The Study Of The Mutual Induction Between Tooth Germ Cells And BMSCs Transfected By BMP4 Gene In Vitro

Objective::To construct lentiviral vector carrying bone morphogenetic protein4and detect the odontogenic effects on rat bone marrow mesenchymal stem cells (BMSCs), and to study the mutual induction between tooth germ cells and bone marrow mesenchymal stem cells with bone morphogenetic protein4 gene in vitro for tissue-engineered tooth seed cells. Methods::hBMP4 cDNA was gained by RT-PCR from the mRNA,which was extracted from mature human placenta. Then it was cloned into plenti6/v5-D-TOPO vector. Lentiviral vectors carrying BMP4 were constructed to transfect rat bone marrow mesenchymal stem cells(BMSCs). And the proliferation ability of BMSCs was detected with MTT method and after transfection. BMSCs and BMSCs tansfected by BMP4 were co-cultured with tooth germ cells (1:1) respectively.The cells were divided into five groups:BMSCs group、BMP4/BMSCs group, tooth germ cells group、BMSCs/tooth germ cells (the mixed groupl)、BMP4/BMSCs/tooth germ cells group (the mixed group2).The five groups were extracted using real-time quantitative PCR and western blotting to detect the mRNA and protein levels of collagen I, enamel protein,dentin matrix protein 1, and distal-less homeobox gene 1. Results(1) After transfection, the proliferation ability of BMSCs was increased.(2) Compared to BMSCs group,the mRNA and protein levels of enamel protein,dentin, matrix protein land distal-less homeobox gene lof BMP4/BMSCs group were higher, and the difference was statistically significant (P<0.05),but the mRNA levels of collagen I BMP4/BMSCs group were no significant difference (.P>0.05),and (2) Compared to the mixed groupl,the mRNA and protein levels of collagen I, enamel protein,dentin matrix protein 1, distal-less homeobox gene 1 of the the mixed group2 were higher, and the difference was statistically significant (P<0.05) Conclusion:BMP4 is able to improve the proliferation and odontogenic differentiation potential of BMSCs in vitro, and the co-culture of tooth germ cells and BMSCs tansfected by BMP4 can increase the expressions of odontogenic genes,and they are suitable as seed cells of tissue-engineered tooth.