Objective Calcific Aortic Valve Disease (CAVD) is the most prevalent type of heart valve disease, affecting ~2% of the US population. CAVD is characterized by the presence of calcific nodules resulting in aortic valve (AoV) stenosis; however, the underlying mechanisms driving disease remain unknown. Studies of human diseased AoV provide initial evidence that BMP signaling, essential for normal bone formation, is activated during CAVD. Mice deficient in Klotho, an FGF23 transmembrane co-receptor, exhibit premature aging and develop AoV calcific nodules as occurs in human CAVD. The role of BMP signaling in the development of CAVD was examined in porcine aortic valve interstitial cells (VICs) and Klotho−/− mice. Approach & Results We show that activation of BMP signaling, as indicated by pSmad1/5/8 expression, precedes and later localizes with AoV calcification in Klotho−/− mice. In addition, cellular and ECM changes resembling features of normal bone formation are accompanied by increased osteochondrogenic gene induction in calcified Klotho−/− AoV. Likewise, osteogenic media (OM) treatment of porcine VICs results in BMP pathway activation, increased osteochondrogenic gene induction and formation of calcific nodules in vitro. We demonstrate that genetic inactivation of the Bmpr1a receptor in Klotho−/− aortic VICs, as well as BMP pathway inhibition of OM-treated aortic VICs in vitro, results in the inhibition of AoV calcification. Discussion BMP signaling and osteochondrogenic gene induction are active in calcified Klotho−/− AoV in vivo and calcified porcine aortic VICs in vitro. Importantly, BMP signaling is required for the development of AoV calcification in vitro and in vivo.

The mature heart valves consist of stratified extracellular matrix (ECM) layers, and heart valve disease is characterized by ECM dysregulation and mineralization. There is increasing evidence that regulatory pathways that control heart valve development also are active in disease. In human diseased valves and mouse models, the expression of valve progenitor markers, including Twist1, Msx1/2 and Snail1/2, is induced. Additional markers of osteogenesis, including Runx2, osteocalcin and bone sialoprotein, also are expressed in calcific aortic valve disease (CAVD) in humans and mice. New mouse models have been developed for studies of valve disease mechanisms. Klotho-null mice are a model for premature aging and exhibit calcified nodules in aortic valves with osteogenic gene induction. Osteogenesis Imperfecta mice, bearing a collagen1a2 mutation, develop features of myxomatous valve disease, including thickening, increased proteoglycan deposition and chondrogenic gene induction. Together, these findings demonstrate specific molecular indicators of valve disease progression, including the identification of early disease markers, which represent potential targets for therapeutic intervention.