DAMP molecule S100A9 acts as a molecular pattern to enhance inflammation during influenza A virus infection: role of DDX21-TRIF-TLR4-MyD88 pathway.

Pathogen-associated molecular patterns (PAMPs) trigger host immune response by activating pattern recognition receptors like toll-like receptors (TLRs). However, the mechanism whereby several pathogens, including viruses, activate TLRs via a non-PAMP mechanism is unclear. Endogenous "inflammatory mediators" called damage-associated molecular patterns (DAMPs) have been implicated in regulating immune response and inflammation. However, the role of DAMPs in inflammation/immunity during virus infection has not been studied. We have identified a DAMP molecule, S100A9 (also known as Calgranulin B or MRP-14), as an endogenous non-PAMP activator of TLR signaling during influenza A virus (IAV) infection. S100A9 was released from undamaged IAV-infected cells and extracellular S100A9 acted as a critical host-derived molecular pattern to regulate inflammatory response outcome and disease during infection by exaggerating pro-inflammatory response, cell-death and virus pathogenesis. Genetic studies showed that the DDX21-TRIF signaling pathway is required for S100A9 gene expression/production during infection. Furthermore, the inflammatory activity of extracellular S100A9 was mediated by activation of the TLR4-MyD88 pathway. Our studies have thus, underscored the role of a DAMP molecule (i.e. extracellular S100A9) in regulating virus-associated inflammation and uncovered a previously unknown function of the DDX21-TRIF-S100A9-TLR4-MyD88 signaling network in regulating inflammation during infection.

The production of monokaryotic hyphae by Cryptococcus neoformans can be induced by high temperature arrest of the cell cycle and is independent of same-sex mating.

Cryptococcus neoformans is a heterothallic fungal pathogen of humans and animals. Although the fungus grows primarily as a yeast, hyphae are produced during the sexual phase and during a process called monokaryotic fruiting, which is also believed to involve sexual reproduction, but between cells of the same mating type. Here we report a novel monokaryotic fruiting mechanism that is dependent on the cell cycle and occurs in haploid cells in the absence of sexual reproduction. Cells grown at 37°C were found to rapidly produce hyphae (∼4 hrs) and at high frequency (∼40% of the population) after inoculation onto hyphae-inducing agar. Microscopic examination of the 37°C seed culture revealed a mixture of normal-sized and enlarged cells. Micromanipulation of single cells demonstrated that only enlarged cells were able to produce hyphae and genetic analysis confirmed that hyphae did not arise from α-α mating or endoduplication. Cell cycle analysis revealed that cells grown at 37°C had an increased population of cells in G2 arrest, with the proportion correlated with the frequency of monokaryotic fruiting. Cell sorting experiments demonstrated that enlarged cells were only found in the G2-arrested population and only this population contained cells able to produce hyphae. Treatment of cells at low temperature with the G2 cell cycle arrest agent, nocodazole, induced hyphal growth, confirming the role of the cell cycle in this process. Taken together, these results reveal a mating-independent mechanism for monokaryotic fruiting, which is dependent on the cell cycle for induction of hyphal competency.

The incidence, success rate, and complications of awake tracheal intubation in 1,554 patients over 12 years: an historical cohort study.

PURPOSE: Awake tracheal intubation is one recommended option to address select situations in the management of a patient with an anticipated difficult airway. A scarcity of data exists on how often awake intubation is performed or whether its use is changing over time, particularly with the increasingly widespread availability of video laryngoscopy. This retrospective database review was undertaken to determine the incidence, success, and complications of awake intubation and the incidence of other tracheal intubation techniques in the operating room over a 12-yr period (2002-2013) at our institution. METHODS: The Anesthesia Information Management System in use at a Canadian tertiary care centre was searched for all awake intubations that occurred during the years 2002-2013. Records were also searched to identify airway methods other than direct laryngoscopy that may have been used after the induction of general anesthesia. Changes in both the incidence of awake intubation and in the use of video laryngoscopy over the 12 years were analyzed using linear regression modelling. RESULTS: Of 146,252 cases performed under general anesthesia with endotracheal intubation, 1,554 intubations (1.06%) were performed awake. There was no significant change in the rate of awake intubation over the studied years (slope -1.4(-4) incidence·year(-1); 95% confidence interval [CI]: -3.0(-4) to 3.0(-5); P = 0.102). The relatively steady rate of awake intubation occurred despite a significant increase in the use of video laryngoscopy over the same time (slope 0.080 incidence·year(-1); 95% CI: 0.076 to 0.083; P < 0.001), particularly from 2009 onwards. Attempted awake intubation failed in 31 (2%) of the cases. Self-reported complications occurred in 15.7% of successful procedures. In addition, in a convenience sample of three years (2011-2013), the rate at which each of 49 attending staff performed awake intubation varied widely from 0-3.4 awake intubations per 100 cases of general anesthesia with endotracheal intubation. CONCLUSIONS: At our tertiary care centre, we did not find a significant change in the use of awake tracheal intubation over the studied years 2002-2013 despite increasing availability and use of video laryngoscopy. It appears that awake tracheal intubation retains an important and consistent role in the management of the difficult airway.

The difficult airway with recommendations for management--part 2--the anticipated difficult airway.

BACKGROUND: Appropriate planning is crucial to avoid morbidity and mortality when difficulty is anticipated with airway management. Many guidelines developed by national societies have focused on management of difficulty encountered in the unconscious patient; however, little guidance appears in the literature on how best to approach the patient with an anticipated difficult airway. METHODS: To review this and other subjects, the Canadian Airway Focus Group (CAFG) was re-formed. With representation from anesthesiology, emergency medicine, and critical care, CAFG members were assigned topics for review. As literature reviews were completed, results were presented and discussed during teleconferences and two face-to-face meetings. When appropriate, evidence- or consensus-based recommendations were made, and levels of evidence were assigned. PRINCIPAL FINDINGS: Previously published predictors of difficult direct laryngoscopy are widely known. More recent studies report predictors of difficult face mask ventilation, video laryngoscopy, use of a supraglottic device, and cricothyrotomy. All are important facets of a complete airway evaluation and must be considered when difficulty is anticipated with airway management. Many studies now document the increasing patient morbidity that occurs with multiple attempts at tracheal intubation. Therefore, when difficulty is anticipated, tracheal intubation after induction of general anesthesia should be considered only when success with the chosen device(s) can be predicted in a maximum of three attempts. Concomitant predicted difficulty using oxygenation by face mask or supraglottic device ventilation as a fallback makes an awake approach advisable. Contextual issues, such as patient cooperation, availability of additional skilled help, and the clinician's experience, must also be considered in deciding the appropriate strategy. CONCLUSIONS: With an appropriate airway evaluation and consideration of relevant contextual issues, a rational decision can be made on whether an awake approach to tracheal intubation will maximize patient safety or if airway management can safely proceed after induction of general anesthesia. With predicted difficulty, close attention should be paid to details of implementing the chosen approach. This should include having a plan in case of the failure of tracheal intubation or patient oxygenation.

The difficult airway with recommendations for management--part 1--difficult tracheal intubation encountered in an unconscious/induced patient.

BACKGROUND: Previously active in the mid-1990s, the Canadian Airway Focus Group (CAFG) studied the unanticipated difficult airway and made recommendations on management in a 1998 publication. The CAFG has since reconvened to examine more recent scientific literature on airway management. The Focus Group's mandate for this article was to arrive at updated practice recommendations for management of the unconscious/induced patient in whom difficult or failed tracheal intubation is encountered. METHODS: Nineteen clinicians with backgrounds in anesthesia, emergency medicine, and intensive care joined this iteration of the CAFG. Each member was assigned topics and conducted reviews of Medline, EMBASE, and Cochrane databases. Results were presented and discussed during multiple teleconferences and two face-to-face meetings. When appropriate, evidence- or consensus-based recommendations were made together with assigned levels of evidence modelled after previously published criteria. CONCLUSIONS: The clinician must be aware of the potential for harm to the patient that can occur with multiple attempts at tracheal intubation. This likelihood can be minimized by moving early from an unsuccessful primary intubation technique to an alternative "Plan B" technique if oxygenation by face mask or ventilation using a supraglottic device is non-problematic. Irrespective of the technique(s) used, failure to achieve successful tracheal intubation in a maximum of three attempts defines failed tracheal intubation and signals the need to engage an exit strategy. Failure to oxygenate by face mask or supraglottic device ventilation occurring in conjunction with failed tracheal intubation defines a failed oxygenation, "cannot intubate, cannot oxygenate" situation. Cricothyrotomy must then be undertaken without delay, although if not already tried, an expedited and concurrent attempt can be made to place a supraglottic device.

Phosphatidylinositol 3-kinase activation attenuates the TLR2-mediated macrophage proinflammatory cytokine response to Francisella tularensis live vaccine strain.

An inadequate innate immune response appears to contribute to the virulence of Francisella tularensis following pulmonary infection. Studies in mice suggest that this poor response results from suppression of proinflammatory cytokine production early during infection, but the mechanisms involved are not understood. PI3K is known to regulate proinflammatory cytokine expression, but its exact role (positive versus negative) is controversial. We sought to clarify the role of PI3K in regulating proinflammatory signaling and cytokine production during infection with F. tularensis live vaccine strain (LVS). In this study, we demonstrate that the induction of TNF and IL-6 expression by LVS in mouse bone marrow-derived macrophages was markedly enhanced when PI3K activity was inhibited by either of the well-known chemical inhibitors, wortmannin or LY294002. The enhanced cytokine expression was accompanied by enhanced activation of p38 MAPK and ERK1/2, both of which were critical for LVS-induced expression of TNF and IL-6. LVS-induced MAPK activation and cytokine production were TLR2- and MyD88- dependent. PI3K/Akt activation was MyD88-dependent, but was surprisingly TLR2-independent. LVS infection also rapidly induced MAPK phosphatase-1 (MKP-1) expression; PI3K and TLR2 signaling were required. Peak levels of MKP-1 correlated closely with the decline in p38 MAPK and ERK1/2 phosphorylation. These data suggest that infection by LVS restrains the TLR2-triggered proinflammatory response via parallel activation of PI3K, leading to enhanced MKP-1 expression, accelerated deactivation of MAPKs, and suppression of proinflammatory cytokine production. This TLR2-independent inhibitory pathway may be an important mechanism by which Francisella suppresses the host's innate immune response.

TLR2/MyD88/NF-κB pathway, reactive oxygen species, potassium efflux activates NLRP3/ASC inflammasome during respiratory syncytial virus infection.

Human respiratory syncytial virus (RSV) constitute highly pathogenic virus that cause severe respiratory diseases in newborn, children, elderly and immuno-compromised individuals. Airway inflammation is a critical regulator of disease outcome in RSV infected hosts. Although "controlled" inflammation is required for virus clearance, aberrant and exaggerated inflammation during RSV infection results in development of inflammatory diseases like pneumonia and bronchiolitis. Interleukin-1ß (IL-1ß) plays an important role in inflammation by orchestrating the pro-inflammatory response. IL-1ß is synthesized as an immature pro-IL-1ß form. It is cleaved by activated caspase-1 to yield mature IL-1ß that is secreted extracellularly. Activation of caspase-1 is mediated by a multi-protein complex known as the inflammasome. Although RSV infection results in IL-1ß release, the mechanism is unknown. Here in, we have characterized the mechanism of IL-1ß secretion following RSV infection. Our study revealed that NLRP3/ASC inflammasome activation is crucial for IL-1ß production during RSV infection. Further studies illustrated that prior to inflammasome formation; the "first signal" constitutes activation of toll-like receptor-2 (TLR2)/MyD88/NF-κB pathway. TLR2/MyD88/NF-κB signaling is required for pro-IL-1ß and NLRP3 gene expression during RSV infection. Following expression of these genes, two "second signals" are essential for triggering inflammasome activation. Intracellular reactive oxygen species (ROS) and potassium (K(+)) efflux due to stimulation of ATP-sensitive ion channel promote inflammasome activation following RSV infection. Thus, our studies have underscored the requirement of TLR2/MyD88/NF-κB pathway (first signal) and ROS/potassium efflux (second signal) for NLRP3/ASC inflammasome formation, leading to caspase-1 activation and subsequent IL-1ß release during RSV infection.

Does postoperative ileus exist?

In the past, it was assumed that normal feeding immediately after major abdominal surgery was impossible because normal intestinal motility was disrupted. Today, such a view is widely rejected. However, there has never been clear agreement on just what is the supposed abnormality in intestinal function. There are major difficulties in studying intestinal function after surgery. Furthermore, a number of other factors such as anesthetic agents and analgesia can influence the situation. There has been very little research on the subject in recent years, and uncertainty remains about this subject, which is of profound importance to patients' comfort and well-being.

TLR-dependent control of Francisella tularensis infection and host inflammatory responses.

BACKGROUND: Francisella tularensis is the causative agent of tularemia and is classified as a Category A select agent. Recent studies have implicated TLR2 as a critical element in the host protective response to F. tularensis infection, but questions remain about whether TLR2 signaling dominates the response in all circumstances and with all species of Francisella and whether F. tularensis PAMPs are predominantly recognized by TLR2/TLR1 or TLR2/TLR6. To address these questions, we have explored the role of Toll-like receptors (TLRs) in the host response to infections with F. tularensis Live Vaccine Strain (LVS) and F. tularensis subspecies (subsp.) novicida in vivo. METHODOLOGY/PRINCIPAL FINDINGS: C57BL/6 (B6) control mice and TLR- or MyD88-deficient mice were infected intranasally (i.n.) or intradermally (i.d.) with F. tularensis LVS or with F. tularensis subsp. novicida. B6 mice survived >21 days following infection with LVS by both routes and survival of TLR1(-/-), TLR4(-/-), and TLR6(-/-) mice infected i.n. with LVS was equivalent to controls. Survival of TLR2(-/-) and MyD88(-/-) mice, however, was significantly reduced compared to B6 mice, regardless of the route of infection or the subspecies of F. tularensis. TLR2(-/-) and MyD88(-/-) mice also showed increased bacterial burdens in lungs, liver, and spleen compared to controls following i.n. infection. Primary macrophages from MyD88(-/-) and TLR2(-/-) mice were significantly impaired in the ability to secrete TNF and other pro-inflammatory cytokines upon ex vivo infection with LVS. TNF expression was also impaired in vivo as demonstrated by analysis of bronchoalveolar lavage fluid and by in situ immunofluorescent staining. CONCLUSIONS/SIGNIFICANCE: We conclude from these studies that TLR2 and MyD88, but not TLR4, play critical roles in the innate immune response to F. tularensis infection regardless of the route of infection or the subspecies. Moreover, signaling through TLR2 does not depend exclusively on TLR1 or TLR6 during F. tularensis LVS infection.