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Background.  Mouse dental mesenchymal cells (mDMCs) from tooth germs of cap or later stages are frequently used in the context of developmental biology or whole-tooth regeneration due to their odontogenic potential.  In vitro -expanded mDMCs serve as an alternative cell source considering the difficulty in obtaining primary mDMCs; however, cultured mDMCs fail to support tooth development as a result of functional failures of specific genes or pathways. The goal of this study was to identify the genes that maintain the odontogenic potential of mDMCs in culture.   Methods.  We examined the odontogenic potential of freshly isolated versus cultured mDMCs from the lower first molars of embryonic day 14.5 mice. The transcriptome of mDMCs was detected using RNA sequencing and the data were validated by qRT-PCR. Differential expression analysis and pathway analysis were conducted to identify the genes that contribute to the loss of odontogenic potential.   Results.  Cultured mDMCs failed to develop into well-structured tooth when they were recombined with dental epithelium. Compared with freshly isolated mDMCs, we found that 1,004 genes were upregulated and 948 were downregulated in cultured mDMCs. The differentially expressed genes were clustered in the biological processes and signaling pathways associated with tooth development. Following  in vitro  culture, genes encoding a wide array of components of MAPK, TGF- β /BMP, and Wnt pathways were significantly downregulated. Moreover, the activities of  Bdnf ,  Vegfα ,  Bmp2 , and  Bmp7  were significantly inhibited in cultured mDMCs. Supplementation of VEGF α , BMP2, and BMP7 restored the expression of a subset of downregulated genes and induced mDMCs to form dentin-like structures  in vivo .   Conclusions.   Vegfα ,  Bmp2 , and  Bmp7  play a role in the maintenance of odontogenic potential in mDMCs.

Insight into the maintenance of odontogenic potential in mouse dental mesenchymal cells based on transcriptomic analysis