Browse Articles

Development of T-cell-based immunotherapy against chronic Hepatitis B infection 2309924

Abstract Introduction Even though T cells are known to be essential for Hepatitis B virus (HBV) clearance, there is currently no FDA-approved immunotherapy against chronic HBV infection, one of the major global causes of liver cancer, hepatocellular carcinoma (HCC). We aimed to characterize and test HBV-specific T cell receptors (TCRs) in a rhesus macaque (RMs) model of acute HBV clearance. Methods We analyzed HBV-specific T cell responses using non-parenchymal liver cells from RMs that cleared HBV by enzyme-linked immunospot and intracellular cytokine staining (ICS). We live-sorted activated HBV-specific CD8+ T cells from an animal with the strongest response using the TNF-α processing inhibitor (TAPI-0) and analyzed their transcriptional profile as well as T cell receptors (TCRs) via single-cell RNAseq. We used the retroviral method of expressing the α and β TCR chains from the top 8 most enriched TCRs in naive CD8+ T cells to confirm their specificity, map the minimal epitope, and identify MHC restriction. Results We identified three animals with strong CD8+ T cell responses against the same HBV surface 15-mer. The transcriptional profile showed that the responding CD8+ T cells primarily expressed αβ TCR chains and were highly activated. Using ICS, we confirmed the anti-viral activity of 3 TCRs. Peptide dilution experiments using T cells transduced with these TCRs showed that the immunogenic peptide is FLLTRILTI, which is known to be presented by HLA-A*02 in humans. We also determined that in RMs the restricting allele for this peptide is Mamu-B*031:01:01. Conclusion One of the main challenges in developing immunotherapy against chronic HBV infection is a lack of pre-clinical animal models. We show that RMs can be used to identify HBV-specific CD8+ T cell responses. Future directions include developing MHC-peptide tetramers to detect and elucidate HBV-specific T cell responses, identifying HBV-specific TCRs against other HBV proteins, and testing T-cell-based immunotherapy efficiency and safety. Funding Source n/a Topic Categories Vaccines and Immunotherapy (VAC)

Elevated TGF-β/IL-10 in platelet-monocyte complexes exacerbates atherosclerosis in people with HIV 2252272

Abstract Introduction People living with HIV (PLWH) are more susceptible to atherosclerosis (AS). Here, we examine the functional consequences of platelet-monocyte complexes (PMCs), which are elevated in PLWH and are hypothesized to contribute to HIV-associated comorbidities. Methods We performed single-cell RNA sequencing (scRNAseq) of monocytes isolated from age-, sex-, and cardiovascular risk score-matched participants categorized based on HIV and early AS status (n = 8 per group). Monocytes and platelets were cocultured ex vivo ± LPS (to simulate chronic inflammation) and analyzed at multiple time points as PMCs (24 hours) and PMC-derived macrophages (Mϕs, six days) via bulk RNA sequencing/RT-qPCR, ELISA, and flow cytometry. PMCs were cocultured with primary human endothelial cells (ECs) between three and six days of macrophage differentiation to assess endothelial to mesenchymal transition (EndMT) via RT-qPCR and ELISA. Results scRNAseq PMCs in PLWH and AS had increased TGF-β/IL-10 signaling compared to either PLWH or people living with AS (PLWAS) alone. Ex-vivo PMCs upregulated expression of genes involved in TGF-β signaling and IL-10 expression at both the monocyte and Mϕ developmental states. LPS stimulation increased IL-10 and TGF-β secretion and reduced TNF-α levels in culture supernatants. PMC-derived Mϕs increased expression of costimulatory molecules CD86 and CD80. When cultured with primary ECs, PMC-derived Mϕs reduced Pecam1 and Cdh5 and increased S100a4 expression. CCL2 and IL-6 were elevated in PMC-derived Mϕ-EC cocultures. LPS stimulation of cocultures increased expression of adhesion molecules Icam1 and Vcam. Conclusion Platelet interactions underscore monocyte-Mα¦ differentiation that promotes EndMT via TGF-β and IL-10, suggesting a pro-AS role of these factors in early disease. Enhanced TGF-β/IL-10 expression in PMCs of PLWH and AS suggests an increased relevance of this pathology in these individuals and defines a mechanistic link between aberrant platelet activation, PMC formation, and AS in PLWH. Funding Source T32AI118689, R01HL160229 Topic Categories Innate Immune Responses and Host Defense: Cellular Mechanisms (INC)

Heterogeneity in the Naïve CD8+ T Pool Shapes the Infection Response 2309970

Abstract Introduction Naïve CD8+ T cells are classically viewed as a uniform population that expands upon infection to protect the host against infection and cancer. However, growing evidence demonstrates that naïve CD8+ T cell heterogeneity influences the infection response. Recent single-cell studies revealed transcriptional heterogeneity among naïve CD8+ T cells, but whether this diversity reflects functionally distinct subsets possessing characteristic responses to infection remains unclear. We hypothesized that the naïve CD8+ T cell pool is composed of distinct subsets with underlying molecular programs that shape response kinetics and fate decisions during infection. Methods We combined single-cell multi-omic profiling (gene expression, surface protein, TCR), conventional and spectral flow cytometry, and in vivo adoptive transfer and infection models to identify, isolate, and functionally interrogate previously uncharacterized naïve CD8+ T cell subsets. Results scRNA-seq analysis identified multiple naïve CD8+ T cell subsets enriched for signatures of quiescence, effector priming, memory potential, interferon responsiveness, NF-κB signaling, regulatory function, and metabolic remodeling. Using subset-defining surface markers, we isolated these populations and assessed their in vivo responses to infection. Individual subsets showed differences in expansion, effector differentiation and memory formation, demonstrating that pre-existing regulatory states influence the infection response. Notably, further multi-omic profiling revealed that the relative abundance of naïve subsets varies with genetic background and age. Conclusion Our findings redefine the naïve CD8+ T cell pool as a heterogeneous compartment composed of functionally distinct subsets with intrinsic fate biases. Thus, variation in naïve pool composition represents a key mechanism that shapes CD8+ T cell immunity and may contribute to age- and genotype-dependent differences in susceptibility to infection and cancer. Funding Source NIH - NIAID Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)

Bioinspired Dynamic Remodeling of Excited‐State Pathways for High‐Performance Stimuli‐Responsive Materials

ABSTRACT Artificial stimuli‐responsive systems play a pivotal role in driving rapid advances at the frontiers of science and technology. However, existing systems are commonly constrained by “single fixed reaction pathways”, making it difficult to replicate the synergistic integration of low‐energy activation, multistable states, and excellent cycling reversibility of biological processes. Inspired by the precise regulation of water in biological photoreceptors, this study introduces a “water‐assisted excited‐state pathway remodeling” (WEPR) mechanism. By designing photoswitchable molecules that functionally mimic rhodopsin chromophores, we employ environmentally benign water as a dynamic regulatory factor to achieve intelligent switching of reaction pathways between high‐ and low‐barrier channels. Materials developed based on this mechanism exhibit exceptional bistability (>7 days), excellent cycling reversibility (>100 cycles), and remarkable long‐term stability (>2 years). These properties make it a compelling candidate for technologies such as dynamic information encryption and sustainable displays featuring on‐demand visualization and instant erasure. Furthermore, leveraging the multiscale similarities between our molecular switch system and biological visual pigments, the potential role of water in visual perception is discussed. This work not only provides a bioinspired paradigm for developing novel intelligent materials but also offers new insights into the fundamental mechanisms of color perception.

The effect of Roy adaptation model-based education on self-care and coping in patients with plaster casts: a single-blind randomized controlled trial

Antigen-independent formation of renal Trm-like CD8 T cells perpetuates hypertension 2259853

Abstract Introduction T cells critically contribute to hypertension, a condition affecting nearly half of American adults and driving cardiovascular mortality. However, the mechanisms by which these cells sustain hypertensive pathology remain unclear. We identified a persistent, memory-like CD8T cell population in the kidneys of hypertensive humans and hypothesize that kidney-resident memory CD8T cells (CD8Trm) establish a renal salt-retention memory that drives the chronic progression and recurrence of hypertension. Methods Kidneys from hypertensive patients and mice were analyzed. Trm-deficient mice (T cell-specific TGFβ receptor knockout; Tsp-TGFβR-KO) were used to assess the role of Trms in hypertension. Flow cytometry, single-cell metabolic profiling, and histopathology characterized renal immune populations. Mechanistic studies employed Rag2OT-I mice and ex vivo stimulation of CD8 T cells with ATP + TGFβ to test antigen-independent Trm formation and effects on distal nephron epithelial cells. Results Abundant renal CD8Trm were identified in both hypertensive patients and mice. In contrast, Tsp-TGFβR-KO mice showed blunted blood pressure in response to chronic hypertensive stimuli. In the salt-memory model, Tsp-TGFβR-KO mice developed initial hypertension but were protected from recurrence upon high-salt rechallenge, confirming Trm involvement in renal salt-retention memory. Renal CD8Trm formation occurred without antigen; Rag2OT-I mice without OVA exposure still generated P2X7+ CD8Trm, implicating ATP-driven CD8T-activation. Ex vivo ATP plus TGFβ treatment induced Trm formation with canonical markers, cytokine production, and distinct metabolic profiles. Co-culture with renal tubule cells enhanced sodium retention, linking immune activation to renal salt handling. Conclusion We identify a renal CD8Trm population that sustains hypertension through ATP+TGFβ-driven activation. This CD8Trm-salt axis anchors immune-hypertensive memory, offering a new immunological target to prevent hypertension recurrence. Funding Source NIH F31HL175932, AHA24PRE1198737, NIH T32 GM150536, NIH R01-HL146713, AHA 23TPA1076467 Topic Categories Immune Mechanisms of Human Disease (HUM)

Dietary Iron Overload Rewires Regulatory T Cells into IL-17—Mediated Proinflammatory Effectors Aggravating Yersinia enterocolitica Infection 2259908

Abstract Introduction Iron is indispensable for host physiology but, when in excess, fuels inflammatory and infectious diseases. The mechanisms by which nutritional iron overload perturbs adaptive immunity remain unclear. Here, we show that a high-iron diet (HID) profoundly reprograms FoxP3+ regulatory T (Treg) cells into IL-17—producing proinflammatory effectors that exacerbate Yersinia enterocolitica infection. Methods Mice were on a high iron diet for 4 weeks to generate iron overload followed by infection with YE. Control mice were fed control diet with normal iron. Changes in immune parametrs were evaluated by microscopy and flow cytometry analysis. Role of specific cell types were evaluated by adoptive transfer assays. Results HID-fed mice displayed a striking loss of reparative ST2+ Tregs and expansion of IL-17+ FoxP3+ Tregs with Th17-like transcriptional and metabolic signatures. Iron-conditioned Tregs exhibited enhanced mTORC1 activation, glycolytic flux, and RORγt expression, driving secretion of IL-17A and CXCL1 that promoted excessive neutrophil infiltration and mucosal damage. Neutralization of IL-17, inhibition of glycolysis (2-DG), or iron chelation (deferoxamine) restored ST2+ Tregs, reduced neutrophilia, and improved survival. Mechanistically, mTOR inhibition with rapamycin prevented iron-induced metabolic reprogramming, confirming an iron—mTOR—glycolysis axis that destabilizes Treg identity. Adoptive transfer experiments demonstrated that IL-17+ Tregs are pathogenic, whereas canonical ST2+ Tregs confer protection. Conclusion These findings uncover a previously unrecognized nutritional-immune checkpoint in which excess dietary iron converts reparative Tregs into proinflammatory effectors, amplifying neutrophil-driven immunopathology. Targeting this metabolic axis offers therapeutic promise for iron-associated inflammatory and infectious disorders. Funding Source RO1AI162670 Topic Categories Microbial, Parasitic, and Fungal Immunology (MPF)

Human Macrophages Modulate Neutrophil Transendothelial Migration in an Inflammation-on-a-Chip Model 2255264

Abstract Introduction Dysregulated neutrophil function contributes to chronic inflammatory diseases, including ANCA-associated vasculitis, yet it remains unclear how macrophages contribute to this dysfunction. We engineered a microfluidic Vasculature-on-a-Chip with primary human neutrophils, macrophages, and a model endothelial blood vessel to address this gap. Methods We adapted our “Inflammation-on-a-Chip” device to embed monocyte-derived macrophages in collagen ECM adjacent to an endothelial lumen. We confirmed macrophage viability and differentiation (M1: M-CSF, IFN-γ, LPS; M2: M-CSF, IL-4). We measured endothelial structure, adhesion markers (ICAM-1, VE-cadherin), and permeability with polarized macrophages. Neutrophil transendothelial migration (TEM) and migration in response to sterile inflammation (IL-8, LTB4) were quantified for 8 h in coculture with M1 or M2 macrophages. Results Macrophages had polarization-dependent effects on both the vessel and the neutrophil response. Without inflammation, M1significantly increased vessel permeability vs M2. With inflammation, M2 co-culture preserved barrier function vs M1. While inflammatory stimuli and co-culture with either M1 or M2 trended toward higher ICAM−1 expression, macrophages imposed a clear polarization-dependent TEM control. M1 resembled cell-free controls, while M2 significantly reduced transmigration. Single-cell tracking showed M2-restricted ECM migration. Conclusion Human macrophages regulate neutrophil access within a 3D vascular microenvironment. Pro-resolution cells strongly inhibit both TEM and post-TEM motility. Ongoing work will identify M2-derived mediators for targeted, pro-resolution therapies. Funding Source National Institutes of Health (NIH) Grant R35 GM146737A T32NIH Biophysics Training Program Topic Categories Cellular Adhesion, Migration, and Inflammation (CAM)

Therapeutic treatment with PD-1 agonist antibody prevents progressive autoantibody increase in murine autoimmune disease models 2258756

Abstract Introduction Autoimmune diseases are chronic inflammatory conditions with no complete cure. Repeated inflammatory episodes against self-antigens gradually accumulates tissue damage leading to morbidities; therefore, the treatment focuses on the management of disease to reduce the frequency and severity of inflammatory episodes. We have identified PD-1 agonist antibodies that can stimulate the immunosuppressive activity of PD-1. PD-1 agonist antibodies could suppress not only effector functions of activated T cells but also antibody development by targeting Tfh cells. The current study is designed to elucidate the efficacy of PD-1 agonists against autoimmune tissue damage by effector cells and autoantibody. Methods The therapeutic efficacy of PD-1 agonist antibody was tested using two autoimmune disease models: chronic graft-versus-host disease and MRL/lpr mice. Chronic graft-versus-host disease was induced in BDF1 mice by the transfer of CD8+-depleted splenocytes from human PD-1-knock-in B6 mice. Human PD-1-knock-in MRL/lpr mice received PD-1 agonist antibody treatment starting at 8 weeks old. Results Chronic graft-versus-host disease is a lupus model where single transfer of allogenic cells sufficiently induced anti-dsDNA antibody production. Treatment after the disease onset with PD-1 agonist antibody prevented further autoantibody increase by the additional transfer of B6 cells. The increase of autoantibody is far more progressive in MRL/lpr mice, but PD-1 agonist antibody starting blocked the increase of autoantibodies as well as splenomegaly for an extended time. Conclusion PD-1 agonists has a potential to be a novel therapeutic approach for the management of autoimmune diseases to keep them under control. Funding Source n/a Topic Categories Therapeutic Approaches to Autoimmunity (THER)

Investing the Role of IFI35 on Cell Viability Following Bacterial and Viral Infection by means of a Novel Human IFI35 Knock Out Respiratory Cell Line 2309817

Abstract Introduction Viral infections, such as Respiratory Syncytial Virus (RSV) or SARS-CoV-2, can trigger a widespread antiviral immune response that are both protective and pathogenic. Patients with respiratory viral infections are often susceptible to secondary bacterial infections, which may lead to further complications. One agent of particular concern according to the World Health Organization, is the gram-negative bacteria Acinetobacter baumannii, which has a proclivity for infecting ventilator-assisted patients. Interferon-induced protein 35 (IFI35) is upregulated during the antiviral interferon response and may modulate the host immune response, however, the specific impact and mechanisms by which IFI35 acts during viral and bacterial infections remains unclear. We sought to investigate the effect of IFI35 on lung epithelial cell viability under viral and bacterial infection conditions. Methods IFI35 knockout cells were generated via CRISPR-Cas9 in a human lung epithelial cell line, A549. We transfected wild-type and novel IFI35 knock out (IFI35KO) A549 cells with poly I:C, a mimic of viral RNA, to simulate a viral infection in the absence of viral proteins that may alter the natural host immune response. An MTT assay was then used to quantify cell viability. Additionally, control and IFI35KO cells were infected with A. baumannii at a MOI of 25 and host cell viability was assayed via AnnexinV/7-AAD at 8 and 24 hours post infection. Results IFI35KO was confirmed at the DNA and protein level in two independent cell lines. Following both viral and bacterial infection conditions, IFI35KO cells displayed increased cell viability relative to wild-type A549s. Conclusion These findings suggest that IFI35 plays a role in cell death during both viral and bacterial host immune responses. Further study of IFI35’s role cell dealth pathways may assist in developing new interventions to prevent complications of severe viral infections or reduce the risk of secondary infection, ultimately improving patient outcomes. Funding Source This work was supported in part by a COVID-19 Research Initiative grant from the Office of the Vice President for Research at the University of South Carolina, USCSOMG Scholars funding (KF, MM) a grant from the University of South Carolina Magellan Schola Topic Categories Microbial, Parasitic, and Fungal Immunology (MPF)

Targeting SERPINA3 in Gliomas Extends Survival and Reprograms the Immune Landscape 2260933

Abstract Introduction Malignant gliomas remain the most lethal cancers, with WHO grade IV having a 5-year survival rate below 7 percent. These tumors are characterized by high proliferation and an immunosuppressive tumor microenvironment (TME) that inhibits anti-tumor immunity. Using the TCGA and CGGA databases, we identified SERPINA3 as a critical, novel tumor vulnerability. Patients with high expression of SERPINA3 have worse survival compared to their low expression counterparts. SERPINA3 inhibits immune-activating proteases, and we hypothesize that it contributes to immune evasion and tumor progression in gliomas. Methods To test this, we screened three murine glioma cell lines for SERPINA3 expression using western blot analysis and found high expression in GL261. We used CRISPR to generate a SERPINA3 knockout (KO) GL261 line and intracranially implanted either wild-type (WT) or KO tumor lines into syngeneic mice. Results Mice bearing KO tumors showed significantly longer survival (MST 68 days) than WT controls (MST 28 days). Ongoing studies are profiling the TME immune landscape to determine how SERPINA3 affects immune cell infiltration and activation using flow cytometry. Conclusion These findings identify SERPINA3 as a novel regulator of glioma progression. Targeting SERPINA3 may enhance anti-tumor response and present a promising immunotherapeutic strategy for malignant gliomas. Funding Source N/A Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)

IL-17A+ γδ T Cells Are Compromised by ICU-Associated Sleep Fragmentation, Increasing Pneumonia Susceptibility 2310058

Abstract Introduction Critically ill patients experience profound sleep disruption and are at high risk for bacterial pneumonia. Using a murine model of ICU-associated sleep fragmentation (ICU-SF) combined with Streptococcus pneumoniae (Sp) infection, we previously showed that ICU-SF impairs bacterial clearance and worsens lung injury. We hypothesized that ICU-SF increases pneumonia susceptibility by selectively compromising antibacterial immunity. To test this, we used our combined ICU-SF and Sp model to characterize pulmonary immune populations involved in host defense. Methods Wildtype mice were subjected to ICU-SF using an orbital shaker and infected with Sp. Control groups included mice exposed to neither ICU-SF nor Sp, Sp alone, or ICU-SF alone. Lungs were harvested, enzymatically digested into single-cell suspensions, stained for surface and intracellular markers, and analyzed by spectral flow cytometry (Sony ID7000). Data were analyzed using FlowJo v10 and Prism 10 (Mann-Whitney or Kruskal-Wallis tests; p < 0.05). Results ICU-SF alone led to a marked reduction in lung γγT cells with no change in macrophage or neutrophil numbers. ICU-SF also selectively decreased lung IL-17A+ γγT cells among IL-17A-producing lymphocytes. Given the vital role of early IL-17A in bacterial clearance and prevention of pneumonia, we tested whether these findings persisted in the presence of Sp. In response to infection, we found a significant increase in pulmonary IL-17A+ γγT cells compared to uninfected controls. However, infected mice exposed to ICU-SF showed significantly decreased numbers of IL-17A+ γγT cells compared to non-ICU-SF infected controls. There was no difference in the number of IL-17A-producing CD4+ T cells, CD8+ T cells, group 3 innate lymphoid cells, or natural killer cells. Conclusion These findings identify a specific immune vulnerability induced by ICU-SF–IL-17A+ γδ T cell reduction–which may represent a key mechanism underlying increased susceptibility to hospital-acquired pneumonia in critically ill patients. Funding Source NIH T32 Training Grant: 5T32HL007534-43 Topic Categories Innate Immune Responses and Host Defense: Cellular Mechanisms (INC)

Living Inorganic Nanomaterials: Design, Preparation, and Biomedical Applications

ABSTRACT Living inorganic nanomaterials are defined as inorganic nanostructures that are integrated with living bacteria, cells, or biologically active cell‐derived components. This unique integration addresses key limitations of traditional inorganic nanomaterials in biomedical applications, including poor biocompatibility and limited ability to cross physiological barriers. This review outlines the developmental trajectory of bioactive inorganic nanomaterials, focusing on their evolution from passive carriers to life‐integrated therapeutic platforms. From biomacromolecule‐conjugated nanoparticles to nanozymes, culminating in living inorganic nanomaterials, we highlight the design of living cell‐ and bacteria‐conjugated inorganic nanomaterials for drug delivery, diagnosis, and therapy. We discuss the clinical translation and artificial intelligence‐aided design of bioactive inorganic nanomaterials. We also discuss the translational challenges and opportunities of living inorganic nanomaterials as a new generation of intelligent biomedical tools.

Reoxygenating the Tumor Microenvironment with Biomimetic Hemoglobin Nanoparticles Enhances Chemo, CAR-T, and Gene Therapies 2260007

Abstract Introduction Tumor hypoxia diminishes antitumor immunity by stabilizing HIF-1α, promoting M2-like macrophage polarization, and impairing cytotoxic T-cell activity. To reverse these effects, we developed hemoglobin-loaded biomimetic nanoparticles (Hb-BNPs) using a modified nanoprecipitation strategy. The nanoparticles encapsulate hemoglobin within a polycaprolactone matrix and are cloaked with RBC/A549 membranes to provide immune camouflage and sustained oxygen release. This study evaluates their ability to reoxygenate the tumor microenvironment and restores immune and therapeutic responses in non-small-cell lung carcinoma (NSCLC). Methods Hb-BNPs were synthesized via modified nanoprecipitation and membrane cloaking. Physicochemical features were assessed by DLS, TEM. Hypoxia was induced in A549 cells, followed by Hb-BNP treatment. qRT-PCR, paclitaxel IC50 assays, 3D spheroids, LunX CAR-T cytotoxicity, and KANK1-transfection studies evaluated immunologic and therapeutic responses (all n = 3, ANOVA/t). Results Under hypoxia, HIF1A, VEGF, BNIP3, ENO1, HK1, PGK1 were upregulated by 5—8-fold (p < 0.01). After Hb-BNP reoxygenation, these genes were downregulated by 4—6-fold (p < 0.001), and dissolved oxygen increased 5.3-fold (p < 0.001). Oxygen recovery improved the immune function as LunX CAR-T cytotoxicity increased 2.3-fold (p < 0.01), and KANK1-driven transgene expression increased 2.7-fold (p < 0.05). Co-treatment with paclitaxel reduced IC50 from 25,612 ng/mL to 781 ng/mL (p < 0.0001) and enhanced 3D spheroid core cell death 2.4-fold (p < 0.01). Hb-BNPs did not elevate IL-6, TNF-α, or IL-8 in THP-1 macrophages, confirming immune tolerance. Conclusion Hb-BNPs reoxygenate hypoxic tumors, suppress HIF-1α signaling, and restore cytotoxic immune function without inducing inflammation. By normalizing oxygen balance, these biomimetic nanoparticles enhance CAR-T activity, gene transfection, and chemotherapy response, offering a scalable, immune-tolerant platform to overcome hypoxia-driven resistance in solid tumors. Funding Source N/A Topic Categories Tumor Immunology: Checkpoints, Prevention, and Treatment (TIPT)

Antigen-driven MAIT cell activation restores skin Treg levels and attenuates lupus-like skin inflammation 2257534

Abstract Introduction MAIT cells are innate-like T cells with both repair and inflammatory capacity. Their role in inflammatory skin disease is unknown. Here, we report a protective role for MAIT cells in lupus skin disease. Methods Lupus and healthy PBMCs were analyzed by spectral cytometry (Dartmouth) and CyTOF (Brigham, 145 SLE, 40 healthy). B6, MAIT-deficient (Mr1-/-), and SLE (MRL/lpr or TLR7 agonist) mice (8-24wks, F) received topical or systemic MAIT antigen (5OPRU, 1mM). MAITs (5OPRU-loaded MR1 tetramer) and Treg were assessed by flow cytometry. ScRNA-seq of MAITs and Treg was conducted using 10X Genomics. Results MAITs are reduced and activated in circulation of SLE patients, compared to healthy donors. In MRL/lpr (MRL) skin, MAITs are reduced ∼10-fold compared to controls (B6, MpJ) and are CD69+PD1+. Topical antigen 5OPRU, but not vehicle or 5OPRU + anti-MR1 IgG (prevents MAIT-MR1 interaction) resolved severe skin lesions. MAIT expansion in non-lesional MRL skin stimulated Treg expansion and suppressed inflammatory and cytotoxic genes (Gzmb, Ifng, Il6, Tnf). In MRL mice with proteinuria, systemic 5OPRU prevented increases in serum inflammatory cytokines (CXCL1, GCSF, IL2, IFNg) and drove systemic expansion of Treg, including in skin. In the TLR7 SLE model, inflammatory (Ifng, Gzmb, Csf1) and interferon-stimulated (Ifi27l2ra, Isg20) genes were increased in Mr1-/- compared to B6 skin. Topical 5OPRU increased Treg:CD8 ratio in B6 but not Mr1-/- skin. ScRNA-seq of 5OPRU-treated skin MAITs revealed a tissue repair transcriptional profile (Furin, Tgfb1, Lgals3). 5OPRU-driven Treg expansion was MAIT-dependent (no expansion in Mr1-/- skin) and heterogeneous, revealing equal expansion of 7 distinct Treg subsets in B6 skin. Conclusion Antigen-driven MAIT activation supports Treg expansion and reduces morbidity in MRL/lpr mice. Conversely, MAIT deficiency heightens inflammation in a second SLE model, further supporting a protective role for MAITs in inflammatory skin disease, likely through a novel MAIT-Treg axis. Funding Source NIAMS R01AR080641 LFA Gina M. Finzi Memorial Fellowship Topic Categories Basic Autoimmunity (BA)

IgM antibodies and B cells are early responders during acute secondary viral infections and contribute to protection from severe disease 2255909

Abstract Introduction IgM antibodies comprise 5% of serum antibodies and are thought to be a primary (1°) and weak immune response. Yet, isolation of IgM monoclonal antibodies shows that IgM can potently neutralize viruses. Dengue viruses (DENV) cause >100 million infections per year. Secondary (2°) infections are a time of risk for severe disease because infection can be enhanced by pre-existing cross-reactive IgG antibodies. Methods Here we tested the role of polyclonal plasma IgM antibodies and IgM+ B cells in early 2° immunity to DENV infection. We conducted paired IgM- vs mock-depletions of acute and early convalescent (EC) sera from dengue patients to quantify the contribution of IgM to plasma neutralizing activity (Groups: 2° acute n = 32, 2° EC n = 19; 1° EC reference n = 18). Also, we used fluorescently labelled DENV to enumerate DENV-specific B cells by flow cytometry in 1° (n = 4) vs 2° (n = 4) dengue cases. Results Half of all 2° dengue cases demonstrated contribution of IgM to plasma neutralizing activity at both acute and EC phases; of these, IgM contributed, on average, to 41% of plasma neutralizing activity. In acute 2° DENV infection, a higher contribution of IgM to plasma neutralizing activity was associated with less severe disease, indicating a protective effect. IgM purified from 2° EC plasma demonstrated broad neutralizing activity against multiple DENV serotypes, unlike IgM from 1° EC, suggesting that 2° IgM may be recalled from memory. Notably, DENV-specific IgM+ B cells were significantly more expanded in acute 2° compared to 1° dengue immune responses. Conclusion IgM antibodies contribute to substantial plasma neutralizing activity in acute 2° DENV infection and confer broadly neutralizing specificities in early convalescence. In addition, IgM+ B cells expand rapidly in acute 2° dengue. Together, this indicates an underappreciated role for IgM antibodies and B cells as early responders in infection that may mitigate progression to severe disease. Funding Source HHMI Hanna H. Gray Fellowship to T.S. and P01 AI106695 to E.H. Topic Categories Viral Immunology (VIR)

Bridging the gap : CD8αß+ γδ T cells at the frontline of neonatal respiratory immunity 2259032

Abstract Introduction The observation that infants are more susceptible to some pathogens but resistant to others has long puzzled our understanding of early life immunity. This dichotomy can be explained by the absence or presence of specific immune components during this developmental period. Here we describe a population of CD8αß+ γδ T cells that are highly represented in the lungs of neonatal mice and assess their contribution to respiratory antiviral immunity. Methods Adult Foxn1α”/α” mutant mice, which harbor a persistently late fetal thymus, and C57BL/6 neonatal mice were used to assess respiratory perinatal T cells via intravascular labeling and flow cytometry. IFNγ production after in vitro αCD3/CD28 or inflammatory cytokine (IL-12/18) stimulation was determined. Mice were also infected with influenza virus and monitored for weight loss, viral load, and T cell activation by flow cytometry. Results Naïve Foxn1α”/α” mice harbored more CD8αß+ γδ T cells in their lung parenchyma compared to Foxn1α”/+ mice. These cells produce IFNγ and are particularly reactive to cytokine stimulation. While CD8αß+ γδ T cells account for 25% of the T cells in the lungs of C57BL/6 pups at post-natal day 0 (P0), their representation declines by P7. Interestingly, P3 pups express the activation marker CD69 48 h after influenza infection. Since Foxn1α”/α” mice are enriched for CD8αß+ γδ T cells and generate few conventional αß T cells, we infected them with influenza to determine whether the CD8αß+ γδT cells were sufficient for protection. Interestingly, Foxn1α”/α” mice were acutely resistance to influenza before eventually succumbing to a persistent infection. Conclusion CD8αß+ γδ T cells are a population of IFNγ producers enriched in the neonatal lung and activated early after influenza infection. We posit these are innate-like T cells unique to the perinatal period and poised to temporarily fill the functional niche of bona fide tissue resident memory T cells, which are lacking in the neonate. Funding Source NIH/NAID Grant# P01AI139449 Topic Categories Mucosal and Regional Immunology (MUC)

Regulatory T Cells Act as Tolerogenic Antigen Presenting Cells to Suppress Autoimmunity 2310009

Abstract Introduction Professional antigen presenting cells (APC) initiate most T cell responses, but human T cells express MHC-II post-activation and present antigen, which studies indicate is tolerogenic. Tregs are immunosuppressive and have a higher activation status than conventional T cells (Tcons) and may therefore be a more potent tolerogenic APC. We used human cell culture and transgenic animal models to investigate the capability of Tregs and Tcons to present antigen and the resulting effect on responder T cells and autoimmunity. Methods Antigen-pulsed human APC T cells were cultured with antigen-specific responder T cells, proliferation and activation measured by flow cytometry and responders profiled by sequencing. Parallel mouse assays were performed using OVA-presenting MHC-II+ T cells from Lck-CIITA mice with OT-II T cells. APC function was evaluated in vivo in experimental autoimmune encephalomyelitis (EAE) by transferring Lck-CIITA Tcons or Tregs presenting MOG35—55 before disease induction with the same peptide and by targeting the peptide to Tregs for presentation with a Treg-specific IL-2—MOG fusion protein. Results Human Tregs from blood exhibited a more APC-like phenotype than Tcons, expressing higher levels of MHC-II and costimulatory molecules. In culture, all T cell subsets drove antigen-specific proliferation, but the response induced by Tregs was the most robust. In mouse culture, CD8+ Tcons and Tregs generated a proliferative response and promoted Foxp3 expression in responder cells. In vivo, treatment with CD4+ Tcons or Tregs presenting MOG prevented EAE, but only Treg APCs converted MOG-specific T cells into Tregs. Targeting of MOG to Tregs during disease onset using a Treg-specific IL-2 also ameliorated EAE. Conclusion From human data, we conclude that Tregs are functional APCs and in mouse models we find that they drive a tolerogenic response. These findings identify Treg antigen presentation as a novel mechanism of immune suppression that may be harnessed for treatment of autoimmunity. Funding Source N/A Topic Categories Classical and Non-Classical Antigen Presenting Cells (APC)

Pharmacological activation of LANCL2 provides immunometabolic protection from disease and lung pathology in mouse models of asthma 2260965

Abstract Introduction Asthma is a chronic, complex, immunological disease that afflicts over 28 million people in the U.S. Current therapies are modestly successful but leave significant subpopulations of patients without adequate response. LANCL2 is a novel, immunoregulatory target for the treatment of asthma. Methods The therapeutic efficacy of novel LANCL2-targeting therapeutics was evaluated in the house dust mite (HDM) and ovalbumin (OVA) plus lipopolysaccharide (LPS) models of asthma. For HDM, mice were immunized with an HDM injection and then sensitized intranasally for 3 wk. For OVA+LPS, mice were immunized with OVA, exposed to OVA by aerosolization and then administered LPS intratracheally. Lung immunophenotype, gene expression profiles and histological lesions were evaluated. Expression of inflammatory mediators and key metabolic genes were assessed in human primary bronchial epithelial cells isolated from asthma patients (DHBE) upon pharmacological activation of LANCL2. Results In the HDM model, oral LANCL2 activation decreased lung histopathological lesions, proinflammatory gene expression (CD68, Ifng, Saa3), and the infiltration of neutrophils. In the OVA + LPS model, LANCL2 activation resulted in decreased goblet cell hyperplasia and infiltration of inflammatory immune subsets (IL4+ CD4+ T cells, IL5+ CD11b+ cells and neutrophils) while increasing the proportion of Treg cells in the lungs. In translational studies with DHBE cells, activation of LANCL2 decreased the expression of proinflammatory cytokines Il6 and Tnf, plus Ldha, the glycolytic enzyme that catalyzes the production of lactate. Conclusion Activation of LANCL2 by novel agonists ameliorates disease severity and immunopathology in mouse models of asthma. LANCL2-targeting oral, once-daily small molecule drugs represent innovative approaches for the treatment of neutrophilic asthma. Funding Source N/A Topic Categories Immediate Hypersensitivity, Asthma, and Allergic Responses (HYP)

ImmuneSpace in 2026: A Centralized Repository for Curated Human Immune-Profiling Data 2310296

Abstract Introduction ImmuneSpace (immunespace.org) is a freely accessible database that hosts curated human immune-profiling data from a wide range of studies. It was created as the central data repository of the Human Immunology Project Consortium (HIPC), a multi-center NIH-funded program to characterize the diverse states of the human immune system and its regulation, using consistently formatted data [PMID: 23648045]. The overarching goal is to investigate human immune perturbations using state-of-the-art systems-level profiling technologies and innovative methodologies, and to make these data available to the scientific community, accessible for both humans and machines. Methods ImmuneSpace hosts data related to immunological exposure, demographics, cytokine profiling, cytometry, and neutralizing antibody assays. It builds on the ImmPort data model, a long-term archive of research and clinical data for the NIH [PMID: 29485622], but implements additional standardization and normalization rules, following the HIPC Data Standards initiative [PMID: 22343568, 26861911, 31272390, 32283555]. Results ImmuneSpace is continuously updated with data and features. Each study undergoes enhanced curation to ensure consistent use of ontology-based terminology, enabling more efficient queries both within and across studies. Study components stored in different repositories, unparsed raw data, and computationally inaccessible elements are integrated through manual curation, such as study timelines and author-determined ‘immune signatures’. Recently, we have added the ‘Finder Feature’ which simplifies complex searches through a hierarchical, tree-based interface that allows users to visually browse, search with autocomplete, or select entire branches of categorized filters while providing definitions, synonyms, and ontology links. Conclusion The HIPC Project and the ImmuneSpace platform demonstrate the feasibility and benefits of a structured approach to representing human immunological studies to elucidate system-level phenomena. Funding Source U01 AI167892 Topic Categories Computational and Systems Immunology (COMP)