| Calcium phosphate ceramics (CaP ceramics) have been widely used in bone grafting. A lot of experiments have proved that certain calcium phosphate biomaterials are osteoinductive if they possess a specific porous structure. These calcium phosphate biomaterials induce bone formation at extra-skeletal sites without the need for additional osteogenic cells or growth factors.Previous studies have found that the following materials factors could affect the osteoinductive ability of these CaP ceramics: (1) porous structure; (2) phase composition; (3) degree of crystallinity and mechanics. In this paper, porous hydroxyapatite/tricalcium phosphate (HA/TCP) biphasic ceramics were employed for their better osteoinductive capacity compared with pure hydroxyapatite.In the osetoinductivity mechanism area, the target cells which involved in bone formation induced by CaP ceramics were unclear, especially the source of stem cells need to further research. Meanwhile, a lot of bone-related genes were expressed while these stem cells differentiate to osteoblasts, in the relation of material-genes, whether the cells could be induced by CaP ceramics to up-regulated the key gene which control bone growth and what kinds of material factors could affect the expression of the genes, these questions were main content in this paper.In the process of osteoinductivity of CaP ceramics, undifferentiated mesenchymal stem cells, early osteoprogenitor, late osteoprogenitor, preosteoblast and osteoblast, those kinds of cells all may be the target cells which could be induced to differentiation towards osteocyte by CaP ceramics. In this paper, osteoblasts, bone marrow stromal cells and myogenic progenitor cells were cultured for mechanism study of osteoinductivity, and for molecular biological data of cell differentiation.Firstly, the osteoinduction of primary cultured bone marrow stromal cells (MSCs) by CaP ceramics was studied, MSCs were separated from mouse marrow, expanded and co-cultured with CaP ceramics. If CaP ceramics had the osteoinductive effect to MSCs, when MSCs co-cultured with CaP ceramics for some days, the formation of bone tissue would be observed. Based on this hypothesis, diffusion chamber model was devised, porous CaP ceramics with MSCs were embedded in diffusion chamber, and were implanted in dog muscle for 6 months, then the implanted samples were taken out for histological observation. The bone like tissue was found to formed, it confirmed that CaP ceramics could induced MSCs to form bone tissue obviously and proved the osteoinductivity of CaP ceramics.Secondly, cell growth, proliferation and bone-related genes of osteoblasts on CaP ceramics were researched in vitro. CaP ceramics with different phase composition or different sintering temperature or different surface topography (bone-like apatite) were used in this work, the results proved that phase composition and sintering temperature could affect the expression of the bone-related genes and the bone-like apatite formed on the surface of CaP ceramics could enhance the mRNA expression of osteocalcin and type I collagen.At last, because of a lot of multipotent cells existed in muscle tissue, so we employed myogenic progenitor cells (C2C12 cell line) as cell model. As we know, In the process of bone formation, stem cells differentiated to osteoblasts gradually with high expression of bone-related gene (Cbfa1, OCN, Type I collagen, Osteopontin etc.). The gene, Cbfa1, had been identified to be an osteoblast-specific transcription factor and a key regulator of osteoblast differentiation. According to real-time RT-PCR, the gene of Cbfa1 was found to up-regulated within the C2C12 cells when they were cultured on the surface of CaP ceramics for 2 days and 5 days, at the same time, Osteocalcin gene also was detected to express highly. The results suggested that CaP ceramics had osteoinductivity property by causing expression of Cbfa1 and other bone special gene. |
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