key: cord-017361-2lrmg6z0
authors: Ballinger, Megan N.; Standiford, Theodore J.
title: Innate Immune Responses in Ventilator-Associated Pneumonia
date: 2012-10-26
journal: Mucosal Immunology of Acute Bacterial Pneumonia
DOI: 10.1007/978-1-4614-5326-0_8
sha: 
doc_id: 17361
cord_uid: 2lrmg6z0

Ventilator-associated pneumonia (VAP) is a common complication of mechanical ventilation, resulting in substantial morbidity, mortality, and health care cost. Early upper airway colonization by pathogenic bacteria and microaspiration are the primary pathogenic events leading to VAP. Patients at risk for VAP have defects in structural/mechanical defenses of the respiratory tract. In addition, critical illness, including sepsis, trauma, and postoperative states, is associated with profound defects in both innate and acquired antibacterial immunity, influencing antimicrobial effector functions of both leukocytes and structural/parenchymal cells. Factors present within the lung microenvironment, including alveolar stretch, cyclical atelectasis, changes in oxygen tension, and respiratory tract microbiota, substantially impact antibacterial host responses. Mechanisms accounting for dysregulated immune homeostasis are incompletely understood, but likely involve: (1) alterations in the balance of pro- and anti-inflammatory cytokines; (2) changes in pathogen recognition receptor and G-protein coupled receptor expression and downstream signaling cascades; and (3) dysregulated cell death responses. Antibiotics and preventive strategies are the mainstay of therapy in patients with VAP. However, novel approaches are needed to reverse immunological reprogramming that occurs during critical illness and/or mechanical ventilation, and to identify patients who are most likely to benefit from immunomodulatory therapy.

The most common cause of VAP is by the Gram-positive bacteria Staphylococcus aureus , with methicillin resistant S. aureus (MRSA) representing over 60% of the S. aureus isolates in VAP. Other VAP-causing pathogens include aerobic Gramnegative bacilli such as Pseudomonas aeruginosa , Klebsiella pneumoniae , Escherichia coli , Enterobacter species, Acinetobacter species, and Stenotrophomonas maltiphila . Legionella pneumophila is an obligate intracellular bacterial pathogen that is an etiologic agent in both community acquired pneumonia (CAP), HAP and VAP. Viruses and fungi are unusual causes of VAP, although these organisms can modulate innate mucosal responses predisposing to the development of VAP. While the bacterial pathogens that cause VAP are similar to those that cause HAP in nonintubated patients, VAP is more frequently caused by pathogens with intrinsic resistance to multiple antimicrobial agents, including P. aeruginosa , Acinetobacter species, S. maltiphila , and MRSA. Mortality is considerably higher in patients with VAP due to P. aeruginosa strains that express the type III secretion system required for the secretion of pseudomonal exotoxins S, T, U, and Y (Roy-Burman et al. 2001 ; Sadikot et al. 2005 ) . A recent and disturbing trend is the increasing prevalence of community acquired stains of MRSA (CA-MRSA) as a cause of nosocomial infections, including VAP (Kashuk et al. 2010 ) . CA-MRSA, which is typically the USA300 strain, produce an array of exotoxins that promote extensive tissue necrosis and cavity formation. The intrinsic antibiotic resistance of these Gram-positive and Gram-negative bacterial strains contributes to increased mortality in patients with VAP (Fujitani et al. 2011 ) . However, these pathogens are generally less virulent and invasive than pathogens that cause pneumonia in otherwise healthy individuals in the community, and tend to be invasive in hosts with anatomic defects in the respiratory tract or substantial impairment in lung mucosal innate immunity. Therefore, the presence of these bacterial species as pathogens identi fi es patients with profound anatomic defects or defects in lung innate immunity.

The vast majority of VAP cases develop as a result of microaspiration of bacteria colonizing the oropharynx (American Thoracic Society; Infectious Diseases Society of America 2005 ) . Oropharyngeal colonization occurs very rapidly in critically ill patients. For example, nearly 75% of patients with underlying lung disease and/or undergoing oropharyngeal intubation were found to be colonized by pathogenic bacteria within 24 h of admission to the intensive care unit (Garrouste-Orgeas et al. 1997 ) . Reservoirs contributing to oropharyngeal colonization include the nasopharynx, sinuses, and stomach. Endotracheal tubes contribute to colonization by directly injuring mucosal surfaces of the upper respiratory tract, which facilitates bacterial adhesion. Organisms that cause VAP, including P. aeruginosa and S. aureus , promote bio fi lm formation with the endotracheal tube lumen, which can function as a nidus for direct inoculation of infected material into the distal airspaces. Less common sources of bacterial inoculation include colonization of the ventilator circuit or direct inoculation via infected aerosols or instrumentation, particularly suction catheters or bronchoscopes. By comparison, hematogenous seeding of the lung as a cause of VAP is considerably less common, accounting for <15% of cases. Notable exceptions are hematogenous seeding from an intravascular S. aureus infection or gut bacterial translocation that can occur in immunocompromised patients with neutropenia.

Microaspiration is a common event in both healthy and critical ill patients. These events rarely result in infection in healthy subjects, primarily due to highly effective means to eradicate infectious or toxic insults of the respiratory tract, which include ef fi cient mucocilliary clearance mechanisms and robust innate mucosal antimicrobial responses. In mechanically ventilated patients, impairments in both mucocilliary transport and innate cellular responses results in the establishment of pulmonary infection. A summary of factors contributing to the pathogenesis of VAP is shown in Fig. 8 .1 .

Ciliated, pseudostrati fi ed columnar epithelial cells line the tracheobronchial tree. These ciliated cells are critical to effective mucocilliary transport and the cephalad movement of mucous, microbes, and acellular debris present within the conducting airways. Damage to ciliated cells can occur as a direct result of endotracheal intubation The end result from a combination of host factors, medication, and instrumentation is the introduction of infectious material into the sterile lung environment. These factors along with the immune state of host, contribute to the development of VAP or conditions that predispose the patient to respiratory failure (Nicholls et al. 2003 ; Piatti et al. 2005 ; Pittet et al. 2010 ) . As discussed previously, denuding of columnar epithelial cells can result from the endotracheal tube or endotracheal tube cuff. Moreover, lung conditions that can result in mechanical ventilation, such as COPD, are associated with impaired mucocilliary transport (Piatti et al. 2005 ) . Moreover, certain forms of infectious lung injury, including severe acute respiratory syndrome (SARS) is characterized by bronchial epithelial denudation and loss of cilia (Nicholls et al. 2003 ) . Similarly, in fl uenza infection predisposes to secondary bacterial infection, which is due not only to impairment in lung innate responses, but also disruption of mucocilliary transport mechanisms (Pittet et al. 2010 ) .

Many forms of critical illness result in a profound state of immune suppression affecting both the cellular and acquired arms of host immunity. This syndrome of immune suppression has been best characterized and is perhaps most severe in sepsis, but has also been described in trauma patients, burn injury patients, and patients during the peri-operative period. Sepsis is a complex clinical syndrome resulting from the interaction between microbe and host. Clinically, it is de fi ned as the systemic in fl ammatory response syndrome (SIRS) with evidence of infection (Members of the American College of Chest Physicians/Society of Critical Care Medicine 2003 ) . Changes in the population at risk for the development of sepsis, including an increase in the number of elderly and immunocompromised patients, has resulted in a steady rise in the incidence of severe sepsis (Martin et al. 2003 ) . Despite improvements in supportive care and immunomodulatory therapies, the mortality rate from severe sepsis remains unacceptably high (Brun-Buisson 2000 ) . Host immune responses in critical illness, including sepsis can be conceptualized as occurring in distinct but overlapping phases. The initial response during critical illness, referred to as the systemic in fl ammatory response syndrome (SIRS), is characterized by the release of a number of pro-in fl ammatory mediators, including early responses cytokines such as tumor necrosis factor-alpha (TNF-a ), interleukin-1 b (IL-1 b ), interleukin-6 (IL-6), interleukin 12 (IL-12) leukocyte-active chemokines, adhesion molecules, and in fl ammatory leukotrienes (Dinarello 2000 ) . SIRS is counter-regulated by the release of inhibitory molecules, including anti-in fl ammatory cytokines (e.g., interleukin 10 (IL-10), transforming growth factor-beta (TGF-b )), suppressors of pathogen recognition signaling cascades, immunomodulatory prostanoids and hormones. This counter-regulatory phase is referred to as the compensatory anti-in fl ammatory response syndrome (CARS) (Wesche et al. 1999 ; Bone 1996 ) . Molecules released during CARS are believed to serve as a functional "brake" on systemic in fl ammation, and the expression of these mediators is induced by both microbial-derived and host-derived signals. SIRS and CARS overlap considerably, hence the overall immune status of the patient is dependent on which response predominates ( Fig. 8 .2 ) (van der Poll and van Deventer 1999 ) . Recent evidence suggests a third response to an in fl ammatory insult, referred to as the mixed antagonist response syndrome (MARS). This response is characterized by the secretion of both pro-and anti-in fl ammatory mediators (speci fi cally IL-6, IL-8, monocyte chemotactic protein (MCP)-1, macrophage in fl ammatory protein (MIP)-1 b , IFN-g , granulocyte-macrophage colony stimulating factor (GM-CSF), and IL-10) (Tamayo et al. 2011 ) . Consistent with this mixed systemic cytokine response, elevated levels of IL-6 in circulation has been shown to predict the development of VAP (Ramirez et al. 2009 ) . Whether the initial SIRS response drives the expression of molecules that contribute to immune suppression or simply a marker of systemic in fl ammation remains to be determined. A summary of innate immune events in critical illness is shown in Fig. 8 .2 .

The compensatory release of anti-in fl ammatory molecules in sepsis is believed to mediate immunosuppression during the peri-septic or post-injury period, during which time immune cell function is substantially impaired (historically referred to as critical illness-induced leukocyte "deactivation" or "immunoparalysis"). Recently, since the altered leukocyte phenotype in critical illness involves selective regulation of some, but not all innate genes, this phenomenon is now more appropriated referred to as reprogramming. Leukocyte reprogramming appears to be of considerable clinical signi fi cance, as higher rates of nosocomial infection and increased mortality are observed in postoperative, burn injury or septic patients who display evidence of monocyte deactivation, either in the form of decreased monocyte HLA-DR expression, ex vivo cytokine production or impaired delayed-type hypersensitivity responses (Appel et al. 1989 ; Munoz et al. 1991 ) . Septic patients are especially susceptible to nosocomial infections of the lung, particularly pneumonia from multidrug-resistant Gram-positive and Gram-negative organisms, including S. aureus and P. aeruginosa (Richardson et al. 1982 ; Mustard et al. 1991 ) . Sepsis-induced immunosuppression is particularly prominent in patients with preexisting de fi ciencies in innate and acquired immunity, including the elderly and patients with chronic medical conditions (Hotchkiss and Karl 2003 ) .

Patients undergoing severe stress, including trauma, massive hemorrhage, burn injury, post-surgery, and sepsis exhibit signi fi cant defects in circulating and resident leukocyte populations. In addition, changes in the pulmonary microenvironment that occur as a result of mechanical ventilation substantially in fl uence lung innate responses. Multiple leukocyte subtypes are affected and speci fi c defects are shown in Fig. 8 .3 .

While sepsis and similar stress-associated events have been shown to in fl uence the effector activity of a variety of immune cells, the majority of studies have focused on peripheral blood monocytes (PBM), and to a lesser extent tissue macrophages. Changes in monocyte/macrophage function in sepsis resemble but are not identical to those observed in endotoxin-tolerized macrophages. Endotoxin tolerance describes the phenomena whereby upon initial exposure to LPS, cells become refractory to a secondary stimulus with LPS. Pathogen-associated molecular patterns (PAMPs) other than LPS can also induce a tolerance phenotype, and PAMPs of one type can induce cross tolerance to a different PAMP. Induction of tolerance results in suppression of multiple in fl ammatory genes, including both NF-k B and mitogen-activated protein kinase (MAPK)-dependent genes (e.g., TNF-a , IL-6, iNOS). Tolerance does not cause global suppression of all genes, as genes encoding certain antimicrobial and phagocytic proteins, including cathelicidin antimicrobial peptide, lipocalin, the scavenger receptor MARCO and the fMLP receptor, are indeed super-induced in response to sequential exposure to LPS (Foster et al. 2007 ) . It is also noteworthy that the induction of this phenotype is not restricted to myeloid cells, as structural cells, including alveolar epithelial cells, have been shown to develop a tolerance response upon repeated exposure to PAMPs. The LPS or PAMP tolerized phenotype is transient in nature and entirely reversible, and has been associated with remodeling of chromatin in the promoter region of several tolerizable genes (Foster et al. 2007 ; Chan et al. 2005 ) . Critical illness, like endotoxin tolerance, leads to inhibition of a broad range of NF-k B-dependent in fl ammatory genes in monocytes. Most notably, a signi fi cant reduction in the ex vivo production of in fl ammatory cytokines, including IL-1 a , IL-1 b , IL-6, and TNF-a has been observed in monocytes isolated from patients with sepsis (Munoz et al. 1991 ) . This change in cytokine production may be a predictor of outcome, as peripheral monocytes isolated from those who survived sepsis regained their ability to produce cytokines in response to LPS stimulation, and There are a variety of cellular, bacterial, and mechanical mediators which contribute to the impaired innate and acquired immune responses during critical illness. ( Upward arrow ) represent effects that enhance expression/function ( downward arrow ) represents effects that reduce expression/function monocytes isolated from the nonsurvivors did not (Munoz et al. 1991 ) . Conversely, the production of certain anti-in fl ammatory proteins, including IL-10, IL-1 receptor antagonist, and the TNF soluble receptor I and II are enhanced in monocytes isolated from sepsis patients or patients with ventilator-induced lung injury (Frank et al. 2006 ) . Patients with sepsis or early trauma have reduced monocyte HLA-DR expression (Appel et al. 1989 ; Adib-Conquy et al. 2006 ) . This reduction in HLA-DR expression has been reported to directly correlate with the magnitude of sepsis (Volk et al. 2000 ) and may partially contribute to impaired cell-mediated immunity observed in patients with critical illness.

Similar critical illness-induced defects have been noted in macrophages residing in various tissues, which in some instances have been associated with evidence of enhanced macrophage apoptosis (Ayala et al. 1992 ; Gallinaro et al. 1994 ) . In particular, alveolar macrophage function has been shown to be impaired in the setting of sepsis. For example, alveolar macrophages recovered from mice with abdominal sepsis (cecal ligation and puncture) display reduced production of in fl ammatory cytokines, chemokines, eicosanoids, nitric oxide, and respiratory burst (Reddy et al. 2001 ; Goya et al. 1992 ) . Importantly, these phenotypic alterations in alveolar macrophage effector function are associated with a markedly enhanced susceptibility to intrapulmonary challenge with both Gram-positive and Gram-negative bacterial pathogens (Steinhauser et al. 1999 ; Deng et al. 2006 ) . Little is known about alveolar macrophage phenotype in critically ill patients at risk for the development of VAP. However, we have performed Affymetrix microarray analysis on adherence puri fi ed alveolar macrophages recovered from patients with sepsis-induced ALI within 3 days of onset of sepsis. Relative to alveolar macrophages recovered from healthy subjects, lung macrophages from sepsis-induced ALI patients displayed a hybrid tolerized/alternatively activated phenotype, as characterized by minimal change or suppression of NF-k B-dependent genes (e.g., TNF-a , IL-1 b , IL-6, iNOS), induction of antimicrobial genes (antimicrobial peptides, chemoattractant, and phagocytosis genes), and expression of makers of alternative (M2) rather than classical (M1) activation (high arginase, CCR2, IL-4R a , MMP expression; low iNOS, interferong , and IFN-inducible chemokine expression) (Gordon and Martinez 2010 ) . Although this expression pattern may partially re fl ect the lung injury response, it is likely that the phenotype is shaped by systemic in fl ammation.

Alterations in neutrophils (PMN), resembling those described in monocyte/ macrophages, are present during the septic response and are predictive of adverse outcomes in these patients. Systemic in fl ammation promotes cytoskeletal changes in PMN cell membrane rigidity and reduced cellular deformability, resulting in impaired recruitment to sites of infection and deleterious accumulation and activation of PMN in vascular beds of distant organs. Directed migration is also impaired by nitric oxide-mediated inhibition of ICAM and VCAM-dependent adhesion and transmigration of PMN, downregulation of the chemokine receptor CXCR2, and inhibition of G-protein coupled receptor signaling (Benjamim et al. 2000 ; Cummings et al. 1999 ; Czermak et al. 1999 ; Huber-Lang et al. 2001 ; Swartz et al. 2000 ) . Microarray analysis of PMN isolated from septic patients within 24 h of onset reveals a global suppression of immune regulation and in fl ammatory response gene clusters, particularly genes regulated in an NF-k B-dependent fashion (Tang et al. 2007 ) . Conversely, the expression of selected suppressive genes was enhanced, including the NF-k B inhibitor NF k BIA.

The discovery of neutrophil extracellular traps (NETs) has provided yet another role for neutrophils in the containment of infection. NETs are complex structures composed of nuclear chromatin, histones, a variety of granular antimicrobial proteins and some cytoplasmic proteins (Urban et al. 2009 ) . Formation occurs in response to exposure of neutrophils to plasma from septic patients (Clark et al. 2007 ) as well as direct contact with microbial pathogens (Remijsen et al. 2011 ) . Neutrophil elastase is released from azurophilic granules, assisting in the formation of NETs via decondensation of nuclear chromatin, which along with other serine proteases confer antimicrobial responses (Papayannopoulos et al. 2010 ) . NETassociated myeloperoxidase directly contributes to bacterial killing of Staphylococcus aureus in the presence of H 2 O 2 (Parker et al. 2012 ) . NETs are capable of physically ensnaring bacteria and facilitating the interactions between bacteria and antimicrobial effectors, ultimately leading to enhanced bacterial killing (Mantovani et al. 2011 ) . Despite their broad antimicrobial capacity, some bacteria express nucleases to degrade NETs, thus avoiding capture and bacterial cell death (Buchanan et al. 2006 ; Berends et al. 2010 ; Young et al. 2011 ) . In some cases, NETs may exert detrimental effects to the host. Increasing evidence links NET formation to excessive in fl ammation and tissue damage in diseases such as sepsis (Clark et al. 2007 ) . NET formation has recently been demonstrated in the alveoli of mice with in fl uenza H1N1 pneumonia, and these structures contribute to acute lung injury responses in these animals (Narasaraju et al. 2011 ) . While the presence of NETs has not been clearly established in experimental bacterial pneumonia or in patients with VAP, it is tempting to speculate that these structures may contribute to lung injury that can occur in this setting.

Dendritic cells (DC) are the most ef fi cient professional antigen-presenting cells (APC) in the lung and have the unique ability to induce primary immune responses in naïve T cells. DC are prevalent centrally within the spleen, lymphatics, and at mucosal surfaces, most notably in gut and respiratory tract. Systemic endotoxin administration in mice results in a brisk depletion in splenic DC by 24 h post-LPS. Similarly, there is a prolonged loss of DC out to 15 days post-induction of abdominal sepsis in both lung and spleen (Wen et al. 2008 ) . In humans with lethal sepsis, follicular DC are substantially diminished early in the course of disease (Hotchkiss et al. 2002 ) .

Similarly, reductions in blood myeloid DC and plasmacytoid DC (27 and 53% of controls, respectively) have been observed in patients admitted to the hospital with pneumonia, and numbers of DC inversely correlated with procalcitonin levels, a marker of systemic in fl ammation (Dreschler et al. 2012 ) . Endotoxin-tolerized DC or DC isolated from animals or humans with sepsis produce low levels of IL-12 and TNF-a , but high levels of IL-10 (Wen et al. 2008 ; Wysocka et al. 2001 ) . This shift in cytokine pro fi les can persist for up to 6 weeks post-abdominal sepsis (CLP), and has been associated with posttranslational epigenetic modi fi cations of histones binding to the IL-12 p35 and p40 promoters and increased susceptibility to pulmonary fungal challenge (Wen et al. 2008 ) . Regulatory DC, or "tolerogenic" DC, are a newly described DC population that can be induced by incubation of bone marrow-derived DC with IL-10, resulting in DC that preferentially secrete IL-10 rather than IL-12, and induce T cell tolerance. A naturally occurring DC reg population has been identi fi ed in spleen (CD11c low , CD45RB high ), and adoptive transfer of this cell population to septic mice diminished in fl ammatory cytokine production and sepsis-induced lethality (Fujita et al. 2006 ) . Changes in the number, distribution, and function of these cells in lung, especially during critical illness, have not yet been explored.

Like other leukocyte populations, various lymphocyte populations are in fl uenced by and likely contribute to the immunosuppressive effects of critical illness. This effect can be directly due to changes in lymphocytes numbers or effector functions, or indirectly due to changes in APC function, most notably DC. Studies consistently show that sepsis or other states of extreme stress (trauma, burn injury) generally result in anergy and a shift in T cell cytokine responses favoring a Th2-, rather than Th1-phenotype response.

Sepsis, trauma, and other critical states result in a substantial drop in the number of circulating lymphocytes. Lymphopenia develops early after the insult, and the persistence and magnitude of lymphopenia correlates with risk of nosocomial infection and death (Hotchkiss et al. 2001 ) . Autopsy studies in septic patients revealed a profound loss of splenic B cells, CD4+ T cells, and follicular dendritic cells. No alterations in numbers of CD8+ T cells were observed. The loss of B and CD4+ T cells was mediated by caspase-9-dependent apoptosis. Similar changes, although not as uniform, could be observed in critically ill patients without sepsis (Hotchkiss et al. 2001 ) .

In addition to changes in the absolute number of lymphocytes, the septic response can induce considerable alterations in lymphocyte effector function. For instance, the memory/effector CD8+/CD45RO+ T lymphocyte subset in nonsurviving septic patients demonstrate signi fi cantly decreased IFN-g synthesis compared with survivors (Zedler et al. 1999 ) . Similarly, T cell proliferative responses and cytokine production (IL-2, TNF-a ) were signi fi cantly depressed in patients with abdominal sepsis, as compared to healthy controls, and the degree of IL-2 and TNF-a suppression directly correlated with patient survival (Heidecke et al. 1999 ) . The proportion of Th2 T cells is increased in patients with sepsis, but not in non-septic critically ill control patients and healthy subjects (Ferguson et al. 1999 ) . Similar observations have been made in animal models of sepsis. Splenocytes isolated from mice undergoing CLP produced less IL-2, IL-12, and IFN-g , and more IL-4 and IL-10 than splenocytes isolated from healthy animals (Ayala et al. 1994 ; O'Sullivan et al. 1995 ) . Given the importance of Th1 phenotype responses in host defense against both intracellular and extracellular microbial pathogens, this shift away from an appropriate Th1-and towards a dysregulated Th2-phenotype response has obvious implications for antimicrobial host immunity.

Regulatory T cells (Treg), are a limited but important population of CD4+, CD25+ T cells that universally express the transcription factor Forkhead box p3 (Foxp3). Treg inhibit CD4+ and CD8+ T cell effector functions, resulting in negative regulation of both innate and acquired immune responses. Suppressive effects of Treg are mediated by both direct cell-cell contact and through the release of soluble mediators, including but not limited to TGF-b and IL-10. An increase in the percentage (but not absolute number) of Treg has been found in blood, lymphatics, or spleen in septic mice and humans with sepsis or trauma (Venet et al. 2008 ; Scumpia et al. 2006 ; Wisnoski et al. 2007 ) . Moreover, there is evidence of enhanced Foxp3 expression and suppressive function of Treg in mice with abdominal sepsis, and adoptive transfer of Treg into septic mice reduced overzealous TNF-a production and improved mortality. However, the depletion of CD4+ CD25+ Treg in mice with polymicrobial sepsis had little impact on sepsis-induced mortality (Scumpia et al. 2006 ; Wisnoski et al. 2007 ) . Thus, the role of Treg in controlling the systemic in fl ammatory response, or as a mediator of impaired innate and acquired immunity in critically ill patients at risk for VAP, is uncertain and requires further study.

A recently described B cell may play a critical role in innate responses during localized and systemic infection (Rauch et al. 2012 ) . Innate response activator B (IRA-B) cells are a population of CD19+, B220+ cells that produce large quantities of GM-CSF during infection. This population expands in bone marrow and spleen in response to systemic LPS administration or abdominal sepsis, and the genetic deletion of these cells resulted in marked reduction of systemic cytokine responses, GM-CSF expression, and the ability to clear abdominal polymicrobial infection.

Microbes and microbial components that initiate the septic response are recognized by both cell surface and intracellular pathogen recognition receptors (PRR), including Toll-like receptors (TLRs) and nucleotide-binding oligomerization domain (NOD)like receptors (NLR). Toll-like receptors are a family of evolutionarily conserved type I transmembrane receptors that respond to PAMPs expressed by a diverse group of infectious microorganisms, resulting in activation of the host's immune system (Aderem and Ulevitch 2000 ; Akira and Hemmi 2003 ) . There exist 13 distinct TLRs (10 in humans and 13 in mice) that have in common an extracellular domain with leucine rich repeats and an intracytoplasmic domain shared with the IL-1 receptor (IL-1R). Binding of ligands to TLRs initiates a signaling cascade involving myeloid differentiation marker 88 (MyD88), IL-1R-associated kinases (IRAK 1 and 4), and TNFR-associated factor 6 (TRAF6), resulting in NF-k B translocation and MAPK activation, culminating in expression of genes involved in antimicrobial host defense (Aderem and Ulevitch 2000 ; Akira and Hemmi 2003 ) . In addition, certain TLRs, such as TLR2, TLR3, and TLR4 can initiated a MyD88-independent signaling cascade that requires the adaptor proteins Toll-IL-1 receptor domain containing adaptor protein inducing interferon (TRIF) and TRIF-related adaptor molecule (TRAM), resulting in the expression of interferon responsive genes. The most relevant TLRs in lung antibacterial host defense include TLR2, which recognizes speci fi c components of Grampositive bacteria and fungi; TLR4, which is the major receptor for LPS; TLR5, which recognizes and is activated by bacterial fl agellin; and TLR9, which is activated by unmethylated CpG motifs present in microbial but not mammalian DNA. In addition to PAMPs, TLRs can be activated by host-derived danger signals, referred to as damage-associated molecular patterns (DAMPs) or alarmins, and include heat shock proteins and matrix components (Ohashi et al. 2000 ) . Also, high-mobility group box 1 protein (HMGB1) is a molecule released during the septic response that has recently been shown to activate TLR2 and TLR4 (Park et al. 2004 ) . This is of particular relevance in the setting of sepsis and acute lung injury. Multiple TLRs participate in lung host immunity against Gram-negative bacteria. For example, TLR4 recognizes the lipid A moiety of LPS, and is the major TLR mediating early innate responses and clearance of non-fl agellated Gram-negative organisms that cause VAP, including K. pneumoniae , H. in fl uenza, and E. coli (Schurr et al. 2005 ; Bhan et al. 2010 ; Wieland et al. 2005 ) . In addition, mice de fi cient in TLR9 display impaired dendritic cell-mediated responses during experimental Klebsiella or Legionella pneumonia, culminating in reduced lung bacterial clearance and decreased survival (Bhan et al. 2007 (Bhan et al. , 2008 . Innate responses to the fl agellated extracellular bacteria P. aeruginosa are mediated by several MyD88dependent TLRs, predominantly TLR4 and TLR5 (Hajjar et al. 2005 ; Ramphal et al. 2008 ; Skerrett et al. 2004 ) . Interestingly, both bone marrow-derived and stromal cells contribute to MyD88-dependent innate responses to P. aeruginosa in the lung (Hajjar et al. 2005 ) .

Toll-like receptors appear to play a lesser role in host defense against S. aureus . For example, while TLR2 has been shown to mediate in fl ammatory responses to the staphylococcal toxin Panton-Valentine Leukocidin, neither TLR2, TLR4, nor MyD88 is required for effective anti-staphylococcal host immunity during respiratory infection (Skerrett et al. 2004 ; Zivkovic et al. 2011 ) . The nucleotide-binding oligodimerization domain (NOD)-like receptors (NLR) NOD1 and NOD2, which recognize the peptidoglycan component muramyl dipeptide (MDP), have been shown to be important in in fl ammatory cytokine release and bacterial eradication in a murine S. aureus skin infection model (Hruz et al. 2009 ; Inohara et al. 2005 ) . More recently, mice de fi cient in RIP2, the shared NOD1/2 adaptor molecule, are considerably more susceptible to intrapulmonary challenge with S. aureus than wild-type mice, an effect which is dependent on downstream activation of in fl ammasomecaspase-1-dependent IL-1 b release (unpublished observations, J. Deng). These later observations suggest that NLRs, rather than TLRs, may be the predominant contributors to anti-staphylococcal immunity in the lung

Signaling Cascades

Some, but not all studies have identi fi ed changes in the cell surface expression of various TLRs during the septic response. In particular, either enhanced or reduced cell surface expression of TLR2 and TLR4 have been described in monocytes from sepsis patients and in tissue macrophages during experimental sepsis (Deng et al. 2006 ; Brunialti et al. 2006 ) . Moreover, changes in monocyte cell surface expression of LPS binding partners MD2, CD14, and CD71 have also been observed in sepsis (Brunialti et al. 2006 ; Wolfs et al. 2008 ; Williams et al. 1998 ) . Disparate fi ndings are likely attributable to temporal differences in assessment of TLR expression and the heterogeneity of patient populations studied and animal models employed. The extracellular domains of certain TLRs can be shed from activated macrophages, and serve as sinks to bind extracellular PAMPs, and as a consequence dampen TLRmediated signal transduction. For instance, soluble TLR2 (sTLR2) is released by human peripheral blood monocytes (PBM) and diminishes the cellular response to the TLR2 agonist Pam3Cys without affecting cellular responses to LPS (LeBouder et al. 2003 ) . Both naturally occurring and recombinant soluble TLR4 have been shown to diminish responses to LPS (Iwami et al. 2000 ; Hyakushima et al. 2004 ) . The contribution of soluble TLR2 and TLR4 to impaired innate responses during critical illness remains to be determined. Illuminating the importance of TLRs in lung innate immunity during critical illness, combined loss of function polymorphisms in both TLR4 and the TLR4 adaptor TIRAP/Mal, or a homozygous TIRAP/Mal polymorphism have been causally linked to reduced circulating in fl ammatory cytokine levels, reduced ex vivo monocyte cytokine expression, and increased risk for serious postoperative infections, including VAP (Ferwerda et al. 2009 ) .

Molecules have been identi fi ed that inhibit TLR signaling at multiple sites downstream of the receptor. Interleukin-1 receptor-associated kinase (IRAK)-1 and -4 are key kinases necessary for both MyD88-dependent and IL-1 receptor-mediated signal transduction. Consequently, interruption of IRAK-1 and -4 phosphorylation or traf fi cking can have profound effects on the downstream expression of both NF-k B and MAPK-dependent in fl ammatory or antimicrobial genes. Interleukin-1 receptor-associated kinase-M (IRAK-M), also named IRAK-3, is a member of the IRAK family. However, IRAK-M differs from IRAK-1 and IRAK-4 in that this protein lacks kinase activity and IRAK-M has been shown to be a negative regulator of TLR signaling by blocking the disassociation of IRAK-1 from the Toll-IL-1 signaling domain. Bone marrow-derived or lung macrophages lacking IRAK-M display enhanced MAPK kinase activation and in fl ammatory cytokine production in response to TLR agonists and live bacteria (Wesche et al. 1999 ; Kobayashi et al. 2002 ) . Importantly, IRAK-M is induced by endotoxin, the NOD-2 ligand muramyl dipeptide (MDP), and other PAMPs and is required for the development of tolerance to endotoxin and peptidoglycan (Kobayashi et al. 2002 ; Hedl et al. 2007 ; Nakayama et al. 2004 ) . We have found that IRAK-M is upregulated in alveolar macrophages during experimental sepsis in a MyD88-dependent fashion, and mediates both the suppression of macrophage cytokine responses and impaired lung clearance of P. aeruginosa in septic mice (Deng et al. 2006 ; Lyn-Kew et al. 2010 ) . IRAK-M has also been shown to suppress TLR-mediated responses in murine primary alveolar epithelial cells (Seki et al. 2010 ) . Emerging data suggests that IRAK-M may be a major mediator and perhaps a biomarker for severity of disease in sepsis. IRAK-M is substantially induced in monocytes from healthy subjects administered LPS intravenously (van't Veer et al. 2007 ) . In patients with Gramnegative sepsis, blood monocytes demonstrate a more rapid and robust expression of IRAK-M when stimulated ex vivo with LPS (Escoll et al. 2003 ) . Additionally, enhanced expression of IRAK-M mRNA has been noted in pediatric patients with sepsis, and high IRAK-M mRNA levels were associated with longer length of intensive care unit (ICU) stay, need for mechanical ventilation and death (Hall et al. 2007 ) . We have also observed high constitutive expression of IRAK-M mRNA in alveolar macrophages and peripheral blood buffy coat cells isolated from patients with sepsis-induced ALI, as compared to similar cell populations from healthy subjects (T. Standiford, unpublished observations). In fact, IRAK-M was the only negative regulator of TLR signaling found to be signi fi cantly induced in this patient population.

Several other molecules have been causally linked with the development of endotoxin tolerance or hyperin fl ammatory responses to LPS in genetically de fi cient mice. Suppression of tumorigenicity 2 (ST2) is a transmembrane protein and soluble secreted protein that is expressed by a variety of cells, including T cells and macrophages. ST2 inhibits MyD88-dependent signaling by interfering with the ability of Mal/TIRAP and MyD88 to interact with downstream signaling molecules. This protein appears to contribute to sepsis-induced impairment in lung antibacterial defense, at least in animal models (Holub et al. 2003 ) . Speci fi cally, CLP-induced impairment in anti-pseudomonal lung host defense is reversed in mice de fi cient in ST2. Interestingly, responsiveness of ST2 −/− AM was not altered, whereas the expression of IFN-g and TNF-a from CD4+ and CD8+ T cells was preserved in ST2 −/− mice in the setting of abdominal sepsis, as compared to similarly treat wildtype animals.

Toll-like receptor signaling can also be modulated by both extracellular and intracellular decoys. Single immunoglobulin IL-1R-related protein (SIGIRR) is a member of the IL-1 receptor superfamily but is unable to signal. However, the extracellular domain of this molecule inhibits Toll-IL-1 signaling by interfering with binding of ligands to TLR4, TLR5, TLR9, and IL-1 receptor I, whereas the intracellular domain interferes with the complexing of IRAK-1 with TRAF-6 ( Thomassen et al. 1999 ; Wald et al. 2003 ; Qin et al. 2005 ) . SIGIRR is expressed predominantly by epithelial cells, including alveolar epithelial cells, but also to a lesser degree in monocytic populations. Mice de fi cient in SIGIRR have enhanced in fl ammatory responses to LPS challenge. Moreover, SIGIRR is upregulated in the PBM of septic patients, and is associated with the development of endotoxin tolerance in these cells (Adib-Conquy et al. 2006 ) . MyD88 short (MyD88s) is an alternatively spliced variant of the parent molecule, MyD88. MyD88s functions as a dominant negative molecule by blocking recruitment of IRAK-4 to the toll-IL-1 signaling domain, resulting in reduced phosphorylation of IRAK-1 (Burns et al. 2003 ; Rao et al. 2005 ) . The expression of MyD88s is induced in monocytes in response to LPS and is constitutively expressed in blood monocytes isolated from patients with sepsis (Adib-Conquy et al. 2006 ) . Tollip disrupts IRAK-1 and IRAK-4 interactions, whereas microRNA 146 (miRNA 146) post-transcriptionally inhibits IRAK-1 and TRAF6 expression (Nahid et al. 2011 ) . The suppressors of cytokine signaling (SOCS) are a family of molecules that predominately inhibit JAK-Stat signaling, but also disrupt TLR signaling cascades through a yet unde fi ned mechanism. While these latter molecules could contribute to suppression of TLR-mediated responses during critical illness, there is no data to show enhanced expression and/or activity in blood monocytes or lung macrophages in patients at risk for the development of VAP.

Initiation of mechanical ventilation (MV) is a vital therapeutic intervention in patients with respiratory failure. A consequence of mechanical ventilation is the inhomogeneous distribution of pressure and volumes to various regions of lung, resulting in excessive stretch in some alveolar units (referred to as volutrauma), and repeated alveolar collapse in other regions (referred to as atelectotrauma) (Pugin et al. 1998 ) . Excessive lung stretch results in activation of several transcriptional pathways, including the NF-kB and the MAPK kinase pathway (Fos, Jun), which contributes to the release of various in fl ammatory mediators such as TNF-a , IL-1 b , IL-6, and IL-8 (Gharib et al. 2009 ; Halbertsma et al. 2005 ; Jaecklin et al. 2011 ) . These cellular mediators not only trigger deleterious lung injury responses and possibly multiple organ dysfunction ) , but may also promote the reprogramming of leukocytes and structural cells that occurs in critical illness. Importantly, MV at moderate to high lung volumes can also prime the lung for enhanced lung injury or systemic organ failure in response to an infectious challenge (e.g., second hit). For instance, as compared to spontaneously breathing animals, the intrapulmonary administration of S. aureus or E. coli to mechanically ventilated mice results in enhanced lung in fl ammation and lung injury, without changes in lung bacterial clearance . Likewise, the i.p. administration of LPS to mice undergoing high tidal volume MV substantially increased lung and systemic cytokine expression and extrapulmonary organ injury, as compared to non-mechanically ventilated controls . Mechanisms accounting for synergistic interactions between lung stretch and infectious challenge have not been clearly de fi ned. However, previous work has shown that stretch of human alveolar epithelial cells increases the expression of TLR2 by sixfold (Charles et al. 2011 ) . Moreover, mechanical ventilation increased the relative expression of TLR2 and TLR4 in lung tissue and increased the generation of endogenous ligands for TLR4 in bronchoalveolar lavage fl uid (Vaneker et al. 2008 ) . Recent work has shown that mechanical ventilation also generates other TLR4-independent and MyD88dependant endogenous TLR ligands (Chun et al. 2010 ) . Hyperin fl ation of the lung with high tidal volume not only promotes a signi fi cant increase in the expression of TLR4, but also paradoxically induces the expression of IRAK-M, an important negative regulator of TLR signaling (Villar et al. 2010 ) . A frequent consequence of mechanical ventilation and diseases that cause acute respiratory failure is alveolar collapse and atelectasis. Alveolar collapse is due, in part, to reductions in surfactant that occur in patients receiving mechanical ventilation and in patients with VAP (Nakos et al. 2003 ) . Atelectasis has been shown to promote bacterial overgrowth, and use of open ventilation strategies and administration of exogenous surfactant can reduce bacterial numbers in an animal model of VAP (van Kaam et al. 2004 ) . Moreover, administration of positive end-expiratory pressure (PEEP) at 5-8 cmH 2 O to non-hypoxemic mechanically ventilated patients can reduce the incidence of VAP (Manzano et al. 2008 ) . Surfactant proteins A and D can agglutinate P. aeruginosa , and SP-D can serve as an opsonin to enhance phagocytosis of P. aeruginosa (McCormack 2006 ) . Pseudomonal elastase has been shown to degrade SP-A and SP-D, and these proteins are decreased in the lungs of patients with cystic fi brosis (Mariencheck et al. 2003 ) . However, changes in SP-A and SP-D levels during mechanical ventilation and/or VAP have not been well characterized.

Administration of high concentrations of oxygen (FIO 2 >50%) used during transient or prolonged mechanical ventilation is a common treatment for patients with respiratory failure (Gore et al. 2010 ) . Although therapeutically necessary, hyperoxia results in the generation of reactive oxygen species (ROS), which promote the breakdown of critical barriers leading to systemic cellular and organ injury (Lee and Choi 2003 ) . In the lung, ROS cause severe cellular damage and death, exposure of the basement membrane and disruption of the alveolar capillary membrane leading to increased pulmonary permeability, in fl ux of in fl ammatory cells, and impaired gas exchange (Bhandari and Elias 2006 ) . Hyperoxic exposure can also exacerbate alveolar epithelial injury and apoptosis in response to infectious challenge with P. aeruginosa or L. pneumophila , resulting in increased bacterial dissemination (Kikuchi et al. 2009 ) . Moreover, high oxygen tensions inhibit the function of innate immune cells. For instance, macrophages exposed to elevated concentration of oxygen both in vitro and in vivo display reduced phagocytosis and killing of Gramnegative bacteria which correlated with changes in cell morphology and actin polymerization (O'Reilly et al. 2003 ) . In addition, in vivo hyperoxia exposure increased the susceptibility to K. pneumoniae lung infections, an effect that was partially attributed to reduced BAL GM-CSF levels and cell surface expression of TLR4 by AM (Baleeiro et al. 2003 ) . Importantly, systemic treatment of these mice with GM-CSF during hyperoxia preserved macrophage functionality and decreased the severity of lung infection (Baleeiro et al. 2006 ) . Taken together, hyperoxia is detrimental to the host by promoting greater alveolarcapillary injury, impairing local antibacterial responses, and increasing the risk of bacterial dissemination.

Emerging clinical and epidemiological data suggests a possible link between colonization with Candida species and susceptibility to P. aeruginosa pulmonary infection. Candida species is among the most common organisms recovered from endotracheal tube bio fi lm and tracheal secretions in patients with VAP (Adair et al. 1999 ) . Historically, Candida has been considered a commensal organisms rather than a true pathogen, and therefore believed to play no role in VAP disease pathogenesis. However, an observational study found a statistical association between airway colonization with Candida species and the development of P. aeruginosa VAP (Azoulay et al. 2006 ) . In a rat model of P. aeruginosa pneumonia, prior bronchial instillation of live but not heat-killed C. albicans resulted in increased susceptibility to subsequent bacterial challenge (Roux et al. 2009 ) . Mechanisms accounting for impaired in vivo clearance responses were not identi fi ed, but C. albicans was found to inhibit AM respiratory burst ex vivo. While these intriguing fi ndings require con fi rmation in other experimental model systems, they do raise the possibility that Candida and perhaps other commensal organisms may contribute meaningful to VAP pathogenesis.

Antibiotics, prophylactic measures to reduce oropharyngeal colonization and microaspiration, and approaches to stimulate mucocilliary transport are the mainstay of therapy to prevent and treat VAP. While these treatments are effective in some patients, adjuvant therapies are needed in others to bolster innate host responses, especially in the elderly and in patients with chronic immunosuppressive therapy. The recognition that critical illness can induce a profound state of immune dysregulation has prompted a reevaluation of potential immunologic approaches being used in the treatment of sepsis and other forms of critical illness (Pockros et al. 2007a ) . Effective immunoadjuvant therapy must necessarily promote antimicrobial effects without exacerbating deleterious lung in fl ammatory responses.

Common features of both endotoxin tolerance and immune dysregulation of critical illness is impaired TLR signaling, NF-k B-dependent responses, reduced APC function, and a shift toward type 2 rather than type 1 immune responses. Two cytokines that have been shown to partially reverse these changes in vitro and in vivo are IFN-g and GM-CSF. In endotoxin-tolerized monocytes, treatment with IFN-g or GM-CSF can reverse the tolerance phenotype, in part by facilitating interactions between IRAK and MyD88, resulting in enhanced downstream activation of NF-k B (Adib-Conquy and Cavaillon 2002 ) . Similarly, ex vivo treatment of blood monocytes from trauma patients with IFN-g or GM-CSF, but not G-CSF, enhanced LPSinduced cytokine production, and HLA-DR expression (Lendemans et al. 2007 ) . These preclinical studies served as the foundation for several small clinical trials in patients with sepsis. Docke and colleagues administered IFN-g to patients with sepsis in an attempt to reverse the cytokine imbalance and restore monocyte function (Docke et al. 1997 ) . In this uncontrolled study, nine patients with evidence of sepsis-induced immunosuppression (decreased blood monocyte HLA-DR expression) were administered IFN-g at a dose of 100 m g subcutaneously daily. Treatment with IFN-g resulted in increased monocyte HLA-DR expression in all patients, along with a restoration of monocyte TNF-a production to levels observed in monocytes isolated from healthy subjects. Resolution of sepsis occurred in eight of the nine treated patients (Docke et al. 1997 ) . In two small single center clinical trials, the i.v. administration of hrGM-CSF to patients with sepsis resulted in improvements in ex vivo effector responses in PBMs or neutrophils (Nierhaus et al. 2003 ; Presneill et al. 2002 ) . Moreover, one of the studies revealed improvements in PaO 2 /FIO 2 ratios, as a measure of pulmonary gas exchange, suggesting reduced lung injury in the GM-CSF treated group (Presneill et al. 2002 ) . Prevention of lung injury may be due, in part, to the fact that GM-CSF is an alveolar epithelial cell mitogen and can protect the alveolar epithelium against hyperoxic and bleomycin-induced injury (Baleeiro et al. 2006 ; Moore et al. 2000 ) and in a murine model of in fl uenza pneumonia (Sever-Chroneos et al. 2011 ) . These preclinical and clinical fi ndings served as the basis for a multicenter randomized placebo controlled trial of subcutaneous GM-CSF administration in 38 patients with severe sepsis and evidence of monocyte deactivation (reduced HLA-DR expression). As compared to the placebo group, GM-CSF administration resulted in improved monocyte function (restored cell surface TLR2/4 expression, TNF production, and HLA-DR expression) and improved clinical outcomes, including reduced APACHE II scores, shorter time of mechanical ventilation, and a trend toward decreased length of ICU and hospital stay. These studies and others suggest that immunostimulatory therapy for treatment of critical illness-induced immune dysregulation or even end-organ injury appears to be a potentially viable therapeutic option that warrants larger controlled trials (Luedke and Cerami 1990 ) . An obvious concern of immunostimulatory therapy in patients with severe sepsis and/or pneumonia is the potential of exacerbating the "cytokine storm" of SIRS. Fortunately, neither IFN-g nor GM-CSF has precipitated worsening of hemodynamic compromise or multiorgan failure, even in patient with severe sepsis or septic shock (Docke et al. 1997 ; Nierhaus et al. 2003 ; Meisel et al. 2009 ) . Additional consideration could be given to compartmentalized immunostimulatory therapy (e.g., aerosolized delivery) to prevent or treat VAP. However, this approach may be limited substantially by ventilation-perfusion mismatching that occurs in patients with lung disease, and the concern that the leukocyte reprogramming that occurs during critical illness is not limited to the lung microenvironment but almost certainly occurs more broadly in leukocyte populations systemically.

Activation of the PI3K/Akt pathway in certain leukocyte populations can lessen NF-k B-mediated pro-in fl ammatory responses while stimulating pro-survival and antimicrobial responses (Williams et al. 2006 ; Wrann et al. 2007 ; Zhang et al. 2007 ) . For example, the administration of selective activators of the PI3K/Akt signaling pathway (e.g., glucan, a -lipoic acid) to LPS-challenged mice or mice undergoing CLP reduced apoptosis, in fl ammatory cytokine release, and improved mortality (Wrann et al. 2007 ; Zhang et al. 2007 ) .

Interleukin 15 is a pleurapotential cytokine that regulates DC, T, and NK cell activation, proliferation, and survival. The administration of IL-15 to mice with abdominal sepsis (CLP) has been shown to block sepsis-induced apoptosis of NK cells, DC, and CD8 T cells, and to restore NK cell production of IFN-g (Inoue et al. 2010 ) . Treatment with IL-15 also mitigated sepsis-induced apoptosis of gut epithelium. Importantly, IL-15 not only reduced mortality in CLP, but also in mice administered P. aeruginosa i.t.

Finally, caspase inhibitors have been shown to reduce lymphocyte apoptosis and increase survival in murine models of sepsis (Hotchkiss et al. 2000 ) . A pan-caspase inhibitor (IDN-6556) have been employed in the treatment of liver disease in patients with Hepatitis C (Pockros et al. 2007b ) . However, trials targeting caspases or other pro-apoptotic molecules or administration of pro-survival factors (e.g., AKT activators, IL-15) in patients with sepsis or nosocomial pneumonia have not yet been reported.

In this review, we have de fi ned the clinical features of VAP, and described the impact of critical illness and microenvironment factors introduced during mechanical ventilation on susceptibility to VAP, with special attention to speci fi c molecules as potential mediators of immunosuppression and tissue injury. Increases in microbial resistance, combined with a burgeoning population of patients at risk, are trends that clearly make VAP a major clinical problem now and in the future. Preventative strategies and optimal ventilator management have been paramount in reducing the incidence of VAP. However, critical illness-induced reprogramming of leukocyte innate immune responses clearly contributes to susceptibility to VAP and VAPinduced tissue injury. Given our past failures, a paradigm shift in how we approach patients with evidence of immune dysregulation is required. In order for novel therapies to proceed, better clinical markers are needed to distinguish a deleterious innate response (e.g. SIRS) from a state of immunoparalysis (CARS) or mixed antagonist response syndrome (MARS) as the in fl ammatory response evolves (Wesche et al. 1999 ) . Differentiating these quite disparate but overlapping responses in a patient-speci fi c fashion will allow for better selection of patients in which immunoadjuvant therapy is more likely to be bene fi cial.

Implications of endotracheal tube bio fi lm for ventilator-associated pneumonia

Toll-like receptors in the induction of the innate immune response

Gamma interferon and granulocyte/monocyte colonystimulating factor prevent endotoxin tolerance in human monocytes by promoting interleukin-1 receptor-associated kinase expression and its association to MyD88 and not by modulating TLR4 expression

Up-regulation of MyD88s and SIGIRR, molecules inhibiting Toll-like receptor signaling, in monocytes from septic patients

Recognition of pathogen-associated molecular patterns by TLR family

Guidelines for the management of adults with hospital-acquired, ventilator-associated, and healthcare-associated pneumonia

Heme oxygenase-1 system, in fl ammation and ventilator-induced lung injury

Experimental and clinical signi fi cance of endotoxin-dependent HLA-DR expression on monocytes

Polymicrobial sepsis selectively activates peritoneal but not alveolar macrophages to release in fl ammatory mediators (interleukins-1 and -6 and tumor necrosis factor)

Polymicrobial sepsis but not lowdose endotoxin infusion causes decreased splenocyte IL-2/IFN-[gamma] release while increasing IL-4/IL-10 production

Candida colonization of the respiratory tract and subsequent pseudomonas ventilator-associated pneumonia

Sublethal hyperoxia impairs pulmonary innate immunity

GM-CSF and the impaired pulmonary innate immune response following hyperoxic stress

Role of nitric oxide in the failure of neutrophil migration in sepsis

Nuclease expression by Staphylococcus aureus facilitates escape from neutrophil extracellular traps

TLR9 is required for protective innate immunity in Gram-negative bacterial pneumonia: role of dendritic cells

Toll-like receptor 9 regulates the lung macrophage phenotype and host immunity in murine pneumonia caused by Legionella pneumophila

Cooperative interactions between TLR4 and TLR9 regulate interleukin 23 and 17 production in a murine model of gram negative bacterial pneumonia

Cytokines in tolerance to hyperoxia-induced injury in the developing and adult lung

Sir Isaac Newton, sepsis, SIRS, and CARS

The epidemiology of the systemic in fl ammatory response

TLR2, TLR4, CD14, CD11B, and CD11C expressions on monocytes surface and cytokine production in patients with sepsis, severe sepsis, and septic shock

DNase expression allows the pathogen group A Streptococcus to escape killing in neutrophil extracellular traps

Inhibition of interleukin 1 receptor/Toll-like receptor signaling through the alternatively spliced, short form of MyD88 is due to its failure to recruit IRAK-4

Endotoxin tolerance disrupts chromatin remodeling and NF-{kappa}B transactivation at the IL-1{beta} promoter

Mild-stretch mechanical ventilation upregulates toll-like receptor 2 and sensitizes the lung to bacterial lipopeptide

Mechanical ventilation modulates Toll-like receptor-3-induced lung in fl ammation via a MyD88-dependent, TLR4-independent pathway: a controlled animal study

Platelet TLR4 activates neutrophil extracellular traps to ensnare bacteria in septic blood

Expression and function of the chemokine receptors CXCR1 and CXCR2 in sepsis

Protective effects of C5a blockade in sepsis

Sepsis-induced suppression of lung innate immunity is mediated by IRAK-M

Mechanical ventilation induces in fl ammation, lung injury, and extra-pulmonary organ dysfunction in experimental pneumonia

Proin fl ammatory cytokines

Monocyte deactivation in septic patients: restoration by IFN-gamma treatment

Altered phenotype of blood dendritic cells in patients with acute pneumonia

Rapid up-regulation of IRAK-M expression following a second endotoxin challenge in human monocytes and in monocytes isolated from septic patients

T helper cell subset ratios in patients with severe sepsis

Functional and genetic evidence that the Mal/TIRAP allele variant 180 L has been selected by providing protection against septic shock

Gene-speci fi c control of in fl ammation by TLRinduced chromatin modi fi cations

Pathogenetic signi fi cance of biological markers of ventilator-associated lung injury in experimental and clinical studies

Regulatory dendritic cells act as regulators of acute lethal systemic in fl ammatory response

Pneumonia due to Pseudomonas aeruginosa: part I: epidemiology, clinical diagnosis, and source

Alteration of mononuclear cell immune-associated antigen expression, interleukin-1 expression, and antigen presentation during intra-abdominal infection

Oropharyngeal or gastric colonization and nosocomial pneumonia in adult intensive care unit patients. A prospective study based on genomic DNA analysis

Noninjurious mechanical ventilation activates a proin fl ammatory transcriptional program in the lung

Alternative activation of macrophages: mechanism and functions

Hyperoxia sensing: from molecular mechanisms to signi fi cance in disease

Characteristics of alveolar macrophages in experimental septic lung

An essential role for non-bone marrow-derived cells in control of Pseudomonas aeruginosa pneumonia

Cytokines and biotrauma in ventilator-induced lung injury: a critical review of the literature

Monocyte mRNA phenotype and adverse outcomes from pediatric multiple organ dysfunction syndrome

Chronic stimulation of Nod2 mediates tolerance to bacterial products

Selective defects of T lymphocyte function in patients with lethal intraabdominal infection

Lymphocyte subset numbers depend on the bacterial origin of sepsis

The pathophysiology and treatment of sepsis

Caspase inhibitors improve survival in sepsis: a critical role of the lymphocyte

Sepsisinduced apoptosis causes progressive profound depletion of B and CD4+ T lymphocytes in humans

Depletion of dendritic cells, but not macrophages, in patients with sepsis

NOD2 contributes to cutaneous defense against Staphylococcus aureus through alpha-toxin-dependent innate immune activation

Role of C5a in multiorgan failure during sepsis

Interaction of soluble form of recombinant extracellular TLR4 domain with MD-2 enables lipopolysaccharide binding and attenuates TLR4-mediated signaling

NOD-LRR proteins: role in host-microbial interactions and in fl ammatory disease

IL-15 prevents apoptosis, reverses innate and adaptive immune dysfunction, and improves survival in sepsis

Cutting edge: naturally occurring soluble form of mouse Toll-like receptor 4 inhibits lipopolysaccharide signaling

Lungderived soluble mediators are pathogenic in ventilator-induced lung injury

Patterns of early and late ventilator-associated pneumonia due to methicillin-resistant Staphylococcus aureus in a trauma population

Hyperoxia exaggerates bacterial dissemination and lethality in Pseudomonas aeruginosa pneumonia

IRAK-M is a negative regulator of Toll-like receptor signaling

Soluble forms of Toll-like receptor (TLR)2 capable of modulating TLR2 signaling are present in human plasma and breast milk

Pathways of cell signaling in hyperoxia

Differential immunostimulating effect of granulocyte-macrophage colony-stimulating factor (GM-CSF), granulocyte colony-stimulating factor (G-CSF) and interferon gamma (IFNgamma) after severe trauma

Interferon-gamma overcomes glucocorticoid suppression of cachectin/ tumor necrosis factor biosynthesis by murine macrophages

IRAK-M regulates chromatin remodeling in lung macrophages during experimental sepsis

Neutrophils in the activation and regulation of innate and adaptive immunity

Positive-end expiratory pressure reduces incidence of ventilator-associated pneumonia in nonhypoxemic patients

Pseudomonas aeruginosa elastase degrades surfactant proteins A and D

The Epidemiology of Sepsis in the United States from 1979 through 2000

New concepts in collectin-mediated host defense at the air-liquid interface of the lung

Granulocytemacrophage colony-stimulating factor to reverse sepsis-associated immunosuppression: a double-blind, randomized, placebo-controlled multicenter trial

American College of Chest Physicians/Society of Critical Care Medicine consensus conference: de fi nitions for sepsis and organ failure and guidelines for the use of innovative therapies in sepsis

GM-CSF regulates bleomycin-induced pulmonary fi brosis via a prostaglandin-dependent mechanism

Innate Immune Responses in Ventilator-Associated Pneumonia

Dysregulation of in vitro cytokine production by monocytes during sepsis

Pneumonia complicating abdominal sepsis. An independent risk factor for mortality

Mechanistic role of microRNA-146a in endotoxin-induced differential cross-regulation of TLR signaling

Involvement of IRAK-M in peptidoglycan-induced tolerance in macrophages

Ventilator-associated pneumonia and atelectasis: evaluation through bronchoalveolar lavage fl uid analysis

Excessive neutrophils and neutrophil extracellular traps contribute to acute lung injury of in fl uenza pneumonitis

Lung pathology of fatal severe acute respiratory syndrome

Reversal of immunoparalysis by recombinant human granulocyte-macrophage colony-stimulating factor in patients with severe sepsis

Cutting edge: heat shock protein 60 is a putative endogenous ligand of the Toll-like receptor-4 complex

Mechanical ventilation interacts with endotoxemia to induce extrapulmonary organ dysfunction

Hyperoxia impairs antibacterial function of macrophages through effects on actin

Major injury leads to predominance of the T helper-2 lymphocyte phenotype and diminished interleukin-12 production associated with decreased resistance to infection

Neutrophil elastase and myeloperoxidase regulate the formation of neutrophil extracellular traps

Involvement of toll-like receptors 2 and 4 in cellular activation by high mobility group box 1 protein

Myeloperoxidase associated with neutrophil extracellular traps is active and mediates bacterial killing in the presence of hydrogen peroxide

Effects of salmeterol on cilia and mucus in COPD and pneumonia patients

In fl uenza virus infection decreases tracheal mucociliary velocity and clearance of Streptococcus pneumoniae

Oral IDN-6556, an antiapoptotic caspase inhibitor, may lower aminotransferase activity in patients with chronic hepatitis C

Oral IDN-6556, an antiapoptotic caspase inhibitor, may lower aminotransferase activity in patients with chronic hepatitis C

A randomized phase II trial of granulocyte-macrophage colony-stimulating factor therapy in severe sepsis with respiratory dysfunction

Activation of human macrophages by mechanical ventilation in vitro

SIGIRR inhibits interleukin-1 receptor-and Toll-like receptor 4-mediated signaling through different mechanisms

Systemic in fl ammatory response and increased risk for ventilator-associated pneumonia: a preliminary study

Control of Pseudomonas aeruginosa in the lung requires the recognition of either lipopolysaccharide or fl agellin

A novel splice variant of interleukin-1 receptor (IL-1R)-associated kinase 1 plays a negative regulatory role in Toll/IL-1R-induced in fl ammatory signaling

Innate response activator B cells protect against microbial sepsis

Alveolar macrophage deactivation in murine septic peritonitis: role of interleukin 10

Dying for a cause: NETosis, mechanisms behind an antimicrobial cell death modality

Pulmonary infection complicating intra-abdominal sepsis

Candida albicans impairs macrophage function and facilitates Pseudomonas aeruginosa pneumonia in rat

Type III protein secretion is associated with death in lower respiratory and systemic Pseudomonas aeruginosa infections

Pathogen-host interactions in Pseudomonas aeruginosa pneumonia

Central role of toll-like receptor 4 signaling and host defense in experimental pneumonia caused by Gram-negative bacteria

Increased natural CD4+CD25+ regulatory t cells and their suppressor activity do not contribute to mortality in murine polymicrobial sepsis

Critical role of IL-1 receptor-associated kinase-M in regulating chemokine-dependent deleterious in fl ammation in murine in fl uenza pneumonia

GM-CSF modulates pulmonary resistance to in fl uenza A infection

Cutting edge: myeloid differentiation factor 88 is essential for pulmonary host defense against Pseudomonas aeruginosa but not Staphylococcus aureus

IL-10 is a major mediator of sepsis-induced impairment in lung antibacterial host defense

Decreased systemic polymorphonuclear neutrophil (PMN) rolling without increased PMN adhesion in peritonitis at remote sites

Innate Immune Responses in Ventilator-Associated Pneumonia

Pro-and antiin fl ammatory responses are regulated simultaneously from the fi rst moments of septic shock

The use of gene-expression pro fi ling to identify candidate genes in human sepsis

Identi fi cation and characterization of SIGIRR, a molecule representing a novel subtype of the IL-1R superfamily

Neutrophil extracellular traps contain calprotectin, a cytosolic protein complex involved in host defense against Candida albicans

Low-tidal-volume mechanical ventilation induces a toll-like receptor 4-dependent in fl ammatory response in healthy mice

Induction of IRAK-M is associated with lipopolysaccharide tolerance in a human endotoxemia model

Regulatory T cell populations in sepsis and trauma

Mechanical ventilation modulates TLR4 and IRAK-3 in a non-infectious, ventilator-induced lung injury model

Clinical aspects: from systemic in fl ammation to 'immunoparalysis'

SIGIRR, a negative regulator of Toll-like receptor-interleukin 1 receptor signaling

Epigenetic regulation of dendritic cell-derived interleukin-12 facilitates immunosuppression after a severe innate immune response

IRAK-M is a novel member of the pelle/interleukin-1 receptor-associated kinase (IRAK) family

The MyD88-dependent, but not the MyD88-independent, pathway of TLR4 signaling is important in clearing nontypeable haemophilus in fl uenzae from the mouse lung

Granulocytemacrophage colony-stimulating factor induces activation and restores respiratory burst activity in monocytes from septic patients

Modulation of the phosphoinositide 3-kinase signaling pathway alters host response to sepsis, in fl ammation, and ischemia/reperfusion injury

The contribution of CD4+ CD25+ T-regulatory-cells to immune suppression in sepsis

Increased release of sMD-2 during human endotoxemia and sepsis: a role for endothelial cells

The phosphatidylinositol 3-kinase signaling pathway exerts protective effects during sepsis by controlling C5a-mediated activation of innate immune functions

IL-12 suppression during experimental endotoxin tolerance: dendritic cell loss and macrophage hyporesponsiveness

Neutrophil extracellular trap (NET)-mediated killing of Pseudomonas aeruginosa: evidence of acquired resistance within the CF airway, independent of CFTR

T-cell reactivity and its predictive role in immunosuppression after burns

Alpha-lipoic acid attenuates LPS-induced in fl ammatory responses by activating the phosphoinositide 3-kinase/Akt signaling pathway

TLR 2 and CD14 mediate innate immunity and lung in fl ammation to staphylococcal Panton-Valentine leukocidin in vivo