HDAC2通过c-Jun/PAI-1信号通路调控炎症因子的表达促进I型麻风反应发生的研究

Leprosy type I reaction (TIR) is a hypersensitivity reaction to M. leprae antigen, is the leading cause of disabilities and deformities in leprosy. Applicant found the gene HDAC2 was highly expressed in the skin lesion of TIR patients. Previous studies have shown that HDAC2 plays an immunoprotective role by blocking the c-Jun/PAI-1 signaling pathway to block bacterial lipopolysaccharide-induced inflammatory responses. The mutations in the upstream signaling pathway of this gene may cause the body fail to finish the above immune response in TIR. However, no experimental evidence is available to support these implications. The applicant further intends to study the gene expression by immunohistochemistry and use cytology experiments to study the effect of this gene on macrophage, Schwann cell associated inflammatory factors and c-Jun/PAI-1 pathway in vitro. Furthermore, knock out of zebrafish Hdac1 by CRISPR/Cas9 and rescue with human HDAC2 mRNA will be performed in mycobacterium marinum infected zebrafish to study the effects in vivo. This study will reveal the molecular pathogenesis of HDAC2 in TIR and will also provide suggestions for the risk prediction and intervention strategy of leprosy.

I型麻风反应（TIR）是机体对麻风分枝杆菌抗原产生的一种超敏反应，可导致患者出现神经损伤，这是麻风患者畸残的主要原因。申请人发现HDAC2基因在TIR患者皮损中高表达。既往研究表明HDAC2通过调节c-Jun/PAI-1信号通路阻断细菌脂多糖诱导的炎症反应发挥免疫保护作用，该基因上游信号通路相关变异的发生可能会导致机体无法发生上述免疫应答致TIR的发生，但具体免疫学机制有待进一步证实。因此，申请人下一步拟采用免疫组化等技术进一步明确该基因在病例中的表达情况；通过体外细胞感染模型研究该基因对巨噬细胞、施旺细胞相关炎症因子及c-Jun/PAI-1通路的影响；同时利用基因敲除斑马鱼模型在体内条件下探讨该基因在TIR炎症反应中的具体机制，为麻风畸残的风险预测与早期干预提供理论依据。

I型麻风反应(TIR)是机体对麻风分枝杆菌抗原产生的一种超敏反应，可导致患者出现神经损伤，这是麻风患者畸残的主要原因。HDAC2基因在TIR患者皮损中高表达。课题组在前期研究基础上，按照项目实验计划进行了相关研究。首先，课题组利用完成了THP-1细胞的培养及细胞HDAC2基因表达的检测，利用qPCR技术检测结果显示HDAC2基因明显表达升高，其表达水平随着加入菌的MOI水平升高而升高，进一步证实了HDAC2基因与分枝杆菌感染的相关性。然后构建了稳定过表达HDAC2基因的THP-1细胞，深入探索HDAC2基因在分枝杆菌感染过程中的作用，其表达的蛋白产物在N端融合了绿色荧光蛋白。利用荧光标记的海分枝杆菌感染上述细胞系，流式细胞实验发现HADAC2基因的敲除和过表达并不影响巨噬细胞的杀菌能力，利用流式细胞术观察HADAC2对于T细胞亚群的影响。通过以上系列研究，初步阐释了HADAC2基因在分枝杆菌感染肉芽肿形成中的作用机制。