Cardiovascular Diseases (CVDs) are the leading cause of death in United States and worldwide. Angiotensin II (Ang II) is a well-known key factor in the development of CVDs. Long noncoding RNAs (lncRNAs), enhancers and enhancer associated lncRNAs have recently been identified as important regulators of gene expression and linked to CVDs. Studies have shown an association between epigenetics and lncRNAs to regulate VSMC functions and lead to CVDs. I propose to study the role of an enhancer associated lncRNA in Ang II mediated VSMC dysfunction. My preliminary data shows that Ang II leads to significant upregulation of a novel lncRNA transcribed from an enhancer enriched locus in rat VSMC which we named Alive (Angiotensin II induced LncRNA In Vascular Cells with Enhancer function). In addition, knockdown of Alive in Ang II treated VSMCs downregulated the expression of genes involved in chondrogenic transformation. VSMC chondrogenesis related arterial stiffening is as a risk factor for atherosclerosis. My specific Aim 1 is Characterize the regulation, function and mechanisms of Alive actions in Ang II mediated VSMC chondrogenesis. Alive and target target gene expression will be analyzed by RT-qPCR and the process of VSMC chondrogenesis by Alcian blue staining after Alive overexpression. ChIP assays will be performed to determine epigenetic marks altered by Alive modulation at its target gene promoters; CRISPR mediated genome editing, 4C-seq; Chromatin isolation by RNA purification will be carried out to determine the functions of enhancer associated with Alive and identify Alive interaction sites on VSMC genome to study the cis- and trans- effects of Alive. Specific Aim 2: Determine the in vivo function of Alive in Ang II infused rats and examine the functional relevance of the human orthologous ALIVE. Sprague-Dawley rats will be infused with Ang II or PBS(control) using osmotic mini pumps for 4 weeks and expression of Alive, chondrogenic genes and hypertension will be determined. To knockdown Alive, these rats will be treated intravenously with Alive targeting GapmeRs. The regulation and function of human orthologous ALIVE in Ang II-induced chondrogenic genes will be examined in by knockdown/overexpression approaches in Human VSMC by RT-qPCR and staining.The completed project provides novel insight into lncRNA and epigenetic mechanisms of Ang II induced VSMC chondrogenesis and could lead to development of unique therapies for CVDs targeting VSMC.
|Program type||Predoctoral Fellowship|
|Effective start/end date||01/01/2019 → 12/31/2020|