Tooth-forming Potential Of Bone Marrow Mesenchymal Stem Cells In The Milieus Of Diverse Growth-Factor Induction And Scaffold In Vivo

Objective:To investigate the tooth-forming potential of bone marrow mesenchymal stem cells(BMSCs) in the milieus of different signaling induction and diverse scaffold in vivo. Methods:BMSCs were inducted via either a single signaling derived from co-culture with dental germ cells(DGCs) isolated from the 4-day-old SD rats, or a mixed multi-signaling from the co-culture of DGCs plus growth factors basic fibroblast growth factor(bFGF) and bone morphogenetic protein 2(BMP-2). After induction the BMSCs were further co-cultured in either Absorbable Gelatin Sponge(AGS), or Polyvinyl Alcohol-double crystal ceramic powder(PVA-DCCP) scaffold constructed by a three dimensions(3D) printer. Then, the 3D co-culture messes were replanted into the renal capsules of 8-week-old SD rats divided into 5 groups: Group A-DGCs-induced BMSCs with AGS; Group B-DGCs+bFGF+BMP-2-induced BMSCs with AGS; Group C-DGCs-induced BMSCs with PVA/DCCP scaffold; Group D-DGCs+bFGF+BMP-2-induced BMSCs with PVA/DCCP scaffold; and Group E-DGCs alone. The replanted tissues were harvested and the real-time quantitative PCR(RT-PCR) of Ameloblastin(AMBN), dentin matrix acidic phosphoprotein 1(DMP1), Collagen-?, and DLX1 mRNA and HE staining were performed at 1, 3, 7, 14 and 28 days post-replantation to assess teeth-forming potentials genetically and historically.Results: The expression of Collagen-? increased gradually with the prolongation of culture time, whereas expression of AMBN, DMP1 and DLX1 decreased, and the difference of expression between these four genes and those in E group was statically significant(P﹤ 0.05). The differences of mRNA expression levels among the four groups mentioned above was statistically significant between different scaffolds(P﹤ 0.0001), and the differences in induction signals do affect the expression of each gene, but not as obvious as that of scaffold material. DGCs can virtually develop into a normal dental structure at 28-day post-replantation, whereas the BMSCs in group B can develop into a tooth-like structure incompletely, and the BMSCs in the group A, C, and D failed to form tooth-like mineralized tissue. The scaffolds in group C and D were absorbed incompletely. Conclusion:The mixed multi-signaling induced BMSCs could proliferate more quickly and differentiate more profoundly, comparing to the single signaling induced ones. Moreover, those BMSCs express more corresponding odontogenic genes promoting the formation of tooth tissue when combined with scaffold. It is of great value in transforming non-odontogenic cells into odontogenic ones to optimize the scaffold and in establishing proper tissue engineering tooth model.