ATF4信号通路激活致小梁网内皮细胞凋亡及眼压升高机制研究

Glaucoma is the second leading cause of irreversible blindness in China and in the world. Primary open-angle glaucoma (POAG) is the most common type. Endoplasmic reticulum (ER) stress has been found to play an important role in the pathogenesis of trabecular meshwork cells dysfunction, apoptosis and then increased intraocular pressure (IOP) in POAG. Activating transcription factor 4 (ATF4) is the major ER stress transcription factor that induced the expression of ER apoptotic protein-CHOP and has been found to mediate the production of multiple inflammatory cytokines. This project will focus on ATF4 signaling pathway as the following: (1) To investigate the inflammatory factors expression and cell apoptosis after over-expression of ATF4 in cultured trabecular meshwork endothelial (TME) cells; to investigate the inflammatory and apoptotic response after inhibition of ATF4 expression in the oxidant treated TME cells; (2)To investigate the expression of inflammatory cytokines, viability of TME cells, and IOP level after over-expression of ATF4 in the trabecular meshwork of mice; to investigate the inflammatory response of trabecular meshwork and IOP level after inhibition of ATF4 expression in the Tunicamycin induced glaucomatic model of mice. Our study will help further dissect the mechanisms of ER stress in the TME cells injuries and dysfunction in POAG. And it may provide critical information for identification of new therapeutic approaches other than lowering IOP or neuroprotective drugs, that is, to prevent or delay the loss of function of trabecular meshwork in POAG patients.

原发性开角型青光眼（POAG）是青光眼中最常见的致盲类型。已发现内质网应激在POAG小梁网细胞功能障碍、凋亡，以致眼压升高中起着重要作用。ATF4是诱导内质网凋亡蛋白CHOP产生的主要转录因子，并可能介导多种炎症因子的产生。本项目以ATF4为轴线进行以下研究：(1)培养小梁网内皮(TME)细胞，探讨ATF4表达上调后炎症因子的表达及细胞凋亡变化；在TME细胞的氧化应激模型中研究抑制ATF4表达后细胞炎症反应及活性改变；(2)体内实验探讨小梁网组织ATF4表达上调后炎症因子的表达、TME细胞的存活，以及眼压改变；在Tunicamycin诱导的小鼠高眼压模型中，探讨ATF4表达下调后小梁网组织炎症反应，以及眼压改变。该研究将有利于进一步阐明内质网应激在POAG小梁网细胞损伤及功能障碍发生中的具体机制，为寻找除了降眼压药以外新的POAG治疗手段、预防或延缓POAG小梁网功能的丧失带来全新的认识。

青光眼是常见的不可逆性致盲眼病。原发性开角型青光眼（POAG）是青光眼中最常见的致盲类型。POAG的发病机制至今尚未阐明。已有研究提示，小梁网细胞功能障碍、死亡，从而导致房水外流阻力的增加、眼压的升高，在POAG的发病中起着关键作用。引起小梁网细胞功能障碍的分子生物学机制尚不清楚。氧化应激是首要危险因素之一。同时，氧化应激可破坏内质网的氧化还原环境，引发内质网应激。而氧化应激和内质网应激在小梁网细胞功能障碍所起到的相互作用及细胞损伤机制未见报道。本研究旨在探讨内质网应激信号通路的主要转录因子——ATF4在氧化应激引起的小梁网内皮细胞功能障碍、细胞损伤中所起的作用，进一步揭示内质网应激信号通路激活引起小梁网细胞炎症、凋亡的具体作用机制，以期为阐明POAG小梁网细胞功能障碍、凋亡，以致眼压升高的发生机制提供思路。.本研究的主要结果包括：（1）在小梁网细胞的氧化应激模型中，叔丁基过氧化氢（TBHP）激活内质网应激通路蛋白，诱导ATF4活化，引发细胞凋亡；（2）ATF4是氧化应激诱导小梁网细胞炎症和细胞凋亡的重要介质。下调ATF4的表达后，TBHP诱导的炎症因子ELAM-1和IL-8的表达明显下降，伴有细胞凋亡的减轻；（3）ATF4介导内质网应激诱发的小梁网细胞炎症及氧化损伤。（4）ATF4参与小梁网细胞吞噬功能的调节。（5）ATF4表达上调足以引起小梁网细胞炎症因子表达的升高及诱发细胞凋亡。上述结果提示，内质网应激及ATF4信号通路在小梁网细胞功能障碍及细胞凋亡中起着关键性的作用。本研究结果有利于进一步阐明POAG小梁网内皮细胞凋亡、以致眼压升高的发生机制，为研究新的延缓或预防小梁网细胞凋亡的抗青光眼药物提供新的思路。