牙胚细胞与骨形态发生蛋白4转染的骨髓间充质干细胞相互诱导的体外实验研究

目的:探讨牙胚细胞和骨形态发生蛋白4(BMP4)转染的骨髓间充质干细胞(BMSCs)相互间的成牙诱导作用。方法:构建骨形态发生蛋白4慢病毒载体,转染SD大鼠骨髓间充质干细胞,并将骨髓间充质干细胞和BMP4转染的骨髓间充质干细胞分别与SD大鼠牙胚细胞按1∶1比例混合培养,同时将细胞分为5组,BMSCs组、BMP4/ BMSCs组、牙胚细胞组、BMSCs/牙胚细胞组(混合组1)、BMP4/ BMSCs/牙胚细胞组(混合组2),分别采用实时荧光定量PCR和western-blotting检测五组细胞Ⅰ型胶原蛋白、成釉蛋白、牙本质基质蛋白1、同源异型盒基因1成牙相关基因mRNA水平和蛋白水平相对表达量的变化。结果:与混合组1相比,混合组2Ⅰ型胶原蛋白、成釉蛋白、牙本质基质蛋白1、同源异型盒基因1 mRNA水平和蛋白水平表达量增多,差异有统计学意义(P<0.05)。结论:牙胚细胞与BMP4转染的骨髓间充质干细胞共培养,促进了成牙相关基因的表达,可作为组织工程牙的备选种子细胞。

Study on the Mutual Induction between Tooth Germ Cells and Bone Marrow Mesenchymal Stem Cells Transfected by Bone Morphogenetic Protein4 Gene in Vitro.

Objective: To study the mutual induction between tooth germ cells and bone marrow mesenchymal stem cells with bone morphogenetic protein4 gene in vitro. Methods: Lentiviral vectors carrying BMP4 were constructed to transfect rat bone marrow mesenchymal stem cells(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 group (the mixed group1), 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 homeobox gene 1. Results: In the mixed group2, mRNA and protein levels of collagen I, enamel protein,dentin matrix protein 1 and homeobox gene 1 were higher, and the difference was statistically significant (P<0.05). Conclusion: 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.