Overexpression of miR-22 attenuates oxidative stress injury in diabetic cardiomyopathy via Sirt 1
Overexpression of miR-22 attenuates oxidative stress injury in diabetic cardiomyopathy via Sirt 1
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DOI：
10.1111/1755-5922.12318
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发表时间：
2018-04-01
影响因子：
3.1
通讯作者：
Wang, WeiDong
中科院分区：
文献类型：
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作者：
Tang, Qinghui;Len, Qiang;Wang, WeiDong
背景/目的氧化应激损伤被认为在糖尿病心肌病中很重要。最近的证据表明miR-22在多种心血管疾病中发挥着重要作用，但miR-22在糖尿病心肌病中的保护作用仍不清楚。方法通过腹腔注射链脲佐菌素联合高脂饮食诱导雄性C57BL/6小鼠糖尿病，转染腺相关病毒后miR-22过度表达。通过超声心动图和心导管系统评估心脏功能。在体外研究中，用正常或高葡萄糖（HG）处理H9c2细胞，并分别使用细胞计数试剂盒8（CCK-8）测定和流式细胞术检测细胞活力或凋亡。在糖尿病小鼠和 H9c2 细胞中也检测到活性氧、丙二醛和超氧化物歧化酶。采用实时荧光定量PCR检测miR-22的表达水平。 Western blotting检测Sirt 1、氧化应激损伤相关蛋白（GRP78、CHOP、ATF 3）和凋亡相关蛋白Bax/Bcl-2、cl-casp-9/casp-9和cl-casp-3/casp-3的蛋白表达。结果H9c2细胞中观察到HG诱导的氧化应激损伤和凋亡，miR-22可改善这些损伤和凋亡。在糖尿病小鼠中也观察到心脏功能障碍和严重改变的心脏结构，并且通过 miR-22 的过度表达而显着逆转。 Sirt 1 的表达在糖尿病小鼠和 HG 处理的 H9c2 细胞中显着降低。 miR-22的过表达恢复了Sirt 1的水平。生物信息学分析预测Sirt 1是miR-22的潜在靶基因。荧光素酶报告基因测定证实 miR-22 通过直接结合 Sirt 1 3 个非翻译重复序列来促进 Sirt 1 表达。在 HG 处理的 H9c2 细胞中，Sirt 1 的上调可以提高细胞活力并减轻氧化应激损伤和细胞凋亡，类似于 miR-22 的作用。然而，敲低Sirt 1会消除miR-22对HG诱导的氧化应激损伤和细胞凋亡的保护作用。结论 miR-22的过表达可以通过体内和体外上调Sirt 1来减轻糖尿病心肌病的氧化应激损伤。
Background/AimsOxidative stress injury is believed to be important in diabetic cardiomyopathy. Recent evidence indicates that miR-22 plays an important role in various cardiovascular diseases, but the protective role of miR-22 in diabetic cardiomyopathy remains undetermined.MethodsDiabetes was induced in male C57BL/6 mice by intraperitoneal injection with streptozotocin combined with a high-fat diet, and miR-22 was overexpressed following transfection with adeno-associated virus. Cardiac function was assessed by echocardiography and a cardiac catheter system. In vitro study, H9c2 cells were treated with normal or high glucose (HG), and cell viability or apoptosis was detected using the Cell Counting Kit-8 (CCK-8) assay and flow cytometry, respectively. Reactive oxygen species, malondialdehyde, and superoxide dismutase were also detected in diabetic mice and H9c2 cells. The expression level of miR-22 was detected by real-time PCR. The protein expression of Sirt 1, oxidative stress injury-related proteins (GRP78, CHOP, ATF 3), and apoptosis-related proteins Bax/Bcl-2, cl-casp-9/casp-9, and cl-casp-3/casp-3 were determined by Western blotting analysis.ResultsHG-induced oxidative stress injury and apoptosis were observed in H9c2 cells, which were ameliorated by miR-22. Cardiac dysfunction and severely altered heart structure were also observed in diabetic mice and were dramatically reversed by overexpression of miR-22. The expression of Sirt 1 decreased significantly in diabetic mice and HG-treated H9c2 cells. Overexpression of miR-22 restored the level of Sirt 1. Bioinformatics analysis predicted that Sirt 1 was a potential target gene of miR-22. Luciferase reporter assay verified that miR-22 promoted Sirt 1 expression by direct binding to the Sirt 1 3untranslated repeats. Upregulation of Sirt 1 could improve cell viability and attenuate oxidative stress injury and apoptosis in the HG-treated H9c2 cells, similar to the effect of miR-22. However, the protective effects of miR-22 against HG-induced oxidative stress injury and apoptosis were abrogated by knockdown of Sirt 1.ConclusionsOverexpression of miR-22 can attenuate oxidative stress injury in diabetic cardiomyopathy by upregulation of Sirt 1 in vivo and in vitro.