| Ovarian carcinoma is one of gynecological malignancies with poor prognosis, and its five-year survival rate of advanced-stage patients is only 15-20%. One of the reasons is that most of patients with ovarian carcinoma present advanced-stage disease at the first diagnosis with wide-spread of cancer cells and great accumulating of ascetic fluid in peritoneal cavity. Vascular endothelial growth factor (VEGF) is an important growth factor associated with progression of ovarian cancer and accumulating of ascetic fluid and plays an important role in invasion, metastasis and ascetic fluid accumulating in ovarian cancer. A lot of evidences suggest that there are high expression of VEGF in many kinds of human tumors including gastric, ovarian, breast, bladder, prostate cancers, etc. The main function of VEGF is promoting angiogenesis through binding with VEGFRs in vascular endothelial cells. The mechanism of VEGF effects involves that VEGF binds with VEGFRs, promotes proliferation and migration of vascular endothelial cells and increases permeability of microvascules. In vascular endothelial cells, VEGF induced various biological effects through several signal pathways including MAPK, PI3K and PLC-γ mediated by VEGFRs. In recent studies, in vitro, someevidences show that VEGF signal transduction in vascular endothelial cells also induces JAK-STAT pathway. VEGF can stimulate STATs phosphorylation and regulate the transcription of downstream targeted genes in vascular endothelial cells.Recent studies show that VEGFRs are also expressed on the surface of various tumor cells, which suggests that there is not only a paracrine loop through vascular endothelial cells to promote angiogenesis for tumor growth but also an autocrine loop in tumor cells for proliferation and anti-apoptosis. But whether VEGF signal pathway in tumor cells follows those in vascular endothelial cells, especially STAT signal pathway, remains unclear. Previous study showed constitutive activation of STAT3 in many kinds of cancers including ovarian carcinoma. Abnormal activations of STAT3 are always associated with occurrence and progression of malignant tumors. The high expression of VEGFRs and phosphorylation of STAT3 in tumor cells suggest phosphorylated STATs may participate in VEGF signal pathways in tumor cells.In the study,levels of p-STAT3 and VEGFR2 induced by VEGF with different concentration and different effect-time were detected by Immunocytochemistry and Western Blot. Furthermore, whether p-STAT3 participated in VEGF signal transduction in ovarian carcinoma cells was observed through blocking the bind of VEGF and VEGFR2 with a peptide specific for VEGFR2.The aim of the study is to investigate the molecular mechanisms by which VEGF directly effects on ovarian cancer cells.Materials and MethodsEpithelial ovarian cancer cell lines SKOV-3, Caov-3 and 3ao were selected for studies. The expression of p-STAT3 and VEGFR2 in three celllines induced by VEGF were detected and the cell lines without VEGF stimulation were as controls. A cell line with the highest response to VEGF incubation in p-STAT3 and VEGFR2 expression was regarded to be a model for further study. P-STAT3 expression induced by VEGF with different concentrations (0, 5, 25,50,100 ng/ml) and different times(0,15,30,45,60min) of VEGF incubation was observed. The suppression of p-STAT3 expression induced by VEGF was further observed when blocked the VEGFR2 by the peptide(ATWLPPR) with different concentrations(0,40,80,160,320 μM) and different effect-times (0,15,30,45,60min). Immunocytochemistry and Western blot were used for detecting expressin of p-STAT3 and VEGFR2. The results were semi-quantitified by grade evaluation of immunocytochemical staining and Quantity One Software for Western blot.Oneway ANOVA and Univariate Analysis of Variance were used for statistical analysis. All data were managed with SPSS 10.0 for windows.Results1, Screening of cell linesVEGFR2 and p-STAT3 were found to be co-expressed in all the three cell l... |
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