Olmesartan alleviates bleomycin-mediated vascular smooth muscle cell senescence via the miR-665/SDC1 axis

Abstract

Olmesartan (OMST) is a new angiotensin II receptor antagonist recently approved by the FDA to treat cardiovascular diseases. We investigated the molecular mechanisms by which OMST regulates vascular senescence. In the present study, bleomycin (BLM) was used to induce senescence in vascular smooth muscle cells (VSMCs); after which, the cells were treated with OMST. The effects of OMST on BLM-mediated cell senescence were evaluated using cell adhesion, NAD ⁺/NADH, and Annevin V/PI double staining assays, as well as by immunofluorescence staining of γH2AX, Edu flow cytometry, and evaluations of senescence-associated β-gal activity. Differentially expressed microRNAs (DEMs) were identified by miRNA microarray assays, and subsequently validated by quantitative real time PCR. Bisulfite sequencing PCR (BSP) was used to detect the methylation status of the miR-665 promoter. The target genes of miR-665 were predicted and confirmed using luciferase reporter assays. We found that miR-665 was upregulated in VSMCs in response to BLM-induced cellular senescence. BSP studies revealed that CpG sites in the promoter region of the miR-665 gene underwent extensive demethylation during BLM-induced cellular senescence, and there was a concomitant up-regulation of miR-665 expression. SDC1 mRNA was identified as a direct target of miR-665. Either miR-665 overexpression or SDC1 knockdown significantly reversed the effects of OMST on BLM-induced VSMC senescence. Moreover, SDC1 overexpression partially reversed the changes that occurred in cells with BLM-induced senescence caused by miR-665 overexpression. Our findings suggest that the miR-665/SDC1 axis functions as a vital modulator of VSMC senescence, and may represent a novel biological target for treating atherosclerosis.