丙二酸二甲酯在脑梗死后的保护作用

Protective Effect of Dimethyl Malonate After Ischemic Stroke

目的探索丙二酸二甲酯(DMM)对脑梗死小鼠的具体保护作用。方法将220只成年雄性C57BL/6小鼠分为假手术+盐水组(50只)、假手术+DMM组(50只)、脑梗死+盐水组(60只)、脑梗死+DMM组(60只)。使用线栓法制备小鼠短暂性大脑中动脉闭塞模型,在缺血前10 min经尾静脉滴注等体积生理盐水或DMM(6 mg·kg^(-1)·min^(-1))直至线栓拔出。术后第1天通过TTC染色和伊文思蓝染色判断DMM对梗死体积和血脑屏障的作用,通过蛋白印迹实验检测脑组织中炎症相关因子肿瘤坏死因子-α(TNF-α)、诱导型一氧化氮合酶(iNOS)、白介素-10(IL-10)和紧密连接蛋白1(ZO-1)的表达水平。术后第3天通过蛋白印迹实验检测脑组织NOD样受体热蛋白结构域相关蛋白3(NLRP3)、白介素-18(IL-18)、白介素-1β(IL-1β)的表达水平,通过免疫荧光检测小鼠大脑梗死周围区iNOS阳性细胞数量的变化。结果术后第1天DMM减小了小鼠脑梗死体积并降低了促炎因子TNF-α、iNOS的表达,上调了抗炎因子IL-10的表达。术后第1天DMM减轻了梗死小鼠伊文思蓝的泄露并降低了脑含水量,ZO-1表达水平上升。术后第3天DMM降低了NLRP3、IL-18和IL-1β的表达水平,减少了梗死周围区iNOS阳性细胞数量。结论丙二酸二甲酯通过抑制神经炎症和保护血脑屏障对脑梗死小鼠产生保护作用。

Objective To explore the protective effect of dimethyl malonate(DMM)on ischemic stroke mice.Methods 220 adult male C57BL/6 mice were divided into sham+saline group(n=50),sham+DMM group(n=50),transient middle cerebral artery occlusion(tMCAO)+saline group(n=60),and tMCAO+DMM group(n=60).The mouse model of tMCAO was prepared by inserting suture,and constant volume of normal saline or DMM(6 mg·kg^(-1)·min^(-1))was dripping through tail vein 10 minutes before ischemia until the suture occlusion was removed.The effect of DMM on infarct volume and BBB was determined by TTC staining and Evans blue staining on day 1 after operation.The expression levels of inflammation-related factors tumor necrosis factor-α(TNF-α),inducible nitric oxide synthase(iNOS),and interleukin 10(IL-10)and zonula occluden 1(ZO-1)in the brain tissues were detected by Western blot on day 1 after operation,and the expression levels of NOD-like receptor thermal protein domain associated protein 3(NLRP3),interleukin 18(IL-18)and interleukin 1β(IL-1β)in the brain tissues were detected by Western blot on day 3 after operation.The number of iNOS-positive cells in the peri-infarct area of mice were detected by immunofluorescence on day 3 after operation.Results On day 1 after operation,DMM significantly decreased the cerebral infarction volume and reduced the expressions of pro-inflammatory cytokines TNF-αand iNOS,and up-regulated the expression of anti-inflammatory cytokine IL-10.On day 1 after operation,DMM significantly alleviated the leakage of Evans blue and reduced brain water content,and the expression level of ZO-1 was also significantly increased.DMM reduced the expression levels of NLRP3,IL-18 and IL-1β,and decreased the number of iNOS-positive cells in the peri-infarct area on day 3 after operation.Conclusion Dimethyl malonate has protective effects on ischemic stroke mice by inhibiting neuroinflammation and protecting blood-brain barrier.