[The comparison of inducing specific CTL by two methods of leukemia cell antigen loading of DCs and fusion of DCs with leukemia cells].

AIM: To compare the potency of the fusion of DCs with leukemia cells and freeze-thawed leukemia antigen-loading DCs in inducing antigen-specific CTLs.

METHODS: Peripheral blood mononuclear cells (PBMC) isolated by HES-Ficoll two-step method were incubated 2 hours to select adherent monocytes. The adherent monocytes were then cultured in the presence of GM-CSF and IL-4 for 5 days and then divided into four groups: DCs were fused with K562 cells or CML cells from CML patients in the presence of 500 g/L PEG-100 mL/L DMSO (Group A), DCs were loaded with lysates from K562 cells or CML cells (group B), DCs were co-cultured with K562 cells or CML cells (group C) and DCs were cultured alone (group D). Before cell fusion, K562 cells were labeled using a red fluorescent dye, PKH26. After fusion, flow cytometry was used to detect hybrids labeled by both PKH26 and FITC-conjugated anti-HLA-ABC mAb to assess the efficiency of fusion. On day 6, TNF-alpha was added to induce the terminal maturation of DCs. Then each group DCs were co-cultured with autologous T cells respectively. The cytotoxicity of CTLs of each group against different target cells was measured using MTT coloremetry.

RESULTS: DCs induced by GM-CSF+IL-4 and TNF-alpha had DC-classical phenotypic characteristics. The efficiency of cell fusion ranged from 17.33% to 29.94%. Both DC-K562 hybrids and DC loaded with leukemic freeze-thaw lysates could stimulate specific CTL responses against target cells. Furthermore, the cytotoxicity induced by the former DCs were much stronger than that induced by lysates-loaded DCs at the same E/T ratio.

CONCLUSION: Compared with freeze-thaw lysates loaded DCs, DC-leukemic hybrids showed higher efficiency in antigen presentation and the stimulation of leukemia-specific CTLs.