Nucleotide Variations amongst VHGenes of AMA-Producing B Cell Clones in Primary Biliary Cirrhosis

Abstract

Primary biliary cirrhosis, a chronic liver disease characterized by progressive inflammatory destruction of intrahepatic bile ducts, is also characterized by the presence of antimitochondrial antibodies (AMA). The predominant autoantibody is directed at the E2 component of pyruvate dehydrogenase (PDC-E2). Recent studies of this autoantibody response have analysed immunoglobulin-variable regions of human monoclonal antibodies and provided evidence for antigen-driven clonal selection. However, the number of clones analysed has been very limited and the presence of somatic mutations not formally proven. In this study, we took advantage of three stable B cell lines producing human IgG anti-PDC-E2 mAbs from a patient with PBC. We analysed the VHand VLgene structure of these reagents and, in addition, analysed 10 VH-D and D-JHsequences over a period of nearly 3 years. The expressed Ig V regions of the heavy chain (VH) and the light chain (VL) genes of mAb18, mAb37, and mAb82 utilized the VHIII-VλI, VHIV-VλIII, and VHIV-VkIV gene families, respectively. The utilized gene elements were Ig gene elements that were found frequently in other antibodies with different specificity and affinity. Presence of somatic point-mutations was confirmed in mAb82 by comparison of the expressed VHgene sequence with that of corresponding germline VHgene obtained from the granulocyte genomic DNA of the same patient. Interestingly, clonally related B cells were consistently found throughout the observation period and nucleotide variations among the VHgenes were very few, ranging from 0.19 to 0.72% per base. These findings suggest that long-lived B cell clones can exist and may contribute, at least in part, to maintenance of autoantibodies in humans.