| Objective: To investigate the proliferation profile and the cytotoxicity against mainfold gastric cancer cells of CIK(cytokine-induced killer) cells from healthy donor in vitro and their anti-tumor affects in Balb/c nude mice bearing OCUM-2MD3 cells strain of ascites tumor from gastric, To investigate variety of content in manifold cytokine and CEA at their anti-tumor affects in vitro and in vivo. Methods1 Generetion of CIK cells: CIK cells were generated as described previously. In brief, nonadherent Ficoll separated human peripheral blood monouclear cells(PBMC) from healthy donors were induced to become CIK cells by multicytokine (CD3McAb,IL-2,IL-1α,IFN-γ)in vitro incubation. PBMC were induced to become CD3AK cells by CD3McAb and IL-2 in vitro incubation. Cells were incubated at 37℃ in a humidified atmosphere of 5% carbon dioxide and subcultured every 3 rd day in fresh complete medium and IL-2 at 3×106 cells ml-1.2 Expanded CIK cells in vitro and its cytotoxicity: Investigate the proliferation profile of CIK cells by TrytanBlue .CIK cultures were analyzed on different time points by fluorescence-activated cell sorter (FACS) and investigate variety of content in manifold cytokine from supernatant fluid by Radioimmuno-detection (RAID) and Enzymelinked immunosorbent assay(ELISA).3 Cytotoxicity of CIK cells were analyzed in vitro by MTT assays, Compared with CD3AK cells . The variety of content manifold cytokine and CEA after added CIK cells were determined in supernatant fluid by RAID and ELISA.4 Balb/c nu/nu female mice, which were 8 wk old, were kept on trimethoprim and sulfamethoxazole, and chlorinated water adlibitum since birth. Aseptic procedures were used routinely. For a dose titration experiment, a total of 32 mice were injected OCUM-2MD3 cells strain of ascites tumor (5×106 /0.2ml/mice) in by abdominocentesis from human gastric, Tumor cells were in log phase of growth when injected. After 4 weeks, 12 mice received 4×107 /0.2ml CIK cells, 10 mice received 4×107 /0.2ml CD3AK cells,10 mice received NS 0.2ml as comparison . In each group were added every 1 day, total 3.After 7 days, blood and ascites were removed, the variety of content manifold cytokine and CEA after abdominocentesis were determined in serum and ascites by RAID and ELISA. Results: 1 CIK cells were generated by culturing PBMC in the presence of IFN-γon day 0 and CD3McAb,IL-2, IL-1αonday 1, CIK cells were expanded 115-fold after 21 days .2 The flow cytometry analysis showed that CD3+CD56+ T cells which were the major cytolytic effectors in CIK cells expanded 1155-fold after 2 weeks in CIK culture; we detected the amount of IFN-γ, IL-2, TNF-α,GM-CSF increased in supernatant fluid after CIK cultures by RAID and ELISA ,highest amounts were in 2 weeks .3 CIK cells had greater cytotoxicity against mainfold gastric tumor cells in vitro than that of CD3AK cells (P<0.05t) and are non-major histocompatibility complex-restricted . IL-2 of content in supernatant fluid distinctness reduced disincrease on the CIK cells at OCUM-2MD3 cells in vitro (P<0.05t) and which content of IFN-γ, TNF-α,GM-CSF in supernatant fluid were not patency(P>0.05t); content of CEA in super-natant fluid distinctness reduced on the CIK cells at OCUM-2MD3 cells in vitro (P<0.05t).4 In addition, CIK cells had a stronger suppressive effect on the tumor growth in Balb/c nude mice bearing OCUM-2MD3 tumor, compared with CD3AK cells in vivo (P<0.05t);The content of manifold cytokine distinctness increase in serum and ascites on CIK group, compared with NS group(P<0.05t); The content of CEA distinctness increase in serum and ascites on CIK group, compared with NS group(P<0.01t). Conclusion: 1 CIK cells were neotype highly efficient cytolytic effector cells which have a stronger expanded.2 The FCAS analysis showed that CD3+CD56+ T cells and CD8+ which were the major cytolytic effectors in CIK cells 3 CIK cells had significant suppression cytotoxicity against mainfold growth of carcinoma of stomach...
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