7,8-二羟基黄酮保护视网膜神经元及促进轴突再生的实验研究及其机制探讨

Retinal ganglion cells (RGCs) death and axon degeneration caused by glaucoma and other retinal degenerative diseases，leads to irreversible visual loss .It is critical to find a neuroprotective drug that is safe, effective and can cross blood-retinal barrier. 7,8-dihydroxyflavone（7,8-DHF）is a recently identified small-molecule TrkB agonist. We have found that 7,8-DHF has potential to protect retinal neurons. Further study is needed to clarify whether 7,8-DHF has neuroprotective effects on injured retinal neurons and even promote axonal regeneration. The three-dimensional retinal culture system and rat optic nerve crush model are employed for observing neuron survival and axon regeneration after 7,8-DHF administration, additional gene therapy for enhance 7,8-DHF rescue efficacy, TrkB gene overexpression caused by in vivo electroporation is designed, and the possible mechanisms explored. Furthermore, we intend to demonstrate the drug permeability across the blood-retinal barrier, and compare the safety and validity of local and systemic administration. We try to provide a scientific evidence to find a new, safe and effective neuroprotective drugs for glaucoma and other retinal degenerative diseases.

青光眼等视网膜变性性疾病由于进行性的神经元凋亡和轴突变性，导致不可逆性视觉障碍乃至失明。因此，研发安全、有效、能通过血眼屏障的神经保护药物迫在眉睫。7,8-二羟基黄酮（DHF）是近来新发现的一种小分子TrkB受体激动剂，我们前期预实验发现7,8- DHF玻璃体腔注射给药后，可减轻视神经挫伤损害。本项目将在视网膜微植片三维立体培养和视神经挫伤模型两个层面，深入研究7,8- DHF对大鼠视网膜神经元的保护效应和轴突再生效果；结合TrkB受体阻断和过表达两种状态下的效果分析，多角度验证7,8-DHF通过激活TrkB受体发生级联反应影响AKT、ERK信号通路的机制假说；进一步在成年SD大鼠视神经挫伤模型上研究7,8-DHF局部及全身给药的安全性、有效性、血眼屏障的通透性，以期为视网膜变性性眼病寻找适宜的神经保护性药物提供科学依据。

本研究通过体内和体外实验观察了7,8二羟基黄酮对大鼠视网膜神经元的保护效应和轴突再生效果。取得结果有：（1）优化了视网膜移植片三维立体培养系统，在此基础上，观察到7,8-DHF能够促视网膜移植片的轴突再生，且在其促轴突再生过程中可以激活TrkB信号通路，而阻滞TrkB信号通路可以抑制7,8-DHF的促轴突再生作用；（2）采用大鼠视神经夹伤模型，分别经玻璃体腔和腹腔注射7,8-DHF，发现均能保护受伤的RGCs；进一步采用电穿孔辅助TrkB基因转染联合7,8-DHF玻璃体腔局部注射，可以提高7,8-DHF的RGCs保护效果。这些研究结果说明，7,8-DHF可以通过血眼屏障，进而激活TrkB信号通路，而发挥保护视网膜神经元、促进视神经轴突再生的作用。