Objective Aortic valve disease, including calcification affects more than 2% of the human population and is caused by complex interactions between multiple risk factors including genetic mutations, the environment and biomechanics. At present, there are no effective treatments other than surgery and this is due to the limited understanding of the mechanisms that underlie the condition. Previous work has shown that valve interstitial cells (VICs) within the aortic valve cusps differentiate towards an osteoblast-like cell and deposit bone-like matrix that leads to leaflet stiffening and calcific aortic valve stenosis. However the mechanisms that promote pathological phenotypes in VICs are unknown. Approach and Results Using a combination of in vitro and in vivo tools with mouse, porcine and human tissue, we show that in VICs, reduced Sox9 expression and nuclear localization precedes the onset of calcification. In vitro, Sox9 nuclear export and calcific nodule formation is prevented by valve endothelial cells (VECs). While in vivo, loss of Tgfβ1 in the endothelium leads to reduced Sox9 expression and calcific aortic valve disease. Conclusions Together, these findings suggest that reduced nuclear localization of Sox9 in VICs is an early indicator of calcification and therefore pharmacological targeting to prevent nuclear export could serve as a novel therapeutic tool in the prevention of calcification and stenosis.

Decellularized allograft heart valves have been used as tissue-engineered heart valve (TEHV) scaffolds with promising results; however, little is known about the cellular mechanisms underlying TEHV neotissue formation. To better understand this phenomenon, we developed a murine model of decellularized pulmonary heart valve transplantation using a hemodynamically unloaded heart transplant model. Furthermore, because the hemodynamics of blood flow through a heart valve may influence morphology and subsequent function, we describe a modified loaded heterotopic heart transplant model that led to an increase in blood flow through the pulmonary valve. We report host cell infiltration and endothelialization of implanted decellularized pulmonary valves (dPV) and provide an experimental approach for the study of TEHVs using mouse models.

Heart valves are complex structures composed of a heterogeneous population of valve interstitial cells (VICs), an overlying endothelium and highly organized layers of extracellular matrix. Alterations in valve homeostasis are characteristic of dysfunction and disease, however the mechanisms that initiate and promote valve pathology are poorly understood. Advancements have been largely hindered by the limited availability of tools for gene targeting in heart valve structures during embryogenesis and after birth. We have previously shown that the transcription factors Sox9 and Scleraxis (Scx) are required for heart valve formation and in this study we describe the recombination patterns of Sox9-and Scx-Cre lines at differential time points in aortic and mitral valve structures. In ScxCre; ROSA26GFP mice, recombination is undetected in valve endothelial cells (VECs) and low in VICs during embryogenesis. However, recombination increases in VICs from post natal stages and by 4 weeks side-specific patterns are observed. Using the inducible Sox9CreERT2 system, we observe recombination in VECs and VICs in the embryo, and high levels are maintained through post natal and juvenile stages. These Cre-drivers provide the field with new tools for gene targeting in valve cell lineages during differential stages of embryonic and post natal maturation and maintenance.