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Objective:The aim of the study was to construct the eukaryotic expression vector of human angiopoietin-like protein 4(ANGPTL4) and observe the effect of ANGPTL4 overexpression on the growth of esophageal carcinoma EC9706 cells.Methods:Total RNA was extracted from normal hepatic tissue,and ANGPTL4 cDNA was amplified by RT-PCR.The PCR product was doubly digested by XbaI and SalI,and then recombined into eukaryotic expression vector.Then,pIRES-GFP-ANGPTL4 was obtained by G418 selection,then pIRES-GFP-ANGPTL4 and pIRES-GFP were transfected into EC9706 cells with lipidosome-packaged method.Meanwhile,the transfected cells were selected by G418,and then stable transfected cell lines were obtained.ANGPTL4 mRNA levels,the cell cycles and growth curves of EC9706 cells in experiment group(transfected with pIRES-GFP-ANGPTL4),empty vector group(transfected with pIRES-GFP) and blank control group(EC9706 cells without transfection) were detected with RT-PCR,flow cytometry and MTT methods,respectively.Results:Eukaryotic ANGPTL4 expression vector pIRES-GFP-ANGPTL4 was successfully constructed.The ANGPTL4 mRNA level(0.21 ± 0.03) in experiment group was significantly higher than that of the empty vector group(0.04 ± 0.008) and the blank control group(0.05 ± 0.007),with significant differences(P < 0.01).The proportion of cells in S phase in experiment group was significantly different with those of the other two groups(P < 0.05).The cell growth of EC9706 cells in experiment group was slower than those of the other two groups.From the third day,the differences began to be significant.Conclusion:ANGPTL4 overexpression in esophageal carcinoma EC9706 cells could inhibit the growth of EC9706 cells.
Objective: The aim of the study was to construct the eukaryotic expression vector of human angiopoietin-like protein 4 (ANGPTL4) and observe the effect of ANGPTL4 overexpression on the growth of esophageal carcinoma EC9706 cells. Methods: Total RNA was extracted from normal hepatic tissue , and ANGPTL4 cDNA was amplified by RT-PCR. The PCR product was doubly digested by XbaI and SalI, and then recombined into eukaryotic expression vector.Then, pIRES-GFP-ANGPTL4 was obtained by G418 selection, then pIRES- GFP- ANGPTL4 and pIRES-GFP were transfected into EC9706 cells with lipidosome-packaged method. Meanwhile, the transfected cells were selected by G418, and then stably transfected cell lines were obtained .ANGPTL4 mRNA levels, the cell cycles and growth curves of EC9706 cells in experiment group ( transfected with pIRES-GFP-ANGPTL4), empty vector group (transfected with pIRES-GFP) and blank control group (EC9706 cells without transfection) were detected with RT-PCR, flow cytometry and MTT methods, respectively .Results: Eukaryotic ANGPTL4 expression vector pIRES-GFP-ANGPTL4 was successfully constructed. ANGPTL4 mRNA level (0.21 ± 0.03) in experiment group was significantly higher than that of the empty vector group (0.04 ± 0.008) and the blank control group (0.05 ± 0.007), with significant differences (P <0.01). The proportion of cells in S phase in experiment group was significantly different from those of the other two groups (P <0.05). The cell growth of EC9706 cells in experiment group was slower than those of the other two groups. From the third day, the differences began to be significant. Confrc: ANGPTL4 overexpression in esophageal carcinoma EC9706 cells could inhibit the growth of EC9706 cells.