development of accurate non-invasive methods for determination of fibrosis. Although no single non-invasive test or model developed to date can match that information obtained from actual histology (i.e., inflammation, fibrosis, steatosis), combinations of 2 modalities of non-invasive methods can reliably differentiate between minimal and significant fibrosis, and thereby avoid liver biopsy in a significant percentage of patients.

Carlson and colleagues (2009) evaluated the clinical and economic outcomes of non-invasive testing strategies in the diagnosis of significant liver fibrosis (Metavir score greater than or equal to 2) compared with liver biopsy. These researchers developed a decision analytic model of non-invasive testing strategies in a hypothetical patient population with genotype 1 hepatitis C virus infection, with no contraindications to liver biopsy. The testing strategies included a testing algorithm using the Fibrosure test, a non-invasive measure of fibrosis, followed by liver biopsy for patients with indeterminate results, Fibrospect II, and Fibroscan. The primary outcomes were sensitivity, specificity, diagnostic accuracy (true positive + true negatives/total patients), and costs, evaluated from the health-care payer perspective. The testing algorithm using Fibrosure was the most accurate non-invasive strategy with a sensitivity, specificity, and overall accuracy of 84 %, 87 %, and 86 %, respectively. In comparison with liver biopsy alone, there was a cost savings of approximately $770/person with the Fibrosure testing algorithm, but a net decrease in accuracy of 14 %. Fibrospect II and Fibroscan had poorer accuracy (decreases of 12 % and 4 %, respectively) and lower costs (-$138 and -$357, respectively) compared with the Fibrosure algorithm. In uncertainty analyses in which biopsy sampling error was considered, the Fibrosure algorithm remained consistently less accurate (5 to 14 % decrease). The authors concluded that the results of this study suggested that compared with liver biopsy, non-invasive testing algorithms can result in short-term cost savings, but the consequences of misdiagnosis in terms of health outcomes and treatment costs might out-weigh the short-term gains in cost and convenience.

Adams (2011) stated that fibrosis prediction is an essential part of the management of patients with chronic liver disease. Serum biomarkers offer a number of advantages over the traditional standard of fibrosis assessment of liver biopsy, including safety, cost-savings and wide spread accessibility. Current biomarker algorithms include indirect surrogate measures of fibrosis, including aminotransaminases and platelet count, or direct measures of fibrinogenesis or fibrinolysis such as hyaluronic acid and tissue inhibitor of metalloproteinase-1. A number of algorithms have now been validated across a range of chronic liver disease including chronic viral hepatitis, alcoholic and non-alcoholic fatty liver disease. Furthermore, several models have been demonstrated to be dynamic to changes in fibrosis over time and are predictive of liver-related survival and overall survival to a greater degree than liver biopsy. Current limitations of biomarker models include a significant indeterminate range, and a predictive ability that is limited to only a few stages of fibrosis. Utilization of these biomarker models requires knowledge of patient co-morbidities which may produce false positive or negative results in a small proportion of individuals. Furthermore, knowledge of the underlying prevalence of fibrosis in the patient population is required for interpretation of the positive or negative predictive values of a test result. Novel proteins identified by proteomic technology and genetic polymorphisms from genome association studies offer the possibility for further refinement and individualization of biomarker fibrosis models in the future.

Sebastiani and associates (2011) examined the effect of etiology and stages of hepatic fibrosis on the performance of fibrosis biomarkers. A total of 2,411 patients with compensated chronic liver disease (CLD) (hepatitis C virus [HCV] = 75.1 %, hepatitis B virus [HBV] = 10.5 %, non-alcoholic steato-hepatitis [NASH] = 7.9 %, HIV/HCV = 6.5 %) were consecutively enrolled in 9 centers. APRI, Forns’index, Lok index, AST-to-ALT ratio, Fib-4, platelets and Fibrotest-Fibrosure were tested against liver biopsy, considered the gold standard. The effect of the stages of hepatic fibrosis to diagnose significant fibrosis and cirrhosis (greater than or equal to F2 and F4, respectively) was investigated through difference between advanced and non-advanced fibrosis stages (DANA). Performance was expressed as observed area under the receiver-operating characteristic (ROC) curve (ObAUROC) and AUROC adjusted for DANA (AdjAUROC). Performance of APRI and Fibrotest-Fibrosure was higher than other biomarkers. In all etiologies, AdjAUROC was higher than ObAUROC. APRI showed its best performance in HCV mono-infected cases, with an AdjAUROC of 0.77 and 0.83 for greater than or equal to F2 and F4, respectively. In HBV and NASH patients, its performance was poor (AdjAUROC < 0.70). Performance of Fibrotest-Fibrosure was good in all etiologies for both greater than or equal to F2 and F4 (AdjAUROC > 0.73), except for greater than or equal to F2 in NASH (AdjAUROC = 0.64). Performance of all biomarkers was reduced in HCV cases with normal ALT. The authors concluded that etiology is a major factor influencing the performance of liver fibrosis biomarkers. Even after correction for DANA, APRI and Fibrotest-Fibrosure exhibit the best performance. However, liver biopsy is not replaceable, especially to diagnose greater than or equal to F2 and in HCV carriers with normal ALT.

Adams et al (2005) stated that staging hepatic fibrosis by liver biopsy guides prognosis and treatment of hepatitis C, but is invasive and expensive. These researchers sought to create an algorithm of serum markers that accurately and reliably predict liver fibrosis stage among hepatitis C patients. A total of 10 biochemical markers were measured at time of liver biopsy in 117 untreated hepatitis C patients (training set). Multi-variate logistic regression and ROC curve analyses were used to create a predictive model for significant fibrosis (METAVIR F2, F3, and F4), advanced fibrosis (F3 and F4), and cirrhosis (F4). The model was validated in 104 patients from other institutions. A model (HepaScore) of bilirubin, GGT, hyaluronic acid (HA), alpha(2)-macroglobulin, age, and sex produced areas under the ROC curves (AUCs) of 0.85, 0.96, and 0.94 for significant fibrosis, advanced fibrosis, and cirrhosis, respectively. In the training set, a score greater than or equal to 0.5 (range of 0.0 to 1.0) was 92 % specific and 67 % sensitive for significant fibrosis, a score of less than 0.5 was 81 % specific and 95 % sensitive for advanced fibrosis, and a score of less than 0.84 was 84 % specific and 71 % sensitive for cirrhosis. Among the validation set, the AUC for significant fibrosis, advanced fibrosis, and cirrhosis were 0.82, 0.90, and 0.89, respectively. A score greater than or equal to 0.5 provided a specificity and sensitivity of 89 % and 63 % for significant fibrosis, whereas scores less than 0.5 had 74 % specificity and 88 % sensitivity for advanced fibrosis, respectively. The authors concluded that a model of 4 serum markers plus age and sex provides clinically useful information regarding different fibrosis stages among hepatitis C patients.

Ngo et al (2006) compared the 5-year prognostic value of the FibroTest with biopsy staging for predicting cirrhosis decompensation and survival in patients with chronic HCV infection. Fibrosis stage was assessed on the same day by FibroTest and biopsy in a prospective cohort of 537 patients. Disease classification at baseline was 157 patients with severe fibrosis (FibroTest >0.58), 137 with moderate fibrosis (FibroTest 0.32-0.58), and 243 with no or minimal fibrosis (FibroTest < 0.32). In 64 untreated patients with severe fibrosis, survival without HCV complications was 73% [95% CI, 59%-086%; 13 complications], and survival without HCV-related death was 85% (95% CI, 73%-96%; 7 HCV deaths). Survival rates were higher in patients with moderate fibrosis, [99% (95% CI, 97%-100%; 1 complication; P <0.001) and 100% (no HCV death; P <0.001) for patients with and without HCV-related complications, respectively], and in patients with minimal fibrosis [100% (no complication; P <0.001 vs severe) and 100% (no HCV death; P <0.001 vs severe), respectively]. FibroTest was a better predictor than biopsy staging for HCV complications, with area under the ROC curves (AUROC) = 0.96 (95% CI, 0.93%-0.97%) vs 0.91 (95% CI, 0.85%-0.94%; P = 0.01), respectively; it was also a better predictor for HCV deaths: AUROC = 0.96 (95% CI, 0.93%-0.98%) vs 0.87 (95% CI, 0.70%-0.94%; P = 0.046), respectively. The prognostic value of FibroTest was still significant (P <0.001) in multivariate analyses after taking into account histology, treatment, alcohol consumption, and HIV coinfection.

Cacoub et al (2008) compared non-invasive biological liver fibrosis scores, as alternatives to liver biopsy, in HIV/HCV co-infected patients. Two hundred and seventy-two HIV/HCV patients, naive for HCV treatment, underwent liver biopsy [197 (72%) men, 39.9 years, fibrosis stage (Metavir) F1 (25%), F2 (40%), F3 (25%), F4 (10%), median CD4 486/mm(3) and median HIV viral load 3.5 log. Fibrotest (FT), Hepascore (HS), Fibrometer (FM), SHASTA, APRI, Forns index, and Fib-4 were tested in order to differentiate patients with mild to moderate fibrosis (F2) and those with advanced fibrosis (F3). The AUROC and the rate of well-classified patients were compared to liver biopsy. FT, HS, and FM were able to stage liver fibrosis in all patients with AUROCs of 0.78, 0.84 and 0.89 for the diagnosis of F2, respectively. The correlation coefficient indexes were 0.37, 0.46 and 0.48, respectively. The rates of well-classified patients were 62%, 68% and 71%, respectively. Fib-4, APRI and the Forn’s index were only able to stage 37-61% of patients and showed lower accuracies. Using a combination of FT, HS and FM did not significantly increase the performance of each test. The investigators concluded that, in HIV/HCV co-infected patients, Fibrometer, Hepascore and Fibrotest outperformed other non-invasive liver fibrosis biomarkers for the prediction of significant liver fibrosis.

Halfon et al (2008) updated a previous meta-analysis of Fibrotest (FT) diagnostic value. For diagnostic value, the main endpoint was the FT area under the ROC curves (AUROCs) for the diagnosis of bridging fibrosis (F2/F3/F4 vs F0/F1), standardized for the spectrum of fibrosis. Sensitivity analysis integrated the non-standardized observed AUROCs, the independency of authors, size (length) of biopsy, prospective design, correctness of procedures, co-morbidities, and time-lag between biopsy and serum sampling. For prognostic value, the main endpoint was the FT AUROC for the prognostic value of liver complications or death related to liver disease. A total of 38 diagnostic studies were included, which pooled 7985 subjects who had undergone both FT and biopsy (4600 HCV, 1580 HBV, 267 NAFLD, 524 ALD and 1014 mixed). The mean standardized AUROC was 0.84 (95% CI, 0.83-0.86), with no differences in terms of causes of liver disease: HCV 0.84 (0.82-0.87); HBV 0.81 (0.78-0.83); NAFLD 0.84 (0.76-0.92); ALD 0.87 (0.82-0.92); and mixed 0.85 (0.81-0.89). Three prognostic studies were also included. FT was found to have higher or similar prognostic value compared with biopsy in patients with chronic hepatitis C (CHC), CHB or ALD.

Sebastiani and Alberti (2012) chronic hepatitis C represents a major cause of progressive liver disease that can eventually evolve into cirrhosis and its end-stage complications. Formation and accumulation of fibrosis in the liver is the common pathway that leads to evolutive liver disease. Precise staging of liver fibrosis is essential for patient management in clinical practice because the presence of bridging fibrosis represents a strong indication for anti-viral therapy, while cirrhosis requires a specific follow-up. Liver biopsy has always represented the standard of reference for assessment of hepatic fibrosis, but it has limitations: it is invasive, costly and prone to sampling errors. Recently, blood markers and instrumental methods have been proposed for the non-invasive assessment of liver fibrosis in hepatitis C. However, international guidelines do not recommend the widespread use of non-invasive methods for liver fibrosis in clinical practice. This is because of, in some cases, unsatisfactory accuracy and incomplete validation of others. Some studies suggested that the effectiveness of non-invasive methods for assessing liver fibrosis may increase when they are combined, and a number of sequential and synchronous algorithms have been proposed for this purpose, with the aim of reducing rather than substituting liver biopsies. This may represent a rational and reliable approach for implementing noninvasive assessment of liver fibrosis in clinical practice. It could allow more comprehensive first-line screening of liver fibrosis in hepatitis C than would be feasible with liver biopsy alone.

Usluer et al (2012) compared the results of 9 non-invasive serum biomarkers with liver biopsies to predict liver fibrosis stage. HCV-RNA-positive, HCV genotype 1, treatment-naive patients with chronic HCV infections were included from 14 centers (n = 77). The platelet count, AST/ALT ratio (AAR), cirrhosis discriminate score (CDS), FIB4, APRI, age-platelet (AP) index, Goteborg University cirrhosis index (GUCI), FibroTest, and ActiTest were calculated and compared to histologic findings. All serum biomarkers, except AAR, were weakly or moderately correlated with liver biopsy results (ISHAK fibrosis score). The mean scores of FibroTest, FIB4, APRI, and AP index were significantly different between F0-F2 and F3-F4 groups and the negative predictive values (NPVs) of the F3-F4 group were 95 %, 85 %, 85 %, and 8 3%, respectively, for these serum biomarkers. The authors concluded that these findings suggested that serum biomarkers may help to diagnose significant fibrosis but inadequate to detect fibrosis in early stages.

Bhogal and Sterling (2012) noted that several blood tests, algorithms, and imaging tests have been studied as non-invasive markers to stage fibrosis in hepatitis C. In patients without suspicion for cirrhosis, 2 non-invasive methods can be used to predict presence of absence of significant liver fibrosis; however, liver biopsy remains the gold standard.

Chladek et al (2013) compared the results of serial measurements of serum fibrosis markers during the remission-induction phase of treatment with methotrexate (MTX) to those of patients on biological therapy and long-term MTX therapy (greater than 2 years). Serum concentrations of HA, N-terminal pro-peptide of collagen type III (PIIINP) and the results of 2 multi-test algorithms FibroTest and HepaScore were evaluated in patients with chronic plaque psoriasis (n = 24, age: 28 to 79 years, baseline Psoriasis Area Severity Index [PASI] 13.5, range of 2.2 to 33) at baseline and weeks 16 and 26 after the start of pharmacokinetically-guided therapy with MTX (Group A). Patients on established therapy with biologics (n = 15, Group B) and long-term MTX users (n = 10, Group C) with the mean baseline PASI scores of 0.9 and 1.2 were studied in parallel cohorts. At baseline, HA, HepaScore and PIIINP were correlated with PASI of Group A patients. At weeks 16 and 26, HA decreased by 48 % and 40 % (p < 0.001) and HepaScore by 31 (p < 0.01) and 20 % (p < 0.05) respectively. PASI75 (greater than or equal to 75 % improvement from baseline PASI) was observed in 76 % of Group A patients by week 26 and the absolute decreases in PASI and both fibrosis markers were correlated (HA: r = 0.49, p = 0.018, HepaScore: r = 0.47, p = 0.022). In contrast, no significant within-group differences were found in HA and HepaScore results of patients in the groups B and C. PIIINP and FibroTest were stable in all groups. The authors concluded that the fibrosis markers HA and HepaScore (the multiple test algorithm that includes HA) are less liver specific and more prone to reflect psoriasis activity than PIIINP and FibroTest.

Salkic et al (2014) systematically reviewed studies describing the diagnostic accuracy of Fibrotest (FT) for predicting chronic hepatitis B (CHB)-related fibrosis. MEDLINE and EMBASE searches and hand searching methods were performed to identify studies that assessed the diagnostic accuracy of FibroTest in HB patients using LB as a reference standard. The investigators used a hierarchical summary receiver operating curves model and the bivariate model to produce summary receiver operating characteristic curves and pooled estimates of sensitivity and specificity. The investigators included 16 studies (N=2494) and 13 studies (N=1754) in the heterogenous meta-analysis for liver fibrosis and cirrhosis, respectively. The area under the hierarchical summary receiver operating curve for significant liver fibrosis and for all included studies was 0.84 (95% confidence interval (CI): 0.78-0.88). At the FT threshold of 0.48, the sensitivity, specificity, and diagnostic odds ratio (DOR) of FT for significant fibrosis were 61 (48-72%), 80 (72-86%), and 6.2% (3.3-11.9), respectively. The area under the hierarchical summary receiver operating curve for liver cirrhosis and for all included studies was 0.87 (95% CI: 0.85-0.90). At the FT threshold of 0.74, the sensitivity, specificity, and DOR of FT for cirrhosis were 62 (47-75%), 91 (88-93%), and 15.7% (8.6-28.8), respectively. The authors concluded that FibroTest is of value in exclusion of patients with CHB-related cirrhosis, but has suboptimal accuracy in the detection of significant fibrosis and cirrhosis. It is necessary to further improve the test or combine it with other noninvasive modalities in order to improve accuracy.

Chou and Wasson (2013) stated that many blood tests have been proposed as alternatives to liver biopsy for identifying fibrosis or cirrhosis. These investigators evaluated the diagnostic accuracy of blood tests to identify fibrosis or cirrhosis in patients with HCV infection. Data sources included MEDLINE (1947 to January 2013), the Cochrane Library, and reference lists. Studies that compared the diagnostic accuracy of blood tests with that of liver biopsy were selected. Investigators abstracted and checked study details and quality by using pre-defined criteria. A total of 172 studies evaluated diagnostic accuracy. For identifying clinically significant fibrosis, the platelet count, age-platelet index, APRI, FibroIndex, FibroTest, and Forns index had median positive likelihood ratios of 5 to 10 at commonly used cut-offs and areas under the ROC curve (AUROCs) of 0.70 or greater (range of 0.71 to 0.86). For identifying cirrhosis, the platelet count, age-platelet index, APRI, and HepaScore had median positive likelihood ratios of 5 to 10 and AUROCs of 0.80 or greater (range of 0.80 to 0.91). The GUCI and the Lok index had slightly lower positive likelihood ratios (4.8 and 4.4, respectively). In direct comparisons, the APRI was associated with a slightly lower AUROC than the FibroTest for identifying fibrosis and a substantially higher AUROC than the aspartate aminotransferase-alanine aminotransferase ratio for identifying fibrosis or cirrhosis. The authors concluded that many blood tests are moderately useful for identifying clinically significant fibrosis or cirrhosis in HCV-infected patients. Drawbacks of this study included only English-language articles were included, and most studies had methodological limitations, including failure to describe blinded interpretation of liver biopsy specimens and inadequate description of enrollment methods.

Rossi et al (2013) noted that serum HA and biochemical models that require HA analysis are commonly used as predictors of liver fibrosis in patients with chronic liver disease, however biological variation data for HA are deficient. Four serial serum samples were obtained at weekly intervals from healthy volunteers and patients with chronic hepatitis B, chronic hepatitis C and non-alcoholic fatty liver disease (NAFLD) (20 in each group). The within-individual week-to-week variation (CVI) and reference change values for HA, α₂-macroglobulin and HepaScore were obtained. HepaScore was calculated from HA, α2-macroglobulin, bilirubin and GGT activity. Hyaluronic acid displayed large within-individual variation, the CVI values were 62 % in healthy subjects, 38 % in hepatitis C, 37 % in hepatitis B, and 36 % in NAFLD patients. HepaScore CVIs were 43 % in healthy subjects, 24 % in hepatitis C, 28 % in hepatitis B, and 39 % in NAFLD patients. Moreover, α₂-Macroglobulin was much less variable with CVIs ranging from 4.4 % to 7.6 %. Bland-Altman plots of week-to-week variations showed rates of significant disagreement for samples collected in any 2 successive weeks varied from 5 % in NAFLD patients to 8.3 % in healthy subjects. The authors concluded that when using non-fasting serum samples, HA and to a lesser extent, the HepaScore model displayed large within-individual variations in both health and chronic liver disease. This information is critical for interpreting the significance of both single measurements and changes in serial measurements.

Grattagliano et al (2013) stated that the diagnostic utilities of ultrasonography (US), fatty liver index (FLI) and an algorithm of 9 serum markers (FibroMAX) were evaluated in family practice to non-invasively characterize patients with NAFLD. A multi-center study was conducted by enrolling 259 consecutively observed patients (age of 51 +/- 10 years) with clinical and US features of NAFLD. Patients had mild (16.2 %), moderate (69.9 %), or severe (13.9 %) liver steatosis and 60.2 % had hyper-transaminasemia. The percent of patients with overweight, obesity, diabetes, hypertension, and dyslipidemia were 42.7 %, 46.5 % (4.2 % severe obesity), 24.7 %, 40.9 %, and 56.4 %, respectively. Lean patients (10.8 %) had normal transaminases in 2/3 of the cases. A multi-variate logistic regression (including age greater than 50 years, body mass index (BMI) greater than 30 kg/m2, homeostasis model assessment [HOMA] greater than 3, and hyper-transaminasemia) identified 12.3 % of patients at risk for steatohepatitis. With a sensitivity of 50 % and specificity of 94.7 %, FibroMAX identified 34 patients (13.1 %) with likely advanced fibrosis and found that over 28 % of patients with moderate (ultrasonographic) steatosis were likely to be carrying severe steatosis. Steatotest score was significantly associated with BMI, waist circumference, ALT, triglycerides, and FLI. FibroTest correlated only with ALT. Fatty liver index identified 73.4 % of patients as likely to be carrying a fatty liver. The authors concluded that NAFLD should be systematically searched and characterized in all patients with metabolic disturbances and cardiovascular risk. Asymptomatic subjects at risk also should be screened for NAFLD. They stated that FibroMAX is a promising non-invasive diagnostic tool in family medicine for identifying patients at risk for NAFLD who require targeted follow-up.

Xu et al (2014) conducted a systematic review on records in PubMed, EMBASE and the Cochrane Library electronic databases, up until April 1st, 2013, in order to systematically assess the effectiveness and accuracy of these biomarkers for predicting HBV-related fibrosis. The questionnaire for quality assessment of diagnostic accuracy studies (QUADAS) was used. Out of 115 articles evaluated for eligibility, 79 studies satisfied the pre-determined inclusion criteria for meta-analysis. The authors final data set for the meta-analysis contained 30 studies. The areas under the SROC curve for APRI, FIB-4, and FibroTest of significant fibrosis were 0.77, 0.75, and 0.84, respectively. For cirrhosis, the areas under the SROC curve for APRI, FIB-4 and FibroTest were 0.75, 0.87, and 0.90, respectively. The heterogeneity of FIB-4 and FibroTest were not statistically significant. The heterogeneity of APRI for detecting significant fibrosis was affected by median age (P = 0.0211), and for cirrhosis was affected by etiology (P = 0.0159). Based on the analysis the authors concluded that FibroTest has excellent diagnostic accuracy for identification of HBV-related significant fibrosis and cirrhosis. FIB-4 has modest benefits and may be suitable for wider scope implementation.

In their review, Mato et al (2019) stated nonalcoholic fatty liver disease (NAFLD) is a heterogeneous and complex disease that is imprecisely diagnosed by liver biopsy. NAFLD covers a spectrum that ranges from simple steatosis, nonalcoholic steatohepatitis (NASH) with varying degrees of fibrosis, to cirrhosis, which is a major risk factor for hepatocellular carcinoma. Lifestyle and eating habit changes during the last century have made NAFLD the most common liver disease linked to obesity, type 2 diabetes mellitus and dyslipidemia, with a global prevalence of 25%. NAFLD arises when the uptake of fatty acids (FA) and triglycerides (TG) from circulation and de novo lipogenesis saturate the rate of FA β-oxidation and very-low density lipoprotein (VLDL)-TG export. Deranged lipid metabolism is also associated with NAFLD progression from steatosis to NASH, and therefore, alterations in liver and serum lipidomic signatures are good indicators of the disease’s development and progression. This review focuses on the importance of the classification of NAFLD patients into different subtypes, corresponding to the main alteration(s) in the major pathways that regulate FA homeostasis leading, in each case, to the initiation and progression of NASH. This concept also supports the targeted intervention as a key approach to maximize therapeutic efficacy and opens the door to the development of precise NASH treatments. The authors state that a challenge in NAFLD research is the identification of which patients with NAFLD will develop NASH and, for those with NASH, how fast the disease will progress. At present, it is premature to conclude which of these blood biomarkers, alone or in combination, would be best to precisely and rapidly diagnose the severity of NASH and monitor the liver’s response to treatment.

In their review, Zhou et al (2019) state with the increasing number of individuals with diabetes and obesity, nonalcoholic fatty liver disease (NAFLD) is becoming increasingly prevalent, affecting one-quarter of adults worldwide. The spectrum of NAFLD ranges from simple steatosis or nonalcoholic fatty liver (NAFL) to nonalcoholic steatohepatitis (NASH). NAFLD, especially NASH, may progress to fibrosis, leading to cirrhosis and hepatocellular carcinoma. NAFLD can impose a severe economic burden, and patients with NAFLD-related terminal or deteriorative liver diseases have become one of the main groups receiving liver transplantation. The increasing prevalence of NAFLD and the severe outcomes of NASH make it necessary to use effective methods to identify NAFLD. Although recognized as the gold standard, biopsy is limited by its sampling bias, poor acceptability, and severe complications, such as mortality, bleeding, and pain. Therefore, noninvasive methods are urgently needed to avoid biopsy for diagnosing NAFLD. This review discusses the current noninvasive methods for assessing NAFLD, including steatosis, NASH, and NAFLD-related fibrosis, and explores the advantages and disadvantages of measurement tools. In addition, the authors analyze potential noninvasive biomarkers for tracking disease processes and monitoring treatment effects, and explore effective algorithms consisting of imaging and non-imaging biomarkers for diagnosing advanced fibrosis and reducing unnecessary biopsies in clinical practice. The authors state there are currently no effective noninvasive biomarkers recommended for diagnosing NASH. Future studies are needed to investigate more efficient noninvasive biomarkers for distinguishing NASH from simple steatosis.

Furthermore, an UpToDate review on “Tests used for the noninvasive assessment of hepatic fibrosis” (Curry and Afdhal, 2013) states that “While tremendous progress has been made in improving the accuracy of serum markers of hepatic fibrosis, they cannot yet supplant direct analysis of the liver. The ideal fibrosis marker is one that is specific, biologically based, noninvasive, easily repeated in all patients, correlates well with disease severity and outcome, and is not confounded by co-morbidities or drugs. Although this ideal has nearly been reached, no serum test has emerged as the perfect marker of fibrosis; all the serum tests have limitations …. Overall, the serum assay approaches remain promising, in part because these tests may represent an integrated readout of liver activity, rather than a minute sampling of the type obtained by conventional liver biopsy”.

In a UpToDate review “Noninvasive assessment of hepatic fibrosis: Overview of serologic and radiographic tests”, (Curry and Afdhal, 2019), the authors state that noninvasive tests of hepatic fibrosis have been used in many clinical scenarios. The majority of studies of serologic markers and radiologic tests have looked at the use of these tests for staging of fibrosis in patients with chronic liver disease. The authors typically consider noninvasive testing for patients presenting for evaluation of chronic viral hepatitis. However, all the serum tests have limitations, such as they typically reflect the rate of matrix turnover, not deposition, and thus tend to be more elevated when there is high inflammatory activity. By contrast, extensive matrix deposition can go undetected if there is minimal inflammation. Also, none of the markers are liver-specific, and concurrent sites of inflammation or fibrosis may contribute to serum levels. Serum levels are affected by clearance rates, which may be impaired either due to sinusoidal endothelial cell dysfunction or impaired biliary excretion. Lastly, serum tests are surrogates, not biomarkers.

Houot et al (2016) selected studies from 2002 to 2014 that directly compared the diagnostic accuracy of FibroTest, aspartate aminotransferase-platelet ratio index (APRI), FIB4 index or transient elastrography (TE), with biopsy as a reference, in patients with CHC or chronic hepatitis B (CHB). Investigators abstracted and checked study details and quality by using pre-defined criteria. Bayesian method in intention to diagnose was the primary outcome. Of 1321 articles identified, 71 studies including 77 groups according to etiology (All-CB) were eligible: 37 Only-C, 28 Only-B and 12 Mixed-C-B. There were 185 direct comparisons between the area under the ROC curves (AUROCs), 99 for the diagnosis of advanced fibrosis and 86 for cirrhosis. In All-CB, Bayesian analyses revealed significant AUROCs differences in identifying advanced fibrosis in favor of FibroTest vs. TE [credibility interval: 0.06(0.02-0.09)], FibroTest vs. APRI [0.05 (0.03-0.07)] and for identifying cirrhosis TE vs. APRI [0.07 (0.02-0.13)] and FIB4 vs. APRI [0.04(0.02-0.05)]. No differences were observed between TE and FibroTest, for identifying cirrhosis in All-CB, and in sub-groups (Only-C, Only-B, Mixed-CB) for both cirrhosis and fibrosis. The investigators concluded that in CHC and CHB, APRI had lower performances than FIB-4, TE and FibroTest. TE had lower performance than FibroTest for identifying advanced fibrosis in All-CB, without significant difference for identifying cirrhosis in all groups.

Neuman et al (2016) noted that chronic liver diseases may cause inflammation and progressive scarring, over time leading to irreversible hepatic damage (cirrhosis). As a result, the need to assess and closely monitor individuals for risk factors of components of matrix deposition and degradation, as well as the severity of the fibrosis using biomarkers, has been increasingly recognized. These investigators reviewed the use of biomarker for diagnosing and defining the severity of liver fibrosis. A systematic literature review was performed using the terms “hyaluronic acid” and “liver fibrosis” as well as the name of each biomarker or algorithm known to be employed. PubMed and Google Scholar were searched, and English language articles indexed between January 2010 and October 2014 in which HA was used as a marker of liver fibrosis were retrieved, regardless of the underlying liver disease. Each author read the publications separately and the results were analyzed and discussed. Biomarkers offer a potential prognostic or diagnostic indicator for disease manifestation, progression, or both. Serum biomarkers, including HA, have been used for many years. Emerging biomarkers such as metalloproteinases have been proposed as tools that provide valuable complementary information to that obtained from traditional biomarkers. Moreover, markers of extracellular matrix degradation provide powerful predictions of risk. In order for biomarkers to be clinically useful in accurately diagnosing and treating disorders, age-specific reference intervals that account for differences in gender and ethnic origin are a necessity.

World Health Organization guidelines on hepatitis B (WHO, 2015) state that “aspartate aminotransferase (AST)-to-platelet ratio index (APRI) is recommended as the preferred non-invasive test (NIT) to assess for the presence of cirrhosis (APRI score >2 in adults) in resource-limited settings. Transient elastography (e.g., FibroScan) or FibroTest may be the preferred NITs in settings where they are available and cost is not a major constraint. (Conditional recommendation, low quality of evidence).”

Guidelines on hepatitis C from the American Association for the Study of Liver Disease (AASLD) (Terrault et al, 2015) state that “[e]valuation for stage of disease using noninvasive methods or liver biopsy is useful in guiding treatment decisions including duration of therapy.” The guidelines explain that: “Serum markers of fibrosis, such as aspartate aminotransferase (AST)-to-platelet ratio index (APRI), FIB-4, FibroTest, and vibration-controlled transient elastography, have only moderate accuracy in identifying persons with significant fibrosis (fibrosis stage 2 or greater on the Metavir scale), but good diagnostic accuracy in excluding advanced fibrosis and may be useful aids in decision making.”

The 2019 hepatitis C guidance update from the American Association for the Study of Liver Diseases (AASLD) and the Infectious Diseases Society of America (IDSA) (Ghany 2020) state assessing liver disease severity is an essential component of the workup for all persons with newly diagnosed chronic hepatitis C as this factor influences initial and follow-up evaluation This assessment (i.e., presence or absence of cirrhosis) can usually be accomplished with noninvasive tests. Liver biopsy is rarely required but is a consideration if other causes of liver disease are suspected. Noninvasive tests to assess liver Disease Severity include: liver-directed physical exam (normal in most patients); routine blood tests (e.g., ALT, AST, albumin, bilirubin, INR, and CBC with platelet count); serum fibrosis marker panels; transient elastography, liver imaging (e.g., ultrasound or computed tomography scan); AST-to-platelet ratio index; and FIB-4 score.

The guidance further states that the simplified HCV treatment algorithm for adults without cirrhosis who have not been previously treated for their infection and do not have evidence of cirrhosis as defined by the noninvasive parameters specified in the HCV guidance. The guidance further describes evidence of cirrhosis as a FIB-4 score > 3.25, or any of the following findings from a previously performed test: transient elastography indicating cirrhosis (e.g., FibroScan [Echosens, Paris, France] stiffness more than 12.5 kPa), noninvasive serologic tests that exceed proprietary cutoffs (e.g., FibroSure [BioPredictive, Paris, France], Enhanced Liver Fibrosis Test [Siemens Healthcare, Erlangen, Germany], etc.), clinical evidence of cirrhosis (e.g., liver nodularity and/or splenomegaly on imaging, platelet count < 150,000/mm3, etc.), and/or prior liver biopsy showing cirrhosis.

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Castiella et al (2010) stated that advances in recent years in the understanding of, and the genetic diagnosis of hereditary hemochromatosis (HH) have changed the approach to iron overload hereditary diseases. The ability to use a radiologic tool (MRI) that accurately provides liver iron concentration (LIC) determination, and the presence of non-invasive serologic markers for fibrosis prediction (serum ferritin, platelet count, transaminases, etc.), have diminished the need for liver biopsy for diagnosis and prognosis of this disease. Consequently, the role of liver biopsy in iron metabolism disorders is changing. Furthermore, the irruption of transient elastography to assess liver stiffness, and, more recently, the ability to determine liver fibrosis by means of MRI elastography will change this role even more, with a potential drastic decline in hepatic biopsies in years to come. These investigators provided a brief summary of the different non-invasive methods available nowadays for diagnosis and prognosis in HH, and pointed out potential new techniques that could come about in the next years for fibrosis prediction, thus avoiding the need for liver biopsy in a greater number of patients. It is possible that liver biopsy will remain useful for the diagnosis of associated diseases, where other non-invasive means are not possible, or for those rare cases displaying discrepancies between radiological and biochemical markers. The authors concluded that based on the advances during the last few years, biochemical markers, LIC determination by MRI (Fibrosis index) and FibroScan and, probably, MR elastography, all constitute reliable non-invasive means for detecting liver fibrosis. The role of liver biopsy in the study of hemochromatosis is decreasing. In future, it appeared that liver biopsy will only be performed for diagnosis of associated diseases, or in patients where discrepancies between radiologic and biochemical markers exist. These investigators believed it is time to take a step forward and to reduce the “faith” in liver biopsy in favor of non-invasive methods for liver fibrosis prediction.

Also, an UpToDate review on “Clinical manifestations and diagnosis of hereditary hemochromatosis” (Schrier and Bacon, 2018) states that “with the availability of ultrasound-based elastography as a test for detecting advanced hepatic fibrosis, it is even less likely that the liver biopsy would be performed in those with HH. These non-invasive ways of detecting fibrosis to identify patients who are candidates for screening for varices and liver cell cancers seem much more reasonable than a liver biopsy”.

Enhanced Liver Fibrosis (ELF) Test

The Enhanced Liver Fibrosis Test by Siemens Healthcare is a blood based analysis of 3 biomarkers (hyaluronic acid [HA], procollagen III amino terminal peptide [PIIINP], and tissue inhibitor of metalloproteinase 1 [TIMP-1]). Sherman et al (2020) state noninvasive fibrosis markers are routinely used in patients with liver disease. Magnetic resonance elastography (MRE) is recognized as a highly accurate methodology, but a reliable blood test for fibrosis would be useful. The authors examined performance characteristics of the Enhanced Liver Fibrosis (ELF) Index compared to MRE in a cohort including those with HCV, HIV, and HCV/HIV. Subjects enrolled in the Miami Adult Studies on HIV (MASH) cohort underwent MRE and blood sampling. The ELF Index was scored and receiver-operator curves constructed to determine optimal cutoff levels relative to performance characteristics. Cytokine testing was performed to identify new markers to enhance noninvasive marker development. The ELF Index was determined in 459 subjects; more than half were male, non-white, and HIV-infected. MRE was obtained on a subset of 283 subjects and the group that had both studies served as the basis of the receiver-operator curve analysis. At an ELF Index of > 10.633, the area under the curve for cirrhosis (Metavir F4, MRE > 4.62 kPa) was 0.986 (95% CI 0.994-0.996; p < 0.001) with a specificity of 100%. For advanced fibrosis (Metavir F3/4), an ELF cutoff of 10 was associated with poor sensitivity but high specificity (98.9%, 95% CI 96.7-99.8%) with an AUC of 0.80 (95% CI 0.749-0.845). ELF Index performance characteristics exceeded FIB-4 performance. HCV and age were associated with increased fibrosis (p < 0.05) in a multivariable model. IP-10 was found to be a promising biomarker for improvement in noninvasive prediction algorithms. The authors concluded that the ELF Index was a highly sensitive and specific marker of cirrhosis, even among HIV-infected individuals, when compared with MRE. IP-10 may be a biomarker that can enhance performance characteristics further, but additional validation is required.

Inadomi et al (2020) state the Enhanced Liver Fibrosis (ELF) test comprises a logarithmic algorithm combining three serum markers of hepatic extracellular matrix metabolism. The authors aimed to evaluate the performance of ELF for the diagnosis of liver fibrosis and to compare it with that of liver stiffness measurement (LSM) by FibroScan in non-alcoholic fatty liver disease (NAFLD). ELF cut-off values for the diagnosis of advanced fibrosis were obtained using receiver operating characteristic (ROC) analysis in patients with biopsy-confirmed NAFLD (training set; n=200). Diagnostic performance was analyzed in the training set and in a validation set (n=166), and compared with that of LSM in the FibroScan cohort (n=224). The area under ROC curve was 0.81 for the diagnosis of advanced fibrosis and the ELF cut-off values were 9.34 with 90.4% sensitivity and 10.83 with 90.6% specificity in the training set, and 89.8% sensitivity and 85.5% specificity in the validation set. There was no significant difference in the area under the ROC curve between ELF and LSM (0.812 and 0.839). A combination of ELF (cut-off 10.83) and LSM (cut-off 11.45) increased the specificity to 97.9% and the positive predictive value, versus ELF alone. Sequential use of the fibrosis-4 index (cut-off 2.67) and ELF (cut-off 9.34) increased the sensitivity to 95.9%. ELF can identify advanced liver fibrosis in NAFLD and its diagnostic accuracy is comparable to that of FibroScan. According to the clinical setting, combinations or sequential procedures using other non-invasive tests complement the diagnostic performance of ELF for the identification of advanced fibrosis.

The July 2016 National Institute for Health and Care Excellence (NICE) guidance for the assessment for advanced liver fibrosis in people with non-alcoholic fatty liver disease (NAFLD) recommends considering using the enhanced liver fibrosis (ELF) test in people who have been diagnosed with NAFLD to test for advanced liver fibrosis (Glen 2016).

FibroTest-ActiTest / HCV-FibroSure for Diagnosing / Monitoring of Primary Biliary Cholangitis

UpToDate reviews on “Clinical manifestations, diagnosis, and prognosis of primary biliary cholangitis (primary biliary cirrhosis)” (Poupon, 2021a), and “Overview of the management of primary biliary cholangitis” (Poupon, 2021b) do not mention FibroTest-ActiTest and HCV-FibroSure as a management tools.

Micro-Fibrillar Associated Glycoprotein 4 (MFAP4)

Molleken et al (2016) noted that several comparable mechanisms have been identified for hepatic and pulmonary fibrosis. The human micro-fibrillar associated glycoprotein 4 (MFAP4), produced by activated myofibroblasts, is a ubiquitous protein playing a potential role in extra-cellular matrix (ECM) turnover and was recently identified as biomarker for hepatic fibrosis in hepatitis C patients. These researchers evaluated serum levels of MFAP4 in patients with pulmonary fibrosis in order to test its potential as biomarker in clinical practice. They also examined if MFAP4 deficiency in mice affects the formation of pulmonary fibrosis in the bleomycin model of lung fibrosis. A total of 91 patients with idiopathic pulmonary fibrosis (IPF), 23 with hypersensitivity pneumonitis (HP) and 31 healthy subjects were studied. In the mouse model, C57BL/6 Mfap4+/+ and Mfap4-/- mice between 6 to 8 weeks of age were studied. Serum levels of MFAP4 were measured by ELISA in patients and in mice. Surfactant protein D (SP-D) and LDH were measured as comparison biomarkers in patients with pulmonary fibrosis. Morphometric assessment and the Sircol kit were used to determine the amount of collagen in the lung tissue in the mouse model. Serum levels of MFAP4 were not elevated in lung fibrosis – neither in the patients with IPF or HP nor in the animal model. Furthermore no significant correlations with pulmonary function tests of IPF patients could be found for MFAP4; MFAP4 levels were increased in BAL of bleomycin-treated mice with pulmonary fibrosis. The authors concluded that MFAP4 is not elevated in sera of patients with pulmonary fibrosis or bleomycin-treated mice with pulmonary fibrosis. This may be due to different pathogenic mechanisms of liver and lung fibro-genesis. They stated that MFAP4 appeared to be useful as serum biomarker for hepatic but not for lung fibrosis. These preliminary findings need to be validated by well-designed studies.

MicroRNA-21

Zhao and colleagues (2014) stated that microRNA-21 (miR-21) plays an important role in the pathogenesis and progression of liver fibrosis. These researchers determined the serum and hepatic content of miR-21 in patients with liver cirrhosis and rats with dimethyl-nitrosamine-induced hepatic cirrhosis and examined the effects of miR-21 on SPRY2 and HNF4α in modulating ERK1 signaling in hepatic stellate cells (HSCs) and epithelial-mesenchymal transition (EMT) of hepatocytes. Quantitative real-time polymerase chain reaction (RT-PCR) was used to determine miR-21 and the expression of SPRY2, HNF4α and other genes. Immunoblotting assay was performed to examine the expression of relevant proteins. Luciferase reporter assay was performed to assess the effects of miR-21 on its predicted target genes SPRY2 and HNF4α. Primary HSCs and hepatocytes were treated with miR-21 mimics/inhibitors or appropriate adenoviral vectors to examine the relation between miR-21 and SPRY2 or HNF4α. The serum and hepatic content of miR-21 was significantly higher in cirrhotic patients and rats. SPRY2 and HNF4α mRNA levels were markedly lower in the cirrhotic liver. MiR-21 over-expression was associated with enhanced ERK1 signaling and EMT in liver fibrosis. Luciferase assay revealed suppressed SPRY2 and HNF4α expression by miR-21. Ectopic miR-21 stimulated ERK1 signaling in HSCs and induced hepatocyte EMT by targeting SPRY2 or HNF4α. Down-regulating miR-21 suppressed ERK1 signaling, inhibited HSC activation, and blocked EMT in TGFβ1-treated hepatocytes. The authors concluded that MiR-21 modulated ERK1 signaling and EMT in liver fibrosis by regulating SPRY2 and HNF4α expression; MiR-21 may serve as a potentially biomarker as well as intervention target for hepatic cirrhosis.

Kitano and Bloomston (2016) noted that miRNAs are small non-coding RNAs that regulate gene expression by either blocking translation or inducing degradation of target mRNA. Hepatic stellate cells play a central role in development of hepatic fibrosis and there are intricate regulatory effects of miRNAs on their activation, proliferation, collagen production, migration, and apoptosis. There are multiple differentially expressed miRNAs in activated HSCs, and these researchers summarized current data on miRNAs that participate in the development of hepatic fibrosis. The authors concluded that miRNAs may serve as biomarkers for diagnosis of liver disease, as well as markers of disease progression. Most importantly, dysregulated miRNAs may potentially be targeted by novel therapies to treat and reverse progression of hepatic fibrosis.

Signal-Induced Proliferation-Associated 1 Like 1 (SIPA1L1)

Marfa et al (2016) noted that currently several procedures are used for staging liver fibrosis. However, these methods may involve clinical complications and/or present diagnostic uncertainty mainly in the early stages of the disease. This study was designed to unveil new non-invasive biomarkers of liver fibrosis in an in-vivo model of fibrosis/cirrhosis induction by CCl4 inhalation by using a label-free quantitative LC-MS/MS approach. These researchers analyzed 94 serum samples from adult Wistar rats with different degrees of liver fibrosis and 36 control rats. Firstly, serum samples from 18 CCl4-treated rats were clustered into 3 different groups according to the severity of hepatic and the serum proteome was characterized by label-free LC-MS/MS. Furthermore, 3 different pooled serum samples obtained from 16 control Wistar rats were also analyzed. Based on the proteomic data obtained, these investigators performed a multivariate analysis that displayed 3 main cell signaling pathways altered in fibrosis. In cirrhosis, more biological imbalances were detected as well as multi-organ alterations. In addition, hemopexin and signal-induced proliferation-associated 1 like 1 (SIPA1L1) were selected as potential serum markers of liver fibro-genesis among all the analyzed proteins. The results were validated by ELISA in an independent group of 76 fibrotic/cirrhotic rats and 20 controls that confirmed SIPA1L1 as a potential non-invasive biomarker of liver fibrosis. In particular, SIPA1L1 showed a clear diminution in serum samples from fibrotic/cirrhotic rats and a great accuracy at identifying early fibrotic stages. The authors concluded that the proteomic analysis of serum samples from CCl4-treated rats has enabled the identification of SIPA1L1 as a non-invasive marker of early liver fibrosis. These preliminary findings of an in-vivo study need to be validated by well-designed studies in human subjects.

Transient Elastography

Ultrasound transient elastography (eg, FibroScan) is a noninvasive, bedside ultrasonic technique to evaluate liver fibrosis by measuring liver stiffness. Transient elastography is based on the theory that fibrosis makes the liver stiffer and that elastic waves move more quickly through stiff tissue than through normal tissue. The device consists of a control unit (computer-based), a low-frequency (50 Hz) vibration emitter and a high-frequency (5 MHz) ultrasound probe. When the vibration emitter is pressed between the ribs on the right side of the body, a low-frequency elastic sheer wave is propagated through the liver. The stiffness is proportional to the square of the velocity of the shear wave, which is measured in kilopascals (kPa). There are approximately five to 10 readings taken and the median is used as the final value. In cirrhotic patients, liver stiffness measurements range from 12.5 to 75.5 kPa (kPa).

In a prospective study, de Ledinghen et al (2006) evaluated the accuracy of liver stiffness measurement for the detection of fibrosis and cirrhosis in HIV/hepatitis C virus (HCV)-coinfected patients and compared its accuracy with other non-invasive methods. These researchers studied 72 consecutive HIV patients with chronic hepatitis C who had a simultaneous liver biopsy and liver stiffness measurement by transient elastography (FibroScan; Echosens, Paris, France) for the assessment of liver fibrosis. Liver stiffness values ranged from 3.0 to 46.4 kPa. Liver stiffness was significantly correlated to fibrosis stage (Kendall tau-b = 0.48; p < 0.0001). The area under the receiver operating characteristic (AUROC) curve of liver stiffness measurement was 0.72 for F > or = 2 and 0.97 for F = 4. For the diagnosis of cirrhosis, AUROC curves of liver stiffness measurement were significantly higher than those for platelet count (p = 0.02), aspartate aminotransferase/ALT ratio (p = 0.0001), Aspartate aminotransferase-to-Platelet Ratio Index (p = 0.01), and FIB-4 (p = 0.004). The authors concluded that liver stiffness measurement is a promising noninvasive method for the assessment of fibrosis in HIV-infected patients with chronic HCV infection. They also noted that its use for the follow-up of these patients should be further evaluated.

Foucher and colleagues (2006) assessed the accuracy of FibroScan for the detection of cirrhosis in patients with chronic liver disease. A total of 711 patients with chronic liver disease were studied. Etiologies of chronic liver diseases were hepatitis C virus or hepatitis B virus infection, alcohol, non-alcoholic steatohepatitis, other, or a combination of the above etiologies. Liver fibrosis was evaluated according to the METAVIR score. Stiffness was significantly correlated with fibrosis stage (r = 0.73, p < 0.0001). Areas under the receiver operating characteristic curve (95 % CI) were 0.80 (0.75 to 0.84) for patients with significant fibrosis (F > 2), 0.90 (0.86 to 0.93) for patients with severe fibrosis (F3), and 0.96 (0.94 to 0.98) for patients with cirrhosis. Using a cut off value of 17.6 kPa, patients with cirrhosis were detected with a positive predictive value and a NPV of 90 %. Liver stiffness was significantly correlated with clinical, biological, and morphological parameters of liver disease. With an NPV greater than 90 %, the cut off values for the presence of esophageal varices stage 2/3, cirrhosis Child-Pugh B or C, past history of ascites, hepatocellular carcinoma, and esophageal bleeding were 27.5, 37.5, 49.1, 53.7, and 62.7 kPa, respectively. The authors concluded that FibroScan is a promising non-invasive method for detection of cirrhosis in patients with chronic liver disease. They noted that its use for the follow-up and management of these patients could be of great interest and should be evaluated further.

Corpechot and associates (2006) assessed the diagnostic performance of liver stiffness measurement (LSM) for the determination of fibrosis stage in chronic cholestatic diseases. A total of 101 patients with primary biliary cirrhosis (PBC, n = 73) or primary sclerosing cholangitis (PSC, n = 28) were prospectively enrolled in a multi-center study. All patients underwent liver biopsy (LB) and LSM. Histological and fibrosis stages were assessed on LB by two pathologists. LSM was performed by FibroScan. Efficiency of LSM for the determination of histological and fibrosis stages were determined by a ROC curve analysis. Analysis failed in 6 patients (5.9 %) because of unsuitable LB (n = 4) or LSM (n = 2). Stiffness values ranged from 2.8 to 69.1 kPa (median of 7.8 kPa). LSM was correlated to both fibrosis (Spearman’s rho = 0.84, p < .0001) and histological (0.79, p < .0001) stages. These correlations were still found when PBC and PSC patients were analyzed separately. Areas under ROC curves were 0.92 for fibrosis stage (F) > or = 2, 0.95 for F > or = 3 and 0.96 for F = 4. Optimal stiffness cutoff values of 7.3, 9.8, and 17.3 kPa showed F > or = 2, F > or = 3 and F = 4, respectively. LSM and serum hyaluronic acid level were independent parameters associated with extensive fibrosis on LB. The authors concluded that FibroScan is a simple and reliable non-invasive means for assessing biliary fibrosis. They stated that it should be a promising tool to assess anti-fibrotic therapies in PBC or PSC.

The Canadian Agency for Drugs and Technologies in Health (CADTH) performed an evaluation on FibroScan for non-invasive assessment of liver fibrosis (Murtagh and Foster, 2006). It stated that the diagnostic performance of FibroScan is good for identifying severe fibrosis or cirrhosis, but it is less accurate for milder presentations. It concluded that FibroScan is a promising technology, but large multi-center studies comparing a range of emerging non-invasive fibrosis staging technologies are needed. An earlier assessment by the French Committee for Evaluation and Diffusion of Innovative Technologies (CEDIT, 2004) reached similar conclusions, stating that the absence of conclusive evidence concerning the diagnostic value of FibroScan argues against its immediate dissemination. More recently, an assessment by the French National Authority for Health (HAS, 2007) concluded that additional studies are necessary to evaluate the comparative cost-effectiveness of different methods of assessing liver fibrosis (e.g., FibroTest, FibroScan, and biopsy). A technology assessment by the Malaysian Ministry of Health (Darus, 2008) reached similar conclusions about the need for additional research for the Fibroscan.

de Franchis et al (2007) stated that transient elastography (Fibroscan) might be of value for the non-invasive diagnosis of cirrhosis; however, its reproducibility needs to be further validated. Furthermore, Berrutti et al (2007) noted that FibroScan is a new, non-invasive method to evaluate liver stiffness and, consequently, the degree of liver fibrosis. Since its use in the clinical setting is of great interest, further studies should define the exact role of this procedure.

de Ledinghen et al (2007) assessed the feasibility of liver stiffness measurement and compared FibroScan, FibroTest, and APRI with liver biopsy for the diagnosis of cirrhosis in children with chronic liver diseases. A total of 116 consecutive children with chronic liver diseases were prospectively included. All except 1 child (58 boys, mean age of 10.7 years) could have non-invasive tests for fibrosis: FibroScan, FibroTest, and APRI, and, when necessary, a liver biopsy (n = 33). FibroScan, FibroTest, and APRI were correlated with clinical or biological parameters of chronic liver diseases, but the FibroScan marker correlated most with all parameters. By histology, the METAVIR fibrosis category score was F1 in 7 cases, F2 in 8 cases, F3 in 6 cases, and F4 in 12 cases. FibroScan, FibroTest, and APRI were significantly correlated with the METAVIR fibrosis score. For the diagnosis of cirrhosis, AUC was 0.88, 0.73, and 0.73 for FibroScan, FibroTest, and APRI, respectively. The authors concluded that these findings indicated that liver stiffness measurement is feasible in children and is related to liver fibrosis. A specific probe dedicated to children and slender patients has thus been developed and is currently under evaluation. The FibroScan equipped with this specific probe could become a useful tool for the management of chronic liver diseases in children.

Shaheen et al (2007) stated that the accurate diagnosis of HCV-related fibrosis is crucial for prognostication and treatment decisions. Due to the limitations of biopsy, non-invasive alternatives including FibroTest and FibroScan have been developed. These investigators systematically reviewed studies describing the accuracy of these tests for predicting HCV-related fibrosis. Studies comparing FibroTest or FibroScan versus biopsy in HCV patients were identified via an electronic search. Random effects meta-analyses and AUC were examined to characterize test accuracy for significant fibrosis (F2 to F4) and cirrhosis. Heterogeneity was explored using meta-regression. A total of 12 studies were identified, 9 for FibroTest (n = 1,679) and 4 for FibroScan (n = 546). In heterogeneous analyses for significant fibrosis, the AUCs for FibroTest and FibroScan were 0.81 (95 % confidence interval [CI] 0.78 to 84) and 0.83 (0.03 to 1.00), respectively. At a threshold of approximately 0.60, the sensitivity and specificity of the FibroTest were 47 % (35 % to 59 %) and 90 % (87 % to 92%). For FibroScan (threshold approximately 8 kPa), corresponding values were 64 % (50 % to 76 %) and 87 % (80 % to 91 %), respectively. Methodological quality, the length of liver biopsy specimens, and inclusion of special populations did not explain the observed heterogeneity. However, the diagnostic accuracy of both measures was associated with the prevalence of significant fibrosis and cirrhosis in the study populations. For cirrhosis, the summary AUCs for FibroTest and FibroScan were 0.90 (95 % CI not calculable) and 0.95 (0.87 to 0.99), respectively. The authors concluded that FibroTest and FibroScan have excellent utility for the identification of HCV-related cirrhosis, but lesser accuracy for earlier stages. They noted that refinements are necessary before these tests can replace liver biopsy.

Sagir et al (2008) noted that transient elastography (also known as FibroScan) is a rapid, non-invasive, and reproducible method for measuring liver stiffness, which correlates with the degree of liver fibrosis in patients with chronic hepatitis. However, whether FibroScan is useful in the detection of pre-existing liver fibrosis/cirrhosis in patients presenting with acute liver damage is unclear. In this study, patients with acute liver damage of different etiologies were analyzed. Liver stiffness was measured during the acute phase of the liver damage and followed-up to the end of the acute phase. A total of 20 patients were included in the study. In 15 of the 20 patients, initial liver stiffness values measured by FibroScan during the acute phase of the liver damage were suggestive of liver cirrhosis. However, none of these 15 patients showed any signs of liver cirrhosis in the physical examination, ultrasound examination, or liver histology [performed in 11 of 15 (73 %) patients]. A significant difference was observed in the initial bilirubin levels (5.8 +/- 6.5 mg/dL versus 15.7 +/- 11.8 mg/dL; p = 0.042) and age (32.4 +/- 17.5 years versus 49.7 +/- 15.8 years; p = 0.042) between patients with liver stiffness below or above 12.5 kPa. Six patients with initially high liver stiffness were followed-up to abatement of the acute hepatic phase; in all of them, liver stiffness values decreased to values below the cut-off value for liver cirrhosis. The authors concluded that transient elastography frequently yields pathologically high values in patients with acute liver damage and is unsuitable for detecting cirrhosis/fibrosis in these patients.

Han and Yoon (2008) stated that although liver biopsy is still the gold standard for assessing hepatic fibrosis, it has some technical limitations and risks. Furthermore, the dynamic process of liver fibrosis resulting from progression and regression can not be quantified by liver biopsy. Thus, alternative, simple, reliable and non-invasive tests are needed to assess the stage of fibrosis. Several non-invasive direct and indirect serum markers able to predict the presence of significant fibrosis or cirrhosis in patients with chronic liver disease with considerable accuracy have been reported. However, since most of these markers require complicated calculations, clinical application is difficult. Transient elastography (FibroScan) is a new method for the evaluation of liver stiffness. It is based on changes in tissue elasticity induced by hepatic fibrosis. The authors noted that based on accumulating clinical data, clinical applications of elastography will increase in the near future.

Sporea and colleagues (2008) stated that evaluation of liver fibrosis can be performed by FibroTest, elastography (FibroScan), and by LB, which is considered to be the “gold standard”. At the present, there are 3 techniques for performing LB: percutaneous, trans-jugular, and laparoscopic. The percutaneous LB can be performed blind, ultrasound (US)-guided or US-assisted. There 2 two main categories of specialists who perform LB: gastroenterologists (hepatologists) and radiologists, and the specialty of the individual who performs the LB determines if the LB is performed under ultrasound guidance or not. There are 2 types of biopsy needles used for LB: cutting needles (Tru-Cut, Vim-Silverman) and suction needles (Menghini, Klatzkin, Jamshidi). The rate of major complications after percutaneous LB ranges from 0.09 % to 2.3 %, but the echo-guided percutaneous liver biopsy is a safe method for the diagnosis of chronic diffuse hepatitis (cost-effective as compared to blind biopsy) and the rate of complications seems to be related to the experience of the physician and the type of the needle used (Menghini type needle seems to be safer). The authors stated that maybe in a few years non-invasive markers of fibrosis will be used, but at this time, most authorities in the field consider LB to be useful and necessary for the evaluation of chronic hepatopathies, despite the fact that it is not a perfect test.

Castera and associates (2008) stated that transient elastography (TE, FibroScan) is a novel non-invasive method that has been proposed for the assessment of hepatic fibrosis in patients with chronic liver diseases, by measuring liver stiffness. It is a rapid and user-friendly technique that can be easily performed at the bedside or in the outpatient clinic with immediate results and good reproducibility. Limitations include failure in approximately 5 % of cases, mainly in obese patients. So far, TE has been mostly validated in chronic hepatitis C, with diagnostic performance equivalent to that of serum markers for the diagnosis of significant fibrosis. Combining TE with serum markers increases diagnostic accuracy and as a result, LB could be avoided for initial assessment in most patients with chronic hepatitis C. These investigators stated that this strategy warrants further evaluation in other etiological types of chronic liver diseases. Transient elastography appears to be an excellent tool for early detection of cirrhosis and may have prognostic value in this setting. As TE has excellent patient acceptance it could be useful for monitoring fibrosis progression and regression in the individual case, but more data are awaited for this application. Guidelines are needed for the use of TE in clinical practice.

In a meta-analysis, Friedrich-Rust et al (2008) examined the performance of TE for the staging of liver fibrosis. Literature data bases and international conference abstracts were searched. Inclusion criteria were as follows: evaluation of TE, LB as reference, and assessment of the area under the receiver operating characteristic curve (AUROC). The meta-analysis was performed using the random-effects model for the AUROC, summary receiver operating curve techniques, as well as meta-regression approaches. A total of 50 studies were included in the analysis. The mean AUROC for the diagnosis of significant fibrosis, severe fibrosis, and cirrhosis were 0.84 (95 % CI: 0.82 to 0.86), 0.89 (95 % CI: 0.88 to 0.91), and 0.94 (95 % CI: 0.93 to 0.95), respectively. For the diagnosis of significant fibrosis a significant reduction of heterogeneity of the AUROC was found when differentiating between the underlying liver diseases (p < 0.001). Other factors influencing the AUROC were the scoring system used and the country in which the study was performed. Age, body mass index, and biopsy quality did not have a significant effect on the AUROC. The authors concluded that TE can be performed with excellent diagnostic accuracy and independent of the underlying liver disease for the diagnosis of cirrhosis. However, for the diagnosis of significant fibrosis, a high variation of the AUROC was found that is dependent on the underlying liver disease. A critique of this meta-analysis by the Centre for Reviews and Dissemination (CRD, 2010) stated that, although sensitivity and specificity data appeared to have been available for many of the studies included in this meta-analysis, the meta-analysis focused on pooled estimates of AUROC. The CRD noted that use of this measure of overall accuracy results in a loss of clinically important information about test performance; it was unclear how many inaccurate test results were due to false positive and how many to false negatives. The CRD critique stated that generation of summary receiver operating characteristic (SROC) curves using a bivariate of hierarchical model may have been more appropriate to this data set; the CRD noted that such models allow generation of summary estimates of sensitivity and specificity as well as potential to assess the significance of sources of heterogeneity. The CRD concluded that these limitations in the analysis mean that the conclusions of this meta-analysis should be interpreted with caution.

Abenavoli et al (2008) noted that in clinical practice there are currently 3 methods for the evaluation of liver fibrosis. First, LB is still considered as the “gold standard” method. Second, serological markers and their mathematical combination were suggested in the last years as an alternative to LB. Third, TE was proposed recently. This technique (TE) is based on the progression speed of an elastic shear wave within the liver. The authors concluded that currently, there are just a few studies capable of evaluating the effectiveness of TE in evaluating chronic liver diseases, mainly in patients infected with HCV. Its application must also be studied in the monitoring of patients suffering from chronic HCV infection and subjected to treatments that can modify their degree of liver fibrosis.

Muñoz et al (2009) evaluated the correlation between values of Fibroscan, liver biopsy, and clinical data among HCV-positive renal transplant patients. A total of 24 HCV/RNA-positive patients with a previous liver biopsy were selected to undergo Fibroscan (transient elastography) and a clinical evaluation of liver function. Fibroscan values were expressed in kilopascals (kPa). As 2 patients were eliminated due to obesity or ascites, these investigators analyzed 22 patients. Thirteen patients (59 %) with fibrosis F0-F1 (METAVIR score) by biopsy and normal liver function showed a mean Fibroscan score of 5.2 kPa (range of 2.3 to 6.8 kPa). Three patients (13.6 %) exhibited F2 by biopsy and normal liver function with a mean Fibroscan score of 8.2 kPa (range of 7.3 to 8.9 kPa). Three patients (13.6 %) with F3 by biopsy and abnormal liver function showed a high mean Fibroscan score of 10.9 kPa (range of 10.5 to 11.6 kPa). The last 3 patients (13.6 %) with F4 (cirrhosis) by biopsy and abnormal clinical data showed the highest mean Fibroscan value of 14.2 kPa (range of 8.9 to 18 kPa). The authors concluded that among renal transplant patients with HCV, the values of Fibroscan seem to correlate with the degree of fibrosis by biopsy and with clinical liver function. Thus, Fibroscan may be useful to follow patients with LD. However, these results should be analyzed with caution due to the small number of cases and retrospective nature of the study.

Andersen and colleagues (2009) stated that liver biopsy is considered the “golden standard” for assessment of hepatic fibrosis. However, the procedure has limitations because of inconvenience and rare but serious complications as bleeding. Furthermore, sampling errors are frequent, and inter-observer variability often poses problems. Recently, transient elastography has been developed to assess fibrosis. The device measures liver elasticity, which correlates well with the degree of fibrosis. Studies have shown that transient elastography is more accurate in diagnosing cirrhosis than minor-to-moderate fibrosis. Most of the studies have been conducted on patients with chronic hepatitis but a few studies have also covered fibrosis and cirrhosis due to other etiologies, and they also demonstrated the high sensitivity and specificity. The authors concluded that transient elastography for assessment of fibrosis may turn out to be a valuable diagnostic procedure and follow-up of patients with chronic liver diseases.

Breton et al (2009) examined the feasibility and reliability of liver stiffness measurement in children with liver diseases. Liver stiffness measurements were performed on 72 children, from 4 to 18 years of age, with potential hepatic fibrosis disease. The clinical, biological, ultrasonographic, and endoscopic parameters were noted to identify children with portal hypertension syndrome. The APRI (ASAT-to-platelet ratio index) test was calculated according to the standard formula. An APRI test score higher than 1.5 indicates significant hepatic fibrosis. METAVIR scoring from 14 liver biopsies was compared to the liver stiffness using the Kappa statistic. A total of 28 patients had viral hepatitis, 20 cystic fibrosis, 16 chronic liver cholestasis, 5 autoimmune hepatitis, and 3 patients had liver fibrosis with uncertain etiology. FibroScan measurements were available in all children. There was good agreement between FibroScan and pathological studies (weighted kappa = 0.814). Only 9 children had portal hypertension syndrome with an average measurement of liver stiffness significantly higher than children without portal hypertension (26.5kPa versus 6.4kPa; p < 0.01). The APRI test for 6 out of 9 patients scored higher than 1.5. The authors concluded that these findings indicate that liver stiffness measurement is feasible in children and seems to be related to liver fibrosis. They stated that larger prospective studies are needed to validate this FibroScan method.

In a meta-analysis of transient elastography for the detection of hepatic fibrosis, Stebbing et al (2010) evaluated its use in comparison with liver biopsy. Studies from the literature were analyzed with a median liver stiffness value in kilopascal given for fibrosis stages according to histopathologic findings on biopsy and best discriminant cut-off levels in kilopascals for significant fibrosis (greater than or equal to F2) and cirrhosis (F4). A total of 22 studies were selected comprising 4,430 patients; chronic hepatitis C infection was the most common etiology of fibrosis. The pooled estimates for significant fibrosis (greater than or equal to F2) measured 7.71 kPa (LSM cut-off value) with a sensitivity of 71.9 % [95 % CI: 71.4 % to 72.4 %] and specificity of 82.4 % (95 % CI: 81.9 % to 82.9 %), whereas for cirrhosis (F4) the results showed a cut-off of 15.08 kPa with a sensitivity of 84.45 % (95 % CI: 84.2 % to 84.7 %) and specificity of 94.69 % (95 % CI: 94.3% to 95 %). The authors concluded that further evaluation of transient elastography to assess LSM is needed in prospective studies to potentially increase the sensitivity and establish its clinical utility.

Myers (2009) noted that non-alcoholic fatty liver disease (NAFLD) is the most common cause of chronic liver disease, affecting about 30 % of Western populations and a frequent indication for liver transplantation. The histological spectrum of NAFLD includes simple steatosis, which has a benign prognosis, and non-alcoholic steatohepatitis, a more aggressive form of liver injury that may progress to cirrhosis and its complications. At present, the only widely accepted means of differentiating these lesions, including the severity of hepatic fibrosis, is liver biopsy. However, due to the invasiveness of this procedure, the rising prevalence of NAFLD, and the expected availability of effective therapies for this condition, the identification of non-invasive tools for the diagnosis and staging of NAFLD has emerged as a major clinical and research priority. The author summarized important advances in this field during the past decade, including the development of biomarkers of hepatic fibrosis, apoptosis, and inflammation; novel imaging techniques such as transient elastography; and high-throughput technologies including proteomics and genomics. Future studies must focus on the development of accurate, inexpensive, and reliable tools that can differentiate the major histologic determinants of NAFLD; are responsive to changes in NAFLD severity due to therapeutic intervention and time; and have prognostic significance. Until such tools are developed, liver biopsy remains an important tool in the assessment of patients with NAFLD.

Myers and co-workers (2010) determined the feasibility and performance of TE in a North American cohort of patients with chronic liver disease. Liver stiffness measurements were obtained using TE in 260 patients with chronic hepatitis B or C, or NAFLD from 4 Canadian hepatology centers. The accuracy of TE compared with liver biopsy for the prediction of significant fibrosis (Metavir fibrosis score of F2 or greater), bridging fibrosis (Metavir fibrosis score of F3 or greater) and cirrhosis (Metavir fibrosis score of F4 ) was assessed using area under ROC curves (AUROCs), and compared with the aspartate aminotransferase-to-platelet ratio index. The influence of ALT levels and other factors on liver stiffness was determined using linear regression analyses. Failure of TE occurred in 2.7 % of patients, while liver biopsies were inadequate for staging in 0.8 %. Among the remaining 251 patients, the AUROCs of TE for Metavir fibrosis scores of F2 and F3 or greater, and F4 were 0.74 (95 % CI: 0.68 to 0.80), 0.89 (95 % CI: 0.84 to 0.94), and 0.94 (95 % CI: 0.90 to 0.97), respectively. Liver stiffness measurement was more accurate than the aminotransferase-to-platelet ratio index for bridging fibrosis (AUROC 0.78) and cirrhosis (AUROC 0.88), but not significant fibrosis (AUROC 0.76). At a cut-off of 11.1 kPa, the sensitivity, specificity, and positive and negative predictive values for cirrhosis (prevalence 11 %) were 96 %, 81 %, 39 % and 99 %, respectively. For significant fibrosis (prevalence 53 %), a cut-off of 7.7 kPa was 68 % sensitive and 69 % specific, and had a positive predictive value of 70 % and a negative predictive value of 65 %. Liver stiffness was independently associated with ALT, body mass index and steatosis. The optimal LSM cut-offs for cirrhosis were 11.1 kPa and 11.5 kPa in patients with ALT levels lower than 100 U⁄L and 100 U⁄L or greater, respectively. For fibrosis scores of F2 or greater, these figures were 7.0 kPa and 8.6 kPa, respectively. The authors concluded that the major role of TE is the exclusion of bridging fibrosis and cirrhosis. However, TE can not replace biopsy for the diagnosis of significant fibrosis. Because liver stiffness may be influenced by significant ALT elevation, BMI,and/or steatosis, tailored liver stiffness cut-offs may be necessary to account for these factors.

Cholongitas et al (2010) systematically reviewed the literature regarding non-invasive tests (NIT) following liver transplantation. These investigators identified 14 studies evaluating NIT based on serum markers and/or liver imaging techniques: 10 studies assessed NIT in recipients with recurrent HCV infection for fibrosis and 4 studies evaluated predictors of progression of fibrosis in recurrent HCV. Transient elastography had good discrimination for significant fibrosis (median AUROC: 0.88). Among the serum NIT, APRI had good performance (median AUROC: 0.75). Transient elastography performed better than serum (direct and indirect) NIT for significant fibrosis with median AUROC 0.88 (versus 0.66, p < 0.001), median sensitivity 0.86 (versus 0.56, p = 0.002), median NPV 0.90 (versus 0.74, p = 0.05) and median positive predictive value (PPV) 0.80 (versus 0.63, p = 0.02). Transient elastography compared to indirect serum NIT, had better performance, but was not superior to APRI score. Finally, direct, compared to indirect NIT, were not significantly different except for specificity: median: 0.83 versus 0.69, respectively, p = 0.04. The authors concluded that NIT could become an important tool in clinical management of liver transplant recipients, but whether they can improve clinical practice needs further evidence. Their optimal combination with liver biopsy and assessment of collagen content requires investigation.

Stebing et al (2010) performed a meta-analysis to further assess the use of TE in comparison with liver biopsy. Studies from the literature were analyzed with a median liver stiffness value in kilopascal given for fibrosis stages according to histopathologic findings on biopsy and best discriminant cutoff levels in kilopascals for significant fibrosis (greater than or equal to F2) and cirrhosis (F4). A total of 22 studies were selected comprising 4,430 patients; chronic hepatitis C infection was the most common etiology of fibrosis. The pooled estimates for significant fibrosis greater than or equal to F2) measured 7.71 kPa (LSM cut-off value) with a sensitivity of 71.9 % [95 % CI: 71.4 % to 72.4 %] and specificity of 82.4 % (95 % CI: 81.9 % to 82.9%), whereas for cirrhosis (F4) the results showed a cut-off of 15.08 kPa with a sensitivity of 84.45 % (95 % CI: 84.2 % to 84.7 %) and specificity of 94.69 % (95 % CI: 94.3 % to 95 %). The authors concluded that further evaluation of TE to assess LSM is needed in prospective studies to potentially increase the sensitivity and establish its clinical utility.

Thabut et al (2011) noted that severe portal hypertension is responsible for complications and death. Although measurement of the hepatic venous pressure gradient is the most accurate method for evaluating the presence and severity of portal hypertension, this technique is considered invasive and is not routinely performed in all centers. Several non-invasive techniques have been proposed to measure portal hypertension. Certain methods evaluate elements related to the pathogenesis of portal hypertension through the measurement of hyperkinetic syndrome, or they examine the development of hepatic fibrosis through the measurement of increased intra-hepatic vascular resistance. Other methods assess the clinical consequences of portal hypertension, such as the presence of esophageal varices or the development of porto-systemic shunts. Methods evaluating increased hepatic vascular resistance are fairly accurate and primarily involve the detection of hepatic fibrosis by serum markers and transient elastography. The radiological assessment of hyperkinetic syndrome probably has value but is still under investigation. The assessment of severe portal hypertension by the presence of varices may be performed with simple tools such as biological assays, computed tomography, and esophageal capsules. More sophisticated procedures seem promising but are still under development. Screening tools for large populations must be simple, whereas more complicated procedures could help in the follow-up of already diagnosed patients. Although most of these non-invasive methods effectively identify severe portal hypertension, methods for diagnosing moderate portal hypertension need to be developed; this shows that further investigation is needed in this field.

Tsochatzis et al (2011) studied the performance of elastography for diagnosis of fibrosis using meta-analysis. Medline, Embase, SCI, Cochrane Library, conference abstracts books, and article references were searched. These investigators included studies using biopsy as a reference standard, with the data necessary to calculate the true- and false-positive, true- and false-negative diagnostic results of elastography for a fibrosis stage, and with a 3-month maximum interval between tests. The quality of the studies was rated with the QUADAS tool. These researchers identified 40 eligible studies. Summary sensitivity and specificity was 0.79 (95 % CI: 0.74 to 0.82) and 0.78 (95 % CI: 0.72 to 0.83) for F2 stage and 0.83 (95 % CI: 0.79 to 0.86) and 0.89 (95 % CI: 0.87 to 0.91) for cirrhosis. After an elastography result at/over the threshold value for F2 or cirrhosis (“positive” result), the corresponding post-test probability for their presence (if pre-test probability was 50 %) was 78 %, and 88 %, respectively, while, if values were below these thresholds (“negative” result), the post-test probability was 21 % and 16 %, respectively. No optimal stiffness cut-offs for individual fibrosis stages were validated in independent cohorts and cut-offs had a wide range and overlap within and between stages. The authors concluded that elastography theoretically has good sensitivity and specificity for cirrhosis (and less for lesser degrees of fibrosis); however, it should be cautiously applied to everyday clinical practice because there is no validation of the stiffness cut-offs for the various stages. They stated that such validation is required before elastography is considered sufficiently accurate for non-invasive staging of fibrosis.

An UpToDate review on “Epidemiology, clinical features, and diagnosis of nonalcoholic steatohepatitis” (Sheth and Chopra, 2012) states that “A potentially useful non-invasive method for excluding advanced fibrosis is measurement of liver stiffness with transient elastography. However, the approach is not widely available and has not been extensively studied in NASH”.

Poca et al (2011) stated that prognostic markers of compensated cirrhosis should mainly investigate factors involved with progression to decompensation because death in cirrhosis is related with decompensation. Portal hypertension plays a crucial role in the pathophysiology of most complications of cirrhosis. Accordingly, hepatic venous pressure gradient (HVPG) monitoring has strong prognostic value. An HVPG of greater than or equal to 10 mm Hg determines a significantly higher risk of developing decompensation. Esophageal varices also can develop when the HVPG is greater than or equal to 10 mm Hg, although an HVPG greater than or equal to 12 mm Hg is required for variceal bleeding to occur. Monitoring the changes induced by the treatment of portal hypertension on HVPG, provides strong prognostic information. In compensated cirrhosis hemodynamic response is appropriate when the HVPG decreased to less than 10 mm Hg or by less than 10 % from baseline, because the incidence of complications such as bleeding or ascites significantly decrease when these targets are achieved. Whether serum markers, such as the FibroTest, may be valuable to predict decompensation should be established. Transient elastography is a promising technique that has shown an excellent accuracy to detect severe portal hypertension. However, whether it can adequately determine clinically significant portal hypertension, and risk of developing varices and decompensation, should be established. Magnetic resonance elastography is also promising.

Lee and colleagues (2013) evaluated and compared the ability of serum HA and human cartilage glycoprotein-39 (YKL-40) values, as well as TE findings, to predict advanced hepatic fibrosis in a cohort from a single pediatric center. Subjects who underwent liver biopsy analysis within 12 months before enrollment were eligible for this prospective study. Hyaluronic acid and YKL-40 measurements were obtained within 1 month of TE. A METAVIR score of F3 or F4 was considered to indicate advanced fibrosis. A total of 128 patients (51 % males) aged 1.4 months to 27.6 years (22 % aged less than 2 years) were enrolled. Thirty-one subjects had data on HA and YKL-40; and 97 subjects had data on both blood tests and TE. For the prediction of advanced fibrosis, the AUC values were 0.83 for TE, 0.72 for HA, and 0.52 for YKL-40. The AUC of 0.83 for TE was statistically significantly greater than the AUCs for HA (p = 0.03) and YKL-40 (p < 0.0001). Optimal cut-off points for predicting F3-F4 fibrosis were 8.6 kPa for TE (p < 0.0001), 43 ng/ml for HA (p < 0.0001), and 26.2 ng/ml for YKL-40 (p = 0.85). The combination of TE and HA was not better than TE alone for predicting advanced fibrosis (p = 0.15). The authors concluded that in this study, which evaluated TE, HA, and YKL-40 to predict liver fibrosis in children in the United States, YKL-40 had no predictive value and TE was superior to HA, but the addition of HA did not improve the performance of TE. They stated that these findings suggested that TE and HA may be useful non-invasive tools for assessing liver fibrosis in children.

An assessment of FibroScan by the Institute for Clinical Effectiveness and Health Policy (IECS) (Pichon Riviere et al, 2012) found: “There is evidence of good methodological quality from systematic reviews and meta-analysis of observational studies on diagnostic accuracy of transient elastography compared with the gold standard: biopsy. FibroScan seems to be a potential non-invasive diagnostic method used as an alternative to liver biopsy to diagnose and stage the degree of fibrosis. It might be useful in patients with contraindications for percutaneous biopsy and to follow-up patients with fibrosis who are under treatment. However, it has several disadvantages that may limit its reproducibility such as processes which increase or decrease liver consistency (steatosis), obesity, ascites or reduced intercostal spaces, and its low accuracy to identify mild to moderate stages of fibrosis. At present, the biopsy is still the diagnostic method of choice for the diagnosis of liver fibrosis.”

An assessment of transient elastography by Health Quality Ontario (Brener, 2015) concluded: “There was evidence to support the diagnostic accuracy of transient elastography compared to liver biopsy for assessing liver fibrosis in the disease areas of interest. There was evidence that the diagnostic accuracy of FibroTest and acoustic force radiation impulse were not significantly different from transient elastography for assessing liver fibrosis in the disease areas of interest. There was evidence to support the diagnostic accuracy of controlled attenuation parameter compared to liver biopsy for assessing steatosis in the disease areas of interest. No evidence was found that assessed the clinical utility of transient elastography (with or without controlled attenuation parameter) versus biopsy, as measured by a change in clinical diagnosis, treatment, or patient outcomes. Beneficial impact could be presumed, given that the accuracy of TE is comparable to that of a biopsy and would have an impact as a noninvasive alternative to diagnose. The clinical utility of CAP is less certain given that treatment for this condition generally consists of providing advice about healthy behaviors”.

Guidelines on the management of hepatitis C from the American Association for the Study of Liver Disease (2014) state that “[n]on-invasive methods frequently used to estimate liver disease severity include a liver-directed physical exam (normal in most patients), routine blood tests (eg, serum alanine transaminase, albumin, bilirubin, international normalized ratio levels, and complete cell blood counts with platelets), serum fibrosis marker panels, liver imaging (eg, ultrasound, computed tomography scan), and liver elastography. . . . Liver elastography can provide instant information regarding liver stiffness at the point-of-care but can only reliably distinguish cirrhosis from non-cirrhosis.”

Transient Elastography for Detection of Esophageal Varices in Individuals with Cirrhosis

Li and colleagues (2016) TE has been used for the prediction of large esophageal varices in cirrhotic patients. However, the conclusions have not been always consistent throughout the different studies. These researchers performed a meta-analysis to evaluate the diagnostic accuracy of TE for the prediction of large esophageal varices. They performed a systematic literature search in PubMed, Embase, Web of Science, and CENTRAL in The Cochrane Library without time restriction. The strategy used was “(FibroScan or transient elastography or stiffness) and esophageal varices”. Accuracy measures such as pooled sensitivity (SEN), specificity (SPE), among others, were calculated using Meta-DiSc statistical software. A total of 20 studies (2,994 patients) were included in this meta-analysis. The values of pooled SEN, SPE, positive and negative likelihood ratios (LRs) and DOR were as follows: 0.81 (95 % CI: 0.79 to 0.84), 0.71 (95 % CI: 0.69 to 0.73), 2.63 (95 % CI: 2.15 to 3.23), 0.27 (95 % CI: 0.22 to 0.34) and 10.30 (95 % CI: 7.33 to 14.47). The AUROC was 0.83. The Spearman correlation coefficient was 0.246 with a p-value of 0.296, indicating the absence of any significant threshold effects. In the subgroup analysis, the heterogeneity could be partially explained by the geographical origin of the study or etiology; or it could be partially explained blindingly, through the appropriate interval and cut-off value of the LS. The authors concluded that TE could be used as a non-invasive screening tool for the prediction of large esophageal varices. However, since LS cut-off values varied throughout the different studies and significant heterogeneity also existed among them, there is a need for more reasonable approaches or flow diagram in order to improve the operability of this technology.

Pu and co-workers (2017) examined the diagnostic accuracy of FibroScan (FS) in detecting esophageal varices (EV) in cirrhotic patients. Through a systemic literature search of multiple databases, these investigators reviewed 15 studies using endoscopy as a reference standard, with the data necessary to calculate pooled SEN and SPE, positive and negative LRs, DOR and AUROC. The quality of the studies was rated by the Quality Assessment of Diagnostic Accuracy studies-2 tool. Clinical utility of FS for EV was evaluated by a Fagan plot. Heterogeneity was explored using meta-regression and subgroup analysis. All statistical analyses were conducted via Stata12.0, MetaDisc1.4 and RevMan5. In 15 studies (n = 2,697), FS detected the presence of EV with the summary sensitivities of 84 % (95 % CI: 81.0 % to 86.0%), specificities of 62 % (95 % CI: 58.0 % to 66.0%), a positive LR of 2.3 (95 % CI: 1.81 to 2.94), a negative LR of 0.26 (95 % CI: 0.19 to 0.35), a DOR of 9.33 (95 % CI: 5.84 to 14.92) and an AUROC of 0.8262. FS diagnosed the presence of large EV with the pooled SEN of 0.78 (95 % CI: 75.0 % to 81.0 %), SPE of 0.76 (95 % CI: 73.0 % to 78.0 %), a positive and negative LR of 3.03 (95 % CI: 2.38 to 3.86) and 0.30 (95 % CI: 0.23 to 0.39), respectively, a summary diagnostic OR of 10.69 (95 % CI: 6.81 to 16.78), and an AUROC of 0.8321. A meta-regression and subgroup analysis indicated different etiology could serve as a potential source of heterogeneity in the diagnosis of the presence of EV group. A Deek’s funnel plot suggested a low probability for publication bias. The authors concluded that using FS to measure liver stiffness could not provide high accuracy for the size of EV due to the various cut-off and different etiologies. They stated that these limitations precluded widespread use in clinical practice at this time; therefore, the results should be interpreted cautiously given its SEN and SPE.

Transient Elastography for Diagnosis of Glycogenic Hepatopathy

Khoury and colleagues (2018) stated that glycogenic hepatopathy (GH) is a disorder associated with uncontrolled diabetes mellitus, most commonly type 1 (T1DM), expressed as right upper quadrant abdominal pain, hepatomegaly and increased liver enzymes. The diagnosis may be difficult, because laboratory and imaging tests are not pathognomonic. Although GH may be suggested based on clinical presentation and imaging studies, the gold standard for diagnosis is a liver biopsy, showing a significant accumulation of glycogen within the hepatocytes. Glycogenic hepatopathy may be diagnosed also after elevated liver enzymes in routine blood tests; GH usually regresses after tight glycemic control. Progression to end-stage liver disease (ESLD) has never been reported. This review aimed to increase the awareness to this disease, to suggest a pathway for investigation that may reduce the use of unnecessary tests, especially invasive ones. These researchers carried out a PubMed database search (up to July 1, 2017) with the words “glycogenic hepatopathy”, “hepatic glycogenosis”, “liver glycogenosis” and “diabetes mellitus-associated glycogen storage hepatopathy”. Articles in which diabetes mellitus-associated liver glycogen accumulation was described were included in this review. A total of 47 articles were found, describing 126 patients with GH. Hepatocellular disturbance was more profound than cholestatic disturbance. No synthetic failure was reported. The authors concluded that GH may be diagnosed conservatively, based on corroborating medical history, physical examination, laboratory tests, imaging studies and response to treatment, even without liver biopsy. In case of doubt about the diagnosis or lack of clinical response to treatment, a liver biopsy may be considered. These investigators stated that there is no role for non-invasive tests like FibroScan or FibroTest for the diagnosis of GH or for differentiation of this situation from NAFLD.

Sherigar and associates (2018) GH is a rare complication of the poorly controlled diabetes mellitus characterized by the transient liver dysfunction with elevated liver enzymes and associated hepatomegaly caused by the reversible accumulation of excess glycogen in the hepatocytes. It is predominantly observed in patients with longstanding T1DM and rarely reported in association with type 2 diabetes mellitus (T2DM). Although it was first observed in the pediatric population, since then, it has been reported in adolescents and adults with or without ketoacidosis. The association of GH with hyperglycemia in diabetes has not been well-established. One of the essential elements in the pathophysiology of development of GH is the wide fluctuation in both glucose and insulin levels; GH and NAFLD are clinically indistinguishable, and the latter is more prevalent in diabetic patients and can progress to advanced liver disease and cirrhosis. Gradient dual-echo MRI can distinguish GH from NAFLD; however, GH can reliably be diagnosed only by liver biopsy. Adequate glycemic control can result in complete remission of clinical, laboratory and histological abnormalities. There has been a recent report of varying degree of liver fibrosis identified in patients with GH. The authors concluded that further research is needed for an ideal non-invasive, rapid diagnostic test to avoid the extensive work-up and associated costs in evaluating suspected cases of GH. For now, a more aggressive pursuit of liver biopsy in the evaluation of elevated transaminases could identify additional cases of GH, allowing for continued elucidation of prevalence and natural history of this entity. Clinicians should also continue to pool patient data from case studies of patients with GH, to better understand the underlying risk factors and characteristics of this disease.

Transient Elastography for Diagnosis of Portal Hypertension

Kim and colleagues (2017) noted that TE has been proposed as a promising non-invasive alternative to hepatic venous pressure gradient (HVPG) for detecting portal hypertension (PH). However, previous studies have yielded conflicting results. These researchers gathered evidence on the clinical usefulness of TE versus HVPG for assessing PH. They conducted a systematic review by searching databases for relevant literature evaluating the clinical usefulness of non-invasive TE for assessing PH in patients with cirrhosis. A literature search in Ovid Medline, Embase and the Cochrane Library was performed for all studies published prior to December 30, 2015. A total of 8 studies (1,356 patients) met inclusion criteria. For the detection of PH (HVPG greater than or equal to 6 mmHg), the summary SEN and SPE were 0.88 (95 % CI: 0.86 to 0.90) and 0.74 (95 % CI: 0.67 to 0.81), respectively. Regarding clinically significant PH (HVPG greater than or equal to 10 mmHg), the summary SEN and SPE were 0.85 (95 % CI: 0.63 to 0.97) and 0.71 (95 % CI: 0.50 to 0.93), respectively. The overall correlation estimate of TE and HVPG was large (0.75, 95 % CI: 0.65 to 0.82, p < 0.0001). The authors concluded that TE showed high accuracy and correlation for detecting the severity of PH. These investigators stated that TE showed promise as a reliable and non-invasive procedure for the evaluation of PH that should be integrated into clinical practice; however, further investigation is needed.

The authors stated that this study had several drawbacks. First, only 8 studies were used to evaluate the usefulness and performance of TE, thus limiting the robustness of the conclusions reached. Second, the characteristics of the included studies, including patient characteristics, cirrhosis etiologies and varying diagnostic thresholds, were not completely consistent. Third, these researchers included only studies written in English, thus language bias might have influenced the results.

In a retrospective study, Kumar and associates (2017) examined the diagnostic accuracy of TE for detecting clinically significant PH (CSPH) in patients with cirrhotic PH. This trial was conducted on consecutive patients with cirrhosis greater than 15 years of age who underwent HVPG and TE from July 2011 to May 2016. Correlation between HVPG and TE was analyzed using the Spearman’s correlation test; ROC curves were prepared for determining the utility of TE in predicting various stages of PH. The best cut-off value of TE for the diagnosis of CSPH was obtained using the Youden index. The study included 326 patients (median age of 52 [range of 16 to 90] years; 81 % males]. The most common etiology of cirrhosis was cryptogenic (45 %) followed by alcohol (34 %). The median HVPG was 16.0 (range of 1.5 to 30.5) mm Hg; 85 % of patients had CSPH. A significant positive correlation was noted between TE and HVPG (rho 0.361, p < 0.001). The AUROC curve for TE in predicting CSPH was 0.740 (95 % CI: 0.662 to 0.818) (p < 0.01). A cut-off value of TE of 21.6 kPa best predicted CSPH with a PPV of 93 %. The authors concluded that TE has a fair positive correlation with HVPG; thus, TE can be used as a non-invasive modality to evaluate the degree of PH. A cut-off TE value of 21.6 kPa identifies CSPH with a PPV of 93 %. These investigators stated that as a non-invasive procedure, TE is a promising tool to translate into routine clinical practice for detecting CSPH. Moreover, they stated that further large prospective studies are needed to prospectively validate the findings of this study and also to examine if TE can be used in monitoring the hemodynamic response and the effect of drugs reducing portal pressure.

The authors stated that his study had 2 main drawbacks. First, it was a retrospective study, so the study may suffer from selection bias. These researchers included only those patients who underwent HVPG and TE during the study period; hence, the patients may not represent the entire population of patients with cirrhosis, as most included patients have moderate-to-severe PH. A prospective study design, which includes all consecutive patients of cirrhosis, regardless of degree of portal hypertension, would have been a better study design and more representative of the cirrhotic population of the community. Second, the lack of follow-up; follow-up data on complications of PH would have further validated these findings of TE cut-off for CSPH.

Transient Elastography for Prognosis of Chronic Hepatitis C

Erman and colleagues (2018) stated that CHC is a leading cause of hepatic fibrosis and cirrhosis. The level of fibrosis is traditionally established by histology, and prognosis is estimated using fibrosis progression rates (FPRs; annual probability of progressing across histological stages). However, newer non-invasive alternatives are quickly replacing biopsy. One alternative, TE, quantifies fibrosis by LSM. Given these developments, these researchers estimated prognosis in treatment-naïve CHC patients using TE-based liver stiffness progression rates (LSPR) as an alternative to FPRs and compared consistency between LSPRs and FPRs. A systematic literature search was performed using multiple databases (January 1990 to February 2016); LSPRs were calculated using either a direct method (given the difference in serial LSMs and time elapsed) or an indirect method given a single LSM and the estimated duration of infection and pooled using random-effects meta-analyses. For validation purposes, FPRs were also estimated. Heterogeneity was explored by random-effects meta-regression. A total of 27 studies reporting on 39 groups of patients (n = 5,874) were identified with 35 groups allowing for indirect and 8 for direct estimation of LSPR. The majority (approximately 58 %) of patients were HIV/HCV-co-infected. The estimated time-to-cirrhosis based on TE versus biopsy was 39 and 38 years, respectively. In uni-variate meta-regressions, male sex and HIV were positively and age at assessment, negatively associated with LSPRs. The authors concluded that non-invasive prognosis of HCV is consistent with FPRs in predicting time-to-cirrhosis, but more longitudinal studies of liver stiffness are needed to obtain refined estimates.

Transient Elastography for Diagnosis of Acute Cellular Rejection Following Liver Transplantation

Nacif and colleagues (2018) noted that TE is a non-invasive technique that measures liver stiffness. When an inflammatory process is present, this is shown by elevated levels of stiffness. Acute cellular rejection (ACR) is a consequence of an inflammatory response directed at endothelial and bile epithelial cells, and it is diagnosed through liver biopsy. In a systematic review, these investigators examined the viability of TE in ACR following liver transplantation. The Cochrane Library, Embase, and Medline PubMed databases were searched and updated to November 2016. The MESH terms used were “liver transplantation,” “graft rejection,” “elasticity imaging techniques” (PubMed), and “elastography” (Cochrane and Embase). A total of 70 studies were retrieved and selected using the PICO (patient, intervention, comparison or control, outcome) criteria; 3 prospective studies were selected to meta-analysis and evaluation. A total of 33 patients with ACR were assessed with TE. One study showed a cut-off point of greater than 7.9 kPa to define graft damage and less than 5.3 kPa to exclude graft damage (ROC 0.93; p < 0.001). Another study showed elevated levels of liver stiffness in ACR patients. However, in this study, no cut-off point for ACR was suggested. The final prospective study included 27 patients with ACR at liver biopsy. Cut-off points were defined as TE greater than 8.5 kPa, moderate-to-severe ACR, with a specificity of 100 % and ROC of 0.924. The measurement of TE of less than 4.2 kPa excluded the possibility of any ACR (p = 0.02). The authors concluded that TE may be an important tool for the severity of ACR in patients following liver transplantation; moreover, these researchers stated that further studies are needed to better define the cut-off points and applicability of this approach.

Transient Elastography (e.g., FibroScan) for Diagnosis of Primary Sclerosing Cholangitis

Halasz et al (2015) examined if expression of selected miRNAs obtained from fibrotic liver biopsies correlated with fibrosis stage. A total of 52 patients were enrolled in the study representing various etiologic backgrounds of fibrosis: 24 cases with chronic hepatitis infections (types B, C), 19 with autoimmune liver diseases (autoimmune hepatitis, primary biliary cirrhosis, primary sclerosing cholangitis [n = 2], overlapping syndrome cases), and 9 of mixed etiology (alcoholic and nonalcoholic steatosis, cryptogenic cases). Severity of fibrosis was determined by both histologic staging using the METAVIR scoring system and non-invasive TE. Following RNA isolation, expression levels of miR-21, miR-122, miR-214, miR-221, miR-222, and miR-224 were determined using TaqMan MicroRNA Assays applying miR-140 as the reference. Selection of miRNAs was based on their characteristic up- or down-regulation observed in HCC. Relative expression of miRNAs was correlated with fibrosis stage and LS value measured by TE, as well as with serum ALT level. The expression of individual miRNAs showed deregulated patterns in stages F1 to F4 as compared with stage F0, but only the reduced level of miR-122 in stage F4 was statistically significant (p < 0.04). When analyzing miRNA expression in relation to fibrosis, levels of miR-122 and miR-221 showed negative correlations with fibrosis stage, and miR-122 was found to correlate negatively and miR-224 positively with LS values (all p < 0.05). ALT levels displayed a positive correlation with miR-21 (p < 0.04). Negative correlations were observed in the fibrosis samples of mixed etiology between miR-122 and fibrosis stage and LS values (p < 0.05), and in the samples of chronic viral hepatitis, between miR-221 and fibrosis stage (p < 0.01), whereas miR-21 showed positive correlation with ALT values in the samples of autoimmune liver diseases (p < 0.03). The results also revealed a strong correlation between fibrosis stage and LS values (p < 0.01) when etiology of fibrosis was not taken into account. The authors concluded that reduced expression of miR-122 in advanced fibrosis and its correlation with fibrosis stage and LS values appeared to be characteristic of hepatic fibrosis of various etiologies.

The authors concluded that the main drawback of this study was its small sample size in the various etiologic groups (total of 52 patients). Thus, further studies are needed to examine if the observed miRNA correlations were also characteristic of the various etiology groups or whether these relationships are only summed characteristics of the fibrosis samples by reason of the various etiologies.

Corpechot et al (2014) assessed the diagnostic performance, reproducibility, longitudinal changes, and prognostic value of liver stiffness measurement (LSM) using vibration-controlled transient elastography (VCTE). In a prospective study, the investigators analyzed percutaneous liver biopsy specimens from 73 consecutive patients with PSC from January 2005 to December 2010. Patients underwent VCTE no more than 6 months after the biopsy specimens were collected. The biopsy specimens were analyzed by a pathologist blinded to the results of VCTE for the stage of fibrosis, and LSM was associated with the stage of fibrosis and other variables using the Kruskal-Wallis and Spearman correlation tests. The cutoff values of LSM were selected based on the accuracy with which they identified the stage of fibrosis on receiver-operating characteristic analysis. The rates of LSM progression were assessed using a linear mixed model, and the association between LSM values and clinical outcomes were evaluated using Cox regression analysis in 168 patients with PSC treated with ursodeoxycholic acid and followed up from November 2004 to July 2013 (mean follow-up period, 4 years). LSM was independently linked to the stage of fibrosis. Cutoff values for fibrosis stages ≥F1, ≥F2, ≥F3, and F4 were 7.4 kPa, 8.6 kPa, 9.6 kPa, and 14.4 kPa, respectively. The adjusted diagnostic accuracy values for severe fibrosis and cirrhosis were 0.83 and 0.88, respectively. The diagnostic performance of LSM was comparable to that of hyaluronic acid measurement but superior to the aspartate aminotransferase/platelet ratio index, FIB-4 score, and Mayo risk score in differentiating patients with significant or severe fibrosis from those without. LSM had a high level of reproducibility between operators for the same measurement site and for the same operator between 2 adjacent sites. LSM increased significantly and exponentially over time. Baseline measurements and rate of LSM progression were strongly and independently linked with patients’ outcomes. VCTE is able to differentiate severe from nonsevere liver fibrosis with high levels of confidence in patients with PSC. Baseline measurements of LSM and longitudinal changes are prognostic factors for PSC.

Krawczyk et al (2017) stated that previous studies demonstrated a close correlation between TE and liver histology in chronic liver diseases. Data on the accuracy of TE in primary sclerosing cholangitis (PSC) remains scarce. These investigators examined the association between TE, serum marker of liver injury and histology of explanted livers in PSC patients. A total of 30 patients were prospectively recruited; TE (FibroScan) and blood sampling were performed during evaluation for liver transplantation (LT); the 2nd blood sampling was performed on the day of LT. Fibrosis of explanted livers according to the 7-point Laennec staging system and liver collagen contents were measured. TE correlated with Laennec stages of fibrosis (p = 0.001), collagen contents (p < 0.001) and with diameter of thickest septa (p = 0.034) in explanted livers. It also correlated with serum indices of liver injury, namely AST, bilirubin as well as FIB-4 and APRI scores (all p < 0.05). In a multi-variate model, only liver fibrosis, according to either Laennec score (p = 0.035) or collagen contents (p = 0.005), was significantly associated with TE. Finally, patients with cirrhosis had increased liver stiffness (p = 0.002) and the TE cut-off of 13.7 kPa showed the best predictive value (AUC = 0.90, 95 % confidence interval [CI]: 0.80 to 1.00, p < 0.001) for detecting cirrhosis. The authors concluded that TE correlated with liver fibrosis and markers of liver injury in patients with PSC. However, liver fibrosis appeared to be the strongest predictor of liver stiffness assessed with TE. Thus, these researchers postulated that TE is a reliable tool for non-invasive monitoring of PSC.

Khoshpouri et al (2019) noted that PSC is a chronic progressive inflammatory disease of the bile ducts that leads to multi-focal bile duct fibrosis, strictures, cholestasis, liver parenchymal changes, and ultimately cirrhosis. It more commonly occurs in young adults, with a variety of clinical and imaging manifestations. The cause of the disease is unknown, however, it has a strong association with inflammatory bowel disease (IBD) and could overlap with other autoimmune diseases, including autoimmune hepatitis and immunoglobulin G4-related disease. Patients are predisposed to various hepatic and extra-hepatic deteriorating complications, such as bile duct and gallbladder calculi, acute bacterial cholangitis, liver abscess, and portal hypertension, as well as malignancies including cholangiocarcinoma (CCA), gallbladder cancer, and colorectal carcinoma (CRC). Imaging has an essential role in diagnosis, surveillance, and detection of complications. MR cholangiopancreatography (MRCP) and endoscopic retrograde cholangiopancreatography (ERCP) have high specificity and sensitivity for detection of primary disease and assessment of disease progression. However, many patients with PSC are still diagnosed incidentally at ultrasonography (US) or computed tomography (CT). Novel imaging techniques such as TE and MR elastography (MRE) are used to survey the grade of liver fibrosis. Annual cancer surveillance is necessary in all PSC patients to screen for CCA and gallbladder cancer. Familiarity with PSC pathogenesis and imaging features across various classic imaging modalities and novel imaging techniques could aid in correct imaging diagnosis and guide appropriate management.

Tafur et al (2020) compared biliary stricture severity on MRCP, MRE, and vibration-controlled TE (VCTE) liver stiffness (LS) for evaluation of risk stratification and prognostication in PSC. A total of 87 patients (age of 31 to 61 years; 34 women/53 men) prospectively underwent biochemical testing, VCTE, MRCP, and MRE between January 2014 and July 2016. Correlation between the MRCP grading of PSC based on biliary stricture severity, LS on MRE and VCTE, and the Mayo Risk Score as well as the Amsterdam Oxford Prognostic Index (AOPI) were evaluated and compared. Stricture severity was classified according to previous classification systems based on ERCP. Spearman’s correlation and Kruskal-Wallis tests were performed. MRE-LS and intra-hepatic stricture severity combined demonstrated higher discriminatory ability among risk categories based on Mayo Risk Score (AUROC = 0.8). MRE-LS alone demonstrated excellent discriminatory ability among risk categories based on AOPI using cut-offs of 1 and 2.7 and was superior to intra-hepatic stricture severity (AUROC = 0.9, AUROC = 0.6 to 0.7). There was a weak correlation between intra-hepatic stricture severity and MRE-LS (rho = 0.3; p = 0.011). VCTE-LS values were not correlated with stricture severity and were non-contributory to differentiate patients across risk groups. Intra-hepatic stricture severity alone was a poor discriminator of advanced liver fibrosis on MRE (AUROC = 0.7); however, combining intra- and extra-hepatic stricture severity and controlling for cholestasis and disease duration improved results (AUROC = 0.9). The authors concluded that the findings of this study demonstrated a significant discriminatory ability of LS values on MRE to distinguish between early to moderate and advanced liver fibrosis; LS values on MRE may add value to risk prognostication and further studies including clinical outcomes are needed.

Guidelines on primary sclerosing cholangitis from the British Society of Gastroenterology (Chapman, et al., 2019) state: “While liver biopsy does provide information on the stage of liver fibrosis, there has been increasing interest in the value of non-invasive assessment in patients with PSC. One retrospective study highlighted the strong correlation between transient elastography and histological stage of liver fibrosis, as well as the prognostic significance. Serological assessment of liver fibrosis using the enhanced liver fibrosis test correlates with elastography and helps to stratify prognosis in patients with PSC. Both these modalities are undergoing further evaluation, and recent reports from a larger cohort suggest they may be effective markers of fibrosis and disease progression. Magnetic resonance elastography is also emerging as a possible non-invasive marker of cirrhosis in PSC. European Association for the Study of the Liver (EASL) clinical practice guidelines recommend the use of non-invasive markers for monitoring the degree of liver fibrosis, but evidence specifically related to patients with PSC is still evolving.”

Furthermore, an UpToDate review on “Primary sclerosing cholangitis in adults: Clinical manifestations and diagnosis” (Kowdley, 2020) states that “Transient elastography measures liver stiffness and is a technique for noninvasively assessing hepatic fibrosis. Studies suggest that it can be used to estimate the degree of hepatic fibrosis in patients with cholestatic liver disease, including PSC. In one study, the area under the receiver operating characteristic curve for transient elastography predicting cirrhosis in patients with cholestatic liver disease was 0.96 when a cutoff of 17.3 kPa was used”. The “Summary and Recommendations” section does not mention transient elastography.

Transient Elastography (e.g., FibroScan) for Evaluation of Alagille Syndrome

Shneider et al (2020) stated that elastographic measurement of liver stiffness is of growing importance in the assessment of liver disease. Pediatric experiences with this technique are primarily single-center and limited in scope. The Childhood Liver Disease Research Network (ChiLDReN) provided a unique opportunity to examine elastography in a well-characterized multi-institutional cohort. Children with biliary atresia (BA), alpha-1 antitrypsin deficiency (A1ATD), or Alagille syndrome (ALGS) followed in a prospective longitudinal network study were eligible for enrollment in a prospective investigation of transient elastography (FibroScan). Studies were carried out in subjects who were non-fasted and non-sedated. Liver stiffness measurements (LSMs) were correlated with standard clinical and biochemical parameters of liver disease along with a research definition of clinically evident portal hypertension (CEPH) graded as absent, possible, or definite. Between November 2016 and August 2019, 550 subjects with a mean age of 8.8 years were enrolled, 458 of whom had valid LSMs (BA, n = 254; A1ATD, n = 104; ALGS, n = 100). Invalid scans were more common in participants less than 2-year old. There was a positive correlation between LSM and total bilirubin, international normalized ratio (INR), aspartate aminotransferase (AST), alanine aminotransferase (ALT), gamma-glutamyl transpeptidase (GGT), GGT to platelet ratio (GPR), pediatric end-stage liver disease score, AST to platelet ratio index, and spleen size, and a negative correlation with albumin and platelet count in BA, with similar correlations for A1ATD (except AST, ALT, and albumin) and ALGS (except for INR, GGT, GPR, and ALT). Possible or definite CEPH was more common in BA compared to ALGS and A1ATD. LSM was greater in definite versus absent CEPH in all 3 diseases. Disease-specific clinical and biochemical characteristics of the different CEPH grades were observed. The authors concluded that it is feasible to obtain LSMs in children, especially over the age of 2 years; LSM correlated with liver parameters and portal hypertension, although disease-specific patterns exist.

The authors stated that FibroScan‐based LSM in children is non-invasive and feasible, although there is a critical need to attend to technical details of the measurements. Its use in un-sedated children under the age of 2 years may be limited. These studies have confirmed in a large cohort of children with 3 distinct types of liver disease that LSM correlated with features of portal hypertension and routine clinical laboratory assessments of liver disease in children. LSM may be influenced by the type of liver disease in ways that are not clear at present, warranting careful interpretation of results and further investigation. Moreover, these researchers stated that future investigations will examine change in LSM over time and the correlation of baseline and change in LSM with clinical events identified in the 3 on‐going, prospective, longitudinal ChiLDReN studies.

Furthermore, an UpToDate review on “Causes of cholestasis in neonates and young infants” (Erlichman and Loomes, 2021) states that “Alagille syndrome, which is characterized by chronic cholestasis with paucity of the interlobular bile ducts on liver biopsy. Associated features found in most patients include cardiac anomalies, butterfly vertebrae, posterior embryotoxon of the eye, and characteristic facial features”. However, this UTD review does not mention transient elastography / FibroScan as a management tool.

Transient Elastography for Evaluation of Alpha-1-Antitrypsin Deficiency

An UpToDate review on “Clinical manifestations, diagnosis, and natural history of alpha-1 antitrypsin deficiency” (Stoller, 2022a) does not mention transient elastography as a management tool.

Furthermore, an UpToDate review on “Extrapulmonary manifestations of alpha-1 antitrypsin deficiency” (Stoller, 2022a) states that “Pending greater clarity on the ideal testing protocol, our practice is to assess serum aminotransferases (e.g., alanine aminotransferase, aspartate aminotransferase), alkaline phosphatase, and bilirubin annually. Ultrasound-based elastography is increasingly used to monitor for fibrosis, although the optimal frequency of screening has not been determined”. Moreover, elastography is not mentioned in the “Summary and Recommendations” section of this UTD review.

Transient Elastography / FibroScan for Evaluation of Gilbert Syndrome

Poynard et al (2008) noted that liver biopsy, owing to its limitations and risks, is an imperfect gold standard for assessing the severity of the most frequent chronic liver diseases chronic hepatitis C (HCV), B (HBV) non-alcoholic (NAFLD) and alcoholic (ALD) fatty liver diseases. These investigators summarized the advantages and the limits of the available biomarkers of liver fibrosis. Among a total of 2,237 references, 14 validated serum biomarkers have been identified between 1991 and 2008; 9 were not patented and 5 were patented. Two alternatives to liver biopsy were the most evaluated FibroTest and FibroScan. For FibroTest, there was a total of 38 different populations including 7,985 subjects with both FibroTest and biopsy (4,600 HCV, 1,580 HBV, 267 NAFLD, 524 ALD, and 1014 mixed). For FibroScan, there was a total of 11 published studies including 2,260 subjects (1,466 HCV, 95 cholestatic liver disease, and 699 mixed). For FibroTest, the mean diagnostic value for the diagnosis of advanced fibrosis assessed using standardized area under the ROC curves was 0.84 (95 % confidence interval [CI]: 0.83 to 0.86), without a significant difference between the causes of liver disease, hepatitis C, hepatitis B, and alcoholic or non-alcoholic fatty liver disease. High-risk profiles of false negative/false positive of FibroTest, mainly Gilbert syndrome, hemolysis and acute inflammation, were present in 3 % of the populations. In case of discordance between biopsy and FibroTest, 50 % of the failures could be due to biopsy; the prognostic value of FibroTest was at least similar to that of biopsy in HCV, HBV and ALD. The authors concluded that this overview of evidence-based data suggested that biomarkers could be used as an alternative to liver biopsy for the 1st-line assessment of fibrosis stage in the 4 most common chronic liver diseases, namely HCV, HBV, NAFLD and ALD. Neither biomarkers nor biopsy alone was sufficient for taking a definite decision in a given patient; all the clinical and biological data must be taken into account. There is no evidence-based data justifying biopsy as a 1st-line estimate of liver fibrosis. Health authorities in some countries have already approved validated biomarkers as the 1st-line procedure for the staging of liver fibrosis.

Furthermore, an UpToDate review on “Gilbert syndrome and unconjugated hyperbilirubinemia due to bilirubin overproduction” (Roy-Chowdhury et al, 2022) does not mention transient elastography / FibroScan as a management option.

Transient Elastography and Hepatitis B

Leibenguth et al (2024) stated that liver diseases of infectious and non-infectious etiology cause considerable morbidity and mortality, especially in low- and middle-income countries (LMICs); however, data on the prevalence of liver diseases and underlying risk factors in LMICs are scarce. These investigators examined the occurrence of infectious diseases among individuals with chronic liver damage in a rural setting of Côte d’Ivoire. In 2021, these researchers screened 696 individuals from 4 villages in the southern part of Côte d’Ivoire for hepatic fibrosis and steatosis, employing transient elastography (TE) and controlled attenuation parameter (CAP). They classified CAP ≥ 248 dB/m as steatosis, TE ≥ 7.2 kPa as fibrosis, and did subgroup analysis for participants with TE ranging from 7.2 kPa to 9.1 kPa. Clinical and microbiologic characteristics were compared to an age- and sex-matched control group (TE < 6.0 kPa; n = 109). Stool samples were subjected to duplicate Kato-Katz thick smears for diagnosis of Schistosoma mansoni. Venous blood samples were examined for hepatitis B and hepatitis C virus. Furthermore, an abdominal ultrasound (US) examination was carried out. Among 684 individuals with valid TE measurements, TE screening identified hepatic pathologies in 149 participants (17 % with fibrosis and 6 % with steatosis); 419 participants were included for further analyses, of which 261 had complete microbiologic analyses available. The prevalence of S. mansoni, hepatitis B, and hepatitis C were 30 %, 14 %, and 7 %, respectively. Logistic regression analysis revealed higher odds for having TE results between 7.2 kPa and 9.1 kPa in individuals with S. mansoni infection (odds ratio [OR] = 3.02, 95 % confidence interval [CI]: 1.58 to 5.76, p = 0.001), while HCV infection (OR = 5.02, 95 % CI: 1.72 to 14.69, p = 0.003) and steatosis (OR = 4.62, 95 % CI: 1.60 to 13.35, p = 0.005) were found to be risk factors for TE ≥ 9.2 kPa. The authors concluded that besides viral hepatitis, S. mansoni also warrants consideration as a pathogen causing liver fibrosis in Côte d’Ivoire. In-depth diagnostic work-up among individuals with abnormal TE findings might be a cost-effective public health strategy.

Xu et al (2024) noted that chronic hepatitis B (CHB) presents a global health challenge due to its potential to cause severe liver conditions such as hepato-cellular carcinoma (HCC) and cirrhosis. Previous research has established a correlation between CHB infection with low-level viremia (LLV) and liver disease progression, such as increased HCC incidence. In a retrospective, cohort study, these investigators examined if LLV during treatment with nucleos(t)ide analogs (NAs) would contribute to the accelerated progression of liver fibrosis (LF). This trial focused on CHB patients undergone NA monotherapy for over 96 weeks. Subjects were categorized into maintained virological response (MVR) and LLV groups based on hepatitis B virus (HBV) DNA levels. The study evaluated LF using various markers and methods, including chitinase 3-like 1 protein (CHI3L1), aspartate aminotransferase-to-platelet ratio index (APRI), fibrosis-4 (FIB-4) score, and TE. Analysis was carried out on 92 CHB patients, categorized into LLV (n = 42) and MVR (n = 50) groups, following the exclusion of 101 patients for various reasons. Significant findings included lower baseline HBV DNA in MVR (less than 20 IU/ml) compared to LLV (67.8 IU/ml, p < 0.001) and different AST/ALT ratios (LLV: 1.1, MVR: 1.36, p = 0.011). LF was assessed using CHI3L1, FIB-4, and APRI, with LLV showing a higher baseline CHI3L1 (LLV:83.3 ng/ml versus MVR: 54.5 ng/ml, p = 0.016) and scores compared to MVR, indicative of fibrosis. CHI3L1 levels in LLV were higher at baseline and weeks 48, 72, and 96 than MVR, with significance at baseline (p = 0.038) and week 48 (p = 0.034). Liver stiffness measurement (LSM) showed a time-dependent decline in both groups but no significant intergroup differences. The authors concluded that non-invasive monitoring of CHB patients who have received treatment indicated that LLV contributed to the progression of LF, necessitating proactive adjustment of anti-viral treatment strategies.

Jin et al (2024) stated that LSM using TE can evaluate fibrotic burden in chronic liver diseases. In a systematic review and meta-analysis, these investigators examined if LSM using TE can predict the risk of development of HCC in CHB patients. They carried out a systematic literature search of the Ovid-Medline, Embase, Cochrane, and KoreaMed databases (from January 2010 to June 2023). Of the 1,345 individual studies identified, 10 studies that used TE were finally registered. Hazard ratios (HRs) and the 95 % CIs were considered summary estimates of treatment effect sizes of greater than or equal to 11 kilopascal (kPa) standard for HCC development. Meta-analysis was carried out using the restricted Maximum Likelihood random effects model. Among the 10 studies, data for risk ratios (RRs) for HCC development could be obtained from 9 studies. When analyzed for the 9 studies, the HR for HCC development was high at 3.33 (95 % CI: 2.45 to 4.54) in CHB patients with a baseline LSM of greater than or equal to 11 kPa compared to patients who did not. In 10 studies included, LSM of greater than or equal to 11 kPa showed the sensitivity and specificity for predicting HCC development were 61 % (95 % CI: 50 % to 71 %) and 78 % (95 % CI: 66 % to 86 %), respectively, and the diagnostic accuracy was 0.74 (95 % CI: 0.70 to 0.77). The authors concluded that the risk of HCC development was elevated in CHB patients with TE-determined LSM of greater than or equal to11 kPa. This finding suggested that TE-determined LSM values may aid the risk prediction of HCC development in CHB patients.

Furthermore, an UpToDate review on “Hepatitis B virus: Overview of management” (Lok, 2024) states that “The initial evaluation of patients with chronic HBV infection should include screening for fibrosis using noninvasive tests (e.g., vibration-controlled transient elastography, serum fibrosis panel) or liver biopsy. Noninvasive assessments of liver fibrosis, notably measurements of liver stiffness, are increasingly used instead of liver biopsies; however, liver stiffness can be influenced by inflammation as well as fibrosis, and therefore, liver stiffness measurements may overestimate liver fibrosis in patients with a high ALT (more than 100 units/L)”.

Quantitative Magnetic Resonance for Analysis of Liver Tissue Composition (e.g., LiverMultiScan)

Quantitative magnetic resonance for analysis of liver tissue composition (e.g., LiverMultiScan) has been developed for non-invasive liver evaluation including tissue composition (e.g., fat, iron, water content). LiverMultiScan employs multi-parametric magnetic resonance imaging (MRI) to quantify liver tissue. The system is designed to provide to aid physicians care for patients with fatty liver disease such as NAFLD and NASH. LiverMultiScan purportedly would reduce the need for liver biopsy and provides the care team with a quantitative report to help monitor therapy. However, there is insufficient evidence to support the clinical value of this approach.

McDonald and colleagues (2018) noted that LiverMultiScan is an emerging diagnostic tool using multi-parametric MRI (mpMRI) to quantify liver disease. In a prospective, 2-center, validation study, 161 consecutive adult patients who had clinically-indicated liver biopsies underwent contemporaneous non-contrast mpMRI at 3.0 Tesla (proton density fat fraction (PDFF), T1 and T2* mapping), TE and ELF test. Non-invasive liver tests were correlated with gold standard histological measures. Reproducibility of LiverMultiScan was examined in 22 healthy volunteers. Iron-corrected T1 (cT1), TE, and ELF demonstrated a positive correlation with hepatic collagen proportionate area (all p < 0·001). TE was superior to ELF and cT1 for predicting fibrosis stage. cT1 maintained good predictive accuracy for diagnosing significant fibrosis in cases with indeterminate ELF, but not for cases with indeterminate TE values. PDFF had high predictive accuracy for individual steatosis grades, with AUROCs ranging from 0.90 to 0.94. T2* mapping diagnosed iron accumulation with AUROC of 0.79 (95 % CI: 0.67 to 0.92) and NPV of 96 %. LiverMultiScan showed excellent test/re-test reliability (coefficients of variation ranging from 1.4 % to 2.8 % for cT1). Overall failure rates for LiverMultiScan, ELF and TE were 4.3 %, 1.9 % and 15 %, respectively. The authors concluded that LiverMultiScan is an emerging point-of-care (POC) diagnostic tool that is comparable with the established non-invasive tests for assessment of liver fibrosis, while at the same time offering a superior technical success rate and contemporaneous measurement of liver steatosis and iron accumulation. These investigators stated that these findings support the further development and refinement of mpMRI technology for evaluation of liver disease.

Yin and co-workers (2019) examined the use of MRE-derived mechanical properties (shear stiffness (|G*|) and loss modulus (G″)) and MRI-derived fat fraction (FF) to predict the NAFLD activity score (NAS) in a NAFLD mouse model. A total of 89 male mice were studied, including 64 training and 25 independent testing animals. An MRI/MRE examination and histologic evaluation were performed. Pairwise, non-parametric comparisons and multi-variate analyses were used to evaluate the relationships between the t3 imaging parameters (FF, |G*|, and G″) and histologic features. A virtual NAS score (vNAS) was generated by combining 3 imaging parameters with an ordinal logistic model (OLM) and a generalized linear model (GLM). The prediction accuracy was evaluated by ROC analyses. The combination of FF, |G*|, and G″ predicted NAS greater than 1 with excellent accuracy in both training and testing sets (AUROC > 0.84). OLM and GLM predictive models misclassified 3/54 and 6/54 mice in the training, and 1/25 and 1/25 in the testing cohort respectively, in distinguishing between “not-NASH” and “definite-NASH”. “Borderline-NASH” prediction was poorer in the training set, and no borderline-NASH mice were available in the testing set. The authors concluded that this preliminary study showed that multiparametric MRI/MRE can be used to accurately predict the NAS score in a NAFLD animal model, representing a promising alternative to liver biopsy for assessing NASH severity and treatment response.

Bachtiar et al (2019) noted that as the burden of liver disease reaches epidemic levels, there is a high unmet medical need to develop robust, accurate, and reproducible non-invasive methods to quantify liver tissue characteristics for use in clinical development as well as in clinical practice. In a prospective, cross-sectional study, these investigators examined the repeatability and reproducibility of iron-corrected T1 (cT1), T2*, and hepatic PDFF quantification with multi-parametric MRI across different field strengths, scanner manufacturers and models. A total of 61 adult subjects with mixed liver disease etiology and those without any history of liver disease underwent multi-parametric MRI on combinations of 5 scanner models from 2 manufacturers (Siemens and Philips) at different field strengths (1.5T and 3T). These researchers reported high repeatability and reproducibility across different field strengths, manufacturers, and scanner models in standardized cT1 (repeatability CoV: 1.7 %, bias -7.5ms, 95 % LoA of -53.6 ms to 38.5 ms; reproducibility CoV 3.3 %, bias 6.5 ms, 95 % LoA of -76.3 to 89.2 ms) and T2* (repeatability CoV: 5.5 %, bias -0.18 ms, 95 % LoA -5.41 to 5.05 ms; reproducibility CoV 6.6 %, bias -1.7 ms, 95 % LoA -6.61 to 3.15 ms) in human measurements. PDFF repeatability (0.8 %) and reproducibility (0.75 %) coefficients revealed high precision of this metric. Similar precision was observed in phantom measurements. Inspection of the ICC model indicated that most of the variance in cT1 could be accounted for by study participants (ICC = 0.91), with minimal contribution from technical differences. The authors concluded that multi-parametric MR-derived metrics, cT1, T2* and PDFF, exhibited good repeatability and reproducibility that could quantify liver tissue characteristics independent of scanner manufacturer (Philips or Siemens) and field strength (1.5T or 3T). Moreover, these investigators stated that multi-parametric MRI is a non-invasive method that does not require additional hardware, and can be completed in less than 15-mins, which will have important implications for routine monitoring and assessment of the liver in clinical practice. The ability to standardize metrics will be important in the clinical trial settings for examining treatment interventions.

Loomba and Adams (2020) noted that liver fibrosis should be examined in all individuals with chronic liver disease as it predicts the risk of future liver-related morbidity; thus, the need for treatment, monitoring and surveillance. Non-invasive fibrosis tests (NITs) overcome many limitations of liver biopsy and are now routinely incorporated into specialist clinical practice. Simple serum-based tests (e.g., Fibrosis Score 4, NAFLD Fibrosis Score) consist of readily available biochemical surrogates and clinical risk factors for liver fibrosis (e.g., age and sex). These have been extensively validated across a spectrum of chronic liver diseases, however, tend to be less accurate than more “complex” serum tests, which incorporate direct measures of fibro-genesis or fibrolysis (e.g., hyaluronic acid, N-terminal pro-peptide of type III collagen). Elastography methods quantify liver stiffness as a marker of fibrosis and are more accurate than simple serum NITs, however, suffer increasing rates of unreliability with increasing obesity. MRE appeared more accurate than US elastography and is not significantly impacted by obesity but is costly with limited availability. NITs are valuable for excluding advanced fibrosis or cirrhosis, however, are not sufficiently predictive when used in isolation. Combining serum and elastography techniques increases diagnostic accuracy and can be used as screening and confirmatory tests, respectively. Unfortunately, NITs have not yet been demonstrated to accurately reflect fibrosis change in response to treatment, limiting their role in disease monitoring. However, recent studies have demonstrated lipidomic, proteomic and gut microbiome profiles as well as microRNA signatures to be promising techniques for fibrosis assessment in the future. The authors listed mpMRI as one of the emerging technologies – new methods using mpMRI incorporate damping ratio at a lower frequency using 3D MRE along with shear wave stiffness on MRE and these may further help refine the detection of NASH and NASH-related fibrosis.

Kim and associates (2020) stated that NASH is a complex disease consisting of various components including steatosis, lobular inflammation, and ballooning degeneration, with or without fibrosis; thus, it is difficult to diagnose NASH with only one imaging modality. In a prospective study, these researchers examined the feasibility of MRI for predicting NASH and developed a non-invasive mpMRI index for the detection of NASH in NAFLD patients. This trial included 47 NAFLD patients who were scheduled to undergo or underwent US-guided liver biopsy within 2 months. Biopsy specimens were graded as NASH or non-NASH. All patients underwent non-enhanced MRI including MRS, MRE, and T1 mapping. Diagnostic performances of MRS, MRE, and T1 mapping for grading steatosis, activity, and fibrosis were evaluated. A mpMRI index combining fat fraction (FF), liver stiffness (LS) value, and T1 relaxation time was developed using linear regression analysis. Receiver operating characteristic (ROC) curve analysis was carried out to evaluate the diagnostic performance of the newly devised MR index. A total of 20 NASH patients and 27 non-NASH patients were included. Using MRS, MRE, and T1 mapping, the mean AUCs for grading steatosis, fibrosis, and activity were 0.870, 0.951, and 0.664, respectively. The mpMRI index was determined as 0.037 × FF (%) + 1.4 × LS value (kPa) + 0.004 × T1 relaxation time (msec) -3.819. ROC curve analysis of the MR index revealed an AUC of 0.883. The cut-off value of 6 had a sensitivity of 80.0 % and specificity of 85.2 %. The authors concluded that MRI showed high diagnostic performance for detecting and grading the NASH components of steatosis, activity (lobular inflammation and ballooning degeneration), and fibrosis. This newly devised mpMRI index combining MRS, MRE, and T1 mapping also demonstrated high diagnostic performance for detecting NASH in NAFLD patients; thus, this mpMRI index may help diagnose NASH in NAFLD patients and potentially reduce the need for liver biopsy.

The authors stated that this study had several drawbacks. First, only a few patients limited to a single tertiary center using 1 MRI scanner were included. Although a small cross-vendor validation study comparing the reproducibility of 2 difference MRI scanners from Philips and GE Healthcare in 13 subjects demonstrated that LS value measurements were reproducible and had good consistency across 2 vendors, a multi-center study with a larger number of patients using several different MRI scanners is needed to develop an mpMRI index that can be widely used in various clinical settings. Second, this newly devised mpMRI index was not validated using a validation group due to the small number of patients. Another prospective study using a validation group of NAFLD patients is needed to validate the findings of this study. Third, it is well known that hepatic iron deposition can be increased with liver disease, and age, gender, as well as menopause for women may affect hepatic iron deposition, which in turn may affect MRI measurements. However, it was not possible to perform subgroup analyses according to age, gender, and menopause due to small number of patients. These investigators hope to study the effect of hepatic iron deposition on MRI measurements in the future with a larger number of patients. Fourth, in this study, a non-invasive index was developed using only MRI parameters and linear regression analysis with 3 fixed MR parameters (FF, LS value, and T1 relaxation time). However, the objective of the study was to develop a simple MRI index for predicting NASH in NAFLD patients. In the future, it is expected to devise a NASH index via a prospective study using multi-variate regression analysis of clinical (demographic and laboratory) data and MRI parameters.

Tonev et al (2020) stated that the rising prevalence of NAFLD and the more aggressive subtype, NASH, is a global public health concern. Left untreated, NAFLD/NASH can result in cirrhosis, liver failure, and death. The current standard for diagnosing and staging liver disease is a liver biopsy, which is costly, invasive, and carries risk for the patient; thus, there is a growing need for a reliable, feasible, and cost-effective, non-invasive diagnostic tool for these conditions. LiverMultiScan is one such promising tool that uses mpMRI to characterize liver tissue and to aid in the diagnosis and monitoring of liver diseases of various etiologies. The primary objective of this trial (RADIcAL1) is to examine the cost-effectiveness of the introduction of LiverMultiScan as a standardized diagnostic test for liver disease in comparison to standard care for NAFLD, in different European Union (EU) territories. RADIcAL1 is a 2-arm, multi-center, randomized control trial (RCT) carried out in 4 EU territories (13 sites, from across Germany, Netherlands, Portugal, and the United Kingdom). In total, 1,072 adult patients with suspected fatty liver disease will be randomized to be treated according to the result of the mpMRI in the intervention arm, so that further diagnostic evaluation is recommended only when values for metrics of liver fat or fibro-inflammation are elevated. Subjects in the control arm will be treated as per center guidelines for standard of care. The primary outcome for this trial is to compare the difference in the proportion of patients with suspected NAFLD incurring liver-related hospital consultations or liver biopsies between the study arms, from the date of randomization to the end of the study follow-up. Secondary outcomes include patient feedback from a patient satisfaction questionnaire, at baseline and all follow-up visits to the end of the study, and time, from randomization to diagnosis by the physician, as recorded at the final follow-up visit. This trial is currently open for recruitment. The anticipated completion date for the study is December 2020. The authors concluded that this RCT will provide the evidence to accelerate decision-making regarding the inclusion of mpMRI-based tools in existing NAFLD/NASH clinical care. RADIcAL1 is among the first and largest European health economic studies of imaging technologies for fatty liver disease. Strengths of the trial include a high-quality research design and an in-depth assessment of the implementation of the cost-effectiveness of the mpMRI diagnostic. If effective, the trial may highlight the health economic burden on tertiary-referral hepatology clinics imposed by unnecessary consultations and invasive diagnostic investigations, and demonstrate that including LiverMultiScan as a NAFLD diagnostic test may be cost-effective compared to liver-related hospital consultations or liver biopsies.

Jayaswal et al (2020) noted that liver cT1, liver T1, TE, and blood-based biomarkers have independently been shown to predict clinical outcomes but have not been directly compared in a single cohort of patients. These researchers compared these tests’ prognostic value in a cohort of patients with compensated chronic liver disease. Patients with unselected compensated liver disease etiologies had baseline assessments and were followed-up for development of clinical outcomes, blinded to the imaging results. The prognostic value of non-invasive liver tests at pre-specified thresholds was assessed for a combined clinical endpoint comprising ascites, variceal bleeding, hepatic encephalopathy, hepatocellular carcinoma (HCC), liver transplantation and mortality. A total of 197 patients (61 % men) with median age of 54 years were followed-up for 693 patient-years (median (IQR) 43 (26 to 58) months). The main diagnoses were NAFLD (41 %), viral hepatitis (VH, 25 %) and alcohol-related liver disease (ArLD; 14 %). During follow-up, 14 new clinical events, and 11 deaths occurred. Clinical outcomes were predicted by liver cT1 greater than 825 ms with HR 9.9 (95 % CI: 1.29 to 76.4, p = 0.007), TE greater than 8kPa with HR 7.8 (95 % CI: 0.97 to 62.3, p = 0.02) and FIB-4 greater than 1.45 with HR 4.09 (95 % CI: 0.90 to 18.4, p = 0.05). In analysis taking into account technical failure and unreliability, liver cT1 greater than 825 ms could predict clinical outcomes (p = 0.03), but TE greater than 8kPa could not (p = 0.4). These investigators carried out multi-variate analysis that indicated liver cT1 to perform as well as T1, better than APRI, but worse than Fibrosis-4 (FIB-4) and aspartate aminotransferase/alanine aminotransferase (AST/ALT) ratio. However, analysis also showed that at the pre-specified cut-offs, liver cT1 performed better than T1 and FIB-4 and similarly to AST/ALT and APRI. Meaningful cut-offs are desirable in clinical use, indicating liver cT1 has independent clinical prognostic relevance. These multi-variate results should be taken with caution as the number of events per variable was lower than the minimum recommended threshold and therefore may yield results with high margin of error. Moreover, these researchers stated that liver cT1 is likely to have a greater impact earlier rather than later in the screening process for the detection of early liver fibroinflammatory disease and aid in monitoring these patients non-invasively. Studies of even larger cohorts may bring out these differences in the future. The authors concluded that liver iron-corrected T1 (cT1), TE and serum-based blood biomarkers can identify patients at risk of developing clinical outcomes in a cohort with mixed CLD etiologies, typical of general hepatology cohorts; however, when taking into account technical failures of MRI and TE, MRI and blood markers performed better than TE. They stated that further multi-center studies are needed to validate these findings.

These researchers stated that drawbacks of this study included no fixed follow-up time-point, which meant that the AUC analyses, sensitivities, specificities, PPVs and NPVs generated in cut-off analysis should be taken with caution because unlike in Cox proportional hazards and Kaplan-Meier analysis, the effect of patients being censored at different time-points was not taken into account. The patient cohort included a wide range of liver disease severity and etiologies (from patients with mild fibrosis to those with cirrhosis). They stated that future studies should examine the prognostic value of liver cT1 alongside other non-invasive tests in pre-specified liver disease severity groups.

Andersson et al (2021) stated that NAFLD is increasing in prevalence worldwide. NAFLD is associated with excess risk of all-cause mortality, and its progression to NASH and fibrosis accounts for a growing proportion of cirrhosis and HCC, and thus is a leading cause of liver transplant worldwide. Non-invasive precise methods to identify patients with NASH and NASH with significant disease activity and fibrosis when the disease is still modifiable are crucial. These researchers examined the clinical utility of cT1 versus MRI liver fat for identification of NASH participants with NAS greater than or equal to 4 and F greater than or equal to 2 (“high-risk” NASH). Data from 5 clinical studies (n = 543) with participants suspected of NAFLD were pooled or used for individual participant data meta-analysis. The diagnostic accuracy of the MRI biomarkers to stratify NASH patients was determined using AUROC. A stepwise increase in cT1 and MRI liver fat with increased NAFLD severity was demonstrated, and cT1 was significantly higher in NASH participants with fibrosis grade greater than or equal to 2 (high-risk NASH). The diagnostic accuracy (AUROC [95 % CI]) of cT1 to identify those with NASH was 0.78 [CI: 0.74 to 0.82], for liver fat was 0.78 [CI: 0.73 to 0.82], and when combined with MRI liver fat was 0.82 [CI: 0.78 to 0.85]. The diagnostic accuracy of cT1 to identify those with high-risk NASH was good (AUROC: 0.78 [CI: 0.74 to 0.82]), was superior to MRI liver fat (AUROC: 0.69 [CI: 0.64 to 0.74]) and was not substantially improved by combining it with MRI liver fat (AUC: 0.79, [CI: 0.75 to 0.83]). The meta-analysis showed similar performance to the pooled analysis for these biomarkers. The authors concluded that these findings showed that both MRI liver fat and cT1 are effective biomarkers for identifying those with NASH; however, cT1 was superior in identifying NASH patients at greatest risk of disease progression, and thus has the potential to reduce unnecessary biopsies by providing an accurate and reliable alternative in the clinical care pathway.

Imajo and co-workers (2021) stated that non-invasive assessment of NASH is increasing in desirability due to the invasive nature and costs associated with the current form of evaluation – liver biopsy. Quantitative mpMRI to measure liver fat (PDFF) and fibro-inflammatory disease (cT1), as well as elastography techniques (VCTE liver stiffness measure), MRE and 2D Shear-Wave elastography (SWE) to measure stiffness and fat (CAP) are emerging alternatives which could be used as safe surrogates to liver biopsy. In a prospective, observational study, these researchers examined the agreement of non-invasive imaging modalities with liver biopsy, and their subsequent diagnostic accuracy for identifying NASH patients. From January 2019 to February 2020, Japanese patients suspected of NASH were recruited and were screened using non-invasive imaging techniques; mpMRI with LiverMultiScan, VCTE, MRE and 2D-SWE. Patients were subsequently biopsied, and samples were scored by 3 independent pathologists. The diagnostic performances of the non-invasive imaging modalities were evaluated using AUC with the median of the histology scores as the gold standard diagnoses. Concordance between all 3 independent pathologists was further explored using Krippendorff’s alpha from weighted kappa statistics. A total of 145 patients with mean age of 60 (SD: 13 years), 39 % women, and 40 % with BMI of greater than or equal to 30 kg/m2 were included in the analysis. For identifying patients with NASH, MR liver fat and cT1 were the strongest performing individual measures (AUC: 0.80 and 0.75, respectively), and the mpMRI metrics combined (cT1 and MR liver fat) were the overall best non-invasive test (AUC: 0.83). For identifying fibrosis of greater than or equal to 1, MRE performed best (AUC: 0.97), compared to VCTE-liver stiffness measure (AUC: 0.94) and 2D-SWE (AUC: 0.94). For assessment of steatosis of greater than or equal to 1, MR liver fat was the best performing non-invasive test (AUC: 0.92), compared to CAP (AUC: 0.75). Assessment of the agreement between pathologists showed that concordance was best for steatosis (alpha = 0.58), moderate for ballooning (alpha = 0.40) and fibrosis (alpha = 0.40), and worst for lobular inflammation (alpha = 0.11). The authors concluded that quantitative mpMRI is an effective alternative to liver biopsy for diagnosing NASH and non-alcoholic fatty liver; and therefore, may offer clinical utility in patient management.

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The authors stated that this study was not without its drawbacks, with the pre-screening step prior to liver biopsy likely truncating the correlations that would be observed across the full disease spectrum. While this may also impact the diagnostic accuracy evaluated, this pre-screening approach is representative of clinical practice. Observed failure rates for the US-based methods also have the potential to skew the results given its dependence on BMI, with patients with high BMI less likely to be included in the study. In practice, this has a bearing on the value proposition of such technologies for screening and monitoring as failed measurements will result in necessary further clinical tests for patients.

Schaapman and colleagues (2021) stated that NAFLD is a common cause of chronic liver disease, affecting more than 25 % of the adult population. NAFLD covers a spectrum including simple steatosis and NASH. Liver biopsy is currently the reference standard to discriminate between hepatic steatosis and steatohepatitis. Since liver biopsy has several disadvantages, non-invasive diagnostic methods with high sensitivity and specificity are desirable for the analysis of NAFLD. Improvements in MRI technology are continuously being implemented in clinical practice, specifically mpMRI methods such as PDFF, T2 *, and T1 mapping, along with MRE. The authors concluded that mpMRI of the liver has a promising role in the clinical management of NAFLD with quantification of fat content, iron load, and fibrosis, which are features in NAFLD. These researchers stated that further studies are needed to refine the sensitivity and specificity of these mpMRI methods, the use of MRI in the non-invasive evaluation in patients suspected for NAFLD, and the evaluation of their prognostic potential. Level of Evidence = V.

Beyer et al (2021) noted that MRI-based proton density fat fraction (PDFF) and the US-derived controlled attenuation parameter (CAP) are non-invasive techniques for quantifying liver fat, which can be used to evaluate steatosis in patients with NAFLD. In a retrospective study, these researchers compared these 2 techniques to histopathological graded steatosis for the evaluation of fat levels in a large pooled NAFLD cohort. This trial pooled n = 581 participants from 2 suspected NAFLD cohorts (mean age (SD) of 56 (12.7) years, 60 % women). Steatosis was graded according to NASH-CRN criteria. Liver fat was measured non-invasively using PDFF (with Liver MultiScan’s Iterative Decomposition of water and fat with Echo Asymmetry and Least-squares estimation method, LMS-IDEAL, Perspectum, Oxford) and CAP (FibroScan, Echosens, France), and their diagnostic performances were compared. LMS-IDEAL and CAP detected steatosis grade of 1 or higher with AUROCs of 1.00 (95 % CI: 0.99 to 1.0) and 0.95 (95 % CI: 0.91 to 0.99), respectively. LMS-IDEAL was superior to CAP for detecting steatosis grade of 2or higher with AUROCs of 0.77 (95 % CI: 0.73 to 0.82] and 0.60 (95 % CI: 0.55 to 0.65), respectively. Similarly, LMS-IDEAL out-performed CAP for detecting steatosis grade 3 or higher with AUROCs of 0.81 (95 % CI: 0.76 to 0.87) and 0.63 (95 % CI: 0.56 to 0.70), respectively. The authors concluded that LMS-IDEAL was able to diagnose individuals accurately across the spectrum of histological steatosis grades. CAP performed well in identifying individuals with lower levels of fat (steatosis grade 1 or higher); however, its diagnostic performance was inferior to LMS-IDEAL for higher levels of fat (steatosis grade 2 or higher and grade 3 or higher).

The authors stated that the large number of subjects with biopsy-paired, non-invasive measures of liver fat was a strength of this study as larger cohort sizes would result in more accurate and representative results; however, pooling together participants from 2 independent studies may also have some drawbacks. Not only were the participants from different geographic populations (U.S. and Japan); they were also selected using 2 slightly different sets of criteria resulting in a pooled cohort with a mixture of liver health profiles. Study 1 only biopsied patients after a preliminary PDFF screening, which in NASH clinical trials is not an uncommon method to reduce screen fails at confirmatory biopsy, but potentially biased the study population by excluding some of the likely false-negatives in the biopsy comparison. While this was a limitation of the analysis, it was mitigated by pooling the data from the 2 studies. In addition, across the pooled cohort, MRI and CAP measurements were not all carried out at the same center, using the same MRI scanner models or US devices, or by the same operators. Another drawback was the use of liver biopsy as the reference for assessing steatosis, which although it remains the gold standard, is associated with sampling errors, as well as intra- and inter-observer variability. Furthermore, in this pooled cohort, biopsies were not centrally read and while the combined sample size was substantial it did not contain a large number of those without steatosis; therefore, linear relationships may not be accurately reflected. Some of these limitations could explain why the correlations between biopsy scores and LMS-IDEAL in this trial tended to be slightly lower than those reported in the literature. For example, the literature correlations between MRI-PDFF and histological scores ranged between r = 0.74 and r = 0.82, compared with 0.54 in the current study. Notably, exploration of the 2 cohorts separately showed that the correlation between LMS-IDEAL and histology for study 1 was r = 0.58 and for study 2 was r = 0.71. It should also be noted that the presence of hepatic fibrosis has been reported to reduce the correlation between biopsy results and MRI PDFF, which was not accounted for in this trial.

Furthermore, an UpToDate review on “Epidemiology, clinical features, and diagnosis of nonalcoholic fatty liver disease in adults” (Sheth and Chopra, 2021) does not mention LiverMultiScan / multiparametric MRI as a management tool.

Arndtz et al (2021) stated that non-invasive monitoring of disease activity in autoimmune hepatitis (AIH) has potential advantages for patients for whom liver biopsy is invasive and with risk. These investigators examined the association of mpMRI with clinical course of patients with AIH. They prospectively recruited 62 patients (median age of 55 years; 82 % women) with clinically confirmed AIH. At recruitment, patients underwent mpMRI with LiverMultiScan alongside clinical investigations, which were repeated after 12 to 18 months. Associations between iron-corrected T1 (cT1) and other markers of disease were examined at baseline and at follow-up. Discriminative performance of cT1, liver stiffness, and enhanced liver fibrosis (ELF) to identify those who failed to maintain remission over follow-up was examined using the AUCs. Baseline cT1 correlated with alanine aminotransferase (Spearman’s correlation coefficient [r S] = 0.28, p = 0.028), aspartate aminotransferase (r S = 0.26, p = 0.038), international normalized ratio (r S = 0.35 p = 0.005), Model for ESLD (r S = 0.32, p = 0.020), ELF (r S = 0.29, p = 0.022), and liver stiffness r S = 0.51, p < 0.001). After excluding those not in remission at baseline (n = 12), 32 % of the remainder failed to maintain remission during follow-up. Failure to maintain remission was associated with significant increases in cT1 over follow-up (AUC, 0.71; 95 % CI: 0.52 to 0.90; p = 0.035) but not with changes in liver stiffness (AUC, 0.68; 95 % CI: 0.49 to 0.87; p = 0.067) or ELF (AUC, 0.57; 95 % CI: 0.37 to 0.78; p = 0.502). cT1 measured at baseline was a significant predictor of future loss of biochemical remission (AUC, 0.68; 95 % CI: 0.53 to 0.83; p = 0.042); neither liver stiffness (AUC, 0.53; 95 % CI: 0.34 to 0.71; p = 0.749) nor ELF (AUC, 0.52; 95 % CI: 0.33 to 0.70; p = 0.843) were significant predictors of loss of biochemical remission. The authors concluded that mpMRI showed underlying fibro-inflammatory activity and the breadth of abnormalities observed within the liver. By demonstrating an ability to identify those who will experience disease progression and loss of biochemical remission, mpMRI has shown promise in the phenotyping and risk stratification of individuals with high‐risk disease who may not be identified using serum biochemistry alone. These researchers stated that this proof-of-concept study identified mpMRI as a disruptive technology and justifies future prospective clinical trials in this area, potentially in combination with liver biopsy, to fully explore its utility. There is the potential to develop this technology further to aid in clinical decision-making, such as to improve identification of patients at risk of flare events and to provide an evidence base for making therapeutic decisions.

The authors stated that this study had several drawbacks. Specifically, the lack of liver histology resulted in the inability to examine the correlations between cT1 and liver histology at both time-points in the study. Previous studies have reported strong correlations between MR‐based metrics and histologic measures of fibrosis and inflammation; however, prospective studies pairing mpMRI techniques and biopsy in AIH are justified to enable further understanding of the associations between cT1 and liver inflammation and fibrosis in this population. On correlation analysis, the strengths and significances of associations between cT1/cT1 IQR and other surrogate biomarkers (apart from ALT and AST) were lower when assessed at the follow‐up visit compared to the analysis at baseline. This difference may partly be explained by the reduction in statistical power resulting from the smaller sample size at follow‐up after excluding those patients who were not in biochemical remission at baseline and those who did not undergo mpMRI at follow‐up. The exclusion of those who were not in biochemical remission at baseline may have introduced selection bias to the latter analysis, with the cohort not being representative of that analyzed at baseline. Furthermore, patients who failed to maintain biochemical remission have been shown to have a higher frequency of cirrhosis and have also shown evidence of portal hypertension. Because 24 % had evidence of cirrhosis with portal hypertension, this could have attributed to the frequency of suboptimal response; thus, future analyses following the same consistent cohort over time might yield a better understanding of the changes associated with the correlation of these markers over time. Nevertheless, from the presented findings, it was evident that cT1 is capable of characterizing liver fibro-inflammatory activity in an orphan disease where there is a growing unmet need due to the decrease in liver biopsy and the variability associated with patient care in clinical practice.

Torres and D’Ippolito (2021) stated that chronic liver disease (CLD) is a major public health problem globally, primarily due to the growing epidemic of obesity, alcohol consumption, and hepatitis. The central changes in the pathogenesis of CLD, such as fibrosis, necro-inflammatory activity, iron over-load, and fat accumulation, have traditionally been detected and monitored by biopsy, an invasive, costly, and limited procedure that is associated with non-negligible morbidity and mortality, making it imperative to replace liver biopsy with options that are more clinically practical, affordable, and accurate. mpMRI is currently one of the most promising tools for this task, because it makes it possible to use a wide range of advanced sequences that cover and analyze the complex spectrum of metabolic and cellular changes in the liver parenchyma, which has led some researchers to consider a future in which virtual liver biopsy will be carried out by mpMRI. Preliminary evidence suggested that, in comparison with biopsy, mpMRI has a better cost-benefit ratio. Advanced MRI sequences for the evaluation of CLD include proton magnetic resonance spectroscopy (1H-MRS); proton density fat fraction measurement; T2 and T2* mapping; T1 mapping; elastography; diffusion-weighted imaging; susceptibility-weighted imaging; and dynamic series of temporal enhancement by para-magnetic contrast agents, to calculate the rate of intracellular uptake by hepatocytes and the extracellular volume fraction. Most studies of CLD have employed such sequences in different combinations to create a multi-parametric protocol. The authors concluded that there have been and will continue to be, in the coming years, an increasing number of exciting new developments the field of mpMRI, with technical refinements of sequences that are expected to be increasingly more accurate in detecting the gamut of parenchymal alterations in diffuse liver disease; thus, avoiding the need for histological evaluation in many situations. In the context of the development and maturation of new treatments, the increasing calls for closer, non-invasive clinical monitoring of patients, and the low specificity of serum biomarkers, mpMRI appeared to have great potential to fill an unmet need for better diagnostic tools and to transform clinical practice. The design and clinical validation of future imaging studies must take into account the challenge posed by the fact that percutaneous liver biopsy is not an ideal reference standard for fibrosis, steatosis, or hemosiderosis, as well as the fact that it may be desirable to correlate the imaging findings with clinically relevant outcomes.

Heneghan et al (2022) noted that in AIH, clinical practice and treatment guidelines frequently diverge as a reflection of disease heterogeneity and challenges in achieving standardized care. These investigators examined the use of mpMRI in patients with AIH, and the impact of this technology on physicians’ decision-making and intended patient management. This study included 82 AIH patients, recruited from 2 sites between June and November 2019 as part of an observational cohort study; they underwent non-contrast MRI alongside their standard clinical investigations. Correlations between cT1 and other markers of disease were examined alongside the use of imaging markers to risk stratify patients in biochemical remission. The impact of mpMRI on clinical decision-making was examined using pair-wise t-tests. The discriminatory ability of the imaging markers was assessed using AUCs. cT1 had a significant impact on clinician intended patient management (p < 0.0001). cT1 correlated with ALT (p = 0.0005), AST (p < 0.001), IgG (p = 0.0005), and liver stiffness (p < 0.0001). Patients in deep biochemical remission (n = 11; AST/ALT less than 50 % upper limit of normal [ULN] and IgG less than 12 g/L) had low cT1, while 7/34 in normal biochemical remission (AST/ALT between 50 % and 100 % of ULN) had high cT1 and were at risk of disease flare. cT1 measures of disease heterogeneity, ALP and bilirubin made the best predictor of those not in biochemical remission (AUC: 0.85). The authors concluded that mpMRI quantitative biomarkers have shown a positive impact on clinicians intended management plan as well as utility in characterizing the fibro-inflammatory status of those in various gradations of biochemical remission. By identifying differences between those in normal biochemical remission, cT1 has shown promise in the phenotyping and risk stratification of individuals with this orphan disease who may not be identified using serum biochemistry and liver stiffness alone. Moreover, these researchers stated that future analyses examining the associations between disease and clinical outcomes should also evaluate the use of markers of disease heterogeneity.

The authors stated that as this was a real world study, concurrent liver biopsy was not included in the study protocol, as this would have deviated from standard of care. Consequently, these investigators acknowledged the limitations to this study; specifically, the inability to examine relationships between cT1 and histological findings. Nevertheless, previous studies have shown significant correlations between cT1 and histology in both adults and pediatrics. This cross-sectional study only covered a single time-point; therefore, following this cohort over time will yield a better understanding of the changes associated with these markers as well as the impact they may have on longitudinal disease monitoring. Finally, as it has been shown that patient management may vary between clinicians, future meta-analyses evaluating large pooled data across multiple trusts may yield useful information that will support the results presented in this investigation. These studies should also gather data regarding compliance to medication, previous relapses (and their frequency), induction treatment regimens, as well as other factors that can affect the remission status of a patient. Statistical analyses following these investigations should control for these co-variates within any multi-variate models generated so as to ensure that any potential confounding effects these factors have will not influence the outcomes observed.

Bajre et al (2022) stated that AIH is a rare chronic progressive liver disease, managed with corticosteroids and immunosuppressants and monitored using a combination of liver biochemistry and histology. Liver biopsy (gold standard) is invasive, costly and has risk of complications. Non-invasive imaging using mpMRI can detect the presence and extent of hepatic fibro-inflammation in a risk-free manner. By means of early economic modelling, these investigators examined the affordability of using mpMRI as an alternative to liver biopsy. Medical test costs associated with following 100 patients over a 5-year time horizon were assessed from a National Health Service payor perspective using tariff costs and average biopsy-related adverse events (AEs) costs. Sensitivity analyses modelling the cost consequences of increasing the frequency of mpMRI monitoring within the fixed cost of liver biopsy were performed. Per 100 moderate/severe AIH patients receiving an annual mpMRI scan (in place of biopsy), early economic modelling showed minimum cost savings of £232,333. Per 100 mild/moderate AIH patients receiving 3 mpMRI scans over 5 years estimated minimum cost savings were £139,400. One-way sensitivity analyses showed increasing the frequency of mpMRI scans from 5 to 10 over 5 years in moderate/severe AIH patients resulted in a cost saving of £121,926.20. In patients with mild/moderate AIH, an increase from 3 to 6 mpMRI scans over 5 years could save £73,155.72. In a minimalistic approach, the use of 5 mpMRI scans was still cost saving (£5,770.48) if they were to replace 2 biopsies over the 5-year period for all patients with moderate/severe or mild/moderate AIH. The authors concluded that the findings of this analysis suggested that the integration of non-invasive mpMRI scanning in AIH patient pathways has the potential to improve the monitoring care pathway and may result in cost savings for AIH patients in secondary care in the NHS in England. Moreover, more frequent, and detailed phenotyping could result in improvements in patient outcomes, such as better titrated immunosuppression for individual patients. These investigators stated that with more research data, from clinical trial and real-world usage monitoring, a more in-depth economic evaluation of the impact on the adoption of mpMRI into the AIH patient monitoring pathway can be produced.

The authors stated that this study had several drawbacks. First, there is heterogeneity in the management of AIH patients; thus, liver biopsy may not be used as routinely by all clinicians as recommended by clinical guidelines. Therefore, some of the assumptions used in the modelling pertaining to liver biopsy frequency may not reflect the costs associated with real-world patient management. Nevertheless, a long-term review of AIH outcomes has shown after achievement of remission (confirmed histologically reduced relapse rates can be achieved, however, there is still a very high relapse rate (up to 80 %) if treatment is stopped after initial remission (biochemical ± histological). Consequently, to compensate for this, a minimalistic model considering replacing 2 biopsies over 5 years with mpMRI scanning was investigated. Although clinical guidelines have been updated more recently to reflect the changes in the use of liver biopsy, it is still recognized as the gold standard for diagnosis and disease monitoring. The need for non-invasive technologies to replace liver biopsy is becoming more widely recognized. Second, although a small number of patients with AIH reach complete remission, during the base case modelling, these researchers assumed 0 cure rates. This assumption was made as the data needed for such analyses, including the healthcare costs associated with AIH patient management, could only be obtained from real-world evaluations implementing the suggested hypothetical model. Thus, as the early economic evaluation suggested that using mpMRI scanning for monitoring in AIH is cost-saving, future investigation should examine the feasibility of implementing this hypothetical monitoring model across different trusts in the NHS. This model would also need to account for discount costs depending on the most appropriate payment mode accepted by the NHS payors. Moreover, additional testing could improve overall patient care, which may have other direct and indirect health service cost savings. Therefore, as non-invasive techniques can significantly improve patient monitoring, these investigators recommended that a full economic evaluation be carried out using trial and real-world data – incorporating analyses evaluating QALY gains into the cost model – to fully quantify the improvement in healthcare the inclusion of mpMRI scanning in the standard of care might bring.

The American Association of Clinical Endocrinology (AACE)’s clinical practice guideline on the “Diagnosis and management of nonalcoholic fatty liver disease” (Cusi et al, 2022) states that “newer imaging techniques are becoming available. Velacur (Sonic Incytes Medical Corp.) is a point-of-care liver assessment device based on Shear Wave Absolute Vibro-Elastography that incorporates elastography and a greater liver volume visualization. LiverMultiScan uses multiparametric MRI to noninvasively quantify liver fat and cT1 signal maps of the liver to assess disease activity (NAFLD activity score [NAS]) and potentially outcomes. These techniques are currently being used largely in research for screening studies, or to assess primary end points in clinical trials for investigational drugs in development for the treatment of NASH. Both have received FDA-approval for use in persons with chronic liver disease and await future work to fully assess their place in the diagnostic algorithm of persons with NAFLD”.

Tan et al (2023) stated that bariatric surgery is the most effective treatment for morbid obesity and reduces the severity of NAFLD in the long-term; however, less is known regarding the effects of bariatric surgery on liver fat, inflammation, and fibrosis during the early stages following bariatric surgery. In an exploratory study, these researchers employed advanced imaging methods to examine NAFLD and fibrosis changes during the early metabolic transitional period following bariatric surgery. A total of 9 subjects with morbid obesity underwent sleeve gastrectomy; mpMRI and MRE were carried out at baseline, during the immediate (1 month), and late (6 months) post-surgery period. Liver fat was measured using PDFF, disease activity using iron-correct T1 (cT1), and liver stiffness using MRE. Repeated measured ANOVA was used to assess longitudinal changes and Dunnett’s method for multiple comparisons. All subjects (age of 45.1 ± 9.0 years, BMI 39.7 ± 5.3 kg/m2) had elevated hepatic steatosis at baseline (PDFF greater than 5 %). In the immediate post-surgery period, PDFF decreased significantly from 14.1 ± 7.4 % to 8.9 ± 4.4 % (p = 0.016) and cT1 from 826.9 ± 80.6 ms to 768.4 ± 50.9 ms (p = 0.047). These improvements continued to the later post-surgery period. Bariatric surgery did not reduce liver stiffness measurements. The authors concluded that the findings of this study supported the use of MRI as a non-invasive tool to monitor NAFLD in patient with morbid obesity during the early stages following bariatric surgery.

The authors stated that this study had several drawbacks. First, although the sample size was small (n = 9) and should be confirmed in larger studies, these findings were consistent with those presented in the literature. Second, none of the subjects had advanced liver fibrosis; therefore, the potential impact of bariatric surgery and subsequent changes in mpMRI and MRE in subjects with more advanced disease will need to be examined in future studies. Future investigation should examine the use of such MRI technologies over longer follow-up periods. Third, all subjects in this trial underwent sleeve gastrectomy, and it remains uncertain if other procedures such as Roux-en-Y gastric bypass would produce similar changes.

Roca-Fernandez et al (2023) stated that CLD is associated with increased cardiovascular disease (CVD) risk. These investigators examined if early signs of liver disease (measured by iron-corrected T1-mapping [cT1]) were associated with an increased risk of major CVD events. Liver disease activity (cT1) and fat (PDFF) were measured using LiverMultiScan between January 2016 and February 2020 in the U.K. Biobank imaging sub-study. Using multi-variable Cox regression, these researchers examined associations between liver cT1 (MRI) and primary CVD (coronary artery disease, atrial fibrillation [AF], embolism/vascular events, heart failure [HF] and stroke), and CVD hospitalization and all-cause mortality. Liver blood biomarkers, general metabolism biomarkers, and demographics were also included. Subgroup analysis was carried out in those without metabolic syndrome (defined as at least 3 of: a large waist, high triglycerides, low high-density lipoprotein cholesterol [HDL-C], increased systolic blood pressure [SBP], or elevated hemoglobin A1c). A total of 33,616 participants (mean age of 65 years, mean BMI of 26 kg/m2, mean hemoglobin A1c of 35 mmol/mol) had complete MRI liver data with linked clinical outcomes (median time to major CVD event onset of 1.4 years [range of 0.002 to 5.1]; follow-up of 2.5 years [range of 1.1 to 5.2]). Liver disease activity (cT1), but not liver fat (PDFF), was associated with higher risk of any major CVD event (HR 1.14; 95 % CI: 1.03 to 1.26; p = 0.008), AF (1.30; 1.12 to 1.51; p < 0.001); HF (1.30; 1.09 to 1.56; p = 0.004); CVD hospitalization (1.27; 1.18 to 1.37; p < 0.001); and all-cause mortality (1.19; 1.02 to 1.38; p = 0.026). FIB-4 index was associated with HF (1.06; 1.01 to 1.10; p = 0.007). Risk of CVD hospitalization was independently associated with cT1 in individuals without metabolic syndrome (1.26; 1.13 to 1.4; p < 0.001). The authors concluded that liver disease activity, as indexed by cT1, was independently associated with a higher risk of incident CVD and all-cause mortality, independent of pre-existing metabolic syndrome, liver fibrosis or fat. These researchers stated that the findings of this study suggested the MRI-derived biomarker cT1 has a promising role to play in risk stratification of those at greatest risk of CVD morbidity and mortality.

These investigators noted that many CVD risk scores exist, including the QRISK score, Framingham score. and ASCVD score, which are already employed clinically. However, considering these results, and the momentum towards appreciating multi-system disease and multi-specialty care, these findings highlighted an opportunity to improve on these risk scores by incorporating the degree of liver-related disease activity. In relation to the FIB-4 index, while these researchers did not observe a robust association with CVD risk, it should be acknowledged that the currently adopted thresholds to rule out or rule in significant liver fibrosis are designed for patients being specifically evaluated for CLD, and may be inappropriate in “healthier” populations where CLD is under-diagnosed or at an earlier, potentially more modifiable, stage. Of course, the fact that these authors did not observe an association with the FIB-4 index was being attributed to the likely absence of fibrosis; however, a notable limitation of the U.K. Biobank imaging study was that there was a delay of approximately 10 years between the blood tests and imaging, which may have prevented meaningful interpretation of blood test results, although correction for this by imputation of annualized change did not alter the main findings. Other notable limitations were the lack of confirmatory biopsy. Furthermore, the study cohort was homogenous with a predominant white ethnicity and was slightly older compared to the whole U.K. Biobank cohort; but had no clinically significant differences in the mean BMI or proportion of males. Low numbers in this imaging cohort or collinearity effects may have prevented full investigation of known CVD risk factors, such as smoking, cholesterol, BMI, or diabetes. Lastly, these analyses had short duration of follow-up for imaging and relied on ICD-10 codes for outcome collection.

The AASLD’s practice guidance on “The clinical assessment and management of nonalcoholic fatty liver disease” (Rinella et al, 2023) stated that “Imaging techniques such as cT1 may also be considered for the identification of “at-risk” NASH. In an individual patient meta-analysis of 543 patients, cT1 performed well (area under the receiver operating characteristic curve: 0.78), although this requires further validation in large independent cohorts. Precise cutoffs have not been validated, and superiority over less expensive, point-of-care techniques remains to be demonstrated”.

Magnetic Resonance Elastography

Magnetic resonance elastography (MRE) uses wave propagation and tissue deformation analysis to assess changes to tissue viscoelasticity caused by disease.14 It purportedly is based on principles similar to ultrasound transient elastography and acoustic radiation force impulse (ARFI); however, MRE supposedly assesses wave propagation and tissue displacement in three dimensions rather than one dimension.14 This form of imaging involves placing a probe against the individual’s back which emits low frequency vibrations that pass through the liver and can reportedly be measured by the MRI spin echo sequence.

Venkatesh et al (2013) stated that many pathological processes cause marked changes in the mechanical properties of tissue. MR elastography (MRE) is a non-invasive MRI based technique for quantitatively assessing the mechanical properties of tissues in-vivo. Magnetic resonance elastography is performed by using a vibration source to generate low frequency mechanical waves in tissue, imaging the propagating waves using a phase contrast MRI technique, and then processing the wave information to generate quantitative images showing mechanical properties such as tissue stiffness. Since its first description in 1995, published studies have explored many potential clinical applications including brain, thyroid, lung, heart, breast, and skeletal muscle imaging. However, the best-documented application to emerge has been the use of MRE to assess liver disease. Multiple studies have demonstrated that there is a strong correlation between MRE-measured hepatic stiffness and the stage of fibrosis at histology. The emerging literature indicated that MRE can serve as a safer, less expensive, and potentially more accurate alternative to invasive liver biopsy, which is currently the gold standard for diagnosis and staging of liver fibrosis.

The British HIV Association’s guidelines on “The management of hepatitis viruses in adults infected with HIV 2013” (Wilkins et al, 2013) suggested hepatic TE (FibroScan™ or ARFI) as the non-invasive investigation of choice (2B) but if unavailable, or when reliable TE readings are not obtained, a blood panel test (aspartate transaminase to platelet ratio index [APRI], FIB-4, enhanced liver fibrosis [ELF], Fibrometer™, Forns Index, FibroTest™) as an alternative (2C)”. It did not mention MR elastography as a management tool.

Tubb (2015) states that “Magnetic resonance elastography is an emerging MRI technology that provides sensitive and semi-quantitative assessment of tissue stiffness. The most promising clinical application for MR elastography is the assessment of liver stiffness as a surrogate marker of liver disease and fibrosis”.

Furthermore, an UpToDate review on “Noninvasive assessment of hepatic fibrosis: Overview of serologic and radiographic tests” (Curry and Afdhal, 2015) states that “Radiologic methods for staging hepatic fibrosis are emerging as promising tools. The methods include ultrasound-based transient elastography, magnetic resonance elastography, acoustic radiation force impulse imaging, and cross-sectional imaging. Ultrasound-based transient elastography is the most studied radiologic method for staging hepatic fibrosis. When ultrasound-based transient elastography is used in a clinical setting, commonly used cutoffs for significant fibrosis and cirrhosis are >7 kPa and >11 to 14 kPa, respectively”.

Magnetic Resonance Elastography for Prediction of Ascites in Persons with Chronic Liver Disease

Abe and colleagues (2018) evaluated the utility of MRE as a non-invasive method for predicting ascites in patients with CLD. A total of 208 CLD patients underwent MRE to measure LS at the authors’ institution from March 2013 to June 2015 were include in this trial. These investigators evaluated the diagnostic performance of MRE for predicting the presence of ascites using ROC curve analysis and compared the performance with that of serum fibrosis markers. Multi-variate logistic regression analysis was performed to identify factors associated with the presence of ascites. The cumulative incidence of ascites was examined in patients without ascites at baseline. The pathological stage of liver fibrosis was evaluated in 81 CLD patients using histopathologic diagnosis. Of the 208 patients, 41 had ascites. The optimal cut-off LS value for the presence of ascites was 6.0 kPa (AU ROC curve = 0.87). The AUROC curve for the presence of ascites was significantly higher for MRE than that for fibrosis markers. Multi-variate analysis revealed that LS greater than 6.0 kPa was an independent risk factor for the presence of ascites. The cumulative incidence of ascites was significantly higher among those with LS values of greater than 6.0 kPa. There was significantly greater diagnostic accuracy for liver fibrosis stage greater than or equal to 4 with MRE than that with fibrosis markers. The authors concluded that compared with serum fibrosis markers, MRE has higher diagnostic performance in predicting the presence of ascites. They stated that MRE-based LS has the potential to predict the presence of ascites in CLD patients.

Magnetic Resonance Elastography for Non-Alcoholic Steatohepatitis (NASH)

Park and associates (2017) stated that MRI techniques and US-based TE can be used in non-invasive diagnosis of fibrosis and steatosis in patients with NAFLD. In a prospective study, these researchers compared the performance of MRE versus TE for diagnosis of fibrosis, and MRI-based proton density fat fraction (MRI-PDFF) analysis versus TE-based controlled attenuation parameter (CAP) for diagnosis of steatosis in patients undergoing biopsy to assess NAFLD. They performed a cross-sectional study of 104 consecutive adults (56.7 % female) who underwent MRE, TE, and liver biopsy analysis (using the histologic scoring system for NAFLD from the Nonalcoholic Steatohepatitis Clinical Research Network Scoring System) from October 2011 through May 2016 at a tertiary medical center. All patients received a standard clinical evaluation, including collection of history, anthropometric examination, and biochemical tests. The primary outcomes were fibrosis and steatosis; secondary outcomes included dichotomized stages of fibrosis and NASH versus no NASH. Receiver operating characteristic curve analyses were used to compare performances of MRE versus TE in diagnosis of fibrosis (stages 1 to 4 versus 0) and MRI-PDFF versus CAP for diagnosis of steatosis (grades 1 to 3 versus 0) with respect to findings from biopsy analysis. MRE detected any fibrosis (stage 1 or more) with an area under the receiver operating characteristic curve (AUROC) of 0.82 (95 % CI: 0.74 to 0.91), which was significantly higher than that of TE (AUROC, 0.67; 95 % CI: 0.56 to 0.78). MRI-PDFF detected any steatosis with an AUROC of 0.99 (95 % CI: 0.98 to 1.00), which was significantly higher than that of CAP (AUROC, 0.85; 95 % CI: 0.75 to 0.96). MRE detected fibrosis of stages 2, 3, or 4 with AUROC values of 0.89 (95 % CI: 0.83 to 0.96), 0.87 (95 % CI: 0.78 to 0.96), and 0.87 (95 % CI: 0.71 to 1.00); TE detected fibrosis of stages 2, 3, or 4 with AUROC values of 0.86 (95 % CI: 0.77 to 0.95), 0.80 (95 % CI: 0.67 to 0.93), and 0.69 (95 % CI: 0.45 to 0.94). MRI-PDFF identified steatosis of grades 2 or 3 with AUROC values of 0.90 (95 % CI: 0.82 to 0.97) and 0.92 (95 % CI: 0.84 to 0.99); CAP identified steatosis of grades 2 or 3 with AUROC values of 0.70 (95 % CI: 0.58 to 0.82) and 0.73 (95 % CI: 0.58 to 0.89). The authors concluded that using prospective, head-to-head comparisons, they showed that MRI-based MRE and MRI-PDFF were significantly more accurate than ultrasound-based TE and CAP, respectively, for diagnosing fibrosis and steatosis in an American cohort of patients with biopsy-proven NAFLD. MRI-based techniques may be preferable to TE for accurate non-invasive assessment of NAFLD. Moreover, these researchers stated that future studies are needed to evaluate the clinical utility of MRI and TE for diagnosing fibrosis and steatosis in a multi-center, longitudinal design, both in observational and intervention studies. The cost-effectiveness of utilizing MRE versus TE and/or biopsy must also be evaluated to develop optimal diagnostic strategies for diagnosing NAFLD-associated fibrosis and steatosis.

The authors stated that this study had several drawbacks. The cross-sectional design of the study did not allow the assessment of MRE and TE for monitoring longitudinal changes in fibrosis. Since this was a single-center study in a highly specialized setting, the generalizability of its findings in other clinical settings was unknown. The median time interval between TE and biopsy was 107 days. A recent meta-analysis of paired liver biopsy studies has shown that the rate of fibrosis progression was slow, with an average progression of one stage to take 14.3 years in patients with NAFL and 7.1 years in patients with NASH. Thus, the time interval observed in this study was reasonable as fibrosis stage was unlikely to change within a year. Furthermore, the analyses showed that the biopsy to imaging time interval did not affect the diagnostic accuracy of MRI and TE. Nevertheless, rapid changes in steatosis were possible, and ideally biopsy and imaging should be performed contemporaneously within 1 week, if feasible. MRI-based techniques, including MRE and MRI-PDFF, were often expensive, although at the authors’ center the cost of MRE was lower than that of biopsy without the associated morbidity. Although TE was more widely available in some parts of the world, MRI techniques are more widely deployed in the United States, therefore MRE could also be made available on commercially available MRI platforms throughout the United States. While TE may be more useful for wide-spread screening, MRE may play a role in clinical trial assessments that require higher accuracy and precision. These investigators stated that further studies are needed to evaluate the cost-effectiveness of MRI over TE for diagnosing NAFLD-related fibrosis and steatosis in before implementing these competing non-invasive approaches in routine clinical practice.

Besutti and colleagues (2019) stated that non-invasive tests to diagnose NASH are urgently needed. In a systematic review, these investigators examined imaging accuracy in diagnosing NASH among NAFLD patients, using liver biopsy as reference. Eligible studies were systematic reviews and cross-sectional/cohort studies of NAFLD patients comparing imaging with histology, considering accuracy and/or associations. Medline, Scopus, Embase and Cochrane Library databases were searched up to April 2018. Studies were screened on title/abstract, then assessed for eligibility on full-text. Data were extracted using a pre-designed form. Risk of bias was assessed using Quality Assessment of Diagnostic Accuracy Studies-2 tool. Of the 641 studies screened, 61 were included in scoping review, 30 of which (with accuracy results) in data synthesis. Imaging techniques included: elastography (ARFI, MRE, TE), computed tomography (CT), MRI, scintigraphy and US. Histological NASH definition was heterogeneous. In 28/30 studies, no pre-specified threshold was used (high risk of bias). AUROCs were up to 0.82 for TE, 0.90 for ARFI, 0.93 for MRE and 0.82 for US scores. MR techniques with higher accuracy were spectroscopy (AUROC = 1 for alanine), susceptibility-weighted imaging (AUROC = 0.91), multi-parametric MRI (AUROC = 0.80), optical analysis (AUROC = 0.83), gadoxetic acid-enhanced MRI (AUROCs = 0.85) and super-paramagnetic iron oxide-enhanced MRI (AUROC = 0.87). Results derived mostly from single studies without independent prospective validation. The authors concluded that there is currently insufficient evidence to support the use of imaging to diagnose NASH. These researchers stated that more studies are needed on US and MR elastography as well as non-elastographic techniques, to-date the most promising methods.

AASLD guidelines describe the role of (MRE in identifying different degrees of fibrosis in patients with NAFLD, performing better than transient elastography for recognizing intermediate stage of fibrosis, but showing a same predictive value for advanced fibrosis stages (Lioni et al, 2018). Therefore AASLD guidelines conclude that MRE and TE are both useful tools for identifying NAFLD patients with advanced liver fibrosis.

Magnetic Resonance Elastography for the Diagnosis of Hepatic Fibrosis

Bi et al (2021) stated that few studies comprehensively compared the performance of MRE and TE in the diagnosis of liver fibrosis. In a systematic review and meta-analysis, these researchers compared the diagnostic effectiveness of these 2 techniques in patients with hepatic fibrosis to gain a better understanding of their overall diagnostic performance and aid in maximizing their clinical utility. They carried out systematic literature searches of the PubMed, Embase, Cochrane Library, and China National Knowledge Infrastructure databases to identify studies that applied MRE and TE in the diagnosis of liver fibrosis. The combined sensitivity, specificity, positive and negative likelihood ratios, and DORs were estimated using a bi-variate random effects model. Review Manager 5.2 was used to analyze the selected articles, and forest plot, sensitivity, and bias analyses were performed for the included literature. To determine the diagnostic efficacy of MRE and TE for liver fibrosis, pooled sensitivity and specificity analyses were conducted. A total of 8 studies met the inclusion criteria. In the diagnosis of stage F0-F1 liver fibrosis, MRE showed higher sensitivity than TE (OR = 0.62, 95 % CI: 0.41 to 0.95, p = 0.03). MRE also showed higher specificity than TE for diagnosing stage F2-F4 liver fibrosis (OR = 0.41, 95 % CI: 0.27 to 0.62, p < 0.0001). There was no difference in the sensitivity of MRE and TE to F2-F4 hepatic fibrosis and the specificity of MRE and TE to F0-F1 hepatic fibrosis. The authors concluded that in terms of sensitivity and specificity, MRE was superior to TE in diagnosing different stages of liver fibrosis to a certain extent. These researchers stated that MRE may be a useful, non-invasive method for the assessment of liver fibrosis in patients with chronic liver disease. Moreover, these investigators stated that there this meta-analysis had several drawbacks. First, it did not take into account comparisons of different age groups. Second, the details of heterogeneity were not analyzed. These researchers stated that future studies will seek to address these drawbacks.

Acoustic Radiation Force Impulse (ARFI)

Acoustic radiation forced impulse (ARFI) (e.g., Virtual Touch Imaging – Acuson S2000-3000) relies on short-duration, high-intensity acoustic pulses to quantify the mechanical properties of tissues, without manual compression, by measuring shear wave velocity induced by acoustic radiation and propagated in the tissue. This technique purportedly provides single one-dimensional measures of tissue elasticity, although the area can be positioned on a two-dimensional B mode image. The measurements are expressed as m/s, which supposedly indicates the shear wave speed traveling perpendicular to the shear wave source.

An assessment of ARFI by the Canadian Agency for Drugs and Technologies in Health (CADTH, 2016) found that studies were “favorable to the use of ARFI in hepatitis C”, but “[n]o literature was identified regarding evidence-based guidelines and clinical effectiveness on ARFI compared with liver biopsy in patients with Hepatitis C” . The review cited an economic analysis that found that ARFI was dominated by less costly and more effective options among chronic hepatitis C patients.

Plasma Cytokeratin-18

Castera and co-workers (2013) noted that a common clinical concern in patients with NAFLD is whether they have NASH or simple steatosis and, more importantly, what the stage of fibrosis is and whether the level of fibrosis has increased over time. Such concern is based on the fact that patients with NAFLD with advanced fibrosis are at greatest risk of developing complications of end-stage liver disease. Although it lacks sensitivity, ultrasonography is an accepted tool for steatosis screening. The controlled attenuation parameter (CAP) seems a promising screening technique, but requires further validation. Cytokeratin-18 (CK-18) has been extensively validated, but it is an imperfect serum marker of NASH. Ultrasonography-based TE can exclude advanced fibrosis and cirrhosis, but its main limitation is its reduced applicability in patients with NAFLD, which is not completely solved by use of the XL probe. Of the non-invasive serum markers, the NAFLD fibrosis score is the most validated and has appropriate accuracy in distinguishing patients with and without advanced fibrosis. The authors concluded that although non-invasive methods require further validation, they could be useful for selecting those patients with NAFLD who require a liver biopsy.

Cusi and associates (2014) stated that liver biopsy is the only reliable way of diagnosing and staging NASH but its invasive nature limits its use. Plasma caspase-generated CK-18 fragments have been proposed as a non-invasive alternative. These researchers studied its clinical value in a large multi-ethnic NAFLD population and examined its relationship to clinical/metabolic/histological parameters. A total of 424 middle-aged subjects were included in this study – these investigators measured adipose tissue, liver and muscle insulin resistance (IR), liver fat by magnetic resonance spectroscopy (MRS; n = 275) and histology (n = 318). Median CK-18 were elevated in patients with versus without NAFLD by MRS (209 [IQR: 137 to 329] versus 122 [IQR: 98 to 155]U/L) or with versus without NASH (232 [IQR: 151 to 387] versus 170 [IQR: 135 to 234]U/L, both p < 0.001). Plasma CK-18 raised significantly with any increase in steatosis, inflammation and fibrosis, but there was a significant overlap across disease severity. The CK-18 AUROC to predict NAFLD, NASH or fibrosis were 0.77 (95 % CI: 0.71 to 0.84), 0.65 (95 % CI: 0.59 to 0.71) and 0.68 (95 % CI: 0.61 to 0.75), respectively. The overall sensitivity/specificity for NAFLD, NASH and fibrosis were 63 % (57 to 70 %)/83 % (69 to 92 %), 58 % (51 to 65 %)/68 % (59 to 76 %) and 54 % (44 to 63 %)/85 % (75 to 92 %), respectively. CK-18 correlated most strongly with ALT (r = 0.57, p < 0.0001) and adipose tissue IR (insulin-suppression of FFA: r = -0.43; p < 0.001), less with steatosis, lobular inflammation and fibrosis (r = 0.28 to 0.34, all p < 0.001), but not with ballooning, BMI, metabolic syndrome or type 2 diabetes mellitus. The authors concluded that plasma CK-18 has a high specificity for NAFLD and fibrosis, but its limited sensitivity makes it inadequate as a screening test for staging NASH. Whether combined as a diagnostic panel with other biomarkers or clinical/laboratory tests may prove useful requires further study.

Kwok and colleagues (2014) reviewed current literature on the use of non-invasive tests to assess the severity of NAFLD. These researchers performed a systematic literature searching identified studies evaluating non-invasive tests of NASH and fibrosis using liver biopsy as the reference standard. Meta-analysis was performed for areas with adequate number of publications. Serum tests and physical measurements like TE have high NPV in excluding advanced fibrosis in NAFLD patients. The NAFLD fibrosis score comprises of 6 routine clinical parameters and has been endorsed by current American guidelines as a screening test to exclude low-risk individuals. The pooled sensitivities and specificities for TE to diagnose F ≥ 2, F ≥ 3 and F4 disease were 79 % and 75 %, 85 % and 85 %, and 92 % and 92 %, respectively. Liver stiffness measurement often fails in obese patients, but the success rate can be improved with the use of the XL probe. A number of biomarkers have been developed for the diagnosis of NASH, but few were independently validated. Serum/plasma CK-18 fragments have been most extensively evaluated and have a pooled sensitivity of 66 % and specificity of 82 % in diagnosing NASH. The authors concluded that current non-invasive tests are accurate in excluding advanced fibrosis in NAFLD patients, and may be used for initial assessment. Moreover, they stated that further development and evaluation of NASH biomarkers are needed.

Furthermore, an UpToDate review on “Epidemiology, clinical features, and diagnosis of nonalcoholic fatty liver disease in adults” (Sheth and Chopra, 2015) does not mention plasma cytokeratin-18 as a diagnostic tool.

Hepatic Artery Resistive Index for Evaluation of Fibrosis Progression in Individuals with NAFLD

Tana and colleagues (2018) stated that US can reveal the presence of steatosis in NAFLD, but its diagnostic accuracy to reveal signs of fibrosis is low except in advanced stages of disease (e.g., cirrhosis). Current guidelines suggest the use of clinical algorithms, such as the NAFLD fibrosis score, and elastography to predict the progression of fibrosis, and the integration of elastography improves the detection accuracy of LS. However, there is a lack of evidence about the correlation between clinical algorithms and conventional US, and elastography is limited by the relative low diffusion, necessity of training, and loss of diagnostic accuracy in patients with high BMI, waist circumference, or increased thickness of parietal walls, with consequent significant rates of failure of measurement of LS. Recently, the measurement of hepatic artery resistive index (HARI) has demonstrated a significant positive correlation with fibrosis degree, as measured with NAFLD fibrosis score, suggesting that the fibrous tissue accumulation may result in increased arterial rigidity and, thus, in a rise of resistance to flow, and that the different tissue composition of the liver (adipose versus fibrous) can influence HARI differently. These investigators stated that these issues should be further examined because some aspects are still unknown. The limited data currently justify the need of larger, prospective studies aimed at examining if HARI correlates with elastography results. In view of their effect on weight loss, serum lipid concentration, and hepatic arterial flow hemodynamics, it could be interesting to evaluate if lifestyle and diet changes can influence significantly HARI values in NAFLD patients.

Serum Angiotensin Converting Enzyme for Liver Fibrosis

Akar and colleagues (2018) examined if any possible relationship between serum angiotensin-converting enzyme (ACE) levels and the stages of liver fibrosis in patients with CHC. A total 100 CHC and 100 healthy subjects were enrolled in this study. The relationship between serum ACE level and the stages liver fibrosis was investigated using 3 different formats: group G-I, classic Ishak’s Score from F1 to F6; group G-II, mild [F1-2], moderate [F3-4] and severe [F5-6]; and group G-III, mild [less than or equal to F2] and advanced [F greater than 2]. The clinical usability of serum ACE level for both groups was also investigated. Median serum ACE levels were higher in the healthy group than in CHC (42.5 [7 to 119] versus 36 [7 to 91] U/I, p = 0.002). There was no statistical difference among the 3 different fibrosis groups (G-I, G-II, G-III, p = 0.797, p = 0.986, and p = 0.874) and no correlation between serum ACE level and the stages of liver fibrosis (r = 0.026, p = 0.923). The usability of serum ACE for evaluated patients with CHC and healthy subjects were calculated as 47 % and 64 %, respectively. The authors concluded that the findings of this study indicated that there was no relationship or correlation between serum ACE levels and stages of liver fibrosis in patients with CHC.

Serum miR-29a and miR-122 for Diagnosis of Non-Alcoholic Fatty Liver Disease

Jampoka and colleagues (2018) stated that NAFLD is an over accumulation of triglyceride in the liver without alcohol consumption, which its major cause is from insulin resistance. Patients with NAFLD can develop liver fibrosis, cirrhosis and HCC. MicroRNAs (miRNAs) are non-coding RNAs that regulate post-transcriptional gene silencing. Previous research reported that miR-29 family (a, b and c) and miR-122 have an important role in regulating insulin resistance related to NAFLD. These investigators examined if miR-29 and miR-122 can be possible biomarkers for non-invasive diagnosis of NAFLD. Serum samples were collected from 58 NAFLD patients and 34 healthy controls; miRNAs were extracted from serum by using miRNA purification kit followed by polyuridylation, reverse transcription and quantitative real-time PCR. Furthermore, these researchers analyzed the correlation between miR-29 and miR-122 and level of liver inflammation in NAFLD patients. They found that serum miR-29a levels in NAFLD patients were significantly lower (p = 0.006) than the control group, while miR-29c levels were unchanged, and miR-29b levels were undetectable. However, these researchers found that serum miR-122 levels in NAFLD patients were significantly higher (p < 0.001) than those found in the control group. For miR-29a, the area under curve (AUC) was 0.679 (p = 0.0065) with 60.87 % sensitivity and 82.35 % specificity. For miR-122, the AUC was 0.831 (p < 0.0001) with 75.00 % sensitivity and 82.35 % specificity. Interestingly, the level of serum miR-122 were significantly different between patients with not steatohepatitis (NAFLD Activity Score [NAS] of less than 4) and steatohepatitis (NAS of greater than or equal to 4), indicating that the levels of miR-122 were related to the severity of NAFLD. The authors concluded that serum levels of miR-29a and miR-122 might be possible biomarkers for non-invasive diagnosis of NAFLD.

Serum miRNA-221 and miRNA-222 for Progression of Liver Fibrosis

Abdel-Al and colleagues (2018) attempted to find highly specific and sensitive miRNA biomarkers that can be used to detect different stages of liver fibrosis. The trial entailed 42 cases of CHC with early-stage fibrosis, 45 cases of CHC with late-stage fibrosis, and 40 healthy subjects with no CHC or fibrosis as controls. Expression patterns of 5 miRNAs (miR-16, miR-146a, miR-214-5p, miR-221, and miR-222) were analyzed in each group using TaqMan real-time PCR. Serum levels of miRNA-16, miRNA-146a, miRNA-221, and miRNA-222 were all significantly up-regulated in early and late stages of liver fibrosis; miRNA-222 had the highest sensitivity and specificity values in early and late fibrosis; miRNA-221 had the 2nd highest sensitivity and specificity with the late-stage fibrosis group. Furthermore, miRNA-221 showed significant positive correlations with both miRNA-16 and miRNA-146a in the early- and late-stage fibrosis groups, with the early stage having a stronger correlation. The authors concluded that these findings indicated that miRNA-16, miRNA-146a, miRNA-221, and miRNA-222 can be used to detect the presence of liver fibrosis. They stated that the high sensitivity and specificity of miRNA-222 and miRNA-221 in late-stage fibrosis indicated promising prognostic biomarkers for HCV-induced liver fibrosis.

Ultrasound Elastography for Screening Portal Hypertension-Related Complications in Children

In a systematic review and meta-analysis, Kim and colleagues (2019) examined the diagnostic performance of US elastography in evaluating portal hypertension in children and compared the liver and spleen stiffness values between the portal hypertension and control groups. Studies in the Medline and Embase databases were selected that investigated the diagnostic performance of US elastography in children with portal hypertension up to December 21, 2017. Pooled sensitivity and specificity data were assessed by hierarchical logistic regression modeling. A total of 11 studies were included in the systematic review, and a meta-analysis could be conducted in 7 of these publications to evaluate the diagnostic performance of US elastography. The summary sensitivity and specificity of this method for liver stiffness were 90 % (95 % CI: 83 % to 94 %) and 79 % (95 % CI: 73 % to 84 %), respectively, and the AUROC was 0.92 (95 % CI: 0.90 to 0.94). A subgroup analysis of 5 TE studies revealed similar diagnostic performance (sensitivity, 90 %; specificity, 78 %). In 10 of the 11 studies that examined liver stiffness and 2 of the 3 studies that also measured spleen stiffness, patients in the portal hypertension group had a significantly higher stiffness value than the control group (p < 0.05). The authors concluded that US elastography exhibited good performance in diagnosing portal hypertension and could identify significant differences in liver and spleen stiffness in children with this condition. These researchers stated that this method thus has considerable potential as a non-invasive tool for screening portal hypertension-related complications in children with chronic liver disease.

Transient Elastography for Follow-Up of Primary Sclerosing Cholangitis

Corpechot et al (2014) noted that PSC is a chronic cholestatic disease that results in extensive liver fibrosis and cirrhosis, which are associated with poor outcome; however, there are no validated non-invasive markers of liver fibrosis in patients with PSC. In a prospective study, these investigators examined the diagnostic performance, reproducibility, longitudinal changes, and prognostic value of LSMs using VCTE. They analyzed percutaneous liver biopsy specimens from 73 consecutive patients with PSC from January 2005 to December 2010. Patients underwent VCTE no more than 6 months after the biopsy specimens were collected. The biopsy specimens were analyzed by a pathologist blinded to the results of VCTE for the stage of fibrosis, and LSM was associated with the stage of fibrosis and other variables using the Kruskal-Wallis and Spearman correlation tests. The cut-off values of LSM were selected based on the accuracy with which they identified the stage of fibrosis on ROC analysis. The rates of LSM progression were evaluated using a linear mixed model, and the association between LSM values and clinical outcomes were examined using Cox regression analysis in 168 patients with PSC treated with ursodeoxycholic acid (UDCA) and followed-up from November 2004 to July 2013 (mean follow-up period of 4 years). LSM was independently linked to the stage of fibrosis. Cut-off values for fibrosis stages ≥ F1, ≥ F2, ≥ F3, and F4 were 7.4 kPa, 8.6 kPa, 9.6 kPa, and 14.4 kPa, respectively. The adjusted diagnostic accuracy values for severe fibrosis and cirrhosis were 0.83 and 0.88, respectively. The diagnostic performance of LSM was comparable to that of hyaluronic acid (HA) measurement but superior to the aspartate aminotransferase/platelet ratio index, FIB-4 score, and Mayo risk score in differentiating patients with significant or severe fibrosis from those without. LSM had a high level of reproducibility between operators for the same measurement site and for the same operator between 2 adjacent sites. LSM increased significantly and exponentially over time. Baseline measurements and rate of LSM progression were strongly and independently linked with patients’ outcomes. The authors concluded that VCTE was able to differentiate severe from non-severe liver fibrosis with high levels of confidence in patients with PSC. These investigators stated that baseline measurements of LSM and longitudinal changes are prognostic factors for PSC.

Ehlken et al (2016) stated that patients with PSC develop progressive liver fibrosis and ESLD; non-invasive and widely available parameters are urgently needed to evaluate disease stage and the risk of clinical progression. Transient elastography has been reported to predict fibrosis stage and disease progression; however, these findings have not been confirmed in an independent cohort and comparison of TE measurement to other non-invasive means is missing. In a retrospective study, these researchers collected data from consecutive PSC patients receiving TE measurements from 2006 to 2014 (n = 139). Data from 62 patients who also underwent a liver biopsy were used to examine the performance of TE and spleen length (SL) measurement for the staging of liver fibrosis. Follow-up data from this cohort (n = 130, Hamburg) and another independent cohort (n = 80, Paris) was used to compare TE and SL as predictors of clinical outcome applying Harrel’s C calculations. TE measurement had a very good performance for the diagnosis and exclusion of higher fibrosis stages (≥ F3: AUROC 0.95) and an excellent performance for the diagnosis and exclusion of cirrhosis (F4 versus < F4: AUROC 0.98). Single-point TE measurement had very similar predictive power for patient outcome as previously published. In a combined cohort of PSC patients (n = 210), SL measurements had a similar performance as TE for the prediction of patient outcome (5 x cross-validated Harrel’s C 0.76 and 0.72 for SL and TE, respectively). The authors concluded that baseline TE measurement exhibited an excellent performance for the diagnosis of higher fibrosis stages in PSC. These investigators noted that baseline measurements of SL and TE had similar usefulness as predictive markers for disease progression in patients with PSC.

Cazzagon et al (2019) noted that magnetic resonance (MR) risk scores and LS have individually been shown to predict clinical outcomes in PSC. In a retrospective study, these investigators examined their complementary prognostic value. Patients with PSC from 3 European centers with a 3-dimensional MR cholangiography available for central reviewing and a valid LSM assessed by VCTE by FibroScan performed within a 6-month interval were included in this longitudinal trial. The MR score (Anali) without gadolinium (Gd) was calculated according to the formula: (1 × dilatation of intrahepatic bile ducts) + (2 × dysmorphy) + (1 × portal hypertension). The primary endpoint was survival without LT or cirrhosis decompensation. The prognostic values of LS and Anali score without Gd were assessed using Cox proportional hazard models. A total of 162 patients were included in this study. Over a total follow-up of 753 patient-years, 40 patients experienced an adverse outcome (4 LTs, 6 liver-related deaths, and 30 cirrhosis decompensations). LS and Anali score without Gd were significantly correlated (ρ = 0.51, p < 0.001) and were independently associated with the occurrence of an adverse outcome. Optimal prognostic thresholds were 10.5 kPa for LS and 2 for the Anali score without Gd. Hazard ratios (HR; 95 % CI) were 2.07 (1.06 to 4.06) and 3.78 (1.67 to 8.59), respectively. The use in combination of these 2 thresholds allowed these researchers to separate patients into low-, medium-, and high-risk groups for developing adverse outcomes. The 5-year cumulative rates of adverse outcome in these 3 groups were 8 %, 16 %, and 38 % (p < 0.001), respectively. The authors concluded that the combined use of MRI and ACTE allowed easy risk stratification of patients with PSC.

Mazhar and Russo (2021) noted that several prognostic tests for PSC have been developed, including biochemical models, elastography and MRI scores. These investigators carried out a systematic review of non-invasive prognostic tests for PSC. It was performed from 1987 to 2020 of blood tests, biochemical models, elastography and imaging scores associated with outcomes in PSC. A total of 40 studies of prognostic tests that collectively included 16,094 subjects with PSC were reviewed, of which 26 studies of non-invasive tests including 13,759 subjects with PSC were included. Normalization or reduction of alkaline phosphatase with or without therapy was associated with transplant-free survival and reduced risk of hepato-biliary cancers but cut-off values for alkaline phosphatase were inconsistent among studies. The most studied prognostic biochemical model was the Mayo Risk Score (MRS) evaluated in 18 studies with a c-statistic from 0.63 to 0.85 for clinical outcomes. One study showed that the UK-PSC score outperforms MRS for predicting clinical outcomes with a c-statistics of 0.81and 0.75, respectively. A TE score of greater than 11.1 kPa was associated with survival and liver-related complications. The Anali score, derived from specific MRI and MRCP features, was associated with the development of cholangiocarcinoma and decompensated cirrhosis. Promising prognostic models include the enhanced liver fibrosis (ELF) score, ELF test and PREsTo scores. The authors concluded that MRS was the most studied prognostic score for clinical outcomes in PSC; however, the UK-PSC score and PREsTo had better test performance.

The European Association for the Study of the Liver’s clinical practice guidelines on “Sclerosing cholangitis” (EASL, 2022) stated that “A consensus process initiated by the International PSC Study Group (IPSCSG) resulted in a short list of 5 candidates for measuring disease progression: ALP, VCTE, histology, combination of ALP + histology, and bilirubin. The 2015 EASL CPG on non-invasive tests stated that noninvasive assessment of fibrosis using VCTE should be considered in people with PSC (grade B2), acknowledging that cholestasis related to untreated dominant biliary strictures influences liver stiffness assessment. The 2021 EASL CPG on noninvasive tests confirmed this recommendation and added that the ELF score could also be used for risk stratification both at baseline and during follow-up”.

Furthermore, an UpToDate review on “Primary sclerosing cholangitis in adults: Clinical manifestations and diagnosis” (Kowdley, 2022) states that “Transient elastography measures liver stiffness and is a technique for noninvasively assessing hepatic fibrosis. Studies suggest that it can be used to estimate the degree of hepatic fibrosis in patients with cholestatic liver disease, including PSC”.

Artificial Intelligence (AI) in Non-Alcoholic Fatty Liver Disease (NAFLD)

Li et al (2022) stated that NAFLD is one of the most important causes of chronic liver disease in the world; it has been found that cardiovascular and renal risks and diseases are also highly prevalent in adults with NAFLD. Evaluation of the severity of NAFLD entails a variety of clinical parameters, how to optimize non-invasive evaluation methods is a necessary issue that needs to be discussed in this field. Artificial intelligence (AI) has become increasingly widespread in healthcare applications, and it has been also brought many new insights into better analyzing chronic liver disease, including NAFLD. These investigators reviewed AI-related investigations in NAFLD field published recently, summarized diagnostic models based on electronic health record and laboratory test, US and radio imaging, and liver histopathological data. Furthermore, these researchers also analyzed present AI models in distinguishing healthy versus NAFLD/NASH, and fibrosis versus non-fibrosis in the evaluation of NAFLD progression. They hope to provide alternative directions for the future research.

These researchers stated that the ambiguity of AI made it problematic for machine learning (ML) systems to be adopted in a sensitive yet critical domains, such as healthcare. As a result, scientific interest in the field of Explainable AI (XAI), a field that is concerned with the development of new methods that explain and interpret ML models, has been tremendously re-ignited over recent years. However, AI provides a new way for disease understanding by extracting the characters of complex data and combining them with the mode of “automatic learning”, which will contribute to an increase in diagnostic quality, facilitate the development of remote medicine, and reduce the costs in the national healthcare. From this point of view, it may have a large enough potential to induce a paradigm shift in the handling of NAFLD. Certainly, ML itself is far from fulfilling its potential in NAFLD research, and researchers have a long way to go to uncover the networked intricacies and complexities of living systems. These investigators noted that accumulation of subjective and objective data as well as long-term follow-up verification are still the most basic, individual factors that need to be considered in the application of AI model. In addition, XAI for NAFLD studies are also needed to be examined.

Su et al (2023) noted that the reported prevalence of NAFLD in studies of lean individuals ranged from 7.6 % to 19.3 %. These researchers developed ML models for the prediction of fatty liver disease in lean individuals. This retrospective study included 12,191 lean subjects with a BMI of less than 23 kg/m2 who had undergone a health checkup from January 2009 to January 2019. Participants were divided into a training (70 %, 8,533 subjects) and a testing group (30 %, 3,568 subjects). A total of 27 clinical features were analyzed, except for medical history and history of alcohol or tobacco consumption. Among the 12,191 lean individuals included in the present study, 741 (6.1 %) had fatty liver. The ML model comprising a 2-class neural network using 10 features had the highest AUROC value (0.885) among all other algorithms. When applied to the testing group, these investigators found the 2-class neural network exhibited a slightly higher AUROC value for predicting fatty liver (0.868, 0.841 to 0.894) compared to the FLI (0.852, 0.824 to 0.81). The authors concluded that the 2-class neural network had greater predictive value for fatty liver than the FLI in lean individuals. Moreover, these researchers stated that further studies with larger sample sizes using other forms of clinical information and image are needed to validate the use of novel ML models in predicting steatosis and fibrosis.

The authors stated that this study had several drawbacks. First, these researchers did not collect data on hepatitis B and C virus status or history of alcohol consumption or diabetes. This trial comprised subjects with fatty liver on US which may have included patients with NAFLD and alcoholic fatty disease. Although patients with viral hepatitis and alcohol liver disease may have been included in the present study, further studies are needed to determine the predictive ability of ML models in detecting steatosis in populations with a single liver disease etiology. Metabolic-associated fatty liver disease (MAFLD) is not a suitable representation of this study subjects, given that these participants did not satisfy the diagnostic criteria for MAFLD. Second, some clinical parameters were not included in the present study, such as uric acid, hemoglobin A1c (HbA1c), C-reactive protein (CRP) and homeostasis model assessment for insulin resistance (HOMA-IR). These parameters have been demonstrated to be associated with lean NAFLD. Third, these investigators did not collect histological data from liver biopsies or other non-invasive imaging techniques, such as the controlled attenuation parameter measured by FibroScan or MRI-derived proton density fat fraction to confirm the degree of the steatosis. The use of US may have resulted in the under-diagnosis of hepatic steatosis in these patients relative to that using histology. Accordingly, the prevalence of fatty liver may have been under-estimated in the present study. This would suggest that the ML model cannot be used to predict mild steatosis in the general population.

Weng et al (2023) stated that fatty liver disease (FLD) is an important risk factor for liver cancer and cardiovascular disease and can lead to significant social and economic burden. However, there is currently no nationwide epidemiological survey for FLD in China, making early FLD screening crucial for the Chinese population. Unfortunately, liver biopsy and abdominal US, the preferred methods for FLD diagnosis, are not practical for primary medical institutions. These researchers developed ML models for screening individuals at high risk of FLD; and provided a new perspective on early FLD diagnosis. This study included a total of 30,574 individuals between the ages of 18 and 70 years who completed abdominal US and the related clinical examinations. Among them, 3,474 individuals were diagnosed with FLD by abdominal US. These investigators employed 11 indicators to build 8 classification models to predict FLD. The model prediction ability was examined by the AUC, sensitivity, specificity, PPV, NPV, and kappa value. Feature importance analysis was assessed by Shapley value or root mean square error loss after permutations. Among the 8 ML models, the prediction accuracy of the extreme gradient boosting (XGBoost) model was highest at 89.77 %. By feature importance analysis, these researchers found that the BMI, triglyceride, and ALT played important roles in FLD prediction. The authors concluded that ML models, especially XGBoost, can predict FLD via demographic indicators, blood glucose, liver function test, and blood lipid profile with high accuracy and good repeatability. Among 11 predictive indicators, BMI, ALT, and TTG are vitally important for most models. With the aid of these ML methods, physicians can now assess a patient’s fatty liver condition in its incipient stages, even in the absence of liver biopsy or imaging evidence. This early assessment can facilitate timely diagnosis and provide a novel avenue for the early screening of fatty liver in primary healthcare facilities. These researchers stated that in future research, they will collect more data from various populations to further modify the model and give it better generalization ability.

The authors stated that this study had several drawbacks. First, although more than 30,000 samples were included in this study, these samples were all from the Second Xiangya Hospital of Central South University, and the population representation was less comprehensive than that of multi-center clinical studies. The generalization ability of the model in different ethnic groups is open to question. Second, the response variables used in the ML model constructed in this study were solely based on the diagnostic results of abdominal US, which have a lower level of evidence compared to liver biopsy and MRI. This may potentially affect the accuracy of the predictions. Third, tumor, hepatitis, and other metabolic diseases (such as diabetes, hyperthyroidism, etc.) were not excluded from the population included in this study. As a result, the potential impact of these factors on the predictive model could not be fully assessed. Fourth, it should be noted that these researchers did not gather data on alcohol consumption and medication history among the study population; thus, they were unable to rule out the potential interference caused by alcohol and drugs. Previous studies have indicated that both factors could influence the development of fatty liver. To improve their model, future studies should gather more detailed information on alcohol intake and medication history.

Serum Ferritin as a Biomarker for Detecting or Monitoring Hepatic Fibrosis

Wang et al (2022) stated that NAFLD has become the most common liver disorder globally, and non-invasive evaluation approaches are in need to evaluate NAFLD disease progression. Serum ferritin has been proposed as one of the biomarkers for NAFLD diagnosis in previous studies. In a systematic review, these investigators examined the association of serum ferritin level with the various stages of NAFLD in the elderly. A total of 3 databases (Medline, Embase, and Scopus) were systematically searched to obtain potentially relevant publications before July 2022. No restrictions were employed to geographical region, study design, publication type and language. The association between serum ferritin level or different ferritin categories and the various stages of NAFLD was the primary outcome of interest. Title and abstract screenings, data extraction and coding, and quality assessment were independently completed by 2 authors with discrepancies resolved through discussion with a 3rd author. A total of 32 studies were included and heterogeneity was considerable. The associations between serum ferritin level and the stages of hepatic steatosis, fibrosis, inflammation and ballooning and the occurrence of NASH were examined; however, inconsistent associations were reported. Most studies identified serum ferritin to be a predictor of advanced NAFLD, while several revealed the opposite. The authors concluded that serum ferritin could be considered to act as a non-invasive biomarker for evaluating various stages of NAFLD. Moreover, these researchers stated that further studies are needed to confirm its predictive value since this study reported inconsistent associations based on the qualitative synthesis.

The authors stated that this review had 2 main drawbacks. First, meta-analysis was not carried out due to the heterogeneity of NAFLD grading standards and statistical analysis methods among the included studies. Moreover, the absence of necessary data in some included studies for conducting meta-analysis prevented these investigators from further analysis. Second, 5 studies were not incorporated in this study since these researchers did not have access to their full texts, which might have resulted in a certain level of bias.

Transient Elastography for Follow-Up of Liver Transplant Recipients

Nacif et al (2018) noted that TE is a non-invasive technique that measures liver stiffness. When an inflammatory process is present, this is shown by elevated levels of stiffness. Acute cellular rejection (ACR) is a consequence of an inflammatory response directed at endothelial and bile epithelial cells, and it is diagnosed via liver biopsy. In a systematic review, these investigators examined the viability of TE in ACR following liver transplantation (LTx). The Cochrane Library, Embase, and Medline PubMed databases were searched and updated to November 2016. The MESH terms used were “Liver Transplantation,” “Graft Rejection,” “Elasticity Imaging Techniques” (PubMed), and “Elastography” (Cochrane and Embase). A total of 70 studies were retrieved and selected using the PICO (patient, intervention, comparison or control, outcome) criteria; 3 prospective studies were selected to meta-analysis and evaluation. A total of 33 patients with ACR were assessed with TE. One study showed a cut-off point of greater than 7.9 kPa to define graft damage and less than 5.3 kPa to exclude graft damage (ROC of 0.93; p < 0.001). Another study reported elevated levels of liver stiffness in ACR patients; however, in that study, no cut-off point for ACR was suggested. The final prospective study included 27 patients with ACR at liver biopsy. Cut-off points were defined as TE of greater than 8.5 kPa, moderate-to-severe ACR, with a specificity of 100 % and ROC curve of 0.924. The measurement of TE of less than 4.2 kPa excluded the possibility of any ACR (p = 0.02). The authors concluded that TE may be an important tool for the severity of ACR in patients following LTx. Moreover, these researchers stated that further investigations are needed to better define the cut-off points and applicability of the examination.

The authors stated that a drawback of this study was that these researchers did not find RCTs, and the selections of the studies were presented in the general post-transplant context (which needed to exclude other causes such as hepatitis, cholangitis, and autoimmune or cholestatic recurrence, virus recurrence, thrombosis), and only 1 study examined the specific topic of ACR. Thus, these investigators knew that more controlled, multi-center studies are needed on the topic. The real benefit of this meta-analysis was to show that researchers are moving toward better evaluation of these patients with less invasive methods.

Nacif et al (2020) there are few studies using elastography in ACR, which is one of the main complications following LTx. The golden pattern diagnostic is by liver biopsy, which is invasive and subject to complications. These investigators examined the use of elastography in ACR; they carried out prospective and comparative study of patients transplanted from January 2017 to March 2019. Comparison group (ACR versus non-ACR) via liver biopsy. The variables analyzed were liver elastography (FibroScan and ARFI), laboratory tests, liver biopsy, and US. Mann-Whitney U test was employed to compare independent samples, and p < 0.05 was considered significant. All tests performed with α of 0.05 and a confidence interval of 95 %, by IBM SPSS 25 software. A total of 40 patients, 25 (62.5 %) with ACR and 15 (37.5 %) without ACR; 5 (20 %) cases with early acute rejection, late acute rejection in 19 cases (76 %), and chronic rejection in 3 (12 %). Comparative ACR versus non-ACR showed results of total bilirubin (p = 0.03), direct bilirubin (p = 0.015), aspartate aminotransferase (0.001), alanine aminotransaminase (0.001), and gamma-glutamyl transferase (p = 0.026). The mean elastography (FibroScan) value in ACR was 12.5 ± 8.2 kPa and without was 8.9 ± 3.7 kPa (p = 0.05). The mean elastography (ARFI) in ACR was 1.9 ± 0.6 m/s and without was 1.6 ± 0.2 m/s (p > 0.05). The ROC curve analysis shows the FibroScan for ACR with AUC 0.688 (95 % CI: 0.511 to 0.865 ; p = 0.049), PPV of 0.76, and NPV of 0.60. The authors concluded that FibroScan (TE) proved to be a good tool for ACR; however, this method still needs to be further studied to determine cut-off values to grade the intensity of the ACR. Furthermore, this examination should be studied to find a way to differentiate ACR from other graft diseases.

The authors noted that this study’s limitation pointed to the small number of studies found. Although the findings demonstrated that elastography could assess rejection, these researchers were unable to define cut-off numbers to determine if rejection is mild, moderate, or severe, as they did not have an adequate number of patients to separate them into these groups. Another limitation of was that these investigators could not guarantee that this liver examination can replace liver biopsy because this method still cannot distinguish between a rejection and a relapse of the underlying disease that led to the transplant. Furthermore, vascular and biliary changes may compromise the elastography results.

Furthermore, UpToDate reviews on “Liver transplantation in adults: Long-term management of transplant recipients” (Gaglio and Cotler, 2023), and “Liver transplantation in adults: Clinical manifestations and diagnosis of acute T-cell mediated (cellular) rejection of the liver allograft” (Reddy, 2023) do not mention transient elastography as a management option.

FIB-4 Index for Assessing Risk in Persons with Risk Factors for NAFLD

A guideline on the diagnosis and management of NAFLD was published in 2022 by the AACE in collaboration with the AASLD recommends the FIB-4 index (a nonproprietary algorithm that incorporates AST, ALT, platelets and the patient’s age) as the preferred method of screening persons with risk factors for non-alcoholic fatty liver disease (NAFLD) and cirrhosis (Cusi, et al., 2022). The AACE guidelines stated that “A combination of the FIB-4 followed by VCTE seems to be the best approach” to further characterize persons whose FIB-4 index places them in an indeterminate or high-risk group. Similarly, an AGA care pathway recommended the FIB-4 test as the preferred test for fibrosis screening in high risk persons (Kanwal, et al., 2021).

Transient Elastography (Fibroscan) for Monitoring of Liver Function in Wilson’s Disease

Sini et al (2012) stated that liver biopsy has always represented the standard of reference in hepatic fibrosis assessment. Recently, blood markers and instrumental methods have been proposed for non-invasive assessment. These investigators validated TE and other non-invasive tests compared to liver histology in Wilson’s Disease (WD). Liver stiffness in 35 patients with WD was evaluated by Fibroscan, serum fibrosis markers (AST-to-platelet-ratio index and FIB-4), and liver biopsy. Compared to liver histology, the FibroScan values increased proportionally with progression of the histological fibrosis stage. Significant fibrosis could be predicted with a Fibroscan cut-off value of 6.6 kPa. Advanced fibrosis could be predicted with a FibroScan cut-off value of 8.4 kPa. Serum fibrosis marker values gave good correlation with hepatic stage. The authors concluded that a FibroScan value of 6.6 kPa was found to be a significant separation limit for differentiating significant fibrosis stages from milder stages and a Fibroscan value of 8.4 kPa was found to be a significant separation limit for differentiating advanced fibrosis stages from milder stages. These researchers stated that FibroScan values were clinically useful for predicting fibrosis stages and helpful in managing chronic therapy in patients with WD.

Poujois and Woimant (2018) noted that WD is characterized by a deleterious accumulation of copper in the liver and brain. It is one of those rare genetic disorders that benefits from effective and life-long treatments that have dramatically transformed the prognosis of the disease. In Europe, its clinical prevalence is estimated at between 1.2 and 2/100,000; however, the genetic prevalence is higher, at around 1/7,000. Incomplete penetrance of the gene or the presence of modifier genes may account for the difference between the calculated genetic prevalence and the number of patients diagnosed with WD. The clinical spectrum of WD is broader as expected with mild clinical presentations and late onset of the disease after the age of 40 years in 6 % of patients. WD is usually suspected when ceruloplasmin and serum copper levels are low and 24-hour urinary copper excretion is elevated. Recently, a major diagnostic advance was achieved with implementation of the direct assay of “free copper”, or exchangeable copper (CuEXC). The relative exchangeable copper (REC) that corresponds to the ratio between CuEXC and total serum copper enables a diagnosis of WD with high sensitivity and specificity when REC is greater than 18.5 %. Moreover, CuEXC values at diagnosis are a marker of extra-hepatic involvement and its severity. A value of greater than 2.08 μmol/L is suggestive of corneal and brain involvement (sensitivity = 86 %, specificity = 94 %), and the disease will be more clinically and radiologically severe as values rise. The use of FibroScan is becoming more wide-spread for evaluating liver stiffness measurements in WD patients; 6.6 kPa is considered to be a threshold value between mild and moderate fibrosis, whereas a value higher than 8.4 is indicative of severe fibrosis. These investigators stated that more studies are needed to confirm the usefulness of Fibroscan in managing chronic therapy for WD patients. Treatment of this disease is based on an initial active and prolonged chelating phase (with d-penicillamine or trientine) followed by maintenance with trientine or zinc salt. The 2 major problems that may be encountered are neurological worsening during the initial phase and non-compliance with treatment during maintenance therapy. Liver transplantation is the recommended therapeutic option in WD with acute liver failure or end-stage liver cirrhosis; its indication should be considered when neurological status deteriorates rapidly despite effective chelation. Regular clinical, biological and liver US follow-up is essential to examine effectiveness, tolerance, and treatment compliance, but also to detect the onset of HCC on a cirrhotic liver.

Yavuz et al (2022) examined parenchymal changes in the liver and pancreas related to copper accumulation using US in pediatric patients with WD, and examined the effectiveness of 2D shear wave elastography in the diagnosis of involvement of these organs. Patients with WD (n = 25) who were treated and followed at the authors’ center were evaluated prospectively. In addition to routine clinical assessments, eye examination, laboratory analyses, as well as abdominal US imaging, all patients underwent tissue stiffness measurements from the liver and pancreas (head, body, and tail) by 2D shear wave elastography. The data obtained from the WD patients were compared with those of age- and sex-matched healthy controls (n = 37). Liver elastography measurements showed significantly increased tissue stiffness in the patient group than in control subjects (p < 0.001). While there was no significant difference between the groups in the tissue thickness of pancreatic head, body, and tail, tissue stiffness was significantly reduced in the patient group (p < 0.001). Disease duration was significantly associated and moderately correlated with liver tissue stiffness (r = 0.417, p = 0.038) but not significantly associated with pancreatic tissue stiffness. The authors concluded that in the early stages of WD, parenchymal changes occur in the liver and pancreas, which could not be detected by conventional US imaging, but may be revealed by 2D shear wave elastography. Ultrasound elastography was an easy-to-use, non-invasive, and promising method that provided numerical data on the early changes in tissue stiffness, allowing for objective monitoring of WD patients who require life-long follow-up.

The 2022 practice guidance on “Wilson disease” from the American Association for the Study of Liver Diseases (Schilsky et al, 2022) stated that “Along with primary treatment of WD, patients with advanced liver disease must be treated for any complications of portal hypertension. Elastography may be informative in hepatic WD … Progression of hepatic fibrosis despite therapy also constitutes treatment failure, due to failure of pharmacotherapy or independent liver injury from an unrelated process. Monitoring can be by elastography or with use of serum‐based assays such as Fibrosis‐4 (Fib‐4) Index or AST/platelet ratio. The gold standard remains liver biopsy”.

Nehring et al (2023) stated that staging of liver fibrosis is of special significance in WD as it determines the patient’s prognosis and treatment. Histopathological examination is a standard method for fibrosis assessment; however, non-invasive methods like TE and SWE are believed to be reliable and repetitive, and are expected to replace liver biopsy in WD.

Guillaud et al (2023) noted that WD and alpha1-antitrypsin deficiency are 2 rare genetic diseases that may impact predominantly the liver and/or the brain, and the liver and/or the lung, respectively. The early diagnosis of these diseases is important in order to initiate a specific treatment, when available, ideally before irreversible organ damage, but also to initiate family screening. These investigators focused on the non-invasive diagnostic tests available for clinicians in both diseases. These tests are crucial at diagnosis to reduce the potential diagnostic delay and evaluate organ involvement. They also play an important role during follow-up to monitor disease progression and assess treatment effectiveness of current or emerging therapies.

Janczyk et al (2023) stated that autoantibodies occur in healthy subjects as well as in children with WD; however, their prevalence and significance are unknown. These investigators examined the prevalence of autoantibodies and autoimmune markers, and their relationship to liver injury in WD children. This trial included 74 children with WD, and 75 healthy children as a control group. Patients with WD underwent TE examinations, as well as determination of liver function tests, copper metabolism markers, and serum immunoglobulins (Ig). In the sera of the WD patients and controls, anti-nuclear (ANA), anti-smooth muscle, anti-mitochondrial, anti-parietal cell, anti-liver/kidney microsomal, anti-neutrophil cytoplasmic autoantibodies, and specific celiac antibodies were determined. Among the autoantibodies, only the prevalence of ANA in children with WD was higher than in the controls. There was no significant relationship between the presence of autoantibodies and liver steatosis or stiffness after TE. However, advanced liver stiffness (E > 8.2 kPa) was related to IgA, IgG, and gamma globulin production. The type of treatment did not influence the prevalence of autoantibodies. The authors concluded that these findings suggested that autoimmune disturbances in WD might not be directly related to liver damage as expressed by steatosis and/or liver stiffness after TE.

Cave et al (2024) noted that WD with acute onset poses a diagnostic challenge because it is clinically indistinguishable from other acute liver diseases. Furthermore, serum ceruloplasmin and urinary copper excretion, the 1st-line diagnostic tools for WD, can show false positive results in the case of acute liver failure, and the diagnostic role of genetic analysis is limited by the time needed to perform it. In the case of fulminant onset, there is a clear indication of LTx. “New Wilson Index” is often used to discriminate between patients who need LTx versus those who can be successfully managed by medical treatment; however, its reliability remains controversial. Timely referral of patients with acute liver failure due to WD may be a key factor in improving patient survival. Although LTx very often represents the only chance for such patients, maximum effort should be made to promote survival with a native liver. Moreover, these investigators stated that for diagnostic purposes, liver biopsy in children is only required if a definite diagnosis of WD is not achieved with non-invasive tests, or if further liver disorders are suspected. In cases where there is coagulopathy, however, the use of liver biopsy is usually precluded.

The OWLiver Panel

The OWLiver Panel (CIMA Sciences, LLC) is a non-invasive diagnostic tool for evaluating all lesional phases of MAFLD (metabolic dysfunction-associated fatty liver disease). This lipidomic analysis of fasting blood samples employs high-resolution liquid chromatography coupled with mass spectrometry (UHPLC-MS) to measure a panel of lipid biomarkers (28 metabolites) reflecting liver fat content, inflammation, and fibrosis. The OWLiver Panel’s 3 algorithms includes lipid concentrations, and clinical data, providing an estimation of MAFLD stage.

Bril et al (2018) examined the use of existing metabolomics scores to classify liver disease in patients with T2DM. A total of 220 patients with T2DM were recruited. Subjects underwent routine laboratory tests, liver proton magnetic resonance spectroscopy (1 H-MRS), a 75-g oral glucose tolerance test, and liver biopsy if 1 H-MRS findings indicated NAFLD. A serum sample was blindly provided to OWL Metabolomics on which to run the OWLiver Care and OWLiver tests. When compared with liver biopsy, the OWLiver Care and OWLiver tests had a suboptimal performance in patients with T2DM (AUROC curve both less than 0.70). Given the discordance of these findings in this heterogeneous, multi-ethnic cohort compared with those of a previous report in predominantly white patients without diabetes, these researchers examined the influence of age, ethnicity, and other variables on test performance. A specific subset of patients was selected to mirror the characteristics of the population used for the development of this model (i.e., white patients without T2DM). Among white patients with good glycemic control (glycated hemoglobin of less than 53 mmol/mol [or less than 7 %]) and without cirrhosis, the AUROC curve was significantly improved (0.79, 95 % CI: 0.68 to 0.90]). Among white patients with lower insulin resistance (homeostatic model assessment of insulin resistance of less than 3) and without cirrhosis, the AUROC was even higher: 0.87 (95 % CI: 0.76 to 0.97). The authors concluded that there was a great need to develop non-invasive approaches to diagnose non-alcoholic steatohepatitis in patients with T2DM; models originally developed for patients without diabetes could not be directly applied to patients with T2DM.

Bril et al (2020) noted that the 2019 Standards of Medical Care in Diabetes suggested that patients with NAFLD should be examined for liver fibrosis; however, the performance of non-invasive clinical models/scores and plasma biomarkers for the diagnosis of NASH and advanced fibrosis has not been carefully evaluated in patients with T2DM. In a cross-sectional study, patients (n = 213) had a liver MRS, and those with a diagnosis of NAFLD underwent a percutaneous liver biopsy. Several non-invasive clinical models/scores and plasma biomarkers were measured to identify NASH and advanced fibrosis (NASH: ALT, cytokeratin-18, NashTest 2, HAIR, BARD, and OWLiver; advanced fibrosis: AST, fragments of pro-peptide of type III procollagen [PRO-C3], FIB-4, APRI, NAFLD fibrosis score, and FibroTest). None of the non-invasive tools assessed for the diagnosis of NASH in patients with T2DM had an optimum performance (all AUCs of less than 0.80). Of note, none of the panels or biomarkers was able to out-perform plasma ALT (AUC 0.78, 95 % CI: 0.71 to 0.84). Performance was better to diagnose advanced fibrosis, in which plasma PRO-C3, AST, and APRI showed better results than the other approaches (AUC 0.90, 95 % CI: 0.85 to 0.95; 0.85, 95 5 CI: 0.80 to 0.91; and 0.86, 95 % CI: 0.80 to 0.91, respectively). Again, none of the approaches did significantly better than plasma AST. Sequential use of plasma AST and other non-invasive tests may aid in limiting the number of liver biopsies needed to identify patients with advanced fibrosis. The authors concluded that performance of non-invasive clinical models/scores and plasma biomarkers for the diagnosis of NASH or advanced fibrosis was suboptimal in patients with T2DM. Combination of multiple tests may provide an alternative to minimize the need for liver biopsies to detect fibrosis in these patients.

Perez-Diaz-Del-Campo et al (2021) stated that NAFLD affects 25 % of the world’s population. The pathogenesis of NAFLD is complex; available data showed that genetics and ascribed interactions with environmental factors may play an important role in the development of this condition. These investigators examined genetic and non-genetic determinants putatively involved in the onset and progression of NAFLD after a 6-month weight loss nutritional treatment. A group of 86 over-weight/obese subjects with NAFLD from the Fatty Liver in Obesity (FLiO) study were enrolled and metabolically evaluated at baseline and after 6 months. A pre-designed panel of 95 genetic variants related to obesity and weight loss was applied and analyzed. Three genetic risk scores (GRS) concerning the improvement on hepatic health evaluated by minimally invasive methods such as the fatty liver index (FLI) (GRSFLI), lipidomic-OWLiver-test (GRSOWL) and MRI (GRSMRI), were derived by adding the risk alleles genotypes. Body composition, liver injury-related markers, and dietary intake were also monitored. A total of 23 SNPs were independently associated with the change in FLI, 16 SNPs with OWLiver-test, and 8 SNPs with MRI, which were specific for every diagnosis tool. After adjusting for gender, age, and other related predictors (insulin resistance, inflammatory biomarkers and dietary intake at baseline) the calculated GRSFLI, GRSOWL and GRSMRI were major contributors of the improvement in hepatic status. Therefore, fitted linear regression models showed a variance of 53 % (adj. R2 = 0.53) in hepatic functionality (FLI), 16 % (adj. R2 = 0.16) in lipidomic metabolism (OWLiver-test), and 34 % (adj. R2 = 0.34) in liver fat content (MRI). The authors concluded that these findings showed that 3 different genetic scores could be useful for the personalized management of NAFLD, whose treatment must rely on specific dietary recommendations guided by the measurement of specific genetic biomarkers. Moreover, these researchers stated that this trial was designed as a proof-of-concept in order to examine if the genetic background linked to NAFLD-related factors may influence hepatic amelioration. Furthermore, examining new causes of disease and the underlying mechanism or alteration in specific pathways and clinical outcomes may be of interest.

The authors stated that this study had several drawbacks. First, liver biopsy results were not available to corroborate the precise diagnosis of patients. However, in this trial, these investigators conducted a complete evaluation of the liver status by means of validated and widely used techniques as well as blood biomarkers and hepatic indexes, which are affordable and practical methods to use in health assessment. Second, the sample size and the enrollment of subjects were not very large. For this reason, these models may be further validated in different populations to establish whether they might represent a reliable and accurate, “non-invasive alternative” to liver biopsy. Furthermore, the role of new SNPs associated with excessive adiposity and accompanying metabolic alterations via a GRS approach needs to be re-examined. Third, type I and type II errors could not be completely ruled out, especially those related to the selection of SNPs to be introduced into the GRS. However, due to the use of less stringent p-value thresholds compared to association studies of single variants, genomic profile risk scoring analyses could tolerate, at balance, some of these biases, as previously reviewed. Fourth, dietary intake was assessed using self-reported information of the subjects, which may have produced some bias on the evaluation of these findings. Fifth, the constructions of the GRS using specific obesity-related SNP was also an important limitation.

Sanyal et al (2023) noted that there are no approved diagnostic biomarkers for at-risk NASH, defined by the presence of NASH, high histological activity and fibrosis stage of 2 or greater, which is associated with higher incidence of liver-related events and mortality. FNIH-NIMBLE is a multi-stakeholder project to support regulatory approval of NASH-related biomarkers. The diagnostic performance of 5 blood-based panels was examined in an observational (NASH CRN DB2) cohort (n = 1,073) with full spectrum of NAFLD. The panels were intended to diagnose at-risk NASH (NIS4), presence of NASH (OWLiver), or fibrosis stages greater than 2, greater than 3 or 4 (enhanced liver fibrosis (ELF) test, PROC3 and FibroMeter VCTE). The pre-specified performance metric was an AUROC of 0.7 or higher, and superiority over ALT for disease activity, and the FIB-4 test for fibrosis severity. Multiple biomarkers met these metrics. NIS4 had an AUROC of 0.81 (95 % CI: 0.78 to 0.84) for at-risk NASH. The AUROCs of the ELF test, PROC3 and FibroMeterVCTE for clinically significant fibrosis (stage 2 or greater), advanced fibrosis (stage 3 or greater) or cirrhosis (stage 4), respectively, were all 0.8 or higher. ELF and FibroMeter VCTE out-performed FIB-4 for all fibrosis endpoints. The authors concluded that multiple biomarker panels met the pre-specified criteria described in the letter of intent for biomarker qualification by the FDA in stage 1 of the circulating work-stream of the NIMBLE project of the FNIH. These findings informed the development of the full qualification package for these biomarkers for diagnostic enrichment in the next stage of the NIMBLE project.

The authors stated that this study had several drawbacks. First, the NASH Clinical Research Network (CRN) was based at tertiary care centers, generating ascertainment bias. The study population was also predominantly White ethnicity; thus, the data were not generalizable to other ethnicities. The curated patient population to ensure a balanced distribution of fibrosis stages to rigorously define sensitivity and specificity did not allow evaluation of the predictive values in populations with variable distribution of disease phenotypes. This will be performed in the final qualification step, and this trial set the stage for the evaluation of these diagnostic cut-offs to be validated in future studies. Second, new biomarker (e.g., FAST, Agile and ADAPT) were not studied in the pre-determined qualification panel. These were, however, not developed at the time when this trial was conceived, and they were undergoing rigorous evaluation and would be reported as post-hoc analyses separately. Third, although the study population was specifically curated to have a relatively even distribution of fibrosis stages to avoid spectrum bias, real-world populations do not have such a distribution, and the PPV and NPV of the tests in populations with varying prevalence may require separate confirmation. Moreover, these researchers stated that it must be noted that the journey from discovery and initial validation of a biomarker to a diagnostic tool that is approved for use by all clinicians is a long one, and entails many steps that could not be combined in 1 study.

Navarro-Masip et al (2024) stated that NAFLD, now termed MAFLD, is a growing health concern associated with obesity and T2DM. Bariatric surgery offers potential benefits; however, its impact on MAFLD remains incompletely understood, with scarce long-term follow-up prospective studies. Furthermore, since liver biopsy is the gold standard for liver condition measurement, the need for non-invasive techniques that allow the evaluation of MAFLD development following bariatric surgery is important. OWLiver Care and OWLiver represent 2 serum lipidomic tests, featuring panels comprising 11 and 20 triglycerides, respectively. These researchers carried out a prospective study entailing 80 Caucasians to examine the effects of bariatric surgery on MAFLD using non-invasive diagnostics and to identify baseline predictors of MAFLD remission. Serum samples were collected before surgery and at a 3-year follow-up. After 3 years, the proportion of patients exhibiting a healthy liver escalated from 5.0 % at baseline to 26.3 %. Conversely, the percentage of steatohepatitis declined from 35.1 % to a mere 7.6 %. Younger age, female gender, and the absence of T2DM were associated with MAFLD remission. However, age stood as the only independent variable associated with this favorable liver evolution (R2 = 0.112). The authors concluded that bariatric surgery showed mid-term benefits in improving MAFLD, with younger age as a baseline predictor of remission. Non-invasive diagnostic methods, such as OWLiver, are valuable tools for monitoring MAFLD evolution. Moreover, these investigators stated that further investigations with larger populations and longer follow-up periods are needed to refine personalized treatment approaches.

Qadri and Yki-Jarvinen (2024) noted that fatty liver plays an important role in the pathogenesis of the metabolic syndrome and T2DM. According to an updated classification, any individual with liver steatosis and 1 or more features of the metabolic syndrome, without excess alcohol consumption or other known causes of steatosis, has MASLD. Up to 60 % to 70 % of all individuals with T2DM have MASLD. However, the prevalence of advanced liver fibrosis in patients with T2DM remains uncertain, with reported estimates of 10 % to 20 % relying on imaging tests and likely over-estimating the true prevalence. All stages of MASLD impact prognosis; however, fibrosis is the best predictor of all-cause and liver-related mortality risk. Patients with T2DM face a 2- to 3-fold increase in the risk of liver-related death and HCC, with 1.3 % progressing to severe liver disease over 7.7 years. Because reliable methods for detecting steatosis are lacking, MASLD mostly remains an incidental finding on imaging. Regardless, several medical societies advocate for universal screening of individuals with T2DM for advanced fibrosis. Proposed screening pathways entail annual calculation of the Fibrosis-4 (FIB-4) index, followed by a secondary test such as TE for intermediate-to-high-risk individuals. However, owing to unsatisfactory biomarker specificity, these pathways are expected to channel approximately 40 % of all individuals with T2DM to TE, and 20 % to tertiary care, with a false discovery rate of up to 80 %, raising concerns regarding feasibility. Therefore, there is a need to develop more effective strategies for surveying the liver in patients with T2DM. Nonetheless, weight loss via lifestyle changes, pharmacotherapy, or bariatric surgery remains the cornerstone of management, proving highly effective not only for metabolic co-morbidities but also for MASLD. The authors concluded that emerging evidence suggests that fibrosis biomarkers may serve as tools for risk-based targeting of weight-loss interventions and potentially for monitoring response to therapy. Moreover, these researchers stated that while many unanswered questions remain as to how patients should be best identified and managed, embracing strategies that are evidence-based and that prevent over-burdening the healthcare system could garner wider acceptance among both healthcare providers and patients.

NASHnext

NASHnext is a non-invasive blood test that reflects both NASH activity and fibrosis in a single score by combining the results of 4 individual NASH-associated biomarkers (serum alpha2-macroglobulin, serum CH13L1/YKl40, serum miR34a, and whole-blood HbA1c). The intended use of this test is for predicting likelihood of at-risk NASH. (NASH is now referred to as metabolic dysfunction-associated steatohepatitis (MASH)). However, there is insufficient evidence to support the use of NASHnext for diagnosis of non-alcoholic steatohepatitis and liver fibrosis.

Harrison et al (2020) stated that non-invasive tests that can identify patients with NASH at higher risk of disease progression are lacking. In a prospective derivation and global validation study, these researchers reported the development and validation of a blood-based diagnostic test to non-invasively rule in and rule out at-risk NASH (defined as non-alcoholic fatty liver disease [NAFLD] activity score [NAS] of 4 or more and fibrosis stage of 2 or more). Blood samples, clinical data, and liver biopsy results from 3 independent cohorts with suspected NAFLD were used to develop and validate a non-invasive blood-based diagnostic test, called NIS4. Derivation was carried out in the discovery cohort, which comprised 239 prospectively recruited patients with biopsy-confirmed NASH (NAFLD NAS 3 or greater; fibrosis stage 0 to 3) from the international GOLDEN-505 phase-IIb clinical trial. A complete matrix based on 23 variables selected for univariate association with the presence of at-risk NASH and avoiding high multi-collinearity was used to derive the model in a bootstrap-based process that minimized the Akaike information criterion. The overall diagnostic performance of NIS4 was externally validated in 2 independent cohorts: RESOLVE-IT diag and Angers. The RESOLVE-IT diag cohort comprised the first 475 patients screened for potential inclusion into the RESOLVE-IT phase-III clinical trial. Angers was a retrospective cohort of 227 prospectively recruited patients with suspected NAFLD and clinical risk factors for NASH or fibrosis stage 2 or more according to abnormal elastography results or abnormal liver biochemistry. Both external validation cohorts were independently analyzed and were combined into a pooled validation cohort (n = 702) to assess clinical performance of NIS4 and other non-invasive tests. The derived NIS4 algorithm comprised 4 independent NASH-associated biomarkers (miR-34a-5p, alpha-2 macroglobulin, YKL-40, and glycated hemoglobin; AUROC of 0.80, 95 % CI: 0.73 to 0.85), and did not require adjustment for age, sex, BMI, or AST concentrations. Clinical cut-offs were established within the discovery cohort to optimize both rule out and rule in clinical performance while minimizing indeterminate results. NIS4 was validated in the RESOLVE-IT diag cohort (AUROC 0.83, 95 % CI: 0.79 to 0.86) and the Angers cohort (0.76, 0.69 to 0.82). In the pooled validation cohort, patients with a NIS4 value of less than 0.36 were classified as not having at-risk NASH (ruled out) with 81.5 % (95 % CI: 76.9 to 85.3) sensitivity, 63.0 % (57.8 to 68.0) specificity, and a NPV of 77.9 % (72.5 to 82.4), whereas those with a NIS4 value of more than 0.63 were classified as having at-risk NASH (ruled in) with 87.1 % (83.1 to 90.3) specificity, 50.7 % (45.3 to 56.1) sensitivity, and a PPV of 79.2% (73.1 to 84.2). The diagnostic performance of NIS4 within the external validation cohorts was not influenced by age, sex, BMI, or AST concentrations. The authors concluded that NIS4 is a novel blood-based diagnostic that provided an effective way to non-invasively rule in or rule out at-risk NASH in patients with metabolic risk factors and suspected disease. Moreover, these researchers stated that the use of NIS4 in clinical trials or in the clinic has the potential to greatly reduce unnecessary liver biopsies in patients with lower risk of disease progression.

The authors stated that the main drawback of this trial was the lack of patients with cirrhosis in the discovery cohort and only a few of these patients (25 [4 %] of 702) in the pooled validation cohort. It was encouraging that most patients with fibrosis stage 4 were appropriately categorized with NIS4, and if VCTE had been used for all patients with indeterminate or high NIS4 scores, these investigators believed almost all patients with cirrhosis would be identified. Unfortunately, clinics have patients with NASH and cirrhosis who have not been diagnosed with either condition yet. All patients with cirrhosis require specific management regardless of the etiology of their cirrhosis, such as screening for HCC, esophagogastroduodenoscopy to screen for varices, and pneumococcal vaccination. The performance of NIS4 to identify patients who need additional testing, such as VCTE, to further identify cirrhosis, needs to be examined in future studies. A small number of patients had advanced fibrosis (fibrosis stage of 3 or more) with a low NAS; therefore, not meeting the criteria for at-risk NASH; alternative diagnostics, such as VCTE, will need to be evaluated for the identification of these patients, since NIS4 was not designed to detect fibrosis without a NASH component. Another drawback was that ELF and VCTE data were only available in a subpopulation of patients in the pooled validation cohort and precluded further analyses.

In a commentary on the afore-mentioned study by Harrison et al (2020), Choy (2021) noted that the authors established that NIS4 is not influenced by patients’ age, sex, BMI, or AST concentrations; however, other potential confounding factors associated with these individual biomarkers did not appear to have been considered. Choy also stated that in the European multi-center assessment of clinical course and biomarkers in severe chronic airway disease (BIOAIR) Trial, serum YKL-40 was significantly increased in individuals with asthma and those with chronic obstructive pulmonary disease (COPD) compared with healthy controls; thus , these underlying respiratory conditions could have been a potential confounding factor. Furthermore, a decrease in blood HbA1c could be caused by increased erythropoiesis (e.g., chronic liver disease), increased erythrocyte destruction (e.g., hemolytic anemia), or abnormal hemoglobin (e.g., hemoglobinopathies). As a consequence, HbA1c assessment might not be appropriate in individuals with these conditions; the possibility of these medical conditions invalidating the HbA1c result should be noted. Additionally, underlying hemoglobin abnormalities are more frequently observed in some ethnic communities; thus, ethnicity should be considered when HbA1c concentration is measured in clinical trials or in the clinic.

In another commentary on the afore-mentioned study by Harrison et al (2020), Zhou et al (2021) stated that the sequential combination of non-invasive tests designed for liver fibrosis detection accurately classifies more than 90 % of patients with advanced NAFLD fibrosis. In a similar way, Zhou et al proposed that the sequential combination of NIS-4, MACK-3, and FAST scores might aid in narrowing the grey zone and further avoid unnecessary liver biopsies. The FAST score relies heavily on serum AST concentrations and thus it accurately classifies patients with advanced fibrosis, but could also place individuals with normal serum liver enzymes in the grey zone. In such instances, Zhou et al proposed that additional testing with NIS4 or MACK-3 might better identify patients who are at risk of NASH, as these 2 blood-based diagnostic tests include in their equations other important parameters associated with NASH. Zhou et al stated that sequential combination of these 3 newly developed non-invasive tests to better identify patients who are at risk of developing NASH warrants further research.

References

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