Neutropenic Animals Research Articles

Experimental murine acid aspiration injury is mediated by neutrophils and the alternative complement pathway.

Acid aspiration may result in the development of the acute respiratory distress syndrome, an event associated with significant morbidity and mortality. Although once attributed to direct distal airway injury, the pulmonary failure after acid aspiration is more complex and involves an inflammatory injury mediated by complement (C) and polymorphonuclear leukocytes. This study examines the injurious inflammatory cascades that are activated after acid aspiration. The role of neutrophils was defined by immunodepletion before aspiration, which reduced injury by 59%. The injury was not modified in either P- or E-selectin-knockout mice, indicating that these adhesion molecules were not operative. C activation after aspiration was documented with immunochemistry by C3 deposition on injured alveolar pneumocytes. Animals in which C activation was inhibited with soluble C receptor type 1 (sCR1) had a 54% reduction in injury, similar to the level of protection seen in C3-knockout mice (58%). However C4-knockout mice were not protected from injury, indicating that C activation is mediated by the alternative pathway. Finally, an additive effect of neutrophils and C was demonstrated whereby neutropenic animals that were treated with sCR1 showed an 85% reduction in injury. Thus acid aspiration injury is mediated by neutrophils and the alternative C pathway.

The Role of Hyperuricemia in the Increased Cytokine Production After Lipopolysaccharide Challenge in Neutropenic Mice

AbstractPatients with severe granulocytopenia are more susceptible to severe infections and sepsis. Proinflammatory cytokines such as tumor necrosis factor-α (TNF ), interleukin-1α (IL-1α), and IL-1β play an important role in the pathophysiology of sepsis. The profile of these proinflammatory cytokines after lipopolysccharide (LPS) challenge in cyclophosphamide-induced neutropenic mice was assessed, and possible mechanisms responsible for the modified cytokine production were studied. After LPS, both circulating concentrations of TNF and IL-1α in neutropenic mice were 50% to 200% higher than those of controls, whereas IL-1β concentrations were not modified. The kinetics of cytokine production were similar in neutropenic and control animals. The susceptibility of neutropenic mice to an LPS challenge was increased. The observed overproduction of TNF and IL-1α was not due to a direct effect of cyclophosphamide treatment. Because circulating concentrations of uric acid were increased in the neutropenic mice, the effect of hypouricemic treatment with allopurinol and sodium bicarbonate was investigated; such treatment in neutropenic mice challenged with LPS was followed by an improved survival and a reduced proinflammatory cytokine production towards the concentrations in control mice. Hyperuricemia induced by repeated administrations of uric acid in normal mice led to an increased TNF production after LPS. In conclusion, neutropenic mice respond with enhanced cytokine production and increased susceptibility to an LPS challenge, and hyperuricemia probably plays an important role in this phenomenon.

Induction of Cytokine-Induced Neutrophil Chemoattractant (CINC) mRNA in the Lungs of Septic Rats

To study cytokine-induced neutrophil chemoattractant (CINC) mRNA induction in lungs of normal, neutropenic, and adrenalectomized rats after intraperitoneal Escherichia coli lipopolysaccharide (LPS) administration and in cultured rat pulmonary cell lines after exposure to mediators of the septic response. Northern blotting was used to assay relative CINC mRNA levels and a colorimetric myeloperoxidase assay was used as a measure of neutrophil infiltration. After a single dose of LPS, rapid induction of CINC mRNA coincided with neutrophil infiltration into lungs, a response that lasted approximately 12 to 24 hours. Multiple LPS treatments resulted in a similar CINC response, but a more prolonged myeloperoxidase response. CINC mRNA induction in lungs was heightened 30% in adrenalectomized animals and 400% in neutropenic ones. LPS and cytokines induced CINC mRNA in cultured endothelial and epithelial cells. Induction of CINC mRNA expression in pulmonary endothelial and/or epithelial cells by systemic LPS or cytokines may play a role in mediating neutrophil infiltration into lungs during sepsis. Markedly increased CINC induction in the lungs of neutropenic animals suggests that neutrophils may act to inhibit expression of this chemoattractant via a negative feedback mechanism.

Neutrophil-independence of the initiation of colonic injury

Although the role of neutrophils in the pathogenesis of several forms of gastrointestinal injury has been well demonstrated, their role in the development of experimental colonic injury is less clear. To examine whether neutrophils play a role in the development of experimental colitides, the effects of a sustained neutropenia on multiple indices of colonic injury in rats was examined 24 hr following the initiation of colitis with the intrarectal application of acetic acid, trinitrobenzene sulfonic acid (TNBS)-ethanol, or the potent proinflammatory agent, phorbol-12-myristate-13-acetate (PMA). In comparison to animals with normal neutrophil counts and colitis induced by any of the three agents, no attenuation in macroscopic damage or histopathologic injury was observed in neutropenic animals exhibiting a greater than 95% reduction in circulating neutrophils and 85% reduction in tissue-associated myeloperoxidase activity. Although the tissue edema associated with acetic acid or PMA-induced colitis was not reduced by neutropenia, the colonic edema associated with TNBS colitis was attenuated by prior neutrophil depletion with anti-neutrophil antiserum. Despite our initial hypothesis that neutrophils played a key role in the genesis of experimental colitis (especially that induced by PMA), the results demonstrated that these cells are not essential for the development of the major pathological features of colitis induced by this agent, acetic acid, or TNBS. Although these results support the proposal that in these models of colitis, inflammation develops secondary to injury (rather than the converse), further studies will be necessary to elucidate the role of inflammatory cells other than neutrophils in the genesis of experimental colitides.

Interleukin-1 and related pro-inflammatory cytokines in the treatment of bacterial infections in neutropenic and non-neutropenic animals.

Bacterial infections in the immunocompromized host cause considerable mortality, and even the recently developed antimicrobial strategies often fail to cure these infections, especially in granulocytopenic patients. Cytokines and hematopoietic growth factors have been shown to stimulate host defense mechanisms in vitro and in vivo. We discuss the possible role of the pro-inflammatory cytokines interleukin-1, tumor necrosis factor-alpha, interleukin-6 and interleukin-8 as modulators of host resistance to bacterial infections. Interleukin-1 has been shown effective in various animal models of potentially lethal bacterial infection, even during severe granulocytopenia. The protective mechanism of interleukin-1 may be mediated via downregulation of cytokine receptors and cytokine production, and via induction of acute phase proteins. Moreover, in subacute and chronic infections interleukin-1 interferes with microbial outgrowth, via mechanisms that have only been partially elucidated.

Salicylate potentiates amikacin therapy in rodent models of Klebsiella pneumoniae infection.

In vitro effects of salicylate on Klebsiella pneumoniae include capsule repression and enhanced aminoglycoside activity. The effects of high-dose salicylate on amikacin therapy were assessed in animal models. In a mouse lethality model, amikacin protective doses were reduced up to 12-fold by salicylate doses of 277 mg/kg. Salicylate alone increased the LD50 for K. pneumoniae in mice by nearly 1 log unit. In a lobar pneumonia model, rats received salicylate-amikacin intravenously 3 days after challenge, and bacteria were quantified from lung tissue on day 7. Rats receiving one-fourth the minimum effective amikacin dose had 1 x 10(6) cfu/g of lung, while no bacteria were detected in those also receiving salicylate (173 mg/kg). At one-eighth the minimum effective dose of amikacin, salicylate prevented the development of pneumonia. Salicylate potentiation of antimicrobial therapy far surpassed that observed in vitro, yet the potentiation was abolished in neutropenic animals. Thus, salicylate may enhance phagocytosis rather than antibiotic action in therapy.

Complement C5a-mediated myocardial ischemia and neutrophil sequestration: two independent phenomena

The intracoronary infusion of complement C5a causes a decrease in coronary blood flow and contractile dysfunction mediated by thromboxane (TxA2) and leukotrienes. Although these effects are accompanied by polymorphonuclear leukocyte (PMN) sequestration, the role of PMNs and the source of these eicosanoids remain unknown. To assess the contribution of PMNs to the C5a-induced myocardial ischemic response, the left anterior descending (LAD) coronary artery of pigs (n = 13) was cannulated and pump perfused at constant pressure with either normal arterial blood or neutropenic arterial blood (PMN count 0.02 x 10(3) cells/microliters) obtained from animals treated with cyclophosphamide (50 mg/kg iv, given 4 days before). The coronary vein draining the LAD region was cannulated for measurement of leukocyte count and TxB2 levels. Two groups of animals were studied: group 1 (n = 7) neutropenic animals were instrumented and normal animals served as blood donors and group 2 (n = 6) normal animals were instrumented and neutropenic animals served as blood donors. The myocardial response to intracoronary C5a (500 ng) was determined in each animal during coronary perfusion with normal arterial blood and also with neutropenic arterial blood. During perfusion with normal arterial blood, C5a decreased coronary flow to 52.3% and contractile function to 58.8% of preinfusion values. This was accompanied by a transient myocardial accumulation of PMNs (arterial-coronary venous gradient of 5.4 x 10(3) cells/microliters) and increased TxB2 levels in coronary venous blood (from 0.31 to 17.5 ng/ml).(ABSTRACT TRUNCATED AT 250 WORDS)

Interleukin-1 as a possible agent for treatment of infection.

Treatment of experimental animals with bacterial products, such as cell wall components of gram-negative bacteria, leads to enhanced resistance to a variety of microorganisms. Since interleukin-1 and other pro-inflammatory cytokines are induced by such bacterial products, it has been investigated whether these cytokines are also capable of enforcing host resistance. It has been possible to demonstrate that a low dose of interleukin-1 protects mice against death from either gram-negative or gram-positive bacteria, Candida albicans and Plasmodium berghei. The protection against lethal bacterial and fungal infection can be produced in both normal and neutropenic animals. Despite extensive investigations, the mechanism of protection is not understood. A possible mechanism, which is currently being investigated, is that interleukin-1 interferes with the deleterious action of the pro-inflammatory cytokines during the lethal phase of the infection.

Role of neutrophils in endotoxin-mediated microvascular injury in hamsters

The purpose of this study was to examine the role of circulating neutrophils in endotoxin-induced increase in microvascular permeability in vivo. Fifteen hamsters were anesthetized, and a plastic chamber was placed in each cheek pouch to observe the microvasculature. Fluorescein-labeled dextran (FITC-D, 150 kDa) was injected intravenously, and changes in leaky sites and FITC-D clearance were measured in three groups: control (saline, n = 4), endotoxin suffusion (n = 6), and endotoxin suffusion after neutropenia induction (n = 5). We found a significant increase in leaky sites and FITC-D clearance with endotoxin (45 +/- 18/cm2 and 20 +/- 6 x 10(-6) ml/min, respectively; mean +/- SD, P less than 0.05) in comparison to control (7 +/- 6/cm2 and 7 +/- 5 x 10(-6) ml/min) and endotoxin suffusion in neutropenic animals (19 +/- 11/cm2 and 12 +/- 4 x 10(-6) ml/min). There was a significant correlation between the number of leaky sites and FITC-D clearance (r = 0.91, P less than 0.01) and between the number of circulating neutrophils and FITC-D clearance (r = 0.87, P less than 0.01). We conclude that endotoxin-mediated increase in microvascular permeability in the peripheral circulation is dependent in part on circulating neutrophils.

Leucocyte depletion does not affect post-ischaemic nerve cell damage in the rat

Leucocytes play an important role in inflammation and immunologic responses. They might be of special significance under pathophysiological conditions of the brain i.e. ischaemia or stroke. It has been shown that neutropenic animals undergoing reversible ischaemia show higher post-ischaemic blood flow, suggesting improved post-ischaemic perfusion. In this study it was investigated therefore, whether polymorphonuclear leucocytes contribute to the nerve cell loss in the hippocampus after a reversible period of ischaemia. Rats were made neutropenic with a specific anti-serum against rat polymorphonuclear leucocytes yielding leucocyte counts less than 10% of normal. The animals were then subjected to 15 min reversible forebrain ischaemia. Quantitative histology was performed after a survival period of 7 days. Nerve cell counts in the frontal cortex and in the CA1 and CA3 sectors of the hippocampus did not reveal any differences between neutropenic rats and animals with normal leucocyte counts. From the results it might be concluded that neutrophils do not significantly contribute to the selective post-ischaemic nerve cell damage in the rat.

The role of neutrophils and platelets in a rabbit model of thromboembolic stroke.

Cerebral ischemia is accompanied by many of the cardinal features of acute inflammation such as neutrophil and platelet activation and accumulation. We sought to determine whether circulating neutrophils or platelets contribute to brain injury in a rabbit model of thromboembolic stroke that includes a fixed duration of superimposed systemic hypotension. We randomized 18 rabbits to receive either antineutrophil antiserum (n = 6), antiplatelet antiserum (n = 5), or nonimmune serum (n = 7). We assessed brain ischemia by measuring cerebral blood flow, intracranial pressure, and infarct size. Following the intracarotid administration of an autologous clot, cerebral blood flow in all groups fell to less than 5 ml/100 g/min during induced hypotension. After restoration of baseline blood pressure, mean cerebral blood flow in neutropenic animals recovered to 20-30 ml/100 g/min while that in control and thrombocytopenic rabbits remained at less than 10 ml/100 g/min. Intracranial pressure in control animals rose steadily to a final value of 241% of baseline, while a much smaller increase (148% of baseline) was noted in the thrombocytopenic group; no change from baseline was evident in the neutropenic group. Infarct size was significantly (p less than 0.05) reduced in the neutropenic group but not in the thrombocytopenic group. These results suggest that neutrophils may be important contributors to ischemia-induced brain injury whereas the role of platelets is more subtle.

Fibronectin fragments containing the RGDS cell-binding domain mediate monocyte migration into the rabbit lung. A potential mechanism for C5 fragment-induced monocyte lung accumulation.

Many inflammatory processes are characterized by an early phase of neutrophil migration and a later phase of monocyte migration into the inflammatory site. Mechanisms that govern the transition between phases are the subject of these investigations. Acute lung inflammation induced by C5 fragments in the rabbit leads to an initial neutrophil influx and plasma leakage into the alveolar space, followed by monocyte influx that we have previously shown to be dependent on prior emigration of neutrophils. Neutrophil enzymes are known to cleave intact fibronectin into fragments that are monocyte chemotaxins in vitro. Accordingly, generation of appropriate fibronectin fragments in situ by proteolytic enzymes from infiltrating neutrophils might represent a potential mechanism for attraction of monocytes into the lung. The studies reported herein demonstrate that a 120-kD fragment of fibronectin containing the RGDS fibroblast cell-binding domain induced monocyte migration into the rabbit lung in vivo. Intact fibronectin was inactive. A significant proportion of the monocyte migration was neutrophil independent. Intact fibronectin was present in bronchoalveolar lavage fluid from C5 fragment-treated animals rendered neutropenic, but absent in lavage from normal C5 fragment-treated animals. Fibronectin fragments were present in bronchoalveolar lavage fluid from both C5 fragment-treated and control rabbits. In addition, the amount of fibronectin was significantly increased in lavage of C5 fragment-treated normal but not neutropenic animals. Monoclonal antibodies directed against an epitope of fibronectin containing the RGDS cell-binding domain significantly inhibited the C5 fragment-induced monocyte migration, but not neutrophil migration. These studies suggest that chemotactic fibronectin fragments may in part be responsible for the recruitment of monocytes into areas of acute lung inflammation.

Efficacy of a Monoclonal Antibody Directed against Tumor Necrosis Factor in Protecting Neutropenic Rats from Lethal Infection with Pseudomonas aeruginosa

A monoclonal antibody directed against murine tumor necrosis factor-alpha (TNF) was studied in a neutropenic rat model to determine its efficacy in protecting animals from lethal infection with Pseudomonas aeruginosa. Anti-TNF monoclonal antibody at a dose of 20 mg/kg given intravenously at 0 and 120 h resulted in a 53% survival rate (8/15) compared with no survival in control animals (0/15) (P less than .005). The combination of anti-TNF monoclonal antibody and oral ciprofloxacin at a suboptimal dose of 2.5 mg/kg/day resulted in a 100% survival rate in neutropenic animals (16/16), while ciprofloxacin alone produced only a 67% survival rate (10/15) during the 7-day period of neutropenia (P less than .05). Thus anti-TNF monoclonal antibody alone or in addition to antimicrobial agents improved survival in neutropenic animals after infection with P. aeruginosa.

Leukotriene B4 potentiates colonic ulceration in the rat

The ability of various leukotrienes, platelet-activating factor and N-formyl-methyl-leucyl-phenylalanine to augment colonic damage induced by 30% ethanol was investigated in the rat. Each of the mediators was tested at a dose of 2 nmol, administered intracolonically in the ethanol vehicle. When colonic damage was assessed 72 hr later, only leukotriene B4 significantly augmented damage compared to the controls. The incidence of ulcers increased from 35% in the control group to 90% in the group receiving leukotriene B4. Leukotriene B4 administration also resulted in significant increases in colonic myeloperoxidase activity and colonic leukotriene B4 synthesis. To assess the possible contribution of infiltrating neutrophils to the increase in colonic leukotriene B4 synthesis that accompanies colonic inflammation, colitis was induced in normal and neutropenic rats by intracolonic administration of trinitrobenzene sulfonic acid. Neutropenia was achieved by treatment with an antineutrophil serum. In the neutropenic animals killed 4 hr after induction of colitis significant changes in leukotriene B4 synthesis were not observed, whereas a fourfold increase was observed in the controls. From these studies we conclude the following: (1) leukotriene B4, at a dose of 2 nmol, can significantly potentiate the colonic ulceration induced by 30% ethanol; (2) this action of leukotriene B4 is not shared by the same dose of the other inflammatory mediators tested; and (3) infiltrating neutrophils are the major source of colonic leukotriene B4 synthesis in a rat model of colitis.

Neutrophilic proteases: Mediators of formyl-methionyl-leucyl-phenylalanine-induced ileitis in rats

N-formyl-methionyl-leucyl-phenylalanine (FMLP), a tripeptide of bacterial origin that activates and attracts neutrophils, increases mucosal permeability when placed in the lumen of rat ileum. Although studies using neutropenic animals demonstrate the essential role of neutrophils in FMLP-induced mucosal injury, the neutrophil-derived chemical mediator of this injury process remains undefined. The objective of this study was to determine whether neutrophilic proteases mediate FMLP-induced increases in mucosal permeability. The blood-to-lumen clearance of 51Cr-ethylenediaminetetraacetate was used to monitor mucosal permeability in the terminal ileum of Sprague-Dawley rats. In control (untreated) animals luminal perfusion with 10−5 M FMLP resulted in twofold and fourfold increases in 51Cr-ethylenediaminetetraacetate clearance after 1 and 2 h of FMLP exposure, respectively. Pretreatment with the nonspecific serine protease inhibitor, soybean trypsin inhibitor (15 mg/kg), significantly attenuated the clearance responses normally observed during luminal perfusion with FMLP. The specific elastase inhibitors MeOSuc-Ala-Ala-Pro-Val-CH2Cl (10 mg/kg) and Eglin c (8 mg/kg) significantly attenuated the FMLP-induced increases in ethylenediaminetetraacetate clearance observed after both 1 and 2 h of exposure. The results of this study indicate that neutrophilic proteases mediate at least part of the increased mucosal permeability induced by luminal exposure to FMLP.

Trehalose dimycolate enhances resistance to infection in neutropenic animals.

Bacterial infections are lethal complications of neutropenia, and antibiotics alone are inadequate therapy for these infections. Irradiated mice become severely neutropenic and remain susceptible to infection for 2 to 3 weeks, depending on the dose and quality of radiation. Some bacterial cell wall derivatives stimulate nonspecific host defense mechanisms against a variety of microbes which might cause postirradiation infection. In this study we determined if the cell wall glycolipid trehalose dimycolate (TDM), derived from Mycobacterium phlei, or a synthetic preparation of TDM was able to (i) enhance survival in mice when given before or after lethal doses of 60Co radiation and (ii) increase nonspecific resistance to postirradiation infection. Treatment with TDM oil-in-water emulsions and with synthetic TDM significantly enhanced survival before and after lethal doses of 60Co irradiation. This result correlated with the ability of TDM to reduce the translocation of intestinal bacteria and to stimulate hematopoiesis. With respect to nonspecific resistance to infection, TDM injected 1 h after sublethal irradiation increased resistance to a lethal Klebsiella pneumoniae challenge (10 50% lethal doses of K. pneumoniae in 30 days [LD50/30]) 4 or 14 days later. Increasing the dose of K. pneumoniae to 5,000 LD50/30 on day 4 overwhelmed the ability of TDM-treated mice to overcome infection. However, TDM treatment 1 h postirradiation combined with ceftriaxone antibiotic therapy (days 5 through 14) enhanced survival, even when the higher dose of bacteria (5,000 LD50/30) was used. These results indicate that in irradiated mice, TDM can be used to enhance survival and, as a potent stimulant of nonspecific resistance to infection in neutropenic mice, can act synergistically with antibiotic therapy to reduce sepsis and mortality.

Multiple Organ Damage Caused by Tumor Necrosis Factor and Prevented by Prior Neutrophil Depletion

The effect of TNF on nonpulmonary multiple organ damage (MOD) was studied. Since polymorphonuclear leukocytes (PMN) are thought to play an important role in septic or TNF-induced MOD, we investigated both neutrophil sufficient (PMN+) and neutropenic (PMN-) guinea pigs. Sepsis was induced by Escherichia coli administration (2 x 10(9)/kg) or recombinant human TNF (1.4 x 10(6) U/kg) was infused into PMN+ and PMN- guinea pigs. During necropsy, the PMN+/TNF and PMN+/E coli animals exhibited marked damage in the adrenal glands, kidneys and liver as evidenced by hemorrhage, congestion, and PMN sequestration on histopathologic examination. There was also increased tissue albumin accumulation in the adrenal glands, kidneys, spleen, heart, and liver as demonstrated by 125I-labeled albumin determinations. In contrast, the PMN-/TNF group did not reveal histopathologic damage in any organ system and there was no abnormal organ accumulation of 125I-albumin. However, in PMN-/E coli animals, marked histopathologic damage in the adrenal glands and liver was evident. Furthermore, there were marked accumulations of 125I-albumin in the adrenals, heart, kidneys, liver, and spleen. Moreover, the PMN-/E coli guinea pigs had a much greater accumulation (p less than 0.01) of 125I-albumin in the kidneys than any other group including the PMN+/E coli group. Thus, nonpulmonary MOD in guinea pigs is caused by TNF administration and can be prevented by PMN depletion. However, while E coli administration also caused marked nonpulmonary MOD in neutrophil sufficient guinea pigs, equivalent or greater damage was produced in neutropenic animals. This suggests that while TNF-induced MOD may be primarily mediated by PMN, E coli-induced MOD seems to be mediated by more than PMN.

Isolation, structure, and immunogenicity of Pseudomonas aeruginosa immunotype 4 high-molecular-weight polysaccharide

A high-molecular-weight, immunogenic form of the lipopolysaccharide O side chain of Pseudomonas aeruginosa Fisher immunotype 4 (type 4, International Antigenic Typing System 1, Lanyi O:6) was isolated and characterized. Analysis by nuclear magnetic resonance spectroscopy confirmed the structural similarity of this high-molecular-weight polysaccharide and the type 4 O side chain. The polysaccharide was immunogenic in rabbits and mice, eliciting opsonophagocytic killing antibodies. Immunization with the polysaccharide produced significant protection against homologous challenge in both burned and granulocytopenic mice. Naturally acquired opsonic killing antibodies to type 4 polysaccharide were present in sera from unimmunized normal adults at levels comparable to postimmunization levels achieved after immunization with other type-specific polysaccharides. The specificity of the naturally occurring antibodies for the O side chain was documented by immunoblot analysis and inhibition studies. Naturally occurring polysaccharide-specific antibodies were comparable in their protective activity against live challenge in neutropenic animals to immunization-induced murine antibodies with similar specificity. These data suggest that naturally occurring serum antibody to P. aeruginosa type 4 lipopolysaccharide O side chains in most adults is not distinguishable in quantity or quality from immunization-induced antibodies in mice; evaluation of type 4-specific vaccines in humans may be complicated by this finding.

In vivo activation of neutrophil function in hamsters by recombinant human granulocyte colony-stimulating factor

The in vivo effect of Escherichia coli-derived recombinant human granulocyte colony-stimulating factor on neutrophil function was studied in golden Syrian hamsters. Significant increases in superoxide generation and specific binding of N-formylmethionyl-leucyl-phenylalanine were observed in neutrophils isolated 4 h following a single subcutaneous injection of the factor (30 micrograms/kg). However, phagocytotic activity was not significantly stimulated in hamsters treated with the factor. Recombinant human granulocyte colony-stimulating factor hastened the recovery of peripheral neutrophil counts in animals made leukopenic by prior treatment with cyclophosphamide. Beginning several hours after infection, resistance to lethal infection following intraperitoneal injection of Staphylococcus aureus was increased when neutropenic animals were treated daily with the factor. This protective effect was associated with increased peritoneal neutrophil counts and a decreased incidence of positive peritoneal bacterial cultures at 24 h after the start of treatment. These results suggest that recombinant human granulocyte colony-stimulating factor may be a useful adjunct in the treatment of bacterial infections in neutropenic patients.

Leucocyte infiltration and cartilage proteoglycan loss in immune arthritis in the rabbit

1. The relationship between phagocytic leucocyte infiltration and cartilage degradation in immune arthritis has been investigated in groups of normal and neutropenic rabbits. 2. Injection of antigen into the knee joints of sensitized control animals induced joint swelling, prostaglandin E2 (PGE2) synthesis, leucocyte accumulation and proteoglycan loss from articular cartilage. 3. Intravenous injection of nitrogen mustard caused a selective depletion of circulating neutrophils and monocytes with little or no effect on platelets or lymphocytes. In neutropenic animals challenged with antigen, there was virtually no joint swelling, PGE2 synthesis or leucocyte infiltration but cartilage proteoglycan loss was unchanged after 1 day and increased by day 4 compared to control animals. 4. The numbers of circulating leucocytes returned to normal 3-4 days after nitrogen mustard treatment and leucocyte infiltration occurred in antigen-challenged joints but this was not accompanied by joint swelling. Subsequent intra-articular injection of PGE2 did, however, cause swelling. 5. Lysosomal enzyme levels in arthritic joint fluids were measured. The levels of beta-glucuronidase, which is released by activated phagocytes, were decreased in neutropenic animals but the levels of N-acetyl-beta-glucosaminidase, which is a marker of tissue damage, were not changed by neutrophil depletion. 6. Intra-articular injections of the cytokine interleukin-1 (IL-1) induced a pattern of leucocyte infiltration and cartilage proteoglycan loss similar to that seen in immune arthritis. In neutropenic animals, IL-1 did not cause significant accumulation of leucocytes in the joint but the loss of proteoglycan from cartilage was unimpaired. 7. These results indicate that both leucocyte infiltration and prostaglandin synthesis are required for joint swelling but that tissue degradation is mediated by resident cells. It is likely that release of IL-1 by synovial cells stimulates the synthesis and activation of metalloproteinases which initiate the process of tissue degradation.