Insulin ameliorates miR-1-induced injury in H9c2 cells under oxidative stress via Akt activation.
Growing evidence indicates that aberrant upregulation of microRNA-1 (miR-1) occurs in ischemic myocardium. In addition, insulin elicits metabolism-independent cardioprotection against cardiovascular diseases. The aim of this study is to determine whether insulin ameliorates miR-1-induced injury in H9c2 cells under oxidative stress and to investigate the underlying mechanisms. By quantitative real-time RT-PCR (qRT-PCR), we show that miR-1 is upregulated in H9c2 cells after treatment with hydrogen peroxide (H(2)O(2)), and this effect is both dose- and time dependent. Furthermore, expression of miR-1 decreased significantly after insulin treatment (4.5 ± 0.1 vs. 3.0 ± 0.2, p < 0.05). To determine the potential role of miR-1 in cellular injury and gene regulation, adenovirus-mediated overexpression of miR-1 was used. Overexpression of miR-1 decreased cell viability by 28 ± 2 % (n = 6, p < 0.05) and damaged Akt activation with or without H(2)O(2) treatment. To further investigate the effect of the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in miR-1-induced injury, H9c2 cells were pretreated with LY294002 (10 μM LY, a specific inhibitor of PI3K) with or without insulin (100 nM) and subjected to H(2)O(2) treatment. LY pretreatment-inhibited Akt activation, lead to increased reactive oxygen species (ROS), and further decreased cell viability induced by miR-1 (n = 6, p < 0.05, n = 9-10 cells/group, p < 0.05 and n = 6, p < 0.05) under oxidative stress. This effect was abolished by insulin. In summary, our findings suggest that miR-1 expression is sensitive to H(2)O(2) stimulation. In addition, insulin decreases miR-1 expression and induces a marked protective effect on miR-1-induced injury under oxidative stress, which may be mediated by the Akt-mediated pathway. These results provide an important, novel clue as to the mechanism of the cardiovascular action of insulin.
Department of Cardiovascular Medicine, First Affiliated Hospital of Medical School, Xi'an Jiaotong University, Xi'an, Shaanxi, China.
Tao
T
Chen
Guoliang
G
Ding
Zhuqing
Z
Jin
Mary B
MB
Wagner
Zuyi
Z
Yuan
D000818:Animals; D002470:Cell Survival; D006861:Hydrogen Peroxide / Q000633:toxicity; D066298:In Vitro Techniques; D007328:Insulin / Q000378:metabolism / Q000494:pharmacology*; D035683:MicroRNAs / Q000235:genetics / Q000378:metabolism; D015428:Myocardial Reperfusion Injury / Q000139:chemically induced / Q000378:metabolism*; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000378:metabolism; D051058:Oncogene Protein v-akt / Q000235:genetics / Q000378:metabolism; D018384:Oxidative Stress; D058539:Phosphatidylinositol 3-Kinase / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D015398:Signal Transduction
D007328:Insulin; C542191:MIRN1 microRNA, rat; D035683:MicroRNAs; D017382:Reactive Oxygen Species; D006861:Hydrogen Peroxide; D058539:Phosphatidylinositol 3-Kinase; D051058:Oncogene Protein v-akt