湖南中医药大学学报2025，Vol.45Issue(9)：1625-1635,11.DOI:10.3969/j.issn.1674-070X.2025.09.006
荷叶碱促进S1P/S1PR1/AMPK来减轻H2O2诱导的血管内皮细胞线粒体氧化应激损伤
Nuciferine alleviating H2O2-induced mitochondrial oxidative stress damage in vascular endothelial cells by promoting S1P/S1PR1/AMPK pathway
摘要
Abstract
Objective To explore the protective effects of nuciferine(NF)on mitochondrial damage caused by H2O2-induced oxidative stress in human umbilical vein endothelial cells(HUVECs)and the related mechanisms.Methods HUVECs were subjected to oxidative damage induced by H2O2 as an experimental model.The cells were divided into the control group,H2O2 group,H2O2+NF group,H2O2+NF+W146 group,H2O2+NF+SEW2871 group,and H2O2+NF+Compound C group.They were pretreated with the S1PR1 inhibitor W146(10 μmol/L),S1PR1 agonist SEW2871(20 nmol/L),and AMPK inhibitor Compound C(dorsomorphin,1 μmol/L)for 30 min,30 min,and 24 h,respectively.Subsequently,10 μmol/L NF was added and incubated for 12 h,followed by co-incubation with 400 μmol/L H2O2 for 2 h.The secretion level of S1P was measured using ELISA.The protein expression levels of S1PR1,S1PR3,AMPK,and p-AMPK were determined by Western blot.The intracellular levels of reactive oxygen species(ROS)were measured using dihydroethidium staining.The ultrastructure of mitochondria was observed using transmission electron microscopy.The mitochondrial membrane potential(MMP)was determined using JC-1 fluorescent probes and flow cytometry.The opening degree of the mitochondrial permeability transition pore(MPTP)was examined using the Calcein AM fluorescent probe.The intracellular ATP levels were measured using an ATP assay kit.Results Compared with the control group,the H2O2 group exhibited increased mitochondrial oxidative stress damage(P<0.01).Compared with the H2O2 group,the H2O2+NF group demonstrated enhanced cell viability(P<0.001),reduced ROS levels,alleviated mitochondrial ultrastructural damage,and decreased MPTP opening degree,as well as elevated MMP and ATP levels(P<0.05).Additionally,the H2O2+NF group showed increased S1P secretion(P<0.05)and elevated protein expression levels of S1PR1(P<0.05)and p-AMPK(P<0.01).Compared with the H2O2+NF group,the H2O2+NF+SEW2871 group exhibited increased p-AMPK protein expression level(P<0.05)and reduced mitochondrial oxidative stress damage(P<0.05).Conversely,the H2O2+NF+W146 and H2O2+NF+Compound C groups showed decreased p-AMPK protein expression level(P<0.05)and elevated mitochondrial oxidative stress level(P<0.05,P<0.01).Conclusion NF alleviates mitochondrial damage caused by H2O2-induced oxidative stress in HUVECs by promoting S1P/S1PR1/AMPK signaling pathway.关键词
氧化应激/线粒体损伤/荷叶碱/人脐静脉内皮细胞/1-磷酸鞘氨醇/1-磷酸鞘氨醇受体1/腺苷酸活化蛋白激酶Key words
oxidative stress/mitochondrial damage/nuciferine/human umbilical vein endothelial cell/sphingosine-1-phosphate/sphingosine-1-phosphate receptor 1/adenosine 5'-monophosphate-activated protein kinase分类
医药卫生引用本文复制引用
刘婉婷,谢玉鑫,李婧,李朝荃,姚慧,付傲妮,杨皓天,易光辉..荷叶碱促进S1P/S1PR1/AMPK来减轻H2O2诱导的血管内皮细胞线粒体氧化应激损伤[J].湖南中医药大学学报,2025,45(9):1625-1635,11.基金项目
国家自然科学基金项目(81770490) （81770490）
湖南省科技计划项目(2020JJ4535). （2020JJ4535）
