Objective: To explore the effects of olmesartan on age-associated migration and invasion capacities and microRNA (miRAN) axis in human aortic vascular smooth muscle cells (HA-VSMCs).
Methods: Cultured HA-VSMCs were divided into control group, bleomycin-mediated senescence (BLM) group and bleomycin + olmesartan treatment group. Wound-healing assay and Boyden chambers invasion assay were used to assess the changes in migration and invasion of the cells, gelatin zymography was used to analyze matrix metalloproteinase-2 (MMP-2) activation in the cells. The differentially expressed miRNAs were identified by miRNA microarray assay and validated by quantitative real-time PCR. MiR-3133 inhibitor was used to examine the effects of molecular manipulation of olmesartan on age-associated migration and invasion and MMP-2 activation in the cells.
Results: Compared with those of the control group, the percentage of the repopulated cells and the number of cells crossing the basement membrane increased significantly in BLM group [(78.43±12.76)% vs (42.47±7.22)%, P < 0.05; 33.33±5.51 vs 13.00±4.36, P < 0.05]. A significant increase of MMP-2 activation was found in BLM group as compared with the control group (1.66 ± 0.27 vs 0.87 ± 0.13, P < 0.05). Olmesartan significantly inhibited BLM-induced enhancement of cell migration and invasion and MMP-2 secretion in the cells. MiR-3133 was significantly downregulated in BLM group and upregulated in olmesartan group. Transfection with miR-3133 inhibitor significantly reversed the effects of olmesartan on age-associated migration and invasion of the cells [(85.87±7.39)% vs (49.77±3.05)%; 34.67±2.31 vs 20.00±4.58, P < 0.05] and MMP-2 activation in the cells (1.76±0.19 vs 0.94±0.10, P < 0.05).
Conclusions: Olmesartan inhibits the migration and invasion of ageassociated HA-VSMCs probably by upregulating of the miR-3133 axis.
目的: 探讨奥美沙坦酯(OMST)抑制衰老相关人主动脉血管平滑肌细胞(HA-VSMCs)迁移和侵袭能力的作用及微小RNA(miRNA)参与的机制。
方法: HA-VSMCs体外培养，分别经博莱霉素(BLM)诱导衰老、OMST干预及瞬时转染miR-3133 NC/miR-3133 inhibitor。分别设置对照组：HA-VSMCs正常培养；BLM组：HA-VSMCs+BLM；OMST组：HA-VSMCs+BLM+OMST；OMST+miR-3133 NC组：HA-VSMCs转染miR-3133 NC+BLM+OMST；OMST+miR-3133 inhibitor组：HA-VSMCs转染miR-3133 inhibitor+BLM+OMST。划痕实验和Boyden小室侵袭实验分别测定HA-VSMCs迁移和侵袭力。明胶酶谱分析测定HA-VSMCs中金属基质蛋白酶-2 (MMP-2)分泌。MiRNA生物芯片测定差异表达miRNA并行qRT-PCR验证。
结果: 与对照组相比，BLM组划痕实验和侵袭实验显示细胞迁移和侵袭力增加([78.43±12.76) % vs(42.47±7.22) %；33.33±5.51 vs 13.00±4.36，P < 0.05]，MMP-2分泌水平增加[1.66±0.27 vs 0.87±0.13，P < 0.05]；而OMST干预可抑制增加的细胞迁移/侵袭力和MMP-2分泌。生物芯片检测显示，以上3组存在差异miRNAs表达。qRT-PCR证实BLM组miR-3133表达下调，而OMST组miR-3133表达上调。与OMST组相比，OMST+miR-3133 inhibitor组HA-VSMC迁移和侵袭能力增加([85.87±7.39) % vs(49.77±3.05) %；34.67±2.31 vs 20.00±4.58，P < 0.05]；MMP-2分泌水平也增加(1.76±0.19 vs 0.94±0.10，P < 0.05)。
结论: OMST能抑制衰老HAVSMC的迁移和侵袭能力，这一过程可能是通过上调miR-3133表达实现的。
Keywords: aging; invasion; microRNA; migration; olmesartan; vascular smooth muscle cells.