The Relationship Between FAK Gene And Gastric Cancer, Liver Cancer Cell Behavior And Related Gene Expression

Backgrounds and Objectives:Gastric cancer and liver cancer are the most common gastrointestinal cancer, new cases and deaths ranks as the first in China. Currently, surgery、chemotherapy、radiation therapy and other methods are the main way to treat gastric cancer and liver cancer, but the effect is poor. Gene diagnosis and treatment as an emerging methods, has obvious advantages in the treatment and prognosis for cancer. It is may be an important aids to fight against cancer in the future. Therefore, the study of gene function associated with the tumor is particularly important. Explore gastric and liver cancer of development mechanism to achieve gene diagnosis and treatment, which may be resolved cancer clinical dilemma. Tumor metastasis, recurrence is an important sign of malignant cancer, which involves cell-to-cell adhesion reduced, intercellular and extracellular matrix adhesion enhanced and extracellular matrix degradation enhancements. In the process of tumor development, cells can be obtained cell malignant phenotype by epithelial-mesenchymal (EMT). EMT process requires cell connections, cell-matrix sites by the integrin receptor mediated focal adhesion formation and other changes. Focal adhesion kinase (FAK) is a key protein plays an important role.Our previous studies suggest that FAK is overexpressed in many cancer cells/ tissues. It is closely related to the process of cancer occurrence and development. In order to reveal the relationship between FAK and malignant progression of gastric cancer and liver cancer, assess their significance in gastric cancer and liver cancer development process, and evaluate the potential value of gastric cancer and liver cancer diagnosis and treatment of target molecule. We inhibited the FAK expression in SGC-7901 and SMMC-7721 cells using RNAi, followed by the analysis of cell growth and motility, as well as explore the relationship of potential relationships with certain related genes.Method1. NCBI database search FAK gene for online design of FAK siRNA, Blast Hit evaluates specific, constructed pU6H1-GFP/FAK-siRNA.2. pU6H1-GFP/FAK-siRNA plasmids were transfected into SGC-7901 cells and SMMC-7721 cells by calcium phosphate transfection methods. siRNAscr and wild type cells as a control. After 72h, RT-PCR and Western blot confirmed suppression efficiency.3. Downregulation of FAK, wound healing assay and Transwell assay were used to detect the motility of SGC-7901 and SMMC-7721 cells.4. Downregulation of FAK, flow cytometry was used to detect the cell proliferation of SGC-7901 and SMMC-7721 cells.5. Downregulation of FAK, RT-PCR and Western blot were used to detect the mRNA and protein expression levels of Src、Ras、Apex1、 Rgnef.6. Downregulation of FAK, immunofluorescence were used to detect the cell levels of Src、Ras、Apex1、Rgnef expression.7. Used bioinformatics analysis of FAK、Src、Ras、Apex1、Rgnef with gastric cancer and liver cancer development potential relationship.Result1. Successfully constructed recombinant plasmid, calcium phosphate transfection efficiency above 80%.2. FAK siRNA plasmid significantly inhibited the expression levels of FAK in cancer cells.3. Wound healing assay, Trans Well assay and flow cytometry results suggest that downregulation of FAK, the motility of SGC-7901 and SMMC-7721 cells were inhibited significantly. SGC-7901 cell proliferation decreased with the Gl time arrest, SMMC-7721 was arrested in S phase, but did not find apoptotic peak.4. RT-PCR、Western blo、immunofluorescence shown that downregulation of FAK, at the mRNA、protein and cell expression of Src、Ras、Apex1 and Rgnef significantly were suppressed.Conclusion1. Downregulation of FAK inhibit SGC-7901, SMMC-7721 cell motility and proliferation. SGC-7901 cell cycle was delayed in the Gl phase; SMMC-7721 cell was delayed in S phase. This results show that FAK play an important role on SGC-7901 and SMMC-7721 cells metastasis, invasion and cancer development process.2. RT-PCR, Western blot, immunofluorescence results show that the expression of FAK was positively correlated with Sr、Ras、Apex1、Rgnef expression.3. Bioinformatics analysis showed that the clinical cases of gastric cancer and liver cancer, FAK、Src、Ras、Apex1、Rgnef for the cancer development is significantly.4. From the perspective of the malignant phenotype of cancer cells and related genes, our researches indicate that FAK involved the occurrence and development of gastric cancer and liver cancer. FAK may be considered as a promising biomarker for the molecular diagnosis and a candidate target for the therapy of human gastric cancer and liver cancer.