β-catenin mediates mechanically regulated, transforming growth factor-β1-induced myofibroblast differentiation of aortic valve interstitial cells.
OBJECTIVE
In calcific aortic valve disease, myofibroblasts and activation of the transforming growth factor-β1 (TGF-β1) and Wnt/β-catenin pathways are observed in the fibrosa, the stiffer layer of the leaflet, but their association is unknown. We elucidated the roles of β-catenin and extracellular matrix stiffness in TGF-β1-induced myofibroblast differentiation of valve interstitial cells (VICs).


RESULTS
TGF-β1 induced rapid β-catenin nuclear translocation in primary porcine aortic VICs in vitro through TGF-β receptor I kinase. Degrading β-catenin pharmacologically or silencing it with small interfering RNA inhibited TGF-β1-induced myofibroblast differentiation without altering Smad2/3 activity. Conversely, increasing β-catenin availability with Wnt3A alone did not induce differentiation. However, combining TGF-β1 and Wnt3A caused greater myofibroblast differentiation than TGF-β1 treatment alone. Notably, in VICs grown on collagen-coated PA gels with physiological stiffnesses, TGF-β1-induced β-catenin nuclear translocation and myofibroblast differentiation occurred only on matrices with fibrosa-like stiffness, but not ventricularis-like stiffness. In diseased aortic valves from pigs fed an atherogenic diet, myofibroblasts colocalized with increased protein expression of Wnt3A, β-catenin, TGF-β1, and phosphorylated Smad2/3 in the fibrosa.


CONCLUSIONS
Myofibroblast differentiation of VICs involves matrix stiffness-dependent crosstalk between TGF-β1 and Wnt signaling pathways and may explain in part why the stiffer fibrosa is more susceptible to disease.
Institute of Biomaterials and Biomedical Engineering, University of Toronto, Toronto, Ontario, Canada.
Jan-Hung
JH
Chen
Wen Li Kelly
WL
Chen
Krista L
KL
Sider
Cindy Ying Yin
CY
Yip
Craig A
CA
Simmons
D021581:Active Transport, Cell Nucleus; D000818:Animals; D001021:Aortic Valve / Q000378:metabolism* / Q000473:pathology; D054338:Cell Transdifferentiation; D002478:Cells, Cultured; D003094:Collagen / Q000378:metabolism; D004195:Disease Models, Animal; D004548:Elasticity; D005109:Extracellular Matrix / Q000378:metabolism*; D006349:Heart Valve Diseases / Q000378:metabolism* / Q000473:pathology; D058628:Myofibroblasts / Q000378:metabolism* / Q000473:pathology; D010766:Phosphorylation; D017346:Protein Serine-Threonine Kinases / Q000378:metabolism; D034622:RNA Interference; D000077293:Receptor, Transforming Growth Factor-beta Type I; D018125:Receptors, Transforming Growth Factor beta / Q000378:metabolism; D012598:Sclerosis; D015398:Signal Transduction; D051899:Smad2 Protein / Q000378:metabolism; D051900:Smad3 Protein / Q000378:metabolism; D013552:Swine; D013997:Time Factors; D053773:Transforming Growth Factor beta1 / Q000378:metabolism*; D051153:Wnt Proteins / Q000378:metabolism; D060508:Wnt3 Protein; D051176:beta Catenin / Q000235:genetics / Q000378:metabolism*
D018125:Receptors, Transforming Growth Factor beta; D051899:Smad2 Protein; D051900:Smad3 Protein; D053773:Transforming Growth Factor beta1; D051153:Wnt Proteins; D060508:Wnt3 Protein; D051176:beta Catenin; D003094:Collagen; D017346:Protein Serine-Threonine Kinases; D000077293:Receptor, Transforming Growth Factor-beta Type I