Autoantibodies are serum markers of autoimmune liver disease (mainly including both autoimmune hepatitis and primary biliary cirrhosis). Anti-nuclear antibodies (ANA), anti-smooth muscle antibodies (SMA), and anti-liver and kidney microsomes type I antibodies (anti-LKM1) are the hallmark antibodies for autoimmune hepatitis (AIH), and anti-mitochondrial antibodies (AMA) are the hallmark antibodies for primary biliary cirrhosis (PBC). However, these antibody titer measurements are not associated with disease activity and do not have the ability to predict treatment outcome. Predictive serum markers could improve the current treatment strategy for autoimmune liver disease through early diagnosis, establishment of individualized treatment regimens, and guidance of withdrawal timing.
1. Predictive markers in autoimmune hepatitis
(1) antibodies to Soluble Liver Antigen (anti-SLA)
anti-SLA liver-specific antibodies, also known as anti-hepatopancreatic antigen (LP) antibodies. Detection methods include radioligand assays, enzyme-linked immunosorbent assays (ELISA), protein blotting, and immunoprecipitation reactions, and the clinical and epidemiological characterization may vary depending on the results obtained by the different methods, with protein immunoblotting having a higher specificity and sensitivity than ELISA for the detection of anti-SLA.
Anti-SLA can be present in ANA, SMA, and anti-LKM1-negative patients, defined as a specific group of AIH types (previously referred to as type III AIH). anti-SLA is associated with the genetic gene DRB1*0301, and anti-SLA-positive type I AIH patients have a higher expression of HLA DR3 compared to ANA and SMA-positive individuals (79 Anti-SLA-positive patients were characterized by severe liver tissue damage, the need for longer-term maintenance therapy, and high rates of reaggravation after withdrawal, liver failure, and liver transplantation.
(2) Anti-actin antibodies (anti-actin)
Anti-actin is a subclass of anti-smooth muscle antibody (SMA). Detection methods include indirect immunofluorescence, convective immunoelectrophoresis, ELISA and actin-myosin functional assay. The predictive value of the different assays differs.
Indirect immunofluorescence assays for anti-actin-positive pediatric patients with AIH are characterized by drug therapy dependence, susceptibility to progression to liver failure, and high demand for liver transplantation.
The rate of mortality from liver failure and the need for liver transplantation was higher in anti-actin-positive adults with early age of onset and poorer response to glucocorticoid therapy than in antinuclear-antibody-positive patients (19% vs. 0%, P=0.03). In addition, the presence of immune responses against both actin and α-coactin suggested that patients had severe clinical or histological disease.
(3) Anti-liver cytosol antigen type 1 (anti-LC1)
The target antigen of anti-LC1 is iminomethyltetrahydrofolate cyclization dehydrogenase (FTCD), an enzyme with dual function in folate metabolism, often present together with anti-LKM1, a marker of type II AIH, mainly found in pediatric and adolescent patients (age <20 years). Detection methods include indirect immunofluorescence, convective immunoelectrophoretic assays, immunoprecipitation reactions, protein immunoblotting, and ELISA.
Anti-LC1 positivity suggests the possible presence of other autoimmune diseases, severe hepatitis and progressive cirrhosis in AIH patients, and it can be the only serum marker in pediatric patients with acute, acute severe, or chronic hepatitis. anti-LC1 is important for the diagnosis of ANA, SMA, and anti-LKM1-negative patients, who respond better to glucocorticoid therapy. Anti-LC1 can be present in AIH patients along with SMA and ANA and can also be detected in 12% of patients with chronic hepatitis B.
(4) Anti-asialoglycoprotein receptor (anti-ASGPR)
Anti-ASGPR can be detected by ELISA and radioimmunofiltration method. Among them, the sensitivity and specificity of anti-ASGPR detected by ELISA using human desialoglycoprotein receptor as substrate is high, but its application is limited by the extraction of target antigens and the lack of supporting reagents. In recent years, convenient and effective ELISA reagents based on rabbit antigen have been developed for the diagnosis of AIH with a sensitivity of 78% and a specificity of 99%.
anti-ASGPR can be present in patients with AIH who are negative for conventional antibodies. anti-ASGPR expression is increased in the presence of apoptosis and is densely expressed in the characteristic histological region around the portal vein of the hepatic lobules, i.e., in interfacial hepatitis, suggesting its association with hepatocyte necrosis and interfacial hepatitis. anti-ASGPR-positive patients have higher serum gamma globulin and immunoglobulin G levels Anti-ASGPR correlates with the histological activity of the disease and can be measured by measuring the persistence of anti-ASGPR after glucocorticoid treatment to reflect the patient’s response to hormonal therapy. The disappearance of anti-ASGPR during treatment suggests remission after withdrawal, while the persistence or reappearance of anti-ASGPR suggests relapse of the disease, a feature that suggests the potential of anti-ASGPR for determining the timing of withdrawal.
(5) Anti-chromatin antibody (anti-bodies to chromatin, anti-chromatin)
The detection rate of anti-chromatin was higher in patients with active AIH than inactive patients (32% vs. 19%, P=0.01), and the relapse rate after withdrawal was also higher in positive patients (91% vs. 66%, P=0.01). 94% of the AIH cases positive for anti-chromatin also had ANA, but only 68% of the cases with ANA also had anti-chromatin. The specificity and positive predictive value of anti-chromatin in predicting relapse after withdrawal was 100%, but the sensitivity was only 42%, and, like other predictive markers, it had a negative predictive value for relapse after withdrawal. The negative predictive value of anti-chromatin for relapse after withdrawal was not high, only 22%, compared with 17% for anti-SLA and 22% for anti-actin. anti-chromatin positivity indicates inflammatory activity, severe disease and poor response to glucocorticoid therapy in AIH.
(6) Antibodies to Cyclic Citrullinated Peptide (anti-CCP)
Anti-CCP is a predictor of progressive rheumatoid arthritis (RA) and erosive joint disease, usually detected by ELISA, and is present in 9-11% of AIH patients, 75% of whom do not have RA. The incidence of cirrhosis was significantly higher in patients with anti-CCP-positive AIH (47% vs. 20%, P=0.01), with almost 100% of those with RA developing cirrhosis and a higher incidence of liver failure (25% vs. 9%, P=0.04). Most anti-CCP-positive AIH patients also had other autoantibodies, including anti-chromatin antibodies (60%), anti-actin antibodies (90%), anti-soluble liver antigen antibodies (27%), and rheumatoid factor (56%). Autoimmunity in a hyperreactive state can produce multiple autoantibodies through epitope expansion and molecular mimicry, further exacerbating immune disorders and leading to the development of aggressive hepatitis, and anti-CCP may serve as a marker for this process. Based on this hypothesis, it is hypothesized that anti-CCP may be one of the multiple expression products of the pathogenic process of immune disorders.
(7) Anti-liver kidney microsomal antibody 3 (anti-LKM3)
The anti-LKM3 target antigen is UDP-glucuronosyltransferase (UGT), which is currently the most highly regarded member of the family of antibodies against UGT and cytochrome P450. anti-LKM3 is present in 13% of patients with chronic hepatitis D, 8% of patients with type II AIH, and a minority of patients with hepatitis B. Unlike anti-LKM2, anti-LKM3 reflects the virus-mediated auto-activation of the microsomal enzyme system. unlike anti-LKM2, anti-LKM3 reflects a virus-mediated autoimmune response to the microsomal enzyme system and is not associated with drug toxicity. Although the predictive value of anti-LKM3 in type II AIH is unclear, its association with viral hepatitis and the classical enzyme system during peptide conversion may provide a perspective on the pathogenic mechanisms of autoimmune response, molecular mimicry, and disease exacerbation.
2. Predictive markers in primary biliary cirrhosis
(1) Anti-Sp100 antibody (anti-Sp100)
anti-Sp100 is also known as anti-MND (anti-multinuclear dot antibody). Its sensitivity is 27% and specificity is 94% for the diagnosis of PBC, and anti-Sp100 is more significant for the diagnosis of PBC in AMA-negative patients. anti-Sp100’s predictive value is far from its diagnostic value, it is mostly found in elderly patients with PBC and is associated with high serum gamma globulin concentration, and after 24 weeks of regular observation, it is found to be associated with the rapid progression of liver tissue lesions in PBC. It was found to be associated with rapid progression of PBC liver tissue lesions after 24 weeks of regular observation. T cells reacting to the mitochondrial pyruvate dehydrogenase E2 subunit were cross-reactive with Sp100 and gp210, suggesting that molecular mimicry can sensitize T cells to derive an autoimmune response. Molecular mimicry of mitochondrial and Sp100 antigens also includes bacterial antigens. Female patients with recurrent urinary tract infections without liver disease express both AMA and anti-Sp100, and the detection rate of anti-Sp100 is much higher in PBC patients with recurrent urinary tract infections than in those without recurrent urinary tract infections (74% vs 5%).
(2) Anti-gp210 antibody (anti-gp210)
anti-gp210 is a kind of anti-nuclear membrane antibody, which is present in 16%-25% of PBC patients. The application of ELISA assay based on recombinant gp210 fusion protein has been established and is important for the diagnosis of AMA-negative PBC patients, which can be present together with anti-Sp100. The target antigen is the nuclear pore protein, and the nuclear pore complex has the role of transporting nucleoplasm to the cytoplasm and maintaining cellular function. theoretically, damage to this site can directly affect cellular function and aggravate the disease, so anti-gp210 has a closer correlation with the severity and prognosis of PBC than anti-Sp100. anti-gp210 is associated with severe interface Anti-gp210 is associated with severe interfacial hepatitis and lobular inflammation, and positivity suggests a tendency to progress to liver failure or more severe cholestasis and hepatic impairment, with a poor prognosis. After treatment with ursodeoxycholic acid (UDCA) for positive anti-gp210 antibodies, the antibodies may turn negative for those who are effective, so the effect of UDCA on PBC can also be predicted by detecting changes in gp210 antibody expression during the course of the disease.
(3) Anti-centromere antibodies
The target antigen of anti-centromere antibody is 80kDa of centromere protein B, which is present in 9%-60% of PBC patients. The specificity of PBC diagnosis is not high because anti-adhesion antibodies can also be found in various rheumatic diseases, chronic hepatitis B and AIH. PBC patients with positive anti-adherent antibodies are more likely to progress to liver failure (48% vs. 26% after 8.9 years of follow-up), have higher alkaline phosphatase levels, more severe bile duct injury, and a higher incidence of portal hypertension compared to negative patients [24].
3. Limitations and perspectives of serum markers for disease prediction
The main limitations of serum markers for disease prediction are: (1) overinterpretation of markers in clinical applications. Autoantibodies are not necessarily the cause of disease and in many cases are not disease-specific. The use of these serum markers to determine disease severity and prognosis must be closely integrated with clinical practice, and their significance is mainly to raise ideological vigilance rather than to modify treatment protocols. ②The negative predictive value of serum markers is low. ③The concentration of antibodies in the serum does not necessarily correlate with the severity and prognosis of the disease, and the disappearance of markers does not mean that the disease is improving. ④Lack of standardized assays and the minimum concentration of markers to test positive have not been determined, and many assays can only be applied in research institutions.
Further elucidation of the pathogenesis of autoimmune liver disease to identify more advantageous serum markers may improve their negative and positive predictive values for disease severity and prognosis. Also, as the impact of genetic factors and individual differences on the clinical features of autoimmune liver disease are further elucidated, autoantibody analysis in combination with genetic and other factors will provide better prediction of disease severity and treatment response. In addition, because patients’ disease is influenced by a combination of genetics, age, the presence of other autoimmune diseases, treatment strategy, and disease stage, individual serum markers can only reflect one causative factor. Therefore, the combined detection and analysis of a series of related serum markers and the establishment of conversion methods for genetic, age, disease stage and other influencing factors can provide individualized disease prediction analysis for patients in the future.