| Part 1 The expression and correlation of CLIC1 and AMOT-p130 in gastric cancerObjective: To explore the expression and relationship of Chloride Intracellular Channel 1(CLIC1)and Angiomotin(AMOT)-p130 in gastric cancer.Methods: Immunohistochemistry was performed to analyze the expression of CLIC1 and AMOT-p130 in GC tissues and adjacent tissues.Western blot,q RT-PCR and immunofluorescence were used to analyze the expression of AMOT-p130 in gastric cancer cells after CLIC1 was knocked out.Results: 1.CLIC1 is mainly expressed in the cytoplasm and nucleus of gastric cancer tissues,while AMOT-p130 is mainly expressed in the cytoplasm of adjacent tissues.The expression of CLIC1 in gastric cancer tissue was higher than that in adjacent tissues(P<0.05),while the expression of AMOT-p130 in gastric cancer tissue was lower than that in adjacent tissues(P<0.05).2.The expression of CLIC1 was not significantly correlated with gender,age,depth of invasion,tumor differentiation,lymphatic metastasis,and tumor size(P>0.05)and it was correlated with vascular invasion and TNM pathological stage(P<0.05).The expression of AMOT-p130 was not significantly correlated with gender,age,tumor differentiation,lymphatic metastasis,and tumor size(P>0.05)and it was correlated with the depth of invasion and TNM pathological stage(P<0.05).3.The correlation between high expression of CLIC1 and low expression of AMOT-p130 was statistically significant(P < 0.05).4.In the q RT-PCR experiment,the m RNA expression of AMOT-p130 was significantly increased in MGC-803 CLIC1 KO and AGS CLIC1KO(P<0.05),but significantly decreased after CLIC1 was overexpressed(MGC-803 OE-CLIC1,AGS OE-CLIC1)(P<0.05).5.Western blot experiments showed that the protein expression of AMOT-p130 was increased in MGC-803 CLIC1 KO and AGS CLIC1KO(P<0.05),but significantly decreased after CLIC1 was overexpressed(MGC-803 OE-CLIC1,AGS OE-CLIC1)(P<0.05).6.Immunofluorescence experiments showed that the expression of AMOT-p130 was significantly increased in MGC-803 CLIC1 KO and AGS CLIC1KO(P<0.05).Conclusions: The expression of CLIC1 in gastric cancer tissue was increased,and it is related to vascular invasion and TNM pathological stage;the expression level of AMOT-p130 in gastric cancer tissue was decreased,and it is related to the depth of tumor invasion and TNM pathological stage.The correlation between high expression of CLIC1 and low expression of AMOT-p130 was statistically significant.CLICl inhibits the expression of AMOT-p130 in gastric cancer cells.Part 2 CLIC1 mediates the effect of AMOT-p130 on the biological behavior of gastric cancer cellsObjective: To explore the effects of CLIC1-mediated AMOT-p130 on the invasion,migration and proliferation of gastric cancer cells.Methods: MGC-803 CLIC1 KO and AGS CLIC1 KO were used to construct stable cell lines with low and high expression of AMOT-p130 for subsequent experiments.Transwell and wound-healing assays were used to detect the migration and invasion ability of gastric cancer cells,and CCK8 and EDU were used to detect the proliferation ability of gastric cancer cells.Results: 1.Compared with the control group,MGC-803 CLIC1 KO and AGS CLIC1 KO have weakened invasion and migration ability(P<0.05),and weakened proliferation ability(P<0.05).2.After silencing the expression of AMOT-p130,the invasion and migration ability of gastric cancer cells was enhanced(P<0.05),and the proliferation ability was enhanced(P<0.05)compared with the control group.After overexpression of AMOT-p130,the invasion and migration ability of gastric cancer cells was decreased(P<0.05),and the proliferation ability was decreased(P<0.05).Conclusions: CLIC1 promotes the invasion,migration and proliferation of gastric cancer cells by regulating the expression of AMOT-p130 in vitro.Part 3 Study on the mechanism of CLIC1 mediating AMOT-p130 to affect the biological behavior of gastric cancer cellsObjective: To explore the relevant mechanism of CLIC1 mediating AMOT-p130 to affect the biological behavior of gastric cancer cells.Methods: Western blot was used to detect the changes of AMOT-p130,E-cadherin,Snail and Vimentin.Results: 1.Compared with the control group,after silencing AMOT-p130,the expression of E-cadherin was downregulated(P<0.05),while the expression of Vimentin and Snail was upregulated(P<0.05)in MGC-803 CLIC1 KO and AGS CLIC1 KO.2.After overexpressing AMOT-p130,the expression of E-cadherin was upregulated(P<0.05),while the expression of Vimentin and Snail were downregulated(P<0.05)in MGC-803 CLIC1 KO and AGS CLIC1 KO.Conclusions: CLIC1 mediates AMOT-p130 to regulate the epithelial-mesenchymal transition of gastric cancer cells. |
