Abstract
BackgroundDental pulp stem cells (DPSCs) play an important role in tissue regeneration. This study compares the growth kinetics and characterization of third molar and first premolar human DPSCs.Material and MethodsDental pulp tissues were isolated from human first premolar and third molar teeth and were digested by treating them with collagenase type I. Single-cell suspensions from each dental pulp were seeded in T25 culture flasks and the media were replaced every 3 days until 70% confluence. The cells were enumerated to determine the population doubling time (PDT). Cells were characterized using flow cytometry, RT-PCR and osteogenic medium for differentiation of DPSCs. Karyotyping assay was also performed till passage 7th.ResultsThe DPSCs had spindle-shaped morphology. There was an increase in PDT in third molar DPSCs when compared to first premolar teeth. Positive expression of CD44, CD73, and CD90 and negative expression of CD34 and CD45 were illustrated. A normal karyotype was visible for all seven passages. The Alizarin red staining was positive for osteogenic induction of DPSCs.ConclusionsWhen DPSCs are needed, third molar teeth can be a good and convenient candidate for cell transplantation, yielding high number of cells with mesenchymal characteristics. They can be a source for further investigations in vitro and work on tissue engineering protocols. Key words:Stem cells, dental pulp, growth kinetics, characterization.
Highlights
Isolation of mesenchymal stem cells (MSCs) has been reported from bone marrow (BM) [1], adipose tissue [2], endometrium [3], periodontal ligament [4], and dental pulp [5]
The comparison of growth curve of first premolar teeth with different initial density of seeded cells of 6×104 and 11×104 cells per well demonstrated that the cell proliferation rate was significantly more in Dental pulp stem cells (DPSCs) isolated from human third molar teeth (Fig. 1D). -Morphology of DPSCs Ten days after expansion of DPSCs, both third molar and first premolar teeth showed a fibroblast‐like, elongated, spindle‐shaped morphology and adherent property under a convert microscope at all passages (Fig. 2). -Characterization by flow cytometry DPSCs isolated from both third molar and first premolar teeth were positive for expression of CD44 and CD90 and negative for CD34 expression (Fig. 3A). -Characterization by RT-PCR Using RT-PCR, it was shown that all cells till passage 7 expressed CD73
Our study showed that DPSCs isolated from both third molar and first premolar teeth were mesenchymal stem cells with spindle shape morphology, multipotency, and expression of mesenchymal and lack of hematopoietic stem cell markers [13]
Summary
Isolation of mesenchymal stem cells (MSCs) has been reported from bone marrow (BM) [1], adipose tissue [2], endometrium [3], periodontal ligament [4], and dental pulp [5]. The growth curve of DPSCs isolated from human third molar teeth while the initial density of seeded cells per well were 3×104 cells revealed that the PDT was 29.8 hours (Fig. 1A). The comparison of growth curve of first premolar teeth with different initial density of seeded cells of 6×104 (green line) and 11×104 (red line) cells per well demonstrated that the cell proliferation rate was significantly more in DPSCs isolated from human third molar teeth (Fig. 1D). -Karyotype analysis The chromosome number of DPSCs isolated from both third molar and first premolar teeth was 2n=46, containing 44 autosomal and 2 sex chromosomes in all passages (Fig. 4A, 4B, respectively). -Osteogenic induction Analysis of alizarin red S stained areas demonstrated the osteogenic potential of DPSCs isolated from both third molar and first premolar teeth at passage 7 (osteogenic medium: Fig. 5A,B). We illustrated that initial number of 6×104 cells per well was an adequate number for cell culture and ex-
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