The role of adrenocorticoids as modulators of immune function in health and disease: neural, endocrine and immune interactions.
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Adrenal Cortex Hormones↗
Biomedical subjects
K W Brunson
Publications and source records attributed to K W Brunson.
Enhanced immunosuppressive activity associated with metastatic lymphoma cells.
Earlier reports from our laboratory showed that Abelson virus-induced, highly malignant and liver metastatic RAW117-H10 cells, but not the parental, less metastatic RAW117-P cells, inhibited both T-cell and B-cell mitogen-induced proliferation of syngeneic normal murine spleen cells. Similar inhibition was also noted when RAW117-H10 cell surface molecules extracted with butanol were used instead of whole tumor cells. In this report we describe the suppressive properties of the butanol-extracted RAW117-H10 cell surface molecules on other immune functions and the isolation/purification of a molecule from RAW117-H10 cell butanol extract which shows inhibitory activity. The immunosuppressive molecules also inhibit natural killer cell-mediated cytotoxicity, lymphokine-activated killer cell-mediated cytotoxicity, and bone marrow colony-forming unit-granulocyte-macrophage colony formation, but not colony-forming unit-fibroblast colony formation. The suppressive molecules inhibit interleukin 2 production by the T-lymphocytes. One of the molecules responsible for some of the immunosuppressive activity has been isolated and purified from butanol extracts of the metastatic RAW117-H10 cells by preparative isoelectrofocusing techniques. The suppressive molecule has an isoelectric point of 4.3 with an approximate molecular weight of 70,000. Metastatic RAW117-H10 lymphoma cells therefore express immunosuppressive molecules, which may facilitate their growth and metastasis in vivo.
Animals↗
Therapeutic agents for treatment of established metastases and inhibitors of metastatic spread: preclinical and clinical progress.
This article reviews recently described agents for the treatment of established metastases or inhibitors of metastatic spread. Recent progress in both preclinical models of experimental metastasis and clinical evaluation is highlighted. Where possible, distinctions are made between therapeutic agents for established metastatic tumors and those that merely interfere with the process of metastasis. The approaches emphasized include radiotherapy, cytotoxic chemotherapy, targeting the metastatic process and tumor metastases (adhesion, invasion, and angiogenesis), and modulation of the immune response, including chemoimmunotherapy. Emphasis has been placed on the necessity of using appropriate animal models of metastatic disease for the discovery and development of novel and effective antimetastatic therapeutic agents. Also addressed is the likelihood that future approaches to the metastasis problem will employ novel combined antimetastatic therapeutic modalities.
Adjuvants, Immunologic↗
Differential cell surface antigen expression on metastatic variant lymphoma cell lines.
In the present investigation we have studied the cell surface antigenicity of a syngeneic murine metastatic lymphoma model system. This system is comprised of a highly malignant and metastatic RAW117-H10 subline and the less malignant/metastatic parental RAW117-P cell line. Using rabbit antisera raised against whole tumor cells we have been able to identify two major glycoprotein antigens which are differentially expressed on the cells. Although these antigens have a similar molecular weight (70 kD), they are antigenically distinct as determined by in vitro cytotoxicity assays. The levels of glycosylation on these glycoproteins were also found to be different. Increased expression of one of the antigenic components on the metastatic RAW117-H10 cells appeared to be associated with metastasis in this tumor model system.
Antigens, Neoplasm↗
Regulated expression of the human mutant ras gene after transfection of BALB/c mouse embryo fibroblast cells.
Four continuous cell lines constructed by transfecting BALB/c mouse fibroblast cells with an expression system that has the mutant c-Ha-ras gene under control of a truncated version of the mouse metallothionein-1 (mt-1) promoter were characterized for zinc-induced phenotype switching. These cells were selected for transformation in the presence of zinc, a known inducer of the mt-1 promoter. When the transfected cells were grown in medium depleted of zinc, there was a dramatic reduction in their soft agar cloning efficiency. Adding zinc back to the medium restored the transformed phenotype in a dose-dependent manner. Analysis of the intracellular p21 levels of induced and uninduced cells showed that zinc was modulating the expression of the transfected ras gene. In vivo studies done with syngeneic mice showed that zinc-induced cells were tumorigenic and formed metastatic lesions in the lungs of the inoculated animals.
Animals↗
Effects of differentiation inducing chemicals on in vivo malignancy and NK susceptibility of metastatic lymphoma cells.
In the present investigation we have determined the effects of the differentiation-inducing chemicals dimethyl sulfoxide (DMSO) and sodium butyrate on the growth and tumorigenicity of a highly malignant/metastatic cell line (RAW117-H10) and its less tumorigenic parental lymphoma cell line (RAW117-P). Both of these agents at doses shown to be nontoxic slowed the growth of these cells in suspension culture and significantly lengthened the doubling time while reducing colony formation in the agar tumor stem cell assay. Corresponding to these observations, the in vivo tumorigenicity of the highly malignant RAW117-H10 line was reduced by both chemicals but particularly by butyrate treatment. In addition, both agents increased the expression of some glycoproteins and the glycolipid asialo GM1. There was also a corresponding increase in the NK susceptibility of the normally NK resistant RAW117-H10 cells. To determine if the decreased malignancy of the highly malignant cells following treatment with the chemical agents was primarily due to the alteration in cell surface glycoconjugates or merely due to concomitant decreased growth potential, we transplanted highly glycosylated membrane fragments from normal syngeneic thymocytes to both RAW117-P and RAW117-H10 cells using a Sendai virus mediated membrane fusion technique. The in vivo tumorigenicity of the membrane altered RAW117-H10 cells was significantly decreased. These results strongly suggest that the decreased in vivo malignancy of RAW117-H10 cells resulting from treatment with chemical differentiation agents is caused by their increased susceptibility to NK cell mediated lysis which in turn results from cell surface changes involving altered, primarily increased, expression of certain glycosylated surface molecules. The cell surface glycoconjugates, such as receptors for certain lectins and glycolipid asialo GM1, can be used as markers for malignant potential and NK sensitivity of malignant lymphoid cells.
Animals↗
Cell surface properties associated with malignancy of metastatic large cell lymphoma cells.
The highly malignant and metastatic RAW117-H10 cell line was developed by in vivo selection from the Abelson leukemia virus induced parental RAW117-P lymphoma. In this study we have characterized these cell lines with regard to their expression of lymphocyte and macrophage differentiation antigens, adherence, phagocytic properties, binding of various lectins, binding of antibodies to glycolipid asialo-monoganglioside, and the role of butanol extractable cell surface molecules to determine if any of these cell surface properties are associated with the malignant potential of RAW117-H10 cells. The only major difference in immunological phenotypes between RAW117-P and RAW117-H10 cells was an increased expression of Thy-1 molecules by the latter. However, the highly malignant RAW117-H10 cells bound significantly less concanavalin A, Ricinia communis agglutinin, succinylated wheat germ agglutinin, and particularly anti-asialomonoganglioside than their parental counterpart and were resistant to natural killer cell mediated cytolysis. Removal of butanol extractable cell surface molecules significantly decreased the malignancy of RAW117-H10 cells and increased their susceptibility to natural killer cell mediated cytolysis. The butanol treated RAW117-H10 cells regained high in vivo malignancy when recultured for 3 days to permit regeneration of their cell surface components. The butanol extracted RAW117-H10 cells still expressed high levels of Thy-1 indicating that this most probably represented "inappropriate" antigen expression. Since the expression of lymphocyte differentiation antigens did not correlate with the malignant behavior of the cells, we postulate that these antigenic differences merely represent phenotypic variation. The decreased malignant potential of the butanol treated RAW117-H10 cells did correlate with increased cell surface anti-asialomonoganglioside binding (glycolipid) and increased natural killer cell susceptibility.
Animals↗
Differential growth characteristics of low and high metastatic variant RAW117 murine lymphosarcoma cells.
The Abelson-virus-induced murine lymphosarcoma cell line RAW117-P and its in vivo selected highly malignant and metastatic variant RAW117-H10 have been studied for their growth in vivo, in vitro and in semi-solid agar. The highly malignant metastatic variant RAW117-H10 cells killed syngeneic Balb/c mice rapidly and formed 100-200 times more gross liver tumor nodules than the less malignant parental RAW117-P cells. On the other hand, there were no differences between the in vitro growth kinetics of these cells as measured by various parameters such as metaphase arrest or cells in DNA synthetic phase on various days after the initiation of the culture. These cells could be grown in semi-solid agar and formed characteristic colonies. The parental cells formed many very small colonies, whereas the highly malignant cells formed fewer, but very large colonies in soft agar. These results suggest that differential interactions of highly malignant RAW117-H10 cells with the host, particularly the host immune system, are much more important for regulating the number of metastases than any intrinsic growth advantages. It appears that growth differences, potentially based on lack of response to feedback inhibition rather than on kinetic parameters, are responsible for highly malignant RAW117-H10 cells forming larger colonies both in agar in vitro and in the liver in vivo.
Animals↗
Cytotoxicity of rabbit macrophage peptides MCP-1 and MCP-2 for mouse tumor cells.
The cytotoxicity of cationic peptides MCP-1 and MCP-2 isolated from rabbit alveolar macrophages was tested against two tumor cell lines of murine lymphosarcoma origin, RAW117-P and RAW117-H10, and a normal mouse connective tissue fibroblast strain, ATCC CCL1.RAW117-H10 is a highly malignant metastatic variant derived from the less malignant RAW117-P. Our results indicate that these peptides possess a cytotoxic activity against the tumor cells tested but not against the normal cells tested. At concentrations of 30 micrograms/ml, these peptides completely killed RAW117-H10 cells in suspension cultures, while inhibiting growth of RAW117-P for but a limited period of time, up to 48 h, after which growth resumed. RAW117-P cells were killed by concentrations of 40 micrograms/ml. These peptides showed little cytotoxicity for normal mouse connective tissue fibroblasts at concentrations of 40 micrograms/ml.
Animals↗
The effects of 5-bromodeoxyuridine on cyclic AMP levels and cytoskeletal organization in malignant melanoma cells.
5-Bromodeoxyuridine (BUdR) causes mouse melanoma cells to develop a flattened morphology and simultaneously adhere tenaceously to the substratum on which they are growing. Experiments were done to determine if these events are coupled to increases in cAMP levels and to rearrangements in the cells' cytoskeleton. Cyclic AMP assays revealed that cell flattening and the increase in adhesive properties caused by BUdR is not accompanied by an increase in the cellular concentration of cyclic AMP. However, electron micrographs of cells grown in the presence of BUdR show a striking increase in the number of organized microtubules and microfilaments. Colchicine binding revealed no difference in the amount of tubulin present in untreated or BUdR-treated cells indicating that the increase in the number of microtubules is due to the polymerization of pre-existing tubulin subunits. These results are discussed in light of possible similar mechanisms of action of BUdR and cyclic AMP in regulating the organization of microtubules and microfilaments and the role these structures play in altering cell morphology and adhesive properties.
Animals↗
Malignancies of metastatic murine lymphosarcoma cell lines and clones correlate with decreased cell surface display of RNA tumor virus envelope glycoprotein gp70.
Variant sublines of the murine lymphosarcoma RAW117 have been derived by sequential cycles of intravenous inoculation of cells and harvesting of solid liver tumors in syngeneic BALB/c mice [Brunson K. W. & Nicolson, G. L. (1978) J. Natl. Cancer Inst. 61, 1499-1503] and also by sequential removal of lectin-reactive cells via repeated adsorption on immobilized-lectins [Reading, C. L. Belloni, P. N. & Nicolson, G. L. (1980) J. Natl. Cancer Inst. 64, 1241-1249]. These cell sublines and their clones were analyzed for abilities to form gross liver tumor metastases after injection intravenously or subcutaneously into syngeneic mice, and this response was related to certain cell surface properties including quantities of viral antigens and lectin-binding sites, exposure of specific cell surface proteins, and quantities of cell surface glycoproteins visualized in gels with 125I-labeled lectins or antibodies. Consistent differences were obtained between RAW117 sublines of low and high malignancy with respect to the amounts or exposures of cell surface glycoprotein components of Mr approximately 70,000 or 69,000 and 71,000, depending on the gel system. Competition radioimmunoassays for RNA tumor virus antigens in the RAW117 lines and clones indicated the presence of Moloney murine leukemic virus antigens gp70, p30, and p12. Enhanced malignancy and metastasis to liver was accompanied by decreases in the cellular contents of viral antigens and loss of gp70 cell surface exposure. Analysis of several clones obtained from sublines selected in vivo and in vitro for high or low malignancy confirmed the inverse relationship between metastasis and expression of gp70 in this system.
Animals↗
Selection of malignant melanoma variant cell lines for ovary colonization.
Murine melanoma line B16-F1, which shows some specificity for metastatic organ colonization of lung but rarely metastasizes to ovary, was used to select variant cell lines with increased preference for experimental ovary metastasis. Ovary-colonizing melanoma cell lines were sequentially selected in syngeneic C57BL/6 mice by repeated intravenous administration and surgical recovery of ovarian melanoma tumors for tissue culture. After ten selections for experimental ovary metastasis, line B16-010 was established which formed experimental metastatic ovary tumors in almost every test animal. In tissue culture B16-010 cells grew in circular colonies with rounded, smooth cell peripheries compared to B16-F1 cells which were flatter, grew in irregular patterns, and exhibited long cellular projections. Ovary-selected B16 lines contained less melanin pigment (B16-010 less than B16-05 less than B16-01 approximately equal to B16-F1) compared to the parental melanoma line. Together with previous cloning and selection data, these results are consistent with the preexistence of highly malignant cells in the parental tumor population that possess the ability to metastasize to specific organs.
Animals↗
Selection and altered properties of brain-colonising metastatic melanoma.
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Animals↗
Lipopolysaccharide effects on sensitive and resistant variant Chinese hamster ovary cell lines.
Chinese hamster ovary (CHO . K1 . PRO) cell growth was inhibited by addition of a gram-negative bacterial lipopolysaccharide (LPS) to the cell culture medium. Growth inhibition began after three or four days of incubation, was dose-dependent up to a maximum at an LPS concentration of 500 microgram/ml and was accompanied by cell shape changes and enhanced cytoplasmic vacuolization. Formation of bizarre CHO . K1 . PRO cell shapes and vacuole formation were most pronounced after seven days of incubation with LPS and could be observed by light and electron microscopy. An LPS-resistant cell population was obtained by intermittent in vitro exposure to high levels of LPS; these variant cells or clones derived from them failed to display growth inhibition in the presence of LPS. A clone from the LPS-resistant variant population showed altered cell properties compared to the parental cell line which included changes in cell morphology, adhesion, and endocytosis. Parental cells was markedly density-inhibited, whereas the cariant clone exhibited considerable growth after confluency. The LPS-resistant variant cells showed a more elongated morphology than the parental line. No significant differences were observed between rates of detachment of parental and variant cells when sparse cultures of either line were removed from tissue culture dishes by ethylenediaminetetracetate (EDTA). However, at confluency approximately 100% of the variant cells versus 35% of the parental cells were removed by EDTA in one hour. Measurements of 125I-ferritin uptake by parental and variant cells showed approximately twenty-fold and twofold increases, respectively, in uptake induced by LPS when compared to untreated control cultures.
Animals↗
Selection and biologic properties of malignant variants of a murine lymphosarcoma.
A syngeneic tumor model system with the RAW117 lymphosarcoma cell line was developed for use in investigations of host and tumor cell properties associated with an enhanced state of malignancy. This BALB/c mouse model was found to be similar to human lymphosarcoma in that liver and spleen were the major organs involved. Sequential in vivo selections were performed by iv tumor cell inoculation, removal of resulting solid liver tumor nodules for growth in vitro, and reinjection of the cultured cells iv. After ten such sequential selections for implantation, invasion, survival, and growth in liver, a lymphosarcoma cell line was obtained (RAW 117-H10) that formed approximately 200-fold more gross liver tumor nodules than did the parental tumor cell line in comparative biologic assays and displayed enhanced malignant properties when monitored by time of host death. Very few lung nodules or tumors were present at other sites with either the parental or any of the selected RAW117 lines, which confirms previous reports in which in vivo sequential selection yielded stable cell lines with preferential tumor colonization at specific sites.
Animals↗
Specificity of arrest, survival, and growth of selected metastatic variant cell lines.
Animal tumor models for blood-borne metastasis have been developed by in vitro cloning or in vivo selection of malignant tumor cell populations to obtain organ-preferring variant tumor cell lines with altered arrest, survival, invasion, and growth properties. Selection and some tumor cell characteristics of lung-, brain-, and ovary-colonizing metastatic B16 melanoma, liver-colonizing RAW114 lymphosarcoma, and lung-colonizing MSV3T3 vasoformative sarcoma variant lines will be discussed along with additional data, suggesting that tumor cells of varying malignant potential preeexist in the unselected tumor population.
Animals↗
Cell interactions in the metastatic process: some cell surface properties associated with successful blood-borne tumor spread.
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Animals↗
Limulus amebocyte lysate reaction with streptococcal pyrogenic exotoxin.
Three immunologically distinct pyrogenic exotoxins derived from group A streptococci were found to cause coagulation of limulus amebocyte lysate. These exotoxins were shown to be several thousand times less active for lysate coagulation than gram-negative endotoxin.
Arthropods↗