单卵双胞胎银屑病患者de novo突变的细胞模型验证及T细胞功能/表观遗传研究

Psoriasis is an immune disorder occurred in genetic background. Monozygotic twin is the most ideal object for study of genetic disease. We have obtained lots of progress in our previous studies on psoriatic T cells (including twins). Recently, it was found that genetic variation in MHC proteins is associated with T cell receptor (TCR) expression biases. In recent years, we have collected 10 of pairs monozygotic twins with more than 10 years of psoriasis history. After collaborative research with The Beijing Genomics Institute, we found de novo mutations and different expression in TCR Vαand Vβbetween twins. Lately, the peripheral blood from the patients of ten pairs twins was measured again to carry on the whole genome sequencing, which verify the existence of de novo mutation. The important progress have been made, so we proposed to study on the following research on previous basis: (1) construct cell model by knocking out de novo mutant genes in order to further validation the mutant at cellular level; (2) investigate and analyze the expression of mRNA, lncRNA and DNA methylation of CD4+ and CD8 + T cells; (3) investigate TCR RNA expression of CD4+ and CD8 + T cells, and analyze the expression combined with information of whole genome sequencing; (4) according to the results of this study and combing with the open psoriasis information database, the relevant information is comprehensive analyzed, as far as possible to find the link between de novo mutant and T cell immune function.

银屑病是在遗传背景下发生的免疫紊乱性疾病。单卵双胞胎是研究遗传疾病最理想的对象。前期我们已在银屑病（包括双胞胎）T细胞研究方面取得不少进展。新近有研究发现MHC基因变异与TCR 表达偏移有关。近年我们收集到10对病史10年以上的单卵双胞胎银屑病家系，经与华大合作研究，发现了de novo突变及差异表达的TCR V链。在此基础上，新近又完成了双胞胎双亲全基因组测序，结果验证了de novo突变的存在。研究已取得了重要进展，故拟在此基础上进行如下研究：①敲除de novo突变的基因构建细胞模型，细胞水平进一步验证；②分选CD4+、CD8+T细胞，进行mRNA、lncRNA及全基因组甲基化检测，并整合分析信息；③RNA 水平检测CD4+、CD8+ T细胞组库，结合前期的全基因组信息进行分析；④结合研究结果和开放的银屑病信息库进行综合分析，尽可能找出de novo突变与T细胞免疫功能间的联系。

银屑病是在遗传背景下发生的免疫紊乱性疾病。单卵双胞胎理论上携带有完全一致的基因，故患病不一致的单卵双胞胎是研究遗传疾病的最理想对象。在前期T细胞与间充质干细胞研究基础上，本项目首先对银屑病患病不一致的单卵双胞胎进行了全基因组测序、甲基化等贯穿组学研究，证实了de novo突变（特别是C3和SPRED1突变）在银屑病发病中的作用，并分析验证了HLA-C*07:02、TCRV与银屑病临床表型的相关性。在项目引申研究中，研究结果显示银屑病皮肤间充质干细胞存在差异表达circular RNA， circular RNA-miRNA-mRNA作用网络分析与基因敲除模型进一步证实了cirRNA在银屑病皮肤间充质干细胞活性异常中的作用。银屑病皮肤间充质干细胞：促进角质形成细胞增殖、分化、迁移和糖代谢能力增强，但促进角质形成细胞连接的能力降低；对T细胞增殖、细胞因子产生和T-bet、ROR-γt表达的抑制效应减弱；银屑病皮损来源的微血管内皮细胞的血管形成和迁移能力增高，增殖和能量代谢水平降低。课题为银屑病de novo突变的研究提供了理论依据，并进一步充实了银屑病间充质干细胞的研究。