Inhibition of Pathological Differentiation of Valvular Interstitial Cells by C-Type Natriuretic Peptide

Abstract

Objective-Calcific aortic valve disease is associated with the differentiation of valvular interstitial cells (VICs) to myofibroblast and osteoblast-like cells, particularly in the fibrosa layer of the valve. Previous studies suggested that C-type natriuretic peptide (CNP) protects against calcific aortic valve disease to maintain homeostasis. We aimed to determine whether CNP inhibits VIC pathological differentiation as a mechanism to explain its protective effects. Methods and Results-CNP expression was prominent in normal porcine aortic valves, particularly on the ventricular side, but reduced in sclerotic valves concomitant with the appearance of pathological VIC phenotypes in the fibrosa. In vitro, CNP inhibited calcified aggregate formation and bone-related transcript and protein expression by VICs grown in osteogenic conditions. Under myofibrogenic culture conditions, CNP reduced ␣-smooth muscle actin expression and cell-mediated gel contraction, indicating inhibition of myofibroblast differentiation. Similar to CNP, simvastatin inhibited VIC osteoblast and myofibroblast differentiation in vitro. Strikingly, simvastatin upregulated CNP expression in VICs cultured under myofibrogenic conditions, and small interfering RNA knockdown of natriuretic peptide receptor-b (a CNP receptor) significantly reduced the antifibrotic effect of simvastatin, suggesting that it acts in part via CNP/NPR-B autocrine/paracrine signaling. 1 The normal VIC population is heterogeneous, consisting primarily of fibroblasts, a subpopulation of which consists of mesenchymal progenitor cells with multilineage differentiation potential, 2 and a small population (Ϸ1% to 5%) of smooth muscle cells and myofibroblasts. [3][4][5][6] VICs undergo myofibroblast and osteoblast differentiation during the progression of CAVD, contributing to fibrosis and calcification. 5,7 Myofibroblasts, which are up to 30% of the total VIC population in sclerotic valves, 3,5 are identified by prominent ␣-smooth muscle actin (␣-SMA) stress fibers and are associated with increased collagen synthesis and cellular contractility. 8 Osteoblast-like cells are also often found in calcified aortic valves, 7,9 accompanied by an increase in the expression of bone-related transcripts and proteins. Conclusion-CNPPathological differentiation of VICs has been shown to be inhibited by statins in vitro 10 -12 ; however, clinical trials with statins have yielded conflicting results. [13][14][15][16][17] Retrospective and prospective cohort studies show that statins delay progression of CAVD, 16,18 whereas randomized, controlled trials such as Simvastatin and Ezetimibe for Aortic Stenosis and Scottish Aortic Stenosis and Lipid Lowering Trial, Impact on Regression found no benefit with statin therapy. 14,15,19 An improved understanding of the molecular determinants leading to pathological differentiation of VICs may provide insights into the potential of statins or other pharmacological therapies to arrest CAVD, which will eventually aid in refining treatment regimens and pat...

Hemodynamic and Cellular Response Feedback in Calcific Aortic Valve Disease

“…93 In culture, CNP prevents VICs from differentiating to the myofibroblasts and osteoblast lineages, suggesting that it plays a homeostatic role in the aortic valve. 94 Indeed, CNP expression is distinctly downregulated in stenotic human valves. 95 The implications from these studies are 2-fold.…”

A time course investigation of the statin paradox among valvular interstitial cell phenotypes

“…Relatively few studies have examined the statin-valve relationship on a cellular level (2,12,23,24). One previous publication documented a "statin paradox," wherein the response of a clonal population of porcine aortic valve myofibroblasts to statin consisted of a decrease in mineralization markers [namely, nodule counts and alkaline phosphatase (ALP) production], whereas murine osteoblast precursor cells responded to statin by increasing these same markers (23).…”

“…Although statin treatment of valve cells has been evaluated in other studies performed to investigate different hypotheses (2,12,24), one limiting factor in all of these studies is that they all have only examined the effects of statin treatment at a single time point, thereby limiting the obtainable information about the relationship between valves and statin treatment.…”

Biophysical Analysis of Dystrophic and Osteogenic Models of Valvular Calcification

“…Osteogenic calcification is present in 13% of valves containing dystrophic calcification and is identified by the pres ence of osteoid matrix reminiscent of active bone formation [6]. At present, there exists only surgical intervention for CAVD, which although effective, includes a 3% mortality rate and is only utilized at the end stage disease [7,8], Efforts to describe the mechanisms of valvular calcification may lead to the development of novel pharmacological treatments for CAVD [9][10][11].…”