ABSTRACT
The possibility of identification of cancer stem cells "side population" in solid tumors by using the flow cytometer equipped with 405 nm violet laser was investigated. Ovarian cancer (Skov-3) and colon cancer (Colo 320) cell lines formed the "side population" after vital staining with Hoechst 33342. Analysis of cells isolated from tumor tissue of malignant melanoma and colorectal cancer, also revealed "side population" that was a result of the fluorescent dye exclusion. The percentage of melanoma cells included in the "side population" was the same as that of cells co-expressing cancer stem cells markers--CD34 and CD44. In contrast, the colon cancer "side population" was significantly smaller than the minor populations of colon cancer cells identified by either CD133 expression or exclusion of Rhodamine 123.
Subject(s)
Colorectal Neoplasms/pathology , Flow Cytometry/methods , Neoplastic Stem Cells/pathology , Ovarian Neoplasms/pathology , AC133 Antigen , Antigens, CD/biosynthesis , Antigens, CD34/biosynthesis , Cell Line, Tumor , Colorectal Neoplasms/metabolism , Female , Glycoproteins/biosynthesis , Humans , Hyaluronan Receptors/biosynthesis , Lasers , Male , Neoplasm Proteins/biosynthesis , Neoplastic Stem Cells/metabolism , Ovarian Neoplasms/metabolism , PeptidesSubject(s)
Chromosomes, Human , Chromosomes , Genomic Library , Hybrid Cells , Animals , Humans , MiceABSTRACT
A new model for the generation of specific antitumor cytotoxic T-lymphocytes (CTL) was proposed. In contrast to other models, it allows to generate effector CTL without immunization in vitro. C57BL/10 mice or/and C57BL/6 mice were immunized by injection with gamma-irradiated syngeneic tumor cells into the footpads. For estimation of cytotoxic activity, chromium-51 release assay was used. It has been shown that effector CTL were absent in the lymph nodes in 1-fold as well as 2-fold immunization. Cytotoxic cells have not been found in 1-fold immunization even after maturation of the lymphocytes in monoculture. Specific CTL were detected only after secondary immunization and subsequent cultivation in vitro. Effector cells had Thy1.2+, Lyt2+, L3T4- phenotypes. Presence in vitro of exogenous IL-2 was needed for the generation of CTL against MX-11 sarcoma but not against EL4 lymphoma. We suggest that the release of IL-2 from lymphomas cells could stimulate generation of the effector cells through activation of the endogenous production of IL-2, or due to some other factors.
Subject(s)
Neoplasms, Experimental/immunology , T-Lymphocytes, Cytotoxic , Animals , Antibodies, Monoclonal/immunology , Antigens, Neoplasm , Cells, Cultured , Culture Media , Immunization , Interleukin-2/metabolism , Lymphocytes , Lymphoma, T-Cell/immunology , Male , Mice , Mice, Inbred C57BL , Sarcoma, Experimental/immunology , Splenic Neoplasms/immunology , Tumor Cells, CulturedABSTRACT
The generation of specific antitumor cytotoxic T-lymphocytes (CTL) via 2-fold immunization in vivo and subsequent cultivation without tumor cells (in monoculture) was previously described. The spleen cells from B10 mice bearing progressively growing MX-11 sarcoma suppressed the maturation of CTL specific to MX-11 but not to EL-4 lymphoma in monoculture. It was observed, that the suppression was not the result of the inhibitory effect of suppressor cells upon the IL-2 production, because suppression took place in the presence of the exogenous IL-2 in monoculture. Since the treatment of the spleen cells with MoAb against both L3T4 and Lyt2.2 antigens plus C' considerably decreased the suppressive activity, it was suggested, that two distinct subsets of T-lymphocytes were required for suppression. It might be possible, that the presence of anti-idiotype on the effector suppressors was the cause of the suppressive specificity in the absence of tumor antigens in vitro.
