ABSTRACT
<p><b>UNLABELLED</b>OBJECTIVE To explore the effects of human macrophage inflammatory protein-1 beta (hMIP-1beta) modification on the in vivo tumorigenicity and vaccine efficacy of tumor cells.</p><p><b>METHODS</b>Murine colorectal adenocarcinoma CT26 cells were transfected with a recombinant adenovirus carring the hMIP-1beta gene (AdhMIP-1beta). The efficacy of gene transfection was tested by X-gal staining. The hMIP-1beta level in the supernatant of hMIP-1beta gene-modified CT26 cells was assayed by ELISA, and the chemotactic activity for CD4+ T cells, CD8+ T cells, NK cells and immature dendritic cells (imDCs) were assayed by a transwell chamber. The changes of growth characteristics and in vivo tumorigenicity of hMIP-1beta gene-modified CT26 cells were also assessed. BALB/c mice were immunized with hMIP-1beta gene-modified CT26 tumor vaccine and the antitumor effect was evaluated.</p><p><b>RESULTS</b>hMIP-1beta gene could be transfected into CT26 cells by AdhMIP-1beta with an efficiency over 95%. The level of hMIP-1beta in the culture supernatant of hMIP-1beta gene-modified CT26 cells was 980 pg/ml and the supernatant displayed ramarkable chemotactic activity to CD4+ T cells, CD8+ T cells, NK cells and imDCs compared with LacZ gene-modified CT26 cells and control. When the hMIP-1beta gene-modifited CT26 cells were subcutaneously inoculated in BALB/c mice, the tumorigencity was delayed and suppressed, and overt necrosis and lymphocyte infiltration were observed in the tumor tissue, but not in those inoculated with LacZ gene-modified CT26 cells or parental CT26 cells. The mice immunized with hMIP-1beta gene-modified CT26 tumor vaccine could induce tumor specific CTL activity and nonspecific NK activity, and exhibited resistance to later challenge with wild-type CT26 cells.</p><p><b>CONCLUSION</b>hMIP-1beta gene-modified CT26 cells exhibit decreased tumorigenicity, and hMIP-1beta gene-modified tumor vaccine may induce a powerful specific and nonspecific antitumor response. The data suggested that hMIP-1beta gene-modified tumor vaccine may play a potent role in prevention of metastasis and recurrence of malignant tumors.</p>
Subject(s)
Animals , Female , Humans , Mice , Adenocarcinoma , Metabolism , Pathology , Adenoviridae , Genetics , CD4-Positive T-Lymphocytes , Allergy and Immunology , CD8-Positive T-Lymphocytes , Allergy and Immunology , Cancer Vaccines , Cell Line, Tumor , Chemokine CCL4 , Genetics , Metabolism , Chemotaxis, Leukocyte , Colonic Neoplasms , Metabolism , Pathology , Cytotoxicity, Immunologic , Dendritic Cells , Allergy and Immunology , Genetic Vectors , Killer Cells, Natural , Allergy and Immunology , Mice, Inbred BALB C , Neoplasm Transplantation , Random Allocation , Recombinant Proteins , Genetics , Metabolism , Transfection , Tumor BurdenABSTRACT
<p><b>OBJECTIVE</b>To construct a dendritic cell vaccine transduced by murine alpha-fetoprotein (mAFP) gene, and evaluate its immunoprotective effect on C57BL/6J mice during the induction of hepatocellular carcinoma by diethylnitrosamines, carbon tetrachloride and ethanol.</p><p><b>METHODS</b>Dendritic cells (DCs) were induced and augmented by murine IL-4 and GM-CSF, and transfected by recombinant adenovirus engineered with mAFP gene. Major MHC class I and II, B7.1 (CD80), B7.2 (CD86), CD18a, and CD54 molecules on DC were analyzed by FACS. 80 C57BL/6J male mice were randomly divided into 4 groups (20 mice per group): Simple DC inoculated group, pAdBM5-mAFP-DC inoculated group, pAdBM5-mAFP plasmid inoculated group, and PBS control group. They were immunized once with 5 x 10(5) DCs (0.1 ml)/mouse administered s. c. in the left flank or 100 mg pAdBMS-mAFP plasmid/mouse administered i. m. in the left tibialis anterior muscle. Inoculation was conducted once a week for 4 weeks after 3 times consecutive immunization initially. At the same time of immunization, DEN/CCl4/ethanol were given to induce hepatocellular carcinoma. Tumor incidence was assessed after 20 weeks.</p><p><b>RESULTS</b>A transgenic DC vaccine was successfully constructed and the mAFP transgenic DCs expressed high level molecules of major MHC class I and II , B7.1, B7.2, CD18a, and CD54. After the 20-week induction, the incidence of primary hepatocellular carcinoma (PLC) was 70.0% in simple DC inoculated group, 25.0% in pAdBMS-mAFP-DC inoculated group, 65.0% in pAdBM5-mAFP plasmid inoculated group, and 75.0% in PBS control group. There was a significant difference between group B and other groups (P < 0.05).</p><p><b>CONCLUSION</b>mAFP transgenic DC tumor vaccine inoculation may induce strong immunoprotection against liver carcinogenesis and tumor development and reduce PLC incidence induced by DEN/CCl4/ethanol.</p>
Subject(s)
Animals , Male , Mice , Adenoviridae , Genetics , B7-1 Antigen , Metabolism , Cancer Vaccines , Carbon Tetrachloride , Cells, Cultured , Dendritic Cells , Cell Biology , Allergy and Immunology , Metabolism , Diethylnitrosamine , Ethanol , Genetic Vectors , Histocompatibility Antigens Class I , Metabolism , Histocompatibility Antigens Class II , Metabolism , Intercellular Adhesion Molecule-1 , Metabolism , Liver Neoplasms, Experimental , Allergy and Immunology , Mice, Inbred C57BL , Random Allocation , Recombinant Proteins , Genetics , Metabolism , Transfection , alpha-Fetoproteins , Genetics , MetabolismABSTRACT
<p><b>OBJECTIVE</b>To investigate the expression of integrin-linked kinase (ILK) in kidneys of mice with unilateral ureteral obstruction and its relevance with the epithelial-mesenchymal transition.</p><p><b>METHODS</b>Mice were randomly divided into two groups, sham operation (C, n = 20) and unilateral ureteral obstruction (UUO, n = 40). The animals were sacrificed at day 1, 3, 7 and 14 respectively after the surgery. Tubulointerstitial fibrosis (TIF) was graded according to Masson staining. The protein level of ILK was examined by Western blot. Tissue/cytological expression for ILK, alpha-SMA and E-cadherin were investigated by immunohistochemistry. The mRNA levels of ILK, alpha-SMA and E-cadherin were analyzed by quantitative real-time PCR.</p><p><b>RESULTS</b>In the control animals (group C), weak staining for ILK was detected mainly in the podocytes. Significant increase of staining for ILK in the experimental mice (UUO group) was detected from day 1 onward (t = 16.5, P < 0.01), reaching the peak at day 7. The protein expression of E-cadherin was continuously down-regulated from day 3 onward after surgery (t = 21.0, P < 0.01), while expression for alpha-SMA was up-regulated. From day 1 to day 7, the protein expression of ILK was positively correlated with alpha-SMA (R = 0.88, P < 0.01), but negatively correlated with E-cadherin (R = -0.87, P < 0.01). The mRNA expression of ILK and alpha-SMA analyzed by real-time PCR increased from postoperative day 1 and 3 respectively, but the mRNA expression of E-cadherin decreased from day 3 onward.</p><p><b>CONCLUSION</b>Increasing expression of ILK occurs in the early phase of UUO mouse and may play an important role in the process of TIF through mediating the epithelial-mesenchymal transition.</p>