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Published online before print August 2, 2006
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Department of Academic Surgery, Cork University Hospital, National University of Ireland (NUI)/University College Cork (UCC), Wilton, Cork, Ireland
1 Correspondence: Department of Academic Surgery, Cork University Hospital, National University of Ireland (NUI)/University College Cork (UCC), Wilton, Cork, Ireland. E-mail: jh.wang{at}ucc.ie
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ABSTRACT |
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Key Words: lipoprotein • lipopolysaccharide • tolerance • cross-tolerance • TLR signaling
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INTRODUCTION |
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and IL-6. The phenomenon of tolerance is an adaptive host response to bacterial infection, which was first described by Beeson [4 ] in 1946. It was found that pre-exposure to low-dose LPS induces a transient state of cellular hyporesponsiveness with decreased production of proinflammatory cytokines [5 ], thereby conferring protection against a subsequent "lethal" LPS challenge, resulting in a significant survival advantage [6 ]. LPS tolerance has been the focus of extensive scientific and clinical research over the last several decades in an attempt to elucidate the sequence of changes in cell surface receptor expression, intracellular signaling proteins, gene expression, and cytokine production, which occur in tolerized cells. The findings to date were the subject of a recent review by Fan and Cook [7 ]. Tolerance to components of gram-positive bacterial cell walls such as bacterial lipoprotein (BLP) and LTA is a much lesser studied, although equally important, phenomenon. The discovery of TLRs and their role in pathogen recognition have led to a resurgence in interest in the mechanisms of gram-positive and -negative tolerance, more correctly described as cellular reprogramming [8 ].
TLRs are a highly conserved group of PRRs, initially described in the fruit fly, Drosophila melanogaster [9 ]. Medzhitov et al. [10 ] described the first human homologue of Toll in 1997. Eleven TLRs, TLR1–11, have now been described. TLRs are members of the IL-1 receptor (IL-1R) superfamily and are transmembrane proteins characterized by an extracellular leucine-rich repeat domain and an intracellular Toll/IL-1R homology domain. Each extracellular leucine-rich repeat is a sequence of 24–29 amino acids, and it is the leucine-rich repeat domain that is involved in pathogen recognition.
TLRs play an essential role in activating signal transduction pathways leading to the induction of the inflammatory response. After ligand binding, TLRs sequentially recruit the adaptor molecules MyD88 [11
], IL-1R-associated kinase (IRAK) [12
], and TNF receptor-associated factor 6 (TRAF6) [13
]. In turn, these adaptor molecules activate the I
B kinase (IKK) complex and MAPKs, JNK, p38, and ERK1/2, leading to the activation of NF-B and AP-1, respectively, and ultimately, to transcription of inflammatory target genes [14
]. Multiple studies have confirmed that LPS signals primarily through TLR4 [15
, 16
], and TLR4-deficient, C3H/HeJ mice are hyporesponsive to LPS [17
]. PAMPs from gram-positive bacteria, including peptidoglycan [18
] and LTA [19
20
21
], activate the inflammatory response via TLR2, and TLR2-deficient mice are highly susceptible to gram-positive bacterial challenge [22
].
This review will focus on cellular reprogramming by gram-positive PAMPs, the observed in vivo and in vitro changes, including survival data, cell surface receptor alterations, and changes in intracellular signaling, gene expression, and cytokine production, and the phenomenon of cross-tolerance.
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GRAM-POSITIVE TOLERANCE AND CROSS-TOLERANCE |
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and IL-6, in mice challenged with a second dose of BLP or LPS. Cross-tolerance to BLP in LPS-pretreated animals, however, was not demonstrated, and there was no survival advantage in LPS-tolerized mice when exposed to lethal BLP challenge.
Induction of tolerance to another TLR2 ligand, MALP-2, was the subject of a study by Sato et al. [27
]. Similar to the above results, MALP-2 was found to induce self-tolerance as measured by TNF- production but also, to induce cross-tolerance to LPS. However, the reverse was not shown to be true, and LPS-tolerized macrophages responded to MALP-2 challenge with increased TNF- production. MALP-2 tolerance and cross-tolerance were independent of changes in surface expression of the LPS receptors TLR4-MD2 and CD14.
In vivo work from Deiters et al. [30
] further confirmed Sato’s in vitro results. They demonstrated that pretreatment of mice with an i.p., sublethal dose of MALP-2 significantly attenuated TNF- and IL-6 production to a second MALP-2 or LPS challenge, although protection was only partial with high doses of LPS. Galactosamine, a compound that sensitizes animals to TNF- [31
], was then used in conjunction with MALP-2 pretreatment, which abrogated the response to TNF- stimulation completely. These effects were demonstrated to be dependent on signaling via TLR2, as MALP-2 failed to induce tolerance in TLR2-deficient mice.
Further work from O’Brien et al. [32
] about cross-tolerance demonstrated that induction of BLP tolerance in TLR4-deficient, C3H/HeJ mice induced resistance to gram-negative, Salmonella typhimurium sepsis, to which these animals are highly susceptible. This was associated with an increase in circulating neutrophils and peritoneal macrophages, up-regulation of surface complement receptor 3 (CR3), and Fc
III/IIR on these cells, along with consequent enhanced bacterial recognition, ingestion, and intracellular killing in the TLR4-deficient, BLP-tolerized mice. An early study by Ausobsky et al. [28
] using MDP found that mice, pretreated with MDP, also believed to signal via TLR2 and exposed to a second septic challenge, had increased survival and displayed a hyperphagocytic state compared with naïve mice.
Lehner et al. [33 ] also examined tolerance and cross-tolerance in vivo using LTA. This group demonstrated, by measuring proinflammatory cytokine levels, that LTA pretreatment of C3H/HeN mice induces tolerance to LTA but also cross-tolerance to LPS, and this is TLR2-dependent. Induction of LPS tolerance not only induced self-tolerance but also cross-tolerance to LTA. LPS tolerance or cross-tolerance to LTA could not be induced in TLR4-deficient mice, suggesting that LPS-induced tolerance and cross-tolerance is TLR4-dependent. In vitro data from Jacinto et al. [26 ] correlates partly with Lehner’s results. In vitro pretreatment of THP-1 monocytes with LPS induced tolerance to a second LPS challenge, as well as cross-tolerance to LTA, which, however, did not induce cross-tolerance to LPS. LTA pretreatment induced tolerance only to a subsequent LTA challenge, and cells responded normally to LPS challenge [26 ].
From the data presented, a number of conclusions can be drawn. Gram-positive bacterial cell wall components, BLP, MALP-2, and LTA can induce tolerance in vivo and in vitro to a second gram-positive septic challenge. Gram-positive PAMP recognition, and induction of tolerance is TLR2-dependent. Data about cross-tolerance to date is, however, inconsistent. Several studies report that gram-positive pretreatment induces cross-tolerance to gram-negative challenge, and the reverse is not proven to be true, and LPS-tolerant cells respond normally to gram-positive challenge. However, there are other studies that not only fail to demonstrate gram-positive-induced cross-tolerance but in addition, demonstrate LPS-induced cross-tolerance to gram-positive bacteria.
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CELL SURFACE RECEPTOR ALTERATIONS |
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CD14 is a 55-kDa glycosylphosphatidylinositol-linked glycoprotein, which exists in membrane-bound and soluble forms [34
, 35
] and recognizes and binds LPS with high-affinity [36
]. CD14 has also been implicated in activation of the inflammatory response to gram-positive bacterial components such as BLP [37
, 38
] and LTA [39
], and augmentation of the production of proinflammatory cytokines was in the presence of CD14. A study by Wang et al. [40
] demonstrated that CD14 was not involved in BLP-induced stimulation of or BLP-induced tolerance in THP-1 monocytes. These cells did not display surface expression of membrane CD14 in response to BLP stimulation. Furthermore, when cells were cultured in serum-free medium to eliminate soluble CD14, BLP-induced production of proinflammatory cytokines was unchanged. Pre-exposure of THP-1 cells to BLP induced tolerance not only to BLP but also cross-tolerance to LPS, an effect, which was maintained when cells were cultured in serum-free medium and when cells were cultured with a CD14-blocking mAb, MEM-18. These results indicate that BLP tolerance and cross-tolerance to LPS are independent of CD14. A study by Sato et al. [27
] also refuted a role for CD14 in the development of gram-positive tolerance. Peritoneal macrophages were tolerized in vitro by MALP-2, and when exposed to a second MALP-2 stimulus, no change in surface expression of CD14 was demonstrated. In contrast, in vitro experiments using a human monocytic cell line, Mono-Mac-6, demonstrated that the use of an anti-CD14 mAb decreased BLP-induced TNF- production by almost 50%, and in addition, pretreatment of cells with BLP for 42 h resulted in dramatically up-regulated expression of CD14 and CD14 mRNA [41
]. Despite these results, it has been demonstrated that gram-positive tolerance can be induced in the absence of CD14, thus making it unlikely that this molecule is a key factor in the development of tolerance.
A far more important receptor and one that has been the subject of many scientific studies is TLR2. The surface alterations in TLR2 expression following exposure to bacterial products have resulted in much debate. Early work by Medvedev et al. [42 ] sought to answer this question by engineering a Chinese hamster ovary fibroblast cell line (CHO/CD14) to overexpress TLR2 and TLR4. Having confirmed that LPS activated cells expressing TLR4 and that the mycobacterial components, lipoarabinomannan (LAM) and soluble tuberculosis factor (STF), activated cells expressing TLR2, they proceeded to elucidate the changes in surface TLR2 and TLR4 expression during tolerance and cross-tolerance. They found that there was no change in TLR2 or TLR4 surface expression after cells were pretreated with LAM, STF, or LPS prior to subsequent stimulus. Cross-tolerance to LPS stimulation in LAM- and STF-pretreated cells was demonstrated, and there was no change in TLR2 or TLR4 surface expression. Similarly, cross-tolerance to LAM/STF in LPS-tolerant cells was also proven to be independent of alterations in TLR surface expression. A study carried out by Wang et al. [40 ] examined TLR2 protein expression in THP-1 monocytes. They demonstrated that induction of tolerance to BLP in THP-1 monocytes, which constitutively express TLR2, strongly inhibited BLP-induced TLR2 overexpression. Stimulation with LPS did not affect TLR2 protein expression.
A study by Jacinto et al. [26 ] about LPS and LTA tolerance in THP-1 monocytes repudiated a role for TLR2 as a regulatory protein in LTA tolerance. This group demonstrated that LTA-tolerized cells retain functional TLR2 signaling, as evidenced by selective expression of the anti-inflammatory soluble IL-1R antagonist protein, and theorize that it is alterations in downstream signaling that are responsible for the decrease in proinflammatory cytokines seen in LTA-tolerized cells. RAW264.7 cells tolerized with peptidoglycan also showed no change in TLR2 surface expression when compared with naïve cells [43 ]. Seidlar et al. [41 ] demonstrated a marginal up-regulation in TLR2 surface expression in Mono-Mac-6 cells tolerized with BLP; however, this was not mirrored by a change in TLR2 mRNA expression, which remained unchanged compared with naïve cells. Li et al. [44 ] also demonstrated unchanged TLR2-mRNA levels in their study, which showed marked down-regulation of TLR2 protein expression in BLP-tolerized monocytes. Further work by Li at al. [44 ] demonstrated that cross-tolerance to LPS in BLP-tolerized cells is not associated with alterations in TLR4 protein expression, a finding that is consistent with an earlier study by Sato et al. [27 ], who found no change in surface expression of TLR4-MD2 in peritoneal macrophages pretreated with MALP-2 and exposed to a second MALP-2 stimulus or LPS. The lack of a TLR4 response in gram-positive, tolerant cells is in contrast to changes in TLR2 signaling in LPS tolerance, and multiple studies show up-regulation of TLR2-mRNA [45 , 46 ] or TLR2 surface receptor expression [47 ] in LPS-tolerized mice or cells.
There is a lack of consistent evidence as to the exact alterations in TLR2 surface expression in cells tolerized to gram-positive bacterial components (Table 1 ). However, it seems that although TLR2 is necessary in the inflammatory response to gram-positive bacteria and in the development of gram-positive tolerance and cross-tolerance, it is not the key factor in the development of the latter phenomena.
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INTRACELLULAR SIGNALING PROTEIN ALTERATIONS |
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 TOPABSTRACTINTRODUCTIONGRAM-POSITIVE TOLERANCE AND...CELL SURFACE RECEPTOR... INTRACELLULAR SIGNALING PROTEIN... INCREASED PHAGOCYTOSIS IN GRAM...GRAM-POSITIVE TOLERANCE: THE...SUMMARYREFERENCES |
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B and subsequent transcription of proinflammatory genes [14
, 48
]. This is underlined further by studies, which have observed that MyD88-deficient mice are unresponsive to LPS [49
], peptidoglycan [50
], and MALP-2 [51
]. In LPS tolerance, there is evidence for reduced association of TLR4-MyD88 [45
] and MyD88-IRAK complex formation [52
]. IRAK was originally described as a signal transducer for the proinflammatory cytokine IL-1 and was later discovered to be involved in signal transduction of other TLR/IL-1R family members. To date, four members have been discovered: IRAK-1, IRAK-2, IRAK-4, and IRAK-M [53
]. IRAKs act downstream of MyD88, and all bind to TRAF6 [54
55
56
]. Investigating changes in gram-positive tolerance, Siedlar et al. [41
] demonstrated no change in MyD88 protein expression in Mono-Mac-6 cells pretreated with BLP but found decreased IRAK-1 levels in BLP-tolerant Mono-Mac-6 cells, although this was not associated with a decrease in IRAK-1 mRNA. This was consistent with observed data from Li et al. [44
], who demonstrated no change in MyD88 mRNA or MyD88 protein in THP-1 monocytes stimulated with BLP or LPS, compared with naïve cells. In naïve THP-1 monocytes challenged with BLP, they observed a decrease in IRAK-1 protein 2–6 h post-BLP stimulation. When cells were pretreated with BLP for 24 h, there was a marked decrease in IRAK-1 protein levels compared with naïve cells, and this was not reversed with high-dose BLP or LPS stimulation. A defect in association between MyD88 and IRAK-1 in BLP-tolerized cells was also observed, and the authors hypothesize that this may in part be a result of the decreased levels of IRAK-1 protein in tolerized cells. Sato et al. [27 ] similarly observed no change in IRAK-1 mRNA and IRAK-1 protein levels in mouse macrophages pretreated with MALP-2. Jacinto et al. [26 ] compared IRAK activity in THP-1 monocytes stimulated with LPS and observed that although LPS causes IRAK-1 activation, phosphorylation, and subsequent degradation, LTA induces IRAK-1 activation but not phosphorylation or subsequent degradation. In LPS-tolerant monocytes, IRAK-1 protein and IRAK kinase activity remained low when cells were challenged with a second dose of LPS or LTA. In contrast, in LTA-tolerized cells, IRAK-1 protein remained high, but IRAK kinase activity was suppressed when exposed to subsequent LTA challenge. However, LPS challenge resulted in IRAK kinase activation and protein degradation. The authors postulate that their results indicate that it is disruption of a signaling protein upstream of IRAK that is responsible for LTA tolerance. Dobrovolskaia et al. [57 ] compared alterations in IRAK using LPS, Porphyromonas gingivalis LPS (PgLPS), and BLP to stimulate peritoneal macrophages from C3H/OuJ mice. Their study demonstrated significantly decreased IRAK-1 activation when mice challenged with a TLR2 agonist were exposed to a second challenge of LPS or PgLPS. There was no difference in IRAK-1 protein expression; therefore, the authors conclude that it is IRAK kinase activity that is suppressed. This is supported by a study from Nakayama et al. [43 ], who examined the mechanisms of peptidoglycan tolerance in RAW264.7 macrophages. They demonstrated that IRAK-1 kinase activity was decreased in peptidoglycan-tolerized macrophages, with no change in IRAK protein levels, suggesting that IRAK-1 was not phosphorylated in these cells. Their study also demonstrated a defect in association between IRAK-1 and MyD88 in tolerized cells.
IRAK-4 has been demonstrated to act upstream of IRAK-1 and is responsible for phosphorylation of IRAK-1 [56 ]. IRAK-4 knockout mice show almost complete impairment in response to bacterial components, which signal via TLR2, TLR4, TLR3, and TLR9 [58 ]. It therefore represents a potential target in the development of tolerance. Li et al. [44 ], however, failed to show any alteration in IRAK-4 mRNA and protein levels in THP-1 monocytes tolerized with BLP.
IRAK-M, in contrast to other IRAK family members, has been demonstrated to negatively regulate TLR signaling [59
]. It prevents dissociation of IRAK-1 and IRAK-4 from MyD88 and inhibits formation of the IRAK-TRAF-6 complex. IRAK-M knockout mice show increased production of proinflammatory cytokines after bacterial challenge. IRAK-M is up-regulated in LPS-tolerant cells [60
], and LPS tolerance is significantly impaired in IRAK-M knockout mice [59
]. Nakayama et al. [43
] sought to elucidate the relationship between IRAK-M and TLR signaling in gram-positive tolerance using RAW264.7 macrophages stimulated with peptidoglycan. Their study demonstrated that IRAK-M is not constitutively expressed in naïve cells but induced in cells pretreated with peptidoglycan. IRAK-M expression was then modified by using cells transfected with IRAK-M small interfering RNA. It was observed that in control cells tolerized to peptidoglycan, TNF- production was reduced. However, in tolerized cells with down-regulated IRAK-M, TNF- production was restored, suggesting that IRAK-M plays a critical role in peptidoglycan-induced tolerance. Alterations in expression of the IRAK proteins in gram-positive tolerance are summarized in Table 2
.
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B activation. TAK-1 phosphorylates the MAPKs and the IKK complex, a cytoplasmic inhibitor of NF-B. IKK is a specific kinase complex composed of two serine/threonine kinases, IKK and IKKß, and a scaffold protein IKK. Activation of IKK complex results in phosphorylation of IB and IBß, ubiquitination, and degradation of IB, and release of the active NF-B complex [61
]. Alterations in the IKK complex in gram-positive tolerance are summarized in Table 3
.
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In gram-positive tolerance, Nayakama et al. [43
] observed almost complete abrogation of IB phosphorylation in peptidoglycan-tolerized RAW264.7 macrophages. There was also marked inhibition of ERK1/2, p38, and JNK in tolerant compared with naïve cells. These findings were echoed in a study by Siedlar et al. [41
], who demonstrated that although IB is degraded in naïve Mono-Mac-6 cells stimulated with BLP, there is no change in IB levels, and no detectable phosphorylation of IB occurs in BLP-tolerized cells.
In THP-1 monocytes pretreated with BLP, Li et al. [44
] demonstrated that IB phosphorylation was markedly suppressed in response to BLP or LPS challenge. IB protein levels were also significantly lower in tolerized compared with naïve cells, and BLP tolerance led to inhibition of IB protein degradation. The authors suggest that one reason for the reduced IB protein levels may be a result of the inhibition of NF-B, as IB is one of the target genes of NF-B [66
]. Dobrovolskaia et al. [57
] demonstrated that IKKß kinase activity is stimulated by TLR2 ligands PgLPS and BLP, as well as LPS. In cells pretreated with PgLPS or BLP, IKKß kinase activity was inhibited significantly, but cells responded with increased IKKß kinase activity if challenged with LPS. A similar result was observed in LPS-tolerant cells. This group also looked at the degradation of the NF-B inhibitor proteins, IB, IBß, and IB
. There was reduced degradation of IB in cells pretreated with PgLPS or BLP and rechallenged with these agonists or with LPS, although to a lesser extent with the latter. In contrast, they observed a marked increase in IB in PgLPS and BLP-tolerized cells, which was maintained not only when cells were restimulated with BLP or PgLPS but also if exposed to LPS. The authors speculate that perhaps elevated IB may contribute to the suppressed NF-B activation in tolerized cells. This group also demonstrated inhibition of JNK activation in macrophages pretreated with PgLPS and restimulated with PgLPS or LPS but with some residual ERK1/2 activation following repeat stimulus.
Wang et al. [40 ] demonstrated that BLP stimulation of THP-1 monocytes induces activation of ERK1/2, JNK, and p38. When THP-1 cells were tolerized with BLP, they observed almost complete suppression of phosphorylation of all three MAPKs when cells were challenged with a second dose of BLP or LPS. Sato et al. [27 ], using MALP-2-pretreated mouse macrophages, demonstrated that JNK activation was suppressed when cells were exposed to MALP-2 or LPS. In contrast, significant JNK activation, although reduced compared with naïve cells, was observed in LPS-tolerant macrophages stimulated with MALP-2.
Phosphorylation of the MAPKs and IB is a necessary step in the signaling cascade leading to the synthesis of proinflammatory cytokines [67
68
69
]. A study by Ropert et al. [70
] investigated the role of phosphatases in tolerant cells. Having first demonstrated that mucin-like glycoprotein of Trypanosoma cruzi (tGPI-mucin), a TLR2 agonist [71
, 72
], induces self-tolerance and cross-tolerance to LPS [68
], they proceeded to demonstrate that MAPK activation, IB phosphorylation and degradation were inhibited in tGPI-mucin- and LPS-tolerant cells. They then extended this a step further by examining the effect of a phosphatase inhibitor okadaic acid on LPS-tolerant macrophages. They observed that phosphatase activity induced by bacterial stimulation coincides with macrophage hyporesponsiveness to subsequent challenge. The increase in phosphatase activity is associated with an impaired ability of macrophages to phosphorylate IRAK-1, MAPKs, and IB. They then demonstrated that the use of a phosphatase inhibitor resulted in up to 80% restoration in TNF- production following second LPS challenge. This was associated with restoration of IRAK-1, MAPK, and IB phosphorylation. Although the authors did not replicate these results using a TLR2 agonist, they did demonstrate that tGPI-mucin, signaling through TLR2, also induces phosphatase activity, similar to LPS.
NF-B
Following activation, NF-B is translocated to the nucleus, where it activates the transcription of proinflammatory genes [67
]. Suppression of NF-B activation has long been a recognized feature of LPS tolerance [73
74
75
]. The NF-B family is composed of several proteins, which can form homodimers and heterodimers. These include p50, p65, p52, RelB, and c-Rel [76
]. The p65 subunit has been demonstrated to be required for gene trans activation [77
], and the p50 subunit has no trans activation domain. Therefore, although the p50/p50 subunit can bind to DNA, it cannot trans activate. It is believed that in endotoxin tolerance, p50/p50 binding prevents p50/p65 binding and thus, activation of proinflammatory gene transcription [78
79
80
].
NF-B activation was not observed in MALP-2-tolerized mouse macrophages when challenged with a second MALP-2 or LPS stimulus by Sato et al. [27
]. However, MALP-2 stimulation of LPS-tolerant cells did result in NF-B activation, with translocation of mainly p50/p65 heterodimers. Wang et al. [40
] observed that induction of BLP tolerance in THP-1 monocytes significantly inhibited NF-B activation and NF-B DNA binding following exposure to a second BLP or LPS challenge.
Siedlar et al. [41 ] demonstrated that BLP-tolerant Mono-Mac-6 cells showed no change in p50/p50 homodimers, but mobilization of p50/p65 heterodimers was blocked after a second challenge. This was in contrast to LPS-tolerant cells, where p50/p65 mobilization remained intact, and p50/p50 homodimerization was induced.
Dobrovolskaia et al. [57
] observed that although there was suppression of NF-B DNA binding in PgLPS-tolerized peritoneal macrophages, there were still high levels of translocated p50/p65 in the nucleus. Suppression of NF-B activity was greater in homotolerance than in cross-tolerance. The authors also observed that DNA binding of another transcription factor, AP-1, remained detectable in PgLPS-tolerized cells, despite upstream inhibition of the MAPKs. This is in contrast to a study about LPS tolerance in RAW264.7 macrophages, which demonstrated decreased AP-1 DNA binding activity on secondary LPS stimulation [74
]. The accumulation of the p50/p65 subunit in the nucleus but its failure to bind DNA was also demonstrated in a recent study about LPS tolerance, where the authors observed that despite activation and translocation of NF-B p65, it failed to bind to the IL-1ß promoter in LPS-tolerized cells [81
].
Using CHO cells engineered to overexpress TLR2 and TLR4, Medvedev et al. [42
] observed the effect of mycobacterial components, LAM/STF, and LPS tolerance and cross-tolerance on NF-B activation. Induction of tolerance using LAM/STF or LPS significantly reduced NF-B activation in response to a second stimulus. LPS-tolerant cells had more marked suppression of NF-B when stimulated with LAM or STF than LAM/STF-tolerant cells subsequently challenged with LPS.
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INCREASED PHAGOCYTOSIS IN GRAM-POSITIVE TOLERANCE |
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TOPABSTRACTINTRODUCTIONGRAM-POSITIVE TOLERANCE AND...CELL SURFACE RECEPTOR...INTRACELLULAR SIGNALING PROTEIN...INCREASED PHAGOCYTOSIS IN GRAM...GRAM-POSITIVE TOLERANCE: THE...SUMMARYREFERENCES |
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III/IIR, have been demonstrated to be up-regulated in response to a sublethal, gram-positive challenge. This was the focus of a study by Wang et al. [29
], who found that induction of BLP tolerance and similarly, LPS tolerance resulted in an increased population of polymorphonuclear neutrophils (PMNs) in the circulation and peritoneal cavity. However, in BLP tolerance, this was associated with a significant up-regulation in CR3 and FcIII/IIR expression on PMNs and peritoneal macrophages, and this was not seen in LPS-tolerant cells. Subsequently, increased bacterial recognition, uptake, and intracellular killing, compared with naïve cells or LPS-tolerant cells, were also observed. A further study by O’Brien et al. [32
] looked at BLP-induced cross-tolerance to gram-negative sepsis in TLR4-deficient mice. This study demonstrated that BLP pretreatment of TLR4-deficient mice results in up-regulation of phagocytic receptors CR3 and FcIII/IIR on PMNs and peritoneal macrophages, enhancing the phagocytic activity of these cells when exposedto lethal, gram-negative challenge and thus conferring a significant survival advantage over naïve mice. A study by Feterowski et al. [82
] demonstrated significantly lower numbers of viable bacteria in peritoneal lavage fluid from mice tolerized with MALP-2 or BLP, compared with naïve mice, in a model of polymicrobial peritonitis. This was associated with increased neutrophil accumulation in the peritoneal cavity of tolerized mice. In vitro, an increase in the production of reactive oxygen metabolites was demonstrated in neutrophils from septic compared with nonseptic mice. However, the production of reactive oxygen metabolites was equivalent in MALP-2-tolerized and naïve neutrophils. Although this suggests that perhaps neutrophil accumulation and activation are important factors in the enhanced bacterial clearance seen in gram-positive tolerance, O’Brien et al. [32
] demonstrated that BLP pretreatment resulted in a statistically significant increase in bacterial clearance from the blood, lungs, and spleen in neutropenic mice. These studies advance the findings of an early study by Ausobsky et al. [28
], which demonstrated that tolerance to MDP was associated with a hyperphagocytic state. Although Wang et al. [29
] did not demonstrate that LPS tolerance enhances intracellular killing of bacteria, increases in circulating neutrophil, macrophage activity, and bacterial clearance have been demonstrated in LPS tolerance [83
84
85
]. The exact mechanisms of improved bacterial clearance in tolerant animals have not been explained definitively. From the work published to date, there appears to be an up-regulation of phagocytic function in animals pre-exposed to sublethal infection, which subsequently enables rapid uptake of bacteria following a second lethal septic challenge. |
GRAM-POSITIVE TOLERANCE: THE ROLE OF IL-10 |
|---|
, IL-6, and IL-1ß. It has also been demonstrated to down-regulate TLR4 surface expression and to enhance phagocytic receptor expression [86
]. Several groups have looked for a potential role for IL-10 in gram-negative and gram-positive tolerance. The role of IL-10 as a key regulator in LPS tolerance has been refuted, and studies demonstrate that LPS tolerance can be induced in IL-10-deficient mice [84
, 87
, 88
]. A regulatory role for IL-10 in gram-positive tolerance is similarly unlikely, and Sato et al. [27
] used IL-10-deficient mice to demonstrate that MALP-2 tolerance and cross-tolerance to LPS are independent of IL-10, which has been shown to increase FcR expression [86
], raising the question of whether it is an increase in IL-10 in tolerized cells that augments bacterial clearance in these cells. Varma et al. [84
] demonstrated that bacterial clearance was in fact further enhanced in IL-10-deficient, LPS-tolerized mice compared with wild-type controls. Moore et al. [86
], in their recent review, found that IL-10 inhibits intracellular killing of bacteria in neutrophils, a phenomenon that was reversed with neutralization of endogenous IL-10. Although bacterial uptake and intracellular killing in IL-10-deficient mice have not been studied specifically in a model of gram-positive tolerance, the findings of Varma et al. [84
] suggest that IL-10 has, in fact, a negative impact on bacterial clearance in tolerized cells. |
SUMMARY |
|---|
|
III/IIR and CR3, is associated with enhanced bacterial clearance in tolerized mice exposed to a second septic challenge, and this perhaps represents one of the mechanisms by which tolerized cells do not succumb to lethal bacterial challenge. It has been demonstrated that BLP-tolerant cells have decreased association between MyD88 and IRAK-1. Decreased IRAK-1 protein expression and decreased IRAK-1 kinase activity, without corresponding change in IRAK-1-mRNA, are features of gram-positive tolerance. In addition, IRAK-M, a negative regulator of IRAK-1, is up-regulated in peptidoglycan-tolerized cells. Thus, the IRAK family of proteins represents an attractive target for further research and potentially, therapeutic modification.
Downstream of IRAK, decreased phosphorylation and degradation of IB have been demonstrated in different gram-positive tolerance models, as has a decrease in IKKß kinase activity and an increase in IB protein in tolerized cells. Suppression of MAPK activation is also a feature of gram-positive tolerance.
NF-B activity is down-regulated in gram-positive tolerance. Evidence suggests that mobilization of the p50/p65 heterodimer is blocked, or its binding to promoter regions of DNA is somehow inhibited. Transcription factor AP-1, a downstream target of MAPK signaling, is also inhibited in gram-positive tolerance.
Gram-positive bacterial components have been demonstrated, not only to induce self-tolerance but also cross-tolerance. The current evidence suggests that cross-tolerance to LPS in gram-positive-tolerized cells is stronger than cross-tolerance in LPS-tolerant cells.
Considerable progress has been made in recent times toward understanding exactly how a cell responds to microbial challenge and how the induction of tolerance in a cell alters this response. Multiple changes in intracellular signaling have been identified in gram-positive and gram-negative tolerance (Fig. 2 ). As yet, the mechanism and significance of cross-tolerance remain to be explained, as does the fundamental question: As we unravel the molecular complexities of tolerance, how does this translate into therapeutic targets in critically ill patients?
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Received May 9, 2006; revised July 7, 2006; accepted July 16, 2006.
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