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Published ahead of print on March 5, 2003, doi:10.1164/rccm.200207-664OC
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American Journal of Respiratory and Critical Care Medicine Vol 167. pp. 1567-1574, (2003)
© 2003 American Thoracic Society

Original Article

Early Alterations in Neutrophil Activation Are Associated with Outcome in Acute Lung Injury

Kuang-Yao Yang, John J. Arcaroli and Edward Abraham

Division of Pulmonary Sciences and Critical Care Medicine, University of Colorado Health Science Center, Denver, Colorado; Chest Department, Taipei Veterans General Hospital, School of Medicine, National Yang-Ming University, Taipei, Taiwan

Correspondence and requests for reprints should be addressed to Dr. Edward Abraham M.D., Division of Pulmonary Sciences and Critical Care Medicine, University of Colorado Health Sciences Center, Mail Code C-272, 4200 East Ninth Avenue, Denver, CO 80262. E-mail: Edward.Abraham{at}UCHSC.edu


   ABSTRACT
TOP
 ABSTRACT
METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 
Persistent elevations of proinflammatory cytokines in the lungs are associated with increased mortality from acute lung injury (ALI), suggesting that the degree of pulmonary inflammation is an important determinant of clinical course in ALI. The transcriptional regulatory factor nuclear factor-{kappa}B (NF-{kappa}B) is involved in modulating the expression of many cytokines and other proinflammatory mediators implicated in the development and progression of ALI. Because neutrophils appear to play a major role in the development of ALI, we examined the relationships between clinical outcome and activation of NF-{kappa}B in peripheral neutrophils from patients (n = 30) with sepsis-induced ALI. We found that nuclear translocation of NF-{kappa}B in this setting was dependent on the activation of p38 and Akt kinases. Diminished activation of NF-{kappa}B or Akt, but not p38, in the early postintubation period was associated with less time on the ventilator and improved survival in critically ill patients with ALI. These results suggest that early alterations in neutrophil activation patterns, particularly involving the ability to accumulate NF-{kappa}B to the nucleus after relevant stimuli, contribute to subsequent clinical course in ALI.

Key Words: neutrophils • transcription factors • signal transduction • cellular activation • lung

Acute lung injury (ALI) is presently defined by clinical criteria, including bilateral pulmonary infiltrates on chest radiographs and hypoxemia (1). Histologic examination of the lungs from patients with ALI demonstrates loss of epithelial integrity, diffuse alveolar damage, increased interstitial edema, as well as the frequent presence of large numbers of activated neutrophils (2, 3). Neutrophils appear to play a major role in the development of ALI. In experimental models, the severity of ALI is diminished when neutrophils are eliminated before initiation of pathophysiologic insults that lead to lung damage such as endotoxemia or hemorrhage (4, 5). Although ALI can develop in patients with neutropenia, worsening of pulmonary function and the occurrence of ALI often occur as neutrophil counts are restored (6, 7).

Increased levels of proinflammatory cytokines, such as interleukin (IL)-8 and tumor necrosis factor-{alpha}, are present in the lungs of patients with ALI, and elevated pulmonary concentrations of IL-8 are found in at-risk patients who subsequently progress to ALI (8, 9). Persistent elevation of proinflammatory cytokines in the lungs correlates with poor outcome in ALI (10). Many of the proinflammatory mediators that appear to be involved in the pathogenesis of ALI, including IL-8 and tumor necrosis factor-{alpha}, are under the regulatory control of the transcriptional factor nuclear factor-{kappa}B (NF-{kappa}B) (11, 12). NF-{kappa}B is normally sequestered in the cytoplasm through association with the inhibitory molecule I{kappa}B-{alpha} (inhibitor of {kappa}B-{alpha}), but after cellular stimulation, I{kappa}B-{alpha} is phosphorylated, ubiquinated, and then degraded, allowing the heterodimeric NF-{kappa}B complex to move to the nucleus, where it can bind to specific promoter sequences, associate with coactivators, and initiate transcription (11, 12).

Increased nuclear translocation of NF-{kappa}B has been demonstrated to be present in the lungs of patients with ALI (13). In experimental models, inhibition of NF-{kappa}B activation diminishes the severity of lung injury and inflammation (14, 15). In clinical studies of patients with sepsis, persistent elevations in the nuclear levels of NF-{kappa}B among circulating mononuclear cell populations is associated with worse outcome, including mortality (16, 17). However, no clinical studies have examined the association that alterations of nuclear NF-{kappa}B levels in the early postintubation period may have with outcome from ALI.

Although multiple intracellular signaling cascades have been implicated in phosphorylation of I{kappa}B-{alpha} and nuclear translocation of NF-{kappa}B, pathways involving the p38 mitogen-activated protein (MAP) or Akt kinases appear to be particularly important in neutrophils. In LPS-stimulated neutrophils, specific inhibition of p38 prevented nuclear accumulation of NF-{kappa}B (18). In recent in vitro and in vivo experiments, blockade of phosphoinositide 3-kinase, the kinase immediately upstream to Akt, diminished translocation of NF-{kappa}B to the nucleus of LPS-treated neutrophils and also diminished the severity of endotoxemia-induced ALI (15).

In the present study, we examined the relationships between activation of NF-{kappa}B, p38, and Akt in peripheral neutrophils and clinical outcome in patients with ALI. We found that nuclear translocation of NF-{kappa}B in this setting was modulated by both p38 and Akt and that diminished nuclear translocation of NF-{kappa}B and phosphorylation of Akt, but not p38, in the early postintubation period were associated with less time on the ventilator and improved survival in such critically ill patients. These results suggest that early alterations in neutrophil activation patterns, particularly involving the ability to accumulate NF-{kappa}B to the nucleus after relevant stimuli, contribute to subsequent clinical course in ALI.


   METHODS
TOP
 ABSTRACT
 METHODS
RESULTS
 DISCUSSION
 REFERENCES
 
Reagents
RPMI 1640 was obtained from Life Technologies (Gaithersburg, MD). Fetal bovine serum and penicillin/streptomycin were purchased from Gemini Bioproducts (Calabasas, CA). LPS (from Escherichia coli O111:B4) was obtained from Sigma Chemical (St. Louis, MO). Poly(dI-dC)·poly(dI-dC) and Percoll were purchased from Amersham-Pharmacia (Piscataway, NJ). The Coomassie-Plus Protein Assay Reagent and bicinchoninic acid (BCA) Protein Assay Reagent were purchased from Pierce (Rockford, IL). SB203580 and LY294002 were purchased from Calbiochem (La Jolla, CA). Antibodies to serine 473 phosphorylated Akt and total Akt were purchased from Upstate Biotech (Lake Placid, NY). Antibodies to threonine 180/tyrosine 182 phosphorylated p38 and total p38 were purchased from Cell Signaling (Beverly, MA).

Patients
Entry criteria for sepsis-induced ALI were essentially as used in previous studies (19). In particular, patients were eligible for inclusion if they were in an intensive care unit, had clinical evidence of infection, required positive pressure ventilation via endotracheal or tracheostomy tube, had acute onset of significantly impaired oxygenation with a PaO2/FIO2 ratio less than 300 (adjusted for barometric pressure [i.e., < 250] in Denver), bilateral infiltrates consistent with pulmonary edema on a frontal chest radiograph, and no clinical evidence of left atrial hypertension or if a pulmonary artery catheter was in place of a pulmonary artery occlusion pressure less than 18 mm Hg. Patients had to be enrolled within 24 hours of developing these criteria. The study was approved by the institutional review board. Consent was obtained from all patients or their surrogates before enrollment.

Exclusion criteria were age below 18 years, neurologic conditions that could impair weaning, severe chronic respiratory disease, severe chronic liver disease (defined as a Child–Pugh score of > 10), burns of 30% or more of the total body surface area, malignancy or other irreversible condition for which 6-month mortality was estimated to be above 50%, use of chronic immunosuppressive therapy (i.e., corticosteroids at a dose of >= 1 mg/kg/day of methylprednisolone or equivalent within the 3 days before intensive care unit admission or cytotoxic agents for immunologic or oncologic disorders), or a history of bone marrow or lung transplantation. Patients were also excluded if the clinicians caring for the patient were not committed to providing aggressive life support at the time of enrollment.

Volunteers
Peripheral blood neutrophils were collected from 10 healthy volunteers, aged 26 to 50 years, for comparison of nuclear translocation of NF-{kappa}B or activation of p38 or Akt with that found in neutrophils isolated from patients with sepsis-induced ALI. The protocol for peripheral blood draw from volunteers was approved by the institutional review board and consent was obtained from all subjects.

Isolation and Culture of Human Neutrophils
Peripheral blood was obtained from patients and volunteers, and neutrophils (purity > 98%) were isolated by plasma-percoll gradients after dextran sedimentation of erythrocytes (18, 20). Neutrophils were resuspended in RPMI 1640, 5% fetal calf serum at a final concentration of 5 x 106 cells/ml and cultured with or without 100 ng/ml LPS for 1 hour. The p38 MAP kinase inhibitor SB203580 (30 µM) or the PI3-K inhibitor LY294002 (100 µM) were added to the neutrophil cultures for 1 hour before LPS stimulation. These doses of SB203580 and LY294002 have previously been shown to block p38 and PI3-K activity in human neutrophils (18, 21).

Electrophoretic Mobility Shift Assay
Nuclear extracts were prepared as previously described (13, 15, 22). Isolated neutrophils were incubated for 15 minutes in buffer A (10 mM N-2-hydroxyethylpiperazine-N'-ethane sulfonic acid [pH 7.9], 1.5 mM magnesium chloride, 10 mM potassium chloride, pH 7.9). After cytoplasm was removed from the nuclei by 15 passages through a 25-gauge needle, the nuclei were collected by centrifugation at 600 x g for 6 minutes at 4°C. The nuclear pellet was incubated on ice for 15 minutes in buffer C (20 mM N-2-hydroxyethylpiperazine-N'-ethane sulfonic acid [pH 7.9], 0.42 M sodium chloride, 1.5 mM magnesium chloride, 0.2 mM ethylenediaminetetraacetic acid, 25% glycerol), after which the extract was centrifuged at 4°C for 10 minutes at 12,000 x g. The supernatant was collected, divided into aliquots, and stored at -86°C. Protein concentration was determined by using the Coomassie-Plus Protein Assay Reagent (Pierce, Rockford, IL) standardized to bovine serum albumin (BSA), according to the manufacturer's protocol.

Nuclear extracts (5 µg) were incubated at room temperature for 15 minutes in 20 µl of reaction buffer containing 10 mM tris(hydroxymethyl) aminoethane–hydrochloric acid (pH 7.5), 1 mM magnesium chloride, 0.5 mM ethylenediaminetetraacetic acid, 0.5 mM dithiothreitol, 50 mM sodium chloride, and 4% glycerol, with [32P]end-labeled, double-stranded oligonucleotide probe specific for the {kappa}B site, 5'-GCCATGGGGGGATCCCCG AAG TCC-3' (Geneka Biotechnology Inc., Montreal, Quebec, Canada) and 1 µg of poly(dI-dC)·poly(dI-dC). The complexes were resolved on 5% polyacrylamide gels in tris(hydroxymethyl) aminoethane–hydrochloric acid (pH 8.0)–borate–ethylenediaminetetraacetic acid buffer at 10 V/cm. Dried gels were exposed with Kodak Biomax MS film (Rochester, NY) for 1 to 24 hours at -70°C. Samples from control, LPS-stimulated, and, where appropriate, SB203580- or LY294002-treated neutrophils were all run on the same gel. Densitometry was performed using an imaging system and analysis software (BioRad, Hercules, CA).

Western Blot Analysis
Whole cell extracts from human neutrophils were collected from cells denatured in ice-cold lysis buffer (50 mM N-2-hydroxyethylpiperazine-N'-ethane sulfonic acid, 150 mM sodium chloride, 10% glycerol, 1% Triton X-100, 1.5 mM magnesium chloride, 1 mM ethyleneglycol-bis-(ß-aminoethyl ether)-N,N'-tetraacetic acid, 1 mM sodium vanadate, 10 mM sodium pyrophosphate, 10 mM sodium flouride, 300 µM p-nitrophenyl phosphate, 1 mM phenylmethylsulfonyl fluoride, 10 µg/ml leupeptin, 10 µg/ml aprotinin, pH 7.3) for 15 minutes. The protein concentration of each sample was assayed using the BCA protein assay kit standardized to BSA, according to manufacturer's protocol.

For Western blots, 70 µg of protein from whole cell extracts were loaded on a 10% tris(hydroxymethyl) aminoethane–hydrochloric acid sodium dodecyl sulfate polyacrylamide gel. Protein was electrotransferred to a nitrocellulose membrane and then blocked with 5% nonfat dry milk, 20 mM tris(hydroxymethyl) aminoethane–buffered saline, with 0.1% Tween. After blocking, the membrane was incubated overnight at 4°C with antibodies to phos-p38, total p38, phos-Akt, or total Akt, using a dilution of 1:1,000 followed by horseradish peroxidase–coupled secondary antibody at a dilution of 1:2,000. After washing five times, bands were detected using chemiluminescence Western blotting detection reagents (ECL Detection System, Amersham-Pharmacia). Densitometry was performed using chemiluminescence system and analysis software (BioRad), and the normalized ratio of phosphorylated to total kinase was determined.

Statistical Analysis
The primary clinical variables collected were survival to Day 28 after study entry and ventilator-free days (VFD) in survivors. VFD were calculated as previously described (19) and were counted as the number of days of unassisted breathing within the 28 days postenrollment period in survivors. Nonsurvivors were considered to have no VFD. Unassisted breathing was defined as either extubation with or without supplemental oxygen, T-tube breathing, tracheostomy mask breathing, or continuous positive airway pressure of less than or equal to 5 cm H2O without pressure support or intermittent mandatory ventilation assistance. Linear regression analysis was performed using the least squares method. Tukey–Kramer multiple group comparison test (for multiple groups) or Student's t test (for two groups) were used for comparisons between groups. Data are presented as mean ± SEM. p Values of less than 0.05 was considered significant.


   RESULTS
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 ABSTRACT
 METHODS
 RESULTS
DISCUSSION
 REFERENCES
 
Patient Characteristics
Demographic characteristics of the patients studied are shown in Table 1 . There were 24 survivors and 6 nonsurvivors, with all deaths occurring within 28 days of intubation and inclusion in this study. ALI continued to be present at the time of death in the nonsurvivors. In all patients, the major acute clinical problem associated with the development of ALI was infection, with 16 patients having a pulmonary source and 14 having an extrapulmonary focus.


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  		TABLE 1. Patient characteristics

 
NF-{kappa}B, Akt, and p38 Activation
There was no statistically significant relationship between clinical outcome, defined as VFD, and nuclear levels of NF-{kappa}B, or phosphorylation of Akt or p38 in unstimulated peripheral blood neutrophils from patients with ALI (Figure 1) . In contrast, in neutrophils cultured with LPS, there were statistically significant correlations between VFD and relative increase in nuclear translocation of NF-{kappa}B or phosphorylation of Akt, but not in phosphorylation of p38, induced by LPS stimulation (Figure 1).



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  		Figure 1. Relationship between nuclear levels of nuclear factor-{kappa}B (NF-{kappa}B) or phosphorylation of Akt or p38 and clinical course, as defined by ventilator-free days (VFD), in unstimulated neutrophils and neutrophils cultured with LPS for 1 hour (i.e., NF-{kappa}B LPS, Akt LPS, or p39 LPS) in patients with acute lung injury (ALI). Peripheral blood neutrophils were collected within 24 hours of intubation. Correlation coefficients and p values are shown.

 
To explore relationships between patterns of neutrophil activation and clinical outcome in ALI, the patient population was divided into two groups: those who died or remained on the ventilator for more than 14 days (i.e., <= 14 VFD) and those who survived and required fewer than 14 days on the ventilator (i.e., > 14 VFD). No differences in PaO2/FIO2 ratios were present at the time neutrophils were collected (i.e., within 24 hours of intubation) in the patients whose subsequent clinical course was associated with prolonged ventilation or death (i.e., <= 14 VFD as compared with those patients who survived and required a shorter period of ventilatory support (i.e., > 14 VFD) (Figure 2) . Age, male/female ratio, Acute Physiology and Chronic Health Evaluation II score, and pulmonary/extrapulmonary source of infection were not significantly different between the two groups. There were no significant differences in the nuclear translocation of NF-{kappa}B or phosphorylation of Akt or p38 in unstimulated neutrophils from patients with subsequent clinical courses characterized by less than or equal to 14 or more than 14 VFD (Figures 3–5) . With the exception of nuclear levels of NF-{kappa}B in ALI patients with 14 VFD values for nuclear levels of NF-{kappa}B or phosphorylated levels of Akt and p38 were significantly greater in patients with ALI than in normal volunteers.



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  		Figure 2. Baseline PaO2/FIO2 ratios are not significantly different in patients with ALI who survived and required less than 14 days of ventilator support, i.e., more than 14 VFD (n = 18), compared with those with a more severe subsequent clinical course, as determined by death or prolonged ventilation, i.e., less than or equal to 14 VFD (n = 12). PaO2/FIO2 values were determined at the same time that neutrophils were collected from patients and were within 24 hours of intubation.

 


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  		Figure 3. NF-{kappa}B levels were significantly increased in nuclear extracts from neutrophils of patients with ALI compared with that found in volunteers. In LPS-stimulated neutrophils of patients with ALI whose clinical course was characterized by prolonged ventilator support or death (<= 14 VFD), nuclear levels of NF-{kappa}B were significantly increased as compared with volunteers or patients with more rapid recovery (> 14 VFD). There were no significant differences in nuclear NF-{kappa}B accumulation between the two patient groups in unstimulated neutrophils. Representative electrophoretic mobility shift assay gels are presented. Densitometry data are shown for unstimulated neutrophils as "NF-{kappa}B Baseline," whereas the relative percentage increase in nuclear NF-{kappa}B concentrations in neutrophils cultured with LPS for 1 hour as compared with those found in unstimulated neutrophils is presented as "NF-{kappa}B LPS." *p values less than 0.05 versus values for volunteers and {dagger}p values less than 0.05 versus patients with ALI with more than 14 VFD.

 


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  		Figure 5. Levels of the active, phosphorylated form of p38 were significantly increased in unstimulated neutrophils of patients with ALI compared with that found in volunteers. However, levels of phosphorylated p38 were no different in LPS-stimulated neutrophils of patients with ALI as compared with volunteers. Representative gels showing levels of phosphorylated and total Akt from unstimulated and LPS-stimulated neutrophils are presented. Densitometric data comparing the levels of phosphorylated (p-p38) to total (t-p38) p38 are shown for unstimulated neutrophils as "p38 Baseline." The relative percentage increase of phosphorylated/total p38 levels in neutrophils cultured with LPS for 1 hour as compared with those found in unstimulated neutrophils is presented as "p38 LPS." *p values less than 0.05 versus values for volunteers.

 


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  		Figure 4. Levels of the active, phosphorylated form of Akt were significantly increased in neutrophils from patients with ALI compared with volunteers. In LPS-stimulated neutrophils of patients with ALI whose clinical course was characterized by prolonged ventilator support or death (<= 14 VFD), amounts of phosphorylated Akt are significantly increased compared with those from patients with more rapid recovery (> 14 VFD) or from volunteers. Representative gels showing levels of phosphorylated (p-Akt) and total (t-Akt) Akt from unstimulated and LPS-stimulated neutrophils are presented. Densitometric data comparing the levels of phosphorylated to total Akt are shown for unstimulated neutrophils as "Akt Baseline." The relative percentage increase of phosphorylated/total Akt levels in neutrophils cultured with LPS for 1 hour as compared with those found in unstimulated neutrophils is presented as "Akt LPS." *p values less than 0.05 versus values for volunteers and {dagger}p values less than 0.05 versus patients with ALI with more than 14 VFD.

 
When nuclear translocation of NF-{kappa}B or phosphorylation of Akt or p38 were examined in neutrophils after culture with LPS, patients who had prolonged periods on the ventilator or died demonstrated significantly higher nuclear translocation of NF-{kappa}B as well as activation of Akt, but not p38, compared with that found in volunteers or in patients who survived and were more rapidly extubated (Figures 3–5). There were no significant differences in LPS-induced activation of NF-{kappa}B, Akt, or p38 in neutrophils from patients who survived and required less than 14 VFD compared with that found in volunteers.

Association between Nuclear Translocation of NF-{kappa}B and Clinical Outcome from ALI
NF-{kappa}B is involved in the transcriptional regulation of proinflammatory and immunomodulatory genes whose products are believed to contribute to the development and progression of ALI (11–15). Because of this, relative activation of NF-{kappa}B may, in itself, play a pathophysiologic role in modulating inflammatory response in the lungs and determining outcome from ALI. If nuclear translocation of NF-{kappa}B does contribute to lung injury and outcome in the setting of infection, then decreased activation of NF-{kappa}B at early time points would be expected to be associated with a better clinical outcome in such critically ill patients.

As shown in Figure 1, although neutrophils from most patients with ALI demonstrated increased nuclear translocation of NF-{kappa}B when stimulated with LPS, there were some patients in whom no apparent increase in nuclear levels of NF-{kappa}B occurred after culture of their neutrophils with LPS. If NF-{kappa}B activation in neutrophils plays a role in progression of ALI, then patients with such nonresponsive neutrophils would be expected to have a more benign subsequent clinical course from ALI. To examine this issue, the study patients were divided into two groups. Nonresponders were defined as patients in whom stimulation of peripheral blood neutrophils with LPS produced no increase in NF-{kappa}B activation, and responders were considered to be patients in whom neutrophils demonstrated any enhancement of NF-{kappa}B nuclear translocation when incubated with LPS.

Among the patients included in these studies, there were nine in whom stimulation of peripheral blood neutrophils with LPS produced no increased nuclear translocation of NF-{kappa}B (nonresponders). Of the patients whose neutrophils showed no increased nuclear levels of NF-{kappa}B after culture with LPS, eight survived and required less than 14 days on the ventilator (i.e., > 14 VFD) and one had a more severe clinical course, being extubated 27 days after intubation (i.e., 1 VDF). The initial PaO2/FIO2 ratio in these patients was no different from that present in patients whose neutrophils demonstrated increased activation of NF-{kappa}B in response to LPS (responders, n = 21) (Figure 6) . However, when the clinical course of patients with nonresponding neutrophils was compared with that of patients whose neutrophils demonstrated increased activation of NF-{kappa}B after culture with LPS, nonresponders spent significantly less time on the ventilator than did patients with LPS-responsive neutrophils (Figure 7) .



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  		Figure 6. PaO2/FIO2 ratios are not significantly different in patients with ALI with neutrophils that increased nuclear translocation of NF-{kappa}B after stimulation with LPS (responder) compared with those that did not (nonresponder). PaO2/FIO2 values were determined at the same time that neutrophils were collected from patients, and were within 24 hours of intubation.

 


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  		Figure 7. Clinical course, as defined by VFD or survival, was significantly worse in patients whose neutrophils showed increased nuclear translocation of NF-{kappa}B after LPS stimulation (responder) compared with those with neutrophils in which there was no alteration in nuclear NF-{kappa}B concentrations after LPS stimulation (nonresponder). **p values less than 0.01 versus values in nonresponders.

 
In neutrophils from patients that did not increase nuclear translocation of NF-{kappa}B in response to LPS, there was significantly decreased phosphorylation of both Akt and p38 compared with levels in neutrophils from patients with ALI where enhanced nuclear localization of NF-{kappa}B was present after culture with LPS (Figure 8) . Such results suggested that Akt and p38 are involved in activating NF-{kappa}B in neutrophils from patients with ALI, and that diminished activation of p38 and Akt in nonresponder neutrophils may contribute to the decreased nuclear translocation of NF-{kappa}B observed in this situation.



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  		Figure 8. Levels of the active, phosphorylated forms of Akt or p38 were significantly increased after LPS stimulation in neutrophils that also increased nuclear accumulation of NF-{kappa}B in response to LPS (responder) as compared with levels of phosphorylated Akt or p38 in neutrophils that did not demonstrate any change in NF-{kappa}B activation after culture with LPS (nonresponder). Densitometric data are presented that show the percent increase in the ratio of phosphorylated to total Akt or p38 after LPS stimulation compared with that present in unstimulated neutrophils. *p values less than 0.05 versus values present in the nonresponder neutrophils.

 
Role of Akt or p38 in Determining NF-{kappa}B Activation in Patients with ALI
Akt and p38 have been described to participate in NF-{kappa}B activation in neutrophils (15, 18). To investigate directly the role that Akt and p38 play in neutrophil activation in the setting of ALI, we added specific inhibitors of activation of Akt (LY294002) or p38 (SB203580) to neutrophil cultures and then examined nuclear translocation of NF-{kappa}B in response to LPS. Six consecutive patients with ALI were studied. The subsequent clinical course of all of these patients was characterized as less than or equal to 14 VFD, with five of the patients requiring prolonged time on the ventilator and one patient subsequently died with multiple organ failure and continued ALI.

Inhibition of p38 or Akt significantly reduced LPS-induced NF-{kappa}B activation (Figure 9) . Although blockade of Akt appeared to reduce NF-{kappa}B nuclear translocation to a greater extent than did inhibition of p38, the differences in effect associated with each of these kinases were not statistically significant.



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  		Figure 9. Inhibition of p38 or Akt reduced LPS-induced nuclear translocation of NF-{kappa}B in neutrophils from patients with ALI (n = 6). Neutrophils were cultured with or without the p38 inhibitor SB203580 or the Akt inhibitor LY294002 for 1 hour and then the relative increase in nuclear accumulation of NF-{kappa}B induced by LPS stimulation for 1 hour was compared with that present in unstimulated neutrophils. *p values less than 0.05 versus values for neutrophils exposed to LPS but not SB203580 or LY294002.

 

   DISCUSSION
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
REFERENCES
 
In the present studies, neutrophil activation patterns in the early postintubation period, particularly those involving the kinase Akt and the transcriptional factor NF-{kappa}B, were found to be associated with subsequent clinical course as determined by time on the ventilator or survival. At the time that the differences in neutrophil responses were determined, pulmonary function as assessed by the PaO2/FIO2 ratio did not differ between patients whose clinical course was characterized by survival with relatively short periods on the ventilator versus those who died or survived but spent prolonged periods on mechanical ventilation. Such results not only indicate that neutrophil phenotypes correlate with outcome in ALI but also suggest that neutrophil activation patterns may contribute to lung injury and clinical course in this setting.

In experimental models, neutrophils appear to contribute to the severity of ALI. Induction of neutropenia before endotoxin administration decreases severity of lung injury (4, 5). In addition, therapies that decrease activation of neutrophils diminish inflammatory lung injury, even if neutrophils continue to accumulate in the lungs (23). Such findings indicate that the ability of neutrophils to take on an activated, proinflammatory phenotype is important in determining the development and severity of ALI. Results from the present study are consistent with such a hypothesis because the clinical course was less severe in patients whose neutrophils did not increase nuclear levels of NF-{kappa}B in response to stimulation with LPS.

Many of the proinflammatory mediators believed to play an important role in the development of ALI are under the regulatory control of NF-{kappa}B (11–15). In particular, transcription of IL-8 and tumor necrosis factor-{alpha} is largely regulated by NF-{kappa}B (24, 25). IL-1ß has been reported to be the major proinflammatory stimulus present in the lungs of patients with ALI (9), and NF-{kappa}B binding sites are present in the promoter of human IL-1ß (26). In previous studies, we found that neutrophils are the major source of IL-1ß in the lungs after endotoxemia (27). Therefore, inability of neutrophils to activate NF-{kappa}B in response to LPS would be expected to diminish pulmonary concentrations of IL-1ß and other neutrophil-derived cytokines, including IL-8 and tumor necrosis factor-{alpha}, thereby providing a potential mechanism for the decreased severity of clinical course in patients with such a neutrophil phenotype.

Alterations in neutrophil response to LPS were found very early in the clinical course of the patients examined in this study, being present in the immediate postintubation period. In particular, the presence of neutrophils that did not increase nuclear translocation of NF-{kappa}B after culture with LPS was associated with the requirement for significantly less ventilator support. Similar findings, using peripheral blood mononuclear cells collected during the first day of intensive care unit admission in patients with sepsis, suggested that decreased LPS-induced nuclear accumulation of NF-{kappa}B was associated with survival, although those results did not achieve statistical significance (28). A possible explanation for the present results is that early acquisition of endotoxin tolerance by neutrophils correlates with improved survival. However, previous investigations of endotoxin tolerance in patients with sepsis found that it was not associated with improved survival and, in fact, may correlate with worse clinical outcome (29, 30). An alternate explanation for the presence of early alterations in neutrophil response to LPS is that the differences in neutrophil phenotype may not have been acquired but rather preceded the development of ALI and that patients whose neutrophils become less activated in response to bacterial products, such as LPS, have a less severe clinical course when infected. Recent information has shown that high and low responder phenotypes exist in human peripheral blood mononuclear cells exposed to LPS and that such phenotypes are stable over prolonged periods of time in healthy volunteers (31). In preliminary studies, we have found similar stable phenotypes among neutrophils.

In the present studies, nuclear translocation of NF-{kappa}B in LPS-stimulated neutrophils from patients with ALI was dependent on p38 and Akt. Previous reports have demonstrated that both of these kinases are involved in modulating NF-{kappa}B activation in neutrophils (15, 18). In particular, inhibition of p38{alpha} MAP kinase activation with SB203580, the same molecule used in our studies, prevented nuclear accumulation of NF-{kappa}B in LPS-stimulated neutrophils (18). Recent results from our laboratory demonstrated that blockade of Akt activation with LY294002, the same phosphoinositide 3-kinase inhibitor used in the present experiments, decreased LPS-induced nuclear translocation of NF-{kappa}B in neutrophils (15). Although there are several proposed mechanisms by which Akt enhances NF-{kappa}B activity, a consistently demonstrated interaction between Akt and enhanced nuclear translocation of NF-{kappa}B centers on the ability of Akt to activate the regulatory I{kappa}B kinase (inhibitor of NF-{kappa} B kinase {alpha}), through phosphorylation of an Akt phosphorylation consensus sequence at Thr23. Akt-dependent activation of IKK{alpha} leads to accelerated degradation of I{kappa}B{alpha}, enhanced translocation of NF-{kappa}B to the nucleus, and increased NF-{kappa}B–dependent transcriptional activity (32–34).

Given the importance of Akt in modulating NF-{kappa}B activation in neutrophils from patients with ALI, it is not surprising that the clinical course was less severe in those situations where decreased activation of this kinase was present. In contrast, although increased p38 activation was present in unstimulated neutrophils from patients with ALI compared with that present among neutrophils from healthy volunteers, no relationship was found in the present studies between LPS-induced activation of p38 in peripheral blood neutrophils and subsequent clinical course. In experimental models of ALI, p38 has been shown to participate in LPS-induced neutrophil migration into the alveolar space (35, 36). However, despite the role that p38 plays in neutrophil chemotaxis, other studies have not demonstrated that p38 inhibition diminishes the severity of endotoxemia-induced ALI (37). Such results are consistent with the present findings and suggest that p38 may be less important in determining activation patterns among peripheral neutrophils than are other pathways leading to NF-{kappa}B activation.

In experimental models, the severity of LPS-induced ALI is reduced when Akt is inhibited (15). Such findings, coupled with those of the present study, indicate that activation of Akt may contribute to the severity of ALI through enhancing nuclear accumulation of NF-{kappa}B in neutrophils, thereby leading to increased transcription of NF-{kappa}B–dependent proinflammatory genes. In addition, because NF-{kappa}B exerts potent antiapoptotic effects through increasing transcription of antiapoptotic proteins, such as A1 and Bcl-xL, enhanced activation of NF-{kappa}B may worsen the severity of ALI by prolonging the presence of activated neutrophils in the lungs (38–40).

In the present study, decreased activation of Akt and NF-{kappa}B in LPS-stimulated neutrophils was associated with a more benign clinical course in patients with ALI. However, at the time that neutrophils were collected and such alterations in response detected, there were no apparent clinical differences between patients with low or high responder neutrophils. These results would suggest that neutrophil phenotypes may not affect the initial development of ALI but rather are involved in determining subsequent course, particularly after activation in vivo through exposure to bacterial products, such as LPS, or other stimuli. Prospective experiments in infected patients at risk for ALI will be necessary to address this issue.

The findings of the present study suggest that neutrophil response profiles may contribute to clinical course and outcome in ALI. In particular, patients whose neutrophils did not increase nuclear translocation of NF-{kappa}B after exposure to LPS spent significantly less time on the ventilator than did those patients with neutrophils showing NF-{kappa}B activation. Because the activation of NF-{kappa}B was dependent on Akt and p38, both of these kinases may represent therapeutic targets for ameliorating the course of ALI. However, such interventions may be of limited use in patients whose neutrophils already show minimal activation of p38 or Akt after stimulation. These findings suggest that there is heterogeneity among signaling mechanisms contributing to ALI and that therapies for this condition may need to be individualized on the basis of the specific pathways activated in each patient. Early determination of alterations in neutrophil activation patterns may be useful not only because of association with subsequent clinical course but also in designing appropriate therapeutic regimens for patients with ALI.


   FOOTNOTES
 
Supported in part by National Institutes of Health grant HL62221 and 1 PO1 HL068743.

Received in original form July 6, 2002; accepted in final form February 26, 2003


   REFERENCES
TOP
 ABSTRACT
 METHODS
 RESULTS
 DISCUSSION
 REFERENCES
 

  1. Bernard GR, Artigas A, Brigham KL, Carlet J, Falke K, Hudson L, Lamy M, Legall JR, Morris A, Spragg R. The American-European Consensus Conference on ARDS: definitions, mechanisms, relevant outcomes, and clinical trial coordination. Am J Respir Crit Care Med 1994;149:818–824.
  2. Pittet JF, Mackersie RC, Martin TR, Matthay MA. Biological markers of acute lung injury: prognostic and pathogenetic significance. Am J Respir Crit Care Med 1997;155:1187–1205.[Medline]
  3. Ware LB, Matthay MA. Medical progress: the acute respiratory distress syndrome. N Engl J Med 2000;342:1334–1339.[Free Full Text]
  4. Abraham E, Carmody A, Shenkar R, Arcaroli J. Neutrophils as early immunologic effectors in hemorrhage or endotoxemia induced acute lung injury. Am J Physiol 2000;279:L1137–L1146.
  5. Chignard M, Balloy V. Neutrophil recruitment and increased permeability during acute lung injury induced by lipopolysaccharide. Am J Physiol 2000;279:L1083–L1090.
  6. Azoulay E, Darmon M, Delclaux C, Fieux F, Bornstain C, Moreau D, Attalah H, Le Gall JR, Schlemmer B. Deterioration of precious acute lung injury during neutropenia recovery. Crit Care Med 2002;30:781–786.[CrossRef][Medline]
  7. Rinaldo JE, Borovetz H. Deterioration of oxygenation and abnormal lung microvascular permeability during resolution of leukopenia in patients with diffuse lung injury. Am Rev Respir Dis 1985;131:579–583.[Medline]
  8. Donnelly SC, Strieter RM, Kunkel SL, Walz A, Robertson CR, Carter DC, Grant IS, Pollok AJ, Haslett C. Interleukin-8 and development of adult respiratory distress syndrome in at-risk patient groups. Lancet 1993;341:643–647.[CrossRef][Medline]
  9. Pugin J, Ricou B, Steinberg KP, Suter PM, Martin TR. Proinflammatory activity in bronchoalveolar lavage fluids from patients with ARDS, a prominent role for interleukin-1. Am J Respir Crit Care Med 1996;153:1850–1856.[Abstract]
  10. Meduri GU, Kohler G, Headley S, Tolley E, Stentz F, Postlethwaite A. Inflammatory cytokines in the BAL of patients with ARDS: persistent elevation over time predicts poor outcome. Chest 1995;108:1303–1314.[Abstract/Free Full Text]
  11. Ghosh S, Karin M. Missing pieces in the NF-{kappa}B puzzle. Cell 2002;109:S81–S96.
  12. Ghosh S, May MJ, Kopp EB. NF-{kappa}B and Rel proteins: evolutionarily conserved mediators of immune responses. Annu Rev Immunol 1998;16:225–260.[CrossRef][Medline]
  13. Moine P, McIntyre R, Schwartz MD, Kaneko D, Shenkar R, Le Tulzo Y, Moore EE, Abraham E. NF-{kappa}B regulatory mechanisms in alveolar macrophages from patients with acute respiratory distress syndrome. Shock 2000;13:85–91.[Medline]
  14. Blackwell TS, Blackwell TR, Holden EP, Christman BW, Christman JW. In vivo antioxidant treatment suppresses nuclear factor-{kappa}B activation and neutrophilic lung inflammation. J Immunol 1996;157:1630–1637.[Abstract]
  15. Yum HK, Arcaroli J, Kupfner J, Shenkar R, Penninger JM, Sasaki T, Yang KY, Park JS, Abraham E. Involvement of PI3-K in neutrophil activation and the development of acute lung injury. J Immunol 2001;167:6601–6608.[Abstract/Free Full Text]
  16. Arnalich F, Garcia-Palomero E, Lopez J, Jimenez M, Madero R, Renart J, Vazquez JJ, Montiel C. Predictive value of nuclear factor {kappa}B activity and plasma cytokine levels in patients with sepsis. Infect Immun 2000;68:1942–1945.[Abstract/Free Full Text]
  17. Bohrer H, Qiu F, Zimmerman T, Zhang Y, Jilmer T, Manner D, Bottinger BW, Stern DM, Waldherr R, Saeger HD, et al. Role of NF-{kappa}B in the mortality of sepsis. J Clin Invest 1997;100:972–985.[Medline]
  18. Nick JA, Avdi NJ, Young SK, Lehman LA, McDonald PP, Frasch SC, Billstrom MA, Henson PM, Johnson GL, Worthen GS. Selective activation and functional significance of p38{alpha} mitogen-activated protein kinase in lipopolysaccharide-stimulated neutrophils. J Clin Invest 1999;103:851–858.[Medline]
  19. The ARDS Network. A trial of traditional tidal volume versus lower tidal volume ventilation in acute lung injury and acute respiratory distress syndrome. N Engl J Med 2000;342:1301–1308.[Abstract/Free Full Text]
  20. Haslett C, Guthrie LA, Kopaniak MM, Johnston RB, Henson PM. Modulation of multiple neutrophil functions by preparative methods or trace concentrations of bacterial lipopolysaccharide. Am J Pathol 1985;119:101–110.[Abstract]
  21. Nijhuis E, Lammers JJ, Koenderman L, Coffer PJ. Src kinases regulate PKB activation and modulate cytokine and chemoattractant-controlled neutrophil functioning. J Leukoc Biol 2002;71:115–124.[Abstract/Free Full Text]
  22. Abraham E, Arcaroli J, Shenkar R. Activation of ERK1/ERK2, NF-{kappa}B, and CREB in lung neutrophils occurs by differing mechanisms after hemorrhage or endotoxemia. J Immunol 2001;166:522–530.[Abstract/Free Full Text]
  23. Abraham E, Bursten S, Shenkar R, Allbee J, Tuder R, Woodson P, Guidot DM, Rice G, Singer JW, Repine JE. Phosphatidic acid signaling mediates lung cytokine expression and lung inflammatory injury following hemorrhage. J Exp Med 1995;181:569–575.[Abstract/Free Full Text]
  24. Udalova IA, Knight JC, Vidal V, Nedospasov SA, Kwiatkowski D. Complex NF-{kappa}B interactions at the distal tumor necrosis factor promoter region in human monocytes. J Biol Chem 1998;273:21178–21186.[Abstract/Free Full Text]
  25. Kunsch C, Rosen CA. NF-{kappa}B subunit-specific regulation of the interleukin-8 promoter. Mol Cell Biol 1993;13:6137–6146.[Abstract/Free Full Text]
  26. Hiscott J, Marois J, Garoufalis J, D'Addario M, Roulston A, Kwan I, Pepin N, Lacoste J, Nguyen H, Bensi G, et al. Characterization of a functional NF-{kappa}B site in the human interleukin 1ß promoter: evidence for a positive autoregulatory loop. Mol Cell Biol 1993;13:6231–6240.[Abstract/Free Full Text]
  27. Parsey MV, Tuder R, Abraham E. Neutrophils are major contributors to intraparenchymal lung IL-1ß expression after hemorrhage and endotoxemia. J Immunol 1998;160:1007–1013.[Abstract/Free Full Text]
  28. Adib-Conquy M, Adrie C, Moine P, Asehnoune K, Fitting C, Pinsky MR, Dhainaut JF, Cavaillon JM. NF-{kappa}B expression in mononuclear cells of patients with sepsis resembles that observed in lipopolysaccharide tolerance. Am J Respir Crit Care Med 2000;162:1877–1883.[Abstract/Free Full Text]
  29. McCall CE, Grosso-Wilmoth LM, LaRue K, Guzman RN, Cousart SL. Tolerance to endotoxin-induced expression of the interleukin-1ß gene in blood neutrophils of humans with the sepsis syndrome. J Clin Invest 1993;91:853–861.
  30. West MA, Heagy W. Endotoxin tolerance: a review. Crit Care Med 2002;30:S64–S73.[CrossRef][Medline]
  31. Park WY, Frevert CW, Wong VA, Martin TR. High and low inflammatory response phenotypes in 101 normal human subjects. Chest 2002;121:87S–88S.[Free Full Text]
  32. Ozes ON, Mayo LD, Gustin JA, Pfeffer SR, Pfeffer LM, Donner DB. NF-{kappa}B activation by tumor necrosis factor requires the Akt serine-threonine kinase. Nature 1999;401:82–85.[CrossRef][Medline]
  33. Romashkova JA, Makarov SS. NF-{kappa}B is a target of Akt in anti-apoptotic PDGF signalling. Nature 1999;401:86–90.[CrossRef][Medline]
  34. Sizemore N, Lerner N, Dombrowski N, Sakurai H, Stark GR. Distinct roles of the I{kappa}B kinase {alpha} and ß subunits in liberating nuclear factor {kappa}B (NF-{kappa}B) from I{kappa}B and in phosphorylating the p65 subunit of NF-{kappa}B. J Biol Chem 2002;277:3863–3869.[Abstract/Free Full Text]
  35. Nick JA, Young SK, Brown KK, Avdi NJ, Arndt PG, Suratt BT, Janes MS, Henson PM, Worthen GS. Role of p38 mitogen-activated protein kinase in a murine model of pulmonary inflammation. J Immunol 2000;164:2151–2159.[Abstract/Free Full Text]
  36. Nick JA, Young SK, Arndt PG, Lieber JG, Suratt BT, Poch KR, Avdi NJ, Malcolm KC, Taube C, Henson PM, et al. Selective suppression of neutrophil accumulation in ongoing pulmonary inflammation by systemic inhibition of p38 mitogen-activated protein kinase. J Immunol 2002;169:5260–5269.[Abstract/Free Full Text]
  37. Arcaroli J, Yum H-K, Park JS, Yang K-Y, Abraham E. Role of p38 MAP kinase in the development of acute lung injury. Clin Immunol 2001;101:211–219.[CrossRef][Medline]
  38. Kupfner JG, Arcaroli J, Yum H-K, Nadler SG, Abraham E. Role of NF-{kappa}B in endotoxemia induced alterations of lung neutrophil apoptosis. J Immunol 2001;167:7044–7051.[Abstract/Free Full Text]
  39. Lee HH, Dadgostar H, Cheng Q, Shu J, Cheng G. NF-{kappa}B-mediated up-regulation of Bcl-x and Bfl-1/A1 is required for CD40 survival signaling in B lymphocytes. Proc Natl Acad Sci USA 1999;96:9136–9141.[Abstract/Free Full Text]
  40. Wang CY, Guttridge DC, Mayo MW, Baldwin AS. NF-{kappa}B induces expression of the Bcl-2 homologue A1/Bfl-1 to preferentially suppress chemotherapy-induced apoptosis. Mol Cell Biol 1999;19:5923.[Abstract/Free Full Text]



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