LncRNA-ANRIL inhibits cell senescence of vascular smooth muscle cells by regulating miR-181a/Sirt1
LncRNA-ANRIL inhibits cell senescence of vascular smooth muscle cells by regulating miR-181a/Sirt1
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DOI：
10.1139/bcb-2018-0126
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发表时间：
2019-10-01
影响因子：
2.9
通讯作者：
Wang, Hai-Qin
中科院分区：
文献类型：
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作者：
Tan, Pan;Guo, Yong-Hong;Wang, Hai-Qin
背景：心血管疾病是威胁人类生命和健康的主要疾病之一，血管老化是其发生的重要原因。INK4位点反义非编码RNA （ANRIL）是一类在细胞衰老过程中起重要作用的长链非编码RNA （lncRNA）。然而，ANRIL在血管平滑肌细胞（VSMCs）衰老中的作用和机制尚不清楚。方法：分别采用MTT法和流式细胞术评价细胞活力和细胞周期。衰老相关(SA)- β -半乳糖苷酶（gal）染色检测细胞衰老。双荧光素酶报告基因检测证实了ANRIL与miR-181a、miR-181a与Sirt1的结合。qRT-PCR检测ANRIL、miR-181a和Sirt1的表达，Western blotting检测sa - β -gal和p53-p21通路相关蛋白的蛋白水平。结果：衰老VSMCs中ANRIL和Sirt1下调，miR-181a上调。在年轻和衰老的VSMCs中，ANRIL过表达可下调miR-181a，上调Sirt1。MTT和sa - β -gal染色分析显示，过表达ANRIL和抑制miR-181a可促进细胞活力，抑制VSMC衰老。双荧光素酶报告试验确定miR-181a直接靶向ANRIL和Sirt1的3'-UTR。此外，ANRIL的过表达抑制了细胞周期阻滞和p53-p21通路。结论：ANRIL可能通过调控miR-181a/Sirt1促进VSMCs细胞活力，抑制衰老，并通过抑制p53-p21通路缓解细胞周期阻滞。这项研究为ANRIL在细胞衰老过程中的作用提供了新的见解。
Background: Cardiovascular disease is one of the major threats to human life and health, and vascular aging is an important cause of its occurrence. Antisense non-coding RNA in the INK4 locus (ANRIL) is a kind of long non-coding RNA (lncRNA) that plays important roles in cell senescence. However, the role and mechanism of ANRIL in senescence of vascular smooth muscle cells (VSMCs) are unclear. Methods: Cell viability and cell cycle were evaluated using an MTT assay and flow cytometry analysis, respectively. Senescence-associated (SA)-beta-galactosidase (gal) staining was used to determine cell senescence. Dual luciferase reporter assays were conducted to confirm the binding of ANRIL and miR-181a, as well as miR-181a and Sirt1. The expression of ANRIL, miR-181a, and Sirt1 was determined using qRT-PCR and protein levels of SA-beta-gal and p53-p21 pathway-related proteins were evaluated by Western blotting. Results: ANRIL and Sirt1 were down-regulated, whereas miR-181a was up-regulated in aging VSMCs. In young and aging VSMCs, over-expression of ANRIL could down-regulate miR-181a and up-regulate Sirt1. MTT and SA-beta-gal staining assays showed that over-expression of ANRIL and inhibition of miR-181a promoted cell viability and inhibited VSMC senescence. The dual-luciferase reporter assay determined that miR-181a directly targets ANRIL and the 3'-UTR of Sirt1. Furthermore, over-expression of ANRIL inhibited cell cycle arrest and the p53-p21 pathway. Conclusion: ANRIL promotes cell viability and inhibits senescence in VSMCs, possibly by regulating miR-181a/Sirt1, and alleviating cell cycle arrest by inhibiting the p53-p21 pathway. This study provides novel insights for the role of ANRIL in the development of cell senescence.