Clonal Characteristics of Circulating B Lymphocyte Repertoire in Primary Biliary Cholangitis

Abstract

Primary biliary cholangitis (PBC) is an autoimmune liver disease characterized by elevated serum anti-mitochondrial Ab and lymphocyte-mediated bile duct damage. This study was designed to reveal the clonal characteristics of B lymphocyte repertoire in patients with PBC to facilitate better understanding of its pathogenesis and better management of these patients. Using high-throughput sequencing of Ig genes, we analyzed the repertoire of circulating B lymphocytes in 43 patients with PBC, and 34 age- and gender-matched healthy controls. Compared with healthy controls, PBC patients showed 1) a gain of 14 new clones and a loss of 8 clones; 2) a significant clonal expansion and increased relative IgM abundance, which corresponded with the elevated serum IgM level; 3) a significant reduction of clonal diversity and somatic hypermutations in class-switched sequences, which suggested a general immunocompromised status; 4) the reduction of clonal diversity and enhancement of clonal expansion were more obvious at the cirrhotic stage; and 5) treatment with ursodeoxycholic acid could increase the clonal diversity and reduce clonal expansion of the IgM repertoire, with no obvious effect on the somatic hypermutation level. Our data suggest that PBC is a complex autoimmune disease process with evidence of B lymphocyte clonal gains and losses, Ag-dependent ogligoclonal expansion, and a generally compromised immune reserve. This new insight into the pathogenesis of PBC opens up the prospect of studying disease-relevant B cells to better diagnose and treat this devastating disease.

Clonal characteristics of paired infiltrating and circulating B lymphocyte repertoire in patients with primary biliary cholangitis

“…For the peripheral repertoire of this cohort (6 PBC, 3 ALD patients and 8 HCs): both PBC and ALD showed preferential IGHV5‐51 use as compared to HC ( P = .045, P = .018, respectively), consistent with our previous report . Preferential use of IGHV2‐26 ( P = .006, .012), and reduced use of IGHV3‐9 ( P = .001, P = .015) , was observed in PBC patients as compared to both ALD and HC, which is inconsistent with previous results . Other IGHV use also deviated from the former PBC cohort, relative to HC, perhaps because of the smaller sample size and terminal disease stage of this cohort.…”

“…In line with most investigators, we defined a B cell clone using the heavy chains . A unique IGH‐CDR3 amino acid sequence represents an existing B cell clone.…”

“…Patients with coexisting infectious disease, other liver diseases, other autoimmune diseases, diabetes, recent inoculation, immunosuppressive treatment or neoplasm were excluded. All patients received routine liver‐supportive treatment that normally would not influence B cell immunity except UDCA …”

“…Multiplex‐PCR products were separated by agar gel electrophoresis, and object fragments between 200‐300 bp were recovered and purified by QIAquick Gel Purification Kit (QIAGEN). DNA library construction, cluster generation, template hybridization, isothermal amplification, linearization, blocking and denaturation, and hybridization of sequencing primers were implemented as described elsewhere . Paired‐end sequencing was performed with a 151 bp read length using the Illumina Hiseq2500 platform.…”

“…Using NGS technology, we recently reported the distinct clonal characteristics of the circulating B lymphocyte repertoire in PBC patients as compared to healthy controls (HCs) . Herein, using liver samples, we first analysed the clonal characteristics of the infiltrating B lymphocyte repertoire of PBC compared to ALD patients, then analysed the evolutionally related B cell clones in paired peripheral and liver samples to see whether monitoring the clonal characteristics of peripheral B cells could reflect the infiltrating ones.…”