赤芍801可能通过下调nNOS及iNOS的表达抑制缺血性神经元凋亡

Inhibition of neuronal apoptosis induced by cerebral ischemia-reperfusion through down-regulation of nNOS and iNOS expression by Propyl Gallate

目的研究赤芍801(PG)对大鼠脑缺血/再灌注模型缺血区周边组织神经元凋亡的抑制作用及其可能机制。方法线栓左侧大脑中动脉(MCAO)建立大鼠短暂局灶性脑缺血模型,腹腔内注射PG(23.5,47,94μmol·kg-1)干预,分别于缺血2h再灌注1、2、4、6、12、24h收集脑组织标本,通过Nissl、TUNEL染色法观察模型鼠缺血区周边组织阳性神经元数量;蛋白免疫印迹、免疫组织化学方法检测活化型Caspase3、神经元型一氧化氮合酶(nNOS)、诱导型一氧化氮合酶(iNOS)的表达情况。结果再灌注1、2h时段nNOS表达明显增强;自1h起iNOS开始表达并逐渐增强,以12h较明显;再灌注6h活化型Caspase3开始表达,于12h达到高峰,24h表达减弱;于12h开始出现TUNEL阳性神经元,24h其数量明显增多,自4h起Nissl染色阳性神经元逐渐减少,以24h为著,神经元凋亡率亦于同期达到高峰。PG干预各剂量组,nNOS(1h)、iNOS(12h)及活化型Caspase3(12h)表达均有不同程度减弱,TUNEL阳性神经元数量明显减少(24h),Nissl阳性神经元增多(24h),神经元凋亡率明显降低,以94μmol·kg-1作用为著。结论PG可能通过下调nNOS及iNOS的表达而抑制缺血性神经元凋亡。

Aim To explore the inhibition effect of propyl gallate （PG） on neuronal apoptosis in the boundary zone of the infarction area induced by cerebral ischemia-reperfusion and its possible mechanisms. Methods Transient focal ischemia models of rats＇ middle cerebral artery occlusion were induced by inserting a filament through left internal carotid artery for 2 h. Rats were grouped as following： control, sham operation, model, vehicle, PG 23.5, 47,94μmol·kg^-1 and received treatment for three days before operation. Coronal brain sections were collected after 1,2, 4, 6, 12, 24 h of reperfusion, Neuronal injury in the boundary zone of the infarction area were evaluated with TUNEL and Nissl staining ; The expression of activated Caspase-3, nNOS and iNOS was investigated with immunohistochemistry and Western-blot . Results After reperfusion, nNOS immunoreactivity increased markedly at 1 h and 2 h in the boundary zone of the infarction area, the expression of iNOS protein appeared at 1 h and enhanced gradually, peaked at vated Caspase-3 immunoreactivity appeared 12 h. Acti- at 6 h and peaked at 12h, TUNEL positive neurons ap 12 h and became more abundant at 24 h; Th peared at e number of Nissl positive neurons decreased gradually and the apoptosis ratio of neurons peaked at 24 h. The expression of nNOS and iNOS reduced markedly in the rats treated with Propyl Gallate. Meanwhile, both the ex- pression of activated Caspase-3 and the TUNEL positive neurons decreased significantly when compared with model group, However, Nissl positive neurons in treatment groups were more than the model group. Conclusion Down-regulation of nNOS and iNOS expression is the possibly associated with the inhibition of neuronal apoptosis induced by cerebral ischemia-reperfusion by Propyl Gallate.