Activation of valvular interstitial cells is mediated by transforming growth factor-β1 interactions with matrix molecules

Strategies for the tissue-engineering of living cardiac valve replacements are limited by a lack of appropriate scaffold materials that both permit cell viability and actively contribute to the growth of functional tissues. Components of the extracellular matrix can localize and modify growth factor signals, and by doing so impart instructional stimuli for direction of cell phenotype. Fibronectin, collagen I, and heparin were explored as affinity matrices for sequestering and presenting soluble signaling molecules to control differentiation of valvular interstitial cells (VICs) to myofibroblasts. VIC differentiation is commonly characterized by expression of stress fibers containing alpha smooth muscle actin (α-SMA), and transforming growth factor-β1 (TGF-β1) is a central mediator of this transition. Both fibronectin and heparin, which are known to possess TGF-β1 binding interactions, were found to increase VIC α-SMA expression (120% and 258% of expression in controls), while VICs cultured on collagen I-modified substrates had diminished α-SMA expression (66% of control). Heparin treatment significantly stimulated VIC production of TGF-β1 at all concentrations tested (50 to 400 μg/ml). Heparin-modified substrates were found to alter cell morphology through increased adsorption of serum proteins, specifically TGF-β1. In sum, heparin produced α-SMA-positive myofibroblasts through both the de novo production of TGF-β1, and its localization in the pericellular environment. The addition of heparin to fibronectin-modified substrates led to a synergistic increase in VIC α-SMA expression, produced by the reciprocal binding of fibronectin, heparin, cell-produced TGF-β1. The characterization of molecules, both soluble and insoluble, that control VIC activation will be important for the development of tailored 3D culture environments for tissue-engineering applications.