高通量测序揭示原发性胆汁性胆管炎患者的T细胞受体图谱特征
High-throughput sequencing reveals the characteristics of T cell receptor profiles in patients with primary biliary cholangitis
刘珍玉 1张俊宁 1杨雪丽 1王广宇 1侯显良1
作者信息
- 1. 桂林医学院第二附属医院 广西慢性病代谢重塑与智能医学工程重点实验室 中心实验室,广西 桂林 541199
- 折叠
摘要
目的 揭示原发性胆汁性胆管炎(primary biliary cholangitis,PBC)患者T细胞受体(T cell receptor,TCR)图谱特征,并揭示大肠埃希菌(Escherichia coli,E.coli)的丙酮酸脱氢酶复合体E2亚基(pyruvate dehydrogenase complex E2,PDC-E2)抗原在PBC疾病的分子模拟机制.方法 采用多重聚合酶链反应和免疫组库测序技术分析PBC患者和健康对照者的CD4+和CD8+记忆性TCR β链互补决定区3(complementarity determining region 3,CDR3)序列的多样性、氨基酸组成及疏水性、共有CDR3序列.体外诱导和扩增人PDC-E2163-176(人PDC-E2)抗原相关T细胞和E.coli PDC-E231-44/134-147/235-248(E.coli PDC-E2)抗原相关T细胞,通过免疫组库测序技术鉴定人(和E.coli)PDC-E2抗原相关TCR β CDR3图谱,并分析其丰度变化.结果 PBC患者组和健康对照组间的CD4+记忆性T细胞、CD8+记忆性T细胞TCRβ CDR3免疫图谱多样性相似,D50指数[CD4+记忆性T细胞:0.028±0.019比0.034±0.015;CD8+记忆性T细胞:(1.86±2.70)×10-3比(4.62±3.89)×10-4]、Shannon指数(CD4+记忆性T细胞:9.473±1.346比9.734±0.933;CD8+记忆性T细胞:6.197±1.519比5.436±1.629)、Gini指数(CD4+记忆性T细胞:0.786±0.048比0.760±0.036;CD8+记忆性T细胞:0.920±0.047比0.939±0.025)等差异均无统计学意义(P均＞0.05).PBC组和健康对照组CD4+记忆性T细胞和CD8+记忆性T细胞间共有CDR3序列百分比差异无统计学意义[(6.47±1.43)%比(6.21±3.18)%;t =-0.21,P = 0.84].健康对照组和PBC组中序列长度为13、14、15的CDR3分子第6位和第7位氨基酸的组成频率中部分存在显著差异,疏水氨基酸组成频率近似.通过细胞培养和免疫组库测序鉴定了一系列人PDC-E2和E.coli PDC-E2抗原刺激后丰度显著上升的TCR序列.结论 该研究鉴定出PBC疾病的TCR图谱特征,从TCR这个新视角阐述了E.coli在PBC疾病中的分子模拟机制.
Abstract
Objective To reveal the characteristics of T cell receptor(TCR)profiles in patients with primary biliary cholangitis(PBC)and to reveal the molecular mimicry mechanism of pyruvate dehydrogenase complex E2(PDC-E2)antigen of Escherichia coli(E.coli)in PBC disease.Methods The sequence diversity,amino acid composition and hydrophobicity,and common sequences of memory TCR β chain complementarity determining region 3s(CDR3s)of PBC patients and healthy volunteers were analyzed by multiplex polymerase chain reaction and immune repertoire sequencing.Human PDC-E2163-176(PDC-E2)antigen-associated T cells and E.coli PDC-E231-44/134-147/235-248(E.coli PDC-E2)antigen-associated T cells were induced and amplified in vitro,and human(E.coli)PDC-E2 antigen-related TCRβ CDR3 repertoire were identified by immune repertoire sequencing.Results TCRβ CDR3 repertoire diversity of CD4+memory T cells and CD8+ memory T cells between PBC patients and healthy controls was similar,there were no statistically significant differences in D50 index[CD4+ memory T cells:0.028±0.019 vs.0.034±0.015;CD8+ memory T cells:(1.86±2.70)×10-3 vs.(4.62±3.89)×10-4],Shannon index(CD4+ memory T cells:9.473±1.346 vs.9.734±0.933;CD8+ memory T cells:6.197±1.519 vs.5.436±1.629),Gini index(CD4+ memory T cells:0.786±0.048 vs.0.760±0.036;CD8+ memory T cells:0.920±0.047 vs.0.939±0.025),etc.There was no significant difference in the percentage of common CDR3 sequence between CD4+ memory T cells and CD8+ memory T cells of PBC patients and healthy controls[(6.47±1.43)%vs.(6.21±3.18)%;t =-0.21,P = 0.84].The percentage of some amino acids at positions 6 and 7(P6 and P7)of TCRβ CDR313～15 amino acids in length in healthy control group and PBC group were significantly different,while the percentage of hydrophobic amino acids were similar.It is noteworthy that a series of human PDC-E2 and E.coli PDC-E2 antigen-related TCR sequences through cell culture and immune repertoire sequencing,which may play an important role in the pathogenesis of PBC were identified.Conclusions This study identified the TCR profile features of PBC disease and elucidated the molecular mimicry mechanism of E.coli in PBC disease from the perspective of TCR.
关键词
原发性胆汁性胆管炎/T细胞受体/高通量测序/丙酮酸脱氢酶复合体E2亚基Key words
Primary biliary cholangitis/T cell receptor/High-throughput sequencing/Pyruvate dehydrogenase complex E2 subunit引用本文复制引用
基金项目
中央引导地方科技发展资金项目(桂科AD20238021)
广西自然科学基金(2024GXNSFAA010096)
中国博士后科学基金(2023M740344)
出版年
2024
NETL
NSTL
万方数据