The Relationship Between FAK Gene Expression And The Development Of Esophageal Cancer

Background and ObjectiveEsophageal carcinoma is a malignant tumor that occurs in the a third of the upper part of the esophagus and the middle of the squamous cell carcinoma and the lower part of the glandular cell carcinoma esophagus,which accounts for almost 2%of all malignancies.The mortality rate is second in all malignant tumour.According to incomplete statistics around the world about 180,000 people died of esophageal cancer each year,and China as the world's highest incidence and mortality of esophageal cancer,nearly 100,000 people die each year.Therefore,the regulation of esophageal cancer diagnosis and treatment can benefit many patients with esophageal cancer,which is an important task for clinicians.At present,the main means of esophageal cancer treatment is surgical treatment,radiotherapy can also used in early esophageal cancer.Although the unilateral chemotherapy effect is not obvious,but the clinic is usually used as adjuvant therapy,surgery,radiotherapy and chemotherapy combined to ensure that the treatment of cancer better.The key to malignant tumor treatment is good or bad is mainly depending on the early diagnosis of the tumor is timely,the sooner the diagnosis,the greater the probability of successful treatment.With the development of science,the use of specific genes for the diagnosis and treatment of tumors has become a new means of biological diagnosis and treatment,the search for specific genes for early diagnosis of the tumor,post-treatment and postoperative recurrence of prevention has important clinical significance.Therefore,the study of tumor-related gene function is quite important.Tumor metastasis and recurrence are important features of cancer.During the development of tumor,Epithelial Mesenchymal Transition process can make some malignant manifestation of tumor cells,which leads to the increase of cell adhesion between cell and cell and extracellular matrix,increased celluar mobility,proliferation and associated with dedifferentiation,no apoptosis and loss of contact inhibition.According to the study of the genes involved in the development and progression of esophageal cancer,it is very possible to solve the clinical difficult problem of esophageal cancer.The study of Focal adhension kinase in the laboratory has revealed that it is highly expressed in various tumor types and is closely related to the occurrence and development of malignant tumor.In view of this,FAK as the research object,to explore its relationship with the occurrence and development of esophageal cancer,to evaluate the role of FAK in the development and progression of esophageal cancer and its potential as a target for the diagnosis and treatment of esophageal cancer.In this study,RNAi was used to inhibit the expression of FAK in esophageal cancer cell line Eca-109,and to study the relationship between FAK and esophageal cancer cell proliferation,motility,apoptosis and cytoskeleton,cell surface microstructure and some aspects of esophageal cancer EMT Related genes.Methods1.The relevant gene information of FAK gene was retrieved in NCBI database and the siRNA targeting FAK was designed online.The specificity was detected and cloned into plasmid pU6H 1-GFP to form pU6H1-GFP/FAK-siRNA.2.The prepared FAK plasmid was transfected into human esophageal cancer cell line Eca-109 by calcium phosphate transfection,using wild cells and missense siRNAscr cells as control,the transfection efficiency was evaluated after transfection for 72 h.3.Injury repair,MTT,soft agar clone formation and Trans Well chamber were used to detect the effect of FAK expression on the proliferation and motility of Eca-109 cells.4.The effect of FAK downregulation on the cytoskeleton and surface microstructure of Eca-109 cells was detected by Coomassie brilliant blue staining and scanning electron microscopy.5.The expression of MMP9,VIM,KRT1 and E-cadherin was detected by Western blot after downregulation the expression of FAK.6.The expression of MMP9,VIM,KRT1 and E-cadherin in cell level was detected by immunocytochemistry after downregulation the expression of FAK.7.The relationship between MMP9,VIM,KRT1,E-cadherin and the development of esophageal carcinoma was analyzed by bioinformatics.Results1.The recombinant plasmid targeting FAK was successfully constructed and the transfection efficiency of the cells was over 85%.2.Western blot results showed that FAK siRNA3 could significantly reduce the expression of FAK in esophageal cancer Eca-109 cells.3.The results of injury repair,Transwell,MTT and soft agar clone showed that the proliferation and motility of Eca-109 cells were significantly decreased after FAK gene expression was down-regulated.4.Coomassie brilliant blue staining,scanning electron microscopy showed that FAK gene expression down-regulated made intercellular contact inhibition has been restored and the cell morphology changed.5.Western blot and immunohistochemistry showed that inhibition of FAK expression could significantly inhibit the protein expression of MMP9 and VIM,meanwhile,enhance the protein expression of KRT1 and E-cadherin.Conclusion1.Downregulation the expression level of FAK can effectively inhibit the movement and proliferation of Eca-109 cells,which indicates that FAK plays an important role in the development of esophageal cancer cells metastasis and invasion.2.Western blot and immunochemistry showed that the expression of FAK gene was positively correlated with the expression of MMP9 and VIM,and negatively correlated with the expression of KRT1 and E-cadherin,indicating that FAK was involved in the EMT process of esophageal cancer.3.Downregulation of FAK expression can effectively restore Eca-109 cell contact inhibition.4.Bioinformatics analysis of the statistical data of clinical cases of esophageal cancer showed that FAK,MMP9,VIM,KRT1 and E-cadherin were significant in the development of cancer in the clinical data of esophageal cancer.5.This study shows that FAK gene expression is closely related to the development andprogression of esophageal cancer by studying the analysis of the behavioral characteristics of esophageal cancer cells and EMT-related genes,which further reflects the importance of FAK gene has significant reference value in the diagnosis and treatment of esophageal cancer.