Mitochondrial network remodeling and the development of the hyper-proliferative and antiapoptotic endothelial phenotype.

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Project Summary Pulmonary vascular disease is responsible for significant morbidity in infants and children with common congenital heart defects that result in increased pulmonary blood flow (PBF) and pressure. There is a lack of effective therapies to limit the shared pathophysiologic features of endothelial dysfunction and vascular remodeling. Our recent studies have demonstrated that metabolic reprogramming and mitochondrial dysfunction, mediated by mechanical stress, is a core regulatory pathway underlying the vascular injury in these children. Further, we have recently identified the presence of a hyper-proliferative, anti-apoptotic endothelial phenotype in our Shunt lamb model of increased PBF and pressure that is involved in an angiogenic response and results in an increase in pulmonary arteriole number. Our data indicate that this endothelial phenotype is associated with increased expression of survivin (an anti-apoptotic protein), mitochondrial fission and increased autophagy/mitophagy. These processes are linked to a loss of NO signaling. The decreased NO signaling in our Shunt lamb model occurs, at least in part, through a decrease in ATP-mediated hsp90 activation. The massive metabolic requirement associated with hyper-proliferation requires a significant consumption of ATP. Based on these data our overall hypothesis is that the ATP consumption required to maintain the hyper- proliferative, anti- apoptotic, endothelial phenotype associated with increased PBF and pressure plays a significant role in the loss of NO signaling by attenuating hsp90 activity. Our data implicate RhoA/ROCK signaling as a master-regulator of these pathways. Studies in our lab have shown that ligation of S1PR3 receptor, induces Rho GTPase signaling and cytoskeletal remodeling. Interestingly, S1PR1 receptor, which exerts a protective effect against mechanical stress, is significantly downregulated in the lungs of our Shunt lamb model, while S1PR3 receptor expression is increased. We hypothesize that a mechanical stress mediated activation of an S1PR3 receptor-RhoA/ROCK axis is responsible for the mitochondrial fission, autophagy/mitophagy, and apoptosis that synergize to produce the hyper-proliferative, anti-apoptotic endothelial phenotype and the loss of NO signaling. Three Specific Aims (SAs) are proposed to test this hypothesis. In Aim 1, we will define the role of mitochondrial fission in the development of a hyper-proliferative endothelial phenotype and determine how this modulates NO signaling. In Aim 2, we will characterize the role played by autophagy/mitophagy in the loss of NO signaling under conditions of increased PBF and pressure and investigate the role played by the anti- apoptotic factor, survivin (Birc5). In Aim 3, we will determine whether targeting mitochondrial fission and autophagy are potential therapeutic targets in our pre-clinical Shunt lamb model. With the completion of this study, we will significantly increase our understanding of the role played by mitochondrial network remodeling and autophagy/mitophagy in the pathogenesis of pulmonary vascular disease associated with increased PBF and pressure while highlighting the application of novel therapeutic interventions.