microRNA在IFN-γ上调角膜上皮细胞MICA分子表达中的作用机制研究

角膜排斥反应是角膜移植术后失败的主要原因。传统观点认为经典MHC抗原分子是参与角膜移植排斥反应的主要免疫分子。我们最近发现：人类角膜上皮细胞使用干扰素IFN-γ处理后表达非经典MHC分子MICA分子显著上调，并且NK细胞对角膜上皮细胞的细胞毒性作用显著增强。最近有学者报道了IFN-γ可以通过上调miRNA-520b下调肿瘤细胞表达MICA分子。这与IFN-γ上调角膜细胞MICA表达正好相反，但后者机制尚不明确。在本研究中，我们拟通过microRNA芯片、流式细胞仪、RT-PCR检测等方法进一步明确IFN-γ处理角膜细胞后microRNA的差异性表达；结合已有文献及数据库筛选出候选microRNA；检测增强及减弱候选microRNA情况下MICA的表达规律，并探讨MICA介导的细胞毒性作用的变化情况，从而明确IFN-γ上调角膜细胞表达MICA的机制，为揭示角膜排斥反应的发病机制提供新的思路。

Though keratoplasty is the most successful transplantation, corneal graft rejection is still the most frequent complication after corneal grafting and often leads to irreversible transplant failure. It is widely accepted that MHC class I and II antigen play crucial role in this rejection. In our recent study, we revealed for the first time that the proinflammatory cytokine, IFN-γ promoted surface MHC class I-related chain A (MICA) expression on corneal epithelium in a dose-dependent manner. IFN-γ also enhanced NK cell-mediated cytotoxicity against the corneal epithelium. On the contrary, Yadav et al. found that one of microRNAs, miR-520b was induced by IFN-γ, leading to a reduction in MICA surface protein levels. miR-520b acted on both the MICA 3'-untranslated region and the promoter region and caused a decrease in the levels of MICA transcript. The exact mechanism involved in the increased expression of MICA on corneal epithelium induced by IFN-γ is still unknown. In the current study, we will explore the changes of microRNAs in the human corneal epithelial cells upon IFN-γ treatment by Agilent whole human genome microarray analysi, flow cytometry, and Invitrogen TaqMan miRNA test. To identify miRNAs that regulate the expression of MICA, we will use the Sanger MicroCosm search engine (microrna.sanger. ac.uk), Gene Ontology (GO) analysis, and microRNA-gene-network analysis. We will then assess the MICA transcript and MICA-mediated cytotoxicity against the corneal epithelium by up-regulation or knock-down of the potential microRNA. Regulation of MICA on human corneal epithelium by IFN-γ-induced microRNAs may lead to a new vision into the pathogenesis of corneal graft rejection.