Statins Block Calcific Nodule Formation of Valvular Interstitial Cells by Inhibiting α-Smooth Muscle Actin Expression

Abstract

Objective Calcific aortic stenosis, characterized by excessive fibrosis and deposition of bone-like calcified tissue, affects roughly 2–3% of the US population over 65. Recent studies have suggested that statins have a positive effect on the progression of aoritic stenosis, likely due to their ability to affect the resident cell population, known as valvular interstitial cells (VICs). VICs are fibroblastic cells that can differentiate to form activated myofibroblasts, displaying increased alpha smooth muscle actin (αSMA) expression, contractility, and collagen production. Methods and Results In culture, VICs spontaneously form multicellular aggregates that subsequently develop into calcified nodules, providing an in vitro model for aortic stenosis. Using real-time microscopic tracking, we observed that confluent VIC monolayers spontaneously contract into rounded nodules, suggesting that myofibroblastic contractility is a critical step in the process of nodule formation. Over-expression of αSMA increased VIC calcific nodule formation and contractility, while knock-down of αSMA with siRNAs reduced these phenotypes, suggesting that the expression and contractile properties of αSMA are essential to the formation of nodules. Statin treatment of VICs reduced αSMA expression, inhibited contractility, and decreased nodule formation. When statins were used to treat preformed nodules, no decrease in the number of calcified nodules was observed, suggesting that statins may play more of a preventative role in aortic stenosis than a cure. Conclusions Our studies provide evidence of a causal relationship between VIC myofibroblastic activity and initial VIC calcific nodule formation. Furthermore we demonstrate that pravastatin inhibition of calcific nodule formation is related to inhibition of myofibroblastic activity.

Multilayer three-dimensional filter paper constructs for the culture and analysis of aortic valvular interstitial cells

“…With valve replacement as the only viable option, recent research has focused on more robust tissue valve replacements and noninvasive methods for treating or preventing valve disease [1,2]. Aortic valvular interstitial cells (VICs) have been targeted in studies on valve calcification because of their role in the production and turnover of leaflet extracellular matrix (ECM) [3,4].…”

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.…”

Apposite Insulin-like Growth Factor (IGF) Receptor Glycosylation Is Critical to the Maintenance of Vascular Smooth Muscle Phenotype in the Presence of Factors Promoting Osteogenic Differentiation and Mineralization

“…This hypothesis is consistent with a recent study showing that atorvastatin promotes osteogenic differentiation and calcium deposition by VSMCs (18) and osteoblast cell lines (19,20) in vitro. However, there are also reports that HMG-CoA reductase inhibitors inhibit the osteoblastic conversion of human VSMCs (21,22) and TGF␤-induced calcific nodule formation by valvular interstitial cells (23,24). The reason for these discrepancies is not clear but may reflect differences in the factors used to stimulate differentiation in these studies.…”