Abstract:

Objective

To investigate the effect of Ad5-Oct4-EGFP transfection on the proliferation and multilineage differentiation capabilities of human dental pulp cells（DPCs）.

Methods

The recombinant adenovirus was constructed by Admax system which carried transcription factor and enhanced green fluorescence protein enhanced green fluorescent protein （EGFP）， and the optimal multiplicity of infection（MOI） was selected. After transfection， CCK-8 was used to examine the proliferation of transfected cells at days 1， 2， 3， 5， 7 and 14. Transfected DPCs were cultured in the odontogenic and adipogenic induction media for 21 days respectively. Odontogenic and adipogenic differentiation was investigated by Alizarin Red staining and Oil Red O staining， and the odontogenic and adipogenic differentiation markers were evaluated by RT-PCR.

Results

The recombinant adenoviral vector of Oct4 gene was successfully constructed and the MOI value of 200 was selected as the optimal MOI for the further study. The CCK-8 test revealed that the OD value in Ad5-Oct4-EGFP transfected cells was higher than that in the control groups at day 2， with significant increase at day 7 and day 14 （P＜0.05）. The results of Alizarin Red staining indicated that Oct4 gene transfection promoted more calcified nodules formation in Ad5-Oct4-EGFP group. DSPP， BSP and collagen I mRNA expression levels were significantly greater in Ad5-Oct4-EGFP transinfected cells than those of the control groups at days 21 （P＜0.05）. Adipogenic differentiation was demonstrated by the large accumulation of neutral lipid vacuoles indicated by the Oil Red O stain in Ad5-Oct4-EGFP group， and further confirmed by the significant up-regulation of LPL and PPARγ2 mRNAs compared with the control groups （P＜0.05）.

Conclusion

Our study has demonstrated Oct4 can improve the proliferation and multilineage differentiation capablity of cultured DPCs.

Key words: Dental pulp cells, Transcription factor Oct4, Proliferation, Multilineage differentiation