基于青光眼类器官模型研究CREB/c-Fos通路在视神经损伤与修复中的作用及机制

高眼压是青光眼视神经损伤的最重要危险因素，缺乏自然动物模型是研究该损伤机制的主要障碍之一，我们在前期研究中创新性地对由人诱导多能干细胞分化的视网膜类器官加压处理，构建出一种高度模拟眼压升高环境的人青光眼类器官模型。基于此模型，通过RNA-seq测序筛选了一个关键基因c-Fos。加压后，c-Fos表达上调，视神经轴突出现损伤；撤去压力后，c-Fos表达显著下降，损伤的轴突呈现部分修复；同时，c-Fos抑制剂能使损伤轴突再次延伸。通过数据库分析，表明c-Fos可能受上游基因CREB转录调控。为此，本项目拟体外通过细胞原代培养、RNA干预、ChIP等实验证明CREB转录调控c-Fos；体内通过小鼠硅油诱导青光眼模型验证其对视网膜功能的影响。该研究不仅揭示高眼压下CREB调控c-Fos转录表达的具体分子机制，而且阐明其在视神经损伤及修复再生中的作用，为青光眼视神经修复与再生治疗提供新靶点。

High intraocular pressure is the most important factor contributing to glaucomatous optic nerve injury. Lacking natural animal glaucoma model is the main obstacle concerning to understanding mechanism of optic nerve injury. In our previous studies, we innovatively treated the HiPS-induced retinal organoid with pressure, constructing a human glaucomatous organoid model which highly simulates the environment of IOP elevation. Based on this model, we performed RNA-seq to screen out a key gene c-Fos. Under hypertension, elevated c-Fos expression induced axonal injury. While pressure was removed, c-Fos expression decreased significantly, damaged axons appeared partial repaired. In addition, c-Fos specific inhibitor was able to extend broken axons. By database analysis, c-Fos expression may be regulated by the transcriptional factor CREB. Thus, this project will in vitro perform primary cell culture, RNA intervention, ChIP and other experiments to prove the role of CREB on c-Fos transcriptional regulation; in vivo verify its effects on retinal function through murine silicone oil-induced glaucomatous model. This study not only reveals the specific molecular mechanism of CREB regulation on c-Fos transcriptional expression under high intraocular pressure, but also clarifies its role in optic nerve injury and nerve repair, which provides a new target for regeneration of glaucomatous optic nerve.