Abstract 14365: Identification of MicroRNAs and Their Target Genes in Microvascular Disease in Diabetes

Abstract

Cardiovascular disease is the leading cause of death in patients with diabetes. Among coronary vascular defects, obstructive coronary artery disease (CAD) is the primary cause of cardiac mortality. However, there is growing evidence that patients with coronary microvascular disease (CMD, also known as non-obstructive coronary disease) also experience chest pain and occasional heart attack. Furthermore, recent studies show that diabetes is the risk factor of CMD. However, the molecular mechanisms by which diabetes develops CMD is not fully understood. MicroRNAs (miRNAs) are a non-coding small RNA that regulates gene expression at the post-transcriptional level, and the change of miRNA profile is implicated in many diseases. In this study, we identified the core miRNAs which are involved in the development of coronary endothelial dysfunction in diabetes. We used an inducible type 2 diabetic (T2D) mouse model generated by high-fat diet and a single injection of low-dose streptozotocin. Our T2D mice did not show detectable atherosclerotic plaque, but developed CMD evidenced by reduced coronary flow velocity reserve. We conducted a miRNA array using mouse coronary endothelial cells (MCECs) isolated from diabetic and control mice, and selected seven miRNAs based on the result. Next, real-time PCR assays were carried out with seven miRNAs and found that miR210 and miR342-5p were downregulated, and miR378 was upregulated in MCECs of diabetic mice compared to the control. To identify the target genes of miR342-5p and miR378, we inhibited miR342-5p or overexpressed miR378 in MCECs. After testing 93 genes that are involved in endothelial functions, we found that inhibition of miR342-5p decreased the levels of Atp2a3 , Opa1 , Sod3 , and Vegfb , whereas miR378 overexpression significantly decreased Aggf1, HK1, Mapk3, Pak1, Panx1, Stim1, Stim2, Vcam1, and Vegfb and increased Casp2 and Gja1 . Western blotting data revealed that the expression levels of Affg1, Opa1, and Pak1 were significantly decreased in the MCECs from diabetic mice compared to the control. In summary, these data suggest that miR342-5p and miR378 could be a potential therapeutic target for CMD in diabetic patients, and Affg1 Opa1 and Pak1 are the downstream mRNAs of miR342-5p or miR378.