Department of Pathology, Schulich School of Medicine and Dentistry and the University of Western Ontario, London, Ontario, Canada.
Diabetes. 2011 Nov;60(11):2975-84. doi: 10.2337/db11-0478. Epub 2011 Sep 1.
MicroRNAs (miRNAs), through transcriptional regulation, modulate several cellular processes. In diabetes, increased extracellular matrix protein fibronectin (FN) production is known to occur through histone acetylator p300. Here, we investigated the role of miR-146a, an FN-targeting miRNA, on FN production in diabetes and its relationship with p300.
miR-146a expressions were measured in endothelial cells from large vessels and retinal microvessels in various glucose levels. FN messenger RNA expression and protein levels with or without miR-146a mimic or antagomir transfection were examined. A luciferase assay was performed to detect miR-146a's binding to FN 3'-untranslated region (UTR). Likewise, retinas from type 1 diabetic rats were studied with or without an intravitreal injection of miR-146a mimic. In situ hybridization was used to localize retinal miR-146a. Cardiac and renal tissues were analyzed from type 1 and type 2 diabetic animals.
A total of 25 mmol/L glucose decreased miR-146a expression and increased FN expression compared with 5 mmol/L glucose in both cell types. miR-146a mimic transfection prevented such change, whereas miR-146a antagomir transfection in the cells in 5 mmol/L glucose caused FN upregulation. A luciferase assay confirmed miR-146a's binding to FN 3'-UTR. miR-146a was localized in the retinal endothelial cells and was decreased in diabetes. Intravitreal miR-146a mimic injection restored retinal miR-146a and decreased FN in diabetes. Additional experiments showed that p300 regulates miR-146a. Similar changes were seen in the retinas, kidneys, and hearts in type 1 and type 2 diabetic animals.
These studies showed a novel, glucose-induced molecular mechanism in which miR-146a participates in the transcriptional circuitry regulating extracellular matrix protein production in diabetes.
微小 RNA（miRNAs）通过转录调控，调节多种细胞过程。在糖尿病中，已知细胞外基质蛋白纤维连接蛋白（FN）的产生增加是通过组蛋白乙酰转移酶 p300 发生的。在这里，我们研究了 FN 靶向 miRNA miR-146a 在糖尿病中 FN 产生中的作用及其与 p300 的关系。
在各种葡萄糖水平下，测量大血管内皮细胞和视网膜微血管中 miR-146a 的表达。检查 FN 信使 RNA 表达和蛋白水平，以及转染 miR-146a 模拟物或反义寡核苷酸后的表达。进行荧光素酶测定以检测 miR-146a 与 FN 3'非翻译区（UTR）的结合。同样，用或不用 miR-146a 模拟物进行玻璃体内注射来研究 1 型糖尿病大鼠的视网膜。用原位杂交法定位视网膜 miR-146a。分析 1 型和 2 型糖尿病动物的心脏和肾脏组织。
与 5mmol/L 葡萄糖相比，25mmol/L 葡萄糖使两种细胞类型中的 miR-146a 表达降低，FN 表达增加。miR-146a 模拟物转染可防止这种变化，而在 5mmol/L 葡萄糖中的细胞中转染 miR-146a 反义寡核苷酸则导致 FN 上调。荧光素酶测定证实 miR-146a 与 FN 3'UTR 结合。miR-146a 定位于视网膜内皮细胞，在糖尿病中减少。玻璃体内 miR-146a 模拟物注射可恢复糖尿病中的视网膜 miR-146a 并减少 FN。进一步的实验表明，p300 调节 miR-146a。在 1 型和 2 型糖尿病动物的视网膜、肾脏和心脏中也观察到类似的变化。
这些研究显示了一种新的葡萄糖诱导的分子机制，其中 miR-146a 参与调节糖尿病中细胞外基质蛋白产生的转录电路。
