The Effects Of Akt1 And Akt2 Gene Transfection On Growth And The Vitro Invasion Ability Of Human Gastric Epithelial Cell GES-1

Objective The eukaryotic expression plasmid vectors of p-LXSN-Aktl and p-LXSN-Akt2 were used to transfect human gastric epithelial GES-1 cells, to observe the effects of Aktl and Akt2 genes transfection on the growth and invasion of human gastric epithelial GES-1 cells.Methods The plasmids of p-LXSN-Aktl which contained the whole Aktl gene sequence (Aktl group) and p-LXSN-Akt2 which contained the whole Akt2 gene sequence (Akt2 group) were used to transfect human gastric epithelial GES-1 cells with lipofectamine, the GES-1 cells which were transfected by the empty vector p-LXSN were used as the blank vector group, and the no transfected ones were used as the control group. Then the positive cell clones were selected with antibiotics G418. The protein expressions of Akt1, Akt2, CyclinDl and Mmp-2 in the GES-1 cells were measured by Western blotting analysis; MTT method was applied to detect the proliferation activity of GES-1 cells; the effect of Aktl and Akt2 genes transfection on the cells cycle of the GES-1 cells was tested by flow cytometry (FCM). Transwell assay was used to analyze the invasion and metastasis capability. The expression of F-actin was detected by immunofluorescence.Results Compared with the blank vector group and control group, the relative expressions of CyclinDl protein and Mmp-2 in Aktl group were significantly increased (P<0.01); Compared with the blank vector group and control group, the relative expressions of Mmp-2 protein in Akt2 group were significantly increased (P<0.01), the relative expressions of CyclinD1 protein were no significant differences (P>0.05);②MTT method was applied to detect the proliferation activity of GES-1 cells. Compared with the blank vector group and the control group, the absorbance (A490) values of Aktl group in the first 1,2 days were no significant differences, but there were statistically significant differences (all P<0.01) in the first 3 to 6 days; the absorbance (A490) values of Akt2 group in the first 1 to 6 days were no significant differences;③Compared with the blank vector group and control group, S phase cells in Aktl group were increased by approximately 20.00% and 21.10%, G2 phase cells were reduced by approximately 20.80% and 15.48%(all P<0.01). S phase cells in Akt2 group were no significant differences (P>0.05).④Transwell assay showed that the invasive ability were significantly increased in Aktl and Akt2 groups, compared with the blank vector group and control group(P<0.01).⑤The expression of F-actin was increased correspondingly in Aktl and Akt2 groups detected by immunofluorescence.Conclusion①Cell models which high expressed Aktl gene (Aktl group) and high expressed Akt2 gene (Akt2 group) were successfully constructed.②Akt1 gene transfection could increase the proportion of S-phase cells and enhance the proliferation ability of the GES-1 cells, but the effects of Akt2 gene was not obvious.③Akt1 and Akt2 gene could increase cell invasion and migration, leading to malignant transformation of GES-1 cells.