Protective Mechanism Of Rho Kinase Inhibitor Y27632 On Acute Retinal Ischemia Reperfusion Injury In Rats

Objective To investigate the protective mechanism of Rho kinase inhibitor Y27632 on retinal ischemia reperfusion injury in order to provide a theoretical basis for the treatment of glaucoma optic nerve protection.Methods SD rats were randomly divided into normal group,ischemia-repeffusion group,saline group,Y27632 group.Animal models of ischemia-repeffusion were made.Twenty-four hours and 126 hours after the induction of ischemia-repeffusion,the rats were killed,retina were stained with HE or ADPase.Histolopathological change of the retina was examined and the thickness of retina wese measured.Twenty-four hours after the induction of ischemia-repeffusion,the rats were killed,the retinas were homogenized.The expression of Caspase-3 protein in retinas was detected by Western Blot.The expression of Bcl-2 mRNA were detected by Real-time PCR.Statistical analysis was performed by using SPSS 17.0 software for ANOVA follwed by LSD test,P<0.05 was considred to be significant difference.Results:1.HE staining suggests that in the normal control group,the retinal structure was clear and the structure of the three layer cells was orderly.Inner limiting membrane was smooth.24 hours after chemia-reperfusion,the thickness of retina increased,the inner and outer layer tissue was loose,the retinal ganglion cells,the inner and outer nuclear layer cells were obvious,and the retinal ganglion cells were decreased.After 168 hours,the retinal edema subsided,the thickness became thin,and the number of ganglion cells and inner and outer nuclear layer cells decreased,and capillaries in the retina and nerve fiber layer were found.2.Measurement of retinal thickness:after 24h of reperfusion,the retinal thickness of rats in ischemia and reperfusion group was thicker than that of the normal control group,the difference is statistically significant(t=2.869,P=0.008);the retinal thickness of rats in Y27632 treatment group was lower than that of the saline control group,the difference is statistically significant(t=2327,P=0.032).After 168 hours of reperfusion,the retinal thickness of rats in ischemia and reperfusion group was lower than that in normal control group,the difference was statistically significant(t=3.174,P=0.003);the retinal thickness of rats in Y27632 treatment group was was thicker than that in saline control group,the difference is statistically significant(t=115.8,P=0.035).3.ADPase staining:normal control group of rat retinal vascular self-esteem nipple issued to was surrounded by a radial uniform distribution,capillary network structure clear.After 168h of reperfusion,acute ischemia again perfusion group of retinal vascular diameter became fine and walked stiff,decrease of the branches,as around the nipple and week retinal visible large area of non perfusion,perfusion seen around the new vessel sprout gradually into the mesh.In the Y27632 treatment group,there was no perfusion area around the optic disc and the peripheral part of the optic disc.The area of 4PD in the posterior pole part was significantly less than that in the ischemia reperfusion group.4.The expression of caspase-3 protein expression by Western Blot:the protein expression of rats in ischemia and reperfusion group was more than that in normal group,the difference has statistical significance(t=3.306,P=0.002).The protein expression of rats in Y27632 treatment group was less than that in normal saline control group,the difference has statistical significance(t=2.364,P=0.025).5.The expression of Bcl-2 mRNA expression by Real-time PCR:after 24 hours of reperfusion,the expression of Bcl-2 of rats in ischemia reperfusion group was more than that in normal group,the difference has statistical significance(t=2.953,P=0.005).The expression of Bcl-2 of rats in Y27632 treated group was more than that in saline control group,the difference has statistical significance(t=2.249,P=0.03).Conclusions:1.Acute ocular hypertension can cause retinal ischemia reperfusion injury,leading to retinal edema and atrophy,and at the same time lead to retinal ganglion cell apoptosis.2.Retinal ischemia reperfusion injury can cause retinal nerve fiber layer neovascularization.3.Y27632 intravitreous injection can reduce retinal ischemia and early reperfusion retinal edema,reduce the apoptosis of retinal ganglion cells,could reduce late reperfusion retinal atrophy and have a neuroprotective role.4.Y27632 intravitreous injection can reduce retinal neovascularization after retinal ischemia reperfusion injury.5.Y27632 intravitreous injection can reduce the expression of Caspase-3 protein induced by retinal ischemia reperfusion injury.6.Y27632 intravitreous injection can enhance the expression of Bcl-2 mRNA after retinal ischemia-reperfusion injury.