MicroRNA-25–5p counteracts oxidized LDL-induced pathological changes by targeting neuronal growth regulator 1 (NEGR1) in human brain micro-vessel endothelial cells

Abstract

MicroRNA-25–5p (miR-25–5p) may be involved in the pathogenesis and processes of vascular diseases. The aim of this study was to investigate the role of miR-25–5p in oxidized low-density lipoprotein (ox-LDL)-treated human brain microvessel endothelial cells (HBMECs) and the underlying mechanisms. RT-qPCR and/or Western blot were used to detect the expression levels of miR-25–5p and neuronal growth regulator 1 (NEGR1). The effect of miR-25–5p overexpression and NEGR1 silencing on cell proliferation, migration, apoptosis and reactive oxygen species (ROS) production of HBMECs were measured by using CCK-8 assay, transwell assay and flow cytometry, respectively. The expression levels of apoptosis-related protein (cleaved caspase-3 and pro-caspase-3) were detected using Western blot, and the nitric oxide (NO) production was measured by a nitric oxide assay kit. The expression level of miR-25–5p was decreased in HBMECs treated with ox-LDL. Compared with the control group, miR-25–5p overexpression significantly promoted the proliferation and migration of HBMECs treated with ox-LDL (p < 0.01). Overexpression of miR-25–5p significantly suppressed cell apoptosis, ROS production and NO reduction of ox-LDL-induced HBMECs (p < 0.01). In addition, the target gene of miR-25–5p was predicted to be NEGR1 through Targetscan online analysis. The effect of NEGR1 silencing on cell proliferation, migration, apoptosis, ROS and NO production of ox-LDL-induced HBMECs was similar to that of miR-25–5p overexpression. Furthermore, miR-25–5p overexpression and NEGR1 silencing significantly downregulated the protein expression levels of JAK2 and STAT3. Thus, miR-25–5p neutralizes the effects of ox-LDL on multiple functions of HBMECs through suppressing the expression of NEGR1 via regulating the JAK/STA signaling pathway.