Zhao Di, Wang Na-Sui, Chen Fu, Li Zheng-Bing, Li Xi-Tao, Zhu Xu-Xin
Department of Orthopedics, the First Affiliated Hospital of Shantou University Medical College, Shantou, China.
Division of Endocrinology and Metabolism, Department of Medicine, the First Affiliated Hospital of Shantou University Medical College, Shantou, China.
Exp Clin Endocrinol Diabetes. 2019 Apr;127(4):255-262. doi: 10.1055/s-0043-125324. Epub 2018 Mar 12.
miR-34a is a multifunctional post-translational modulator, which is involved in several diabetes-related complications. However, miR-34a remains to be fully elucidated in the diabetic endothelium from rats. In this study, the role of miR-34a/NOTCH1 signaling in the progression of hyperglycemia-vascular endothelial dysfunction was investigated.
In intravenous injection of miR-34a mimics and inhibitors in streptozotocin (STZ)-induced diabetic rats, the biomarkers of endothelial dysfunction was measured. The targeted genes were predicted by a bioinformatics algorithm and confirmed by a dual luciferase reporter assay. The mRNA and protein levels were assayed by qRT-PCR and western blotting, respectively. Immunohistochemical staining was performed to measure NOTCH1 expression in the diabetic endothelium.
miR-34a was significantly up-regulated, and NOTCH1 down-regulated, in the thoracic aorta from STZ-induced diabetic rats compared with control group. As compared to model group, the mRNA of NOTCH1 was significantly decreased or increased by miR-34a mimics or inhibitors ex vivo, respectively. Bioinformatics methods further demonstrated that NOTCH1 was a potential target of miR-34a, which was confirmed by dual-luciferase reporter assay. Moreover, both serum ET and NO were significantly increased in diabetic rats as compared to control group. miR-34a inhibitors ex vivo treatment resulted in significant down-regulation ofserum ET and NO levels in diabetic rats as compared to model group.
These results provide evidence to support the use of miR-34a inhibitors as a therapeutic approach attenuating hyperglycemia-induced vascular endothelial dysfunction.
miR-34a是一种多功能的翻译后调节剂，参与多种糖尿病相关并发症。然而，miR-34a在大鼠糖尿病内皮细胞中的作用仍有待充分阐明。本研究探讨了miR-34a/NOTCH1信号通路在高血糖诱导的血管内皮功能障碍进展中的作用。
对链脲佐菌素（STZ）诱导的糖尿病大鼠静脉注射miR-34a模拟物和抑制剂，检测内皮功能障碍的生物标志物。通过生物信息学算法预测靶基因，并通过双荧光素酶报告基因检测进行验证。分别采用qRT-PCR和蛋白质印迹法检测mRNA和蛋白质水平。进行免疫组织化学染色以测量糖尿病内皮细胞中NOTCH1的表达。
与对照组相比，STZ诱导的糖尿病大鼠胸主动脉中miR-34a显著上调，NOTCH1下调。与模型组相比，miR-34a模拟物或抑制剂在体外分别显著降低或增加NOTCH1的mRNA水平。生物信息学方法进一步证明NOTCH1是miR-34a的潜在靶标，双荧光素酶报告基因检测证实了这一点。此外，与对照组相比，糖尿病大鼠血清ET和NO均显著升高。与模型组相比，miR-34a抑制剂体外处理导致糖尿病大鼠血清ET和NO水平显著下调。
这些结果为使用miR-34a抑制剂作为减轻高血糖诱导的血管内皮功能障碍的治疗方法提供了证据。
