硫化氢对缺血导致视网膜损伤和钙超载的作用及其机制

Retinal ischemia is a serious pathological damage, which induced apoptosis in retinal ganglion cells(RGC)，and the injury of RGCs directly cause blindness of paients. Hydrogen sulfide(H2S) was consided as a newly discovered gas signaling molecules, some research has shown that H2S reduced the injury of retina caused by ischemia. Our preliminary study has shown that the hydrogen sulfide synthase CBS existed in the retina of rats and H2S inhibited the calciun overload in RGCs caused by NMDA, also reduced the damage of ischemic retina. However, the existence of endogenous hydrogen sulfide in the retina and the role of hydrogen sulfide reduced the injury of ischemia retina are unknown. Accordingly, we intend to design the experiments to prove the existence of endogenous hydrogen sulfide in the retina first, and then find the evidence that Exogenous and endogenous hydrogen sulfide could inhibit the calcium overload through the activatiion of the ATP sensitive potassium channel and further more supress the injury of RGCs using a variety of experimental techniques,such as electrophysiological technique (Patch Clamp), calcium imaging and molecular biology technology. The research results is not only help to further clarify the physiological function of H2S in retina, but also first reveal the protective role of hydrogen sulfide in retina injured by ischemia , and provide an important basis for the pathological changes and treatment of ischemic retina.

视网膜缺血是临床常见的病理损伤，可引起视网膜神经节细胞凋亡，导致病人失明。硫化氢是新发现的一种气体信号分子，已有研究证实硫化氢对神经元缺血损伤具有保护作用。我们预实验发现正常大鼠存在硫化氢合成酶CBS，并且硫化氢可减轻缺血导致的视网膜损伤，抑致兴奋性中毒导致的视网膜神经节细胞钙超载，但视网膜上是否存在内源性硫化氢，以及硫化氢减轻缺血视网膜的具体机制还不清楚。申请人以此为依据，拟设计实验首先证明视网膜上是否存在内源性硫化氢，然后建立在体和离体视网膜缺血模型，利用膜片钳、钙成像及分子生物学等多种实验技术进一步证明外源性和内源性硫化氢能否激活视网膜神经节细胞上的ATP敏感钾通道，进而抑制缺血视网膜神经节细胞钙超载并减轻其损伤。研究结果不仅进一步揭示硫化氢在视网膜中的生理功能，而且首次揭示硫化氢在视网膜缺血损伤及保护中的作用及其机制，为探讨视网膜缺血的病理变化和治疗提供重要实验和理论依据。

视网膜缺血是临床常见病理生理过程，其中视网膜神经节细胞损伤可导致不可逆性视力缺失。硫化氢是新发现的一种神经调质，可提高谷胱甘肽浓度保护氧化应激导致的RGC-5损伤，还可通过激活KATP通道逆转缺血导致的HT22细胞损伤。我们的研究设想硫化氢是否可以通过激活离体或在体缺血视网膜节细胞上的KATP通道减轻其损伤。我们的结果显示（1）视网膜上存在CBS酶并能合成硫化氢；（2）外源性和内源性硫化氢可减轻离体或在体缺血视网膜节细胞钙超载，降低细胞死亡率；（3）利用膜片钳技术，我们发现硫化氢可通过激活KATP通道降低视网膜节细胞膜电位和兴奋性，减轻其缺血损伤；（4）视网膜节细胞上还存在Kv7.4通道，其激活也可导致细胞膜超极化，降低其兴奋性。这些结果提示钾通道调节细胞兴奋性可做为保护缺血视网膜节细胞免受损伤的药物作用靶点和治疗方案。