Abstract
Atherosclerosis is the leading cause of disability and death worldwide. The whole spectrum of atherosclerosis develops through series of highly specific cellular and molecular events that lead to the formation and progression of atherosclerotic plaque and finally its complications. Coronary artery disease (CAD), most common among cardiovascular diseases, is characterized by insufficient oxygen supply to the heart muscle which primarily occurs due to coronary artery atherosclerosis. During last three decades, percutaneous coronary intervention has become the major strategy to treat coronary artery disease but, restenosis (re-narrowing of the vessel after an interventional procedure) is the major limitation of this approach. Although, the rate of restenosis is significantly reduced by stent implantation, especially drug eluting stents, there is a concern that drug eluting stents might increase the rate of in-stent thrombosis, a potentially fatal complication. Neointimal hyperplasia, a cell proliferation and differentiation process, is the predominant mechanism in the development of in-stent restenosis. |Vitamin D is a secosteroid which functions through vitamin D receptor (VDR), a transcription factor, and directly or indirectly controls more than 200 heterogeneous genes including genes that regulate cellular differentiation, proliferation, and angiogenesis. Vitamin D receptors are distributed in a variety of tissues including, vascular smooth muscle cells (VSMCs), cardiomyocytes, endothelium, and cells of immune system. The growth suppressant and immunomodulatory effects of calcitriol are of great interest because of their potential use in the management of disorders, including post-interventional restenosis, atherosclerosis and post-transplant vasculopathy in which the underlying pathological mechanisms are uncontrolled cell growth and remodeling in the vascular wall. |The central hypothesis is that calcitriol inhibits proliferation, migration and phenotypic modulation of porcine coronary artery smooth muscle cells (PCASMCs) through VDR and that vitamin D supplementation reduces the incidence of restenosis by decreasing neointimal hyperplasia after coronary artery intervention in coronary artery disease. |In this study, the effect of calcitriol stimulation on the expression of VDR and vitamin D metabolizing enzymes including CYP24A1 and CYP27B1 was examined in cultured PCASMCs. Further, the effect of calcitriol stimulation on cell proliferation, migration, phenotypic modulation and apoptosis was investigated in cultured PCASMCs. Next, expression of TNF-α and VDR in the neointimal lesions in postintervention hypercholesterolemic swine coronary arteries were examined. Finally, the effect of vitamin D deficiency and vitamin D supplementation on the development of post-intervention restenosis was investigated in a well-controlled atherosclerotic swine model of coronary restenosis. |PCASMCs express VDR and vitamin D metabolizing enzymes. Treatment of PCASMCs with calcitriol significantly increased the mRNA and protein expression of VDR and CYP24A1 in a dose-dependent manner while expression of CYP27B1 was significantly decreased as compared to control. Calcitriol treatment significantly decreased serum-induced proliferation of PCASMCs and has no effect on the apoptosis in these cells. Calcitriol also decreased PDGF-BB-induced proliferation, migration and phenotypic modulation in PCASMCs. In vivo morphometric analysis of tissues revealed that coronary intervention in hypercholesterolemic Yucatan miniature swine induced significant restenosis. Histological evaluation of post-intervention swine coronary arteries showed expression of smooth muscle α-actin and significantly increased expression of TNF-α in neointimal lesions. Interestingly, there was significantly decreased expression of VDR in PCASMCs of neointimal region compared to normal media. Vitamin D deficiency increased the magnitude of restenosis and PCNA-positive cells in neointimal tissue of post-intervention coronary arteries, which is suppressed by supplementation of vitamin D post-intervention. Vitamin D supplementation significantly downregulated the levels of TNF-α and IFN-γ, upregulated the levels of IL-10, and had no effect on serum IL-6 levels. |These data suggest that calcitriol inhibits proliferation in PCASMCs through VDR and there is significant downregulation of VDR in proliferating PCASMCs of neointimal lesions. Thus, downregulation of VDR in VSMCs of post-interventional arteries due to high concentration of TNF-α could be a potentially contributing factor for uncontrolled growth of VSMCs in injured arteries leading to neointimal hyperplasia and restenosis. Expression of VDR is increased by VDR ligands and growth inhibitory and immunomodulatory actions of VDR may likely be enhanced in the presence of vitamin D ligands. Therefore, anti-proliferative effect of VDR ligands can prevent/decrease VSMC proliferation after mechanical injury to the artery and attenuate restenosis. This could be an inexpensive and safe therapeutic approach for reduction in cardiovascular disease burden.