Abstract 3530: Wnt3a Modulates Human Valve Interstitial Cell Growth and Phenotype: Relevance to Valve Calcification
Valve Calcification is a complex process which involves the transformation of valve cells into an osteogenic phenotype. The Wnt signalling pathway has been shown to play an important role in valve development and bone formation through binding of the Wnt proteins to the frizzled receptors (FZDs) and/or co-receptors LRP5/6. We have previously demonstrated the expression of Wnts and their receptors, FZDs, and several secreted Wnt inhibitors such as SFRP-1,3,4 and DKK-1,2,3,4 in valve interstitial cells (ICs). However, the exact role of Wnt pathway in valve calcification remains unknown. Primary cultures of human aortic valve ICs were treated with 2, 4, 10 and 20 ng/ml of Wnt3a for 24 hrs and demonstrated a concentration-dependent increase of 213% in proliferation from 359±47.1 cpm (2 ng/ml) to 768±28.6 cpm (20 ng/ml) (P<0.01, n=3). Treatment with higher concentration of Wnt3a (40ng/ml), human valve ICs differentiated into an osteogenic cell phenotype with increased expression of the osteoblast cell markers, alkaline phosphatase (ALP) and osteocalcin. There was a significant 5-fold increase in the activity of ALP in response to Wnt3a (40ng/ml) compared to untreated cells (P<0.05, n=3). Immunocytochemical staining of human valve ICs also showed that Wnt3a treatment (40ng/ml) increased the expression of osteocalcin in comparison to untreated cells (n=3). In addition, treatment of valve ICs with atorvastatin (100μM) in the presence of Wnt3a (40ng/ml) reduced the ALP activity from 9.4±1.04 nmol/min/mg protein in the osteogenic treated cells to 2.4±1.9 nmol/min/mg protein (P<0.04, n=3) and inhibited the expression of osteocalin after 21 days. This study demonstrates the importance of Wnt3a in modulating growth and in changing human valve ICs phenotype towards osteoblasts. These findings suggest a key role of Wnt3a in the calcification process and possibly identify new therapeutic targets.