从CD40多态调控DC角度探讨Behcet病的发生及其机制

Behcet's disease (BD) is a multi-systemic autoinflammation disease mainly mediated by genetic predisposition, infections and immunological abnormalities. However the mechanism is still undetermined. Recently we showed that CD40 was identified as a susceptibility gene of BD in a Han Chinese population. It was reported that rs1883832 SNP in CD40 gene controlled CD40 expression. Besides the priming of T cells with its ligand, CD40 controlled the activation of macrophages and dendritic cell (DC) and contributed to initiation of innate immune responses as well as the subsequent induction of adaptive immune responses. Therefore, we propose that CD40 polymorhpims control CD40 expression, subsequent DC activation and contribute to the BD development. To test this hypothesis, we determine to study the role of CD40 polymorphism in activation of DC in the development of BD. The project mainly includes 4 parts as follow. (1) Expression of CD40 will be investigated to study influence of rs1883832 variant on CD40 translations. (2) Phenotype, mature and functional properties of DC will be explored to determine the role of CD40 in DC activation. (3) Activation status and interaction of Btk with CD40 and major histocompatibility complex (MHC) class II molecules, binding of Btk with MyD88 and TRIF, and effect of Btk on the activation of a reporter for NF-κB or IRF3 will be determined to study the underlying mechanism of CD40 on the activation of DC in Behcet's diseases. (4) The incidence and severity of experimental autoimmune uveitis (EAU),phenotype,mature and functional of DC, and freqencies of Th17 and Treg will be assessed to study the role of CD40 deficiency in EAU. These studies are expected to gain a deep insight into the initiating mechanisms involved in BD from DC and to provide a new strategy in the prevention and treatment for this disease.

Behcet病是一种多系统、多器官受累的自身炎症性疾病，遗传易感、感染和免疫紊乱是其致病的核心要素，但具体机制并不完全清楚。最近我们发现CD40 是Behcet病易感基因，文献报道rs1883832变异影响CD40蛋白翻译。除与其配体作用激活T细胞外，CD40激活巨噬、树突状细胞(DC)而触发天然和适应性免疫应答。据此推测CD40多态可能调控该蛋白表达及后续DC分化、成熟和功能而始动疾病发生。为验证此假设，本研究以rs1883832多态为切入点，以Behcet病为研究对象，探讨该多态对CD40蛋白及后续DC分化、成熟和功能影响；通过评价TLR信号中Btk活性及其与CD40、MHC-Ⅱ结合状态及Btk对MyD88、TRIF、NF-κB和IRF3激活的影响，探讨该多态影响DC的机制；探讨干预此因素对葡萄膜炎动物模型防治作用。旨在从DC层面找到诱发Behcet病始动因素，为其防治研究提供新策略。

Behcet病是一种多系统、多器官受累的自身炎症性疾病，遗传易感、感染和免疫紊乱是其致病的核心要素，但具体机制并不完全清楚。前期我们和其它研究发现，CD40基因是Behcet病易感基因。CD40蛋白可通过激活巨噬、树突状细胞(DC)而触发天然和适应性免疫应答。在Behcet病中CD40是否通过DC细胞参与Behcet病的发生及其参与机制尚不清楚。基于上述背景，在国家自然科学基金委的支持下，我们进行了以下两方面研究：一方面，通过与CD40基因相关的NF-κB途径（REL，PRKCQ）基因多态与中国汉族Behcet病、Vogt-小柳原田综合征相关性研究，发现REL rs842647与我国Behcet病的发生以及皮肤损伤有关，未发现PRKCQ基因的rs947474、rs4750316、rs11258747三个位点与我国Behcet病相关；发现PRKCQ基因与我国Vogt-小柳原田综合征的易感性相关，未发现REL基因与我国Vogt-小柳原田综合征相关；揭示出REL基因可能是我国汉族Behcet病患者的易感因素，PRKCQ基因可能是我国汉族Vogt-小柳原田综合征患者的易感因素。另一方面，我们通过光感受器维生素A类结合蛋白IRBP161-180肽段诱导B10.RIII小鼠建立了实验性自身免疫性葡萄膜炎小鼠模型和复发性实验性自身免疫性葡萄膜炎小鼠模型，探讨了CD40参与葡萄膜炎发生的作用及其机制，并拓展性地研究了免疫抑制中成药火把花根对EAU葡萄膜炎的预防和治疗作用及其机制；总结出EAU和复发性EAU葡萄膜炎发生、临床表现以及病理学特征，发现EAU和复发性EAU葡萄膜炎的病理特征与人类葡萄膜炎病理特征相似，复发性EAU葡萄膜炎具有与人类慢性复发性的病程；发现CD40通过抑制DC细胞分泌促炎性因子和对淋巴细胞增殖反应减轻葡萄膜炎的炎症严重程度，发现火把花根通过抑制DC细胞中CD40以及抑制DC细胞功能发挥其预防和治疗作用，揭示出CD40参与葡萄膜炎的作用机制与途径，为葡萄膜炎的治疗提供新的治疗方法与策略。通过上述研究，我们完成了本项目的研究任务，发表SCI收录期刊论文1篇（PLoS ONE）和CSCD收录期刊论文1篇（已接受，第三军医大学学报），此外还形成论文拟投稿SCI期刊论文2篇。参加学术会议6次，培养副研究员1名，硕士研究生2名。