Objective: To explore the differential expressions of microRNA (miRNA) between young and senescent endothelial cells.
Methods: Young and senescent aorta endothelial cells (EC) were isolated and cultured in young and old male C57BL/6J mice. Immunostaining of VIII factor was performed to identify the endothelial cells. The method of diphenyl tetrazolium bromide (MTT) was employed to compare the cell growth. Microarray was used to detect the differential expression of microRNA between young and senescent endothelial cells and the microarray results were confirmed by real-time polymerase chain reaction (PCR). The expression of endothelial nitric oxide synthase (eNOS) was detected by Western blot.
Results: Primarily cultured endothelial cells were confirmed by the VIII immunostaining factor. Senescent ECs grew more rapidly than young ECs in lower serum ex vivo. Excluding gender difference, miR-135a, miR-182, miR-96, miR-31, miR-126-3p and miR-362-5p were up-regulated over 2 folds in young ECs, and miR-335-3p and miR-335-5p up-regulated over 2 folds in senescent ECs by miRNA microarray and RT-PCR. The up-regulation of miR335-3p in old ECs and the up-regulation of miR-135a, miR-96 in the young ECs might contribute to a lower expression of eNOS in senescent ECs.
Conclusion: The expression of miRNAs changes with advancing age and may result in differential expressions of downstream genes.