Activated p300 acetyltransferase activity modulates aortic valvular calcification with osteogenic transdifferentiation and downregulation of Klotho Shao-Jung Li a,b , Yu-Hsun Kao a,c, ⁎, Cheng-Chih Chung a,d , Wei-Yu Chen e,f , Wan-li Cheng a , Yi-Jen Chen a,d, ⁎ a Grarduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan b Division of Cardiovascular Surgery, Department of Surgery, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan c Department of Medical Education and Research, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan d Division of Cardiovascular Medicine, Department of Internal Medicine, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan e Department of Pathology, School of Medicine, College of Medicine, Taipei Medical University, Taipei, Taiwan f Department of Pathology, Wan Fang Hospital, Taipei Medical University, Taipei, Taiwan abstract article info Article history: Received 2 August 2016 Received in revised form 30 December 2016 Accepted 3 January 2017 Available online xxxx Background: The calcific aortic valve (AV) disease is a common disease with the unclear mechanism, and optimal pharmacological treatment remains unavailable. Epigenetic modulation by histone acetyltransferase (HAT) plays a critical role in osteogenic transdifferentiation and atherosclerosis. The purposes of this study were to investigate whether HAT contributes to the pathophysiology of AV calcification and assess the therapeutic potential of HAT inhibition. Methods: Porcine valvular interstitial cells (VICs) were treated with osteogenic medium (10 ng/mL of tumor ne- crosis factor-α and 4 mmol/L of high phosphate) for 7 days. We analyzed the RNA and protein expression of myofibroblastic (α-SMA, vimentin, collagen 1A1, collagen 3, Egr-1, MMP2, MMP9) and osteoblastic markers (osteocalcin and alkaline phosphatase) in VICs, and studied the effects of a p300 inhibitor (C646, 10 μmol/L) on calcification (Alizarin Red S staining), osteogenesis, HAT activity, the mitogen-activated protein kinase (MAPK) and Akt pathway, and Klotho expression on VICs. Results: Osteogenic medium treated VICs had higher expressions of osteocalcin, alkaline phosphatase and acety- lated lysine-9 of histone H3 (ac-H3K9) than control cells. C646 attenuated osteogenesis of VICs with simulta- neous inhibition of the HAT activity of p300. There was neither significant increase of p300 protein nor p300 transcript during the osteogenesis process. Additionally, osteogenic medium treated VICs decreased the expres- sion of Klotho, which is attenuated by C646. Conclusions: Activated HAT activity of p300 modulates AV calcification through osteogenic transdifferentiation of VICs with Klotho modulation. P300 inhibition is a potential therapeutic target for AV calcification. © 2017 Elsevier B.V. All rights reserved. Keywords: Aortic valve calcification Klotho Osteogenesis P300 Valvular interstitial cells 1. Introduction Calcific aortic valve (AV) disease (CAVD) is a leading valvular heart disease in older adults and has a poor prognosis [1]. Diseased AV leaflets finally form a restricted opening of the outflow tract of the left ventricle, which leads to congestive heart failure or sudden cardiac death. CAVD arises from an active disease process with interactions between fibrosis and calcification through the mediation of extracellular matrix (ECM) remodeling, inflammation, cell apoptosis, and dystrophic calcification. Currently, no effective medication can prevent or reverse the disease progression of CAVD [2]. Invasive surgical procedures with AV replace- ment or transcatheter AV implantation are the gold standard treatment of CAVD, but both are associated with variable risks. Moreover, the cru- cial pathophysiological process of CAVD continually damages the im- planted bioprosthesis even after the native valve is exenterated [3]. International Journal of Cardiology xxx (2017) xxx–xxx Abbreviations: ac-H3K9, histone acetylation at histone H3 Lys 9; AV, aortic valve; CAVD, calcific aortic valve disease; ECM, extracellular matrix; Egr-1, early growth response-1; ERK, extracellular signal-regulated kinase; HAT, histone acetyltransferase; HDAC, histone deacetylase; HP, high phosphate; IκB, inhibitor of kappa B; MAPK, mitogen-activated protein kinase; MMP, matrix metalloproteinase; NF-κB, nuclear factor kappa-light-chain-enhancer of activated B cells; p300, EP300 or E1A binding protein p300 (adenovirus early region 1A); CBP or CREBBP, CREB (cAMP response element- binding protein) -binding protein; Pi, phosphate; TGF, transforming growth factor; TNAP, tissue non-specific alkaline phosphatases; TNF, tumor necrosis factor; RXR, retinoid-x receptor; VIC, valve interstitial cell; VDR, vitamin D (1,25-dihydroxy vitamin D3) receptor; VDRE, vitamin D response element. ⁎ Corresponding authors at: Graduate Institute of Clinical Medicine, College of Medicine, Taipei Medical University, Taiwan, 250 Wu-Xing Street, Taipei 11031, Taiwan. E-mail addresses: yuhsunkao@gmail.com (Y.-H. Kao), a9900112@ms15.hinet.net (Y.-J. Chen). IJCA-24370; No of Pages 9 http://dx.doi.org/10.1016/j.ijcard.2017.01.005 0167-5273/© 2017 Elsevier B.V. All rights reserved. Contents lists available at ScienceDirect International Journal of Cardiology journal homepage: www.elsevier.com/locate/ijcard Please cite this article as: S.-J. Li, et al., Activated p300 acetyltransferase activity modulates aortic valvular calcification with osteogenic transdifferentiation and downreg..., Int J Cardiol (2017), http://dx.doi.org/10.1016/j.ijcard.2017.01.005