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Reconstitution of human bronchial epithelial cells in humanized mice with autologous immune system 2335217
Abstract Introduction Humanized mouse models were developed to bridge fundamental differences between murine and human immune systems and to overcome the limitations of in vitro models. However, existing humanized mice fail to fully recapitulate human mucosal immunity in the lung due to the absence of human airway epithelial cells within their native tissue architecture. Ths, we sought to develop a novel model with human immune cells alongside a human lung epithelium within the mouse lung, provides a physiologically relevant platform to investigate human epithelial—immune crosstalk in vivo. Methods NSGF6 mice, NSG‑based immunodeficient mice carrying a murine Flt3 knockout and human IL6 knock‑in, were engrafted with donor‑matched human bone marrow—derived CD34+ hematopoietic stem and progenitor cells and bronchial epithelial progenitor cells. Engraftments were performed at different time points to synchronize immune and epithelial development. Human immune cell and lung epithelial cell reconstitution were assessed by flow cytometry and immunofluorescence microscopy. Mice were infected intranasally with live influenza A virus to evaluate epithelial functionality and human immune responses. Results Following transplantation, human airway epithelial cells differentiated within their appropriate physiological niche in the murine lung and were permissive to human influenza A virus infection. Spatial analysis revealed a distinct compartmentalization, with human airway epithelial cells localized along the airway lining, whereas human immune cells were distributed throughout lung tissues, with sites of epithelial—immune interaction observed. In response to acute human influenza A virus infection, a significant increase in human CD14+CD16+ monocytes were detected in the lung. Conclusion Our model enables in situ reconstitution of human immune and airway epithelial compartments, providing a physiologically relevant platform to investigate human lung mucosal immunity, respiratory infection, and immune‑mediated lung disease. Funding Source R21 OD032454 and R01 CA219880 Topic Categories Transplantation Immunology (TRAN)
Inflammasome Activation Limits Radiotherapy-Induced Antitumor Immunity and Shapes CD8+ Responses in a Sex-Dependent Manner 2260523
Abstract Introduction Radiation therapy (RT) induces immunogenic cell death and antitumor immunity. We demonstrated that myeloid MyD88 signaling restricts CD8+ T cell responses after RT and hypothesized that inflammasome activation downstream of MyD88 contributed to this phenotype. Methods We cocultured bone marrow—derived macrophages and PK5L1940 pancreatic tumor cells in the presence of specific inhibitors to assess the mechanism of inflammasome activation following RT. WT or ASC-mCitrine reporter mice bearing subcutaneous tumors were treated with RT and the caspase-1 inhibitor VX765, with immune profiling and inflammasome activity assessed by flow cytometry. Results RT activated the NLRP3 inflammasome in macrophages dependent on ER Ca2+ release, and mitochondrial ROS. VX765 improved post-RT survival in a CD8+ T cell—dependent manner, with efficacy observed in males. VX765 enhanced the expansion of T-bet+Eomes+Ki67+ CD8+ effector cells and increased tumor antigen—specific T cells. RT also induced IL-23 expression and Th17 response, while VX765 further upregulated IL-23R expression on CD4+ T cells. In females, estrogen and progesterone suppressed inflammasome activation, and VX765 efficacy required hormonal blockade. Conclusion NLRP3 inflammasome activation limits RT-induced antitumor immunity by restricting CD8+ responses. VX765 enhances effector CD8+ T cell expansion and survival in males post-RT. Sex hormones modulate inflammasome activity, impacting efficacy of its blockade in combination with RT. Funding Source Collins Medical Trust K22 CA251643 Providence Foundation Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Monoclonal antibody therapy promotes survival by recalibrating neuroinflammation during Australian Bat Lyssavirus Infection 2308563
Abstract Introduction Australian Bat Lyssavirus (ABLV), a neurotropic relative of Rabies virus, causes fatal encephalitis. Effective treatment for symptomatic infections remains elusive. Our prior work showed that an anti-ABLV monoclonal antibody (mAb) F11 or A6 can prevent disease even after the establishment of central nervous system (CNS) infection. Survival requires CD4 T cell-dependent immunity. mAb therapy alters CNS leukocyte composition, suggesting mAb-driven immune restructuring. The precise mechanism of protection is unclear; specifically, whether mAb protection results from enhancing overall immunity or from moderating inflammation. We hypothesized that F11 or A6 therapy promotes survival by recalibrating, rather than enhancing, CNS immune activation. Methods Mice were infected with a lethal dose of ABLV in the rear footpad. On day 5 post-infection, mice received mAb (F11 or A6), an isotype control mAb, or mock treatment. Brain tissue was analyzed on day 13 or 14 post-infection using multiplex cytokine assays, bulk RNA sequencing, and flow cytometry to compare immune signatures across treatment groups. Results Mock-treated mice showed elevated levels of inflammatory cytokines, type I/II interferons, and chemokines associated with myeloid, NK, and T cell recruitment, with transcriptomic enrichment for neuroinflammation and interferon/STAT pathways. In contrast, F11 or A6 therapy produced distinct transcriptional signatures marked by coordinated downregulation of inflammatory pathways and reduced cytokine and chemokine expression. Despite reduced inflammation, treated mice displayed increased numbers of CD11b+MHCII+ myeloid cells and a higher CD4:CD8 T cell ratio. Conclusion Therapy with F11 or A6 dampens interferon-driven inflammation while promoting balanced immunity in the CNS. Survival correlates with reduced pro-inflammatory cytokine expression and altered phenotypes of infiltrating immune cells, suggesting that excessive inflammation is detrimental to survival during ABLV infection. Funding Source US Department of Defense Topic Categories Vaccines and Immunotherapy (VAC)
Characterization of myelin lipid changes across disease stages in experimental autoimmune encephalomyelitis 2302268
Abstract Introduction Multiple sclerosis (MS) is an autoimmune demyelinating disease of the central nervous system. The myelin sheath of the CNS is a highly specialized, lipid-rich membrane that acts as the primary target of autoimmunity in MS, leading to demyelination, axonal injury, and consequent neuronal dysfunction. While the immunopathogenesis of MS has been extensively studied, particularly autoreactive lymphocytes, cytokine networks, and myelin sheath targeted immune responses, the contribution of lipid metabolism to disease progression mechanisms remains relatively underexplored. Here, we investigated whether neuroinflammation promoted alterations in the myelin lipidome during the initial attack of disease and during disease relapses that could contribute to MS progression. Methods We used mass spectrometry imaging (MSI), thiobarbituric acid reactant (TBARS) assays, and integrated immunohistochemistry to characterize lipid profiles, peroxidation, and spatial distribution in relapsing-remitting experimental autoimmune encephalomyelitis (EAE) models across disease stages. Results We identified significant lipid peroxidation during the acute phase of disease, with levels remaining elevated throughout remission, spatial heterogeneity of myelin sheath sulfatide distribution in the central nervous system, and disease-specific lipid modifications corresponding to inflammatory infiltrates, supporting the pathogenic role of oxidized lipids in MS. Conclusion These findings provide novel insights into the dynamics of the CNS lipidome during neuroinflammation and suggest that myelin lipid alterations could contribute to MS progression through mechanisms independent of neuroinflammation. Funding Source R21 NS137101-01 Topic Categories Neuroimmunology (NEUR)
Impact of Epstein-Barr Virus LMP2A on Toll-Like Receptor 9 (TLR9) expression and function 2257860
Abstract Introduction Toll-like receptor 9 (TLR9) in an important mechanism by which both innate and adaptive immune cells sense unmethylated DNA of bacteria and viruses. Epstein-Barr virus (EBV) is a double-stranded DNA enveloped virus that infects both epithelial cells and B cells. Since B cells express TLR9, we sought to determine if the latency protein, LMP2A, from EBV would impact TLR9 expression and/or function. Methods BJAB B cell lymphoma cells stably transfected with either vector, wildtype LMP2A or forms of LMP2A that harbor mutations in the N-terminal signaling tail were analyzed for TLR9 levels by flow cytometry. Additionally, these cells were stimulated with the TLR9 agonist, agatolimod (ODN2006), and the production of IL-6 was assessed by ELISA. Results The data indicate that LMP2A does not consistently alter TLR9 levels, but does enhance TLR9-induced cytokines, such as IL-6. The mechanism by which LMP2A confers this enhancement is currently under investigation. Conclusion TLR9 is an important innate sensing mechanism to identify the presence of pathogen’s DNA intracellularly. The ability of EBV to modulate signaling through these receptors may provide insight into mechanisms by which latency proteins regulate latency and B cell activation. Funding Source n/a Topic Categories Viral Immunology (VIR)
CD2 x IL2RG bispecific antibodies synergistically modulate lymphocyte activation, resulting in suppression of inflammation and prevention of GVHD 2258985
Abstract Introduction The common γ chain (γc, IL2RG) is a shared receptor subunit for IL-2, IL-4, IL-7, IL-9, IL-15, and IL-21 that regulates lymphocyte survival and activation. Mutations in IL2RG cause X-linked severe combined immunodeficiency (XSCID), underscoring its essential role in immune homeostasis. Dysregulated lymphocyte activation drives autoimmune and inflammatory diseases, yet current therapies such as corticosteroids and JAK inhibitors have limited efficacy in many patients and are often poorly tolerated. We developed CD2 x IL2RG bispecific antibodies that block IL2RG-mediated γc cytokine signaling and CD2-dependent co-stimulation, providing a dual-mechanism approach that combines direct inhibition of both pathways with CD2-targeted selectivity for IL2RG blockade on CD2+ lymphocytes (T and NK cells). Methods Bispecific CD2 x IL2RG antibodies were engineered with blocking arms for IL2RG and CD2 to inhibit γc cytokine and co-stimulatory pathways selectively on T and NK cells. These molecules were evaluated for inhibition of γc signaling, suppression of T-cell cytotoxicity and cytokine release in vitro, and mitigation of GVHD in vivo. Results By combining γc- and CD2-blocking arms, CD2 x IL2RG antibodies exhibited up to a three-log increase in potency for inhibiting γc-mediated signaling compared with monovalent or bivalent anti-IL2RG antibodies. The bispecifics synergistically suppressed T-cell—mediated lysis and reduced IFN-γ, and IL-2 production. In a human PBMC-engrafted NSG GVHD model, treatment initiated one week after PBMC transfer and continued weekly or biweekly for five weeks prevented mortality and weight loss, maintaining durable survival until study end. Conclusion Dual targeting of IL2RG and CD2 enables selective, avid blockade of cytokine and co-stimulatory signaling, leading to potent suppression of effector activity and GVHD. This approach may provide durable immune control while minimizing systemic immunosuppression from broad γc pathway inhibition. Funding Source n/a Topic Categories Therapeutic Approaches to Autoimmunity (THER)
Sunlight‐Driven Concurrent Synthesis of Jet Fuel Precursors and Hydrogen From Biomass‐Derived Furfuryl Alcohol Over Oxygen‐Doped Zn <sub>2</sub> In <sub>2</sub> S <sub>5</sub>
ABSTRACT Photoreforming of biomass‐derived furfuryl alcohol (FA) offers a green and economic route for the concurrent production of jet fuel precursor hydrofuroin (HF) and hydrogen. However, conventional FA photoreforming is generally hampered by low quantum yield and poor HF selectivity. Herein, an oxygen‐doping strategy to modify the Zn 2 In 2 S 5 (ZIS) photocatalyst is reported, which markedly enhances both quantum yield and HF selectivity. Oxygen dopants fulfill multiple critical roles: engineering shallow donor levels to facilitate photocarrier separation and extend their lifetime, accelerating the conversion of the furfural (FF) byproduct, and strengthening FA adsorption. Notably, the O‐doped Zn 2 In 2 S 5 (ZIS‐O) achieves 95% HF selectivity under simulated sunlight alongside high apparent quantum yields (AQY) for HF: 37.6% at 380 ± 20 nm and 12.4% at 420 ± 20 nm. Pilot‐scale tests under natural sunlight reveal a long‐term average HF selectivity of 90% with a turnover number (TON) of 1390. Theoretical investigations confirm that oxygen dopants thermodynamically and kinetically favor the FA‐to‐HF conversion. These findings provide valuable mechanistic insights for the rational design of high‐performance photocatalysts tailored for the selective conversion of biomass‐derived platform molecules into value‐added chemicals.
Metasurface‐Enabled Light‐Driven Liquid Crystal Elastomer Actuators
ABSTRACT Soft actuators based on liquid crystal elastomers (LCEs) offer programmable shape‐morphing capabilities for biomedical devices and microrobotics. However, conventional photothermal LCE actuators lack spatial selectivity in actuation, suffer from low photothermal conversion efficiency and poor wavelength versatility, which severely limit their performance, especially at the challenging 980 nm window. Here, we introduce a plasmonic metasurface strategy that enables spatially selective, high‐efficiency photothermal actuation. By patterning designed gold nanostructures onto LCEs, we create programmable “hot spots” that drive localized deformations under NIR illumination. This platform achieves unprecedented combined performance at 980 nm, including high photothermal conversion efficiency, low driving power, and competitive response times, while enabling patterned actuation. Moreover, we employ photoluminescence thermography to directly visualize and quantify thermal gradients generated across the LCE film, providing crucial insights into the spatiotemporal thermal dynamics. We also provide a comprehensive analysis from nanoscale plasmonic heating to macroscopic heat transfer. By integrating metasurface patterning with photoalignment, we demonstrate diverse actuation modes, from simple 2D bending to complex 3D morphing. This work establishes a new paradigm for high‐performance soft actuators, opening avenues for advanced applications in soft robotics, adaptive optics, and intelligent responsive systems.
Between Nodes and Niches: Tracing Tumor-Associated B Cell Trafficking and Effector Maturation in NSCLC 2260893
Abstract Introduction B cells support anti-tumor immunity via antibody production, antigen presentation, and immunomodulation. Somatic hypermutation (SHM) diversifies receptors to recognize tumor antigens. The origin of tumor-infiltrating B cells (TIBs) is unclear: they may arise from germinal centers (GCs) in tumor-draining lymph nodes (tdLNs) or tertiary lymphoid structures (TLSs) within tumors. Understanding B cell migration between these sites, maturation differences, and transcriptional programs driving differentiation and affinity maturation is key to defining their anti-tumor role and improving immunotherapy and tumor resection strategies. Methods B cells from paired tumors and tdLNs of NSCLC patients treated with neoadjuvant PD-L1 blockade were isolated by flow cytometry and analyzed with single-cell RNA and BCR sequencing (10x Genomics). We profiled subtypes, class switching, SHM levels, and clonal lineages using the Immcantation pipeline. Transcriptional profiles assessed activation, effector functions, and maturation states. Results Shared B cell lineages across tumors and tdLNs suggest dynamic migration. Many lineage trees link antibody-secreting tumor cells to GC B cells in tdLNs, indicating maturation there before tumor infiltration. Tumor GC B cells show distinct transcriptional signatures and SHM levels, implying unique maturation stages. Clonal overlap between tumor and tdLNs varies by donor, reflecting personalized B cell responses. Conclusion Our findings reveal patient-specific B cell clonal diversity and migration between GCs in tdLNs and TLSs, highlighting anti-tumor B cell complexity. Mapping these trajectories clarifies sites of B cell maturation and functional adaptation, informing strategies to enhance anti-tumor responses, refine tumor resection, and optimize immunotherapy. Future studies adding spatial resolution will clarify B cell niches and tumor microenvironment interactions and link migratory patterns to tumor antigen affinity. Funding Source NA Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Spatial and vascular determinants of tumor-infiltrating lymphocyte expansion in glioblastoma 2260218
Abstract Introduction Adoptive transfer of tumor-infiltrating lymphocytes (TILs) has shown promise in solid tumors but remains limited in glioblastoma (GBM), where immune dysfunction and spatial exclusion constrain TIL outgrowth. We profiled GBM tumors that did or did not yield TIL cultures to identify microenvironmental features predictive of successful expansion. Methods TIL expansion was attempted in 40 tumors; 24 (60%) achieved successful TIL expansion following optimized rapid expansion protocol (REP) culture. Expanded TILs were profiled by spectral flow cytometry and TCR sequencing. A subset of TIL+ (n = 5) and TIL⁻ (n = 6) tumors underwent single-cell RNA sequencing, CODEX multiplex proteomics, and Xenium spatial transcriptomics to characterize immune states, distict niches, receptor—ligand interactions, and vascular topology. Results Expanded TILs proliferated 100—1000-fold from pre-REP to REP, were CD4+-enriched, and dominated by effector-memory phenotypes in both CD4+ and CD8+ compartments with minimal Tregs or naïve subsets. Overall T-cell infiltration was low (&lt; 3% of total cells), yet TIL+ tumors contained more stem-like TCF7+ CD8+ cells and lower HAVCR2 and TOX expression. Spatial analysis revealed 2.3-fold higher CD31+ endothelial density and shorter T cell—endothelium distances (p &lt; 0.05) in TIL+ tumors, suggesting greater vascular accessibility. TIL⁻ tumors showed 1.6-fold stronger inhibitory TAM—microglia signaling, whereas TIL+ regions demonstrated enriched T cell—endothelial and TAM—macrophage interactions, indicating more permissive immune niches. Conclusion These findings identify vascular architecture and immune topology as key determinants of TIL expansion potential. Spatial-immune profiling may guide patient selection and inform strategies to recondition the GBM microenvironment for improved TIL and combination immunotherapies. Funding Source n/a Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Targeting Myeloid-Derived Suppressor Cell-Mediated Immunotherapy Resistance in Pediatric Brain Cancer 2260592
Abstract Introduction Despite the decrease in childhood cancer mortality, a cure for certain tumors remains elusive. The survival rates for high-grade medulloblastoma are disappointing, pointing to the immediate need for creative therapeutic strategies. In the tumor microenvironment (TME), suppressor cells inhibit the immune system, which includes myeloid-derived suppressor cells- a heterogeneous population of immature myeloid cells that play a role in cancer progression and metastasis. Two subsets of these cells, granulocytic MDSCs (G-MDSCs) and monocytic MDSCs (M-MDSCs), promote this immunosuppressive milieu by inhibiting anti-tumor T cell activity. Methods Monocytes were co-cultured with HTB-186 cells in a transwell system to generate tumor-educated MDSCs. CFSE-labeled T cells were co-cultured with MDSCs and stimulated with anti-CD3/CD28 beads. Proliferation was analyzed by flow cytometry (FACSCanto, FlowJo v10). Data were analyzed using GraphPad Prism 8.0 (t-test, mean ± SEM). Results Our study revealed that M-MDSCs serve as a biomarker for high-grade, metastatic pediatric cancers, including medulloblastoma. Our findings suggest that high concentrations of the soluble factors GM-CSF and IL-6, which are produced by HTB-186 medulloblastoma cells, lead to the induction of medulloblastoma M-MDSCs. M-MDSCs from medulloblastomas are highly immunosuppressive and exert their suppressive function by inducing reactive oxygen species (ROS). Our data indicate that GM-CSF from medulloblastoma cells induces ROS production in M-MDSCs via the coordinated activation of p47phox, a critical subunit of the NOX2 NAPDH oxidase, by STAT3 and NF-κB, resulting in inhibition of T cells leading to immune suppression in the TME. Conclusion In conclusion, our study identifies M-MDSCs as key drivers of immune suppression in high-grade pediatric medulloblastoma. These findings highlight M-MDSCs as promising therapeutic targets to overcome immune suppression and improve outcomes in pediatric brain tumors. Funding Source PON2 728 2300001864 and PON2728 2400001585 to KY Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Gut Microbial Translocation may drive NK cell Reprogramming in Adolescents Living with Perinatally Acquired HIV 2308891
Abstract Introduction Natural killer (NK) cell dysregulation contributes to chronic inflammation in people living with HIV, yet underlying mechanisms remain unclear. Gut microbial translocation (GMT), driven by impaired gut barrier integrity, is a source of systemic immune activation in HIV. We explored whether GMT influences NK cell phenotype and function in adolescents with perinatally acquired HIV (PHIV) on long-term antiretroviral therapy. Methods PHIV (n = 10) were compared to age-matched healthy controls (HC, n = 11). High dimensional flow cytometry was employed for immune profiling, including stimulation with HIV gag peptide pool. Plasma biomarkers were measured by ELISA. UMAP and FlowSOM were used for Phenotypic clustering. Data were analyzed using Mann-Whitney U test and Pearson correlation coefficient. Results Six of 15 NK clusters expressing TLR2, TLR4 and HLA-DR were upregulated in PHIV; two KLRG1+ clusters were downregulated (p &lt; 0.05). Conventional gating revealed increased TLR2 in CD16+CD56- NK subset with elevated HLA-DR (p &lt; 0.05) alongside NKG2D in CD16+CD56bright subset (p &lt; 0.005), indicating persistent activation. KLRG1+ effector memory-like NK cells were downregulated (p &lt; 0.05) in three of five NK subsets. Upon stimulation, NK cells from PHIV secrete more TNFα (p &lt; 0.05), while KLRG1+ NK cells produced less Granzyme B (p &lt; 0.005). This reflects a pro-inflammatory yet less cytotoxic phenotype. Plasma intestinal fatty acid binding protein, a marker of gut integrity, was elevated in PHIV (p &lt; 0.05) and positively correlated with TLR2+/TLR4+/HLA-DR+ NK subsets (R &gt; 0.5) and inversely correlated with KLRG1+ NK subsets (R &lt; -0.5). Conclusion Our findings suggest that GMT may promote NK cell activation via TLR2/TLR4 signaling, driving a pro-inflammatory yet less cytotoxic phenotype. This may contribute to the development of HIV-associated comorbidities. Strategies to restore gut barrier integrity or/and modulation of NK cells could mitigate chronic inflammation and improve long-term outcomes in PHIV. Funding Source NIH 1 U01 AI168630-01 Topic Categories Viral Immunology (VIR)
Exocyst complex regulates fungal-mediated IL-33 release and facilitates type 2 immune response 2304790
Abstract Introduction Recent studies overwhelmingly show that the fungal microbiome (mycobiome) is pervasive in tumors and plays a key role in reprogramming the tumor microenvironment. However, the molecular mechanism of fungal-mediated tumorigenesis remains unknown. Our recent study on pancreatic ductal adenocarcinoma (PDAC) shows that the intratumoral fungal mycobiome facilitates IL33 release from PDAC cells, thereby promoting PDAC tumorigenesis. However, the mechanism by which fungal infection mediates IL-33 release from PDAC cells remains unknown. Methods We used proteomic profiling, immunological techniques, orthotopic and genetically modified animal models, histology of human PDAC tissue microarray, and comprehensive molecular biology methods to identify the IL-33 secretion machinery and its role in PDAC progression. Results A high-throughput integrated proteomic profiling and protein-protein interaction study identified an Exocyst protein complex interacting with IL-33 in PDAC cells. The Exocyst complex consists of eight subunit proteins that tether the secretory vesicles to the plasma membrane and are implicated in cellular exocytosis. Further screening identified EXOC6B, a component of the Exocyst protein complex, which modulates IL-33 release via the exocytosis pathway from the PDAC cells. Genetic depletion of EXOC6B blocked IL-33 release and significantly abrogated PDAC tumor progression by suppressing type 2 immune response. Mechanistically, fungal mycobiome activates a non-canonical exocytosis pathway, thereby driving IL-33 nucleocytoplasmic transport and secretion from PDAC cells. Conclusion This study provides insights into a previously unrecognized mechanism and pathway of IL-33 release. Moreover, our findings bear important implications for identifying new therapeutic targets in various pathophysiological conditions, such as allergic airway disease and cancer, wherein fungal infections are known to increase IL-33 release and exacerbate the disease process. Funding Source Roswell Park Alliance foundation Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Influence of the absence of TRPM5 in CD4 T cells during the inflammatory response in autoimmune colitis induced by adoptive cell transfer 2258085
Abstract Introduction Inflammatory Bowel Disease (IBD) is a chronic inflammatory disorder of the gastrointestinal tract, with an increased incidence in several countries. IBD is developed with the help of different leukocyte types, among which are CD4 T cells. It is known that the activation and differentiation of CD4 T cells is dependent on calcium. Since the transient receptor potential melastatin 5 channel (TRPM5) controls the concentration of intracellular calcium, we believe that this may influence the polarization and activation of CD4 T cells. Therefore, our hypothesis is that the selective absence of TRPM5 in CD4 T cells would result in greater susceptibility to intestinal inflammation, due to changes in their activation and differentiation. Methods For this, autoimmune colitis was induced by adoptive transfer of CD4+CD45RB+ T cells isolated from spleen and lymph nodes of WT or TRPM5 KO mice into Rag1 KO mice recipients (5 × 10⁵ cells i.p.). Mice were monitored for up to 7 weeks. Clinical, immunological and histological parameters were evaluated, and subjected to rigorous statistical analysis. Results Our results indicate that the absence of TRPM5 in CD4 T cells partially attenuated the severity of adoptive transfer—induced colitis, delaying the onset of clinical signs of the disease. Immunophenotyping of the colonic lamina propria revealed that mice receiving mice CD4+CD45RB+ T cells from TRPM5 KO mice showed a trend toward reduced CD69+CD4+ T cells, along with increased memory and PD-1+CD4+ T cells. A similar reduction in CD69+CD4+ T cells was also detected in the intestinal epithelium. These changes correlated with milder clinical symptoms, suggesting that TRPM5 contributes to CD4 T cell activation and proinflammatory responses during chronic colitis. Conclusion Therefore, TRPM5 plays an important role in CD4 T cells, and its absence may influence the activation and differentiation profile of these cells, prominently in the intestine, impacting the inflammatory response in colitis. Funding Source CAPES (Coordenação de Aperfeiçoamento de Pessoal de Nível Superior); FAPESP (Fundação de Amparo à Pesquisa do Estado de São Paulo) Topic Categories Mucosal and Regional Immunology (MUC)
Age and Activity-Dependent Differences in Intraocular Immune Profile of Chronic Autoimmune Uveitis 2259109
Abstract Introduction Autoimmune uveitis is a chronic inflammatory disorder of the eye. Uveitis relapse occurs in 85% of patients who discontinue medications, necessitating lifelong immunosuppression, yet the pathogenesis of chronic, relapsing eye inflammation is poorly understood. Methods We obtained intraocular aqueous humor (AQH) samples from patients with clinically inactive uveitis (n = 29) or non-inflammatory controls (n = 16) undergoing eye surgery. Olink proteomic profiling and 10X single-cell RNA sequencing were performed on each sample. Results 41% of all uveitis patients displayed subclinical elevation of inflammatory cytokines within their eyes. Even more striking, &gt;85% of children had subclinical immune activity. Single-cell RNA sequencing revealed distinct immune cell populations associated with patient age and clinical disease activity. Namely, we found a higher prevalence of monocytes in adult uveitis, with none detected in pediatric samples. This was supported by proteomic data as well, which showed an association with adult uveitis and myeloid-related inflammatory cytokines. There was no correlation between monocytes and disease activity. In contrast, subclinical immune activity was associated with increased CD8+ T cells, specifically resident memory T cells (TRM), and decreased CD4+ T cell populations. Proteomic analysis confirmed a direct correlation between IL-15, which supports CD8+ T cells and TRM survival, and subclinical disease activity. Conclusion Our results provide further insights into chronic autoimmune uveitis, highlighting distinctions between pediatric and adult disease and activity-dependent immunity in the ocular microenvironment. Funding Source Rheumatology Research Foundation and Joint Biology Consortium microgrant off parent grant NIH/NIAMS P30AR070253 Topic Categories Immune Mechanisms of Human Disease (HUM)
Synergistically Enhancing Light Harvesting and Mechanical Flexibility for Ultra‐Flexible Organic Biosensors
ABSTRACT Ultra‐flexible organic optoelectronic biosensors, as key components of next‐generation wearable electronics, require devices that sufficiently harvest light, tolerate oblique illumination, and maintain mechanical compliance under bending or stretching. Herein, we report a solvent‐vapor spin‐coating (SVS) strategy combining chloroform:methanol mixed solutions, which simultaneously induces a shallow island‐like surface modulation and optimizes molecular packing. The former improves light harvesting and reduces angular sensitivity, whereas the latter contributes to improved optoelectronic performance; together, they are also associated with enhanced mechanical flexibility. Consequently, organic photovoltaics (OPVs) based on this strategy achieve a power conversion efficiency (PCE) of 20.28% on rigid substrates; more importantly, ultra‐flexible devices exhibit a record PCE of 19.03%, accompanied by improved mechanical robustness and reduced angular sensitivity. For organic photodetectors (OPDs), the enhanced light‐harvesting translates to a high specific detectivity ( D *) exceeding 10 13 Jones across the 320–920 nm range, and a response time of < 10 µs. Finally, for the first time, we demonstrate a top‐illumination/top‐emission ultra‐flexible photoplethysmography (PPG) sensor by integrating an OPV module, an organic light‐emitting diode (OLED), and an OPD, which accurately records on‐skin pulse signals. This study provides a promising route to co‐optimize the power output, detection sensitivity, and mechanical ductility of organic optoelectronics for practical applications.
The Ubiquitin Ligase Marchf1 Controls cDC Development to Sustain CD8+ T Cell Antitumor Immunity and Response Checkpoint Blockade 2259196
Abstract Introduction Conventional dendritic cells (cDCs) are essential for sustaining CD8+ T cell-mediated tumor immunity, yet the developmental mechanisms controlling their homeostasis remain unclear. We identify the E3 ubiquitin ligase Marchf1 as a critical regulator of cDC development with direct implications for tumor immune surveillance. Methods Using mice with DC-specific Marchf1 deletion (Marchf1ΔDC) or controls (Marchf1fl/fl) in murine melanoma and colon carcinoma models, we integrated scRNA-seq, flow cytometry, and immunofluorescence analyses of the TME, draining lymph nodes and bone marrow. We further analyzed human MARCHF1 expression in PBMCs from healthy individuals or melanoma patients, and analyzed the Cancer Genome Atlas (TCGA) melanoma cohort for correlations between MARCHF1 expression and clinical outcomes. Results Marchf1 deficiency led to accelerated tumor growth despite increased MHCII and CD86 expression on the DCs. The TME was characterized by a reduced immune infiltration, with selective loss of intratumoral cDC1s and impaired CD8+ T cell proliferation and cytokine production. Mechanistically, Marchf1 ablation induced premature MHCII upregulation in the bone marrow pre-cDCs, activating an apoptotic checkpoint that impaired cDC differentiation, confirmed by a competitive bone marrow chimeras. Remaining cDC1s retained cross-presentation, indicating numerical rather than functional insufficiency. Systemic Flt3L administration restored pre-cDC and cDC1 homeostasis, rescuing CD8+ T cell—mediated tumor control. In human melanoma, high MARCHF1 expression correlated with improved survival and checkpoint blockade responses, while circulating cDC1s from patients displayed selective MARCHF1 downregulation. Consistently, Marchf1ΔDC mice failed to respond to anti—PD-1/CTLA-4 therapy. Conclusion These findings identify Marchf1 as a developmental checkpoint linking cDC ontogeny to antitumor immunity and highlight MARCHF1 as a biomarker and therapeutic target to enhance DC-mediated cancer immunotherapy. Funding Source n/a. Topic Categories Classical and Non-Classical Antigen Presenting Cells (APC)
Assessment of Transcription Factors and Cytokines of CD4+ T Cells in beta2-Adrenergic Receptor Deficient Stressed Mice during Chlamydia Infection 2260245
Abstract Introduction Genital infection by Chlamydia trachomatis is the most common bacterial sexually transmitted disease worldwide. However, the relationship between stress, chlamydia genital disease, and its influence on the host immune response remains unknown. It is reported that the beta2-adrenergic receptor (β2-AR) expressed on immune cells is the target of the stress hormone, norepinephrine, during stress. Previously, we demonstrated that cold-induced stress (CIS) enhances the severity of chlamydia genital infection in mice by downregulating T-bet gene expression and interferon-gamma (IFN-γ) production, while upregulating GATA-3 gene expression and the secretion patterns of Thelper cytokines. However, the roles of Thelper17 and Tregulatory cells are unknown in our stress model. Methods This study aimed to determine the cytokine production by CD4+ T cells and the gene expression profiles of transcription factors, retinoic acid-related orphan receptor γt (RORγt) and FoxP3, using quantitative PCR and ELISA. Chronic stress was produced by immersing mice in cold water for 5 minutes daily over 21 days. Results Decreased gene expression of RORγt and FoxP3 was observed in β2-AR KO stressed mice compared to stressed mice (P &lt; 0.05). Furthermore, the secretion of IFN-γ, interleukin (IL)-4, IL-5, IL-10, IL-13, IL-17, and IL-23 was high in CD4+ cells of β2-AR knockout (KO) mice compared to that of stressed wildtype (P &lt; 0.05). Moreover, differential expressions of surface markers of naïve, effector, and memory cells of stress and non-stress mice were observed. Conclusion The obtained data show that a deficiency in β2-AR leads to the restoration of protective immune responses in stressed mice. The animal model study may give insight into how to control the effect of stress on the high prevalence of chlamydia in the low socioeconomic status population. Funding Source WV-INBRE P20GM103434 and NIH grant #R15AI172169 Topic Categories Microbial, Parasitic, and Fungal Immunology (MPF)
Post-operative IL-15 enhances the immunosuppressive effects of myeloid derived suppressor cells in sarcoma 2258326
Abstract Introduction Myeloid derived suppressor cells (MDSCs) inhibit NK and T cells leading to negative outcomes in cancer. Interleukin 15 (IL-15) is a stimulatory cytokine with anti-tumor activity that is limited by systemic toxicity. We have previously shown clinical benefit of IL-15 in dogs with metastatic osteosarcoma. We recently reported results of a phase II canine oncology trial of inhaled IL-15 after surgery for primary osteosarcoma demonstrating paradoxically worse outcomes with IL-15. Here, we sought to evaluate the effects of post-operative IL-15 on MDSC phenotype and function in dog and mouse sarcoma. Methods We performed single cell RNA sequencing of peripheral blood mononuclear cells from dogs with osteosarcoma treated with surgery and IL-15. Immune populations were identified and MDSC frequencies and gene expression were compared before and after surgery. C57BL/6 mice underwent excision of syngeneic fibrosarcoma flank tumors and received post-operative IL-15. Splenic MDSCs were co-cultured with naïve wild type splenocytes stimulated with anti-CD3:CD28. MDSC phenotype and T cell proliferation were evaluated by flow cytometry. Results MDSC frequencies and IL-15 receptor alpha (IL-15RA) expression were enriched in dogs after surgery. In mice, surgery increased MDSC frequencies, IL-15RA expression, and CD155 expression. Post-operative IL-15 increased PD-L1 and arginase expression on intratumoral MDSCs. Surgery increased MDSC suppression across multiple suppressor:responder ratios, and post-operative IL-15 further increased MDSC suppression. Conclusion In canine patients and mouse sarcoma models, surgery increased MDSC frequencies and IL-15RA expression, and post-operative IL-15 increased MDSC suppressive phenotype and function. These findings suggest surgery may limit the anti-tumor benefit of IL-15 in the post-operative setting. As surgery remains the cornerstone of treatment for sarcoma, targeting MDSCs in the setting of post-operative IL-15 immunotherapy is likely to have high translational impact. Funding Source NIH 1T32CA251007, UC Cancer Research Coordinating Committee Grant, National Leiomyosarcoma Foundation Grant Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)
Innate immune sensing of dietary alcohol ignites inflammation to drive alcohol-related disease 2309590
Abstract Introduction Alcohol is a leading cause of global morbidity and mortality, yet the innate immune sensors that detect alcohol-induced danger signals remain poorly defined. We identify ZBP1 as an innate immune sensor that integrates alcohol exposure with sterile- or infection-induced interferon signaling to trigger inflammatory cell death and promote the development of alcoholic liver disease. Methods Primary murine bone marrow—derived macrophages, human monocytic and liver cell lines, and genetically modified mice were treated with ethanol in combination with interferons. Cell death pathways were assessed using biochemical, molecular, and imaging-based assays. Z-RNA accumulation was quantified by immunofluorescence, dot blotting, and Z-RNA—specific RNA immunoprecipitation. Alcoholic liver disease was examined using murine ethanol-feeding models and human transcriptomic datasets. Results Alcohol synergized with sterile- or infection-induced interferon signaling to induce rapid inflammatory cell death, cytokine release, and liver injury. Combined ethanol and interferon stimulation triggered multiple regulated cell death pathways, including pyroptosis, apoptosis, and necroptosis. The innate immune sensor ZBP1 was essential for this response, as genetic ablation of the innate immune sensor ZBP1 protected cells and mice from cell death and alcoholic liver pathology. Mechanistically, interferon induced ZBP1 expression, whereas ethanol suppressed the RNA-editing protein ADAR1, promoting cytosolic Z-RNA accumulation. Alcohol and interferon converged on JNK signaling to induce endogenous Z-RNA, which activated ZBP1 via its Zα2 domain to drive inflammatory cell death. Conclusion These findings identify ZBP1 as a central innate immune sensor that integrates alcohol-induced stress and interferon signaling to drive Z-RNA—dependent inflammatory cell death and alcoholic liver disease, providing mechanistic insight and highlighting potential therapeutic targets for alcohol-related disease. Funding Source n/a Topic Categories Innate Immune Responses and Host Defense: Molecular Mechanisms (INM)