Bone Morphogenetic Protein‐2 Decreases MicroRNA‐30b and MicroRNA‐30c to Promote Vascular Smooth Muscle Cell Calcification

Abstract

Background Vascular calcification resembles bone formation and involves vascular smooth muscle cell ( SMC ) transition to an osteoblast‐like phenotype to express Runx2 , a master osteoblast transcription factor. One possible mechanism by which Runx2 protein expression is induced is downregulation of inhibitory micro RNA s (mi R ). Methods and Results Human coronary artery SMCs ( CASMCs ) treated with bone morphogenetic protein‐2 ( BMP ‐2; 100 ng/mL) demonstrated a 1.7‐fold ( P <0.02) increase in Runx2 protein expression at 24 hours. A mi R microarray and target prediction database analysis independently identified mi R ‐30b and mi R ‐30c (mi R ‐30b‐c) as mi R s that regulate Runx2 expression. Real‐time–polymerase chain reaction confirmed that BMP ‐2 decreased mi R ‐30b and mi R ‐30c expression. A luciferase reporter assay verified that both mi R ‐30b and mi R ‐30c bind to the 3′‐untranslated region of R unx2 m RNA to regulate its expression. CASMCs transfected with antagomirs to downregulate mi R ‐30b‐c demonstrated significantly increased R unx2, intracellular calcium deposition, and mineralization. Conversely, forced expression of mi R ‐30b‐c by transfection with pre–mi R ‐30b‐c prevented the increase in Runx2 expression and mineralization of SMCs . Calcified human coronary arteries demonstrated higher levels of BMP ‐2 and lower levels of mi R ‐30b than did noncalcified donor coronary arteries. Conclusions BMP ‐2 downregulates mi R ‐30b and mi R ‐30c to increase Runx2 expression in CASMCs and promote mineralization. Strategies that modulate expression of mi R ‐30b and mi R ‐30c may influence vascular calcification.