Chemical preconditioning escalates chondrogenic activity in explant cultured human dental pulp stem cell study model for future temporomandibular joint regeneration

Abstract

Context: Human dental pulp stem cells (hDPSC) derived from dental pulp in conducive environment activated by chemicals can enhance chondrogenic cells for future animal model temporomandibular joint model. Aim: The study aims at evaluating the chemicals preconditioning (curcumin and rapamycin) efficacy toward chondrogenic proliferation of human dental pulp stem cells. Settings and Design: The in vitro study model with 10 premolar teeth extirpated pulp was processed under sterile chemical conditions. The cells viability was checked with calorimetric assay for adipogenic and chondrogenic, osteogenic lineages. The viability of the cells and the concentration of curcumin (CU) and rapamycin (RP) required for cell differentiation toward chondrogenic lineage were assessed. Material and Methods: The hDPSC was evaluated after explant long-term cultivation with characterization and chemical conditioning with dimethyl sulfoxide (DMSO) as control. MTT assay was used for cytotoxicity evaluation, cell viability, and proliferation. The dose optimization was observed with RP and CU. Chondrogenic proliferation was assessed with standard staining method of 0.1% Safranin O and 0.1% Alcian blue. Statistical Design: The flow cytometry analysis revealed good results for CD 90 compared to others. The intergroup analysis was done by ANOVA, and intragroup analysis was done by Post hoc Tukey’s test. The intragroup analysis showed P value < 0.05 for RP in comparison between the various preconditioning agents CU and RP. The dosage of 10 µg/ml RP was considered statistically significant. Results: The flow cytometer analysis revealed good results for CD 90 compared to other surface markers. The dosage of 10 µg/ml RP was having good chondrogenic cell proliferation. The intragroup analysis showed P value < 0.05 for RP in comparison between the various preconditioning agents CU and RP. The calorimetric assay (MTT) quantitative analysis of the chondrogenic cells with Safranin O stain the standard deviation (SD = 0.017 for rapamycin), Alcian blue (SD = 0.49 for RP) in comparison to DMSO (control) and CU. Conclusion: RP activates mTOR pathway and hence stabilizes the stem cell maintenance of human dental pulp stem cell and the dose quantified can be used for future animal temporomandibular joint animal model.