MicroRNA profiling reveals new potential modulators of insulin-resistance and cardiovascular risk in type 2 diabetes

Background: Type 2 Diabetes (T2DM) is a chronic disease characterized by an inadequate beta-cell response to the progressive insulin resistance. The tiny mechanism(s) underlying insulin-resistance and increased atherosclerosis burden in T2DM patients are not fully understood. MicroRNAs (miRNAs) are short (20–22 nucleotides of length), endogenous, non-coding, RNAs representing a new class of regulators of gene expression. Remarkably, they are found in all cell type and their presence is clearly detectable in peripheral blood. Currently miRNAs involvement has been demonstrated in many diseases such as cancer, inflammatory and cardiovascular diseases. However, the role of miRNAs in T2DM in humans is not fully elucidated, thus aim of this study was to investigate the plasma miRNAs profile of diabetic patients. Materials and Methods: Blood samples were collected from 11 diabetic patients and 11 matched control patients. T2DM diagnosis was formulated according currently available American Diabetes Association guidelines. We enrolled only drug-naı¨ve patients before starting specific treatment. Patients with active inflammatory diseases, chronic kidney disease and cancer were excluded. RNA was extracted according with previously validated methods, quantified and pooled and a wide microRNA expression profiling was performed (miRNome). Then, some of the miRNAs that were differently expressed between two groups were validated by RealTime-PCR (RT-PCR). Data analyses were performed with deltadelta Ct method. Finally, bioinformatics was used in order to identify the potential targets of these miRNAs. Results: Microarray analysis showed that 4 miRNAs were upregulated whereas 21 miRNAs were downregulated in diabetic patients. Interestingly, RT-PCR validation confirmed a significant downregulation of let-7a (p = 0.023) and let-7f (p = 0.049). Moreover, we found a significant upregulation of miR-326 (p = 0.006). Furthermore, an interesting trend supporting down-regulation of miR-16, miR-21 and let-7 g in diabetic patients was found, despite these values did not reach statistical significance probably due the small study population. In silico analysis of predicted targets confirmed that these miRNAs may modulate genes greatly involved in insulin-signaling (including Adiponectin, IGF-1 receptor and others), endothelial function (including PTEN) and linked to cardiovascular risk (including VLDL receptor, TGF-beta). Conclusion: This study demonstrated that diabetes is associated with a modulation of the expression of plasma miRNA that are involved in insulin resistance and endothelial function. Further studies on these specific miRNAs’ targets are required to understand the molecular read-out of this modulation. If confirmed, these findings will contribute to improve our knowledge on diabetes pathophysiology and lead to the identification of new innovative therapeutic strategies for T2DM.