Abstract 328: High-Density Lipoproteins Increase miR-223 Levels in Human Endothelial Cells, Which Regulate Scavenger Receptor Class B Member 1

Abstract

Background: MicroRNAs (miRNAs) are recently discovered small non-encoding RNAs that control and regulate gene expression. We and others have recently demonstrated that miRNAs play key roles in lipoprotein metabolism. Here we aim to determine if miR-223 regulates scavenger receptor class B member 1 (SR-B1) in human endothelial cells. Methods and results: In silico target prediction studies suggest that SR-BI harbors a miR-223 target site within its 3’ untranslated region (3’UTR). This possibility was investigated by determining the miRNA profile of human coronary artery endothelial cells (HCAECs) using TaqMan arrays. Mature miR-223 levels were found in HCAECs and confirmed by real-time PCR. Here we demonstrate that incubation of HCAECs with native HDL (1 mg protein/ml) for 16 h increased miR-223 levels by 4-fold increase (p<0.007) as assessed by TaqMan Assay and decreased SR-B1 mRNA levels by 29±6% (p<0.05) as measured by real-time PCR. To determine if endogenous levels of miR-223 in HCAECs are sufficient to repress SR-BI mRNA, we used hairpin inhibitors (100 nM) to block miR-223 activity. Inhibition of miR-223 resulted in an increase in SR-B1 mRNA levels by 33±11% (p<0.05). Furthermore, transfection of HCAECs with miR-223 mimic (100 nM) decreased SR-B1 mRNA levels by 27±4% (p<0.01). Transient transfection of HCAECs with miR-223 mimics also reduced SR-BI protein levels by 30±9%. Using gene reporter (luciferase) assays, we determined that miR-223 directly targets the 3’UTR of SR-B1. Conclusion: miR-223 directly targets and represses endothelial cell SR-B1 levels, and may regulate HDL-induced cholesterol efflux and lipoprotein metabolism.