HIF-1/TWIST1信号通路在病理性视网膜新生血管中的作用机制

Retinal vascular diseases are characterized by pathologic neovascularization, which could be initiated by elevated hypoxia-inducible factor-1 (HIF-1). Previous research found that conditional knockout of Twist1 in endothelial cells significantly decreased neovascularization in mouse model of oxygen induced retinopathy (OIR). TWIST1 was also found directly regulated by HIF-1 in promoting tumor angiogenesis. Therefore, we hypothesize that a HIF-1-mediated TWIST1 signaling pathway might play an important role in retinal neovascularization. In our previous study, we found that several angiogenic factors were potential target genes of TWIST1. In this study, we plan to verify these target genes of TWIST1 by chromatin precipitation and luciferase report assays; then, to investigate the role of HIF-1/TWIST1/GeneX signaling pathway in regulating of angiogenesis of human retinal vessel endothelial cells by manipulating HIF-1 and TWIST1 gene expression. Finally, the therapeutic effective of retinal neovascularization in mice will be evaluated by knocking down Twist1. Our results hopefully benefit our understanding of the mechanism of retinal neovascularization, which has a profound theoretical and clinical significance for retinal vascular diseases.

病理性视网膜新生血管是视网膜新生血管相关性疾病的基本病理特征，缺氧诱导因子-1（HIF-1）水平升高是其始动因素。既往发现TWIST相关蛋白1（TWIST1）缺失可显著抑制缺氧诱导的小鼠视网膜新生血管生成。而HIF-1可通过调控TWIST1的转录水平参与肿瘤新生血管生成。据此，我们推测HIF-1介导的TWIST1相关信号通路也在视网膜新生血管生成中起重要作用。本研究前期发现TWIST1的多个潜在靶基因（GeneX）编码促血管生成活性因子或受体，现拟进一步通过染色质免疫沉淀及荧光素酶报告实验等手段鉴定小鼠TWIST1下游靶基因；再经体外正负向调控干预，证实HIF-1/TWIST1/GeneX介导促人视网膜血管内皮细胞增殖及成血管活性；随后在体内探索以TWIST1为负向干预靶点抑制小鼠视网膜新生血管生成的作用。研究成果有望进一步揭示视网膜新生血管生成的分子机制，具有重要理论及临床意义。

病理性视网膜新生血管是视网膜新生血管相关性疾病的基本病理特征，既往发现TWIST相关蛋白1（TWIST1）缺失可显著抑制缺氧诱导的小鼠视网膜新生血管生成，且在病理新生血管中TWIST1表达增加。为验证TWIST1在视网膜病理新生血管生成中的作用，我们在分别在人视网膜血管内皮细胞和小鼠视网膜内皮细胞中过表达了TWIST1。并通过视网膜铺片，冰冻切片免疫荧光等技术对其表型进行了验证。我们的数据表明TWIST1在视网膜血管新生中发挥着关键作用，过表达TWIST1后小鼠视网膜血管发育变缓，血管通透性增加。通过RNAseq、Western Blot等实验对其作用机制进行了探索，揭示了其通过影响PI3K-AKT等信号通路影响了视网膜血管的发育。研究成果有望进一步揭示视网膜新生血管生成的分子机制，具有重要理论及临床意义。