Effects of activating and inhibiting Notch signaling pathway on epidermal stem cell proliferation and differentiation

Abstract

Objective To observe the effects of activating and inhibiting Notch signaling pathway on epidermal stem cell (ESC) proliferation and differentiation.Methods ESCs of SD rats were isolated,cultured and purified.Recombinant rat Jagged1/FC chimera (1000 μg/L),an activator of the Notch signaling pathway,and DAPT (16 μmol/L),an inhibitor of the Notch signaling system,were added into the culture medium respectively.The cells in the medium added with phosphate buffered saline were used as control group.After co-culture for 48 h,the proliferation of ESCs was observed.The morphological changes and clone counting in each group were determined after culture.Methyl thiazol tetrazolium (MTT) assay was used to calculate the growth curve.Integrinβ1 (CD29) was identified by flow cytometry.The expression level of cytokeratin (CK) 19 and CK10 in each group was detected by using immunohistochemistry.Results There was no significant difference in morphological changes.After culture for 5 days,the number of colony formations in Jaggedl/FC,DAPT and control groups was 45.7 ± 3.8,16.9 ± 2.1 and 33.4 ± 3.1 respectiely.There was significant difference in every two groups (P ＜ 0.05).MTT assay showed that activated Notch signaling significantly promoted the proliferation of the ESCs (P ＜0.05).The percentage of CD29 positive cells in Jaggedl/FC group was significantly higher than that in DAPT group and control group[(96.42±2.57)％ vs.(72.58 ±3.87)％ and (88.75 ±3.14)％,both P＜0.05].The expression of CK19 was significantly higher in Jaggedl/FC group than in control group (P ＜ 0.05).The expression of CK19 in DAPT group was significantly lower than in control group (P ＜0.05).On the other hand,the expression of CK10 was significantly lower in Jaggedl/FC group than in control group (P ＜0.05).The expression of CK10 in DAPT group was significantly higher than in control group (P ＜ 0.05).Conclusion Notch signaling pathway may promote the ESC proliferation and maintain the undifferentiated state.When the Notch signaling pathway is inhibited,ESCs have the tendency to differentiate into epithelial cells. Key words: Epidermal stem cell; Notch signaling; Proliferation; Differentiation