Objective: To analyze the characteristics of immunoglobulin heavy chain complementarity-determining region (IgH-CDR3) repertoire of peripheral B cells in a patient with primary biliary cholangitis (PBC) and to investigate the diversity of the immune system. Methods: Arm-PCR was used to amplify the IgH-CDR3 region of circulating B cells isolated from a PBC patient, and high-throughput sequencing was used to analyze the amplified product. The characteristics of immune repertoire were analyzed by bioinformatics. Results: In total, 329219 sequence reads were generated from the sample, with 325540 total CDR3 sequences and 72774 distinct CDR3 sequences, and the D50 of IGH-CDR3 was 7.7. The dominant CDR3 length of the sample was 45 nt (9.6%); the N addition with the highest frequency ranged from 13 to 14 nt (5.25%); the J trimming with the highest frequency was 0 nt (12.7%); the three most frequent V alleles were V4-59 (9.5%), V3-23 (8.1%), and V1-69 (6.4%). Conclusion: The diversity of IgH-CDR3 repertoire is relatively low in this patient with PBC, with several B-cell clonal expansions. The specificity needs to be further verified after increasing the sample size.
目的: 分析原发性胆汁性胆管炎(PBC)患者个体水平外周血B细胞受体免疫球蛋白重链互补决定区3(IGH-CDR3)免疫组库特征,了解其免疫系统多样性的特点。 方法: 采用扩增子救援多重PCR(arm-PCR)技术特异性扩增一例PBC患者外周血B细胞受体,即膜表面免疫球蛋白重链IGH-CDR3区核苷酸序列,通过第二代高通量测序技术进行测序,结合生物信息学分析其免疫组库特征。 结果: 共测得序列数329 219个,总CDR3序列为325 540个,独特CDR3序列为72 774个,D50为7.7;IGH-CDR3区长度主要为45个核苷酸(nt)(9.6%),N区添加的核苷酸数量频率最高的为13~14 nt(5.25%),J基因片段末端去除的核苷酸数主要是0(12.7%);V基因使用频率最高的前3个等位基因是V4-59(9.5%),其次为V3-23(8.1%)和V1-69(6.4%)。 结论: 该PBC患者IGH-CDR3免疫组库多样性程度较低,存在个别克隆异常增殖情况,该特异性有待扩大样本量进一步验证。.
Keywords: B cell receptor; IGH-CDR3 repertoire; Liver cirrhosis, biliary.