The NOD-like receptors (NLRs) are an important part of the innate immune system, capable of recognizing or sensing pathogens or cell homeostasis imbalance, and play an important role in pathogen ...removal and body protection. NOD2 is one of the important members in NLR family. In present study, NOD2 was cloned and identified from Nile tilapia (Oreochromis niloticus). The Nile tilapia NOD2 ORF was 2961 bp in length encoding a peptide of 986 amino acid residues with a typical structure of CARD-CARD-NACHT-LRRs. Phylogenetic and synteny analysis illustrated that NOD2 was highly conserved between different species. Tissue distribution of NOD2 mRNA showed that it was widely expressed and highly expressed in immune-related tissues (blood, head kidney, liver and spleen). Post Streptococcus agalactiae challenge, NOD2 mRNA expression levels were altered in immune organs or tissues (liver, head kidney, spleen and blood). Subcellular localization of Nile tilapia NOD2 illuminated that it was an intracellular PRR like mammalian counterparts, most of which was co-localized with mitochondria, and a few of which was co-localized with endosome and endoplasmic reticulum. In addition, Nile tilapia NOD2-mediated signaling pathway and ligand recognition were also studied, it illuminated that Nile tilapia NOD2 could recognize MDP intracellularly and activate NF-κB signaling pathway. In conclusion, this study indicated that the Nile tilapia NOD2 was conservative and had a significant role in innate immune defense against bacterial infection.
•The recognition mechanism between fish NOD2 and its specific ligand was determined.•Nile tilapia NOD2 could significantly activate the NF-κB, type I interferon and MAPK signaling pathway.•Nile tilapia NOD2 was mainly co-localized with mitochondria.•NOD2 was highly conserved between different species.
Pattern recognition receptor nucleotide-binding oligomerization domain 2 (NOD2) is well investigated in immunity, but its expression and function in platelets has never been explored.
Using reverse ...transcription polymerase chain reaction and Western blot, we show that both human and mouse platelets express NOD2, and its agonist muramyl dipeptide induced NOD2 activation as evidenced by receptor dimerization. NOD2 activation potentiates platelet aggregation and secretion induced by low concentrations of thrombin or collagen, and clot retraction, as well. These potentiating effects of muramyl dipeptide were not seen in platelets from NOD2-deficient mice. Plasma from septic patients also potentiates platelet aggregation induced by thrombin or collagen NOD2 dependently. Using intravital microscopy, we found that muramyl dipeptide administration accelerated in vivo thrombosis in a FeCl3-injured mesenteric arteriole thrombosis mouse model. Platelet depletion and transfusion experiments confirmed that NOD2 from platelets contributes to the in vivo thrombosis in mice. NOD2 activation also accelerates platelet-dependent hemostasis. We further found that platelets express receptor-interacting protein 2, and provided evidence suggesting that mitogen activated-protein kinase and nitric oxide/soluble guanylyl cyclase/cGMP/protein kinase G pathways downstream of receptor-interacting protein mediate the role of NOD2 in platelets. Finally, muramyl dipeptide stimulates proinflammatory cytokine interleukin-1β maturation and accumulation in human and mouse platelets NOD2 dependently.
NOD2 is expressed in platelets and functions in platelet activation and arterial thrombosis, possibly during infection. To our knowledge, this is the first study on NOD-like receptors in platelets that link thrombotic events to inflammation.
The nucleotide-binding oligomerization domain 2 (NOD2) has been identified as an important sensor for microorganic invasion in both mammals and teleost fishes. In this study, two splicing variants of ...NOD2 (NOD2-v1 and NOD2-v2) were identified as truncating the functional domains of wild-type NOD2 in the teleost fish Schizothorax prenanti. NOD2-v1 included an intron sequence that terminated within the third leucine-rich repeat (LRR) domain, while NOD2-v2 incorporated an insertion of one and half intron sequences and truncated within the second caspase activation and recruitment domain (CARD). NOD2, NOD2-v1 and NOD2-v2 genes were ubiquitously expressed. All three genes positively responded to exposure of Aeromonas hydrophila and lipopolysaccharide stimulation in varying degrees. Using luciferase activity assays in HEK293T cells, our results revealed that NOD2 activated the NF-κB signal and recognized muramyl dipeptide (MDP). NOD2-v1 exhibited deficiency in the LRR domains and could not sense MDP, but maintained the ability to activate NF-κB and enhanced NOD2-mediated MDP recognition. Given the significant change to the functional structure, NOD2-v2 lost its capacity for NF-κB activation, but interestingly repressed NOD2-mediated MDP sensing and NF-κB activation, and even NOD2-v1-induced NF-κB activation. Altogether, our study reveals a novel pattern of signal regulation by splicing variants in teleost fishes.
•Human-like NOD2 splicing variants were firstly identified in S. prenanti, named NOD2, NOD2-v1 and NOD2-v2.•NOD2 and NOD2-v1 but NOD2-v2 can induce NF-κB activation.•NOD2-v1 and NOD2-v2 lose the function of sensing specific ligand MDP.•NOD2-v2 represses NOD2-mediated MDP sensing and NF-κB activation, and even NOD2-v1-induced NF-κB activation.
Irregular mitochondria structure and reduced ATP in some patients with IBD suggest that metabolic stress contributes to disease. Loss-of-function mutation in the nucleotide-binding oligomerization ...domain (NOD)-2 gene is a major susceptibility trait for IBD. Hence, we assessed if loss of NOD2 further impairs the epithelial barrier function instigated by disruption of mitochondrial ATP synthesis via the hydrogen ionophore dinitrophenol (DNP). NOD2 protein (virtually undetectable in epithelia under basal conditions) was increased in T84 (human colon cell line) cells treated with noninvasive
+ DNP (16 h). Increased intracellular bacteria in wild-type (WT) and NOD2 knockdown (KD) cells and colonoids from NOD2
mice were mediated by reactive oxygen species (ROS) and the MAPK ERK1/2 pathways as determined by cotreatment with the antioxidant mitoTEMPO and the ERK inhibitor U0126: ROS was upstream of ERK1/2 activation. Despite increased
in DNP-treated NOD2 KD compared with WT cells, there were no differences in the internalization of fluorescent inert beads or dead
particles. This suggests that lack of killing in the NOD2 KD cells was responsible for the increased numbers of viable intracellular bacteria; a conclusion supported by evidence of reduced autophagy in NOD2 KD T84 epithelia. Thus, in a two-hit hypothesis, decreased barrier function due to dysfunctional mitochondrial is amplified by lack of NOD2 in transporting enterocytes: subsequently, greater numbers of bacteria entering the mucosa would be a significant inflammatory threat especially since individuals with NOD2 mutations have compromised macrophage and Paneth cell responses to bacteria.
Increased internalization of bacteria by epithelia with dysfunctional mitochondria (reduced ATP) is potentiated if the cells lack nucleotide-binding oligomerization domain 2 (NOD2), mutations in which are inflammatory bowel disease-susceptibility traits. Uptake of bacteria was dependent on reactive oxygen species and MAP-kinase activity, and the increased viable intracellular bacteria in NOD2
cells likely reflect a reduced ability to recognize and kill bacteria. Thus a significant barrier defect occurs with NOD2 deficiency in conjunction with metabolic stress that could contribute to inflammation.
Digestive and cardiodigestive forms of Chagas' disease are observed in 2% to 27% of the patients, depending on their geographic location, Trypanosoma cruzi strain and immunopathological responses. ...The aim of this work was to evaluate the role of NOD2 innate immune receptor in the pathogenesis of the digestive system in Chagas' disease. Patients with digestive form of the disease showed lower mRNA expression of NOD2, higher expression of RIP2 and α-defensin 6, compared to indeterminate form, detected by Real-time PCR in peripheral blood mononuclear cells. In addition, there was a negative correlation between the expression of NOD2 and the degree of dilation of the esophagus, sigmoid and rectum in those patients. The infection of NOD2-/- mice with T. cruzi strain isolated from the digestive patient induced a decrease in intestinal motility. Histopathological analysis of the colon and jejunum of NOD2-/- and wild type C57BL/6 animals revealed discrete inflammatory foci during the acute phase of infection. Interestingly, during the chronic phase of the infection there was inflammation and hypertrophy of the longitudinal and circular muscular layer more pronounced in the colon and jejunum from NOD2-/- animals, when compared to wild type C57BL/6 mice. Together, our results suggest that NOD2 plays a protective role against the development of digestive form of Chagas' disease.
Reduced microbial diversity has been associated with inflammatory bowel disease (IBD) and probiotic bacteria have been proposed for its prevention and/or treatment. Nevertheless, comparative studies ...of strains of the same subspecies for specific health benefits are scarce. Here we compared two Bifidobacterium longum ssp. longum strains for their capacity to prevent experimental colitis.
Immunomodulatory properties of nine probiotic bifidobacteria were assessed by stimulation of murine splenocytes. The immune responses to B. longum ssp. longum CCM 7952 (Bl 7952) and CCDM 372 (Bl 372) were further characterized by stimulation of bone marrow-derived dendritic cell, HEK293/TLR2 or HEK293/NOD2 cells. A mouse model of dextran sulphate sodium (DSS)-induced colitis was used to compare their beneficial effects in vivo.
The nine bifidobacteria exhibited strain-specific abilities to induce cytokine production. Bl 372 induced higher levels of both pro- and anti-inflammatory cytokines in spleen and dendritic cell cultures compared to Bl 7952. Both strains engaged TLR2 and contain ligands for NOD2. In a mouse model of DSS-induced colitis, Bl 7952, but not Bl 372, reduced clinical symptoms and preserved expression of tight junction proteins. Importantly, Bl 7952 improved intestinal barrier function as demonstrated by reduced FITC-dextran levels in serum.
We have shown that Bl 7952, but not Bl 372, protected mice from the development of experimental colitis. Our data suggest that although some immunomodulatory properties might be widespread among the genus Bifidobacterium, others may be rare and characteristic only for a specific strain. Therefore, careful selection might be crucial in providing beneficial outcome in clinical trials with probiotics in IBD.
The mammalian commensal gut microbiota is highly diverse and displays an individual-specific composition determined by host genotype and environmental factors. The temporal development of ...host-microbial homeostasis in the digestive tract is recognised as a major function of the immune system. However, the underlying cellular and molecular mechanisms are just beginning to come to light. Nucleotide-binding, oligomerisation domain 2 (NOD2) recognises bacterial muramyl dipeptide and is regarded as a pivotal sensor molecule of the intestinal barrier. The aim of this study was to investigate its influence on the development and composition of the intestinal microbiota using a Nod2-deficient mouse model.
The dynamics of faecal and ileal microbial composition were investigated in Nod2(+/+)and Nod2(-/-) mice on a C57BL/6J background. We assessed microbial diversity and composition using 16S ribosomal RNA gene-based clone library sequencing and high throughput pyrosequencing and quantified the observed changes by real-time PCR. Changes in the major bacterial phyla were investigated in human samples by quantitative real-time PCR.
We found that adult Nod2-deficient mice display a substantially altered microbial community structure and a significantly elevated bacterial load in their faeces and terminal ileum compared to their wild-type counterparts. Interestingly, we demonstrate that these findings are also present in weaning mice, indicating a profound influence of Nod2 on the early development and composition of the intestinal microbiota. We demonstrate that NOD2 genotypes also influence the microbial composition in humans.
Our results point to an essential role of Nod2 for the temporal development and composition of the host microbiota, both in mice and in humans, which may contribute to the complex role of NOD2 for the aetiopathogenesis of Crohn's disease.
NOD1 {nucleotide-binding oligomerization domain 1; NLRC NOD-LRR (leucine-rich repeat) family with CARD (caspase recruitment domain) 1} and NOD2 (NLRC2) are among the most prominent members of the NLR ...(NOD-LRR) family -proteins that contain nucleotide-binding NACHT domains and receptor-like LRR domains. With over 20 members identified in humans, NLRs represent important components of the mammalian innate immune system, serving as intracellular receptors for pathogens and for endogenous molecules elaborated by tissue injury. NOD1 and NOD2 proteins operate as microbial sensors through the recognition of specific PG (peptidoglycan) constituents of bacteria. Upon activation, these NLR family members initiate signal transduction mechanisms that include stimulation of NF-κB (nuclear factor-κB), stress kinases, IRFs (interferon regulatory factors) and autophagy. Hereditary polymorphisms in the genes encoding NOD1 and NOD2 have been associated with an increasing number of chronic inflammatory diseases. In fact, potential roles for NOD1 and NOD2 in inflammatory disorders have been revealed by investigations using a series of animal models. In the present review, we describe recent experimental findings associating NOD1 and NOD2 with various autoimmune and chronic inflammatory disorders, and we discuss prospects for development of novel therapeutics targeting these NLR family proteins.
The intracellular sensor Nod2 is activated in response to bacteria, and the impairment of this response is linked to Crohn's disease. However, the function of Nod2 in host defense remains poorly ...understood. We found that
Nod2
−/− mice exhibited impaired intestinal clearance of
Citrobacter rodentium, an enteric bacterium that models human infection by pathogenic
Escherichia coli. The increased bacterial burden was preceded by reduced CCL2 chemokine production, inflammatory monocyte recruitment, and Th1 cell responses in the intestine. Colonic stromal cells, but not epithelial cells or resident CD11b
+ phagocytic cells, produced CCL2 in response to
C. rodentium in a Nod2-dependent manner. Unlike resident phagocytic cells, inflammatory monocytes produced IL-12, a cytokine that induces adaptive immunity required for pathogen clearance. Adoptive transfer of Ly6C
hi monocytes restored the clearance of the pathogen in infected
Ccr2
−/− mice. Thus, Nod2 mediates CCL2-CCR2-dependent recruitment of inflammatory monocytes, which is important in promoting bacterial eradication in the intestine.
► Nod2
−/− mice exhibit impaired
C. rodentium clearance and CCL2 production ► Reduced CCL2 was associated with impaired Gr1
+ monocyte influx to the colon ► Intestinal stromal cells were the main producers of CCL2 ► Adoptive transfer of monocytes restored the clearance of
C. rodentium in Ccr2
−/− mice
Chlamydial species are common intracellular parasites that cause various diseases, mainly characterized by persistent infection, which lead to inflammatory responses modulated by pattern recognition ...receptors (PRRs). The best understood PRRs are the extracellular Toll-like receptors, but recent significant advances have focused on two important proteins, NOD1 and NOD2, which are members of the intracellular nucleotide-binding oligomerization domain receptor family and are capable of triggering the host innate immune signaling pathways. This results in the production of pro-inflammatory cytokines, which is vital for an adequate host defense against intracellular chlamydial infection. NOD1/2 ligands are known to derive from peptidoglycan, and the latest research has resolved the paradox of whether chlamydial species possess this bacterial cell wall component; this finding is likely to promote in-depth investigations into the interaction between the NOD proteins and chlamydial pathogens. In this review, we summarize the basic characteristics and signal transduction functions of NOD1 and NOD2 and highlight the new research on the roles of NOD1 and NOD2 in the host defense against chlamydial infection.
This review summarizes the basic characteristics and signal transduction functions of NOD1 and NOD2 and highlights the roles of NOD1 and NOD2 in the host defense against chlamydial infection.
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