Calcific aortic valve disease is increasingly recognized as an active cellular process that is regulated by a distinct molecular program. Epidemiologic studies have identified numerous clinical risk factors, which result in endothelial cell dysfunction marked by reduced nitric oxide bioavailability, that predispose the valve to calcification. In addition, there is an increasing amount of evidence that valve interstitial cells respond to endothelial cell dysfunction by transforming into osteoblast-like cells, resulting in calcium deposition on aortic valve cusps. Currently, a significant knowledge gap exists in our molecular understanding of how endothelial cells communicate with the underlying valve interstitial cells (VICs) to promote osteogenic-like changes. This knowledge deficit hinders the development of new therapies for calcific aortic valve disease. The basis of this proposal is the discovery by our group that mutations in NOTCH1 cause calcific aortic valve disease in humans. Furthermore we have shown that reduced nitric oxide signaling from the valve endothelium promotes the calcification of valve interstitial cells by a Notch1-mediated mechanism. In addition, the NO and Notch1 signaling pathways display genetic interaction as mice homozygous null for endothelial nitric oxide synthase (eNOS, or Nos3) and heterozygous for Notch1 have aortic valve thickening and abnormal hemodynamics similar to humans, at ~100% penetrance. Our long-term goal is to utilize this novel molecular pathway for the development of novel therapeutics to prevent or slow calcific aortic valve disease. The overall objective of this application is to define the mechanisms by which defects in endothelial NO signaling regulate Notch1 in valve interstitial cells and promote osteogenic changes and calcification. The central hypothesis is: In CAVD, decreased endothelial nitric oxide bioavailability decreases nitrosylation of the Notch1 receptor and Notch1 intracellular domain (N(1)ICD nuclear localization in valve interstitial cells. As a result, Notch activity is reduced, causing calcification. Guided by our publications and preliminary data, the central hypothesis will be tested by pursuing three Specific Aims: 1) Define the temporal development of aortic valve disease in Notch1+/-;Nos3-/- mice, and determine the therapeutic potential of NO donor treatment; 2) Determine if endothelial cell-Jag1 is required to regulate Notch1 activity in VICs and determine its requirement in valve homeostasis in vivo; 3) Define the molecular mechanisms by which endothelial cell-derived nitric oxide (NO) regulates Notch1 signaling in VICs to prevent calcification. Success of this proposal will open multiple new avenues for potential therapies for calcific aortic valve disease. The proposed research is significant because it not only characterizes a novel molecular pathway linking the valve endothelium to the process of calcification of valve interstitial cells but also establishes a new mouse model for longitudinal studies of aortic valve calcification. The knowledge gained will be used to improve prevention measures and develop new therapies for calcific aortic valve disease.
The proposed study is relevant to public health as it addresses the molecular mechanisms that lead to calcific aortic valve disease, a disease process that is becoming increasing prevalent with an aging population. The planned research has the potential to increase the fundamental understanding of disease pathogenesis and discover new molecular pathways that may lead to novel therapeutic targets to prevent or treat the disease.
