title: Inhibition of calcifying nodule formation in cultured porcine aortic valve cells by nitric oxide donors.
abstract: Calcific aortic stenosis displays some similarities to atherosclerosis including evidence of endothelial dysfunction. Whether nitric oxide (NO), which is produced by valvular endothelium, has direct protective effects extending to calcification processes in aortic valve cells has not previously been examined. In vitro calcifying nodules in porcine aortic valve interstitial cell cultures, formed in response to transforming growth factor-beta1 (TGF-beta1) 5 ng/ml, were inhibited by NO donors DETA-NONOate 5-100 microM, and sodium nitroprusside (SNP) 3 microM. Raising intracellular cGMP concentrations, via 8-bromo cGMP 1 mM or via brain natiuretic peptide and C-type natiuretic peptide 0.1 microM, inhibited TGF-beta1-induced nodule formation, potentially implicating the cGMP pathway in the NO effect. Stimulation of interstitial cells with substance P or calcium ionophone (A23187) caused NO release and increased intracellular cGMP respectively. However in the presence of TGF-beta1 basal levels of NO production via nitric oxide synthase (NOS) were insufficient to affect nodule formation. Increased dihydroethidium (DHE) fluorescence in response to TGF-beta1, which was inhibited by DETA-NONOate and TEMPOL, suggested a role for intracellular superoxide in TGF-beta1 signalling. Moreover, nodule formation was suppressed by superoxide scavengers TEMPOL, hydralazine and polyethylene glycol-superoxide dismutase (PEG-SOD), but not SOD. In conclusion, NO donors, or agents raising intracellular cGMP levels, may protect aortic valve interstitial cells from early events leading to calcification.
authors.affiliation: Cardiology Unit, The Queen Elizabeth Hospital, Woodville South, South Australia, Australia. Jenny.Kennedy@nwahs.sa.gov.au
authors.forename: Jennifer A
authors.initials: JA
authors.lastname: Kennedy
authors.forename: Xiang
authors.initials: X
authors.lastname: Hua
authors.forename: Kumaril
authors.initials: K
authors.lastname: Mishra
authors.forename: Geraldine A
authors.initials: GA
authors.lastname: Murphy
authors.forename: Anke C
authors.initials: AC
authors.lastname: Rosenkranz
authors.forename: John D
authors.initials: JD
authors.lastname: Horowitz
mesh_terms: D000818:Animals; D001021:Aortic Valve / Q000166:cytology* / Q000378:metabolism / Q000473:pathology; D002114:Calcinosis / Q000378:metabolism* / Q000473:pathology*; D002478:Cells, Cultured; D006152:Cyclic GMP / Q000378:metabolism; D016166:Free Radical Scavengers / Q000494:pharmacology; D007476:Ionophores / Q000494:pharmacology; D018696:Neuroprotective Agents / Q000494:pharmacology; D009569:Nitric Oxide / Q000378:metabolism; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D009599:Nitroprusside / Q000494:pharmacology; D013552:Swine; D053773:Transforming Growth Factor beta1 / Q000037:antagonists & inhibitors / Q000378:metabolism; D014665:Vasodilator Agents / Q000494:pharmacology*
chemical_list: D016166:Free Radical Scavengers; D007476:Ionophores; D018696:Neuroprotective Agents; D020030:Nitric Oxide Donors; D053773:Transforming Growth Factor beta1; D014665:Vasodilator Agents; D009599:Nitroprusside; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D006152:Cyclic GMP