Isolation and Differentiation of Adipose-Derived Stem Cells into Odontoblast-Like Cells: A Preliminary In Vitro Study

(Pages: 288-293)
Saber Khazaei, D.D.S., M.Sc., Ph.D., 1,2Abbasali Khademi, M.Sc., 1,*Mohammad Hossein Nasr Esfahani, Ph.D., 3Mozafar Khazaei, Ph.D., 4,*Mohammad Hossein Nekoofar, D.D.S., M.Sc., Ph.D., 5Paul M. H. Dummer, B.D.S., M.Sc.D., Ph.D., D.D.Sc., F.D.S.R., 6
Department of Endodontics, School of Dentistry and Dental Research Centre, Dental Research Institute, Isfahan University of Medical Sciences, Isfahan, Iran
Department of Endodontics, School of Dentistry, Kermanshah University of Medical Sciences, Kermanshah, Iran
Department of Animal Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran
Fertility and Infertility Research Centre, Health Technology Institute, Kermanshah University of Medical Sciences, Kermanshah, Iran
Department of Endodontics, School of Dentistry, Tehran University of Medical Sciences, Tehran, Iran
School of Dentistry, College of Biomedical and Life Sciences, Cardiff University, Cardiff, UK
Department of Endodontics, School of Dentistry and Dental Research Centre, Dental Research Institute, Isfahan University of Medical Sciences, Isfahan, Iran
Department of Endodontics, School of Dentistry, Kermanshah University of Medical Sciences, Kermanshah, Iran
Department of Animal Biotechnology, Reproductive Biomedicine Research Center, Royan Institute for Biotechnology, ACECR, Isfahan, Iran
Fertility and Infertility Research Centre, Health Technology Institute, Kermanshah University of Medical Sciences, Kermanshah, Iran
Department of Endodontics, School of Dentistry, Tehran University of Medical Sciences, Tehran, Iran
School of Dentistry, College of Biomedical and Life Sciences, Cardiff University, Cardiff, UK
*Corresponding Addresses: P.O.Box: 8174673461 Department of Endodontics School of Dentistry and Dental Research Centre Dental Research Institute Isfahan University of Medical Sciences Isfahan Iran P.O.Box: 6714869914 Fertility and Infertility Research Centre Health Technology Institute Kermanshah University of Medical Sciences Kermanshah Iran Emails:a_khademi@dnt.mui.ac.ir,mkhazaei1345@yahoo.com
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Khazaei Saber, Khademi Abbasali, Nasr Esfahani Mohammad Hossein, Khazaei Mozafar, Nekoofar Mohammad Hossein, Dummer Paul M. H.. Isolation and Differentiation of Adipose-Derived Stem Cells into Odontoblast-Like Cells: A Preliminary In Vitro Study. Cell J. 2021; 23(3): 288-293.

Abstract

Objective

The aim of present study was to isolate and differentiate human adipose-derived stem cells (ASCs) into odontoblast-like cells.

Materials and Methods

In this experimental study, human adipose tissues were taken from the buccal fat pad of three individuals (mean age: 24.6 ± 2.1 years). The tissues were transferred to a laboratory in a sterile culture medium, divided into small pieces and digested by collagenase I (2 mg/mL, 60-90 minutes). ASCs were isolated by passing the cell suspension through cell strainers (70 and 40 µm), followed by incubation at 37ºC and 5% CO2in Dulbecco’s modified eagle medium (DMEM) supplemented with fetal bovine serum (FBS 5%) and penicillin/streptomycin (P/S). After three passages, the ASCs were harvested. Subsequently, flow cytometry and reverse transcriptase polymerase chain reaction (RT-PCR) were used to detect expression levels of NANOG and OCT4 to evaluate stemness. Then, a differentiation medium that included high-glucose DMEM supplemented with 10% FBS, dexamethasone (10 nM), sodium β-glycerophosphate (5 mM) and ascorbic acid (100 µM) was added. The cells were cultivated for four weeks, and the odontogenic medium was changed every two days. Cell differentiation was evaluated with Alizarin red staining and expressions of collagen I (COL1A1), dentin sialophosphoprotein (DSPP) and dentin matrix protein-1 (DMP1).

Results

The ASCs were effectively and easily isolated. They were negative for CD45 and positive for the CD105 and CD73 markers. The ASCs expressed OCT4 and NANOG. Differentiated cells highly expressed DSPP, COL1A1 and DMP1. Alizarin red staining revealed a positive reaction for calcium deposition.

Conclusion

ASCs were isolated successfully in high numbers from the buccal fat pad of human volunteers and were differentiated into odontoblast-like cells. These ASCs could be considered a new source of cells for use in regenerative endodontic treatments.