Study on regulation of osteogenesis through BMP antagonists

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In this study, we aimed at understanding how BMP antagonists regulate osteogenesis. For this purpose, we first examined how many BMP antagonists are expressed in response to BMP in osteoblasts and then investigated function of each antagonist in osteogenesis.1)Search for BMP antagonists induced by BMP ligandsOsteoblastic cell line MC3TE1 cells were treated with BMP2.5.7, and 5 plus 7. We then examined mRNA expression of BMP antagonists using each specific primers. Noggin, Sost (Sclerostin) and PRDC were specifically expressed in response to BMP whereas expression levels of DAN, Tsg, USAG1, cereberus and Gremlin were not altered.2)Expression and function of SostIn situ hybridization study revealed that Sost mRNA is localized to osteogenic front of calvaria in close to the area of osterix expression in vivo. During osteoblastic differentiation, Sost mRNA was increased and this increase was blocked by adenoviral infection of Noggin into osteoblasts. Suppression of osterix resulted in decrease in Sost mRNA expression in osteoblasts.3)Suppression of Noggin and PRDC expression using siRNAWe designed several siRNA to suppress expression of Noggin and PRDC. Of the siRNAs, Noggin2 siRNA was more effective than Noggin4 and PRDC 880 siRNA is more effective than PRDC 373 and 367.Suppression of Noggin by siRNA did not change ALP activity and expression in osteoblast.4)Expression and function of PRDC.By in situ hybridization, we found that PRDC mRNA was expressed in primordia of vertebrae and skull during skeletogenesis. Over expression of PRDC using adenovirus in osteoblasts resulted in suppression of mRNA expression of ALP and osteocalcin in osteoblasts. ALP activity and numbers of mineralized nodules were also suppressed by PRDC expressing adenovirus5)Suppression of PRDC by siRNASuppression of endogenous expression PRDC by PRDC 880 siRNA resulted in elevation of ALP activity and expression in osteoblast. Numbers of mineralized nodules were also increased by si RNA.