bluuebunny/crossref_metadata_2025_split huggingface dataset:
Leveraging outbred mice to identify genetic predictors of durable response to anti-tumor antibody
Authors: Glassbrook James E , Hackett Justin , Muñiz Maria , Gibson Heather 
Published: February 2023

Abstract
Trastuzumab is one of many monoclonal antibody (mAb) therapies targeting tumor-associated antigens (TAA) that have markedly improved cancer outcomes. Trastuzumab targets epidermal growth factor 2 (HER2). While it has long been considered the standard-of-care for HER2+ breast cancer treatment, differential response between patients suggests that host genetic background may contribute to an effective response to therapy. We utilize genetically heterogeneous Diversity Outbred (DO) mice to identify genetic elements that influence host response to TAA-mAb therapy. In a cohort of (DOxBALB/c)F1 mice bearing established BALB/c-syngeneic Neu+ (a HER2 homolog) TUBO mammary tumors, we find a divergent response to αNeu mAb therapy (clone 7.16.4). Several phenotypes were collected to compare immunological function to projected tumor doubling time (TDT). These include activation of peripheral anti-tumor T cells and IgG, natural killer cell ADCC competence, bone marrow derived macrophage ADCP/trogocytosis efficacy, and tumor immune cell infiltrate proportions. Contrary to the prospective mechanism of action of trastuzumab, we show that neither capacity for ADCC or ADCP correlate with a robust response to therapy. Following genetic linkage analysis, we identify a locus on Chr 10 that associates with TDT. To verify this locus, we use recombinant inbred collaborative cross (CC) mouse strains (CCxBALB/c) F1 mice that contain or lack this driver genotype. We find pooled survivorship between prospective responder/non-responder strains 2 months post tumor implantation differs by ~8%. This locus alone can’t fully account for a durable response to mAb therapy highlighting the multigenic nature of host immune response.
Multicellular immune hubs and their organization in MMRd and MMRp colorectal cancer
Authors: Chen Jonathan H , Pelka Karin , Hofree Matan , Giannakis Marios , Boland Genevieve M , Aguirre Andrew J , Anderson Ana C , Rozenblatt-Rosen Orit , Regev Aviv , Hacohen Nir 
Published: February 2023

Abstract
Immune responses to cancer are highly variable, with mismatch repair-deficient (MMRd) tumors exhibiting more anti-tumor immunity than mismatch repair-proficient (MMRp) tumors. To understand the rules governing these varied responses, we transcriptionally profiled 371,223 cells from colorectal tumors and adjacent normal tissues of 28 MMRp and 34 MMRd patients. Analysis of 88 cell subsets and their 204 associated gene expression programs revealed extensive transcriptional and spatial remodeling across tumors. To discover hubs of interacting malignant and immune cells, we identified expression programs in different cell types that co-varied across patient tumors and used spatial profiling to localize coordinated programs. We discovered a myeloid cell-attracting hub at the tumor-luminal interface associated with tissue damage, and an MMRd-enriched immune hub within the tumor, with activated T cells together with malignant and myeloid cells expressing T-cell-attracting chemokines. By identifying interacting cellular programs, we thus reveal the logic underlying spatially organized immune-malignant cell networks.
Targeted protein degradation via NEDD8 conjugation does not enhance direct MHC class I antigen presentation.
Authors: Vijayasimha Kartikeya , Leestemaker-Palmer Amy Leigh , Gibb James , Yewdell Jonathan W , Dolan Brian P 
Published: February 2023

Abstract
Knowledge of different pathways contributing to peptide generation for direct MHC class I antigen presentation is important to the field of immunotherapy. Here we investigated the role of the ubiquitin-like protein NEDD8 in antigen presentation. We show that proteins tagged N-terminally with NEDD8 undergo rapid degradation via the proteasomal, and autophagy-lysosomal pathways. To determine if protein NEDDylation increased peptide presentation, we fused NEDD8 to the N-terminus of a cytosolic form of ovalbumin (OVA) and measured presentation of the SIINFEKL peptide via the murine MHC class I molecule K b , and compared the results to ovalbumin N-terminally modified with ubiquitin. While NEDD8-OVA was processed for degradation via the proteasomal and autophagosomal pathway Ub-OVA was only degraded via the proteasome. Peptide presentation from NEDD8-OVA was dependent on NEDD8-ultimate buster 1 (NUB1), while autophagy did not contribute peptides antigen presentation. Overall, we found that N-terminal Ub conjugation was far more effective at generating antigenic peptides than N-terminal NEDD8 conjugation. However, upon inhibition of NEDDYylation by treatment with MLN4924, we did detect a decrease in peptide presentation from a model antigen used specifically to track Defective Ribosomal Product (DRiP) presentation. These data demonstrate that while NEDD8 can cause the rapid degradation of substrates it is not as efficient as Ubiquitin for generating antigenic peptides. Our data provides a basis for understanding the role of NEDD8 in protein degradation and antigen presentation, especially in conditions of NEDD8 dysregulation, such as in some cancers and protein folding pathologies.
Epithelial antigen presentation regulates CD4+ TRM cell locations, functions and activities
Authors: Shenoy Anukul T. , Arafa Emad I , De Ana Carolina Lyon , Barker Kimberly A , Korkmaz Filiz T , Ramanujan Aditya , Etesami Neelou S , Martin Ian MC , Tilton Brian R , Hinds Anne , Goltry Wesley N , Kathuria Hasmeena , Jones Matthew R , Quinton Lee J , Belkina Anna C , Mizgerd Joseph P 
Published: February 2023

Abstract
Barrier tissues are sentinelled by CD4+ TRM cells with potent anti-microbial activities and considerable lineage plasticity. We hypothesized that local antigen presentation by lung epithelial cells (LECs) instruct CD4+ TRM cell activities. Pneumococcal infections in transgenic mice, flow- and spectral-cytometry, computational biology, and immunofluorescence were used to study this biology. All LECs including a novel alveolar surfactant protein C (SPC)low LEC were adept at antigen presentation. Temporal analysis of LECs for MHC-II and costimulatory/coinhibitory molecules revealed that airway club cells were T-cell stimulatory via CD40 while alveolar LECs expressed T-cell inhibitory PD-L1. This anatomical segregation of LEC antigen presentation correlated with deposition of CD4+ TRM cells around airways such that ablation of LEC MHC-II disrupted CD4+ TRM niches and blockade of CD40 signals prevented accumulation of CD4+ TRM cells. Recurrent memory recalls in absence of LEC MHC-II led to expansion of unconventional CD4+ TRM cells co-expressing classically incompatible lineage-defining transcription factors, changing their cytokine repertoire and leading to dysregulated immunity that phenocopied clinical features of checkpoint blockade therapy. Consequently, a tight correlation between MHC-II and PD-L1 was confirmed in mouse and human LECs. We discovered that LEC MHC-II functions in post-translational trafficking lockstep with PD-L1 to exert its restraints on TRM cell activities. Our results identify epithelial antigen presentation as critical instructors of CD4+ TRM cell locations, phenotypes and activities and establish epithelial-CD4+ TRM cell immunological synapses as key components of barrier immunity.
Sex hormones play a critical role in modulating the immune response to peanut
Authors: Vininski McKenna S , Dolence Joseph J. 
Published: February 2023

Abstract
The prevalence of peanut (PN) allergy is increasing rapidly. While PN allergy is a major medical and economic problem, our knowledge of the immunological mechanisms involved in the development of the disease remains limited. Sexual dimorphisms have been observed in allergic asthma and in autoimmune diseases such as systemic lupus erythematosus. However, how sex hormones regulate the immune pathways that lead to the development of PN allergy is unknown. This study aims to better understand how sex hormones impact the development of allergic responses to PN. Male and female wild type C57Bl/6 mice were sensitized to PN in our established four-week PN inhalation mouse model. Wild type (WT) males had significantly less IgE levels than WT females. When WT male and female C57Bl/6 mice were gonadectomized and implanted with capsules containing opposite sex hormone (e.g. β-estradiol in males and dihydrotestosterone in females), data showed females treated with testosterone displayed reduced allergic reactions to PN. Taken together, this data strongly suggests that sexual differences exist in the development of PN allergy and furthermore, supports that testosterone plays a role in modulating allergic immune responses to PN.
IL-9 regulates type-2 CD4 tissue resident memory cell mediated allergic airway recall responses
Authors: Kharwadkar Rakshin P , Ulrich Benjamin J , Chu Michelle , Fu Yongyao , Pajulas Abigail , Kaplan Mark H 
Published: February 2023

Abstract
Allergic asthma is a chronic inflammatory lung disease with intermittent flares where patients experience symptoms mediated by memory T cells. Allergen-specific CD4+ tissue resistant memory (Trm) cells in peripheral tissues have been reported to facilitate mucosal barrier immunity. However, the precise role of Trm CD4+ T cells in allergic exacerbations is not clearly defined. In this study we identified interleukin-9 (IL-9)-secreting allergen-specific CD4 T cells with a memory phenotype that regulate recall response to Aspergillus fumigatus. Inhibition of circulating T cells by administering FTY720 in the last month of rest, diminished the total CD4 T cells population in the lung, while the IL-9 secreting CD4 tissue resident cells remained stable. Single-cell RNA-seq and ATAC-seq confirmed the presence of multi-cytokine producing IL-9 secreting CD4 Trms in early recall responses to the allergen. Blockade of IL-9 prior to recall challenge phase reduced overall allergic lung inflammation observed through both flow-cytometry and single cell-RNA-seq analysis. IL-9 neutralization reduced cellularity of several populations in the lung including granulocytes and lymphocytes. Moreover, the expression of genes associated with mucus metaplasia including Muc2, Muc5ac, Muc5b, and Bpifb1 were significantly reduced in type-2 alveolar and epithelial cells with anti-IL-9 treatment. These findings demonstrate that IL-9-producing Trms play an important role in mediating allergic memory responses. IL-9 could be a promising therapeutic target specifically in patients showing symptoms of intermittent allergic response.
Suppression of T-cell Immune responses by Neutrophils in a direct contact manner
Authors: Soyemi Olamipejo O. , Moreira Josimar Dornelas , Samten Buka 
Published: February 2023

Abstract
Neutrophils are protective against early Mycobacterium tuberculosis (Mtb) infection, but overactive neutrophil response is often associated with chronic lung inflammation, tissue damage and compromised T-cell immunity, implying potential immunosuppression by neutrophils in Mtb infection. To test this, we examined the effect of peripheral blood neutrophils on the immune responses of T-cells from the peripheral blood mononuclear cells (PBMC) of persons with latent tuberculosis infection. Co-cultures of PBMC with autologous neutrophils at different ratios inhibited Mtb-stimulated T-cell proliferation and cytokines IFN-γ, IL-17 and IL-10 production dose dependently without affecting cell viability as determined by resazurin assay for cell metabolism and annexin V/PI staining for cell death. Neutralization of IL-10 and TGF-β failed to rescue neutrophil suppression of T-cell IFN-γ production. Neutrophils suppressed IFN-γ mRNA expression as determined by qPCR. Neutrophils inhibited purified T-cells stimulated with immobilized anti-CD3 and anti-CD28 mAbs, suggesting direct effect on T-cells without the involvement of antigen presenting cells. Trans-well study showed that cell-cell contact is required for this suppression. Further assessment of granulocyte derived suppressor cell phenotype and blocking the interaction between immune checkpoint molecules, PD-1 and PD-L1, refuted their roles in suppression of T-cells responses in Mtb infection. Together, these data suggest that neutrophils suppress T-cell immune responses by direct cell-cell contact mechanism and identification of molecular details of such suppression will shed new insights into the pathogenesis of tuberculosis and other infectious diseases.
Immunosuppressive function of DPP4, a proinflammatory molecule, in inflammatory colitis
Authors: Zhong Jixin , Wei Yingying , Rao Xiaoquan 
Published: February 2023

Abstract
We and others have shown that dipeptidyl peptidase-4 (DPP4) promotes inflammation in a number of conditions. However, it’s role in inflammatory colitis is not fully understood. Here we examined the role of DPP4 in dextran sulfate sodium (DSS)-induced colitis model and T cell transfer model of chronic colitis. The expression of DPP4 was significantly decreased in the colon tissue of wild-type (WT) mice with colitis. The mortality and disease activity score were markedly increased in mice deficient for DPP4 (Dpp4−/−), compared to WT mice. Chimeric mice with Dpp4−/− bone marrow showed a similar disease severity to whole body DPP4 knockout mice in DSS induced colitis, suggesting that bone marrow-derived immune cells were responsible for the protective effect of DPP4 on colitis. Flow cytometry analysis indicated that DPP4 was mainly expressed in T cells and dendritic cells (DCs) in mesenteric lymph nodes. Using T cell transfer model of chronic colitis in Rag2−/− mice or Dpp4−/− Rag2−/− mice, we found that innate immune cells other than adaptive immune cells were responsible for the protective effect of DPP4. Subsequent flow cytometric analysis showed that DPP4 was mainly expressed on a DC subset that was responsible for the induction of regulatory T cells (Tregs) in the gut. RNA sequencing data also indicated that expression of genes in TGFβ signaling pathway was significantly reduced in Dpp4−/− mice, accompanied by an increased expression of multiple inflammatory genes. Co-transfer of WT Tregs protected Rag2−/− mice, but not Dpp4−/− Rag2−/− mice, from developing colitis after transfer of CD4+ naive T cells. Our data suggest that DPP4 expression on innate immune cells may be responsible for maintaining Treg’s suppressive function in colitis.
The Ubiquitin Ligase Itch Regulates Germinal Center Selection
Authors: Moser Emily K 
Published: February 2023

Abstract
More than 23 million Americans suffer from autoimmune diseases driven by somatically mutated autoantibodies. High levels of somatic hypermutation of immunoglobulin genes occurs in germinal center (GC) B cells. Some of these mutations render autoreactivity, but cells bearing autoreactive receptors are eliminated due to strict selection of GC B cells by mechanisms that are still poorly defined. Inappropriate positive selection signals can rescue autoreactive cells from elimination and drive autoimmune disease. The E3 ubiquitin ligase Itch prevents the emergence of autoimmune disease and autoantibodies in humans and mice, and patients lacking Itch develop debilitating, multi-faceted, potentially fatal, autoimmune disease; yet how Itch regulates GC B cell fate or function is not well understood. By studying Itch deficient mice, we have recently shown that Itch directly limits B cell activity to shape antibody responses. While Itch-deficient mice displayed normal numbers of pre-immune B cell populations, they showed elevated numbers of GC B cells. Mixed bone marrow chimeras revealed that Itch acts within B cells to skew the proportions of light zone and dark zone GC B cells. Proteomic profiling of GC B cells uncovered that Itch deficient cells exhibit hallmarks of positive selection. These results support a novel role for Itch in limiting positive selection of GC B cells to restrict GC size. In this role, Itch may shape the GC-derived antibody repertoire to prevent autoimmunity.
Lessons from humanized mice: Is Foxp3 a bona fide marker of human regulatory T cells?
Authors: Morina Lyra , Ward Kirstin , Shevach Ethan M , Buszko Maja 
Published: February 2023

Abstract
Regulatory T cells (Tregs) suppress conventional T cells (Tconv) and are critical for homeostasis of the immune system. Tregs are identified by the transcription factor Foxp3 and, to a large extent, by the transcription factor Helios (expressed by 80% of human Tregs). One confounding factor in the study of human Tregs is that Tconv cells upregulate Foxp3 expression upon activation in vitro. However, it remains unclear whether this is the case in vivo. To investigate differences in phenotype between Tregs and activated Tconv cells, peripheral blood mononuclear cells (PBMCs) from healthy human donors were activated in vitro and in vivo and analyzed for expression of activation markers and cytokines using flow cytometry. For in vitro activation, PBMCs were stimulated in xenogeneic reactions with mouse splenocytes or with anti-CD3. For in vivo activation, human PBMC-engrafted NOD-scid IL2Rgnull (NSG) mice were used. NSG mice develop xenogeneic graft-versus-host disease (GVHD) as human cells begin to attack mouse tissues. During in vitro activation, there was an increase in Foxp3 expression by IL-2 producing, Helios− Tconv cells. On the contrary, during the course of GVHD in vivo, activated Tconv cells, identified as CD25+ and Foxp3−, produced IL-2 but remained Foxp3− and Helios−. Foxp3+ Tregs did not produce IL-2 and remained predominantly Helios+. In conclusion, while expression of Foxp3 is not a reliable marker of activated human Tregs in vitro, it appears to be a specific marker of activated human Tregs in vivo as observed in this humanized mouse model. Using Foxp3 to distinguish human Tregs can improve diagnosis and treatment for autoimmune disorders and cancer.
Critical role for adhesion and degranulation promoting adaptor protein in homeostasis and function of invariant natural killer T cells
Authors: Das Rupali , Bahal Devika , Gohl Trevor , Lee Hyun Hee 
Published: February 2023

Abstract
Invariant natural killer T cells (iNKTs) are innate T lymphocytes that are critical in modulating immune response in cancer, infection and inflammation. However, the mechanisms that regulate iNKT cell development and function are not fully known. In the current study, we demonstrate that adhesion and degranulation promoting adaptor protein (ADAP) is dispensable for iNKT cell thymic development but is required for their peripheral maintenance. Strikingly, ADAP-deficient (Adap−/−) mice have significantly reduced iNKTs in the periphery that is associated with decreased surface expression of the critical homing receptor (CXCR6), homeostatic proliferation and increased apoptosis. Consistently, Adap−/− iNKTs have markedly reduced expression of pro-survival factors (PLZF, ICOS, GATA-3 and Id2) but increased levels of pro-apoptotic molecules (Bax, Bad and Bim). Functionally, Adap−/− iNKTs have significantly reduced cytokine production and bystander immune cell activation as well as poor anti-tumor response, both in vitro and in vivo. In agreement with these findings, ADAP-silenced human iNKTs have remarkably impaired cytokine production and cytotoxicity against leukemia targets in vitro. Mechanistically, preformed transcripts for interleukin (IL)-4 and IFN-g as well as several microRNAs are reduced in Adap−/− iNKTs. Additionally, these cells exhibit reduced capacity to form conjugates and have diminished actin polymerization at the immunological synapse. Collectively, our findings establish a novel and pivotal role for ADAP in iNKT cell homeostasis and function.
Vitamin C Potentiates Plasma Cell Differentiation via TET-mediated DNA Modifications at a Novel Prdm1 Distal Element
Authors: Chen Heng-Yi , Alamonte-Loya Ana , Lay Fang-Yun , Johnson Eric , Gonzalez-Avalos Edahi , Yin Jieyun , Wozniak Daniel J. , Ghoneim Hazem , Harrison Fiona , Lio Chan-Wang Jerry 
Published: February 2023

Abstract
Ascorbate (vitamin C) is an essential micronutrient in humans. Chronic severe ascorbate deficiency results in a condition termed Scurvy with increased susceptibility to infections. How ascorbate affects the immune system at the cellular and molecular levels remained unclear. In this study, we showed that ascorbate is pivotal for the IL-21-dependent differentiation of BLIMP1+ plasma cells. Ascorbate significantly increases the proportion of B cells expressing BLIMP1 upon IL-21 stimulation without affecting the IL-21-STAT3 signaling. As a co-factor for TET DNA 5-methylcytosine oxidases, ascorbate boosters the enzymatic activity of TET, promoting the oxidation of 5-methylcytosines into 5-hydroxymethylcytosine (5hmC), an intermediate for DNA demethylation. Correspondingly, TET2/3 are required for the enhancement of plasma cell differentiation by ascorbate. Using 5hmC enrichment analysis, we identified ascorbate promotes plasma cell lineage via a novel distal element at the BLIMP1/Prdm1 locus. This ascorbate-responsive element (EAR) is normally methylated and is hydroxymethylated in the presence of ascorbate. Notably, the 5hmC-modified CpG exists within the STAT3 binding motif and the pattern is evolutionarily conserved, suggesting the ascorbate-assisted DNA modification is required for STAT3 recruitment to EAR and the subsequent Prdm1 activation. This study provides an example of how micronutrients may influence immune response via epigenetic regulation. Our finding also implies that epigenetic enzymes can function as sensors to gauge the availability of metabolites and influence cell-fate decisions.
Tonic sympathetic nervous system signaling regulates the maturation of natural killer cells
Authors: Geyer Rory , Moussa Marmar , Mandoiu Ion I. , Srivastava Pramod K. , Nevin James T. 
Published: February 2023

Abstract
Natural killer (NK) cells play a critical role in immune responses to certain viral infections and cancers by directly killing infected or transformed cells and producing cytokines that may enhance the function of other immune cells. NK cell effector function depends on the acquisition of a variety of activating and inhibitory receptors on their surface. NK cells that have acquired the full complement of these surface receptors are considered “mature.” While maturation of NK cells is essential for their proper function, knowledge of specific signals that contribute to their maturation is lacking. Here, we find that one such signal comes from the sympathetic nervous system (SNS). Single-cell RNA-sequencing and flow cytometric characterizations of mice injected with the sympatholytic compound 6-hydroxydopamine (6-OHDA) reveal that they have reduced frequencies of mature CD27−CD11b+ NK cells and increased frequencies of their immature precursors, CD27+CD11b− and CD27+CD11b+ NK cells in the spleen and bone marrow. Similarly, NK cell recovery after depletion is altered by SNS ablation, with the NK cells that recover in 6-OHDA-treated mice being unable to reach full maturity. SNS ablation also leads to accelerated growth of RMA/S tumors, the rejection of which depends on NK cells. The data presented here, along with further characterization of the effects of SNS signaling on NK cells will enhance our understanding of how the nervous system and immune system communicate with each other and how this communication influences immune responses to infection and cancer.