IL-35基因修饰间充质干细胞通过JAK-STAT通路治疗免疫性肝损伤的作用及机制研究

Immunological regulation of acute liver injury is one of the major topics of today’s immunology and liver disease research areas. Recent study showed that CD4+CD25+Treg play protective role through IL-10 or TGF-b in Con A induced liver injury in mice. IL-35 is a newly described Treg-specific cytokine, which is required for maximal regulatory activity of Tregs in vitro and in vivo. IL-35 can induce conventional CD4+CD25- T cells (Tconv) convert into suppressive regulatory T cell population termed iTr35. iTr35 may play a significant physiological contribution to the regulatory milieu established by Treg and become the research hotspot in autoimmune diseases. The question we care about is that which role played by IL-35 in the Con A induced liver injury in mice. Mesenchymal stem cells(MSCs) are considered to maintain the high self-renewal capacity, have the potential to differentiate into all cells of three germinal layers, and be useful in the fields of stem cell transplanting therapy and tissue engineering. Researches indicated that MSCs can be transplanted into mice successfully via tail vein and can specifically migrate to the mucosa of the injured tissue. In this study, we use BM-MSCs serve as vehicle carrying IL-35 expressing gene targeted injured liver to evaluate the therapeutic effect of exogenous IL-35 and iTr35 by using Con A incued liver injury model. Through analyzing its effect on the local immune microenvironment,the number or function of NKT cells and changes of the downstream signal transduction pathway,the possible mechanism of exogenous IL-35 on liver injury will be dissected.

免疫性肝损伤见于自身免疫性肝炎、病毒性肝炎及药物性肝损伤，严重危害人类的健康。白介素-35（IL-35）在多种自身免疫病模型中起保护作用，但其与天然免疫、感染免疫的关系及其在器官损伤防护中的作用尚不清楚。课题组前期研究证实IL-35基因修饰的间充质干细胞（MSC-IL-35）具有更强的免疫抑制功能，并能诱导Treg的产生及增殖。本课题拟利用刀豆蛋白A(ConA)诱导的小鼠肝损伤模型观察MSC-IL-35对免疫性肝损伤的治疗作用。通过检测淋巴器官及肝脏局部CD4+T、NKT、Treg细胞比例、活化状态及功能，以及MSCs细胞因子分泌功能对T细胞亚群构成及功能的影响，研究MSC-IL-35在维持机体及肝脏局部免疫稳态中的作用。通过检测IL-35下游信号通路关键分子及转录因子的表达/磷酸化，着重从IL-35的角度阐明其治疗免疫性肝损伤的细胞及分子机制，为肝损伤的免疫调理治疗提供新策略和理论依据。

免疫性肝损伤见于自身免疫性肝炎、病毒性肝炎及药物性肝损伤，严重危害人类的健康。白细胞介素-35（Interleukin-35，IL-35）在多种自身免疫性疾病模型中起保护作用，但其与天然免疫、感染免疫的关系及其在器官损伤防护中的作用尚不清楚。间充质干细胞（Mesenchymal stem cells，MSCs）因其低免疫原性和能够向受损器官组织定向迁移的特性而备受关注，成为众多免疫治疗的基因载体。本课题组采用慢病毒感染的方式将IL-35 的编码基因整合到MSC的基因组中，体外实验证实其能够成功表达大量具有抑制功能的IL-35。然后利用刀豆蛋白A(Concanavalin A， ConA)诱导的小鼠免疫性肝损伤模型，通过静脉输注的方式给予IL-35基因修饰的间充质干细胞（IL-35-MSCs）进行基因治疗。通过血清学检查和病理学诊断评价IL-35-MSCs对免疫性肝损伤的治疗作用，结果发现IL-35-MSCs能够明显减轻小鼠肝脏损伤的范围和程度，治疗组小鼠肝细胞坏死和凋亡都较对照组相比明显减轻。通过对肝脏淋巴细胞亚群比例变化和功能的分析，证实MSC-IL-35主要通过JAK1-STAT1/STAT4通路抑制CD4+ T细胞分泌干扰素-g，从而减轻Con A诱导的小鼠免疫性肝损伤程度。为临床上的自身免疫性肝炎、终末性肝炎等难治性肝脏疾病新疗法的开发提供了实验依据。