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The Journal of Immunology, 2004, 172: 5727-5733.
Copyright © 2004 by The American Association of Immunologists

Phosphoinositide 3-Kinase and Akt Occupy Central Roles in Inflammatory Responses of Toll-Like Receptor 2-Stimulated Neutrophils1

Derek Strassheim2, Karim Asehnoune, Jong-Sung Park, Jae-Yeol Kim, Qianbin He, Donald Richter, Katherine Kuhn, Sanchayita Mitra and Edward Abraham

Division of Pulmonary Science and Critical Care Medicine, Department of Medicine, University of Colorado Health Sciences Center, Denver, CO 80262


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Materials and Methods
 Results
 Discussion
 References
 
Neutrophils are critical initiators and effectors of the innate immune system and express Toll-like receptor 2 (TLR2) and TLR4. Although signaling through pathways involving phosphoinositide 3-kinase (PI3-K) and the downstream kinase Akt (protein kinase B) plays a central role in modulating neutrophil chemotaxis and superoxide generation in response to engagement of G protein-coupled receptors, the importance of these kinases in affecting inflammatory responses of neutrophils stimulated through TLR2 has not been examined. In these experiments, we found activation of Akt in neutrophils stimulated with the TLR2-specific ligands peptidoglycan and the lipopeptide tri-palmitoyl-S-glyceryl-Cys-Ser-(Lys)4 that occurred earlier and was of greater magnitude than that present after exposure to the TLR4 agonist LPS. The release of the proinflammatory mediators TNF-{alpha} and macrophage inflammatory protein-2 was inhibited in a dose-dependent manner by PI3-K blockade. The IC50 for inhibition of peptidoglycan-stimulated Akt activation and macrophage inflammatory protein-2 release correlated closely, indicating linkage of these two events. PI3-K blockade did not inhibit nuclear translocation of NF-{kappa}B, but did prevent Ser536 phosphorylation of the p65 subunit of NF-{kappa}B, an event required for maximal transcriptional activity of NF-{kappa}B. Inhibition of PI3-K also prevented activation of p38 mitogen-activated protein kinase and extracellular receptor-activated kinase 1/2 in TLR2-stimulated neutrophils. These results demonstrate that the PI3-K-Akt axis occupies a central role in TLR2-induced activation of neutrophils.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Materials and Methods
 Results
 Discussion
 References
 
Phosphoinositide 3-kinases (PI3-K)3 and the downstream serine/threonine kinase Akt/protein kinase B are involved in modulating neutrophil activation, chemotaxis, and apoptosis (1, 2, 3, 4, 5). PI3-K catalyzes the addition of a phosphate molecule to the inositol ring of phosphoinositides generating phosphoinositol-3,4,5-P3, which, after binding to the pleckstrin homology domain of Akt, permits association of the phosphatidylinositol-dependent kinases PDK1 and PDK2 and activation of Akt through phosphorylation of Thr308 and Ser473 (1).

Neutrophils play a central role in acute inflammatory responses, including those associated with Gram-positive and Gram-negative bacterial infections (6). Exposure of neutrophils to LPS engages the membrane-based Toll-like receptor 4 (TLR4), resulting in activation of the transcriptional factor NF-{kappa}B and production of proinflammatory cytokines, such as macrophage inflammatory protein-2 (MIP-2) and TNF-{alpha}, that are under the regulatory control of NF-{kappa}B (7, 8). Activation of NF-{kappa}B is a multistep process, involving a number of protein kinases, including the I{kappa}B kinase complex, that not only facilitates movement of NF-{kappa}B heterodimers from the cytoplasm to the nucleus, but can also phosphorylate the NF-{kappa}B p65 subunit (6, 8, 9, 10). Although the involvement of Akt in TLR2 signaling has not been examined in neutrophils, Akt has been shown to participate in NF-{kappa}B activation produced by stimulation of neutrophils with LPS (11) and in other cells cultured with LPS (12, 13). Additionally, Akt-dependent activation of NF-{kappa}B has been described in a variety of cell types stimulated with platelet-derived growth factor (14), TNF-{alpha} (15, 16, 17, 18), IL-1 (19, 20), as well as other agonists (21, 22, 23). In several studies Akt appeared to enhance nuclear translocation of NF-{kappa}B through phosphorylation and activation of I{kappa}B kinase (24, 25, 26), resulting in enhanced degradation of I{kappa}B-{alpha},. Other experiments have found that Akt did not affect nuclear translocation of NF-{kappa}B, but, rather, influenced NF-{kappa}B-dependent transcription through a mechanism dependent on phosphorylation of the p65 NF-{kappa}B subunit (15, 27). However, the involvement of Akt in NF-{kappa}B activation has not consistently been found, as it has been demonstrated that platelet-derived growth factor (28), TNF-{alpha} (29, 30), IL-1 (29, 30, 31), and LPS (24, 32, 33) can induce NF-{kappa}B and cytokine responses independently of Akt. Additionaly, P13-K-Akt axis has been reported to have negative feedback functions in some inflammatory circuits (34, 35, 36).

Ten TLRs have been characterized, and neutrophils express all of them, with the exception of TLR3 (37). TLR2, a receptor for Gram-positive bacterial-derived peptidoglycan and lipoproteins, is expressed in higher concentrations on neutrophils than is TLR4 (38). TLR2-induced neutrophil responses are likely to have important clinical consequences, as Gram-positive organisms are now known to be an increasingly important cause of severe infection associated with organ dysfunction, including septic shock (39).

Despite the clinical importance of Gram-positive infections, only limited attention has been paid to the role of PI3-K or Akt in cellular activation induced by TLRs other than TLR4. In HEK 293 cells transfected with an exogenous TLR2 receptor, Akt has been linked to TLR2-dependent NF-{kappa}B activation induced by cellular exposure to Staphylococcus aureus (40). However, no studies have specifically explored the question of how interactions between TLR2, PI3-K, and Akt might affect neutrophil cytokine/chemokine release.

In the present experiments we examined the involvement of the PI3-K/Akt pathway in modulating TLR2-induced neutrophil activation. These studies show that Akt has a central role in TLR2-associated potentiation of neutrophil responses, specifically in the production of proinflammatory cytokines and chemokines. Although previous work, including our own with LPS-stimulated neutrophils (11), had suggested that the mechanism by which Akt could induce such effects involved regulation of nuclear translocation of NF-{kappa}B, this did not appear to be the case in neutrophils activated by engagement of TLR2. Rather, the present experiments indicate that Akt modulates NF-{kappa}B-dependent transcription in TLR2-stimulated neutrophils through modifying phosphorylation of the p65 subunit. Such results, showing that Akt affects NF-{kappa}B activity through different mechanisms in neutrophils activated by TLR2 compared with TLR4, provide insight into the distinct cellular responses that have been observed in neutrophils and other cell populations after engagement of TLR2 vs TLR4.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Materials and Methods
Results
 Discussion
 References
 
Mice

Male BALB/c mice, 8–12 wk of age, were purchased from Harlan Sprague Dawley (Indianapolis, IN). The mice were kept on a 12-h light, 12-h dark cycle with free access to food and water. All experiments were conducted in accordance with institutional review board-approved protocols.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Materials and Methods
Results
 Discussion
 References
 
Isoflurane was obtained from Abbott Laboratories (Chicago, IL). Escherichia coli 0111:B4 endotoxin (LPS) and C5a were purchased from Sigma-Aldrich (St. Louis, MO). Peptidoglycan (PGN) and tri-palmitoyl-S-glyceryl-Cys-Ser-(Lys)4 (PAM) were purchased from Invitrogen (San Diego, CA). RPMI 1640/25 mM HEPES/L-glutamine was obtained from BioWhittaker (Walkersville, MD), and FBS and penicillin/streptomycin were purchased from Gemini Bioproducts (Calabasas, CA). Bicinchoninic acid protein assay reagent was purchased from Pierce (Rockford, IL). Activation-specific Abs for phospho-Thr202/Tyr204 extracellular receptor-activated kinase 1 (ERK1), phospho-Thr183/Tyr185 ERK2, phospho-Thr180/Tyr182 p38 mitogen-activated protein kinase (p38), phospho-Ser473 Akt, phospho-Ser536 NF-{kappa}B, and total ERK1/2, p38, Akt, p65 NF-{kappa}B were purchased from Cell Signaling Technologies (Beverly, MA). HRP-labeled anti-rabbit Abs and chemiluminescence reagents were purchased from Bio-Rad (Hercules, CA). All other reagents were purchased from Sigma-Aldrich unless otherwise noted in the text. Custom mixture Abs and columns for neutrophil isolation were purchased from Stem Cell Technologies (Vancouver, Canada).

Isolation and culture of bone marrow-derived mouse neutrophils

Bone marrow neutrophils were isolated as described previously (11). To obtain the bone marrow cell suspension, the femur and tibia of a mouse were flushed with RPMI 1640. Tissue fragments were removed by rapid filtration through a glass-wool column, and cells were collected by centrifugation. The cell pellets were resuspended in RPMI 1640/1% FCS and then incubated with primary Abs specific for cell surface markers F4/80, CD4, CD45R, CD5, and TER119 for 15 min at 4°C. This custom mixture (StemCell Technologies) is specific for T and B cells, RBC, monocytes, and macrophages. After a 15-min incubation, 100 µl of antibiotin tetrameric Ab complexes were added, and the cells were incubated for 15 min at 4°C. Then 60 µl of colloidal magnetic dextran iron particles were added to the suspension and incubated for 15 min at 4°C. The entire cell suspension was then placed into a column surrounded by a magnet. The T cells, B cells, RBC, monocytes, and macrophages were captured in the column, allowing the neutrophils to pass through by negative selection methods. Neutrophil purity, as determined by Wright’s-stained cytospin preparations, was >97%. Bone marrow neutrophils (2 x 106/0.5 ml) were cultured in RPMI 1640/0.2% FCS.

Cytokine ELISA

Immunoreactive TNF-{alpha} and MIP-2 were quantified using commercially available ELISA kits (R&D Systems, Minneapolis, MN), according to the manufacturer’s instructions and as described previously (11).

EMSA

Nuclear extracts were prepared and assayed by EMSA as previously described (11). For analysis of NF-{kappa}B, the {kappa}B-DNA sequence of the Ig gene was used. Synthetic double-stranded sequences (with enhancer motifs underlined) were filled in and labeled with [{alpha}-32P]dATP using Sequenase DNA polymerase as follows: {kappa}B, 5'-TTTTCGAGCTCGGGACTTTCCGAGC-3' and 3'-GCTCGAGCCCTGAAAGGCTCGTTTT-5'. For AP1, the consensus oligonucleotides used were 5'-CGC TTG ATG AGT CAG CCG GAA-3' and 3'-GCG AAC TAC TCA GTC GGC CTT-5' (Promega, Madison, WI).

Western blot analysis

Western blots for phosphorylated and total kinases were performed as described previously (11). Parallel samples for total protein kinase were run with samples for activation-specific phosphorylation analysis. Densitometry was performed using a chemiluminescence system and analysis software (Bio-Rad, Hercules, CA) to determine the ratio between phosphorylated and total kinase.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Materials and Methods
 Results
Discussion
 References
 
TLR2 engagement induces cytokine release and activation of Akt in neutrophils

Activation of neutrophil TLR2 resulted in rapid release of TNF-{alpha} and MIP-2, as shown in Fig. 1A. To determine the ability of TLR2 engagement to activate Akt, we cultured murine bone marrow-derived neutrophils with the TLR2 ligands PGN and PAM, the TLR4 ligand LPS, and the TLR-independent agents fMLP and C5a. TLRs, like most other receptors, appear to function as dimers and heterodimerize (7). PGN is reported to be a specific agonist of TLR2/TLR6 heterodimers, whereas PAM is specific for TLR1/TLR2 heterodimers, with distinct signaling and cellular responses reported for TLR1/TLR2 and TLR2/TLR6 engagement (41, 42). For this reason we examined the responses of both TLR1/TLR2- and TLR2/TLR6-specific engagement. Fig. 1 shows that exposure of neutrophils to PAM or PGN resulted in Akt activation that was both greater in intensity and more prolonged than that found after LPS-induced TLR4 stimulation. Although the agonists fMLP and C5a induced robust Akt activation, the duration of this effect was shorter than that found after exposure to TLR2 or TLR4 ligands.



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  		FIGURE 1. TLR2 agonists induce MIP-2 and TNF-{alpha} release as well as Akt activation in neutrophils. A, Neutrophils were stimulated with 50 µg/ml PGN for the times shown, and concentrations of MIP-2 and TNF-{alpha} in supernatants were determined by ELISA. B, Neutrophils were stimulated with 10 nM C5a, 50 µg/ml PGN, 50 µg/ml PAM, or 1 µg/ml LPS for increasing times, and the reactions were terminated with SDS-PAGE sample buffer. Cell lysates were subjected to SDS-PAGE, and Western blot analysis with specific Abs for phospho-Ser473 Akt and total Akt were performed. The fold increase in Akt phospho-Ser473 is plotted vs time (minutes). The data shown are representative of three independent experiments. The mean ± SEM are presented.

 
TLR2-stimulated MIP-2 and TNF-{alpha} release is dependent on Akt activation

Neutrophils stimulated with PGN or PAM secreted increased amounts of MIP-2 (Fig. 2) and TNF-{alpha} (Fig. 3) in a dose-dependent manner, with an IC50 similar to that reported for PGN and PAM stimulation of TLR2 responses by other investigators (41). Addition of the specific PI3-K inhibitor wortmannin to the neutrophil cultures diminished PAM- or PGN-induced MIP-2 and TNF-{alpha} production (Figs. 2 and 3).



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  		FIGURE 2. TLR2-stimulated MIP-2 release is reduced by PI3-K inhibition. Neutrophils were pretreated with 10 nM wortmannin or vehicle (0.1% DMSO) for 10 min, then cultured with increasing concentrations of PGN (A) or PAM (B) for 90 min. MIP-2 concentrations in the supernatants were determined by ELISA. The data shown are representative of three independent experiments. The mean ± SEM are presented.

 


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  		FIGURE 3. TLR2-stimulated TNF-{alpha} release is reduced by PI3-K inhibition. Neutrophils were pretreated with 10 nM wortmannin or vehicle (0.1% DMSO) for 10 min, then cultured with increasing concentrations of PGN (A) or PAM (B) for 90 min. The reactions were terminated by centrifugation, and the concentrations of TNF-{alpha} in the supernatants were determined by ELISA. The data shown are representative of three independent experiments. The mean ±SEM are presented.

 
To confirm that activation of Akt was involved in the TLR2-induced production of proinflammatory cytokines, we determined the IC50 for inhibition of Akt activation and production of MIP-2 using two specific, structurally distinct inhibitors, wortmannin and LY294002, that block PI3-K (43). As shown in Fig. 4, similar values were obtained for the IC50 of wortmannin- or LY294002-associated inhibition of Akt activation and MIP-2 secretion. In particular, with wortmannin the IC50 was 5 nM for Akt activation and 14 nM for MIP-2 secretion, whereas with LY294002 the IC50 was 18 µM for Akt activation and 12 µM for MIP-2 secretion. Of note, these IC50 values for Akt inhibition by wortmannin and LY294002 are similar to those reported by other investigators for PI3-K-dependent responses in neutrophils (5) and other cell populations (44), indicating that the effects of wortmannin and LY294002 on MIP-2 production are a result of specific actions on PI3-K-Akt.



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  		FIGURE 4. PI3-K blockade inhibits TLR2-stimulated MIP-2 release and Akt phosphorylation with similar kinetics. Neutrophils were pretreated with increasing concentrations of wortmannin (A) or LY294002 (B) for 10 min and subsequently stimulated with 50 µg/ml PGN for 60 min. The reactions were terminated by centrifugation, and the concentrations of MIP-2 in the supernatants were determined by ELISA. Cell pellets were solubilized with SDS-PAGE sample buffer and subjected to SDS-PAGE and Western blot analysis with specific Abs for phospho-Ser473 Akt and total Akt. Representative gels are shown, and the fold increase in Akt phospho-Ser473 is plotted vs concentrations of inhibitors. The data shown are representative of three independent experiments. The mean ± SEM are presented.

 
Effects of TLR2-induced Akt activation on NF-{kappa}B translocation and phosphorylation

Binding elements for NF-{kappa}B are present in the promoters of TNF-{alpha} and MIP-2, and activation of NF-{kappa}B occupies a central role in regulating the production of these proinflammatory cytokines. In previous studies (11) we showed that inhibition of PI3-K-dependent activation of Akt in neutrophils stimulated with the TLR4 ligand LPS resulted in decreased nuclear accumulation of NF-{kappa}B as well as expression of MIP-2 and TNF-{alpha}. To determine whether a similar Akt-dependent pathway was involved in neutrophils activated via TLR2, nuclear translocation of NF-{kappa}B was examined in PGN-stimulated neutrophils cultured with or without wortmannin. As shown in Fig. 5A, the increased nuclear accumulation of NF-{kappa}B induced by stimulation of neutrophils with PGN was not affected by concentrations of wortmannin that produced total inhibition of Akt activation (see Fig. 4).



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  		FIGURE 5. Inhibition of PI3-K prevents PGN-induced phosphorylation of p65 NF-{kappa}B, but not the nuclear translocation of NF-{kappa}B. A, Neutrophils were pretreated with vehicle control (0.1% DMSO) or 33 nM wortmannin for 10 min, then stimulated for increasing times for up to 60 min with 50 µg/ml PGN. Nuclear extracts were prepared and subjected to EMSA. B, For analysis of phospho-p65 Ser536, neutrophils were pretreated for 10 min with increasing concentrations of wortmannin and subsequently stimulated with 50 µg/ml PGN for 45 min. The cell pellets were solubilized with SDS-PAGE sample buffer, then analyzed by Western blot with specific Abs for phospho-p65 Ser536 and total p65. C, Neutrophils were pretreated with either 0.1% DMSO vehicle control or 10 nM wortmannin for 10 min, then stimulated with 50 µg/ml PGN for the times shown. Reactions were terminated with SDS-PAGE sample buffer and analyzed by Western blot with specific Abs for phospho-p65 Ser536 and total p65. The data shown are representative of three independent experiments.

 
The inability of PI3-K inhibition to prevent nuclear translocation of NF-{kappa}B after TLR2 engagement indicates that other mechanisms are responsible for the inhibitory effects of PI3-K blockade on NF-{kappa}B-dependent cytokine expression. Recent data have shown that phosphorylation of the p65 subunit of NF-{kappa}B is an important event in modulating the transcriptional activity of NF-{kappa}B. In particular, phosphorylation of p65 is required for maximal NF-{kappa}B-dependent transcription and expression of NF-{kappa}B-dependent genes (10, 24, 45, 46). To determine whether TLR2-induced activation of Akt participates in this mechanism for regulating NF-{kappa}B-dependent transcription, we examined the levels of p65 phosphorylation in PGN-stimulated neutrophils with or without blockade of PI3-K. As shown in Fig. 5, B and C, phosphorylation of Ser536 in p65, an event that is known to be essential for maximal NF-{kappa}B activity (10, 24, 45, 46), was decreased in neutrophils in which PI3-K was blocked with wortmannin. Importantly, the inhibitory effects of wortmannin on p65 phosphorylation occurred at the same concentrations as those blocking Akt activation and secretion of MIP-2 (Fig. 4), indicating that such effects are linked.

Inhibition of Akt prevents TLR2-induced activation of ERK1/2 and p38

Previous studies (2, 3) demonstrated that PI3-K and Akt can be involved in modulating activation of p38 or ERK1/2, but such interactions have not been demonstrated in cells stimulated via the TLR2 pathway. To address this issue, we examined phosphorylation of p38 and ERK1/2 in PGN-stimulated neutrophils treated with increasing concentrations of wortmannin. Fig. 6 shows that the IC50 values for inhibition of PGN-stimulated phosphorylation of ERK1/2 and p38 were consistent with those for inhibition of MIP-2 secretion (Figs. 2 and 4) and Akt activation (Fig. 4), indicating that Akt is directly involved in modulating the activation of ERK1/2 and p38 in TLR2-stimulated neutrophils.



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  		FIGURE 6. Blockade of PI3-K inhibits TLR2-induced ERK1/2 and p38 mitogen-activated protein kinase activation. A, Neutrophils were pretreated with 0.1% DMSO vehicle control or 10 nM wortmannin for 10 min and subsequently stimulated with 50 µg/ml PGN for increasing times. The cell pellets were solubilized with SDS-PAGE sample buffer and subjected to SDS-PAGE analysis, and Western blot was performed with specific Abs for phospho-ERK1(Thr202/Tyr204), phospho-ERK2(Thr183/Tyr185), and phospho-p38(Thr180/Tyr182). B, Neutrophils were pretreated with increasing concentrations of wortmannin for 10 min, then stimulated with 50 µg/ml PGN for 30 min. The cell pellets were solubilized with SDS-PAGE sample buffer and subjected to SDS-PAGE analysis, and Western blot was performed with specific Abs for phospho-ERK1, phospho-ERK2, and phospho-p38. The data shown are representative of three independent experiments.

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Materials and Methods
 Results
 Discussion
References
 
In neutrophils, fMLP, IL-8, GM-CSF, leukotriene B4, and LPS have been demonstrated to activate Akt (2, 3), but the ability of TLR2 engagement to affect Akt has not been examined previously. In the present studies we found that Akt plays a central role in the activation of downstream kinases, such as p38 and ERK1/2, and transcriptional regulatory factors, including NF-{kappa}B, contributing to the generation of proinflammatory cytokines in TLR2-stimulated neutrophils.

Although the intracellular signaling pathways associated with TLR2 or TLR4 have been reported to be similar, previous experiments have demonstrated marked differences in the patterns of neutrophil functions after engagement of TLR2 or TLR 4, raising the issue of the mechanisms modulating such distinct patterns in neutrophil reactions. For example, neutrophil survival appears to be primarily dependent on TLR4 (47, 48). Recent data have shown the existence of MyD88-independent pathways that result in STAT1 tyrosine phosphorylation and the production of IFN-{beta} after TLR4, but not TLR2, activation (49). The present studies, showing that the kinetics and magnitude of activation of Akt were markedly different after stimulation of neutrophils via TLR4 vs TLR2, provide a potential mechanism, involving differential activation of Akt, that may play an important role in determining neutrophil responses to different TLR agonists.

Interactions between TLR and PI3-K-Akt signaling appear to be tissue and receptor specific (7, 41). LPS-induced NF-{kappa}B activation is Akt dependent in some tissues (11, 12, 13), but not in others (24, 32, 33). TLR1, –2, and –6, but not TLR3, –4, and –5, contain a consensus binding motif (YXXM) (1) for the PI3-K p85 subunit (7). A putative consensus binding motif has also been found in the TLR system adapter protein MyD88 (7). The presence of the putative PI3-K binding site in TLR1, –2, and –6 suggests that these receptors might be directly involved in activating PI3-K, whereas TLR3, –4, and –5 may be able to do so only through indirect mechanisms. In the present experiments the time course of Akt activation by LPS in neutrophils was found to be much slower than that classically seen with other PI3-K-Akt activators, such as fMLP and C5a, that signal through GPCRs (1, 2, 3). The TLR2 agonists, PGN and PAM, also produced rapid activation of Akt, similar to that found with GPCRs. Unlike the short duration of Akt activation present after neutrophil exposure to GPCR ligands, that induced through TLR2 was prolonged. Of note, in contrast to the present findings of dependence of MIP-2 and TNF-{alpha} generation on Akt activation after TLR2 stimulation, GPCR engagement by C5a results in Akt-dependent superoxide production in neutrophils, but little or no cytokine release in neutrophils (2, 3).

Although the classic paradigm for modulation of NF-{kappa}B-dependent transcription involves regulation of translocation of the p65:p50 active heterodimer into the nucleus, there is reason to believe that phosphorylation events involving the p65 subunit are important in this process (10, 24, 45). In particular, recent studies demonstrate that nuclear translocation of NF-{kappa}B is not sufficient to activate NF-{kappa}B-dependent transcription (10, 45). Rather, phosphorylation of the p65 subunit at Ser276 (46), Ser529, or Ser536 (24) is required for maximal transcriptional activity of NF-{kappa}B (10, 46). The molecular mechanism by which p65 phosphorylation increases NF-{kappa}B activity is not well defined, but may involve reversible acetylation, which is linked to the duration of NF-{kappa}B activation (50). Data from the present experiments, showing that blockade of PI3-K did not inhibit TLR2-dependent nuclear translocation of NF-{kappa}B, but did decrease Ser536 phosphorylation of p65, suggest that regulation of this phosphorylation event is responsible for the involvement of PI3-K and Akt in TLR2-dependent NF-{kappa}B transcription.

These experiments also found that TLR2-induced activation of p38 and ERK1/2 was dependent on PI3-K. In previous studies a connection between PI3-K and ERK1/2 and p38 has been demonstrated for other receptors, but not TLR2. In particular, activation of p38 and ERK1/2 initiated by C5a, chemokines, Fc{gamma}RIIa, or Fc{gamma}RIIIb is attenuated by inhibition of PI3-K (51, 52). Similarly, the stimulation of ERK1/2 produced by phagocytosis of plasma-opsonized Staphyloccocus aureus is dependent on PI3-K (4). Although p38 clearly participates in regulating NF-{kappa}B-dependent transcription in neutrophils and other cells (9, 53, 54, 55, 56, 57), the mechanisms involved in these effects on NF-{kappa}B activation are less well defined. Although some studies have suggested that p38 directly affects nuclear translocation of NF-{kappa}B (56), others have found that the primary mechanism by which p38 modulates NF-{kappa}B-dependent transcription is through phosphorylation of NF-{kappa}B-associated coactivator proteins, such as TATA-binding protein (9). The demonstration that PI3-K participates in TLR2-induced activation of p38 and phosphorylation of p65 is consistent with such p38-dependent phosphorylation events.

In summary, the present studies demonstrate that Akt occupies a central role in modulating TLR2-induced proinflammatory cytokine release by neutrophils. Unlike TLR4 activation, in which the kinetics of Akt activation are delayed, and the proinflammatory effects of Akt involve enhancing nuclear translocation of NF-{kappa}B, TLR2 stimulation in neutrophils produces rapid increases in Akt activation that appear to affect NF-{kappa}B-dependent transcription not through increasing nuclear accumulation of this transcriptional factor, but, rather, by phosphorylating the p65 subunit. These results highlight the distinct intracellular events activated by TLR2 vs TLR4 in neutrophils and provide insight into mechanisms that may result in differing neutrophil responses after TLR2 or TLR4 activation.


   Footnotes
 
1 This work was supported by National Institutes of Health Grants HL62221 and PO1HL68743 (to E.A.), French Ministry of Foreign Affairs (LAVOISIER Program), and French Society of Anesthesiology and Critical Care Medicine: SFAR (to K.A.). Back

2 Address correspondence and reprint requests to Dr. Derek Strassheim, Division of Pulmonary Science and Critical Care Medicine, University of Colorado Health Sciences Center, Box C272, 4200 East 9th Avenue, Denver, CO 80262. E-mail address: derek.strassheim{at}uchsc.edu Back

3 Abbreviations used in this paper: PI3-K, phosphoinositide 3-kinase; ERK, extracellular receptor-activated kinase; GPCR, heterotrimeric (GTP)-binding protein (G protein)-coupled receptors; MIP, macrophage inflammatory protein; p38, p38 mitogen-activated protein kinase; PAM, tri-palmitoyl-S-glyceryl-Cys-Ser-(Lys)4; PGN, peptidoglycan; PIMIP-2, macrophage inflammatory protein-2. Back

Received for publication November 24, 2003. Accepted for publication February 19, 2004.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Materials and Methods
 Results
 Discussion
 References
 

  1. Cantley, L. C.. 2002. The phosphoinositide 3-kinase pathway. Science 296:1655.[Abstract/Free Full Text]
  2. Hirsch, E., V. L. Katanaev, C. Garlanda, O. Azzolino, L. Pirola, L. Silengo, S. Sozzani, A. Mantovani, F. Altruda, M. P. Wymann. 2000. Central role for G protein-coupled phosphoinositide 3-kinase {gamma} in inflammation. Science 287:1049.[Abstract/Free Full Text]
  3. Wymann, M. P., S. Sozzani, F. Altruda, A. Mantovani, E. Hirsch. 2000. Lipids on the move: phosphoinositide 3-kinases in leukocyte function. Immunol. Today 21:260.[Medline]
  4. McLeish, K. R., J. B. Klein, P. Y. Coxon, K. Z. Head, R. A. Ward. 1998. Bacterial phagocytosis activates extracellular signal-regulated kinase and p38 mitogen-activated protein kinase cascades in human neutrophils. J. Leukocyte Biol. 64:835.[Abstract]
  5. Capodici, C., S. Hanft, M. Feoktistov, M. H. Pillinger. 1998. Phosphatidylinositol 3-kinase mediates chemoattractant-stimulated, CD11b/CD18-dependent cell-cell adhesion of human neutrophils: evidence for an ERK-independent pathway. J. Immunol. 160:1901.[Abstract/Free Full Text]
  6. Abraham, E.. 2003. Neutrophils and acute lung injury. Crit. Care Med. 31:S195.[Medline]
  7. Akira, S.. 2003. Toll-like receptor signaling. J. Biol. Chem. 278:38105.[Free Full Text]
  8. Ghosh, S., M. Karin. 2002. Missing pieces in the NF-{kappa}B puzzle. Cell 109:(Suppl.):S81.
  9. Carter, A. B., K. L. Knudtson, M. M. Monick, G. W. Hunninghake. 1999. The p38 mitogen-activated protein kinase is required for NF-{kappa}B-dependent gene expression: the role of TATA-binding protein (TBP). J. Biol. Chem. 274:30858.[Abstract/Free Full Text]
  10. Vermeulen, L., G. De Wilde, S. Notebaert, W. Vanden Berghe, G. Haegeman. 2002. Regulation of the transcriptional activity of the nuclear factor-{kappa}B p65 subunit. Biochem. Pharmacol. 64:963.[Medline]
  11. Yum, H. K., J. Arcaroli, J. Kupfner, R. Shenkar, J. M. Penninger, T. Sasaki, K. Y. Yang, J. S. Park, E. Abraham. 2001. Involvement of phosphoinositide 3-kinases in neutrophil activation and the development of acute lung injury. J. Immunol. 167:6601.[Abstract/Free Full Text]
  12. Manna, S. K., B. B. Aggarwal. 2000. Wortmannin inhibits activation of nuclear transcription factors NF-{kappa}B and activated protein-1 induced by lipopolysaccharide and phorbol ester. FEBS Lett. 473:113.[Medline]
  13. Li, X., J. C. Tupper, D. D. Bannerman, R. K. Winn, C. J. Rhodes, J. M. Harlan. 2003. Phosphoinositide 3 kinase mediates Toll-like receptor 4-induced activation of NF-{kappa}B in endothelial cells. Infect. Immun. 71:4414.[Abstract/Free Full Text]
  14. Romashkova, J. A., S. S. Makarov. 1999. NF-{kappa}B is a target of Akt in anti-apoptotic PDGF signalling. Nature 401:86.[Medline]
  15. Mayo, M. W., L. V. Madrid, S. D. Westerheide, D. R. Jones, X. J. Yuan, A. S. Baldwin, Jr, Y. E. Whang. 2002. PTEN blocks tumor necrosis factor-induced NF-{kappa}B-dependent transcription by inhibiting the transactivation potential of the p65 subunit. J. Biol. Chem. 277:11116.[Abstract/Free Full Text]
  16. Osawa, Y., M. Nagaki, Y. Banno, D. A. Brenner, T. Asano, Y. Nozawa, H. Moriwaki, S. Nakashima. 2002. Tumor necrosis factor {alpha}-induced interleukin-8 production via NF-{kappa}B and phosphatidylinositol 3-kinase/Akt pathways inhibits cell apoptosis in human hepatocytes. Infect. Immun. 70:6294.[Abstract/Free Full Text]
  17. Ozes, O. N., H. Akca, L. D. Mayo, J. A. Gustin, T. Maehama, J. E. Dixon, D. B. Donner. 2001. A phosphatidylinositol 3-kinase/Akt/mTOR pathway mediates and PTEN antagonizes tumor necrosis factor inhibition of insulin signaling through insulin receptor substrate-1. Proc. Natl. Acad. Sci. USA 98:4640.[Abstract/Free Full Text]
  18. Reddy, S. A., J. H. Huang, W. S. Liao. 2000. Phosphatidylinositol 3-kinase as a mediator of TNF-induced NF-{kappa}B activation. J. Immunol. 164:1355.[Abstract/Free Full Text]
  19. Reddy, S. A., J. H. Huang, W. S. Liao. 1997. Phosphatidylinositol 3-kinase in interleukin 1 signaling: physical interaction with the interleukin 1 receptor and requirement in NF{kappa}B and AP-1 activation. J. Biol. Chem. 272:29167.[Abstract/Free Full Text]
  20. Sizemore, N., S. Leung, G. R. Stark. 1999. Activation of phosphatidylinositol 3-kinase in response to interleukin-1 leads to phosphorylation and activation of the NF-{kappa}B p65/RelA subunit. Mol. Cell. Biol. 19:4798.[Abstract/Free Full Text]
  21. Pianetti, S., M. Arsura, R. Romieu-Mourez, R. J. Coffey, G. E. Sonenshein. 2001. Her-2/neu overexpression induces NF-{kappa}B via a PI3-kinase/Akt pathway involving calpain-mediated degradation of I{kappa}B-{alpha} that can be inhibited by the tumor suppressor PTEN. Oncogene 20:1287.[Medline]
  22. Yang, C. H., A. Murti, S. R. Pfeffer, J. G. Kim, D. B. Donner, L. M. Pfeffer. 2001. Interferon {alpha}/{beta} promotes cell survival by activating nuclear factor {kappa}B through phosphatidylinositol 3-kinase and Akt. J. Biol. Chem. 276:13756.[Abstract/Free Full Text]
  23. Kitaura, J., K. Asai, M. Maeda-Yamamoto, Y. Kawakami, U. Kikkawa, T. Kawakami. 2000. Akt-dependent cytokine production in mast cells. J. Exp. Med. 192:729.[Abstract/Free Full Text]
  24. Yang, F., E. Tang, K. Guan, C. Y. Wang. 2003. IKK{beta} plays an essential role in the phosphorylation of RelA/p65 on serine 536 induced by lipopolysaccharide. J. Immunol. 170:5630.[Abstract/Free Full Text]
  25. Madrid, L. V., M. W. Mayo, J. Y. Reuther, A. S. Baldwin, Jr. 2001. Akt stimulates the transactivation potential of the RelA/p65 subunit of NF-{kappa}B through utilization of the I{kappa}B kinase and activation of the mitogen-activated protein kinase p38. J. Biol. Chem. 276:18934.[Abstract/Free Full Text]
  26. Madrid, L. V., C. Y. Wang, D. C. Guttridge, A. J. Schottelius, A. S. Baldwin, Jr, M. W. Mayo. 2000. Akt suppresses apoptosis by stimulating the transactivation potential of the RelA/p65 subunit of NF-{kappa}B. Mol. Cell. Biol. 20:1626.[Abstract/Free Full Text]
  27. Koul, D., Y. Yao, J. L. Abbruzzese, W. K. Yung, S. A. Reddy. 2001. Tumor suppressor MMAC/PTEN inhibits cytokine-induced NF{kappa}B activation without interfering with the I{kappa}B degradation pathway. J. Biol. Chem. 276:11402.[Abstract/Free Full Text]
  28. Rauch, B. H., A. Weber, M. Braun, N. Zimmermann, K. Schror. 2000. PDGF-induced Akt phosphorylation does not activate NF-{kappa}B in human vascular smooth muscle cells and fibroblasts. FEBS Lett. 481:3.[Medline]
  29. Madge, L. A., J. S. Pober. 2000. A phosphatidylinositol 3-kinase/Akt pathway, activated by tumor necrosis factor or interleukin-1, inhibits apoptosis but does not activate NF{kappa}B in human endothelial cells. J. Biol. Chem. 275:15458.[Abstract/Free Full Text]
  30. Delhase, M., N. Li, M. Karin. 2000. Kinase regulation in inflammatory response. Nature 406:367.[Medline]
  31. Chen, B. C., W. T. Wu, F. M. Ho, W. W. Lin. 2002. Inhibition of interleukin-1{beta}-induced NF-{kappa}B activation by calcium/calmodulin-dependent protein kinase kinase occurs through Akt activation associated with interleukin-1 receptor-associated kinase phosphorylation and uncoupling of MyD88. J. Biol. Chem. 277:24169.[Abstract/Free Full Text]
  32. Guha, M., N. Mackman. 2002. The phosphatidylinositol 3-kinase-Akt pathway limits lipopolysaccharide activation of signaling pathways and expression of inflammatory mediators in human monocytic cells. J. Biol. Chem. 277:32124.[Abstract/Free Full Text]
  33. Ojaniemi, M., V. Glumoff, K. Harju, M. Liljeroos, K. Vuori, M. Hallman. 2003. Phosphatidylinositol 3-kinase is involved in Toll-like receptor 4-mediated cytokine expression in mouse macrophages. Eur. J. Immunol. 33:597.[Medline]
  34. Fukao, T., S. Koyasu. 2003. PI3K and negative regulation of TLR signaling. Trends Immunol. 24:358.[Medline]
  35. Fukao, T., M. Tanabe, Y. Terauchi, T. Ota, S. Matsuda, T. Asano, T. Kadowaki, T. Takeuchi, S. Koyasu. 2002. PI3K-mediated negative feedback regulation of IL-12 production in DCs. Nat. Immunol. 3:875.[Medline]
  36. Monick, M. M., P. K. Robeff, N. S. Butler, D. M. Flaherty, A. B. Carter, M. W. Peterson, G. W. Hunninghake. 2002. Phosphatidylinositol 3-kinase activity negatively regulates stability of cyclooxygenase 2 mRNA. J. Biol. Chem. 277:32992.[Abstract/Free Full Text]
  37. Hayashi, F., T. K. Means, A. D. Luster. 2003. Toll-like receptors stimulate human neutrophil function. Blood 102:2660.[Abstract/Free Full Text]
  38. Kurt-Jones, E. A., L. Mandell, C. Whitney, A. Padgett, K. Gosselin, P. E. Newburger, R. W. Finberg. 2002. Role of toll-like receptor 2 (TLR2) in neutrophil activation: GM-CSF enhances TLR2 expression and TLR2-mediated interleukin 8 responses in neutrophils. Blood 100:1860.[Abstract/Free Full Text]
  39. Cockerill, F. R., III, J. G. Hughes, E. A. Vetter, R. A. Mueller, A. L. Weaver, D. M. Ilstrup, J. E. Rosenblatt, W. R. Wilson. 1997. Analysis of 281,797 consecutive blood cultures performed over an eight-year period: trends in microorganisms isolated and the value of anaerobic culture of blood. Clin. Infect. Dis. 24:403.[Medline]
  40. Arbibe, L., J. P. Mira, N. Teusch, L. Kline, M. Guha, N. Mackman, P. J. Godowski, R. J. Ulevitch, U. G. Knaus. 2000. Toll-like receptor 2-mediated NF-{kappa}B activation requires a Rac1-dependent pathway. Nat. Immunol. 1:533.[Medline]
  41. McCurdy, J. D., T. J. Olynych, L. H. Maher, J. S. Marshall. 2003. Cutting edge: distinct Toll-like receptor 2 activators selectively induce different classes of mediator production from human mast cells. J. Immunol. 170:1625.[Abstract/Free Full Text]
  42. Ozinsky, A., D. M. Underhill, J. D. Fontenot, A. M. Hajjar, K. D. Smith, C. B. Wilson, L. Schroeder, A. Aderem. 2000. The repertoire for pattern recognition of pathogens by the innate immune system is defined by cooperation between toll-like receptors. Proc. Natl. Acad. Sci. USA 97:13766.[Abstract/Free Full Text]
  43. Walker, E. H., M. E. Pacold, O. Perisic, L. Stephens, P. T. Hawkins, M. P. Wymann, R. L. Williams. 2000. Structural determinants of phosphoinositide 3-kinase inhibition by wortmannin, LY294002, quercetin, myricetin, and staurosporine. Mol. Cell 6:909.[Medline]
  44. Su, T. Z., M. Wang, L. J. Syu, A. R. Saltiel, D. L. Oxender. 1998. Regulation of system A amino acid transport in 3T3–L1 adipocytes by insulin. J. Biol. Chem. 273:3173.[Abstract/Free Full Text]
  45. Vermeulen, L., G. De Wilde, P. Van Damme, W. Vanden Berghe, G. Haegeman. 2003. Transcriptional activation of the NF-{kappa}B p65 subunit by mitogen- and stress-activated protein kinase-1 (MSK1). EMBO J. 22:1313.[Medline]
  46. Okazaki, T., S. Sakon, T. Sasazuki, H. Sakurai, T. Doi, H. Yagita, K. Okumura, H. Nakano. 2003. Phosphorylation of serine 276 is essential for p65 NF-{kappa}B subunit-dependent cellular responses. Biochem. Biophys. Res. Commun. 300:807.[Medline]
  47. Yang, K. Y., J. Arcaroli, J. Kupfner, T. M. Pitts, J. S. Park, D. Strasshiem, R. P. Perng, E. Abraham. 2003. Involvement of phosphatidylinositol 3-kinase {gamma} in neutrophil apoptosis. Cell. Signal. 15:225.[Medline]
  48. Sabroe, I., L. R. Prince, E. C. Jones, M. J. Horsburgh, S. J. Foster, S. N. Vogel, S. K. Dower, M. K. Whyte. 2003. Selective roles for Toll-like receptor (TLR)2 and TLR4 in the regulation of neutrophil activation and life span. J. Immunol. 170:5268.[Abstract/Free Full Text]
  49. Rhee, S. H., B. W. Jones, V. Toshchakov, S. N. Vogel, M. J. Fenton. 2003. Toll-like receptors 2 and 4 activate STAT1 serine phosphorylation by distinct mechanisms in macrophages. J. Biol. Chem. 278:22506.[Abstract/Free Full Text]
  50. Chen, L., W. Fischle, E. Verdin, W. C. Greene. 2001. Duration of nuclear NF-{kappa}B action regulated by reversible acetylation. Science 293:1653.[Abstract/Free Full Text]
  51. Coxon, P. Y., M. J. Rane, D. W. Powell, J. B. Klein, K. R. McLeish. 2000. Differential mitogen-activated protein kinase stimulation by Fc{gamma} receptor IIa and Fc{gamma} receptor IIIb determines the activation phenotype of human neutrophils. J. Immunol. 164:6530.[Abstract/Free Full Text]
  52. Rane, M. J., P. Y. Coxon, D. W. Powell, R. Webster, J. B. Klein, W. Pierce, P. Ping, K. R. McLeish. 2001. p38 Kinase-dependent MAPKAPK-2 activation functions as 3-phosphoinositide-dependent kinase-2 for Akt in human neutrophils. J. Biol. Chem. 276:3517.[Abstract/Free Full Text]
  53. Carter, A. B., M. M. Monick, G. W. Hunninghake. 1999. Both Erk and p38 kinases are necessary for cytokine gene transcription. Am. J. Respir. Cell Mol. Biol. 20:751.[Abstract/Free Full Text]
  54. Nick, J. A., S. K. Young, K. K. Brown, N. J. Avdi, P. G. Arndt, B. T. Suratt, M. S. Janes, P. M. Henson, G. S. Worthen. 2000. Role of p38 mitogen-activated protein kinase in a murine model of pulmonary inflammation. J. Immunol. 164:2151.[Abstract/Free Full Text]
  55. Saccani, S., S. Pantano, G. Natoli. 2002. p38-Dependent marking of inflammatory genes for increased NF-{kappa}B recruitment. Nat. Immunol. 3:69.[Medline]
  56. Nick, J. A., N. J. Avdi, S. K. Young, L. A. Lehman, P. P. McDonald, S. C. Frasch, M. A. Billstrom, P. M. Henson, G. L. Johnson, G. S. Worthen. 1999. Selective activation and functional significance of p38{alpha} mitogen-activated protein kinase in lipopolysaccharide-stimulated neutrophils. J. Clin. Invest. 103:851.[Medline]
  57. Nick, J. A., S. K. Young, P. G. Arndt, J. G. Lieber, B. T. Suratt, K. R. Poch, N. J. Avdi, K. C. Malcolm, C. Taube, P. M. Henson, et al 2002. Selective suppression of neutrophil accumulation in ongoing pulmonary inflammation by systemic inhibition of p38 mitogen-activated protein kinase. J. Immunol. 169:5260.[Abstract/Free Full Text]



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