The Culture And Differentiation Of The Mesenchymal Stem Cells Of Tooth Germ Of The Fetal Mice In Different Conditions

To isolate and culture the mesenchyme stem cell of the 1st lower molar tooth germ of the fetal mice, and to identify their characteristics under the culturing situation in vitro. To enlarge the quantity of the cells in vitro and to identify the qualification by which the mesenchyme stem cell of tooth germ could differentiate to express the cell markers of odontoblast cells. And, to prepare the 3-dimensional porous scaffold adapt to the adhesion and growth of the mesenchyme stem cell of tooth germ, in order to seed the cells on which to be cultured in vitro and in vivo.The fetal mice of 15. 5 days was used to isolate the the 1st lower molar tooth germ, and the dental papillae and sac were isolated mechanically and cells were treated enzymatically and cultured in vitro. The phase-contrast microscope was applied to record the morphology and the growth rate, and the cell characteristics were identified by immunohistochemistry method. Leukemia inhibitory factor (LIF) was used to maintain the undifferentiated station of the cells , characteristics were also identified by immunohistochemistry method to ensure the cell differentiation state. Polylactic-co-glycolic acid (PLGA) porous scaffold of different consistency was manufactured to select the better kind by considering theirphysiochemical and mechanical property. The expanded mesenchymal stem cells were seeded on the scaffold to be cultured in vitro, and the cells-scaffold complex was implanted under the skin of nude mice to realize the capability of minerilization and differentiation of the cells seeded on the scaffold.The cultured mesenchymal stem cells of the 1st lower molar tooth germ of the fetal mice were spindle-shaped or multiangular cells, of which the high activity of proliferation were observed , CD57/HNK-1 and Vimentin positive, and most of the cells express collagen type I and telomerase. D/F12 culture medium with 10% fetal cattle serum could induce the stem cells to express DSPP, which is the differentiation marker of odontoblast, While LIF showed powerful ability to inhibit the progress and maintain the activity of the cells to proliferate. The scaffolds of 88.8% and 92.2% consistency were manufactured successfully, and the latter showed better physiochemical and mechanical property. The ideal adhesion ability between the seeded cells and scaffold and the cell differentiation toward odontoblasts were observed in vitro. The implantation study displayed the degradation of the materials and the formation of the new-born hard tissue. However , the typical structure of tooth dentine was not observed.The mesenchymal stem cells of tooth germ of fetal mice were successfully cultured, which could be expanded to a huge quantity. The induction of the cells was successfully applied both in the plain culture situation and the 3D environment, of which the differentiation markers were DSPP expression andthe morphology characteristics. Anyhow, the mesenchymal stem cells of tooth germ and the manufactured porous degradable PLGA scaffold possessed fine prospect to be employed in the study of tissue engineering of tooth.