Browse Articles

Trehalose Enables Surfactant Protein D—Dependent Immune Modulation in Early Cryptococcus neoformans Infection 2309235

Abstract Introduction C. neoformans (C.n), a major fungal pathogen, infects the lungs and can disseminate to the brain leading to life threatening meningoencephalitis. Due to limited antifungals, there is a need to identify novel drug targets. Tps1 is required for the biosynthesis of Trehalose, a fungal-specific sugar that has been identified as a promising target for drug development. Mice infected with tps1-KO (tps1Δ) C.n exhibit increased survival, an immediate decrease in lung fungal burden, and clearance by day 7. Infection kinetics suggest innate or resident immunity factors drive this response. Surfactant protein (SP) D is a collectin endogenously expressed in the alveolar space, and its role in several pulmonary infections has been well characterized. Therefore, we hypothesized SP-D could contribute to tps1Δ clearance. Methods We infected C57BL/6 (WT) or SP-D -/- mice with either tps1Δ or H99 C.n Mice were sacrificed at 2 days post infection (dpi) and lung fungal burdens and immune cell populations were investigated by flow cytometry analysis. Results Tps1Δ infected mice had reduced fungal burdens compared to H99-infected mice, consistent with previous work. Interestingly, pulmonary fungal burdens were not affected by the loss of SP-D on 2dpi. Our results indicate that SP-D moderately decreased CD45+ immune cell recruitment to the lung in H99-infected mice whereas tps1Δ -infected mice saw no change. Additionally, alveolar macrophages from SP-D-/- H99-infected mice saw an increase in CD86 and CD206 expression compared to H99-infected WT mice, whereas tps1Δ -infected mice saw no difference in macrophage activation regardless of the presence or absence of SP-D. Conclusion These results support that WT C.n exploits SP-D for its benefit by promoting a non-protective immune response. This exploitation appears to require trehalose, as it is lost in tps1Δ infections, suggesting a novel effect linked to Tps1-targeting. Further molecular and cellular pathways underpinning these interactions are under investigation. Funding Source U.S. Department of Veterans Affairs grants (Merit Award I01BX000656 and Research Career Scientist Award IK6BX003615-01 to MO, and CDA-2 award IK2BX006208 to KG); Rackham Merit Fellowship Recipient Topic Categories Mucosal and Regional Immunology (MUC)

Distinct Effect of Type III Interferon on Trex1KO autoimmune model 2307705

Abstract Introduction The Tree-prime Repair Exonuclease 1 (Trex1) is a 3’→5’ exonuclease that degrades cytosolic nucleic acids. Mutations in Trex1 are associated with a broad spectrum of autoimmune diseases, such as AGS and SLE. Trex1 deficiency leads to accumulation of cytosolic self-DNA, which induces hyper-activation of the immune system including systemic inflammation, auto-antibody production, and tissue damage. Type I IFN responses in Trex1 deficiency models have been well-studied. However, the function of type III interferon is incompletely understood. Methods We generated Ifnlr1KO and cGASKO mice on the genetic background of Trex1KO mice. Results In our studies, we found that MNoV infection is prevented in Trex1-/- mice but not in Trex1-/-cGAS-/- and Trex1-/-Ifnlr1-/- mice. This indicates that the type III interferon signaling pathway activated in the intestine of Trex1-/- mice is dependent on cGAS-STING. In addition, Trex1-/-Ifnlr1-/- partially alleviates the upregulation of retroelements and the induction of robust interferon stimulated genes (ISG) in intestinal tissues from Trex1-/- mice. Furthermore, the production of auto-antibodies and subsequent tissue damage seen in Trex1-/- mice is alleviated in Trex1-/-Ifnlr1-/- mice. These ameliorations from the loss of Ifnlr1 rescues the survival of Trex1-/- mice. Conclusion We conclude that the accumulation of mislocated nucleic acids activates type III interferon signaling through cGAS-STING in Trex1-/- mice. The type III interferon signaling activation protects the mice from MNoV infection but contributes to hyperinflammation and auto-antibody production, which accelerates tissue damage and increases mortality in Trex1-/- mice. Since interferon lambda receptor’s expression is restricted to gut epithelial cells, it implies the local intestine plays a complex role in exacerbating systemic inflammation in the auto-immune disease model. Blockage of type III interferon signaling might be a therapeutic target for attenuating the symptoms of autoimmune Funding Source NIH Topic Categories Basic Autoimmunity (BA)

Cavity‐Defined van der Waals Plasmonic Nanolasers Enabled by Out‐Of‐Plane Dipole Coupling

ABSTRACT Plasmonic nanolasers enable deeply subwavelength coherent light sources but are often limited by rigid cavity designs and demanding gain conditions. Here, a cavity‐defined plasmonic nanolaser platform is realized through van der Waals integration of layered Indium Selenide (InSe) gain media with lithographically engineered Au cavities separated by an ultrathin hexagonal boron nitride (hBN) spacer. Owing to the direct bandgap and predominantly out‐of‐plane (OP) emission dipole of InSe, efficient coupling to surface plasmon polariton (SPP) modes is achieved. Clear near‐infrared (NIR) lasing characteristics, including a well‐defined threshold, pronounced linewidth narrowing, and a drastic reduction of carrier lifetime, are observed under optical pumping. A plasmonic lasing regime below the optical‐mode cutoff is identified, with lasing sustained for InSe thicknesses down to 79 nm, well beyond the limit of conventional photonic‐mode lasers. Spatially resolved spectroscopy and numerical simulations reveal that the lasing feedback is governed by cavity‐defined SPP modes. In addition, deterministic control of lasing behavior is achieved by engineering Au cavities with diverse geometries, establishing a versatile and general plasmonic nanolaser architecture.

Correction: Glyceraldehyde-3-phosphate dehydrogenase/1,3-bisphosphoglycerate-NADH as key determinants in controlling human retinal endothelial cellular functions: Insights from glycolytic screening

Physiology and reproductive responses of disease-free soybean plants to triazole, strobilurin, and carboxamide fungicides under greenhouse conditions

IgG Autoantibodies and Fc Signaling: A New Axis of Neuropathic Pain in Endometriosis 2265840

Abstract Introduction Endometriosis is a chronic inflammatory disease redefined as a neuroimmune pain syndrome. Pain arises from neuroimmune crosstalk, driving peripheral and central sensitization. Sensory nerves release SP and CGRP, amplifying inflammation and lesion growth. This abstract proposes that B cell—derived IgG autoantibodies trigger neuropathic pain through the FcγR axis, linking neurons, macrophages, and microglia–offering a new immunomodulatory target. Methods Therapeutic strategies target the B cell—FcγR axis to modulate neuroimmune pain in endometriosis. Approaches include B cell depletion with anti-CD20 antibodies, FcγR or BTK inhibition, and blockade of neuropeptides such as CGRP and SP. Additional options include TRPV1 antagonists and NGF/BDNF neutralization to reduce hyperinnervation, inflammation, and neuropathic pain. Results Immunomodulatory and neuro-targeted therapies show promise beyond hormonal treatments. Targeting the B cell—FcγR axis with anti-CD20 antibodies, BTK inhibitors, or FcγR antagonists may reduce cytokine release and IgG-mediated neuropathic pain. Blocking CGRP-RAMP1 signaling lessens hyperalgesia and lesion growth, while TRPV1, NGF, and BDNF inhibition can desensitize nerves and reduce inflammation. Conclusion Endometriosis involves intertwined neuroimmune and inflammatory pathways driving chronic pain. Neurogenic inflammation, hyperinnervation, and immune dysregulation sustain sensitization. B cell—derived IgG autoantibodies acting through the FcγR axis may trigger neuropathic pain and glial activation, linking peripheral inflammation to central sensitization and chronic pelvic pain. Funding Source n/a Topic Categories Neuroimmunology (NEUR)

FFAR2 and FFAR3 Modulate Cigarette Smoke-induced Emphysema through Distinct Inflammatory Pathways 2307232

Abstract Introduction Despite the lack of effective treatments for chronic obstructive pulmonary disease (COPD), increasing evidence from studies on the gut—lung axis demonstrates the role of short-chain fatty acids (SCFAs) in respiratory immune regulation. SCFAs exert anti-inflammatory effects in the lung by activating GPCRs (FFAR2 and FFAR3) and inhibiting histone deacetylase2 (HDAC2). However, the predominant mechanism responsible for COPD alleviation remains unclear. This study aims to evaluate the differential roles of FFAR2 and FFAR3 activation in modulating COPD progression. Methods Mice were exposed to cigarette smoke (CS) for emphysema model with/without propionate, 4-CMTB (selective FFAR2 agonist), or AR420626 (selective FFAR3 agonist). Results CS induced airspace enlargement and mechanical dysfunction, which were improved by agonist treatment. AR420626-treated group restored airway resistance and compliance, accompanied by broad suppression of inflammatory and remodeling genes by qPCR. In contrast, 4-CMTB-treated group improved tissue elastance and reduced neutrophil and monocyte infiltration in bronchoalveolar lavage fluid. Cytokine profiling showed receptor-specific modulation: FFAR3 activation decreased TNF-α and IL-18 while increasing IL-10, whereas FFAR2 reduced IL-6 and IL-17A. Proteomics showed suppression of Th17-associated proteins, with FFAR3 reducing IL-9/IL-17F and FFAR2 reducing IL-9/CXCL12. Flow cytometry showed that FFAR2 activation reduced Th1/Th17 populations and increased M2 macrophage polarization. Western blot and TUNEL assay confirmed that FFAR3 inhibited NLRP3-caspase signaling and alveolar apoptosis, whereas FFAR2 attenuated MAPK/NF-kB inflammatory pathways. Conclusion Selective activation of FFAR3 restored alveolar architecture and suppressed apoptotic signaling, whereas FFAR2 attenuated inflammatory cytokine production and immune cell infiltration. These findings reveal distinct receptor-specific SCFA pathways and highlight FFAR2 and FFAR3 as distinct therapeutic targets in COPD. Funding Source This study was supported by NRF grant (RS-2023-NR077159, RS-2023-00222687, RS-2022-NR067421) and NIH grant (No.2024ER080601) funded by Korean government. Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)

PD1-independent CD8 T cell exhaustion driven by the loss of UFL-1 2260778

Abstract Introduction Cytotoxic CD8 T cells play a crucial role in eliminating infections and cancer. However, chronic antigen exposure induces their development into exhausted T (TEX) cells. TEX cells exhibit increased inhibitory receptors or checkpoints expression, defective effector functions and thus unable to eliminate the virus-infected cells and cancers. Antibodies inhibiting signaling through checkpoints PD1 or CTLA4 (immune checkpoint blockade or ICB) induce differentiation of TEX cells into effectors that eliminate cancer and viral infected cells. PD1 blockade is the leading ICB against several tumor types. However, > 50% of the cancer patients either do not respond or get durable benefits from PD1 blockade. Therefore, there is an urgent need to understand the PD1-independent mechanism to improve immunotherapy against tumors resistant to PD1 blockade. Methods In order to address the role of UFL-1 in CD8 T cell exhaustion, we used LCMV acute and chronic infection models in control and CD4 specific UFL-1 knockout mice. Different subset of CD8 T cells and their differentiation status and functional response upon activation were analyzed using flow cytometry and ScRNA-seq. Results Acute exposure to antigens, is not known to cause T cell exhaustion. We found that UFL1-deficient CD8 T cells undergo exhaustion during acute viral infection. During chronic antigen presence, such as in tumor models and chronic viral infections, UFL1 deficiency worsens CD8 T cell exhaustion. Additionally, the PD1 blockade fails to improve the functionality of UFL1-deficient TEX cells. Consistent with this, UFL1 expression in tumor-infiltrating CD8 T cells is positively correlated with the progression-free survival of melanoma patients treated with PD1 blockade. Conclusion These findings suggest that exhaustion in UFL1-deficient CD8 T cells is PD1-independent. Therefore, understanding how UFL1 prevents T cell exhaustion would help discover new ways to improve immunotherapies, especially for cancers that do not respond to PD1 blockade. Funding Source NIH Funding - NIAID00054 Topic Categories Immune Response Regulation: Molecular Mechanisms (IRM)

A secondary Bile Acid regulates IgE+ germinal center B cell differentiation and IgE-mediated immune homeostasis 2308423

Abstract Introduction Immunoglobulin E (IgE) mediates rapid protective immune responses but also contributes to the development of allergic diseases. Thus, stringent regulation of IgE production is essential for maintaining immune homeostasis. Recent studies have revealed that IgE-mediated immune responses are dysregulated in the absence of the gut microbiome. Although secondary bile acids (SBAs) have been documented to directly regulate T cell functions, their role in IgE homeostasis remains unclear. We therefore hypothesized that specific SBAs regulate IgE production through B cell intrinsic manner. Methods We investigated the role of immune modulatory SBAs in IgE antibody production using in vitro B cell differentiation system and in vivo food allergy mouse model. To evaluate contribution of SBAs in IgE homeostasis, IgE germinal center (GC) B cell responses were analyzed by flow cytometry, and antigen-specific IgE antibody levels were measured using ELISA-based assays. In addition, molecular docking analyses and transcriptomic profiling were conducted to identify putative molecular targets of SBAs and underlying molecular mechanisms. Results We identified isoalloLCA as a negative regulator of IgE production through inhibition of sequential class switching from IgG1 to IgE. IsoalloLCA treatment significantly reduced the accumulation of IgE GC B cells and the generation of high-affinity antigen-specific IgE antibodies in a food allergy mouse model. We identified NR4A1 as a potential target of isoalloLCA in B cells. Mechanistically, we found that efficient sequential class switching from IgG1 to IgE required robust proliferation of IgG1 GC B cells, and isoalloLCA constrained this proliferative expansion specifically. Conclusion We identify that isoalloLCA as a key regulator of IgE homeostasis that constrains the emergence of IgE effector B cells from IgG1 GC B cells. Our study uncovers a previously unrecognized proliferation dependent checkpoint governing sequential class switch from IgG1 to IgE during GC reactions. Funding Source This study was supported by the Basic Science Research Program through the National Research Foundation of Korea (NRF) funded by the Ministry of Science and ICT (RS-2025-24523224), and by the Regional Innovation System & Education (RISE) program through t Topic Categories Immediate Hypersensitivity, Asthma, and Allergic Responses (HYP)

Development of an NS1-based dengue vaccine that protects via non-neutralizing antibodies and evades antibody-dependent enhancement 2249046

Abstract Introduction Dengue is a mosquito-borne virus that threatens one half of the global population. While primary infection is usually mild or asymptomatic, secondary infection with another of the four serotypes increases the risk for severe disease due to a phenomenon called antibody-dependent enhancement (ADE). Through ADE, non-neutralizing antibodies that target dengue structural proteins facilitate viral entry and replication and worsen disease severity and outcomes. Current dengue vaccines are live-attenuated and do not eliminate the possibility of ADE upon breakthrough infection. Novel vaccines that do not induce ADE are consequently needed. NS1 is a secreted non-structural protein that contributes to dengue pathogenesis by mediating vascular leakage. NS1 is a promising vaccine target because anti-NS1 antibodies are non-neutralizing, thereby evading ADE, and they have also been shown to protect mice from disease. Methods Here, we are developing a DENV2 NS1 mRNA-LNP vaccine that is safe, highly immunogenic, and effective in mice. Results This DENV2 NS1 mRNA-LNP vaccine induces non-neutralizing antibodies that protect against severe disease and mortality in a lethal challenge model. Conclusion In future directions, this work will investigate the mechanisms of non-neutralizing anti-NS1-based protection, including the involvement of affinity maturation and Fc effector functions. Together, we present a promising dengue vaccine strategy that evades ADE and protects against lethal disease. Funding Source Jordan Polster is supported by the HHMI Gilliam Fellows Program. David Martinez is supported by the HHMI Hanna Gray Fellows Program. Topic Categories Vaccines and Immunotherapy (VAC)

Novel chemokine-peptide-MHC II CAR-T cell treatment alleviates inflammation in chronic beryllium disease 2309489

Abstract Introduction In chronic beryllium disease (CBD), elevated levels of the inflammatory chemokines CCL3 and CCL4 in the lungs coincide with expanded populations of CD4+ T cells specific to beryllium (Be)-modified peptides derived from these chemokines. Here, we generated HLA-DP2 transgenic (Tg) CCL3-deficient mice (CCL3-/-) that also lack CCL4 to investigate their role in disease development. Methods Mice strains: HLA-DP2 Tg WT, FVB-N, CCL3, and TNF-α. Mice were intratracheally administered with BeO (100µg) on days 0,1, and 2 and euthanized at day 12 or boosted at days,14,15,18,19, and euthanized on day 21. BAL fluid (BALF) was obtained from the instillation of 1 ml PBS. BeO-exposed WT HLA-DP2 Tg (CD45.2) mice were injected intravenously with DR-a or CCL4-CAR- T cells on day 8. Flow cytometry, q-PCR, IFN-γ and IL-2 ELISPOT, Histology (H&E), Tetramer staining, RNA-sequencing, and CAR-T cell design and preparation. Results Be-exposed CCL3-/- mice maintained normal numbers of lung macrophages and dendritic cells (DCs) but exhibited significantly reduced total and HLA-DP2-CCL/Be tetramer-specific CD4+ T cells, IFN-γ-producing CD4+ T cells, and peribronchovascular aggregates, consistent with attenuated inflammation. CCL3 was predominantly expressed in macrophages and DCs, and bone marrow chimera studies confirmed that hematopoietic-derived DCs are the key regulators of CCL/Be-specific CD4+ T cell responses. RNA sequencing of lung-resident CCL4/Be tetramer-positive CD4+ T cells revealed a transcriptional profile enriched for inflammatory and cholesterol metabolism pathways, with elevated IFN-γ, TNF-α, and IL-17a gene expression. Moreover, Be-exposed HLA-DP2 Tg mice lacking TNF-α or treated with peptide-MHC II CAR-T cells targeting CCL4/Be-specific CD4+ T cells showed reduced T cell responses. Conclusion These findings demonstrate that CCL3 and CCL4 promote CCL/Be-specific CD4+ T cell responses and highlight peptide-MHC II CAR-T cells as a novel strategy for depleting self-peptide/Be-specific CD4+ T cells in CBD. Funding Source NIH R01 HL152756 Topic Categories Immune Mechanisms of Human Disease (HUM)

Development of a safe and effective nanoparticle-based vaccine against porcine epidemic diarrhea virus 2308286

Abstract Introduction Porcine epidemic diarrhea virus (PEDV), introduced into the U.S. in 2013, causes severe diarrhea and nearly 100% mortality in neonatal piglets, leading to an estimated annual economic loss near $1.8 billion. Current inactivated vaccines provide only partial protection, while live-attenuated vaccines pose safety concerns. To address this, we are developing a hybrid nanoparticle (hNP)-based spike protein vaccine with enhanced safety, immunogenicity, and protective efficacy. Methods The nanovaccine incorporates the C-terminal domain (CTD) of the PEDV USA/CO/2013/29619 spike protein. CTD is expressed in E. coli, purified, and conjugated to lipid-coated poly(lactic-co-glycolic acid) nanoparticles. The hNP platform enables incorporation of molecular adjuvants within both the polymer core and lipid layer. Nanoparticles are characterized for size, polydispersity, antigen loading, and endotoxin levels. The immunogenicity of the nanovaccines is first evaluated in mice. Results Recombinant CTD was expressed at a yield of 30 mg/L in bacterial culture and purified under denaturing conditions with subsequent refolding. Endotoxin was reduced to below 1.0 EU/mL. hNPs exhibited a size under 200 nm with a polydispersity index below 0.1, reflecting high formulation uniformity. CTD was conjugated efficiently at 35 µg/mg of hNPs. Multiple formulations have been produced with various adjuvants, including CpG oligodeoxynucleotide, resiquimod, and monophosphoryl lipid A. The nanovaccines are evaluated in eliciting systemic IgG, mucosal IgA, and cellular immune responses relevant to PEDV protection. Conclusion This modular hNP platform allows precise antigen display and flexible multi-adjuvant delivery. This platform represents a valuable approach for advancing PEDV vaccine development and improving swine health outcomes. Funding Source USDA National Institute of Food and Agriculture, AFRI project 2021-08581. Topic Categories Vaccines and Immunotherapy (VAC)

An isotype-specific phosphorylation of Hsp90 rewires co-chaperone regulations

Transient recovery voltage for load current switching test of MV load break switch

The Role of gC1qR and C1q in Tumor Cell Hijacking of Mitochondria from T Cells 2267216

Abstract Introduction Tumor cells are in constant need of energy and it has been shown that mitochondrial transfer from T cells to tumor cells can provide that. Complement protein C1q is present on tumor cell surface while the receptor for its globular heads, gC1qR, is present both on the surface and within the mitochondria of T cells. Methods The aim of the present study was therefore to investigate whether the interaction between gC1qR on T cells and C1q on the tumor cell facilitates hijacking of T cell mitochondria, resulting in T cell exhaustion. To this end, breast cancer cell lines SKBR3, MDA-MB-231, and MDA-MB-468 were treated with ethidium bromide (EtBr) to remove mitochondria, adding uridine (uri) as an alternate energy source. Conditions examined were: untreated, EtBr no uri, EtBr low uri, and EtBr high uri. At timepoints, tumor cell mitochondria were stained with fluorescent mitochondrial dye MitoRed and examined by fluorescence microscopy. Results By day five, untreated tumor cells exhibited increased fluorescence while cells treated with EtBr showed a decrease. Once mitochondria were depleted, MOLT4 T cells were stained with MitoRed and co-cultured with unstained tumor cells. On day 13, tumor cells co-cultured with MitoRed-stained T cells showed fluorescence in both tumor and T cells. Conclusion Fluorescence in tumor cells was attributed to T cell mitochondrial taken up by tumor cells, confirming the transfer of mitochondria from T cells to tumor cells. Future studies will examine the roles of C1q and gC1qR in mitochondrial transfer. Funding Source n/a Topic Categories Tumor Immunology: Cellular Responses and Tumor Microevironment (TIME)

Optimizing T Cell Priming Using TLR4/9 Combination Adjuvant For Robust Cell-Mediated Immunity Against Influenza 2307646

Abstract Introduction Subunit vaccines are safe but rarely induce robust, long-lasting immunity, especially cell-mediated immunity (CMI). Here, we investigated the potential of IVAX-1, a combination of TLR4 (MPLA) and TLR9 (CpG 1018) agonists formulated in a squalene oil-in-water emulsion (AddaVax), to mediate protection and generate CMI. Methods We leveraged a murine H5N1 influenza model with fate-map reporters, conditional knock-out mice, multiphoton imaging, and flow cytometry to elucidate early spatiotemporal features of IVAX-1-driven immunity. Antigen-specific CMI was evaluated via ELISpot and recall assays. Results Immune priming with IVAX-1 enhanced dendritic cell (DC) maturation and recruitment of myeloid cells to the injection site, resulting in higher numbers of neutrophils, monocytes, and antigen-presenting DCs in the draining lymph node (DLN), independent of antigen clearance kinetics. This induced islands of IFN-γ+ cells in the DLN, suggesting clonal expansion. In addition, IVAX-1 generated a robust antigen-specific functional (IFN-γ+) T cell response. Deletion of Type 1 conventional DCs (XCR1+) cells abrogated IVAX-1-mediated T cell response. Furthermore, immunization with IVAX-1 generated significantly higher numbers of flu-specific memory T cells in the lungs and spleen. IVAX-1 conferred durable protection and superior viral clearance from the lungs up to 60 days after immunization. Conclusion By leveraging synergistic innate immune activation through multiple TLR pathways, the combination optimizes T cell priming without relying on antigen depot effects. Our findings identify key early innate immune events and cellular mediators that drive robust CMI and demonstrate the potential of TLR agonist combinations to elicit durable T-cell immunity. Funding Source NIH/NIAID grants U01AI160397 & R01AI168063; Adjuvant Comparison and Characterization contract BAA-DAIT-75N93020R00022; and grant from the Defense Threat Reduction Agency #HDTRA-18-1-0036; UC Irvine UROP Research Experience Fellowship; NIH-MARC U-STAR trai Topic Categories Vaccines and Immunotherapy (VAC)

Immune-Privileged Landscapes and Endothelial Heterogeneity in Endocrine Organs 2335129

Abstract Introduction Endocrine glands rely on specialized vasculature to orchestrate hormone trafficking and tissue homeostasis. However, the transcriptomic landscape of endothelial cells (ECs) as gatekeepers of the immune microenvironment remains poorly defined. We aimed to map the molecular programs governing endocrine EC specialization and their role in regulating organ-specific immune communication. Methods We constructed a single-cell transcriptomic atlas of ECs from four endocrine organs (adrenal gland, pituitary, thyroid, kidney), using liver and brain as structural and immunological references. We categorized ECs by architectural features and quantified endothelial-immune crosstalk via ligand-receptor interaction analysis across these distinct vascular beds. Results Our analysis revealed distinct organotypic programs: while adrenal and kidney ECs showed vascular remodeling signatures, pituitary and thyroid ECs exhibited high stress-responsiveness. Crucially, we identified a stark divergence in immunological signatures. While the adrenal, pituitary, kidney, and liver were enriched in pro-inflammatory gene sets, the thyroid and brain were uniquely prominent in anti-inflammatory programs. This reflects a specialized “immune-shielding” interface in the thyroid, characterized by significant downregulation of leukocyte adhesion molecules and pro-inflammatory signaling, mirroring the immune-privileged phenotype of the blood-brain barrier. Strikingly, this protective endothelial interface is significantly compromised during aging and tumorigenesis, leading to a breakdown of the immune-restricted vascular state. Conclusion These findings define the molecular basis of endocrine vascular specialization and identify a unique endothelial-immune communication restriction in the thyroid. The loss of this immune-privileged phenotype is a hallmark of thyroid aging and cancer, suggesting that restoring the endothelial “immune-shielding” capacity could be a novel therapeutic strategy for endocrine pathologies. Funding Source RS-2023-00214581 Topic Categories Immune Response Regulation: Cellular Mechanisms (IRC)

Next-Generation Mpox Vaccine: Enhancing MVA-Based Immunity Through PD-1 Checkpoint Modulation 2309232

Abstract Introduction The 2022 Mpox outbreak exposed gaps in orthopoxvirus vaccine durability, as the licensed two-dose Modified Vaccinia Ankara (MVA) regimen provides only moderate and waning protection. Durable antiviral immunity requires coordinated humoral and CD8+ T-cell responses, with CD8+ T cells playing an important role in controlling infected cells at sites of viral replication. However, current vaccine strategies incompletely sustain these cellular responses. To address this limitation, MVA-X was engineered as an MVA-based vaccine in which a PD-1 antagonist peptide is encoded directly within the viral genome, enabling localized checkpoint modulation during antigen presentation. Methods Age- and sex-matched C57BL/6 mice received formulation buffer, single-dose MVA, two-dose MVA, or single-dose MVA-X. Mice were challenged with lethal vaccinia virus (VACV-WR) at days 55, 90, or 150 post-prime. Vaccine efficacy and immune responses were evaluated by survival, lung viral titers and genome burden, lung histopathology and immunohistochemistry, serum neutralization and binding antibody assays, and virus-specific CD8+ T-cell responses. Results A single dose of MVA-X conferred complete survival at all challenge time points, matching or exceeding two doses of MVA protection. At day 90 post-challenge, MVA-X vaccination resulted in > 97% reductions in lung viral titers and genome copies, preservation of alveolar architecture, and localized peribronchiolar immune infiltrates, consistent with efficient tissue-level viral control. Despite declining binding antibody responses, animals remained protected, consistent with sustained virus-specific CD8+ T-cell responses detectable through day 150 post-prime. Conclusion These findings show that MVA-X supports durable cellular immune responses following a single immunization. This vaccine-intrinsic checkpoint modulation strategy offers a modular framework with broader applicability for enhancing cellular immunity in viral vaccine development. Funding Source GeoVax,Inc Topic Categories Vaccines and Immunotherapy (VAC)

Human CD127+CD45RA+CCR7⁻CD8+ T cells represent an early-memory population governed by FOXP1 2250340

Abstract Introduction Following viral infection, naïve CD8+ T cells differentiate into effector and memory populations that eliminate infected cells and establish long-term immunity. Yet, the regulatory mechanisms directing the differentiation toward a memory fate remain unclear. Here, we aim to delineate the differentiation trajectories of antigen-specific human CD8+ T cells following cognate antigen stimulation, and to identify an early-memory subset with a durable memory potential. Methods Sequential peripheral blood samples were collected from healthy donors, vaccinated with BNT162b2 vaccine who subsequently experienced breakthrough infection, allowing longitudinal profiling of antigen-specific responses. Single-cell RNA sequencing integrated with DNA-barcoded MHC-I multimers was used to identify SARS-CoV-2 spike-specific CD8+ T cells. To further asses epigenetic regulation, parallel single-nucleus RNA and ATAC sequencing in a murine influenza A infection model was conducted. Results Regulatory network and trajectory analyses revealed a distinct early-memory population, uniquely exhibiting transcription factor FOXP1, within a spike-specific effector CD8 cluster. The analogous population in the murine model exhibited increased chromatin accessibility and FOXP1 motif enrichment. Phenotypically, FOXP1+ early-memory population showed a CD45RA+ effector memory T (TEMRA)-like surface expression with a loss of CCR7 expression while uniquely expressing CD127, suggesting functional heterogeneity within the TEMRA population. Functionally, FOXP1 knockdown reduced the expression level of TCF-1 in TCR-stimulated CD8+ T cells, implicating the role of FOXP1 in maintaining T-cell stemness. Conclusion Our results identify that the CD127+CD45RA+CCR7⁻CD8+ T cell subset represents an early-memory population with the potential to generate durable memory responses, and FOXP1 is a key regulator of this population. Our findings provide mechanistic insights into transcriptional and epigenetic regulation of memory CD8+ T-cell generation. Funding Source N/A Topic Categories Lymphocyte Differentiation and Peripheral Maintenance (LYM)

A Method to Detect Differences in Heterogeneous Cell Populations Without Assuming a Specific Data Distribution leveraging SOM-based clustering 2309518

Abstract Introduction Flow cytometry (FCM) is often used to identify specific cell populations that exhibit changes in abundance or count under different conditions. Recent advances in high-dimensional FCM have increased the number of simultaneously measurable parameters, making manual gating impractical and leading to the use of clustering-based analysis to detect differential abundance in cell populations across groups. There are several challenges with its current usage and methodology. First, even though the underlying distribution model of cluster frequencies is typically unknown, parametric statistical tests such as t-test are widely used. Second, existing methods do not adequately account for within-group heterogeneity. These challenges result in user-dependent trial-and-error in statistical analysis. Methods To address these issues, we developed a new differential abundance method for comparing two groups using a non-parametric test, simultaneously quantifying within-group heterogeneity. The concatenated sample data from the two groups are analyzed using SOM-based clustering to construct a sample × cluster matrix, which is then decomposed by Nonnegative Matrix Factorization (NMF). Differentially abundant clusters are identified with basis matrix via a permutation test, and within-group heterogeneity are quantified with coefficient matrix. Results We applied our method to 45-color human PBMC datasets acquired using a Sony ID7000™ Spectral Cell Analyzer (n = 8). Compared with manual analysis, our approach successfully detected differentially abundant clusters, which were also noted by manual analysis. In addition, by quantitatively evaluating within-group heterogeneity at the sample level, our method identified samples with intermediate or unanticipated patterns of cell abundance, which were discussed in manual analysis. Conclusion This approach has the potential to streamline the multi-sample analysis workflows and enhance the interpretability of differential abundance analyses in FCM research. Funding Source n/a Topic Categories Computational and Systems Immunology (COMP)