A Phase 3 Trial of Seladelpar in Primary Biliary Cholangitis.

BACKGROUND: Effective treatments for patients with primary biliary cholangitis are limited. Seladelpar, a peroxisome proliferator-activated receptor delta agonist, has potential benefits. METHODS: In this phase 3, 12-month, double-blind, placebo-controlled trial, we randomly assigned (in a 2:1 ratio) patients who had had an inadequate response to or who had a history of unacceptable side effects with ursodeoxycholic acid to receive oral seladelpar at a dose of 10 mg daily or placebo. The primary end point was a biochemical response, which was defined as an alkaline phosphatase level less than 1.67 times the upper limit of the normal range, with a decrease of 15% or more from baseline, and a normal total bilirubin level at month 12. Key secondary end points were normalization of the alkaline phosphatase level at month 12 and a change in the score on the pruritus numerical rating scale (range, 0 [no itch] to 10 [worst itch imaginable]) from baseline to month 6 among patients with a baseline score of at least 4 (indicating moderate-to-severe pruritus). RESULTS: Of the 193 patients who underwent randomization and treatment, 93.8% received ursodeoxycholic acid as standard-of-care background therapy. A greater percentage of the patients in the seladelpar group than in the placebo group had a biochemical response (61.7% vs. 20.0%; difference, 41.7 percentage points; 95% confidence interval [CI], 27.7 to 53.4, P<0.001). Normalization of the alkaline phosphatase level also occurred in a greater percentage of patients who received seladelpar than of those who received placebo (25.0% vs. 0%; difference, 25.0 percentage points; 95% CI, 18.3 to 33.2, P<0.001). Seladelpar resulted in a greater reduction in the score on the pruritus numerical rating scale than placebo (least-squares mean change from baseline, -3.2 vs. -1.7; least-squares mean difference, -1.5; 95% CI, -2.5 to -0.5, P = 0.005). Adverse events were reported in 86.7% of the patients in the seladelpar group and in 84.6% in the placebo group, and serious adverse events in 7.0% and 6.2%, respectively. CONCLUSIONS: In this trial involving patients with primary biliary cholangitis, the percentage of patients who had a biochemical response and alkaline phosphatase normalization was significantly greater with seladelpar than with placebo. Seladelpar also significantly reduced pruritus among patients who had moderate-to-severe pruritus at baseline. The incidence and severity of adverse events were similar in the two groups. (Funded by CymaBay Therapeutics; RESPONSE ClinicalTrials.gov number, NCT04620733; EudraCT number, 2020-004348-27.).

Circulating Monocyte Chemoattractant Protein-1 (MCP-1) in Patients with Primary Biliary Cholangitis.

Primary biliary cholangitis (PBC) is a chronic autoimmune liver disease that leads to the destruction of the intrahepatic bile ducts. While the inflammatory process can be mediated by monocyte chemotactic protein-1 (MCP-1), the importance of circulating MCP-1 as a biomarker is unclear. Our aim was to assess the diagnostic significance of the serum concentrations of MCP-1 in PBC patients. We compared circulating MCP-1 with biochemical, immunological and histological parameters. Serum samples were collected from 120 PBC patients, 60 pathologic controls and 30 healthy donors. MCP-1 levels were determined by using commercial enzyme-linked immunosorbent assay (ELISA). Elevated serum MCP-1 levels were detected in 66% of PBC patients with a specificity of 97%. Significantly higher levels of MCP-1 protein were found in the sera of patients with PBC than in the group of healthy individuals-410.2 pg/mL vs. 176.0 pg/mL, p < 0.01). Patients with higher concentrations of alkaline phosphatase also had higher levels of MCP-1 (r = 0.4, p < 0.01). In accordance with Ludwig's classification, a positive correlation of serum MCP-1 concentration with the degree of fibrosis was observed, OR = 6.1, p = 0.0003. We compared the MCP-1 with procollagen type III, hyaluronic acid (HA), FIB-4 index, APRI and collagen type IV when predicting the advance of liver fibrosis. Circulating MCP-1 is better correlated with liver fibrosis and is also associated with the occurrence of specific antimitochondrial autoantibodies and specific anti-nuclear autoantibodies-anti-gp210. MPC-1 can be considered to be a tool for diagnosing the degree of fibrosis in PBC, and combinations of MCP-1 and other specific biomarkers could support the diagnosis of PBC.

Efficacy and Safety of Elafibranor in Primary Biliary Cholangitis.

BACKGROUND: Primary biliary cholangitis is a rare, chronic cholestatic liver disease characterized by the destruction of interlobular bile ducts, leading to cholestasis and liver fibrosis. Whether elafibranor, an oral, dual peroxisome proliferator-activated receptor (PPAR) α and δ agonist, may have benefit as a treatment for primary biliary cholangitis is unknown. METHODS: In this multinational, phase 3, double-blind, placebo-controlled trial, we randomly assigned (in a 2:1 ratio) patients with primary biliary cholangitis who had had an inadequate response to or unacceptable side effects with ursodeoxycholic acid to receive once-daily elafibranor, at a dose of 80 mg, or placebo. The primary end point was a biochemical response (defined as an alkaline phosphatase level of <1.67 times the upper limit of the normal range, with a reduction of ≥15% from baseline, and normal total bilirubin levels) at week 52. Key secondary end points were normalization of the alkaline phosphatase level at week 52 and a change in pruritus intensity from baseline through week 52 and through week 24, as measured on the Worst Itch Numeric Rating Scale (WI-NRS; scores range from 0 [no itch] to 10 [worst itch imaginable]). RESULTS: A total of 161 patients underwent randomization. A biochemical response (the primary end point) was observed in 51% of the patients (55 of 108) who received elafibranor and in 4% (2 of 53) who received placebo, for a difference of 47 percentage points (95% confidence interval [CI], 32 to 57; P<0.001). The alkaline phosphatase level normalized in 15% of the patients in the elafibranor group and in none of the patients in the placebo group at week 52 (difference, 15 percentage points; 95% CI, 6 to 23; P = 0.002). Among patients who had moderate-to-severe pruritus (44 patients in the elafibranor group and 22 in the placebo group), the least-squares mean change from baseline through week 52 on the WI-NRS did not differ significantly between the groups (-1.93 vs. -1.15; difference, -0.78; 95% CI, -1.99 to 0.42; P = 0.20). Adverse events that occurred more frequently with elafibranor than with placebo included abdominal pain, diarrhea, nausea, and vomiting. CONCLUSIONS: Treatment with elafibranor resulted in significantly greater improvements in relevant biochemical indicators of cholestasis than placebo. (Funded by GENFIT and Ipsen; ELATIVE ClinicalTrials.gov number, NCT04526665.).

Protein-losing gastroenteropathy complicated with asymptomatic primary biliary cholangitis, refractory to immunosuppressant, and improved by Helicobacter pylori eradication: a case report.

BACKGROUND: Protein-losing gastroenteropathy (PLGE) is a syndrome with a chief complaint of hypoalbuminemia, which occurs due to plasma protein leakage in the gastrointestinal tract, leading to general edema, ascites, and pleural effusions. CASE PRESENTATION: A 71-year-old woman visited another hospital for evaluation of hypoalbuminemia and systemic edema. She was hospitalized for a close inspection of hypoalbuminemia and was diagnosed with PLGE. Steroid and azathioprine therapy was prescribed; however, hypoalbuminemia did not improve, and the patient's condition worsened due to anasarca. As hospitalization was prolonged, the patient was transferred to our hospital. She was infected with Helicobacter pylori, and we performed H. pylori eradication. Following H. pylori eradication, her edema improved remarkably. CONCLUSION: We present the first case wherein H. pylori eradication successfully improved protein leakage in the lower gastrointestinal tract in a patient diagnosed with PLGE complicated with refractory to immunosuppressant treatment. H. pylori eradication should be considered in patients with PLGE complicated with H. pylori infection, without specific endoscopic finding or refractory to immunosuppressants.

Characteristics of peripheral blood Gamma-glutamyl transferase in different liver diseases.

ABSTRACT: Gamma-glutamyl transferase (GGT) is a marker of oxidative stress and cholestasis. Because of its low specificity, clinicians usually ignore its diagnostic value.To compare and analyze the clinical features of GGT in primary biliary cholangitis (PBC), drug-induced liver injury (DILI), alcoholic liver disease (ALD), and non-alcoholic fatty liver disease (NAFLD) from the perspective of different causes instead of the severity of the disease.We observed the distribution characteristics and the rate of abnormality of GGT in the above 4 diseases. The relationship between GGT and alanine aminotransferase (ALT), aspartate transaminase (AST), alkaline phosphatase (ALP), total serum bilirubin, triglyceride (TG), total cholesterol (TC), low-density lipoprotein cholesterol, high-density lipoprotein cholesterol was analyzed using Spearman correlation.The highest level of GGT was up to 1000.00 to 2000.00 U/L in PBC and DILI, and the highest level of GGT was more than 2000.00 U/L in ALD, yet the difference was not statistically significant (P > .05). The highest level of GGT was only about 200.00 U/L in NAFLD and was the lowest in 4 liver diseases. Also, GGT was positively correlated with ALP, TC in PBC and DILI. Also, in ALD, GGT was positively correlated with ALT, AST, ALP, TG, and TC. In NAFLD, GGT was positively correlated with ALT, AST, and TG.The abnormal GGT in PBC and cholestasis DILI was associated with cholestasis; in ALD, it was associated with oxidative stress and cholestasis, and in NAFLD, it was associated with oxidative stress. GGT levels had different characteristics in different liver diseases, which were closely related to the pathogenesis of liver diseases.

Increased sensitivity of gp210 autoantibody detection using a newly designed gp210 antigen.

OBJECTIVES: The detection of autoantibody to glycoprotein 210 (gp210 Ab) against a 15 amino-acid peptide epitope by enzyme-linked immunosorbent assay (ELISA) has been widely used in the diagnosis of primary biliary cholangitis (PBC). However, this small peptide antigen presents spatial limitations for antibody access, which reduces the sensitivity of autoantibody detection. A recombinant gp210 antigen was constructed for increased sensitivity in antibody detection is described here. METHODS: The gp210 C terminal 18 amino acid coding sequence was ligated to the modified C-terminal 108 amino acid coding sequence of human serum albumin (mHSA108) and produced as a recombinant gp210 antigen mHSA108-gp210-C18. Measurements of gp210 Ab using the gp210 C-terminal 25 amino acid peptide (gp210-C25) and mHSA108-gp210-C18 by in-house ELISA were compared. ELISAs with mHSA108-gp210-C18 and commercial INOVA kit for gp210 Ab detection were also compared in PBC patients and healthy controls. The correlation between the two assays was analyzed and their efficiency in diagnosing was compared. RESULTS: Of 86 PBC samples, 35 (40.70%) and 44 (52.33%) positive samples were detected for anti-gp210 Ab using gp210-C25 and mHSA108-gp210-C18, respectively. Of 252 samples from PBC, 114 (45.24%) were positive for mHSA108-gp210-C18 ELISA whereas 94 (37.3%) for commercial ELISA (INOVA). All positive samples detected with commercial ELISA kit were also tested positive in mHSA108-gp210-C18 ELISA. Among 374 patients with other autoimmune diseases, anti-gp210 Ab were detected by mHSA108-gp210-C18 ELISA in 0.95% systemic lupus erythematosus (SLE) patients (2/210), 13.04% rheumatoid arthritis (RA) patients (13/97), and 1.47% of Sjögren's Syndrome (SS) patients (1/67). CONCLUSIONS: Compared to the gp210 peptide antigen, the sensitivity of the ELISA system using mHSA108-gp210-C18 antigen was improved. The novel gp210 antigen could be useful for screening patients known to be at increased risk of developing PBC.

Alterations in gut microbiota and elevated serum bilirubin in primary biliary cholangitis patients treated with ursodeoxycholic acid.

BACKGROUND: The prognosis of primary bile cholangitis (PBC) is linked to gut microbiota dysbiosis. This study investigated the association between the gut microbiome and elevated total bilirubin (TB) level in PBC patients treated with ursodeoxycholic acid (UCDA). METHODS: A total of 47 PBC patients with 12 months of UCDA treatment were enrolled. Patients were divided into the TB (+) (TB>1× upper limit of the normal range [ULN]; n = 20) and TB(-) (TB≤1× ULN; n = 27) groups. Stool and serum specimens were collected, and microbiota composition and functional characteristics in the 2 groups were evaluated by 16S RNA gene sequencing and bioinformatic analysis. RESULTS: Bacterial diversity was lower in the TB(+) group than in the TB(-) group, although there was no significant difference in bacterial community profile. The phylum Saccharibacteria showed differential abundance in the 2 groups. Meanwhile, the TB(-) group had lower abundance of the Gemmiger, Blautia, Anaerostipes and Coprococcus genera than the TB(+) group, whereas Holdemania was absent. The abundance of Gemmiger formicillis and Coprococcus eutactus was positively correlated with that of Faecalibacterium prausnitzii, while Blautia, Anaerostipes and Coprococcus were negatively correlated with total bile acid level. CONCLUSION: TB level in PBC patients treated for 12 months with UCDA is associated with a distinct gut microbiome profile.

Enhanced liver fibrosis score as a surrogate of liver-related complications and mortality in primary biliary cholangitis.

ABSTRACT: The presence of bridging fibrosis predicts survival of primary biliary cholangitis (PBC). This study aimed to compare serum parameters for the estimation of liver fibrosis and prediction of clinical outcomes in PBC.Out of 392 patients with PBC, 102 who underwent liver biopsy and in whom fibrosis indices, platelet count, hyaluronic acid, type IV collagen 7 second domain, procollagen type III amino-terminal peptide, tissue inhibitor of metalloproteinases 1, Mac-2 binding protein glycosylation isomer, N-terminal type III collagen propeptide levels; fibrosis index based on 4 factors, aspartate aminotransferase-to-platelet ratio index, and enhanced liver fibrosis (ELF) score were determined, were included. The correlation of histological stages based on both Scheuer and Nakanuma classifications with fibrosis indices was investigated. The Nakanuma system comprises grading for liver fibrosis and bile duct loss. Diagnostic performances of 10 fibrosis indices were evaluated to identify patients with poor prognosis. Moreover, correlations of those with PBC clinical manifestation and survival were also investigated.Enhances liver fibrosis (ELF) score had the highest correlation coefficient for liver fibrosis evaluated according to either the Scheuer or Nakanuma classification among 10 serum fibrosis indices. It also had the highest diagnostic performance in estimating Scheuer stage III and Nakanuma fibrosis score 2, both of which represent portal-bridging fibrosis. Patients with an ELF score of ≥10.0 had shorter survival and presented more frequently clinical complications than those with an ELF score of <10.0.ELF score determines the severity of liver fibrosis and predicts the occurrence of complications and survival in patients with PBC.

Adjunct Fenofibrate Up-regulates Bile Acid Glucuronidation and Improves Treatment Response For Patients With Cholestasis.

Accumulation of cytotoxic bile acids (BAs) during cholestasis can result in liver failure. Glucuronidation, a phase II metabolism pathway responsible for BA detoxification, is regulated by peroxisome proliferator-activated receptor alpha (PPARα). This study investigates the efficacy of adjunct fenofibrate therapy to up-regulate BA-glucuronidation and reduce serum BA toxicity during cholestasis. Adult patients with primary biliary cholangitis (PBC, n = 32) and primary sclerosing cholangitis (PSC, n = 23), who experienced an incomplete response while receiving ursodiol monotherapy (13-15 mg/kg/day), defined as serum alkaline phosphatase (ALP) ≥ 1.5 times the upper limit of normal, received additional fenofibrate (145-160 mg/day) as standard of care. Serum BA and BA-glucuronide concentrations were measured by liquid chromatography-mass spectrometry. Combination therapy with fenofibrate significantly decreased elevated serum ALP (-76%, P < 0.001), aspartate transaminase, alanine aminotransferase, bilirubin, total serum BAs (-54%), and increased serum BA-glucuronides (+2.1-fold, P < 0.01) versus ursodiol monotherapy. The major serum BA-glucuronides that were favorably altered following adjunct fenofibrate include hyodeoxycholic acid-6G (+3.7-fold, P < 0.01), hyocholic acid-6G (+2.6-fold, P < 0.05), chenodeoxycholic acid (CDCA)-3G (-36%), and lithocholic acid (LCA)-3G (-42%) versus ursodiol monotherapy. Fenofibrate also up-regulated the expression of uridine 5'-diphospho-glucuronosyltransferases and multidrug resistance-associated protein 3 messenger RNA in primary human hepatocytes. Pearson's correlation coefficients identified strong associations between serum ALP and metabolic ratios of CDCA-3G (r2  = 0.62, P < 0.0001), deoxycholic acid (DCA)-3G (r2  = 0.48, P < 0.0001), and LCA-3G (r2  = 0.40, P < 0.001), in ursodiol monotherapy versus control. Receiver operating characteristic analysis identified serum BA-glucuronides as measures of response to therapy. Conclusion: Fenofibrate favorably alters major serum BA-glucuronides, which correlate with reduced serum ALP levels and improved outcomes. A PPARα-mediated anti-cholestatic mechanism is involved in detoxifying serum BAs in patients with PBC and PSC who have an incomplete response on ursodiol monotherapy and receive adjunct fenofibrate. Serum BA-glucuronides may serve as a noninvasive measure of treatment response in PBC and PSC.

Diagnosis and Therapeutic Management of Liver Fibrosis by MicroRNA.

Remarkable progress has been made in the treatment and control of hepatitis B and C viral infections. However, fundamental treatments for diseases in which liver fibrosis is a key factor, such as cirrhosis, alcoholic/nonalcoholic steatohepatitis, autoimmune hepatitis, primary biliary cholangitis, and primary sclerosing cholangitis, are still under development and remain an unmet medical need. To solve this problem, it is essential to elucidate the pathogenesis of liver fibrosis in detail from a molecular and cellular perspective and to develop targeted therapeutic agents based on this information. Recently, microRNAs (miRNAs), functional RNAs of 22 nucleotides, have been shown to be involved in the pathogenesis of liver fibrosis. In addition, extracellular vesicles called "exosomes" have been attracting attention, and research is being conducted to establish noninvasive and extremely sensitive biomarkers using miRNAs in exosomes. In this review, we summarize miRNAs directly involved in liver fibrosis, miRNAs associated with diseases leading to liver fibrosis, and miRNAs related to complications of cirrhosis. We will also discuss the efficacy of each miRNA as a biomarker of liver fibrosis and pathology, and its potential application as a therapeutic agent.

Glycan biomarkers of autoimmunity and bile acid-associated alterations of the human glycome: Primary biliary cirrhosis and primary sclerosing cholangitis-specific glycans.

We have recently introduced multiple reaction monitoring (MRM) mass spectrometry as a novel tool for glycan biomarker research and discovery. Herein, we employ this technique to characterize the site-specific glycan alterations associated with primary biliary cirrhosis (PBC) and primary sclerosing cholangitis (PSC). Glycopeptides associated with disease severity were also identified. Multinomial regression modelling was employed to construct and validate multi-analyte diagnostic models capable of accurately distinguishing PBC, PSC, and healthy controls from one another (AUC = 0.93 ± 0.03). Finally, to investigate how disease-relevant environmental factors can influence glycosylation, we characterized the ability of bile acids known to be differentially expressed in PBC to alter glycosylation. We hypothesize that this could be a mechanism by which altered self-antigens are generated and become targets for immune attack. This work demonstrates the utility of the MRM method to identify diagnostic site-specific glycan classifiers capable of distinguishing even related autoimmune diseases from one another.

Inhibition of autotaxin by bile salts and bile salt-like molecules increases its expression by feedback regulation.

BACKGROUND: Autotaxin is an enzyme that converts lysophospholipid into lysophosphatidic acid (LPA), a highly potent signaling molecule through a range of LPA receptors. It is therefore important to investigate which factors play a role in regulating ATX expression. Since we have reported that ATX levels increase dramatically in patients with various forms of cholestasis, we embarked on a study to reveal factors that influence the enzyme activity ATX as well as its expression level in vitro and in vivo. METHODS: Bile from cholestatic patients was fractionated by HPLC and analyzed for modulation of ATX activity. ATX expression was measured in fibroblasts upon stimulation or inhibition of LPA signaling. RESULTS: Surprisingly, ATX activity was stimulated by most forms of its product LPA, but it was inhibited by bile salts and bile salt-like molecules, particularly by 3-OH sulfated bile salts and sulfated progesterone metabolites that are known to accumulate during chronic cholestasis and cholestasis of pregnancy, respectively. Activation of fibroblasts by LPA decreased ATX expression by 72%. Conversely, inhibition of LPA signaling increased ATX expression 3-fold, indicating strong feedback regulation by LPA signaling. In fibroblasts, we could verify that inhibition of ATX activity by bile salts induces its expression. Furthermore, induction of cholestasis in mice causes increased plasma ATX activity. CONCLUSIONS: Multiple biliary compounds that accumulate in the systemic circulation during cholestasis inhibit ATX activity and thereby increase ATX expression through feedback regulation. This mechanism may contribute to increased serum ATX activity in patients with cholestasis.

Changes of Serum IgG Glycosylation Patterns in Primary Biliary Cholangitis Patients.

Objective: Primary biliary cholangitis (PBC) is an autoimmune cholestatic liver disease whose diagnosis is based significantly on autoantibody detection. This study aims to investigate the glycosylation profile of serum IgG in PBC patients using high-throughput lectin microarrays technology. Method: Lectin microarray containing 56 lectins was used to detect and analyze the expression of serum IgG glycosylation in 99 PBC patients, 70 disease controls (DCs), and 38 healthy controls (HCs). Significant differences in PBC from control groups as well as across PBC subgroups positive for various autoantibodies were explored and verified by lectin blot technique. Results: Lectin microarray detection revealed that compared to DC and HC groups, the specific glycan level of serum IgG sialic acid in PBC patients was increased. For each PBC subgroup, glycan levels of IgG mannose and galactose were decreased in AMA-M2 positive PBC patients compared to the AMA-M2 negative group. IgG N-Acetylgalactosamine (GalNAc) and fucose were decreased in anti-sp100 positive patients. IgG galactose was increased in anti-gp210 positive patients. IgG mannose was decreased in ACA-positive patients. Although the difference in overall sialic acid level was not observed using lectin blot, all results among the above PBC subgroups were consistent with the results of the technique. Conclusion: Lectin microarray is an effective and reliable technique for analyzing glycan structure. PBC patients positive for different autoantibody exhibits distinct glycan profile. Altered levels of glycosylation may be related to the occurrence and development of the disease, which could provide a direction for new biomarker identification.

Predictors for incomplete response to ursodeoxycholic acid in primary biliary cholangitis. Data from a national registry of liver disease.

BACKGROUND: The current standard of treatment in primary biliary cholangitis (PBC) is ursodeoxycholic acid (UDCA), although a considerable proportion of patients show incomplete response resulting in disease progression. OBJECTIVE: This study aimed to assess the prevalence of incomplete response to UDCA and determine associated patients' characteristics. METHODS: Patients with PBC as main diagnosis were included from a national multicentric patient registry-Liver.pt. Main endpoints included incomplete response to UDCA treatment according to Barcelona, Paris I and Paris II criteria, Globe and UK PBC scores and the association between baseline characteristics and incomplete response according to Paris II criteria. RESULTS: A total of 434 PBC patients were identified, with a mean age of 55 years and 89.2% females. Nearly half of patients were asymptomatic at diagnosis and 93.2% had positive anti-mitochondrial antibodies. Almost all patients (95.6%) had been prescribed at least one drug for PBC treatment. At the last follow-up visit, 93.3% were under treatment of which 99.8% received UDCA. Incomplete response to UDCA was observed in 30.7%, 35.3%, 53.7% and 36.4% of patients according to Barcelona, Paris I, Paris II criteria and Globe score, respectively. After adjusting for age and sex, and accordingly to Paris II criteria, the risk for incomplete biochemical response was 25% higher for patients with cirrhosis at diagnosis (odds ratio [OR] = 1.25; 95% confidence interval [95%CI]: 1.02-1.54; p = 0.033) and 35% (95%CI:1.06-1.72; p = 0.016) and 5% (OR = 1.05; 95%CI:1.01-1.10; p = 0.013) for those with elevated gamma-glutamyl transferase (GGT) and alkaline phosphatase (ALP). CONCLUSION: A considerable proportion of patients showed incomplete biochemical response to UDCA treatment according to Paris II criteria. Cirrhosis, elevated GGT and ALP at diagnosis were identified as associated risk factors for incomplete response. Early identification of patients at risk of incomplete response could improve treatment care and guide clinical decision to a more careful patient monitorization.

The Serum Proteome and Ursodeoxycholic Acid Response in Primary Biliary Cholangitis.

BACKGROUND AND AIMS: Stratified therapy has entered clinical practice in primary biliary cholangitis (PBC), with routine use of second-line therapy in nonresponders to first-line therapy with ursodeoxycholic acid (UDCA). The mechanism for nonresponse to UDCA remains, however, unclear and we lack mechanistic serum markers. The UK-PBC study was established to explore the biological basis of UDCA nonresponse in PBC and identify markers to enhance treatment. APPROACH AND RESULTS: Discovery serum proteomics (Olink) with targeted multiplex validation were carried out in 526 subjects from the UK-PBC cohort and 97 healthy controls. In the discovery phase, untreated PBC patients (n = 68) exhibited an inflammatory proteome that is typically reduced in scale, but not resolved, with UDCA therapy (n = 416 treated patients). Nineteen proteins remained at a significant expression level (defined using stringent criteria) in UDCA-treated patients, six of them representing a tightly linked profile of chemokines (including CCL20, known to be released by biliary epithelial cells (BECs) undergoing senescence in PBC). All showed significant differential expression between UDCA responders and nonresponders in both the discovery and validation cohorts. A linear discriminant analysis, using serum levels of C-X-C motif chemokine ligand 11 and C-C motif chemokine ligand 20 as markers of responder status, indicated a high level of discrimination with an AUC of 0.91 (CI, 0.83-0.91). CONCLUSIONS: UDCA under-response in PBC is characterized by elevation of serum chemokines potentially related to cellular senescence and was previously shown to be released by BECs in PBC, suggesting a potential role in the pathogenesis of high-risk disease. These also have potential for development as biomarkers for identification of high-risk disease, and their clinical utility as biomarkers should be evaluated further in prospective studies.

MicroRNA-34a Promotes EMT and Liver Fibrosis in Primary Biliary Cholangitis by Regulating TGF-ß1/smad Pathway.

BACKGROUND AND AIMS: Primary biliary cholangitis (PBC) is an autoimmune cholestatic liver disease. We found microRNA-34a (miR-34a), as the downstream gene of p53, was overexpressed in some of fibrogenic diseases. In this study, we sought to explore whether miR-34a plays a role in the fibrosis of PBC. METHODS: The peripheral blood of PBC patients and controls was collected to analyze the level of miR-34a. Human intrahepatic biliary epithelial cells (HIBEC) were cultured. The expression of miR-34a was regulated by miR-34a mimics and inhibitor. The biomarkers of epithelium-mesenchymal transition (EMT), fibrogenesis, inflammation, and transforming growth factor- (TGF-) ß1/smad pathway were analyzed. RESULTS: We found that miR-34a was overexpressed in the peripheral blood in PBC patients. In vitro, overexpressed miR-34a increased the EMT and fibrogenesis activity of HIBEC. Transforming growth factor-beta type 1 receptor (TßR1), TGF-ß1, and p-smad2/3 were upregulated by miR-34a. Inflammatory factors such as IL-6 and IL-17 were also upregulated. Finally, we showed that miR-34a promoted EMT and liver fibrosis in PBC by targeting the TGF-ß1/smad pathway antagonist transforming growth factor-beta-induced factor homeobox 2 (TGIF2). CONCLUSIONS: Our findings show that miR-34a plays an important role in the EMT and fibrosis of PBC through the TGF-ß1/smad pathway by targeting TGIF2. This study suggests that miR-34a may be a new marker of fibrogenesis in PBC. Inhibition of miR-34a may be a promising strategy in treating PBC and improving the prognosis of the disease.

CXCL12-CXCR4-Mediated Chemotaxis Supports Accumulation of Mucosal-Associated Invariant T Cells Into the Liver of Patients With PBC.

Objectives: To explore the potential role of CD3+CD8+CD161high TCRVα7.2+ mucosal-associated invariant T (MAIT) cells in the pathogenesis of primary biliary cholangitis (PBC). Methods: We enrolled 55 patients with PBC, 69 healthy controls (HCs), and 8 patients with hepatic hemangioma. Circulating MAIT cells and their chemokine receptor profiles and cytokine production were quantified using flow cytometry. Liver-resident MAIT cells were examined by immunofluorescence staining. CXCL12-mediated chemotaxis of MAIT cells was measured using a transwell migration assay. Plasma interleukin (IL)-18 was measured using ELISA, and cytokine production in IL-18-stimulated MAIT cells was detected using flow cytometry. Result: Peripheral MAIT cells were found to be significantly lower in patients with PBC (3.0 ± 3.2% vs. 9.4 ± 8.0%, p < 0.01) and negatively correlated with alkaline phosphatase (ALP) levels (r = -0.3209, p < 0.05). Liver immunofluorescence staining suggested that MAIT cells might accumulate in PBC liver. MAIT cells from patients with PBC expressed higher levels of CXCR4 (84.8 ± 18.0% vs. 58.7 ± 11.4%, p < 0.01), and the expression of CXCL12 was higher in PBC liver. CXCL12 promoted MAIT cell chemotaxis (70.4 ± 6.8% vs. 52.2 ± 3.5%, p < 0.01), which was attenuated by CXCR4 antagonist. MAIT cells from PBC produced significantly more interferon-γ (IFN-γ) (88.3 ± 4.2% vs. 64.2 ± 10.1%, p < 0.01), tumor necrosis factor-α (TNF-α) (93.0 ± 1.1% vs. 80.1 ± 5.3%, p < 0.01), Granzyme B (89.3 ± 3.3% vs. 72.1 ± 7.0%, p < 0.01), and perforin (46.8 ± 6.6% vs. 34.8 ± 7.7%, p < 0.05). MAIT cells from PBC expressed higher levels of IL18-Rα (83.8 ± 10.2% vs. 58.3 ± 8.7%, p < 0.01). Plasma IL-18 was more abundant in patients with PBC (286.8 ± 75.7 pg/ml vs. 132.9 ± 78.1 pg/ml, p < 0.01). IL-18 promoted IFN-γ production in MAIT cells (74.9 ± 6.6% vs. 54.7 ± 6.7%, p < 0.01), which was partially attenuated by blocking IL-18R (68.6 ± 8.3% vs. 43.5 ± 4.2%, p < 0.01). Conclusion: Mucosal-associated invariant T cells from patients with PBC accumulated in the liver via CXCL12-CXCR4-mediated chemotaxis, produced pro-inflammatory cytokines, and contributed to portal inflammation, which was potentially mediated by elevated IL-18. Targeting MAIT cells might be a therapeutic approach for PBC.

Association of serum CCL20 levels with pulmonary vascular involvement and primary biliary cholangitis in patients with systemic sclerosis.

AIM: Systemic sclerosis (SSc) is a chronic autoimmune disease resulting in vasculopathy and fibrosis of the skin and major internal organs. Especially, interstitial lung disease and pulmonary arterial hypertension are the leading causes of mortality. C-C motif ligand 20 (CCL20) is known as a homeostatic and inflammatory chemokine, which is associated with fibrosis and angiogenesis and constantly expressed in organs involved in SSc. Therefore, we investigated the potential contribution of CCL20 to the development of SSc. METHOD: We conducted cross-sectional analyses of 67 SSc patients and 20 healthy controls recruited in a single center for 9 years. Serum CCL20 levels were measured by enzyme-linked immunosorbent assay. Statistical analyses were performed with the Mann-Whitney U test, the Kruskal-Wallis test followed by Dunn's multiple comparison test, Fisher's exact probability test and the Spearman's rank correlation coefficient. RESULTS: SSc patients had significantly higher serum CCL20 levels than healthy controls. In SSc patients, serum CCL20 levels correlated inversely with the percentage of predicated diffusion lung capacity for carbon monoxide and positively with mean pulmonary artery pressure (mPAP). In addition, SSc patients with increased serum CCL20 levels had anti-mitochondrial antibody M2 titer significantly elevated relative to those with normal levels, and SSc patients with asymptomatic primary biliary cholangitis (PBC) possessed higher serum CCL20 levels than those without. Importantly, serum CCL20 levels were associated positively with mPAP values and PBC presence by multivariate regression analysis. CONCLUSION: Serum CCL20 levels may be involved in the development of pulmonary vascular involvement leading to pulmonary arterial hypertension and asymptomatic PBC in SSc patients.