The Role Study Of CEBPα In Skeletaldevelopment By Osteoblast Conditional Knockout Mice

Bone is an important component of the body,and bone development is closely related to the differentiation of chondrocytes and osteoblasts.Osteoblast is an important component of human bone tissue,and is the main functional cell involved in bone formation,and play a key role in bone growth and bone mass maintenance.It is derived from mesenchymal stem cells(MSCs),which is with pluripotent differentiation potential.MSCs grow into osteocytes by cell proliferation,extracellular matrix synthesis,maturation and mineralization,and thus promoting bone formation and maintaining bone mass.MSCs are important member of the stem cell family,derived from the early development phase of mesoderm,belong to pluripotent stem cells and have self-replication ability and strong differentiation potential.MSCs are multipotent cells that are able to differentiate into several cell lineages,including adipogenic,chondrogenic and osteogenic precusor.MSCs form osteoblasts in the bone resorption site by migration,proliferation and differentiation,and mediate new bone formation.The osteogenic differentiation of MSCs is the key process of bone formation.Once the osteoblast differentiation is abnormal,affecting bone development and bone reconstruction,it will cause a variety of bone metabolism diseases,such as osteoporosis.Therefore,it is very important to study the differentiation mechanism of osteoblasts.At present,there is still a blind spot on molecular mechanism of osteogenic differentiation,and thus the treatment of bone metabolism related diseases is limited.In recent years,MSCs osteogenic differentiation mechanism has become a hot topic in both clinical and basic research areas,including research of the regulation signal and some important osteogenic transcription factors.C/EBP(CCAT/enhancer binding protein,C/EBP)is a kind of eukaryotic transcription factors,and many scholars have made studies on its structure and function.Previous studies have found that C/EBPα is an important transcription factor that plays an important role in adipocyte differentiation.In recent years,some scholars found C/EBPb was involved in not only adipocyte differentiation but also osteoblast differentiation process.However,study of C/EBPα in the field of osteoblast differentiation is basically blank.In our preliminary study,we found that similar to C/EBPβ,C/EBPαt will affect the development of chondrocytes and the process of osteogenesis,if it is knocked out from chondrocytes.To study the role of C/EBPα in skeletal development and MSCs differentiation,we constructed C/EBPα osteoblast-conditional knockout mice model(C/EBPα f/f;Osx-cre)by conditional knockout technique.Compare to the wild type mice,C/EBPα f/f;Osx-cre mice reduced bone mineral density,cortical bone becomed thinner and trabecular bone becomed smaller.We used new born mice calvarial cells to do osteoblast primary culture,it showed that C/EBPαf/f;Osx-cre mice reduced alkaline phosphatase activity and inhibited mineralization in vitro.The result of qRT-PCR and western-blot indicated that compare to the Wild type cells,the gene expression of major osteoblast differentiation marker was down regulated in C/EBPαf/f;Osx-cre osteoblasts.In conclusion,C/EBPα is an important transcription factor during osteoblasts differentiation and bone formation.These findings laid the foundation for the follow-up study,but the specific regulatory mechanism is not clear and need further study.