Regulation of immune responses in primary biliary cholangitis: a transcriptomic analysis of peripheral immune cells

BACKGROUND AIMS: In patients with primary biliary cholangitis (PBC), the serum liver biochemistry measured during treatment with ursodeoxycholic acid-the UDCA response-accurately predicts long-term outcome. Molecular characterization of patients stratified by UDCA response can improve biological understanding of the high-risk disease, thereby helping to identify alternative approaches to disease-modifying therapy. In this study, we sought to characterize the immunobiology of the UDCA response using transcriptional profiling of peripheral blood mononuclear cell subsets. METHODS: We performed bulk RNA-sequencing of monocytes and TH1, TH17, TREG, and B cells isolated from the peripheral blood of 15 PBC patients with adequate UDCA response ("responders"), 16 PBC patients with inadequate UDCA response ("nonresponders"), and 15 matched controls. We used the Weighted Gene Co-expression Network Analysis to identify networks of co-expressed genes ("modules") associated with response status and the most highly connected genes ("hub genes") within them. Finally, we performed a Multi-Omics Factor Analysis of the Weighted Gene Co-expression Network Analysis modules to identify the principal axes of biological variation ("latent factors") across all peripheral blood mononuclear cell subsets. RESULTS: Using the Weighted Gene Co-expression Network Analysis, we identified modules associated with response and/or disease status (q<0.05) in each peripheral blood mononuclear cell subset. Hub genes and functional annotations suggested that monocytes are proinflammatory in nonresponders, but antiinflammatory in responders; TH1 and TH17 cells are activated in all PBC cases but better regulated in responders; and TREG cells are activated-but also kept in check-in responders. Using the Multi-Omics Factor Analysis, we found that antiinflammatory activity in monocytes, regulation of TH1 cells, and activation of TREG cells are interrelated and more prominent in responders. CONCLUSIONS: We provide evidence that adaptive immune responses are better regulated in patients with PBC with adequate UDCA response

Amino acid residues in five separate HLA genes can explain most of the known associations between the MHC and primary biliary cholangitis

Primary Biliary Cholangitis (PBC) is a chronic autoimmune liver disease characterised by progressive destruction of intrahepatic bile ducts. The strongest genetic association is with HLA-DQA1*04:01, but at least three additional independent HLA haplotypes contribute to susceptibility. We used dense single nucleotide polymorphism (SNP) data in 2861 PBC cases and 8514 controls to impute classical HLA alleles and amino acid polymorphisms using state-of-the-art methodologies. We then demonstrated through stepwise regression that association in the HLA region can be largely explained by variation at five separate amino acid positions. Three-dimensional modelling of protein structures and calculation of electrostatic potentials for the implicated HLA alleles/amino acid substitutions demonstrated a correlation between the electrostatic potential of pocket P6 in HLA-DP molecules and the HLA-DPB1 alleles/amino acid substitutions conferring PBC susceptibility/protection, highlighting potential new avenues for future functional investigation

Regulation of immune responses in primary biliary cholangitis: a transcriptomic analysis of peripheral immune cells

Copyright \ua9 2023 The Author(s). Published by Wolters Kluwer Health, Inc. on behalf of the American Association for the Study of Liver Diseases.Background &amp; Aims: In patients with primary biliary cholangitis (PBC), the serum liver biochemistry measured during treatment with ursodeoxycholic acid-the UDCA response-accurately predicts long-term outcome. Molecular characterization of patients stratified by UDCA response can improve biological understanding of the high-risk disease, thereby helping to identify alternative approaches to disease-modifying therapy. In this study, we sought to characterize the immunobiology of the UDCA response using transcriptional profiling of peripheral blood mononuclear cell subsets. Methods: We performed bulk RNA-sequencing of monocytes and TH1, TH17, TREG, and B cells isolated from the peripheral blood of 15 PBC patients with adequate UDCA response (“responders”), 16 PBC patients with inadequate UDCA response (“nonresponders”), and 15 matched controls. We used the Weighted Gene Co-expression Network Analysis to identify networks of co-expressed genes (“modules”) associated with response status and the most highly connected genes (“hub genes”) within them. Finally, we performed a Multi-Omics Factor Analysis of the Weighted Gene Co-expression Network Analysis modules to identify the principal axes of biological variation (“latent factors”) across all peripheral blood mononuclear cell subsets. Results: Using the Weighted Gene Co-expression Network Analysis, we identified modules associated with response and/or disease status (q &lt; 0.05) in each peripheral blood mononuclear cell subset. Hub genes and functional annotations suggested that monocytes are proinflammatory in nonresponders, but antiinflammatory in responders; TH1 and TH17 cells are activated in all PBC cases but better regulated in responders; and TREG cells are activated-but also kept in check-in responders. Using the Multi-Omics Factor Analysis, we found that antiinflammatory activity in monocytes, regulation of TH1 cells, and activation of TREG cells are interrelated and more prominent in responders. Conclusions: We provide evidence that adaptive immune responses are better regulated in patients with PBC with adequate UDCA response

Critical shortfalls in the management of PBC: Results of a UK-wide, population-based evaluation of care delivery

\ua9 2023 The Author(s)Background &amp; Aims: Guidelines for the management of primary biliary cholangitis (PBC) were published by the British Society of Gastroenterology in 2018. In this study, we assessed adherence to these guidelines in the UK National Health Service (NHS). Methods: All NHS acute trusts were invited to contribute data between 1 January 2021 and 31 March 2022, assessing clinical care delivered to patients with PBC in the UK. Results: We obtained data for 8,968 patients with PBC and identified substantial gaps in care across all guideline domains. Ursodeoxycholic acid (UDCA) was used as first-line treatment in 88% of patients (n = 7,864) but was under-dosed in one-third (n = 1,964). Twenty percent of patients who were UDCA-untreated (202/998) and 50% of patients with inadequate UDCA response (1,074/2,102) received second-line treatment. More than one-third of patients were not assessed for fatigue (43%; n = 3,885) or pruritus (38%; n = 3,415) in the previous 2 years. Fifty percent of all patients with evidence of hepatic decompensation were discussed with a liver transplant centre (222/443). Appropriate use of second-line treatment and referral for liver transplantation was significantly better in specialist PBC treatment centres compared with non-specialist centres (p &lt;0.001). Conclusions: Poor adherence to guidelines exists across all domains of PBC care in the NHS. Although specialist PBC treatment centres had greater adherence to guidelines, no single centre met all quality standards. Nationwide improvement in the delivery of PBC-related healthcare is required. Impact and implications: This population-based evaluation of primary biliary cholangitis, spanning four nations of the UK, highlights critical shortfalls in care delivery when measured across all guideline domains. These include the use of liver biopsy in diagnosis; referral practice for second-line treatment and/or liver transplant assessment; and the evaluation of symptoms, extrahepatic manifestations, and complications of cirrhosis. The authors therefore propose implementation of a dedicated primary biliary cholangitis care bundle that aims to minimise heterogeneity in clinical practice and maximise adherence to key guideline standards

Dense fine-mapping study identifies new susceptibility loci for primary biliary cirrhosis

We genotyped 2,861 cases of primary biliary cirrhosis (PBC) from the UK PBC Consortium and 8,514 UK population controls across 196,524 variants within 186 known autoimmune risk loci. We identified 3 loci newly associated with PBC (at P 0.8) with the most associated variant at the locus. We found multiple independent common, low-frequency and rare variant association signals at five loci. Of the 26 independent non-human leukocyte antigen (HLA) signals tagged on the Immunochip, 15 have SNPs in B-lymphoblastoid open chromatin regions in high LD (r > 0.8) with the most associated variant. This study shows how data from dense fine-mapping arrays coupled with functional genomic data can be used to identify candidate causal variants for functional follow-up. © 2012 Nature America, Inc. All rights reserved

Improved Statistics for Genome-Wide Interaction Analysis

Recently, Wu and colleagues [1] proposed two novel statistics for genome-wide interaction analysis using case/control or case-only data. In computer simulations, their proposed case/control statistic outperformed competing approaches, including the fast-epistasis option in PLINK and logistic regression analysis under the correct model; however, reasons for its superior performance were not fully explored. Here we investigate the theoretical properties and performance of Wu et al.'s proposed statistics and explain why, in some circumstances, they outperform competing approaches. Unfortunately, we find minor errors in the formulae for their statistics, resulting in tests that have higher than nominal type 1 error. We also find minor errors in PLINK's fast-epistasis and case-only statistics, although theory and simulations suggest that these errors have only negligible effect on type 1 error. We propose adjusted versions of all four statistics that, both theoretically and in computer simulations, maintain correct type 1 error rates under the null hypothesis. We also investigate statistics based on correlation coefficients that maintain similar control of type 1 error. Although designed to test specifically for interaction, we show that some of these previously-proposed statistics can, in fact, be sensitive to main effects at one or both loci, particularly in the presence of linkage disequilibrium. We propose two new “joint effects” statistics that, provided the disease is rare, are sensitive only to genuine interaction effects. In computer simulations we find, in most situations considered, that highest power is achieved by analysis under the correct genetic model. Such an analysis is unachievable in practice, as we do not know this model. However, generally high power over a wide range of scenarios is exhibited by our joint effects and adjusted Wu statistics. We recommend use of these alternative or adjusted statistics and urge caution when using Wu et al.'s originally-proposed statistics, on account of the inflated error rate that can result

Lack of association between the CARD10 rs6000782 polymorphism and type 1 autoimmune hepatitis in a Japanese population

Background: Previous genome-wide association studies have evaluated the impact of common genetic variants and identified several non-HLA risk loci associated with autoimmune liver diseases. More recent genome-wide association studies and replication analyses reported an association between variants of the CARD10 polymorphism rs6000782 and risk of type 1 autoimmune hepatitis (AIH). In this case-control study, we genotyped 326 Japanese AIH patients and 214 control subjects. Results: Genomic DNA from 540 individuals of Japanese origin, including 326 patients with type-1 AIH and 214 healthy controls, was analyzed for two single nucleotide polymorphisms (SNPs) in the CARD10 gene. We selected CARD10 rs6000782 SNPs and genotyped these using PCR-RFLP method and direct sequencing. The Chi square test revealed that the rs6000782 variant alle (c) was not associated with the susceptibility for AIH in a Japanese population [p = 0.376, odds ratio (OR) 1.271, 95 % confidence interval (CI) 0.747-2.161] in an allele model. Our data also showed that CARD10 rs6000782 variants were not associated with AIH or with the clinical parameters of AIH. Conclusions: In this study we examined an association between rs6000782 SNPs in the CARD10 gene and type-1 AIH. Results showed no significant association of rs62000782 with type-1 AIH in a Japanese population. This study demonstrated no association between CARD10 rs6000782 variants and AIH in a Japanese population

The Dichotomous Pattern of IL-12R and IL-23R Expression Elucidates the Role of IL-12 and IL-23 in Inflammation

IL-12 and IL-23 cytokines respectively drive Th1 and Th17 type responses. Yet, little is known regarding the biology of these receptors. As the IL-12 and IL-23 receptors share a common subunit, it has been assumed that these receptors are co-expressed. Surprisingly, we find that the expression of each of these receptors is restricted to specific cell types, in both mouse and human. Indeed, although IL-12Rβ2 is expressed by NK cells and a subset of γδ T cells, the expression of IL-23R is restricted to specific T cell subsets, a small number of B cells and innate lymphoid cells. By exploiting an IL-12- and IL-23-dependent mouse model of innate inflammation, we demonstrate an intricate interplay between IL-12Rβ2 NK cells and IL-23R innate lymphoid cells with respectively dominant roles in the regulation of systemic versus local inflammatory responses. Together, these findings support an unforeseen lineage-specific dichotomy in the in vivo role of both the IL-12 and IL-23 pathways in pathological inflammatory states, which may allow more accurate dissection of the roles of these receptors in chronic inflammatory diseases in humans

Unraveling the Regulatory Mechanisms Underlying Tissue-Dependent Genetic Variation of Gene Expression

It is known that genetic variants can affect gene expression, but it is not yet completely clear through what mechanisms genetic variation mediate this expression. We therefore compared the cis-effect of single nucleotide polymorphisms (SNPs) on gene expression between blood samples from 1,240 human subjects and four primary non-blood tissues (liver, subcutaneous, and visceral adipose tissue and skeletal muscle) from 85 subjects. We characterized four different mechanisms for 2,072 probes that show tissue-dependent genetic regulation between blood and non-blood tissues: on average 33.2% only showed cis-regulation in non-blood tissues; 14.5% of the eQTL probes were regulated by different, independent SNPs depending on the tissue of investigation. 47.9% showed a different effect size although they were regulated by the same SNPs. Surprisingly, we observed that 4.4% were regulated by the same SNP but with opposite allelic direction. We show here that SNPs that are located in transcriptional regulatory elements are enriched for tissue-dependent regulation, including SNPs at 3′ and 5′ untranslated regions (P = 1.84×10−5 and 4.7×10−4, respectively) and SNPs that are synonymous-coding (P = 9.9×10−4). SNPs that are associated with complex traits more often exert a tissue-dependent effect on gene expression (P = 2.6×10−10). Our study yields new insights into the genetic basis of tissue-dependent expression and suggests that complex trait associated genetic variants have even more complex regulatory effects than previously anticipated

An international genome-wide meta-analysis of primary biliary cholangitis: Novel risk loci and candidate drugs

Backgrounds & Aims Primary biliary cholangitis (PBC) is a chronic liver disease in which autoimmune destruction of the small intrahepatic bile ducts eventually leads to cirrhosis. Many patients have inadequate response to licensed medications, motivating the search for novel therapies. Previous genome-wide association studies (GWAS) and meta-analyses (GWMA) of PBC have identified numerous risk loci for this condition, providing insight into its aetiology. We undertook the largest GWMA of PBC to date, aiming to identify additional risk loci and prioritise candidate genes for in silico drug efficacy screening. Methods We combined new and existing genotype data for 10,516 cases and 20,772 controls from 5 European and 2 East Asian cohorts. Results We identified 56 genome-wide significant loci (20 novel) including 46 in European, 13 in Asian, and 41 in combined cohorts; and a 57th genome-wide significant locus (also novel) in conditional analysis of the European cohorts. Candidate genes at newly identified loci include FCRL3, INAVA, PRDM1, IRF7, CCR6, CD226, and IL12RB1, which each play key roles in immunity. Pathway analysis reiterated the likely importance of pattern recognition receptor and TNF signalling, JAK-STAT signalling, and differentiation of T helper (TH)1 and TH17 cells in the pathogenesis of this disease. Drug efficacy screening identified several medications predicted to be therapeutic in PBC, some of which are well-established in the treatment of other autoimmune disorders. Conclusions This study has identified additional risk loci for PBC, provided a hierarchy of agents that could be trialled in this condition, and emphasised the value of genetic and genomic approaches to drug discovery in complex disorders. Lay summary Primary biliary cholangitis (PBC) is a chronic liver disease that eventually leads to cirrhosis. In this study, we analysed genetic information from 10,516 people with PBC and 20,772 healthy individuals recruited in Canada, China, Italy, Japan, the UK, or the USA. We identified several genetic regions associated with PBC. Each of these regions contains several genes. For each region, we used diverse sources of evidence to help us choose the gene most likely to be involved in causing PBC. We used these ‘candidate genes’ to help us identify medications that are currently used for treatment of other conditions, which might also be useful for treatment of PBC