Effect of Concentrated Growth Factor on the Regenerative Potential of Dental Pulp Stem Cells Cultured Under Inflammatory Conditions

Abstract

Background: The concentrated growth factors (CGF) has been illustrated to be a promising factor that induces the regenerative features in Human dental pulp stem cells (DPSCs) through short-term chronic inflammation condition. However, the long-term therapeutic effects of CGF on the regenerative and osteogenic properties of DPSCs have not been elucidated. Then, this study was aimed to investigate the effects of long-term exposure to CGF on DPSCs, either alone or in presence of lipopolysaccharide (LPS) as pro-inflammatory agent. Methods: DPSCs were cultivated with CGF, LPS (0.1, 1 and 10 µg / ml) and the combination CGF of LPS (1 µg / ml) for 21 days. Consequently, MTT and scratch assays were performed to evaluate the cell viability and migration. Also, ALP staining and Sirius red staining were employed to investigate the osteogenic stimulation in treatment groups by evaluating ALP activity and collagen levels, respectively. The expression levels DMP-1 (dentin matrix protein 1)،DSPP (dentin sialophosphoprotein)، OPN (osteopontin)، Runx2 (Runt-related transcription factor 2) and OCN (osteocalcin) were quantified using qRT-PCR method. Data were analyised by ANOVA (one-way analysis of variance) and Tukey test due to normal distribution and Significance level was set as less than 0.05. Statistical analysis was performed by GraphPad Prism ver9.2.0 software.

Results: The obtained results showed that exposure of DPSCs to CGF, in long-term condition, could reduce the cell death induced by LPS. Besides, CGF and LPS (1 µg / ml), separately or in combination, improved the migratory ability of DPSCs and led to a significant increase in osteogenic stimulation through upregulation of ALP activity and collagen levels. Furthermore, CGF treatment significantly upregulated the expression of levels of RUNX2, DSPP, OCN and OPN osteogenic markers while LPS 1 µg / ml only led to OCN overexpression. Conclusion: Taken together, the findings of this study illustrated that CGF, separately or combined with LPS, could be an effective therapeutic agent to induce dentin–pulp complex healing through long-term chronic inflammation.