Cardiovascular disease is the leading cause of morbidity and mortality in the Western and developing world, and the incidence of cardiovascular disease is increasing with the longer lifespan afforded by our modern lifestyle. Vascular diseases including coronary heart disease, high blood pressure, and stroke comprise the majority of cardiovascular diseases, and therefore represent a significant medical and socioeconomic burden on our society. It may not be surprising that these conditions overlap and potentiate each other when we consider the many cellular and molecular similarities between them. These intersecting points are manifested in clinical studies in which lipid lowering therapies reduce blood pressure, and anti-hypertensive medications reduce atherosclerotic plaque. At the molecular level, the vascular smooth muscle cell (VSMC) is the target, integrator, and effector cell of both atherogenic and the major effector protein of the hypertensive signal Angiotensin II (Ang II). Together, these signals can potentiate each other and prime the artery and exacerbate hypertension and atherosclerosis. Therefore, VSMCs are the fulcrum in progression of these diseases and, therefore, understanding the effects of atherogenic stimuli and Ang II on the VSMC is key to understanding and treating atherosclerosis and hypertension. In this review, we will examine studies in which hypertension and atherosclerosis intersect on the VSMC, and illustrate common pathways between these two diseases and vascular aging.

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Introduction: Hypertension is a major risk factor for cardiovascular disease and stroke. Optimally treated hypertensive patients have a 50% greater cardiovascular risk than untreated normotensive subjects, presenting a need for additional targets. Vascular smooth muscle cells (VSMC) play a critical role in vascular contractility and regulation of blood pressure. Fragile X-related protein (FXR1) is a muscle-enhanced RNA binding protein. In addition to containing mRNA binding domains, FXR1 has an agenet-like domain for protein-protein interactions and four WH2 motif domains to mediate actin dynamics. The structure of FXR1 supports its role in post-transcriptional regulation and though largely ignored, cytoskeletal dynamics via potential protein-protein interactions. This study will test the hypothesis that FXR1 regulates vascular contractility by RNA stability and protein interactions. Results: RNA immuno-precipitation sequencing (RIPseq) analysis in human VSMC identified that FXR1 binds to mRNA that participate in VSMC contractility and cytoskeletal reorganization, and FXR1 depletion decreases mRNA abundance and stability of these transcripts. Mass-spectrometry and co-immunoprecipitation identified that FXR1 interacts with members of the WAVE complex, a five-subunit protein complex involved in the formation of the actin cytoskeleton, including CYFIP1 (Cytoplasmic FMR1-interacting protein 1) and Actin Related Protein 2/3 complex (ARP2). The WAVE complex is activated by an ARP2-mediated interaction with Rac1, promoting actin remodeling and cytoskeletal reorganization in a GTP-dependent manner. FXR1 depletion decreases small GTPases RAC1 and CDC42 activation in VSMC. Additionally, FXR1 depletion decreases VSMC lamellipodia formation, adhesion, migration and collagen gel contraction. Novel, VSMC-specific FXR1 conditional knock out mice show decreased diastolic (P < 0.05) blood pressure at baseline compared to controls. Conclusion: These data are the first to suggest that FXR1 regulates mRNA stability of contractile proteins and interacts with members of the WAVE complex. Deletion of FXR1 abrogates VSMC cytoskeletal reorganization, resulting in a hypotensive phenotype in VSMC-specific conditional knockout mice.

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Introduction: Despite the advent of stents, intimal hyperplasia subsequent to vascular interventional procedures remains a major obstacle. Vascular smooth muscle cells (VSMC) play a critical role in the pathogenesis of intimal hyperplasia; therefore regulation of VSMC gene expression is a logical intervention point. FXR1 is a muscle-enhanced RNA binding protein and expression is increased in injured arteries. We have shown that modulation of FXR1 levels affects stability and abundance of inflammatory transcripts in VSMC, suggesting that FXR1 is a negative regulator of inflammation. This drives our hypothesis that FXR1 is involved in mitigating vascular disease and regulating inflammatory and proliferative mRNA in VSMC. Approach/Results: We developed a novel VSMC-specific conditional knockout mouse (FXR1 VSMC/VSMC ). In a carotid artery ligation model of intimal hyperplasia FXR1 VSMC/VSMC mice, have significantly reduced neointima formation (p<0.001) post-ligation compared to controls. To determine the mechanism of these effects, we knocked down FXR1 in human VSMC and observed decreased proliferation (p<0.05) as well as increased beta galactosidase (p<0.05) and gamma H2AX (p<0.01) staining, indicative of senescence. Senescent cells exhibit a senescence associated secretory phenotype (SASP) with characteristic gene expression leading to increased inflammation in the tissue microenvironment. RIP-sequencing demonstrated that FXR1 interacts with transcripts involved in cell cycle control, and stability of these transcripts is decreased with FXR1 KO. qPCR analysis from FXR1 KO mouse VSMC show increased transcripts associated with senescence (p21, p16, p53) as well as increased SASP-associated mRNA. Furthermore, wild-type human VSMC cultured in conditioned media from cells transfected with FXR1 siRNA show increased SASP mRNA and increased proliferation compared with cells cultured in conditioned media from scrambled control cells. Summary & Conclusions: Our results are the first to suggest that in addition to destabilization of inflammatory transcripts, FXR1 may stabilize cell cycle related genes in VSMC, and absence of FXR1 leads to induction of a senescent phenotype, an increase in SASP genes, and reduction of intimal hyperplasia.