Regulation of VSMCs Phenotypic Transformation Mediated by LncRNA SLC12A5-AS1-CDCA3 Axis is Implicated with the Occurrence and Rupture of Intracranial Aneurysm

Abstract

Background: Numerous studies have demonstrated the function of long non-coding RNA (lncRNA) in the regulation of gene expression and its implication in the development of intracranial aneurysm (IA). However, the function and underlying mechanisms through which the lncRNA affects the occurrence and rupture of IA need to be further elucidated. The current study aimed to investigate the lncRNA and mRNA expression profiles in ruptured and unruptured IA patients and control individuals using RNA transcriptome sequencing (RNA-seq) and explore the underlying molecular mechanisms in the onset and rupture of IA mediated by lncRNA-mRNA axis. Methods: RNA-seq was performed to evaluate the lncRNA and mRNA expression profiles in ruptured IA (IA group), unruptured IA (UIA group), and superficial temporal arteries (control group), followed by differential gene expression and functional enrichment analysis. The selected targets were then verified in an expanded cohort of 30 patients per group. Furthermore, the roles of target genes were evaluated in vascular smooth muscle cells (VSMCs) by transfecting them with an overexpression vector for lncRNA SLC12A5-AS1 (SLC12A5-AS1), either alone or in combination with small interfering RNA (si)-cell division cycle associated 3 (CDCA3), si-SLC12A5-AS1, or their corresponding negative controls. The cell proliferation, apoptosis, proteins expression, levels of pro-inflammatory factors, and the angiopoiesis of VSMCs were then determined. Results: RNA-seq showed that SLC12A5-AS1 and CDCA3 gradually increased in a gradient pattern from control donors to UIA to IA patients. Furthermore, receiver operating characteristic (ROC) curve analysis confirmed SLC12A5-AS1 and CDCA3 as valuable biomarkers of UIA and IA patients. Moreover, pathway enrichment analysis of the overlapped differentially expressed (DE) mRNAs in UIA and IA patients revealed that they were mainly enriched in the nuclear factor-kappa B (NF-κB) pathway. Additionally, it was observed that SLC12A5-AS1 overexpression inhibited proliferation, angiopoiesis, and α-smooth muscle actin (α-SMA) and smooth muscle-22α (SM-22α) expression of VSMCs, whereas promoted apoptosis, levels of proinflammatory factors (interleukin-1β (IL-1β), interleukin 6 (IL-6), and tumour necrosis factor-alpha (TNF-α)), matrix metalloproteinase 9 (MMP9) and CDCA3 proteins expression, as well as phosphorylation of p65 and nuclear factor-kappa B inhibitor alpha (IKBα). Conversely, inhibition of CDCA3 had opposite effects on VSMCs, which can be overcome with SLC12A5-AS1 overexpression. Conclusion: Our current findings indicated that the SLC12A5-AS1 is involved in IA occurrence and rupture by regulating the phenotypic transformation of VSMCs through modulating CDCA3.