Inactivation of platelet-derived TGF-β1 attenuates aortic stenosis progression in a robust murine model

Aortic stenosis (AS) is a degenerative heart condition characterized by fibrosis and narrowing of aortic valves (AV), resulting in high wall shear stress (WSS) across valves. AS is associated with high plasma levels of transforming growth factor-b1 (TGF-b1), which can be activated by WSS to induce organ fibrosis, but the cellular source of TGF-b1 is not clear. Here, we show that platelet-derived TGF-b1 plays an important role in AS progres- sion. We first established an aggressive and robust murine model of AS, using the existing Ldlr2/2Apob100/100 (LDLR) breed of mice, and accelerated AS progression by feeding them a high-fat diet (HFD). We then captured very high resolution images of AV movement and thickness and of blood flow velocity across the AV, using a modified ultrasound imaging technique, which revealed early evidence of AS and distinguished different stages of AS progression. More than 90% of LDLR animals developed AS within 6 months of HFD. Scanning electron microscopy and whole-mount immunostaining imaging of AV identified activated platelets physically attached to valvular endothelial cells (VEC) expressing high phosphorylated Smad2 (p-Smad2). To test the contribution of platelet- derived TGF-b1 in AS, we derived LDLR mice lacking platelet TGF-b1 (TGF-b1platelet-KO-LDLR) and showed reduced AS progression and lower p-Smad2 and myofibroblasts in their AV compared with littermate controls fed the HFD for 6 months. Our data suggest that platelet-derived TGF-b1 triggers AS progression by inducing signaling in VEC, and their subsequent transformation into collagen-producing-myofibroblasts. Thus, inhibiting platelet-derived TGF-b1 might attenuate or prevent fibrotic diseases characterized by platelet activation and high WSS, such as AS.