[The effect of different factors on forming-dentin differentiation of rat dental mesenchymal cells].

Purpose: To investigate the effects of combined use of bFGF, IGF1, BMP4 and TGF-β1 on forming-dentin differentiation of rat dental mesenchymal cells (rDMCs).

Methods: Enzyme and differential digestions were performed to isolate and culture rDECs and rDMCs, and immunofluorescence staining against cytokeratin and vimentin were carried out to identify cell sources. Then alizarin red staining and Gomori calcium-cobalt method were used to detect the mineralization ability of rDMCs after mineralized induction. Immunohistochemistry, image analysis, real-time PCR and Western blot were utilized to determine the expression differences of DSPP/CAP/OPN/OCN in rDMCs after induction by bFGF+IGF1 (group 1), TGF-β1+BMP4 (group 2) and bFGF+IGF1+TGF-β1+BMP4 (group 3), respectively. Statistical analysis was performed with SPSS 14.0 software package.

Results: The rDECs and rDMCs were isolated, cultured and identified successfully. Calcium nodus and ALP staining were positive in cytoplasms of rDMCs after being induced by mineralization liquid. In groups 1 and 2, the expression levels of DSPP/CAP/OPN/OCN mRNA and protein were notably higher than those of control group, significant differences were found between groups (P<0.01). Among them, the expression levels of CAP/OCN in group 1 and DSPP/OPN in group 2 were the highest, respectively.

Conclusions: The rDMCs possess osteogenesis property after mineralization induction. bFGF+IGF1 can notably promote the expressions of CAP/OCN, and accelerate rDMCs to differentiate into cementoblast and osteoblast, and the mineralization of cementum matrix and bone matrix. TGF-β1+BMP4 can markedly increase the expressions of DSPP/OPN, and quicken rDMCs to differentiate into odontoblast and osteoblast, and the mineralization of dentinal matrix and bone matrix which display osteogenesis trend. Combined use of four factors had no significant synergism.