摘要
目的 探讨瑞马唑仑对OGD/R诱导的神经细胞自噬和凋亡的影响及作用机制.方法 体外培养小鼠海马神经元细胞(HT22)并进行神经细胞氧糖剥夺/再复氧(OGD/R),筛选实验用瑞马唑仑浓度;将HT22细胞分为对照组、OGD/R组、瑞马唑仑组、2-ME2组、瑞马唑仑+2-ME2组;CCK8法检测5组HT22细胞活力;流式细胞术检测5组HT22细胞凋亡率;透射电子显微镜观察5组HT22细胞自噬小体的形成;Western blot检测5组HT22细胞HIF-1α、BNIP3、LC3-Ⅱ/LC3-Ⅰ的表达.结果 确定实验用瑞马唑仑浓度为50 μg/mL;与对照组比较,OGD/R组HT22细胞OD450值、HIF-1α、BNIP3、LC3-Ⅱ/LC3-I蛋白水平下调,凋亡率上调(P＜0.05);与OGD/R组比较,瑞马唑仑组HT22细胞自噬小体增加,OD450值、HIF-1α、BNIP3、LC3-Ⅱ/LC3-I蛋白水平上调,凋亡率下调(P＜0.05);2-ME2组HT22细胞OD450值、HIF-1a、BNIP3、LC3-Ⅱ/LC3-I蛋白水平下调,凋亡率上调(P＜0.05).与瑞马唑仑组比较,瑞马唑仑+2-ME2组HT22细胞自噬小体数量减少,OD450值、HIF-1α、BNIP3、LC3-Ⅱ/LC3-Ⅰ蛋白水平下调,凋亡率上调(P＜0.05);与2-ME2组比较,瑞马唑仑+2-ME2组HT22细胞OD450值、HIF-1α、BNIP3、LC3-Ⅱ/LC3-Ⅰ蛋白水平上调,凋亡率下调(P＜0.05).结论 瑞马唑仑可通过激活HIF-1α/BNIP3信号通路促进OGD/R诱导的神经细胞自噬,抑制细胞凋亡,从而减轻OGD/R诱导的神经细胞损伤.
Abstract
Objective To explore the effect and underlying mechanism of remimazolam on oxygen glucose deprivation/reoxygenation(OGD/R)-induced neuronal ischemia-reperfusion injuries of autophagy and apoptosis.Methods The mouse hippocampal neuron cell line HT22 was cultured in vitro,and subjected to OGD/R.The concentration of remimazolam used in this experiment was screened.HT22 cells were induced with blank control,OGD/R,remimazolam,2-ME2 and 2-ME2+remimazolam.Cell viability,apoptosis,autophagosomes were detected by CCK-8 assay,flow cytometry and transmission electron microscopy(TEM),respectively.Protein expressions of hypoxia inducible factor-1(HIF-1α),BCL2 interacting protein 3(BNIP3)and light chain 3(LC3)-Ⅱ(LC3-Ⅱ)/LC3-Ⅰ were detected by Western blot.Results The concentration of remimazolam used in this experiment was determined to be 50μg/ml.Compared with those of blank control,OGD/R-induced HT22 cells presented with significantly lower OD450 and protein expressions of HIF-1α,BNIP3 and LC3-Ⅱ/LC3-Ⅰ,but significantly higher apoptotic rate(P＜0.05).Compared with those induced with OGD/R,HT22 cells induced with remimazolam presented with significantly higher number of autophagosomes,OD450 and protein expressions of HIF-1α,BNIP3 and LC3-Ⅱ/LC3-Ⅰ,but significantly lower apoptotic rate(P＜0.05).Compared with those induced with OGD/R,HT22 cells induced with 2-ME2 presented with significantly lower number of autophagosomes,OD450 and protein expressions of HIF-1α,BNIP3 and LC3-Ⅱ/LC3-Ⅰ,but significantly higher apoptotic rate(P＜0.05).Compared with those induced with remimazolam,HT22 cells induced with remimazolam+2-ME presented with significantly lower number of autophagosomes,OD450 and protein expressions of HIF-1α,BNIP3 and LC3-Ⅱ/LC3-Ⅰ but significantly higher apoptotic rate(P＜0.05).Compared with those induced with 2-ME,HT22 cells induced with remimazolam+2-ME presented with significantly higher number of autophagosomes,OD450 and protein expressions of HIF-1α,BNIP3 and LC3-Ⅱ/LC3-Ⅰ,but significantly lower apoptotic rate(P＜0.05).Conclusion Remimazolam can promote OGD/R-induced autophagy and inhibit apoptosis of neural cells by activating the HIF-1α/BNIP3 signaling pathway,thereby alleviating OGD/R-induced neuronal damage.
基金项目
新疆维吾尔自治区自然科学基金(2021D01C207)
NETL
NSTL
维普
万方数据