PROJECT SUMMARY/ABSTRACT Thoracic aortic aneurysm and dissection (TAAD) is a lethal disease and there is no medical treatment to prevent or reverse TAAD. Transforming growth factor (TGF-β) plays a critical role in syndromic and non- syndromic TAAD. The detailed mechanism is unknown. The mutations of genes coding TGF-β receptor I and II (TGFBR1 and TGFBR2), and SMAD3 causing TAAD are known or predicted to be Loss-of-function mutations resulting in decrease of TGF-β signaling. However, a paradoxical elevation of TGF-β signaling was found in the end-stage aneurysmal tissue from the patients with those mutations and Marfan Syndrome (MFS). The increasing of TGF-β signaling in the end aneurysmal tissue was found to be independent of TGF- or genetic background, but due to epigenetic control. Losartan, which blocks the elevation of TGF-β signaling and prevents TAAD in mouse models of MFS and Tgfbr1/2 mutations, does not show any more efficacy of reducing aneurysm in MFS patients than β-blockade in a multicenter clinical trial. On the other hand, conditional interruption of TGF-β signaling in the early life of mice results in decreased TGF-β signaling and severe TAAD. It is unknown how the TGF-β signaling changes in smooth muscle cells (SMCs) in the early life of human with gene mutation of TGFBR1/2 and SMAD3. It is also unknown how those mutations alter human SMC phenotype which directly contribute to the integrity of the aorta and related to TAAD. The only model to answer those questions is to use human induce pleuripotent stem (iPS) cells harboring those mutations to model the TAAD of different part of aorta by differentiating iPS cells into different lineage in vivo and in vitro. The long term goals of this project are to: 1) use a human cellular model harboring mutations of TGF-β pathway and animal model harboring tissue engineered mutant vessel made of patients SMCs to dissect cellular and molecular mechanisms of TAAD and 2) develop therapeutic strategies to prevent or reverse the progression of thoracic aortic aneurysms. The objective of this proposal is to determine the effect of mutations of TGFBR1/2 and SMAD3 on the differentiation, contractility and secretory function of vascular SMCs and the underlying mechanisms in vivo and in vitro. The central hypothesis is that the mutations of TGFBR1/2 and SMAD3 cause defective differentiation of SMCs, resulting in an impaired contractile apparatus and secretion of extracellular matrix (ECM) in SMCs, and aneurysm of the mutant vessels. We predict these effects will be reversed by driving differentiation of the SMCs through pathways distinct from TGF-β, such as rapamycin.