AIM2基因与汉族人银屑病发病机制相关性研究

Psoriasis is an autoimmune disease caused by combined action of genetic and environmental factors which is difficult to effect a radical cure and affects physical and mental health of patients seriously. Our previous study identified a SNP rs2276405 within AIM2 gene significantly associated with psoriasis in Chinese Han population(stop-gained, P=3.22×10-9) and was predicted to impact protein structure. AIM2, a receptor for cytosolic DNA, can form AIM2 inflammasomes activated by cytosolic DNA to mediate immune responses. The object of this study is to find all the causal variants of AIM2 by sequencing the whole length of this gene. Meanwhile, we plan to explore the function of AIM2 deeply in the level of transcription, expression, cells and in vivo animal model with RT-PCR, siRNA interference, luciferase assay, chromatin immunoprecipitation, electrophoretic mobility shift assay, protein subcellular location, epigenetic modification as well as AIM2 transgenic and knock-out animal models to reveal the role of AIM2 gene on the pathogenesis of psoriasis. It will provide theoretical basis for predicting disease risk as well as identifying susceptible population and genetic targets, which will lay a good foundation for prevention, diagnosis, therapy and antenatal diagnosis of psoriasis and drive the process of translational medicine.

银屑病俗称“牛皮癣”，是一种由遗传、环境和共同介导的自身免疫性疾病，难以根治，严重影响患者身心健康。申请人前期发现AIM2基因功能性编码变异rs2276405（Stop-gained, P=3.22×10-9）与汉族人银屑病显著关联，功能预测提示该变异为有害变异。AIM2为胞质DNA传感器，感知细胞质内DNA，诱导炎性复合体形成，参与免疫反应。本课题拟基于前期研究，对AIM2基因进行全基因及调控区域测序，搜寻AIM2基因所有致病变异。借助RT- PCR 技术、siRNA干扰技术、荧光素酶实验、染色体免疫沉淀实验、EMSA试验、蛋白亚细胞定位、表观修饰以及转基因和基因敲除动物模型在转录、表达、细胞及动物模型等不同层面开展功能研究，揭示AIM2基因在银屑病发病过程中的作用，为确定易感人群，遗传靶标及风险预测提供理论依据，进而为疾病预防，临床诊疗及遗传咨询、产前诊断奠定基础，推动转化医学进程。

本课题是基于前期的遗传学研究发现，首先对AIM2基因功能性变异进行寻找和验证。通过两轮11245例银屑病人和11177例健康人样本外显子芯片分析，发现AIM2蛋白的两种突变对健康人群具有保护作用，体外实验表明该突变引起AIM2形成炎症小体能力降低。随后对临床样本和，通过实时荧光定量定量PCR、western、免疫组化、免疫荧光共标、组织芯片和表达谱芯片检测表明AIM2介导的炎症小体途径在银屑病人中被激活。我们又建立了AIM2通路激活的银屑病小鼠模型，并对多个相关指标进行了检测。后面我们对该通路的多个靶点进行了干预实验，包括使用刺激物、基因敲降和中和等方法和手段对通路进行加强和减弱效应，均发现该通路调节了银屑病重要相关细胞因子il17的分泌，表明该通路在银屑病的发病中具有重要调控作用。我们又建立了角质细胞和免疫细胞互作模型，在细胞水平验证了AIM2通路在银屑病中的激活机制。对健康皮肤样本和银屑病人的皮损组织的DNA甲基化水平进行分析，结果表明皮损组织AIM2通路的多个基因的启动子甲基化水平显著降低，表明相关基因被激活。.本课题从细胞、动物和临床样本，体外表达纯化蛋白、mRNA和蛋白水平，借助激光共聚焦显微镜、流式细胞术、表达谱芯片、甲基化芯片和组织芯片等多种技术手段在不同水平和层次解析了AIM2通路激活在银屑病发病中的作用及机制。结果表明AIM2为银屑病的重要正向调控基因。，揭示AIM2基因在银屑病发病过程中的作用，为确定易感人群，遗传靶标及风险预测提供理论依据，进而为疾病预防，临床诊疗及遗传咨询、产前诊断奠定基础，推动转化医学进程。