Uncovering Dysregulated Micro‐RNA Expression in Senescent Human Vascular Smooth Muscle Cells: Impacts on Cellular Communication and Vascular Aging

Abstract

Vascular aging is highly associated with cardiovascular morbidity and mortality and vascular smooth muscle cell (VSMC) senescence is one of its key contributors. However, intracellular and extracellular signaling and communication of senescent VSMCs are not fully elucidated. Understanding micro‐RNA (miRNA) regulation during VSMC senescence may elucidate the molecular mechanistic link between vascular aging and early adverse vascular remodeling. This study aimed to identify dysregulated miRNA expression within senescent VSMCs as well as senescent VSMC‐derived exosomes. Healthy and senescent human VSMCs (hVSMCs) were cultured in vitro and exosomes secreted by these cells were isolated via ultracentrifugation. Subsequently, cellular and exosomal miRNAs were extracted and used for library preparation of whole‐genome small RNA next‐generation sequencing (NGS). Post‐sequencing and differential expression analyses compared the miRNA profiles between healthy and senescent hVSMCs as well as their derived exosomes. Thereafter, NGS data were validated using real‐time polymerase chain reaction (qPCR). Additionally, target prediction and pathway analysis were performed. In senescent hVSMCs, eight significant differentially expressed mature miRNAs were identified (n= 4, q <0.05). Bioinformatic analyses showed correlation between upregulated miRNAs and activated ubiquitin proteasome pathway, a senescence hallmark. Meanwhile, downregulated miRNAs might associate with cell cycle arrest at G1/S phase via the TGF‐β/SMAD signaling pathway, promoting hVSMC entry into senescence. On the other hand, three significant differentially expressed miRNAs were found in senescent hVSMC‐derived exosomes. Data suggested that the upregulated exosomal miRNA was involved in the formation of advanced glycation end‐products, known to play a role in vascular complications. Concurrently, downregulation of exosomal miRNAs promoted cellular senescence, thus advanced aging. Interestingly, there was a consistent observation of hsa‐miR‐155‐5p downregulation in both senescent hVSMCs and their secreted exosomes. Further analyses with the other dysregulated miRNAs suggested hsa‐miR‐155‐5p involvement in the TGF‐β/NF‐κB signaling pathway. The dysregulated miRNAs in senescent hVSMCs and senescent hVSMC‐derived exosomes offered valuable insights on the mechanisms involved in hVSMC intracellular and extracellular signaling during senescence, thus contributing to our understanding of vascular aging in humans.Support or Funding InformationThe study was funded by Ministry of Higher Education Malaysia grant, FRGS/1/2015/SKK08/UNIM/03/2 to YFP.