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Objective To explore the role of CD95 system in chemotherapeutic sensitivity of pancreatic cancer cells, in an attempt to enhance the efficacy of chemotherapy by transfection CD95 gene, and to provide the evidence for the immunological treatment of pancreatic cancer. Methods CD95 gene was transfected into the pancreatic cancer cell line SW1990 by lipofectamine. The transfected cells were selected by G418. CD95 expressions of the transfected cells were detected by Northern blot and Western blot. MTT assay was used to analyze the response of the transfected cells to 5 fluorouracil, adriamycin (ADM), gemcitabine and in combination with anti CD95 monoclonal antibody (mAb). The drug induced apoptosis of transfected cells was measured by flow cytometry. Results The transfected pancreatic cancer cell SW1990 could overexpress CD95 stably. The CD95 mRNA and protein expressions were significantly increased in the transfected cells than in the controls. Anti CD95 mAb could inhibit the growth of the transfected cells. In addition, transfected cells were more sensitive to clinically relevant concentrations of chemotherapeutic drugs than non transfected cells. Anti CD95 mAb addition could enhance the cytotoxic effect of chemotherapeutic drugs. Drug induced apoptosis in ADM treated transfected cells more pronounced than in non transfected cells. Conclusions CD95 transfection could increase the sensitivity of pancreatic cancer cell SW1990 to, and partly reverse the resistance to, chemotherapeutic drugs. The combination of chemotherapeutic drugs with anti CD95 mAb showed a synergistic cytotoxicity to pancreatic cancer cells.
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Objective The anti tumor and anti metastasis effects of angiocytotoxic therapy (TNP 470/Gemcitabine) were investigated using a model of human pancreatic carcinoma by surgical orthotopic implantation (SOI). Methods The SOI model was developed by suturing small pieces of SW1990 tumors into the tail of pancreas in nude mice. Twenty four male mice were randomly divided into control group, G100 group receiving 100 mg/kg Gemcitabine intraperitoneally injection on days 0,3,6 and 9 after transplantation, and T30 group receiving 30 mg/kg TNP 470 subcutaneous injection on alternate days for 8 weeks. Another thirty two male mice were randomly divided into control group, T15 group, G50 group and combination group (TNP 470 30 mg/kg+ Gemcitabine 50 mg/kg). Animals were sacrificed ten weeks after transplantation. Results G100 group had a significant inhibitory effect on tumor growth of pancreatic carcinoma compared to T30 group, while the metastasis of tumor was significantly inhibited by T30 group compared to G100 group. Neither G100 group nor T30 group showed a significant improvement on survival rate. T15 group and G50 group alone had no significant inhibitory effect on the tumor growth and its metastasis. Mean while a significant anti tumor, anti metastatic effect and a significant improvement on the survival rate were observed in combination group. The inhibitory effect of G50 group was enhanced by 2 times with T30, and 2/8 of the tumors bearing animals were cured by the combination therapy. The level of microvessel density in T30 group was significantly lower than that in T15 group and control group ( P
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AIM: The current study was designed to construct eukaryotic expression vector containing NK4 gene and transfect it into human pancreatic cancer cell lines.METHODS: The recombinant of pcDNA3/hNK4 was digested by restriction enzyme, the NK4 gene was cloned into a high effective eukaryotic expressing plasmid which contains CMV2 immediate early gene promoter and then transiently introduced into the pancreatic cancer cell line SW1990 by lipofectamine and clonal cell lines that secrete high levels of NK4 protein were isolated.The expression of NK4 was observed by RT-PCR and Western blot, in vitro the vascular endothelial cell proliferation inhibiting activity of NK4 was examined by 3-[4,5-dimethylthiazolzyl]-2,5-diphenyl tetrazolium bromide(MTT) method. RESULTS: A specific expression of NK4 gene mRNA by lipofectamine-mediated transfer exhibited only in SW1990/NK4 cells,Western blot analysis demonstrated that there was positive expression of NK4 protein(50 kD).The NK4 inhibited proliferation of the vascular endothelial cells in vitro. CONCLUSION: The recombinant of pRC/CMV2-hNK4 is a high effective expressing eukaryotic vector.The bio-engineering product of the NK4 is an angiogenesis inhibitor and may play an important role in the gene therapy for tumor.
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AIM: To elucidate the pattern of 5-fluorocytosine(5-FC) induced apoptosis and its role in gene therapy for human pancreatic cancer. METHODS: The human pancreatic cancer SW1990 cells(CEA-producing) were infected with recombinant adenoviruses(Adex1CEA-prCD or Adex1CEA-prZ).Cytosine deaminase(CD) expression was examind by western blot. Apoptosis induced by 5-FC in human pancreatic cancer SW1990 cells genetically modified to express cytosine deaminase was investigated by applying electron microscopy, DNA electrophoresis and flow cytometry analysis techniques. RESULTS: The SW1990 cells infected with Adex1 CEA-prCD were treated with 5-FC at 100 ?mol?L -1 for 48 h, cell apoptosis occurred. Typical apoptosis morphological feature appeared and DNA ladder could be demonstrated on DNA electrophoresis. Apoptosis peak was also showed by flow cytometry. Apoptotic cells accounted for 34.6% of the cell population. Cells in G 1, S and G 2/M phase of cell cycle were 64%, 11% and 7%, respectively. CONCLUSION: The apoptosis induced by 5-FC may be a primary mechanism in CD gene therapy for pancreatic cancer.
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AIM: To investigate the expression of MAT1 protein in pancreatic cancers and the relationship between MAT1 and clinicopathological features of pancreatic cancers. METHODS: 94 surgical specimens, including 70 pancreatic cancers, 10 pancreatic benign tumors, 14 chronic pancreatitis and 10 autopsy normal pancreas tissues, were analyzed immunohistochemically, and then MAT1 expression and clinicopathological features were compared. RESULTS: MAT1 was expressed mainly in the cancer cells,and also in the fibroblasts, where it was localized within the cytoplasm and nuclear envelope. MAT1 expression was found in 75.7% (53/70) of the cancers, but not detected or weakly expressed in control tissues. There was a significant difference in expression of MAT1 among the above four tissues (P