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PubMed1984

Interferons as modulators of human monocyte-macrophage differentiation. I. Interferon-gamma increases HLA-DR expression and inhibits phagocytosis of zymosan.

The development of HLA-DR (Ia) expression in the presence and absence of interferon-gamma was monitored in monocyte-macrophage cultures. Overnight incubation with doses as low as 5 U/ml gave elevated values for Ia expression and the maximum increase was obtained with 200 U/ml. In contrast interferon-alpha had only a slight effect on the expression of Ia at doses as high as 2000 U/ml. The increase seen at 24 hr was maintained during the first 2 days of culture. The interferon-gamma-treated cells expressed four to five times more Ia than fresh monocytes. During the same time, monocytes cultured in the absence of interferon expressed approximately two times the amount of fresh monocytes. When the surface density of Ia was calculated, the interferon-gamma-treated monocytes expressed twice that of the untreated cells. Major changes in morphology and size occurred between days 3 and 4 of monocyte to macrophage development. Consequently a rapid increase in Ia expression took place; however, when the surface density was calculated this value increased only slightly when the monocytes matured to macrophages. The interferon-gamma-treated cells continued to express more total Ia as well as having increased surface density of this antigen. Interferon-gamma was also added to monocyte-macrophages several days after culture initiation (days 3, 4, and 5). Despite being in different stages of maturation, the cells responded to the interferon with increased Ia expression and surface density. The phagocytic activity of opsonized zymosans was also monitored. In contrast to Ia expression, this activity was downregulated by interferon-gamma, and the lower levels of phagocytosis were maintained through the 7 days of observation. Thus, interferon-gamma appears to change the differentiation pathway of the monocyte. The signal stimulates an increased level of Ia that may assist in the initiation of immune responses, and at the same time downregulates the scavenger role of removing opsonized particles. Once the monocyte has received this specific signal it continues to develop in a pathway different from that of the nontreated monocytes.

Cell Differentiation↗
PubMedDate not supplied

[The influence of cyclophosphane and phenthyrine and their combination with zymosan and serotonin on the functional features distingusihing peritoneal macrophages in mice].

Radiometric analysis into phagocytic activity of macrophages in intact and tumourous mice following introduction to them of phenthyrine and cyclophosphane showed the former to suppress to a greater extent the absorption function and the latter--that of digestion. A combined application of phenthyrine and cyclophosphane together with serotonin helps re-establish the microphages function in tumourous animals and to heighten the antineplastic activity of these compounds. Zymosan and its combination with phenthyrine and cyclophosphane do not produce any positive effect on the peritoneal cells.

Animals↗
PubMed1984

The effect of a single infusion of zymosan-activated plasma on the pulmonary microcirculation of sheep. Structure-function relationships.

Activation of the complement cascade is one of the mechanisms through which endotoxin may cause acute lung damage. The structural and functional changes following infusion of complement-activated plasma are described. In five anesthetized open-chest sheep, the authors monitored pulmonary and systemic artery pressures for 1 hour before and for 4 hours following the start of zymosan-activated plasma (ZAP) infusion (2 ml/min over a 20-minute period). Cardiac output, blood gases, and the number of circulating white cells were also measured. In addition, we took lung biopsy tissue at baseline, 7.5, 15, 30, 60, 120, 180, and 240 minutes following the start of infusion. Lung lymph flow and protein concentration were also monitored in 2 sheep. Following ZAP infusion there was an early phase of leukopenia and marked pulmonary hypertension, followed by a phase characterized by a modest increase in the flow of protein-rich lung lymph. By light microscopy pulmonary sequestration of granulocytes was evident just 7.5 minutes following the start of ZAP infusion. Peripheral lung granulocytes increased threefold above control values by 7.5 minutes, increasing to sevenfold by 30 minutes. Electron-microscopic studies showed that some of the granulocytes were disrupted, and specific and azurophilic granules were seen in the lumen. By 15 minutes endothelial damage was apparent, and intravascular monocytes were surrounded by a proteinaceous coat. Edema accumulation and an infiltration of inflammatory cells in the lungs' connective tissue regions increased to 2 hours. From 2 hours, lung injury was less marked, and the number of peripheral lung granulocytes, fewer. Sequestration of granulocytes occurred with the onset of pulmonary hypertension and leukopenia, and was most marked when lung injury was most severe. Transient endothelial damage and edema preceded the physiologic changes interpreted as an increase in pulmonary vascular permeability. Although pulmonary sequestration of granulocytes was at least as great as that with endotoxemia, unlike endotoxemia, ZAP caused only transient endothelial injury and modest changes in vascular permeability.

Animals↗
PubMed1984

The effect of an infusion of zymosan-activated plasma on hemodynamic and pulmonary function in sheep.

To assess the integrated cardiopulmonary response to acute microvascular lung injury, we infused zymosan-activated plasma (ZAP) into mature, awake sheep. ZAP infusion resulted in an immediate increase in the mean pulmonary artery pressure (PAP) from 17.5 +/- 4.5 mm Hg to a maximum of 33.0 +/- 5.4 mm Hg by 5 min (p less than 0.01) of a 90-min infusion. The pulmonary artery hypertension (PAH) was sustained throughout ZAP infusion, and for up to 30 min after infusion was completed (p less than 0.05). Lung lymph flow increased immediately, but lymph to plasma total protein ([L/P]) ratios remained unchanged, suggesting increased pulmonary microvascular permeability. The PaO2 fell to a minimum by 5 min (106.9 +/- 17 to 88.3 +/- 14 mm Hg; p less than 0.01), but recovered toward preinfusion values in the final 60 minutes of ZAP infusion. The cardiac index (CI) and stroke volume index (SVI) also fell significantly for 15 minutes (p less than 0.01), and then recovered to levels not different from baseline by 30 min. In parallel with all of these changes, the polymorphonuclear leukocyte count (PMN) in arterial blood fell immediately [baseline to 5 min, delta 4.9 +/- 2.5 (X 10(3))/mm3; (p less than 0.01)] due to sequestration within the pulmonary microvasculature, since a baseline PMNs were being released from the lung, -6.0 +/- 19%, while by 15 min the PMNs were being sequestered, +58.4 +/- 31% (p less than 0.01). Therefore, ZAP infusion reproduced the clinical picture of acute microvascular lung injury in humans, specifically acute PAH and increased pulmonary microvascular permeability. These two changes were due to a cascade of events associated with the pulmonary microvascular trapping of PMNs.

Animals↗
PubMed1984

Neutrophil-mediated cellular cytotoxicity induced by zymosan-activated serum.

Human neutrophils, incubated in the presence of zymosan-activated serum ( ZAS ), were able to damage ox red blood cells ( ORBC ) as assessed by a 4-hr 51Cr-release assay. Cells from a patient with chronic granulomatous disease were incapable of mediating ZAS -induced cytolysis. Cytotoxicity by normal neutrophils was prevented by N-ethyl-maleimide as well as by catalase, but heat-inactivated catalase and superoxide dismutase were completely ineffective. Heme enzyme inhibitors, azide and cyanide, only partially reduced the target cell lysis. The results indicate the involvement of products of the oxidative metabolism, mainly hydrogen peroxide, as cytolytic mediators. These mediators released by neutrophils may contribute to the tissue injury occurring during inflammation.

Azides↗
PubMed1983

[Effect of monochromatic red helium-neon laser on the morphology of zymosan arthritis in rats].

Histological and electron microscopy findings suggesting the effect of monochromatic coherent polarized red light on zymosan arthritis in rats are presented. The data obtained have confirmed that treatment of inflamed joints with laser rays exerts an evident therapeutic effect which is determined by activated function of macrophages and fibroblasts of the synovial membrane, resulting in a more rapid change of inflammation phases. Analysis of the cases where the rats were treated with ordinary red light has revealed no essential differences in the joints of the control and irradiated groups.

Animals↗
PubMed1982

Bovine peripheral blood polymorphonuclear neutrophil chemotactic response to Pasteurella haemolytica or zymosan-activated serum.

The chemotactic influence of Pasteurella haemolytica and of products of its growth was evaluated in vitro, using bovine peripheral blood polymorphonuclear neutrophils (PMN) in an underagarose migration assay system. Pasteurella haemolytica was cultured, quantitated, killed, and lysed by freeze-thawing. The PMN directional migration toward P haemolytica lysate was not significantly (P less than 0.9) different from spontaneous random PMN migration. Also, there was no chemotactic effect of viable 6-hour cultures of P haemolytica when tested in the in vitro underagarose system. Directional migration (1.70 +/- 0.53 mm) of PMN to zymosan-activated pooled bovine serum was observed and determined to be significantly (P less than 0.001) greater than was spontaneous random PMN migration (0.84 +/- 0.21 mm).

Animals↗
PubMed1994

Effects of PAF, FMLP and opsonized zymosan on the release of ECP, elastase and superoxide from human granulocytes.

Platelet-activating factor (PAF) is a potent chemoattractant for human eosinophils and neutrophils and causes eosinophil and neutrophil recruitment into animal airways. Since eosinophils and eosinophil cationic proteins are thought to play an important role in the pathophysiology of asthma, we have examined the hypothesis that PAF may also stimulate eosinophil cationic protein (ECP) release from human granulocytes. Granulocytes (93% neutrophils, 3% eosinophils) were isolated from the blood of normal volunteers, using metrizamide density gradients, and stimulated in vitro with PAF, L-formyl-L-methionyl-L-leucyl-L-phenylalanine (FMLP) or opsonized zymosan (OPZ). Superoxide generation was measured colorimetrically, granulocyte degranulation by a fluorimetric assay for elastase, and eosinophil activation by specific radioimmunoassay (RIA) for ECP. Granulocyte chemotaxis was also measured. Whilst both PAF and FMLP were potent chemoattractants for human mixed granulocytes (concentrations producing half the maximal effect (EC50s) ca 10 nM), PAF at concentrations below 10 microM was a poor stimulus to superoxide generation, elastase release or ECP release from the same cell population. In contrast, FMLP was a potent stimulus to both superoxide generation (EC50 48 nM) and ECP (EC50 ca 100 nM) and elastase release (EC50 ca 1 microM). OPZ was a potent stimulus to superoxide generation, but was a poor stimulus to ECP or elastase release. Thus, although PAF is a potent chemoattractant for human granulocytes, our results suggest that it alone may not stimulate their subsequent activation and release of cytotoxic products.

Blood Proteins↗
PubMed1977

Macrophage responses to mouldy hay dust, Micropolyspora faeni and zymosan, activators of complement by the alternative pathway.

Mouse peritoneal macrophages in culture exposed to mouldy hay dust, Micropolyspora faeni or glycopeptide or protein/glycoprotein fractions from this organism show marked biochemical changes. For comparison the interaction of cultured macrophages with zymosan has been investigated. All these agents induce the release of hydrolytic enzymes from macrophages, even in the absence of serum in the medium. The release is time- and dose-dependent and is not associated with loss of the cytoplasmic enzyme lactate dehydrogenase or any other sign of cell death. The parallelism between the capacity of these agents to activate the complement system via the alternative pathway and to induce inflammatory responses in vivo and selective lysosomal enzyme secretion from cultures of macrophages is discussed. The in vitro phenomena seen with mouldy hay dust, M. faeni, the protein/glycoprotein and the glycopeptide derived from it, may be relevant to understanding the role of mononuclear phagocytes in the disease farmer's lung and other inflammatory reactions.

Animals↗
PubMed1979

Augmentation of antigen-stimulated histamine release from human basophils by serum-treated zymosan particles. II. Dependence on IgE-mediated release.

It was observed previously that serum-treated zymosan particles (Zx) augmented antigen and anti-IgE stimulated histamine release. With most of the enhancement attributed to an increased rate of release, this suggested that Zx was active only during the course of IgE-mediated release. This association between IgE-mediated histamine release and responsiveness to Zx was examined further in the present report. Addition of Zx at various time intervals after release had been initiated indicated that the basophil responsiveness to Zx was limited in duration; maximum responsiveness to Zx correlated closely with the period in which the rate of IgE-mediated histamine release was maximum. The time-dependent decline in sensitivity to Zx paralleled the kinetics for desensitization to antigen. Addition of Zx failed to cause release from basophils desensitized in vitro or from basophils of a donor who failed to release histamine upon challenge with anti-IgE. In contrast to the enhancement of IgE-mediated release, Zx did not augment histamine release caused by C5a or the synthetic peptide f-Met-Leu-Phe. It is concluded that an obligatory link exists between ongoing IgE-mediated histamine release and enhancement by Zx.

Antigens↗
PubMed1996

Fusion of Leishmania amazonensis parasitophorous vacuoles with phagosomes containing zymosan particles: cinemicrographic and ultrastructural observations.

Studies on fixed preparations have shown that vacuoles containing zymosan (Z) particles internalized by infected macrophages can selectively fuse with the large parasitophorous vacuoles (PVs) that shelter Leishmania amazonensis. To examine the kinetics of vacuolar fusion in individual cells, particles were followed by time-lapse cinemicrography from their uptake to their entry in a PV. Newly formed Z-containing vacuoles moved centripetally and, if they contacted a PV, the two vacuoles remained closely apposed for variable, often extended, periods of time before they eventually fused. Transmission electron microscopy confirmed that the cytoplasm separating the partner vacuoles could be reduced to a very thin layer. Initiation of fusion was indicated by reduced refractility of the boundary between Z vacuoles and target PVs. Within a few minutes the PV enlarged and encompassed the Z particles, which remained immobile throughout. The interval between phagocytosis and fusion, 50 +/- 7.4 min (N = 17; range, 4 to 108 min), suggests that most but not all Z vacuoles underwent significant maturation by the time of fusion. Some particles were transferred singly, others entered PVs in groups of 2 or more, and additional clustered transfers to the same vacuole were also observed. These observations provide a baseline for studies of the biochemical mechanisms and the pharmacological control of the fusion of Leishmania PVs, and for the comparison of the fusion behavior of the PVs with that of other phagocytically derived vacuoles.

Animals↗
PubMed1997

Bridging of neutrophils to target cells by opsonized zymosan enhances the cytotoxicity of neutrophil-produced H2O2.

Hydrogen peroxide (H2O2) is a well-established cytotoxic agent released by activated neutrophils into the extracellular environment. However, a maximum of only 5 microM H2O2 was detected in the medium when 10(6) neutrophils/ml were activated with opsonized zymosan (OZ), more than 50-fold lower than the concentration of exogenous H2O2 required to produce equivalent killing of a cell line. In addition PMA-activated neutrophils were noncytotoxic, despite the capacity of PMA to generate two- to fourfold as much H2O2 for five times longer. The basis for this discrepancy was explored. NaN3 increased cytotoxicity to >90% only when neutrophils were activated with OZ due in part to inhibition of myeloperoxidase-mediated hydrolysis of H2O2, while catalase completely prevented cytotoxicity of OZ-activated neutrophils. These results indicate that H2O2 was solely responsible for the observed cytotoxicity. OZ-mediated cytotoxicity was prevented by intermittent agitation of the cultures or by the addition of soluble complement receptor type 1, suggesting that a physical association between neutrophils and target cells mediated by OZ was required to generate a cytotoxic environment. Significant numbers of neutrophil-target cell aggregates were observed by microscopic examination only under low hydrodynamic shear conditions. We conclude that the cytotoxic potency of H2O2 produced by neutrophils activated with OZ was due to a localized high concentration of H2O2 to which the target cells were exposed as a result of their labile adherence to OZ. This phenomenon may reflect a mechanism that neutrophils have acquired for maximizing the antimicrobial power of extracellular oxidants toward microbes that escape phagocytotosis.

Catalase↗
PubMed1997

Opsonized zymosan stimulates the redistribution of protein kinase C isoforms in human neutrophils.

We examined the ability of opsonized zymosan (OPZ) to stimulate translocation of protein kinase C (PKC) isoforms in human neutrophils. Neutrophils express five PKC isoforms (alpha, betaI, betaII, delta, and zeta), but little is known of their individual roles in neutrophil activation. As determined by immunoblotting, OPZ caused a time-dependent translocation of the predominant PKC isoforms (betaII, delta, and zeta) to neutrophil membranes, with a concomitant loss from the cytosol. Maximal translocation of all three isoforms occurred by 3 min. No PKC immunoreactivity was observed in a crude nuclear fraction, but PKC-delta and -zeta were found in the granule fraction after degranulation (10 min). PKC activity (Ca2+-dependent and -independent) increased 50- and 19-fold, respectively, by 10 min in the granules from OPZ-stimulated cells. Curiously, no immunoreactive cPKC (alpha and beta(I/II)) could be localized in the granule fraction to account for the Ca2+-dependent PKC activity. Localization of PKC isoforms in the neutrophil membranes and granules suggests their involvement in the regulation of functional responses triggered by OPZ. PKC isoform translocation to membranes from OPZ-stimulated cells preceded both p47phox (a cytosolic component of the NADPH oxidase) translocation and NADPH oxidase assembly. The presence of both PKC isoforms and p47phox in the membrane was transient, with the loss of p47phox occurring sooner than either the loss of membrane-associated PKC or that of NADPH oxidase activity. The apparent EC50 values for PKC translocation and NADPH oxidase assembly were similar. These data suggest that PKC isoforms regulate the assembly and activation of NADPH oxidase induced by OPZ.

Biological Transport↗
PubMed2000

Activation of cytosolic phospholipase A2 by opsonized zymosan in human neutrophils requires both ERK and p38 MAP-kinase.

The present study demonstrates that stimulation of human neutrophils with opsonized zymosan (OZ), which binds to Fc gamma receptors (Fc gamma Rs) and C3b receptors, activates both ERK and p38 MAP-kinase. Thus, the relative role of both types of MAP-kinase, ERK and p38, in activation of cPLA2 by OZ was studied. cPLA2 activation by OZ was detected 15 sec after stimulation, maintained a plateau for 10 min and decreased thereafter. p38 MAP-kinase activation exhibited kinetics similar to that of cPLA2, while ERK activation was detected within 15 sec but decreased significantly in less than 5 min after stimulation. Pretreatment of the cells with the MEK inhibitor, PD-098059, or the p38 MAP-kinase inhibitor, SB-203580 resulted in total inhibition of ERK or p38 MAP-kinase activity, respectively. Each inhibitor caused a partial inhibition during the time course of cPLA2 activity, while their combination caused a total inhibition. Compared to OZ, inactivated OZ, which does not contain the complement proteins, induced an identical time-dependent stimulation of ERK and p38 MAP-kinase as well as a similar cPLA2 activity, suggesting that the role of the C3b receptors in this system is negligible. It is concluded that OZ activates both ERK and p38 MAP-kinase and that the two isotypes are required for the onset and maintenance of cPLA2 activity.

Cytosol↗