肠道树突状细胞亚群microRNA155表达变化对炎症性肠病发病的影响及机制研究

Intestinal inflammation in inflammatory bowel disease(IBD)is characterized by a dysregulated intestinal immune response, initiation and perpetuation of inflammation in experimental colitis by mechanism has not been known. In our preliminary study, we found that bone marrow derived CD11c-miR155KI mice dendritic cell(DC) which cultured in vitro showed spontaneous maturation compared to WT, and increased ability to stimulate T cell activation and proliferation, the ratio of CD11bhiDC subset in the spleen and mesenteric lymphoid node was dramatically decreased. CD11c-miR155KI mice showed a substantial weight loss and its colitis inflammation was more servious in the process of IBD. Accordingly we made the scientific hypothesis as follows: Because of over-expression or deficient of miR155, intestinal DC can be reprogrammed a new DC subset, intracellular molecules in DC will be alterated, these reactions will result in that the ability of DC polarizing T cell response will be changed. In this study, we will do experiments with CD11c-miR155KI or miR155-deficient mice model to address the role of miR155 in the development and function of enteric DC by virtue of modern immunological techniques. Further study will confirm the effect of enteric DC subsets which are different with the change of miR155 expression on the development IBD as a colitis model. The study will contribute to understand deeply the role of intestinal DC which regulated by miR155 expression in IBD, to elucidate the underlying pathogenesis of autoimmue diseases such as IBD. It will provide a protential and theoretical basis for exploiting new immunoregulatory drug in the future.

炎症性肠病（IBD）是免疫相关的肠道炎症疾病，发病机制尚不清楚。研究表明microRNA155（miR155）表达变化可调控树突状细胞（DC）的功能，我们的预实验发现DC选择性miR155过表达转基因鼠（CD11c-miR155KI）与野生型小鼠相比，IBD发病时炎症明显加重。据此推测：DCmiR155的表达变化可使肠道DC亚群"重新编程"，内部信号分子发生改变，促使DC在IBD发病中介导T细胞应答类型发生变化，最终导致疾病发生。本项目以现代免疫学方法结合CD11c-miR155转基因和基因缺陷鼠模型为实验平台，深入探讨IBD发病中miR155调控肠道DC亚群的分子机理和信号通路，分析肠道DC亚群miR155表达变化对IBD发病的作用机制。本研究有助于深刻理解miR155调控的肠道DC亚群在IBD中的作用，以阐明炎症性肠病等自身免疫性疾病的发病机理，为开发出新型免疫调节药物提供理论依据。

炎症性肠病（Inflammatory bowel disease，IBD）包括溃疡性结肠炎（ulcerative colitis, UC）和克罗恩病（Crohn’s disease, CD），已成为我国消化系统常见疾病和慢性腹泻的主要原因，长期炎症性病变可导致结肠癌的发生，因此日益受到重视。本项目原计划围绕“肠道DC亚群miR155表达变化对IBD发病的影响及调控机制”这一科学问题开展研究”但由于肠道LP中DC数量较少，导致一些实验开展困难，同时发现肠道巨噬细胞的数量也发生了变化，因而课题组适时对一些研究内容进行了调整，并通过一些细胞系进行体外实验，探讨了稳态下和DSS诱导IBD疾病过程中miR155的表达变化对小肠LP中DC亚群发育的影响及其调控机制，以及巨噬细胞与树突状细胞在死细胞相关抗原诱导的免疫反应中的相互作用机制，以期对将来临床干预炎症性肠病的机制研究以及临床治疗提供理论依据。研究成果总结如下：.目前认为肠粘膜的免疫反应失控是IBD的特征性表现，但其发生的具体免疫调控机制目前尚不清楚。本研究应用miR155基因缺陷鼠探讨miR155表达水平的变化对小鼠肠道DC亚群的发育的影响，并利用IBD 疾病模型验证miR155缺陷对调控的DC亚群在疾病发生、发展过程中作用及调控机制。我们发现：并且在DSS诱导的IBD模型中，与WT小鼠相比，miR155缺陷鼠肠炎症状减轻，且小肠LP中CD103+CD11b+DC数量减少约40%-50%，但是Annexin V和Ki67分析细胞的凋亡和增殖没有明显变化，此结果表明miR155参与了肠炎的发病，且miR155缺失可影响CD103+CD11b+DC亚群的发育。进一步研究发现：在DSS诱导IBD模型中，与WT相比，miR155缺陷鼠的小肠LP中CD103+CD11b+DC的IRF4和Notch2 mRNA表达水平明显下调。此结果进一步显示IBD发病过程中miR155参与了CD103+CD11b+DC亚群的发育。为确定是IBD炎性环境影响了LP中CD103+CD11b+DC亚群数量发生变化，还是此群细胞的发育自身发育就收到miR155的调控进而影响IBD的发生和发展，因此，我们检测了稳态下miR155缺失对LP中CD103+CD11b+DC亚群发育的影响，结果发现：稳态下，miR155缺失导致小鼠小肠LP中CD103+CD11b