188-LB: MicroRNA-126-3p Impacts Insulin Signaling in Smooth Muscle Cells

Abstract

Human umbilical vein endothelial cells (HUVEC) are a rich source of miRNA-126-3p. HUVEC package miR-126-3p into exosomes and secrete them into the extracellular space, potentially influencing other cells. Smooth muscle cells (SMC) take up miR-126-3p in a co-culture model with endothelial cells. miR-126-3p targets mRNAs in the insulin signaling pathway impacting their protein abundance. To test the hypothesis that miRNA-126-3p would decrease abundance of IRS1 and Akt, targets of miR-126-3p, SMC were isolated from the umbilical veins of infants born to women with (DM group; n=2) and without (control group; n=3) diabetes during pregnancy. miR-126-3p or a non-coding control mimic (NTC; 50 nM per miRNA) was transfected into SMC. After 48 h, SMC were stimulated for 10 min with 0 or 10 nM insulin. For analysis, abundance of all proteins was set relative to the NTC-transfected, unstimulated control group SMC (set at 100%). The fold change in IRS1 abundance was 85% +/- 72% and 85% +/- 30% for the control and DM groups, respectively. Abundance of Akt was 107% +/- 32% and 112% +/- 19% for the control and DM groups, respectively. pAkt abundance was 67% +/- 21% and 40% +/- 9% for the control and DM groups, respectively. Upon insulin stimulation, pAkt abundance in the control group increased to 114% +/- 29% of NTC, while in the DM group, abundance only reached 68% +/- 27% of control NTC. Furthermore, pAS160, necessary for GLUT4 translocation, was largely unchanged in the control group with or without insulin stimulation (96% +/- 23% and 99% +/- 45%, respectively). However, pAS160 abundance in the DM group was 45% +/- 4% of NTC and this reduction was not rescued by insulin stimulation (42% +/- 28%). These data suggest that miRNA-126-3p can decrease abundance of insulin signaling pathway proteins, and these decreases are more substantial in DM-exposed SMC. miRNA-126-3p appears to exert its affects through a decrease in IRS1, rather than through Akt. Future studies are planned to examine the impact of miRNA-126-3p on SMC function. Disclosure A.M. Teague: None. J.B. Tryggestad: None. Funding National Institutes of Health (1K23DK106533-01A1 to J.B.T.); Children's Medical Research Institute Metabolic Research Program; Harold Hamm Diabetes Center; University of Oklahoma Health Sciences Center