title: Boldine improves endothelial function in diabetic db/db mice through inhibition of angiotensin II-mediated BMP4-oxidative stress cascade.
abstract: OBJECTIVE
  Boldine is a potent natural antioxidant present in the leaves and bark of the Chilean boldo tree. Here we assessed the protective effects of boldine on endothelium in a range of models of diabetes, ex vivo and in vitro.
  
  
  METHODS
  Vascular reactivity was studied in mouse aortas from db/db diabetic and normal mice. Reactive oxygen species (ROS) production, angiotensin AT1 receptor localization and protein expression of oxidative stress markers in the vascular wall were evaluated by dihydroethidium fluorescence, lucigenin enhanced-chemiluminescence, immunohistochemistry and Western blot respectively. Primary cultures of mouse aortic endothelial cells, exposed to high concentrations of glucose (30 mmol L(-1) ) were also used.
  
  
  RESULTS
  Oral treatment (20 mg kg(-1) day(-1) , 7 days) or incubation in vitro with boldine (1 μmol L(-1) , 12 h) enhanced endothelium-dependent aortic relaxations of db/db mice. Boldine reversed impaired relaxations induced by high glucose or angiotensin II (Ang II) in non-diabetic mouse aortas while it reduced the overproduction of ROS and increased phosphorylation of eNOS in db/db mouse aortas. Elevated expression of oxidative stress markers (bone morphogenic protein 4 (BMP4), nitrotyrosine and AT1 receptors) were reduced in boldine-treated db/db mouse aortas. Ang II-stimulated BMP4 expression was inhibited by boldine, tempol, noggin or losartan. Boldine inhibited high glucose-stimulated ROS production and restored the decreased phosphorylation of eNOS in mouse aortic endothelial cells in culture.
  
  
  CONCLUSIONS
  Boldine reduced oxidative stress and improved endothelium-dependent relaxation in aortas of diabetic mice largely through inhibiting ROS overproduction associated with Ang II-mediated BMP4-dependent mechanisms.
authors.affiliation: Department of Pharmacology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia.
authors.forename: Yeh Siang
authors.initials: YS
authors.lastname: Lau
authors.forename: Xiao Yu
authors.initials: XY
authors.lastname: Tian
authors.forename: Mohd Rais
authors.initials: MR
authors.lastname: Mustafa
authors.forename: Dharmani
authors.initials: D
authors.lastname: Murugan
authors.forename: Jian
authors.initials: J
authors.lastname: Liu
authors.forename: Yang
authors.initials: Y
authors.lastname: Zhang
authors.forename: Chi Wai
authors.initials: CW
authors.lastname: Lau
authors.forename: Yu
authors.initials: Y
authors.lastname: Huang
mesh_terms: D000804:Angiotensin II; D000818:Animals; D000975:Antioxidants / Q000494:pharmacology*; D001013:Aorta, Thoracic / Q000166:cytology / Q000187:drug effects* / Q000502:physiology; D001060:Aporphines / Q000494:pharmacology*; D055415:Bone Morphogenetic Protein 4 / Q000378:metabolism*; D002478:Cells, Cultured; D003924:Diabetes Mellitus, Type 2 / Q000378:metabolism* / Q000503:physiopathology; D042783:Endothelial Cells / Q000187:drug effects / Q000378:metabolism; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology; D066298:In Vitro Techniques; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D052250:Nitric Oxide Synthase Type III / Q000378:metabolism; D018384:Oxidative Stress; D044140:Receptor, Angiotensin, Type 1 / Q000378:metabolism; D014664:Vasodilation / Q000187:drug effects
chemical_list: D000975:Antioxidants; D001060:Aporphines; C525810:Bmp4 protein, mouse; D055415:Bone Morphogenetic Protein 4; D044140:Receptor, Angiotensin, Type 1; D000804:Angiotensin II; C011686:boldine; D052250:Nitric Oxide Synthase Type III; C496315:Nos3 protein, mouse
keywords: boldine; diabetic mice; endothelial dysfunction; oxidative stress