Toll-like Receptor Research Articles

The Nephroprotective Effect of Punica granatum Peel Extract on LPS-Induced Acute Kidney Injury

Sepsis is an exaggerated immune response resulting from systemic inflammation, which can damage tissues and organs. Acute kidney injury has been detected in at least one-third of patients with sepsis. Sepsis-associated acute kidney injury increases the risk of a secondary infection. Rapid diagnosis and appropriate initiation of antibiotics can significantly reduce mortality and morbidity. However, microorganisms are known to develop resistance to antibiotics. Estimations indicate that the annual casualties caused by microbial resistance will surpass cancer fatalities by 2050. The prevalence of bacterial infections and their growing antibiotic resistance has brought immediate attention to the search for novel treatments. Plant-derived supplements contain numerous bioactive components with therapeutic potential against a variety of conditions, including infections. Punica granatum peel is rich in phenolic compounds. The purpose of this study was to determine the anti-inflammatory and anti-bacterial properties of P. granatum peel extract (PGPE) on lipopolysaccharide (LPS)-induced acute kidney injury. Experimental groups were Control, LPS (10 mg/kg LPS, intraperitoneally), PGPE100, and PGPE300 (100 and 300 mg/mL PGPE via oral gavage, respectively, for 7 days). According to biochemical results, serum blood urea nitrogen (BUN), creatinine (Cr) and C-reactive protein (CRP), kidney tissue thiobarbituric acid reactive substances (TBARS), and reduced glutathione (GSH) levels significantly decreased in the PGPE groups compared to the LPS group. Histopathological and immunohistochemical findings revealed that toll-like receptor 4 (TLR4) level and nuclear factor kappa B (NF-κB) expression increased in the LPS group compared to the Control group. In addition, the anti-Gram-negative activity showed a dose-dependent effect on Acinetobacter baumannii, Escherichia coli, and Pseudomonas aeruginosa with the agar well diffusion method and the minimal inhibitory concentration (MIC). The MIC value was remarkable, especially on A. baumannii. We conclude that PGPE has the potential to generate desirable anti-bacterial and anti-inflammatory effects on LPS-induced acute kidney injury in rats.

Sirt1/Nrf2/TNFα; TLR4/Myd88/NF-κB signaling pathways are involved in mediating hepatoprotective effect of bupropion in rat model of myocardial infarction

Background The aim of the current study is to identify the possible protective effect of bupropion (BUP) on liver injury in rat model of myocardial infarction (MI). BUP was administered in the presence and absence of MI. Materials and methods Thirty-two Wistar adult male rats were randomly arranged into four groups: control, BUP (30 mg/kg/day, intraperitoneal) for 28 days, isoproterenol (ISO) was injected subcutaneous (85 mg/kg) in the 26th and 27th days and BUP/ISO groups. Cardiac and hepatic enzymes were measured, also Hepatic oxidative stress indicators, as well as inflammatory and apoptotic biomarkers, were evaluated. Cardiac and hepatic histopathological examination and hepatic nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) immunohistochemical study were also detected. Results ISO significantly increased cardiac and hepatic enzymes, hepatic oxidative stress, inflammatory, apoptotic, with a histopathological picture of cardiac and hepatic damage and high hepatic NF-κB immunoexpression were detected. BUP significantly normalized the upraised oxidative, inflammatory, and apoptotic parameters, with an impressive improvement in the histopathological picture and a reduction in hepatic NF-κB immunoexpression. Conclusion BUP protects against liver injury on top of MI in rat model via modulation of Sirtuin type 1 (Sirt1)/Nuclear factor (erythroid-derived 2)-like 2 (Nrf2)/tumor necrosis factor α (TNFα); Toll-like receptor 4 (TLR4)/Hepatic myeloid differentiation primary response 88(Myd88)/NF-κB signaling pathways.

Machine learning-derived peripheral blood transcriptomic biomarkers for early lung cancer diagnosis: Unveiling tumor-immune interaction mechanisms.

Lung cancer continues to be the leading cause of cancer-related mortality worldwide. Early detection and a comprehensive understanding of tumor-immune interactions are crucial for improving patient outcomes. This study aimed to develop a novel biomarker panel utilizing peripheral blood transcriptomics and machine learning algorithms for early lung cancer diagnosis, while simultaneously providing insights into tumor-immune crosstalk mechanisms. Leveraging a training cohort (GSE135304), we employed multiple machine learning algorithms to formulate a Lung Cancer Diagnostic Score (LCDS) based on peripheral blood transcriptomic features. The LCDS model's performance was evaluated using the area under the receiver operating characteristic (ROC) curve (AUC) in multiple validation cohorts (GSE42834, GSE157086, and an in-house dataset). Peripheral blood samples were obtained from 20 lung cancer patients and 10 healthy control subjects, representing an in-house cohort recruited at the Sixth People's Hospital of Chengdu. We employed advanced bioinformatics techniques to explore tumor-immune interactions through comprehensive immune infiltration and pathway enrichment analyses. Initial screening identified 844 differentially expressed genes, which were subsequently refined to 87 genes using the Boruta feature selection algorithm. The random forest (RF) algorithm demonstrated the highest accuracy in constructing the LCDS model, yielding a mean AUC of 0.938. Lower LCDS values were significantly associated with elevated immune scores and increased CD4+ and CD8+ T-cell infiltration, indicative of enhanced antitumor-immune responses. Higher LCDS scores correlated with activation of hypoxia, peroxisome proliferator-activated receptor (PPAR), and Toll-like receptor (TLR) signaling pathways, as well as reduced DNA damage repair pathway scores. Our study presents a novel, machine learning-derived peripheral blood transcriptomic biomarker panel with potential applications in early lung cancer diagnosis. The LCDS model not only demonstrates high accuracy in distinguishing lung cancer patients from healthy individuals but also offers valuable insights into tumor-immune interactions and underlying cancer biology. This approach may facilitate early lung cancer detection and contribute to a deeper understanding of the molecular and cellular mechanisms underlying tumor-immune crosstalk. Furthermore, our findings on the relationship between LCDS and immune infiltration patterns may have implications for future research on therapeutic strategies targeting the immune system in lung cancer.

Employing single-cell RNA sequencing coupled with an array of bioinformatics approaches to ascertain the shared genetic characteristics between osteoporosis and obesity.

This study examined the relationship between obesity (OB) and osteoporosis (OP), aiming to identify shared genetic markers and molecular mechanisms to facilitate the development of therapies that target both conditions simultaneously. Using weighted gene co-expression network analysis (WGCNA), we analyzed datasets from the Gene Expression Omnibus (GEO) database to identify co-expressed gene modules in OB and OP. These modules underwent Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment and protein-protein interaction analysis to discover Hub genes. Machine learning refined the gene selection, with further validation using additional datasets. Single-cell analysis emphasized specific cell subpopulations, and enzyme-linked immunosorbent assay (ELISA), protein blotting, and cellular staining were used to investigate key genes. WGCNA revealed critical gene modules for OB and OP, identifying the Toll-like receptor (TLR) signalling pathway as a common factor. TLR2 was the most significant gene, with a pronounced expression in macrophages. Elevated TLR2 expression correlated with increased adipose accumulation, inflammation, and osteoclast differentiation, linking it to OP development. Our study underscores the pivotal role of TLR2 in connecting OP and OB. It highlights the influence of TLR2 in macrophages, driving both diseases through a pro-inflammatory mechanism. These insights propose TLR2 as a potential dual therapeutic target for treating OP and OB.

Short Link N Modulates Inflammasome Activity in Intervertebral Discs Through Interaction with CD14.

Intervertebral disc degeneration and pain are associated with the nucleotide-binding domain, leucine-rich repeat, and pyrin domain-containing 3 (NLRP3) inflammasome activation and the processing of interleukin-1 beta (IL-1β). Activation of thehm inflammasome is triggered by Toll-like receptor stimulation and requires the cofactor receptor cluster of differentiation 14 (CD14). Short Link N (sLN), a peptide derived from link protein, has been shown to modulate inflammation and pain in discs in vitro and in vivo; however, the underlying mechanisms remain elusive. This study aims to assess whether sLN modulates IL-1β and inflammasome activity through interaction with CD14. Disc cells treated with lipopolysaccharides (LPS) with or without sLN were used to assess changes in Caspase-1, IL-1β, and phosphorylated nuclear factor kappa-light-chain-enhancer of activated B cells (NFκB). Peptide docking of sLN to CD14 and immunoprecipitation were performed to determine their interaction. The results indicated that sLN inhibited LPS-induced NFκB and Caspase-1 activation, reducing IL-1β maturation and secretion in disc cells. A significant decrease in inflammasome markers was observed with sLN treatment. Immunoprecipitation studies revealed a direct interaction between sLN and the LPS-binding pocket of CD14. Our results suggest that sLN could be a potential therapeutic agent for discogenic pain by mitigating IL-1β and inflammasome activity within discs.

TLR2 and NLRP3 Orchestrate Regulatory Roles in Escherichia coli Infection-Induced Septicemia in Mouse Models.

Introduction Escherichia coli (E. coli) is a significant commensal gram-negative bacterium that can give rise to various diseases. The roles of Toll-like receptor 2 (TLR2) and the NLR pyrin domain-containing 3 (NLRP3) inflammasome in sepsis induced by E. coli infection remain unclear. Methods In vivo, we investigated differences in mortality, production of inflammatory mediators, organ damage, neutrophil count, and bacterial load during E. coli infection in C57BL/6J mice, as well as in mice deficient in TLR2 or NLRP3. In vitro, we investigated the impact of E. coli on the activation of TLR2 and NLRP3 in macrophages and the influence of TLR2 and NLRP3 on the activation of inflammatory signaling pathways and the secretion of inflammatory mediators in macrophages induced by E. coli infection. Results TLR2-deficient (TLR2-/-) and NLRP3-deficient (NLRP3-/-) mice exhibit significantly increased mortality and organ damage after E. coli infection. These mice also show elevated levels of TNF-α and IL-10 in serum and peritoneal lavage fluid (PLF). Additionally, TLR2-/- and NLRP3-/- mice display heightened neutrophil recruitment, increased bacterial load in the blood. Furthermore, macrophages from these mice demonstrate a significant reduction in the activation of the MAPK signaling pathway. Conclusion TLR2 and NLRP3 play crucial roles in modulating inflammatory mediator expression, immune cell recruitment, and bactericidal activity, thereby preventing excessive tissue damage and reducing mortality in E. coli-induced sepsis.

Salmonella Typhimurium infection inhibits macrophage IFNβ signaling in a TLR4-dependent manner.

Type I Interferons (IFNs) generally have a protective role during viral infections, but their function during bacterial infections is dependent on the bacterial species. Legionella pneumophila, Shigella sonnei and Mycobacterium tuberculosis can inhibit type I IFN signaling. Here we examined the role of type I IFN, specifically IFNβ, in the context of Salmonella enterica serovar Typhimurium (STm) macrophage infections and the capacity of STm to inhibit type I IFN signaling. We demonstrate that IFNβ has no effect on the intracellular growth of STm in infected bone marrow derived macrophages (BMDMs) derived from C57BL/6 mice. STm infection inhibits IFNβ signaling but not IFNγ signaling in a murine macrophage cell line. We show that this inhibition is independent of the type III and type VI secretion systems expressed by STm and is also independent of bacterial phagocytosis. The inhibition is Toll-like receptor 4 (TLR4)-dependent as the TLR4 ligand, lipopolysaccharide (LPS), alone is sufficient to inhibit IFNβ-mediated signaling. Cells downregulated their surface levels of IFNα/β receptor 1 (IFNAR1) in response to LPS, which may be mediating our observed inhibition. Lastly, we examined this inhibition in the context of TLR4-deficient BMDMs as well as TLR4 RNA interference and we observed a loss of inhibition with LPS stimulation as well as STm infection. In summary, we show that macrophages exposed to STm have reduced IFNβ signaling via crosstalk with TLR4 signaling, which may be mediated by reduced host cell surface IFNAR1, and that IFNβ signaling does not affect cell-autonomous host defense against STm.

A modern overview of the process of platelet formation (thrombocytopoiesis) and its dependence on several factors.

Structural and functional alterations in platelets are an actual problem that requires more attention. The treatment of these illnesses proves challenging, inefficient and heavily relies on platelet donations. A difficult task confronting science is producing platelets in vitro, which calls for meticulous examination of factors affecting platelet generation. It is known that megakaryocytes produce platelets in vitro and in vivo differently: in the laboratory we can get a smaller number of platelets compared to the human body. This review primarily examines the stages of megakaryocyte maturation and the processes involved in platelet formation. The article reflects the results of both fundamental research on the problem and the new results obtained over the past decade. Currently, most scientists accept the pro-platelets theory of platelet formation. This review aims to explore in detail each stage of pro-platelet formation and the platelet formation process. It explains on the processes of polyploidization, endomitosis, and apoptosis, as well as the functions of structural cell components (microtubules, mitochondria, T- and α-granules) and pro-platelet migration. The microenvironment influence is acknowledged for the osteoblastic and vascular niches that affect thrombocytopoiesis. The additional aspect is the contribution of specific proteins to thrombocytopoiesis such as RhoA, β1-tubulin, cytokines IL-6, IL-8, Toll-like receptors, etc.

Protective Effects of a Dihydrodiazepine Against Endotoxin Shock Through Suppression of TLR4/NF-κB/IRF3 Signaling Pathways.

Sepsis and septic shock are life-threatening systemic inflammatory conditions and among the most frequent causes of morbidity and mortality globally. Preclinical evidence has identified a number of diazepine-based compounds with therapeutic potential in inflammatory diseases. However, the potential anti-inflammatory properties of diazepines in the overwhelming immune response during sepsis have been rarely examined. Thus, the current study aimed to identify a new diazepine compound with therapeutic potential in sepsis. Assessing the inflammatory response of macrophages to Lipopolysaccharides (LPS) in vitro identified 2-[7-(trifluoromethyl)-2,3-dihydro-1H-1,4-diazepin-5-yl]phenol (2-TDDP) as a potential anti-inflammatory agent. It reduced secretion of Interleukin-1β (IL-1β), IL-6, IL-12p70, IL-18, Tumor necrosis factor-α (TNF-α), Interferon-γ (IFN-γ), IFN-β, and increased the secretion of IL-10. In a mouse model of LPS-induced endotoxin shock, 2-TDDP reduced mortality and attenuated inflammation-induced tissue injury in the spleen, liver, kidney, and lung. This was accompanied by reduced serum levels of IL-1β, IL-6, IL-12p70, TNF-α, IFN-γ, IFN-β, and increased levels of IL-10. Importantly, 2-TDDP suppressed the Toll-like receptor 4 (TLR4)/Nuclear factor-κB (NF-κB) and TLR4/Interferon regulatory factor 3 (IRF3) signaling pathways through a reduction in the expression of TLR4, Myeloid differentiation primary response 88 (MyD88), P65, and TNF receptor-associated factor 3 (Traf3). Moreover, 2-TDDP suppressed the expression of CD86, Programmed death-ligand 1 (PD-L1) and C5a receptor (C5aR), but not Major histocompatibility complex II (MHCII). Analysis of splenic lymphocyte populations revealed a decrease in the number of CD4+, CD8+, and B cells. Collectively, these findings introduced the dihydrodiazepine 2-TDDP as a new anti-inflammatory agent with potent therapeutic potential in endotoxin shock, paving an avenue for future clinical application.

Monosodium glutamate induces hypothalamic–pituitary–adrenal axis hyperactivation, glucocorticoid receptors down-regulation, and systemic inflammatory response in young male rats: Impact on miR-155 and miR-218

Abstract Young children are attracted to flavored foods with enhancers, particularly monosodium glutamate (MSG). Experimental studies have proven that MSG can alter the hypothalamic–pituitary–adrenal (HPA) axis response in neonates. We, therefore, investigated the modulation of microRNAs (miRNAs) by dietary MSG and its association with the stimulation of the HPA axis and inflammatory response in young male rats. One-month-old male rats were fed chow enriched with MSG (3 g/kg) for 16 weeks. Feeding MSG to rats markedly up-regulated hypothalamic miR-218, Toll-like receptors-4, and nuclear factor-kB but down-regulated miR-155 and glucocorticoid receptors (GR). In addition, it triggered a remarkable elevation in adrenocortical lipid peroxidation and depletion of antioxidants. These changes were coupled with increased plasma levels of the HPA axis hormones, comprising corticotropin-releasing hormone, adrenocorticotropic hormone, corticosterone levels, and serum pro-inflammatory cytokines. Taken together, current findings indicated that MSG caused an activation of the HPA axis, a down-regulation of GRs, and a systemic inflammatory response. These disturbances were associated with modulating hypothalamic miRNAs, encompassing miR-218 and 155.

Evaluation and Analysis of Traditional Chinese Medicine Treatment of Bovine Viral Diarrhea/Mucosal Disease Based on Network Pharmacology and In Vitro Studies

Background Bovine viral diarrhea/mucosal disease (BVD-MD) is widely distributed worldwide. The disease causes serious economic losses to animal husbandry every year. Finding targeted antiviral drugs proves to be an effective strategy. Purpose This study was based on network pharmacology and in vitro studies to analyze the potential of traditional Chinese medicine (TCM) in the treatment of BVD-MD. Methods The intersection targets between TCM and the disease were identified. Network topology analysis and protein–protein interaction (PPIs) networks were performed. Intersection targets were analyzed for gene ontology (GO) and Kyoto encyclopedia of genes and genomes (KEGG) enrichment. The molecular docking and molecular dynamics simulation methods were used to reveal the degree of binding of core components to key target genes. Characterization of the anti-Bovine viral diarrhea virus (BVDV) effect of TCM by cytotoxicity and in vitro studies. Results The results revealed the selection of five key Chinese medicines, along with 206 targets associated with BVD-MD. The toll-like receptor, PI3K-AKT, and tumor necrosis factor (TNF) signaling pathways were closely related to the five TCMs. AKT1, EGFR, HSP90AA1, and MAPK1 had good binding with quercetin, the core component of Chinese medicine. Molecular dynamics analysis showed that quercetin exhibited complex stability with AKT1. In vitro studies have demonstrated that the inhibitory effect of quercetin on BVDV is mainly in the initial phase. Quercetin inhibited the expression of AKT1. Conclusion The mechanism of BVDV inhibition by the core Chinese medicines is closely related to MAPK1, AKT1, TNF, and HSP90AA1, with the reduction of AKT1 ultimately affecting viral expression.

Neonatal overfeeding attenuates microgliosis and hippocampal damage in an infant rat model of pneumococcal meningitis.

Pneumococcal meningitis (PM) triggers apoptotic neuronal and progenitor cell death in the hippocampal dentate gyrus (DG), resulting in subsequent cognitive impairment. Microglia play a crucial role in PM-induced hippocampal damage. While the lasting effects of neonatal nutrition on health are well documented, the influence of early-life overfeeding on the host response to neuroinfections remains uncertain. This study aimed to examine whether neonatal overfeeding affects the outcome of PM in the hippocampus (HC). Overfeeding was induced by adjusting litter size immediately after birth. On the eleventh day of life, rats were intracisternally injected with Streptococcus pneumoniae or saline, followed by euthanasia after 24 hours for brain dissection. Histological analysis evaluated apoptosis in the DG and the extent of inflammatory infiltrate in the hippocampal fissure, while microgliosis was assessed by immunohistochemistry. The hippocampal transcriptome was analyzed using RNAseq, and the mRNA levels of specific inflammatory biomarkers were evaluated via RT-qPCR. Overfed rats exhibited 40.5% greater body mass compared to their normal-fed counterparts. Intriguingly, PM-induced apoptosis in the DG was 50% lower in overfed rats. This effect was accompanied by significant alterations in the hippocampal transcriptional profile, particularly the lack of activation of the Programmed cell death pathway in overfed infected animals. RT-qPCR analysis of Aif1 and examination of Iba1-immunostained cells revealed mild microgliosis in the HC of infected-overfed animals. This reduced microglial reaction may be attributed to the diminished activation of interferon signaling pathways, as disclosed by the transcriptome analysis, potentially preventing microglial priming. Additionally, evidence of reduced neuroinflammation in overfed rats with PM was observed through the milder activation of pathways associated with Toll-like receptors, interleukins, and chemokine signaling. Furthermore, overfed animals exhibited increased transcription of proinflammatory Il6 and anti-inflammatory Il10 genes, with the latter showing higher expression even in the absence of PM, suggesting a priming effect of overfeeding on hippocampal immune cells. This study sheds light on the complex interplay between early-life overfeeding, immune response, and neuroprotection in infant rats with PM. The findings demonstrate the neuroprotective impact of early-life overfeeding in the context of PM, linked to the modulation of microglial function.

Regulation of versican expression in macrophages is mediated by canonical type I interferon signaling via ISGF3.

Growing evidence supports a role for versican as an important component of the inflammatory response, with both pro- and anti-inflammatory roles depending on the specific context of the system or disease under investigation. Our goal is to understand the regulation of macrophage-derived versican and the role it plays in innate immunity. In previous work, we showed that LPS triggers a signaling cascade involving Toll-like receptor (TLR)4, the Trif adaptor, type I interferons, and the type I interferon receptor, leading to increased versican expression by macrophages. In the present study, we used a combination of chromatin immunoprecipitation, siRNA, chemical inhibitors, and mouse model approaches to investigate the regulatory events downstream of the type I interferon receptor to better define the mechanism controlling versican expression. Results indicate that transcriptional regulation by canonical type I interferon signaling via interferon-stimulated gene factor 3 (ISGF3), the heterotrimeric transcription factor complex of Irf9, Stat1, and Stat2, controls versican expression in macrophages exposed to LPS. This pathway is not dependent on MAPK signaling, which has been shown to regulate versican expression in other cell types. The stability of versican mRNA may also contribute to prolonged versican expression in macrophages. These findings strongly support a role for macrophage-derived versican as a type I interferon-stimulated gene and further our understanding of versican's role in regulating inflammation.NEW & NOTEWORTHY We report the novel finding that versican expression is regulated by the interferon-stimulated gene factor 3 (ISGF3) arm of canonical type I Ifn signaling in LPS-stimulated macrophages. This pathway is distinct from mechanisms that control versican expression in other cell types. This suggests that macrophage-derived versican may play a role in limiting a potentially excessive inflammatory response. The detailed understanding of how versican expression is regulated in different cells could lead to unique approaches for enhancing its anti-inflammatory properties.

Pentraxin-3 is a novel marker for the diagnosis of endometriosis

Introduction: Endometriosis is one of the most common reproductive disorders affecting women. The pathophysiology of endometriosis is also quite complicated, and many mechanisms have been put forward. Pentraxin-3 is a glycoprotein released by leukocyte and myeloid dendritic cells in peripheral blood with the stimulation of some cytokines, especially IL-1 and TNF-α, Toll-Like Receptor agonists, or microbial components. It has been shown that pentraxin-3 is also released from ectopic endometrial cells. The aim of this study is to investigate serum pentraxin-3 levels and their correlation with endometriosis in women with endometriosis and healthy women without endometriosis. Methods: This case-control study evaluated patients diagnosed with endometriosis based on imaging techniques or laparoscopy between 2018 and 2020. Serum samples of 71 patients who had been diagnosed with endometriosis as a case group and 56 healthy patients who had undergone laparoscopic tubal ligation and were proven to be endometriosis-free as a control group were compared for pentraxin-3 level. Results: Serum pentraxin-3 level was significantly higher in the endometriosis group compared with the control group ( p < 0.05). Parity and gravida were significantly higher in the control group than in the endometriosis group. When patients with endometriosis surgery history were excluded, there was no significant change in serum pentraxin-3 levels in patients with medical treatment for endometriosis ( p > 0.05). Both the sensitivity and specificity of serum pentraxin-3 were 73.2%. Conclusions: These results demonstrate a relationship between serum pentraxin-3 and endometriosis. Serum pentraxin-3 might be used as a non-invasive method for the diagnosis of endometriosis.

RIPK2 is crucial for the microglial inflammatory response to bacterial muramyl dipeptide but not to lipopolysaccharide.

Receptor-interacting serine/threonine protein kinase 2 (RIPK2) is a kinase that plays an essential role in the modulation of innate and adaptive immune responses. As a downstream signaling molecule for nucleotide-binding oligomerization domain 1 (NOD1), NOD2, and Toll-like receptors (TLRs), it is implicated in the signaling triggered by recognition of microbe-associated molecular patterns by NOD1/2 and TLRs. Upon activation of these innate immune receptors, RIPK2 mediates the release of pro-inflammatory factors by activating mitogen-activated protein kinases (MAPKs) and nuclear factor-kappa B (NF-κB). However, whether RIPK2 is essential for downstream inflammatory signaling following the activation of NOD1/2, TLRs, or both remains controversial. In this study, we examined the role of RIPK2 in NOD2- and TLR4-dependent signaling cascades following stimulation of microglial cells with bacterial muramyl dipeptide (MDP), a NOD2 agonist, or lipopolysaccharide (LPS), a TLR4 agonist. We utilized a highly specific proteolysis targeting chimera (PROTAC) molecule, GSK3728857A, and found dramatic degradation of RIPK2 in a concentration- and time-dependent manner. Importantly, the PROTAC completely abolished MDP-induced increases in iNOS and COX-2 protein levels and pro-inflammatory gene transcription of Nos2 , Ptgs2 , Il-1β , Tnfα , Il6 , Ccl2 , and Mmp9 . However, increases in iNOS and COX-2 proteins and pro-inflammatory gene transcription induced by the TLR4 agonist, LPS, were only slightly attenuated with the GSK3728857A pretreatment. Further findings revealed that the RIPK2 PROTAC completely blocked the phosphorylation and activation of p65 NF-κB and p38 MAPK induced by MDP, but it had no effects on the phosphorylation of these two mediators triggered by LPS. Collectively, our findings strongly suggest that RIPK2 plays an essential role in the inflammatory responses of microglia to bacterial MDP but not to LPS.

Catecholamines Attenuate LPS-Induced Inflammation through β2 Adrenergic Receptor Activation- and PKA Phosphorylation-Mediated TLR4 Downregulation in Macrophages.

Inflammation is a tightly regulated process involving immune receptor recognition, immune cell migration, inflammatory mediator secretion, and pathogen elimination, all essential for combating infection and restoring damaged tissue. However, excessive inflammatory responses drive various human diseases. The autonomic nervous system (ANS) is known to regulate inflammatory responses; however, the detailed mechanisms underlying this regulation remain incompletely understood. Herein, we aimed to study the anti-inflammatory effects and mechanism of action of the ANS in RAW264.7 cells. Quantitative PCR and immunoblotting assays were used to assess lipopolysaccharide (LPS)-induced tumor necrosis factor α (TNFα) expression. The anti-inflammatory effects of catecholamines (adrenaline, noradrenaline, and dopamine) and acetylcholine were examined in LPS-treated cells to identify the receptors involved. Catecholamines inhibited LPS-induced TNFα expression by activating the β2 adrenergic receptor (β2-AR). β2-AR activation in turn downregulated the expression of Toll-like receptor 4 (TLR4) by stimulating protein kinase A (PKA) phosphorylation, resulting in the suppression of TNFα levels. Collectively, our findings reveal a novel mechanism underlying the inhibitory effect of catecholamines on LPS-induced inflammatory responses, whereby β2-AR activation and PKA phosphorylation downregulate TLR4 expression in macrophages. These findings could provide valuable insights for the treatment of inflammatory diseases and anti-inflammatory drug development.

Interleukin-1 Receptor-Associated Kinase 1 in Cancer Metastasis and Therapeutic Resistance: Mechanistic Insights and Translational Advances.

Interleukin-1 Receptor Associated Kinase 1 (IRAK1) is a serine/threonine kinase that plays a critical role as a signaling transducer of the activated Toll-like receptor (TLR)/Interleukin-1 receptor (IL-1R) signaling pathway in both immune cells and cancer cells. Upon hyperphosphorylation by IRAK4, IRAK1 forms a complex with TRAF6, which results in the eventual activation of the NF-κB and MAPK pathways. IRAK1 can translocate to the nucleus where it phosphorylates STAT3 transcription factor, leading to enhanced IL-10 gene expression. In immune cells, activated IRAK1 coordinates innate immunity against pathogens and mediates inflammatory responses. In cancer cells, IRAK1 is frequently activated, and the activation is linked to the progression and therapeutic resistance of various types of cancers. Consequently, IRAK1 is considered a promising cancer drug target and IRAK1 inhibitors have been developed and evaluated preclinically and clinically. This is a comprehensive review that summarizes the roles of IRAK1 in regulating metastasis-related signaling pathways of importance to cancer cell proliferation, cancer stem cells, and dissemination. This review also covers the significance of IRAK1 in mediating cancer resistance to therapy and the underlying molecular mechanisms, including the evasion of apoptosis and maintenance of an inflammatory tumor microenvironment. Finally, we provide timely updates on the development of IRAK1-targeted therapy for human cancers.

Toll-like receptor 7 protects against intestinal inflammation and restricts the development of colonic tissue-resident memory CD8+ T cells.

The maintenance of intestinal homeostasis depends on a complex interaction between the immune system, intestinal epithelial barrier, and microbiota. Alteration in one of these components could lead to the development of inflammatory bowel diseases (IBD). Variants within the autophagy gene ATG16L1 have been implicated in susceptibility and severity of Crohn's disease (CD). Individuals carrying the risk ATG16L1 T300A variant have higher caspase 3-dependent degradation of ATG16L1 resulting in impaired autophagy and increased cellular stress. ATG16L1-deficiency induces enhanced IL-1β secretion in dendritic cells in response to bacterial infection. Infection of ATG16L1-deficient mice with a persistent strain of murine norovirus renders these mice highly susceptible to dextran sulfate sodium colitis. Moreover, persistent norovirus infection leads to intestinal virus specific CD8+ T cells responses. Both Toll-like receptor 7 (TLR7), which recognizes single-stranded RNA viruses, and ATG16L1, which facilitates the delivery of viral nucleic acids to the autolysosome endosome, are required for anti-viral immune responses. However, the role of the enteric virome in IBD is still poorly understood. Here, we investigate the role of TLR7 and ATG16L1 in intestinal homeostasis and inflammation. At steady state, Tlr7-/- mice have a significant increase in large intestinal lamina propria (LP) granzyme B+ tissue-resident memory CD8+ T (TRM) cells compared to WT mice, reminiscent of persistent norovirus infection. Deletion of Atg16l1 in myeloid (Atg16l1ΔLyz2 ) or dendritic cells (Atg16l1ΔCd11c ) leads to a similar increase of LP TRM. Furthermore, Tlr7-/- and Atg16l1ΔCd11c mice were more susceptible to dextran sulfate sodium colitis with an increase in disease activity index, histoscore, and increased secretion of IFN-γ and TNF-α. Treatment of Atg16l1ΔCd11c mice with the TLR7 agonist Imiquimod attenuated colonic inflammation in these mice. Our data demonstrate that ATG16L1-deficiency in myeloid and dendritic cells leads to an increase in LP TRM and consequently to increased susceptibility to colitis by impairing the recognition of enteric viruses by TLR7. In conclusion, the convergence of ATG16L1 and TLR7 signaling pathways plays an important role in the immune response to intestinal viruses. Our data suggest that activation of the TLR7 signaling pathway could be an attractive therapeutic target for CD patients with ATG16L1 risk variants.

MiR-24 regulates obstructive pulmonary disease in rats via S100A8

Purpose Chronic obstructive pulmonary disease (COPD) is a persistent inflammatory disorder characterized by minor airway inflammation and emphysema involving various cell types and cytokines. MicroRNAs (miRNAs) have emerged as critical regulators in the pathogenesis of lung diseases. This study investigates the impact of microRNA-24 (miR-24) on airway inflammatory responses in a rat model of COPD. Materials and methods The model was established by combining cigarette smoke exposure and lipopolysaccharide stimulation, and rat lung tissues were transfected with adeno-associated viruses overexpressing miR-24. Pathological changes in the lung were assessed using hematoxylin and eosin staining. Levels of pro-inflammatory cytokines, including tumor necrosis factor-alpha, interleukin-6, and interleukin-8, were measured using enzyme-linked immunosorbent assay. Expression of miR-24 and S100A8 was detected through quantitative reverse transcription PCR, while protein levels of S100A8, Toll-like receptor 4 (TLR4), and myeloid differentiation primary response 88 (MyD88) were assessed using western blotting. Bioinformatics analysis and dual-luciferase reporter assay were performed to determine the relationship between S100A8 and miR-24. Results The results demonstrated the downregulation of miR-24 in rats with COPD, and its overexpression resulted in a significant decrease in S1008 mRNA levels. Additionally, the protein level of S100A8 was significantly increased in the lung tissues of COPD rats. The upregulation of miR-24, however, not only inhibited the protein expression of S100A8, TLR4, and MyD88 in lung tissues but also reduced the release of pro-inflammatory cytokines in the plasma and bronchoalveolar lavage fluid, thereby attenuating inflammatory responses and pathological injuries in the lung. Conclusions Our data suggest that miR-24 attenuates airway inflammatory responses in COPD by inhibiting the TLR4/MyD88 pathway via targeting S100A8.

Influence of Donor-Specific Characteristics on Cytokine Responses in H3N2 Influenza A Virus Infection: New Insights from an Ex Vivo Model.

Influenza A virus (IAV) is known for causing seasonal epidemics ranging from flu to more severe outcomes like pneumonia, cytokine storms, and acute respiratory distress syndrome. The innate immune response and inflammasome activation play pivotal roles in sensing, preventing, and clearing the infection, as well as in the potential exacerbation of disease progression. This study examines the complex relationships between donor-specific characteristics and cytokine responses during H3N2 IAV infection using an ex vivo model. At 24 h post infection in 31 human lung explant tissue samples, key cytokines such as interleukin (IL)-6, IL-10, tumor necrosis factor-alpha (TNF-α), and interferon-gamma (IFN-γ) were upregulated. Interestingly, a history of lung cancer did not impact the acute immune response. However, cigarette smoking and programmed death-ligand 1 (PD-L1) expression on macrophages significantly increased IL-2 levels. Conversely, age inversely affected IL-4 levels, and diabetes mellitus negatively influenced IL-6 levels. Additionally, both diabetes mellitus and programmed cell death protein 1 (PD-1) expression on CD3+/CD4+ T cells negatively impacted TNF-α levels, while body mass index was inversely associated with IFN-γ production. Toll-like receptor 2 (TLR2) expression emerged as crucial in mediating acute innate and adaptive immune responses. These findings highlight the intricate interplay between individual physiological traits and immune responses during influenza infection, underscoring the importance of tailored and personalized approaches in IAV treatment and prevention.