Toll-like Receptor Research Articles

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.

Macrophages as Potential Therapeutic Targets in Acute Myeloid Leukemia.

Acute myeloid leukemia (AML) is a heterogenous malignant hemopathy, and although new drugs have emerged recently, current treatment options still show limited efficacy. Therapy resistance remains a major concern due to its contribution to treatment failure, disease relapse, and increased mortality among patients. The underlying mechanisms of resistance to therapy are not fully understood, and it is crucial to address this challenge to improve therapy. Macrophages are immune cells found within the bone marrow microenvironment (BMME), of critical importance for leukemia development and progression. One defining feature of macrophages is their plasticity, which allows them to adapt to the variations in the microenvironment. While this adaptability is advantageous during wound healing, it can also be exploited in cancer scenarios. Thus, clinical and preclinical investigations that target macrophages as a therapeutic strategy appear promising. Existing research indicates that targeting macrophages could enhance the effectiveness of current AML treatments. This review addresses the importance of macrophages as therapeutic targets including relevant drugs investigated in clinical trials such as pexidartinib, magrolimab or bexmarilimab, but also provides new insights into lesser-known therapies, like macrophage receptor with a collagenous structure (MACRO) inhibitors and Toll-like receptor (TLR) agonists.

Lipid-lowering and antioxidant properties of probiotic Bifidobacterium animalis MSMC83 in rats on a high-fat diet.

Hyperlipidaemia, the abnormally high concentration of lipids such as cholesterol in the body, has a series of deleterious effects on health that are least in part are due to increased inflammation and oxidative stress. Probiotics are living microorganisms that possess the efficacy to improve health. Among the many effects that have been ascribed to probiotics is the potential to lower the body lipid content. Here, we used a rat model of induced hyperlipidaemia to assess the lipid-lowering and antioxidant properties of the probiotic strain Bifidobacterium animalis MSMC83 as well as its impact on intestinal barrier immunity and the intestinal microbiota. Oral probiotic intake led to a reduction of body weight, fasting blood glucose, and lipid levels, and increased expression of cholesterol-7α-hydroxylase and antioxidant enzymes. Additionally, B. animalis MSMC83 decreased the levels of liver enzymes and pro-inflammatory cytokines, leading to reduced hepatic steatosis. Furthermore, it re-established intestinal barrier integrity as shown by restoration of the tight junction protein zonula occludens-1 amount and reduced pathogen-induced inflammation in the intestinal epithelium as shown by readjusted expression of toll-like receptors (TLRs). Moreover B. animalis MSMC83 contributed to the maintenance of a balanced, diverse microbiome. Thus, our results indicate that B. animalis MSMC83 alleviates risk factors associated with hyperlipidaemia, suggesting its use as a probiotic to counter the effects associated with unhealthy diets.

Metabolic dysfunction-associated steatotic liver disease exhibits sex-specific microbial heterogeneity within intestinal compartments.

Evidence suggests that the gastrointestinal microbiome plays a significant role in the biology of metabolic dysfunction-associated steatotic liver disease (MASLD). However, it remains unclear whether disparities in the gut microbiome across intestinal tissular compartments between the sexes lead to MASLD pathogenesis. Sex-specific analyses of microbiome composition in two anatomically distinct regions of the gut, the small intestine and colon, were performed using an experimental model of MASLD. The study involved male and female spontaneously hypertensive rats and the Wistar-Kyoto control rat strain, which were fed either a standard chow diet or a high-fat diet for 12 weeks to induce MASLD (12 rats per group). High-throughput 16S sequencing was used for microbiome analysis. There were significant differences in the overall microbiome composition of male and female rats with MASLD, including variations in topographical gut regions. The beta diversity of the jejunal and colon microbiomes was higher in female rats than in male rats (PERMANOVA p-value=0.001). Sex-specific analysis and discriminant features using LEfSe showed considerable variation in bacterial abundance, along with distinct functional properties, in the jejunum and colon of animals with MASLD. Significantly elevated levels of lipopolysaccharide and protein expression of Toll-like receptor 4 were observed in the livers of male rats with MASLD compared with their female counterparts. This study uncovered sexual dimorphism in the gut microbiome of MASLD and identified microbial heterogeneity within intestinal compartments. Insights into sex-specific variations in gut microbiome composition could facilitate customised treatment strategies.

TLR4 Downregulation Identifies High-Risk HPV Infection and Integration in H-SIL and Squamous Cell Carcinomas of the Uterine Cervix.

Growing scientific evidence suggests a link between the expression of toll-like receptor 4 (TLR4) and cervical cancer carcinogenesis. Specifically, a close relation between TLR4 expression and FIGO stage, lymph node metastases, and tumor size has been reported in cervical cancer. In the present study, we aimed to evaluate the relationship between TLR4 expression levels and human papillomavirus (HPV) infection and/or high-risk (hr) HPV integration status in patients with a histological diagnosis of high-grade squamous intraepithelial lesion (H-SIL), and squamous cell carcinoma (SCC) of the uterine cervix. Sixty biopsies of cervical neoplasia, comprising H-SIL (n = 20) and SCC (n = 40), were evaluated for TLR4 expression by immunohistochemistry. All samples were positive for high-risk HPV as confirmed by in situ hybridization (ISH) and broad-spectrum PCR followed by Sanger sequencing analysis. The intensity of TLR4 staining was higher in tissues negative for intraepithelial lesion or malignancy (NILM) than in H-SIL, and further reduced in SCC. Moreover, statistically significant differences have been observed in the percentage of TLR4 expression between NILM and H-SIL and between H-SIL and SCC, with higher percentages of expression in H-SIL than in SCC. Our results showed a significant downregulation of TLR4 in HPV-related H-SIL and SCC, compared to NILM. These data support the hypothesis that TLR4 expression is suppressed in HPV-driven oncogenesis.

TLR4/TRIF/Caspase-8/Caspase-1 Pathway in Choroidal Endothelial Cells Promotes Choroidal Neovascularization

Purpose The purpose of this study was to investigate the role and mechanism of caspase-8 in the development of choroidal neovascularization induced by age-related macular degeneration, with the aim of identifying a potential therapeutic target for neovascular age-related macular degeneration. Methods Mouse models of laser photocoagulation-induced choroidal neovascularization and hypoxic human choroidal endothelial cells were utilized to examine the involvement of caspase-8 in choroidal neovascularization development. The toll-like receptor 4/TIR domain-containing adaptor molecule 1/caspase-8 pathway was explored in hypoxic human choroidal endothelial cells to elucidate its contribution to pathological angiogenesis. Various experimental techniques, including inhibition assays and immunoblotting analysis, were employed to assess the effects and mechanisms of caspase-8 activation. Results Inhibition of caspase-8 demonstrated attenuated choroidal neovascularization development in mice subjected to laser photocoagulation. Activation of the toll-like receptor 4/TIR domain-containing adaptor molecule 1/caspase-8 pathway was observed in hypoxic human choroidal endothelial cells. Upon activation by the toll-like receptor 4/TIR domain-containing adaptor molecule 1 axis, caspase-8 directly cleaved caspase-1, leading to the cleavage of interleukin-1β and interleukin-18 by caspase-1. Consequently, activation of interleukin-1β and interleukin-18 through the toll-like receptor 4/TIR domain-containing adaptor molecule 1/caspase-8/caspase-1 pathway promoted the proliferative, migratory, and tube-forming abilities of hypoxic human choroidal endothelial cells. Conclusion The findings of this study indicate that caspase-8 plays a crucial role in promoting choroidal neovascularization by activating interleukin-1β and interleukin-18 through the toll-like receptor 4/TIR domain-containing adaptor molecule 1/caspase-8/caspase-1 pathway in choroidal endothelial cells. Therefore, targeting caspase-8 may hold promise as a therapeutic approach for neovascular age-related macular degeneration.

G-protein–coupled estrogen receptor 30 regulation of signaling downstream of protein kinase Cε mediates sex dimorphism in hyaluronan-induced antihyperalgesia

Abstract High molecular weight hyaluronan (HMWH) inhibits hyperalgesia induced by diverse pronociceptive inflammatory mediators and their second messengers, in rats of both sexes. However, the hyperalgesia induced by ligands at 3 pattern recognition receptors, lipopolysaccharide (a toll-like receptor 4 agonist), lipoteichoic acid (a toll-like receptor 2/6 agonist), and nigericin (a NOD-like receptor family, pyrin domain containing 3 activator), and oxaliplatin and paclitaxel chemotherapy–induced peripheral neuropathy are only attenuated in males. After gonadectomy or intrathecal administration of an antisense to G-protein–coupled estrogen receptor 30 (GPER) mRNA, HMWH produces antihyperalgesia in females. In nociceptors cultured from rats that had been treated with oxaliplatin, HMWH reverses nociceptor sensitization from male and GPER antisense–treated female, but not from gonad intact females. G-protein–coupled estrogen receptor–dependent sex dimorphism for HMWH-induced antihyperalgesia was also observed for the prolongation of prostaglandin E2 (PGE2)-induced hyperalgesia in primed nociceptors. While in primed rats, HMWH inhibits early, protein kinase A-dependent hyperalgesia, 30 minutes post PGE2 injection, in both sexes; measured 4 hours post-PGE2, HMWH inhibits the protein kinase Cε (PKCε)-dependent prolongation of PGE2 hyperalgesia only in males and GPER antisense–treated females. In females, hyperalgesia induced by PKCε agonist, ψεRACK, in control but not in primed nociceptors, was inhibited by HMWH. Inhibitors of 2 GPER second messengers, extracellular-regulated kinase 1/2 and nonreceptor tyrosine kinase, also unmasked HMWH antihyperalgesia in females with oxaliplatin chemotherapy–induced peripheral neuropathy, a condition in which nociceptors are primed as well as sensitized. Our results support GPER-dependent sex dimorphism in HMWH-induced antihyperalgesia for pain induced by pattern recognition receptor agonists, and chronic inflammatory and neuropathic pain, mediated by changes in signaling downstream of PKCε in primed nociceptors.

Linker Chemistry and Connectivity Fine-Tune the Immune Response and Kinetic Solubility of Conjugated NOD2/TLR7 Agonists.

There is a growing interest in developing novel immune potentiators capable of eliciting a cellular immune response. We tackle this challenge by harnessing the synergistic cross-activation between two innate immune receptors─the nucleotide-binding oligomerization domain-containing protein 2 (NOD2) and Toll-like receptor 7 (TLR7). Herein, we investigate the structure-activity relationship of a series of novel conjugated NOD2/TLR7 agonists incorporating a variety of flexible aliphatic, poly(ethylene glycol)-based and triazole-featuring linkers. Our findings reveal potent immune-enhancing properties of conjugates in human primary peripheral blood mononuclear cells, characterized by a Th1/Th17 polarized cytokine response. Importantly, we demonstrate that both the chemistry of the linker and the site of linkage affect the immune fingerprint and the kinetic solubility of these conjugated agonists. These results shed further light on the immunostimulatory potential of NOD2/TLR7 cross-activation and provide insights for designing innovative immune potentiators.

Exercise Promotes Hippocampal Neurogenesis in T2DM Mice via Irisin/TLR4/MyD88/NF-κB-Mediated Neuroinflammation Pathway.

Neuroinflammation is a major feature of type 2 diabetic mellitus (T2DM), adversely affecting hippocampal neurogenesis. However, the precise mechanism is not fully understood, and therapeutic approaches are currently lacking. Therefore, we determined the effects of exercise on neuroinflammation and hippocampal neurogenesis in T2DM mice, with a specific focus on understanding the role of the irisin and related cascade pathways in modulating the beneficial effects of exercise in these processes. Ten-week exercise significantly decreased T2DM-induced inflammation levels and markedly promoted hippocampal neurogenesis and memory function. However, these positive effects were reversed by 10 weeks of treatment with cyclo RGDyk, an inhibitor of irisin receptor signaling. Additionally, exercise helped reduce the M1 phenotype polarization of hippocampal microglia in diabetic mice; this effect could be reversed with cyclo RGDyk treatment. Moreover, exercise markedly increased the levels of fibronectin type III domain-containing protein 5 (FNDC5)/irisin protein while decreasing the expression of Toll-like receptor 4 (TLR4), myeloid differential protein-88 (MyD88), and nuclear factor kappa-B (NF-κB) in the hippocampus of T2DM mice. However, blocking irisin receptor signaling counteracted the down-regulation of TLR4/MyD88/NF-κB in diabetic mice undergoing exercise intervention. Conclusively, exercise appears to be effective in reducing neuroinflammation and enhancing hippocampal neurogenesis and memory in diabetes mice. The positive effects are involved in the participation of the irisin/TLR4/MyD88/NF-κB signaling pathway, highlighting the potential of exercise in the management of diabetic-induced cognitive decline.

MiR-5195-3p predicts clinical prognosis and represses colorectal cancer progression by targeting TLR4/MyD88 signaling

BackgroundRecent studies have highlighted the role of miR-5195-3p in suppressing cell growth in various cancers. However, the specific functional impact of miR-5195-3p in colorectal cancer (CRC) remain to be fully clarified. The importance of miR-5195-3p in CRC was evaluated, aiming to uncover its underlying molecular mechanism and identify it as a potential therapeutic target for CRC.ResultsOur research has shown that miR-5195-3p is markedly under-expressed in CRC tissues and cell cultures, with its reduced presence associated with a higher TNM stage, lymphatic invasion, and unfavorable survival outcome. Ectopic miR-5195-3p expression curtailed proliferation, migration, and invasion of SW1116 and HT29 cells. Additionally, we discovered that miR-5195-3p directly targets and negatively influences Toll-like receptor 4 (TLR4) in CRC cells. Moreover, an inverse relationship was noted between miR-5195-3p and TLR4 expression in CRC tissue samples. Notably, restoring TLR4 expression counteracted miR-5195-3p’s suppressive impact on cell growth, motility, invasiveness, epithelial-mesenchymal transition (EMT), and the TLR4/MyD88 signaling pathway in SW1116 and HT29 cells.ConclusionsMiR-5195-3p suppresses the CRC cellular functions through the downregulation of TLR4/MyD88 signaling, indicating that targeting miR-5195-3p might offer a viable therapeutic strategy for CRC.

Inhalation of Microplastics Induces Inflammatory Injuries in Multiple Murine Organs via the Toll-like Receptor Pathway.

Previous studies have detected microplastics (MPs) in human biological samples, such as lungs, alveolar lavage fluid, and thrombus. However, whether MPs induce health effects after inhalation are unclear. In this study, fluorescent polystyrene microplastics (PS-MPs) were found in the thymus, spleen, testes, liver, kidneys, and brain on day 1 or day 3 after one intratracheal instillation. Furthermore, mice showed inflammation in multiple organs, manifested as obvious infiltration of neutrophils and macrophages, increased Toll-like receptors (TLRs), myeloid differentiation primary response protein 88 (MyD88) and nuclear factor-κB (NF-κB), as well as proinflammatory cytokines (tumor necrosis factor (TNF)-α and interleukin (IL)-1β) in the lungs, thymus, spleen, liver, and kidneys after four intratracheal instillations of PS-MPs at once every 2 weeks. Hepatic and renal function indexes were also increased. Subsequently, the inflammatory response in multiple murine organs was significantly alleviated by TLR2 and TLR4 inhibitors. Unexpectedly, we did not find any elevated secretion of monocyte chemotactic protein (MCP)-1 or TNF-α by RAW264.7 macrophages in vitro. Thus, PS-MPs induced inflammatory injuries in multiple murine organs via the TLRs/MyD88/NF-κB pathway in vivo, but not macrophages in vitro. These results may provide theoretical support for healthy protection against PS-MPs and their environmental risk assessment.

Tumor-Selective Nano-Dispatcher Enforced Cancer Immunotherapeutic Effects via Regulating Lactate Metabolism and Activating Toll-Like Receptors.

The development of tumors relies on lactate metabolic reprogramming to facilitate their unchecked growth and evade immune surveillance. This poses a significant challenge to the efficacy of antitumor immunity. To address this, a tumor-selective nano-dispatcher, PIMDQ/Syro-RNP, to enforce the immunotherapeutic effect through regulation of lactate metabolism and activation of toll-like receptors is developed. By using the tumor-targeting properties of c-RGD, the system can effectively deliver monocarboxylate transporters 4 (MCT4) inhibitor (Syro) to inhibit lactate efflux in tumor cells, leading to decreased lactate levels in the tumor microenvironment (TME) and increased accumulation within tumor cells. The reduction of lactate in TME will reduce the nutritional support for regulatory T cells (Tregs) and promote the effector function of T cells. The accumulation of lactate in tumor cells will lead to tumor death due to cellular acidosis. In addition, it will also reduce the uptake of glucose by tumor cells, reduce nutrient plunder, and further weaken the inhibition of T cell function. Furthermore, the pH-responsive release of Toll-like receptors (TLR)7/8 agonist IMDQ within the TME activates dendritic cells (DCs) and promotes the infiltration of T cells. These findings offer a promising approach for enhancing tumor immune response through targeted metabolic interventions.

Influence of polymorphisms on the phenotype of TLR1, TLR4 and TLR9 genes and their association with cervical cancer: Bioinformatics prediction analysis and a case-control study

Susceptibility to cervical cancer has been associated with Toll-like receptors (TLRs), which is an important component of innate immunity. According to previous studies, polymorphisms in TLRs genes can affect immune response pathways and lead to the development of cervical cancer. The present study aims to evaluate the functionality of polymorphisms in TLR1, TLR4 and TLR9 genes and their associations with cervical cancer. To identify the functionality of polymorphisms, we used the following tools: MUpro, ChimeraX, SNP2TFBS and GTEx. A case-control study including 57 cases (11 High-grade Intraepithelial Lesion - HSIL and 46 cervical cancer) and 67 clinically healthy controls was conducted in the Brazilian population. Polymorphisms genotyping was performed by real-time PCR, using TaqMan probes, using the allelic discrimination method. Bioinformatics prediction showed that the TLR1 rs4833095 [NM_003263.4 (TLR1):c.743T>C (p.Asn248Ser)] and TLR4 rs4986790 [NM_138554.5 (TLR4):c.896A>G (p.Asp299Gly)] polymorphisms alter the structure and stability of their respective proteins. TLR9 rs187084 [NM_017442.3(TLR9):c.-1486A>G] polymorphism seems to affect the THAP1 binding site and modify gene expression. In the case-control study, the c.743TC heterozygous genotype of the rs4833095 SNP in the TLR1 gene was associated with an increased risk for HSIL/cervical cancer. No association of TLR4 rs4986790 and TLR9 rs187084 SNPs with HSIL/cervical cancer was found in the studied population. Allelic combination CAG (rs4833095/ rs4986790/ rs187084) increased the risk of cervical cancer. In conclusion, the present study identified that polymorphisms in TLRs genes can affect the phenotype of their respective genes and contribute to the development of HSIL or cervical cancer.

Expression profile of Toll-like receptors and cytokines in the cecal tonsil of chickens challenged with Eimeria tenella.

Chicken coccidiosis, caused by Eimeria spp., seriously affects the development of the poultry breeding industry. Currently, extensive studies of chicken coccidiosis are mostly focused on acquired immune responses, while information about the innate immune response of chicken coccidiosis is lacking. Toll-like receptor (TLR), the key molecule of the innate immune response, connects innate and adaptive immune responses and induces an immune response against various pathogen infections. Therefore, the quantitative real-time PCR was used to characterize the expression profile of chicken TLRs (chTLRs) and associated cytokines in the cecal tonsil of chickens infected with Eimeria tenella. The results showed that the expression of chTLR1a, chTLR2a, and chTLR5 was significantly upregulated at 3h post-infection, while chTLR1b, chTLR2b, chTLR3, chTLR7, chTLR15 and chTLR21 was significantly downregulated (p < 0.05). In addition, chTLR1a expression rapidly reached the peaked expression at 3h post-infection, while chTLR2b and chTLR15 peaked at 168h post-infection, and chTLR2a expression was highest among chTLRs, peaking at 48h post-infection (p < 0.05). For cytokines, interleukin (IL)-6 and tumor necrosis factor (TNF)-α peaked at 96h post-infection, IL-4 and IL-12 peaked at 144h post-infection, and interferon-γ expression was highest among cytokines at 120h post-infection. In addition, IL-12 and IL-17 were markedly upregulated at 6h post-infection (p < 0.05). These results provide insight into innate immune molecules during E. tenella infection in chickens and suggest that innate immune responses may mediate resistance to chicken coccidiosis.

Exenatide improves cisplatin induced ovarian damage through NLRP3, Nrf-2, and TLR4 pathways.

Cisplatin (CP) toxicity causes ovarian damage by oxidative stress, inflammation and fibrosis. The aim of the present study is to investigate the possible beneficial effects of exenatide on the experimental ovarian damage model produced by CP. For 14 rats, CP was administered by intraperitoneally (i.p) twice a week for 5 weeks. No drug was administered to the remainder of rats (n = 7) (Group 0). The rats taken CP were divided into two groups. Group 1 rats (n = 7) were given 1 mL/kg/day saline i.p., and Group 2 rats (n = 7) was given with 20 μg/kg/day exenatide. The number of primordial, primary, secondary, and tertiary follicle was significantly lower in Group1 compared with Group 0 and Group 2. The ovarian fibrosis percent was significantly higher in Group 1 than Group 0 and 2. The plasma anti-Mullerian hormone value was lower in Group1compared with Group 0 and 2. Over Nuclear factor-erythroid factor 2-related factor 2 level, Over Toll-like receptor 4 level and over nucleotide-binding domain leucine-rich repeat and pyrin domain containing receptor 3 were higher in Group 1 compared with Group 0 and 2. Exenatide has possible beneficial effect on ovarian damage induced by CP by anti-inflammatory actions and can be a promising candidate for ovarian damage caused by CP.