MiR-32-5p as potential key mediator of vascular remodeling enhances cell proliferation of vascular smooth muscle cells

Abstract Funding Acknowledgements Type of funding sources: Public grant(s) – National budget only. Main funding source(s): University Hospital Halle, Martin-Luther-University Halle-Wittenberg Background and purpose The dysfunctionality of endothelial cells (EC) and smooth muscle cells (SMC) contributes to the progression of vascular remodeling and subsequent cardiovascular pathologies. Several microRNAs (miRNA) have been revealed as key regulators of EC and SMC function, therefore suggesting them as potential targets in the modulation of vascular remodeling. Here, we investigate the potential role of miR-32-5p in vascular cell function and its potential as a therapeutic target in vascular remodeling. Material and Methods Initial screenings by qRT-PCR analysis for miR-32-5p expression were performed in murine femoral artery tissue on day 10 and 21 after wire-induced injury in C57Bl6 mice, serving as restenosis model. MiR-32-5p expression was assessed using qRT-PCR in human vascular endothelial (HVECs) and human vascular smooth muscle cells (HVSMC). The role of miR-32-5p on cellular functions, as migration and proliferation, was determined by performing scratch wound and BrdU assays. Potential miR-32-5p downstream targets were identified from literature and their expression levels upon interference with miR-32-5p evaluated on mRNA and protein levels. Results MiR-32-5p expression was decreased over time in the wire-induced injury model. In vitro, miR-32-5p expression was significantly decreased in replicative senescent HVSMC and HVEC (p0,05) and relatively higher in HVSMC than in HVEC for all conditions. Under growth conditions an elevated expression of miR-32-5p in HVEC and significantly increased expression in HVSMC was detected (p0,01). Proliferation in HVSMC increased significantly (p0,05) by pre-miR-32-5p transfection, but remained unaffected in HVEC. Migration of HVECs and HVSMCs was improved by miR-32-5p and decelerated after antagomiR interference. Systemic literature target research revealed several potential targets in both cell types, of which the following were significantly downregulated in HVSMC after pre-miR transfection including PIK3r3 (p0,05), KLF4 (p0,05), CD69 (p0,01), MAP2K4 (p0,05) and BCL211 (p0,01). Additionally, CD69 and KLF4 expression increased significantly either in HVECs (p0,05). Conclusion Our results suggest that miR-32-5p may be a substantial regulator in HVEC and HVSMC senescence and function and subsequently in vascular remodeling. To fully evaluate its role in the pathogenesis of atherosclerosis and therefore, its therapeutic potential in cardiovascular diseases, further investigations are needed. Additional experiments with the purpose of investigating miR-32-5p-target interactions and apoptotic effects will be performed shortly.