Search Results (783)

We investigated the role of the intermediate-conductance, Ca²⁺-activated K⁺ channel K_Ca3.1 and volume-regulatory anion channel LRRC8A in regulating C-C motif chemokine ligand 2 (CCL2) expression in THP-1-differentiated M₂ macrophages (M₂-MACs), which serve as a useful model for studying tumor-associated macrophages (TAMs). CCL2 is a potent chemoattractant involved in the recruitment of immunosuppressive cells and its expression is regulated through intracellular signaling pathways such as ERK, JNK, and Nrf2 in various types of cells including macrophages. The transcriptional expression of CCL2 was suppressed in M₂-MACs following treatment with a K_Ca3.1 activator or an LRRC8A inhibitor via distinct signaling pathways: ERK–CREB2 and JNK–c-Jun pathways for K_Ca3.1, and the NOX2–Nrf2–CEBPB pathway for LRRC8A. Under in vitro conditions mimicking the elevated extracellular K⁺ concentration ([K⁺]_e) characteristic of the tumor microenvironment (TME), CCL2 expression was markedly upregulated, and this increase was reversed by treatment with them in M₂-MACs. Additionally, the WNK1–AMPK pathway was, at least in part, involved in the high [K⁺]_e-induced upregulation of CCL2. Collectively, modulating K_Ca3.1 and LRRC8A activities offers a promising strategy to suppress CCL2 secretion in TAMs, potentially limiting the CCL2-induced infiltration of immunosuppressive cells (TAMs, T_regs, and MDSCs) in the TME. Full article

Hibiscus syriacus L. (HS), native to Eastern and Southern Asia, has been traditionally used in Asian herbal medicine for its anticancer, antimicrobial, and anti-inflammatory properties. Despite these recognized bioactivities, its potential cardioprotective effects, particularly in the setting of heart failure (HF), remain largely unexplored. This study aimed to investigate the effects of HS extracts and its bioactive constituents on angiotensin II (Ang II)-induced cardiac injury using an in vitro model with H9c2 rat cardiomyocytes. Cells exposed to Ang II were pretreated with HS extracts, and assays were performed to assess cell viability, reactive oxygen species (ROS) generation, protein synthesis, and secretion of inflammatory mediators, including tumor necrosis factor-alpha, interleukin 1β (IL-1β), and interleukin 6 (IL-6), as well as chemokine (CCL20) and HF-related biomarkers, such as brain natriuretic peptide (BNP) and endothelin-1. The results demonstrated that HS extracts significantly and dose-dependently attenuated Ang II-induced ROS accumulation and suppressed the secretion of pro-inflammatory cytokines, chemokines, BNP, and endothelin-1. Additionally, HS and its purified components inhibited Ang II-induced protein synthesis, indicating anti-hypertrophic effects. Collectively, these findings highlight the antioxidative, anti-inflammatory, and antihypertrophic properties of HS in the context of Ang II-induced cardiac injury, suggesting that HS may represent a promising adjunctive therapeutic candidate for HF management. Further in vivo studies and mechanistic investigations are warranted to validate its clinical potential. Full article

Kynurenic acid (KYNA), a tryptophan metabolite, possesses immunomodulatory properties, although the molecular mechanism of this action has not yet been resolved. In the present study, the effects of KYNA on the secretion of selected cytokines and chemokines by macrophages derived from the human THP-1 cell line are investigated. Furthermore, the involvement of the aryl hydrocarbon receptor (AhR) and the G protein-coupled receptor 35 (GPR35) in mediating the effects of KYNA was examined. In lipopolysaccharide (LPS)-stimulated THP-1-derived macrophages, KYNA significantly reduced IL-6 and CCL-2, but increased IL-10 and M-CSF levels. AhR antagonist CH-223191 reduced the KYNA influence on IL-6, CCL-2, and M-CSF production, while the GPR35 antagonist, ML-145, blocked KYNA-induced IL-10 production. Furthermore, it was shown that THP-1 derived macrophages were capable of synthesizing and releasing KYNA and that its production was increased in the presence of LPS. These findings suggest that THP-1-derived macrophages are a source of KYNA and that KYNA modulates inflammatory responses predominantly through AhR and GPR35 receptors. Our study provides further evidence for the involvement of macrophages in immunomodulatory processes that are dependent on AhR and GPR35 receptors, as well as the potential role of KYNA in these phenomena. Full article

IgG antibodies, signaling via the inhibitory receptor, FcγRIIb, are potent inhibitors of IgE-mediated mast cell activation. We have previously reported that in addition to blocking mast cell degranulation, inhibitory IgG signals shut down a proinflammatory transcriptional program in which mast cells produce cytokines and chemokines known to drive type 2 tissue inflammation. To determine whether such effects of allergen-specific IgG can modulate allergic inflammation in vivo, we examined the airways of mice sensitized to ovalbumin (OVA) by intraperitoneal injection and then challenged with intranasal OVA. Pretreatment with allergen-specific IgG significantly reduced the recruitment of inflammatory cells, including macrophages and eosinophils, into the lungs of OVA-sensitized mice. The bronchoalveolar lavage fluid of OVA-challenged mice contained elevated levels of chemokine ligands (CCL2 and CCL24) and interleukin-5, a response that was markedly blunted in animals receiving allergen-specific IgG. IgG-treated animals exhibited attenuated allergen-induced production of IgE, IL-4, and IL-13, along with impaired OVA-induced goblet cell hyperplasia and Muc5ac expression and suppressed airway hyperresponsiveness, consistent with a shift away from a Th2 response. Using mice with a lineage-specific deletion of FcγRIIb, we demonstrated that each of these protective effects of IgG was dependent upon the expression of this receptor on mast cells. Overall, our findings establish that allergen-specific IgG can reduce allergen-driven airway inflammation and airway hyperresponsiveness and point to a mechanistic basis for the therapeutic benefit of aeroallergen-specific IgG therapy. Full article

Ruxolitinib, a clinically approved JAK1/2 inhibitor used in the treatment of hematologic malignancies and inflammatory conditions, has been shown to interfere with the function of cytotoxic T lymphocytes (CTLs). Previous studies supported the involvement of the multidrug resistance transporter P-glycoprotein (Pgp/ABCB1) in CTL biology; however, the nature of its regulation remains unclear. To address this, we investigated the impact of ruxolitinib on Pgp expression and function in human CD8⁺ T cells. We demonstrate that CD8⁺ T lymphocytes express Pgp dynamically at both the mRNA and protein levels across naïve, short-term, and long-term activation states. Ruxolitinib increased the calcein accumulation in human Pgp-overexpressing NIH-3T3 cells and in CTLs and directly modulated Pgp function by increasing its basal ATPase activity in a concentration-dependent manner (10–100 μM), similar to the effect of the known Pgp substrate/modulator verapamil. Although measurable ATPase stimulation and transport inhibition were observed at supratherapeutic concentrations of ruxolitinib, its Pgp-mediated efflux may also occur at therapeutically relevant concentrations. In contrast, at therapeutically relevant plasma concentrations (1–3 μM), ruxolitinib significantly stabilized the mRNA expression of Pgp during early T-cell receptor (TCR) activation and inhibited the TCR-induced upregulation of Pgp, CD8, and PD-1 surface markers, suggesting its interference with activation-associated differentiation. At these same concentrations, ruxolitinib also impaired CCL19-directed transmigration of CTLs across human umbilical vein endothelial cell (HUVEC) monolayers, indicating disruption of lymphoid homing cues. Collectively, these findings demonstrate that ruxolitinib modulates Pgp at both the transcriptional and functional levels, with distinct concentration dependence. The ability of ruxolitinib to alter CTL activation and migration at clinically relevant plasma concentrations highlights the need for careful evaluation of JAK inhibitor–mediated immunomodulation and its implications for vaccination, transplantation, and T cell-based immunotherapies. Full article

Cancer metastasis often accounts for the primary cause of cancer-related mortality, with the lymphatic system playing a pivotal role in the dissemination of malignant cells. While hematogenous vessel spread is commonly associated with distant organ metastasis, the lymphatic system serves as an early conduit for tumor cell invasion and dissemination. The process of lymphatic metastasis is a highly coordinated sequence of events that involves cancer cell invasion, intravasation into lymphatic vessels, survival, transport, and colonization of regional lymph nodes (LNs). Cancerous cells then establish micro-metastases at the colonized sites and expand in the new microenvironment, ultimately resulting in the generation of secondary tumors. Tumor-secreted factors, such as vascular endothelial growth factors (VEGF-C and VEGF-D), contribute to metastasis through lymphangiogenesis, the formation of new lymphatic vessels. In addition, cancer cells utilize pre-existing chemokine signaling pathways by expressing chemokine receptors, such as CCR7, which bind to chemokine ligands, such as CCL19 and CCL21, to facilitate targeted migration into the lymphatic vessels. LNs are often the initial sites for metastasis and therefore are indicators of distant organ involvement. It is well established that the location and extent of LN involvement provides significant prognostic information, although the optimal treatment approach for LN metastases remains a subject of debate. Understanding the mechanisms of lymphatic metastasis offers potential therapeutic targets to mitigate cancer progression. Full article

Purpose: To profile tear cytokine changes in Allogeneic Hematopoietic Stem Cell Transplant (HSCT) patients after instillation of daily topical cyclosporine-A 0.1% cationic emulsion. Methods: Participants in a longitudinal study were given cyclosporine eyedrops daily from 3 to 5 weeks before and 3 months, 6 months, and 12 months post-HSCT. The outcomes included tear cytokine concentration assayed by the Proximity Extension Assay O-linked target 96 platform. The patients were divided into two groups: Group 1 (n = 8 conjunctival CD4 cells responding to cyclosporine) and Group 2 (n = 5 conjunctival CD4 cells not suppressed after cyclosporine, where patients were non-compliant with cyclosporine). All participants had a standardized clinical examination, including meibomian gland evaluation and tear breakup times. Results: The levels of 38 cytokines/chemokines showed significant changes (p < 0.05) over time, and in many, the elevation was marked at one year. These include gamma-interferon, CXCL9, CCL3, and CCL4 (all p < 0.0001). For gamma-interferon, there was significant interaction between group and time at 1 year (p = 0.022), where the cytokine was significantly suppressed in Group 1. Four other cytokines showed significant group and time interaction at 1 year: FGF23, FGF5, LIFR, and Enrage (all p < 0.05). All patients had either withdrawal or a reduction in systemic immunomodulation between 6 months and 1 year. We found several cytokines to be associated with changes in tear osmolarity or symptom scores. Conclusions: HSCT induces significant elevation of 38 tear cytokines/chemokines even without the occurrence of ocular graft-versus-host disease when systemic immunosuppression is reduced within the first year. Topical daily cyclosporine eyedrops can reduce some pro-inflammatory tear cytokines. Full article

Metabolic syndrome (MetS), characterized by obesity, insulin resistance, and dyslipidemia, is a major risk factor for renal injury. Oxidative stress (OxS) plays a pivotal role in its progression; however, the underlying molecular mechanisms are not fully understood. In this study, we established a rat model of MetS using a high-fat diet combined with a single-dose streptozotocin injection in male Wistar rats. MetS rats exhibited systemic OxS, evidenced by elevated circulating levels of free oxygen radicals and decreased antioxidant defense capacity, as well as hypertension, renal lipid peroxidation, glomerular hyperfiltration, and renal tubular injury. Transcriptomic profiling of renal tissue revealed significant downregulation of six OxS-related genes: C-C motif chemokine ligand 5 (CCL5), glutamate-cysteine ligase catalytic subunit, glutathione peroxidase 6, recombination activating gene 2, NAD(P)H: quinone oxidoreductase 1, and selenoprotein P-1. Among these downregulated genes, CCL5 was further confirmed to be repressed at both mRNA and protein levels across intrarenal and systemic compartments. Given its documented functions in immune signaling and redox homeostasis, CCL5 downregulation may contribute to enhanced oxidative damage in MetS-associated renal injury. These findings highlight the role of redox gene dysregulation in the pathogenesis of MetS-related kidney disease and support the potential of CCL5 as a biomarker for oxidative renal injury. Full article

Hepatitis B virus (HBV) infection is one of the most dangerous viral diseases, with innate immunity representing the first line of defense against the virus. In this branch of the immune system, neutrophils are considered key cellular mediators. To better understand the implication of neutrophils in the distinct stages of the disease, HBV-infected patients were enrolled in this study and categorized into three groups: patients with acute infection, chronic infection under treatment, and at early cirrhotic stage. To elucidate the role of inflammatory mediators and cellular mechanisms of neutrophilic origin in the course of the infection, both ex vivo and in vitro studies were performed. Increased levels of C-C motif chemokine ligand 2 (CCL2), interleukin (IL)-18, IL-33, and citrullinated histone H3 (CitH3)—an accurate marker of neutrophil extracellular traps (NETs)—were detected in the circulation of patients with acute infection or early cirrhosis. In parallel, sera from the aforementioned patient groups induced the formation of IL-1b-bearing NETs in neutrophils from healthy individuals. These inflammatory NETs affected primary fibroblasts towards acquiring a pro-fibrotic phenotype. These results suggest that NETs could be regarded as mediators in hepatitis B manifestations, while their therapeutic targeting could enhance the management of early-stage cirrhotic patients. Full article

Canine chronic enteropathy (CE) is a gastrointestinal disorder characterized by persistent or recurrent digestive symptoms lasting more than three weeks. It shares similarities with human inflammatory bowel disease but its immunopathogenesis remains poorly characterized in dogs. The aim of this study was to characterize the local and systemic immune profile of dogs with CE by assessing cytokine and chemokine expression in serum and intestinal tissue, as well as the mRNA expression of immune-related receptors such as integrins, chemokine receptors, and cytokines. Duodenal biopsies and blood samples were collected from five dogs diagnosed with a CE and five healthy controls. Serum concentrations of cytokines and chemokines were determined by multiplex ELISA, and mRNA expression in the intestinal mucosa was analyzed by quantitative PCR. Dogs with a CE showed increased expression of pro-inflammatory cytokines, including TNF-α and IFN-γ, and increased concentrations of chemokines such as CXCL10 and CCL2 in both serum and tissue samples. Increased mRNA expression of the chemokine receptor CCR9 and the adhesion molecule MAdCAM-1 were also observed in intestinal samples. These findings provide new insights into the immune response involved in CE and may aid the development of future diagnostic biomarkers and targeted therapies for canine chronic enteropathies. Full article

Mycobacterium marinum is an opportunistic pathogen prevalent in aquatic environments, causing significant morbidity in fish, including Coho salmon (Oncorhynchus kisutch), a species increasingly cultured in Chinese salmonid aquaculture. This study investigated the immune response of Coho salmon to M. marinum infection and the bacterial proliferation dynamics in the liver and kidney. Transcriptome analysis revealed 5028 differentially expressed genes (DEGs) in the kidney and 3419 DEGs in the liver at 6 weeks post-infection. Gene Ontology and KEGG enrichment analysis highlighted pathways such as cytokine–cytokine receptor interaction, metabolic pathways, and Toll-like receptor signaling in the kidney, while the DEGs in the liver were enriched in metabolic pathways, immune system processes, and stress and defense responses. The temporal expression profiling of 15 immune-related genes, including acute-phase proteins (serum amyloid A-5 and hepcidin), cytokines (TNF-α, IL-1β, IL-17A), chemokines (CXCL13 and CCL19), pattern recognition receptors (Toll-like receptor 13), and other immune-related genes, showed significant upregulation against M. marinum infection, with stronger responses in the liver. Furthermore, it was found that there was a progressive proliferation of M. marinum in the infected liver and kidney from approximately 2.5 log₁₀ cfu/g at week 2 to about 6 log₁₀ cfu/g by 6 weeks, with a significantly higher load in the liver. These findings provide critical insights into the immune mechanisms of Coho salmon against M. marinum and the pathogen’s tissue-specific proliferation, offering a foundation for developing targeted control strategies against M. marinum in aquaculture. Full article

Background/Objectives: Cellular aging represents a crucial element in the progression of musculoskeletal diseases, contributing to muscle atrophy, functional decline, and alterations in bone turnover, which promote fragility fractures. However, knowledge about expression patterns of factors potentially involved in aging and senescence at the tissue level remains limited. Our pilot study aimed to characterize the expression profile of cell cycle regulators, factors released by aged cells, and sirtuin 1 (SIRT1) in the muscle tissue of 26 elderly patients undergoing hip arthroplasty, including 13 with low-energy fracture and 13 with osteoarthritis (OA). Methods: The mRNA expression levels of cyclin-dependent kinase inhibitor 1A (CDKN1A), cyclin-dependent kinase inhibitor 1B (CDKN1B), cyclin-dependent kinase inhibitor 2A (CDKN2A), p53, tumor necrosis factor alpha (TNF-α), interleukin-1 beta (IL-1β), interleukin-6 (IL-6), interleukin-15 (IL-15), chemokine (C-C motif) ligand 2 (CCL2), chemokine (C-C motif) ligand 3 (CCL3), growth differentiation factor 15 (GDF15), and SIRT1 were evaluated in muscle tissue by qRT-PCR. In addition, immunohistochemistry and Western blotting analysis were conducted to measure the protein levels of SIRT1. Results: A marked muscle atrophy was observed in fractured patients compared to the OA group, in association with an up-regulation of cell cycle regulators and factors released by the aged cells. The expression of matrix metallopeptidase 3 (MMP3), plasminogen activator inhibitor 1 (PAI-1), and fas cell surface death receptor (FAS) was also investigated, although no significant differences were observed between the two experimental groups. Notably, SIRT1 expression was significantly higher in OA patients, confirming its role in maintaining muscle health during aging. Conclusions: Further studies will be needed to clarify the role of SIRT1 in the senescence characteristic of age-related musculoskeletal disorders, counteracting the muscle atrophy that predisposes to fragility fractures. Full article

Background/Objectives: Metabolic dysfunction-associated steatotic liver disease (MASLD) is the most prevalent chronic liver disease worldwide, contributing to metabolic dysfunction and increased healthcare costs. The green Mediterranean diet reduces intrahepatic fat and elevates the plasma levels of 2,5-dihydroxybenzoic acid (2,5-DHBA), suggesting a mechanistic role for 2,5-DHBA in hepatic lipid metabolism. This study aimed to evaluate the therapeutic potential of 2,5-DHBA in MASLD and elucidate its molecular mechanism. Methods: Lipid accumulation was assessed in oleic acid-treated HepG2 cells and a high-fat diet (HFD)-induced MASLD mouse model. RNA sequencing, molecular docking, and immunohistochemical staining were performed to investigate the molecular mechanisms, focusing on the chemokine (C-C motif) ligand 2 (CCL2)–CCL2 receptor (CCR2) axis. Results: 2,5-DHBA significantly reduced hepatic lipid accumulation in both HepG2 cells and HFD-fed mice in a dose-dependent manner. RNA sequencing revealed the marked downregulation of CCL2, a key proinflammatory mediator in MASLD pathogenesis. Molecular docking predicted that 2,5-DHBA competed with CCL2 for binding at the CCR2 axis. Immunohistochemistry further confirmed that 2,5-DHBA treatment lowered hepatic CCL2 expression, suppressed nuclear factor-κB activation, and reduced inflammatory cell infiltration. These findings suggest that 2,5-DHBA exerted anti-steatotic effects by modulating the CCL2-CCR2 signaling pathway. Conclusions: This is the first study to demonstrate that 2,5-DHBA attenuates hepatic steatosis via targeting the CCL2-CCR2 axis. These findings highlight its potential as a novel nutraceutical strategy for MASLD treatment. Full article

Respiratory syncytial virus (RSV) is the leading cause of respiratory infections in children. Extracellular vesicles (EVs), released by airway epithelial cells, contain proteins and different families of non-coding RNAs (EV cargo) that can modulate the responses of target cells to viral infection. Nasal mucosa is a primary site of viral entry and the source of EVs present in the upper airway secretions. In this study we characterized proteins, including inflammatory mediators and cytokines, and the piwi-interacting RNA (piRNAs) cargo of EVs isolated from pediatric human nose organoids (HNO) and nasopharyngeal secretions (NPS) positive for RSV. Using Proximity Extension Assay (PEA) and Luminex multi-target arrays, we found significant enrichment in several chemokines and other mediators/biomarkers, including CCL2, CCL20, CXCL5, CX3CL1, CXCL6, MMP-1, MMP-10, uPA, Flt3L, ARNT and CD40 in EVs secreted by RSV-infected HNO compared to control mock HNO. Analysis of NPS samples from RSV infected children revealed that CCL3, CCL20, CXCL8, uPA, VEGFA, were concentrated in the NPS-EV fraction. LC-MS/MS and Gene Ontology indicated that RSV positive NPS-EVs originate from different cellular sources, with the most abundant proteins from neutrophils and epithelial cells. A total of 490 piRNAs were detected by NGS sequencing of small RNA libraries obtained from NPS-EVs, which has not been reported prior to this study. Identification of inflammatory mediators and small non-coding RNAs which are compartmentalized in EVs contributes to understanding mechanisms of virus-mediated pathogenesis in RSV infections. Full article

Patients with head and neck squamous cell carcinoma (HNSCC) suffer from severe morbidity and mortality. Immunotherapy represents a novel promising treatment option. Therefore, a better understanding of the immune niche is needed. This study focuses on the spatial distribution and prognostic value of different T cell subtypes in 84 HNSCC specimens as well as chemokine and cytokine levels associated with spatial T cell infiltration. Density of T helper (T_H), cytotoxic (CTL), and regulatory T cells (T_reg) was quantified by multicolor tissue cytometry on a single cell level in whole tissue sections, discriminating between T cells located in epithelial tumor cell nests or tumor stroma, respectively. In addition, quantitative levels of 27 immune-related factors were assessed. Survival analysis of patients with p16-negative HNSCC revealed higher stromal T_reg densities to be an independent prognostic factor for better progression-free and overall survival. Furthermore, high levels of CXCL10, IL-9, and CCL4 were associated with significantly higher numbers of T cells, especially for CTL with direct contact to tumor cells, whereas for VEGF the opposite effect was observed in the tumor stroma. In conclusion, T_reg cell infiltration as well as distinct cytokine levels could serve as new immune biomarkers in p16-negative HNSCC to predict survival and the spatial distribution of T cells. Full article