title: Miltirone protects human EA.hy926 endothelial cells from oxidized low-density lipoprotein-derived oxidative stress via a heme oxygenase-1 and MAPK/Nrf2 dependent pathway.
abstract: BACKGROUND
  Oxidized low-density lipoprotein (ox-LDL) is an underlying cause of endothelial dysfunction, which is an early event in the pathogenesis of atherosclerosis. In our previous study, we established an ARE-driven luciferase reporter system and screened out several potential Nrf2 activators from Salvia miltiorrhiza Bunge.
  
  
  OBJECTIVE
  Since miltirone showed the most potent ARE-driven luciferase activity, the aim of this study was to test the protective role of miltirone against oxidative stress in endothelial cell and to investigate the underlying mechanistic signaling pathways.
  
  
  METHODS
  In the present study, miltirone increased the expression of nuclear translocation and transcriptional activities of NF-E2-related factor 2 (Nrf2), which led to augmented expression of antioxidant-response element (ARE)-dependent heme oxygenase-1 (HO-1) and NAD(P)H-quinone oxidoreductase 1 (NQO1). Inhibition of Nrf2/HO-1 by RNA interference abolished miltirone-induced cytoprotective effects against ox-LDL, which suggested that Nrf2 and the downstream expression of HO-1 are required for the functional effects of miltirone. Ox-LDL-stimulated mitogen-activated protein kinase activation, ROS production, and miltirone dramatically inhibited synthesis of ROS, as well as decreased SOD and glutathione S-transferase (GST) in human EA.hy926 endothelial cells.
  
  
  RESULTS
  Miltirone-induced Nrf2 and HO-1 expression was related to mitogen-activated protein kinase (MAPK) pathways. The activation of MAPK was partially dependent on the phosphorylation of the c-Jun NH2-terminal kinase (JNK) and extracellular signal-regulated kinase (ERK) pathways, but not P38 MAPK signaling. However, miltirone-induced Nrf2/HO-1 expression can only be effectively blocked by JNK inhibitor SP600125.
  
  
  CONCLUSIONS
  Our findings reveal that miltirone exerts protective functions on endothelial cells in response to ox-LDL-induced oxidative stress, and does so via Nrf2/HO-1, which provides novel insights into the antioxidant capacity of miltirone.
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China.
authors.forename: Liu
authors.initials: L
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China.
authors.forename: Hui
authors.initials: H
authors.lastname: Zhang
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China.
authors.forename: Xueyan
authors.initials: X
authors.lastname: Li
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China.
authors.forename: Bingjie
authors.initials: B
authors.lastname: Jia
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China.
authors.forename: Yuyu
authors.initials: Y
authors.lastname: Yang
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China.
authors.forename: Ping
authors.initials: P
authors.lastname: Zhou
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China. Electronic address: liping2004@126.com.
authors.forename: Ping
authors.initials: P
authors.lastname: Li
authors.affiliation: State Key Laboratory of Natural Medicines (China Pharmaceutical University), No. 24 Tongjia Lane, Nanjing 210009, China; State key Laboratory of Drug Research, Shanghai Institute of Materia Medica, Chinese Academy of Sciences, Shanghai 201203, China. Electronic address: chenj2002cpu@126.com.
authors.forename: Jun
authors.initials: J
authors.lastname: Chen
mesh_terms: D061745:Antioxidant Response Elements; D000975:Antioxidants / Q000494:pharmacology; D050197:Atherosclerosis / Q000378:metabolism; D002460:Cell Line; D042783:Endothelial Cells / Q000187:drug effects* / Q000378:metabolism; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D005982:Glutathione Transferase / Q000378:metabolism; D051547:Heme Oxygenase-1 / Q000378:metabolism*; D006801:Humans; D048031:JNK Mitogen-Activated Protein Kinases / Q000378:metabolism; D008077:Lipoproteins, LDL / Q000378:metabolism*; D020935:MAP Kinase Signaling System / Q000187:drug effects*; D051267:NF-E2-Related Factor 2 / Q000378:metabolism*; D018384:Oxidative Stress / Q000187:drug effects*; D010616:Phenanthrenes / Q000494:pharmacology*; D010936:Plant Extracts / Q000494:pharmacology; D027561:Salvia miltiorrhiza / Q000737:chemistry; D015398:Signal Transduction / Q000187:drug effects; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D000975:Antioxidants; D008077:Lipoproteins, LDL; D051267:NF-E2-Related Factor 2; D010616:Phenanthrenes; D010936:Plant Extracts; C099252:oxidized low density lipoprotein; C068880:miltirone; D051547:Heme Oxygenase-1; D005982:Glutathione Transferase; D048049:Extracellular Signal-Regulated MAP Kinases; D048031:JNK Mitogen-Activated Protein Kinases; D048051:p38 Mitogen-Activated Protein Kinases
keywords: Endothelial dysfunction; HO-1; Miltirone; Nrf2; Oxidative stress