ABSTRACT
Objective:To investigate the proliferation,odontogenic differentiation and mineralization of human dental pulp cells (HDPCs) on bioactive glass(BG) and extracted dentin proteins(EDP).Me-thods: Primary HDPCs were isolated from third molars by enzyme digestion and were cultured in Dulbecco's minimum essential medium (DMEM).Then the 4th generation of HDPCs was cultured with DMEM,which contained BG-EDP,BG,and EDP,respectively.Meanwhile HDPCs were cultured in DMEM as control group.Proliferation of HDPCs was evaluated by 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2-H-tetrazolium bromide(MTT) colorimetric assay.Odontogenic differentiation was determined by alkaline phosphatase (ALP) activity assay and real-time PCR.Mineralization was investigated by Alizarin red staining and cetylpyridinium chloride (CPC) assay.Results: The proliferation of HDPCs was increased significantly in BG-EDP group on 3,7,and 9 d(optical density value:1.36±0.06,2.52±0.20,2.72±0.29) compared with BG(optical density value: 1.20±0.26,2.33±0.26,2.50±0.30),EDP(optical density value: 1.13±0.15,2.10±0.13,2.38±0.22) and control group(optical density va-lue: 0.84±0.17,1.84±0.18,1.95±0.19),P0.05).After 14 days,ALP activity of BG-EDP group (56.67±1.83) was significantly upregulated compared with EDP group (41.98±9.71) and control group (30.82±6.70),P0.05;DSPP gene expression was upregulated significantly in BG-EDP group (5.79±1.94) compared with the other groups (P0.05).The alizarin red staining showed more mineral nodules in BG-EDP group,the cetylpyridinium chloride semi-quantification presented higher calcification in BG-EDP group (0.27±0.01) compared with the other groups (P<0.05).Conclusion: Compared with either BG or EDP,BG-EDP significantly promotes the proliferation,odontogenic differentiation and mineralization of HDPCs.
ABSTRACT
Objective:Positive effects of bioactive glass (BG) on proliferation,mineralization,and differentiation of human dental pulp cells (hDPCs) was already verified in various former studies.The Arg-Gly-Asp-Ser sequence (RGDS) was confirmed of affecting cell adhesion.Before further investigation,the objective of this study is to investigate whether RGDS can affect the effects of BG on the adhesion,proliferation and mineralization of hDPCs.Methods: hDPCs were harvested from third molars of 18-25-year-old individuals after informed consent.Enzyme digestion technique was used.The 4th to 6th ge-neration of hDPCs were used for all experiments.The cells of the experimental groups were cultured in Dulbecco minimum essential medium (DMEM) containing ionic dissolution products of BG and RGDS of seve-ral concentrations (12.5 mg/L,25.0 mg/L,50.0 mg/L,100.0 mg/L,200.0 mg/L).DMEM containing ionic dissolution products of BG without RGDS was used for cell culture as control group.Cell adhesion was tested 4 h after cell seeding by MTT assay.Cell proliferation was examined at 1,3,5,7,and 9 d after cell seeding by MTT assay.Cell mineralization was investigated on days 14 and 28 by alizarin red staining.After being stained and dried,mineralized nodules were dissolved by cetylpyridinium chloride (CPC) for semi-quantitative test.Results were statistically analyzed by one way ANOVA,SPSS (version 19.0) and P<0.05 was considered to be significant.Results: Cell adhesion in BG group showed no difference from that in DMEM group.Compared with BG group,hDPCs in BG+RGDS groups suggested weaker cell adhesion.When the concentration of RGDS increased,the adhered cell number decreased.hDPCs cultured with BG and RGDS showed lower proliferation activity in the early stage,while no significant difference was observed after 3 d.BG group promoted the mineralization of hDPCs compared with positive control group,negative control group and RGDS group.No significant difference was observed between BG+RGDS group and BG group or between RGDS group and positive control group.Conclusion: BG promotes proliferation and mineralization without affecting cell adhesion of hDPCs.Unbounded RGDS inhibits cell adhesion,but has no influence on the positive effects of BG on the proliferation and mineralization of hDPCs.