title: Exenatide can inhibit calcification of human VSMCs through the NF-kappaB/RANKL signaling pathway.
abstract: BACKGROUND
  Arterial calcification is an important pathological change of diabetic vascular complication. Osteoblastic differentiation of vascular smooth muscle cells (VSMCs) plays an important cytopathologic role in arterial calcification. The glucagon-like peptide-1 receptor agonists (GLP-1RA), a novel type of antidiabetic drugs, exert cardioprotective effects through the GLP-1 receptor (GLP-1R). However, the question of whether or not GLP-1RA regulates osteoblastic differentiation and calcification of VSMCs has not been answered, and the associated molecular mechanisms have not been examined.
  
  
  METHODS
  Calcifying VSMCs (CVSMCs) were isolated from cultured human arterial smooth muscle cells through limiting dilution and cloning. The extent of matrix mineralization was measured by Alizarin Red S staining. Protein expression and phosphorylation were detected by Western blot. Gene expression of receptor activator of nuclear factor-κB ligand (RANKL) was silenced by small interference RNA (siRNA).
  
  
  RESULTS
  Exenatide, an agonist of GLP-1 receptor, attenuated β-glycerol phosphate (β-GP) induced osteoblastic differentiation and calcification of human CVSMCs in a dose- and time-dependent manner. RANKL siRNA also inhibited osteoblastic differentiation and calcification. Exenatide decreased the expression of RANKL in a dose-dependent manner. 1,25 vitD3 (an activator of RANKL) upregulated, whereas BAY11-7082 (an inhibitor of NF-κB) downregulated RANKL, alkaline phosphatase (ALP), osteocalcin (OC), and core binding factor α1 (Runx2) protein levels and reduced mineralization in human CVSMCs. Exenatide decreased p-NF-κB and increased p-AMPKα levels in human CVSMCs 48 h after treatment. Significant decrease in p-NF-κB (p-Ser(276), p-Ser(536)) level was observed in cells treated with exenatide or exenatide + BAY11-7082.
  
  
  CONCLUSIONS
  GLP-1RA exenatide can inhibit human VSMCs calcification through NF-κB/RANKL signaling.
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. zhanjunkun@163.com.
authors.forename: Jun-Kun
authors.initials: JK
authors.lastname: Zhan
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. tanpan929@aliyun.com.
authors.forename: Pan
authors.initials: P
authors.lastname: Tan
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. wyj_xy@163.com.
authors.forename: Yan-Jiao
authors.initials: YJ
authors.lastname: Wang
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. cdchenhao@139.com.
authors.forename: Yi
authors.initials: Y
authors.lastname: Wang
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. xyeyyhjy@gmail.com.
authors.forename: Jie-Yu
authors.initials: JY
authors.lastname: He
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. jacksontzy@163.com.
authors.forename: Zhi-Yong
authors.initials: ZY
authors.lastname: Tang
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. huangwu_2008@yeah.net.
authors.forename: Wu
authors.initials: W
authors.lastname: Huang
authors.affiliation: Department of Geriatrics, Institute of Aging and Geriatrics, The Second Xiangya Hospital, Central South University, 139 Middle Renmin Road, Changsha, Hunan, 410011, P.R. China. Liuyoushuo@yeah.net.
authors.forename: You-Shuo
authors.initials: YS
authors.lastname: Liu
mesh_terms: D002113:Calcification, Physiologic / Q000187:drug effects*; D002454:Cell Differentiation / Q000187:drug effects; D000077270:Exenatide; D000067757:Glucagon-Like Peptide-1 Receptor; D006801:Humans; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000187:drug effects / Q000378:metabolism*; D032389:Myocytes, Smooth Muscle / Q000187:drug effects; D016328:NF-kappa B / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology*; D053245:RANK Ligand / Q000378:metabolism*; D034741:RNA, Small Interfering / Q000235:genetics; D018027:Receptors, Glucagon / Q000187:drug effects; D015398:Signal Transduction / Q000187:drug effects*; D061205:Vascular Calcification / Q000188:drug therapy*; D014688:Venoms / Q000494:pharmacology*
chemical_list: C000593833:GLP1R protein, human; D000067757:Glucagon-Like Peptide-1 Receptor; D016328:NF-kappa B; D010455:Peptides; D053245:RANK Ligand; D034741:RNA, Small Interfering; D018027:Receptors, Glucagon; D014688:Venoms; D000077270:Exenatide