Abstract 17909: The P75NTR-induced MicroRNA-30c-2* Impairs Endothelial Cell Survival and Angiogenesis
MicroRNAs (miRs) negatively regulate the expression of targeted mRNAs. Diabetes induces expression of the anti-angiogenic and pro-apoptotic p75 neurotrophin receptor (p75NTR) in endothelial cells (ECs). Here, we aim to elucidate the contribution of p75NTR-induced miR30c2* in endothelial dysfunction in diabetes.
A miR array (Exiqon) was done on human umbilical vein ECs (HUVECs) transduced to overexpress p75NTR (control: Null). miR30c2* expression was further measured by Taqman PCR in HUVECs and CD146+ ECs sorted from the adductor muscles of mice with/out streptozotocin-induced diabetes mellitus. Next, we studied the impact of miR-30c-2* on apoptosis (caspase 3/7 assay) and angiogenesis (Matrigel) in HUVECs cultured under normal conditions or exposed to high D-glucose (HG) and low growth factors (LGF), thus mimicking advanced diabetes, when hyperglycaemia is accompanied by tissue starvation. HUVEC miR-30c2* expression was modulated by pre-miR precursor, anti-miR inhibitor or negative control (Ambion). Predicted miR target genes were searched (Microcosm and TargetScan). p75NTR increased miR30c2* in HUVECs. Moreover, miR-30c-2* expression was upregulated under LGF, HG and a LGF/HG combination (p<0.01). In addition, miR-30c-2* was three-fold upregulated in muscular ECs of diabetic mice (p<0.01). miR-30c-2* overexpression induced HUVEC apoptosis and impaired angiogenesis. Conversely, miR-30c-2* inhibition prevented LGF- and LGF/HG-induced apoptosis and impaired angiogenesis. The cell cycle regulator minichromosome maintenance complex component 7 (MCM7) is a putative miR-30c-2* target. MCM-7 mRNA levels were decreased in p75NTR-HUVECs (p<0.05 vs. Null) and in HUVECs cultured in HG/LGF (p<0.05). miR-30c-2* overexpression reduced MCM7 mRNA (p<0.01).
In conclusion, the p75NTR-induced miRNA miR-30c-2* represses EC survival and angiogenesis and may prove to be a novel therapeutic target for diabetes-induced endothelial damage in limbs.
- © 2013 by American Heart Association, Inc.