Optimizing a serum-free/xeno-free culture medium for culturing and promoting the proliferation of human dental pulp stem cells.

Background: Dental pulp stem cells (DPSCs) hold great promise for utilization in tissue repair and regenerative medicine. Routinely, culture media used for culturing stem cells are supplemented with animal serum for promoting growth and successful maintenance of stem cells. However, there is a growing demand for optimizing a well-defined culture media that could safely increase the efficacy and reproducibility of the cultured cells. In this study, we aimed at optimizing a serum-free/xeno-free culture medium.

Methods: A cocktail of various supplements intended to enrich DPSCs proliferation in defined concentrations was designed. It consisted of recombinant human basic fibroblast growth factor (hbFGF), insulin transferrin selenium (ITS), ascorbic acid (vitamin C), Beta mercaptoethanol and cholesterol. The effect of this optimized media on the proliferation of DPSCs was assessed by MTT assay and flow cytometric analysis (FACS) of early apoptotic marker annexin V. Expression of stemness-related genes (OCT4, SOX and NANOG) was assessed by qRT-PCR.

Results: Proliferation results by MTT illustrated a significant increase in the proliferation rate of DPSCs cultured in the proposed media. FACS analysis of annexin V expression was nil. Expression of OCT4, SOX and NANOG genes was also up-regulated.

Conclusions: The proposed combination of supplements utilized in the proposed culture media successfully increased the proliferation potential of DPSCs in addition to enhancing the stemness properties. Thus, it can be considered a promising and safe substitute to traditional animal derived supplements like fetal bovine serum (FBS).