Studying in vitro phagocytosis of apoptotic cancer cells by recombinant GMCSF-treated RAW 264.7 macrophages (original) (raw)
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A potential role of macrophage activation in the treatment of cancer
Critical Reviews in Oncology/Hematology, 2002
One of the functions of macrophages is to provide a defense mechanism against tumor cells. In the last decades the mechanism of tumor cell killing by macrophages have been studied extensively. The tumor cytotoxic function of macrophages requires stimulation either with bacterial cell wall products such as lipopolysaccharide (LPS) or muramyldipeptide (MDP) or with cytokines such as interferon-g (IFN-g) and granulocyte-macrophage colony-stimulating factor (GM-CSF). Activated macrophages secrete several substances that are directly involved in tumor cell killing i.e. tumor necrosis factor (TNF) and nitric oxide (NO). On the other hand, substances are secreted that are able to stimulate tumor cell growth, depending on the stage and the nature of the tumor. Several clinical trials have been performed aiming at the activation of macrophages or dendritic cells, a subpopulation of the macrophages. In this review we will summarize and discuss experimental studies and clinical trials based on the activation of macrophages.
Macrophage activation and anti-tumor activity of biologic and synthetic agents
International Journal of Cancer, 1976
Systemic administration of the synthetic immunopotentiator pyran, was as effective as the use of the biologic immunopotentiator BCG in activating macrophages and in inhibiting the Lewis lung carcinoma and MCA 2182 sarcoma. Several other synthetic polyanions also activated macrophages and exhibited some anti-tumor activity, but none were as effectives as pyran. Cell-wall fractions such as the Ribi vaccine and MER were considerably less effective than BCG. The anti-tumor activity of pyran against the virtually non-immunogenic Lewis lung carcinoma involved non-specifically activated macrophages, and both anti-tumor activity and macrophage activating ability persisted over a 100-fold range of drug from 0.5 mg/kg to 50 mg/kg. The ability of activated macrophages to destroy tumor cells was abrogated by treatment with trypan blue, an inhibitor of macrophage lysosomal enzymes. In addition, preincubation of tumor cells with activated peritoneal cells at effector-cell:target-cell ratios of 20:1 and 5:1 markedly decreased tumor incidence and mortality. Glycogen-stimulated or unstimulated peritoneal cells were completely inactive in inhibiting tumor growth in vivo or exhibiting cytotoxicity in vitro, demonstrating the requirement for activated macrophages selective for tumor-cell destruction.
Biological and antitumor effects of recombinant human macrophage colony-stimulating factor in mice
Cancer research, 1991
The administration of recombinant human macrophage colony-stimulating factor (M-CSF) i.p. in doses of 25 or 100 micrograms twice daily for 5 consecutive days to non-tumor-bearing C57BL/6 mice resulted in a dose-dependent infiltration of mononuclear cells in the livers but not the lungs of these treated animals. Immunohistochemical examination of fixed liver tissue with the murine macrophage-specific monoclonal antibody, F4/80, revealed a greater than 5-fold increase in the number of hepatic macrophages. Quantification of F4/80-positive cells in a mononuclear single cell suspension derived from liver revealed a greater than 25-fold expansion in the number of hepatic macrophages compared to control mice. These cells were then tested in 18-h 51Cr release assays for tumoricidal activity, after an 18-h incubation with or without gamma-interferon, against cultured P815 targets. Significant tumor cell lysis by these liver-associated mononuclear cells occurred, which was enhanced by gamma-i...
International Journal of Cancer, 1986
Macrophage-mediated cytolysis of thymidine-prelabelled murine A9 fibrosarcoma cells was compared to the level of cytolytic factor (CF) present in the cultures by assaying supernatant aliquots on actinomycin (AcD)-treated A9 fibrosarcoma cells. A good correlation between the level of A9 killing and CF titer was observed when different concentrations of lipopolysaccharide (LPS) were added to various macrophage populations: murine peritoneal cells, short-term bonemarrow (BM)-derived macrophages and JEM@ macrophage lines. Optimal A9 killing and CF secretion, equivalent to the killing of about 1000 AcD-pretreated A9 cells by a single macrophage, were obtained following activation of JEM@ by LPS. CF production by EM-derived macrophages was enhanced in serum-free medium when compared to its release in the presence of fetal calf serum. The LPS-activated macrophages could be restimulated by the activating agent to produce CF following one week of propagation in the absence of LPS. On the other hand, CF activity was absent from the supernatants of activated macrophages co-cultured with normal embryonic fibroblasts, which are resistant to macrophage-mediated killing. This effect could be attributed to a factor, secreted by normal fibroblasts but not by A9 cells, which suppressed CF release from the activated macrophages. Our data strongly support earlier observations, suggesting that CF [which appears t o resemble the tumor necrosis factor (TNF)] is responsible for LPS-induced macrophage-mediated tumor cell lysis. It is suggested that suppression of the latter process by the fibroblast-derived factor proceeds via inhibition of CF/TNF production from the macrophage.
Effect of colony stimulating factor on murine macrophages. Induction of antitumor activity
Journal of Clinical Investigation, 1982
stimulating factor (CSF) was assessed for its capacity to stimulate antitumor activity in macrophages. Murine peritoneal macrophages incubated with CSF for 48 h inhibited [3H]thymidine (TdR) incorporation by P815 tumor cells to-20% of control values. Inhibition of CSF-stimulated macrophages was significantly greater than inhibition by unstimulated macrophages (P < 0.001). CSF had little direct effect on the proliferation of either tumor cells or macrophages alone, indicating that the antitumor activity of CSF was mediated by macrophages. It is unlikely that impurities in the CSF preparations were responsible for the effect since CSF that had been purified to homogeneity was as active as crude preparations. Furthermore the activity of CSF on macrophages was blocked by addition of purified anti-CSF antibodies. In addition to being tumoristatic, CSFstimulated macrophages were tumoricidal as determined by a tumor colony growth assay. Tumor cells that had been incubated with CSF-stimulated macrophages showed a significant reduction in tumor colony-forming units (P < 0.01). Thus, in addition to its effect on hemopoietic stem cells, CSF induces certain effector functions in mature macrophages that may enhance endogenous antitumor host defenses. Address reprint requests to Dr. Edward J. Wing.
Journal of Leukocyte Biology, 2000
The clearance of apoptotic cells is crucial to avoid chronic inflammation and autoimmunity. Little is known about the factors that regulate it in vivo. We show that granulocyte-macrophage colony-stimulating factor (GM-CSF) administration to carcinoma patients confers to their leukocytes a significantly higher ability to phagocytose apoptotic cells than before (P < 0.005). GM-CSF increased the concentration of monocytes and polymorphonuclear leukocytes in the peripheral blood and activated circulating polymorphonuclear leukocytes. Both effects abated early after treatment, whereas phagocytosis of apoptotic cells was still significantly higher after 18 days compared with basal values (P < 0.005 and P < 0.025 for monocytes and polymorphonuclear leukocytes, respectively). On in vitro phagocytosis of apoptotic cells monocytes, but not polymorphonuclear leukocytes, up-regulated MHC class II membrane expression. These findings are consistent with the possibility that GM-CSF endows...
1988
Bone marrow-derived cells from C3H/HeJ mice were cultured in the presence of recombinant murlne granulocyte-macrophage colony-stimulating factor (rGM-CSF) or highly purified murine macrophage colony-stimulating factor (CSF-l) for 7 days. Following this 7-day culture period, mature macrophages were harvested and replated at precise densities in the absence of exogenous rGM-CSF or CSF.l, and assayed in a two-signal tumoricidal assay. Cultures were stimulated with medium only or with combinations of recombinant lnterferon-y (rIFN-.y) as the "priming" signal, and/or butanol-extracted lipopolysaccharlde (But-LPS) as the "triggering" signal for 24 hr. At this time, 51Crlabeled, P815 tumor target cells were added, and the percent tumor cell cytotoxicity was determined after 16 hr. Macrophages derived under the influence of rGM-SF exhibited significant tumoricidal capacity with medium alone (16 ± 5%). The addition of "priming" signal only (i.e., rIFN-y, 10.0 U/mI) significantly increased tumoricidal capacity to 31 ± 9%. Treatment with But-LPS alone did not alter the basal tumoricidal activity of rGM-CSF-derived macrophages. Combinations of rlFN-(10.0 U/mI) and But-LPS (0.5-5.0 g/ ml) generated highly tumoricidal macrophages (50-60% tumor cell cytotoxicity). In contrast, medium-treated CSF-l-derived macrophages exhibited a significantly lower basal level of tumor cytotoxicity (6 ± 3%). Unlike rGM-CSF-derived macrophages, treatment of CSF-1-derived macrophages with high concentrations of rlFN-y alone did not increase significantly the level of cytotoxicity above that of medium-treated cultures. However, CSF-1-derived macrophages responded to the highest concentrations of But-LPS (5.0 g/ml) to increase tumoricidal activIty from 6 ± 3% to 17 ± 5%. OptImal tumoricldal activity (44 ± 17%) was observed when CSF-1-derived macrophages were treated simultaneously with high concentrations of both r1FN-y and But-LPS. Thus, macrophages derived from bone marrow progenitors in either rGM-CSF or CSF-1 exhibited tumoricidal capacities that differed in basal activity as well as in their requirements for and sensitivities to "priming" and "triggering" signals.
Cellular Immunology, 1978
MCF has been shown to be produced by CON A stimulated lymphoblasts. Its activity was tested and titrated on lymphocyte-free bone-marrow macrophages. The cytotoxic effect of activated macrophages on various normal and malignant cells has been tested. For that purpose bone-marrow macrophages from a 9-day old culture, which are never preactivated, were activated with 100 pl of a titrated MCF preparation. Thus the effector macrophages were of an identical stage of maturation and activation throughout the experiments. The data show that both normal and malignant cells are affected by cytotoxic macrophages. However the degree of cytotoxicity differs markedly among the various cell types tested : Fibroblasts and mature macrophages are resistant, whereas lymphocytes and blast cells of various origin are susceptible to the cytotoxic action of activated macrophages. By far the highest susceptibility has been observed with tumor target cells.