EMT Marker Expression In Gastric Cancer Cell Line With Its Invasion、VM Formation And Chemotherapy Sensitivity In Vitro Study

Objective:In the first part of this study, Understand the ability of migration and invasive of three gastric cancer cell line MGC803, BGC823, SGC7901 through scarification, transwell experiment, then investigating the relationship between EMT, VM and gastric cancer invasion according to this theory and whether there is a correlation between the two.The second part of experiment is aim to inquire whethe EMT, VM is associate with chemotherapy sensitivity. Methods:1、Delineating the area, After 12 h,Observating the number of three groups gastric cancer cells moving into the area,understanding the migration of the gastric cancer cell line.2、Inoculating the concentration of 2×105/ml of each cell suspension 30 ul on the Transwell upper room pore. After 40 h,Counting the number of cells through the DAPI staining which pass through the membrane, understanding the invasion of the gastric cancer cell line.3、Testing EMT markers: E-cadherin, Vimenin expression difference in MGC803, BGC823, SGC7901 through the Western-blot experiment, so as to assessment of EMT difference.4、Inoculating the concentration of 5×105/ml of each cell suspension 1ml in 300 ul Matrigel for three-dimensional culture. After 12 h,Observating whether the formation of VM pipeline, then randomly selecting five field from microscopy to counte VM. Statistical analysis to understand the difference of three groups of gastric cancer cells.Then combine with the experiment of scratch test,Transwell and western-blot in order to investigate the relationship between EMT, VM and gastric cancer invasion through Western-blot、TDCC and whether there is a correlation between the two.5、1ug/ml, 5ug/ml, 10ug/ml docetaxel handle 1000 gastric cancer cell for 24 h, Determination OD after the CCK-8 assay to understand the difference of the gastric cancer cell line chemotherapy-susceptibility,so as to inquire whethe EMT, VM is associate with gastric cancer resist to chemotherapy. Results:1、Scratch test showed that the migration of MGC803, BGC823, SGC7901 decreased in turn.2、Transwell experiment results showed that MGC803(72.25±2.75), BGC823(44.25±2.50), SGC7901(22.50±2.08), Pairwise comparison, the difference was statistically significant(P<0.05). the invasiness of MGC803, BGC823, SGC7901 decreased in turn.3、MGC803:E-cadherin no expression, the highest expression of Vimentin; SGC7901:the highest expression of E-cadherin, but Vimentin expression is the lowest. The degree of EMT of BGC823, SGC7901,MCG803 were decreased in turn.4、MGC803, BGC823, SGC7901 can form VM pipeline, MGC803(11.42 ±8.67), BGC823(102.40 ±8.41), SGC7901(91.00 ±6.82), Pairwise comparison, the difference was statistically significant(P<0.05). The ability of VM formation of MGC803, BGC823, SGC7901 decreased in turn.5、Docetaxel chemotherapy:OD show that MGC803:1ug/ml(0.714±0.005), 5ug / ml(0.464±0.008), 10 ug / ml(0.352±0.009); BGC823:1ug/ml(0.652±0.009), 5ug / ml(0.432±0.007), 10 ug / ml(0.326±0.003); SGC7901:1ug/ml(0.503±0.010), 5ug / ml(0.312±0.005), 10 ug / ml(0.226±0.003) value. Gastric cancer cells of each group and the OD value of docetaxel showed dose-effect relationship; under the same concentration of chemotherapy, the chemotherapy sensitivity of MGC803、BGC823、SGC7901 rise in turn, and the difference was statistically significant(P < 0.05). Conclusion:1、The ability of migration and invasion of MGC803, BGC823, SGC7901 were decreased in turn, EMT, VM are positive correlation with gastric tumor invasion,and Gastric cancer cells which have higher degree of EMT can form VM easily.2 、 The occurrence of EMT and The formation of VM can weak the chemotherapy- sensitivity of gastric cancer.