摘要
目的 探讨大鼠局灶性脑缺血再灌注后多聚ADP核糖多聚酶 (PARP)不同时空的表达改变及其与凋亡的关系。方法 运用免疫组织化学和分子生物学技术观察大鼠局灶性脑缺血再灌注后PARP蛋白表达与降解、凋亡的时空动态改变。结果 (1)脑缺血再灌注诱导PARP蛋白表达增强 ,与凋亡的时间变化规律相似 ,但范围大于并涵盖凋亡的范围 ,凋亡分布区外侧的缺血区表达也明显增加。 (2 )同时 ,PARP蛋白出现降解 ,随着缺血或再灌注时间的延长 ,降解逐渐增强。结论 脑缺血 /再灌注损伤可诱导神经细胞DNA修复蛋白PARP的表达。以上结果提示 :轻度缺血时 ,PARP可修复受损DNA ,神经细胞耐受缺血而存活 ;缺血损害重时 ,PARP被降解 ,DNA修复机制受损 ,细胞凋亡程序启动。
Objective To address the relationship between the DNA repair protein, PARP and apoptosis, we examined the expression of PARP and their proteolytic cleavage following reversible focal cerebral ischemia in rats. Methods Adult Wistar rats were subjected to 30 min and 2 h of transient focal cerebral ischemia respectively induced by intraluminal blockade of the left middle cerebral artery, followed by reperfusing after 1-48 h. The expression and cleavage of PARP were analyzed by immunohistochemistry and Western Blot analysis. Apoptosis was evaluated by terminal deoxynucleotidyltranferase (TdT)-mediated dUTP-flourescein nick end-labeling (TUNEL) assay in adjacent sections. Results The protein of PARP was increased after 30 min of occlusion with 1 h of reperfusion. The expression was ischemia- and reperfusion-time dependent. Apoptosis was observed in the inner boundary zone of infarction, with a temporal profile similar to that of PARP-containing cells. In the outer ischemic area, there were few apoptotic cells, even though the expression of PARP was increased at all time points studied. There was a time-course of PARP cleavage with similar kinetics as apoptosis. Conclusions Our findings provide evidence that the expression of DNA repair protein, PARP, is elevated following transient focal cerebral ischemia. The result is consistent with a model in which the amount of DNA damage is small, after mild ischemic insult, DNA repair capability overwhelms DNA damage, and then neurons survive from ischemia, and after severe ischemia injury, the amount of DNA damage is greater and the DNA repair system is impaired, then cell apoptosis programs are initiated.
基金
国家自然科学基金重点项目 (3 973 0 171)
国家自然科学基金项目 (3 9770 810 )
国家自然科学基金项目 (3 0 0 70 82 5 )
