磷酸肌酸的脑保护作用研究

Cerebral protective effect of phosphocreatine in ischemia and reperfusion

关键词:  磷酸肌酸  脑缺血缺氧  PC12细胞

Key words:  phosphocreatine  cerebral ischemia anoxia  PC12 cell

摘要:

[目的]建立小鼠脑缺血、脑缺血再灌注模型以及细胞缺氧缺糖模型，研究损伤时磷酸肌酸（PCr）对脑的保护作用及其机制。[方法](1) 小鼠尾静脉注射PCr后，进行双侧颈总动脉结扎，以4 h为限记录小鼠的生存时间；濒死小鼠断头取脑组织，测定丙二醛（MDA）含量。(2) 小鼠尾静脉注射PCr，夹闭两侧颈总动脉，夹闭30 min后复灌注30 min，断头取脑，测定MDA含量。(3) 用含连二亚硫酸钠(Na2S2O4)的无糖Earle’s液诱导大鼠嗜铬细胞瘤PC12细胞建立缺氧缺糖损伤模型。用MTT法检测细胞活性；通过检测培养液上清中乳酸脱氢酶（LDH）的释放量来研究PCr对PC12细胞缺氧缺糖损伤的保护作用，以相同浓度的肌酸作对照。[结果]PCr能延长颈总动脉结扎小鼠的平均生存时间，降低脑缺血、脑缺血再灌注小鼠脑组织MDA水平，与模型组比较差异有显著性意义（P<0.01）。在缺氧缺糖PC12细胞损伤模型中，PCr能提高细胞的存活率， PCr浓度为5 mmol/L 及以上时，培养液中LDH水平与模型组比较差异有显著性意义（P<0.01），其作用与同浓度肌酸相比差异无显著性意义。[结论]PCr对缺血小鼠脑组织及缺氧缺糖培养条件下的PC12细胞均具有一定的保护作用。

Abstract:

[Objective] To investigate the protective effect of phosphocreatine (PCr) against brain injury by means of mouse cerebral ischemia/reperfusion model and oxygen/glucose deprivation model on PC12 cells.[Method] （1）The cerebral ischemia model in mice was made by means of ligating bilateral common carotid arteries. The survive rate of mice during 4 hs was observed and malondialdehyde (MDA) in the the brain tissue was measured. （2）After being iv PCr through caudal veins, cerebral ischemia reperfusion model of the mice was made by clipping bilateral common carotid arteries for 30 mins and then loosen for 30 mins. The mice were terminated and brain tissue was taken out for MDA detection. （3）The oxygen/glucose deprivation model on PC12 cells was induced by using sodium hydrosulfite (Na2S2O4) and non-sugar Earle’s fluid respectively. Using creatine as a control, we studied the effect of PCr on the oxygen/glucose deprivation injuries of PC12 cells by determining cell viability (MTT assay) and lactic dehydrogenase (LDH) respectively. [Results] The PCr prolonged the survival time of mice ligated with bilateral carotid arteries strictly and decreased MDA level of the brain tissue in cerebral ischemia mice obviously as compared with those in the injury model group (P<0.01). The PCr had significant protective effects on cultured PC12 cells against oxygen/glucose deprivation injuries, with markedly elevated cell viability and reduced LDH level . When the concentration of PCr was 5 mmol/L and above, the strictly attenuated LDH level was observed compared with those of injury group(P<0.01). There had no obviously difference between PCr and creatine at the same concentration. [Conclusion] PCr has protective effect on brain in cerebral ischemia/perfusion models of mice and PC12 cells in the oxygen/glucose deprivation injury model.