ABSTRACT
Objective:Mineral trioxide aggregate (MTA),a wildly used pulp capping material,could affect the proliferation and differentiation of dental pulp cells.The aim of this study is to study the roles of Notch signalling and autophagy in MTA induced human dental pulp cells (hDPCs) proliferation promotions.Methods:Healthy human third molars were collected and hDPCs were isolated by a combined digestion of collagenase Ⅰ and dispase Ⅱ.MTA extracts of different concentrations (0.5,1.0,2.0,5.0,10.0 mg/mL) were used to test the cytotoxicity by cells counting kit (CCK-8) assays and to select the optimum concentration for hDPCs survival..Expressions of Notch1,Hes1,LC3Ⅱ / Ⅰ and p62 in wild type and MTA treated hDPCs were detected by western blotting.Results:MTA extracts in a concentration of 1.0 mg/mL exerted most profoundly promotion effects on the proliferation of hDPCs among all concentrations tested.MTA of high concentration (10 mg/mL) was toxically to cells.Compared with that of wild type hDPCs,the expressions of Notch1 and Hes1(P＜0.05),or p62 and LC3Ⅱ/Ⅰ (P＜0.01) in MTA treated hDPCs were significantly increased.Much lower expression of Notch1 was detected in hDPCs when autophagy was induced by Earle's balanced salt solution (EBSS) starvation for 24 h.Conclusions:MTA could up-regulated hDPCs proliferation with highly relevant in stimulating Notch1-Hes1 signalling and inhibition of autophagy.The study is supposed to provide new insight in unrevealing the mechanisms of MTA mediated dental pulp cells proliferation.
ABSTRACT
Objective@# The aim of this study is to investigate the roles of Notch signaling and autophagy on mineral trioxide aggregate (MTA) induced differentiation of human dental pulp cells (hDPCs). @*Methods @#Third molars from healthy human were collected and hDPCs were isolated by a combined digestion of collagenase Ⅰ and dispaseⅡ. Real time PCR were used to test the mRNA expression levels of alkaline phosphatase (ALP), runt-related transcription factor 2 (Runx2) and dentin sialophoprotein (DSPP) in MTA treated hDPCs in different time (24 h, 3 d and 7 d). The mineralization nodules formed by hDPCs with or without MTA treatment were detected by Von Kossa staining. Expressions of Notch1, Jagged1, Hes1, LC3Ⅱ/LC3 Ⅰand p62 in wild type and MTA treated hDPCs were detected by western blotting.@*Results@#MTA extracted in a concentration of 0.1 mg/mL could promote the differentiation of hDPCs. Compared with that of wild type hDPCs, the expressions of Notch1, Hes1, or Jagged1 and p62 (P<0.01) in MTA treated hDPCs were significantly increased. MTA treatment showed inhibition effects on autophagy flux similar to Bafilomycin A1, a specific inhibitor of fusion between autophagosomes and lysosomes. @*Conclusion@#MTA could promote hDPCs differentiation with highly relevant in stimulating Notch1-Jagged1-Hes1 signaling and inhibition of autophagy flux.