Smooth muscle cells hold a central position in vascular homeostasis and during the development of vascular remodelling. As such, they also contribute in pathological processes induced by vascular aging and senescence like i.e. atherosclerosis. Screening for differentially regulated microRNAs, we identified miR-92b-3p as a robustly regulated miRNA in senescent SMC in vitro and in vivo.

Expression analysis of miR92b-3p both in senescent and non-senescent cells using qPCR was performed in primary human coronary artery smooth muscle cells cells (SMCs) and primary cultured human umbilical vein endothelial cells (HUVECS). Further methods included multiple functional assays to assess the effect of induced alterations in miR-92b-3p expression, including wound scratch- and proliferation assays as well as cell death assays by annexin staining. Furthermore, possible targets were identified by structured literature search and target binding simulation. Targets were subsequently confirmed by qPCR and Western Blotting. In vivo, the expression of miR-92b-3p and their respective targets was assessed in different organs in both young and aged (18 months) C57BL6 mice.

We demonstrate that miR-92b-3p expression levels in SMCs were downregulated during replicative senescence. On the other hand, the expression increased during replicative senescence in HUVECS. A similar change of miRNA 92b-3p expression was observed in murine ECs and SMCs.

Reconstitution of miR-92b-3p levels in senescent cells using miR-92b-3p mimics also enhanced and restored the migrational capacity in these cells (p<0.05). In contrast, further degradation of the reduced expression levels of miR-92b-3p in SMCs by the use of specific antimiRs resulted in a further reduction in the proliferative capacity of senescent SMC, determined by WST (p<0,05) and BRDU-assays (<0,05). Similar effects were observed for the migrational capacity of senescent SMC, where overexpression enhanced, but knock down further reduced migration of SMC (p<0.05). Furthermore, overexpression reduced the rate of apoptotic cells, whereas knock down resulted in a further increase in apoptosis (p<0.05). Also, the before identified target genes were confirmed to be altered following over expression or knock down respectively.

In conclusion we identified miR-92b-3p as a microRNA which is differentially expressed during cellular senescence in different vascular cell types. Moreover, miR-92b-3p seems to regulate senescence progression in SMC, since reconstitution of miR-92b-3p levels in SMC partially reduces senescence-induced functional impairments of their functional properties. This might allow future therapeutic approaches to affect vascular aging and senescence.