Author/Authors :
Raoof, Maryam Department of Endodontics - Kerman Oral and Dental Diseases Research Center - Kerman University of Medical Science, Kerman , Yaghoobi, Mohammad Mehdi Department of Biotechnology - Institute of Science and High Technology and Environmental Sciences - Graduate University of Advanced Technology, Kerman , Derakhshani, Ali Department of Pathology - Stem Cell Research Center - Kerman University of Medical Sciences, Kerman , Kamal-abadi, Ali Mohammadi gradaute , Ebrahimi, Behnam Department of Biotechnology - Institute of Science and High Technology and Environmental Sciences - Graduate University of Advanced Technology, Kerman , Abbasnejad, Mehdi Department of Biology - Shahid Bahonar University of Kerman, Kerman , Shokouhinejad, Noushin Department of Endodontics - School of Dentistry/Dental Research Center - Tehran University of Medical Sciences - and Iranian Center for Endodontic Research, Tehran
Abstract :
Background: Dental pulp stem cells can be used in regenerative endodontic therapy. The aim of
this study was to introduce an efficient method for dental pulp stem cells isolation.
Materials and Methods: In this in-vitro study, 60 extracted human third molars were split and
pulp tissue was extracted. Dental pulp stem cells were isolated by the following three different
methods: (1) digestion of pulp by collagenase/dispase enzyme and culture of the released cells;
(2) outgrowth of the cells by culture of undigested pulp pieces; (3) digestion of pulp tissue pieces
and fixing them. The cells were cultured in minimum essential medium alpha modification (αMEM)
medium supplemented with 20% fetal bovine serum(FBS) in humid 37°C incubator with 5% CO2.
The markers of stem cells were studied by reverse transcriptase polymerase chain reaction
(PCR). The student t-test was used for comparing the means of independent groups. P <0.05 was
considered as significant.
Results: The results indicated that by the first method a few cell colonies with homogenous
morphology were detectable after 4 days, while in the outgrowth method more time was needed
(10-12 days) to allow sufficient numbers of heterogeneous phenotype stem cells to migrate out
of tissue. Interestingly, with the improved third method, we obtained stem cells successfully with
about 60% efficiency after 2 days. The results of RT-PCR suggested the expression of Nanog, Oct-4,
and Nucleostemin markers in the isolated cells from dental pulps.
Conclusion: This study proposes a new method with high efficacy to obtain dental pulp stem
cells in a short time.
Keywords :
Dental pulp stem cells , isolation , method , molar
