Inflammasomes and Atherosclerosis: a Mixed Picture.

The CANTOS (Canakinumab Anti-inflammatory Thrombosis Outcome Study) and colchicine trials suggest an important role of inflammasomes and their major product IL-1ß (interleukin 1ß) in human atherosclerotic cardiovascular disease. Moreover, studies in mouse models indicate a causal role of inflammasomes and IL-1ß in atherosclerosis. However, recent studies have led to a more granular view of the role of inflammasomes in atherosclerosis. Studies in hyperlipidemic mouse models suggest that prominent activation of the NLRP3 inflammasome requires a second hit such as defective cholesterol efflux, defective DNA repair, clonal hematopoiesis or diabetes. Similarly in humans some mutations promoting clonal hematopoiesis increase coronary artery disease risk in part by promoting inflammasome activation. Recent studies in mice and humans point to a wider role of the AIM2 (absent in melanoma 2) inflammasome in promoting cardiovascular disease including in some forms of clonal hematopoiesis and diabetes. These developments suggest a precision medicine approach in which treatments targeting inflammasomes or IL-1ß might be best employed in clinical settings involving increased inflammasome activation.

Urine biomarkers individually and as a consensus model show high sensitivity and specificity for detecting UTIs.

BACKGROUND: Current diagnoses of urinary tract infection (UTI) by standard urine culture (SUC) has significant limitations in sensitivity, especially for fastidious organisms, and the ability to identify organisms in polymicrobial infections. The significant rate of both SUC "negative" or "mixed flora/contamination" results in UTI cases and the high prevalence of asymptomatic bacteriuria indicate the need for an accurate diagnostic test to help identify true UTI cases. This study aimed to determine if infection-associated urinary biomarkers can differentiate definitive UTI cases from non-UTI controls. METHODS: Midstream clean-catch voided urine samples were collected from asymptomatic volunteers and symptomatic subjects ≥ 60 years old diagnosed with a UTI in a urology specialty setting. Microbial identification and density were assessed using a multiplex PCR/pooled antibiotic susceptibility test (M-PCR/P-AST) and SUC. Three biomarkers [neutrophil gelatinase-associated lipocalin (NGAL), and Interleukins 8 and 1ß (IL-8, and IL-1ß)] were also measured via enzyme-linked immunosorbent assay (ELISA). Definitive UTI cases were defined as symptomatic subjects with a UTI diagnosis and positive microorganism detection by SUC and M-PCR, while definitive non-UTI cases were defined as asymptomatic volunteers. RESULTS: We observed a strong positive correlation (R2 > 0.90; p < 0.0001) between microbial density and the biomarkers NGAL, IL-8, and IL-1ß for symptomatic subjects. Biomarker consensus criteria of two or more positive biomarkers had sensitivity 84.0%, specificity 91.2%, positive predictive value 93.7%, negative predictive value 78.8%, accuracy 86.9%, positive likelihood ratio of 9.58, and negative likelihood ratio of 0.17 in differentiating definitive UTI from non-UTI cases, regardless of non-zero microbial density. NGAL, IL-8, and IL-1ß showed a significant elevation in symptomatic cases with positive microbe identification compared to asymptomatic cases with or without microbe identification. Biomarker consensus exhibited high accuracy in distinguishing UTI from non-UTI cases. CONCLUSION: We demonstrated that positive infection-associated urinary biomarkers NGAL, IL-8, and IL-1ß, in symptomatic subjects with positive SUC and/or M-PCR results was associated with definitive UTI cases. A consensus criterion with ≥ 2 of the biomarkers meeting the positivity thresholds showed a good balance of sensitivity (84.0%), specificity (91.2%), and accuracy (86.9%). Therefore, this biomarker consensus is an excellent supportive diagnostic tool for resolving the presence of active UTI, particularly if SUC and M-PCR results disagree.

Amino Sugar-Enriched Fraction of Korean Red Ginseng Extract Induces the Priming Step of NLRP3 Inflammasome.

Intracellular protein complexes, known as inflammasomes, activate caspase-1 and induce the secretion of pro-inflammatory cytokines, namely interleukin (IL)-1ß and -18. Korean Red Ginseng extract (RGE) is a known immunomodulator and a potential candidate for the regulation of inflammasomes. The saponins, such as ginsenosides, of RGE inhibit inflammasome signaling, while non-saponin substances containing amino sugars promote the priming step, up-regulating inflammasome components (pro-IL-1ß, NLRP3, caspase-1, and Asc). In this study, the amino sugar-enriched fraction (ASEF), which increases only non-saponin components, including amino sugars, without changing the concentration of saponin substances, was used to investigate whether saponin or non-saponin components of RGE would have a greater impact on the priming step. When murine macrophages were treated with ASEF, the gene expression of inflammatory cytokines (IL-1α, TNFα, IL-6, and IL-10) increased. Additionally, ASEF induced the priming step but did not affect the inflammasome activation step, such as the secretion of IL-1ß, cleavage of caspase-1, and formation of Asc pyroptosome. Furthermore, the upregulation of gene expression of inflammasome components by ASEF was blocked by inhibitors of Toll-like receptor 4 signaling. Maltol, the main constituent of ASEF, promoted the priming step but inhibited the activation step of the inflammasome, while arginine, sugars, arginine-fructose-glucose, and fructose-arginine, the other main constituents of ASEF, had no effect on either step. Thus, certain amino sugars in RGE, excluding maltol, are believed to be the components that induce the priming step. The priming step that prepares the NLRP3 inflammasome for activation appears to be induced by amino sugars in RGE, thereby contributing to the immune-boosting effects of RGE.

Signaling metabolite L-2-hydroxyglutarate activates the transcription factor HIF-1α in lipopolysaccharide-activated macrophages.

Activated macrophages undergo metabolic reprogramming, which not only supports their energetic demands but also allows for the production of specific metabolites that function as signaling molecules. Several Krebs cycles, or Krebs-cycle-derived metabolites, including succinate, α-ketoglutarate, and itaconate, have recently been shown to modulate macrophage function. The accumulation of 2-hydroxyglutarate (2HG) has also been well documented in transformed cells and more recently shown to play a role in T cell and dendritic cell function. Here we have found that the abundance of both enantiomers of 2HG is increased in LPS-activated macrophages. We show that L-2HG, but not D-2HG, can promote the expression of the proinflammatory cytokine IL-1ß and the adoption of an inflammatory, highly glycolytic metabolic state. These changes are likely mediated through activation of the transcription factor hypoxia-inducible factor-1α (HIF-1α) by L-2HG, a known inhibitor of the HIF prolyl hydroxylases. Expression of the enzyme responsible for L-2HG degradation, L-2HG dehydrogenase (L-2HGDH), was also found to be decreased in LPS-stimulated macrophages and may therefore also contribute to L-2HG accumulation. Finally, overexpression of L-2HGDH in HEK293 TLR4/MD2/CD14 cells inhibited HIF-1α activation by LPS, while knockdown of L-2HGDH in macrophages boosted the induction of HIF-1α-dependent genes, as well as increasing LPS-induced HIF-1α activity. Taken together, this study therefore identifies L-2HG as a metabolite that can regulate HIF-1α in macrophages.

The H240R Protein of African Swine Fever Virus Inhibits Interleukin 1ß Production by Inhibiting NEMO Expression and NLRP3 Oligomerization.

The H240R protein (pH240R), encoded by the H240R gene of African swine fever virus (ASFV), is a 241-amino-acid capsid protein. We previously showed that the deletion of H240R from the ASFV genome, creating ASFV-ΔH240R, resulted in an approximately 2-log decrease in infectious virus production compared with the wild-type ASFV strain (ASFV-WT), and ASFV-ΔH240R induced higher interleukin 1ß (IL-1ß) production in porcine alveolar macrophages (PAMs) than did ASFV-WT, but the underlying mechanism remains to be elucidated. Here, we demonstrate that the activation of the NF-κB signaling and NLRP3 inflammasome was markedly induced in PAMs upon ASFV-ΔH240R infection compared with ASFV-WT. Moreover, pH240R inhibited NF-κB activation by interacting with NEMO and promoting the autophagy-mediated lysosomal degradation of NEMO, resulting in reduced pro-IL-1ß transcription. Strikingly, NLRP3 deficiency in PAMs inhibited the ASFV-ΔH240R-induced IL-1ß secretion and caspase 1 activation, indicating an essential role of NLRP3 inflammasome activation during ASFV-ΔH240R replication. Mechanistically, pH240R interacted with NLRP3 to inhibit its oligomerization, leading to decreased IL-1ß production. Furthermore, the inhibition of the NF-κB signaling and NLRP3 inflammasome activation promoted ASFV-ΔH240R replication in PAMs. Taken together, the results of this study reveal an antagonistic mechanism by which pH240R suppresses the host immune response by manipulating activation of the NF-κB signaling and NLRP3 inflammasome, which might guide the rational design of live attenuated vaccines or therapeutic strategies against ASF in the future. IMPORTANCE African swine fever (ASF), a lethal hemorrhagic disease, is caused by African swine fever virus (ASFV). There are no commercially available vaccines or antivirals for the disease. Here, we showed that ASFV with a deletion of the H240R gene exhibits high-level expression of interleukin 1ß (IL-1ß), a proinflammatory cytokine, in porcine alveolar macrophages and that the H240R protein (pH240R) exhibits robust inhibitory effects on IL-1ß transcription and production. More specifically, pH240R inhibited NF-κB activation via the autophagy-mediated lysosomal degradation of NEMO, leading to the decrease of pro-IL-1ß transcription. In addition, pH240R interacted with NLRP3 to inhibit its oligomerization, leading to decreased IL-1ß production. Our results indicate that pH240R is involved in the evasion of host innate immunity and provide a novel target for the development of a live attenuated vaccine against ASF.

Apelin is found in human sperm and testis and is raised in inflammatory pathological conditions.

Apelin/APJ receptor (R) is involved in many oxidative stress-induced pathological conditions. Since this system is not yet explored in male reproduction, we studied apelin/APJ-R in human semen and testis. Semen of 41 infertile patients with varicocele, genitourinary infections, unexplained infertility and 12 fertile men was analysed (WHO guidelines, 2021). Apelin was quantified by ELISA in seminal fluid and spermatozoa, interleukin (IL)-1ß in seminal fluid. Apelin/APJ-R were immunolocalized in spermatozoa and testis. Apelin was present in spermatozoa and its levels were negatively correlated with normal sperm morphology% (r = -0.857; p < 0.001), and positively with IL-1ß levels (r = 0.455; p < 0.001). Apelin and IL-1ß concentrations were increased in patients' samples with varicocele (apelin p < 0.01; IL-1ß p < 0.05) and infections (apelin p < 0.01; IL-1ß p < 0.001). By logistic regression analysis, apelin (OR 1.310; p = 0.011) and IL-1ß (OR 1.572; p = 0.005) were predictors of inflammatory diseases (varicocele, infections). Apelin and APJ-R immunofluorescence labels were weak in sperm tail of fertile men and intense along tail, cytoplasmic residues and post-acrosomal sheath of sperm from infertile men. In testis, apelin and APJ-R labels were evident in Leydig cells and weak inside the seminiferous tubule. Apelin/APJ-R system is present in human spermatozoa and testicular tissue and probably involved in human fertility.

YY1 (Yin-Yang 1), a transcription factor regulating systemic inflammation, is involved in cognitive impairment of depression.

AIM: Clinical and preclinical studies suggest that alterations in the peripheral and brain immune system are associated with the pathophysiology of depression, also leading to changes in local glucose metabolism in the brain. Here, the authors identified Yin-Yang 1 (YY1), a transcription factor closely associated with central and peripheral inflammation. METHODS: Plasma levels of YY1, interleukin (IL) 6, and IL-1ß in major depressive disorder (MDD) were collected before and after treatment with vortioxetine, and correlation with clinical and cognitive scores was studied. Chronic unpredictable mild stress was treated with vortioxetine. Micropositron emission tomography (microPET) was used to analyze glucose metabolism and mRNA, and the protein level of the YY1-nuclear factor κB (NF-κB)-IL-1ß inflammatory pathway were measured in related brain regions. RESULTS: Plasma levels of YY1 and IL-1ß were significantly increased in MDD and decreased after treatment with vortioxetine. Meanwhile, the level of YY1 in plasma was negatively correlated with cognitive functions in patients with MDD and positively correlated with the level of IL-1ß in plasma. Compared with the control group, in chronic unpredictable mild stress rats, (microPET) analysis showed that the decrease of glucose metabolism in the hippocampus, entorhinal cortex, amygdala, striatum, and medial prefrontal cortex was reversed after treatment. mRNA and protein level of related molecular in YY1-NF-κB-IL-1ß inflammatory pathway decreased in the hippocampus and was reversed by vortioxetine. CONCLUSION: The current study suggests that the YY1-NF-κB-IL-1ß inflammatory pathway may play an essential role in both mood changes and cognitive impairment in depression, and may be associated with changes in glucose metabolism in emotion regulation and cognition. These findings provide new evidence for the inflammatory mechanisms of depression.

Photobiomodulation, alone or combined with adipose-derived stem cells, reduces inflammation by modulation of microRNA-146a and interleukin-1ß in a delayed-healing infected wound in diabetic rats.

Diabetic wounds are categorized by chronic inflammation, leading to the development of diabetic foot ulcers, which cause amputation and death. Herewith, we examined the effect of photobiomodulation (PBM) plus allogeneic diabetic adipose tissue-derived stem cells (ad-ADS) on stereological parameters and expression levels of interleukin (IL)-1ß and microRNA (miRNA)-146a in the inflammatory (day 4) and proliferation (day 8) stages of wound healing in an ischemic infected (with 2×107 colony-forming units of methicillin-resistant Staphylococcus aureus) delayed healing wound model (IIDHWM) in type I diabetic (TIDM) rats. There were five groups of rats: group 1 control (C); group 2 (CELL) in which rat wounds received 1×106 ad-ADS; group 3 (CL) in which rat wounds received the ad-ADS and were subsequently exposed to PBM(890 nm, 80 Hz, 3.5 J/cm2, in vivo); group 4 (CP) in which the ad-ADS preconditioned by the PBM(630 nm + 810 nm, 0.05 W, 1.2 J/cm2, 3 times) were implanted into rat wounds; group 5 (CLP) in which the PBM preconditioned ad-ADS were implanted into rat wounds, which were then exposed to PBM. On both days, significantly better histological results were seen in all experimental groups except control. Significantly better histological results were observed in the ad-ADS plus PBM treatment correlated to the ad-ADS alone group (p<0.05). Overall, PBM preconditioned ad-ADS followed by PBM of the wound showed the most significant improvement in histological measures correlated to the other experimental groups (p<0.05). On days 4 and 8, IL-1 ß levels of all experimental groups were lower than the control group; however, on day 8, only the CLP group was different (p<0.01). On day 4, miR-146a expression levels were substantially greater in the CLP and CELL groups correlated to the other groups, on day 8 miR-146a in all treatment groups was upper than C (p<0.01). ad-ADS plus PBM, ad-ADS, and PBM all improved the inflammatory phase of wound healing in an IIDHWM in TIDM1 rats by reducing inflammatory cells (neutrophils, macrophages) and IL-1ß, and increasing miRNA-146a. The ad-ADS+PBM combination was better than either ad-ADS or PBM alone, because of the higher proliferative and anti-inflammatory effects of the PBM+ad-ADS regimen.

Evaluating the relationship between ankylosing spondylitis and periodontal disease: a case-control study.

OBJECTIVES: This study aimed to determine the possible relationship between periodontal disease and ankylosing spondylitis (AS) by evaluating clinical periodontal measurements and gingival crevicular fluid (GCF) levels of sclerostin, interleukin-1ß (IL-1ß), and matrix metalloproteinase-8 (MMP-8) levels. MATERIALS AND METHODS: Twenty-eight patients with AS (AS group) and 28 systemically healthy controls (C group) were enrolled in this study. Full-mouth periodontal measurements: plaque index, bleeding on probing (BOP), probing pocket depth (PPD), and clinical attachment level (CAL) measurements were obtained from all patients. AS-related parameters were included in the data analyses. An enzyme-linked immunosorbent assay determined GCF IL-1ß, MMP-8, and sclerostin levels. RESULTS: There were no significant differences in the clinical periodontal measurements between the two groups (p > 0.05). Interestingly, patients with AS had significantly lower GCF sclerostin levels than the C group (p < 0.05). But there were no statistical differences in the GCF levels of IL-1ß and MMP-8 between the two groups (p > 0.05). Serum C-reactive protein (CRP) levels strongly correlated with both BOP (r = 0.497, p < 0.05) and PPD (r = 0.570, p < 0.05) in the AS group. Bath AS Metrology Index (BASMI) also positively correlated with both BOP (r = 0.530, p < 0.05) and CAL (r = 0.568, p < 0.05). Similarly, Maastricht Ankylosing Spondylitis Enthesis Score (MASES) strongly correlated with both BOP (r = 0.487, p < 0.05) and CAL (r = 0.522, p < 0.05). CONCLUSION: These results suggest that the patient's systemic condition may influence local sclerostin levels in GCF, and the strong correlations between periodontal measurements and AS-related parameters may indicate an interrelationship between inflammatory periodontal disease and AS. CLINICAL RELEVANCE: The present study provides important information concerning the relationship between periodontal disease and ankylosing spondylitis. TRIAL REGISTRATION: Thai Clinical Trials.gov (TCTR20200908001) (08. September 2020).

Gut Region-Specific Interleukin 1ß Induction in Different Myenteric Neuronal Subpopulations of Type 1 Diabetic Rats.

Interleukin 1ß (IL1ß) is a pro-inflammatory cytokine that may play a crucial role in enteric neuroinflammation in type 1 diabetes. Therefore, our goal is to evaluate the effects of chronic hyperglycemia and insulin treatment on IL1ß immunoreactivity in myenteric neurons and their different subpopulations along the duodenum-ileum-colon axis. Fluorescent immunohistochemistry was used to count IL1ß expressing neurons as well as the neuronal nitric oxide synthase (nNOS)- and calcitonin gene-related peptide (CGRP)-immunoreactive myenteric neurons within this group. Tissue IL1ß level was measured by ELISA in muscle/myenteric plexus-containing homogenates. IL1ß mRNA was detected by RNAscope in different intestinal layers. The proportion of IL1ß-immunoreactive myenteric neurons was significantly higher in the colon than in the small intestine of controls. In diabetics, this proportion significantly increased in all gut segments, which was prevented by insulin treatment. The proportion of IL1ß-nNOS-immunoreactive neurons only increased in the diabetic colon, while the proportion of IL1ß-CGRP-immunoreactive neurons only increased in the diabetic ileum. Elevated IL1ß levels were also confirmed in tissue homogenates. IL1ß mRNA induction was detected in the myenteric ganglia, smooth muscle and intestinal mucosa of diabetics. These findings support that diabetes-related IL1ß induction is specific for the different myenteric neuronal subpopulations, which may contribute to diabetic motility disturbances.

Accumulation of Inflammatory Mediators in the Normal Pericardial Fluid.

There is paucity of studies that focus on the composition of pericardial fluid under resting conditions. The purpose of this study is to determine the levels of inflammatory mediators in pericardial fluid and their correlation with plasma levels in patients undergoing elective cardiac surgery. We conducted a prospective cohort study on candidates for elective aortic valve replacement surgery. Pericardial fluid and peripheral venous blood samples were collected after opening the pericardium. Levels of interleukin 1α (IL-1α); interleukin 1ß (IL-1ß); interleukin 2 (IL-2) interleukin 4 (IL-4); interleukin 6 (IL-6); interleukin 8 (IL8); interleukin 10 (IL10); tumor necrosis factor-α (TNF-α), interferon-γ (IFN-γ), vascular endothelial growth factor (VEGF), monocyte chemotactic protein-1 (MCP-1) epidermal growth factor (EGF), soluble E-selectin, L-selectin, P-selectin, intercellular adhesion molecule-1 (ICAM-1), and vascular cell adhesion molecule-1 (VCAM-1) were determined in both pericardial fluid and serum samples. A total of 45 patients with a mean age of 74 years were included of which 66% were males. Serum levels of all study mediators were within normal limits. Serum and pericardial levels of IL-1 α, IL-1 ß, IL-2, IL-4, and IL-10 were similar. Levels of VEGF, EGF, VCAM-2, ICAM 1, E-selectin, P-selectin, and L-selectin were significantly lower in pericardial fluid than in serum. However, levels of IL-6, IL-8, TNF-α, IFN-γ, MCP-1, and MCP-1 were significantly higher in the pericardial fluid than in serum. Under normal conditions, the pattern of distribution of different inflammatory mediators in the pericardial fluid does not reflect serum levels. This may either reflect the condition of the underlying myocardium and epicardial fat or the activity of the mesothelial and mononuclear cells present in pericardial fluid.

Recombinant Human Prolidase (rhPEPD) Induces Wound Healing in Experimental Model of Inflammation through Activation of EGFR Signalling in Fibroblasts.

The potential of recombinant human prolidase (rhPEPD) to induce wound healing in an experimental model of IL-1ß-induced inflammation in human fibroblasts was studied. It was found that rhPEPD significantly increased cell proliferation and viability, as well as the expression of the epidermal growth factor receptor (EGFR) and downstream signaling proteins, such as phosphorylated PI3K, AKT, and mTOR, in the studied model. Moreover, rhPEPD upregulated the expression of the ß1 integrin receptor and its downstream signaling proteins, such as p-FAK, Grb2 and p-ERK 1/2. The inhibition of EGFR signaling by gefitinib abolished rhPEPD-dependent functions in an experimental model of inflammation. Subsequent studies showed that rhPEPD augmented collagen biosynthesis in IL-1ß-treated fibroblasts as well as in a wound healing model (wound closure/scratch test). Although IL-1ß treatment of fibroblasts increased cell migration, rhPEPD significantly enhanced this process. This effect was accompanied by an increase in the activity of MMP-2 and MMP-9, suggesting extracellular matrix (ECM) remodeling during the inflammatory process. The data suggest that rhPEPD may play an important role in EGFR-dependent cell growth in an experimental model of inflammation in human fibroblasts, and this knowledge may be useful for further approaches to the treatment of abnormalities of wound healing and other skin diseases.

Biological determinants of blood-based cytokines in the Alzheimer's disease clinical continuum.

Converging translational and clinical research strongly indicates that altered immune and inflammatory homeostasis (neuroinflammation) plays a critical pathophysiological role in Alzheimer's disease (AD), across the clinical continuum. A dualistic role of neuroinflammation may account for a complex biological phenomenon, representing a potential pharmacological target. Emerging blood-based pathophysiological biomarkers, such as cytokines (Cyt) and interleukins (ILs), have been studied as indicators of neuroinflammation in AD. However, inconsistent results have been reported probably due to a lack of standardization of assays with methodological and analytical differences. We used machine-learning and a cross-validation-based statical workflow to explore and analyze the potential impact of key biological factors, such as age, sex, and apolipoprotein-E (APOE) genotype (the major genetic risk factor for late-onset AD) on Cyt. A set of Cyt was selected based on previous literature, and we investigated any potential association in a pooled cohort of cognitively healthy, mild cognitive impairment (MCI), and AD-like dementia patients. We also performed explorative analyses to extrapolate preliminary clinical insights. We found a robust sex effect on IL12 and an APOE-related difference in IL10, with the latter being also related to the presence of advanced cognitive decline. IL1ß was the variable most significantly associated with MCI-to-dementia conversion over a 2.5 year-clinical follow-up. Although preliminary, our data support further clinical research to understand whether plasma Cyt may represent reliable and noninvasive tools serving the investigation of neuroimmune and inflammatory dynamics in AD and to foster biomarker-guided pathway-based therapeutic approaches, within the precision medicine development framework.

FGF2 and IL-1ß - explorers of unconventional secretory pathways at a glance.

Fibroblast growth factor 2 (FGF2) and interleukin 1ß (IL-1ß) were among the earliest examples of a subclass of proteins with extracellular functions that were found to lack N-terminal secretory signal peptides and were shown to be secreted in an ER- and Golgi-independent manner. Many years later, a number of alternative secretory pathways have been discovered, processes collectively termed unconventional protein secretion (UPS). In the course of these studies, unconventional secretion of FGF2 and IL-1ß were found to be based upon distinct pathways, mechanisms and molecular machineries. Following a concise introduction into various pathways mediating unconventional secretion and transcellular spreading of proteins, this Cell Science at a Glance poster article aims at a focused analysis of recent key discoveries providing unprecedented detail about the molecular mechanisms and machineries driving FGF2 and IL-1ß secretion. These findings are also highly relevant for other unconventionally secreted cargoes that, like FGF2 and IL1ß, exert fundamental biological functions in biomedically relevant processes, such as tumor-induced angiogenesis and inflammation.

Recruitment of α4ß7 monocytes and neutrophils to the brain in experimental colitis is associated with elevated cytokines and anxiety-like behavior.

BACKGROUND: Behavioral comorbidities, such as anxiety and depression, are a prominent feature of IBD. The signals from the inflamed gut that cause changes in the brain leading to these behavioral comorbidities remain to be fully elucidated. We tested the hypothesis that enhanced leukocyte-cerebral endothelial cell interactions occur in the brain in experimental colitis, mediated by α4ß7 integrin, to initiate neuroimmune activation and anxiety-like behavior. METHODS: Female mice treated with dextran sodium sulfate were studied at the peak of acute colitis. Circulating leukocyte populations were determined using flow cytometry. Leukocyte-cerebral endothelial cell interactions were examined using intravital microscopy in mice treated with anti-integrin antibodies. Brain cytokine and chemokines were assessed using a multiplex assay in animals treated with anti-α4ß7 integrin. Anxiety-like behavior was assessed using an elevated plus maze in animals after treatment with an intracerebroventricular injection of interleukin 1 receptor antagonist. RESULTS: The proportion of classical monocytes expressing α4ß7 integrin was increased in peripheral blood of mice with colitis. An increase in the number of rolling and adherent leukocytes on cerebral endothelial cells was observed, the majority of which were neutrophils. Treatment with anti-α4ß7 integrin significantly reduced the number of rolling leukocytes. After anti-Ly6C treatment to deplete monocytes, the number of rolling and adhering neutrophils was significantly reduced in mice with colitis. Interleukin-1ß and CCL2 levels were elevated in the brain and treatment with anti-α4ß7 significantly reduced them. Enhanced anxiety-like behavior in mice with colitis was reversed by treatment with interleukin 1 receptor antagonist. CONCLUSIONS: In experimental colitis, α4ß7 integrin-expressing monocytes direct the recruitment of neutrophils to the cerebral vasculature, leading to elevated cytokine levels. Increased interleukin-1ß mediates anxiety-like behavior.

The zebrafish NLRP3 inflammasome has functional roles in ASC-dependent interleukin-1ß maturation and gasdermin E-mediated pyroptosis.

The NLR family pyrin domain containing 3 (NLRP3) inflammasome is one of the best-characterized inflammasomes in humans and other mammals. However, knowledge about the NLRP3 inflammasome in nonmammalian species remains limited. Here, we report the molecular and functional identification of an NLRP3 homolog (DrNLRP3) in a zebrafish (Danio rerio) model. We found that DrNLRP3's overall structural architecture was shared with mammalian NLRP3s. It initiates a classical inflammasome assembly for zebrafish inflammatory caspase (DrCaspase-A/-B) activation and interleukin 1ß (DrIL-1ß) maturation in an apoptosis-associated speck-like protein containing a caspase-recruitment domain (ASC)-dependent manner, in which DrNLRP3 organizes DrASC into a filament that recruits DrCaspase-A/-B by homotypic pyrin domain (PYD)-PYD interactions. DrCaspase-A/-B activation in the DrNLRP3 inflammasome occurred in two steps, with DrCaspase-A being activated first and DrCaspase-B second. DrNLRP3 also directly activated full-length DrCaspase-B and elicited cell pyroptosis in a gasdermin E (GSDME)-dependent but ASC-independent manner. These two events were tightly coordinated by DrNLRP3 to ensure efficient IL-1ß secretion for the initiation of host innate immunity. By knocking down DrNLRP3 in zebrafish embryos and generating a DrASC-knockout (DrASC-/-) fish clone, we characterized the function of the DrNLRP3 inflammasome in anti-bacterial immunity in vivo The results of our study disclosed the origin of the NLRP3 inflammasome in teleost fish, providing a cross-species understanding of the evolutionary history of inflammasomes. Our findings also indicate that the NLRP3 inflammasome may coordinate inflammatory cytokine processing and secretion through a GSDME-mediated pyroptotic pathway, uncovering a previously unrecognized regulatory function of NLRP3 in both inflammation and cell pyroptosis.

Multiple sclerosis in a patient with cryopyrin-associated autoinflammatory syndrome: Evidence that autoinflammation is the common link.

The co-existence of an autoinflammatory syndrome with a demyelinating disorder is a very rare occurrence raising the question whether there is a pathophysiological connection between them. We describe the case of a man with symptoms of cryopyrin-associated periodic syndrome (CAPS) since infancy who later developed multiple sclerosis (MS). As CAPS was genetically confirmed, the inhibition of interleukin-1 (IL-1) with anakinra led to a swift resolution of the CAPS symptoms and also, in combination with teriflunomide, to a clinical and imaging improvement of MS. In vitro studies showed that, upon a CAPS flare, the patient's peripheral neutrophils released neutrophil extracellular traps (NETs) decorated with IL-1ß, while NET release was markedly decreased following anakinra-induced remission of CAPS. Taking into account the growing evidence on the involvement of IL-1ß in experimental models of MS, this rare patient case suggests that the role of neutrophils/NETs and IL-1ß in MS should be further studied.

Smek1 deficiency exacerbates experimental autoimmune encephalomyelitis by activating proinflammatory microglia and suppressing the IDO1-AhR pathway.

BACKGROUND: Experimental autoimmune encephalomyelitis (EAE) is an animal disease model of multiple sclerosis (MS) that involves the immune system and central nervous system (CNS). However, it is unclear how genetic predispositions promote neuroinflammation in MS and EAE. Here, we investigated how partial loss-of-function of suppressor of MEK1 (SMEK1), a regulatory subunit of protein phosphatase 4, facilitates the onset of MS and EAE. METHODS: C57BL/6 mice were immunized with myelin oligodendrocyte glycoprotein 35-55 (MOG35-55) to establish the EAE model. Clinical signs were recorded and pathogenesis was investigated after immunization. CNS tissues were analyzed by immunostaining, quantitative polymerase chain reaction (qPCR), western blot analysis, and enzyme-linked immunosorbent assay (ELISA). Single-cell analysis was carried out in the cortices and hippocampus. Splenic and lymph node cells were evaluated with flow cytometry, qPCR, and western blot analysis. RESULTS: Here, we showed that partial Smek1 deficiency caused more severe symptoms in the EAE model than in controls by activating myeloid cells and that Smek1 was required for maintaining immunosuppressive function by modulating the indoleamine 2,3-dioxygenase (IDO1)-aryl hydrocarbon receptor (AhR) pathway. Single-cell sequencing and an in vitro study showed that Smek1-deficient microglia and macrophages were preactivated at steady state. After MOG35-55 immunization, microglia and macrophages underwent hyperactivation and produced increased IL-1ß in Smek1-/+ mice at the peak stage. Moreover, dysfunction of the IDO1-AhR pathway resulted from the reduction of interferon γ (IFN-γ), enhanced antigen presentation ability, and inhibition of anti-inflammatory processes in Smek1-/+ EAE mice. CONCLUSIONS: The present study suggests a protective role of Smek1 in autoimmune demyelination pathogenesis via immune suppression and inflammation regulation in both the immune system and the central nervous system. Our findings provide an instructive basis for the roles of Smek1 in EAE and broaden the understanding of the genetic factors involved in the pathogenesis of autoimmune demyelination.

Andrographolide inhibits IL-1ß release in bone marrow-derived macrophages and monocyte infiltration in mouse knee joints induced by monosodium urate.

Andrographolide (AND) is the major diterpenoid in A. paniculata with wide clinical application and has been shown to be a potent anti-inflammatory agent. Gout is the leading inflammatory disease of the joints, and the deposition of urate in the articular cavity attracts immune cells that release inflammatory cytokines. Monosodium urate (MSU) is known to be one of the activators of the NLRP3 (NLR family pyrin domain containing 3) inflammasome. After activation, the NLRP3 inflammasome releases interleukin-1ß (IL-1ß), which causes the development of many inflammatory diseases. The aim of the present study was to investigate whether AND attenuates the release of IL-1ß mediated by the NLRP3 inflammasome. The effects of AND were studied in bone marrow-derived macrophages (BMDMs) treated with lipopolysaccharide (LPS) and MSU and in mice with MSU-induced joint inflammation. AND suppressed MSU phagocytosis dose-dependently and markedly inhibited LPS- and MSU-induced IL-1ß release in BMDMs. Moreover, AND pretreatment inhibited the LPS-induced NLRP3 inflammasome priming stage by inhibiting the IKK/NFκB signaling pathway, which resulted in decreased protein expression of NLRP3 and proIL-1ß. AND induced HO-1 protein expression in a dose-dependent manner and attenuated MSU-induced ROS generation. Silencing HO-1 mitigated AND inhibition of LPS/MSU-induced IL-1ß release in J774A.1 cells. In addition, AND decreased MSU-mediated ASC binding to NLRP3. Oral administration of AND attenuated MSU-induced monocyte infiltration in mouse knee joints. These results suggest that the working mechanisms by which AND down-regulates MSU-induced joint inflammation might be via HO-1 induction and attenuation of ROS-mediated NLRP3 inflammasome assembly and subsequent IL-1ß release.

The association between serum interleukin-1 beta and heparin sulphate in diabetic nephropathy patients.

Inflammation is considered an important mechanism in the development of diabetes mellitus (DM) and persists for a long time before the occurrence of diabetic nephropathy (DN). Many studies have demonstrated that a decrease in the endothelial glycocalyx (EG) is negatively correlated with proteinuria. To elucidate whether EG damage induced by inflammasomes in DM patients leads to the occurrence of microalbuminuria (MA) and accelerates the progression of DN, this study screened 300 diagnosed DM patients. Finally, 70 type 2 diabetes patients were invited to participate in this study and were divided into two groups: the T2DM group (patients with normal MA and without diabetic retinopathy, n = 35) and the T2DN group (patients with increased MA and diabetic retinopathy, n = 35). Circulating heparin sulphate (HS, EG biomarkers) and interleukin-1 beta (IL-1ß, inflammasome biomarkers) of the patients were measured by ELISA. Laboratory data were measured using routine laboratory methods. Patients in the T2DN group had increased serum HS, increased IL-1ß, increased CRP, decreased haemoglobin, and increased neutrophils compared to patients in the T2DM group (all P < 0.05). Increased HS and decreased haemoglobin were independently associated with T2DN patients. ROC curves showed that the AUC of HS for the prediction of T2DN was 0.67 (P < 0.05). The combination of HS and haemoglobin yielded a significant increasement in the AUC (0.75, P < 0.001) with optimal sensitivity (71.2%) and specificity (79%). Furthermore, serum IL-1ß was positively correlated with HS and was an independent associated factor of HS in the T2DN group. The relationship between HS and IL-1ß was not significant in the T2DM group. Our findings surgessed the inflammasome may be associated with and promote damage to the EG during the disease course of DN that manifests as increased MA.