血必净注射液抑制eNOS脱耦联减轻脓毒症血管内皮细胞氧化应激

Xuebijing injection reduces oxidative stress of sepsis vascular endothelial cells by inhibiting eNOS decoupling

目的:探讨血必净注射液改善脓毒症血管内皮细胞(VECs)氧化应激的机制。方法:选用EA.hy926细胞建立脓毒症细胞模型(LPS组)、血必净注射液干预模型(XBJ+LPS组)和对照组(XBJ组),孵育12 h后裂解细胞,分别检测总超氧化物歧化酶(T-SOD)活性、丙二醛(MDA)浓度、一氧化氮(NO)浓度及一氧化氮合酶(NOS)活性;采用质谱分析法检测细胞裂解液的四氢叶酸(BH4)和左旋精氨酸(L-Arg)浓度;荧光探针法和ELISA法分别检测正常功能的内皮源性一氧化氮合成酶(eNOS)和总的eNOS浓度,计算脱偶联的eNOS的水平。结果:与XBJ组相比,LPS组培养液中的MDA水平均明显升高(P<0.05),NO的含量均明显降低(P<0.05);而XBJ+LPS组培养液中的MDA水平和T-SOD活性均没有明显变化(P>0.05),仅有NO水平明显升高(P<0.05)。各组BH4和L-Arg含量基本保持稳定。然而,与XBJ组相比,LPS组细胞内的总eNOS并未明显变化(P>0.05),正常功能性eNOS下降,脱偶联的eNOS相应增加(P<0.05);XBJ+LPS组与LPS组比较,正常功能性eNOS升高,脱偶联的eNOS明显减少(P<0.05)。结论:血必净注射液能够减轻脓毒症VECs氧化应激,其可能机制与降低eNOS脱偶联有关。

Objective:To investigate the mechanism of Xuebijing injection in improving oxidative stress in septic vascular endothelial cells(VECs).Methods:The EA.hy926 cells were divided into three groups:sepsis cell model(LPS group),Xuebijing injection(XBJ+LPS group)and control group(XBJ group).After incubated for 12 hours,the following indicators were detected:total superoxide dismutase(T-SOD)activity,malondialdehyde(MDA)concentration,nitric oxide(NO)concentration and nitric oxide synthase(NOS)concentration were detected by detection kit.The concentrations of tetrahydrobiopterin(BH4)and L-arginine(L-Arg)were detected by mass spectrometry.The concentrations of eNOS in normal function and total eNOS were detected by fluorescence probe method and ELISA method respectively,and the level of uncoupling eNOS was calculated.Results:Compared with XBJ group,MDA level in LPS group was significantly increased(P<0.05),and NO level was significantly decreased(P<0.05),while MDA level and T-SOD activity in XBJ+LPS group were not significantly changed(P<0.05),only NO level was significantly increased(P<0.05).The contents of BH4 and L-Arg remained stable.However,compared with XBJ group,the total eNOS in LPS group did not change significantly(P<0.05),the normal functional eNOS decreased,and the uncoupling eNOS increased(P<0.05);compared with the LPS group,the normal functional eNOS increased and the uncoupling eNOS decreased significantly in XBJ+LPS group(P<0.05).Conclusion:Xuebijing injection can reduce the oxidative stress of VECs in sepsis,and the possible mechanism is to reduce eNOS decoupling.