Overexpression Of Vascular Endothelial Growth Factor-C Gene Promoted Human Gastric Cancer Cells Proliferation And Colony Formation And Endothelial Tube Formation

Background and objective:Gastric cancer is one of gastrointestinal cancers with the serious threat to the physical and mental health of human, and the morbidity and mortality of gastric cancer have been in the second place of malignant tumors. The lymph node metastasis in gastric cancer is the early distant metastasis of patients, but also is an important cause eventually leading to the death. The current view is that lymphangiogenesis in tumor adjacent tissue is the most crucial factor of tumor cells lymph node metastasis, but the specific mechanism remains unclear. VEGF-C (Vascular endothelial growth factor-C) is a member of the VEGF family, and one of the biological factors can promote lymphangiogenesis has accepted. Related findings, VEGF-C can combine specific receptor VEGFR-3(vascular endothelial growth factor receptor-3) in lymphatic endothelial cell surface, mediate and activate tyrosine kinase signaling system of the receptor, which can contribute to the proliferation and migration of lymphatic endothelial cells, regulate lymphangiogenesis of tumor. There are numerous studies have showed that VEGF-C in gastric cancer tissues has significant increase in the levels of expression of genes and proteins, by regulating expression of VEGF-C gene and interaction of VEGF-C and its receptor VEGFR-3, promote micro lymphatic formation in the periphery of tumor, which contribute to tumor cells distant metastasis through the lymphatic. Some recent studies have confirmed the VEGFR-3receptor not only expressed in the lymphatic vessels mesothelial cells, but aslo there are different levels of expression in human malignant cells, such as gastric cancer, colorectal cancer, breast cancer, esophageal cancer, and lung cancer. Although a large number of studies on the VEGF-C gene, but its biological effects in the tumor occurrence and development are still not very clear. In this study, by constructing and packaging the overexpression of VEGF-C gene lentiviral, and infecting human gastric cancer cell MKN-45, observed the effects of VEGF-C gene in human gastric cancer cell proliferation, colony formation and endothelial cell tube forming, explored the biological function in the development of gastric cancer tissues, provided a theoretical basis for the VEGF-C gene study in tumor development.Methods:The differences of the VEGF-C mRNA and protein expression levels among MKN-45、MKN-28、NCI-N87、BGC-823、AGS and SGC-7901gastric cancer cell lines were were tested by the SYBR Green relative quantitative reverse transcriptase polymerase chain reaction (QRT-PCR) method and protein immune imprinting method (Western blot method). We assayed cell growth activity by CCK-8, colony formation in soft agar, and endothelial tube formation by constructing lentiviral plasmid of human vascular endothelial growth factor-C gene overexpression. The whole experiment was divided into the experimental group (MKN-45-GV/C), the control group (MKN-45-GV) and the blank group (MKN-45)Results:1.VEGF-C gene and protein expression in all of the six gastric cancer cell lines was detected with different levels by QRT-PCR and Western Blot, the minimum expression of VEGF-C was found in MKN-45gastric cancer cell line as0.45±1.44, and the maximum expression in SGC-7901cell line as31.13±0.75in which;2.The cell proliferation assay through CCK-8indicated:compared with control group, the experimental group showed a significant increase of cell proliferation volume (P<0.01)3.The results of colony formation in soft agar demonstrated:experimental group and control group had the cloned number of6.23±0.32and1.40±1.67respectively per average view. The efficiency of colony formation was85%and31%respectively (P<0.05)4.The endothelial tube formation experiment showed:the experimental group, control group and blank group had tubular number of8.14±1.25,12.76±0.79and18.46±0.72respectively, the length of tube was98.56±3.71,105.17±1.34and151.62±2.51respectively.Conclusions:1.Recombinant lentiViral human vascular endothelial growth factor-C gene plasmid was successfully constructed;2.The gastric cancer cell line of VEGF-C overexpression was established successfully;3.VEGF-C gene promoted the cell growth, colony formation, and endothelial tube formation.