Gallium arsenide exposure impairs processing of particulate antigen by macrophages: modification of the antigen reverses the functional defect

Gallium arsenide (GaAs), a semiconductor used in the electronics industry, causes systemic immunosuppression in animals. The chemical's impact on macrophages to process the particulate antigen, sheep red blood cells (SRBC), for a T cell response in culture was examined after in vivo exposure of mice. GaAs-exposed splenic macrophages were defective in activating SRBC-primed lymph node T cells that could not be attributed to impaired phagocytosis. Modified forms of SRBC were generated to examine the compromised function of GaAs-exposed macrophages. SRBC were fixed to maintain their particulate nature and subsequently delipidated with detergent. Delipidation of intact SRBC was insufficient to restore normal antigen processing in GaAs-exposed macrophages. However, chemically exposed cells efficiently processed soluble sheep proteins. These findings suggest that the problem may lie in the release of sequestered sheep protein antigens, which then could be effectively cleaved to peptides. Furthermore, opsonization of SRBC with IgG compensated for the macrophage processing defect. The influence of signal transduction and phagocytosis via Fcgamma receptors on improved antigen processing could be dissociated. Immobilized anti-Fcgamma receptor antibody activated macrophages to secrete a chemokine, but did not enhance processing of unmodified SRBC by GaAs-exposed macrophages. Restoration of normal processing of particulate SRBC by chemically exposed macrophages involved phagocytosis through Fcgamma receptors. Hence, initial immune responses may be very sensitive to GaAs exposure, and the chemical's immunosuppression may be averted by opsonized particulate antigens.     

The phagocytic capacity of macrophages in the S phase of the cell cycle

An inflammatory reaction was induced in the peritoneal cavity of mice. Two days later, the peritoneal macrophages, containing a proportion of S-phase (DNA-synthesizing) cells, were harvested and adhered to glass. Then the S-phase macrophages were labeled with [³H]thymidine (radioautography) and the macrophage monolayers were tested with regard to their ability to phagocytose immunoglobulincoated sheep red blood cells (SRBC). The percentages of S-phase macrophages which had phagocytosed SRBC were a little lower than those found for G-phase (G₁ + G₂) cells. Otherwise, the number of phagocytosed SRBC per macrophage was about equal for macrophages in both phases, and they both responded well by increasing the phagocytosis when the SRBC: macrophage ratio was increased. The S-phase macrophages also phagocytosed latex beads and zymosan particles efficiently.     

A reactivity to polyclonal stimulation of immunogenesis with salmozan after its repeated administration

The injection of 100 micrograms of salmozan (polysaccharide isolated from Salmonella typhi somatic O-antigen) or 50 micrograms of Escherichia coli lipopolysaccharide into mice induced a considerable increase in the number of antibody-forming cells in the spleen in response to the injection of sheep red blood cells (SRBC) 2-3 days later. This polyclonal effect was essentially weaker if the animals previously received 500 micrograms of salmozan (9-10 days prior to the injection of SRBC). The absence of reactivity was not linked with antibodies to salmozan or with some other serum factor. The lymphocytes of nonreactive mice proved to be capable of polyclonal response in the adoptive system, and at the same time the polyclonal response of intact lymphocytes to salmozan in the body of nonreactive irradiated mice was essentially weakened. The features making the above phenomenon similar to, as well as different from, the so-called "endotoxin tolerance" are analyzed.     

Characteristics of the reaction of immunologically tolerant mice to the polyclonal stimulant salmozan

The nonspecific stimulant of the immune response salmozan (Sal) increases the number of PFC against SRBC in intact mice, B mice (thymectomized, irradiated and reconstituted with embryonic liver cells), and in mice pretreated with cyclophosphamide (CY). This effect was decreased in mice pretreated with SRBC and CY. The subsequent injections of SRBC and Sal into tolerant mice did not increase the response under study. It is concluded that the effect observed is due to partial alteration of antigen-specific B cells of tolerant mice and this alteration cannot be explained by the lack of the regeneration of membrane immunoglobulins.     

Metabolic changes in the peritoneal macrophages of mice differing in the H-2 locus of histocompatibility after intraperitoneal administration of salmozan

The activity of 5'-nucleotidase in mouse peritoneal macrophages differing by histocompatibility locus H-2 after intraperitoneal injection of salmozan, an immunostimulating agent, has been studied. The character of changes in the activity of 5'-nucleotidase in peritoneal exudate macrophages after the intraperitoneal injection of salmozan has proved to be unrelated to the genotype of mice. The injection of salmozan induces a deep and prolonged decrease in the activity of 5'-nucleotidase in these macrophages. In mice of different strains changes in the activity of 5'-nucleotidase after the intraperitoneal injection of salmozan are of a linear type and can be approximated by a linear regression model.     

Immunomodulating properties of salmozan and its effect on the functional activity of macrophages

The effect of salmozan on the resistance of mice to Listeria monocytogenes infection, the formation of delayed hypersensitivity (DH) to sheep red blood cells in the animals, as well as changes in some functional activity characteristics of macrophages have been studied. The study has revealed that salmozan enhances anti-infectious resistance, suppresses the dermal manifestations of DH, and decreases the level of 5'-nucleotidase in peritoneal macrophages, stimulating their phagocytic activity. The intensity of the drug action depends on the time of its administration. The most pronounced immunomodulating action and maximal changes in the function of macrophages have been registered simultaneously after the treatment of the animals with salmozan.     

The effect of bilirubin on the fc receptor expression and phagocytic activity of mouse peritoneal macrophages

The effect of bilirubin on the phagocytic activity of mouse peritoneal macrophages and on the expression of Fc receptors and receptors for SRBC was studied. Intraperitoneally administered bilirubin influenced the expression of Fc receptors for IgM, IgG2B, IgA and IgE, whereas the expression of other receptors as well as the phagocytic activity of peritoneal macrophages remained unchanged. The possible mechanism of the effect of bilirubin on Fc receptors is discussed.     

The immune response in the hamster. VII. Studies on cytophilic immunoglobulin.

Hamster 7S IgG1 and IgG2 antibodies to hen-egg albumin (HEA) were tested for their capacity to bind to macrophage cytophilic Ig receptors. Both IgG1 and IgG2 were cytophilic for hamster macrophages though the membrane receptor had a predominant specificity for IgG1. Hamster IgG1 bound primarily to homologous macrophages whereas IgG2 bound to macrophages from other rodent species as well. The binding of hamster Ig to hamster macrophages was inhibited by a wide range of heterologous rodent sera. The only exception was guinea pig serum since guinea pig IgG2 was found to bind only to homologous macrophages. Sheep red blood cells (SRBC) coated with hamster IgG2 were ingested by macrophages more readily than those coated with hamster IgG1. Thus, there appeared to be a paradoxical relationship between the apparently strong affinity of IgG1 for the hamster macrophage Ig receptor and its reactivity weak ingestion promoting activity. Implications of this phenomenon are discussed.     

Opsonic effect of antiserum and complement on phagocytosis by macrophages from the peritoneal cavity of the Japanese eel, Anguilla japonica

Macrophage exudates in the Japanese eel, Anguillu japonica, were induced by intraperitoneal injection with a mixture of Edwnrdsiella bacterin, proteose peptone and liquid paraffin, and the opsonic effect of antiserum and complement on the phagocytic activity of the macrophages was studied.
The macrophages contained a round nucleus with a nucleolus, and an agranular cytoplasm with projecting processes. These cells showed phagocytosis of sheep red blood cells (SRBC). An opsonic effect of antiserum was recognized in terms of a significant increase of macrophage phagocytic activity against SRBC treated with antiserum relative to that against SRBC treated with inactivated normal serum. Complement, however, did not enhance the phagocytic activity of macrophages.     

IL-1 gene expression in lymphoid tissues

We examined the expression of IL-1 mRNA in vivo by in situ hybridization. RNA probes for murine IL-1 alpha and IL-1 beta were used to detect IL-1 mRNA in frozen sections of spleen, lymph node, and thymus of mice injected with Salmonella typhi LPS or SRBC. No IL-1 was detected in lymphoid tissues from un-injected mice. This lack of expression correlated with the absence of IL-1 biologic activity. However, after LPS injection, IL-1 alpha and beta mRNA expression was found in macrophages of the red pulp and marginal zone of the spleen. The periarteriolar lymphoid sheath contained cells that only expressed IL-1 beta mRNA. These cells were not lymphocytes and did not stain with the macrophage marker F4/80. A similar cellular response was found after SRBC injection. Scattered macrophages in lymph nodes and thymus were positive, but only after LPS or SRBC injection. The spleens of mice injected with LPS had megakaryocytes containing IL-1 mRNA.     

Modulation of the immune response by anaphylatoxin in the microenvironment of the interacting cells

It was shown that the anaphylatoxins C3a and C5a can modulate in vitro immunological reactivities. C3a suppresses both the in vitro polyclonal antibody response and the specific antibody response to sheep red blood cells (SRBC) of both mouse spleen cells and human peripheral blood cells. The target cell in the mouse for C3a appears to be an Lyt-1+2- suppressor-inducer cell and macrophages appear not to be required. In contrast to C3a, C5a enhances in vitro responses of mice. Both the response to SRBC and the mixed lymphocyte reaction are enhanced by C5a. This enhancement appears to be through an Ia- macrophage that contains receptors for C5a. It appears that enhancement may be brought about by interleukin 1, which is released when Ia- macrophages are pulsed with C5a. It is suggested that these anaphylatoxins, when present in high concentrations in the microenvironment of the interacting cells of the immune system, play a dynamic role in the regulation of the immune response. Peptide fragments cleaved from the Fc portion of antibody, complexed with antigen in this microenvironment, may have a similar regulating role.     

Granulocyte-macrophage colony-stimulating factor augments the primary antibody response by enhancing the function of antigen-presenting cells

Purified, recombinant-derived murine granulocyte-monocyte colony-stimulating factor was found to enhance the primary in vitro immune response to SRBC by murine spleen cells. In determining the mechanism of this augmentation, it was found that only splenic adherent cells and neither resting nor activated T cells nor B cells expressed specific receptors for GM-CSF. When splenic adherent cells were pulsed briefly with GM-CSF before addition to macrophage-depleted cultures, they reconstituted the PFC response to a significantly greater degree than did control macrophages. Splenic adherent cells incubated overnight with SRBC plus GM-CSF were also more efficient antigen-presenting cells than splenic adherent cells incubated with antigen alone. The mechanism of this enhanced antigen presentation was found to be due to a GM-CSF-dependent increase in the level of IL 1 secretion and Ia antigen expression. Consistent with these data was the finding that GM-CSF augmented IL 2 production by splenic T cells in response to suboptimal concentrations of Con A. Finally, the day 5 in vivo antibody response (as measured by serum titers) of mice immunized with a low dose of SRBC was enhanced by two daily inoculations of GM-CSF. Thus, the role that GM-CSF plays in augmenting immune responses may not be solely accounted for by its ability to cause the proliferation or differentiation of macrophages, but more than likely includes its ability to enhance the function of antigen-presenting macrophages.     

Lectin-like molecules on the murine macrophage cell surface

Lectin-like molecules on the surface of murine peritoneal exudate macrophages induced by thioglycolate or an anti-tumor streptococcal preparation, OK-432, were investigated and isolated. Furthermore, their sugar-binding specificities and their role in macrophage-mediated tumor cytotoxicity were examined. A neoglycoprotein, D-galactose (Gal)-bovine serum albumin, bound to these murine peritoneal macrophages. This binding of Gal-bovine serum albumin was inhibited by D-galactose, and by complex-type oligosaccharides (unit B) and high mannose-type oligosaccharides (unit A) prepared from porcine thyroglobulin. When thioglycolate-elicited macrophages were activated by lipopolysaccharide and/or the culture supernatant of concanavalin A-activated mouse spleen cells, they became tumoricidal and the number of the lectin-like molecules on the macrophage surface was found to increase. Since the binding and cytotoxic activities of these tumoricidal macrophages toward tumor cells were partially inhibited by D-galactose, the D-galactose-binding lectin-like molecules on the surface of tumoricidal macrophages might play an important role in macrophage-mediated cytotoxicity. These lectin-like molecules were then isolated from solubilized murine peritoneal exudate cells labeled with pyridoxal 5'-phosphate and sodium [3H]borohydride by affinity chromatography on columns of asialo unit B oligosaccharide-Sepharose 4B and/or beta-D-galactose-Bio-Gel P-100. The proteins bound to the asialo unit B oligosaccharide-Sepharose 4B column and eluted specifically were found to have approximate molecular weights of 79 000 and 18 000, and the protein bound to and eluted from the beta-D-galactose-Bio-Gel P-100 column had an approximate molecular weight of 77 000. These isolated proteins bound to the surface of glutaraldehyde-fixed tumor cells, and their binding was inhibited by D-galactose and also by D-mannose. Since most of the 77 kDa protein bound to the asialo unit B oligosaccharide-Sepharose 4B, this protein was assumed to be identical with the 79 kDa protein. These results suggest that the lectin-like molecules on murine macrophages have wide specificity and that one lectin-like molecule can bind both D-galactose and D-mannose.     

Adenosine inhibits macrophage colony-stimulating factor-dependent proliferation of macrophages through the induction of p27kip-1 expression.

Adenosine is produced during inflammation and modulates different functional activities in macrophages. In murine bone marrow-derived macrophages, adenosine inhibits M-CSF-dependent proliferation with an IC50 of 45 microM. Only specific agonists that can activate A2B adenosine receptors such as 5'-N-ethylcarboxamidoadenosine, but not those active on A1 (N6-(R)-phenylisopropyladenosine), A2A ([p-(2-carbonylethyl)phenylethylamino]-5'-N-ethylcarboxamido adenosine), or A3 (N6-(3-iodobenzyl)adenosine-5'-N-methyluronamide) receptors, induce the generation of cAMP and modulate macrophage proliferation. This suggests that adenosine regulates macrophage proliferation by interacting with the A2B receptor and subsequently inducing the production of cAMP. In fact, both 8-Br-cAMP (IC50 85 microM) and forskolin (IC50 7 microM) inhibit macrophage proliferation. Moreover, the inhibition of adenylyl cyclase and protein kinase A blocks the inhibitory effect of adenosine and its analogues on macrophage proliferation. Adenosine causes an arrest of macrophages at the G1 phase of the cell cycle without altering the activation of the extracellular-regulated protein kinase pathway. The treatment of macrophages with adenosine induces the expression of p27kip-1, a G1 cyclin-dependent kinase inhibitor, in a protein kinase A-dependent way. Moreover, the involvement of p27kip-1 in the adenosine inhibition of macrophage proliferation was confirmed using macrophages from mice with a disrupted p27kip-1 gene. These results demonstrate that adenosine inhibits macrophage proliferation through a mechanism that involves binding to A2B adenosine receptor, the generation of cAMP, and the induction of p27kip-1 expression.     

Quantifying complement-mediated phagocytosis by human monocyte-derived macrophages

The present study demonstrates that SRBC can be opsonized with untreated human serum such that lysis by active complement components is minimal but sufficient opsonization occurs to permit high rates of complement-mediated phagocytosis. Phagocytosis of SRBC opsonized with 2% whole human serum by human monocyte-derived macrophages was quantified in a colourimetric assay. Ingestion of SRBC was shown to occur solely via complement receptors because no phagocytosis was observed when SRBC were coated with heat- inactivated human serum, phagocytosis was augmented by the phorbol ester, PMA, and phagocytosis was inhibited by a protein kinase C (PKC)-specific inhibitor RO 31-8220. This method was used to demonstrate directly that HIV-1 infection of human monocyte-derived macrophages inhibits complement-mediated phagocytosis and will provide a useful tool for pharmacological investigations on complement-mediated phagocytosis by adherent macrophages.     

Influence of changes in composition of the cerebrospinal fluid on the secretory activity of the subcommissural organ in Rana esculenta

Summary Normally the lymphatic sinuses of the lymph node are loosely packed with lymphocytes and free macrophages as well as with macrophages adhering to the fibrocellular trabeculae. After immunization with SRBC cluster formation occurs in the medullary sinuses of rats between a central macrophage and peripherally located lymphocytes. These rosette-like clusters are nearly identical with the clusters found during primary and secondary immune response against SRBC in vitro and seem to be the in vivo equivalent for the same immune response.     

In vitro immune response to sheep erythrocytes in macrophage depleted cultures. Restoration with interleukine 1 or a monokine from resident macrophages and stimulation by N-acetyl-muramyl-L-alanyl-D-isoglutamine (MDP)

The involvement of macrophages in the adjuvanticity of N-acetyl-muramyl-L-alanyl-D-isoglutamine (MDP) has been examined. The stimulation of the in vitro primary immune response to sheep red blood cells (SRBC) has been studied, because it is known that macrophages cooperate through the mediation of soluble compounds for the induction of the anti-SRBC response. The cultures depleted of macrophages by passing spleen cells on Sephadex G-10 were unable to give any response to SRBC. Their immune responsiveness was fully restored by the addition of either Interleukine 1 (IL 1) obtained from P388D1 cells or a factor able to replace macrophages (FRM) obtained from resident peritoneal macrophages. MDP alone, at any dose, was unable to induce any response in such macrophage depleted cultures, but it was able to enhance the antibody response of these cultures reconstituted with monokines, with the same characteristics in dose effect and timing dependence than in whole spleen cells.     

Augmentation of macrophage complement receptor function in vitro. V. Studies on the mechanisms of ligation of macrophage Fc receptors required to trigger macrophages to signal T lymphocytes to elaborate the lymphokine that activates macrophage C3 receptors for phagocytosis

We previously described a unique lymphokine that activates macrophage C3 receptors for phagocytosis. The lymphokine is generated when T lymphocytes receive a signal from macrophages that have ingested IgG-coated material. In the present work, we examined the mechanisms by which macrophage Fc receptors must be engaged for macrophages to signal lymphocytes to elaborate the lymphokine. We found that ingestion mediated by any of the three classes of murine macrophage Fc receptors was sufficient to trigger macrophages, and that engagement of macrophage Fc receptors by immobilized immune complexes was effective as well. We also found that ligation of Fc receptors by an anti-Fc receptor IgG antibody or by its F(ab')2 or Fab fragments also triggered macrophages. The ability of monovalent ligation of the receptor to elicit biologic activity suggests that this system may be of value in elucidating general mechanisms by which ligand binding of receptors is transduced into biologic effects.