Experiment Studies On Expression Of 15-LOX-1 Gene In Human Gastric Cancer AGS Cells

Objective : The study was to investigate the role of 15-lipoxygenase-1(15-LOX-1) in regulating proliferation and apoptosis of human gastric cancer AGS cells.Methods :AGS cells were divided into three groups: AGS/15-LOX-1 group transfected with pcDNA3.1(+)/15-LOX-1,vector control group transfected with pcDNA3.1(+) and blank control group without transfection.Plasmids of pcDNA3.1(+)/15-LOX-1 encoding 15-LOX-1 gene were transiently transfected into AGS cells.The expression of 15-LOX-1 at mRNA and protein levels was measured by reverse transcription polymerase chain reaction(RT-PCR) and western blot. Then light microscopy was used to measure the effects of 15-LOX-1 expression on cell morphology. The proliferation,cell cycle and apoptosis of AGS cells were assessed by thiazolyl blue tetrazolium bromide(MTT) assay,flow cytometry ,AnnexinV/PI double staining and TdT-mediated dUTP Nick-End Labeling (TUNEL),respectively.Results: 15-LOX-1 mRNA and protein were only expressed in AGS/15-LOX-1 cells but not expressed in AGS and AGS /pcDNA3.1 cells detected by RT-PCR and western blot. Morphological changes of AGS cells which were transfected with pcDNA3.1(+)/15-LOX-1 were obvious compared with control group under inverted microscope There was a significant decline in cell proliferation ability of AGS/15-LOX-1 in comparision with control groups measured by MTT assay.Cell survival rates were 72.07%,64.53% and 43.07%,respectively,48 , 72 and 96h after transfection.Also,flow cytometry analysis revealed that the number of AGS/15-LOX-1cells in G0/G1 phrase increased markedly from 38.21%±4.73% to 54.54%± 3.21%,while the S phrase decreased from 41.93%±5.47% to 24.26%±1.88%(p<0.05). AnnexinV/PI and TUNEL confirmed that 15-LOX-1 could induce apoptosis of AGS cells,showing a statistically significant differences in comparision with control groups(p<0.01).Conclusion:The expression of 15-LOX-1 can strongly suppress the proliferation and induce apoptosis in human gastric cancer AGS cells,which provides the experimental evidence for investigating the mechanism and treatment of gastric cancer.