Dishing up bone formation
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《细胞学杂志》
In 1979, he and his colleagues started establishing cell lines that differentiated into osteoblasts (bone-forming cells) rather than odontoblasts (dentin-forming cells). At that time, only a few primary bone cell cultures had been reported to show hints of in vitro bone formation (Binderman et al., 1979; Nijiweide et al., 1982).
Kodama (RIKEN, Wako Saitama, Japan), says the secret to success was using the same cell culture method used to make the immortalized mouse fibroblast 3T3 cell line (Todaro and Green, 1963; see "A cell line that is under control" JCB 168:988). This meant repeated subcultivation of newborn mouse skull bone cells under 3T3 conditions—3 days to transfer at a factor of 3 cell plating density.
When one of the lines that arose, called MC3T3-E1, became confluent, it exhibited properties of osteoblasts, including high alkaline phosphatase activity and staining for calcified secretions (Sudo et al., 1983). At day 21 of culture, calcified nodes appeared and then grew in number and size to eventually fuse with one another. By day 30, white nodes in the dish were visible to the naked eye. But because calcium is easily deposited under basic culture conditions, the team had to show that they were seeing true bone formation.
Making bone in a dish: osteoblasts (Ob) surround a nodule of young osteoclasts (YOc) embedded in mineralized matrix.
KODAMA
Mineralization proceeded in much the same way it did in vivo, by the secretion of matrix vesicles containing crystals, which were deposited along collagen fibrils. Electron diffraction defined the crystals as hydroxyapatite, the chemical that forms bone matrix.
But in vitro bone formation, Kodama notes, was not easily reproducible by other groups until the discovery that bone morphogenetic protein acts as a potent inducer of osteoblast differentiation (Yamaguchi et al., 1991). The MC3T3 cell line remains an important tool for studying bone cell differentiation today, with renewed focus on mesenchymal stem cell differentiation and the need for better osteoporosis therapies, like bone-building agents. In fact, the line was the most widely distributed by the RIKEN Bioresource Center (Tsukuba, Japan) in 2003. KP
Binderman, I., et al. 1979. Science. 206:222–225.
Nijiweide, P.J., et al. 1982. J. Cell Biol. 93:318–323.
Sudo, H., et al. 1983. J. Cell Biol. 96:191–198.
Todaro, G.J., and H. Green. 1963. J. Cell Biol. 17:299–313.
Yamaguchi, A., et al. 1991. J. Cell Biol. 113:681–687.(Hiroaki Kodama knew little about dental )
Kodama (RIKEN, Wako Saitama, Japan), says the secret to success was using the same cell culture method used to make the immortalized mouse fibroblast 3T3 cell line (Todaro and Green, 1963; see "A cell line that is under control" JCB 168:988). This meant repeated subcultivation of newborn mouse skull bone cells under 3T3 conditions—3 days to transfer at a factor of 3 cell plating density.
When one of the lines that arose, called MC3T3-E1, became confluent, it exhibited properties of osteoblasts, including high alkaline phosphatase activity and staining for calcified secretions (Sudo et al., 1983). At day 21 of culture, calcified nodes appeared and then grew in number and size to eventually fuse with one another. By day 30, white nodes in the dish were visible to the naked eye. But because calcium is easily deposited under basic culture conditions, the team had to show that they were seeing true bone formation.
Making bone in a dish: osteoblasts (Ob) surround a nodule of young osteoclasts (YOc) embedded in mineralized matrix.
KODAMA
Mineralization proceeded in much the same way it did in vivo, by the secretion of matrix vesicles containing crystals, which were deposited along collagen fibrils. Electron diffraction defined the crystals as hydroxyapatite, the chemical that forms bone matrix.
But in vitro bone formation, Kodama notes, was not easily reproducible by other groups until the discovery that bone morphogenetic protein acts as a potent inducer of osteoblast differentiation (Yamaguchi et al., 1991). The MC3T3 cell line remains an important tool for studying bone cell differentiation today, with renewed focus on mesenchymal stem cell differentiation and the need for better osteoporosis therapies, like bone-building agents. In fact, the line was the most widely distributed by the RIKEN Bioresource Center (Tsukuba, Japan) in 2003. KP
Binderman, I., et al. 1979. Science. 206:222–225.
Nijiweide, P.J., et al. 1982. J. Cell Biol. 93:318–323.
Sudo, H., et al. 1983. J. Cell Biol. 96:191–198.
Todaro, G.J., and H. Green. 1963. J. Cell Biol. 17:299–313.
Yamaguchi, A., et al. 1991. J. Cell Biol. 113:681–687.(Hiroaki Kodama knew little about dental )