| There are many outstanding issues in cancer immunotherapy. LAK, TIL, CIK, et al are all adoptive immunotherapy and have certain effect. Debdritic cells (DC) are considered the most potent antigen-presenting cells (APC) and are thus promising new tools for the immunotherapy of cancer. DC can stimulate the primary activation of T cells due to their enhanced capacity of presenting immunogenic peptides in association with self-major histocompatibility complex I and II molecules. DC can also process both exogenous proteins and intracellular protein to T cells and can directly modulate B-cell growth and differentiation. To use DC as "nature's adjuvants" for the potentiation of immune reactivity against tumors, these cells are modified by pulsing them with tumor lysates, tumor proteins, tumor peptides, or are transfected with cDNA encoding tumor antigens. Various studies in animal models have clearly shown that DC pulsed with tumor antigens in vitro and then re-injected in vivo induce immune responses thatlead either to protection against lethal tumor challenge or to regression of established tumors. Clinical trials with modified DC have confirmed the value of this approach. This might hold true especially in the treatment of minimal residual disease after therapy for primary tumors.We obtain DC from PBMC stimulated by GM-CSF, IL-4, TNF- a and PGE, in vitro. High potent and specific anti-tumor effect is induced using DC and high agglomerative staphylococcin (HAS), a kind of superantigen.1. In vitro generation of DCPBMC were isolated from leukocyte-enriched buffy coats by standard density gradient centrifugation on Ficoll-Paque, resuspended in complete RPMI 1640 medium. PBMC were allowed to adhere in cell culture flasks. Nonadherent cells were removed and adherent cells were cultured in medium containing GM-CSF and IL-4. After 5-6 day of culture, cells were washed and recultured in medium containing TNF- a and PGE2. Cells were analyzed for surface antigen by FACS. The cultured cells exhibit the typical morphological features of DC and express high levels of HLA- I ,HLA- II and CD45 molecules. Positive rates of CD la and CD83 are 71% and 50%, respectively.2. Effector cells culture and cytotoxicity assay in vitroNonadherent PBMC were resuspended in complete medium supplemented with HAS and cultured at 37 癈 and 5% CO2(named HASL). Culture medium were exchanged every 4 days. GLC-82 antigen pulsed DC and HASL were cocultured and the obtained cells were named DC-HASL. DC-L was DC cocultured with lymphocytes. Cytotoxicity of HASL, DC-L and DC-HASL were determined by MTT assay. In brief, effector cells(HASL, DC-L, DC-HASL) and target cells(GLC-82, A549,moser and MCF-7) were seeded in 96-well microtiter plates at an appropriate ratio. Cytotoxic rates of7DC-HASL to GLC-82 cells reached 100% and the cytotoxic rates to other cancer cell lines A549, moser and MCF-7 were 94.2%, 73.5% and 71.2%, respectively; Cytotoxic rates of HASL and DC-L to the above four cell lines were 48%, 63%, 60%, 45% and 46.4%, 42.1%,39%,25.6%, respectively. 3. Growth inhibition test of lung cancer nude mice modelEvery nude mouse was inoculated with 5 X 105 GLC-82 cells at thigh. All mice were divided into 4 groups:(1)control group,(2)DC-HASL therapy group, (3)DC-L therapy group,4HASL therapy group.Nude mice in 3 therapy groups were injected 1 X 107 effector cells on day 4,10,15 after tumor inoculation, respectively. Nude mice were killed on day 20 and tumor tissues were resected. The tumor volume of four groups were 298.40?7.58,18.5?14.2,43.15?9.44 and 70.67?0.51 mm3, respectively. Tumor inhibition rates of 3 therapy groups compared with control group is 93.8%, 85.5% and 76.3%, respectively.Our research showed lymphocyte can proliferate to a certain extent stimulated by HAS in vitro and HASL can lyse tumor cells. High potent and specific antitumor immune response can be induced when combining DC with HASL.We combine DC mediated tumor active immunotherapy with super antigen mediated immunotherapy in our research...
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