[Recombinant human transforming growth factor ß1 promotes dental pulp stem cells proliferation and mineralization].
Beijing Da Xue Xue Bao Yi Xue Ban
; 49(4): 680-684, 2017 08 18.
Article
em Zh
| MEDLINE
| ID: mdl-28816288
ABSTRACT
OBJECTIVE:
To explore suitable concentration of recombinant human transforming growth factor ß1 (rhTGF-ß1) usage and study the effect of rhTGF-ß1 on differentiation of dental pulp stem cells (DPSCs).METHODS:
DPSCs were isolated from the undiseased third molars of people aged 18-25 years and cultured according to instructions in vitro. Different concentrations (1 , 6 , 10 µg/L) of rhTGF-ß1 were added to the culture medium to examine DPSCs proliferation by CCK-8 (cell counting kit-8) assay.The suitable concentration was then selected. For differentiation, the DPSCs were incubated for 7 or 14 days with rhTGF-ß1 supplemented with osteo/odontoblastic induction medium containing 10 nmol/L dexamethasone, 10 mmol/L b-glycerophosphate, 50 g/L ascorbate phosphate, 10 nmol/L 1,25-dihydroxyvitamin D3 and 10% fetal bovine serum. The cells were then washed 3 times with phosphate-buffered saline and sonicated with 1%Triton X-100 for 30 minutes on ice. Cellular alkaline phosphatase (ALP) activity was assayed with p-nitrophenyl phosphate as the substrate. The enzyme activity was expressed as p-nitrophenyl produced per milligram of protein [bicinchoninic acid (BCA) protein assay kit]. To examine mineral nodule formation, the cultured cells were fixed in 4% paraformaldehyde and washed in water, and the mineralization of the extracellular matrix was assayed by 1% alizarin red S staining and elution of staining was examined as optical density (D) under microplate reader. The mean difference was considered significant at 0.05 and 95% confidence interval.RESULTS:
The DPSCs had typical fibroblast morphology and could form mineral nodules after being cultured with osteo/odontoblstic induction medium for 14 days. 6 µg/L rhTGF-ß1 significantly promoted the DPSCs proliferation on the 3rd and 5th days. After the incubation of osteo/odontoblastic induction medium, the DPSCs with the 6 µg/L rhTGF-ß1 increased ALP activities compared with the control; D values in the 6 µg/L rhTGF-ß1 group was 0.31±0.03, while the control group was 0.02±0.01 (P<0.05). The total protein content in the 6 µg/L rhTGF-ß1 group was (2 775.46±83.54) mg/L, and the control group was (1 432.20±110.83) mg/L (P<0.05). To eliminate the cells proliferation influence, relative ALP activities, which was defined as the total ALP divided by the total protein content, the 6µg/L rhTGF-ß1 group was 6 times higher than the control group. Alizarin red S staining showed increased mineral nodule formation in the rhTGF-ß1 group. The elution of staining under microplate reader also showed more optical density in the 6 µg/L rhTGF-ß1-treated cells (0.83±0.02) than that in the control groups (0.55±0.05, P<0.05).CONCLUSION:
6 µg/L rhTGF-ß1 could significantly promote DPSCs proliferation and odontoblastic differentiation in vitro.
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Base de dados:
MEDLINE
Assunto principal:
Células-Tronco
/
Proteínas Recombinantes
/
Polpa Dentária
/
Proliferação de Células
/
Fator de Crescimento Transformador beta1
Limite:
Adolescent
/
Adult
/
Humans
Idioma:
Zh
Revista:
Beijing Da Xue Xue Bao Yi Xue Ban
Assunto da revista:
MEDICINA
Ano de publicação:
2017
Tipo de documento:
Article
País de afiliação:
China