Intracellular Pattern Recognition Receptors Research Articles

Clinical characteristics of patients with NOD2 high risk allele variants

The nucleotide-binding oligomerization domain 2 (NOD2) is an intracellular pattern recognition receptor. NOD2 gene variants are associated with inflammatory bowel disease (IBD) as well as Blau and Yao syndromes. IBD is associated with extraintestinal manifestations, including autoinflammation, which may be the presenting symptoms. The clinical symptoms may depend on the specific gene variants. This retrospective study was undertaken to examine the clinical manifestations of patients with pathogenic NOD2 variants in our institution. This study was approved by our Institutional Review Board.From January 2020 to February 2022, we identified 203 patients that had genetic testing that included the NOD2 gene. Forty-one had high risk alleles variants (HRA) that segregated in three mutations: HRA1 c.2104C>T (n = 24); HRA2 c.3019dup (n = 11); and HRA3 c.2722G > C (n = 6).The reported age of initial symptoms ranged from 21 days to 62 years, with most patients being < 12 years of age. All three variants had clinical evidence of immune dysregulation. Common manifestations (listed respectively for the three variants) included asthma (HRA1 = 21%, HRA2 = 9%, HRA3 = 33%), reflux/gastritis (29%, 18%, 33%), irritable bowel syndrome (21%, 18%, 7%), chronic diarrhea (8%, 18%, 0%), IBD (13%, 27%, 50%), recurrent fevers (38%, 45%, 0%), cytokine storm (4%, 9%, 17%), arthralgia/arthritis (33%, 45%, 17%), skin rashes (42%, 36%, 0%), malignancy (13%, 0%, 17%), bacterial infections (38%, 36%, 67%), fungal infections (4%, 18%, 0%), other autoimmune (21%, 9%, 0%), autoinflammatory syndromes (13%, 45%, 0%). Autoinflammatory/autoimmune disorders diagnosed within the cohort included PFAPA (HRA1 = 0, HRA2 = 1, HRA3 = 0), Yao syndrome (1, 0, 0), juvenile idiopathic arthritis (1, 1, 0), SAPHO syndrome (0, 1, 0), relapsing polychondritis (0,1,0), lupus (0,1,0), Sjogren’s syndrome (0,1,0), Raynaud’s syndrome (1, 0, 0), vitiligo (0, 1, 0), type I diabetes (2, 0, 0) and hypothyroidism (2, 1, 0). Detailed information is in table 1.Our chart review demonstrates a broad spectrum of extraintestinal clinical manifestations of patients with NOD2 variants which do not always include IBD. NOD2 is important in the protection of infection and autoinflammation and its role in human disease needs to be examined further.

Unexpected Need of the Epidermal Growth Factor Receptor Tyrosine Kinase Activity for Signaling by Intracellular Pattern Recognition Receptors of Nucleic Acids.

Many pattern recognition receptors in mammalian cells initiate signaling processes that culminate in mounting an innate protective response mediated by induced synthesis of a large number of proteins including type I interferons and other cytokines. Many of these receptors are not located on the plasma membrane but on the membranes of intracellular organelles such as endosomes, mitochondria, and the endoplasmic reticulum; they primarily recognize microbial or cellular nucleic acids. In the course of biochemical analyses of the signaling pathways triggered by these receptors, we discovered that they require tyrosine phosphorylation by the protein kinase activity of the epidermal growth factor receptor (EGFR), which is located not only on the plasma membrane but also on the intracellular membranes. Here, we discuss how specific members of this family of receptors, such as TLR3, TLR9, or STING, interact with EGFR and other protein tyrosine kinases and what are the functional consequences of their post-translational modifications. The article highlights an unexpected functional link between a growth factor receptor and cellular innate immune response.

Pattern-recognition receptors in endometriosis: A narrative review.

Endometriosis is closely associated with ectopic focal inflammation and immunosuppressive microenvironment. Multiple types of pattern recognition receptors (PRRs) are present in the innate immune system, which are able to detect pathogen-associated molecular patterns (PAMPs) and danger-associated molecular patterns (DAMPs) in both intracellular and external environments. However, the exact role of PRRs in endometriosis and the underlying molecular mechanism are unclear. PRRs are necessary for the innate immune system to identify and destroy invasive foreign infectious agents. Mammals mainly have two types of microbial recognition systems. The first one consists of the membrane-bound receptors, such as toll-like receptors (TLRs), which recognize extracellular microorganisms and activate intracellular signals to stimulate immune responses. The second one consists of the intracellular PRRs, including nod-like receptors (NLRs) and antiviral proteins retinoic acid-inducible gene I (RIG-I) and melanoma differentiation-associated gene 5 (MDA-5) with helix enzyme domain. In this review, we mainly focus on the key role of PRRs in the pathological processes associated with endometriosis. PRRs recognize PAMPs and can distinguish pathogenic microorganisms from self, triggering receptor ligand reaction followed by the stimulation of host immune response. Activated immune response promotes the transmission of microbial infection signals to the cells. As endometriosis is characterized by dysregulated inflammation and immune response, PRRs may potentially be involved in the activation of endometriosis-associated inflammation and immune disorders. Toll-like receptor 2 (TLR2), toll-like receptor 3 (TLR3), toll-like receptor 4 (TLR4), nod-like receptor family caspase activation and recruitment domain (CARD) domain containing 5 (NLRC5), nod-like receptor family pyrin domain containing 3 (NLRP3), and c-type lectin receptors (CLRs) play essential roles in endometriosis development by regulating immune and inflammatory responses. Absent in melanoma 2 (AIM2)-like receptors (ALRs) and retinoic acid-inducible gene I-like receptors (RLRs) may be involved in the activation of endometriosis-associated immune and inflammation disorders. PRRs, especially TLRs, may serve as potential therapeutic targets for alleviating pain in endometriosis patients. PRRs and their ligands interact with the innate immune system to enhance inflammation in the stromal cells during endometriosis. Thus, targeting PRRs and their new synthetic ligands may provide new therapeutic options for treating endometriosis.

Cytosolic DNA sensors and glial responses to endogenous DNA.

Genomic instability is a key driving force for the development and progression of many neurodegenerative diseases and central nervous system (CNS) cancers. The initiation of DNA damage responses is a critical step in maintaining genomic integrity and preventing such diseases. However, the absence of these responses or their inability to repair genomic or mitochondrial DNA damage resulting from insults, including ionizing radiation or oxidative stress, can lead to an accumulation of self-DNA in the cytoplasm. Resident CNS cells, such as astrocytes and microglia, are known to produce critical immune mediators following CNS infection due to the recognition of pathogen and damage-associated molecular patterns by specialized pattern recognition receptors (PRRs). Recently, multiple intracellular PRRs, including cyclic GMP-AMP synthase, interferon gamma-inducible 16, absent in melanoma 2, and Z-DNA binding protein, have been identified as cytosolic DNA sensors and to play critical roles in glial immune responses to infectious agents. Intriguingly, these nucleic acid sensors have recently been shown to recognize endogenous DNA and trigger immune responses in peripheral cell types. In the present review, we discuss the available evidence that cytosolic DNA sensors are expressed by resident CNS cells and can mediate their responses to the presence of self-DNA. Furthermore, we discuss the potential for glial DNA sensor-mediated responses to provide protection against tumorigenesis versus the initiation of potentially detrimental neuroinflammation that could initiate or foster the development of neurodegenerative disorders. Determining the mechanisms that underlie the detection of cytosolic DNA by glia and the relative role of each pathway in the context of specific CNS disorders and their stages may prove pivotal in our understanding of the pathogenesis of such conditions and might be leveraged to develop new treatment modalities.

The microbiome in HLA-B27-associated disease: implications for acute anterior uveitis and recommendations for future studies.

The pathogenesis of human leukocyte antigen (HLA)-B27-associated diseases such as acute anterior uveitis (AAU) and ankylosing spondylitis (AS) remains poorly understood, though Gram-negative bacteria and subclinical bowel inflammation are strongly implicated. Accumulating evidence from animal models and clinical studies supports several hypotheses, including HLA-B27-dependent dysbiosis, altered intestinal permeability, and molecular mimicry. However, the existing literature is hampered by inadequate studies designed to establish causation or uncover the role of viruses and fungi. Moreover, the unique disease model afforded by AAU to study the gut microbiota has been neglected. This review critically evaluates the current literature and prevailing hypotheses on the link between the gut microbiota and HLA-B27-associated disease. We propose a new potential role for HLA-B27-driven altered antibody responses to gut microbiota in disease pathogenesis and outline recommendations for future well-controlled human studies, focusing on AAU.

The role of NOD2 in intestinal immune response and microbiota modulation: A therapeutic target in inflammatory bowel disease.

As an intracellular pattern recognition receptor (PPR), the nucleotide-binding oligomerization domain-containing protein 2 (NOD2) triggers a cascade of immune responses. Previous studies of NOD2 regarding inflammatory bowel disease (IBD) mainly focused on the relevance of NOD2 mutations and loss within the disease onset and progression. With increasing research, more studies are exploring other functional roles and clinical applications of NOD2. In this review, we discuss the role of NOD2 in intestinal immune response and microbiota modulation in IBD and explore its clinical potential as a therapeutic target for IBD.

Novel Evidence That NADPH Oxidase 2 (Nox-2) Enhances Trafficking of Hematopoietic Stem Progenitor Cells (HSPCs) By Involving ROS Mediated Activation of Nlrp3 Inflammasome

Background. NADPH oxidase 2 (Nox-2) is a superoxide-generating enzyme that forms reactive oxygen species (ROS) to regulate the redox state involved in the self-renewal of hematopoietic stem/progenitor cells (HSPCs). Moreover, ROS are released from Nox-2 expressed by cell surface and mitochondria membranes and are potent activators of intracellular pattern recognition receptor Nlrp3 inflammasome, which regulates several aspects of HSPCs biology, including their trafficking, proliferation, and metabolism. Our previous research demonstrated a novel role of Nlrp3 inflammasome in the migration, mobilization, homing, and engraftment of HSPCs (Leukemia 2022;36(1):23-32;Stem Cell Rev Rep. 2020 16(5):954-967). The role of ROS was extensively studied in the proliferation of hematopoietic cells; however, its role in the migration of HSPCs is not very well understood. Aim of the study.Based on the role of Nox-2 in activating Nlrp3 inflammasome, we hypothesized that activation of Nox-2-ROS-Nlrp3 inflammasome axis in both HSPCs and BM microenvironment orchestrates trafficking of HSPCs.Materials and Methods. First, we studied in Transwells chemotaxis of Nox-2 deficient cells to bone marrow (BM) chemoattractants, including SDF-1, S1P, and eATP. Next, to shed more light on the role of Nox2 in the trafficking of HSPCs, we performed in Nox-2-KO animals G-CSF- and AMD3100-induced pharmacological mobilization. We also studied homing and engraftment of Nox-2-KO cells in BM of wild-type (WT) animals and homing and engraftment o WT BM cells in Nox-2-KO recipients. Nox-2-KO mice were also sublethal irradiated, and we followed the recovery of peripheral blood counts in these animals. Activation of Nlrp3 inflammasome in Nox-2-KO murine Sca-1+c-kit+lineage- HSPCs was evaluated by Glow assay. To ''bypass'' Nox-2-ROS activation in control experiments, nigericin has been employed as a direct activator of Nlrp3 inflammasome. Since membrane lipid rafts (MLRs) play a crucial role optimal migration of HSPCs, we evaluated the formation of membrane lipid rafts by employing confocal analysis in Nox-2-KO cells stimulated with SDF-1. We also studied the expression of enzymes (SREBP2, HMGCs, and HMGCR) involved in providing lipid components for MLRs assemble in activated Nox-2-KO cells. Results. We noticed that Nox-2-KO animals have a profound defect in the trafficking of HSPCs as evidenced by impaired chemotaxis to BM chemoattractants, decrease in pharmacological mobilization, and defect in homing and engraftment after transplantation. Nox-2-KO mice have also defect in recovery from sublethal irradiation. At the molecular level, HSPCs from Nox-2-KO mice displayed a defect in activation of Nlrp3 inflammasome and in cholesterol and sphingolipids synthesizing enzymes, which provide lipid components required for optimal assemble of MLRs. These defects were partially ameliorated by using direct activator of Nlrp3 inflammasome that is nigericin. Conclusions. We provide for the first time evidence that Nox-2-ROS-Nlrp3 inflammasome axis operates in normal HSPCs and is crucial for regulating trafficking and post-irradiation recovery of these cells. These effects are mediated by providing components for MLRs required for optimal migration and signaling in HSPCs

Novel evidence that the P2X1 purinergic receptor-Nlrp3 inflammasome axis orchestrates optimal trafficking of hematopoietic stem progenitors cells.

Our previous research demonstrated P2X purinergic receptors as important extracellular adenosine triphosphate (eATP) sensing receptors promoting the trafficking of hematopoietic stem progenitor cells (HSPCs). Accordingly, mice deficient in expression of P2X4 and P2X7 receptors turned out to mobilize poorly HSPCs. Similarly, defective expression of these receptors on transplanted HSPCs or in the bone marrow (BM) microenvironment of graft recipient mice led to defective homing, engraftment, and delayed hematopoietic reconstitution. This correlated with decreased activation of intracellular pattern recognition receptor Nlrp3 inflammasome. The P2X receptor family consists of seven purinergic receptors (P2X1-7) and we noticed that in addition to P2X4 and P2X7, HSPCs also highly express rapidly signaling the P2X1 receptor. Therefore, we asked if P2X1 receptor is also involved in HSPCs trafficking. We employed in vitro and in vivo murine models to study the role of P2X1 receptor blocked on HSPCs or bone marrow microenvironment cells by specific small molecular inhibitor NF499. First, we performed in vitro cell migration assays of bone marrow mononuclear cells (BMMNCs) isolated from normal mice that were exposed to NF499 and compared them to unexposed control cells. Next, in experiments in vivo we mobilized mice exposed to NF499 with G-CSF or AMD3100 and compared mobilization to control unexposed animals. Flow cytometry was employed to identify cell populations and clonogenic assays to enumerate the number of mobilized clonogenic progenitors. Similarly, in homing and engraftment experiments BMMNCs or recipient mice were exposed to NF499 and we evaluated homing and engraftment of transplanted cells by enumerating the number of cells labeled with fluorochromes in recipient mice BM and by evaluating the number of clonogenic progenitors in BM and spleen 24 hours and 12 days after transplantation. We also evaluated the potential involvement of Nlrp3 inflammasome in P2X1 receptor-mediated HSPCs trafficking. We report that the functional P2X1 receptor is highly expressed on murine and human HSPCs. We could demonstrate that the P2X1 receptor promotes the trafficking of murine cells in Nlrp3 inflammasome-dependent manner. Mice after exposure to P2X1 receptor inhibitor poorly mobilized HSPCs from the bone marrow into the peripheral blood. Mice transplanted with BMNNCs exposed to NF499 or recipient mice pretreated with this inhibitor demonstrated defective homing and engraftment as compared to control animals transplanted with cells not exposed to P2X1 inhibitor. Similar effects were noticed for control recipient mice that were not exposed to NF499. This study demonstrates for the first time the novel role of the P2X1 receptor in HSPCs trafficking in the mouse. Furthermore, it supports an important role of purinergic signaling engaging its downstream target Nlrp3 inflammasome in the mobilization, homing and engraftment of HSPCs.

Viral proteases activate the CARD8 inflammasome in the human cardiovascular system.

Nucleotide-binding oligomerization domain (NBD), leucine-rich repeat (LRR) containing protein family (NLRs) are intracellular pattern recognition receptors that mediate innate immunity against infections. The endothelium is the first line of defense against blood-borne pathogens, but it is unclear which NLRs control endothelial cell (EC) intrinsic immunity. Here, we demonstrate that human ECs simultaneously activate NLRP1 and CARD8 inflammasomes in response to DPP8/9 inhibitor Val-boro-Pro (VbP). Enterovirus Coxsackie virus B3 (CVB3)-the most common cause of viral myocarditis-predominantly activates CARD8 in ECs in a manner that requires viral 2A and 3C protease cleavage at CARD8 p.G38 and proteasome function. Genetic deletion of CARD8 in ECs and human embryonic stem cell-derived cardiomyocytes (HCMs) attenuates CVB3-induced pyroptosis, inflammation, and viral propagation. Furthermore, using a stratified endothelial-cardiomyocyte co-culture system, we demonstrate that deleting CARD8 in ECs reduces CVB3 infection of the underlying cardiomyocytes. Our study uncovers the unique role of CARD8 inflammasome in endothelium-intrinsic anti-viral immunity.

Activation and regulation mechanisms of NOD-like receptors based on structural biology.

Innate immunity is a primary defense system against microbial infections. Innate immune pattern recognition receptors (PRRs) play pivotal roles in detection of invading pathogens. When pathogens, such as bacteria and viruses, invade our bodies, their components are recognized by PRRs as pathogen-associated molecular patterns (PAMPs), activating the innate immune system. Cellular components such as DNA and RNA, acting as damage-associated molecular patterns (DAMPs), also activate innate immunity through PRRs under certain conditions. Activation of PRRs triggers inflammatory responses, interferon-mediated antiviral responses, and the activation of acquired immunity. Research on innate immune receptors is progressing rapidly. A variety of these receptors has been identified, and their regulatory mechanisms have been elucidated. Nucleotide-binding and oligomerization domain (NOD)-like receptors (NLRs) constitute a major family of intracellular PRRs and are involved in not only combating pathogen invasion but also maintaining normal homeostasis. Some NLRs are known to form multi-protein complexes called inflammasomes, a process that ultimately leads to the production of inflammatory cytokines and induces pyroptosis through the proteolytic cascade. The aberrant activation of NLRs has been found to be associated with autoimmune diseases. Therefore, NLRs are considered targets for drug discovery, such as for antiviral drugs, immunostimulants, antiallergic drugs, and autoimmune disease drugs. This review summarizes our recent understanding of the activation and regulation mechanisms of NLRs, with a particular focus on their structural biology. These include NOD2, neuronal apoptosis inhibitory protein (NAIP)/NLRC4, NLR family pyrin domain containing 1 (NLRP1), NLRP3, NLRP6, and NLRP9. NLRs are involved in a variety of diseases, and their detailed activation mechanisms based on structural biology can aid in developing therapeutic agents in the future.

Extracellular Adenosine (eAdo) - A2B Receptor Axis Inhibits in Nlrp3 Inflammasome-dependent Manner Trafficking of Hematopoietic Stem/progenitor Cells

We postulated that mobilization, homing, and engraftment of hematopoietic stem/progenitor cells (HSCPs) is facilitated by a state of sterile inflammation induced in bone marrow (BM) after administration of pro-mobilizing drugs or in response to pre-transplant myeloablative conditioning. An important role in this phenomenon plays purinergic signaling that by the release of extracellular adenosine triphosphate (eATP) activates in HSPCs and in cells in the hematopoietic microenvironment an intracellular pattern recognition receptor (PPR) known as Nlrp3 inflammasome. We reported recently that its deficiency results in defective trafficking of HSPCs. Moreover, it is known that eATP after release into extracellular space is processed by cell surface expressed ectonucleotidases CD39 and CD73 to extracellular adenosine (eAdo) that in contrast to eATP shows an anti-inflammatory effect. Based on data that the state of sterile inflammation promotes trafficking of HSPCs, and since eAdo is endowed with anti-inflammatory properties we become interested in how eAdo will affect the mobilization, homing, and engraftment of HSPCs and which of eAdo receptors are involved in these processes. As expected, eAdo impaired HSPCs trafficking and this occurred in autocrine- and paracrine-dependent manner by direct stimulation of these cells or by affecting cells in the BM microenvironment. We report herein for the first time that this defect is mediated by activation of the A2B receptor and a specific inhibitor of this receptor improves eAdo-aggravated trafficking of HSPCs. To explain this at the molecular level eAdo-A2B receptor interaction upregulates in HSPCs in NF-kB-, NRF2- and cAMP-dependent manner heme oxygenase-1 (HO-1), that is Nlrp3 inflammasome inhibitor. This corroborated with our analysis of proteomics signature in murine HSPCs exposed to eAdo that revealed that A2B inhibition promotes cell migration and proliferation. Based on this we postulate that blockage of A2B receptor may accelerate the mobilization of HSPCs as well as their hematopoietic reconstitution and this approach could be potentially considered in the future to be tested in the clinic.Graphical

1329-P: A RIPK2 Inhibitor Protects against Saturated-Fat–Induced Glucose Intolerance in Mice

Background: Obesity and type 2 diabetes (T2D) are associated with chronic low-grade inflammation. Elevated free fatty acids (FFAs) , as seen in obesity and T2D, can act as damage associated molecular patterns (DAMPs) activating the innate immune system. Recent studies have identified nucleotide binding oligomerization domain 1 (NOD1) , an intracellular pattern recognition receptor, can be activated by saturated fatty acids (sFA) . NOD1 activates inflammatory kinase RIPK2, leading to increased cytokine production by the innate immune system. Our study aimed to determine whether wild type mice treated with GSK583, a RIPK2 inhibitor, are protected against FFA-induced glucose intolerance. Material and Methods: C57Bl/6J mice were surgically cannulated in their jugular veins and were infused with ethylpalmitate (EtP) for 48h, which results in increased plasma palmitate levels and mimics the elevated FFA levels seen in vivo. Furthermore, mice were also infused with EtP and GSK583 for 48h. After infusion, the mice were subjected to a hyperglycemic clamp and the glucose infusion rate was measured to assess glucose tolerance that under hyperglycemic clamp conditions, is mainly determined by the beta cell ability to compensate for FFA-induced insulin resistance. Results: Preliminary data revealed that mice infused with EtP and GSK583 had a higher glucose infusion rate (GIR; mg/kgmin) (83+20, mean+SEM) than mice infused with EtP alone (57+4) . The EtP and GSK583 was comparable to the GIR of control (EtP vehicle infused) mice (84+7) . Conclusions: Our data indicate that pharmacological inhibition of NOD1 signaling protects against FFA-induced glucose intolerance in mice. Targeting NOD1 may be an effective strategy for treating beta cell dysfunction in T2D. Disclosure N.Tsakiridis: None. A.Ivovic: None. S.Rahman: None. Y.Tan: None. A.Giacca: None.

1322-P: Role of Macrophage NOD1 in Fat-Induced ß-Cell Dysfunction

In individuals with obesity, free fatty acids (FFA) are chronically raised resulting in ß-cell dysfunction, leading to type 2 diabetes (T2D) . Recent data implicate the role of inflammation in developing ß-cell dysfunction, which is in part mediated by nucleotide-oligomerization domain 1 (NOD1) , an intracellular pattern recognition receptor. NOD1 is activated by peptidoglycans of bacterial cell walls; however, in vitro studies have documented that NOD1 can also be activated by saturated FFA. Previous data from our lab have shown that palmitate-induced ß-cell dysfunction was prevented in whole body and ß-cell specific NOD1 null mice. We have seen earlier reports on increased macrophage infiltration in islets after elevation in circulating palmitate, and much higher expression of NOD1 in macrophages than ß-cells. Hence, we hypothesized that macrophage NOD1 receptors play a pivotal role in FFA-induced ß-cell dysfunction. We have generated myeloid specific NOD1 null mice to target macrophage NOD1 receptors by crossing NOD1 floxed mice with Lyz2 Cre+ mice. Both knockouts and their controls (floxed and Cre) were infused for 48 h with ethylpalmitate (hydrolyzed to palmitate and ethanol in plasma) , to achieve elevated circulating FFA in a non-toxic manner. Ethanol vehicle was infused as control treatment. To assess in vivo ß-cell function, hyperglycemic clamp was performed following the 48 h infusion, and the disposition index (DI) was calculated. Ethylpalmitate resulted in elevated plasma FFA compared to vehicle infusion. Ethylpalmitate caused marked reduction in DI in control mice, while knockout mice displayed no significant changes in DI between ethylpalmitate and vehicle infused groups. These results suggest that macrophage NOD1 plays a causal role in palmitate-induced ß-cell dysfunction in vivo. Overall, our data reveal NOD1 as a key factor and a potential therapeutic target for obesity-associated diabetes. Disclosure S.Rahman: None. J.Yung: None. Y.Zhang: None. A.Giacca: None.

Essential role of Rnd1 in innate immunity during viral and bacterial infections

Intracellular and cell surface pattern-recognition receptors (PRRs) are an essential part of innate immune recognition and host defense. Here, we have compared the innate immune responses between humans and bats to identify a novel membrane-associated protein, Rnd1, which defends against viral and bacterial infection in an interferon-independent manner. Rnd1 belongs to the Rho GTPase family, but unlike other small GTPase members, it is constitutively active. We show that Rnd1 is induced by pro-inflammatory cytokines during viral and bacterial infections and provides protection against these pathogens through two distinct mechanisms. Rnd1 counteracts intracellular calcium fluctuations by inhibiting RhoA activation, thereby inhibiting virus internalisation. On the other hand, Rnd1 also facilitates pro-inflammatory cytokines IL-6 and TNF-α through Plxnb1, which are highly effective against intracellular bacterial infections. These data provide a novel Rnd1-mediated innate defense against viral and bacterial infections.

Palmitoylation facilitates inflammation through suppressing NOD2 degradation mediated by the selective autophagy receptor SQSTM1

ABSTRACT The intracellular pattern recognition receptor NOD2 senses bacterial peptidoglycan to drive proinflammatory and antimicrobial responses. Dysregulation of NOD2 signaling confers susceptibility to several immunological and inflammatory diseases. Although palmitoylation of NOD2 is required for its membrane recruitment and activation, whether palmitoylation can modulate the stability of NOD2 to orchestrate inflammation remains unclear. Recently, we have revealed that S-palmitoylation restricts SQSTM1-mediated selective macroautophagic/autophagic degradation of NOD2, and identified a gain-of-function R444C variant of NOD2 short isoform (NOD2sR444C) in autoinflammatory disease, which induces excessive inflammation through its enhanced S-palmitoylation level and decreased autophagic degradation.

Genomic structure and molecular characterization of NLRC3-like from Siberian sturgeon (Acipenser baerii) and expression response to Streptococcus iniae and pathogen-associated molecular patterns

NLRs are important intracellular pattern recognition receptors and play an essential role in innate immunity for vertebrates. However, research on sturgeon NLRs is still scarce. In this study, the NLRC3-like gene from Siberian sturgeon (Acipenser baerii) (AbNLRC3-like) was cloned and characterized. The AbNLRC3-like full-length cDNA was composed of 3310 bp with a 166 bp of 5′-UTR, 2838 bp open reading frame (ORF) and 306 bp 3′-UTR, encoding 945 amino acids. Structural analysis of AbNLRC3-like showed the typical NLRs structure, including a central NACHT domain and a C-terminal 12 LRR motifs. Quantitative real-time PCR (qRT-PCR) analysis revealed that AbNLRC3-like was widely distributed in all tested tissues with a relatively high expression level in the mid-kidney, head-kidney and spleen. After Streptococcus iniae infection, the mRNA expression of AbNLRC3-like was significantly up-regulated at the pre-mortality period and the recovering period in valvula intestine, but it was significantly down-regulated around the mortality period in the duodenum and spleen. In splenic leukocyte, the mRNA expression of AbNLRC3-like was significantly induced by LPS, PGN and Poly(I:C). These results suggested that AbNLRC3-like may play a critical role in the immune response of Siberian sturgeon during pathogen challenge. To our best knowledge, this is the first report of the NLRC3 gene in sturgeon.

Attenuation of apoptotic cell detection triggers thymic regeneration after damage.

SUMMARYThe thymus, which is the primary site of T cell development, is particularly sensitive to insult but also has a remarkable capacity for repair. However, the mechanisms orchestrating regeneration are poorly understood, and delayed repair is common after cytoreductive therapies. Here, we demonstrate a trigger of thymic regeneration, centered on detecting the loss of dying thymocytes that are abundant during steady-state T cell development. Specifically, apoptotic thymocytes suppressed production of the regenerative factors IL-23 and BMP4 via TAM receptor signaling and activation of the Rho-GTPase Rac1, the intracellular pattern recognition receptor NOD2, and micro-RNA-29c. However, after damage, when profound thymocyte depletion occurs, this TAM-Rac1-NOD2-miR29c pathway is attenuated, increasing production of IL-23 and BMP4. Notably, pharmacological inhibition of Rac1-GTPase enhanced thymic function after acute damage. These findings identify a complex trigger of tissue regeneration and offer a regenerative strategy for restoring immune competence in patients whose thymic function has been compromised.

NOD2 Genotypes Affect the Symptoms and Mortality in the Porcine Circovirus 2-Spreading Pig Population.

The nucleotide oligomerization domain (NOD)-like receptor 2 (NOD2) is an intracellular pattern recognition receptor that detects components of peptidoglycans from bacterial cell walls. NOD2 regulates bowel microorganisms, provides resistance against infections such as diarrhea, and reduces the risk of inflammatory bowel diseases in humans and mice. We previously demonstrated that a specific porcine NOD2 polymorphism (NOD2-2197A > C) augments the recognition of peptidoglycan components. In this study, the relationships between porcine NOD2-2197A/C genotypes affecting molecular functions and symptoms in a porcine circovirus 2b (PCV2b)-spreading Duroc pig population were investigated. The NOD2 allele (NOD2-2197A) with reduced recognition of the peptidoglycan components augmented the mortality of pigs at the growing stage in the PCV2b-spreading population. Comparison of NOD2 allele frequencies in the piglets before and after invasion of PCV2b indicated that the ratio of NOD2-2197A decreased in the population after the PCV2b epidemic. This data indicated that functional differences caused by NOD2-2197 polymorphisms have a marked impact on pig health and livestock productivity. We suggest that NOD2-2197CC is a PCV2 disease resistant polymorphism, which is useful for selective breeding by reducing mortality and increasing productivity.

NLRP3 promotes endometrial receptivity by inducing epithelial-mesenchymal transition of the endometrial epithelium.

Endometrial receptivity is crucial for successful embryo implantation. It is regulated by multiple factors which include ovarian steroid hormones and the immune microenvironment among others. Nod-Like Receptor Pyrins-3 (NLRP3) is a key intracellular pattern-recognition receptor and a critical component of the inflammasome, which plays an essential role in the development of inflammation and of immune responses. However, the physiological functions of NLRP3 in the endometrium remain largely unclear. This study investigated the physiological and pathological significance of NLRP3 in human endometrial epithelial cell during the implantation window. NLRP3 is highly expressed during the mid-proliferative and mid-secretory phases of the human endometrium and transcriptionally up-regulated by estradiol (E2) through estrogen receptor β (ERβ). In addition, NLRP3 promotes embryo implantation and enhances epithelial-mesenchymal transition (EMT) of Ishikawa (IK) cells via both inflammasome-dependent and inflammasome-independent pathways, which might provide a novel insight into endometrial receptivity and embryo implantation. Our findings suggest that NLRP3, which is transcriptionally regulated by E2, induces epithelial-mesenchymal transition of endometrial epithelial cells and promotes embryo adhesion.

HBsAg Dampened STING Associated Activation of NK Cells in HBeAg-Negative CHB Patients.

NK cells play crucial roles in defending against persistent HBV. However, NK cells present dysfunction in chronic hepatitis B virus (CHB) infection, and the associated mechanism is still not fully understood. Except for the regulatory receptors, NK cells could also be regulated by the surface and intracellular pattern recognition receptors (PRRs). In the present study, we found that the level of the adaptor of DNA sensor STING in NK cells was significantly decreased in HBeAg-negative CHB patients, and it was positively associated with the degranulation ability of NK cells. Compared to NK cells from healthy donors, NK cells from HBeAg-negative CHB patients displayed a lower responsiveness to cGAMP stimulation. Further investigation showed that HBsAg could inhibit the STING expression in NK cells and suppress the response of NK cells to cGAMP. Significantly, STAT3 was identified to be a transcription factor that directly regulated STING transcription by binding to the promoter. In addition, STAT3 positively regulated the STING associated IFN-α response of NK cells. These findings suggested that STING is an important adaptor in NK cell recognition and activation, while HBsAg disturbs NK cell function by the STAT3-STING axis, providing a new mechanism of NK disability in HBeAg-negative CHB infection.