Résumé
Objective: To compare the cytotoxic activity of mammaglobin A-specific CD8+ cytotoxic T lymphocytes (CTLs) and cytokine-induced killers (CIKs) against breast cancer cells in vitro. Methods: PBMCs were isolated in vitro from the peripheral blood of healthy volunteers. Then DCs, CTLs and CIKs were isolated, induced and cultured from PBMCs in vitro. CD8+ CTLs were purified with immunomagnetic beads from CTLs. DCs were infected with recombinant adenovirus encoding mammaglobin A(Ad-MGBA). CTLs and CIKs were co-cultured with DCs being infected with Ad-MGBA. The cytotoxic activity of mammaglobin A-specific CD8+ CTLs and CIKs against breast cancer cells was compared by flow cytometry. Results: The apoptosis rate of breast cancer cell MDA-MB-415, which expressed MGBA was 63.07% by CD8+ CTL killing and 48.35% by CIK killing (P<0.05). The apoptosis rate of breast cancer cell MDA-MB-231, which could express MGBA was 14.62% by CD8+CTL killing and 29.29% by CIK killing (P<0.05). Conclusion: The cytotoxic activity of antigen-specific CD8+ CTLs for the same antigen-expressing oncology cells is higher than CIKs. However, for different antigen-expressing oncology cells, the cytotoxic activity of CD8+CTLs is lower than that of CIKs.
Résumé
Objective: To investigate the effect of RetroNectin on CIKs cells and the related mechanisms. Methods: Peripheral blood mononuclear cells (PBMC) were collected from patients and divided into two groups: group Ⅰ and group Ⅱ. Samples in group Ⅰ were seeded into culture flask precoated with RetreNec-tin and CD3mAb to induce CIKs. While samples in group Ⅱ were seeded into common culture flask. The pro-liferation of CIKs was detected by cytometric analysis. The cytotoxic activity of CIKs was determined by LDH assays. The phenotype changes and cell cycle of CIKs were identified by flow cytometry. The apoptosis of cells was detected by Annexin V/PI. Western blot was employed to detect the level of protein Vav1. The CD49d and CD49e were blocked by anti-CD49d and anti-CD49e and the proliferation of cells was tested by cytometric analysis after the blockage. The phenotype changes of cells were identified by flow cytometry after the blockage. Results: RetroNectin enhanced the proliferation of CIKs (P<0.05). Flow cytometric analysis showed that RetroNectin significantly increased the number of CD25+ T cells (P<0.05). RN-CIK was more ac-tive than CIK in killing HCT-8 cell lines in vitro (P<0.05). RetroNectin could block the CIKs at G_1 phase (P<0.05) and resist apoptosis. There was no significant difference in the proliferation between the two groups af-ter the blockage with CD49d and CD49e (P>0.05). The expression of protein Vavl was associated with CD25+T cells. Conclusion: RetroNectin enhances the proliferation of CIKs by influencing the cell cycle, resist-ing apoptosis possibly through the site of CD49d and CD49e, and inducing T cell activation as the second sig-naling through Vav1.