丹参酮ⅡA预处理对氧糖剥夺-复氧复糖N9细胞NF-κB信号通路的影响

Effect of Tanshinonne ⅡA preconditioning on nuclear factor kappa B signaling pathway in N9 cells subjected to oxygen-glucose deprivation and restoration

目的评价丹参酮ⅡA预处理对氧糖剥夺-复氧复糖N9细胞NF-κB信号通路的影响。方法体外培养的N9细胞,采用随机数字表法分为4组(n=16):对照组(C组)、氧糖剥夺-复氧复糖组(OGD/R组)、溶剂组(V组)和丹参酮ⅡA预处理(TP组)。采用氧糖剥夺4 h,复氧复糖24 h的方法制备N9细胞氧糖剥夺-复氧复糖模型。C组不做任何处理;OGD/R组行氧糖剥夺4 h,复氧复糖24 h;TP组采用含丹参酮ⅡA(终浓度5 mg/L)培养液培养24 h后制备模型;V组培养液中加入二甲基亚砜。于复氧24 h时采用分光光度法测定培养液LDH及NO水平,采用ELISA法测定IL-β和TNF-α水平;采用TUNEL法观察细胞凋亡情况,采用Western blot法检测胞核NF-κB的表达。结果与C组比较,OGD/R组、V组和TP组细胞培养液LDH、IL-β、TNF-α、NO水平和细胞凋亡率升高,胞核NF-κB表达上调(P<0.05);与OGD/R组比较,TP组细胞培养液LDH、IL-β、TNF-α、NO水平和细胞凋亡率降低,胞核NF-κB表达下调(P<0.05),V组上述指标差异无统计学意义(P>0.05)。结论丹参酮ⅡA预处理减轻N9细胞氧糖剥夺-复氧复糖损伤的机制与抑制NF-κB信号通路有关。

Objective To evaluate the effect of Tanshinonne ⅡA preconditioning on nuclear factor kappa B (NF-κB) signaling pathway in N9 cells subjected to oxygen-glucose deprivation and restoration (OGD/R). Methods N9 cells cultured in vitro were divided into 4 groups (n=16 each) by using a random number table method: control group (C group), OGD/R group, vehicle group (V group) and Tanshinone IIA preconditioning group (TP group). N9 cells were subjected to O2-glucose deprivation for 4 h followed by restoration of O2-glucose supply for 24 h to establish the OGD/R model.Group C receive no treatment.In group TP, N9 cells were cultured in medium containing Tanshinone ⅡA (final concentration 5 mg/L) for 24 h before the model was established.Dimethyl sulfoxide was added to the culture medium in group V. At the 24 h of reoxygenation, the level of lactate dehydrogenase (LDH) and nitric oxide(NO)in culture medium was detected by spectrophotometry, cell apoptosis was assessed by using TUNEL, and the expression of NF-κB in nucleus was detected by Western blot.The apoptosis rate was calculated. Results Compared with group C, the levels of LDH, IL-β, TNF-α and NO in culture medium and apoptosis rate were significantly increased, and the expression of NF-κB in nucleus was up-regulated in OGD/R, V and TP groups (P<0.05). Compared with group OGD/R, the levels of LDH, IL-β, TNF-α and NO in culture medium and apoptosis rate were significantly decreased, and the expression of NF-κB in nucleus was down-regulated in group TP (P<0.05), and no significant change was found in the parameters mentioned above in group V (P>0.05). Conclusion The mechanism by which Tanshinonne ⅡA preconditioning reduces OGD/R injury to N9 cells is related to inhibiting NF-κB signaling pathway.