Objective: To observe dynamically the development of fetal long bone and detect the expression and distribution of HIF-1alpha,to investigate the expression pattern and possible effects of hypoxia inducible factor-1alpha (HIF-1alpha) in fetal long bone development of mouse.
Methods: E12.5, E13.5, E14.5, E15.5, E16.5 and E17.5 pregnant C57BL6 mice were sacrificed. After sacrifice, the embryos were delivered by caesarean section. The development of fetal long bone was dynamically observed by stereoscopic microscope, and the distributional expression of HIF-1alpha protein was detected by using method of immunohistochemistry. The expression of HIF-1alpha mRNA and osteoblast marker gene at various stage were also detected by using methods of reverse transcription-polymerase chain reaction (RT-PCR).
Results: The cartilaginous long bone began to form and joints outline arised at E13.5, then the primary ossification center was observed at E14.5, showing opaque ossification under stereoscopic microscope,and then the osteogenesis expanded and extended to both sides. Immunohistochemistry demonstrated lots of HIF-1alpha protein positive chondrcytes in the center of primary ossification at E14.5, then they decreased dramatically. HIF-1alpha mRNA expressed at high level from E13.5 to E15.5, and then decreased to low level.
Conclusion: Fetal long bone development pattern appeared to be endochondral osteogenisis process, existing hypoxia microenviroment may increase HIF-1alpha mRNA expression and thus initiate the cascade of endochondral osteogenisis.