TNFR2内化修饰在糖尿病视网膜病变中的作用及分子机制

The formation of retinal neovascularization(NV) arises from the dysfuction of endothelial cells (EC). Tumor necrosis factor receptor 2(TNFR2）, which is an EC-specific cytokine and a prognostic biomarker of diabetic complication, plays an important role in ischemia-mediated cell survival, migration, proliferation, arteriogenesis and angiogenesis.In our previous study, TNFR2 can also elicit cell apoptosis signaling pathway in endothelial cells, which dependented on TNFR2 internalized .The internalization of TNFR2 highly expressed in EC, which was cultured in high glucose.Suggesting TNFR2 displays internalization and EC function switching. However, the function and molecular mechanism of TNFR2 internalization in diabetic retinopathy is largely unknown. Based on our previous finding, in this project, we will investigate internalized modification regulation mechanism of TNFR2 domains via double-Immunofluorescence assay, ChIP assay and DNA methylation analysis. Moreover, we will determine the function of TNFR2 internalization in diabetic retinopathy in vivo and intro by gene gain-of-function and loss-of function strategy,and explore molecular mechanism. Furthermore, we will confirm expression and internalized modification of TNFR2 domains in retinal neovascularization tissues of diabetic retinopathy via isoform-specific antibodies. This project will ont only provide a novel insight into elucidating the function and regulation of TNFR2 internalized modification in diabetic retinopathy, but provide important theoretical foundation for identifying molecular marker in The formation of retinal neovascularization and developing specific-target intervention.

新生血管形成源于血管内皮细胞（EC）功能改变，肿瘤坏死因子受体2（TNFR2）作为EC特异性细胞因子和判断糖尿病并发症预后的生物标记，在缺血诱导的EC迁移、增殖、血管生成和再生中具有重要作用。我们前期研究发现，TNFR2还具有诱导EC凋亡的新功能。该功能依赖于TNFR2内化，并受高糖调控，提示TNFR2存在内化修饰和对EC功能的转换，但其内化调控机制及在糖尿病视网膜病变（DR）中的作用尚未明确。本课题拟在前期研究基础上，通过双重免疫荧光定位染色、定量染色质免疫沉淀技术、基因功能获得和功能缺失策略，从分子-细胞-动物-人群水平分析TNFR2内化修饰介导EC功能转换的分子及调控机制，探讨其在DR发展中的作用；揭示TNFR2内化修饰的分子调控机制及在DR发展中的作用机制；验证TNFR2内化在不同分期的DR玻璃体组织中的表达。本研究将为发现新生血管形成的新分子标记和开展特异性靶向干预提供新策略。

新生血管形成源于血管内皮细胞（EC）功能改变，肿瘤坏死因子受体2（TNFR2）作为EC特异性细胞因子和判断糖尿病并发症预后的生物标记，在缺血诱导的EC迁移、增殖、血管生成和再生中具有重要作用。我们的研究发现，TNFR2还具有诱导EC凋亡的新功能。该功能依赖于TNFR2内化，并受高糖调控，提示TNFR2存在内化修饰和对EC功能的转换，高糖条件下，TNFR2去内化结构域TNFR2-59失活，Akt/NF-κB信号通路关闭，EC存活、增殖受到抑制，TNFR2内化增加，内化结构域TNFR2-84介导的AIP1-JNK死亡信号通路激活，发挥EC凋亡的作用我们发现结构域 TNFR2-59主要在细胞膜上表达，而TNFR2-84可由细胞膜上内化到胞质内表达，表明TNFR2-84有其特定的信号通路。AIP1-JNK信号表达的变化和TNFR2内化转录功能的关系密切，TNFR2／ASK1信号通路在视网膜内皮细胞的生长和迁移中发挥重要作用，缺血损伤可以激活视网膜血管内皮细胞TNFR2，TNFR2激动剂对于视网膜血管内皮细胞具有保护作用。本课题相关研究发表SCI论文11篇，结果将为发现新生血管形成的新分子标记和开展特异性靶向干预提供新策略。