Abstract 14268: MiR-133 Modulates Vascular Smooth Muscle Cell Phenotypic Switch in vitro and in vivo
Background: MicroRNA (mi-R)-1 and -133 are involved in various cardiovascular processes such as cardiac differentiation, development, hypertrophy and proliferation. However, the role of miR-1 and miR-133 in vascular disease is currently unknown. Thus, the aim of the present study was to evaluate the role, if any, of miR-1 and miR-133 in vascular smooth muscle cell (VSMC) phenotypic switch from quiescent/contractile to proliferative/synthetic state in vitro and in vivo.
Methods: Primary culture of VSMCs were transfected in vitro with adenoviral vector constructs carrying mir-1 or miR-133 or with the specific Antago-miRs. In the in vivo protocol, balloon injury of the right carotid was produced in male Wistar rats. Straight after the vascular injury, the balloon-dilated arteries were randomly transfected with adenoviral vectors carrying miR-1, miR-133 or green fluorescent protein (GFP). An empty adenoviral vector was transfected as additional control. Total RNAs were extracted from cultured VSMCs treated with the specific adeno-miRs and from rat carotid artery before and after vascular injury. MiR-1 and miR-133 were detected by quantitative real-time RT-PCR. In situ hybridizations (ISH) of miR-1 and miR-133 and co-immunofluorescence with the smooth muscle marker SM-actin were performed in carotid sections. Potential targets were analyzed by qRT-PCR, luciferase assay and western blot.
Results: Freshly isolated aortic VSMC expressed significant levels of miR-133 but not of mi-R−1. miR-133 levels decreased as VSMCs changed morphology and started to proliferate in vitro. Loss and gain of function experiments show that miR-133 regulates VSMC switch from the quiescent to proliferative state in vitro. miR-133 is down-regulated in VSMCs 2 days after balloon injury and increase then 14 days after vascular injury. Importantly, the in vivo transfection of adeno-miR-133 significantly reduced neointimal formation after balloon injury in rats. We identified Moesin and Sp-1 as specific targets of miR-133, which are key intermediary of miR-133 effects in VSMCs.
Conclusions: Our data show that miR-133 is a key regulator of VSMC phenotypic switch in vitro and in vivo, suggesting its potential therapeutic application for vasculo-proliferative diseases.
- © 2010 by American Heart Association, Inc.