The autoimmune regulator (Aire) gene is essential for the establishment of central tolerance by promoting the ectopic expression of tissue-restricted antigens in medullary thymic epithelial cells (mTECs). Although Aire deficiency impairs negative selection, its impact on thymic architecture, output, and early T cell development remains incompletely understood. Here, we used a murine Aire-deficient model carrying a CRISPR-Cas9-induced deletion within exon 6, disrupting the SAND domain, to investigate the consequences of Aire loss on thymic structure and function. Aire-deficient mice exhibited focal alopecia and marked disorganization of the thymic medulla. T cell receptor excision circle (TREC) analyses revealed altered peripheral TREC dynamics and a reduced sj/betaTREC ratio, indicating altered thymic output and impaired intrathymic precursor expansion. Complementary flow-cytometric analysis showed a significant reduction in the proportion of CD4+CD8+ double-positive thymocytes in heterozygous mutants, supporting altered intrathymic maturation. Serum IL-27, IL-28B, and IL-31 were decreased in homozygous mutants, whereas IL-22 and IL-23 remained unchanged. Despite these alterations, TCRVbeta repertoire sequencing revealed no significant differences in rearrangements, clonality, or V/J gene usage. Together, these findings indicate that Aire deficiency disrupts thymic architecture and output, while preserving overall TCR repertoire diversity.
Secondary immunodeficiency or immunoparesis is an acquired impairment of the immune system which is a recognised feature of multiple myeloma (MM). Although classically described as a reduced production of polyclonal immunoglobulins, MM induces global immune dysfunction with defects in innate immune and T cell effector function. Immunoparesis can be detected in premalignant states, including monoclonal gammopathy of uncertain significance. Patients with immunoparesis have a poorer prognosis and present with severe, recurrent infection, which is associated with increased morbidity and mortality especially in the first 3 months following diagnosis, with relapse or progression and with the use of novel therapeutics. Bacterial infections, and specifically pneumonia, are the commonest presentation although there are increased risks of both viral reactivation and invasive fungal infection. Management of immunoparesis in MM includes the use of antimicrobial prophylaxis, and immunoglobulin replacement therapy particularly during high-risk periods. Although vaccination against encapsulated organisms, reactivation viruses, and circulating seasonal viruses is recommended, the response in MM may be suboptimal.
BACKGROUND:Epidermal growth factor receptor tyrosine kinase inhibitors (EGFR-TKIs) frequently cause gastrointestinal adverse events, including diarrhea, and are associated with alterations in intestinal mucosal defense. Previous studies have shown that afatinib alters Paneth cell-associated α-defensin 5 (DEFA5)-related antimicrobial defense and epithelial transport-related molecular readouts, including cystic fibrosis transmembrane conductance regulator (CFTR) expression. OBJECTIVE:We investigated whether S-equol, a gut microbiota-derived isoflavone metabolite, shifts selected afatinib-associated mucosal immune-epithelial and promoter methylation readouts toward their corresponding control values under defined post-exposure and post-dose intervention schedules. METHODS:Mature Caco-2 monolayers were exposed to 20 µM afatinib for 24 h, washed, and subsequently treated with either 600 µM S-equol or 600 µM R-equol for 24 h. Rats received afatinib at 5.2 mg/kg once daily for 14 consecutive days, followed 2 h after the final afatinib dose by a single oral dose of S-equol at 7.8 mg/kg. Tissues were collected 24 h after the final afatinib dose, corresponding to 22 h after S-equol administration. In a separate differentiation-stage experiment, Caco-2 cells were exposed to 1000 nM afatinib for 24 h starting Day 1, treated with 60 µM S-equol for 24 h starting Day 7, and harvested on Day 14. RESULTS:In mature Caco-2 monolayers, afatinib reduced DEFA5, WNT3, and LGR5 protein abundance relative to that under the vehicle-control condition. Subsequent S-equol treatment increased the abundance of all three proteins relative to that under the afatinib-only condition and shifted these readouts toward their corresponding control values. R-equol produced similar directional shifts in WNT3 and LGR5 protein abundance but not in DEFA5 protein abundance. In rats, post-dose S-equol shifted afatinib-reduced intestinal Defa5 and Pigr mRNA expression and lipopolysaccharide-stimulated Peyer's patch IgA output toward their corresponding control values, while attenuating afatinib-associated increases in CFTR protein abundance, Peyer's patch IgA protein abundance, and salivary IgA concentration. Salivary IgA concentration was positively associated with IgA protein abundance in Peyer's patch across treatment groups. In the differentiation-stage experiment, S-equol attenuated afatinib-associated increases in the DEFA5 and SOX9 promoter methylation indices and shifted afatinib-reduced DEFA5 protein abundance toward the control value. CONCLUSIONS:Across complementary epithelial and animal models, S-equol shifted selected afatinib-associated mucosal immune-epithelial and promoter methylation readouts toward their corresponding control values. These findings identify molecular responses that warrant further investigation using direct intestinal functional measurements and models that evaluate compatibility with afatinib-mediated tumor control.
Innate lymphoid cells (ILCs) lack antigen-specific receptors and are, consequently, distinct from conventional T and B lymphocytes. Nevertheless, they play a critical role in immune regulation through the rapid secretion of effector cytokines. Due to these functional properties, ILCs exhibit remarkable similarities to CD4+ T helper lymphocytes. With the inclusion of natural killer (NK) cells in the current ILC classification, ILCs are now considered the innate immune system counterparts of adaptive T lymphocytes. Recent studies have demonstrated that ILCs are essential for maintaining tissue homeostasis, particularly at mucosal surfaces, and their dysregulation is closely linked to the pathogenesis of inflammatory, autoimmune and allergic diseases. Beyond their roles in chronic inflammation, ILCs act as pivotal integrators of dietary, microbial, and neuroendocrine signals within specialized tissue microenvironments. In allergic diseases and asthma, ILC2s initiate early type 2 responses, while in the intestinal mucosa, plasticity between ILC3 and ILC1 subsets critically regulates the balance between homeostasis and inflammatory bowel disease (IBD). Additionally, ILCs are emerging as significant players in tumor immunology, exhibiting both pro- and anti-tumorigenic roles depending on the tumor microenvironment. This review provides a comprehensive overview of ILC biology, their tissue-specific functions, and their involvement in immune-mediated disorders. Moreover, we discuss the therapeutic potential of targeting ILCs, including cell-based approaches and their significance as biomarkers, to establish a framework for next-generation precision immunotherapies.
Obesity is characterized by chronic inflammation and sustained hyperleptinemia, conditions known to disrupt immune homeostasis. Natural killer (NK) cells are particularly sensitive to metabolic cues, and obese individuals frequently display impaired NK cell activity. Here, we investigated how leptin modulates NK cell frequency, activation, effector function, and intracellular signaling in normal-weight (NW) and obese (OB) adults. Obesity was associated with reduced frequencies of circulating NK cells, particularly among women, along with decreased expression of the activating receptors CD16 and NKp46. Upon leptin exposure, NK cells from obese donors exhibited impaired effector function, characterized by reduced granzyme B intensity and diminished IFN-γ production. Leptin further modulated these responses in a weight-dependent manner: whereas high-concentration leptin (50 nM) enhanced IFN-γ production in NK cells from normal-weight individuals, it induced a suppressive response in cells from obese donors. At the signaling level, NK cells from obese individuals showed reduced phosphorylation of ERK1/2 and p38 following leptin stimulation, which correlated with impaired cytotoxic and cytokine responses. In contrast, mTOR phosphorylation was intrinsically reduced in NK cells from obese donors, independent of leptin exposure. Sex-stratified analyses revealed higher mTOR activation and granzyme B expression in NK cells from lean men compared to lean women under high leptin conditions, whereas leptin-mediated suppression of IFN-γ was more pronounced in men across both NW and OB groups. Together, these findings suggest that leptin exerts divergent effects on NK cells depending on metabolic and sex-related context, contributing to the functional impairment observed in obesity.
As immune cells develop and migrate throughout the body, they encounter diverse biochemical, mechanical and osmotic environments. These environments can induce rapid changes in cell volume through the activation or inhibition of ion channels and transporters. Recent work has highlighted that volume regulation contributes to key aspects of immune-cell function, including migration, activation and proliferation. Focusing on neutrophils, macrophages, dendritic cells and T cells, the cell types for which the most direct experimental evidence is currently available, this review summarises our current understanding of the underlying mechanisms and examines the evidence linking changes in cell volume to immune-cell behaviour, including the extent to which such changes are passive consequences of environmental conditions or active regulators of immune-cell function. We also highlight the limited use of direct volume measurements and discuss how this constrains our current understanding, will review available approaches to measure cell volume, and outline key directions for future work.
Influenza virus remains a major global public health threat due to its high transmissibility, ability to cause severe disease, and pandemic potential. Although vaccination is the main strategy for reducing the burden of influenza, it has notable limitations. Therefore, novel therapeutic and immunomodulatory approaches are urgently needed.Here, using plasmid-driven reverse genetics, we engineered a defective recombinant influenza virus encoding interleukin 7 (Flu:IL-7), to investigate the role of IL-7 in influenza pathogenesis and immunomodulation in a murine model. Our findings demonstrate that Flu:IL-7 is safe and effectively attenuates disease severity caused by a wild-type replicative influenza virus (PR8), improving clinical recovery and reducing disease-associated morbidity.Notably, local expression of IL-7 through the Flu:IL-7 vector at the time of PR8 infection significantly reduced the severity of secondary pneumococcal pneumonia. These protective effects were associated with earlier and more coordinated pulmonary immune response, characterized by enhanced activation and expansion of innate and adaptive immune cells, early formation of iBALT-like aggregates, and reduced tissue damage.Collectively, our findings suggest that transient local IL-7 expression temporally reprograms the pulmonary immune response, promoting early antiviral immunity followed by timely resolution of inflammation. In summary, Flu:IL-7 provides a useful experimental model for investigating the immunomodulatory role of IL-7 during influenza infection. Our findings support transient local IL-7 expression as a promising host-directed strategy to improve disease outcome and reduce influenza-associated complications. These concepts may also be applicable to other diseases in which modulation of the local immune response is desirable, including lung cancer and COVID-19.
Polymorphonuclear leukocytes (PMNs), predominantly neutrophil granulocytes, are key components of the innate immune system that eliminate invading pathogens through phagocytosis and clear apoptotic cells through efferocytosis. Beta-2-microglobulin (β2m) is best known as the light chain of major histocompatibility complex class I (MHC I), where it is required for antigen presentation to CD8⁺ T cells. However, emerging evidence suggests that extracellular β2m may also regulate innate immune responses. Here, we show that extracellular β2m enhances neutrophil phagocytosis and efferocytosis. Addition of soluble β2m (50 µg/ml) increased phagocytosis of latex beads by PMNs from 23% to 31%, whereas the proteolytically cleaved variant desLys58-β2m (dK58β2m) had no effect. In contrast, both β2m and dK58β2m enhanced phagocytosis of the Gram-positive bacterium Streptococcus pyogenes and the Gram-negative bacterium Acinetobacter baumannii by >3.6-fold. Furthermore, both β2m variants promoted efferocytosis of apoptotic Jurkat cells in a dose-dependent manner, resulting in up to a two-fold increase that was comparable to the effect of GM-CSF. Cytochalasin D abolished β2m-mediated uptake of apoptotic cells. Pre-incubation of latex beads with β2m followed by washing did not enhance phagocytosis, and pre-incubation of PMNs with β2m followed by washing did not enhance subsequent efferocytosis of apoptotic cells. These findings indicate that β2m does not act by coating phagocytic targets or by inducing sustained neutrophil priming. Collectively, these findings identify extracellular β2m as a regulator of neutrophil-mediated phagocytosis and efferocytosis and demonstrate that proteolytic processing differentially influences these activities.
BACKGROUND:Langerhans cells (LCs) play a crucial role in sensing and processing stimuli from the skin and the external environment. They are implicated in various skin disorders, acting either as pro-inflammatory agents or as regulatory elements. However, the exact function of LCs in the pathogenesis of psoriasis remains unclear. Psoriasis is characterized by a significant Th17/Treg immune imbalance. To elucidate the independent effect of Treg dysfunction on psoriatic inflammation, we established a Treg-inhibited psoriasis group using the Foxp3 inhibitor CMD178, in conjunction with LC depletion models to investigate the LC-Th17/Treg regulatory axis METHODS: Psoriasis models were established through the topical application of IMQ. Female C57BL/6 N mice were divided into three groups: control, IMQ-induced psoriasis, and Treg-depleted psoriasis. LCswere depleted using diphtheria toxin in Langerin-DTR mice, while Tregs were inhibited by CMD178. Inflammation was assessed through measurements of ear thickness, histopathology, apoptosis, and cytokine production. Additionally, RT-qPCR was utilized to detect the expression of S100A7, S100A8, and MMP2. Flow cytometry was employed to analyze the functional status of lymphocytes and T cell phenotypes. RESULTS:IMQ-induced psoriasis-like lesions exhibit activated LCs, an increase in Th17 cells, a decrease in Tregs, elevated levels of IL-17A and IL-22, and reduced levels of TGF-β and PD-L1. Depletion of LCs alleviated the lesions and restored the Th17/Treg balance, whereas depletion of Tregs exacerbated the inflammation. CONCLUSION:LCs play a central regulatory role in the psoriatic skin environment by influencing the Th17/Treg.
A high-fat diet (HFD) has been associated with dysregulated immunity both at the systemic and mucosal levels in mouse models. The underlying mechanisms are partially known. We analysed the effect of diet on immunity in HLA-DQ8 (DQ8) transgenic mice, a specific model of gluten sensitivity, and investigated a possible interplay between gluten and an HFD. Our aim was to further dissect the impact of a long-term HFD regimen on both systemic and intestinal immunity. Adult DQ8 mice (n = 6/group) were fed a gluten-free diet (GFD), an HFD, or an HFD containing 8% gluten (HFD+G) for 23 weeks. We assessed clinical parameters, the phenotype and function of splenic and lamina propria (LP) T cells by flow cytometry, and multiparametric quantitation of cytokines induced in vitro. An HFD for 23 weeks increased body weight as expected, paralleled by a significant shortening of the caecum (P < 0.001). FACS analysis revealed that an HFD drastically increased the number of potentially cytotoxic (CD8+) T cells in the LP but not in the spleen. An HFD+G significantly reduced, but did not abolish, the percentage of these cells (p < 0.01). Induction of cytokine secretion in vitro showed that LP cells from both HFD- and HFD+G-fed mice did not secrete any cytokines, whereas both innate (TNF-α) and adaptive immunity (IL-4, IFN-γ and IL-17) cytokines were induced in the LP of GFD-fed mice. In conclusion, we found that a long-term HFD regimen impaired the phenotype of immune cells and their effector functions distinctly in the small intestine of gluten-sensitive DQ8 mice. The presence of gluten in the diet changed the phenotype but did not abolish the block of effector activity. Our findings could help further elucidate the functional status of the immune system under the HFD regimen.
Iron deficiency, but also iron overload (as observed in Friedreich's ataxia), are associated with an increased prevalence of restless legs syndrome (RLS). The pathophysiological mechanisms linking these conditions to RLS remain unknown. Here, we propose that elevated circulating levels of interleukin-6 (IL-6) may represent a common underlying factor linking changes in iron homeostasis to the development of RLS. Peripheral IL-6 affects systemic iron distribution, central dopamine synthesis, and sensory neuron sensitivity, thereby contributing to the motor and sensory manifestations of RLS. We suggest that IL-6-mediated signaling may represent a potential therapeutic target for RLS.
The advent of single-cell technologies has provided unprecedented resolution of dendritic cell (DC) heterogeneity, yet it has paradoxically fueled conceptual fragmentation and conflicting nomenclatures. The field is currently divided by divergent models of conventional type 2 DC (cDC2) ontogeny and the debated identity of the DC3 subset, whether it constitutes a distinct hematopoietic lineage or a transient activation state. In this Perspective, I critically analyze these conflicting classifications, focusing on the cDC2A/DC2A developmental dichotomy and the integration of newly described populations such as transitional DC-derived DC2s (tDC2s). I emphasize that these disputes extend beyond semantics, profoundly impacting our mechanistic understanding of disease and therapeutic targeting, as evidenced by the distinct roles of pro-DC3s in viral myocarditis and tDC2s in immune tolerance. I argue that the immunology community urgently requires a consensus framework based on rigorous ontogenetic and functional criteria to harmonize DC classification and translate high-resolution mapping into actionable clinical insights.
Cancer is a disease of the self. Unlike infectious threats that come from outside, malignancies arise from within, when ordinary cells abandon their cooperative roles and pursue unchecked proliferation. This betrayal of biological solidarity offers profound metaphors for understanding interpersonal and social dysfunction. Drawing on the "hallmarks of cancer," the concept of the tumor microenvironment, and advances in immunotherapy, this essay argues that cancer mirrors many relational pathologies: the violation of boundaries, exploitation of shared resources, the masking of harm, and the spread of dysfunction across contexts. Just as tumors evade immune surveillance and exploit systemic vulnerabilities, individuals and groups in social systems manipulate tolerance and exploit complicity. Yet cancer biology also offers lessons in resilience. Therapies reveal the importance of precise, context-sensitive intervention; the biology of remission emphasizes the possibility of repair without erasure. By reflecting on malignancy as metaphor, we see that the health of human relationships requires balance between growth and restraint, vigilance and tolerance, repair and prevention. Cancer thus becomes a biomedical frontier and an ethical and social teacher, offering insights into how individuals and communities can recognize, contain, and heal the malignant dynamics that arise from within.
BACKGROUND:Antineutrophil cytoplasmic antibody (ANCA)-associated vasculitis (AAV) is a systemic autoimmune disorder characterized by vascular inflammation and the activation of neutrophils. Complement component 5a (C5a) is pivotal in neutrophil priming and ANCA-mediated activation. Although progranulin (PGRN) is recognized for its involvement in inflammatory processes, yet its specific role in ANCA-associated vasculitis (AAV) remains poorly understood This study investigates the functional interplay between PGRN and C5a in enhancing neutrophil activation in response to ANCA stimulation. METHODS:Neutrophils were primed with recombinant PGRN and subsequently stimulated with myeloperoxidase (MPO)-ANCA or proteinase 3 (PR3)-ANCA-positive immunoglobulin G. The respiratory burst was evaluated through dihydrorhodamine oxidation, while degranulation was quantified by measuring lactoferrin release. Additionally, the effects of PGRN-neutralizing antibodies on C5a-primed neutrophils were evaluated. RESULTS:PGRN significantly upregulated membrane-bound proteinase 3 expression in neutrophils compared to untreated controls (368.0 ± 18.5 vs. 178.0 ± 14.7, p < 0.001) and enhanced MPO release in the culture supernatants (1462.8 ± 202.2 vs. 526.8 ± 118.8, p < 0.001). PGRN-primed neutrophils demonstrated increased respiratory burst activity (p < 0.001) and elevated lactoferrin release (p < 0.001) compared to non-primed cells. Inhibition of PGRN significantly diminished ANCA-mediated oxygen radical production (p < 0.001) and degranulation (p < 0.001) in C5a-primed neutrophils. CONCLUSIONS:PGRN functionally enhances C5a-mediated neutrophil activation, suggesting a cooperative effect but not a direct molecular interaction. This in vitro study using human neutrophils explores the cooperative effects of PGRN and C5a in ANCA-induced activation. Future research should investigate the use of PGRN inhibitors to mitigate inflammation in AAV.
Autoimmune thyroid diseases (AITDs), including Graves' disease (GD) and Hashimoto's disease (HD), are thyroid-specific autoimmune diseases; however, predicting prognosis is difficult. Follicular helper T (Tfh) cells play a critical role in the differentiation of B cells and can be divided into three distinct subsets (Tfh1, Tfh2, and Tfh17), each with different abilities to regulate B cell responses. To elucidate the roles of circulating Tfh cells in the pathogenesis and prognosis of AITDs, we determined their proportions in the peripheral blood of patients with AITD and genotyped single-nucleotide variants (SNVs) in the CXCR5 gene. The proportion of circulating Tfh1 cells in Tfh cells was significantly higher in patients with GD and HD compared with control subjects. In contrast, the proportion of circulating Tfh2 cells was significantly lower in patients with GD and HD compared with control subjects. The G allele of CXCR5 SNV3 was significantly more frequent in patients with severe HD compared with those with mild HD. Overall, increased circulating Tfh1 cells and decreased circulating Tfh2 cells may play an important role in the pathogenesis of AITDs. The G allele of CXCR5 SNV3 may have a role in HD severity.
BACKGROUND:Rheumatoid arthritis (RA) is a common chronic systemic autoimmune disease. Accumulating evidence reveals that lactate plays a vital role in progression of RA. This study aimed to identify lactate-related genes (LRGs) associated with RA and investigate their potential pathological correlation with immune-infiltrating cells and their potential as diagnostic markers in RA. METHODS:We downloaded four RA-related datasets containing 51 RA and 36 healthy synovium samples from the Gene Expression Omnibus (GEO) database and extracted the expression profiles of lactate-related genes to identify differentially expressed genes (DEGs). Subsequently, weighted gene co-expression network analysis (WGCNA) and least absolute shrinkage and selection operator (LASSO) were employed to identify the hub genes for RA diagnosis. Receiver operating characteristic (ROC) curves were used to validate the specificity and sensitivity of the hub genes. Additionally, we analyzed the infiltration levels of 28 immune cells in the expression profiles and their relationship with the hub genes using single-sample gene set enrichment analysis. Finally, the expression levels and function of hub genes were verified. RESULTS:We identified 373 differentially expressed LRGs (232 upregulated and 141 downregulated genes) between RA and healthy controls. WGCNA and LASSO analysis results identified TPK1 and IBA57 as potential diagnostic biomarkers for RA. The combination of the two hub genes showed great diagnostic ability (AUC > 0.8) in multiple independent validation dataset. Additionally, IBA57 may involved in the immune mechanism of DC cells, whereas TPK1 may be involved in regulating macrophage function in the immune response to RA. Also, TPK1 is highly expressed in RA, and knockdown TPK1 attenuated RA-FLS invasion and migration. CONCLUSIONS:These data indicate that lactate-related gene TPK1 and IBA57 are valuable diagnostic biomarkers for RA and are closely associated with immune cell infiltration, and TPK1 may play an important role in RA progression. These findings provide new insights into the pathogenesis of RA and potential diagnostic targets.
BACKGROUND:Allergic asthma is a chronic respiratory condition characterized by persistent airway inflammation and dysregulated macrophage activation. Although aerobic exercise is known to exert anti-inflammatory effects, its influence on macrophage polarization and mitochondrial dynamics in asthma remains poorly defined METHODS: Using a rat model of ovalbumen (OVA)-induced allergic asthma, we investigated the impact of aerobic exercise on macrophage polarization and mitophagy regulation. Animals were divided into four experimental groups: control, OVA-induced asthma, OVA with aerobic exercise intervention, and OVA with Drp1 inhibitor (Mdivi-1) treatment. We evaluated airway inflammation, macrophage phenotypes, mitochondrial function, and key mitophagy-related proteins RESULTS: Aerobic exercise significantly attenuated allergic airway inflammation, as evidenced by reduced inflammatory cell infiltration, decreased mucus production, and a shift in macrophage polarization from the M2 towards the M1 phenotype. At the molecular level, exercise suppressed mitophagy activation, reduced Drp1 phosphorylation, and downregulated the expression of mitophagy-related proteins. These effects were mirrored by Drp1 inhibition with Mdivi-1, confirming the crucial role of Drp1-mediated mitophagy in exercise-induced modulation of macrophage polarization CONCLUSION: Our findings indicate that aerobic exercise alleviates allergic airway inflammation by inhibiting Drp1-dependent mitophagy and rebalancing macrophage polarization. These results provide novel mechanistic insights into the therapeutic potential of exercise in asthma and highlight mitophagy as a promising target for inflammatory respiratory diseases.
