OBJECTIVEmicroRNAs (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.RESEARCH DESIGN AND METHODSmiR-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.RESULTSA 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.CONCLUSIONSThese studies showed a novel, glucose-induced molecular mechanism in which miR-146a participates in the transcriptional circuitry regulating extracellular matrix protein production in diabetes.

In diabetes, some of the cellular changes are similar to aging. We hypothesized that hyperglycemia accelerates aging-like changes in the endothelial cells (ECs) and tissues leading to structural and functional damage. We investigated glucose-induced aging in 3 types of ECs using senescence associated β-gal (SA β-gal) staining and cell morphology. Alterations of sirtuins (SIRTs) and their downstream mediator FOXO and oxidative stress were investigated. Relationship of such alteration with histone acetylase (HAT) p300 was examined. Similar examinations were performed in tissues of diabetic animals. ECs in high glucose (HG) showed evidence of early senescence as demonstrated by increased SA β-gal positivity and reduced replicative capacities. These alterations were pronounced in microvascular ECs. They developed an irregular and hypertrophic phenotype. Such changes were associated with decreased SIRT (1–7) mRNA expressions. We also found that p300 and SIRT1 regulate each other in such process, as silencing one led to increase of the others’ expression. Furthermore, HG caused reduction in FOXO1’s DNA binding ability and antioxidant target gene expressions. Chemically induced increased SIRT1 activity and p300 knockdown corrected these abnormalities slowing aging-like changes. Diabetic animals showed increased cellular senescence in renal glomerulus and retinal blood vessels along with reduced SIRT1 mRNA expressions in these tissues. Data from this study demonstrated that hyperglycemia accelerates aging-like process in the vascular ECs and such process is mediated via downregulation of SIRT1, causing reduction of mitochondrial antioxidant enzyme in a p300 and FOXO1 mediated pathway.

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Glycyrrhizin is quite effective against hyperglycaemia, hyperlipidaemia and associated oxidative stress, and may be a potential therapeutic agent for diabetes treatment.