Influence of FGF23 silence on differentiation of rat calvarial osteoblastic cells by parathyroid hormone invitro

Objective To investigate the roles of fibroblast growth factor 23 (FGF23) in osteoblast by RNA interference (RNAi) and its influence on cell differentiation by parathyroid hormone (PTH) in vitro. Methods The primary rat calvarial osteoblasts were cultured in MEM medium containing 10% charcoal stripped fetal bovine serum(CSFBS) and treated with rhPTH1-34 for 3 days.The changes of proliferation and alkaline phosphates (ALP) activity were measured by MTT and PNPP methods respectively,and the ALP and OCN mRNA levels were determined by real-time RT-PCR.Further FGF23 was transiently silenced by shRNA method in osteoblasts and the expression levels of FGF23,ALP and OCN were determined in transcriptional levels after rhPTH1-34 treatment. Results rhPTH1-34 prompted osteoblasts proliferation while less effect on its differentiation.The cellsproliferations were increased 31.6%~50.5% (P0.05) by rhPTH1-34 in range of 1×10-10-1×10-8 mol/L,while the cell ALP activities (calculated by D405/D570) were slightly but no significantly changed.Meanwhile,the mRNA levels of ALP and OCN were slightly increased 35% (P0.05) and 16% (P0.05) by 1×10-9 mol/L rhPTH1-34 in 2 hours treatment.The FGF23 expression was up-regulated (about 4 folds) in osteoblasts by 1×10-9 mol/L rhPTH1-34 in transcriptional level and which was turned down by the transfection of shRNA of FGF23.Further,the marked stimulating effects of rhPTH1-34 on ALP and OCN expression (about 1.8 and 5.8 folds respectively) were showed in the transfected osteoblasts. Conclusions The expression of FGF23 might involve in osteogenesis regulation by PTH.The up-regulation of FGF23 expression by PTH could suppress its stimulating effect on osteoblasts differentiation,and the underlying mechanisms remain to be clarified.