| Objectives To observe the effect of butylphthalide on the expression of apoptotic factor p53 and Bcl-2 after retinal ischemia-reperfusion injury in rabbits,which can provide a theoretical basis for clinical treatment.Methods A total of 80 healthy New-Zealand white rabbits were randomly divided into control group(n=10),RIRI group(n=35)and drug group(n=35).The control group was subdivided into 2 subgroups: 6h and 24 h subgroup(5 rabbits each subgroup).RIRI group and drug group were further divided into 7 subgroups(5 rabbits each subgroup): 1h,6h,12 h,24h,48 h,72h and 7d after reperfusion.The rabbit retinal ischemia-reperfusion injury model was established by high intraocular pressure perfusion method,and the butylphthalide soft capsule was dissolved into 10 mg?m L-1 with edible sesame oil,the dose was 40mg?kg-1.Butylphthalide(head higher then feet and side position)was administered to the drug group immediately after the model was successfully established,and the same dose of edible sesame oil was administered to RIRI group immediately after the model was successfully established,once a day(before meals)until death.The control group was given the same dose of sesame oil as RIRI group at the same observation time point.Results 1 HE staining method to observe the retinal tissue and RGCs morphology The retinal morphology of the control group was similar to that of human,with clear layer boundary,smooth and complete inner limiting membrane,large number of cells in each layer,good morphology,slight edema of a small number of cells,uniform density and neat and tight arrangement.In RIRI group,only local tissues of retinal nerve fiber layer and inner plexiform layer begin to appear slight edema,tissue thickness increased slightly,cytoplasm began to appear vacuolation in 1h after reperfusion.From 6h to 24 h,edema of retinal nerve fiber layer and inner plexiform layer gradually increased,RGCs and inner core layer cells gradually appeared disorder of arrangement,decreased number,and vacuolization of cytoplasm increased.At 24 h,the whole retinal layer showed high edema,local folds of the inner limiting membrane,outward bulge,disordered arrangement of the inner core layer cells,greatly reduced rgcs number,obvious cell degeneration,vacuolar degeneration and unclear boundary.After 48 h and 72 h,the edema gradually decreased.at 7d,the retinal edema almost completely disappeared,but the retinal inner layer was degenerated and atrophied,and rgcs was lost.The change trend of injury degree at each observation time point in the drug group was similar to that in the RIRI group,and the injury peak also appeared at the 24 h time point,but the retinal injury degree in the drug group was lighter than that in the RIRI group at the same time point.2 Expression of RGCs p53 and Bcl-2 was detected by immunohistochemical method Few positive stained cells were detected in the control group,which was significantly lower than RIRI group at the same time point(P<0.05).There was no significant difference in the expression rate of positive cells at 6h or 24 h in the control group(P>0.05),indicating that ocular hypertension perfusion model can cause retinal injury and exclude the time and environmental factors.The positive expression rate of RIRI group was increased before 24h(P<0.05)and decreased after 24h(P<0.05).At each time point,the positive expression rate of RGCs bcl-2 in the drug group was higher than that in RIRI group(P<0.05),and the positive expression rate of p53 in the drug group was lower than that in RIRI group(P<0.05),indicating that butylphthalide could up-regulate the expression of Bcl-2 and down-regulate the expression of p53.3 Apoptosis was detected by TUNEL method Few apoptosis positive stained cells were detected in the control group,which was significantly lower than that in RIRI group at the same time point(P<0.05).There was no significant difference in the expression rate of positive cells at 6h or 24 h in the control group(P<0.05),indicating that ocular hypertension perfusion model can cause retinal injury and exclude time and environmental factors.The positive expression rate of RIRI group was increased before 24h(P<0.05)and decreased after 24h(P<0.05).At each time point,the RGCs apoptosis positive rate of the drug group was less than that of RIRI group at the same time point,the difference was statistically significant except for 7d(P<0.05),indicating that butylphthalide can effectively inhibit cell apoptosis.There was no significant difference in the positive expression of apoptosis between the two groups at 7d(P>0.05),which may be related to the long observation time,the gradual reduction of RIRI damage to retina and the gradual enhancement of retinal self-repair.Conclusions Butylphthalide has a protective effect on maintaining the normal morphology of RIRI retinal tissue and RGCs in rabbits.Butylphthalide can upregulate the expression of Bcl-2 and downregulate the expression of p53.Butylphthalide can inhibit RGCs apoptosis in RIRI to some extent. |
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