bluuebunny/crossref_metadata_2025 huggingface dataset:
Nonspecific and Candida-Specific Immune Responses in Mice Suppressed by Chronic Administration of Anti-μ
Authors: Kuruganti Uma , Henderson Lee A , Garner Ronald E , Asofsky Richard , Baker Phillip J , Domer Judith E
Published: January 2018

CBA/J mice were immunosuppressed by repeated administration of goat antibody specific for μ chain of immunoglobulin M (IgM) and tested for nonspecific and Candida albicans-specific immune responses. Immunosuppression was demonstrated by a dramatic reduction in the number of antibody-forming cells in the spleens of anti-μ-treated mice when immunized with sheep erythrocytes, by greatly reduced in vitro responsiveness of both spleen and lymph node lymphocytes from anti-μ-treated mice to lipopolysaccharide, and by a large reduction in the number of splenic IgM-posttive cells. T cell function, on the other hand, appeared to be relatively unaltered in anti-μ-treated animals, in the cytotoxic T lymphocyte activity against an allogeneic target was similar in splenocyte cultures from anti-μ- and mock-treated animals, and splenic and lymph node lymphocytes proliferated in response to concanavalin A in a lymphocyte stimulation assay. Moreover, Candida-specific delayed hypersensitivity to two different Candida antigens, one cell wall-derived (GP) and the other cell membrane-derived (BEX), was of comparable intensity in immunosuppressed and normal animals. When anti-μ- and mock-treated mice were immunized by the cutaneous inoculation of viable C. albicans blastospores and then challenged intravenously to assess the development of protective immunity, only mock-treated animals, male and female, had significant (p ≤ 0.05) protective responses demonstrable by reduction in the number of colony-forming units cultured from their kidneys 28 days after intravenous challenge. If consideration was given to the number of animals which had cleared Candida completely from the kidney, however, there appeared to be protective responses operative in the female anti-μ-treated animals as well. Neither anti-μ-treated males nor females, when immunized and challenged with C. albicans, produced Candida-specific antibody detectable by counte-rimmunoelectrophoresis, whereas all immunized and challenged mock-treated animals produced antibody. The data are consistent with the hypothesis that anti-μ treatment has little effect on multiple cellular immune functions, including those specific for C. albicans, and the combination of antibody, cell-mediated immunity and innate defenses are responsible for solid systemic defense against the fungus.
Are Serine Proteases Involved in Immune Complex Activation of Neutrophils?
Authors: Mark Deiren E , Lazzari Kristina G , Simons Elizabeth R
Published: January 2018

Soybean polypeptides and diisopropylfluorophosphate (DFP) have been reported to inhibit neutrophil functions such as the oxidative buret, chemotaxis, and/or phagocytosis in response to soluble stimuli; these observations would be compatible with the involvement of an active serine protease in neutrophil stimulation. We have investigated the possibility of such involvement when particulate stimuli such as immune complexes are utilized. The depolarization of the neutrophils’ membrane potential, one of the earliest indicators of stimulation, and superoxide production, which is detectable 30-45 sec later, were our indicators of neutrophil response to immune complexes. The neutrophils were equilibrated with, and after 5 min washed free of, up to 60 mM DFP, a potent covalent serine protease inhibitor. At DFP concentrations below 24 mM, such treatment did not perturb neutrophil activation as measured by either of the above parameters, nor did F- alone under comparable conditions. Additionally, the immune complex induced responses of neutrophils preincubated for 3 min with N-α-p-tosyl-L-lysine chloromethylketone (TLCK), L-1-tosylamido-2-phenyl-ethyl-chloromethylketone (TPCK), or phenyl-methyl-sulfonyl-fluoride (PMSF), covalent serine protease inhibitors which have, however, been shown to function in other capacities, e.g., as superoxide dismutases; 1 mM PMSF or 0.5 mM TLCK consistently reduced the observed membrane depolarization, one of the earliest consequences of neutrophil activation, by 20-30%, while 0.I mM TLCK and 0.01 mM TPCK had little or no effect. The inhibition of superoxide production, a slightly later stimulus response, by PMSF, TLCK, and TPCK was more profound (50-75%); these compounds have, however, been shown to have activities other than serine protease inhibitors—for example, as superoxide dismutases. Since DFP is purely a serine protease inhibitor, and since the three other compounds do not affect depolarization (the earlier and superoxide independent event), our results indicate that active serine proteases do not appear to be necessary for immune-complex-initiated neutrophil stimulation. Other stimuli, which are known to activate neutrophils by different pathways, were not investigated.
Graunulocyte-Macrophage Colony Stimulating Factor (GM-CSF) and Macrophage Colony Stimulating Factor (CSF-1) Synergize to Stimulate Progenitor Cells With High Proliferative Potential
Authors: Falk Lydia A , Vogel Stefanie N
Published: January 2018

Department of Microbiology, Uniformed Services University of the Health Sciences, Bethesda, Maryland The ability to expand a population of bone marrow progenitors capable of forming macrophage colonies of high proliferative potential (HPP-CFC) was achieved using a culture system and signal requirements not previously shown to support the growth of HPP-CFC. Using bone marrow cells from untreated animals (not 5-FU-treated), culture conditions designed primarily for the detection of GM-CFC or M-CFC progenitors, and a more stringent criteria for HPP-CFC colony size (≥ 2 mm diameter), HPP-CFC progenitor expansion was demonstrated following simultaneous addition of rGM-CSF and CSF-1 to bone marrow cultures. Examination of the sequence and temporal requirements for rGM-CSF and CSF-1 addition necessary for the development of HPP-CFC-like colonies revealed that addition of the second factor could be delayed for up to 5 days and still result in the development of significant numbers of HPP-CFC colonies. Based on a comparison with human spleen cell conditioned medium (HSCM) and interleukin 1 (rIL 1), as sources of “synergistic activity” (SA) for the development of HPP-CFC-like colonies in combination with CSF-1, the combination of GM-CSF and CSF-1 appears to represent a novel pathway for stimulating the expansion of HPP-CFC progenitors with high proliferative potential.
Synergistic Interactions Between IFN-γ and IFN-β in Priming Murine Macrophages for Tumor Cell Killing
Authors: Pace Judith L
Published: January 2018

The ability of MulFN-β and MulFN-γ to potentiate the development of tumoricidal activity in proteose peptone-elicited murine peritoneal macrophages was investigated. Macrophages were stimulated with increasing concentrations of either MulFN-β or MulFN-γ, alone and in combination, in the presence of 1 ng/ml lipopolysaccharide (LPS). The priming activities attributable to the interferons (IFNs) were quantified using the dose-response curves obtained for these samples. The priming activity observed for mixtures of MulFN-β and MulFN-γ was greater than that expected if MulFN-β and MulFN-γ had acted in an additive manner. Isobologram analysis of data obtained when macrophages were stimulated with combinations of IFNs demonstrated that MulFN-β and MulFN-γ acted synergistically to prime macrophages for tumor cell killing. The greatest degree of synergy was observed when macrophages were stimulated with suboptimal and nearly equivalent concentrations of each class of IFN. Further studies demonstrated that macrophages stimulated with combinations of MulFN-β and MulFN-γ were more sensitive to the trigger signal provided by LPS than were cells primed with either IFN alone. Thus, the synergistic effects observed were quantitative in nature in that macrophages perceived combinations of MulFN-β and MulFN-γ as having higher priming activities than expected.
Interleukin-1 and Interleukin-6 Stimulate Acute-Phase Protein Production in Primary Mouse Hepatocytes
Authors: Prowse Karen R , Baumann Heinz
Published: January 2018

Primary mouse hepatocytes were treated with the acute-phase mediators interleukin-1, Interleukin-6, and glucocorticoids, singularly and in combination, in order to delineate the spectrum of proteins induced by the stimulatory factors. As found for rat and human liver cells, mouse hepatocytes responded to the cytokines by increasing production of acute-phase proteins, which in mice include haptoglobin, α1-acid glycoprotein, complement C-3, serum amyloid A, and hemopexin. Serum amyloid A was unusual in that only the acidic peptide form responded to treatment with IL-1 and IL-6; the more basic form remained unchanged. In addition, an unidentified secretory protein was induced only by mixtures containing IL-6. The present study shows that a combination of IL-1, IL-6, and glucocorticoids is required for regulation of acute-phase plasma protein production in mouse liver cells.
In Vivo Neutrophil Emigration in Response to Interleukin-1 and Tumor Necrosis Factor-Alpha
Authors: Mason Margaret J , van Epps Dennis E
Published: January 2018

The migration of polymorphonuclear leukocytes (PMN) in response to recombinant interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF), C5a, and f-met-leu-phe-lys (FMLPL) in vivo was studied using a mouse subcutaneous sponge implantation model. In this model sponges were implanted in C3H/OUJ mice, and 2 days later they were injected with the test sample. After varying times, sponges were removed and digested with collagenase, and total cell counts and differentials were enumerated. IL-1 was found to stimulate a significant influx of PMN, which peaked at 6 hr and declined to near baseline levels by 24 hr. This response was dose-dependent, with the greatest response observed when 5 units of IL-1 were injected. When the IL-1 concentration was increased to 10 U, the total number of PMN migrating into the sponge was decreased, compared with that observed with 5 U of IL-1. The overall number of PMN migrating into the sponge 6 hr after injecting 5 U of IL-1 averaged 269% of the number of PMN migrating randomly into the sponge. No difference in the total number of macrophages or lymphocytes in control or IL-1-injected sponges was observed in this time frame. Heat treatment of the IL-1 at 90°C for 30 min ablated the response. Similar studies with TNF and C5a showed that both of these agents also stimulated an influx of PMN that peaked 6 hr postinjection. In contrast, FMLPL did not stimulate a PMN response. When IL-1 and TNF were injected simultaneously, an additive response was observed. These data indicate that IL-1, TNF, and C5a can all stimulate a PMN response in vivo and support the hypothesis that these substances are actively involved in the mobilization of PMN to inflammatory sites in vivo.
Comparative Study of Cytotoxicity, Tumor Necrosis Factor, and Prostaglandin Release After Stimulation of Rat Kupffer Cells, Murine Kupffer Cells, and Murine Inflammatory Liver Macrophages
Authors: Decker Thomas , Lohmann-Matthes Marie-Luise , Karck Ulrich , Peters Thomas , Decker Karl
Published: January 2018

Macrophages (MØ) and MØ-depleted (nonadherent) nonparenchymal cells (NPC) of the liver were examined for their cytotoxic potential against tumor cells, production of tumor necrosis factor (TNF), and release of prostaglandins (PG) following stimulation by lipopolysaccharide (LPS), interferon-γ (IFNγ), and zymosan.

Resident murine liver macrophages had no natural cytotoxicity for the TNF-resistant target cell line P815. Activation of these cells was only obtained by a combination of IFNγ and LPS. Inflammatory murine macrophages were in a primed stage and could be activated by LPS alone in the absence of IFNγ. Rat resident macrophages resembled functionally the inflammatory macrophages of the mouse liver rather than the resident macrophages. They displayed natural cytotoxicity against all targets tested and were further activated by LPS in the absence of IFNγ. Similar results were obtained with respect to macrophage-depleted nonadherent NPC: Mouse NPC had a low level of NK activity against Yac-1 cells. Treatment with pyran copolymer resulted in a strong increase of cytotoxicity against Yac-1; furthermore, a TNF-dependent killing of Wehi 164 and TNF-independent cytotoxicity against P815 cells were now acquired. In the rat NPC prepared from unstimulated animals expressed high levels of natural cytotoxicity against all targets.

No major differences could be observed between inflammatory MØ and Kupffer cells of rat and mouse liver with regard to TNF production and TNF-dependent killing of Wehi 164 tumor cells. The same was true for the spectrum of secreted prostanoids. Upon activation of all cell populations a marked shift toward the production of PGE2 occurred. Experiments involving the cyclooxygenase inhibitor indomethacin showed enhanced TNF-dependent tumor cell killing by nonactivated MØ in the absence of prostanoid production.
Neutrophil Membrane Sulfhydryl Groups Are Involved in Stimulated Neutrophil Adherence to Endothelium
Authors: Schwartz Barbara R , Harlan John M
Published: January 2018

We investigated the role of membrane sulfhydryl groups in adherence of stimulated polymorphonuclear neutrophils to cultured endothelial cells. Treatment of neutrophils with p-chloromercuriphenyl sulfonate (PCMPS), a slowly penetrating sulfhydryl reagent, inhibited phorbol myristate acetate (PMA)- or calcium ionophore A23187-stimulated adherence to cultured human or bovine endothelial cells. At concentrations which completely blocked PMA-stimulated adherence, PCMPS did not cause release of lactic dehydrogenase, inhibit PMA-mediated degranulation or hydrogen peroxide production, or prevent the PMA-induced increased surface expression of CD11b/CD 18 (Mac-1). Coincubation with a competing reduced sulfhydryl compound protected neutrophils from inhibition of PMA-stimulated adherence by PCMPS, whereas coincubation with an oxidized sulfhydryl compound did not. Monobromotrimethylammoniobimane, a nonpenetrating sulfhydryl reagent that is structurally unrelated to PCMPS, also inhibited stimulated neutrophil adherence to endothelium cultures. We conclude that stimulated neutrophil adherence to endothelium involves neutrophil membrane protein sulfhydryl groups.
Effect of Interleukin 1 on the Induction and Maintenance of B Cell Tolerance In Vitro
Authors: Duan Ji-Ming , Habicht Gail S
Published: January 2018

The effects of interleukin 1 (IL-1) on induction and maintenance of immune tolerance in vitro have been studied by using splenocytes from C57BL/6 male mice. B cell tolerance to the hapten trinitrophenol (TNP) was induced with TNP-human gamma globulin (HGG) and the cells were challenged with TNP-Ficoll. To determine the tolerance (or immunity), antibody concentrations in the supernatant fluids were measured by using a TNP-specific ELISA assay. Partially purified murine IL-1 abrogated tolerance induction, and when it was added at challenge phase, it also abrogated tolerance. In addition, partially purified IL-1 converted TNP-HGG from a tolerogen into an immunogen without any additional exposure to antigen. Similar results were obtained when recombinant human IL-1 α was used in place of partially purified natural IL-1. IL-1 is most likely acting directly on B cells rather than through the agency of T cells because purified B cells failed to become tolerant in the presence of IL-1. Studies of IL-1 production by antigen- or tolerogenstimulated splenocytes or purified B cells showed that only antigen could elicit IL-1 production in these cells. That tolerance abrogation is unique to IL-1 is suggested by studies which show that TNF, IL-2, and INF γ, alone, in combination with each other, or in combination with subeffective concentrations of IL-1 failed to effect tolerance abrogation.
Binding of Metastatic Colon Carcinoma Cells to Liver Macrophages
Authors: Gjøen Tor , Seljelid Rolf , Kolset Svein Olav
Published: January 2018

The liver is frequently colonized by metastatic tumor cells despite its dense population of macrophages (Kupffer cells). We have studied the interactions between metastatic colon carcinoma cells (DHD) and syngeneic Kupffer cells under different experimental conditions in vitro. In an adhesion assay the binding of DHD cells to Kupffer cell monolayers was shown to be time and temperature dependent, reaching a maximum level after about 90 min of incubation at 37°. in contrast, only a low level of binding could be observed at 4°. The level of binding could be increased by pretreatment of the Kupffer cells with phorbol 12-myristate 13-acetate. A firm interaction between the two cell types was shown to be dependent on the presence of calcium- and trypsin-sensitive structures on the surface of the Kupffer cells. Pretreatment of the macrophages with the cytoskeletal inhibitors colchicine and cytochalasin B was also found to reduce significantly the binding of tumor cells. This binding was also inhibited to a large extent by D-mannose and N-acetyl-D-galactosamine. The Kupffer cells were not cytotoxic against the colon carcinoma cells.
Structural Alterations in Fibroblast Monolayers Caused by Mast Cell Degranulation
Authors: Ginsburg Haim , Amira Milly , Padawer Jacques , Davidson Sima
Published: January 2018

Populations of mature, long-lived, nondividing mast cells develop on embryonic fibroblast monolayers after 1 mo growth of lymph node cells taken from mice immunized with horse serum. Total mast cell degranulation with 80-90% histamine release has been obtained by monoclonal anti-2,4-dinitrophenol (anti-DNP) IgE and the antigen. This degranulation process was studied by time-lapse cinematography and scanning electron microscopy. Excitation of the mast cells began as early as 10 s after addition of the antigen and lasted for about 15 s. Consequently, the fibroblast cytoplasm was displaced by short 5-10 s movements. Before degranulation, due to an extracellular film that coated the cells and the extracellular fibers, the monolayer appeared as a continuous, uninterrupted layer. After degranulation and fibroblast cytoplasm displacement, the fibrous network was exposed. Several inhibitors and antagonists of mast cell mediators were introduced to the cultures prior to addition of the antigen. So far, only with soybean trypsin inhibitor was the cytoplasm dislocation inhibited. Histamine H1 and H2 and serotonin receptor antagonists, as well as indomethacin, Cortisol, aprotinin, and phenylmethylsulfonyl fluoride, did not inhibit. These results suggest that chymase, which constitutes the greater part of the mast cell granule protein, is the causative agent.
Thermal Regulation of FMLP Receptors on Human Neutrophils
Authors: Coles Rosalind B , Ranney Richard R , Freer Richard J , Carchman Richard A
Published: January 2018

Polymorphonuclear leukocytes (PMNs) from subjects diagnosed as having iuvenile periodontitis (JP) have been categorized on the basis of their chemotactic (CTX) response to f-met-leu-phe (FMLP) when assayed concurrently with PMNs from periodon-tally healthy subjects (HP). When PMNs from JP groups demonstrating depressed CTX were assayed for lysosomal enzyme secretion (LES) in response to FMLP, there were no significant differences with respect to rate or amount. Significant differences were observed between HP and chemotactically depressed JP cells when assessed for FMLP receptor ligand binding at 23°, but not at 4°. Receptor differences observed at 23° in HP cells included an increase in amount of total binding, number of receptors, and available dispiaceable binding sites, compared with the chemotactically depressed JP PMNs, whereas the receptor affinities were similar. These data suggest that differences in FMLP receptor density in JP PMN that are chemotactically depressed may be related to processes that modulate receptor mobility and/or expression.
Induction of Interferon-Gamma and Tumor Necrosis Factor by Legionella pneumophila: Augmentation of Human Neutrophil Bactericidal Activity
Authors: Blanchard D Kay , Friedman Herman , Klein Thomas W , Djeu Julie Y
Published: January 2018

We have previously reported that Legionella pneumophila antigens can induce inter-feron-gamma (IFN-gamma) and tumor necrosis factor (TNF) in vitro and in vivo in mice. Furthermore, treatment of murine polymorphonuclear leukocyte (PMN) cultures with these cytokines resulted in augmented killing of the bacteria in vitro. The purpose of the present study was to determine if these findings could be extended to human responses. Here we report that Legionella antigens induced IFN-gamma and TNF in nonimmune human leukocytes cultures, and that these cytokines were able to stimulate the bactericidal activity of isolated PMN against L. pneumophila in vitro. Furthermore, optimal production of IFN-gamma was found in cultures which were enriched for large granular lymphocytes (LGL). The phenotype of IFN-producing cells was determined to be CD11 +, CD16+, CD2+, and negative for CD4, CD8, CD14, and Leu 7. Additionally, Legionella-infected monocytes were found to produce TNF in a dose-dependent response to the number of infecting bacteria, and the addition of recombinant IFN-gamma to infected monocytes resulted in augmented production of TNF in a synergistic manner. Finally, treatment of PMN with recombinant IFN-gamma and recombinant TNF augmented their bactericidal activity against Legionella in a dose dependent response. Thus, cytokines which can be induced by L. pneumophila antigens are able to stimulate PMN function in vitro, suggesting that resistance to infection results from a complex interaction of cytokines and cell responses.
Hematologic Interactions of Endotoxin, Tumor Necrosis Factor Alpha (TNFα), Interleukin 1, and Adrenal Hormones and the Hematologic Effects of TNFα in Corynebacterium parvum-Primed Rats
Authors: Ulich Thomas R , del Castillo Juan , Ni Rong-Xiang , Bikhazi Nadim
Published: January 2018

Endotoxin reduces the release among other cytokines of tumor necrosis factor (TNF) and interleukin 1 (IL-1) and causes peripheral lymphopenia and a dose response-dependent initial neutropenia followed by a monophaslc neutrophilia. TNF alone induces lymphopenia and an initial neutropenia followed by a biphasic neutrophilia. IL-1 alone induces lymphopenia and a monophaslc neutrophilia. TNF-plus-IL-1 caused a greater lymphopenia than either monokine alone, suggesting that both monokines contribute to LPS-Induced lymphopenia. TNF-plus-IL-1 induced neutropenia similar in magnitude to that induced by TNF alone and induced a neutrophilia significantly greater than that induced by either monokine alone, suggesting that LPS-induced neutropenia is caused by TNF, while LPS-induced neutrophilia is due to the combined effects of TNF and II-1. TNF and IL-1 were administered together with LPS to simulate the in vivo condition of endogenous monokine release during gram-negative bacteremia. TNF combined with LPS increased both the duration and magnitude of LPS-induced lymphopenia, LPS-induced neutropenia, and LPS-induced neutrophilia. TNF-plus-LPS treated rats at 2 hours after injection exhibited a striking 93% decrease in bone marrow neutrophils even though no peripheral neutrophilia was yet apparent, suggesting that the subsequent neutrophilia was due to demargination and recirculation of neutrophils sequestered in the peripheral vasculature immediately after their release from the bone marrow. Epinephrine, which causes neutrophilia by demargination but not by release of marrow neutrophils, reversed the initial neutropenia in TNF-plus-LPS-treated rats and increased the neutrophilia. IL-1 combined with LPS increased LPS-induced neutrophilia, suggesting that endogenous IL-1 also contributed to LPS-induced neutrophilia. Corynebacterium parvum-primed rats with hyperplasia of the monocyte macrophage system and treated with TNF differed from naive rats treated with TNF in that the second peak was as great as the initial peak of neutrophilia, supporting the hypothesis that the second peak of TNF-induced neutrophilia is due to the release of endogenous monokines. In conclusion, exogenous TNF, IL-1, and adrenal hormones affect circulating numbers of lymphocytes and neutrophils in a fashion consistent with their postulated endogenous role in the regulation of leukocyte trafficking during bacterial infection.
Natural Killer Cell Function in Patients With Acquired Immunodeficiency Syndrome and Related Diseases
Authors: Brenner Bluma G , Dascal André , Margolese Richard G , Wainberg Mark A
Published: January 2018

This review describes current knowledge of changes in natural killer (NK) cell function in acquired immunodeficiency syndrome (AIDS)-related disorders, vis-à-vis associated abnormalities in NK cytolytic function, NK cell subset distribution, NK cytopathology, and lymphokine regulation. NK cells, which are closely associated with large granular lymphocytes, are spontaneously cytotoxic to tumor and virally infected targets. As such, they may play a role in natural resistance to human immunodeficiency virus type 1 (HIV-1)-associated disorders and other opportunistic infections. Yet, peripheral blood NK activity is frequently reduced in patients with HIV-1-induced disease. NK cells are heterogeneous both with respect to their expression of serologically defined membrane antigens and functional activity. In AIDS-related syndromes, there appears to be a diminution of the NK pool (CD16+ cells) involved in cytolytic function, while there is an elevation of the NK pool that coexpresses NK (Leu 7+) and T (CD8+) cell markers, which show little or no involvement in cytolytic function. The impairment of in vitro NK function is not associated with a reduced frequency of lytic conjugates of effectors and target cells nor with the recycling capacity of these effector cells but rather is associated with defects in the NK cell lytic machinery following formation of such conjugates. NK cells in AIDS patients show an impairment in effector cell microtubule rearrangement following target cell interaction. The causes of NK cell dysfunction in AIDS-related disorders remain unknown. NK cells do not appear to express the CD4 epitope of the HIV receptor, nor have they been demonstrated to be susceptible to infection by HIV-1. There appears to be a preponderance of immature NK cells and a lymphokine imbalance in patients with HIV-1 associated disease. Interleukin-2 can partially restore diminished in vitro NK function. Elucidation of the involvement of the NK compartment in natural resistance to HIV-1 merits further investigation.
Chemiluminescence in a Macrophage Cell Line Modulated by Biological Response Modifiers
Authors: Tosk Jeff , Lau Benjamin H S , Lui Paul , Myers R Craig , Torrey Robert R
Published: January 2018

We have studied a murine macrophage cell line, J774, and found these cells capable of a zymosan-triggered chemiluminescent oxidative burst. Such activity was enhanced by preincubation with Corynebacterium parvum (CP), bacillus Calmette-Guerin, and Iipopolysaccharide (LPS). Under similar conditions, CP and LPS were shown to enhance J774-mediated tumor cell lysis. We have also demonstrated that murine interferon α + β rendered J774 cells more sensitive to the actions of CP and LPS. These results indicate that J774 cells may be useful for the in vitro evaluation of biological response modifiers as well as the study of oxygen radical production by macrophages.
Functional Analysis of Monocyte-Macrophages Derived From Nonadherent Cord Blood Progenitor Cells: Correlation With the Ontogeny of Cell Surface Proteins
Authors: Santiago-Schwarz Frances , McHugh Donna M , Fleit Howard B
Published: January 2018

In this study we examine some of the phenotypic and functional characteristics that accompany the differentiation of monocyte-macrophages from nonadherent precursors present in cord blood. Class II major histocompatibility complex (MHC) molecules identified by monoclonal antibody (mAb) L243 were the earliest monocyte-macrophage-associated antigens to be expressed. CD14 molecules, identified by mAb MO.2, and the transferrin receptor, identified by mAb OKT9, increased linearly over the 21-day culture period. In contrast, FcγR were not expressed on these cells until 14 days of culture. FcγRI was present on approximately 80% of the cells, whereas FcγRII and FcγRIII were present on 30% and 20% of the cells, respectively.

Functional activity of the nonadherent cell-derived monocyte-macrophages paralleled the phenotypic maturation of these cells. The capacity to stimulate a mixed leukocyte reaction paralleled the expression of class II MHC molecules. Similarly, the capacity to phagocytose Escherichia coli and Staphylococcus aureus paralleled the appearance of CR3. Indeed, phagocytosis of these organisms was partially blocked by mAb OKM-1. These studies demonstrate that monocyte-macrophages derived from cord blood nonadherent mononuclear cells are a useful population with which to characterize human macrophage differentiation and functional and phenotypic heterogeneity.
Proliferation of Macrophage Subpopulations in the Adult Rat: Comparison of Various Lymphoid Organs
Authors: Westermann Jürgen , Ronneberg Sabine , Fritz Franz Joachim , Pabst Reinhard
Published: January 2018

Adult male Lewis rats received a single intravenous injection of 5-bromo-2′-deoxyuridine (BRDU) to label all proliferating cells in the S-phase of the cell cycle. Various lymphoid organs were removed 1 and 24 hr after injection to assess local proliferation and migration of newly formed cells, respectively. In ceil suspensions, surface staining was performed for macrophage subsets (ED1, ED2, ED3), and the DNA label BRDU was detected by a monoclonal antibody. Local proliferation of ED1+ macrophages occurred in all organs investigated with the exception of the blood. Bone marrow outweighed the other organs by far; in addition to the proliferating ED1+ promonocytes, the bone marrow also contained BRDU-labeled ED2+ macrophages. Newly formed ED1+ monocytes migrated into lymphoid organs such as the mesenteric lymph nodes and spleen where they comprised about 90% of newly formed macrophages. In the spleen, ED3+ macrophages seemed to be renewed by local proliferation, whereas in the mesenteric lymph nodes these cells were replaced by immigration. The heterogeneity of macrophages was further demonstrated by the different renewal of splenic macrophages. ED1+ and ED3+ cells were replaced in a matter of days, whereas it would probably take several months to renew ED2+ cells.
Studies of the Mechanism of Natural Killer (NK) Degranulation and Cytotoxicity
Authors: Atkinson Eric A , Gerrard Jon M , Hildes G Elske , Greenberg Arnold H
Published: January 2018

Exocytosis of cytotoxic granule contents towards bound target cells is thought to be of central importance in natural killer (NK) cell-mediated killing. Although cellular cytotoxicity involving degranulation is thought to be calcium-dependent, the biochemical mechanisms that mediate this granule mobilization are unknown. Inositol-1,4,5-tris-phosphate (IP3), which acts to elevate internal calcium levels, and 1,2-diacylglycerol (DAG), which activates protein kinase C, are potent second messengers that have been shown to synergistically mediate secretion in other cell types. Production of these products of inositol phospholipid metabolism has previously been demonstrated in a rat NK cell line RNK upon exposure to susceptible tumor targets. We therefore Investigated the role of IP3 and DAG in NK-mediated cytotoxicity, specifically at the level of degranulation. Pretreatment of RNK cells with neomycin, a drug that interferes with the hydrolysis of inositol phospholipids and thus inhibits the formation of second messengers, inhibited RNK cytotoxicity against a susceptible tumor target and also inhibited RNK production of DAG in response to a similar target Natural killing exhibited by normal rat nylon wool-nonadherent spienocytes was also inhibited by neomycin. Phorbol-12-myristate,13-acetate (PMA), a phorbol ester that acts like DAG to activate protein kinase C, markedly enhanced lysis of a susceptible target cell by RNK. We evaluated whether modulation of lysis by these drugs was associated with effects on RNK degranulation by assaying the release of a granule-specific serine esterase (BLTE) in response to PMA and the calcium ionophore A23187. These agents synergized to promote the release of BLTE, and the extent of release was dependent on the concentrations of both agents. D2O and cytochalasin B, which enhance secretion in other cells, both enhanced BLTE release from RNK cells, indicating that we were detecting BLTE released via granule secretion and not due to nonspecific causes such as cell lysis. Our findings lead us to propose that NK cells form IP3 and DAG in response to susceptible target cells and that a major function of these second messengers is to mediate the exocytosis of cytotoxic granules towards the bound target cells.
Antigen Presentation by the CD4 Positive Monocyte Subset
Authors: Szabo G , Miller C L , Kodys K
Published: January 2018

Although CD4 antigen is expressed on monocytes (MØ), its functional role is uncharacterized. In this study, isolated human MØ were separated into CD4+ and CD4- MØ subsets and assessed for presentation of tetanus toxoid. The CD4- MØ subset had decreased antigen presenting cell (APC) capacity as well as increased PGE2 production when compared to the CD4+ MØ subset. Addition of a cyclo-oxygenase inhibitor (Indo-methacin) did not restore the CD4- MØ subset's APC capacity to that of the similarly treated CD4+ MØ subset, eliminating differential PGE2 production as the primary cause of differential APC capacity. Production of monokines such as IL-1 and plasminogen activator, which affect APC capacity, was similar in the CD4 MØ subsets. However, tumor necrosis factor (TNF) production (IFNγ plus MDP-induced) of the CD4+ MØ subset was slightly greater than that of the CD4- MØ. CD4- MØ's lower APC capacity is not totally explained by their differential IL-1, TNF, or PGE2 production.
Immune Dysfunction Expressed Selectively on L3T4+ T Cells in the Tumor-Bearing State
Authors: Tada Tsuyoshi , Sano Haruo , Sato Soichiro , Shima Junko , Fujiwara Hiromi , Hamaoka Toshiyuki
Published: January 2018

Spleen cells from normal C3H/He or BALB/c mice generate cytotoxic T-lymphocyte (CTL) responses to both trinitrophenyl (TNP)-modified syngeneic cells (TNP-self) and allogeneic cells. In contrast, cells from these strains of mice bearing a syngeneic tumor failed to induce anti-TNP-self-CTL responses, although portions of the same responding cells generated comparable anti-allo-CTL responses to those induced by normal responding cells. Although anti-allo-CTL responses were inducible from only Lyt-2+ T-cell subset of responding cells from normal or tumor-bearing mice, induction of TNP-CTL responses required the participation of L3T4+ T-cell subset as well as Lyt-2+ CTL precursors. In addition, the fact that the addition of concanavalin A-stimulated culture supernatant to cultures of responding cells from tumor-bearing mice resulted in the induction of appreciable anti-TNP-self CTL responses demonstrated the defect of L3T4+ T-cell function in the tumor-bearing state. Such a functional defect was ascribed neither to the loss of L3T4+ T cells nor to the generation of suppressor cells in spleen cells of tumor-bearing mice. It was found on one hand that in vitro stimulation of antigen-presenting cell (APC)-depleted normal responding population with TNP-self prepared from cells of normal or tumor-bearing mice produced comparable anti-TNP CTL responses. On the other hand, APC-depleted responding cells from tumor-bearing mice were unable to induce anti-TNP CTL responses under conditions in which APC-depleted normal responding cells induced an effective TNP-CTL response. These results indicate that selective impairment of L3T4+ T cell-mediated immunity is induced in the tumor-bearing state and that such an impairment is ascribed to the dysfunction of L3T4+ T cells themselves, but not of APC required for the activation of L3T4+ T-cell subset.
Activation of Neutrophils by Cachectin/Tumor Necrosis Factor: Priming of N-Formyl-Methionyl-Leucyl-Phenylalanine-Induced Oxidative Responsiveness Via Receptor Mobilization Without Degranulation
Authors: Tennenberg Steven D , Solomkin Joseph S
Published: January 2018

Human recombinant cachectin/tumor necrosis factor (TNF) was shown to prime neutrophils (PMNs), in a dose-dependent fashion, for subsequent oxidative responsiveness toward n-formyl-methionyl-leucyl-phenylalanine (FMLP). One basis for this phenomenon appeared to be TNF-mediated FMLP receptor mobilization. The maximal observed priming response was associated with a nearly twofold increase in the expression of PMN FMLP surface receptors, without changes in receptor affinity. Priming was not seen following stimulation with phorbol myristate acetate, possibly eliminating a role for the protein kinase C-dependent transductional components of FMLP-induced oxidative activity in the priming process. FMLP receptor mobilization occurred without significant degranulation as evident by an absence of increased granular enzyme release. These data support a potential role of macrophage-derived TNF in the augmentation of PMN host-defense during infectious and inflammatory challenge. TNF-mediated PMN oxidative priming may also promote oxidant tissue injury as seen in septic shock, adult respiratory distress syndrome, and multiple system organ failure.
Effect of Idarubicin and Epirubicin on In Vitro Polymorphonuclear Function: Diminished Superoxide Radical Formation Compared to Their Parent Compounds Daunorubicin and Doxorubicin
Authors: Cairo Mitchell S , Toy Cynthia , Sender Leonard , van de Ven Carmella
Published: January 2018

Anthracyclines are active against a variety of malignancies. The present study examined the effect of epirubicin and idarubicin on in vitro polymorphonuclear (PMN) function including chemotaxis, aggregation, bacteriocidal activity degranulation, and superoxide generation. We also studied the effects of cyclosporin (100 ng/ml) and verapamil (500 ng/ml), two membrane active biological response modifiers, on their potential role in modulating anthracycline-induced oxygen radical formation in the human PMN. Older anthracyclines (doxorubicin and daunorubicin) did not affect superoxide generation in the human PMN. However, the newer anthracyclines, both epirubicin and idarubicin, profoundly inhibited human superoxide generation (P < .02) and (P < .01), respectively. This inhibition was not agonist specific but occurred with multiple agonists including FMLP, PMA, A23187, and Zymosan-activated serum (ZAS). Last, cyclosporin and verapamil did not modulate the anthracycline effects of PMN superoxide generation. This study suggests that two of the newest anthracyclines, epirubicin and idarubicin, inhibit more PMN superoxide radical formation compared to their parent compounds. This reduction in oxyradical formation may account in part for their difference in anthracyline cellular cytotoxic activity.
Immunologic Consequences of Taurine Deficiency in Cats
Authors: Schuller-Levis Georgia , Mehta Pankaj D , Rudelli Raul , Sturman John
Published: January 2018

Our results show that a lack of taurine in the diet of cats results in a significant leukopenia, a shift in the percentage of polymorphonuclear and mononuclear leukocytes, an increase in the absolute count of mononuclear leukocytes, and a change in the sedimentation characteristics of white cells. Functional studies of polymorphonuclear cells isolated from cats fed taurine-free diets show a significant decrease in the respiratory burst as measured by chemiluminescence as well as a decrease in phagocytosis of Staphylococcus epidermis compared to cats fed the same diet containing taurine. In addition, serum gamma globulin in cats fed taurine-free diets was significantly increased compared to taurine-supplemented cats, indicating that other immune cells may be affected by taurine deficiency. Histological examination of lymph nodes and spleen revealed regression of follicular centers with depletion of reticular cells, mature and immature lymphocytes (B cell areas), as well as mild extravascular hemolysis. These results indicate that there are profound immunologic consequences in cats with prolonged taurine deficiency.
Purification of Tryptase From a Human Mast Cell Line
Authors: Butterfield Joseph H , Weiler Deborah A , Hunt Loren W , Wynn Susan R , Roche Patrick C
Published: January 2018

The neutral protease tryptase has been isolated from a human mast cell line, HMC-1. The HMC-1 line was established from the peripheral blood of a patient with mast cell leukemia and maintained as continuously proliferating clones in vitro and as solid mast cell tumors in nude mice. HMC-1-derived tryptase was purified by sequential chromatography on Dowex 1, DEAE 5 PW, and heparin-agarose. Purified tryptase has an apparent molecular weight of 150,000, as determined by molecular sieve HPLC, but migrates as a doublet of bands of 32/35,000 on SDS-PAGE gels. Maximal enzymatic activity was observed at pH 8.5. Cleavage of tosyl-L-arginine methyl ester by purified tryptase was inhibited by dansyl-L-glutamyl-glycyl-L-arginine chloromethyl ketone 2 HCI, HgCI2, tosyl-L-lysine chloromethyl ketone, leupeptin, and PMSF but not by benzamidine, aprotinin, tosyl-L-phenylalanine chloromethyl ketone, soybean trypsin inhibitor, human plasma, ovomucoid inhibitor, or lima bean trypsin inhibitor. Microsequencing of purified tryptase yielded an amino terminal sequence that was identical to that previously reported for human pituitary-derived tryptase.
Flow Cytometric Analysis of Respiratory Burst Activity in Phagocytes With Hydroethidine and 2′,7′-Dichlorofluorescin
Authors: Rothe Gregor , Valet Günter
Published: January 2018

Hydroethidine (HE) and 2′,7′-dichlorofluorescin (DCFH) were used for the flow cytometric measurement of reactive oxygen metabolites in leukocytes. Hydroethidine and DCFH were both rapidly oxidized in a cell-free cuvette assay to ethidium bromide (EB) and 2′,7′-dichlorofluorescein (DCF) by H2O2 and peroxidase, but not by H2O2 alone, while only HE was oxidized by KO2, a source of O-2. Quiescent lymphocytes, monocytes, and neutrophils spontaneously oxidized HE to EB, while DCFH was only oxidized to a low degree. Neutrophils increased 6.9-fold in EB red fluorescence and 12.5-fold in DCF green fluorescence during the respiratory burst induced by phorbol 12-myristate 13-acetate or 6.1-fold and 4.7-fold, respectively, during the respiratory burst induced by Escherichia coll bacteria. The HE or DCFH oxidation during the respiratory burst, unlike the spontaneous HE oxidation, was not inhibitable by 10 mM NaN. indicating a non-mitochondrial source of cellular oxidants during the respiratory burst such as NADPH oxidase, which produces O-2. The oxidation of DCFH, but not of HE, was decreased in stimulated neutrophils, which were simultaneously loaded with HE and DCFH. Intracellular DCFH oxidation induced by incubation of resting neutrophils with extracellular H2O2 was not influenced by the presence of HE. This indicates that HE is oxidized at an earlier step in the reactive oxygen metabolism of neutrophils than DCFH, i.e., by early oxygen metabolites like O2-, while DCFH is oxidized in part by H2O2 and phagosomal peroxidases. The differential oxidation of HE and DCFH during simultaneous cellular staining permits the analysis of up to three functionally different neutrophil populations in septic patients. This is of interest for the determination of disease-related alterations of oxygen metabolism in quiescent and stimulated leukocytes.
Development, Differentiation, and Maturation of Macrophages in the Fetal Mouse Liver
Authors: Naito Makoto , Takahashi Kiyoshi , Nishikawa Shin-lchi
Published: January 2018

Primitive macrophages emerged in the sinusoidal lumen of the fetal mouse liver at 10 days of gestation before the Initiation of hepatic hematopoiesis and matured into fetal macrophages. In the culture of cell suspensions from the fetal liver with LP3-conditioned medium, monocyte colonies were formed, but monocytopoiesis was poor in the early stage of hepatic hematopoiesis in vivo. In the culture of cell suspensions obtained from the fetal liver at 10 days of gestation on the monolayer of a mouse bone marrow stromal cell line, ST2, primitive/fetal macrophage colonies were formed before the development of monocyte/macrophage colonies and showed differentiation of primitive macrophages into fetal macrophages without passing through the stage of promonocytes and monocytes. At this time, the fetal cardiovascular system was connected with the vitelline vein just before the formation of the liver. With the progress of gestation, a monocytic cell series was observed to develop and form a monocyte/macrophage population. This was confirmed by in vitro studies with an LP3-conditioned medium and on a monolayer of ST2. Thus, it appears that there exist two different macrophage populations, a primitive/ fetal macrophage population and a monocyte/macrophage population in hepatic hematopoiesis. It also appears that fetal macrophages are differentiated from primitive macrophages which are colonized into the fetal liver from the yolk sac or which develop in loco, presumably from hematopoietic stem cells.
Lipopolysaccharide (LPS) Alters Phosphatidylcholine Metabolism in Elicited Peritoneal Macrophages
Authors: Grove Robert I , Allegretto Nick J , Kiener Peter A , Warr Glenn A
Published: January 2018

We investigated the effects of LPS on mouse peritoneal macrophage phospholipids using radiolabeled precursors. LPS (200 ng/ml) stimulated incorporation of [32P] into all classes of phospholipids within 0.5 hr, and after 2 hr the increase was 60% greater than controls. Separation of the phospholipid classes by thin-layer chromatography revealed a selective increase in incorporation of label into phosphatidylcholine (PC) (90% increase compared to approximately 50% in the other phospholipids). In macrophages labeled with [3H]-choline, LPS stimulated both the incorporation of label into PC and the release of incorporated label into the medium. The time dependencies of stimulated [3H] release and [32P] incorporation were similar. These data are consistent with the hypothesis that LPS activates macrophages via a PC-specific phospholipase-dependent mechanism.