Osteoblast-specific Knockout Of The Smad4 Gene Results In Early Onset Osteoporosis

The transforming growth factor-β(TGF-β) superfamily plays important roles during vertebrate development by regulating cell proliferation, differentiation, migration and apoptosis. A large amount of TGF- s are stored in the bone matrix and serve as important regulators of skeletal development and bone remodelling. Smad4 is a central mediator of TGF- signals and is expressed in many tissues during embryogenesis and adulthood. It was previously shown that loss of Smad4 results in lethality at embryonic days 6-7 and its role in osteoblast remains unclear. To further study functions of Smad4 in osteoblast proliferation and differentiation, we generated an osteoblast-specific Smad4 knockout mouse using the Cre-loxP system.The mice with disrupted Smad4 in osteoblast were phenotypically nomal at birth and developed a short stature 2 weeks later. Mutant mice can be distinguished from littermates by their small size and light weight from the third postnatal week. Their teeth were shorter and less mineralized. Adolescent and adult mutant mice exhibited a reduction in total bone mineral density characterized by decreased trabecular bone and marked thinning of the parietal bones of the skull. The osteoclasts on tibia sections of 16-day-old mutants were significantly reduced and became more obvious when the mutant mice aged. The growth plates of 3-week-old mutant mice were shorter. The results of in vivo fluorescent labelling showed that the bone formation was decreased. The results from in vitro cell culture indicated osteoblast differentiation was not affected, however, the data clearly indicated that the mutant osteoblasts with inactivated Smad4 gene could not response to both BMP-2 and TGF-1. The results of in situ hibrydization revealed osteoblasts were less and the expression of Runx2/Cbfal and many extracellular genes were reduced significantly. On the other hand, abolish of Smad4 gene in osteobiast may influence osteoclastformation and differentiation, resulted in decreased osteoclast number and bone resorption.In conclusion, our study showed that deletion of Smad4 gene in osteoblast gave rise to reduced bone formation and bone resorption, and resulted in early onset osteoporosis. These results suggested that Smad4 is a crucial regulator of modulating the normal osteoblast function and bone mass. The osteoporosis mouse model generated by disruption of Smad4 in osteoblast could be used for further study of the function and molecular mechanism of Smad4 mediated TGF-P signals in maitaining normal function of osteoblast and the precess of osteoporosis. The mouse model could also be used for screening the therapeutic targets and evaluating the treatments of osteoporosis.