原发性胆汁性胆管炎相关肠道菌群通过调控肠道滤泡辅助性T细胞影响肝脏免疫微环境的机制研究

Primary biliary cholangitis (PBC) is characterized by the destruction of intrahepatic small bile ducts. The role of gut-liver axis has been increasingly appreciated in the pathogenesis of PBC. Recently, our group has found gut microbial profile was altered in PBC patients, suggesting the important role of gut microbiota in PBC. However, it remains unclear whether the changes are cause or effect of the disease. Our preliminary studies found that Klebsiella pneumoniae was enriched in PBC patients, compared with healthy controls. Interestingly, the serum level of total bilirubin was significantly higher in PBC patients with high-abundance of Klebsiella pneumoniae than patients with low-abundance of K. pneumoniae. Furthermore, we observed that extracts from Klebsiella pneumoniae expanded T follicular helper (Tfh) cell populations in peripheral blood mononuclear cells from healthy donors. Notably, our genome-wide association study has identified that several key molecules for regulating Tfh development and function including CXCR5, IL21 and ICOS, were PBC susceptible genes. Therefore, we hypothesize that PBC-associated gut microbe, Klebsiella pneumoniae, may trigger bile duct damage by inducing Tfh cell differentiation and migration into liver, leading to increased production of auto-antibody by B cells and altered microenvironment in the liver. Our study will elucidate the mechanism of gut microbial dysbiosis in the disease development and provide potential therapeutic target for PBC.

原发性胆汁性胆管炎（PBC）是以肝内小叶间胆管损伤为特征的自身免疫性肝病，“肠-肝对话”是PBC发病机制研究的重要方向。我们前期研究表明，肠道菌群失衡与PBC发病密切相关，但两者的因果关系尚不明确。进一步分析发现，肺炎克雷伯菌的丰度在PBC患者肠道中明显增加，且与血清胆红素水平显著相关；肺炎克雷伯菌提取物可上调外周血滤泡辅助性T细胞（Tfh）的比例。值得一提的是，我们的全基因组关联分析发现，Tfh细胞关键分子CXCR5、IL21和ICOS基因的遗传变异显著增加PBC易感性。在此基础上，我们提出如下假设：PBC相关细菌肺炎克雷伯菌在肠道定植后，调控肠道Tfh细胞分化、成熟和肝脏迁移；肝内Tfh细胞促进B细胞成熟并产生特异性自身抗体，引发肝内体液免疫反应和胆管损伤。本研究拟从临床样本、细胞及小鼠模型等多个水平探讨肠道菌群失衡参与PBC发生发展的免疫学机制，有望为PBC治疗提供新的思路和策略。

原发性胆汁性胆管炎（PBC）病因复杂，涉及遗传易感性、环境因素及免疫应答异常等。肠道菌群作为一种重要的环境因素与宿主免疫系统和疾病的密切联系受到广泛关注。通过宏基因组测序，我们发现韦荣氏球菌在PBC患者中变化最为显著。更为重要的是，韦荣氏球菌与疾病严重程度呈正相关，且在UDCA非应答者中的丰度显著高于应答者，提示韦荣氏球菌可能在PBC发生发展中发挥重要作用。有意思的是，韦荣氏球菌与滤泡辅助性T细胞（Tfh）的频率呈正相关；而肺炎克雷伯菌与Tfh细胞未见明显相关性。进一步研究发现，PBC患者肝内存在一群组织驻留特性的Tfh样细胞，这一群细胞是PBC肝内驱动体液免疫信号的主要效应细胞。值得一提的是，具有干性的Tfh样CD4+ Tscm细胞是PBC中Tfh细胞的主要来源。体外实验发现，韦荣氏球菌提取物可促进Tfh细胞分化；而肺炎克雷伯菌不能提高Tfh细胞的诱导效率。进一步研究发现，韦荣氏球菌可促进小鼠自身免疫性肝损伤的发生发展。除了菌体及其自身成分与宿主细胞的直接接触作用外，紊乱的肠道菌群影响机体生理病理的另一重要途径是肠菌来源的代谢产物。我们发现，PBC患者胆汁酸谱和短链脂肪酸发生显著改变，且与疾病严重程度和治疗应答有关。机制研究发现，短链脂肪酸通过调控髓系源性抑制细胞（MDSCs）的分化和免疫抑制功能，进而影响肝内免疫微环境。本项目不仅建立了肠道细菌与宿主免疫细胞及临床表型的相关性，而且明确肠道细菌与疾病发生的潜在因果关系，对开展基于肠道菌群的靶向治疗具有重要的意义。