The Expression Of Tissue Factor Pathway Inhibitor-2Gene Effects On Cell Proliferation SHI-1

[Objective]In this study, we transfected well constructed eukaryotic expression vectorFEGFP-N1-TFPI-2into acute mononuclear leukemia SHI-1cell line (SHI-1), observe theTFPI-2gene expression in SHI-1cell, and the proliferation and growth of SHI-1cell.[Methods]1. Well constructed eukaryotic expression vector FEGFP-N1-TFPI-2gene, and usingJM109cells plasmid amplification, than by using the plasmid extraction technology forpurified eukaryotic expression FEGFP-N1-TFPI-2gene.2. Well constructed eukaryotic expression vector FEGFP-N1-TFPI-2gene and emptycarrier FEGFP-N1using lipofectamine2000mediated gene transfection technique,transfected them into acute mononuclear leukemia cell lines SHI-1. SHI-1cell successfultransfected eukaryotic expression vector FEGFP-N1-TFPI-2gene named SHI-1-TFPI-2group, as experimental group. SHI-1cell and transfect of empty carrier FEGFP-N1cells ascontrol groups, divided into SHI-1-P group and SHI-1-V group.3. Western blot analyzed the TFPI-2protein expression in the three groups.4. Compare the average growth rate of cells in the experimental group and two controlgroups every day by cell count method. The growth curve was depicted by the averagequantity of three groups′cells which were in counted every24hours.5. To detect cell proliferation, analyzed for three groups′cells relative growth rate of24,48,72hours by MTT colorimetry.[Results]1. The purpose of plasmid containing antibiotic resistance genes after the conversion,and thus can survive on LB culture medium containing antibiotics, and form a single colony.2. Western Blot results: SHI-1-TFPI-2groups can see TFPI-2protein expression, but notexpress in the other two groups. 3. Using the cell count technology count three groups cells in seven days, counting toSHI-1-TFPI-2, SHI-1-V, SHI-1-P groups average growth rate were0.6306,0.8348,0.8760,suggesting-TFPI-2-SHI-1average cell proliferation at a slower pace, difference betweenexperimental group and control groups had statistical significance (P<0.05). Seeing from thecell growth curve, the SHI-1-TFPI-2group was flatter than those of the other two controlgroups.4. Determined by MTT colorimetric method to compare SHI-1-P, SHI-1-V, SHI-1-TFPI-2relative growth rate (%) of three groups of cells, cell relative growth rate (%) ofvisible24hours divided into (97±16.1), respectively (91.8±10.0),(92.5±9.8); cell relativegrowth rate (%) of visible48hours divided into (97±14.6),(96.9±9.3),(87.3±10.2); cellrelative growth rate (%) of visible72hours divided into (97±11.5),(94.4±10),(60.7±12.9).Using SPSS13.0analysis of variance,24hours later TFPI-2-SHI-1, SHI-1-P, SHI-1-V grouprelative growth rate was decreased, but the difference was no statistically significant (P>0.05).48hours and72hours later TFPI-2-SHI-1, SHI-1-P, SHI-1-V group relative growth ratecomparison the difference was statistically significant (P<0.05). Show that in each timeperiod experimental cell relative growth rate of decline, with the passage of time thedifference was more obvious,48hours and72hours later the differences between groups wasstatistically significant (P <0.05).[Conclusion]1.Successfully constructed FEGFP-N1-TFPI-2-SHI-1cell line, and TFPI-2gene expressed stably in cells.2. TFPI-2gene expression can make SHI-1cell proliferate at a slower pace, and relativegrowth rate reduced in a time dependence;...