Effect Of Anti-cervical Cancer Immunity Induced By Cord Blood-derived Dendritic Cells Loaded With HPV E7_44-62 Peptide And Activated By Poly I:C

Part â…  Study on the effect of anti-cervical cancer immunity induced by cord blood-derived dendritic cells loaded with HPV E744-62peptide and activated by Poly â… :C in vitro.Objective:To investigate the effect of anti-cervical cancer immunity induced by cord blood-derived dendritic cells loaded with HPV E744-62peptide and activated by Poly â… :C in vitro.Methods:Cord blood was collected from healthy term delivery mother.Cord blood mononuclear cells were isolated by density centrifugation. The adherent fraction was cultured in presence of rhGM-CSF/IL-4for7days. DCs were activated by HPV E744,62peptide, Poly â… :C, and HPV E744.62peptide combined with Poly I:C, respectively, with DCs without loading antigen as control group. The yield of mature DCs was calculated and the morphological changes of the cultured cells were observed by light microscopy. Surface expressions of CD1a, HLA-DR, CD40and CD83were analysized by flow cytometry analysis. IL-12and IL-10levels of DCs were detected using ELISA method. The ability of DCs to stimulate the proliferation of lymphocytes was determined by mixed lymphocyte reaction. IFN-γ level of T cells was detected using ELISA method. MTT assay was employed to test the inhibition rates of tumor specific cytotoxic T lymphocyte on human cervical cancer cell line CaSki and human hepatocarcinoma cell line HepG2.Results:A certain amount of DCs with typical morphology and phenotype were harvested from all groups. Antigen-loaded DCs had higher expression of CD1a, HLA-DR, CD40, and CD83, ability of inducing proliferation of allogeneic T cells, secretion of IL-12from DCs, secretion of IFN-γ from T cells, specific antitumor effect of CTL on CaSki cervical cancer cells, and lower secretion of IL-10, than that of DCs without loading antigen, especially in group of DCs activated by HPV16E744-62peptide combined with Poly â… :C, while CTL had no specific antitumor effect to HepG2.Conclusion:It can be induced the specific anti-cervical cancer immunity by cord blood-derived dendritic cells loaded with HPV E744-62peptide in vitro. Poly â… :C might act as a adjuvant to enhance antitumor effect. Part II Study on the effect of anti-cervical cancer immunity in vivo induced by cord blood-derived dendritic cells loaded with HPV E744-62peptide and activated by Poly I:C in vivo.Objective:To evaluate the effect of anti-cervical cancer immunity induced by cord blood-derived dendritic cells loaded with HPV E744.62peptide and activated by poly I:C in bearing human cervical cancer nude mice with immune reconstitution of human T cells.Methods:Cell immune reconstitution of nude mice was established by tail vein injection of T cells from cervical cancer patients. Then24hours later, those nude mice were injected subcutaneously with CaSki cells to establish bearing human cervical cancer model. And one week later, those nude mice were inoculated with different vaccines by subcutaneously injection, and randomly divided into5groups according to the type of vaccines:group Ctrl (vaccinated with cord blood-derived dendritic cells loaded without any antigen), group E (vaccinated with cord blood-derived dendritic cells loaded with HPV E744-62peptide), group P (vaccinated with cord blood-derived DCs activated by poly â… :C), group E+P (vaccinated with cord blood-derived DCs activated by HPV E744-62peptide and poly I:C), and group NS (vaccinated with NS, as control group). The second injection was given two weeks interval. Biological behavior of nude mice, tumorigenecity, latent period, tumor volume, and the survival period of nude mice were observed. At35days after nude mice bearing human cervical cancer, the percentage of human CD3+, CD4+, CD8+T cells in mice blood and the level of IFN-γ in mice serum were detected.Results:The tumorigenecity was100%and the latent period is5to8days. Human CD3+/CD4+, CD3+/CD8+T cells could detected in all mice blood, indicating that T cell immune reconstitution of nude mice was established successfully. Compared with group NS, others groups could elevate the percentage of human CD3+/CD4+and CD3+/CD8+cells, increase IFN-γ production, inhibit the tumor growth, and prolong survival period of nude mice, especially in group E+P. Conclusion:Anti-tumor immune response could be induced by cord blood-derived dendritic cells loaded with HPV E744₆2peptide and activated by poly â… :C in bearing human cervical cancer nude mice model with immune reconstitution of human T cells.