Abstract 2014: Long Non-coding RNA LINC-PINT Regulates Smooth Muscle Cell Plasticity In Developing Abdominal Aortic Aneurysms

Abstract

Background: Long noncoding RNAs (lncRNAs) play pivotal roles as molecular regulators in diverse vascular biological processes and diseases. This study utilized multi-transcriptomic profiling to investigate the contribution of lncRNAs in Abdominal Aortic Aneurysm (AAA) disease. Materials & Methods: Utilizing bulk and single-cell RNA sequencing, we identified lncRNA long intergenic non-protein coding RNA, p53 induced transcript (LINC-PINT) as a significantly deregulated transcript in AAA compared to non-dilated aortas. Deconvolution of single-cell and bulk RNA-sequencing via the BayesPrism tool identified plasticity of smooth muscle cells (SMCs) as the predominant shifted condition between dilated vs. non-dilated aortas. Hybridization-based RNA in situ sequencing (HybRISS) confirmed the presence of LINC-PINT in de-differentiating SMCs. Results: Experimental knock-down of LINC-PINT , using site-specific antisense oligonucleotides (LNA-GapmeRs) markedly decreased proliferation rates of cultured human aortic smooth muscle cells (AoSMCs), but also increased their apoptosis, whereas overexpression of LINC-PINT had the opposite effect. Transcriptomic profiling of LINC-PINT -silenced AoSMCs indicated down-regulation of contractility markers, accompanied by an increase of inflammation and extracellular matrix (ECM) degradation. In line with this, LINC-PINT knock-down abolished the transforming growth factor beta (TGF-β)- induced AoSMCs polarization towards a contractile phenotype. Notably, RNA immunoprecipitation (RIP) demonstrated an interaction between LINC-PINT and p65, inhibiting its translocation to the promoter region of inflammatory genes. LINC-PINT knock-down increased p65 binding to its coactivator p50, which was confirmed by Co-immunoprecipitation (CO-IP). Combined in silico prediction and luciferase reporter assays further revealed that LINC-PINT expression was controlled by the ZNF93 and ZNF263 transcription factors. Conclusion: We provide evidence of LINC-PINT being a novel regulator influencing SMC dynamics and fate decisions of central importance to AAA pathophysiology. Currently ongoing in vivo studies in Lncpint -deficient mice explore its relevance in experimental AAA development.