Chronic liver diseases of differing etiologies are among the leading causes of mortality and morbidity worldwide. Establishing accurate staging of liver disease is very important for enabling both therapeutic decisions and prognostic evaluations. A liver biopsy is considered the gold standard for assessing the stage of hepatic fibrosis, but it has many limitations. During the last decade, several noninvasive markers for assessing the stage of hepatic fibrosis have been developed. Some have been well validated and are comparable to liver biopsy. This paper will focus on the various noninvasive biochemical markers used to stage liver fibrosis.
Chronic liver diseases of differing etiologies are among the leading causes of morbidity and mortality worldwide [
(1) The liver biopsy does not efficiently reflect the fibrotic changes occurring in the entire liver because an optimally sized biopsy contains 5–11 complete portal tracts and reflects only 1/50000 the volume of the liver. (2) The process of hepatic fibrosis is not linear, and biopsies from different areas have shown different stages of fibrosis. (3) Several reports have shown that cirrhosis may be missed in 10–30% of patients. (4) A liver biopsy cannot differentiate between early and advanced end-stage cirrhosis; thus, it cannot be used as an ideal prognostic predictor. (5) Disagreements between pathologists occur, which may correlate with the experience of the pathologist. (6) There is a risk of complications arising from liver biopsy, and they can vary from mild symptoms, such as mild abdominal pain, to severe hemorrhage and injury to the biliary system. (7) Due to the risk of complications, some patients may refuse liver biopsy. (8) In hospital observation for 4–6 hours is usually required after liver biopsy. Furthermore, the use of ultrasound or the development of complications increases the cost of treatment and may also prolong hospitalization [
NIMs are helpful in assessing the stage of fibrosis in patients with no clear indication for a liver biopsy, such as patients with chronic hepatitis B (CHB) and persistently normal serum alanine aminotransferase (ALT), patients with chronic hepatitis C (CHC) or CHB and who require follow-up assessment of the stage of fibrosis during or after treatment [
The ideal NIM for assessing hepatic fibrosis must be simple, readily available, reliable, inexpensive, safe, and well validated in different forms of chronic liver disease. It must also be useful in assessing the progression of liver disease [
The typical mechanism underlying the development of hepatic fibrosis is an imbalance between the deposition and removal of extracellular matrix (ECM). Hepatic stellate cells are the predominant producers of ECM, and their activation and proliferation are mediated by different cytokines during the process of liver injury [
NIBMs for liver fibrosis are grouped into two main categories: class 1 fibrosis markers, or direct biomarkers, and class 2 fibrosis markers, or indirect biomarkers [
Procollagen is a collagen precursor. It is cleaved by two different enzymes at its carboxy-terminal (type 1 (PC1CP)) and amino-terminal (type III (PCIIINP)), leading to the production of collagen. Mature collagen integrates into ECM [ The PCICP terminal peptide is major component of the connective tissue [ PCIIINP or PIIINP is another major component of connective tissue that has been extensively studied. Serum levels of PCIIINP reflect the stage of hepatic fibrosis [
YXL-40 chondrex is a member of the chitinase family, and it is involved in the remodeling and degradation of the ECM [
(1) Serum alanine aminotransferase (ALT) is one of the oldest markers used to assess liver disease [
(2) The aspartate aminotransferase (AST)/ALT (AAR) ratio is one of the eldest markers of liver fibrosis that is easily available and applicable. It has been validated in different forms of liver disease, [
(3) The AST/platelet ratio (APRI) was developed by Wai et al. in 2003 [
(4) The Forns index: this index was described by Forns et al. in 2002. It is calculated based on the age of the patient and three routine laboratory tests, namely, platelet count, cholesterol level, and
(5) The PGA index was proposed by Poynard et al. in 1977 as a marker to assess alcoholic liver disease. It is generated via a combination of GGT, the prothrombin index, and apolipoprotein A [
(6) Fibro test and Fibrosure: these tests are identical but are marketed under different names [
ACTI test: the Acti test is a modification of the Fibro test in which ALT values are added. It reflects both necroinflammatory activity and liver fibrosis [
(7) The Fibro index: this index was developed in 2007 by Koda et al. to assess hepatic fibrosis in CHC [
(8) The FIB-4 score: This score is calculated based on age, platelet count, AST, and ALT. It was first developed by Sterling et al. to assess fibrosis in HIV/HCV coinfected patients at a cutoff value of 3.25; 87% of patients were correctly classified, with an AUC of 0.765 for significant fibrosis [
(9) The FibroQ test: this test was proposed by Hsieh et al. in 2009. It is calculated based on age, AST, prothrombin time (PT-INR), platelet count, and ALT [
(10) Currently, with the increase in the incidence of metabolic syndromes, NAFLD is considered the most frequent cause of liver disease in the world [
(11) Steato test: this test was proposed by Poynard et al. to assess NAFLD. It incorporates the five components of the Fibro test (
(1) The Fibrometer test was described by Calès et al. in 2005. It is performed by combining the platelet count, prothrombin index, aspartate aminotransferase,
(2) Fibrospect II test combines three parameters: hyaluronic acid, TIMP-1, and
(3) SHASTA index is based on serum hyaluronic acid, AST, and albumin. In a study of 95 HIV/HCV coinfected patients, an index of 0.3 showed a sensitivity of >88% and a negative predictive value of >94%, and a level of 0.8 showed a specificity of 100% and a positive predictive value of 100% for detection of severe fibrosis of F3 or more [
(4) The Hepascore model was proposed by Adams et al. in 2005. It combines age, gender, serum bilirubin, GGT, hyaluronic acid, and
(5) European liver fibrosis panel (ELF) test was proposed by the ELF panel [
(1) 13C-methacetin breath test (MBT) and 13C-caffeine breath test (CBT) are tests that assess cytochrome P450-dependent hepatocellular function [
(2) Proteomics and glycomics: proteins and glycoproteins are assessed using mass spectrometry. Using serum samples [
(3) Kam et al. proposed the Fibro-Glyco index for assessing liver fibrosis; it is based on the N-glycome level determined using mass spectrometry. They reported a significant correlation between this index and the degrees of liver fibrosis
(4) King’s score: this score is the most recently proposed noninvasive index [
(5) Noninvasive hepatitis C-related cirrhosis early detection (NIHCED) index was suggested by Bejarano-Redondo et al. in 2009 for the detection of F2–F4 fibrosis. It is assessed based on age (≥60 years), prothrombin time (≥1.1), platelets (≤100,000), and AST/ALT (≥1). In addition to the presence of right hepatic lobe atrophy, caudate lobe hypertrophy is observed upon ultrasound examination. This test at score of more than 6 shows an accuracy of 72% and an AUC of 0.787 [
(6) Two Chinese models that involve different NIBMs have been suggested for assessing CHB [
(7) More recently, data on the use of surface-enhanced laser desorption/ionization time of flight mass spectrometry (SELDI-ATAOF-MS) in HCC identified a panel of serum proteins of value in differentiating HCC patients from those with cirrhosis or normal controls [
As mentioned above SHASTA index was used for HIV/CHC coinfected patients [
Several authors have attempted to combine NIMs to assess hepatic fibrosis, and they have suggested that these combinations improved sensitivity. In 2006, Sebastiani et al. proposed the SAFE algorithm (sequential algorithm for fibrosis evaluation) for use in CHC patients. In that study, 190 CHC patients were assessed using the APRI, Forns index, and Fibro test at the time of liver biopsy. The authors observed that the optimal combination was APRI followed by the Fibro test. Using this algorithm, advanced fibrosis and cirrhosis were diagnosed with accuracies >94% and 95%, respectively, and the requirement for liver biopsy was reduced by 60–70% [
Several studies were conducted that compared different algorithms that combined direct NIBMs. The majority of studies showed comparable results for different combinations of NIBMs. Furthermore, the combination algorithms showed significantly better performances compared with individual markers [
Several studies have compared the accuracies of different NIBMs for detecting advanced fibrosis and cirrhosis. In Bourliere’s study, the Fibro test and Hepascore showed similar diagnostic profiles for fibrosis of stages F2–F4 [
Stefanescu et al. recently validated the guidelines for the use of NIBMs in detecting large varices compared with endoscopy. They used the APRI, FIB-4, Forns index, and Lok score in addition to the Fibroscan. They concluded that a combination of the Lok score and the Fibroscan was optimal for detecting large varices [
Summary of NIBMs for assessment of different forms of liver disease (Tables
AUROC for the direct markers that have been used in assessment of fibrosis in various liver diseases.
Marker | Liver disease evaluated by the markers | AURCO for advanced fibrosis | AURCO for cirrhosis | References | |||
---|---|---|---|---|---|---|---|
CHC | CHB | NAFLD | ALD | ||||
PCICP | NA | — | — | NA | NA | NA | [ |
PCIIINP | 0.69–0.78 | — | NA | 0.67–0.867 | 0.67–0.867 | 0.734 | [ |
Type IV collagen | 0.73–0.83 | — | 0.82 | NA | 0.583–0.83 | NA | [ |
HA* | 0.821–0.92 | 0.98 | 0.97 | 0.69–0.93 | 0.69–0.98 | 0.85–0.93 | [ |
Laminin | 0.542–0.82 | — | NA | NA | 0.46–0.82 | NA | [ |
YXL-40 | 0.7–0.81 | NA | NA | 0.7–0.81 | 0.79 | [ | |
MMP-2 | 0.59 | 0.59 | 0.97 | [ | |||
MMP-3 | 0.88 | — | — | — | 0.88 | NA | [ |
MMP-9 | ** | — | — | — | NA | NA | [ |
TIMP-1 | 0.71–0.773 | — | — | — | 0.68–0.73 | 0.91 | [ |
TIMP-2 | 0.73 | — | — | — | 0.73 | NA | [ |
TGF- |
NA | — | — | — | — | — | [ |
TGF- |
NA | — | — | — | — | — | [ |
PDGF | NA | — | — | — | — | — | [ |
**Negative association, MMP-9 decreases with the progression of fibrosis.
NA: Area under receiver operating characteristic (AUROC) is not available.
AUROC for the indirect markers that have been used in assessment of fibrosis in various liver diseases.
Marker | Liver disease evaluated by various markers | AURCO for advanced fibrosis | AURCO for cirrhosis | References | |||
---|---|---|---|---|---|---|---|
CHC | CHB | NAFLD | ALD | ||||
ALT (2.25) the normal | 0.716–0.815 | 0.716–0.815 | — | [ | |||
AST/ALT ratio | 0.54–0.709 | NA | 0.742–0.83 | NA | 0.54–0.83 | 0.67 | [ |
APRI | 0.65–0.87 | 0.67–0.72 | 0.564–0.866 | — | 0.564–0.87 | 0.75–0.92 | [ |
Frons index | 0.78–0.86 | NA | — | — | 0.78–0.86 | [ | |
PGAA index | — | — | — | NA | — | — | [ |
Fibro test | 0.72–0.87 | 0.76–0.85 | 0.82–0.89 | 0.83–0.91 | 0.72–0.87 | 0.77–0.94 | [ |
Acti test | NA | 0.77 SH | [ | ||||
Fibro index | 0.804–0.83- | NA | — | — | 0.82 | 0.845 | [ |
Fib-4* | 0.785–0.86 | 0.81 | — | — | 0.785–0.86 | — | [ |
FibroQ | 0.789 | NA |
— | — | 0.783 | 0.791 | [ |
The simple test (NAFLD) fibrosis score | — | — | 0.82–0.89 | — | 0.82–0.89 | — | [ |
Steato test | — | — | 0.799–0.86 | 0.799–0.86 | — | [ | |
SELDI-TOF protein chip | 0.88 | 0.926 | — | — | 0.88–0.906 | 0.921 | [ |
13C metacetin breath test | 0.83- | — | — | — | 0.83 | 0.96 | [ |
NA: studied but AUROC is not available.
AUROC for liver fibrosis biomarkers that are a mix of direct and indirect markers.
Marker | Liver disease evaluated by various markers | AURCO for advanced fibrosis | AURCO for cirrhosis | References | |||
---|---|---|---|---|---|---|---|
CHC | CHB | NAFLD | ALD | ||||
The Fibrometer | 0.892** | 0.943 | 0.83–0.962 | 0.883–0.962 | 0.94 | [ | |
Fibrospect II | 0.77–0.831 | NA | — | 0.83 | 0.77–0.831 | [ | |
SHASTA index*** | 0.878 | — | — | — | 0.878 | — | [ |
Hepascore | 0.82 | 0.83 | 0.82–0.9 | 0.89–0.92 | [ | ||
ELF | 0.773 | 0.93–0.98 | 0.873 | 0.944 | 0.773–0.98 | [ |
|
— | — |
***HVC and HIV co-infected patients.
#1021 subjects recruited; the numbers in each diagnostic category were as follows: chronic hepatitis C, 496; ALD, 64; fatty liver, 61; hepatitis B, 61; primary biliary cirrhosis or primary sclerosing cholangitis, 53; recurrent disease after liver transplantation, 48; autoimmune hepatitis, 45; hemochromatosis, 32; cryptogenic cirrhosis, 19; hepatitis B and C, 4; 138 patients with other causes of liver disease like granuloma and abnormal liver enzymes from unknown cause.
NA: studied but AUROC is not available.
AUROC for performance of combination algorithms in assessing liver fibrosis.
Combination algorithm | Liver disease evaluated by various markers | AURCO for advanced fibrosis | AURCO for cirrhosis | References | |||
---|---|---|---|---|---|---|---|
CHC | CHB | NAFLD | ALD | ||||
SAFE biopsy | 0.89–1 | — | — | — | 0.97–1 | 0.87 | [ |
Stepwise SAFE algorithm + biopsy | — | NA | — | 0.96 | 0.95 | ||
Castera | 0.97 | — | — | — | 0.97 | 0.93 | [ |
Bourliere algorithms | NA | — | — | — | — | — | [ |
NA: studied but AUROC is not available.
NIBMs are advantageous compared with liver biopsies because of the following reasons. They are noninvasive and can be measured in outpatient departments. They cost less compared with liver biopsies. They can be easily repeated for confirmation. If they are well validated, they may be used for follow-up and monitoring in the future. They are not associated with the liver biopsy morbidity and mortality risks.
Limitations of NIBMs: Some of markers like APRI, Hepascore, and Fibrospect II need more validation in intermediate stages of liver fibrosis [
In spite that the effectiveness of NIBM in assessment of liver fibrosis was demonstrated by many studies some studies had shown that they may not be of diagnostic value in the detection of liver fibrosis [ They remain of limited value in assessing the development of complications, like esophageal varices and chance of variceal bleeding [ Both direct and indirect markers of liver fibrosis are not liver-specific and can be altered by pathological conditions in other organs. Some of the biomarkers lack standardization due to variable values and the different upper-normal ranges used by different laboratories. All studies that evaluated the accuracy of NIBMs used the liver biopsy as the gold standard reference; this protocol is also a limitation because even the best liver biopsy retains a risk of sampling error. A selection bias of the studied population may have biased the results; for example, if a larger number of patients with advanced or minimal fibrosis are included, this bias will affect the accuracy of the markers [ In a large population of patients with liver diseases, for example, patients with autoimmune liver disease, NIBMs remain poorly evaluated and validated. The majority of the direct markers that have been evaluated are not routinely available in all laboratories. Investigators must work to overcome the limitations of NIBMs for liver fibrosis. Several studies of marker combinations or stepwise algorithms have shown improved performance compared with the performance of individual markers [
NIBMs can be used to assess disease progression and to predict complications and survival of liver disease patients. NIBMs can be used to monitor treatment responses. Because of its complications, a liver biopsy cannot be used for the screening of high-risk groups, such as CHB patients with normal liver enzymes and obese or diabetic patients with expected NAFLD. Thus, NIBMs can be used to screen these patients. Proposing new combinations of direct and indirect markers may be a goal of future studies to avoid the limitations of each type of marker and to increase diagnostic accuracy. NIBMs for assessing liver fibrosis have not yet been validated in other less common liver diseases, such as AIH and PBC.
Considering the above-mentioned limitations and patients who fall in the gray areas using the noninvasive markers, liver biopsy is still required to diagnose some patients, for example, patients with viral hepatitis B or C and secondary diagnosis like AIH, NAFLD, or ALD [
Currently, a perfect NIBM for liver histology is unavailable. However, utilization of noninvasive biomarkers for liver histology can significantly reduce, but not completely replace, the requirement for liver biopsies in patients with chronic viral hepatitis and NAFLD. For the other types of liver disease, NIBMs are not well validated and more studies are required. Furthermore, future studies on the currently available NIBMs may reveal more important prognostic capabilities of these markers.
Noninvasive biomarkers
Nonalcoholic fatty liver disease
Alcoholic liver disease
Extracellular matrix
Area under receiver operating characteristic.
The author declares that there is no conflict of interests regarding the publication of this paper.