局灶性脑梗死大鼠半胱氨酸天冬氨酸蛋白酶3和9的表达:空间分布特征及时间演变规律

Expressions of caspase3 and caspase9 in rats with focal cerebral infarction:characteristics of spatial distribution and law of time development

目的:观察大鼠局灶性脑梗死后半胱氨酸天冬氨酸蛋白酶3和9表达的时间和空间分布的动态演变。方法:实验于2004-03/06在山东省泰山医学院微循环重点实验室完成。选择Wistar大鼠84只,随机分为正常组6只、假手术组6只、局灶性脑梗死组72只,局灶性脑梗死组根据光照后时间分为30min,3,6,9,12,24,36,48h,3,4,5,6d共12组,每组6只。采用光化学方法制作大鼠局灶性脑梗死模型。动物处死前1h注射10g/L伊文思兰1mL。正常组不作任何处理,假手术组仅暴露完整的颅骨,按要求时间点麻醉处死大鼠取脑组织,观测半胱氨酸天冬氨酸蛋白酶3和9的表达采用免疫组织化学分析。结果:参加实验的84只大鼠全部进入结果分析。①脑梗死后30min即有半胱氨酸天冬氨酸蛋白酶3和9阳性细胞出现,呈半球状向四周放射样扩展,随脑梗死时间的延长逐渐增多,二者均在48h表达至高峰,但前者表达更显著(P<0.05),阳性细胞局限于脑梗死区周围。②正常对照组、假手术组均未发现半胱氨酸天冬氨酸蛋白酶3和9的阳性表达。结论:①局灶性脑梗死后半胱氨酸天冬氨酸蛋白酶3和9表达部位一致,以坏死区为中心呈放射状空间分布特征。②二者均在30min开始表达,48h时半胱氨酸天冬氨酸蛋白酶3较半胱氨酸天冬氨酸蛋白酶9表达增多,这种时间变化规律说明细胞凋亡的级联反应最终通路有放大作用。

AIM:To observe the development of time and spatial distribution of the expressions of caspase3 and caspase9 following focal cerebral infarction in rats.METHODS:The experiment was carried out in the Key Laboratory of Microcirculation,Taishan Medical College between March and June 2004. Eightyfour Wistar rats were randomly divided into normal group(n=6), shamoperated group(n=6) and focal cerebral infarction group(n=72), and the last one was subdivided into 12 groups with 6 rats in each group according to the time of light shining:30minute group,3,6,9,12,24,36 and 48hour groups and 3,4,5 and 6day groups respectively.Rat models of focal cerebral infarction were established with the method of photochemical initiation of thrombosis.The rats were injected with 10 g/L Evans blue 1 hour before being killed.Rats in the normal group received no treatment,and those in the shamoperated group only treated with exposure of complete skull;The rats were anesthetized and killed at required time points,and then brain tissue was taken;Immunohistochemical analysis was used to detect the expressions of caspase3 and caspase9.RESULTS:All the 84 rats were involved in the analysis of results.①Thirty minutes after cerebral infarction,positive cells of caspase3 and caspase9 were observed,which were expanded radiatively as hemisphere,and the number of the cells gradually increased with the prolongation of cerebral infarction;Both reached the peak values at 48 hours,but the former expressed more significantly(P< 0.05),and the positive cells were limited around the infarcted regions.②No positive expressions of caspase3 and caspase9 were observed in the normal control group and shamoperated group.CONCLUSION:①The expressed locations of caspase3 and caspase9 following focal cerebral infarction are coincident.Radiative spatial distribution takes the infarcted region as the center.②Both caspase3 and caspase9 begin to express at 30 minutes,and the expression of caspase3 is more than that of caspase9 at 48 hours,and this law of time change indicates that final pass of apoptosis cascade reaction has the role of enlargement.