Role Of Runx1and Its Co-transcrption Factor CBFβ In Osteoblast Differentiation And Bone Development

Bone is constructed and maintained by two processes, bone modeling and remodeling. Bone modeling, also known as bone development, can divide into two types, intramembranous ossification and endochondral ossification. Chondrocytes and osteoblasts are the two major cells involving in the two processes. Bone remodeling, also known as bone turnover, is a complicated process which is carrying on constantly throughout the life of vertebrate organism, and maintaining skeletal homeostasis. Bone remodeling is consisted of bone formation osteoblasts and bone resorption by osteoclasts. Imbalance caused by disorder of osteoblasts differentiation will cause bone metabolism disease such as osteoporosis. Thus, we study the role of Runx1and its co-transcrption factor Cbfβ in osteoblast differentiation and bone development Mesenchymal stem cell can differentiate into osteoblasts, chondrocytes, adipocytes, and other cell types. Runxl belongs to Runx family. Runx2, another member of Runx family, is one of the major transcription factors of osteoblast differentiation. Though the role of Runxl in chondrocytes proliferation has been reported recently, however, the role of Runxl in osteoblast differentiation is not clear. Runxl deficient mice died before born. Thus, we used FloxP-cre system to construct Runxl osteoblast-conditional knockout model to study the role of Runxl in skeletal development and mesenchymal stem cell differentiation. Compare with the wild type mice,8-week old Runxlf/f;Osx-cre mice have reduced bone mineral density, thinner cortical bone, thinner trabecular bone and more adipocytes. Osteoblast primary culture using new born mice calvarial cells showed that Runxlf/f deficiency reduced alkaline phosphatase activity and inhibited mineralization in vitro. More adipocytes formation was observed in Runxl f/f;Osx-cre cells. qRT-PCR and western-blot indicated that the major osteoblast differentiation marker gene expression is downregulated in Runxlf/f;Osx-cre osteoblasts compare to the Wild type cells. And adipocytes transcription factors C/EBPa and PPARβ expression is upregulated in Runxl deficient osteoblasts. We also proved that C/EBPa expression is inhibited by Runxl and Cbfβ interaction in mesenchymal stem cells. Therefore, Runxl can inhibit adipocyte differentiation and promote osteoblast differentiation, so as to promote bone formation and contribute to the balance of bone turnover.The β subunit of core binding factor can form a heterodimer with Runx family protein, which has an important role in bone development. Chondrocyte differentiation and bone ossification are inhibited in Cbfp deficient mice. Cbfβ doesn’t bind to DNA itself but it enhances the DNA-binding affinity of Runx family protein and promote its transcript activity. We construct Cbfβ mesenchymal-stem-cell conditional knockout mice (Cbfβf/f;Dermo-cre mice) by crossing the Cbfβ Flox mice with Dermo-Cre mice. Cbfβ is expressed in all growth plate zones while Runxl is mainly expressed in proliferation zone and Runx2is expressed in hypertrophic zone. Mice with Cbfβ deficient in mesenchymal cells displays dwarfism with shortened limbs and delayed ossification. Shortened long bones, elongated growth plates and disorganized proliferation cells are displayed in Cbfβf/f;Dermo-Cre embryos mice. Cbfβ promotes cartlige development through regulating chondrocyte proliferation and ColX expression. Cbfβ not only regulates chondrocytes proliferation but also regulates osteoblasts differentiation. Runx2expression is normal in Cbfpf/f;Dermo-Cre mice femur while Osterix expression is decreased. Cbfβ interact with Runx2to promote osteoblast specific transcription factor Osterix expression and regulate bone development.In summary, Runxl is an important transcription factor during osteoblasts differentiation. Runxl deficient mice have lowered bone mass with increased adipocytes in bone marrow. Cbfβ regulates bone development though promoting chondrocyte proliferation and osteoblast differentiation. Our finding provides Runxl and Cbfp for potential pharmaceutical targets for bone development and density diseases.