The Role And Mechanism Of Transcription Factor FOXO4in The Proliferation And Metastasis Of Gastric Cancer

Gastric cancer is one of the most important malignant tumors who threatened ourhealth. Recently, the major methods for gastric cancer therapy have included surgicaltreatment, radiotherapy,chemotherapy, biological therapy and so on. The more effective ofsurgeryand adjuvant therapy is main for gastric carcinoma in situ, and the lethality ofgastric cancer mostly due to tumor metastasis. Therefore, exploring and finding the keymolecular involved in gastric growth and metastasis and its potential mechanism may helpdiagnosing GC and predict its progress, to reduce the fatality rate of gastric cancer.TheFOXO transcription factor family is thought to play an important role in the gastriccarcinogenesis. In our previous study, we found that transcription factor FOXO4had alower expression in metastatic colon cancer tissue. We supposed that whether FOXO4is akey molecular in gastric carcinogenesis and what its specific mechanism is. So, to studythe function and mechanism of transcription factor FOXO4in progression of gastriccancer is beneficial of our understanding of gastric carcinogenesis. Aims:1. To study the exprseeion change of transcription factor FOXO4in gastric cancer tissuesand cell lines.2. To build the overexpression and down-rugulated expression of gastric cancer cellsublines and research theeffect of FOXO4on the metastasis of gastric cancer in vitroand in vivo.3. To further explore the effect of FOXO4on gastric cancer metastasis and the potentialmechanisms.Methods:1. To verify the expressional change of FOXO4in gastric cancer tissues and gastriccancer cell lines by immunehistochemistry, western blot, tissue microarray, andreal-time PCR respectively.2. To build the over-expression of FOXO4lentivirus vector and down-regulatedexpression oligo-nucleotide vector of FOXO4, and transfect them into gastric cell linesSGC-7901and BGC-823respectively, then using western blot and RT-PCR to verifythe transfection efficiency.3. Through cell proliferation assay(MTT), plate clone and wound healing assay to verifythe effect of FOXO4on gastric cancer growth and metastasis in vitro, and high contentscreening assay help real-time monitoring the movement of SGC-7901and7901-FOXO4cell lines.4. By subcutaneous tumorigenecity in nude mice and tail vein metastatic assay (injectingthe SGC-7901and7901-FOXO4cell lines respectively), and bioluminescentimaging(BLI) of tail vein injected nude mice was performed periodly to study theinfluence of FOXO4on gastric cancer proliferation, metastasis and prognosis in vivo.5. Through bioinformatic methods to predict the potential mechanism of FOXO4ongastric cancer, and test it by performing RT-PCR, western blot andimmunocytofluorescence technique.Results:1. There is a lower expression of FOXO4in tissues and cell lines of gastric cancer compared to normal gastric mucosal tissue and cell line, and a lower expression ofFOXO4was also found in tissues of gastric cancer with lymph node metastasisrelative to carcinoma in situ(P<0.05).2. After tranfection of FOXO4overexpression lentivirus, there is a obviously increaseexpression of FOXO4in gastric cancer cell line SGC-7901; After transfection ofdown-regulated expression of FOXO4siRNA, it is tested by western blot and RT-PCR,FOXO4was significantly down regulated in gastric cancer cell line BGC-823.3. The ability of proliferation and metastasis for FOXO4overexpression of gastric cancercell line7901-FOXO4declined obviously, and opposite result was shown in FOXO4siRNA transfected gastric cancer cell line BGC-823.4. Through injecting gastric cancer cells subcutaneously in nude mice, it is shown that7901-FOXO4has a lower tumorigenecity than SGC-7901, and it is found that there isa weaker potency of distant metastasis in7901-FOXO4than gastric cancer SGC-7901by tail vein injection animal experiment.5. RT-PCR and cell immunofluorescence results confirmed:7901-FOXO4has adecreased expression of vimentin, and a increased expression of E-cadherin.Conclusions:1. Transcription factor FOXO4expressed decreasely in gastric cancer tissues and celllines.2. Studies have confirmed that transcription factor FOXO4has obviously inhibited theproliferation and metastasis of gastric cancer in vitro and in vivo.3. The mechanism of FOXO4inhibiting gastric carcinogenesis is partly by restraining thegenesis of EMT.