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目的探讨丹参酮ⅡA对小胶质细胞BV-2糖氧剥夺/再灌注(oxygen-glucose deprivation and reperfusion,OGD/R)损伤的保护作用及其机制。方法取对数生长期的BV-2细胞OGD 3h后再灌注4~24h,采用Western blot筛选细胞内Nod受体蛋白3(NLPR3)表达水平最高的再灌注时间。将实验组细胞OGD处理3h后分别加入终质量浓度为0~2.5μg/mL的丹参酮ⅡA,CCK8法检测细胞存活率,筛选丹参酮ⅡA的最大有效质量浓度。对BV-2细胞OGD 3h后分别加入0、0.5、1.0、2.0μg/mL丹参酮ⅡA,再灌注12h,Western blot检测BV-2细胞内NLRP3和凋亡蛋白caspase-1的表达水平,ELISA检测BV2细胞白介素(IL)-1β和IL-18的质量浓度。结果 OGD 3h后,再灌注12h时NLPR3蛋白表达水平最高。CCK8法显示丹参酮ⅡA最大有效质量浓度为2.0μg/mL。NLRP3、caspase-1、IL-1β和IL-18表达均随着丹参酮ⅡA质量浓度的升高而降低。结论丹参酮ⅡA能抑制OGD/R处理后BV-2细胞内NLRP3炎症体信号通路分子的表达,这可能是其保护OGD/R细胞的分子机制之一。
Objective To investigate the protective effect of tanshinone ⅡA on microglial cells during oxygen glucose deprivation and reperfusion (OGD / R) injury and its mechanism. Methods BV-2 cells in logarithmic growth phase were reperfused for 3 hours and then reperfused for 4-24 hours. Western blot was used to screen the highest reperfusion time of Nod receptor 3 (NLPR3). The cells in the experimental group were treated with OGD for 3 hours and then treated with tanshinone IIA at the final concentration of 0 ~ 2.5μg / mL. The cell viability was determined by CCK8 assay, and the maximum effective concentration of tanshinone ⅡA was screened. BV-2 cells were treated with 0, 0.5, 1.0 and 2.0 μg / mL tanshinone ⅡA for 3 hours and reperfusion for 12 hours respectively. Western blot was used to detect the expression of NLRP3 and caspase-1 in BV-2 cells. Mass concentrations of interleukin (IL) -1β and IL-18. Results After 3h of OGD, NLPR3 protein expression was the highest at 12h after reperfusion. CCK8 method shows the maximum effective concentration of tanshinone Ⅱ A 2.0μg / mL. The expression of NLRP3, caspase-1, IL-1β and IL-18 decreased with the increase of tanshinone Ⅱ A concentration. Conclusion Tanshinone ⅡA can inhibit the expression of NLRP3 inflammasome signal transduction molecules in BV-2 cells after OGD / R treatment, which may be one of the molecular mechanisms that protect OGD / R cells.