Abstract
Coronary in-stent restenosis is a late complication of angioplasty. It is a multifactorial process that involves vascular smooth muscle cells (VSMCs), endothelial cells, and inflammatory and genetic factors. In this study, the transcriptomic landscape of VSMCs’ phenotypic switch process was assessed under stimuli resembling stent injury. Co-cultured contractile VSMCs and endothelial cells were exposed to a bare metal stent and platelet-derived growth factor (PDGF-BB) 20 ng/mL. Migratory capacity (wound healing assay), proliferative capacity, and cell cycle analysis of the VSMCs were performed. RNAseq analysis of contractile vs. proliferative VSMCs was performed. Gene differential expression (DE), identification of new long non-coding RNA candidates (lncRNAs), gene ontology (GO), and pathway enrichment (KEGG) were analyzed. A competing endogenous RNA network was constructed, and significant lncRNA–miRNA–mRNA axes were selected. VSMCs exposed to “stent injury” conditions showed morphologic changes, with proliferative and migratory capacities progressing from G0-G1 cell cycle phase to S and G2-M. RNAseq analysis showed DE of 1099, 509 and 64 differentially expressed mRNAs, lncRNAs, and miRNAs, respectively. GO analysis of DE genes showed significant enrichment in collagen and extracellular matrix organization, regulation of smooth muscle cell proliferation, and collagen biosynthetic process. The main upregulated nodes in the lncRNA-mediated ceRNA network were PVT1 and HIF1-AS2, with downregulation of ACTA2-AS1 and MIR663AHG. The PVT1 ceRNA axis appears to be an attractive target for in-stent restenosis diagnosis and treatment.
Highlights
Coronary in-stent restenosis (SR) continues to challenge the scientific community, despite the emergence of new biomedical materials [1] and its association with medications (DES) [2]
According to the Competing Endogenous RNA (ceRNA) theory, a pair of co-expressed long non-coding RNA candidates (lncRNAs) and mRNA compete for miRNA target sites; lncRNAs serve as endogenous sponges that target one or more miRNAs, affecting the post-transcriptional regulation of mRNAs
Our in vitro model of stent restenosis has been previously validated; there are several limitations: (1) while indirect contact of Human Umbilical Vein Endothelial Cells (HUVEC) and Human umbilical artery smooth muscle cells (HUASMC) allows molecule and microvesicle traffic, it impedes cell–cell contact [69]; (2) rheological factors such as shear stress and flow play an important role in vascular cell biology and regulation [29]; (3) alternative strategies such as in vivo animal models [70] and in vitro blood vessel constructs [71] closely resemble real live behavior during stenting; to the best of our knowledge there are no precedents for next-generation RNA sequencing of vascular smooth muscle cells (VSMCs) co-cultured with HUVECs in a stent-induced injury model combining bare metal stent and PDGF-BB
Summary
Coronary in-stent restenosis (SR) continues to challenge the scientific community, despite the emergence of new biomedical materials [1] and its association with medications (drug eluting stents) (DES) [2]. In vascular endothelial cells (ECs) treated with oxLDL, lnc-MKI67IP-3 acted as a sponge for let-7e, suppressing its pro-inflammatory effects and thereby upregulating IκBβ expression, forming a positive feedback loop to aggravate inflammation This ceRNA axis might play important roles in the inflammatory responses of ECs and development of atherosclerosis [21]. The growing importance of lncRNAs in the pathogenesis of cardiovascular diseases along with their tissue-specific expression motivated us to evaluate the expression profile of these transcripts during the phenotypic switch of VSMCs induced by bare metal stent and PDGF-BB exposition This will contribute to understanding the mechanisms that rule the change in smooth muscle cell phenotype and represents the first step in a pipeline of genetic biomarker discovery to allow for early diagnosis of post-angioplasty coronary stent restenosis.
Sign-in/Register to view highlights & summary Sign-in/Register to access full text options 
Free) 
Talk to us
Join us for a 30 min session where you can share your feedback and ask us any queries you have
Schedule a call
