拮抗P2X7受体在保护大鼠脑缺血/再灌注损伤中的作用

目的：探讨拮抗P2X7受体(P2X7 receptor，P2X7R)在保护大鼠脑缺血/再灌注(ischemia/reperfusion，I/R)损伤中的作用及其机制。方法：采用改良线栓法建立大鼠大脑中动脉栓塞(middle cerebral artery occlusion，MCAO)模型；随后对其进行缺血2 h再灌注24 h处理，完成大鼠大脑局灶性脑I/R损伤模型的制备。采用Longa五分法对大鼠进行神经行为学评分；2,3,5-氯化三苯基四氮唑(TTC)染色法检测大鼠大脑梗死体积的变化；Western印迹检测细胞外信号调节激酶(extracellular signal-regulated kinase，ERK)，磷酸化细胞外信号调节激酶(phosphorylation extracellular signal-regulatedkinase，p-ERK)，缝隙连接蛋白43(connexin 43，Cx43)，Bax，Bcl-2及cleaved caspase-3蛋白表达的变化。结果：Longa五分法与TTC染色法结果显示：与假手术组比较，I/R组大鼠神经行为学评分(P<0.05)和大脑梗死体积(P<0.01)均明显增加。与I/R组大鼠相比，P2X7R拮抗剂明亮蓝(brilliant blue G，BBG)或ERK抑制剂PD98059均可明显降低大鼠神经行为学评分(P<0.01)和大鼠大脑梗死体积(P<0.05)。在阻断P2X7R的基础上使用PD98059抑制ERK活性后，大鼠神经行为学评分和大脑梗死体积进一步降低(P<0.05)；Western印迹结果显示：BBG或PD98059均可降低p-ERK，Cx43，Bax/Bcl-2及cleaved caspase-3蛋白表达量(P<0.05)。在阻断P2X7R的基础上使用PD98059抑制ERK活性后，p-ERK，Cx43，Bax/Bcl-2及cleaved caspase-3等蛋白表达量进一步降低(P<0.05)。结论：拮抗P2X7R可减轻大鼠脑I/R损伤，其机制可能与抑制ERK激活，进而降低Cx43和cleaved caspase-3蛋白表达及Bax/Bcl-2比值有关。

Protective effects of P2X7 receptor inhibition in cerebral ischemia/reperfusion injury in rats

Objective: To investigate the protective effects of P2X7 receptor (P2X7R) inhibitor againstcerebral ischemia/reperfusion (I/R) injury in rats and the possible mechanismsMethods: The neurological deficit of rats was evaluated by Longa score. The infarct volumewas examined by 2, 3, 5-triphenyltetrazolium chloride (TTC) staining. The expression levels ofextracellular signal-regulated kinase (ERK), phosphorylation extracellular signal-regulated kinasp-ERK), connexin 43 (Cx43), Bax, Bcl-2 and cleaved caspase-3 were detected by Western blot.Results: Compared with sham group, the neurobehavioral score (P<0.05) and cerebral infarctvolume (P<0.01) of rats in I/R group was increased. Compared with I/R group, brilliant blueG (BBG, the antagonist of P2X7R) or PD98059 (the inhibitor of EKR kinase) could reducethe neurobehavioral score (P<0.01) and cerebral infarct volume significantly (P<0.05). Theneurobehavioral score and cerebral infarct volume was further decreased (P<0.05) when ratswere treated with both BBG and PD98059. Compared with I/R group, the expression levels ofp-ERK, Cx43, cleaved caspase-3 and the ratio of Bax/Bcl-2 were decreased by BBG or PD98059pretreatment, and the protective effects were further enhanced when rats were treated with bothBBG and PD98059 (P<0.05).Conclusion: Inhibition of P2X7R reduces the cerebral I/R injury of rats. ERK inhibition isprobably involved in the protective effects of P2X7R inhibitor against cerebral I/R injury, whichmay be related to the reduction of Cx43 and cleaved caspase-3, and the ratio of Bax/Bcl-2.