Introduction

Top Abstract Introduction Materials and Methods Results Discussion References

Neutrophils (polymorphonuclear leukocytes [PMNs]) are summoned into pulmonary air spaces by a variety of stimuli, including infection by pathogens, inhalation of air pollutants, and aspiration of gastric contents. Cytocidal and phagocytic activities of airway PMNs are essential for successful resolution of many bacterial and viral infections (1). In addition, PMNs play a critical role in the inflammatory responses to inhaled particles (4) and for the normal resolution of inflammation and tissue injury in response to irritants such as ozone (5).

Recently, dysfunctional PMN migratory responses in lungs have been demonstrated in endotoxemic rats. PMN migration in response to intratracheally instilled endotoxin (lipopolysaccharide [LPS]) isolated from Gram-negative bacteria is blocked when rats are made endotoxemic by intravenous administration of LPS (6). Furthermore, endotoxemic rats with bacterial pneumonia have higher mortality due to lack of pulmonary PMN migration and clearance of organisms (7, 8). Humans with clinical endotoxemia display altered PMN adhesion profiles (9). Accordingly, the defect in PMN responses during endotoxemia has been proposed as a mechanism for the development of nosocomial pneumonia in some at-risk human patients. It is possible that endotoxemia also affects PMN emigration to other airway stimuli and thereby compromises normal inflammatory processes necessary for the resolution of infection and of inhalation toxicoses. The ability of intravenous LPS to influence PMN emigration elicited by other airway stimuli has not been reported.

The mechanism of pulmonary PMN migration in rats given intratracheal LPS has been partially characterized and involves production by airway cells of inflammatory cytokines such as tumor necrosis factor (TNF) and interleukin (IL)-1 as well as neutrophil-specific chemoattractants known as CXC chemokines, which in rats include the family of cytokine-induced neutrophil chemoattractants (CINCs) and macrophage inflammatory protein-2 (MIP-2; CINC-3) (10). Expression of CD18 adhesion molecules on the PMN surface can be induced by exposure to LPS or TNF (13, 14) and to chemokines such as CINC-1 and MIP-2 (15, 16). Likewise, endothelial cell adhesion proteins can be mobilized by several inflammatory mediators including TNF and IL-1 (17, 18). These CD18 integrins play obligatory roles for full PMN migration responses to Gram-negative pulmonary stimuli in mice and rabbits (19, 20). In rats, treatment with antibodies to CD11a/CD18 or CD11b/CD18, as well as to their intercellular adhesion molecule (ICAM)-1 endothelial cell ligand, attenuated the migration of PMNs to intrapulmonary LPS or Gram-negative bacteria (21, 22). Furthermore, expression of airway and vascular ICAM-1 after intratracheal LPS administration to rats depends on the production of airway TNF and IL-1 (22). Accordingly, airway LPS leads to the production of soluble mediators that can activate specific adhesion molecules on PMNs and endothelial cells (ECs).

Studies using other airway stimuli revealed that PMN emigration into airways may either require CD18 or work by as-yet-undefined, CD18-independent mechanisms. For example, airway instillation of Gram-positive bacteria, the chemotactic peptide C5a, or hydrochloric acid (HCl) elicits pulmonary PMN migration that is largely independent of CD18 (23). Conversely, emigration in response to airway administration of Gram-negative bacteria, LPS, IL-1, or phorbol myristate acetate is due primarily to pathways involving CD18 (23). In a rabbit model of bacterial pneumonia, IL-8 and TNF production was greater in airways after treatment with a CD18-dependent stimulus than with a stimulus that elicits CD18-independent pathways (27). This result suggests that the profile of specific cytokines and chemokines that is elicited by an intrapulmonary stimulus might influence the adhesion molecule requirements for PMN migration.

In this study, we tested the hypothesis that the ability of circulating LPS to inhibit PMN migration into airways depends on the stimulus for migration. The effect of intravenous LPS on pulmonary PMN migration to stimuli that differed in their requirement for CD18 was evaluated. Finally, we determined whether the appearance in airways of MIP-2 and TNF, two mediators involved in PMN migration, was associated with the stimulus-specific inhibition of pulmonary PMN trafficking by systemic LPS exposure.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results Discussion References

Animals

Male Sprague-Dawley rats (CD-Crl:CD[SD]Br; Charles River Laboratories, Portage MI) weighing 225 to 275 g were used for all studies. Rats were maintained on a 12-h light/dark cycle under controlled temperature and humidity and breathed high-efficiency particulate filtered air until experimental protocols commenced. Food (Harlan Teklad 22/5 Rodent Diet 8640; Harlan, Madison, WI) and tap water were supplied ad libitum.

Intratracheal Instillations

Rats were anesthetized with ketamine (100 mg/kg intraperitoneally). An incision was made to expose the trachea, and inflammatory agents dissolved in saline were instilled into the trachea via a polyethylene catheter. Preparation of airway stimuli were as follows: LPS (Escherichia coli; serotype 0128:B15, activity: 25 × 10⁶ endotoxin units/mg; Sigma Chemical, St. Louis, MO), 500 µg dissolved in 0.5 ml sterile saline (0.9%); IL-1 (human, recombinant; Genzyme, Cambridge, MA), 5 ng dissolved in 0.5 ml saline; HCl, 0.1 N final concentration in 0.25 ml saline; and lipoteichoic acid (LTA) isolated from Staphylococcus aureus (Sigma), 250 µg dissolved in 0.5 ml saline. Blood was drawn from rats and used as a source of serum for zymosan activation and for control serum. To prepare zymosan-activated serum (ZAS), fresh serum was incubated with zymosan (5 mg/ml, Type A; Sigma) for 30 min at 37°C and spun in a centrifuge for 10 min at 250 × g. The supernatant fluid was collected and spun again to remove any residual zymosan. Control serum (CS) was prepared by incubating freshly isolated rat serum at 70°C for 30 min to inactivate complement. Rats were treated with 0.5 ml of 15% solutions of ZAS or CS in saline. A dose of LPS was used that produced maximal airway PMN response in preliminary experiments. Doses of IL-1 and LTA that produced responses similar to that of LPS were used, and doses of ZAS and HCl that produced maximal PMN emigration without causing lethality or overt hemorrhage were selected (data not shown).

Intravenous Administration of LPS

Immediately following intratracheal administrations, saline or LPS dissolved in saline (2 mg/ml) was injected into tail veins of rats (1 ml/kg body weight).

Collection of Bronchoalveolar Lavage Fluid and Lungs

Preliminary studies using intratracheal LPS showed that airway PMN accumulation was maximal by 3 h (our unpublished observations). All animals regardless of treatment were killed 3 h after intratracheal and intravenous treatments. Rats were anesthetized with sodium pentobarbital (50 mg/kg, intraperitoneally). The trachea was exposed and cannulated, a midline laparotomy was performed, and animals were killed by exsanguination. After the thoracic cavity was opened, the heart and lungs were carefully removed en bloc. The bronchus to the left lung was clamped, and 5 ml of sterile saline was instilled through the tracheal cannula and withdrawn to recover bronchoalveolar lavage fluid (BALF) from the right lung lobes. A second saline lavage was performed and combined with the first. After lavage, the left bronchial clamp was released, and the lungs were inflated with 10% buffered formalin at 30 cm water pressure for 2 h. Total leukocytes in the BALF were enumerated with a hemocytometer, and BALF PMNs were determined from the fraction of PMNs in a cytospin sample. Protein content in lavage fluid was determined using the bicinchoninic acid (BCA) assay (Pierce, Rockford, IL).

BALF TNF

TNF activity in BALF samples was determined with a cytotoxicity assay using WEHI 1640 fibrosarcoma cells (28) and compared with a standard curve using human recombinant TNF (R&D Systems, Minneapolis, MN) as described previously (29).

BALF MIP-2

BALF MIP-2 (CINC-3) concentration was determined using an ELISA kit, which used peroxidase and tetramethyl benzidine as a detection method (BioSource International, Camarillo, CA). Plates were read at 450 nm on a microplate reader (Biotek Instruments, Winooski, VT), and dilutions of BALF were compared with a standard curve made of rat MIP-2.

Statistical Analysis

Results are expressed as means ± SEM. Data for BALF PMNs and protein were analyzed using a completely randomized analysis of variance. Multiple comparisons were made by the least significant difference post hoc test. The Pearson product moment correlation was used to evaluate the relationship between BALF concentrations of PMNs, MIP-2, and TNF, and the ability of intravenous LPS to inhibit PMN accumulation in BALF. Criterion for significance was taken to be P < 0.05.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

LPS

Intratracheal LPS caused pronounced PMN accumulation in BALF compared with intratracheal treatment with saline (Figure 1). Increases in BALF PMN numbers were completely blocked when animals received concurrent administration of intravenous LPS.

View larger version (18K): [in this window] [in a new window]   Figure 1.   Inhibition by intravenous LPS of airway PMN accumulation elicited by airway LPS administration. Rats were given either sterile saline or 0.5 mg LPS intratracheally and immediately given saline or LPS (2 mg/kg) intravenously. Three hours later rats were killed, lungs were lavaged with saline, and PMNs were enumerated as described in MATERIALS AND METHODS. Results are expressed as mean ± SEM; n = 4. a Significantly different from respective group receiving saline intravenously. b Significantly different from respective group receiving saline intratracheally.

IL-1

The ability of intravenous LPS treatment to inhibit pulmonary PMN recruitment was tested in animals in which a PMN-eliciting inflammagen other than LPS was instilled into airways. PMNs are recruited into air spaces of rabbit lungs in response to intrapulmonary administration of IL-1 by mechanisms that require CD18 adhesion molecules (25). IL-1 (5 ng) caused accumulation of PMNs in BALF after intratracheal administration in rats (Figure 2). In rats treated intratracheally with IL-1, administration of LPS intravenously completely inhibited the increase in BALF PMNs at 3 h when compared with rats treated intravenously with saline.

View larger version (16K): [in this window] [in a new window]   Figure 2.   Inhibition by intravenous LPS of airway PMN accumulation elicited by airway IL-1 administration. Rats were given either sterile saline or IL-1 (5 ng) intratracheally and immediately given saline or LPS (2 mg/kg) intravenously. Three hours later rats were killed, lungs were lavaged with saline, and PMNs were enumerated as described in MATERIALS AND METHODS. Results are expressed as mean ± SEM; n = 4. a Significantly different from respective group receiving saline intravenously. b Significantly different from respective group receiving saline intratracheally.

LTA

Intralobar inoculation of rabbits with S. aureus or other Gram-positive organisms having membranous LTA induces airway PMN emigration, which is partially inhibited by antibodies to CD18 adhesion molecules (20, 24). Treatment of rats intratracheally with 250 µg LTA purified from S. aureus induced a significant increase in BALF PMN numbers (Figure 3). When rats were cotreated with intravenous LPS, PMN emigration was significantly reduced (58%) but not blocked completely.

View larger version (17K): [in this window] [in a new window]   Figure 3.   Effects of intravenous LPS on airway PMN accumulation elicited by airway LTA administration. Rats were given either saline or 250 µg LTA intratracheally and immediately given saline or LPS (2 mg/kg) intravenously. Three hours later rats were killed, lungs were lavaged with saline, and PMNs were enumerated as described in MATERIALS AND METHODS. Results are expressed as mean ± SEM; n = 4. a Significantly different from respective group receiving saline intravenously. b Significantly different from respective group receiving saline intratracheally.

HCl

PMNs emigrate into air spaces of rabbits in response to intralobar instillation of HCl by pathways that are independent of CD18 adhesion molecules (23, 26). Introduction into rat airways of 0.1 N HCl caused significant PMN accumulation in BALF, but to a lesser degree than that caused by intratracheal LPS or IL-1 (Figure 4). When animals were cotreated with intravenous LPS and HCl, the PMN accumulation in BALF was unaffected.

View larger version (17K): [in this window] [in a new window]   Figure 4.   Effects of intravenous LPS on airway PMN accumulation elicited by airway HCl administration. Rats were given either saline or 0.1 N HCl and immediately given saline or LPS (2 mg/kg) intravenously. Three hours later rats were killed, lungs were lavaged with saline, and PMNs were enumerated as described in MATERIALS AND METHODS. Results are expressed as mean ± SEM; n = 4. a Significantly different from respective group receiving saline intravenously. b Significantly different from respective group receiving saline intratracheally.

ZAS

Intrabronchial administration to rabbits of complement fragment C5a (human) induces airway recruitment of PMNs, which is mostly independent of CD18 (25). When zymosan-activated rat serum was used as a source of C5a, intratracheal administration caused significant elevation in BALF PMN numbers compared with that induced by complement-inactivated CS (Figure 5). Cotreatment of rats with intravenous LPS reduced the BALF PMN response to airway ZAS by 40%.

View larger version (16K): [in this window] [in a new window]   Figure 5.   Effects of intravenous LPS on airway PMN accumulation elicited by airway ZAS administration. Rats were given either CS or ZAS and immediately given saline or LPS (2 mg/kg) intravenously. Three hours later rats were killed, lungs were lavaged with saline, and PMNs were enumerated as described in MATERIALS AND METHODS. Results are expressed as mean ± SEM; n = 4. a Significantly different from respective group receiving saline intravenously. b Significantly different from respective group receiving saline intravenously.

Airway Protein Accumulation

Instillation into airways of LPS, IL-1, HCl, and LTA caused significant accumulation of protein in BALF compared with saline controls (Table 1). Protein concentrations in BALF after CS or ZAS were also significantly elevated compared with saline-treated rats. However, inasmuch as CS and ZAS preparations contained 15% serum, the increase in BALF protein can be explained by the serum protein that was introduced into the airways. Protein accumulation after intratracheal LPS or IL-1 was reduced to control levels by cotreating rats with intravenous LPS. Elevations in airway protein caused by LTA, HCl, CS, or ZAS were unaffected by intravenous LPS treatment.

Inflammatory Mediators Recovered in BALF

Certain inflammatory mediators may be associated with specific requirements for adhesion molecule expression during PMN emigration into airways. Accordingly, correlation analyses were performed to evaluate the relationship between PMN migratory behavior and MIP-2 or TNF appearance in BALF. Intrapulmonary administration of all stimuli induced production of MIP-2 in airways (Table 2), and concentrations of MIP-2 correlated with the numbers of PMNs recovered in BALF (r = 0.846; P < 0.05). TNF activity was evident in BALF from rats treated intratracheally with LPS or LTA, but it was negligible in rats receiving IL-1, HCl, or ZAS. LPS induced more than twice the TNF production of LTA, which corresponded to the greater PMN migratory response. TNF activity in BALF was significantly correlated with BALF PMN numbers (r = 0.911; P < 0.05).

Analyses were also performed to determine whether inhibition of migration by intravenous LPS was selective for intrapulmonary stimuli that preferentially result in the appearance of TNF or MIP-2. Correlation analysis revealed a significant association between MIP-2 appearance and degree of inhibition by intravenous LPS (r = 0.659; P < 0.05). No association was evident for TNF (r = 0.413; P > 0.05); however, omission of data from IL-1- treated rats resulted in a significant association between BALF TNF and inhibition of PMN migration by intravenous LPS (r = 0.752; P < 0.05).

The concentration of BALF MIP-2 was also measured in some rats that received intravenous LPS and an intratracheal stimulus (Figure 6). In rats given intratracheal HCl or ZAS, BALF MIP-2 concentration was significantly increased when animals were also treated with intravenous LPS. In rats given intratracheal LTA, treatment with intravenous LPS did not affect MIP-2 concentrations in BALF.

View larger version (22K): [in this window] [in a new window]   Figure 6.   Effect of intravenous LPS on airway production of MIP-2 induced by intrapulmonary HCl, ZAS, and LTA. Either 0.1 N HCl, ZAS, or LTA was instilled intratracheally into rats, and then saline or LPS (2 mg/kg) was immediately injected intravenously. Three hours later, animals were killed, lungs were lavaged with saline, and MIP-2 concentrations in BALF were determined as described in MATERIALS AND METHODS. Results are expressed as mean ± SEM; n = 4. a Significantly different from respective group receiving saline intravenously.

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

The results of this study demonstrate that the degree to which endotoxemia inhibits pulmonary PMN recruitment depends on the intrapulmonary stimulus. Migration of PMNs in response to airway LPS or IL-1 was blocked completely by intravenous LPS, emigration after LTA or ZAS was partially inhibited, and the response to HCl was unaffected. Furthermore, the selectivity of inhibition appears to be related to the putative involvement of CD18 adhesion molecules in the migratory response; specifically, intravenous LPS treatment was more effective at blunting PMN migration, which has been shown in other animal models to require CD18.

The most extensive characterizations of CD18 involvement in the migration of pulmonary PMNs have been in rabbit models (20, 23). PMN migration in response to airway E. coli (20), endotoxin isolated from E. coli (23, 24), or IL-1 (25) was inhibited in rabbits by antibodies directed against CD18. In rats, evidence suggests that PMN migration to intrapulmonary LPS relies on CD18 adhesion pathways (21, 22). By contrast, after HCl aspiration, edema and vascular sequestration of PMNs occur independently of CD18 (30). In addition, intrapulmonary IL-1 mediates PMN recruitment during immune complex injury in rats (31), a model in which PMN migration requires CD11a/CD18 and CD11b/CD18 pathways (32). Furthermore, studies in mice using the same intrapulmonary stimuli used in rabbits suggest that pulmonary PMN migratory responses might be the same across species (19). Our results in rats confirm that intravenous LPS inhibits accumulation of PMNs in airways in response to LPS or IL-1, two stimuli that initiate CD18-dependent migration.

Tuomanen and coworkers (33) instilled various cell wall components of Gram-positive bacteria, including teichoic acid peptidoglycans, into rabbit airways to create leukocytic infiltration. We induced pulmonary PMN emigration in rats by administration of S. aureus-derived LTA and found migration to be inhibited by 60% by intravenous LPS treatment (Figure 3). In comparison, antibodies to CD18 reduced PMN emigration in response to S. pneumonia- or S. aureus-induced pneumonia in rabbits by 45% (20, 24). Thus, both CD18 antibody treatment and intravenous LPS inhibit, by similar degrees, the PMN migration to Gram-positive stimuli in rabbit and rat airways, respectively.

In rats instilled with ZAS, two components of airway PMN emigration were observed, one that was reduced by intravenous LPS treatment (40% inhibition) and one that was unaffected (Figure 5). The chemotactic potential of ZAS is primarily due to complement activation products, especially C5a. In rabbits, an antibody to CD18 reduced only 20 to 30% of the PMN emigration into airways in response to human recombinant C5a, but this effect was not statistically significant (25). Our observation that intravenous LPS failed to inhibit most of the migratory response to ZAS is consistent with the possibility that intravenous LPS inhibits CD18-dependent PMN trafficking.

Instillation of HCl into rabbit lungs causes PMN emigration, which is unaffected by treatment with antibodies to CD18 (23, 26). Likewise, intravenous LPS had no effect on PMN emigration in response to HCl in rats (Figure 4). Generally, the degree to which intravenous LPS treatment inhibited PMN trafficking into airways correlated with the degree to which the various intratracheal stimuli caused CD18-dependent migration.

Burns and Doerschuk (34) have shown that CD18 expression on PMNs in the pulmonary circulation is unchanged prior to migration to a CD18-dependent stimulus in airways. Curiously, CD18 is upregulated on PMNs before they adhere and migrate toward an airway stimulus that does not require CD18. LPS exposure to isolated PMNs causes shedding of L-selectin and increased expression of CD18 in vitro (14), and similar changes are observed on PMNs isolated from endotoxemic animals (6). Thus, LPS-induced CD18 upregulation on circulating PMNs might be a premature step for competent CD18- dependent migration, but it is a normal PMN response in the process of CD18-independent pathways. An untimely expression of CD18 on circulating PMNs may violate the stepwise sequence of rolling, firm adhesion, and diapedesis, and may result in aborted or dysregulated interactions between PMNs and endothelial cells. This possibility is consistent with the ability of LPS to inhibit transendothelial migration, which requires CD18 in vitro (35).

PMN migration into rat airways is often accompanied by an increase in vascular permeability (30, 36, 37). We measured protein accumulation in BALF as a marker of vascular leakage. In rats cotreated with intravenous LPS and intrapulmonary LPS, IL-1, or HCl, decreases in BALF PMN accumulation were associated with reduced BALF protein concentration (Figures 1, 2, and 4, and Table 1). That is, protein accumulation was blocked completely after intrapulmonary IL-1 and LPS but was unaffected after HCl. When PMN emigration was partially inhibited by intravenous LPS treatment in rats given LTA or ZAS, elevations in BALF protein accumulation were unaffected (Figures 3 and 5, and Table 1). Pulmonary PMNs in the microvasculature and alveoli have been implicated as edemagenic mediators in rat models of adult respiratory distress syndrome (38, 39). However, the inhibition by intravenous LPS of BALF protein accumulation after intratracheal LPS or IL-1 was not due to an inhibition of pulmonary PMN localization. Light microscopic evaluation of lung sections revealed that treatment with intravenous LPS caused sequestration of PMNs within alveolar walls irrespective of intratracheal administration (data not shown). Thus, PMN extravasation may be linked to protein leakage in air spaces, but the mere presence of PMNs in the pulmonary microvasculature is not sufficient to cause leakage.

At least one study suggests that whether or not CD18 is involved in pulmonary PMN emigration depends on the type of cytokine mediators produced by the stimulus for migration (27). In this paradigm, inflammatory mediators that activate CD18 on PMNs and/or ICAM-1 on endothelial cells may preferentially induce CD18-dependent migration. Expression of CD18 on PMNs can be induced by either TNF or MIP-2, and ICAM-1 is upregulated on endothelial cell monolayers after exposure to TNF (13, 15, 18). Accordingly, we measured TNF and MIP-2 concentrations in lavage fluid of rats that were given the various intrapulmonary stimuli used in our model (Table 2). All intratracheal treatments resulted in the appearance of airway MIP-2 , and this correlated with the accumulation of PMNs in BALF. Despite the lack of TNF in BALF after IL-1, HCl, or ZAS treatments, the appearance of TNF in airways was also associated with BALF PMN accumulation. TNF is a strong promoter of CD18 and ICAM-1 expression and may preferentially mediate CD18-dependent migration into air spaces. When all groups were considered, the appearance of TNF in BALF correlated poorly (r = 0.413; P < 0.05) with the ability of intravenous LPS to inhibit pulmonary PMN emigration, suggesting at first glance that inhibition is not related to the requirement for CD18. However, IL-1 by itself is capable of promoting expression of both ICAM-1 on cultured ECs and CD18 on isolated PMNs. In rats treated with intrapulmonary IL-1, the presence of TNF therefore may not be necessary to induce CD18 migratory pathways. Indeed, elimination of the results with IL-1 from the analysis resulted in a much stronger correlation between inhibition of PMN migration and BALF TNF (r = 0.752; P < 0.05).

Concentrations of MIP-2 in BALF also correlated with the ability of intravenous LPS to inhibit PMN emigration. MIP-2 is a potent PMN chemoattractant that can promote the surface expression of CD18 on PMNs and may preferentially promote CD18-dependent migration. That intravenous LPS can inhibit MIP-2-dependent migration is supported by the results in HCl- and ZAS-treated rats. Coadministration of intravenous LPS caused an increase in the appearance of MIP-2 in the airways of rats given HCl or ZAS (Figure 6), yet this increase did not result in further PMN accumulation in BALF. Indeed, PMN migration was reduced in ZAS-treated rats (Figure 5) and failed to increase in rats treated with HCl (Figure 4). Thus, the increase in PMNs expected with an increase in MIP-2 in airways was blocked by intravenous LPS treatment. A recent report suggests that LPS can directly downregulate expression of chemokine receptors on human PMNs (40). If MIP-2 receptors on rat PMNs are similarly affected, circulating endotoxin could inhibit PMN migratory responses to intrapulmonary stimuli that rely on MIP-2 pathways.

Production of specific chemokines has not been systematically correlated with CD18-dependent or -independent airway stimuli. Our results indicate that distinct profiles of cytokine production occur in response to different intrapulmonary stimuli, and these mediators might dictate specific adhesion molecule requirements for PMN migration. Intravenous LPS in rats strongly inhibited the intra-alveolar accumulation of PMNs in response to agents that require CD18 for PMN migration and had no or more limited effects on PMN accumulation with materials that are not CD18 dependent. Selective inhibition by intravenous LPS may be due to premature expression of CD18 on circulating PMNs, an event that could be detrimental to CD18-dependent migration. Furthermore, LPS itself can bind specifically to CD18 integrins on PMNs, where it may block ligand binding and activate intracellular pathways of activation (41). Thus, immune suppression and PMN dysfunction that occur during endotoxemia in humans might be due to the inability of PMNs to respond to inflammatory stimuli that require competent CD18 function. Explaining the mechanism of endotoxemia-related PMN dysfunction may suggest therapeutic options for complications such as nosocomial infections and PMN-mediated organ injuries.