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Bone Morphogenetic Proteinâ2 Decreases MicroRNAâ30b and MicroRNAâ30c to Promote Vascular Smooth Muscle Cell Calcification
Background: Vascular calcification resembles bone formation and involves vascular smooth muscle cell (SMC) transition to an osteoblastâlike phenotype to express Runx2, a master osteoblast transcription factor. One possible mechanism by which Runx2 protein expression is induced is downregulation of inhibitory microRNAs (miR). Methods and Results: Human coronary artery SMCs (CASMCs) treated with bone morphogenetic proteinâ2 (BMPâ2; 100 ng/mL) demonstrated a 1.7âfold (P<0.02) increase in Runx2 protein expression at 24 hours. A miR microarray and target prediction database analysis independently identified miRâ30b and miRâ30c (miRâ30bâc) as miRs that regulate Runx2 expression. Realâtimeâpolymerase chain reaction confirmed that BMPâ2 decreased miRâ30b and miRâ30c expression. A luciferase reporter assay verified that both miRâ30b and miRâ30c bind to the 3âČâuntranslated region of Runx2 mRNA to regulate its expression. CASMCs transfected with antagomirs to downregulate miRâ30bâc demonstrated significantly increased Runx2, intracellular calcium deposition, and mineralization. Conversely, forced expression of miRâ30bâc by transfection with preâmiRâ30bâc prevented the increase in Runx2 expression and mineralization of SMCs. Calcified human coronary arteries demonstrated higher levels of BMPâ2 and lower levels of miRâ30b than did noncalcified donor coronary arteries. Conclusions: BMPâ2 downregulates miRâ30b and miRâ30c to increase Runx2 expression in CASMCs and promote mineralization. Strategies that modulate expression of miRâ30b and miRâ30c may influence vascular calcification