Jian Shen1,2, Ning Zhang1,2, Yi-Nuo Lin1,2, PingPing Xiang1,2, Xian-Bao Liu1,2, Peng-Fei Shan3, Xin-Yang Hu1,2, Wei Zhu1,2, Yao-Liang Tang4, Keith A Webster5, Renzhi Cai5,6,7, Andrew V Schally5,6,7, Jian'an Wang1,2, Hong Yu8,2. 1. From the Departments of Cardiology (J.S., N.Z., Y.-N.L., P.P.X., X.-b.L., X.-y.H., W.Z., J.W., H.Y.). 2. Second Affiliated Hospital, College of Medicine, Zhejiang University, Hangzhou, PR China; Cardiovascular Key Laboratory of Zhejiang Province, Hangzhou, PR China (J.S., N.Z., Y.-N.L., P.P.X., X.-b.L., X.-y.H., W.Z., J.W., H.Y.). 3. Endocrinology and Metabolism (P.-f.S.). 4. Vascular Biology Center, Georgia Regents University, Augusta (Y.-l.T.). 5. Department of Molecular and Cellular Pharmacology and the Vascular Biology Institute (K.A.W., R.C., A.V.S.). 6. Divisions of Hematology/Oncology, Department of Medicine (R.C., A.V.S.). 7. Miller School of Medicine, University of Miami, FL; and Endocrine, Polypeptide and Cancer Institute, Veterans Affairs Medical Center, Miami, FL (R.C., A.V.S.). 8. From the Departments of Cardiology (J.S., N.Z., Y.-N.L., P.P.X., X.-b.L., X.-y.H., W.Z., J.W., H.Y.) yuvascular@zju.edu.cn wangjianan111@zju.edu.cn.
Abstract
RATIONALE: Vascular calcification (VC) is a marker of the severity of atherosclerotic disease. Hormones play important roles in regulating calcification; estrogen and parathyroid hormones exert opposing effects, the former alleviating VC and the latter exacerbating it. To date no treatment strategies have been developed to regulate clinical VC. OBJECTIVE: The objective of this study was to investigate the effect of growth hormone-releasing hormone (GHRH) and its agonist (GHRH-A) on the blocking of VC in a mouse model. METHODS AND RESULTS: Young adult osteoprotegerin-deficient mice were given daily subcutaneous injections of GHRH-A (MR409) for 4 weeks. Significant reductions in calcification of the aortas of MR409-treated mice were paralleled by markedly lower alkaline phosphatase activity and a dramatic reduction in the expression of transcription factors, including the osteogenic marker gene Runx2 and its downstream factors, osteonectin and osteocalcin. The mechanism of action of GHRH-A was dissected in smooth muscle cells isolated from human and mouse aortas. Calcification of smooth muscle cells induced by osteogenic medium was inhibited in the presence of GHRH or MR409, as evidenced by reduced alkaline phosphatase activity and Runx2 expression. Inhibition of calcification by MR409 was partially reversed by MIA602, a GHRH antagonist, or a GHRH receptor-selective small interfering RNA. Treatment with MR409 induced elevated cytosolic cAMP and its target, protein kinase A which in turn blocked nicotinamide adenine dinucleotide phosphate oxidase activity and reduced production of reactive oxygen species, thus blocking the phosphorylation of nuclear factor κB (p65), a key intermediate in the ligand of receptor activator for nuclear factor-κ B-Runx2/alkaline phosphatase osteogenesis program. A protein kinase A-selective small interfering RNA or the chemical inhibitor H89 abolished these beneficial effects of MR409. CONCLUSIONS: GHRH-A controls osteogenesis in smooth muscle cells by targeting cross talk between protein kinase A and nuclear factor κB (p65) and through the suppression of reactive oxygen species production that induces the Runx2 gene and alkaline phosphatase. Inflammation-mediated osteogenesis is thereby blocked. GHRH-A may represent a new pharmacological strategy to regulate VC.
RATIONALE: Vascular calcification (VC) is a marker of the severity of atherosclerotic disease. Hormones play important roles in regulating calcification; estrogen and parathyroid hormones exert opposing effects, the former alleviating VC and the latter exacerbating it. To date no treatment strategies have been developed to regulate clinical VC. OBJECTIVE: The objective of this study was to investigate the effect of growth hormone-releasing hormone (GHRH) and its agonist (GHRH-A) on the blocking of VC in a mouse model. METHODS AND RESULTS: Young adult osteoprotegerin-deficient mice were given daily subcutaneous injections of GHRH-A (MR409) for 4 weeks. Significant reductions in calcification of the aortas of MR409-treated mice were paralleled by markedly lower alkaline phosphatase activity and a dramatic reduction in the expression of transcription factors, including the osteogenic marker gene Runx2 and its downstream factors, osteonectin and osteocalcin. The mechanism of action of GHRH-A was dissected in smooth muscle cells isolated from human and mouse aortas. Calcification of smooth muscle cells induced by osteogenic medium was inhibited in the presence of GHRH or MR409, as evidenced by reduced alkaline phosphatase activity and Runx2 expression. Inhibition of calcification by MR409 was partially reversed by MIA602, a GHRH antagonist, or a GHRH receptor-selective small interfering RNA. Treatment with MR409 induced elevated cytosolic cAMP and its target, protein kinase A which in turn blocked nicotinamide adenine dinucleotide phosphate oxidase activity and reduced production of reactive oxygen species, thus blocking the phosphorylation of nuclear factor κB (p65), a key intermediate in the ligand of receptor activator for nuclear factor-κ B-Runx2/alkaline phosphatase osteogenesis program. A protein kinase A-selective small interfering RNA or the chemical inhibitor H89 abolished these beneficial effects of MR409. CONCLUSIONS:GHRH-A controls osteogenesis in smooth muscle cells by targeting cross talk between protein kinase A and nuclear factor κB (p65) and through the suppression of reactive oxygen species production that induces the Runx2 gene and alkaline phosphatase. Inflammation-mediated osteogenesis is thereby blocked. GHRH-A may represent a new pharmacological strategy to regulate VC.
Authors: Xianyang Zhang; Tengjiao Cui; Jinlin He; Haibo Wang; Renzhi Cai; Petra Popovics; Irving Vidaurre; Wei Sha; Janine Schmid; Barbara Ludwig; Norman L Block; Stefan R Bornstein; Andrew V Schally Journal: Proc Natl Acad Sci U S A Date: 2015-10-16 Impact factor: 11.205
Authors: Victoria Florea; Sonia S Majid; Rosemeire M Kanashiro-Takeuchi; Ren-Zhi Cai; Norman L Block; Andrew V Schally; Joshua M Hare; Claudia O Rodrigues Journal: Proc Natl Acad Sci U S A Date: 2014-11-17 Impact factor: 11.205
Authors: Renzhi Cai; Andrew V Schally; Tengjiao Cui; Luca Szalontay; Gabor Halmos; Wei Sha; Magdolna Kovacs; Miklos Jaszberenyi; Jinlin He; Ferenc G Rick; Petra Popovics; Rosemeire Kanashiro-Takeuchi; Joshua M Hare; Norman L Block; Marta Zarandi Journal: Peptides Date: 2013-12-25 Impact factor: 3.750