补体C5a/C5aR途径促Fgl2表达和C5b-9形成的机制及其在暴发性肝衰竭中的作用

The study to provide significant insights into the mechanisms underlying the pathogenesis of the fulminant liver failure will ultimately be very useful in the prophylaxis and therapy of viral hepatitis.The role of complement in fulminant viral hepatitis is unclear. The clinical syndrome of acute liver failure produced by fulminant viral hepatitis can be reproduced in mice by infection with murine hepatitis virus strain 3 (MHV-3), so it becomes an ideal animal model in the study. We found that mice mortality rate and C5b-9 deposition in liver tissue in C5aR KO mice was significant lower than that of WT mice in this murine model of fulminant viral hepatitis. In further studies, we found that C5a/C5aR pathway played an important role in the modulation fgl2 expression. We therefore hypothesized that C5a/C5aR pathway mediates hepatocellular injury in viral hepatitis via the potentiation of Fgl2 induced coagulation and feedback regulation on C5 activation. In the present studies, we will define cellular mechanisms involved in C5a/C5aR regulation in fgl2 expression by series of in vivo and in vitro experiments, confirm the feedback loop of C5a/C5aR pathway on C5 activation, and explore the effect of C5b-9 on the induction of fulminant hepatic failure. Anticipative results will help us further unravel the precise intercommunication between the complement and coagulation systems and the pathogenesis of fulminant liver failure, which may ultimately suggest novel treatment strategies for clinical disease.

研究暴发性肝衰的发病机制，无疑对其预防和治疗有重要意义。补体在病毒诱导的暴发性肝衰中的作用未见报道。MHV-3诱导的暴发性肝衰模型，是最常用的动物模型。利用该模型我们发现，C5aR基因敲除小鼠死亡率显著下降，C5b-9的沉积显著减少。进一步实验发现，C5a/C5aR可调控Fgl2的表达。我们提出，C5a/C5aR可能通过影响Fgl2表达来调节凝血系统的活化并正反馈调节补体C5的活化，促进暴发性肝衰的发生。本课题拟通过体内和体外实验探讨C5a/C5aR促进Fgl2表达的分子机制；通过对Fgl2和凝血酶等的干预实验证实C5a/C5aR通路调节C5活化的正反馈途径；通过对C6的干预来探讨C5b-9在暴发性肝衰中的作用。本课题的结果无疑可使我们深入了解补体和凝血这两个复杂系统的相互作用以及补体自身调节机制；同时可加深对暴发性肝衰发病机理的认识，为通过干预补体活化来减少暴发性肝衰的风险提供理论依据。

肝炎病毒感染所致的暴发性肝衰是严重危害人类健康的疾病之一,其死亡率高达80 %。缺乏对暴发性肝炎的病理发生的了解严重阻碍了有效临床治疗策略的发展。鼠Ⅲ型肝炎病毒(murine hepatitis virus strain 3, MHV-3)感染近交系小鼠建立的暴发性肝衰模型，可重现暴发性病毒性肝炎所诱发的急性肝衰临床综合症，是近年研究病毒诱导的暴发性肝衰发病机制最重要的动物模型。从补体组分C5基因的不同与小鼠对MHV-3敏感性的关联，我们推测过度的补体活化在暴发性肝炎的发生发展起着重要的作用。此研究中，我们发现MHV-3感染敏感型小鼠后，引发补体的大量活化，血清中补体活性片段C5a水平快速升高；肝组织中C5aR表达显著上调；小鼠迅速发展为暴发性肝炎。C5aR基因缺陷或用C5aR阻断剂处理可显著衰减小鼠疾病的进展，伴随着引起感染肝脏坏死的标志蛋白Fgl2的明显降低。与之一致的是，暴发性肝炎病人肝组织Fgl2表达明显上调，与C5aR的上调表达呈正相关。体外实验中，加入重组C5a可加速MHV-3诱导的巨噬细胞Fgl2的产生，而且抑制ERK1/2和p38的活化可有效阻止C5a介导的Fgl2的产生。结论：这些结果证实小鼠对MHV-3诱发的暴发性肝炎的敏感性依赖于补体C5a/C5aR的相互作用，并且C5aR信号通路通过激活ERK1/2和p38 MAPK通路参与Fgl2的产生。抑制C5a/C5aR的相互作用有望为临床暴发性肝炎病人的治疗提供帮助。