Zitman-Gal Tali, Green Janice, Pasmanik-Chor Metsada, Golan Eliezer, Bernheim Jacques, Benchetrit Sydney
Renal Physiology Laboratory, Department of Nephrology and Hypertension, Meir Medical Center, Kfar Saba 44281, Israel.
Cardiovasc Diabetol. 2014 Jan 7;13:8. doi: 10.1186/1475-2840-13-8.
High blood and tissue concentrations of glucose and advanced glycation end-products are believed to play an important role in the development of vascular complications in patients with diabetes mellitus (DM) and chronic kidney disease. MicroRNAs (miRNA) are non-coding RNAs that regulate gene expression in a sequence specific manner. MiRNA are involved in various biological processes and become novel biomarkers, modulators and therapeutic targets for diseases such as cancer, atherosclerosis, and DM. Calcitriol (the active form of vitamin D) may inhibit endothelial proliferation, blunt angiogenesis, and be a cardioprotective agent. Calcitriol deficiency is a risk factor for DM and hypertension. The aim of this project was to study the miRNA microarray expression changes in human umbilical vein endothelial cells (HUVEC) treated in a diabetic-like environment with the addition of calcitriol.
HUVEC were treated for 24 h with 200 μg/ml human serum albumin (HSA) and 100 mg/dl glucose (control group) or 200 μg/ml AGE-HSA, and 250 mg/dl glucose (diabetic-like environment), and physiological concentrations (10-10 mol/l) of calcitriol. miRNA microarray analysis and real time PCR to validate the miRNA expression profile and mRNA target gene expression were carried out.
Compared to control, 31 mature human miRNA were differentially expressed in the presence of a diabetic-like environment. Addition of physiological concentrations of calcitriol revealed 39 differentially expressed mature human miRNA. MiR-181c, miR-15a, miR-20b, miR-411, miR-659, miR-126 and miR-510 were selected for further analysis because they are known to be modified in DM and in other biological disorders. The predicted targets of these miRNA (such as KLF6, KLF9, KLF10, TXNIP and IL8) correspond to molecular and biological processes such as immune and defense responses, signal transduction and regulation of RNA.
This study identified novel miRNA in the field of diabetic vasculopathy and might provide new information about the effect of vitamin D on gene regulation induced by a diabetic-like environment. New gene targets that are part of the molecular mechanism and the therapeutic treatment in diabetic vasculopathy are highlighted.
高血糖和组织中高浓度的葡萄糖及晚期糖基化终产物被认为在糖尿病（DM）和慢性肾脏病患者血管并发症的发生发展中起重要作用。微小RNA（miRNA）是一类非编码RNA，可通过序列特异性方式调控基因表达。miRNA参与多种生物学过程，成为癌症、动脉粥样硬化和DM等疾病的新型生物标志物、调节剂和治疗靶点。骨化三醇（维生素D的活性形式）可能抑制内皮细胞增殖、抑制血管生成，是一种心脏保护剂。骨化三醇缺乏是DM和高血压的危险因素。本项目旨在研究在添加骨化三醇的糖尿病样环境中处理的人脐静脉内皮细胞（HUVEC）中miRNA芯片表达的变化。
用200μg/ml人血清白蛋白（HSA）和100mg/dl葡萄糖（对照组）或200μg/ml糖基化终产物修饰的人血清白蛋白（AGE-HSA）和250mg/dl葡萄糖（糖尿病样环境）以及生理浓度（10-10mol/l）的骨化三醇处理HUVEC 24小时。进行miRNA芯片分析和实时PCR以验证miRNA表达谱和mRNA靶基因表达。
与对照组相比，在糖尿病样环境下有31种成熟的人miRNA差异表达。添加生理浓度的骨化三醇后，有39种成熟的人miRNA差异表达。选择miR-181c、miR-15a、miR-20b、miR-411、miR-659、miR-126和miR-510进行进一步分析，因为已知它们在DM和其他生物学紊乱中会发生改变。这些miRNA的预测靶标（如KLF6、KLF9、KLF10、TXNIP和IL8）对应于免疫和防御反应、信号转导和RNA调控等分子和生物学过程。
本研究在糖尿病血管病变领域鉴定出新型miRNA，并可能提供有关维生素D对糖尿病样环境诱导的基因调控作用的新信息。突出了作为糖尿病血管病变分子机制和治疗靶点一部分的新基因靶点。
