InVivoMAb rat IgG2b isotype control, anti-keyhole limpet hemocyanin

CD4 tissue-resident memory T cells (TRM) are crucial adaptive immune components involved in preventing influenza A virus (IAV) infection. Despite their importance, their physiological role in the upper respiratory tract, the first site of contact with IAV, remains unclear. Here, we find that, after IAV infection, antigen-specific CD4 TRM persist in the nasal tissue (NT) compartment after infection and provide protection upon heterosubtypic challenge. Single-cell RNA-sequencing analysis reveals that NT CD4 TRM are heterogeneous and transcriptionally distinct as compared with their lung counterparts. Mechanistically, we demonstrate that the CXCR6-CXCL16 axis promotes CD4 TRM residency in the NT. Furthermore, we show that the NT of mice and humans contains a high frequency of Th17 CD4 TRM that aid in local viral clearance and in reducing tissue damage. Collectively, our results support a robust physiological role for NT CD4 TRM in local protection during heterosubtypic IAV infection.

Pancreatic ductal adenocarcinoma (PDAC) is a lethal malignancy with a dense desmoplastic stroma and an immunosuppressive tumor microenvironment that contribute to therapeutic resistance. Here, we identify plasminogen activator inhibitor 1 (PAI1) as a stroma-derived mediator of immune evasion and tumor progression in PDAC. PAI1 is predominantly produced and secreted by cancer-associated fibroblasts, and its genetic ablation in the stromal compartment impairs tumor growth. Mechanistically, hypoxia induces PAI1 expression in fibroblasts, which in turn shifts macrophages toward immunosuppressive phenotypes and suppresses CD8+ T cell infiltration and function. We further show that tissue plasminogen activator (tPA), a direct PAI1 target, is also secreted by fibroblasts and supports antitumor CD8+ T cell responses. Notably, elimination of stromal tPA promotes immunosuppressive macrophage phenotypes, reduces CD8+ T cell infiltration, and accelerates PDAC progression. These findings define a previously unrecognized PAI1-tPA regulatory axis within the tumor stroma that modulates antitumor immunity. Targeting this pathway may provide a therapeutic opportunity to overcome stroma-driven immune suppression in PDAC.

Chronic liver diseases (CLD) account for more than 2% of deaths worldwide. Extensive research has been conducted to better understand CLD, generating vast amounts of data. However, only a small fraction of raw preclinical data are publicly available, posing a significant challenge for transparency, reproducibility, and data reuse. Therefore, we built a preclinical liver imaging dataset, the first of its kind to our knowledge. The database contains longitudinal liver MRI scans from mice with hepatocellular carcinoma, metabolic dysfunction-associated steatohepatitis (MASH, formerly NASH), and fibrosis, as well as CT scans of mice with MASH and mice carrying a dysfunctional ICAM-1 gene. Superimposable MRI and CT scans bridge the gap between the modalities. Some of the 222 murine scans have annotated segmentations. Metadata containing both scan and mouse parameters are organized using a tailored metadata profile in ISA-Tabs. This dataset enables advanced image analysis, such as building tools for automated segmentation, train radiomics analysis tools, or can be used as a reference control dataset.

Immunosuppression and metastasis are critical hallmarks of breast cancer, often linked to poor patient outcomes. The secreted cytokine chitinase-3-like 1 (CHI3L1) is frequently overexpressed in breast cancer samples and promotes an immunosuppressed tumor microenvironment. Notably, CHI3L1 expression is elevated in metastatic patient samples when compared with the matched primary breast tumor. To investigate its role in breast cancer metastasis, we generated an inducible genetically engineered mouse model that overexpresses CHI3L1 in the mammary epithelium. Ectopic expression of CHI3L1 in the polyomavirus middle T (PyMT) mouse model of breast cancer suppressed antitumor immune responses, accelerated mammary tumor onset, and enhanced lung metastasis. Mechanistically, elevated CHI3L1 expression in the mammary epithelium enhanced neutrophil recruitment, which subsequently degraded the extracellular matrix and increased the number of circulating tumor cells. These findings reveal a key mechanism driving metastatic dissemination and argue that therapeutically targeting Chi3l1 could enhance antitumor immunity and suppress metastasis.

Bone marrow adipocytes are known to have a critical role within the bone marrow niche. However, our understanding of bone marrow adipose tissue expansion with obesity and the role it plays in immune cell regulation and osteoclastogenesis is limited. Here, we showed the expansion of bone marrow adipocytes promoted osteoclast differentiation and subsequently led to obesity-related trabecular and cortical bone loss through a stimulatory effect of the PD-1/PD-L1 axis. Bone marrow adipocytes isolated from obese mice had increased Mcp-1 expression, a key regulator of osteoclastogenesis and myeloid cell accumulation. With the increase in bone marrow adipose tissue-derived Mcp-1, we found an increase in the number of PD-L1+ myeloid cells. While these cells inhibited activated T-cells, we found evidence of a stimulatory osteoclastogenic effect of PD-L1+ myeloid cells on PD-1-expressing osteoclast precursors. The inhibition of PD-1/PD-L1 signaling during early osteoclastogenesis prevented myeloid cell commitment and resulted in decreased cell fusion, supporting the role of PD-1/PD-L1 signaling in osteoclastogenesis. Using a bone marrow adipocyte depletion mouse model (BMAd-Pparg KO), we demonstrated that obese BMAd-Pparg KO mice had a reduced number of bone marrow PD-L1+ myeloid cells, accompanied by a decrease in PD-1+ osteoclast precursors. The reduction in these precursors resulted in fewer osteoclasts, subsequently leading to improved trabecular bone volume. Since osteoclasts are myeloid cell-derived, these results suggest that bone marrow adipocytes are critical for the commitment and differentiation of myeloid cells into osteoclasts. Targeting bone marrow adipogenesis could ameliorate enhanced osteoclastogenesis and provide a novel approach to treat obesity-related bone loss. Obesity-induced expansion of BM adipocytes leads to PD-1/PD-L1-driven osteoclastogenesis and subsequent bone loss in obese, HFD-fed (OB-HFD) mice. After 12 weeks on a HFD, OB-HFD mice had a significant increase in BM adiposity and BMAT-derived Mcp-1 expression. The increase in BMAT-specific Mcp-1 expression was coupled with an increase in PD-1+ osteoclast (OC) precursors and PD-L1+ myeloid cells. In the context of obesity, the PD-1/PD-L1 axis has a stimulatory effect that enhances osteoclastogenesis and leads to trabecular and cortical bone loss. By depleting BM adipocytes with obesity, BMAT-derived Mcp-1 expression was decreased, as well as a decrease in PD-1+ OC precursors and PD-L1+ myeloid cells. This prevented obesity-related trabecular bone loss. Overall, this work demonstrated a strong correlation between BMAT expansion and PD-1/PD-L1-driven osteoclastogenesis as a mechanism for obesity-induced bone loss. (This image was created using BioRender).

Escherichia coli is a leading cause of neonatal sepsis, with infection occurring in approximately one in every 1,000 live births1,2. However, with E. coli colonization beginning soon after birth3-5 and defects in neonatal host defence maturation6-9, an alternative consideration is why infection does not occur even more frequently. Here we show that newborn babies with E. coli sepsis have selectively reduced vertically transferred natural antibodies that recognize E. coli, mechanistically explaining their susceptibility to infection. Complementary preclinical studies show that preconceptual intestinal colonization with probiotic E. coli Nissle 1917 (EcN)10 primes anti-E. coli immunoglobulin G (IgG) antibodies with broad cross-reactivity to clinical isolates responsible for neonatal sepsis that override the inherent susceptibility of neonatal mice. Outer membrane protein A (OmpA) is a target of maternal IgG and is also essential for EcN colonization-induced serological immunogenicity. Upon vertical transfer to neonates, colonization-primed anti-E. coli IgG uniquely protects against infection via opsonization, requiring both complement and IgG Fc receptors. Compared with specimens from sex and gestational age-matched healthy control babies without infection, dried blood spot specimens collected one day after birth from 100 babies with E. coli sepsis show consistently reduced IgG titres to pooled E. coli clinical isolates and OmpA, along with impaired IgG-dependent antibacterial opsonization. Together, these results demonstrate that natural infection susceptibility of neonates is efficiently rescued by anti-E. coli IgG and identify defects in pathogen-targeted vertically transferred immunity as a primary risk factor for severe invasive infection in newborn babies.

Metastasis is the major cause of death for patients with triple-negative breast cancer and other solid malignancies. Metastases arise from cancer cells that disseminate from the original tumour, survive systemic immune surveillance and colonize new organs1. Little is known about how initial disseminated tumour cells (DTCs) overcome anti-tumour immunity after seeding a new organ. Here we use a visible antigen in a model of triple-negative breast cancer with cognate CD8+ T cells to study the mechanisms of immune evasion in early metastatic seeding. Analysis of surviving DTCs revealed glucocorticoid receptor (GR) activation as a key driver of resistance to both CD8+ T cells and natural killer cells. Niche profiling using an optimized labelling tool identified FAS-FASL as a key pan-cytotoxic pathway against DTCs, which is repressed by GR activation. Pharmacological inhibition of GR in combination with immunotherapy reduced metastatic burden and expanded lifespan in mice. Thus, we identified a mechanism of immune evasion that operates specifically in DTCs, illustrating the unique immune-cancer interactions at this stage in the metastatic cascade. Our findings suggest that there are therapeutic opportunities to eliminate DTCs, separately from treatments aimed at primary tumours, and GR inhibition is one promising target.

Immune checkpoint blockade (ICB) faces limitations owing to high cost and restricted efficacy. This study identifies SNX17 as a key mediator of ICB resistance. Elevated SNX17 correlates with poor anti-PD-1 response in humans and mice. SNX17 deletion in tumor cells inhibits tumor growth via CD8+ T cell-dependent mechanisms. SNX17 reduces uridine in the tumor microenvironment (TME), suppressing IFN-γ and upregulating PD1 in CD8+ T cells. Exogenous uridine shows antitumor efficacy comparable to anti-PD-1/PD-L1 in low-SNX17 tumors and overcomes resistance in high-SNX17 models. Uridine enhances CD8+ T cell function by promoting CD45 N-glycosylation and LCK phosphorylation. Mechanistically, SNX17 stabilizes RUNX2, promoting UPP1 transcription and uridine degradation in the TME. These findings position SNX17 as an ICB response biomarker and nominate uridine as a cost-effective immunotherapeutic strategy.

Prevotella melaninogenica is enriched in the lungs of people with cystic fibrosis (pwCF), yet its functional impact on respiratory tract homeostasis remains incompletely understood. Prior studies identified immune modulatory effects following lung exposure to Prevotella, but the relevance of these findings for CF infections is unknown.

Immunotherapy remains ineffective for a wide variety of solid tumors due to the existence of tumor immune evasion. Although the transcription factor ETV5 is recognized for its oncogenic roles in tumor progression, its role in remodeling the immunosuppressive microenvironment remains largely unexplored. Here, we reveal that tumor-intrinsic ETV5 drives immune evasion and immune checkpoint inhibitor (ICI) resistance by enhancing the expansion and recruitment of polymorphonuclear myeloid-derived suppressor cells (PMN-MDSCs). Genetic silencing of ETV5 in murine tumor models suppressed PMN-MDSCs differentiation from myeloid progenitors, reduced their tumor infiltration, and attenuated immunosuppressive function, resulting in enhanced cytotoxic T cell activity and delayed tumor progression. Mechanistically, ETV5 directly binds to the JH1 domain of JAK2, inducing its dimerization and phosphorylation, which activates STAT3 to transcriptionally upregulate CCL2 and recruit PMN-MDSCs. Therapeutically, ETV5 ablation synergized with anti-PD-L1 therapy to enhance tumor control, mirroring clinical observations where high ETV5 expression predicted immunotherapy resistance. Our study uncovers a non-canonical, transcription-independent role of ETV5 in orchestrating the JAK2/STAT3/CCL2 axis to sustain PMN-MDSC-mediated immune evasion, proposing ETV5 as a druggable target to overcome ICI resistance in solid tumors.

Cancer cells activate the integrated stress response (ISR) to adapt to stress and resist therapy1. ISR signals converge on activating transcription factor 4 (ATF4), which controls cell-intrinsic transcriptional programs that are involved in metabolic adaptation, survival and growth2,3. However, whether the ISR-ATF4 axis influences anti-tumour immune responses remains mostly unknown. Here we show that loss of ATF4 decreases tumour progression considerably in immunocompetent mice, but not in immunocompromised ones, by enhancing T cell-dependent anti-cancer immune responses. An unbiased genetic screen of ATF4-regulated genes identifies lipocalin 2 (LCN2) as the principal ATF4-dependent effector that impairs anti-tumour immunity by favouring infiltration with immunosuppressive interstitial macrophages. Furthermore, we find that LCN2 promotes T cell exclusion and immune evasion in preclinical mouse models, and correlates with decreased T cell infiltration in patients with lung and pancreatic adenocarcinomas. Anti-LCN2 antibodies promote robust anti-tumour T cell responses in mouse models of aggressive solid tumours. Our study shows that the ATF4-LCN2 axis has a cell-extrinsic role in suppressing anti-cancer immunity, and could pave the way for an immunotherapy approach that targets LCN2.

Discovering more targets is of great importance for developing alternative interventions for tumor therapy. The roles of transmembrane protein 175 (TMEM175) in neurodegeneration diseases have been reported, however its functions in tumor immune surveillance are not known. We show that TMEM175 conditional knockout in macrophages inhibits the tumor growth and metastasis through promoting the anti-tumor immunity in the tumor microenvironment (TME), including elevated M1-like polarization, reduced M2-like polarization, and facilitated recruitment and activation of T cells and nature killer cells (NKs). The anti-tumor immunity is abrogated by caspase-1 inhibitor VX-765, anti-IL-1β, and anti-IL-18. Tmem175-/- bone marrow-derived macrophages (BMDMs) show enhanced tumor antigen cross-presentation that is further strengthened by IL-1β and IL-18. NLRP3 is robustly elicited in Tmem175-/- BMDMs by the tumor cell debris through lysosomal permeabilization and cathepsin B leakage. Finally, Tmem175-/- mice are more responsive to anti-PD-1. Our works implies TMEM175 to be a potential target for immunotherapy.

Oral lichen planus (OLP) is a recalcitrant inflammatory disease with potential for malignant transformation, involving a cytotoxic CD8+ T cell-mediated basal keratinocyte apoptosis. However, it lacks an appropriate mouse model for study. Here we developed an OLP-like mouse model using topical oxazolone to induce a delayed-type hypersensitivity-mediated oral lichenoid reaction. Histological and ultrastructural analysis confirmed hallmark pathological features of OLP, including band-like CD8+ T cell infiltration and basal cell damage as well as the presence of Civatte bodies. Comparative transcriptomic analysis revealed significant similarity between RNA-Seq profiles of the mouse model and human OLP lesions, highlighting shared upregulated genes and enriched pathways, particularly those related to IFN-γ signaling and cytotoxic T cell activity. Functional studies demonstrated that the OLP phenotype depended on IFN-γ, with local priming by IFN-γ intensifying the disease through upregulation of major histocompatibility complex class I. Additionally, the absence of Langerhans cells exacerbated disease severity in vivo. Therapeutic evaluation showed that the JAK inhibitors baricitinib and ruxolitinib effectively reduced disease burden and provided mechanistic insights. In conclusion, this OLP-like mouse model recapitulates key immunopathological and transcriptomic features of human OLP, offering a robust platform for dissecting disease mechanisms and evaluating novel therapeutic strategies.

During vascular injury, platelets are essential for halting bleeding and recruiting neutrophils to prevent microbial invasion. However, in antibody-mediated autoimmune diseases occurring without vascular damage, neutrophils infiltrate tissues and contribute to pathology. Here, we investigated whether the dependence of neutrophils on platelets is conserved in the context of antibody-driven inflammation. Using human cells from individuals with rheumatoid arthritis and a microfluidic system mimicking physiological shear over IgG-containing immune complexes, we demonstrate that despite expressing Fc receptors, neutrophils required platelets to stably adhere to immune complexes under flow. Platelet Fcγ receptor 2a (FcγRIIA) binding was critical for resisting shear stress, while neutrophils used FcγRIIA and FcγRIIIB for immune complex recognition. Platelet P-selectin binding to neutrophil P-selectin glycoprotein ligand 1 (PSGL-1) was essential for recruitment, whereas macrophage-1 antigen (Mac-1) was dispensable. In a mouse model of autoantibody-mediated arthritis, intravital imaging confirmed that neutrophil recruitment relied on PSGL-1. Importantly, expression of FcγRIIA aggravated arthritis, and blockade of PSGL-1, but not Mac-1, in these mice abrogated both the platelet and neutrophil interactions and disease. These findings identify key molecular interactions in platelet-neutrophil cooperation and reveal that platelets are essential enablers of FcR-mediated neutrophil adhesion in antibody-driven inflammation.

Background: Lung metastasis is a leading cause of breast cancer (BC)-related mortality, driven by the immunosuppressive traits of the metastatic tumor microenvironment. However, the mechanisms underlying cell-cell crosstalk in shaping immune evasion within the metastatic niche remain poorly defined. Neutrophil extracellular traps (NETs) and their associated proteins, such as cathelicidin, have emerged as key mediators of metastatic regulation in cancer. Here, we aimed to decipher the interaction between a neutrophil subset characterized by high expression of lymphocyte antigen 6 complex locus g (Ly6ghigh) and cluster of differentiation 8-positive T lymphocytes (CD8+ T cells), mediated via cathelicidin embedded in NETs, as well as their synergistic mechanism and cooperative role in promoting lung metastasis of BC. Methods: We characterized neutrophil heterogeneity and functional dynamics by performing single-cell RNA sequencing and flow cytometry on lung tissues derived from murine models of BC lung metastasis. We utilized cathelicidin-related antimicrobial peptide (Cramp) knockout mice to dissect the role of cathelicidin in NETs. The spatial colocalization of apoptotic CD8+ T cells and NETs was analyzed using multiplex immunofluorescence, and the molecular interactions were probed by protein binding assays. Results: Neutrophils in the lung metastatic niche were classified into 2 subsets based on the Ly6g expression: Ly6ghigh and Ly6glow neutrophils. Ly6glow neutrophils, which were recruited in the macrometastatic stage, exhibited myeloid-derived suppressor cell-like characteristics. Notably, Ly6ghigh neutrophils induced CD8+ T cell apoptosis through NET formation, with apoptotic CD8+ T cells spatially clustered within NET-rich areas. Mechanistically, NET-derived cathelicidin (Cramp in mice) directly bound to mitochondrial adenine nucleotide translocator 1 (Ant1) in CD8+ T cells, triggering conformational changes and complex formation with voltage-dependent anion channel 1 (Vdac1). These events resulted in the opening of the mitochondrial permeability transition pore and loss of mitochondrial membrane potential. Conclusions: Our study demonstrates that Ly6ghigh neutrophils play a critical role in immunosuppression and immune evasion through NET-induced apoptosis of CD8+ T cells. These findings underscore the importance of NETs and cathelicidin in BC lung metastasis, suggesting their potential as therapeutic targets in restoring antitumor immunity and in preventing metastatic progression.

While natural killer (NK) cells and B cells arise from common lymphoid progenitors, the existence and nature of cells co-expressing markers of both lineages (NK-B cells) remain controversial. Here, this work identifies a significant population of CD3-NKp46-CD19⁺NK1.1⁺ cells, termed NK-B cells, enriched in phosphoinositide-dependent protein kinase-1(PDK1)-deficient mice but also present in wild-type bone marrow. Single-cell RNA sequencing and high-dimensional flow cytometry reveal these NK-B cells reside within early B cell developmental stages (pro-B/pre-B). Functionally, upon adoptive transfer into lymphocyte-deficient hosts, NK-B cells preferentially differentiated into B cells. Unlike conventional B cell precursors, NK-B cells exhibit innate characteristics, including the capacity to secrete interferon-gamma (IFN-γ) and transforming growth factor-beta (TGF-β), and expressed CD122 (IL-2/15Rβ) and Toll-like receptor 9 (TLR9). Using a novel E4 promoter-binding protein 4 (E4BP4) reporter model, this work demonstrates that both interleukin-15 (IL-15) signaling (via CD122) and TLR9-mediated sensing of the gut microbiota cooperatively sustain E4BP4 expression within these progenitors. Consequently, germ-free mice and mice deficient in IL-15 or E4BP4 exhibit a profound loss of NK-B cells. These findings unveil a distinct E4BP4-expressing, innate-like B cell progenitor pathway regulated by microbiota and IL-15.

Duchenne muscular dystrophy (DMD) is a severe neuromuscular disorder with progressive muscle degeneration and cardiomyopathy leading to heart failure. The inflammatory environment in dystrophic skeletal muscle is well-studied, but little is known about inflammation in DMD cardiomyopathy due to the lack of adequate animal models. We recently developed the Fiona/dko mouse model, deficient for both dystrophin and utrophin, but containing a skeletal muscle-specific expressing utrophin transgene allowing progression of dystrophic cardiomyopathy. This Fiona/dko model is the first DMD cardiomyopathy model to reproducibly progress to reduced cardiac contractile function by 9 mo. In this study, we compared immune cell composition between Fiona/dko mice and their milder littermates that develop cardiac pathology, but do not demonstrate whole heart dysfunction. Flow cytometry analysis revealed that T cells constitute a significant proportion of the immune cell population in dystrophic hearts, in contrast to the known predominantly myeloid signature in dystrophic skeletal muscles. T cell infiltration precedes the development of cardiac fibrosis and dysfunction in Fiona/dko mice. RNA sequencing of whole hearts after cardiac dysfunction shows increased expression of 68 genes related to T cell signaling in Fiona/dko compared with their milder littermates. Furthermore, depletion of circulating CD3+ T cells with a neutralizing antibody ameliorates early pathology in Fiona/dko hearts. Together, these data suggest a role for T cells in the initiation and persistence of dystrophic cardiomyopathy. These findings highlight the distinct inflammatory environment in the dystrophic heart and provide new insights into DMD cardiomyopathy, paving the way for the future development of targeted anti-inflammatory therapies.NEW & NOTEWORTHY Heart failure has become the leading cause of death in Duchenne muscular dystrophy, a progressive degenerative disease of striated muscles. This study highlights an inflammatory environment in dystrophic heart with a high proportion of T-cells that is distinct from the predominant myeloid inflammation in dystrophic skeletal muscles. T-cell-related signaling is associated with the severity of cardiomyopathy, and T-cells contribute to dystrophic cardiomyopathy onset. This data will inform optimal patient treatments that target cardiac inflammation.

The gut microbiome has emerged as a critical modulator of cancer immunotherapy response. However, the mechanisms by which gut-associated metabolites influence checkpoint blockade efficacy in prostate cancer (PC) remain not fully explored. The study aimed to explore how gut metabolites regulate death-ligand 1 (PD-L1) blockade via exosomes and boost immune checkpoint inhibitors (ICIs) in PC.

Fibrosis is a hallmark of organ failure observed after chronic epithelial injury and inflammation. The transforming growth factor beta (TGF-β) is the master regulator of fibrogenesis, so blockade of the TGF-β pathway is a potential treatment strategy for fibrosis; however, the therapeutic potential of pan-TGF-β blockade is limited by side effects.

Decorin (DCN) predominantly produced by fibroblasts is a small leucine-rich proteoglycan with tumor-suppressive property. However, whether DCN has a role in shaping the tumor immune microenvironment remains elusive.