The NLRP6 Inflammasome Recognizes Lipoteichoic Acid and Regulates Gram-Positive Pathogen Infection.

The activator and composition of the NLRP6 inflammasome remain poorly understood. We find that lipoteichoic acid (LTA), a molecule produced by Gram-positive bacteria, binds and activates NLRP6. In response to cytosolic LTA or infection with Listeria monocytogenes, NLRP6 recruited caspase-11 and caspase-1 via the adaptor ASC. NLRP6 activation by LTA induced processing of caspase-11, which promoted caspase-1 activation and interleukin-1ß (IL-1ß)/IL-18 maturation in macrophages. Nlrp6-/- and Casp11-/- mice were less susceptible to L. monocytogenes infection, which was associated with reduced pathogen loads and impaired IL-18 production. Administration of IL-18 to Nlrp6-/- or Casp11-/- mice restored the susceptibility of mutant mice to L. monocytogenes infection. These results reveal a previously unrecognized innate immunity pathway triggered by cytosolic LTA that is sensed by NLRP6 and exacerbates systemic Gram-positive pathogen infection via the production of IL-18.

naRNA-LL37 composite DAMPs define sterile NETs as self-propagating drivers of inflammation.

Neutrophil extracellular traps (NETs) are a key antimicrobial feature of cellular innate immunity mediated by polymorphonuclear neutrophils (PMNs). NETs counteract microbes but are also linked to inflammation in atherosclerosis, arthritis, or psoriasis by unknown mechanisms. Here, we report that NET-associated RNA (naRNA) stimulates further NET formation in naive PMNs via a unique TLR8-NLRP3 inflammasome-dependent pathway. Keratinocytes respond to naRNA with expression of psoriasis-related genes (e.g., IL17, IL36) via atypical NOD2-RIPK signaling. In vivo, naRNA drives temporary skin inflammation, which is drastically ameliorated by genetic ablation of RNA sensing. Unexpectedly, the naRNA-LL37 'composite damage-associated molecular pattern (DAMP)' is pre-stored in resting neutrophil granules, defining sterile NETs as inflammatory webs that amplify neutrophil activation. However, the activity of the naRNA-LL37 DAMP is transient and hence supposedly self-limiting under physiological conditions. Collectively, upon dysregulated NET release like in psoriasis, naRNA sensing may represent both a potential cause of disease and a new intervention target.

Tirap controls Mycobacterium tuberculosis phagosomal acidification.

Progression of tuberculosis is tightly linked to a disordered immune balance, resulting in inability of the host to restrict intracellular bacterial replication and its subsequent dissemination. The immune response is mainly characterized by an orchestrated recruitment of inflammatory cells secreting cytokines. This response results from the activation of innate immunity receptors that trigger downstream intracellular signaling pathways involving adaptor proteins such as the TIR-containing adaptor protein (Tirap). In humans, resistance to tuberculosis is associated with a loss-of-function in Tirap. Here, we explore how genetic deficiency in Tirap impacts resistance to Mycobacterium tuberculosis (Mtb) infection in a mouse model and ex vivo. Interestingly, compared to wild type littermates, Tirap heterozygous mice were more resistant to Mtb infection. Upon investigation at the cellular level, we observed that mycobacteria were not able to replicate in Tirap-deficient macrophages compared to wild type counterparts. We next showed that Mtb infection induced Tirap expression which prevented phagosomal acidification and rupture. We further demonstrate that the Tirap-mediated anti-tuberculosis effect occurs through a Cish-dependent signaling pathway. Our findings provide new molecular evidence about how Mtb manipulates innate immune signaling to enable intracellular replication and survival of the pathogen, thus paving the way for host-directed approaches to treat tuberculosis.

Resistance Mechanisms to Immune-Checkpoint Blockade in Cancer: Tumor-Intrinsic and -Extrinsic Factors.

Inhibition of immune regulatory checkpoints, such as CTLA-4 and the PD-1-PD-L1 axis, is at the forefront of immunotherapy for cancers of various histological types. However, such immunotherapies fail to control neoplasia in a significant proportion of patients. Here, we review how a range of cancer-cell-autonomous cues, tumor-microenvironmental factors, and host-related influences might account for the heterogeneous responses and failures often encountered during therapies using immune-checkpoint blockade. Furthermore, we describe the emerging evidence of how the strong interrelationship between the immune system and the host microbiota can determine responses to cancer therapies, and we introduce a concept by which prior or concomitant modulation of the gut microbiome could optimize therapeutic outcomes upon immune-checkpoint blockade.

Enterococcus hirae and Barnesiella intestinihominis Facilitate Cyclophosphamide-Induced Therapeutic Immunomodulatory Effects.

The efficacy of the anti-cancer immunomodulatory agent cyclophosphamide (CTX) relies on intestinal bacteria. How and which relevant bacterial species are involved in tumor immunosurveillance, and their mechanism of action are unclear. Here, we identified two bacterial species, Enterococcus hirae and Barnesiella intestinihominis that are involved during CTX therapy. Whereas E. hirae translocated from the small intestine to secondary lymphoid organs and increased the intratumoral CD8/Treg ratio, B. intestinihominis accumulated in the colon and promoted the infiltration of IFN-γ-producing γδT cells in cancer lesions. The immune sensor, NOD2, limited CTX-induced cancer immunosurveillance and the bioactivity of these microbes. Finally, E. hirae and B. intestinihominis specific-memory Th1 cell immune responses selectively predicted longer progression-free survival in advanced lung and ovarian cancer patients treated with chemo-immunotherapy. Altogether, E. hirae and B. intestinihominis represent valuable "oncomicrobiotics" ameliorating the efficacy of the most common alkylating immunomodulatory compound.

Use of 2,6-diaminopurine as a potent suppressor of UGA premature stop codons in cystic fibrosis.

Nonsense mutations are responsible for around 10% of cases of genetic diseases, including cystic fibrosis. 2,6-diaminopurine (DAP) has recently been shown to promote efficient readthrough of UGA premature stop codons. In this study, we show that DAP can correct a nonsense mutation in the Cftr gene in vivo in a new CF mouse model, in utero, and through breastfeeding, thanks, notably, to adequate pharmacokinetic properties. DAP turns out to be very stable in plasma and is distributed throughout the body. The ability of DAP to correct various endogenous UGA nonsense mutations in the CFTR gene and to restore its function in mice, in organoids derived from murine or patient cells, and in cells from patients with cystic fibrosis reveals the potential of such readthrough-stimulating molecules in developing a therapeutic approach. The fact that correction by DAP of certain nonsense mutations reaches a clinically relevant level, as judged from previous studies, makes the use of this compound all the more attractive.

Harnessing the Vnn1 pantetheinase pathway boosts short chain fatty acids production and mucosal protection in colitis.

OBJECTIVE: In the management of patients with IBD, there is a need to identify prognostic markers and druggable biological pathways to improve mucosal repair and probe the efficacy of tumour necrosis factor alpha biologics. Vnn1 is a pantetheinase that degrades pantetheine to pantothenate (vitamin B5, a precursor of coenzyme A (CoA) biosynthesis) and cysteamine. Vnn1 is overexpressed by inflamed colonocytes. We investigated its contribution to the tolerance of the intestinal mucosa to colitis-induced injury. DESIGN: We performed an RNA sequencing study on colon biopsy samples from patients with IBD stratified according to clinical severity and modalities of treatment. We generated the VIVA mouse transgenic model, which specifically overexpresses Vnn1 on intestinal epithelial cells and explored its susceptibility to colitis. We developed a pharmacological mimicry of Vnn1 overexpression by administration of Vnn1 derivatives. RESULTS: VNN1 overexpression on colonocytes correlates with IBD severity. VIVA mice are resistant to experimentally induced colitis. The pantetheinase activity of Vnn1 is cytoprotective in colon: it enhances CoA regeneration and metabolic adaptation of colonocytes; it favours microbiota-dependent production of short chain fatty acids and mostly butyrate, shown to regulate mucosal energetics and to be reduced in patients with IBD. This prohealing phenotype is recapitulated by treating control mice with the substrate (pantethine) or the products of pantetheinase activity prior to induction of colitis. In severe IBD, the protection conferred by the high induction of VNN1 might be compromised because its enzymatic activity may be limited by lack of available substrates. In addition, we identify the elevation of indoxyl sulfate in urine as a biomarker of Vnn1 overexpression, also detected in patients with IBD. CONCLUSION: The induction of Vnn1/VNN1 during colitis in mouse and human is a compensatory mechanism to reinforce the mucosal barrier. Therefore, enhancement of vitamin B5-driven metabolism should improve mucosal healing and might increase the efficacy of anti-inflammatory therapy.

NOD1 sensing of house dust mite-derived microbiota promotes allergic experimental asthma.

BACKGROUND: Asthma severity has been linked to exposure to gram-negative bacteria from the environment that are recognized by NOD1 receptor and are present in house dust mite (HDM) extracts. NOD1 polymorphism has been associated with asthma. OBJECTIVE: We sought to evaluate whether either host or HDM-derived microbiota may contribute to NOD1-dependent disease severity. METHODS: A model of HDM-induced experimental asthma was used and the effect of NOD1 deficiency was evaluated. Contribution of host microbiota was evaluated by fecal transplantation. Contribution of HDM-derived microbiota was assessed by 16S ribosomal RNA sequencing, mass spectrometry analysis, and peptidoglycan depletion of the extracts. RESULTS: In this model, loss of the bacterial sensor NOD1 and its adaptor RIPK2 improved asthma features. Such inhibitory effect was not related to dysbiosis caused by NOD1 deficiency, as shown by fecal transplantation of Nod1-deficient microbiota to wild-type germ-free mice. The 16S ribosomal RNA gene sequencing and mass spectrometry analysis of HDM allergen, revealed the presence of some muropeptides from gram-negative bacteria that belong to the Bartonellaceae family. While such HDM-associated muropeptides were found to activate NOD1 signaling in epithelial cells, peptidoglycan-depleted HDM had a decreased ability to instigate asthma in vivo. CONCLUSIONS: These data show that NOD1-dependent sensing of HDM-associated gram-negative bacteria aggravates the severity of experimental asthma, suggesting that inhibiting the NOD1 signaling pathway may be a therapeutic approach to treating asthma.

MAVS deficiency induces gut dysbiotic microbiota conferring a proallergic phenotype.

Prominent changes in the gut microbiota (referred to as "dysbiosis") play a key role in the development of allergic disorders, but the underlying mechanisms remain unknown. Study of the delayed-type hypersensitivity (DTH) response in mice contributed to our knowledge of the pathophysiology of human allergic contact dermatitis. Here we report a negative regulatory role of the RIG-I-like receptor adaptor mitochondrial antiviral signaling (MAVS) on DTH by modulating gut bacterial ecology. Cohousing and fecal transplantation experiments revealed that the dysbiotic microbiota of Mavs-/- mice conferred a proallergic phenotype that is communicable to wild-type mice. DTH sensitization coincided with increased intestinal permeability and bacterial translocation within lymphoid organs that enhanced DTH severity. Collectively, we unveiled an unexpected impact of RIG-I-like signaling on the gut microbiota with consequences on allergic skin disease outcome. Primarily, these data indicate that manipulating the gut microbiota may help in the development of therapeutic strategies for the treatment of human allergic skin pathologies.

Role of NLRs in the Regulation of Type I Interferon Signaling, Host Defense and Tolerance to Inflammation.

Type I interferon signaling contributes to the development of innate and adaptive immune responses to either viruses, fungi, or bacteria. However, amplitude and timing of the interferon response is of utmost importance for preventing an underwhelming outcome, or tissue damage. While several pathogens evolved strategies for disturbing the quality of interferon signaling, there is growing evidence that this pathway can be regulated by several members of the Nod-like receptor (NLR) family, although the precise mechanism for most of these remains elusive. NLRs consist of a family of about 20 proteins in mammals, which are capable of sensing microbial products as well as endogenous signals related to tissue injury. Here we provide an overview of our current understanding of the function of those NLRs in type I interferon responses with a focus on viral infections. We discuss how NLR-mediated type I interferon regulation can influence the development of auto-immunity and the immune response to infection.

The regenerating family member 3 ß instigates IL-17A-mediated neutrophil recruitment downstream of NOD1/2 signalling for controlling colonisation resistance independently of microbiota community structure.

OBJECTIVE: Loss of the Crohn's disease predisposing NOD2 gene results in an intestinal microenvironment conducive for colonisation by attaching-and-effacing enteropathogens. However, it remains elusive whether it relies on the intracellular recruitment of the serine-threonine kinase RIPK2 by NOD2, a step that is required for its activation of the transcription factor NF-κB. DESIGN: Colonisation resistance was evaluated in wild type and mutant mice, as well as in ex-germ-free (ex-GF) mice which were colonised either with faeces from Ripk2-deficient mice or with bacteria with similar preferences for carbohydrates to those acquired by the pathogen. The severity of the mucosal pathology was quantified at several time points postinfection by using a previously established scoring. The community resilience in response to infection was evaluated by 16S ribosomal RNA gene sequence analysis. The control of pathogen virulence was evaluated by monitoring the secretion of Citrobacter-specific antibody response in the faeces. RESULTS: Primary infection was similarly outcompeted in ex-GF Ripk2-deficient and control mice, demonstrating that the susceptibility to infection resulting from RIPK2 deficiency cannot be solely attributed to specific microbiota community structures. In contrast, delayed clearance of Citrobacter rodentium and exacerbated histopathology were preceded by a weakened propensity of intestinal macrophages to afford innate lymphoid cell activation. This tissue protection unexpectedly required the regenerating family member 3ß by instigating interleukin (IL) 17A-mediated neutrophil recruitment to the intestine and subsequent phosphorylation of signal transducer and activator of transcription 3. CONCLUSIONS: These results unveil a previously unrecognised mechanism that efficiently protects from colonisation by diarrhoeagenic bacteria early in infection.

Card9 mediates susceptibility to intestinal pathogens through microbiota modulation and control of bacterial virulence.

OBJECTIVE: In association with innate and adaptive immunity, the microbiota controls the colonisation resistance against intestinal pathogens. Caspase recruitment domain 9 (CARD9), a key innate immunity gene, is required to shape a normal gut microbiota. Card9-/- mice are more susceptible to the enteric mouse pathogen Citrobacter rodentium that mimics human infections with enteropathogenic and enterohaemorrhagic Escherichia coli. Here, we examined how CARD9 controls C. rodentium infection susceptibility through microbiota-dependent and microbiota-independent mechanisms. DESIGN: C. rodentium infection was assessed in conventional and germ-free (GF) wild-type (WT) and Card9-/- mice. To explore the impact of Card9-/-microbiota in infection susceptibility, GF WT mice were colonised with WT (WT→GF) or Card9-/- (Card9-/- →GF) microbiota before C. rodentium infection. Microbiota composition was determined by 16S rDNA gene sequencing. Inflammation severity was determined by histology score and lipocalin level. Microbiota-host immune system interactions were assessed by quantitative PCR analysis. RESULTS: CARD9 controls pathogen virulence in a microbiota-independent manner by supporting a specific humoral response. Higher susceptibility to C. rodentium-induced colitis was observed in Card9-/- →GF mice. The microbiota of Card9-/- mice failed to outcompete the monosaccharide-consuming C. rodentium, worsening the infection severity. A polysaccharide-enriched diet counteracted the ecological advantage of C. rodentium and the defective pathogen-specific antibody response in Card9-/- mice. CONCLUSIONS: CARD9 modulates the susceptibility to intestinal infection by controlling the pathogen virulence in a microbiota-dependent and microbiota-independent manner. Genetic susceptibility to intestinal pathogens can be overridden by diet intervention that restores humoural immunity and a competing microbiota.

Genetic Factors Interact With Tobacco Smoke to Modify Risk for Inflammatory Bowel Disease in Humans and Mice.

BACKGROUND & AIMS: The role of tobacco smoke in the etiology of inflammatory bowel disease (IBD) is unclear. We investigated interactions between genes and smoking (gene-smoking interactions) that affect risk for Crohn's disease (CD) and ulcerative colitis (UC) in a case-only study of patients and in mouse models of IBD. METHODS: We used 55 Immunochip-wide datasets that included 19,735 IBD cases (10,856 CD cases and 8879 UC cases) of known smoking status. We performed 3 meta-analyses each for CD, UC, and IBD (CD and UC combined), comparing data for never vs ever smokers, never vs current smokers, and never vs former smokers. We studied the effects of exposure to cigarette smoke in Il10-/- and Nod2-/- mice, as well as in Balb/c mice without disruption of these genes (wild-type mice). Mice were exposed to the smoke of 5 cigarettes per day, 5 days a week, for 8 weeks, in a ventilated smoking chamber, or ambient air (controls). Intestines were collected and analyzed histologically and by reverse transcription polymerase chain reaction. RESULTS: We identified 64 single nucleotide polymorphisms (SNPs) for which the association between the SNP and IBD were modified by smoking behavior (meta-analysis Wald test P < 5.0 × 10-5; heterogeneity Cochrane Q test P > .05). Twenty of these variants were located within the HLA region at 6p21. Analysis of classical HLA alleles (imputed from SNP genotypes) revealed an interaction with smoking. We replicated the interaction of a variant in NOD2 with current smoking in relation to the risk for CD (frameshift variant fs1007insC; rs5743293). We identified 2 variants in the same genomic region (rs2270368 and rs17221417) that interact with smoking in relation to CD risk. Approximately 45% of the SNPs that interact with smoking were in close vicinity (≤1 Mb) to SNPs previously associated with IBD; many were located near or within genes that regulate mucosal barrier function and immune tolerance. Smoking modified the disease risk of some variants in opposite directions for CD vs UC. Exposure of Interleukin 10 (il10)-deficient mice to cigarette smoke accelerated development of colitis and increased expression of interferon gamma in the small intestine compared to wild-type mice exposed to smoke. NOD2-deficient mice exposed to cigarette smoke developed ileitis, characterized by increased expression of interferon gamma, compared to wild-type mice exposed to smoke. CONCLUSIONS: In an analysis of 55 Immunochip-wide datasets, we identified 64 SNPs whose association with risk for IBD is modified by tobacco smoking. Gene-smoking interactions were confirmed in mice with disruption of Il10 and Nod2-variants of these genes have been associated with risk for IBD. Our findings from mice and humans revealed that the effects of smoking on risk for IBD depend on genetic variants.

LppM impact on the colonization of macrophages by Mycobacterium tuberculosis.

Mycobacterium tuberculosis produces several bacterial effectors impacting the colonization of phagocytes. Here, we report that the putative lipoprotein LppM hinders phagocytosis by macrophages in a toll-like receptor 2-dependent manner. Moreover, recombinant LppM is able to functionally complement the phenotype of the mutant, when exogenously added during macrophage infection. LppM is also implicated in the phagosomal maturation, as a lppM deletion mutant is more easily addressed towards the acidified compartments of the macrophage than its isogenic parental strain. In addition, this mutant was affected in its ability to induce the secretion of pro-inflammatory chemokines, interferon-gamma-inducible protein-10, monocyte chemoattractant protein-1 and macrophage inflammatory protein-1α. Thus, our results describe a new mycobacterial protein involved in the early trafficking of the tubercle bacillus and its manipulation of the host immune response.

Hemidesmosome integrity protects the colon against colitis and colorectal cancer.

OBJECTIVE: Epidemiological and clinical data indicate that patients suffering from IBD with long-standing colitis display a higher risk to develop colorectal high-grade dysplasia. Whereas carcinoma invasion and metastasis rely on basement membrane (BM) disruption, experimental evidence is lacking regarding the potential contribution of epithelial cell/BM anchorage on inflammation onset and subsequent neoplastic transformation of inflammatory lesions. Herein, we analyse the role of the α6ß4 integrin receptor found in hemidesmosomes that attach intestinal epithelial cells (IECs) to the laminin-containing BM. DESIGN: We developed new mouse models inducing IEC-specific ablation of α6 integrin either during development (α6ΔIEC) or in adults (α6ΔIEC-TAM). RESULTS: Strikingly, all α6ΔIEC mutant mice spontaneously developed long-standing colitis, which degenerated overtime into infiltrating adenocarcinoma. The sequence of events leading to disease onset entails hemidesmosome disruption, BM detachment, IL-18 overproduction by IECs, hyperplasia and enhanced intestinal permeability. Likewise, IEC-specific ablation of α6 integrin induced in adult mice (α6ΔIEC-TAM) resulted in fully penetrant colitis and tumour progression. Whereas broad-spectrum antibiotic treatment lowered tissue pathology and IL-1ß secretion from infiltrating myeloid cells, it failed to reduce Th1 and Th17 response. Interestingly, while the initial intestinal inflammation occurred independently of the adaptive immune system, tumourigenesis required B and T lymphocyte activation. CONCLUSIONS: We provide for the first time evidence that loss of IECs/BM interactions triggered by hemidesmosome disruption initiates the development of inflammatory lesions that progress into high-grade dysplasia and carcinoma. Colorectal neoplasia in our mouse models resemble that seen in patients with IBD, making them highly attractive for discovering more efficient therapies.

Enterocyte loss of polarity and gut wound healing rely upon the F-actin-severing function of villin.

Efficient wound healing is required to maintain the integrity of the intestinal epithelial barrier because of its constant exposure to a large variety of environmental stresses. This process implies a partial cell depolarization and the acquisition of a motile phenotype that involves rearrangements of the actin cytoskeleton. Here we address how polarized enterocytes harboring actin-rich apical microvilli undergo extensive cell remodeling to drive injury repair. Using live imaging technologies, we demonstrate that enterocytes in vitro and in vivo rapidly depolarize their microvilli at the wound edge. Through its F-actin-severing activity, the microvillar actin-binding protein villin drives both apical microvilli disassembly in vitro and in vivo and promotes lamellipodial extension. Photoactivation experiments indicate that microvillar actin is mobilized at the lamellipodium, allowing optimal migration. Finally, efficient repair of colonic mechanical injuries requires villin severing of F-actin, emphasizing the importance of villin function in intestinal homeostasis. Thus, villin severs F-actin to ensure microvillus depolarization and enterocyte remodeling upon injury. This work highlights the importance of specialized apical pole disassembly for the repolarization of epithelial cells initiating migration.

CCL17 production by dendritic cells is required for NOD1-mediated exacerbation of allergic asthma.

RATIONALE: Pattern recognition receptors are attractive targets for vaccine adjuvants, and polymorphisms of the innate receptor NOD1 have been associated with allergic asthma. OBJECTIVES: To elucidate whether NOD1 agonist may favor allergic asthma in humans through activation of dendritic cells, and to evaluate the mechanisms involved using an in vivo model. METHODS: NOD1-primed dendritic cells from allergic and nonallergic donors were characterized in vitro on their phenotype, cytokine secretion, and Th2 polarizing ability. The in vivo relevance was examined in experimental allergic asthma, and the mechanisms were assessed using transfer of NOD1-conditioned dendritic cells from wild-type or CCL17-deficient mice. MEASUREMENTS AND MAIN RESULTS: NOD1 priming of human dendritic cells promoted a Th2 polarization profile that involved the production of CCL17 and CCL22 in nonallergic subjects but only CCL17 in allergic patients, without requiring allergen costimulation. Moreover, NOD1-primed dendritic cells from allergic donors exhibited enhanced maturation that led to abnormal CCL22 and IL-10 secretion compared with nonallergic donors. In mice, systemic NOD1 ligation exacerbated allergen-induced experimental asthma by amplifying CCL17-mediated Th2 responses in the lung. NOD1-mediated sensitization of purified murine dendritic cells enhanced production of CCL17 and CCL22, but not of thymic stromal lymphopoietin and IL-33, in vitro. Consistently, adoptive transfer of NOD1-conditioned dendritic cells exacerbated the Th2 pulmonary response in a CCL17-dependent manner in vivo. CONCLUSIONS: Data from this study unveil a deleterious role of NOD1 in allergic asthma through direct induction of CCL17 by dendritic cells, arguing for a need to address vaccine formulation safety issues related to allergy.

Pseudomonas aeruginosa type-3 secretion system dampens host defense by exploiting the NLRC4-coupled inflammasome.

RATIONALE: Pseudomonas aeruginosa, a major problem pathogen responsible for severe infections in critically ill patients, triggers, through a functional type-3 secretion system (T3SS), the activation of an intracellular cytosolic sensor of innate immunity, NLRC4. Although the NLRC4-inflammasome-dependent response contributes to increased clearance of intracellular pathogens, it seems that NLRC4 inflammasome activation decreases the clearance of P. aeruginosa, a mainly extracellular pathogen. OBJECTIVES: We sought to determine the underlying mechanisms of this effect of the activation of NLRC4 by P. aeruginosa. METHODS: We established acute lung injury in wild-type and Nlrc4(-/-) mice using sublethal intranasal inocula of P. aeruginosa strain CHA expressing or not a functional T3SS. We studied 96-hour survival, lung injury, bacterial clearance from the lungs, cytokine secretion in bronchoalveolar lavage, lung antimicrobial peptide expression by quantitative polymerase chain reaction, and flow cytometry analysis of lung cells. MEASUREMENTS AND MAIN RESULTS: Nlrc4(-/-) mice showed enhanced bacterial clearance and decreased lung injury contributing to increased survival against extracellular P. aeruginosa strain expressing a functional T3SS. The mechanism involved decreased NLRC4-inflammasome-driven IL-18 secretion attenuating lung injury caused by excessive neutrophil recruitment. Additionally, in the lungs of Nlrc4(-/-) mice secretion of IL-17 by innate immune cells was increased and responsible for increased expression of lung epithelial antimicrobial peptides. Furthermore, IL-18 secretion was found to repress IL-17 and IL-17-driven lung antimicrobial peptide expression. CONCLUSIONS: We report a new role of the T3SS apparatus itself, independently of exotoxin translocation. Through NLRC4 inflammasome activation, the T3SS promotes IL-18 secretion, which dampens a beneficial IL-17-mediated antimicrobial host response.

Lactobacillus acidophilus modulates intestinal pain and induces opioid and cannabinoid receptors.

Abdominal pain is common in the general population and, in patients with irritable bowel syndrome, is attributed to visceral hypersensitivity. We found that oral administration of specific Lactobacillus strains induced the expression of mu-opioid and cannabinoid receptors in intestinal epithelial cells, and mediated analgesic functions in the gut-similar to the effects of morphine. These results suggest that the microbiology of the intestinal tract influences our visceral perception, and suggest new approaches for the treatment of abdominal pain and irritable bowel syndrome.

Nod-like receptor pyrin domain-containing protein 6 (NLRP6) controls epithelial self-renewal and colorectal carcinogenesis upon injury.

The colonic epithelium self-renews every 3 to 5 d, but our understanding of the underlying processes preserving wound healing from carcinogenesis remains incomplete. Here, we demonstrate that Nod-like receptor pyrin domain-containing protein 6 (NLRP6) suppresses inflammation and carcinogenesis by regulating tissue repair. NLRP6 was primarily produced by myofibroblasts within the stem-cell niche in the colon. Although NLRP6 expression was lowered in diseased colon, NLRP6-deficient mice were highly susceptible to experimental colitis. Upon injury, NLRP6 deficiency deregulated regeneration of the colonic mucosa and processes of epithelial proliferation and migration. Consistently, absence of NLRP6 accelerated colitis-associated tumor growth in mice. A gene-ontology analysis on a whole-genome expression profiling revealed a link between NLRP6 and self-renewal of the epithelium. Collectively, the integrity of the epithelial barrier is preserved by NLRP6 that may be manipulated to develop drugs capable of preventing adenoma formation in inflammatory bowel diseases.