Infusion Of Zymosan-activated Plasma Research Articles

Evidence for Early Bacteremia/Endotoxemia and Inflammatory Responses after Trauma – Hemorrhage and Complement Activation in a Nonhuman Primate Model

There is criticism that the major line of evidence regarding posttraumatic inflammatory response and bacterial translocation has mainly come from experiments in rodents. The aim of this study was therefore to investigate the very early bacteremia/endotoxemia and inflammatory response in nonhuman primates. Six baboons were subjected to hemorrhagic shock (40 mm Hg mean arterial pressure) preceded by infusion of zymosan-activated plasma (to simulate trauma-associated complement activation) followed by reinfusion for 1 h. Measurements of complement and granulocyte (PMN) activation, cytokines of the inflammatory network, soluble cytokine receptors, and bacteria/endotoxin translocation (BT) were performed. There were significantly increased levels of PMN elastase, interleukin-6, and soluble tumor necrosis factors (TNF) receptors, but not of TNF. BT was evident from positive blood cultures, and from trendwise increased endotoxin plasma levels. Early liver damage was shown from elevated glutathione S-transferase (GST) plasma levels. Complement activation and hemorrhagic shock lead to an early inflammatory response together with bacteremia/endotoxemia as evidence of translocation from the gut already during the shock period in subhuman primates.

Changes in myocardial blood flow rates during hyperdynamic sepsis with induced changes in arterial perfusing pressures and metabolic need

To determine whether hyperdynamic sepsis is associated with dysregulation in the control of myocardial blood flow rates unrelated to hypotension or the use of anesthetic agents. Prospective, nonrandomized, controlled trial. Experimental laboratory. Fifteen mature male sheep (34 to 61 kg). Data were recorded in study subjects before and after the induction of sepsis following cecal ligation and perforation. Data were then recorded during: a) an infusion of prostaglandin E1 (PGE1), which decreased mean arterial perfusing pressure; and b) an infusion of zymosan-activated plasma, which increased mean pulmonary arterial pressures. Myocardial blood flow rates were measured by the radiolabeled microsphere technique and cardiac index was measured by the thermodilution technique. Cardiac index (change delta) postcecal ligation and perforation minus baseline (+2.3 +/- 1.0 L/min/m2; p < .01) was increased in the septic study. Blood flow rate to the left ventricle was simultaneously increased, and was not further affected when the PGE1 infusion decreased the mean arterial perfusing pressures (-19 +/- 4%). During the infusion of zymosan-activated plasma, mean pulmonary arterial pressures increased (50 +/- 30%) and right ventricular blood flow was increased (zymosan minus postcecal ligation and perforation study: delta 17.8 +/- 50 mL/100 g/min; p < .01). In this model of hyperdynamic sepsis, increases in blood flow to both the left and right ventricles were positively coupled to changes in respective ventricular work. From the interventional PGE1 and zymosan-activated plasma infusion studies, we found no evidence to support previous suggestions that the regulation of myocardial blood flow rates according to changes in perfusing pressure and/or metabolic oxygen need is significantly altered during hyperdynamic sepsis.

Physical and cytochemical properties of neutrophils activated in situ in the lung during ZAP infusion in sheep.

Increased retention of activated neutrophils in the lungs contributes to endothelial cell injury. However, characterization of the morphological changes that occur in neutrophils during activation in the pulmonary microcirculation has not been fully determined in vivo. Therefore, the present study was designed to determine structural and cytochemical properties of neutrophils in situ in pulmonary arterioles and alveolar capillaries during the infusion of zymosan-activated plasma (ZAP) or plasma (control) in anesthetized sheep. Quantitative morphological methods showed that ZAP infusion caused significant retention of neutrophils in alveolar capillaries [2.19 +/- 0.40 (SD) x 10(8) neutrophils/ml of capillary blood volume] and pulmonary arterioles (1.02 +/- 0.46 x 10(8) neutrophils/ml of arterial blood volume) compared with plasma infusion (1.03 +/- 0.15 and 0.30 +/- 0.10 x 10(8) neutrophils/ml, respectively; P < 0.05). Harmonic mean diameter of ZAP-activated neutrophils in situ (7.19 +/- 0.44 microns) was significantly greater than the diameter of neutrophils in plasma-treated sheep (6.29 +/- 0.17 microns; P < 0.05). Neutrophil cross-sectional area (54 +/- 3 microns2) and volume (248 +/- 27 microns3) in situ in alveolar capillaries were also significantly greater in ZAP-treated sheep than in control sheep (41 +/- 4 microns2 and 184 +/- 9 microns3, respectively; P < 0.05). Similarly, microvascular neutrophils in ZAP-treated sheep were vacuolated and elongated, filamentous actin was redistributed peripherally, and the cells were degranulated. We conclude that during ZAP infusion, neutrophils become enlarged and degranulated in pulmonary microvessels, especially in alveolar capillaries. The structural and cytochemical changes that occur are consistent with the hypothesis that neutrophil activation is accompanied by alterations in neutrophil physical properties, alterations that may facilitate retention and contribute to endothelial cell injury.

Infusion of zymosan-activated plasma affects neutrophils in peripheral blood and bone marrow in sheep.

Intravenous infusion of zymosan-activated plasma (ZAP) in sheep results in acute lung injury mediated in part by free radical release from stimulated neutrophils that are retained in increased numbers in the pulmonary microcirculation. ZAP infusion is also associated with long-term reduction in elicited superoxide anion generation from segmented neutrophils in the circulating and bone marrow pools for at least 24 h after the infusion. The purpose of our study was to investigate the effect of ZAP infusion on leukocyte counts and neutrophil granule-associated enzyme activity in the circulating and bone marrow pools. Cytochemical methods were used to characterize enzyme activity in primary granules (acid phosphatase and myeloperoxidase) and secondary granules (alkaline phosphatase). During the infusion period, neutrophil differential counts decreased less in venous blood than in matched arterial blood. Release from bone marrow stores was evident as increased numbers of circulating band neutrophils during and after the infusion. Neutrophils in venous and arterial blood smears were degranulated acutely during and 1-3 h after the infusion was stopped. Band and segmented neutrophils in bone marrow also appeared slightly degranulated 1-2 h after the infusion. In vitro experiments showed that band and segmented neutrophils in bone marrow degranulated in response to ZAP incubation. Immature polymorphonuclear leukocytes did not degranulate. Five to 24 h after ZAP infusion, enzyme activity in circulating and bone marrow neutrophils was at baseline levels, suggesting that new cells were being released into the circulating pool because degranulated neutrophils do not synthesize new granules. Another indication of a long-term effect in bone marrow were slight decreases in the percentage of immature polymorphonuclear leukocytes and band neutrophils and a significant decrease in the percentage of eosinophils, all of which coincided with a significant increase in the percentage of segmented neutrophils. Our results demonstrate that circulating complement anaphylatoxins affect neutrophils in bone marrow as well as the vascular space.

Downregulation of blood and bone marrow neutrophils decreases expression of acute lung injury in sheep.

We have shown that infusion of zymosan-activated plasma (ZAP) in sheep causes acute lung injury and downregulates peripheral blood neutrophils in that elicited superoxide release is reduced for at least 24 h after the infusion. The present study was designed to test the following hypotheses: 1) peripheral blood neutrophils are representative of neutrophils marginated in the pulmonary circulation, 2) blood neutrophils are downregulated because neutrophils developing in bone marrow are similarly affected, and 3) downregulated neutrophils have a reduced capacity to produce tissue injury. In a series of experiments in 21 sheep, we showed that elicited superoxide release was similar in peripheral blood neutrophils and in marginated neutrophils washed out of the pulmonary vascular bed. Measurements of superoxide release from blood and bone marrow neutrophils collected 2-24 h after ZAP infusion revealed progressive downregulation with time and greater downregulation of superoxide release in bone marrow neutrophils compared with peripheral blood neutrophils. Finally, after downregulating peripheral blood neutrophils, subsequent infusion of ZAP in conscious sheep produced sequestration of neutrophils in the pulmonary circulation but failed to produce a sustained increase in lung lymph protein clearance. The results suggest that neutrophil downregulation, as measured in vitro, is expressed in vivo as reduced ability of neutrophils to produce tissue injury when challenged by an activating agent.

Effect of zymosan-activated plasma on the deformability of rabbit polymorphonuclear leukocytes.

Intravascular infusion of inflammatory mediators causes a sudden neutropenia due to the sequestration of polymorphonuclear leukocytes (PMN) within the microvasculature of the lung and other organs. This sequestration could be due to a decrease in the ability of PMN to deform and pass through the narrow capillary bed. The purpose of this study was to determine if the complement fragments present in zymosan-activated plasma (ZAP) caused a rapid stiffening of PMN. The PMN deformability was determined by measuring the pressure required to pass PMN through a polycarbonate filter containing 5-micron pores at a constant flow rate as well as the extraction of PMN compared with red blood cells and 125I-labeled albumin by the filter. The role of the cytoskeleton in PMN deformation was examined in studies where F-actin formation was inhibited using cytochalasin B or microtubule assembly was inhibited using colchicine. The results showed that treatment with ZAP induced a rapid decrease in PMN deformability. Inhibiting the formation of F-actin made the unstimulated PMN more deformable and reduced the stiffening induced by ZAP. In contrast, inhibition of microtubule reassembly did not alter either normal deformability or the ZAP-induced decrease in deformability. In vivo, colchicine increased normal PMN margination but did not inhibit the rapid sequestration of PMN induced by infusion of ZAP. These studies indicate that ZAP induces a rapid decrease in PMN deformability that is mediated through the cytoskeleton. They suggest that this decrease is due to the polymerization of F-actin.

The role of CD18-mediated adhesion in neutrophil sequestration induced by infusion of activated plasma in rabbits.

Infusion of activated plasma induces neutropenia and sequestration of neutrophils within the microvasculature. This study examined the role of the adhesion glycoprotein complex, CD11/CD18, in this sequestration. Rabbits pretreated with either the anti-CD18 monoclonal antibody (mAb) 60.3 or saline were given infusions of zymosan-activated plasma (ZAP) or saline. The effect of mAb 60.3 on the changes in circulating neutrophil counts, radiolabeled neutrophil kinetics in the lung, and the pulmonary microvascular accumulation of neutrophils induced by ZAP infusion was determined. The data show that pretreatment with mAb 60.3 did not inhibit either the rate of onset or the severity of the neutropenia but prevented the sustained neutropenia. In addition, mAb 60.3 completely prevented the ZAP-induced changes in radiolabeled neutrophil kinetics and largely inhibited the accumulation of neutrophils within the capillaries and the small vessels when evaluated after 15 min of ZAP infusion. We conclude that neutrophil accumulation is a two-step process, the first occurring through a CD18-independent mechanism that may involve a stimulus-induced decrease in neutrophil deformability and acts to slow neutrophil transit through the lung. The second step requires CD18-dependent adhesion and is needed for prolonged accumulation of neutrophils within the pulmonary microvasculature.

PGE1 inhibited PMN attachment to air emboli in vivo during infusion of ZAP without preventing lung injury.

Prostaglandin E1 (PGE1) treatment of neutrophils inhibits their adherence to substrates in vitro, including endothelial cell monolayers. Demonstration that PGE1 inhibits neutrophil adherence in vivo in the lung, however, is complicated by PGE1 effects on cells other than neutrophils, such as endothelial cells. To determine whether PGE1 inhibits neutrophil adherence properties in vivo, we used air emboli as intravascular targets for neutrophil attachment. Four experimental conditions were studied in anesthetized and awake sheep that were treated with 1) PGE1 and air emboli, 2) saline and air emboli, 3) PGE1 and zymosan-activated plasma (ZAP) + air emboli, and 4) saline and ZAP + air emboli. PGE1 (30 ng.kg-1.min-1) or saline was infused continuously 1 h before and 1 h during the infusion of air emboli (group 1; n = 13 sheep) or ZAP + air emboli (group 2; n = 13 sheep). The number of neutrophils (PMNs) attached to air emboli in four anesthetized sheep per condition was significantly less in sheep given PGE1 and ZAP + air emboli [8 +/- 3 (SD) PMNs/mm of embolus perimeter] than in the other three conditions (14-21 PMNs/mm; P less than 0.05). Repeated experiments in five awake sheep per group showed that PGE1 treatment did not prevent increased lung lymph protein clearance in either group compared with saline treatment. We conclude that PGE1 specifically inhibited attachment of ZAP-activated neutrophils to air emboli in vivo. The lack of pathophysiological protection suggests that PGE1-induced alterations in neutrophil attachment properties were independent of other cellular activation responses.

Neutrophil-associated lung injury after the infusion of activated plasma

Previous studies from our laboratory have shown that the infusion of zymosan-activated plasma (ZAP) caused large numbers of neutrophils (PMN) to accumulate in the lung. Although PMN are known to be activated by ZAP, it is unclear whether PMN delayed in the lung by ZAP infusion actually cause lung injury. The present study was designed to examine this question by measuring airway epithelial and endothelial injury. Airway epithelial injury was determined by depositing a known dose of fluorescein isothiocyanate-labeled dextran in the lung and measuring its appearance in the blood, and endothelial injury was measured by injecting colloidal carbon and measuring its accumulation in the microvasculature of the lung. The data show that ZAP infusion caused a mild epithelial and endothelial injury that did not increase either extravascular water or protein. This injury could be prevented either by depleting the animals of PMN or by pretreating them with indomethacin. In addition, the effect of ZAP infusion could be partially restored by transfusing donor PMN into the PMN-depleted animals. We conclude that ZAP infusion produces a mild lung injury that is dependent on PMN and the products of the cyclooxygenase pathway of arachidonic acid metabolism.

Oxidant-mediated, CD18-dependent microvascular dysfunction induced by complement-activated granulocytes.

To determine the mechanisms whereby complement-activated granulocytes induce microvascular dysfunction in skeletal muscle, we examined the effect of antineutrophil serum (ANS), IB4 (a monoclonal antibody that inhibits CD18-dependent neutrophil adherence), xanthine oxidase inhibition or inactivation, deferoxamine, and catalase on the increase in canine gracilis muscle microvascular permeability induced by intravascular administration of zymosan-activated plasma (ZAP). Changes in vascular permeability were assessed by measurement of the solvent drag reflection coefficient (sigma) for total plasma proteins, and the extent of neutrophil infiltration was estimated by assessing muscle myeloperoxidase activity. ZAP infusion was associated with a marked increase in vascular permeability compared with control muscles that received no treatment or to muscles treated with zymosan heat-inactivated plasma (ZIP) (sigma = 0.51 +/- 0.04, 0.89 +/- 0.02, and 0.90 +/- 0.01, respectively). Estimates of sigma in animals rendered neutropenic with ANS, or treated with IB4, deferoxamine, or catalase before ZAP infusion were not significantly different from values obtained in control or ZIP-treated muscles (sigma = 0.96 +/- 0.02, 0.88 +/- 0.03, 0.85 +/- 0.02, and 0.79 +/- 0.01, respectively). However, xanthine oxidase inactivation or inhibition provided no protection from this ZAP-induced microvascular dysfunction (sigma = 0.58 +/- 0.02 and 0.58 +/- 0.01, respectively). In addition, neutropenia and inhibition of neutrophil adherence also prevented ZAP-induced increases in vascular resistance and tissue neutrophil infiltration.(ABSTRACT TRUNCATED AT 250 WORDS)

Failure of therapy with 2,3-dihydroxybenzoic acid to modify the course of sepsis-induced lung injury

Oxidant-induced injury of the pulmonary microvasculature reportedly contributes to an increase in microvascular permeability and pulmonary hypertension, both of which are principal features of acute lung injury (ALI). We tested the hypothesis that antioxidant therapy with 2,3-dihydroxybenzoic acid (DHB), initiated in awake sheep after the development of sepsis-induced ALI, would ameliorate the progression of these lesions. DHB has many actions that suggested to us the potential for demonstrating benefit in ALI complicating sepsis; it is a nontoxic hydroxyl-radical scavenger that also inhibits the cyclooxygenase pathway and acts as a weak iron chelator. In preliminary experiments, we demonstrated that pretreatment with DHB prevented an increase in mean pulmonary arterial pressure, plasma thromboxane A2, measured as its metabolite thromboxane B2, and lymph total protein clearance that otherwise followed an infusion of zymosan-activated plasma (ZAP) in sheep. In subsequent experiments, 12 additional sheep were rendered septic by cecal ligation and perforation. Twenty-four to 36 h after cecal ligation and perforation, an increase in lung microvascular permeability was confirmed, because pulmonary lymph flow had increased by 82% while the mean lymph-to-plasma total protein ratio was unchanged from baseline. At this point, six sheep were then treated with parenteral DHB and six with DHB vehicle for the subsequent 24 h. In contrast to the demonstrated benefit of DHB pretreatment in preventing ALI secondary to an infusion of ZAP, the progressive increase in lymph total protein clearance that complicated septic lung injury in the DHB vehicle group throughout this 24-h study period was not ameliorated in the DHB treatment group. However, DHB did prevent a modest increase in mean pulmonary arterial pressures that was demonstrated in the DHB vehicle group throughout this 24-h treatment period. Although pretreatment prevented ALI after a ZAP infusion, we conclude that DHB only incompletely modified disease progression when administered after the onset of sepsis-induced ALI because it ameliorated the pulmonary hypertensive response without concurrently modifying an increase in lung microvascular fluid flux.

Zymosan-activated plasma-mediated thromboxane production by the perfused rabbit liver and isolated hepatocytes: Involvement of calcium

The present study investigates the mechanism of zymosan-activated plasma (ZAP)-mediated eicosanoid production by the isolated, perfused rabbit liver and describes ZAP-induced eicosanoid stimulation in cultured hepatocytes. Perfused livers receiving untreated plasma demonstrated no significant changes in portal venous pressure or the rates of release of lactic dehydrogenase or acid phosphatase activity (indicator of cellular injury). The control group livers demonstrated stable rates of release for 6-keto PGF1α and thromboxane B2 (TXB2). In contrast, the infusion of ZAP alone resulted in a rapid but transient releaes of TXB2 from the livers. No significant changes in perfusion pressure or enzyme release were observed following ZAP administration. Perfusion of livers with a calcium-free buffer decreased the basal rates of both 6-keto PGF1α and TXB2 production and significantly, but not completely, attenuated the ZAP-mediated increase in hepatic TXB2 production. Perfusion of livers with nifedipine (3 μM) had no effect on ZAP-mediated TXB2 production in this model. Isolated hepatocytes responded to ZAP-treatment with significant increases in TXB2 production. These data suggest that activated fluid phase complement components induce thromboxane production by specific cells within the liver and that this stimulation is partially dependent upon the release of intracellular calcium but independent of complement-mediated cellular injuiry.

Zymosan-activated plasma causes prolonged decreases in PMN superoxide release in sheep

It is well established that activation of neutrophils within the pulmonary circulation produces acute lung injury in which adherence of neutrophils to endothelial cells is an obligatory step in the mechanism of injury. The effects of in vivo activation of neutrophils on the in vitro responses of these cells to stimulation have not been determined, although such information may be important in understanding how different etiological factors may interact to produce infection or acute respiratory failure. By using an assay to sequentially measure superoxide anion (O2-) release from adherent neutrophils stimulated with phorbol myristate acetate (PMA), we measured the in vitro activation response of peripheral blood neutrophils isolated before and 24 h after infusion of zymosan-activated plasma (ZAP; or untreated plasma as a control), air bubbles, or PMA in awake, instrumented sheep. Each of the three inflammatory agents produced an increase in lung microvascular permeability characteristic of acute lung injury; control plasma did not. For the in vivo ZAP experiments, stimulated O2- release in vitro by using PMA was approximately 50% lower (P less than 0.05) for neutrophils isolated 24 h after the in vivo infusion (4.3 +/- 0.8 nmol/500,000 cells) than before (8.1 +/- 0.2 nmol/500,000 cells). For the air emboli or PMA in vivo experiments, there were no changes in neutrophil activation responses in vitro. Similarly, infusion of control plasma did not result in reduced neutrophil O2- release. These results show that alterations in the inflammatory potential of neutrophils may occur in vivo and that such alterations appear to be dependent on the mechanism and agent by which lung injury is produced.

Neutrophil kinetics in rabbits during infusion of zymosan-activated plasma.

Complement activation in vivo produces neutropenia and pulmonary sequestration of neutrophils (PMNs) whereas in vitro activation increases PMN adherence and decreases PMN deformability. The present study examined PMN kinetics in vivo to determine if this sequestration was specific to the lung. Venous or arterial injections of radiolabeled PMNs were given to animals receiving infusions of zymosan-activated plasma (ZAP) or saline, and the PMN distribution was evaluated 10 min later. In control animals, the relative size of the marginated and circulating PMN pools was similar after venous or arterial injection and regional PMN retention increased as blood velocity slowed. ZAP infusion produced threefold increases in PMNs within pulmonary capillaries after venous injection and PMN retention was independent of blood velocity. After arterial injection, ZAP infusion produced PMN sequestration in all organs. Rigid (glutaraldehyde-fixed) PMNs injected into control rabbits showed increased lung recoveries similar to those of fresh PMNs injected into ZAP-treated rabbits. We conclude that activation of the complement system causes PMN sequestration in both the pulmonary and the systemic microvasculature and that the decrease in PMN deformability that occurs with activation of the PMN may be important in the genesis of PMN sequestration.

Characteristics of oedema formation induced by N‐formyl‐methionyl‐leucyl‐phenylalanine in rabbit skin

1. The characteristics of N-formyl-methionyl-leucyl-phenylalanine (FMLP)-induced oedema formation were investigated in vivo in rabbit skin. 2. FMLP injected intradermally alone induced a small increase in plasma leakage, but marked synergism with prostaglandin E2 (PGE2) in producing oedema responses was observed. In the presence of PGE2, FMLP was equiactive with C5a des Arg and 100-1000 times more active than histamine in terms of permeability-increasing activity. The response to FMLP was not dependent on endogenous histamine release. 3. FMLP-induced responses were of long duration (t1/2 approximately 40-50 min) when compared with bradykinin (t1/2 approximately 4-5 min). 4. The activity of a range of N-formyl peptides in increasing vascular permeability in skin correlated well with their activity as neutrophil stimulants in vitro. 5. Intravenous infusion of zymosan-activated plasma (ZAP) resulted in transient neutropenia and inhibition of oedema formation induced by FMLP and C5a des Arg in the skin. Responses to bradykinin were unaffected by the infusion of ZAP. 6. Intravenous injection of the non-steroidal antiinflammatory drug, ibuprofen, resulted in an inhibition of FMLP-induced, but not histamine-induced, oedema formation. This effect was independent of cyclo-oxygenase inhibition and the drug did not induced neutropenia. 7. Intravenous injection of the microtubule blocking agent colchicine inhibited FMLP-induced oedema formation. Responses to bradykinin were unaffected. When colchicine was administered after intradermal FMLP, subsequent plasma leakage was abolished. 8. The inference that receptors have evolved to bacterial secretions (i.e. FMLP) and products of the interaction of bacterial cell walls with tissue fluid (i.e. C5a des Arg), is consistent with the hypothesis that oedema formation is fundamentally a functional process concerned with regulating microbial lysis and opsonisation in an infected tissue.

Effects of hypoxia and complement activation on lung hemodynamics and fluid exchanges in unanesthetized goats.

The effects of hypoxia and zymosan-activated plasma (ZAP) on lung hemodynamics and fluid exchange were studied in unanesthetized goats. Hypoxia produced a sustained increase in pulmonary arterial pressure (PPA), but had no effect on lung lymph flow and protein content. 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. When ZAP was infused at a simulated altitude of 4,000 m, there was a further increase in PPA followed by pulmonary vasodilatation, and about 1 h later it was almost back to the control level. No obvious difference in lung lymph flow and protein content was observed between the groups treated with ZAP alone and ZAP plus hypoxia. We conclude that infusion of ZAP into hypoxic animals reduces pulmonary vasoconstriction and that the increase in pulmonary microvascular permeability associated with ZAP is not enhanced by hypoxic conditions occurring at a simulated altitude of 4,000 m.

Complement-mediated pulmonary edema in sheep

Incubation of plasma with zymosan results in complement activation. Infusion of zymosan-activated plasma (ZAP) into the pulmonary circulation in sheep results in a transient rise in pulmonary vascular resistance, accompanied by hypoxemia. This is associated with production of the vasoactive prostaglandin metabolite, thromboxane. We hypothesized that ZAP infusion caused pulmonary edema and may transiently alter pulmonary permeability to protein. Two sets of experiments were conducted involving six sheep. Three sheep underwent open lung biopsies during ZAP infusion, and electron microscopy documented early reversible interstitial pulmonary edema in response to ZAP infusion. Three other sheep were prepared with lung lymph fistulae and subjected to a similar infusion of ZAP. These studies documented a rapid rise in lymph flow and lymph protein clearance, occurring immediately upon infusion of ZAP. These experiments demonstrate a rapid reversible interstitial edema in sheep lung in response to ZAP infusion and suggest that permeability to protein may be transiently altered as a result of this injury.

The inflammatory activity of activated complement in ovine and bovine mammary glands.

Infusion of zymosan-activated plasma (ZAP), a source of activated complement, induced inflammation in non-lactating mammary glands but not in lactating glands of ewes and cows. Interstitial injection of ZAP in lactating ovine glands also failed to induce inflammation as assessed by the neutrophil content of milk. The inflammatory activity of ZAP in non-lactating glands was inhibited by infusion of whole milk but not by skimmed milk. These results suggest that components of whole milk mask the inflammatory activity of activated complement. In addition, lactating mammary tissue of ewes and cows may lack sensitivity to the inflammatory effect of activated complement. Thus, activated complement cannot be an important mediator of inflammation in the lactating gland.

Species variability in the cardiovascular and hematologic effects of zymosan-activated plasma infusion

The anaphylatoxins have been implicated in the pathogenesis of endotoxin shock and the adult respiratory distress syndrome. Both endotoxin and zymosan activate the complement pathway. Because there are marked species differences in the cardiovascular and hematologic effects of endotoxin infusion, the purpose of this study was to compare the effects of zymosan-activated plasma (ZAP) infusion in dogs, sheep, and baboons. ZAP was infused (0.11 ml/kg/min for 60 min) into dogs ( n = 5), baboons ( n = 5), and sheep ( n = 3). The infusion of ZAP resulted in significant changes in heart rate (HR) ( P < 0.03), mean arterial pressure (MAP) ( P < 0.002), pulmonary artery pressure (PAP) ( P < 0.004), cardiac index (CI) ( P < 0.034), and extra vascular lung water (EVLW) ( P < 0.001). A specific difference between the species' response to ZAP infusion was present when evaluating the effect of ZAP on MAP ( P < 0.02), HR ( P < 0.003). EVLW ( P < 0.001), platelet count ( P < 0.01), and white blood cell count ( P < 0.01). The main species differences in the changes in MAP, HR, and platelet count were an increase in MAP, decrease in HR, and decrease in platelet count that occurred in dogs. The species difference in the WBC count was the result of ZAP-induced neutropenia in sheep versus a leukocytosis in dogs. Unlike dogs and baboons, sheep developed an increase in EVLW. Like endotoxin, the cardiovascular and hematologic effects of ZAP infusion are species dependent. Of the three species studied, dogs were distinctly different. Except for the changes in EVLW sheep and baboons responded in a qualitatively similar fashion, although the magnitude of the sheep's responses was always greater.

Amphotericin B causes aggregation of neutrophils and enhances pulmonary leukostasis.

The influence of amphotericin B (AmB) on the aggregation of polymorphonuclear leukocytes (PMN) was examined by means of in vitro aggregometry as well as in an in vivo model of pulmonary leukostasis (PL). The AmB caused a dose-dependent aggregation of PMN that was partially blocked by addition of serum to the drug prior to its reaction with the PMN. No aggregation of PMN was seen after the addition of nystatin, a similar polyene antibiotic. In vivo studies were conducted in rabbits, where PL was induced by an intravenous infusion of zymosan-activated plasma (ZAP) or phorbol myristate acetate (PMA), and the degree of resulting leukostasis was expressed as the number of PMN per high power field (HPF). Animals in the control group had 6 +/- 3 PMN/HPF. This number increased to 12.4 +/- 5.6 when ZAP was infused and postmortem examination was performed 1 h later and to 17.4 +/- 4 if the examination was performed after 24 h. These numbers increased more than twice when AmB (1 mg/kg) was infused together with the ZAP. Increased PL after the infusion of ZAP and AmB was not attenuated by prior administration of methylprednisolone (30 mg/kg) to the animals. Infusions of AmB per se caused no significant PL but did cause increased pinocytosis in the pulmonary endothelium. Enhancement of PL by AmB was also examined in a model of PMA-induced PL.(ABSTRACT TRUNCATED AT 250 WORDS)