Published online Oct 15, 2001. doi: 10.3748/wjg.v7.i5.690
Revised: August 6, 2001
Accepted: August 12, 2001
Published online: October 15, 2001
AIM: To prepare a cancer vaccine (H22-DC) expressing high levels of costimulatory molecules based on fusions of hepatocarcinoma cells (H22) with dendritic cells (DC) of mice and to analyze the biological characteristics and induction of specific CTL activity of H22-DC.
METHODS: DCs were isolated from murine spleen by metrizamide density gradient centrifugation, purified based on its characteristics of semi-adhesion to culture plates and FcR-, and were cultured in the medium containing GM-CSF and IL-4. A large number of DC were harvested. DCs were then fused with H22 cells by PEG and the fusion cells were marked with CD11c MicroBeads. The H22-DC was sorted with Mimi MACS sorter. The techniques of cell culture, immunocytochemistry and light microscopy were also used to test the characteristics of growth and morphology of H22-DC in vitro. As the immunogen, H22-DC was inoculated subcutaneously into the right armpit of BALB/C mice, and their tumorigenicity in vivo was observed. MTT was used to test the CTL activity of murine spleen in vitro.
RESULTS: DC cells isolated and generated were CD11c+ cells with irregular shape, and highly expressed CD80, CD86 and CD54 molecules. H22 cells were CD11c- cells with spherical shape and bigger volume, and did not express CD80, CD86 and CD54 molecules. H22-DC was CD11c+ cells with bigger volume, being spherical, flat or irregular in shape, and highly expressed CD80, CD86 and CD54 molecules, too. H22-DC was able to divide and proliferate in vitro, but its activity of proliferation was significantly decreased as compared with H22 cells and its growth curve was flatter than H22 cells. After subcutaneous inoculation over 60 d, H22-DC showed no tumorigenecity in mice, which was significantly different from control groups (P < 0.01). The spleen CTL activity against H22 cells in mice implanted with fresh H22-DC was significantly higher than control groups (P < 0.01).
CONCLUSION: H22-DC could significantly stimulate the specific CTL activity of murine spleen, which suggests that the fusion cells have already obtained the function of antigen presenting of parental DC and could present H22 specific antigen which has not been identified yet, and H22-DC could induce antitumor immune response; although simply mixed H22 cells with DC could stimulate the specific CTL activity which could inhibit the growth of tumor in some degree, it could not prevent the generation of tumor. It shows that the DC vaccine is likely to become a helpful approach in immunotherapy of hepatocarcinoma.