Respiratory burst activity is impaired during phagocytosis of gelatinized fixed erythrocytes by inflammatory macrophages

The ability of immune and nonimmune opsonized gelatin-coated particles to stimulate respiratory burst activity by inflammatory macrophages was studied. The uptake and phagocytosis of⁵¹Cr-labeled gelatin-coated fixed erythrocytes, opsonized with either specific IgG or purified plasma fibronectin, was measured in monolayer cultures of rat inflammatory peritoneal macrophages. Respiratory burst activity was evaluated in monolayers of rat inflammatory peritoneal macrophages by measuring: (1) luminol-dependent chemiluminescence and (2) the production of¹⁴CO₂ from the oxidation of [1-¹⁴C] glucose. Uptake of opsonized gelatin-coated, fixed erythrocytes resulted in no stimulation of chemiluminescence and only a limited stimulation of [1-¹⁴C] glucose oxidation. Respiratory burst activity produced by phorbol myristate acetate was not inhibited during the uptake of opsonized gelatincoated particles. These data suggest that metabolic processes associated with macrophage respiratory burst activity may not be coupled to the ingestion of opsonized gelatin-coated fixed erythrocytes.     

Changes in the ultrastructure of human erythrocytes and in their content of free fatty acids during incubation with hydroperoxide and calcium ionsin vitro

It is shown that incubation of a suspension of human erythrocytes with H₂O₂ and Ca²⁺ mainly results in echinocytic transformation and hemolysis; incubation with H₂O₂ in the absence of Ca²⁺ is attended by polymorphous changes in erythrocytes: discocyte swelling, formation of stomatocytes and echinocytes and their hemolysis. The level of free fatty acids in human erythrocytes increases for incubation with Ca²⁺ and calcimycin under anaerobic conditions and drops for H₂O₂-induced activation of lipid peroxidation. Translated fromByulleten' Eksperimental'noi Biologii i Meditsiny, Vol. 118, N ^o 8 pp. 207–211, August, 1994 Presented by N. K. Permyakov, Member of the Russian Academy of Medical Sciences     

Mechanisms of human eosinophil activation by complement protein C5a and platelet-activating factor: similar functional responses are accompanied by different morphologic alterations

The complement system is an important amplification system for the propagation of allergic as well as pseudoallergic inflammatory reactions. In the present study, the effect of the major anaphylatoxin C5a was compared with that of platelet-activating factor (PAF) on highly purified eosinophils (≥95%) by functional as well as morphologic criteria. Upon stimulation with C5a, eosinophils maintained their spheric structure, developing short, pseudopodia-like protrusions, whereas PAF induced the generation of a number of digitating protrusions. As shown by functional and ultrastructural assay systems, both stimuli provoked significant extracellular and intracellular H₂O₂ production in eosinophils, which was inhibited by cytochalasin B. With C5a, a pronounced H₂O₂ production was detected within the small cytoplasmic vesicles, whereas PAF-induced H₂O₂ production was observed on the outer surface of the plasma membrane in the contact zones between adjacent cells. Morphologic signs of degranulation induced by C5a and PAF were accompanied by the significantly increased release of eosinophil cationic protein and eosinophil peroxidase in the presence of cytochalasin B. Like PAF, C5a induced a significant production of reactive oxygen species in eosinophils. as measured by lucigenin-dependent chemiluminescence (CL) responses in eosinophils. Maximal responses, comparable with those of interleukin-5 (100 U/ml), were observed at concentrations of 10^5--10^-6 and 10^-7-10_-8 M for PAF and C5a, respectively. Separation of eosinophils by discontinuous density gradients revealed the existence of two hypodense eosinophil populations, one of them showing significantly reduced CL responses upon stimulation with C5a and PAF. In addition, CL responses upon stimulation with C5a and PAF were abrogated by cytochalasin B, staurosporine, and wortmannin, and were almost completely blocked by pertussis toxin. In conclusion, these data indicate that C5a induces events in human eosinophils comparable to those induced by PAF in the assay systems tested. Thus, C5a, generated after activation of the complement system, may be of major importance for the eosinophil activation observed in eosinophil-related disease.     

Neutrophils may directly synthesize both H2O2 and O2 − since surface stimuli induce their release in stimulus-specific ratios

Many stimuli induce neutrophils to undergo an oxidative burst and generate toxic oxygen metabolites. The major products are O₂ ^– and H₂O₂, the latter being presumed to arise by spontaneous dismutation of the former. If H₂O₂ were indeed derived exclusively from released O₂ ^– according to the equation 2O₂ ^– + 2H⁺ H₂O₂ + O₂, one would expect that relationship to be reflected in the ratio of the two metabolites detectable in the extracellular mileu of stimulated neutrophils. A second corollary is that H₂O₂ should not form when cytochromec is present to scavenge O₂ ^– before it can dismutate. Although H₂O₂ cannot be measured directly in the presence of cytochromec because it is consumed in reoxidizing reduced cytochromec, its presence can be detected indirectly by the ability of catalase to improve the apparent yield of reduced cytochromec. We found that the relative amounts of extracellular H₂O₂ and O₂ ^– that could be measured in the environment of stimulated neutrophils varied with the stimulus and that catalase protected reduced cytochromec from H₂O₂ oxidation when some stimuli were used but not with others. For example, the ratio of O₂ ^– to H₂O₂ produced by neutrophils exposed to PMA was about 2:1, the expected result if H₂O₂ were derived from O₂ ^–. However when cytochalasin B was added to the cells before the stimulus, the yield of H₂O₂ was reduced but not the yield of O₂ ^–. When cells were allowed to settle and spread on tissue culture plastic they produced equimolar amounts of O₂ ^– and H₂O₂. Coating the plastic with IgG doubled cytochromec reduction without effecting H₂O₂. In contrast, coating with albumin reduced H₂O₂ without effecting cytochromec reduction. Soluble IgG aggregates induced production of mostly O₂ ^– whereas immune complexes resulted in release of both metabolites.FMLP and A23187 were similar to the soluble IgG aggregates in their effects and induced release of proportionately more O₂ ^– than H₂O₂. The addition of catalase to the cytochromec solution improved the yield of reduced cytochromec when PMA or IgG was used to stimulate the cells but not when FMLP was used. These and other data suggest that H₂O₂ release is not a linear function of the amount of O₂ ^– generated and that either a variable fraction of O₂ ^– spontaneously dismutates to H₂O₂ or the neutrophil NADPH oxidase, in a manner analogous to xanthine oxidase, is capable, under some circumstances, of producing H₂O₂ as well as O₂ ^–. If the latter were true, the pathologic consequences of neutrophil activation would vary depending on whether O₂ ^– was the primary product (chemotactic activation) or whether H₂O₂ was released as well (immune complex stimulation).     

Specificity of Fc-Receptors on Lymphocytes and Monocytes for Guinea-Pig IgG1 and IgG2: Induction and Inhibition of Cytolysis or Phagocytosis of Erythrocytes

The capacity of guinea-pig IgG1 and IgG2 antibodies to induce lymphocyte (K-cell) mediated lysis or monocyte/macrophage mediated phagocytosis of erythrocytes was studied with both human and guinea-pig effector cells. For both species, induction of K-cell mediated lysis was restricted to IgG2 whereas both IgG1 and IgG2 could induce monocyte/macrophage mediated phagocytosis. In competitive inhibition experiments, only complexed IgG2 inhibited lysis mediated by K-cells. The results suggest that the fc-receptors on K-cells only recognize IgG2. In contrast, complexes of both subclasses inhibited phagocytosis by human monocytes, regardless of the subclass of the inducing antibodies. Inhibition of guinea-pig macrophage mediated phagocytosis by IgG2 complexes was also independent of the inducing antibody. Hence, Fc-receptors common for IgG1 and IgG2 seem to be involved in themonocyte/macrophage mediated effector reaction. Free IgG1 was significantly less ingibitory than free IgG2 for human monocytes and hardly at all for guinea-pig macrophages. However, free IgG2, which was cytophilic for these cells, was more aggregated than IgG1. Thus, both molecular structure and state of aggregation determine interaction of IgG with cellular Fc-receptors.     

Aging of the erythrocyte XVII. Binding of autologous immunoglobulin G

Increased amounts of autologous immunoglobulin (Ig) G were found on older bovine erythrocytes employing ¹²⁵I-labelled Protein A, ¹²⁵I-labelled anti(bovine IgG) antibodies and hemolysis in the presence of anti(bovine IgG) antibodies + complement. In vitro, desialylation, proteolysis and action of an O₂$\text{·}$ source but not glycosylation induced binding of extraneous IgG to the erythrocytes.     

Evidence that C1q,a Subcomponent of the First Component of Complement,is an Fc Receptor of Peritoneal and Alveolar Macrophages

Guinea pig peritoneal macrophages were cultured for 24 h in the presence of two inhibitors of the biosynthesis of collagen-like molecules such as C1q : 10^-3 M 3,4-dehydroproline or 10^-4 M 2,2′-dipyridyl. Their Fc-receptor activity was measured by rosette formation, using sheep erythrocytes (E) coated with rabbit anti-sheep IgG (EA_IgG). The Fc-receptor activity was decreased by 40 to 70% of control cultures depending on the amount of IgG on the E. The activity of a second receptor on the macrophages, mediating the binding of C3b coated E, was not altered by this treatment.Rat alveolar macrophages were depleted of their Fc-receptor activity by pronase treatment (1.5 mg/ml) in the presence of 2,2′-dipyridyl. After washing the cells, the EA_IgG-binding activity was restored to about half of the initial level within 2 h. With 2,2′-dipyridyl also present during the second incubation, the re-expression of the Fc-receptor activity was suppressed further.Preincubation of guinea pig peritoneal macrophages with anti-C1q-F(ab')₂ for 45 min at 37°C caused a dose-dependent reduction of the Fc-receptor activity, but not C3b receptor activity.These results support our hypothesis that C1q synthesized and secreted by macrophages serves as an Fc-receptor in the membrane during the secretion.     

Scavengers of reactive oxygen intermediates do not mediate the depression of macrophage hydrogen peroxide production caused by erythrocyte phagocytosis

Our previous studies have shown that a phagocytic challenge with IgG-coated erythrocytes (EIgG) depressed macrophage triggered H₂O₂ production in vitro, and in vivo there was a decrease in the survival rate following bacteremia. The phagocytosis of an equal number of IgG-coated erythrocyte ghosts had none of these effects, indicating that the contents of the erythrocytes are important for these effects. The present study evaluated the role of the scavengers of reactive oxygen intermediates within erythrocytes in the depression of H₂O₂ production triggered with phorbol myristate acetate following a phagocytic challenge with EIgG. Elicited rat peritoneal macrophages (PM) were challenged with EIgG prepared from normal E or E with inactivated catalase, depleted glutathione, hemoglobin converted to methemoglobin, or fixed with formaldehyde. The depression of triggered H₂O₂ production was similar when equal numbers of normal EIgG and EIgG with inactivated scavengers were phagocytized. When the phagocytic challenge with normal EIgG was carried out in the presence of cytochalasin B, no depression of triggered H₂O₂ production was observed. Cytochalasin B partially blocked the phagocytosis of EIgG, so that with larger doses of EIgG there was sufficient ingestion of EIgG to depress H₂O₂ production in untreated PM. These results indicate that the scavengers of reactive oxygen intermediates present in erythrocytes are neither required nor sufficient to depress H₂O₂ production by macrophages.     

Effect of corticosteroids on the production of superoxide and hydrogen peroxide and the appearance of chemiluminescence by phagocytosing polymorphonuclear leukocytes

Human polymorphonuclear leukocytes were incubated with either methylprednisolone sodium succinate, hydrocortisone sodium succinate, or distilled water, and then latex spherules were added as target particles for phagocytosis. At low concentrations of these corticosteroids (0.04–0.22 mM), no effect was observed on O₂ ^–· production, H₂O₂ production, or chemiluminescence. At high concentrations of these steroids (2.7 mM), a significant inhibition was observed in both O₂ ^–· production and H₂O₂ production. At 2.7 mM, methylprednisolone sodium succinate significantly decreased chemiluminescence, whereas hydrocortisone sodium succinate was without effect on chemiluminescence.     

Susceptibility of Entamoeba histolytica to oxygen intermediates

To explore the susceptibility of the extracellular protozoan, Entamoeba histolytica, to toxic oxygen intermediates, trophozoites were exposed to fluxes of O₂^?, H₂O₂, and OH · generated enzymatically by the glucose oxidase and xanthine oxidase reactions. HM-1 trophozoites were resistant to O₂^?, but were readily killed by H₂O₂ alone. OH · and ¹O₂ were not required for effective amebicidal activity. The addition of a peroxidase and halide enhanced trophozoite killing by H₂O₂. Sonicates of amebae contained virtually no catalase and little glutathione peroxidase activity which may contribute to susceptibility to H₂O₂. Coupled with our previous studies with Toxoplasma gondii and Leishmania spp. these observations indicate that there is a broad spectrum of susceptibility of intra- and extracellular pathogenic protozoa to killing by oxygen intermediates.     

Hydrogen Peroxide-Releasing Function of Chemically Elicited and Immunologically Activated Macrophages: Differential Response to Wheat Germ Lectin and Concanavalin A

Various types of mouse peritoneal macrophages were studied for H₂O₂ release in the presence of wheat germ lectin or phorbol myristate acetate. Macrophages elicited 3 days before harvest by a single injection of thioglycolate, zymosan A, or a streptococcal preparation (OK-432) were highly responsive to wheat germ lectin, resulting in a marked increase in H₂O₂ release. However, immunologically activated macrophages induced by double injections of live and heat-killed BCG at 15 and 3 days before harvest or by double injections of zymosan A or OK-432 at 20 and 3 days before harvest did not show any significant response to wheat germ lectin. On the other hand, all macrophages tested responded well to phorbol myristate acetate by augmentation of H₂O₂ release. Concanavalin A inhibited wheat germ lectin- and phorbol myristate acetate-triggered H₂O₂ release from all types of macrophages, but inhibition was much more marked in the case of wheat germ lectin-stimulated H₂O₂ release. Succinylated concanavalin A (divalent concanavalin A) showed only slight suppressive action against macrophage H₂O₂ release, and prostaglandin E₁ and dibutyryl cyclic adenosine 3′, 5′-monophosphate caused depression of H₂O₂ release from OK-432-induced macrophages.     

The use of protein A and concanavalin A to examine the possible role of the carbohydrate of IgG in the binding of Clq

The suggestion that Fc-carbohydrate of rabbit IgG becomes accessible to conA on cross-linking by Staphylococcal protein A and can play a role in fixing complement (Langone et al., 1978a) was examined specifically for its relevance to Clq binding.On chromatography of pooled rabbit IgG on conA-Sepharose, 80 ± 5% of IgG shows no interaction with conA—Sepharose. Five per cent of IgG is bound and eluted with 0.1 M 1-O-methyl-α-d-glucopyranoside.The apparent K_d of the unretarded fraction for conA was less than 140 μM, whereas that of pooled IgG was 14 μM, as measured by competition for ¹²⁵I-labelled conA with guinea pig erythrocytes.No change in the K_d of the unretarded fraction of IgG for conA could be detected in the presence of protein A. A change of up to two-fold was observed in pooled IgG, and must be a result of heterogeneity of the carbohydrate in either the Fab or the Fc region of IgG.The fraction of IgG which did not bind conA showed the same Clq-binding activity as pooled IgG in ¹²⁵I-labelled Clq binding assays, so that there can be no direct relation between conA-binding activity and Clq binding.The binding of Clq to IgG was not affected by concentrations of conA (1mg/ml) and human transferrin glycopeptide (2 mM) under conditions where inhibition would be expected if complete exposure of the Fc-carbohydrate to solution was important for binding Clq.     

NADPH oxidases and MMP-9/TIMP-1 balance in human glomeruli: putative links to diabetic nephropathy

Study aim Glomerular basement membrane thickening, the hallmark of diabetic nephropathy, is thought to be related to an enhanced oxidative stress and reduced matrix proteolysis. Our study concerned the mRNA and protein expression of NADPH oxidase (NOX) components, MMP‐2, MMP‐9 and TIMP‐1 in freshly isolated human glomeruli as well as enzymatic activities and their modulation by glucose, H₂O₂ and angiotensin‐2. Material and methods NOX, cytosolic and membrane‐bound associated proteins and mRNA were analysed by RT‐PCR and Western blotting after glomerular extraction. Oxidase activity was identified by cytochrome c reduction and chemiluminescence. Gelatinases and inhibitors were semiquantitatively assessed by RT‐PCR, gelatin zymography and ELISA in a model of glomerular conditioned survival. Results NOX‐2, NOX‐4 and membrane‐bound and cytosolic factors could be observed in freshly extracted glomeruli (RNA + protein). p40^phox, p67^phox and p47^phox molecular weights were increased compared to their phagocytic counterparts advocating for specific glomerular analogues, and a slight specific oxidase activity was retrieved in isolated glomeruli. Also, mRNA coding for MMP‐2, ‐9 and TIMP‐1, ‐2 were detected. High glucose concentrations (25 mm) reduced TIMP‐1 release in glomerular survival media and MMP‐2 activity in glomerular extracts. On the opposite, angiotensin‐2 significantly induced MMP‐2 and ‐9 activities in the survival media as well as H₂O₂ in glomerular extracts, while addition of 25 mm glucose blunted these findings. Conclusion Glomerular matrix remodelling, the backbone of renal fibrosis in diabetic patients, could be induced by H₂O₂ from specific glomerular NADPH oxidases under the influence of extra‐cellular glucose and angiotensin‐2 and could participate in the control of MMP activities.     

Acute hyperoxia increases lipid peroxidation and induces plasma membrane blebbing in human U87 glioblastoma cells

Atomic force microscopy (AFM), malondialdehyde (MDA) assays, and amperometric measurements of extracellular hydrogen peroxide (H₂O₂) were used to test the hypothesis that graded hyperoxia induces measurable nanoscopic changes in membrane ultrastructure and membrane lipid peroxidation (MLP) in cultured U87 human glioma cells. U87 cells were exposed to 0.20 atmospheres absolute (ATA) O₂, normobaric hyperoxia (0.95 ATA O₂) or hyperbaric hyperoxia (HBO₂, 3.25 ATA O₂) for 60 min. H₂O₂ (0.2 or 2 mM; 60 min) was used as a positive control for MLP. Cells were fixed with 2% glutaraldehyde immediately after treatment and scanned with AFM in air or fluid. Surface topography revealed ultrastructural changes such as membrane blebbing in cells treated with hyperoxia and H₂O₂. Average membrane roughness (R_a) of individual cells from each group (n=35 to 45 cells/group) was quantified to assess ultrastructural changes from oxidative stress. The R_a of the plasma membrane was 34±3, 57±3 and 63±5 nm in 0.20 ATA O₂, 0.95 ATA O₂ and HBO₂, respectively. R_a was 56±7 and 138±14 nm in 0.2 and 2 mM H₂O₂. Similarly, levels of MDA were significantly elevated in cultures treated with hyperoxia and H₂O₂ and correlated with O₂-induced membrane blebbing (r²=0.93). Coapplication of antioxidant, Trolox-C (150 μM), significantly reduced membrane R_a and MDA levels during hyperoxia. Hyperoxia-induced H₂O₂ production increased 189%±5% (0.95 ATA O₂) and 236%±5% (4 ATA O₂) above control (0.20 ATA O₂). We conclude that MLP and membrane blebbing increase with increasing O₂ concentration. We hypothesize that membrane blebbing is an ultrastructural correlate of MLP resulting from hyperoxia. Furthermore, AFM is a powerful technique for resolving nanoscopic changes in the plasma membrane that result from oxidative damage.     

Endothelin-1 does not prime polymorphonuclear leukocytes for enhanced production of reactive oxygen metabolites

The in vitro effect of endothelin-1 (ET-1) on the capacity of polymorphonuclear leukocytes (PMNLs) to generate reactive oxygen species (ROS) was investigated. Human PMNLs were separated from healthy volunteers and preincubated for 10 min. at 37°C with varying concentrations (10^–7–10^–12 M) of ET-1. After subsequent stimulation with FMLP (10^–7 M) or opsonized zymosan (0.5 mg/ ml) the intra- and extracellular generation of ROS was assessed by luminol-amplified chemiluminescence, superoxide radical (·O ₂ ^– ) and hydrogen peroxide (H₂O₂) production.Results: ET-1 alone failed to stimulate ROS generation. Neither the capacity for extracellular generation of oxygen metabolites nor the production of ROS with an intracellular origin was changed after preincubation of PMNLs with ET-1. ET-1 did not cause a shift of the ·O ₂ ^– /H₂O₂ production ratio after stimulation of PMNLs with FMLP. These findings suggest that ET-1 in vitro does not prime human PMNLs for enhanced production of ROS.     

Effect of vitamin D on humoral and cell-mediated immune response and functional activity of peritoneal macrophages

1,25-Dihydroxyvitamin D₃, an active metabolite of vitamin D, decreases the number of antibody-producing cells in C57B1/6 mice immunized with sheep erythrocytes, suppresses lymphocytes proliferation in response to stimulation with pokeweed mitogen and concanavalin A, and stimulates functional activity of macrophages. Translated fromByulleten’ Eksperimental’noi Biologii i Meditsiny, Vol. 123, No. 1, pp. 63–65, January, 1997     

Assessment of ferrocytochrome C oxidation by hydrogen peroxide

The reduction of ferricytochrome C is commonly employed for the quantitation of O ₂ ^– . H₂O₂ arising from the dismutation of O ₂ ^– is capable of oxidizing ferrocytochrome C. In order to assess whether this may interfere with O ₂ ^– quantitation, the amount of H₂O₂ required for the oxidation of ferrocytochrome C was determined. While H₂O₂ concentrations below 10^–5 M were ineffective, one half of the reduced cytochrome was oxidized by 5×10^–5 M H₂O₂ within 15 min. H₂O₂ in the concentration range at which ferrocytochrome C is oxidized is generated upon interaction of hypoxanthine with xanthine oxidase and upon stimulation of human polymorphonuclear neutrophilic granulocytes by phorbol myristate acetate or the phagocytosis of opsonized zymosan. It is suggested that O ₂ ^– quantitation by cytochrome C reduction is routinely performed in the presence of catalase.     

Diperoxovanadate can substitute for H(2)O(2) at much lower concentration in inducing features of premature cellular senescence in mouse fibroblasts (NIH3T3)

Stress induced premature senescence (SIPS) in mammalian cells is an accelerated ageing response and experimentally obtained on treatment of cells with high concentrations of H₂O₂, albeit at sub-lethal doses, because H₂O₂ gets depleted by abundant cellular catalase. In the present study diperoxovanadate (DPV) was used as it is known to be stable at physiological pH, to be catalase-resistant and to substitute for H₂O₂ in its activities at concentrations order of magnitudes lower. On treating NIH3T3 cells with DPV, SIPS-like morphology was observed along with an immediate response of rounding of the cells by disruption of actin cytoskeleton and transient G2/M arrest. DPV could bring about growth arrest and senescence associated features at 25 μM dose, which were not seen with similar doses of either H₂O₂ or vanadate. A minimal dose of 150 μM of H₂O₂ was required to induce similar affects as 25 μM DPV. Increase in senescent associated markers such as p21, HMGA2 and PAI-1 was more prominent in DPV treated cells compared to similar dose of H₂O₂. DPV-treated cells showed marked relocalization of Cyclin D1 from nucleus to cytoplasm. These results indicate that DPV, stable inorganic peroxide, is more efficient in inducing SIPS at lower concentrations compared to H₂O₂.     

Use of an Attenuated Leishmanial Parasite as an Immunoprophylactic and Immunotherapeutic Agent against Murine Visceral Leishmaniasis

The ability of the leishmanial parasite UR6 to act as an immunoprophylactic and immunotherapeutic agent against Leishmania donovani infection in BALB/c mice was investigated. Unlike the virulent L. donovani AG83 (MOHOM/IN/1983/AG83), UR6 given through intracardiac route failed to induce visceral infection, but when it was injected subcutaneously, UR6 induced a short-lived and localized self-healing skin lesion. Priming of peritoneal macrophages with UR6 in vitro induced superoxide (O₂⁻) generation, whereas similar experiments with virulent AG83 inhibited O₂⁻ generation. It was observed that priming of mice with either live or sonicated UR6 in the absence of any adjuvant provided strong protection against subsequent virulent challenge. Further, UR6-primed infected mice not only displayed a strong antileishmanial delayed-type hypersensitivity (DTH) response but also showed an elevated level of the serum antileishmanial immunoglobulin G2a (IgG2a) isotype, whereas infected mice failed to mount any antileishmanial DTH response and showed an elevated level of IgG1. This indicates that UR6 priming and subsequent L. donovani infection allowed the expansion of Th1 cells. Our studies indicate that UR6 has potential to be used as an immunoprophylactic and immunotherapeutic agent against experimental visceral leishmaniasis.     

Tuftsin stimulates the release of oxygen radicals and thromboxane from macrophages

The physiologically occurring tetrapeptide, tuftsin (Thr Lys-Pro-Arg), was examined for its effects on guinea pig peritoneal macrophages. Adherent macrophages (mø) were exposed to tuftsin at concentrations ranging from 10⁸ to 10⁶ M, and the release of oxygen radicals and the arachidonic acid cyclo-oxygenation product, thromboxane B₂ (TXB₂), was studied. Tuftsin causes both albumin-elicited and C. parvum-activated mø to set free superoxide anion (O₂) and hydrogen peroxide (H₂O₂). It also stimulates the liberation of TXB₂ from albumin-elicited mø. The demonstrated induction of the release of toxic oxygen species from mø by tuftsin may explain how tuftsin-augmented mø-mediated cytotoxicity is accomplished.