Background: Marfan syndrome, caused by mutations in the gene for fibrillin-1, leads to thoracic aortic aneurysms (TAAs). Phenotypic modulation of vascular smooth muscle cells (SMCs) and ECM (extracellular matrix) remodeling are characteristic of both nonsyndromic and Marfan aneurysms. The ECM protein FN (fibronectin) is elevated in the tunica media of TAAs and amplifies inflammatory signaling in endothelial and SMCs through its main receptor, integrin α5β1. We investigated the role of integrin α5-specific signals in Marfan mice in which the cytoplasmic domain of integrin α5 was replaced with that of integrin α2 (denoted α5/2 chimera). Methods: We crossed α5/2 chimeric mice with Fbn1 ^mgR/mgR mice (mgR model of Marfan syndrome) to evaluate the survival rate and pathogenesis of TAAs among wild-type, α5/2, mgR, and α5/2 mgR mice. Further biochemical and microscopic analysis of porcine and mouse aortic SMCs investigated molecular mechanisms by which FN affects SMCs and subsequent development of TAAs. Results: FN was elevated in the thoracic aortas from Marfan patients, in nonsyndromic aneurysms, and in mgR mice. The α5/2 mutation greatly prolonged survival of Marfan mice, with improved elastic fiber integrity, mechanical properties, SMC density, and SMC contractile gene expression. Furthermore, plating of wild-type SMCs on FN decreased contractile gene expression and activated inflammatory pathways whereas α5/2 SMCs were resistant. These effects correlated with increased NF-kB activation in cultured SMCs and mgR aortas, which was alleviated by the α5/2 mutation or NF-kB inhibition. Conclusions: FN-integrin α5 signaling is a significant driver of TAA in the mgR mouse model. This pathway thus warrants further investigation as a therapeutic target.

Aortic dissection and rupture are triggered by decreased vascular wall strength and/or increased mechanical loads. We investigated the role of mTOR signaling in aortopathy using a well-described model of angiotensin II–induced dissection, aneurysm, or rupture of the suprarenal abdominal aorta in Apoe -deficient mice. Although not widely appreciated, nonlethal hemorrhagic lesions present as pseudoaneurysms without significant dissection in this model. Angiotensin II–induced aortic tears result in free rupture, contained rupture with subadventitial hematoma (forming pseudoaneurysms), dilatation, or healing, while the media invariably thickens regardless of mural tears. Medial thickening results from smooth muscle cell hypertrophy and extracellular matrix accumulation, including matricellular proteins. Angiotensin II activates mTOR signaling in vascular wall cells, and inhibition of mTOR signaling by rapamycin prevents aortic rupture but promotes dissection. Decreased aortic rupture correlates with decreased inflammation and metalloproteinase expression, whereas extensive dissection correlates with induction of matricellular proteins that modulate adhesion of vascular cells. Thus, mTOR activation in vascular wall cells determines whether aortic tears progress to dissection or rupture. Previous mechanistic studies of aortic aneurysm and dissection by angiotensin II in Apoe -deficient mice should be reinterpreted as clinically relevant to pseudoaneurysms, and mTOR inhibition for aortic disease should be explored with caution.

The zinc finger protein ZFYVE21 is involved in immune signaling. Using humanized mouse models, primary human cells, and patient samples, we identified a T cell–autonomous role for ZFYVE21 in promoting chronic vascular inflammation associated with allograft vasculopathy. Ischemia-reperfusion injury (IRI) stimulated endothelial cells to produce Hedgehog (Hh) ligands, which in turn induced the production of ZFYVE21 in a population of T memory cells with high amounts of the Hh receptor PTCH1 (PTCH ^hi cells, CD3 ⁺ CD4 ⁺ CD45RO ⁺ PTCH1 ^hi PD-1 ^hi ), vigorous recruitment to injured endothelia, and increased effector responses in vivo. After priming by interferon-γ (IFN-γ), Hh-induced ZFYVE21 activated NLRP3 inflammasome activity in T cells, which potentiated IFN-γ responses. Hh-induced NLRP3 inflammasomes and T cell–specific ZFYVE21 augmented the vascular sequelae of chronic inflammation in mice engrafted with human endothelial cells or coronary arteries that had been subjected to IRI before engraftment. Moreover, the population of PTCH ^hi T cells producing high amounts of ZFYVE21 was expanded in patients with renal transplant–associated IRI, and sera from these patients expanded this population in control T cells in a manner that depended on Hh signaling. We conclude that Hh-induced ZFYVE21 activates NLRP3 inflammasomes in T cells, thereby promoting chronic inflammation.

BACKGROUND: Smooth muscle cells (SMCs) of cardiac and neural crest origin contribute to the developing proximal aorta and are linked to disease propensity in adults. METHODS: We analyzed single-cell transcriptomes of aortic SMCs from adult mice to determine basal states and changes after disrupting TGFβ (transforming growth factor-β) signaling necessary for aortic homeostasis. RESULTS: A minority of Myh11 lineage-marked SMCs differentially expressed genes suggestive of embryological origin. Additional analyses in Nkx2-5 and Wnt1 lineage-marked SMCs derived from cardiac and neural crest progenitors, respectively, showed both lineages contributed to a major common cluster and each lineage to a minor distinct cluster. Common cluster SMCs extended from root to arch, cardiac subset cluster SMCs from root to ascending, and neural crest subset cluster SMCs were restricted to the arch. The neural crest subset cluster had greater expression of a subgroup of TGFβ-dependent genes. Nonetheless, conditional deletion of TGFβ receptors resulted in similar transcriptional changes among all SMC clusters. Several disease-associated transcriptional responses were comparable among SMC clusters in a mouse model of Marfan syndrome aortopathy, while many embryological markers of murine aortic SMCs were not detected in adult human aortas. CONCLUSIONS: There are multiple subtypes of cardiac-derived and neural crest–derived SMCs with shared or distinctive transcriptional profiles; neural crest subset cluster SMCs with increased expression of certain TGFβ-inducible genes are not spatially linked to the aortic root predisposed to aneurysms from aberrant TGFβ signaling; and loss of TGFβ responses after receptor deletion is uniform among SMC clusters.

Hypertension and transient increases in blood pressure from extreme exertion are risk factors for aortic dissection in patients with age-related vascular degeneration or inherited connective tissue disorders. Yet, a common experimental model of angiotensin II–induced aortopathy in mice appears independent of high blood pressure, as lesions do not occur in response to an alternative vasoconstrictor, norepinephrine, and are not prevented by cotreatment with a vasodilator, hydralazine. We investigated vasoconstrictor administration to adult mice following 1 week of disrupted TGF-β signaling in smooth muscle cells (SMCs). Norepinephrine increased blood pressure and induced aortic dissection by 7 days and even within 30 minutes (as did angiotensin II) that was prevented by hydralazine. Initial medial injury manifested as blood extravasation among SMCs and fibrillar matrix, progressive delamination from accumulation of blood, and stretched or ruptured SMCs with persistent attachments to elastic fibers. Altered regulatory contractile molecule expression was not of pathological importance. Rather, reduced synthesis of extracellular matrix yielded a vulnerable aortic phenotype by decreasing medial collagen, most dynamically basement membrane–associated multiplexin collagen, and impairing cell-matrix adhesion. We conclude that transient and sustained increases in blood pressure can cause dissection in aortas rendered vulnerable by inhibition of TGF-β–driven extracellular matrix production by SMCs.