Phagocytosis of Candida albicans enhances malignant behavior of murine tumor cells.

Murine tumor cells were induced to phagocytize either Candida albicans or group A streptococcal cells. The presence of microbial particles within the tumor cell cytoplasm had no effect on in vitro tumor cell growth. However, when Candida albicans-infected tumor cells were injected into syngeneic mice, they formed tumors that grew faster, invaded the surrounding normal tissue more rapidly and metastasized more rapidly than control tumor cells. Tumor cells infected with group A streptococcal particles did not grow faster or show increased malignant behavior. These data indicate that the in vivo behavior of malignant tumor cells can be modulated by microbial particles, which are often present in the microenvironment of the growing tumor.

Systemic complement activation, lung injury, and products of lipid peroxidation.

Previously we have demonstrated that systemic activation of the complement system after intravenous injection of cobra venom factor (CVF) results in acute lung injury as reflected by increases in the vascular permeability of the lung as well as by morphologic evidence of damage to lung vascular endothelial cells. In using the vascular permeability of the lung as the reference, the current studies show a quantitative correlation between lung injury and the appearance in plasma of lipid peroxidation products (conjugated dienes) as well as increased concentrations of lactic dehydrogenase (LDH) and one of its isoenzymes (LDH-4). After injection of CVF, extracts of lungs also showed elevated levels of conjugated dienes, whereas no elevations were found in extracts of liver, kidney, and spleen. There was no evidence in CVF-injected rats of renal or hepatic injury as reflected by the lack of development of proteinuria and the failure to detect increased serum levels of liver-related enzymes. Other peroxidation products identified in plasma of CVF-injected rats involved hydroperoxides and fluorescent compounds with features of Schiff bases. Not surprisingly, malondialdehyde was not found to be a reliable plasma indicator of lipid peroxidation associated with oxygen radical-mediated lung vascular injury. In using a model of oxygen radical-independent lung injury induced by oleic acid, although large amounts of LDH and LDH-4 were found in the plasma, no increases in plasma levels of conjugated dienes were detected. In CVF-injected animals treated with interventions protective against lung injury (neutrophil depletion, catalase, hydroxyl radical scavengers, or iron chelators), there were striking reductions in the plasma levels of conjugated dienes, hydroperoxides, and fluorochromic products. Morphometric analysis of lung sections revealed that the protective interventions did not interfere with the accumulation of neutrophils in lung interstitial capillaries after systemic activation of complement. In vitro studies with phorbol-stimulated neutrophils failed to demonstrate appearance of conjugated dienes, suggesting that the dienes appearing in plasma of CVF-injected animals are not the result of autotoxic changes in neutrophils. The data presented in this paper suggest that acute lung injury mediated by oxygen radicals derived from phagocytic cells can be monitored by the appearance in plasma of products of lipid peroxidation.

Suppression of immune complex vasculitis in rats by prostaglandin.

Immune complex-induced vascular damage can be markedly suppressed by treatment of rats with either prostaglandin (PG)E1 or its stable derivative, 15-(S)-15-methyl PGE1, but not with PGF2 alpha. In addition, PGD2 and PGE2 also show suppressive effects. The PGE1 derivative is considerably more effective than PGE1 and shows potent anti-inflammatory activity even after oral administration. Suppression of the vasculitis reaction is reflected by a greatly diminished increase in vasopermeability, indicating little or no vascular damage. In suppressed animals, the infiltration of neutrophils is greatly reduced, and those leukocytes that have appeared at tissue sites fail to show phagocytic uptake of immune complexes. In suppressed animals, the skin sites nevertheless show deposits of immune complexes and C3 fixation in vascular walls. Neutrophils harvested from the blood of rats treated with PGE1 show depressed responsiveness in chemotaxis and in enzyme secretion after incubation with chemotactic peptide. These studies indicate that certain PG have potent anti-inflammatory activity, which may be related to their effects on leukocytes.

Chemokine expression during hepatic ischemia/reperfusion-induced lung injury in the rat. The role of epithelial neutrophil activating protein.

The liver is highly susceptible to a number of pathological insults, including ischemia/reperfusion injury. One of the striking consequences of liver injury is the associated pulmonary dysfunction that may be related to the release of hepatic-derived cytokines. We have previously employed an animal model of hepatic ischemia/reperfusion injury, and demonstrated that this injury causes the production and release of hepatic-derived TNF, which mediates a neutrophil-dependent pulmonary microvascular injury. In this study, we have extended these previous observations to assess whether an interrelationship between TNF and the neutrophil chemoattractant/activating factor, epithelial neutrophil activating protein-78 (ENA-78), exists that may be accountable for the pathology of lung injury found in this model. In the context of hepatic ischemia/reperfusion injury, we demonstrated the following alterations in lung pathophysiology: (a) an increase in pulmonary microvascular permeability, lung neutrophil sequestration, and production of pulmonary-derived ENA-78; (b) passive immunization with neutralizing TNF antiserum resulted in a significant suppression of pulmonary-derived ENA-78; and (c) passive immunization with neutralizing ENA-78 antiserum resulted in a significant attenuation of pulmonary neutrophil sequestration and microvascular permeability similar to our previous studies with anti-TNF. These findings support the notion that pulmonary ENA-78 produced in response to hepatic-derived TNF is an important mediator of lung injury.

Tumor necrosis factor participates in the pathogenesis of acute immune complex alveolitis in the rat.

We have examined the role of intrapulmonary TNF in a rat model of acute immune complex-triggered alveolitis. Intratracheal instillation of IgG anti-bovine serum albumin (anti-BSA) followed by intravenous infusion of BSA results in acute alveolitis. Over the 4-h course of evolving lung injury, a 10-fold increase in TNF activity occurred in bronchoalveolar lavage (BAL) fluid. Immunohistochemical analysis of lung sections and BAL cells revealed that alveolar macrophages are the chief source of TNF. Antibodies that specifically neutralize rat TNF activity were raised in rabbits immunized with recombinant mouse TNF alpha. When administered into the lungs with anti-BSA, anti-TNF resulted in a marked reduction (up to 61%) in lung injury. Intratracheal instillation of exogenous TNF alone, or in combination with anti-BSA, resulted in an increase in lung injury compared to controls. Morphometric analysis and measurements of myeloperoxidase activities in whole lung extracts from rats treated with anti-TNF revealed a marked reduction in neutrophils compared to positive controls. The anti-TNF antibody preparation did not inhibit in vitro complement activation or diminish neutrophil chemotactic activity present in activated rat serum. These data indicate that intrapulmonary TNF activity is required for the full development of acute immune complex-triggered alveolitis, that alveolar macrophages are the primary source of this cytokine, and that TNF participates in the pathogenesis of immune complex alveolitis through a mechanism involving neutrophil recruitment.

C-C chemokine-induced eosinophil chemotaxis during allergic airway inflammation.

The production of eosinophil-specific chemotactic factors during allergic airway responses may be a pivotal event resulting in eosinophil accumulation, activation, and airway damage. Recent studies have identified specific chemokines that may play crucial roles in recruitment of eosinophils to the site of allergic reactions. In this study we have utilized an established model of schistosome egg antigen (SEA) -mediated allergic responses to examine the role of specific C-C chemokines [macrophage inflammatory protein-1alpha (MIP-1alpha), RANTES, and monocyte chemoattractant protein-1 (MCP-1)] in eosinophil recruitment. We have previously identified a role for MIP-1alpha in eosinophil accumulation in the lung and airway during allergic airway inflammation. We extend those studies using in vitro eosinophil chemotaxis to establish that both MIP-1alpha and RANTES are potent eosinophil chemotactic factors in lungs during allergic airway responses. Morphometric analysis demonstrated a peribronchial accumulation of eosinophils within the lungs beginning at 8 h, peaking at 24 h, and plateauing at 48-96 h after allergen (SEA) challenge. Utilizing whole-lung homogenates from allergen-challenged mice, in vitro eosinophil chemotactic assays demonstrated significant increases in eosinophil chemotactic activity with 8-h lung homogenates and peak activity with samples from 24-h lung homogenates. These data correlated with the morphometric analysis of peribronchial eosinophil accumulation in situ. When lung homogenates from allergen-challenged mice were preincubated in vitro with antibodies specific for MIP-1alpha, RANTES, or MCP-1, a significant reduction in eosinophil chemotaxis was observed with only MIP-1alpha and RANTES neutralization. Altogether, these studies indicate that RANTES and MIP-1alpha are major eosinophil chemotactic factors produced during allergic airway responses.

Mechanisms of hyperacute rejection in porcine liver transplantation. Antibody-mediated endothelial injury.

Hyperacute rejection after OLT is an unusual event. We have demonstrated previously that hepatic hyperacute rejection can occur in the presence of high titers of donor-specific cytotoxic antibody. This study addresses the issues of antibody localization within the allograft and the consequences of antibody deposition and complement activation within the transplanted organ. A porcine model of liver transplantation was used and 3 experimental groups were studied. Group I recipients (n = 6) were specifically sensitized to their liver donors with skin grafts from their liver donors before hepatic grafting. Group II recipients (n = 6) underwent third-party skin graft sensitization before liver transplantation. Group III recipients (n = 6) underwent liver grafting without prior sensitization. After liver transplantation, serum complement (CH50) levels declined promptly in all groups; a statistically significant drop as compared with control animals was seen only in group I (P < 0.05). Liver biopsies from donor-specific sensitized recipients showed massive injury by light microscopy within 30 min of revascularization, demonstrating fibrinoid necrosis of vessels and neutrophil influx. On immunofluorescent examination, liver specimens from donor-specific sensitized animals showed intense IgG, IgM, and C3 deposition in the vessels of the portal triads. Antibody deposition was not seen in third-party sensitized animals or control animals. On electron microscopy, control animals and third-party sensitized animals showed minimal ultrastructural alterations. In comparison, livers from donor-specific sensitized animals showed severe microvascular injury with destruction of endothelial cells, edema, hemorrhage, and hepatocyte necrosis. In a passive serum transfer experiment carried out by infusing serum from a skin graft-sensitized pig directly into the portal vein of the skin graft donor, severe liver injury was evident, with identification of antibody deposition by light and electron microscopy and by immunofluorescence. These studies demonstrate that tissue injury in hyperacute hepatic rejection is mediated by antibody deposition within the grafted liver and is associated with systemic activation of the complement cascade.

The role of strain variation in murine bleomycin-induced pulmonary fibrosis.

In the present study, mice have been used to study the effect of strain variation on the development of pulmonary fibrosis induced by a single intratracheal administration of bleomycin. The fibrotic response was quantitated by biochemical measurement of collagen synthesis and deposition. Results indicate that variations in these parameters of fibrosis are evident depending on the strain being tested. The C57BL/6 were high responders. DBA/2 and Swiss mice were intermediate responders, and BALB/c mice were low responders. These findings indicate that genetic variation influences the development of bleomycin-induced pulmonary fibrosis. Also, because certain aspects of the fibrotic response in this model are similar to those found in other conditions, this model may be useful as a basis to study the role of genetics in general fibrotic mechanisms.

Pulmonary alveolar macrophage function during acute inflammatory lung injury.

Pulmonary alveolar macrophages (PAM) are present during acute lung inflammation, yet the functional role of these cells in both the initiation and resolution of lung injury is not well defined. To better understand the relationship between PAM functional responses and the evolution of acute reversible lung injury, we examined the ability of both unstimulated and stimulated (PMA, zymosan) PAM to secrete reactive oxygen metabolites (superoxide anion O2-) and lysosomal enzymes (lysozyme, N-acetyl-B-D-glucosaminidase) at specific time points (0, 6, 12, 24, 48, and 72 h) after initiation of acute lung injury via reverse passive Arthus reaction in pathogen-free Sprague-Dawley rats. After acute lung injury, stimulated PAM produced increasing amounts of O2- compared with PAM from noninjured lungs. Maximal O2- production by PAM occurred at 24 h after lung injury, at which time a 3.5-fold and 50% increase in O2- production by PAM was observed when PAM were stimulated with PMA and zymosan, respectively. The amount of O2- generated by these cells slowly decreased during the next 48 h. Enhanced generation of O2- by PAM from injured lungs was not due to altered enzymatic activity of the O2--producing NADPH oxidase, nor was it due to an absolute increase in the NADPH oxidase in "activated" PAM. These observations suggest that increased O2- generation by PAM from injured lungs is due to enhancement of mechanisms responsible for induction of oxidase activity. In addition, a differential accumulation and secretion of lysozyme and N-acetyl-B-D-glucosaminidase activity by PAM was observed after acute lung injury.(ABSTRACT TRUNCATED AT 250 WORDS)

Potentiation of alpha-naphthyl thiourea-induced lung injury by prostaglandin E1 and platelet depletion.

alpha-Naphthyl thiourea ( ANTU ) produces pulmonary endothelial injury, pulmonary edema, and pleural effusions in rats in a dose-dependent manner. Since prostaglandins of the E series have been shown to modulate inflammatory responses in vivo and neutrophil and platelet function in vitro we investigated the effects of prostaglandin E1 (PGE1) on ANTU -induced lung injury. Systemic administration of 15-(S)-15-methyl-PGE1 (15-M-PGE1), a stable analog of PGE1, potentiated lung injury induced by ANTU in a dose- and time-dependent manner. 15-M-PGE1 (1 mg/kg, subcutaneously) administered 1 hour prior to ANTU treatment (1 mg/kg, intraperitoneally) resulted in a 164% increase (p less than 0.001) in pleural effusion formation and a 42% increase (p less than 0.02) in wet lung weight at 4 hours after ANTU administration. This was associated with increased pulmonary endothelial cell blebbing and gap formation with a decrease in the number of platelet thrombi in 15-M-PGE1-treated animals compared with controls. 15-(S)-15-methyl-prostaglandin F2 alpha, was less effective than 15-M-PGE1 in potentiating ANTU -induced lung injury. Platelet depletion, but not neutrophil depletion, also potentiated ANTU -induced lung injury, suggesting a protective role for platelets. Platelets isolated from 15-M-PGE1-treated animals demonstrated an approximately 50% decreased aggregation response to adenosine diphosphate. 15-M-PGE1 (1 mg/kg) treatment combined with platelet depletion resulted in a 1.7-fold increase (p less than 0.01) in pleural effusions in ANTU -treated (1 mg/kg) animals compared with platelet depletion alone. These studies indicate that systemic treatment of rats with 15-M-PGE1 will potentiate ANTU -induced lung injury. This injury may be in part secondary to the ability of 15-M-PGE1 to inhibit platelet function. However, platelet depletion studies suggest that 15-M-PGE1 has additional effects, possibly on endothelial cells and/or vascular smooth muscle cells that contribute to the potentiation of ANTU -induced lung injury.

Injury of rat pulmonary alveolar epithelial cells by H2O2: dependence on phenotype and catalase.

In a variety of inflammatory lung diseases, type I alveolar epithelial cells are more likely to be injured than are type II cells. Because oxidants have been implicated as a cause of injury in various inflammatory lung diseases, we evaluated the effects of differentiation on alveolar epithelial cell susceptibility to H2O2-induced injury. With the use of isolated rat type II cells in culture, we found that the cytotoxic effect of H2O2 increased between days 2 and 7, when type II cells are known to lose their distinctive type II properties and assume a more type I-like appearance. We previously reported that type II cells utilized both intracellular catalase and glutathione-dependent reactions to protect against H2O2. We therefore examined whether alterations in either of these protective mechanisms were responsible for the differentiation-dependent changes in sensitivity to H2O2. We found that catalase activity within alveolar epithelial cells decreased between 2 and 7 days in culture, whereas no changes were detected in glutathione-dependent systems. We then used a histochemical technique that detects catalase activity and found that type II cells within rat lungs possessed numerous catalase-containing peroxisomes, whereas very few were detected in type I cells. These findings demonstrate that as type II cells assume a type I-like phenotype, they become more susceptible to H2O2-induced injury. This increased susceptibility is associated with reductions in intracellular catalase activity, both in vitro and in vivo.

Glomerular injury and proteinuria in rats after intrarenal injection of cobra venom factor. Evidence for the role of neutrophil-derived oxygen free radicals.

The purpose of these studies was to determine how intravascular complement activation could lead to glomerular injury. Cobra venom factor (CVF) infused into the renal artery of rats resulted in increased excretion of protein in urine, which was maximal over the first 24 hours (51.2 +/- 6.0 mg/24 hours in CVF versus 14.1 +/- 0.9 mg/24 hours in saline-treated animals; P less than 0.001). Depletion of circulating neutrophils with anti-neutrophil serum significantly reduced the CVF-induced proteinuria in the first 24 hours (neutrophil depleted rats 22.7 +/- 2.8 mg/24 hours versus 63.4 +/- 9.9 mg/24 hours in neutrophil intact rats; P less than 0.005). Morphologic abnormalities (which were quantitated morphometrically) included accumulation of neutrophils in glomerular capillary loops, blebbing of endothelial cells, and epithelial cell foot process fusion. The increased protein excretion was reduced by 70% by simultaneous administration of catalase (23 +/- 4.3 mg/24 hours in CVF plus catalase versus 52.1 +/- 10 mg/24 hours in CVF alone; P less than 0.05). Catalase reduced glomerular endothelial cell blebbing and epithelial cell foot process fusion but not neutrophil accumulation in glomeruli as assessed by morphometry. In similar experiments superoxide dismutase, dimethyl sulfoxide, and deferoxamine did not prevent CVF-induced proteinuria. These studies, therefore, suggest that intravascular activation of complement in the rat causes glomerular injury and proteinuria which is dependent on neutrophils and upon the generation of hydrogen peroxide and/or its metabolites.

Mediation of IgA induced lung injury in the rat. Role of macrophages and reactive oxygen products.

Acute lung injury in the rat has been induced by the instillation of affinity-purified mouse monoclonal IgA antibody with specific reactivity to the hapten dinitrophenol coupled to albumin. As previously reported, this model of lung injury requires an intact complement system but is independent of neutrophils. In contrast to macrophages obtained by bronchoalveolar lavage from rats receiving IgA into the airways in the absence of intravenously injected antigen, macrophages obtained from the lungs of rats developing IgA immune complex-induced lung injury were significantly increased in number, showed greater spontaneous generation of O2.-, and demonstrated significantly enhanced O2.- responses in the presence of an added stimulus, phorbol ester. Inhibition studies in vivo suggested that the IgA-induced lung injury is mediated by oxygen radicals generated from lung macrophages. Pretreatment of animals with superoxide dismutase, catalase, the iron chelator, deferoxamine, or the hydroxyl radical scavenger, dimethyl sulfoxide, suppressed the development of lung injury. Morphologically the lungs of protected animals showed increased numbers of mononuclear cells within the alveolar compartment but little evidence of alveolar or capillary injury, in contrast to unprotected animals in which there was evidence of severe injury, both to microvascular interstitial endothelial cells as well as to alveolar lining epithelial cells. These studies suggest that acute lung injury in the rat induced by IgA immune complexes is mediated by oxygen radical formation and that the macrophage may be the principle effector cell, as compared to IgG immune complex induced lung injury, which is also oxygen radical mediated but in which the neutrophil is the effector cell.

Complement and neutrophil-mediated injury of perfused rat lungs.

It has previously been shown that systemic complement activation in rats leads to acute lung injury. We have now employed an ex vivo model of perfused rat lung which allows a more detailed examination of the role of circulating blood elements and pulmonary vascular pressure changes in the acute lung injury that occurs following cobra venom factor (CVF)-induced complement activation. In whole blood perfused lungs CVF infusion resulted in lung leukocyte sequestration, pulmonary vascular constriction, and acute lung injury. Lung injury was measured by quantitating the accumulation of [125I]-bovine serum albumin in lung parenchyma and alveolar lavage fluid. A histologic analysis of CVF-induced lung injury revealed the development of intravascular aggregates of platelets and accumulation of neutrophils, extensive bleb formation of pulmonary capillary interstitial endothelial cells, and interstitial and intraalveolar edema and intraalveolar hemorrhage. Experiments in lungs perfused with salt solution to which various blood elements were added showed that the development of lung injury was dependent on neutrophils and the interaction of CVF with heat-labile plasma components. The lung injury was not dependent on the presence of platelets or the pulmonary artery pressor response. Addition of catalase or erythrocytes to the lung perfusate significantly attenuated the acute lung injury. Like the development of lung injury, the pulmonary artery pressor response was dependent on the interaction of CVF with heat labile plasma components and neutrophils. The increase in PA pressure was not attenuated by the addition of catalase to the lung perfusate. These studies suggest that intravascular activation of the complement system leads to an acute microvascular injury which is dependent on neutrophils and the production of toxic oxygen metabolites. Pulmonary vascular constriction occurs independently of the associated lung injury and does not appear to be dependent on hydrogen peroxide production.

Phorbol myristate acetate produces injury to isolated rat lungs in the presence and absence of perfused neutrophils.

Because lung injury induced by phorbol myristate acetate (PMA) has been reported by some to be mediated by blood neutrophils (PMN) and by others to occur independently of PMN, we examined the PMN dependency of PMA-induced injury in isolated, perfused lungs of rats. Depending on dose, PMA added to medium perfusing isolated rat lungs produced injury in the presence or in the absence of added PMN. When a high concentration of PMA (57 ng/ml) was added to medium devoid of added PMN, perfusion pressure and lung weight increased. Superoxide dismutase (500 U/ml) and catalase (400 U/ml) added to the medium prior to PMA had no effect on the increases in lung weight or perfusion pressure. When a concentration of PMA (21 ng/ml or less) that did not by itself cause lungs to accumulate fluid was added to perfusion medium containing PMN (1 X 10(8)), superoxide was produced, perfusion pressure increased, and lungs accumulated fluid. Addition of superoxide dismutase and catalase to this preparation prevented the increase in lung weight, but not the increase in perfusion pressure. We conclude that high concentrations of PMA produce lung injury which is independent of neutrophils and oxygen radicals and that lower concentrations produce injury which is neutrophil-dependent and mediated by oxygen radicals. These results may explain why PMA-induced lung injury has variously been reported to be PMN-dependent in some systems and PMN-independent in others.

Role of oxygen radicals in phorbol myristate acetate-induced glomerular injury.

Phorbol myristate acetate (PMA) is known to be a potent activator of neutrophils and macrophages resulting in the generation of large amounts of oxygen-free radicals by these cells. When injected into the left renal artery of 250 to 300 g male Sprague-Dawley rats, PMA caused significant proteinuria compared to control rats which received normal saline (35.4 +/- 4 mg/24 hr in PMA treated vs. 14.1 +/- 0.9 mg/24 hr in saline control, P less than 0.02). The proteinuria was associated with evidence of glomerular injury. These PMA-induced alterations were not prevented by complement depletion but were prevented by prior depletion of neutrophils. The coinstillation of catalase prevented the development of the proteinuria (catalase + PMA 12.7 +/- 2.3 mg/24 hr vs. PMA alone 38.2 +/- 5.7 mg/24 hr, P less than 0.001) suggesting that H2O2 and/or its metabolites derived from neutrophils were important in the PMA-induced proteinuria. In contrast, superoxide dismutase (SOD) had no effect. We conclude that, following the intra-arterial injection of PMA, neutrophil-derived hydrogen peroxide and/or its metabolic products are capable of causing acute proteinuria in association with morphological alterations in glomeruli of rats.

Comparative O2-. responses of lung macrophages and blood phagocytic cells in the rat. Possible relevance to IgA immune complex induced lung injury.

IgA immune complex-induced lung injury in the rat is oxygen radical mediated and partially complement-dependent but develops fully after neutrophil depletion. The extent to which monocytes, lung interstitial macrophages, and alveolar macrophages may be involved in the development of lung injury in this model is unclear. To further understand the pathogenesis of IgA lung injury, we have examined the capacity of phagocytic cells isolated from different anatomic compartments of the lung to produce toxic oxygen-derived metabolites. [3H]Thymidine pulse labeling and autoradiography as well as in vivo phagocytosis studies were used to distinguish macrophages isolated from the alveolar and interstitial compartments. Lung interstitial macrophages were characterized ultrastructurally, cytochemically, and functionally. Interstitial macrophages were relatively uniform in size, had blunt pseudopodia, and contained almost no intracytoplasmic lamellar inclusions compared to alveolar macrophages. Similar to monocytes and alveolar macrophages, interstitial macrophages contained nonspecific esterase activity and exhibited the capacity to phagocytize latex and opsonized zymosan particles. Lung interstitial and alveolar macrophages incubated with IgA immune complexes, IgG immune complexes, or phorbol ester (PMA) produced similar amounts of O2-. in a dose-dependent manner. In contrast, peripheral blood neutrophils responded to IgG immune complexes and PMA but not to IgA immune complexes. Monocytes produced a small amount of O2-. in response to PMA but almost no O2-. in response to IgA or IgG immune complexes. These data are consistent with recent in vivo studies which indicate that IgA immune complex lung injury is neutrophil independent. The data provide direct in vitro evidence that lung interstitial and alveolar macrophages produce O2-. following incubation with PMA, IgA, or IgG immune complexes and may therefore contribute to the development of oxygen radical mediated lung injury.

Acute in vivo effects of human recombinant tumor necrosis factor.

Tumor necrosis factor is a peptide cytokine that induces hemorrhagic necrosis of some tumors and is responsible for the severe cachexia observed in advanced infectious diseases. We evaluated the acute effects of intravenous administration of purified human recombinant tumor necrosis factor in mice. With as little as 0.01 microgram/mouse (0.00045 mg/kg) a peripheral blood lymphopenia and neutrophilia developed as determined by flow cytometric analysis. At 1 microgram/mouse, the lymphopenia was both relative (21 +/- 3% versus 65 +/- 3%; p less than 0.001 treated versus control) and absolute (62 +/- 10 versus 229 +/- 29 X 10(4) cells/ml p less than 0.001). The neutrophilia was also relative (79 +/- 3% versus 34 +/- 3%; p less than 0.001 treated versus control) and absolute (237 +/- 26 versus 110 +/- 13 X 10(4) cell/ml; p less than 0.001). The neutrophilia was due to an increase in both mature and immature cells. At the higher doses the animals developed hypovolemic shock with an increased hematocrit and watery diarrhea occurred. Microscopic examination of the small bowel disclosed necrosis of the villi. Ultrastructural studies of the small bowel confirmed the necrosis and also showed severe endothelial cell damage, pyknotic nuclei, exocytosis of Paneth cell granules, and extravasation of red blood cells and neutrophils into the interstitium. A vascular leak syndrome developed with preferential fluid loss into the small and large bowel. These data demonstrate the potent in vivo effects of purified human recombinant tumor necrosis factor.

Evidence for the role of oxygen radicals in acute nephrotoxic nephritis.

Acute glomerular injury in the rat has been induced by the intrarenal, intraarterial infusion of sheep antibody to glomerular basement membrane (antiglomerular basement membrane). The antiglomerular basement membrane antibody has been verified to be of the variety that is complement and neutrophil dependent for the induction of acute proteinuria, which peaks during the first 24 hours. Following injection of the antibody, acute, intense, glomerular injury resulted, with the denuding of glomerular vascular basement membrane associated with extensive damage or destruction of glomerular endothelial cells and fusion of epithelial cell foot processes. Treatment of animals with catalase produced, in a dose-dependent manner, as much as 75% protection against glomerular injury, as assessed by reduction in the proteinuria. Treatment of animals with superoxide dismutase caused a small reduction in the degree of glomerular injury, again assessed by a reduction in proteinuria. However, this protective effect of superoxide dismutase was not found to be statistically significant. The hydroxyl radical scavenger, dimethyl sulfoxide, which has been shown to protect against endothelial cell injury following systemic activation of complement, was not protective in the anti-GBM model. Morphologically, glomeruli from catalase-protected rats showed numerous neutrophils but little or no evidence of injury of either glomerular endothelial or epithelial cells. These data suggest that acute glomerular injury produced by antiglomerular basement membrane is related to H2O2 production from activated neutrophils.

Acute hypotension due to platelet serotonin-induced chemoreflexes after intravenous injection of dextran sulfate in the rabbit.

The hypotension and bradycardia observed after intravenous injection of dextran sulfate in rabbits was prevented by prior depletion of circulating platelets, but was not prevented by depletion of the third component of complement or Hageman factor. Dextran sulfate injection caused immediate thrombocytopenia with temporary localization of platelets within lungs. Morphological analysis revealed platelet aggregates in lung capillaries. The platelets had changed shape and were in the process of degranulating. Serotonin and histamine levels in blood increased approximately 5-fold and 7-fold, respectively, after dextran sulfate injection. The cardiovascular events following dextran sulfate injection were mimicked by intravenous serotonin but not by intravenous histamine injection, although a combination of serotonin and histamine reproduced the pattern of blood pressure changes better than did either agent alone. Quantification of platelets trapped in lung revealed that the potential release of serotonin from trapped platelets could account for the rise in plasma serotonin concentration and the hemodynamic changes observed. Both the dextran sulfate-induced cardiovascular effects and serotonin-induced hypotension were markedly diminished by cutting vagus and depressor nerves, and were virtually abolished by carotid ligation in addition to nerve section. These results support the concept that platelet activation within rabbit lungs may cause hypotension via serotonin-induced chemoreflexes.