bluuebunny/crossref_metadata_2025_split huggingface dataset:
Selective Recruitment of Lethal Pro-inflammatory Macrophages in Sepsis by MIF but not D-DT (MIF-2)
Authors: Tilstam Pathricia Veronica , Schulte Wibke , Holowka Thomas , Kim Bong-Sung , Piecychna Marta , Pantouris Georgios , Lolis Elias , Leng Lin , Bernhagen Juergen , Bucala Richard 
Published: January 2023

Abstract
Sepsis is a leading cause of mortality with an estimated global incidence of 30 million cases annually. Current paradigms emphasize the role of an overwhelming or dysregulated inflammatory response in sepsis lethality, but to date interventions aimed at particular inflammatory pathways have been unsuccessful. Macrophage migration inhibitory factor (MIF) is of interest because MIF promoter polymorphisms predict mortality in different infections and anti-MIF antibody improves survival in experimental models when administered eight hours after infectious insult. The recent description of a second MIF superfamily member, D-dopachrome tautomerase (D-DT/MIF-2), prompts closer investigation of MIF-dependent responses. This study provides new insight into the pathophysiology of polymicrobial sepsis by revealing a direct role for MIF, but not its close structural homolog D-DT/MIF-2, in activating and recruiting a lethal macrophage subset. We subjected Mif −/− and Mif-2−/− mice to polymicrobial sepsis and observed a survival benefit with Mif but not Mif-2 deficiency. Survival was associated with lower levels of small peritoneal macrophages (SPMs) that, in contrast to large peritoneal macrophages (LPMs), are recruited to site of peritoneal inflammation during sepsis. LPMs produce higher quantities of MIF than SPMs, but SPMs express higher levels of inflammatory cytokines and MIF receptors CD74 and CXCR2. Adoptive transfer of SPMs into Mif −/− hosts abrogated the protective effect of Mif deficiency. Notably, MIF-2 lacks the pseudo-(E)LR motif present in MIF that mediates CXCR2 engagement and SPM migration, supporting MIF’s selective role in the recruitment and accumulation of lethal inflammatory SPMs.
A critical role for IL-37 in regulatory T cell maintenance and CD4+ T cell inhibition
Authors: Osborne Douglas Grant , Domenico Joanne , Dinarello Charles A , Fujita Mayumi 
Published: January 2023

Abstract
Regulatory T (Treg) cells express the transcription factor Foxp3 and play a vital role in suppressing immune responses and maintaining tolerance and homeostasis. Foxp3 expression is induced and maintained in Treg cells during thymic development and adaptive immune insults through three signaling pathways including TCR signaling and costimulation, IL-2 signaling, and TGF-β signaling, as well as Foxp3 expression itself. Recent studies, however, have suggested that these signaling pathways involved in the induction and maintenance of Foxp3 expression in the thymus and during activation are more dispensable in a steady state. To identify additional intracellular signaling molecules that induce/maintain Foxp3 expression and support Treg cell homeostasis and stability, we have screened proteins interacting with the three Foxp3 transcription factors NFAT, Stat5 and Smad3 for their expression in Treg cells and regulation of Foxp3 transcription. Using this protein screen we discovered IL-1 family member IL-37 plays a significant role in Foxp3 expression. By using human T cells from healthy donors and transgenic mice expressing IL-37, we uncovered a novel role for endogenous (expressed) and extracellular (secreted) IL-37 in Foxp3 expression and CD4+ T cells immunity. Treg cells that expressed IL-37 had elevated Foxp3 expression and were more suppressive in vitro and in vivo compared to Treg cells lacking IL-37. Extracellular IL-37, secreted by Treg cells, directly inhibited the canonical T cell signaling pathways through surface receptors. These results characterize an important mechanism for Treg directed immune suppression and Foxp3 expression.
Development of a high sensitivity 10-plex human cytokine assay using Simoa Planar Array technology
Authors: Lambert Jeremy , Zarozinski Christopher , Johnson Joseph , Yan Yu-xin , Rissin David , Stadler Patrizia , Ball Andrew 
Published: January 2023

Abstract
Approaches to quantify multiple cytokines have potential to improve understanding of mechanisms and progression of disease, including cancer pathogenesis. However, in healthy humans many cytokines are present at levels below the limits of quantification for established multiplex assays.
We developed the Simoa Planar Array to enable protein quantification in biological samples with greater sensitivity than traditional ELISAs. In this study we applied Simoa Planar technology to simultaneous multiplex detection of cytokines, using minimal sample volumes, to achieve sensitivity greater than established multiplex immunoassays.
Capture antibodies were printed in a circular pattern in microwells, with biotinylated detector antibodies and chemiluminescent detection used in a sandwich format. Antibody pairs were screened for ability to quantify analytes with minimal cross-reactivity. Pairs targeting IFNγ, IL-1β, IL-4, IL-5, IL-6, IL-8, IL-10, IL-12p70, IL-22 and TNFα were chosen, then assay conditions optimized to minimize matrix interferences. Analytical validation was performed, and circulating cytokine levels measured in normal human serum and plasma.
12.5 μL of human serum/plasma provided sufficient sample volume per replicate. Sub-picomolar LOD was achieved for all analytes; LLOQs ranged from 0.1 – 1.4 pg/mL. In normal human serum and plasma, &gt; 80% normal samples were quantifiable (above LLOQ) for most analytes. Analytical validation confirmed sensitivity, precision, specificity and dilution linearity.
The 10-plex Simoa Planar assay described here combines ultra-sensitivity with powerful multiplexing, enabling simultaneous detection of 10 cytokines in normal biological samples with superior sensitivity.
Bone Morphogenic Protein Receptor 1α and its ligands as a signaling circuit modulating immune response
Authors: Kraj Piotr , Kuczma Michal , Simms Colin , Pietrzak Maciej , Ignatowicz Leszek , Muranski Pawel , Browning Lauren 
Published: January 2023

Abstract
Despite increased understanding of how the transforming growth factor-β (TGF-β) regulates T cell functions, the immunomodulatory roles of other members of the TGF-β cytokine family, especially bone morphogenetic proteins, remain largely unknown. We have found that Bone Morphogenic Protein Receptor 1α (BMPR1α, Alk-3) expressed by activated effector and regulatory CD4+ T cells, modulates functions of both of these cell types by promoting generation of adaptive TR and inhibiting generation of Th17 cells from naive CD4+ T cells. Mice where BMPR1α is deleted in T cells (BMPR1αT− mice) had a decreased proportion of thymic derived TR cells. Activation of BMPR1α deficient (BMPR1α−) CD4+ T cells leads to generation of Th1/Th17-like effector cells expressing high levels of inflammatory cytokines, including IFN-γ, IL-17 and TNF family proteins. Effector CD4+ T cells activated by dendritic cells expressing BMP inhibitors expressed lower levels of PD1. Immunization of BMPR1αT− mice induced vigorous inflammatory response and mice were able to better control B16 melanoma tumors. Tumor infiltrate had very few TR cells and higher proportion of CD8+ T cells compared to tumors in wild type mice. These data demonstrate that TGF-β mediated immunosuppression in the course of tumor growth can be bypassed by inhibition of BMPR1α signaling.
Transcriptome analysis of wild type and BMPR1α− CD4+ Th cells revealed differential expression of transcription factors, cytokines and cytokine receptors and signaling molecules essential to establish regulatory networks supporting Th17 or TR cells. This suggests that BMPR1α is a potential target to augment effector Th cell responses.
Development of group-3 innate lymphoid cells via Tox2-dependent pathway in gut
Authors: Das Arundhoti , Wang Yueqiang , Lu Xiaoxiao , Harly Christelle , Kaye Jonathan , Bhandoola Avinash 
Published: January 2023

Abstract
Innate lymphoid cells (ILCs) are immune cells that lack specific antigen receptors but possess similar effector functions as T cells. Interestingly, ILCs and T cells express many of the same transcription factors. One such factor is TOX, a sequence non-specific nuclear transcriptional regulator that is crucial for development of both CD4 T cells and ILCs. TOX-deficient mice have significantly reduced number of early innate lymphoid progenitors (EILP) and ILC progenitors (ILCP). Consequently, the mature NK cells, ILC1, ILC2 and ILC3 subsets are greatly reduced in Tox−/− mice. Interestingly ILC3 in gut are not affected by the absence of TOX. We have discovered that TOX2, another member of the TOX family of HMG-box proteins, is highly expressed in ILC precursors. We generated Tox2−/− mice and found they have reduced ILC3 in gut; however, ILC3 cells in spleen or lymph node are intact. We also observed putative ILCP in fetal gut were significantly reduced in Tox2−/− mice. Hence our results indicate the existence of at least two pathways of ILC development: a central pathway in bone marrow and fetal liver that is TOX dependent; and a peripheral pathway of development of ILC3 in gut that requires TOX2.
Additionally, we observed that TOX2 is not required for development of T cells. However, Tox2−/− mice showed higher frequency of PD1 expressing CD8 effector memory cells. Tox2−/− mice also showed greater Th1 responses in vitro. Together, our results suggest that TOX is important centrally, for T cells and ILCs; whereas TOX2 is important for peripheral ILC3 generation or maintenance, and may further play an important role in effector T cells in tissues.
Distinct CD28 signaling requirements determine the alternative fates of thymic agonist selection
Authors: Watanabe Masashi , Alkhaleel Farrah , Hodes Richard J 
Published: January 2023

Abstract
During thymic T cell development, T cells strongly reactive to self-Ag are eliminated from the conventional T cell repertoire by clonal deletion. Alternatively, however, a number of T cell subsets are positively selected by strong recognition of self-Ag/MHC by TCR in thymus, termed agonist selection. Those T cell subsets include 1) Foxp3+ Treg cells, 2) invariant NKT (iNKT) cells, and 3) αβTCR+ PD-1+ CD4− CD8− precursors of CD8αα IELs (pIEL). In addition to a strong TCR signal, B7-CD28 co-stimulation has a role in these agonist selections. This role is context dependent. For instance, B7-CD28 has a positive role for thymic Treg and iNKT cell generation, while it has a negative role for thymic pIEL generation. These differences may indicate involvement of distinct CD28 signaling in these events. To test this hypothesis, we analyzed CD28 cytoplasmic tail mutant mice in which tyrosine 170 residue was substituted to phenylalanine (Y170F), proline residues in PYAP motif were substituted to alanine (AYAA), or both mutation were present (Y170F/AYAA). Generation of self-Ag-specific Treg, defined by pMHCII tetramer binding, was attenuated in AYAA mice but not in Y170F mice, consistent with the reported role of the PYAP motif for total Treg generation. Meanwhile, interestingly, the negative role of CD28 for self-Ag-specific pIEL generation was not affected by Y170F and/or AYAA substitution, indicating that this CD28 effect was mediated by motif(s) other than Y170 and PYAP. Analysis of thymic iNKT cell (NKT1, 2 and 17) development in CD28 mutant mice is currently in progress. These results suggest that the differing roles of B7-CD28 costimulation for agonist selection fates are mediated via qualitatively distinct CD28 signaling.
Regulation of thymic type I IFN expression and its role in T cell development
Authors: Salgado Barrero Oscar C , Lee Seungeun Thera , Breed Elise R , Hogquist Kristin Ann 
Published: January 2023

Abstract
Thymocytes undergo a complex maturation process after being positively selected. Recently, the involvement of type I IFN signaling during these late stages was suggested; however, the regulation of intra-thymic type I IFN production and its role in T cell development and central tolerance are relatively unknown. We found that type I IFN, especially IFN-beta, regulate the expression of the maturation marker Qa2, the transcription factor STAT1 and the cell surface protein Ly6A in mature thymocytes. Using a published luciferase reporter mouse strain, Ifnbluc, we found that IFN-beta is expressed in the thymus in the steady state, and its levels peak at around 2–3 weeks of age, when the expression quickly returns to basal levels. The cell source of IFN-beta was mainly medullary thymic epithelial cells (mTEC) expressing high levels of MHC-II and CD80. We showed that IFN-beta expression on mTEC was completely dependent on AIRE and partially dependent on RANKL and CD40 signaling pathways. Additionally, we evaluated the impact of thymic IFN-beta expression on T cell tolerance. Our data suggest that T cells from mice lacking IFN-beta or AIRE show a higher reactivity to IFN-stimulated antigen presenting cells compared to T cells from wild-type mice. We hypothesize that thymic type I IFN provide a mechanism to promote T cell tolerance to IFN-induced peptides.
In vitro modeling of human B cell development can identify distinct physiopathological patterns in primary antibody deficiency (PAD)
Authors: Troilo Arianna , Wehr Claudia , Janowska Iga , Rawluk Justyna , Thiel Jens , Venhoff Nils , Herget Georg , Ochs Björn Gunnar , Konstantinidis Lukas , Warnatz Klaus , Salzer Ulrich , Rizzi Marta 
Published: January 2023

Abstract
Bone marrow (BM) analysis show that 20% of PAD patients harbor a block at an early stage of B cell development. The aim of this work is to study the mechanisms of this developmental block. We set up a feeder free cultivation system that in healthy donors leads to the development of lymphocyte progenitors until the stage of Immature B cells.
BM derived CD34+ cells are expanded in cytokine cocktail for 2 weeks. From day 14 to day 49 cells are cultivated in cytokine-free medium and analyzed once a week by flow cytometry and RNA is collected for PCR assays.
BM derived CD34+ cells from 15 PAD patients and 2 Btk-deficient patients were tested in vitro for their capability to develop into IgM+ Immature B cells. CD34+ cells from patients showing a normal B cell development in vivo (6/17) developed in vitro until the stage of Immature B cells. Among PAD patients presenting a block at early B cell stage, 5/11, including the Btk-deficient patients, could not reach the Immature B cell stage, while 6/11 could develop in vitro until the stage of IgM+ B cells. Among this last group, reaching Immature B cell stage in vitro but not in vivo, 2/6 patients showed a rapid in vitro development of Immature B cells at day 14, but presented a progressive exhaustion of the common lymphoid progenitors (CLP) compartment. In patients with a block in B cell development (in vivo and in vitro), PCR analysis revealed adelayed expression of transcription factors E2A, CD79a and PAX5, reflecting an impaired lineage commitment.
Combining BM phenotyping and in vitro modeling we identified four patients’ groups characterized by (1) an intrinsic B cell defect, (2) a defect in the BM microenvironment impairing early stages of B cell development, (3) a defect in repopulation of CLP and (4) normal B cell development.
TRAF3IP3 at the trans-Golgi network regulates NKT2 maturation via MEK/ERK signaling pathway
Authors: Ge Qing , Zhang Xinwei 
Published: January 2023

Abstract
Thymic natural killer T (NKT)2 cells are a subset of invariant NKT cells with PLZFhiGATA3hiIL-4+. The differentiation of NKT2 cells is not fully understood. We report in the present study an important role of TRAF3-interacting protein 3 (TRAF3IP3) in the functional maturation and expansion of committed NKT2s in thymic medulla. Mice with T cell-specific deletion of TRAF3IP3 had decreased thymic NKT2 cells, decreased IL-4 producing peripheral iNKTs, and defects in response to α-galactosylceramide. Positive selection and high PLZF expression in CD24+CD44− and CCR7+CD44− immature iNKTs were not affected. Only CD44hiNK1.1− iNKTs in Traf3ip3−/− mice showed reduced expression of Egr2, PLZF, IL-17RB, decreased proliferation, and decreased IL-4 production upon stimulation. Such Egr2 and IL-4 expression were augmented by MEK1/ERK activation in iNKTs and TRAF3IP3 at the trans-Golgi network recruited MEK1 and facilitated ERK phosphorylation and nuclear translocation. These data demonstrate an important functional maturation process in NKT2 differentiation that is regulated by MEK/ERK signaling at the trans-Golgi network.
Selective Serotonin Reuptake Inhibitors Suppress Mast Cell Function
Authors: Haque Tamara , Ryan John 
Published: January 2023

Abstract
Mast cells are tissue resident innate immune cells activated by IgE and many other ligands, including purines. Upon activation, they release histamine, cytokines, chemokines, proteases, and lipid mediators evoking allergic symptoms. New ways of targeting mast cells could greatly benefit allergic disease therapy. One plausible target is activation by purinergic receptors binding ligands such as ATP. However, there are currently no clinically available therapies targeting purinergic signaling. Because anti-depressants in the SSRI family can have off-target effects on purinergic receptors, we tested the hypothesis that SSRIs can attenuate mast cell function. We found that the SSRI fluoxetine blocks IgE- and ATP-induced cytokine production, degranulation, and calcium influx in murine bone marrow derived mast cells (BMMC). Furthermore, fluoxetine suppresses IgE-mediated anaphylaxis in vivo, and reduces ATP cellular uptake. Because adding serotonin to BMMC or deleting the serotonin transporter has no effect on fluoxetine-mediated suppression, an off-target effect seemed likely. We show that inhibitors of the ATP receptor P2X3 attenuate IgE-mediated activation similarly to fluoxetine, and combining fluoxetine with these inhibitors does not result in additive effects. Using siRNAs to knock down P2X3, we find that P2X3 is needed for suppression by fluoxetine. These data support the possibility of targeting purinergic signaling to treat allergic diseases, and suggest that this may be possible by repurposing FDA-approved SSRIs.
The HMGCR Inhibitor Fluvastatin Induces Apoptosis and Autophagy in Primary and Neoplastic Mast Cells
Authors: Barnstein Brian O , Paez Patrick A , Dailey Jordan M. , Finnegan Ryan M. , Ryan John J 
Published: January 2023

Abstract
Mast cell hyperplasia is noted in many inflammatory and allergic diseases. Mast cells can also generate a subtype of acute myelogenous leukemia. Understanding and treating mast cell hyperplasia and leukemia could be beneficial to both inflammatory and malignant diseases. Recently the family of statin drugs, widely employed as HMG CoA Reductase (HMGCR) inhibitors to lower serum cholesterol, have been found to be immunosuppressive. However, statin effects on mast cell survival are not known. We report that fluvastatin induces apoptosis in mouse bone marrow-derived mast cells (BMMC) and mastocytoma cells. Apoptosis was suppressed in p53 KO and Bcl-2 Tg BMMC, consistent with detectable mitochondrial damage and Caspase-9 activation. Because apoptosis induction could be mimicked by suppressing geranylgeranyl transferase (GGT), and fluvastatin effects were reversed by adding the product of the GGT reaction, loss of GGT-mediated protein lipidation is the likely mechanism of cell death. In addition to apoptosis, fluvastatin also induced autophagy. Chemical blockade of autophagy enhanced fluvastatin-mediated apoptosis in primary mast cells, but reduced it in mastocytomas. These data suggest that the FDA-approved statin drugs may be useful in suppressing mast cell hyperplasia and leukemia.
Comparing mast cell immunometabolism shifts induced by IgE mediated and non-IgE mediated degranulation
Authors: Mendoza Ryan P , Anderson Colin C , Roede James R , Brown Jared M 
Published: January 2023

Abstract
Mast cells are immune effector cells with the ability to immediately release an array of pre-formed mediators in response to various stimuli, a process termed degranulation. Degranulation is generally divided into two categories: IgE mediated and non-IgE mediated. Non-IgE mast cell degranulation occurs in response to various environmental agents without prior IgE sensitization often making it difficult to identify the causative agent in patients. To better understand non-IgE mast cell degranulation, we characterized and compared metabolic shifts in response to both mechanisms of degranulation (IgE vs non-IgE) in bone marrow-derived mast cells (BMMCs) grown from C57BL/6 mice. Compound 48/80 and silver nanoparticles were used to trigger non-IgE degranulation. To explore metabolic changes in BMMCs, we used Seahorse XF technology to measure: 1) respiratory mitochondrial metabolism, 2) anaerobic glycolytic metabolism, and 3) performed a phenotype stress test to define metabolic pathways. The mito stress test revealed a decrease in respiration for both mechanisms of degranulation but was consistently lower for non-IgE triggers. Secondly, we observed an increase in glycolysis from both mechanisms which was more prominent in non-IgE degranulation. However, there was a complete depletion of glycolytic reserve with non-IgE degranulation that did not occur with IgE mediated degranulation. The cell phenotype test revealed that BMMCs shift towards glycolysis when activated via a non-IgE pathway more prominently than via an IgE-pathway. In conclusion, mast cell metabolism varies significantly between IgE and non-IgE degranulation with an apparent shift towards glycolytic dependence for non-IgE activated mast cells.
Perinatal androgens drive sex differences in mast cell phenotype and severity of mast cell disease
Authors: Mackey Emily , Moeser Adam J. 
Published: January 2023

Abstract
Highly prevalent and debilitating mast cell (MC)-associated immune disorders, such as allergy and irritable bowel syndrome exhibit a female sex bias in childhood and adulthood. The mechanisms underlying sex differences in MC disorders are unknown. Previously, we found adult female mice exhibited greater serum histamine and more severe MC-associated pathophysiology that coincided with an increased capacity to synthesize and store immune mediators, compared with males. Further, recent data showed sex differences in the MC exist prior to puberty, suggesting a mechanism independent of adult sex hormones. Here, we investigated the role of perinatal androgens, a key early life component of sexual differentiation, in sex differences in MC phenotype and MC disease. Testosterone proprionate (100 μg sc qd) or vehicle was given to pregnant female mice E16-P0 and directly to pups PN1-7. IgE-mediated anaphylaxis was performed in gonadectomized control male (CM), control female (CF) and perinatally androgenized female (AF) offspring at 8 wk of age. Serum histamine levels (CM 768 ± 106, CF 1406 ± 200, AF 676 ± 110 ng/mL) and hypothermia responses (Δ temp: CM −4.87 ± 0.45, CF − 7.28 ± 0.48, AF − 4.8 ± 0.39 °C) were ~2x lower in AF than CF mice, while AF and CM mice had similar responses. Further, bone marrow MC from AF mice had lower histamine content (AF 1305 ± 82: CF 2082 ± 94 ng/106cells) and 47% lower histamine release to IgE-FcɛRI stimulus compared to CF mice. Together, we demonstrate that sex differences in MC phenotype and disease susceptibility are established early in life by perinatal androgens. These findings provide insight into the mechanisms of sex differences in MC-associated immune disorders that could lead to new therapeutic targets for MC diseases.