Expression of multiple stem cell markers in dental pulp cells cultured in serum-free media.
BACKGROUND
Stem cell lines are usually grown in medium containing animal products. Fetal bovine serum (FBS) is an important additive for cell growth; however, the allergenic potential and the possibility of contamination when we use a medium containing serum would be a barrier to transplantation and consequently to the introduction of cell therapy methods into clinical applications.


METHODS
Dental mesenchymal cells were isolated and expanded in vitro and maintained in 4 different serum-free media (SFMs): SFM#1 (ITS-X, embryotrophic factor [ETF]); SFM#2 (ITS-X); SFM#3 (ETF); and SFM#4 (ETF, sodium pyruvate, ascorbic acid, fibroblast growth factor [FGF-a], acidic). Viability, proliferative, and immunocytochemical tests for the cells were performed by using 4 stem cell markers (CD44H, CK19, nestin, and P63) for ectoderm, mesoderm, and endoderm.


RESULTS
Viability tests showed a significant difference between the control and SFMs in both deciduous tooth pulp cells (DTPCs) and wisdom tooth pulp cells (WTPCs). However, all SFMs demonstrated 84%-90% viability, whereas the control showed 90%-93%. In both DTPCs and WTPCs, SFM#1 had the highest proliferation rate among the 4 SFMs. Immunocytochemistry stained positive stem cell markers most intensely in cells cultured with SFM#1. Furthermore, all stem cell markers for ectoderm, mesoderm, and endoderm were expressed in the cells cultured with SFM#1.


CONCLUSIONS
SFM#1 showed an acceptable survival rate, the highest proliferation rate, and the strongest expression of all the stem cell markers. SFM#1 proved to be a suitable medium for the culture of human dental pulp stem cells and to preserve pluripotency in differentiation.
Hirata
Thais Miyuki
TM
Department of Oral Health, Nippon Dental University, School of Life Dentistry at Tokyo, Tokyo, Japan.
Ishkitiev
Nikolay
N
Yaegaki
Ken
K
Calenic
Bogdan
B
Ishikawa
Hiroshi
H
Nakahara
Taka
T
Mitev
Vanyo
V
Tanaka
Tomoko
T
Haapasalo
Markus
M
D000818:Animals; D001205:Ascorbic Acid / Q000494:pharmacology; D015415:Biomarkers / Q000032:analysis; D002417:Cattle; D002452:Cell Count; D018929:Cell Culture Techniques; D049109:Cell Proliferation / Q000187:drug effects; D002470:Cell Survival / Q000187:drug effects; D003470:Culture Media; D016895:Culture Media, Serum-Free; D003782:Dental Pulp / Q000166:cytology* / Q000187:drug effects; D004475:Ectoderm / Q000166:cytology; D004707:Endoderm / Q000166:cytology; D016220:Fibroblast Growth Factor 1 / Q000494:pharmacology; D006801:Humans; D018960:Hyaluronan Receptors / Q000032:analysis; D007328:Insulin / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology; D007381:Intermediate Filament Proteins / Q000032:analysis; D053539:Keratin-19 / Q000032:analysis; D059630:Mesenchymal Stem Cells / Q000187:drug effects / Q000502:physiology*; D008648:Mesoderm / Q000166:cytology; D008964:Molar, Third / Q000166:cytology; D009419:Nerve Tissue Proteins / Q000032:analysis; D064231:Nestin; D019289:Pyruvic Acid / Q000494:pharmacology; D012643:Selenium / Q000494:pharmacology; D014094:Tooth, Deciduous / Q000166:cytology; D015534:Trans-Activators / Q000032:analysis; D014157:Transcription Factors; D014168:Transferrin / Q000494:pharmacology; D025521:Tumor Suppressor Proteins / Q000032:analysis
D015415:Biomarkers; C497483:CD44 protein, human; D003470:Culture Media; D016895:Culture Media, Serum-Free; D018960:Hyaluronan Receptors; D007328:Insulin; D036341:Intercellular Signaling Peptides and Proteins; D007381:Intermediate Filament Proteins; D053539:Keratin-19; C577484:NES protein, human; D009419:Nerve Tissue Proteins; D064231:Nestin; C115051:TP63 protein, human; D015534:Trans-Activators; D014157:Transcription Factors; D014168:Transferrin; D025521:Tumor Suppressor Proteins; D016220:Fibroblast Growth Factor 1; D019289:Pyruvic Acid; D012643:Selenium; D001205:Ascorbic Acid