| BACKGROUND AND OBJECTIVES: Gastric cancer is one of the common malignant tumors, of which the morbidity and mortality are located in the forefront. At the same time, the 5-year survival rate is only 5% to 15%. Current studies suggested that the main reason for failure treatment is early metastasis of gastric cancer. Transformation of epithelial mesenchymal is hot research topic of molecular mechanism. It is obvious that EMT plays a significant role in cancer progression and metastasis. Important EMT characteristics include the loss of cell-to-cell contacts and increased cell motility, which is implicated in the aggressive invasion and metastasis of late stage tumors. Therefore, there is an urgent need for an improved understanding of the molecular mechanisms underlying the aggressive and invasive metastasis of gastric cancer.In this study, at first, we tested the expression level of Fbxw7 in gastric cancer tissues and corresponding normal tumor-adjacent tissues, gastric cancer tissues without metastasis and gastric cancer tissues with serosal invasion or vascular invasion or lymph node metastasis. Then we analyzed the relationship between reduced Fbxw7 expression and clinical significance. Secondly, we tested the cytological features of gastric cancer cells and the molecular markers of EMT in Fbxw7 over-expression or knock-down gastric cancer cells using lentivirus transduction. Finally, we analyzed the relationship between Fbxw7 and RhoA in gastric cancer tissues and cells. Moreover, we further explore the specific molecular mechanism. We tested the cytological features of gastric cancer cells and the molecular markers of EMT before and after cotransfection of Fbxw7 and RhoA.METHODS:(1) Immunohistochemical staining was performed to determine the Fbxw7 expression in 60 gastric cancer tissues and corresponding normal tumor-adjacent tissues. Western-blot was used to test the Fbxw7 expression in gastric cancer tissues and corresponding normal tumor-adjacent tissues, gastric cancer tissues without metastasis and gastric cancer tissues with serosal invasion or vascular invasion or lymph node metastasis. Then we analyzed the relationship between reduced Fbxw7 expression and clinical significance.(2) The biological changes and molecular markers were detected in different gastric cancer cells with or without Fbxw7 over-expression or knockdown by Western Blot, co-transfection, CCK8 cell proliferation, cell invasion and migration, apoptosis, immunofluorescence and other experimental methods and we further explored the possible molecular mechanisms that the Fbxw7 how to regulate EMT.(3) We analyzed the relationship between Fbxw7 and RhoA via Immunohistochemical staining, Western Blot and RT-PCR. Then, we used co-IP to further explore the Fbxw7 how to regulate the expression of RhoA. Moreover, we evaluated the molecular mechanism of the biological changes and the variation of molecular markers alone with Fbxw7 changed via Western Blot, co-transfection, CCK8 cell proliferation, cell invasion and migration, apoptosis, immunofluorescence methods.RESULTS:(1) Our study showed that Fbxw7 plays an important role in gastric cancer, and the expression of Fbxw7 in gastric cancer tissues was significantly lower than that in normal tumor-adjacent tissues. The expression of Fbxw7 in gastric cancer tissues without metastasis was significantly higher than that in gastric cancer tissues with serosal invasion or vascular invasion or lymph node metastasis. We observed that the level of Fbxw7 protein expression significantly correlated with tumor size, TNM stage, serosal invasion, vascular invasion and lymph node metastasis, but not with other clinicopathological factors, including patient age, gender, and histological type. There was an intimate connection between Fbxw7 and gastric cancer prognosis via 5-year survival analysis.(2) Fbxw7 over-expression could induced the cell apoptosis and growth arrest and suppressed the invasion and migration of gastric cancer cells, thereby decreasing the expression of Snail1 and ZEB1, which act as important stimulating factors of the epithelial-to-mesenchymal transition(EMT).(3) We found that there was a significantly negative correlation between Fbxw7 and RhoA in protein level. Meanwhile, Fbxw7 over-expression could down-regulate the expression level of RhoA and GTP-RhoA. We further explored that Fbxw7 regulate RhoA through ubiquitination and proteasomal degradation. Finally, we found that Fbxw7 regulates tumor apoptosis, growth arrest and the epithelial-to-mesenchymal transition in part through the Rho A signaling pathway in gastric cancer via co-transfection technology.CONCLUSIONS: These findings demonstrate that Fbxw7 could regulate the expression of RhoA and regulates tumor apoptosis, growth arrest and the epithelial-to-mesenchymal transition in part through the RhoA signaling pathway in gastric cancer... |
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