枸杞多糖通过核因子κB信号通路对过氧化氢诱导的神经细胞损伤保护作用的研究

Protective effect of Lycium barbarum polysaccharide on HO-induced neuronal injury via NF-κB signaling pathway

目的研究枸杞多糖(LBP)对过氧化氢(H_(2)O_(2))诱导神经细胞PC12损伤的保护作用及分子机制。方法 PC12细胞分为正常对照组(Con)、H_(2)O_(2)诱导模型组(H_(2)O_(2))、H_(2)O_(2)+75 mg/L LBP组(H_(2)O_(2)+LBP 75)、H_(2)O_(2)+150 mg/L LBP组(H_(2)O_(2)+LBP 150)、H_(2)O_(2)+300 mg/L LBP组(H_(2)O_(2)+LBP 300)、H_(2)O_(2)+300 mg/L LBP+丙二醇甲醚醋酸酯(PMA)组(H_(2)O_(2)+LBP 300+PMA)。CCK-8法检测细胞存活率,DCFH-DA探针法检测细胞内活性氧簇(ROS)水平,比色法检测细胞丙二醛(MDA)、超氧化物歧化酶(SOD)和一氧化氮(NO)含量,流式细胞术检测细胞凋亡率,Western blot检测细胞中核因子κB(NF-κB)信号通路相关蛋白表达水平。结果与Con组比较,H_(2)O_(2)组细胞存活率降低[(100.00±5.25)%vs(53.37±2.57)%,P<0.05]、ROS水平升高[(20.05±2.14)%vs(46.24±3.27)%,P<0.05],MDA水平升高[(8.17±0.41)vs(20.06±1.06)mmol/L,P<0.05],SOD活性降低[(19.39±1.17)vs(8.42±0.53)U/ml,P<0.05],凋亡率升高[(4.82±1.01)%vs(33.62±3.18)%,P<0.05]。与H_(2)O_(2)组比较,不同浓度LBP能缓解H_(2)O_(2)诱导的氧化损伤,减少细胞凋亡,而NF-κB信号通路激活剂PMA可逆转LBP对PC12细胞损伤保护作用。结论 LBP通过抑制NF-κB信号通路活化,对H_(2)O_(2)诱导的神经细胞损伤有保护作用。

Objective To investigate the protective effect and molecular mechanism of Lycium barbarum polysaccharide(LBP)on the PC12 injury in nerve cells induced by hydrogen peroxide(H_(2)O_(2)).Methods PC12 cells were divided into normal control group(Con),H_(2)O_(2)-induced model group(H_(2)O_(2)),H_(2)O_(2)+75 mg/L LBP group(H_(2)O_(2)+LBP 75),H_(2)O_(2)+150 mg/L LBP group(H_(2)O_(2)+LBP 150)and H_(2)O_(2)+300 mg/L LBP group(H_(2)O_(2)+LBP 300).Cell counting kit(CCK-8)method was adopted to detect cell viability.DCFH-DA probe method was used to detect intracellular reactive oxygen species(ROS)levels,and colorimetric method to detect cellular malondialdehyde(MDA)and superoxide dismutase(SOD)and nitric oxide(NO)content.Flow cytometry was used to detect the rate of apoptosis,and Western blot was used to detect the expression levels of proteins related to the nuclear transcription factor kappa B(NF-κB)signaling pathway.Results Compared with the Con group,the cell survival rate decreased[(100.00±5.25)%vs(53.37±2.57)%,P<0.05],the level of ROS increased[(20.05±2.14)%vs(46.24±3.27)%,P<0.05],MDA content increased[(8.17±0.41)vs(20.06±1.06)mmol/L,P<0.05],SOD activity decreased[(19.39±1.17)vs(8.42±0.53)U/ml,P<0.05],the rate of apoptosis increased[(4.82±1.01)%vs(33.62±3.18)%,P<0.05]in the H_(2)O_(2)induced model group.Compared with the H_(2)O_(2)induced model group,LBP at different concentrations could significantly alleviate the oxidative damage induced by H_(2)O_(2)and reduce cell apoptosis,while the NF-κB signaling pathway activator PMA could reverse the protective effect of Lycium barbarum polysaccharides on PC12 cells.Conclusion LBP had a protective effect on H_(2)O_(2) induced neuronal injury by inhibiting the activation of NF-κB signaling pathway.