BMP2 and GSK126 synergize to induce expression of osteogenic genes. BMP2 and GSK126 synergize to induce expression of osteogenic genes

BMP2 and GSK126 synergize to induce expression of osteogenic genes Overall design: MC3T3 sc4 pre-osteoblasts were treated with BMP2 and GSK126 (Ezh2 inhibitor). Our analysis shows that BMP2 and GSK126 synergistically active expression of osteogenic genes. MC3T3 sc4 cells were maintained in α-minimal essential medium without ascorbic acid (Gibco) containing 10% fetal bovine serum (Atlanta Biologicals), 100 units/ml penicillin and 100 μg/ml streptomycin (Gibco). Cells were seeded in maintenance medium at a density of 10,000 cells/cm2. Next day, cells were treated with vehicle, 5µM GSK126 (Xcess Biociences Inc.), 10 and 50ng/ml BMP2 (R&D Systems), and combination of GSK126 and BMP2 in osteogenic medium (maintenance medium supplementend with 50μg/ml ascorbic acid (Sigma) and 4mM β-glycerol phosphate (Sigma)). Three days later, old osteogenic medium was replaced with a fresh batch of osteogenic medium supplemented with vehicle, GSK126, BMP2, and combination of GSK126/BMP2. Total RNA was isolated on days one and six of osteogenic differentiation using the Direct-zol™ RNA kit (Zymo Research) and TRIzol reagent (Invitrogen).