Abstract 3383: MicroRNA Let-7f Augments Angiogenic Properties of Endothelial Progenitor Cells through AMPK Activation and MnSOD Induction in Type 1 Diabetes
Introduction: Endothelial progenitor cells (EPCs) play an essential role in angiogenesis, but are dysfunctional in diabetes featuring excessive oxidative stress. Activation of AMP-activated protein kinase (AMPK) via its phosphorylation improves endothelial function and angiogenesis in diabetes. Emerging evidence suggests that microRNAs (miRNAs) may serve as novel regulators of angiogenesis. However, whether AMPK and miRNAs regulate EPC function in diabetes is unknown. Thus, we tested the hypothesis that miRNA let-7f improves EPC-mediated angiogenesis through AMPK activation and mitochondrial superoxide dismutase (MnSOD) induction.
Methods and Results: Bone marrow-derived EPCs from adult male streptozotocin (STZ)-induced type 1 diabetic mice were used (glucose 456.0±24.46 vs. 165.3±7.35 mg/dl, n=36, p<0.01). Endogenous let-7f expression was decreased by >60% in diabetic vs. normal EPCs (real-time PCR, n=4 –5, p<0.05). Transfection of diabetic EPCs with let-7f mimic in vitro resulted in a significant improvement of their angiogenic (2.1 fold by Matrigel tube formation assay) and adhesion (3 fold) functions (n=5, p<0.05 vs. scramble), both were retarded by the selective AMPK inhibitor compound C. Let-7f mimic transfection also increased AMPK phosphorylation (1.4 fold) and MnSOD (2.0 fold) in diabetic EPCs (Western blot analysis, n=5– 8, p<0.05 vs. scramble). Compound C pretreatment decreased AMPK phosphorylation (by 37%, n=4, p<0.01) and MnSOD induction (by >50%, n=4, p<0.05) induced by let-7f mimic transfection. Furthermore, elevated mitochondrial ROS level in diabetic EPCs was significantly reduced after let-7f mimic transfection (fluorescent intensity of MitoSOX Red dye, n=5, p<0.05 vs. scramble), an effect that was prevented by compound C. Protein phosphatase 2A (PP2A), a key enzyme for AMPK dephosphorylation and inactivation, was increased 1.2 fold in diabetic EPCs (n=5, p<0.05 vs. normal), which was inhibited by let-7f mimic transfection (n=5, p<0.05 vs. scramble). Finally, let-7f inhibitor transfection significantly impaired normal EPC angiogenic and adhesion functions (n=5, p<0.05 vs. scramble).
Conclusion: MicroRNA let-7f improves EPC angiogenesis through AMPK activation and MnSOD induction in type 1 diabetes.