穿山龙多糖影响NADPH氧化酶/ROS信号通路抗H2O2诱导的HUVECs细胞损伤作用研究

Rhizoma Dioscoreae Nipponicae polysaccharides protective effect onH2O2-induced HUVECs injury model by interfering NADPH oxidase/ROS pathway

关键词:  穿山龙多糖  Nox4  ROS  p22phox  动脉粥样硬化

Key words:  Rhizoma Dioscoreae Nipponicae polysaccharides (RDNP)  Nox4  ROS  p22phox  atherosclerosis

摘要:

目的　检测穿山龙多糖（RDNP）是否可以通过影响NADPH氧化酶/ROS信号通路对H_{2O2诱导的人脐静脉内皮细胞（HUVECs）氧化损伤产生保护作用。方法　H2O2作用HUVECs 4 h造成损伤，分别给予25、50、100、200、400、800 μg/mL RDNP进行干预，选取适当浓度进行后续实验。采用MTT法测定细胞活力；试剂盒检测 LDH、MDA、SOD、T-AOC、GSH-Px、ROS水平；Western Bolt及 qRT-PCR法检测Nox4、p22phox的蛋白及mRNA表达。结果　加入50、100和200 μg/mL浓度的RDNP与H2O2模型组间相比，活力增加，差异有统计学意义（P<0.05或P<0.01）。因此选择50、100、200 μg/mL RDNP剂量组进行实验。不同浓度RDNP均可以显著改善H2O2损伤，使细胞形态逐渐趋于正常，以200 μg/mL RDNP给药组的效果最为显著。RDNP可以降低H2O2损伤HUVECs模型中LDH、MDA及ROS的水平，并增强细胞内SOD、 GSH-Px、T-AOC活力；且从mRNA水平及蛋白水平抑制Nox4、p22phox的表达。结论　RDNP可以通过干扰NADPH氧化酶/ROS信号通路抑制H2O2造成的HUVECs氧化损伤，保护内皮细胞。}

Abstract:

Objective To investigate effect of Rhizoma Dioscoreae Nipponicae polysaccharides (RDNP) on protecting HUVECs from H_{2O2-induced injury by interfering NADPH oxidase/ROS pathway. Methods HUVECs were treated with RDNP in the presence/absence of H2O2 for 4 h. Then cell activity was detected by MTT method. LDH, MDA, SOD, T-AOC, GSH-Px and ROS were detected by reagent kits to evaluate the cell injuries and the antioxidant activity. Western blot and qRT-PCR was used to evaluate the protein expression and the mRNA expression of Nox4, p22phox. Results LDH, MDA, ROS levels decreased and intracellular SOD, T-AOC as well as GSH-Px activities enhanced in H2O2-treated HUVECs when RDNP was used, especially with 200 μg/mL. Simultaneously, Nox4, p22phox expression were both decreased in protein and mRNA levels in HUVECs. Conclusions RDNP could protect HUVECs from H2O2-induced injury by interfering NADPH oxidase/ROS pathway.}