Toll-like Receptor Responses Research Articles

Toll-like receptors in neuroinflammation, neurodegeneration, and alcohol-induced brain damage.

Toll-like receptors (TLRs) or pattern recognition receptors respond to pathogen-associated molecular patterns (PAMPs) or internal damage-associated molecular patterns (DAMPs). TLRs are integral membrane proteins with both extracellular leucine-rich and cytoplasmic domains that initiate downstream signaling through kinases by activating transcription factors like AP-1 and NF-κB, which lead to the release of various inflammatory cytokines and immune modulators. In the central nervous system, different TLRs are expressed mainly in microglia and astroglial cells, although some TLRs are also expressed in oligodendroglia and neurons. Activation of TLRs triggers signaling cascades by the host as a defense mechanism against invaders to repair damaged tissue. However, overactivation of TLRs disrupts the sustained immune homeostasis-induced production of pro-inflammatory molecules, such as cytokines, miRNAs, and inflammatory components of extracellular vesicles. These inflammatory mediators can, in turn, induce neuroinflammation, and neural tissue damage associated with many neurodegenerative diseases. This review discusses the critical role of TLRs response in Alzheimer's disease, Parkinson's disease, ischemic stroke, amyotrophic lateral sclerosis, and alcohol-induced brain damage and neurodegeneration.

Microbial recognition regulates intestinal epithelial growth in homeostasis and disease.

The intestine is constantly exposed to a dynamic community of microbes. Intestinal epithelial cells respond to microbes through evolutionarily conserved recognition pathways, such as the immune deficiency (IMD) pathway of Drosophila, the Toll-like receptor (TLR) response of flies and vertebrates, and the vertebrate nucleotide-binding oligomerization domain (NOD) pathway. Microbial recognition pathways are tightly controlled to respond effectively to pathogens, tolerate the microbiome, and limit intestinal disease. In this review, we focus on contributions of different model organisms to our understanding of how epithelial microbe recognition impacts intestinal proliferation and differentiation in homeostasis and disease. In particular, we compare how microbes and subsequent recognition by the intestine influences barrier integrity, intestinal repair and tumorigenesis in Drosophila, zebrafish, mice, and organoids. In addition, we discuss the importance of microbial recognition in homeostatic intestinal growth and discuss how immune pathways directly impact stem cell and crypt dynamics.

Phospholipase A 2 : A Paradigm for Allosteric Regulation by Membranes and Cell Signaling

Our laboratory discovered and demonstrated that membranes interact allosterically with enzymes to regulate cell signaling and metabolic pathways leading to inflammation1. We have recently employed substrate lipidomics coupled with molecular dynamics to reveal enzyme specificity linked to highly specific hydrophobic binding sites for the sn-2 fatty acyl chains in membrane phospholipid substrates2. We discovered unexpected headgroup and acyl chain specificity for each of the major human phospholipase A2 (PLA2) enzymes that explains the observed specificity at a new atomic level. A unique hydrophobic binding site — and not each enzyme's catalytic residues or polar headgroup binding site — dominates each enzyme's specificity. Each PLA2 shows unique specificity for its required fatty acid ranging from pro-inflammatory omega-6 arachidonic acid or anti-inflammatory fish oil omega-3 EPA and DHA; others favor membrane remodeling linolenic acid, or antibacterial saturated fatty acids, or oxidized fatty acids in LDL. Each PLA2 releases a specific fatty acid after the enzyme associates allosterically with membranes and extracts a single phospholipid substrate into its catalytic site. Stereospecific inhibitors3 have been designed for the specific sites. After decades of advances in lipid research, we can now correlate PLA2 specificity and inhibition potency with molecular structure and physiological function using a novel lipidomics platform that provides a paradigm for protein-membrane lipid interactions in general. From the cell signaling and inflammation perspective, the omics revolution began with genomics, proteomics and metabolomics, but lipidomics now dominates as the largest number of cellular metabolites are lipids, many playing critical roles in cell signaling, and over 40,000 distinct molecular species have been identified by the LIPID MAPS Consortium (www.lipidmaps.org). We have also employed lipidomics analysis to characterize the in vivo role of specific PLA2 enzymes in initiating the inflammatory response and cellular lipid signaling of Toll-like (TLR4) and purinergic (P2X7) receptors in stimulated macrophages as models of bacterial infection and inflammation4. We can now explain in vivo cellular specificity by in vitro enzyme specificity, inhibition and pro-inflammatory and pro-resolution lipid mediator formation pathways and inhibition5. 1Dennis EA (2016) REFLECTIONS: Liberating chiral lipid mediators, inflammatory enzymes and LIPID MAPS from biological grease, J Biol Chem, 291, 24431–48. 2Mouchlis VC, Chen Y, McCammon JA, Dennis EA (2018) Membrane allostery and unique hydrophobic sites promote enzyme substrate specificity, J Am Chem Soc, 140, 3285-91. 3Mouchlis VD, Armando AM, Dennis EA (2019) Substrate specific inhibition constants for phospholipase A2 acting on unique phospholipid substrates in mixed micelles and membranes using lipidomics. J Med Chem, 62, 1999-2007. 4Dennis EA, Norris PA (2015) Eicosanoid storm in infection and inflammation. Nature Immunology Reviews, 15, 511-523. 5Navratil AR, Shchepinov MS, Dennis EA (2018) Lipidomics reveals dramatic physiological kinetic isotope effects during the enzymatic oxygenation of polyunsaturated fatty acids ex vivo. J Am Chem Soc, 140, 235-43.

Evaluation of Molecules Involved in Intestinal Changes Induced by Lawsonia intracellularis Infection

Lawsonia intracellularis (LI)is an obligate intracellular bacterium that causes proliferative enteropathy (PE) in various species including pigs, horses, rabbits, dogs and hamsters. Infection by LI results in the expansion of proliferative SOX9 positive cells and reduction of enterocytes and goblet cells. In a previous study, RNAseq analysis of laser micro-dissected ilea of infected pigs showed up-regulation of ADAMTS1 metallopeptidase which codes for surface proteins with adhesion and protease activity. The immune related marker ­unc-93 homolog (UNC93B1, necessary for toll-like receptors response) and the tetraspanin CD63, that participates in cell differentiation, were downregulated. We hypothesize that LI regulates cell proliferation and differentiation through ADAMTS1 and CD63 genes and avoids immune response through UNC93B1. In the current study, swine enteroids were used to model LI infection. We analyzed gene expression of MUC2 (marker for goblet cells), SOX9, ADAMTS1, CD63 and UNC93B1 in swine enteroids infected with LI for 3, 5, 7, 14 and 21 days. Published studies showed that expression of MUC2 and SOX9 in infected pigs decreased on day 7 and was followed by a decreased SOX9 expression at days 14 and 21. We observed no changes in the expression of MUC2 in infected enteroids. SOX9 expression in infected compared with uninfected enteroids at the same time points reached a maximum of 44% increase on day 7 followed by a decrease in expression by 68% and 33% on days 14 and 21 respectively. Both infected enteroids (fold change: 6.63) and tissue samples (fold change: 1.4) from infected pigs showed increased expression of ADAMTS1 on day 5. Expression of UNC93B1 in infected tissues increased from days 5 (19%) to 15 (43%) while no change was observed on day 5 and increased expression was observed on day 7 (fold change: 2.98) in enteroids. CD63 expression in the tissues increased from day 5 (45%) to 15 (22%) while increase in expression on day 5 (fold change: 1.23) and day 7 (fold change: 3.28) were also observed in enteroids. Overall, swine enteroids replicated most of the changes observed in pigs for MUC2, SOX9, ADAMTS1 and UNC93B1, suggesting infected swine enteroids are a viable model to study PE-associated changes. A probable explanation for the differences in response of enteroids to LI infection in MUC2 and SOX9 expression can be the permissiveness of the culture system for infection since there is no bacterial competition, passage through the gastric environment, or anti-microbial products present in the in vitro system.

Toll-Like Receptor and Cytokine Responses to Infection with Endogenous and Exogenous Koala Retrovirus, and Vaccination as a Control Strategy.

Koala populations are currently declining and under threat from koala retrovirus (KoRV) infection both in the wild and in captivity. KoRV is assumed to cause immunosuppression and neoplastic diseases, favoring chlamydiosis in koalas. Currently, 10 KoRV subtypes have been identified, including an endogenous subtype (KoRV-A) and nine exogenous subtypes (KoRV-B to KoRV-J). The host’s immune response acts as a safeguard against pathogens. Therefore, a proper understanding of the immune response mechanisms against infection is of great importance for the host’s survival, as well as for the development of therapeutic and prophylactic interventions. A vaccine is an important protective as well as being a therapeutic tool against infectious disease, and several studies have shown promise for the development of an effective vaccine against KoRV. Moreover, CRISPR/Cas9-based genome editing has opened a new window for gene therapy, and it appears to be a potential therapeutic tool in many viral infections, which could also be investigated for the treatment of KoRV infection. Here, we discuss the recent advances made in the understanding of the immune response in KoRV infection, as well as the progress towards vaccine development against KoRV infection in koalas.

Targeting Macrophage Migration Inhibitory Factor in Acute Pancreatitis and Pancreatic Cancer.

Macrophage migration inhibitory factor (MIF) is a pleiotropic cytokine implicated in the pathogenesis of inflammation and cancer. It is produced by various cells and circulating MIF has been identified as a biomarker for a range of diseases. Extracellular MIF mainly binds to the cluster of differentiation 74 (CD74)/CD44 to activate downstream signaling pathways. These in turn activate immune responses, enhance inflammation and can promote cancer cell proliferation and invasion. Extracellular MIF also binds to the C-X-C chemokine receptors cooperating with or without CD74 to activate chemokine response. Intracellular MIF is involved in Toll-like receptor and inflammasome-mediated inflammatory response. Pharmacological inhibition of MIF has been shown to hold great promise in treating inflammatory diseases and cancer, including small molecule MIF inhibitors targeting the tautomerase active site of MIF and antibodies that neutralize MIF. In the current review, we discuss the role of MIF signaling pathways in inflammation and cancer and summarize the recent advances of the role of MIF in experimental and clinical exocrine pancreatic diseases. We expect to provide insights into clinical translation of MIF antagonism as a strategy for treating acute pancreatitis and pancreatic cancer.

Transcriptome-wide identification and characterization of toll-like receptors response to Vibrio anguillarum infection in Manila clam (Ruditapes philippinarum)

The Manila clam (Ruditapes philippinarum), one of the major marine aquaculture species in China, is susceptible to infection with the pathogen Vibrio, which results in massive mortality and economic losses. Toll-like receptors (TLRs) are significant pattern recognition receptors (PRRs) of innate immunity that are involved in immune regulation against pathogenic invasion. Molecular characterization of Manila clam TLRs and investigations of their immune functions are essential to prevent and control Vibrio infection. In the present research, eight cDNA sequences of R. philippinarum TLRs (RpTLRs) were identified from previous transcriptome libraries and then classified into four groups, namely, P-TLR (one sequence), V-TLR (one sequence), Ls-TLR (two sequences) and sP-TLR (four sequences), based on the corresponding LRR domain arrangement of their protein structures within the typical TLR motifs. A selective pressure test firstly suggested that the molluscan P-TLR, V-TLR, Ls-TLR and sP-TLR families underwent positive selection, and different numbers of positive selection sites (PSSs) were identified in different domains of the four types of RpTLRs, as determined by PAML and analysis of website data. These findings indicated that the evolution of RpTLRs may be associated with their immune recognition and function. Furthermore, tissue-specific expression analysis showed that all RpTLRs were ubiquitously expressed in all test tissues and were dominant in hemocytes. Quantitative real-time PCR revealed that the cDNA expression of all eight RpTLRs was upregulated after injection with Vibrio anguillarum (P < 0.01) in R. philippinarum hemocytes, revealing that these RpTLRs play important roles in responding to pathogenic stimulation. In summary, these findings provide a foundation for future investigations of the molecular classification and evolutionary patterns of Toll-like receptors in invertebrates, and the innate immune responses of TLR signaling pathways in Mollusca.

The transcription factors GFI1 and GFI1B as modulators of the innate and acquired immune response.

GFI1 and GFI1B are small nuclear proteins of 45 and 37kDa, respectively, that have a simple two-domain structure: The first consists of a group of six c-terminal C2H2 zinc finger motifs that are almost identical in sequence and bind to very similar, specific DNA sites. The second is an N-terminal 20 amino acid SNAG domain that can bind to the pocket of the histone demethylase KDM1A (LSD1) near its active site. When bound to DNA, both proteins act as bridging factors that bring LSD1 and associated proteins into the vicinity of methylated substrates, in particular histone H3 or TP53. GFI1 can also bring methyl transferases such as PRMT1 together with its substrates that include the DNA repair proteins MRE11 and 53BP1, thereby enabling their methylation and activation. While GFI1B is expressed almost exclusively in the erythroid and megakaryocytic lineage, GFI1 has clear biological roles in the development and differentiation of lymphoid and myeloid immune cells. GFI1 is required for lymphoid/myeloid and monocyte/granulocyte lineage decision as well as the correct nuclear interpretation of a number of important immune-signaling pathways that are initiated by NOTCH1, interleukins such as IL2, IL4, IL5 or IL7, by the pre TCR or -BCR receptors during early lymphoid differentiation or by T and B cell receptors during activation of lymphoid cells. Myeloid cells also depend on GFI1 at both stages of early differentiation as well as later stages in the process of activation of macrophages through Toll-like receptors in response to pathogen-associated molecular patterns. The knowledge gathered on these factors over the last decades puts GFI1 and GFI1B at the center of many biological processes that are critical for both the innate and acquired immune system.

TLR4 Response to LPS Is Reinforced by Urokinase Receptor.

GPI-anchored uPAR is the receptor for the extracellular serine protease urokinase-type plasminogen activator (uPA). Though uPAR role in inflammatory processes is documented, underlying mechanisms are not fully understood. In this study we demonstrate that uPAR is a part of Toll-like receptor 4 (TLR4) interactome. Downregulation of uPAR expression resulted in diminished LPS-induced TLR4 signaling, less activation of NFκB, and decreased secretion of inflammatory mediators in myeloid and non-myeloid cells in vitro. In vivo uPAR−/− mice demonstrated better survival, strongly diminished inflammatory response and better organ functions in cecal ligation and puncture mouse polymicrobial sepsis model. Mechanistically, GPI-uPAR and soluble uPAR colocalized with TLR4 on the cell membrane and interacted with scavenger receptor CD36. Our data show that uPAR can interfere with innate immunity response via TLR4 and this mechanism represents a potentially important target in inflammation and sepsis therapy.

Phosphatidylinositol-4-kinase IIα licenses phagosomes for TLR4 signaling and MHC-II presentation in dendritic cells

Toll-like receptor (TLR) recruitment to phagosomes in dendritic cells (DCs) and downstream TLR signaling are essential to initiate antimicrobial immune responses. However, the mechanisms underlying TLR localization to phagosomes are poorly characterized. We show herein that phosphatidylinositol-4-kinase IIα (PI4KIIα) plays a key role in initiating phagosomal TLR4 responses in murine DCs by generating a phosphatidylinositol-4-phosphate (PtdIns4P) platform conducive to the binding of the TLR sorting adaptor Toll-IL1 receptor (TIR) domain-containing adaptor protein (TIRAP). PI4KIIα is recruited to maturing lipopolysaccharide (LPS)-containing phagosomes in an adaptor protein-3 (AP-3)-dependent manner, and both PI4KIIα and PtdIns4P are detected on phagosomal membrane tubules. Knockdown of PI4KIIα-but not the related PI4KIIβ-impairs TIRAP and TLR4 localization to phagosomes, reduces proinflammatory cytokine secretion, abolishes phagosomal tubule formation, and impairs major histocompatibility complex II (MHC-II) presentation. Phagosomal TLR responses in PI4KIIα-deficient DCs are restored by reexpression of wild-type PI4KIIα, but not of variants lacking kinase activity or AP-3 binding. Our data indicate that PI4KIIα is an essential regulator of phagosomal TLR signaling in DCs by ensuring optimal TIRAP recruitment to phagosomes.

Toll-Like Receptor-Dependent Antiviral Responses at the Maternal-Fetal Interface

The maternal–fetal interface is a key barrier to protect the fetus from infection. Toll-like receptors (TLRs) at the maternal–fetal interface are involved in antiviral responses. TLRs are expressed in both maternal decidua and fetal trophoblasts. Virus-induced activation of TLR signaling pathways triggers the release of interferon-related antiviral molecules and other inflammatory cytokines and/or chemokines by the host innate immune system, which may disrupt immune tolerance at the maternal–fetal interface and lead to pregnancy complications. In this review, we summarize the state of knowledge on the most common viral infections during pregnancy, antiviral TLR responses at the maternal–fetal interface, and TLR-associated pregnancy complications.

Effect of bacterial LPS, poly I:C and temperature on the immune response of coelomocytes in short term cultures of red sea urchin (Loxechinus albus)

In echinoderms, the immune system plays a relevant role in defense against infection by pathogens. Particularly, in sea urchins, the immune system has been shown to be complex, especially in terms of the variety of immune genes and molecules described. A key component of the response to external pathogens are the Toll-like receptors (TLRs), which are a well-characterized class of pattern recognition receptors (PRRs) that participate in the recognition of pathogen-associated molecular patterns (PAMPs). Despite the fact that TLRs have been described in several sea urchin species, for the red sea urchin (Loxechinus albus), which is one of the most important sea urchins across the world in terms of fisheries, limited information on the TLR-mediated immune response exists. In the present study, for the first time, we evaluated the effect of thermal stress, LPS and poly I:C treatment on the coelomocyte immune response of Loxechinus albus to determine how these factors modulate TLR and strongylocin (antimicrobial peptides of echinoderms) responses. We show that the tlr3-like, tlr4-like, tlr6-like and tlr8-like transcripts are modulated by poly I:C, while LPS only modulates the tlr4-like response; there was no effect of temperature on TLR expression, as evaluated by RT-qPCR. Additionally, we showed that strongylocin-1 and strongylocin-2 are modulated in response to simulated viral infection with poly I:C, providing the first evidence of strongylocin expression in L. albus. Finally, we determined that temperature and LPS modify the viability of coelomocytes, while poly I:C treatment did not affect the viability of these cells. This study contributes to the knowledge of immune responses in sea urchins to improve the understanding of the role of TLRs and strongylocins in echinoderms.

The second coming: The comeback of the live vaccines.

Live vaccines are gradually replaced by protein-based vaccines given the latter's better safety. But live vaccines seem to be more efficient via stimulation of the innate immune system.

Genetic and Functional Characterization of Toll-Like Receptor Responses in Immunocompetent Patients With CMV Mononucleosis.

Background: Human cytomegalovirus (CMV) modulates both innate and adaptive immune responses. However, limited data are available on the role of receptors of innate immunity, such as toll-like receptors (TLRs) in contributing to antiviral responses and inflammation.Objectives: The aim of this translational study was to characterize TLR responses in immunocompetent patients with primary and symptomatic CMV infection.Study Design: The study population consisted of 40 patients suffering from CMV mononucleosis and 124 blood donors included as controls. We evaluated the association between TLR2, 3, 4, 7 and 9 gene single nucleotide polymorphism (SNP) and susceptibility to symptomatic CMV infection in immunocompetent adults. Additionally, functional TLR-mediated cytokine responses in supernatants of short-term cultures of whole blood from patients with CMV mononucleosis and blood donors were evaluated.Results: TLR2 and TLR7/8 responses were altered in CMV infected patients as compared to healthy donors and were associated with the release of higher levels of the pro-inflammatory cytokines IL-6 and TNF-α, but not of the anti-inflammatory mediator IL-10. The analysis on the TLR SNPs indicated no difference between patients with CMV infection and the control group.Conclusions: No variation in the TLR2,3,4,7 and 9 genes was associated to the development of symptomatic CMV infection in immunocompetent adults. Nevertheless, TLR-mediated responses in CMV-infected patients appeared to be skewed toward a pro-inflammatory profile, which may contribute to the development of inflammatory symptoms during the CMV mononucleotic syndrome.

Leukocyte toll-like receptor expression in pathergy positive and negative Behçet's disease patients.

ObjectivesTo investigate whether the auto-inflammatory nature and the pathergic reaction in Behçet’s disease (BD) are driven by a disturbed toll-like receptor (TLR) response.MethodsWe compared both TLR expression by flow-cytometry and TLR response by stimulation assay in 18 BD patients (both pathergy positive and pathergy negative) with 15 healthy controls.ResultsExpression of TLR1 and 2 was significantly elevated in B-lymphocytes of BD patients compared with healthy controls. TLR1, 2 and 4 were significantly more highly expressed in both CD4+ and CD8+ T-lymphocytes of BD patients. Granulocytes of BD patients displayed significantly higher expression of TLR1, 2, 4 and 6. TLR2, 4 and 5 expression was significantly increased on classical monocytes of BD patients. Intermediate monocytes of BD patients showed an increase in expression of TLR2. Furthermore, TLR2 and 5 were significantly more highly expressed in non-classical monocytes of BD patients. In pathergy positive patients, TLR5 was even more highly expressed compared with pathergy negative patients on B- and T-lymphocytes and granulocytes. Furthermore, TLR2 and 5 showed an elevated TNF-α response to stimulation with their cognate ligands.ConclusionImmune cells of BD patients overexpress TLR1, 2, 4, 5 and 6. Furthermore, after stimulation of TLR2 and 5, BD patients demonstrate a more potent TNF-α response. Although this is a small cohort, in the pathergy positive patients, TLR5 expression is even further augmented, suggesting that a microbial (flagellin) or damage (HMGB1) associated signal may trigger the exaggerated immune response that is characteristic for the pathergy phenomenon in BD. In conclusion, these results point to an exaggerated TLR response in the auto-inflammatory nature of BD.

Chemical modification of pro-inflammatory proteins by peroxynitrite increases activation of TLR4 and NF-κB: Implications for the health effects of air pollution and oxidative stress

Environmental pollutants like fine particulate matter can cause adverse health effects through oxidative stress and inflammation. Reactive oxygen and nitrogen species (ROS/RNS) such as peroxynitrite can chemically modify proteins, but the effects of such modifications on the immune system and human health are not well understood. In the course of inflammatory processes, the Toll-like receptor 4 (TLR4) can sense damage-associated molecular patterns (DAMPs). Here, we investigate how the TLR4 response and pro-inflammatory potential of the proteinous DAMPs α-Synuclein (α-Syn), heat shock protein 60 (HSP60), and high-mobility-group box 1 protein (HMGB1), which are relevant in neurodegenerative and cardiovascular diseases, changes upon chemical modification with peroxynitrite.For the peroxynitrite-modified proteins, we found a strongly enhanced activation of TLR4 and the pro-inflammatory transcription factor NF-κB in stable reporter cell lines as well as increased mRNA expression and secretion of the pro-inflammatory cytokines TNF-α, IL-1β, and IL-8 in human monocytes (THP-1). This enhanced activation of innate immunity via TLR4 is mediated by covalent chemical modifications of the studied DAMPs.Our results show that proteinous DAMPs modified by peroxynitrite more potently amplify inflammation via TLR4 activation than the native DAMPs, and provide first evidence that such modifications can directly enhance innate immune responses via a defined receptor. These findings suggest that environmental pollutants and related ROS/RNS may play a role in promoting acute and chronic inflammatory disorders by structurally modifying the body's own DAMPs. This may have important consequences for chronic neurodegenerative, cardiovascular or gastrointestinal diseases that are prevalent in modern societies, and calls for action, to improve air quality and climate in the Anthropocene.

Role for antimalarials in the management of COVID-19.

The current review highlights recent insights into direct antiviral effects by antimalarials against severe acute respiratory syndrome (SARS)-CoV-2 and other viruses and their potential indirect effects on the host by avoiding exaggerated immune responses (reduced cytokine release, Toll-like receptor response, antigen presentation related to lysosomal processing). Currently, there is a large debate on the use of antimalarials for prophylaxis and treatment of SARS-CoV-2-induced disease based on preclinical in-vitro data, small case series and extrapolation from earlier studies of their effect on intracellular pathogens, including many viruses. Hydroxychloroquine (HCQ) or chloroquine have not demonstrated robust efficacy in prior randomized controlled studies against several other viruses. In-vitro data indicate a reduced viral replication of SARS-CoV-2. Especially immunomodulatory effects of antimalarials might also contribute to a clinical efficacy. For SARS-CoV-2 various large studies will provide answers as to whether antimalarials have a place in prophylaxis or treatment of the acute virus infection with SARS-CoV-2 but compelling data are missing so far. In-vitro data provide a theoretical framework for an efficacy of antimalarials in SARS-CoV-2-induced disease but clinical proof is currently missing.

Diminished toll-like receptor response in febrile infection-related epilepsy syndrome (FIRES)

BackgroundDefective human TLR3 signaling causes recurrent and refractory herpes simplex encephalitis/encephalopathy. Children with febrile infection-related epilepsy syndrome with refractory seizures may have defective TLR responses. MethodsChildren with febrile infection-related epilepsy syndrome were enrolled in this study to evaluate TLR1-9 responses (IL-6, IL-8, IL-12p40, INF-α, INF-γ, and TNF-α) in their peripheral blood mononuclear cells (PBMCs) and monocyte-derived dendritic cells (MDDCs), compared to those with febrile seizures and non-refractory epilepsy with/without underlying encephalitis/encephalopathy. ResultsAdenovirus and enterovirus were found in throat cultures of enrolled patients (2–13 years) as well as serologic IgM elevation of mycoplasma pneumonia and herpes simplex virus, although neither detectable pathogens nor anti-neural autoantibodies in the CSF could be noted. Their PBMCs and MDDCs trended to have impaired TLR responses and significantly lower in cytokine profiles of TLR3, TLR4, TLR7/8, and TLR9 responses but not other TLRs despite normal TLR expressions and normal candidate genes for defective TLR3 signaling. They also had decreased naïve T and T regulatory cells, and weakened phagocytosis. ConclusionChildren with febrile infection-related epilepsy syndrome (FIRES) could have impaired TLR3, TLR4, TLR7/8, and TLR9 responses possibly relating to their weakened phagocytosis and decreased T regulatory cells.

Inflammaging as the basis of age-associated diseases

Aging is one of the most complex biological phenomena that affects all human physiological systems, including the immune system. Immunosenescence is understood as structural and functional changes in both adaptive and innate immunity systems. The so-called inflammaging is among manifestations of immune aging. It is an age-related increase in inflammatory mediators and development of an inflammatory phenotype. An important role in development of inflammaging is assigned to chronic stimulation of immune system by exogenous and endogenous danger signals (pathogen-associated molecular pattern, PAMP and damage-associated molecular pattern, DAMP), which include viruses, microbiota of the gastrointestinal tract, free radicals, etc. PAMP and DAMP are recognized by the innate immunity system cells through the pattern recognition receptors (PRR), e.g., Toll-like receptors (TLR), RIG-I-like receptors (RLR), NODlike receptors (NLR), lectin receptors. Stimulation of PRR leads to activation of intracellular signaling and increased expression of pro-inflammatory factors. PAMPs are the most powerful activators of PRR and inflammation triggers; DAMPs can activate the same receptors and signaling pathways, causing the development of a sterile inflammatory response. The NF-kB signaling pathway is considered as a key signaling pathway for inflammaging. NLR stimulation also leads to formation of inflammasome. Its function is to transform the pro-inflammatory cytokines to a biologically active form, which is an important for the formation of a pro-inflammatory phenotype and development of inflammaging. This process is considered an important risk factor for morbidity and mortality among older people. Chronic inflammation underlies pathogenesis of many age-related diseases, such as osteoporosis, atherosclerosis, Alzheimer’s disease, Parkinson’s disease, type 2 diabetes. Various chronic diseases associated with age are directly related to PAMP and DAMP-induced TLR or NLRP3-mediated inflammatory response. Hence, these ligands and their receptors can be suggested as biomarkers and interventional targets for age-related disorders. Despite numerous studies in age-associated pathology, there are only few works on the contribution of innate immunity in healthy aging. It remains unclear whether the inflammatory phenotype is a manifestation of healthy aging, or it is associated with development of age-related pathology. Further study of the mechanisms of inflammatory aging will reveal biomarkers of healthy aging and potential targets for the treatment of age-associated diseases.

The neovaginal microbiome of transgender women post-gender reassignment surgery

BackgroundGender reassignment surgery is a procedure some transgender women (TW) undergo for gender-affirming purposes. This often includes the construction of a neovagina using existing penile and scrotal tissue and/or a sigmoid colon graft. There are limited data regarding the composition and function of the neovaginal microbiome representing a major gap in knowledge in neovaginal health.ResultsMetaproteomics was performed on secretions collected from the neovaginas (n = 5) and rectums (n = 7) of TW surgically reassigned via penile inversion/scrotal graft with (n = 1) or without (n = 4) a sigmoid colon graft extension and compared with secretions from cis vaginas (n = 32). We identified 541 unique bacterial proteins from 38 taxa. The most abundant taxa in the neovaginas were Porphyromonas (30.2%), Peptostreptococcus (9.2%), Prevotella (9.0%), Mobiluncus (8.0%), and Jonquetella (7.2%), while cis vaginas were primarily Lactobacillus and Gardnerella. Rectal samples were mainly composed of Prevotella and Roseburia. Neovaginas (median Shannon’s H index = 1.33) had higher alpha diversity compared to cis vaginas (Shannon’s H = 0.35) (p = 7.2E−3, Mann-Whitney U test) and were more similar to the non-Lactobacillus dominant/polymicrobial cis vaginas based on beta diversity (perMANOVA, p = 0.001, r2 = 0.342). In comparison to cis vaginas, toll-like receptor response, amino acid, and short-chain fatty acid metabolic pathways were increased (p < 0.01), while keratinization and cornification proteins were decreased (p < 0.001) in the neovaginal proteome.ConclusionsPenile skin-lined neovaginas have diverse, polymicrobial communities that show similarities in composition to uncircumcised penises and host responses to cis vaginas with bacterial vaginosis (BV) including increased immune activation pathways and decreased epithelial barrier function. Developing a better understanding of microbiome-associated inflammation in the neovaginal environment will be important for improving our knowledge of neovaginal health.A1pw-5DXJFMq35mSKHH2fxVideo