MicroRNA-24 Attenuates Neointimal Hyperplasia in the Diabetic Rat Carotid Artery Injury Model by Inhibiting Wnt4 Signaling Pathway.
MicroRNA-24 Attenuates Neointimal Hyperplasia in the Diabetic Rat Carotid Artery Injury Model by Inhibiting Wnt4 Signaling Pathway.
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MicroRNA-24 通过抑制 Wnt4 信号通路减轻糖尿病大鼠颈动脉损伤模型中的新内膜增生
DOI：
10.3390/ijms17060765
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发表时间：
2016-05-24
影响因子：
5.6
通讯作者：
Chen L
中科院分区：
文献类型：
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作者：
Yang J;Fan Z;Yang J;Ding J;Yang C;Chen L
长期的高血糖刺激大大增加了血管成形术后血管再狭窄(RS)的发生率。血管损伤后新生内膜增生是RS的病理原因，但其机制尚不清楚。MicroRNA-24(miR-24)在糖尿病大鼠颈动脉损伤处低表达。然而，miR-24在血管系统中的作用尚不清楚。在这项研究中，我们探讨了miR-24的过表达是否可以抑制链脲佐菌素(STZ)诱导的糖尿病大鼠的新生内膜形成。用腺病毒(Ad-miR-24-GFP)将miR-24基因转移到糖尿病大鼠的颈动脉损伤处。苏木精-伊红(HE)染色检测新生内膜增生程度。用增殖细胞核抗原(增殖细胞核抗原)免疫组织化学方法检测新生内膜中血管平滑肌细胞(VSMC)的增殖情况。用实时定量聚合酶链式反应检测miR-24、增殖细胞核抗原、无翅型MMTV整合位点家族成员4(WNT4)、脱发-1(DVL-1)、β-连环蛋白和细胞周期相关分子(Cyclin D1、p21)的基因表达水平。免疫印迹法检测增殖细胞核抗原、WNT4、DVL-1、β-连环蛋白、细胞周期蛋白D1和p21蛋白的表达。STZ能降低SD大鼠血浆胰岛素水平，升高空腹血糖。糖尿病大鼠颈动脉球囊损伤后miR-24表达降低，腺病毒载体(Ad-miR-24-GFP)上调miR-24表达。糖尿病大鼠14d时，miR-24的过表达抑制了VSMC的增殖和新生内膜的增殖。此外，与假手术组相比，糖尿病大鼠颈动脉球囊损伤后增殖细胞核抗原、WNT4、DVL-1、β-连环蛋白和细胞周期蛋白D1mRNA和蛋白水平均显著上调。有趣的是，miR-24的上调显著降低了上述分子的mRNA和蛋白质水平。相反，球囊损伤后p21mRNA和蛋白水平的变化趋势与之相反。然而，基因传递后miR-24的过度表达增加了p21的mRNA和蛋白水平。结论：miR-24过表达可抑制糖尿病大鼠血管损伤后VSMC的增殖和新生内膜的增生。这一结果可能与WNT4/DVL-1/β-连环蛋白信号通路调控细胞周期蛋白D1和p21的表达有关。
The long-term stimulation of hyperglycemia greatly increases the incidence of vascular restenosis (RS) after angioplasty. Neointimal hyperplasia after vascular injury is the pathological cause of RS, but its mechanism has not been elucidated. MicroRNA-24 (miR-24) has low expression in the injured carotid arteries of diabetic rats. However, the role of miR-24 in the vascular system is unknown. In this study, we explore whether over-expression of miR-24 could attenuate neointimal formation in streptozotocin (STZ)-induced diabetic rats. Adenovirus (Ad-miR-24-GFP) was used to deliver the miR-24 gene to injured carotid arteries in diabetic rats. The level of neointimal hyperplasia was examined by hematoxylin-eosin (HE) staining. Vascular smooth muscle cell (VSMC) proliferation in the neointima was evaluated by immunostaining for proliferating cell nuclear antigen (PCNA). The mRNA levels of miR-24, PCNA, wingless-type MMTV integration site family member 4 (Wnt4), disheveled-1 (Dvl-1), β-catenin and cell cycle-associated molecules (Cyclin D1, p21) were determined by Quantitative Real-Time PCR (qRT-PCR). PCNA, Wnt4, Dvl-1, β-catenin, Cyclin D1 and p21 protein levels were measured by Western blotting analysis. STZ administration decreased plasma insulin and increased fasting blood glucose in Sprague-Dawley (SD) rats. The expression of miR-24 was decreased in the carotid artery after a balloon injury in diabetic rats, and adenoviral transfection (Ad-miR-24-GFP) increased the expression of miR-24. Over-expression of miR-24 suppressed VSMC proliferation and neointimal hyperplasia in diabetic rats at 14 days. Furthermore, compared with Sham group, the mRNA and protein levels of PCNA, Wnt4, Dvl-1, β-catenin, and Cyclin D1 were strikingly up-regulated in the carotid arteries of diabetic rats after a balloon injury. Interestingly, up-regulation of miR-24 significantly reduced the mRNA and protein levels of these above molecules. In contrast, the change trend in p21 mRNA and protein levels was opposite after a balloon injury. However, over-expression of miR-24 after gene delivery increased the mRNA and protein levels of p21. We conclude that over-expression of miR-24 could attenuate VSMC proliferation and neointimal hyperplasia after vascular injuries in diabetic rats. This result is possibly related to the regulation of the expression of Cyclin D1 and p21 through the Wnt4/Dvl-1/β-catenin signaling pathway.