Isorhamnetin prevent endothelial cell injuries from oxidized LDL via activation of p38MAPK.
The present investigation was undertaken to determine the protective effects of isorhamnetin on endothelial cell line EA.hy926 injuries induced by oxidized low-density lipoprotein (ox-LDL) and to uncover some of the underlying mechanisms of these effects. Indices such as cell viability, lactate dehydrogenase (LDH), and nitric oxide (NO) release were measured to evaluate the protective effects of isorhamnetin. 2, 2-diphenyl-1-picrylhydrazyl (DPPH) radical scavenging activity, superoxide dismutase (SOD), superoxide and reactive oxygen species (ROS) generation were also detected to evaluate the antioxidant effects of isorhamnetin. Reverse Transcription-Polymerase Chain Reaction (RT-PCR) was used to confirm the expression of endothelial nitric oxide synthase (eNOS) mRNA and lectin-like ox-LDL receptor-1 mRNA. Western blotting was used to evaluate the protein expression of this receptor and eNOS, as well as p38-mitogen-activated protein kinase (p38MAPK) phosphorylation and NF-kappaB p65 translocation. As a result, cell viability decreased significantly (P<0.01) after 24 h treatment with ox-LDL, accompanied with apparent secretion disorders such as NO reduction and LDH increase. Pretreatment with isorhamnetin resulted in remarkable increase of cell viability (P<0.05) and modulation of secretion disorders mediated by ox-LDL in a concentration-dependent manner. Besides, ox-LDL led to upregulation of lectin-like ox-LDL receptor-1, phosphorylation of p38MAPK, translocation of NF-kappaB, and downregulation of the eNOS expression in endothelial cells. Isorhamnetin pretreatment inhibited the ox-LDL-induced downregulation of eNOS, upregulation of lectin-like ox-LDL receptor-1, phosphorylation of the p38MAPK and translocation of NF-kappaB. Moreover, isorhamnetin exhibited strong antioxidant activity, which was shown by its inhibition effects on ox-LDL-induced superoxide, ROS overproduction and significant SOD reduction. The data indicated the protective effects of isorhamnetin on endothelial cell line EA.hy926 from ox-LDL-induced cell injuries. These effects were obtained via inhibition of lectin-like ox-LDL receptor-1 upregulation, interference of ox-LDL-mediated intracellular signaling pathway (p38MAPK activation, NF-kappaB nuclear translocation, eNOS expression) and the antioxidant activity of isorhamnetin.
Bao
Meihua
M
Institute of Pharmacology and Toxicology and Biochemical Pharmaceutics, College of Pharmaceutical Sciences, Zhejiang University, 353 Yan'an Road, Hangzhou 310031, China.
Lou
Yijia
Y
D021581:Active Transport, Cell Nucleus / Q000187:drug effects; D001713:Biphenyl Compounds / Q000037:antagonists & inhibitors / Q000378:metabolism; D002460:Cell Line; D002467:Cell Nucleus / Q000187:drug effects / Q000378:metabolism; D002470:Cell Survival / Q000187:drug effects; D004305:Dose-Response Relationship, Drug; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism / Q000473:pathology; D004789:Enzyme Activation / Q000187:drug effects; D044948:Flavonols / Q000494:pharmacology*; D016166:Free Radical Scavengers / Q000494:pharmacology; D015870:Gene Expression / Q000187:drug effects; D006801:Humans; D006834:Hydrazines / Q000037:antagonists & inhibitors / Q000378:metabolism; D007770:L-Lactate Dehydrogenase / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000494:pharmacology*; D009569:Nitric Oxide / Q000378:metabolism; D052250:Nitric Oxide Synthase Type III / Q000235:genetics; D010766:Phosphorylation / Q000187:drug effects; D010851:Picrates; D011794:Quercetin / Q000031:analogs & derivatives; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D051131:Receptors, Oxidized LDL / Q000235:genetics; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013482:Superoxide Dismutase / Q000378:metabolism; D013481:Superoxides / Q000378:metabolism; D051996:Transcription Factor RelA / Q000378:metabolism; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism*
D001713:Biphenyl Compounds; D044948:Flavonols; D016166:Free Radical Scavengers; D006834:Hydrazines; D008077:Lipoproteins, LDL; D010851:Picrates; D012333:RNA, Messenger; D017382:Reactive Oxygen Species; D051131:Receptors, Oxidized LDL; D051996:Transcription Factor RelA; C099252:oxidized low density lipoprotein; C047368:3-methylquercetin; D013481:Superoxides; D009569:Nitric Oxide; D011794:Quercetin; C004931:1,1-diphenyl-2-picrylhydrazyl; D007770:L-Lactate Dehydrogenase; D052250:Nitric Oxide Synthase Type III; D013482:Superoxide Dismutase; D048051:p38 Mitogen-Activated Protein Kinases