Key result
Culture of porcine valvular interstitial cells in a fibroblast media formulation on collagen coatings significantly reduced markers of activation, generating quiescent cells that mimic a healthy phenotype.
A novel culture method using fibroblast media and collagen coatings generates quiescent valvular interstitial cells, enabling more accurate in vitro modeling of early calcific aortic valve disease.
May advance physiologic in vitro VIC models; leaves open translation to calcific aortic valve disease.
BACKGROUND: Valvular interstitial cells (VICs) in the healthy aortic valve leaflet exhibit a quiescent phenotype, with <5% of VICs exhibiting an activated phenotype. Yet, in vitro culture of VICs on tissue culture polystyrene surfaces in standard growth medium results in rapid transformation to an activated phenotype in >90% of cells. The inability to preserve a healthy VIC phenotype during in vitro studies has hampered the elucidation of mechanisms involved in calcific aortic valve disease. This study describes the generation of quiescent populations of porcine VICs in 2-dimensional in vitro culture and their utility in studying valve pathobiology. METHODS AND RESULTS: Within 4 days of isolation from fresh porcine hearts, VICs cultured in standard growth conditions were predominantly myofibroblastic (activated VICs). This myofibroblastic phenotype was partially reversed within 4 days, and fully reversed within 9 days, following application of a combination of a fibroblast media formulation with culture on collagen coatings. Specifically, culture in this combination significantly reduced several markers of VIC activation, including proliferation, apoptosis, α-smooth muscle actin expression, and matrix production, relative to standard growth conditions. Moreover, VICs raised in a fibroblast media formulation with culture on collagen coatings exhibited dramatically increased sensitivity to treatment with transforming growth factor β1, a known pathological stimulus, compared with VICs raised in either standard culture or medium with a fibroblast media formulation. CONCLUSIONS: The approach using a fibroblast media formulation with culture on collagen coatings generates quiescent VICs that more accurately mimic a healthy VIC population and thus has the potential to transform the study of the mechanisms of VIC activation and dysfunction involved in the early stages of calcific aortic valve disease.
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Porras et al. (2017) studied In vitro culture of valvular interstitial cells (n=29). Fibroblast media formulation with culture on collagen coatings (FIB-Coll) vs. Standard growth medium (Control) and FIB media alone was evaluated on Reversal of myofibroblastic phenotype (quiescence) measured by markers of activation. Culture of porcine valvular interstitial cells in a fibroblast media formulation on collagen coatings significantly reduced markers of activation, generating quiescent cells that mimic a healthy phenotype.