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Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures

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 a...

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Autores principales: Porras, Ana M., van Engeland, Nicole C. A., Marchbanks, Evelyn, McCormack, Ann, Bouten, Carlijn V. C., Yacoub, Magdi H., Latif, Najma, Masters, Kristyn S.
Formato: Online Artículo Texto
Lenguaje:English
Publicado: John Wiley and Sons Inc. 2017
Materias:
Acceso en línea:https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5524027/
https://www.ncbi.nlm.nih.gov/pubmed/28292746
http://dx.doi.org/10.1161/JAHA.116.005041
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author Porras, Ana M.
van Engeland, Nicole C. A.
Marchbanks, Evelyn
McCormack, Ann
Bouten, Carlijn V. C.
Yacoub, Magdi H.
Latif, Najma
Masters, Kristyn S.
author_facet Porras, Ana M.
van Engeland, Nicole C. A.
Marchbanks, Evelyn
McCormack, Ann
Bouten, Carlijn V. C.
Yacoub, Magdi H.
Latif, Najma
Masters, Kristyn S.
author_sort Porras, Ana M.
collection PubMed
description 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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spelling pubmed-55240272017-08-15 Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures Porras, Ana M. van Engeland, Nicole C. A. Marchbanks, Evelyn McCormack, Ann Bouten, Carlijn V. C. Yacoub, Magdi H. Latif, Najma Masters, Kristyn S. J Am Heart Assoc Original Research 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. John Wiley and Sons Inc. 2017-03-14 /pmc/articles/PMC5524027/ /pubmed/28292746 http://dx.doi.org/10.1161/JAHA.116.005041 Text en © 2017 The Authors. Published on behalf of the American Heart Association, Inc., by Wiley Blackwell. This is an open access article under the terms of the Creative Commons Attribution‐NonCommercial‐NoDerivs (http://creativecommons.org/licenses/by-nc-nd/4.0/) License, which permits use and distribution in any medium, provided the original work is properly cited, the use is non‐commercial and no modifications or adaptations are made.
spellingShingle Original Research
Porras, Ana M.
van Engeland, Nicole C. A.
Marchbanks, Evelyn
McCormack, Ann
Bouten, Carlijn V. C.
Yacoub, Magdi H.
Latif, Najma
Masters, Kristyn S.
Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures
title Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures
title_full Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures
title_fullStr Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures
title_full_unstemmed Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures
title_short Robust Generation of Quiescent Porcine Valvular Interstitial Cell Cultures
title_sort robust generation of quiescent porcine valvular interstitial cell cultures
topic Original Research
url https://www.ncbi.nlm.nih.gov/pmc/articles/PMC5524027/
https://www.ncbi.nlm.nih.gov/pubmed/28292746
http://dx.doi.org/10.1161/JAHA.116.005041
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