Induction Of Cell Cycle G1 Phase Arrest By Epigallocatechin-3-gallate Via Activating PI3K-Akt Signaling Pathway In Human Gastric Cancer BGC-823 Cells

Objective: To elucidate the mechanism of cell cycle arrest induced by Epigallocatechin-3-gallate (EGCG) on human gastric cancer BGC-823 cells via PI3K-Akt pathway.Methods: The survival rate of BGC-823 cells was detected by MTT assay.The ability of grappling growth was tested through the number of colonies.Flow cytometry PI staining was used to test the change of cell cycle in BGC-823 which induced by EGCG. The expression of Akt,pAkt in BGC-823 cells were analyze by immunocytochemical method after treatment with EGCG. Western blot was used to analyze the change of proteins'expression such as Akt,pAkt,cyclinD1,PCNA in BGC-823 cells which were related with PI3K-Akt signal transduction pathway ,after EGCG treatment.Results:EGCG could significantly inhibited the growth of BGC-823 cells in a time- and dose- dependent manner. BGC-823 cells were treated with 25μg/mL,50μg/mL,100μg/mL EGCG for five days,the numbers of colonies were decreased in a dose- dependent manner . PI staining FCM detection showed that Insulin-like Growth Factor-1(IGF-1) as an activator of PI3K-Akt signal transduction pathway could significantly increase G1 phase arrest in BGC-823 cells; BGC-823 cells were exposed to EGCG (at various concentrations of 25μg/ml,50μg/ml,100μg/ml) for 48h or 50μg/ml EGCG for 12h,24h,48h with IGF-I 25μg/mL ,the cells of G1 were decreased, in a time- and dose- dependent manner. The specificity blocking agent of PI3K-Akt signal transduction pathway (LY294002) in a dose of 50μmol/L was used to test BGC-823 cells for 48h,it was also can induced the decreation of G1 cells;and the effection of LY294002 were stranger than EGCG 100μg/ml. The expression of phosphorylation of Akt was down-regulated in a time- and dose- dependent manner while the hole Akt was no changes by Immunocytochemical method after exposure to EGCG 25μg/ml,50μg/ml,100μg/ml for 48h and 50μg/ml for 12h,24h,48h.Western blot confirmed that BGC-823 cells were pre-treated by EGCG at various concentrations of 25μg/ml,50μg/ml,100μg/ml for 48h and 50μg/ml for 12h,24h,48h followed by a 30 min treatment with IGF-I 25μg/mL The expression of pAkt,cyclinD1,PCNA was down-regulated ,in a time- and dose- dependent manner .The results above indicated that EGCG could inhabit PI3K-Akt signal transduction pathway.Conclusion: EGCG can inhibit Proliferation and induced cell cycle G1 phase arrest of BGC-823 cells via activating PI3K-Akt signal transduction pathway.