The Effect Of KGF On Apoptosis And Proliferation Of Human Oral Mucosal Peithelial Cells

Objective:Oral mucosal diseases are closely connected to the apoptosis, Recently studies have found that keratinocyte growth factor (KGF) could not promote the proliferation and differentiation of epithelial cells but inhibit the apoptosis of epithelial cells. Our preliminary studies found that the expressions of KGF and KGF mRNA in oral lichen planus were lower than tose in normal mucosa, and the mRNA and protein expressions of KGF in oral leukoplakia were higher than those in normal mucosa. Based on the previous researches, we speculate that KGF may have an effect on the proliferation and apoptosis of oral epithelial cells.To detect the effects of KGF on apoptosis and proliferation of human oral mucosal epithelial cell, Cell morphology was observed respectively under different concentrations of KGF in vitro, flow cytometry was used to detect the expression of the genes related to the apoptosis and proliferation,This study may provide the basis for further investigation of the role of KGF in the occurrence and development of oral mucosal diseases.Methods:1. Selected5～10th generation cells; when cultured for70～80%, added the D-KFSM with different concentrations of KGF such as0ng/ml(control group),5ng/ml(experimental group1),25ng/ml(experimental group2),50 ng/ml(experimental group3). The morphology of oral mucosal cells co-cultured was observed after12h,24h,48h respectively;2. The apoptosis rate of cells in different groups was detected by flow cytometry;3. The mRNA expressions of apoptosis related genes Bcl-2and Bax of cells co-cultured with different concentrations of KGF were detected by Real-time RT-PCR;4. The mRNA expressions of gene PCNA related to proliferation of cells co-cultured with different concentrations of KGF were detected by Real-time RT-PCR;5. Statistical methods were applied to analysis the mRNA expression of Bcl-2,Bax and PCNA.Results:1. After co-cultured with different concentrations of KGF, the cells in experimental groups were adherent more tightly and with enhanced shading than cells in control group at the same time. Cells in experimental group3showed obvious nuclei at48h;2. The apoptosis rate was decreased with the increase of the concentrations of KGF after48h(P<0.05);3. Results of quantitative real-time PCR of apoptosis showed that:(1) there was no difference of Bcl-2mRNA expression between experimental group1and2and control group at12h, Bcl-2mRNA expressions of experimental group3was higher than control group (P<0.05) at12h;(2) Bcl-2mRNA expressions of all experimental groups were higher than control group (P<0.05) at24h, but there was no difference among experimental groups (P>0.05);(3) Bcl-2mRNA expressions of all experimental groups were higher than control group (P<0.05) and the increase was concentration-dependent at48h;(4) There was no difference of Bax mRNA expression between experimental group1and2and control group (P>0.05) at12h, the Bax mRNA expressions of experimental group3was decreased compared with control group; Bax mRNA expression of experimental groups were decreased compared with control group (P<0.05) at24h and48h;4. Results of quantitative real-time PCR of proliferation showed that:PCNA mRNA expressions of experimental groups were higher than control group(P<0.05)(1) PCNA mRNA expressions of experimental groups were decreased with concentration of KGF at12;(2) at24h, while there was no difference among experimental groups (P>0.05);(3) PCNA mRNA expressions of experimental groups were increased with concentration of KGF (P<0.05) at48h.Conclusion:The study found that the apoptosis rates of epithelial cells decreased with the elevation of concentration of KGF. Exogenous KGF could increase the mRNA expression of Bcl-2and PCNA while decrease the mRNA expression of Bax in epithelial cells., indicating that KGF may inhibit epithelial cell apoptosis and promote epithelial cell proliferation through regulating gene expressions of Bcl-2, Bax and PCNA. The results provide a basis for further investigation of the role of KGF in the occurrence and development of oral mucosal diseases.