Abstract
Objective: This study aimed to isolate the cells from the dental pulp tissue of human primary teeth, study the capacity of proliferation, characterize the cells and standardize the technique of culture and expansion to create a cell banking. Material and Methods: Primary teeth with no caries and orthodontic reasons were extracted for pulp tissue obtainment. The cells were extracted from the pulp cells, isolated and cultured under ideal conditions until full expansion. Results: After consecutive passages, the cultured cells were characterized using immunofluorescence technique and frozen between the 2nd and 6th passage, thus creating a biorepository of dental pulp cells from human primary teeth. Conclusion: The creation of a cell banking from dental pulp cells from human primary teeth enables the easy application of cells in laboratorial studies, reducing the cost and time for obtaining the samples, avoid the involvement of new subjects and allow a fast reproducibility of the researches. KeywordsCell culture techniques; Tooth, deciduous; Pulp; Fibroblasts; Cryopreservation.
Highlights
The development of cell cultures has been proposed since the 50s to evaluate microscopically normal cell events
The collected pulp tissue was placed into a Petri plate (100 mm in diameter x10 mm height), cut at small pieces with the aid of 15c scalpel blade (Figure 1A), and immersed in the supplemented medium containing Dulbecco’s modified Eagle’s medium (DMEM) 20% Fetal Bovine Serum (FBS) supplemented with 600 μL of penicillin (Gibco, Thermo Fisher Scientific in Waltham, MA, USA), 300μL of gentamicin (Gibco, Thermo Fisher Scientific in Waltham, MA, USA), and 100 μL of amphotericin B (Gibco, Thermo Fisher Scientific in Waltham, MA, USA) (Figure1B, 1C and 1D), and incubated at 37oC and 5% CO2 for 40 minutes (Figure 1E)
The pulp tissue obtained from the extracted primary teeth allowed the primary cell culture
Summary
The development of cell cultures has been proposed since the 50s to evaluate microscopically normal cell events. This methodology is largely explored in researches on many knowledge areas because the obtained results are very close or equal to those observed in living organisms [12]. Recent researches demonstrate a great potential of the pulp tissue from exfoliating primary teeth because this tissue is rich in undifferentiated mesenchymal cells, which raise great scientific interest to evaluate the bioactive potential and possible clinical applications [811]. The easy access to pulp tissue, obtained from a non-vital organ that is normally discarded after extraction, are very attractive in research. Notwithstanding, limitations in obtaining, culturing and controlling of the cell proliferation emphasize the search for new sources, techniques, and applications [7,12,13]
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