Activation Of Pattern Recognition Receptors Research Articles

Pattern-recognition receptors and immunometabolic reprogramming: what we know and what to explore.

Evolutionarily, immune response is a complex mechanism that protects the host from internal and external threats. Pattern recognition receptors (PRRs) recognize MAMPs, PAMPs, and DAMPs to initiate a protective pro-inflammatory immune response. PRRs are expressed on the cell membranes by TLR1, 2, 4, and 6 and in the cytosolic organelles by TLR3, 7, 8, and 9, NLRs, ALRs, and cGLRs. We know their downstream signaling pathways controlling immunoregulatory and pro-inflammatory immune response. However, the impact of PRRs on metabolic control of immune cells to control their pro- and anti-inflammatory activity has not been discussed extensively. Immune cell metabolism or immunometabolism critically determines immune cells' pro-inflammatory phenotype and function. The current article discusses immunometabolic reprogramming (IR) upon activation of different PRRs, such as TLRs, NLRs, cGLRs, and RLRs. The duration and type of PRR activated, species studied, and location of immune cells to specific organ are critical factors to determine the IR-induced immune response. The work herein describes IR upon TLR, NLR, cGLR, and RLR activation. Understanding IR upon activating different PRRs is critical for designing better immune cell-specific immunotherapeutics and immunomodulators targeting inflammation and inflammatory diseases.

Bifunctional HDAC and DNMT inhibitor induces viral mimicry activates the innate immune response in triple-negative breast cancer

Triple-negative breast cancer (TNBC) is a unique breast cancer subtype characterized by a lack of estrogen receptor (ER), progesterone receptor (PR), and human epidermal growth factor receptor 2 (HER2) expression. Since TNBC lacks ER, PR, and HER2, there are currently no drugs that specifically target TNBC. Therefore, the development of new drugs or effective treatment strategies to target TNBC has become an urgent clinical need. Research has shown that the application of histone deacetylase (HDAC) inhibitors and DNA methyltransferase (DNMT) inhibitors leads to genomic and epigenomic instability. This, in turn, triggers the activation of pattern recognition receptors (PRRs) and subsequently activates downstream interferon (IFN) signalling pathways. In this study, the bifunctional HDAC and DNMT inhibitor J208 exhibited antitumour activity in TNBC cell lines. J208 effectively induced apoptosis and cell cycle arrest at the G0/G1 phase, inhibiting cell migration and invasion in TNBC. Moreover, this bifunctional inhibitor induced the expression of endogenous retroviruses (ERVs) and elicited a viral mimicry response, which increased the intracellular levels of double-stranded RNA (dsRNA) to activate the innate immune signalling pathway in TNBC. In summary, we demonstrated that the bifunctional inhibitor J208, which is designed to inhibit HDAC and DNMT, has potent anticancer effects, providing a new research basis for reactivating antitumour immunity by triggering innate immune signalling and offering a promising strategy for TNBC treatment.

Mitochondrial damage and activation of NLRP3 induced by Yersinia ruckeri infection play an important role in the occurrence of diffuse acute inflammation of channel catfish (Ictalurus punctatus)

Yersinia ruckeri, as a facultative intracellular pathogen, can cause two different infection phenotypes of fish. An important phenotype is diffuse acute inflammation (DAI), which is mainly characterized by systemic hemorrhage and multiple tissues damage. Another phenotype is enteric redmouth symptom (EM) with predominantly digestive system damage, while slight damage to other tissues and organs. At present, the causes and mechanisms of heterogeneity of Y. ruckeri infection are not clear. According to our previous research and related bioinformatics analysis, it is suggested that the heterogeneity of infection of Y. ruckeri may be related to the activation of intracellular pattern recognition receptor (NLRP3). Furthermore, combined with the transmission electron microscope observation and ultrastructural pathological analysis of DAI and EM phenotypes, it is hypothesized that Y. ruckeri infection may mediate the occurrence of DAI by inducing mitochondrial damage and activation of NLRP3. In this study, we established DAI and EM phenotypes of channel catfish (Ictalurus punctatus) through the artificial infection experiment. Subsequently, we observed the pathological changes and characteristics of different tissues in DAI and EM phenotypes, such as liver, spleen, kidney, intestine and gill. In addition, the mitochondrial damage of head kidney mediated by Y. ruckeri infection was confirmed by transmission electron microscope observation. Furthermore, by detecting the expression of proinflammatory factors and related pattern recognition receptors in various tissues, we preliminarily clarified the molecular characteristics of immune response in DAI and EM phenotype fish. Moreover, the head kidney macrophages isolated from DAI and EM were observed through transmission electron microscope, and the expression levels of NLRP3, Caspase-1 and IL-1β were detected. Finally, the correlation between mitochondrial damage induced by Y. ruckeri infection and NLRP3 signal transduction was preliminary revealed. Taken together, our results showed that the inflammatory DAI phenotype of fish mediated by Y. ruckeri infection is closely related to mitochondrial damage and NLRP3 activation, which will provide theoretical basis for further study on the pathogenic mechanism of Y. ruckeri infection and clinical diagnosis of the disease, and also provide some insights into the role of NLRP3 in the natural immunity of aquatic animals.

Abstract 4295: Apoptotic DNase DFFB-induced DNA damage within cancer persister cells underlies acquired resistance to targeted therapies across multiple tumor types

Abstract Cancer acquired drug resistance contributes to a large proportion of patient deaths. A major obstacle to the development of effective therapies which prevent resistance is our lack of understanding of the relevant fundamental molecular adaptive processes. Here we report the discovery that tumor cells rely upon sublethal engagement of apoptotic death machinery and apoptotic DNase DFFB to acquire resistance to targeted therapies in multiple tumor types in cell culture and in vivo. We found that while DFFB wild type tumor cells readily acquire resistance to targeted therapies by escaping from the quiescent “persister” state into proliferating resistant cells, persister cells lacking functional DFFB remain indefinitely in a quiescent state on treatment. Mechanistically, we found that DFFB, which is activated by apoptotic caspases in surviving cancer persister cells, induces persistent DNA damage which promotes resistance through acquired mutations and nongenetic resistance through suppression of growth-arresting interferon signaling. During drug treatment acute interferon induction arises from sublethal mitochondrial outer membrane permeabilization and cytoplasmic exposure of mitochondrial nucleic acids which activate pattern recognition receptors. However, during longer treatments we found that DFFB knockout persister cells exhibit strikingly elevated levels of STAT1 and STAT2 and interferon stimulated gene sets. This interferon signaling is growth arresting because treatment of DFFB knockout persister cells with a JAK1/2 inhibitor rescues their ability to regrow. Furthermore, in DFFB wild type persister cells we found that DFFB-dependent DNA damage activates stress response transcription factor ATF3 which blocks the transcriptional activation of interferon stimulated genes following initial interferon induction. This is consistent with prior data in other contexts demonstrating that ATF3 is induced by DNA damage and that ATF3 directly binds to promoters of multiple interferon genes repressing their expression. These findings reveal that DFFB activation in persister cells prevents chronic interferon signaling and enables escape into proliferating drug resistant tumor cells. Therefore, DFFB-mediated interferon suppression represents a novel mechanism that is fundamental to acquired drug resistance. In addition, high expression of DFFB and ATF3 correlates with worse overall survival in a pan-cancer analysis and DFFB knockout mice develop normally indicating DFFB is a nonessential gene in normal tissue. Therefore, we propose that DFFB is a promising therapeutic target to prevent acquired resistance to targeted therapy. Citation Format: August Finley Williams, David A. Gervasio, Claire E. Turkal, Anna E. Stuhlfire, Michael X. Wang, Brandon E. Mauch, Ariel H. Nguyen, Michelle H. Paw, Mehrshad Hairani, Cooper P. Lathrop, Sophie H. Harris, Jennifer L. Page, Matthew J. Hangauer. Apoptotic DNase DFFB-induced DNA damage within cancer persister cells underlies acquired resistance to targeted therapies across multiple tumor types [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 4295.

A Polymer-Based Antigen Carrier Activates Two Innate Immune Pathways for Adjuvant-Free Subunit Vaccines.

The activation of multiple Pattern Recognition Receptors (PRRs) has been demonstrated to trigger inflammatory responses and coordinate the host's adaptive immunity during pathogen infections. The use of PRR agonists as vaccine adjuvants has been reported to synergistically induce specific humoral and cellular immune responses. However, incorporating multiple PRR agonists as adjuvants increases the complexity of vaccine design and manufacturing. In this study, we discovered a polymer that can activate both the Toll-like receptor (TLR) pathway and cyclic GMP-AMP synthase (cGAS)-stimulator of interferon genes (STING) pathway. The polymer was then conjugated to protein antigens, creating an antigen delivery system for subunit vaccines. Without additional adjuvants, the antigen-polymer conjugates elicited strong antigen-specific humoral and cellular immune responses. Furthermore, the antigen-polymer conjugates, containing the Receptor Binding Domain (RBD) of the Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) Spike Protein or the Monkeypox Antigen M1R as the antigens, were found to induce potent antigen-specific antibodies, neutralizing antibodies, and cytotoxic T cells. Immunization with M1R-polymer also resulted in effective protection in a lethal challenge model. In conclusion, this vaccine delivery platform offers an effective, safe, and simple strategy for inducing antigen-specific immunity against infectious diseases.

Role of pattern recognition receptors and microbiota-derived ligands in obesity

Obesity is associated with activation of low-grade inflammation in tissues metabolically relevant for the regulation glucose homeostasis. The gut microbiota has been extensively linked to the inflammatory responses observed during obesity emphasizing the interconnection between host immunity and metabolism during obesity. Gut microbiota together with alteration of the gut barrier functions provide a myriad of circulating ligands for the pattern recognition receptors (PRRs) expressed in innate immune cells and nonimmune cells. PRR-dependent signalling drives the expression of a wide range of genes beyond the inflammatory response depending on the specific functions of the targeted cells and on the physiological context. PRRs activation can have opposite effects on host metabolic inflammation. Nucleotide-binding oligomerization domain 1 (NOD1) or NOD-like Receptor pyrin domain containing 3 (NLRP3) activation promote metabolic inflammation and insulin resistance while NOD2 activation improves insulin sensitivity and glucose homeostasis during obesity. Toll-like receptors (TLRs) 2, 4 and 5 also display specific effects on metabolic tissues. TLR5 deficient mice are prone to obesity and inflammation in response to high fat diet, while injection of TLR5 ligand, flagellin, has a protective effect toward diet-induced obesity. To the opposite TLR2 and 4 activations are associated with deleterious metabolic outcome during obesity. TLR4 activation enhances metabolic inflammation and insulin resistance and TLR2 via its activation by molecules derived from the gut microbiota favours the onset of obesity. It is now clear that activation of PRRs by bacterial derived molecules plays a key role in the host metabolic regulation. PRRs are expressed in various cell types complicating the understanding of the mechanisms underlying the relationship between PRRs activation/silencing and metabolic inflammation in obesity context. This review presents an overview of the current understanding of the interrelationship between the gut microbiota and PRRs, with a focus on its consequences for obesity and related metabolic diseases.

Preferential Induction of Canonical IMD and Toll Innate Immune Receptors by Bacterial Challenges in Triatoma Pallidipennis Primed with Gram-Negative and Gram-Positive Bacteria, Respectively

Insects lack an adaptive immune defense against invading microorganisms, but they possess humoral and cellular response similar to that of vertebrates. The Immune Deficiency (IMD) and Toll are the major signaling pathways to produce humoral antimicrobial peptides AMPs. Pathogen molecular patterns (PAMs) of Gram-negative bacteria activate Pattern recognition receptors (PRR) of the IMD pathway, while PAMS of Gram-positive activate PRR of the Toll pathway. Although the IMD pathways is incomplete in Hemipterans, in Triatoma pallidipennis, there is a preferential participation of the IMD pgrp-lc and toll receptors in the responses to Gramnegative and Gram-positive bacteria, respectively. Still, as in other insects, cross induction was observed. An enhanced protection after a previous exposure to a pathogen, termed priming, functionally homologous to the adaptive immune memory of vertebrates, has been documented in several insect Orders but not in Hemiptera, and the participation of the components of the immune signaling cascades remains poorly explored. We present evidence for immune priming to Micrococcus luteus (Gram-positive) and Escherichia coli (Gramnegative) bacteria in T. pallidipennis. The preferential participation of receptors of the IMD and Toll pathways in the responses to each bacterial challenge was recorded

Antiviral responses versus virus-induced cellular shutoff: a game of thrones between influenza A virus NS1 and SARS-CoV-2 Nsp1.

Following virus recognition of host cell receptors and viral particle/genome internalization, viruses replicate in the host via hijacking essential host cell machinery components to evade the provoked antiviral innate immunity against the invading pathogen. Respiratory viral infections are usually acute with the ability to activate pattern recognition receptors (PRRs) in/on host cells, resulting in the production and release of interferons (IFNs), proinflammatory cytokines, chemokines, and IFN-stimulated genes (ISGs) to reduce virus fitness and mitigate infection. Nevertheless, the game between viruses and the host is a complicated and dynamic process, in which they restrict each other via specific factors to maintain their own advantages and win this game. The primary role of the non-structural protein 1 (NS1 and Nsp1) of influenza A viruses (IAV) and the pandemic severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), respectively, is to control antiviral host-induced innate immune responses. This review provides a comprehensive overview of the genesis, spatial structure, viral and cellular interactors, and the mechanisms underlying the unique biological functions of IAV NS1 and SARS-CoV-2 Nsp1 in infected host cells. We also highlight the role of both non-structural proteins in modulating viral replication and pathogenicity. Eventually, and because of their important role during viral infection, we also describe their promising potential as targets for antiviral therapy and the development of live attenuated vaccines (LAV). Conclusively, both IAV NS1 and SARS-CoV-2 Nsp1 play an important role in virus-host interactions, viral replication, and pathogenesis, and pave the way to develop novel prophylactic and/or therapeutic interventions for the treatment of these important human respiratory viral pathogens.

The Sixth Sense: Self-nucleic acid sensing in the brain.

Our innate immune system uses pattern recognition receptors (PRRs) as a first line of defense to detect microbial ligands and initiate an immune response. Viral nucleic acids are key ligands for the activation of many PRRs and the induction of downstream inflammatory and antiviral effects. Initially it was thought that endogenous (self) nucleic acids rarely activated these PRRs, however emerging evidence indicates that endogenous nucleic acids are able to activate host PRRs in homeostasis and disease. In fact, many regulatory mechanisms are in place to finely control and regulate sensing of self-nucleic acids by PRRs. Sensing of self-nucleic acids is particularly important in the brain, as perturbations to nucleic acid sensing commonly leads to neuropathology. This review will highlight the role of nucleic acid sensors in the brain, both in disease and homeostasis. We also indicate the source of endogenous stimulatory nucleic acids where known and summarize future directions for the study of this growing field.

Single-cell transcriptomics of peripheral blood mononuclear cells indicates impaired immune and inflammatory responses in alcohol-associated hepatitis

Alcohol-associated hepatitis (AH) is often diagnosed at advanced stages, and severe AH usually carries poor prognosis and high short-term mortality. In addition, it is challenging to understand the molecular mechanisms of immune dysregulation and inflammation in AH due to the cellular complexity and heterogeneity. Using single-cell RNA sequencing, previous studies found that AH causes dysfunctional innate immune response in monocytes, involving activation of pattern recognition receptors (PRRs) and cytokine signaling pathways. To better understand the coordinated systemic immune response in AH patients, we performed combined single-cell transcriptome, cell-surface protein, and lymphocyte antigen receptor analysis of peripheral blood mononuclear cell (PBMC) samples. Our results showed inflammatory cytokines and chemokines were highly expressed in AH, including IL-2, IL-32, CXC3R1 and CXCL16 in monocytes and NK cells, whereas HLA-DR genes were reduced in monocytes. In addition, we also found altered differentiation of T-helper cells (TH1 and TH17), which could further lead to neutrophil recruitment and macrophage activation. Lastly, our results also suggest impaired NK-cell activation and differentiation in AH with reduced gene expression of KLRC2 and increased gene expression of KLRG1. Our findings indicate different mechanisms may be involved in impaired immune and inflammatory responses for the cellular subtypes of the PBMCs in AH.

Toll-like receptor 7 and RIG-I-like receptors expression in peripheral blood mononuclear cells of naïve patients with hepatitis C

BackgroundThe proper function of Pattern Recognition Receptors (PRRs) as a part of the host immune system can eliminate numerous pathogens from the body. However, some viruses can manipulate PRRs to escape the innate immune system. As there is controversy in the activation of PRRs in patients infected with HCV, we decided to evaluate the gene expression changes of PRRs in HCV cases compared to the healthy control.MethodsIn this study, the relative expression of Toll-like receptor 7, RIG-I, and MAD-5 in peripheral mononuclear blood cells of twenty HCV patients and twenty healthy controls of the same gender and age were analyzed by quantitative Real-time PCR.ResultsOur results showed that the expression of RIG-I and MAD-5 significantly increased in HCV-infected samples compared to the controls (P value:0.01; P value:0.05), while the expression of TLR7 was similar between the case and the control group (P value:0.1).ConclusionIt seems in suppressing HCV, RIG-I and MAD-5 receptors are likely to be more activated than TRL7 in HCV patients. The lack of TLR7 gene expression might reflect the defect of the host in the stimulation of the innate immune system through the TLR7 pathway.

Substantial heterogeneity of inflammatory cytokine production and its inhibition by a triple cocktail of toll-like receptor blockers in early sepsis.

Early sepsis is a life-threatening immune dysregulation believed to feature a "cytokine storm" due to activation of pattern recognition receptors by pathogen and danger associated molecular patterns. However, treatments with single toll-like receptor (TLR) blockers have shown no clinical benefit. We speculated that sepsis patients at the time of diagnosis are heterogeneous in relation to their cytokine production and its potential inhibition by a triple cocktail of TLR blockers. Accordingly, we analyzed inflammatory cytokine production in whole blood assays from early sepsis patients and determined the effects of triple TLR-blockade. Whole blood of 51 intensive care patients sampled within 24h of meeting Sepsis-3 criteria was incubated for 6h without or with specific TLR2, 4, and 7/8 stimuli or suspensions of heat-killed S. aureus or E. coli bacteria as pan-TLR challenges, and also with a combination of monoclonal antibodies against TLR2 and 4 and chloroquine (endosomal TLR inhibition), subsequent to dose optimization. Concentrations of tumor necrosis factor (TNF), Interleukin(IL)-6, IL-8, IL-10, IL-1α and IL-1β were measured (multiplex ELISA) before and after incubation. Samples from 11 sex and age-matched healthy volunteers served as controls and for dose-finding studies. Only a fraction of sepsis patient samples revealed ongoing cytokine production ex vivo despite sampling within 24h of first meeting Sepsis-3 criteria. In dose finding studies, inhibition of TLR2, 4 and endosomal TLRs reliably suppressed cytokine production to specific TLR agonists and added bacteria. However, inflammatory cytokine production ex vivo was only suppressed in the high cytokine producing samples but not in the majority. The suppressive response to TLR-blockade correlated both with intraassay inflammatory cytokine production (r=0.29-0.68; p<0.0001-0.04) and cytokine baseline concentrations (r=0.55; p<0.0001). Upon meeting Sepsis-3 criteria for less than 24h, a mere quarter of patient samples exhibits a strong inflammatory phenotype, as characterized by increased baseline inflammatory cytokine concentrations and a stark TLR-dependent increase upon further ex vivo incubation. Thus, early sepsis patient cohorts as defined by Sepsis-3 criteria are very heterogeneous in regard to inflammation. Accordingly, proper ex vivo assays may be useful in septic individuals before embarking on immunomodulatory treatments.

Hypoxia dampens innate immune signalling at early time points and increases Zika virus RNA levels in iPSC-derived macrophages.

Type I interferons (IFNs) are the major host defence against viral infection and are induced following activation of cell surface or intracellular pattern recognition receptors, including retinoic-acid-inducible gene I (RIG-I)-like receptors (RLRs). All cellular processes are shaped by the microenvironment and one important factor is the local oxygen tension. The majority of published studies on IFN signalling are conducted under laboratory conditions of 18% oxygen (O2), that do not reflect the oxygen levels in most organs (1-5 % O2). We studied the effect of low oxygen on IFN induction and signalling in induced Pluripotent Stem Cell (iPSC)-derived macrophages as a model for tissue-resident macrophages and assessed the consequence for Zika virus (ZIKV) infection. Hypoxic conditions dampened the expression of interferon-stimulated genes (ISGs) following RLR stimulation or IFN treatment at early time points. RNA-sequencing and bio-informatic analysis uncovered several pathways including changes in transcription factor availability, the presence of HIF binding sites in promoter regions, and CpG content that may contribute to the reduced ISG expression. Hypoxic conditions increased the abundance of ZIKV RNA highlighting the importance of understanding how low oxygen conditions in the local microenvironment affect pathogen sensing and host defences.

Role of toll-like receptors and nod-like receptors in acute lung infection.

The respiratory system exposed to microorganisms continuously, and the pathogenicity of these microbes not only contingent on their virulence factors, but also the host's immunity. A multifaceted innate immune mechanism exists in the respiratory tract to cope with microbial infections and to decrease tissue damage. The key cell types of the innate immune response are macrophages, neutrophils, dendritic cells, epithelial cells, and endothelial cells. Both the myeloid and structural cells of the respiratory system sense invading microorganisms through binding or activation of pathogen-associated molecular patterns (PAMPs) to pattern recognition receptors (PRRs), including Toll-like receptors (TLRs) and NOD-like receptors (NLRs). The recognition of microbes and subsequent activation of PRRs triggers a signaling cascade that leads to the activation of transcription factors, induction of cytokines/5chemokines, upregulation of cell adhesion molecules, recruitment of immune cells, and subsequent microbe clearance. Since numerous microbes resist antimicrobial agents and escape innate immune defenses, in the future, a comprehensive strategy consisting of newer vaccines and novel antimicrobials will be required to control microbial infections. This review summarizes key findings in the area of innate immune defense in response to acute microbial infections in the lung. Understanding the innate immune mechanisms is critical to design host-targeted immunotherapies to mitigate excessive inflammation while controlling microbial burden in tissues following lung infection.

Transcriptiomics analysis reveals immue regulation and cell death regulation of IFNg in Pelodiscus sinensis STA cells

Our previous study showed that overexpression interferon gamma (IFNg) in soft-shelled turtle (Pelodiscus sinensis) artery cell line (STA cells) could induce soft-shelled turtle iridovirus (STIV)-infected cell death and inhibit STIV replication. To understand the molecular basis, four samples (named as Ctrl, Ctrl_STIV, IFNg and IFNg_STIV) including cells treatment with recombinant IFNg (rIFNg) or not, and infected or mock-infected with STIV were used to transcriptome analysis. A total of 77.18 Gb clean data were obtained and the clean data of each sample was over 5.97 Gb. Compared to the Ctrl group, there are 2172, 1192 and 2638 DEGs were detected, with 1127, 580 and 1349 up-regulated genes and 1045, 612, and 1259 down-regulated genes, in STA cells after treatment with IFNg, Ctrl_STIV and IFNg_STIV respectively. Subsequently, DEGs were annotated and enriched to GO items and the KEGG pathway, the result showed that immune activation of rIFNg in STA cells including Jak-STAT signaling activation, pattern recognition receptor (PRR) mediated innate immunity activation and cell death regulation etc. The activation of PRR mediated innate immunity may contribute to antiviral activity of IFNg, and it was not affected by STIV infection. From the aspect of cell death, rIFNg could regulated a series of cell death associated Go items including positive regulation and negative regulation, however some GO items including negative regulation of cell death were no longer enriched after STIV infection. The enriched death-related KEGG pathways including “Apoptosis” and “Necroptosis” were enriched in STA cells after rIFNg treatment, and however, besides “Necroptosis”, “Cellular senescence”, “p53 signaling pathway” were enriched after STIV infection. All of these indicate that rIFNg could build an antiviral activity in STA cells and may induce STIV-infected cell death through limiting the negative regulation of cell death and inducing necroptosis, apoptosis and cellular senescence.

Alcohol: Immunomodulatory Effects and Cancer.

Alcohol consumption has been linked to numerous pathologic conditions, including infectious diseases and several types of cancer. Alcohol exerts its modulatory effects on the immune system (IS) in a dose- and time-dependent manner. Numerous studies indicate that these alterations affect responses such as peripheral inflammation or decreased antibody production and promote chronic inflammation, leading to cell death. The molecular mechanisms underlying these effects involve generating an oxidative tissue environment, producing cell damage-associated molecular patterns (DAMPs), and activating pattern recognition receptors. In particular, toll-like receptors and their signaling system emerge as central elements whose activity is altered by alcohol intake. There is also some epidemiological evidence demonstrating the causal role of alcohol in the development of various types of cancer, such as head-and-neck cancer, esophageal cancer, colorectal cancer, liver cancer, and breast cancer. Most recent evidence suggests that factors related to alcohol consumption and cancer include increased levels of acetaldehyde, production of reactive oxygen species, alteration in DNA methylation, and modifications in retinoid metabolism. In addition, changes associated with alcohol use on the IS and intestinal microbiota may favor the growth of some types of tumors.

The Cytomegalovirus M35 Protein Directly Binds to the Interferon-β Enhancer and Modulates Transcription of Ifnb1 and Other IRF3-Driven Genes.

Induction of type I interferon (IFN) gene expression is among the first lines of cellular defense a virus encounters during primary infection. We previously identified the tegument protein M35 of murine cytomegalovirus (MCMV) as an essential antagonist of this antiviral system, showing that M35 interferes with type I IFN induction downstream of pattern-recognition receptor (PRR) activation. Here, we report structural and mechanistic details of M35's function. Determination of M35's crystal structure combined with reverse genetics revealed that homodimerization is a key feature for M35's immunomodulatory activity. In electrophoretic mobility shift assays (EMSAs), purified M35 protein specifically bound to the regulatory DNA element that governs transcription of the first type I IFN gene induced in nonimmune cells, Ifnb1. DNA-binding sites of M35 overlapped with the recognition elements of interferon regulatory factor 3 (IRF3), a key transcription factor activated by PRR signaling. Chromatin immunoprecipitation (ChIP) showed reduced binding of IRF3 to the host Ifnb1 promoter in the presence of M35. We furthermore defined the IRF3-dependent and the type I IFN signaling-responsive genes in murine fibroblasts by RNA sequencing of metabolically labeled transcripts (SLAM-seq) and assessed M35's global effect on gene expression. Stable expression of M35 broadly influenced the transcriptome in untreated cells and specifically downregulated basal expression of IRF3-dependent genes. During MCMV infection, M35 impaired expression of IRF3-responsive genes aside of Ifnb1. Our results suggest that M35-DNA binding directly antagonizes gene induction mediated by IRF3 and impairs the antiviral response more broadly than formerly recognized. IMPORTANCE Replication of the ubiquitous human cytomegalovirus (HCMV) in healthy individuals mostly goes unnoticed but can impair fetal development or cause life-threatening symptoms in immunosuppressed or -deficient patients. Like other herpesviruses, CMV extensively manipulates its hosts and establishes lifelong latent infections. Murine CMV (MCMV) presents an important model system as it allows the study of CMV infection in the host organism. We previously showed that during entry into host cells, MCMV virions release the evolutionary conserved protein M35 protein to immediately dampen the antiviral type I interferon (IFN) response induced by pathogen detection. Here, we show that M35 dimers bind to regulatory DNA elements and interfere with recruitment of interferon regulatory factor 3 (IRF3), a key cellular factor for antiviral gene expression. Thereby, M35 interferes with expression of type I IFNs and other IRF3-dependent genes, reflecting the importance for herpesviruses to avoid IRF3-mediated gene induction.

Arjunolic acid modulate pancreatic dysfunction by ameliorating pattern recognition receptor and canonical Wnt pathway activation in type 2 diabetic rats

BackgroundArjunolic acid (AA) is a potent phytochemical with multiple therapeutics effects. In this study, AA is evaluated on type 2 diabetic (T2DM) rats to understand the mechanism of β-cell linkage with Toll-like receptor 4 (TLR-4) and canonical Wnt signaling. However, its role in modulating TLR-4 and canonical Wnt/β-catenin crosstalk on insulin signaling remains unclear during T2DM.AimThe current study is aimed to examine the potential role of AA on insulin signaling and TLR-4-Wnt crosstalk in the pancreas of type 2 diabetic rats. MethodMultiple methods were used to determine molecular cognizance of AA in T2DM rats, when treated with different dosage levels. Histopathological and histomorphometry analysis was conducted using masson trichrome and H&E stains. While, protein and mRNA expressions of TLR-4/Wnt and insulin signaling were assessed using automated Western blotting (jess), immunohistochemistry, and RT-PCR. ResultsHistopathological findings revealed that AA had reversed back the T2DM-induced apoptosis and necrosis caused to rats pancreas. Molecular findings exhibited prominent effects of AA in downregulating the elevated level of TLR-4, MyD88, NF-κB, p-JNK, and Wnt/β-catenin by blocking TLR-4/MyD88 and canonical Wnt signaling in diabetic pancreas, while IRS-1, PI3K, and pAkt were all upregulated by altering the NF-κB and β-catenin crosstalk during T2DM. ConclusionOverall results, indicate that AA has potential to develop as an effective therapeutic in the treatment of T2DM associated meta-inflammation. However, future preclinical research at multiple dose level in a long-term chronic T2DM disease model is warranted to understand its clinical relevance in cardiometabolic disease.

IL-10 Modulates the Expression and Activation of Pattern Recognition Receptors in Mast Cells.

Mast cells (MCs) are involved in several immune-related responses, including those in bacterial infections, autoimmune diseases, inflammatory bowel diseases, and cancer, among others. MCs identify microorganisms by pattern recognition receptors (PRRs), activating a secretory response. Interleukin (IL)-10 has been described as an important modulator of MC responses; however, its role in PRR-mediated activation of MC is not fully understood. We analyzed the activation of TLR2, TLR4, TLR7 and Nucleotide-binding oligomerization domain-containing protein 2 (NOD2) in mucosal-like MCs (MLMCs) and peritoneum-derived cultured MCs (PCMCs) from IL-10-/- and wild-type (WT) mice. IL-10-/- mice showed a reduced expression of TLR4 and NOD2 at week 6 and TLR7 at week 20 in MLMC. In MLMC and PCMC, TLR2 activation induced a reduced secretion of IL-6 and TNFα in IL-10-/- MCs. TLR4- and TLR7-mediated secretion of IL-6 and TNFα was not detected in PCMCs. Finally, no cytokine release was induced by NOD2 ligand, and responses to TLR2 and TLR4 were lower in MCs at 20 weeks. These findings indicate that PRR activation in MCs depends on the phenotype, ligand, age, and IL-10.

Viperin from the dromedary camel: First report of an antiviral interferon-responsive gene from camelids

Viral infections activate pattern recognition receptors in the host, triggering an innate immune response that involves the production of interferons, which, in turn, stimulates the expression of antiviral effector genes. Viperin is one of the most highly induced interferon-stimulated genes and displays broad antiviral activity, especially against tick-borne viruses. Of late, camelid-borne zoonotic viruses have been on the rise in the Arabian Peninsula, but research into camelid antiviral effector genes has been limited. This is the first report of an interferon-responsive gene from the mammalian suborder Tylopoda to which modern camels belong. From camel kidney cells treated with dsRNA mimetic, we cloned viperin cDNA encoding 361 amino acid protein. Sequence analysis of camel viperin reveals high levels of amino acid conservation, particularly within the RSAD domain. Compared to kidney, the relative mRNA expression of viperin was higher in blood, lung, spleen, lymph nodes, and intestines. The in-vitro expression of viperin was induced by poly(I:C) and interferon treatment in camel kidney cell lines. Viperin expression was subdued in camel kidney cells infected with the camelpox virus during the early stages of infection, suggesting possible suppression by the virus. Overexpression of camel viperin through transient transfection significantly enhanced the resistance of cultured camel kidney cell lines to infection with camelpox virus. Research into the role of viperin in host immunity against emerging viral pathogens of camels will provide insight into novel mechanisms of antiviral activity of the protein, viral immune evasion strategies, and enable the development of better antivirals.