Immune profiling reveals umbilical cord blood mononuclear cells from South India display an IL-8 dominant, CXCL-10 deficient polyfunctional monocyte response to pathogen-associated molecular patterns (PAMPs) that is distinct from adult blood cells.

Neonate responses to pathogen-associated molecular patterns (PAMPS) differ from adults; such understanding is poor in Indian neonates, despite recognised significant infectious risk. Immune profiling analysis was undertaken of ten secreted mediators contextualised with cellular source induced by six PAMPs in umbilical cord (CB; n=21) and adult-blood (PBMC, n=14) from a tertiary care hospital in South India. Differential cytokine expression analysis (minimum log2-fold difference; adj p-value<0.05) identified bacterial PAMPs induced higher concentrations of IL-1ß, IL-10, TNF-α in adults versus IL-8, GM-CSF, IFN-γ and IL-2 in CB. CB responded to poly I:C and SARS-CoV-2 lysate with a dominant IL-8 response, whereas, in PBMC, CXCL-10 dominated poly I:C, but not SARS-CoV-2, responses, highlighting potential IL-8 importance, in absence of Type I Interferons, in antiviral CB immunity. Candida albicans was the only PAMP to uniformly induce higher secretion of effectors in CB. The predominant source of IL-8/IL-6/TNF-α/IL-1ß in both CB and PBMC was polyfunctional monocytes and IFN-γ /IL-2/IL-17 from innate lymphocytes. Correlation matrix analyses revealed IL-8 to be the most differentially regulated, correlating positively in CB versus negatively in PBMC with IL-6, GM-CSF, IFN-γ, IL-2, consistent with more negatively regulated cytokine modules in adults, potentially linked to higher anti-inflammatory IL-10. Cord and adult blood from India respond robustly to PAMPs with unique effector combinations. These data provide a strong foundation to monitor, explore, mechanisms that regulate such immunity during the life course, an area of significant global health importance given infection-related infant mortality incidence.

Infection and disruption of placental multidrug resistance (MDR) transporters: Implications for fetal drug exposure.

P-glycoprotein (P-gp, encoded by the ABCB1 gene) and breast cancer resistance protein (BCRP/ABCG2) are efflux multidrug resistance (MDR) transporters localized at the syncytiotrophoblast barrier of the placenta and protect the conceptus from drug and toxin exposure throughout pregnancy. Infection is an important modulator of MDR expression and function. This review comprehensively examines the effect of infection on the MDR transporters, P-gp and BCRP in the placenta. Infection PAMPs such as bacterial lipopolysaccharide (LPS) and viral polyinosinic-polycytidylic acid (poly I:C) and single-stranded (ss)RNA, as well as infection with Zika virus (ZIKV), Plasmodium berghei ANKA (modeling malaria in pregnancy - MiP) and polymicrobial infection of intrauterine tissues (chorioamnionitis) all modulate placental P-gp and BCRP at the levels of mRNA, protein and or function; with specific responses varying according to gestational age, trophoblast type and species (human vs. mice). Furthermore, we describe the expression and localization profile of Toll-like receptor (TLR) proteins of the innate immune system at the maternal-fetal interface, aiming to better understand how infective agents modulate placental MDR. We also highlight important gaps in the field and propose future research directions. We conclude that alterations in placental MDR expression and function induced by infective agents may not only alter the intrauterine biodistribution of important MDR substrates such as drugs, toxins, hormones, cytokines, chemokines and waste metabolites, but also impact normal placentation and adversely affect pregnancy outcome and maternal/neonatal health.

Genome-wide identification and characterization of Toll-like receptors (TLR) genes in Haliotis discus hannai, H. rufescens, and H. laevigata.

Toll-like receptors (TLRs) play essential roles in the innate immune system and have been extensively studied in mollusks. In this study, through a genome-wide search, TLR genes were identified as 29 in Haliotis discus hannai, 33 in H. rufescens, and 16 in H. laevigata. Domain analysis indicated that these TLR genes contain leucine-rich repeat (LRR) and Toll/IL-1 receptor (TIR) domains and exons ranging from 1 to 5. Polymorphism analysis showed that the TLRs in abalones did not have high diversities with 143 SNPs and no Indel in H. discus hannai, 92 SNPs and 3 Indels together with 6 missense mutations in H. rufescens, and no SNP or Indel in H. laevigata. The expression of 8 TLR genes in H. discus hannai was confirmed in the hepatopancreas, gill, hemolymph, gonads, intestine, muscle, and mantle. The expression of five TLR genes (out of 8) in gills (p < 0.05), three in hepatopancreas (p < 0.05), and three in hemolymph (p < 0.05) was upregulated separately in response to the infection caused by Vibrio parahaemolyticus. The findings in this study would contribute to a better understanding of the molecular immune mechanism of H. discus hannai against stimulation by V. parahaemolyticus and provide a basis for the study of TLRs in abalones.

COVID-19-Induced Myocarditis: Pathophysiological Roles of ACE2 and Toll-like Receptors.

The clinical manifestations of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection responsible for coronavirus disease 2019 (COVID-19) commonly include dyspnoea and fatigue, and they primarily involve the lungs. However, extra-pulmonary organ dysfunctions, particularly affecting the cardiovascular system, have also been observed following COVID-19 infection. In this context, several cardiac complications have been reported, including hypertension, thromboembolism, arrythmia and heart failure, with myocardial injury and myocarditis being the most frequent. These secondary myocardial inflammatory responses appear to be associated with a poorer disease course and increased mortality in patients with severe COVID-19. In addition, numerous episodes of myocarditis have been reported as a complication of COVID-19 mRNA vaccinations, especially in young adult males. Changes in the cell surface expression of angiotensin-converting enzyme 2 (ACE2) and direct injury to cardiomyocytes resulting from exaggerated immune responses to COVID-19 are just some of the mechanisms that may explain the pathogenesis of COVID-19-induced myocarditis. Here, we review the pathophysiological mechanisms underlying myocarditis associated with COVID-19 infection, with a particular focus on the involvement of ACE2 and Toll-like receptors (TLRs).

Antigens from the parasitic nematode Trichuris suis induce metabolic reprogramming and trained immunity to constrain inflammatory responses in macrophages.

Regulation of macrophage (Mɸ) function can maintain tissue homeostasis and control inflammation. Parasitic worms (helminths) are potent modulators of host immune and inflammatory responses. They have evolved various strategies to promote immunosuppression, including redirecting phagocytic cells toward a regulatory phenotype. Although soluble products from the whipworm Trichuris suis (TSPs) have shown significant effects on Mɸ function, the mechanisms underlying these modulatory effects are still not well understood. In this study, we find that TSPs suppressed inflammatory cytokines (TNF and IL-6) in Mɸs stimulated with a broad panel of TLR agonists, whilst inducing IL-10. Moreover, M1 markers such as MHCII, CD86, iNOS, and TNF were downregulated in TSP-treated Mɸs, without polarizing them towards an M2-like phenotype. We showed that TSPs could establish a suppressed activation state of Mɸs lasting at least for 72 h, indicating an anti-inflammatory innate training. Moreover, we found that TSPs, via repression of intracellular TNF generation, decreased its secretion rather than interfering with the release of surface-bound TNF. Metabolic analysis showed that TSPs promote oxidative phosphorylation (OXPHOS) without affecting glycolytic rate. Collectively, these findings expand our knowledge on helminth-induced immune modulation and support future investigations into the anti-inflammatory properties of TSPs for therapeutic purposes.

Oxy210, a Semi-Synthetic Oxysterol, Exerts Anti-Inflammatory Effects in Macrophages via Inhibition of Toll-like Receptor (TLR) 4 and TLR2 Signaling and Modulation of Macrophage Polarization.

Inflammatory responses by the innate and adaptive immune systems protect against infections and are essential to health and survival. Many diseases including atherosclerosis, osteoarthritis, rheumatoid arthritis, psoriasis, and obesity involve persistent chronic inflammation. Currently available anti-inflammatory agents, including non-steroidal anti-inflammatory drugs, steroids, and biologics, are often unsafe for chronic use due to adverse effects. The development of effective non-toxic anti-inflammatory agents for chronic use remains an important research arena. We previously reported that oral administration of Oxy210, a semi-synthetic oxysterol, ameliorates non-alcoholic steatohepatitis (NASH) induced by a high-fat diet in APOE*3-Leiden.CETP humanized mouse model of NASH and inhibits expression of hepatic and circulating levels of inflammatory cytokines. Here, we show that Oxy210 also inhibits diet-induced white adipose tissue inflammation in APOE*3-Leiden.CETP mice, evidenced by the inhibition of adipose tissue expression of IL-6, MCP-1, and CD68 macrophage marker. Oxy210 and related analogs exhibit anti-inflammatory effects in macrophages treated with lipopolysaccharide in vitro, mediated through inhibition of toll-like receptor 4 (TLR4), TLR2, and AP-1 signaling, independent of cyclooxygenase enzymes or steroid receptors. The anti-inflammatory effects of Oxy210 are correlated with the inhibition of macrophage polarization. We propose that Oxy210 and its structural analogs may be attractive candidates for future therapeutic development for targeting inflammatory diseases.

Role of genetic variants of Vitamin D receptor, Toll-like receptor 2 and Toll-like receptor 4 in extrapulmonary tuberculosis.

BACKGROUND: Tuberculosis (TB) is a chronic infectious disease caused by Mycobacterium tuberculosis (MTB). Vitamin D deficiency and vitamin D receptor (VDR) gene abnormalities confer susceptibility to tuberculosis. Toll-like receptors (TLRs), such as TLR-2, are also important mediators of inflammatory response against Mycobacterium tuberculosis. We evaluated VDR, TLR-2 and TLR-4 gene polymorphisms in patients with extrapulmonary tuberculosis (EPTB). OBJECTIVES: To find out a possible association of Vitamin D receptor (VDR) (rs731236), TLR-2 (196-174 Ins > Del) and TLR-4 (Thr399Ile) gene polymorphisms with extrapulmonary tuberculosis in ethnic Kashmiri population. METHODS: A total of 100 extrapulmunary tuberculosis cases and 102 healthy controls were analyzed for Vitamin D receptor (VDR) (rs731236), TLR-2 (196-174 ins > del) and TLR-4 (Thr399Ile) gene polymorphisms using PCR-RFLP and Allele-Specific PCR methods. RESULTS: We found increased frequency of TLR-4 Thr/Ile heterozygous genotype in cases as compared with healthy controls (22% vs 5.8%). Thus acting as a risk factor for extrapulmonary tuberculosis, as was elucidated from statistical analysis [OR, 4.5; 95% CI (1.74-11.68); P < 0.001]. In case of TLR-2 (196-174 ins > del) we observed significant differences in the homozygous variant (Del/Del) genotype of cases and controls (28% in cases & 2.94% in controls). Thus, TLR-2 (Del/Del) genotype acts as a strong risk factor for extrapulmonary tuberculosis predisposition [OR, 12.2; 95% CI (3.5-42.69); P < 0.001]. We did not find any significant differences in the genotypic distribution of (VDR) (rs731236) T > C SNP between cases and controls (P > 0.05). CONCLUSION: TLR-4 (Thr/Ile) and TLR-2 (Del/Del) act as significant risk factors for extrapulmonary tuberculosis predisposition in ethnic Kashmiri population.

Expatiating the molecular approaches of HMGB1 in diabetes mellitus: Highlighting signalling pathways via RAGE and TLRs.

Diabetes mellitus (DM) has become one of the major healthcare challenges worldwide in the recent times and inflammation being one of its key pathogenic process/mechanism affect several body parts including the peripheral and central nervous system. High-mobility group box 1 (HMGB1) is one of the major non-histone proteins that plays a key role in triggering the inflammatory response. Upon its release into the extracellular milieu, HMGB1 acts as an "alarmin" for the immune system to initiate tissue repair as a component of the host defense system. Furthermore, HMGB1 along with its downstream receptors like Toll-like receptors (TLRs) and receptors for advanced glycation end products (RAGE) serve as the suitable target for DM. The forthcoming research in the field of diabetes would potentially focus on the development of alternative approaches to target the centre of inflammation that is primarily mediated by HMGB1 to improve diabetic-related complications. This review covers the therapeutic actions of HMGB1 protein, which acts by activating the RAGE and TLR molecules to constitute a functional tripod system, in turn activating NF-κB pathway that contributes to the production of mediators for pro-inflammatory cytokines associated with DM. The interaction between TLR2 and TLR4 with ligands present in the host and the activation of RAGE stimulates various immune and metabolic responses that contribute to diabetes. This review emphasizes to elucidate the role of HMGB1 in the initiation and progression of DM and control over the inflammatory tripod as a promising therapeutic approach in the management of DM.

The Immunology of Syncytialized Trophoblast.

Multinucleate syncytialized trophoblast is found in three forms in the human placenta. In the earliest stages of pregnancy, it is seen at the invasive leading edge of the implanting embryo and has been called primitive trophoblast. In later pregnancy, it is represented by the immense, multinucleated layer covering the surface of placental villi and by the trophoblast giant cells found deep within the uterine decidua and myometrium. These syncytia interact with local and/or systemic maternal immune effector cells in a fine balance that allows for invasion and persistence of allogeneic cells in a mother who must retain immunocompetence for 40 weeks of pregnancy. Maternal immune interactions with syncytialized trophoblast require tightly regulated mechanisms that may differ depending on the location of fetal cells and their invasiveness, the nature of the surrounding immune effector cells and the gestational age of the pregnancy. Some specifically reflect the unique mechanisms involved in trophoblast cell-cell fusion (aka syncytialization). Here we will review and summarize several of the mechanisms that support healthy maternal-fetal immune interactions specifically at syncytiotrophoblast interfaces.

Human labour is associated with altered regulatory T cell function and maternal immune activation.

During human pregnancy, regulatory T cell (Treg ) function is enhanced and immune activation is repressed allowing the growth and development of the feto-placental unit. Here, we have investigated whether human labour is associated with a reversal of the pregnancy-induced changes in the maternal immune system. We tested the hypothesis that human labour is associated with a decline in Treg function, specifically their ability to modulate Toll-like receptor (TLR)-induced immune responses. We studied the changes in cell number, activation status and functional behaviour of peripheral blood, myometrial (myoMC) and cord blood mononuclear cells (CBMC) with the onset of labour. We found that Treg function declines and that Treg cellular targets change with labour onset. The changes in Treg function were associated with increased activation of myoMC, assessed by their expression of major histocompatibility complex (MHC) class II molecules and CBMC inflammatory cells. The innate immune system showed increased activation, as shown by altered monocyte and neutrophil cell phenotypes, possibly to be ready to respond to microbial invasion after birth or to contribute to tissue remodelling. Our results highlight changes in the function of the adaptive and innate immune systems that may have important roles in the onset of human labour.

Inflammation and joint destruction may be linked to the generation of cartilage metabolites of ADAMTS-5 through activation of toll-like receptors.

OBJECTIVE: Links between pain and joint degradation are poorly understood. We investigated the role of activation of Toll-like receptors (TLR) by cartilage metabolites in initiating and maintaining the inflammatory loop in OA causing joint destruction. METHODS: Synovial membrane explants (SMEs) were prepared from OA patients' synovial biopsies. SMEs were cultured for 10 days under following conditions: culture medium alone, OSM + TNFα, TLR2 agonist - Pam2CSK4, Pam3CSK4 or synthetic aggrecan 32-mer, TLR4 agonist - Lipid A. Release of pro-inflammatory and degradation biomarkers (acMMP3 and C3M) were measured by ELISA in conditioned media along with IL-6. Additionally, human cartilage was digested with ADAMTS-5, with or without the ADAMTS-5 inhibiting nanobody - M6495. Digested cartilage solution (DCS) and synthetic 32-mer were tested for TLR activation in SEAP based TLR reporter assay. RESULTS: Western blotting confirmed TLR2 and TLR4 in untreated OA synovial biopsies. TLR agonists showed an increase in release of biomarkers - acMMP3 and C3M in SME. Synthetic 32-mer showed no activation in the TLR reporter assay. ADAMTS-5 degraded cartilage fragments activated TLR2 in vitro. Adding M6495 - an anti-ADAMTS-5 inhibiting nanobody®, blocked ADAMTS-5-mediated DCS TLR2 activation. CONCLUSION: TLR2 is expressed in synovium of OA patients and their activation by synthetic ligands causes increased tissue turnover. ADAMTS-5-mediated cartilage degradation leads to release of aggrecan fragments which activates the TLR2 receptor in vitro. M6495 suppressed cartilage degradation by ADAMTS-5, limiting the activation of TLR2. In conclusion, pain and joint destruction may be linked to generation of ADAMTS-5 cartilage metabolites.

Toll-Like Receptors Signaling in the Tumor Microenvironment.

The involvement of inflammation in cancer progression is well-established. The immune system can play both tumor-promoting and -suppressive roles, and efforts to harness the immune system to help fight tumor growth are at the forefront of research. Of particular importance is the inflammatory profile at the site of the tumor, with respect to both the leukocyte population numbers, the phenotype of these cells, as well as the contribution of the tumor cells themselves. In this regard, the pro-inflammatory effects of pattern recognition receptor expression and activation in the tumor microenvironment have emerged as a relevant issue both for therapy and to understand tumor development.Pattern recognition receptors (PRRs) were originally recognized as components of immune cells, particularly innate immune cells, as detectors of pathogens. PRR signaling in immune cells activates them, inducing robust antimicrobial responses. In particular, toll-like receptors (TLRs) constitute a family of membrane-bound PRRs which can recognize pathogen-associated molecular patterns (PAMPs) carried by bacteria, virus, and fungi. In addition, PRRs can recognize products generated by stressed cells or damaged tissues, namely damage-associated molecular patterns or DAMPS. Taking into account the role of the immune system in fighting tumors together with the presence of immune cells in the microenvironment of different types of tumors, strategies to activate immune cells via PRR ligands have been envisioned as an anticancer therapeutic approach.In the last decades, it has been determined that PRRs are present and functional on nonimmune cells and that their activation in these cells contributes to the inflammation in the tumor microenvironment. Both tumor-promoting and antitumor effects have been observed when tumor cell PRRs are activated. This argues against nonspecific activation of PRR ligands in the tumor microenvironment as a therapeutic approach. Therefore, the use of PRR ligands for anticancer therapy might benefit from strategies that specifically deliver these ligands to immune cells, thus avoiding tumor cells in some settings. This review focuses on these aspects of TLR signaling in the tumor microenvironment.

Revisiting Platelets and Toll-Like Receptors (TLRs): At the Interface of Vascular Immunity and Thrombosis.

While platelet function has traditionally been described in the context of maintaining vascular integrity, recent evidence suggests that platelets can modulate inflammation in a much more sophisticated and nuanced manner than previously thought. Some aspects of this expanded repertoire of platelet function are mediated via expression of Toll-like receptors (TLRs). TLRs are a family of pattern recognition receptors that recognize pathogen-associated and damage-associated molecular patterns. Activation of these receptors is crucial for orchestrating and sustaining the inflammatory response to both types of danger signals. The TLR family consists of 10 known receptors, and there is at least some evidence that each of these are expressed on or within human platelets. This review presents the literature on TLR-mediated platelet activation for each of these receptors, and the existing understanding of platelet-TLR immune modulation. This review also highlights unresolved methodological issues that potentially contribute to some of the discrepancies within the literature, and we also suggest several recommendations to overcome these issues. Current understanding of TLR-mediated platelet responses in influenza, sepsis, transfusion-related injury and cardiovascular disease are discussed, and key outstanding research questions are highlighted. In summary, we provide a resource-a "researcher's toolkit"-for undertaking further research in the field of platelet-TLR biology.

Comparison of PPAR Ligands as Modulators of Resolution of Inflammation, via Their Influence on Cytokines and Oxylipins Release in Astrocytes.

Neuroinflammation is a key process of many neurodegenerative diseases and other brain disturbances, and astrocytes play an essential role in neuroinflammation. Therefore, the regulation of astrocyte responses for inflammatory stimuli, using small molecules, is a potential therapeutic strategy. We investigated the potency of peroxisome proliferator-activated receptor (PPAR) ligands to modulate the stimulating effect of lipopolysaccharide (LPS) in the primary rat astrocytes on (1) polyunsaturated fatty acid (PUFAs) derivative (oxylipins) synthesis; (2) cytokines TNFα and interleukin-10 (IL-10) release; (3) p38, JNK, ERK mitogen-activated protein kinase (MAPKs) phosphorylation. Astrocytes were exposed to LPS alone or in combination with the PPAR ligands: PPARα (fenofibrate, GW6471); PPARß (GW501516, GSK0660); PPARγ (rosiglitazone, GW9662). We detected 28 oxylipins with mass spectrometry (UPLC-MS/MS), classified according to their metabolic pathways: cyclooxygenase (COX), cytochrome P450 monooxygenases (CYP), lipoxygenase (LOX) and PUFAs: arachidonic (AA), docosahexaenoic (DHA), eicosapentaenoic (EPA). All tested PPAR ligands decrease COX-derived oxylipins; both PPARß ligands possessed the strongest effect. The PPARß agonist, GW501516 is a strong inducer of pro-resolution substances, derivatives of DHA: 4-HDoHE, 11-HDoHE, 17-HDoHE. All tested PPAR ligands decreased the release of the proinflammatory cytokine, TNFα. The PPARß agonist GW501516 and the PPARγ agonist, rosiglitazone induced the IL-10 release of the anti-inflammatory cytokine, IL-10; the cytokine index, (IL-10/TNFα) was more for GW501516. The PPARß ligands, GW501516 and GSK0660, are also the strongest inhibitors of LPS-induced phosphorylation of p38, JNK, ERK MAPKs. Overall, our data revealed that the PPARß ligands are a potential pro-resolution and anti-inflammatory drug for targeting glia-mediated neuroinflammation.

Inhibitor of Hyaluronic Acid Synthesis 4-Methylumbelliferone as an Anti-Inflammatory Modulator of LPS-Mediated Astrocyte Responses.

Astrocytes are glial cells that play an important role in neuroinflammation. Astrocytes respond to many pro-inflammatory stimuli, including lipopolysaccharide (LPS), an agonist of Toll-like receptor 4 (TLR4). Regulatory specificities of inflammatory signaling pathways are still largely unknown due to the ectodermal origin of astrocytes. Recently, we have shown that hyaluronic acid (HA) may form part of astrocyte inflammatory responses. Therefore, we tested 4-methylumbelliferone (4-MU), a specific inhibitor of HA synthesis, as a possible regulator of LPS-mediated responses. Rat primary astrocytes were treated with LPS with and without 4-MU and gene expression levels of inflammatory (interleukins 1ß, (IL-1ß), 6, (IL-6), tumor necrosis factor alpha TNFα,) and resolution interleukin 10 (IL-10) markers were evaluated via real-time PCR and western blot. The release of cytokines and HA was determined by ELISA. Oxylipin profiles were measured by ultra-performance liquid chromatography-tandem mass spectrometry (UPLC-MS/MS) analysis. Our data show that 4-MU (i) has anti-inflammatory effects in the course of TLR4 activation, decreasing the cytokines level TNFα, IL-6 and IL-1ß and increasing IL-10, (ii) downregulates prostaglandin synthesis but not via cyclooxygenases COX-1 and COX-2 pathways, (iii) modulates HA synthesis and decreases LPS-induced HA synthase mRNA expression (HAS-1, HAS-2) but does not have an influence on HAS-3, HYAL1 and HYAL2 mRNAs; (iv) the effects of 4-MU are predominantly revealed via JNK but not p38, ERK mitogen-activated protein kinases (MAPKs) or nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB) pathways. For the first time, it is shown that 4-MU possesses the useful potential to regulate an inflammatory astrocyte response.

The immunopathogenesis of fibrosis in systemic sclerosis.

Systemic sclerosis (SSc) is an idiopathic systemic autoimmune disease. It is characterized by a triad of hallmarks: immune dysfunction, fibrosis and vasculopathy. Immune dysfunction in SSc is characterized by the activation and recruitment of immune cells and the production of autoantibodies and cytokines. How immune abnormalities link the fibrosis and vasculopathy in SSc is poorly understood. A plethora of immune cell types are implicated in the immunopathogenesis of SSc, including T cells, B cells, dendritic cells, mast cells and macrophages. How these different cell types interact to contribute to SSc is complicated, and can involve cell-to-cell interactions and communication via cytokines, including transforming growth factor (TGF)-ß, interleukin (IL)-6 and IL-4. We will attempt to review significant and recent research demonstrating the importance of immune cell regulation in the immunopathogenesis of SSc with a particular focus on fibrosis.

Patients with tumour necrosis factor (TNF) receptor-associated periodic syndrome (TRAPS) are hypersensitive to Toll-like receptor 9 stimulation.

Tumour necrosis factor receptor-associated periodic syndrome (TRAPS) is a hereditary autoinflammatory disorder characterized by recurrent episodes of fever and inflammation. It is associated with autosomal dominant mutations in TNFRSF1A, which encodes tumour necrosis factor receptor 1 (TNF-R1). Our aim was to understand the influence of TRAPS mutations on the response to stimulation of the pattern recognition Toll-like receptor (TLR)-9. Peripheral blood mononuclear cells (PBMCs) and serum were isolated from TRAPS patients and healthy controls: serum levels of 15 proinflammatory cytokines were measured to assess the initial inflammatory status. Interleukin (IL)-1ß, IL-6, IL-8, IL-17, IL-22, tumour necrosis factor (TNF)-α, vascular endothelial growth factor (VEGF), interferon (IFN)-γ, monocyte chemoattractant protein 1 (MCP-1) and transforming growth factor (TGF)-ß were significantly elevated in TRAPS patients' sera, consistent with constitutive inflammation. Stimulation of PBMCs with TLR-9 ligand (ODN2006) triggered significantly greater up-regulation of proinflammatory signalling intermediates [TNF receptor-associated factor (TRAF 3), IL-1 receptor-associated kinase-like 2 (IRAK2), Toll interacting protein (TOLLIP), TRAF6, phosphorylated transforming growth factor-ß-activated kinase 1 (pTAK), transforming growth factor-ß-activated kinase-binding protein 2 (TAB2), phosphorylated TAK 2 (pTAB2), IFN-regulatory factor 7 (IRF7), receptor interacting protein (RIP), nuclear factor kappa B (NF-κB) p65, phosphorylated NF-κB p65 (pNF-κB p65) and mitogen-activated protein kinase kinase (MEK1/2)] in TRAPS patients' PBMCs. This up-regulation of proinflammatory signalling intermediates and raised serum cytokines occurred despite concurrent anakinra treatment and no overt clinical symptoms at time of sampling. These novel findings further demonstrate the wide-ranging nature of the dysregulation of innate immune responses underlying the pathology of TRAPS and highlights the need for novel pathway-specific therapeutic treatments for this disease.

Innate immunity in the postmortem brain of depressed and suicide subjects: Role of Toll-like receptors.

Abnormalities of Toll-like receptors (TLRs) have been implicated in the pathophysiology of depression and suicide. Interactions of TLRs with pathogen-associated molecular patterns (PAMP) and damage-associated molecular patterns (DAMP) initiate signaling through myeloid differentiation primary response-88 (MyD88) and produce cytokines through the activation of the transcription factor nuclear factor kappa beta (NF-kB). We have earlier shown an increase in the protein and mRNA expression of TLR3 and TLR4 in the prefrontal cortex (PFC) of depressed suicide (DS) subjects compared with normal control (NC) subjects. To examine if other TLRs are altered in postmortem brain, we have now determined the protein and mRNA expression of other TLRs (TLR1, TLR2, TLR5, TLR6, TLR7, TLR8, TLR9 and TLR10) in the PFC of DS, depressed non-suicide (DNS), non-depressed suicide (NDS) and NC subjects. We determined the protein expression by Western blot and mRNA expression levels by real-time PCR (qPCR) in the PFC of 24 NC, 24 DS, 12 DNS and 11 NDS subjects. Combined with our previous study of TLR3 and TLR4, we found that the protein expression of TLR2, TLR3, TLR4, TLR6 and TLR10, and mRNA expression of TLR2 and TLR3 was significantly increased in the DS group compared with NC group. This study demonstrated that certain specific TLRs are altered in DS subjects, and hence those TLRs may be appropriate targets for the development of therapeutic agents for the treatment of suicidal behavior.

Innate immune response in patients with acute Zika virus infection.

Innate immunity receptors (Toll-like receptors/TLRs and RIG-like receptors/RLRs) are important for the initial recognition of Zika virus (ZIKV), modulation of protective immune response, and IFN-α and IFN-ß production. Immunological mechanisms involved in protection or pathology during ZIKV infection have not yet been determined. In this study, we evaluated the mRNA expression of innate immune receptors (TLR3, TLR7, TLR8, TLR9, melanoma differentiation-associated protein 5/MDA-5, and retinoic acid inducible gene/RIG-1), its adapter molecules (Myeloid Differentiation Primary Response Gene 88/Myd88, Toll/IL-1 Receptor Domain-Containing Adaptor-Inducing IFN-ß/TRIF), and cytokines (IL-6, IL-12, TNF-α, IFN-α, IFN-ß, and IFN-γ) in the acute phase of patients infected by ZIKV using real-time PCR in peripheral blood. Patients with acute ZIKV infection had high expression of TLR3, IFN-α, IFN-ß, and IFN-γ when compared to healthy controls. In addition, there was a positive correlation between TLR3 expression compared to IFN-α and IFN-ß. Moreover, viral load is positively correlated with TLR8, RIG-1, MDA-5, IFN-α, and IFN-ß. On the other hand, patients infected by ZIKV showed reduced expression of RIG-1, TLR8, Myd88, and TNF-α molecules, which are also involved in antiviral immunity. Similar expressions of TLR7, TLR9, MDA-5, TRIF, IL-6, and IL-12 were observed between the group of patients infected with ZIKV and control subjects. Our results indicate that acute infection (up to 5 days after the onset of symptoms) by ZIKV in patients induces the high mRNA expression of TLR3 correlated to high expression of IFN-γ, IFN-α, and IFN-ß, even though the high viral load is correlated to high expression of TLR8, RIG-1, MDA-5, IFN-α, and IFN-ß in ZIKV patients.

High and Low Molecular Weight Hyaluronic Acid Differentially Influences Oxylipins Synthesis in Course of Neuroinflammation.

Hyaluronic acid (HA), a major glycosaminoglycan of the extracellular matrix, has cell signaling functions that are dependent on its molecular weight. Anti-inflammatory effects for high-molecular-weight (HMW) HA and pro-inflammatory effects for low-molecular-weight (LMW) HA effects were found for various myeloid cells, including microglia. Astrocytes are cells of ectodermal origin that play a pivotal role in brain inflammation, but the link between HA with different molecular weights and an inflammatory response in these cells is not clear. We tested the effects of LMW and HMW HA in rat primary astrocytes, stimulated with Poly:IC (PIC, TLR3 agonist) and lipopolysaccharide (LPS, TLR4 agonist). Oxylipin profiles were measured by the UPLC-MS/MS analysis and metabolites HDoHEs (from docosahexaenoic acid), -HETEs, prostaglandins (from arachidonic acid), DiHOMEs and HODEs (from linoleic acid) were detected. Both, HMW and LMW HA downregulated the cyclooxygenase-mediated polyunsaturated fatty acids metabolism, LMW also reduced lipoxygenase-mediated fatty acid metabolism. Taken together, the data show that both LMW and HMW (i) influence themselves on cytokines (TNFα, IL-6, IL-10), enzymes iNOS, COX-2, and oxylipin levels in extracellular medium of cultured astrocytes, (ii) induced cellular adaptations in long-term applications, (iii) modulate TLR4- and TLR3-signaling pathways. The effects of HMW and LMW HA are predominantly revealed in TLR4- and TLR3- mediated responses, respectively.