title: Modulation of human valve interstitial cell phenotype and function using a fibroblast growth factor 2 formulation.
abstract: Valve interstitial cells (VICs) are fibroblastic in nature however in culture it is widely accepted that they differentiate into a myofibroblastic phenotype. This study assessed a fibroblast culture media formulation for its ability to maintain the phenotype and function of VICs as in the intact healthy valve. Normal human VICs were cultured separately in standard DMEM and in fibroblast media consisting of FGF2 (10 ng/ml), insulin (50 ng/ml) and 2% FCS for at least a week. Cell morphology, aspect ratio, size, levels and distribution of protein expression, proliferation, cell cycle, contraction and migration were assessed. Some VICs and some valve endothelial cells expressed FGF2 in valve tissue and this expression was increased in calcified valves. VICs in DMEM exhibited large, spread cells whereas VICs in fibroblast media were smaller, elongated and spindly. Aspect ratio and size were both significantly higher in DMEM (p<0.01). The level of expression of α-SMA was significantly reduced in fibroblast media at day 2 after isolation (p<0.01) and the expression of α-SMA, SM22 and EDA-fibronectin was significantly reduced in fibroblast media at days 7 and 12 post-isolation (p<0.01). Expression of cytoskeletal proteins, bone marker proteins and extracellular matrix proteins was reduced in fibroblast media. Proliferation of VICs in fibroblast media was significantly reduced at weeks 1 (p<0.05) and 2 (p<0.01). Collagen gel contraction was significantly reduced in fibroblast media (p<0.05). VICs were found to have significantly fewer and smaller focal adhesions in fibroblast media (p<0.01) with significantly fewer supermature focal adhesions in fibroblast media (p<0.001). Ultrastructurally, VICs in fibroblast media resembled native VICs from intact valves. VICs in fibroblast media demonstrated a slower migratory ability after wounding at 72 hours (p<0.01). Treatment of human VICs with this fibroblast media formulation has the ability to maintain and to dedifferentiate the VICs back to a fibroblastic phenotype with phenotypic and functional characteristics ascribed to cells in the intact valve. This methodology is fundamental in the study of normal valve biology, pathology and in the field of tissue engineering.
authors.affiliation: Imperial College, Heart Science Centre, Harefield Hospital, Harefield, Middx, UB9 6JH, United Kingdom; QCRC, Qatar Cardiovascular Research Centre, Qatar Foundation, Doha, Qatar.
authors.forename: Najma
authors.initials: N
authors.lastname: Latif
authors.affiliation: Département de Biologie, École Normale Supérieure de Lyon, Université de Lyon, UCB Lyon1, 46 Allée d'Italie, Lyon, France.
authors.forename: Alfred
authors.initials: A
authors.lastname: Quillon
authors.affiliation: Imperial College, Heart Science Centre, Harefield Hospital, Harefield, Middx, UB9 6JH, United Kingdom.
authors.forename: Padmini
authors.initials: P
authors.lastname: Sarathchandra
authors.affiliation: Imperial College, Heart Science Centre, Harefield Hospital, Harefield, Middx, UB9 6JH, United Kingdom.
authors.forename: Ann
authors.initials: A
authors.lastname: McCormack
authors.affiliation: Imperial College, Heart Science Centre, Harefield Hospital, Harefield, Middx, UB9 6JH, United Kingdom.
authors.forename: Alec
authors.initials: A
authors.lastname: Lozanoski
authors.affiliation: Imperial College, Heart Science Centre, Harefield Hospital, Harefield, Middx, UB9 6JH, United Kingdom; QCRC, Qatar Cardiovascular Research Centre, Qatar Foundation, Doha, Qatar.
authors.forename: Magdi H
authors.initials: MH
authors.lastname: Yacoub
authors.affiliation: Imperial College, Heart Science Centre, Harefield Hospital, Harefield, Middx, UB9 6JH, United Kingdom; QCRC, Qatar Cardiovascular Research Centre, Qatar Foundation, Doha, Qatar.
authors.forename: Adrian H
authors.initials: AH
authors.lastname: Chester
mesh_terms: D000199:Actins / Q000378:metabolism; D000293:Adolescent; D000328:Adult; D001021:Aortic Valve / Q000166:cytology*; D002465:Cell Movement / Q000187:drug effects; D049109:Cell Proliferation / Q000187:drug effects; D002478:Cells, Cultured; D002648:Child; D003470:Culture Media / Q000494:pharmacology; D016222:Fibroblast Growth Factor 2 / Q000378:metabolism*; D005434:Flow Cytometry; D022001:Focal Adhesions / Q000187:drug effects / Q000378:metabolism; D006801:Humans; D056885:Interstitial Cells of Cajal / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D008875:Middle Aged; D010641:Phenotype; D013997:Time Factors; D055815:Young Adult
chemical_list: C541116:ACTA2 protein, human; D000199:Actins; D003470:Culture Media; D016222:Fibroblast Growth Factor 2