bluuebunny/crossref_metadata_2025_split huggingface dataset:
Modulation of mast cell function by TIM-3 signaling (HYP3P.351)
Authors: Phong Binh , Kane Lawrence 
Published: January 2023

Abstract
Transmembrane, immunoglobulin and mucin domain-3 (Tim-3) is an inducible negative regulator of Th1 function and anti-tumor immune responses. Recent studies found that Tim-3 is constitutively expressed on mast cells and that an anti-Tim-3 pAb enhanced secretion of pro-inflammatory cytokines by antigen-stimulated bone marrow-derived mast cells (BMMC). However, the mechanism(s) by which Tim-3 modulates mast cell function are unknown. Here we show that Tim-3 enhances FcεRI-derived signals to co-stimulate NF-AT, AP1, NF-κB, and IL-6 promoter transcriptional activation. This activity requires specific tyrosines in the Tim-3 cytoplasmic tail. Furthermore, various antibodies to Tim-3 significantly enhanced IgE+Ag-stimulated tyrosine phosphorylation and up-regulated BMMC cytokine secretion. Conversely, we observed impaired IgE+Ag-dependent activation of mast cells in which Tim-3 expression was knocked down. Using BMMC generated from Nur77-GFP reporter mice, we found that cross-linking Tim-3 synergizes with the antigen receptor FcεRI to enhance Nur77-GFP expression. Our work suggests that Tim-3 acts proximal to the antigen receptor, leading to enhancement of the NF-κB, NF-AT, and AP-1 pathways. Thus, these studies are consistent with Tim-3 being a positive regulator of mast cell function. Mast cells contribute to both innate and adaptive immune responses, so Tim-3 may serve as an important modulator of mast cell effector functions in autoimmune and allergic diseases.
Effect of NADPH oxidase 2 (Nox2) deficiency on the function of Th2 cells (HYP4P.415)
Authors: Yuk Chae Min , Kwon Bo-In , Lee Seung-Hyo 
Published: January 2023

Abstract
Nox2/gp91phox, one of phagocyte NADPH oxidase subunits, is mainly expressed on phagocytes. They transport electrons across the plasma membranes and kill pathogens by producing ROS. Many evidences have shown that Nox2 deficiency causes susceptibility to infections and immunosuppression. Recent studies have identified functional phagocyte-type Nox2 is also expressed in helper T cells through TCR stimulation. So, many papers attempted to define the role of Nox2 on Th1 and Th17 cell differentiation, but the exact function of Nox2 in Th2 differentiation remains unclear. To elucidate this, we adopted experimental allergic asthma that Th2 cells have been considered as a pivotal mediator for asthma pathogenesis. Here, we revealed that Nox2 deficient mice showed enhanced allergic asthma phenotypes including increased airway hyper responsiveness, eosinophilia, glycoprotein secretion and IL-4 production in bronchoalveolar lavage through augmented Th2 differentiation, regardless of challenge routes and allergen types. Based on our results, we conclude that Nox2 could be a molecular target for controlling Th2 cell differentiation and allergic asthma.
Altered neutrophil homeostasis and DARC regulate CXCL5-mediated pulmonary immune responses (INM2P.435)
Authors: Mei Junjie , Dai Ning , Liu Yuhong , Hudock Kristin , Liu Peihui , Deshmukh Hitesh , Lee Janet , Worthen G. Scott 
Published: January 2023

Abstract
CXCL5 is a neutrophil chemokine, predominantly expressed by epithelial cells and platelets. We recently demonstrated the critical role of CXCL5 in chemokine scavenging and innate immunity to severe bacterial pneumonia, and in regulating neutrophil homeostasis through IL-17/G-CSF axis. Here we show that CXCL5-inhibited chemokine scavenging and innate immune response to severe E.coli pneumonia is completely dependent on G-CSF-mediated increases in BM neutrophils and CXCR2 concentrations, but not on DARC. However, DARC functions as a chemokine reservoir rather than a sink in this pneumonia model. Darc-/- mice showed increased neutrophils in the BM, with normal blood neutrophil counts under basal conditions, and defective neutrophil release from BM upon systemic CXCL2 stimulation. Furthermore, DARC deficiency altered neutrophil homeostasis independently of G-CSF and exhibited additive effect with CXCL5 deficiency on increasing BM neutrophil percentages in Cxcl5-/-Darc-/- mice. These data provide the first evidence for a beneficial role of delicate regulation of neutrphil homeostasis in chemokine scavenging and innate immunity to bacterial pneumonia. Moreover, these results substantiate a mechanistic role for DARC deficiency in neutrophil homeostasis, which may provide insight into benign ethnic neutropenia and other inflammatory diseases seen in Duffy null humans.
Vagal modulation of catecholamines in the adrenal gland controls systemic inflammation in experimental sepsis (INM2P.437)
Authors: Ulloa Luis , Inclan Rico Juan Manuel , Morcillo Patrick , Peña Geber , Torres Rosas Rafael 
Published: January 2023

Abstract
Sepsis is the leading cause of mortality in non-coronary Intensive Care Units accounting for 9.3% of overall deaths in the USA. Sepsis and other critical conditions like hemorrhage, resuscitation, shock, and trauma are characterized by overzealous inflammatory responses that cause lethal multiple organ failure. Despite its recent identification, multiple investigators have already reported that the vagus nerve controls systemic inflammation in experimental ischemia and reperfusion, hemorrhage and resuscitation, pancreatitis, colitis, endotoxemia, septic shock and severe sepsis. Vagus nerve stimulation controls systemic inflammation, but its clinical use is precluded by the surgical exposure of the vagus nerve, the use of anesthetics, and its temporal effect(1,2). Transcutaneous activation of the vagus nerve improves survival in experimental sepsis without surgery. Transcutaneous activation of the vagus nerve inhibits the production of multiple inflammatory cytokines including TNFα, IL6 and INFγ. This anti-inflammatory potential has a lasting effect and does not merely delay the pathogenesis of sepsis. Transcutaneous activation of the vagus nerve controls systemic inflammation by modulating catecholamine's production in the adrenal gland. Studies funded by NIH-GM084125 to LU. REFERENCES 1. Wang, H., et al. Nature medicine 10, 1216-1221 (2004). 2. Wang, H., et al. Nature 421, 384-388 (2003).
Galectin-12 in macrophage polarization (INM2P.425)
Authors: Wan Lei , Hsu Yu-An 
Published: January 2023

Abstract
Galectin-12 (Gal12) is a member of a family of animal lectins. Gal12 is expressed in adipocytes and it has been shown to play an important role in adipocyte differentiation. We found that Gal12 is expressed in macrophages and the ablation of Gal12 did not influence bone marrow cells to differentiate into macrophages. The ablation of Gal12 reduced the phagocytic activity against E. coli and lowered the secretion of nitro oxide. Gal12 KO demonstrated the polarization of macrophages to M2 macrophages, as indicated by the increases in the expression levels of FIZZ1 and Ym1 as well as the reduction in the expression levels of TNF-α, IL-6, IL-12, IL-1β, MCP-1, and CXCL1 pro-inflammatory cytokines. We also detected a higher expression level of IL-10, an anti-inflammatory cytokine, in Gal12 KO macrophages. Moreover, the activation of IKKα/β, Akt and ERK was lower in Gal12 KO macrophages, which subsequently reduced the activation of NF-κB and AP1, resulting in lower expression levels of pro-inflammatory cytokines. The activation of STAT1 and STAT6 was lower in Gal12 KO macrophages, indicating that Gal12 may modulate the activation of the IRF3 and subsequently lower the expression of type I interferon by LPS treatment. The activation of STAT3 was much higher in Gal12 KO macrophages activated by LPS, which indicated higher expression levels of IL-10 in Gal12 KO macrophages. Our data indicates Gal12 KO predisposed macrophage to polarize into M2 macrophage.
Nlrc4 inflammasome is dispensable in vitro for IL-1β production and caspase-1 activation upon infection by paracoccidioides brasiliensis. (INM3P.443)
Authors: Carneiro Natália , Rocha Fernanda , Silva Grace , Zamboni Dario , Silva João 
Published: January 2023

Abstract
The fungus Paracoccidioides brasiliensis (Pb), causative agent of chronic systemic mycosis endemic in rural areas of Latin America, is sensed by the TLR-2, 4 and dectin-1 receptors. However, during this infection, the upstream pathways responsible for the cytokines production by macrophages as IL-1β and IL-18 are completely unknown. In Nlrp3-/- macrophages (BMMs), although amounts of IL-1β were reduced, it is still detectable when compared with uninfected macrophages, suggesting that there are additional receptors required for caspase-1 activation and IL-1β production. Then, we evaluate if Nlrc4 could induce IL-1β production and caspase-1 activation in vitro. Strikingly, we observed higher IL-1β release and caspase-1 activation in Nlrc4-/- BMMs than WT cells. To characterize the role of Nlrc4 during experimental infection, WT and Nlrc4-/- mice were infected intravenously with 1x10^6 yeasts forms of Pb18 and fungal burdens in the lung were analyzed at day 15 and 30 post infection. Whereas at 15 dpi WT and Nlrc4-/- mice had similar fungal counts in the lung tissue, at 30 dpi the lung homogenates from Nlrc4-/- mice contained lower numbers of Pb colony-forming units. Moreover, we showed that the reduction in number of viable yeasts was followed by compact and well defined granulomas formation. Taken together, these results suggest that Nlrc4 mediates efficient granulomatous organization and host resistance to P. brasiliensis infection. Financial support: FAPESP, CNPq, CAPES
The nervous system modulates CD8+ T cell function through the beta2-adrenergic receptor (IRC2P.456)
Authors: Estrada Leonardo , Agac Didem , Farrar David 
Published: January 2023

Abstract
There is growing evidence to suggest that various aspects of immune function are controlled by neural networks. Both cholinergic and adrenergic receptors are expressed by various major immune cell types. Specifically, expression of the β2-adrenergic receptor (ADRB2) has been characterized in different cells of the immune system; however, our lab has demonstrated for the first time a differential expression of this receptor in subsets of effector and memory CD8+ T cells. We found that the ADRB2 is upregulated in human effector memory CD8+ T cells compared to central memory/naïve counterparts. Further, human CD8+ T cells that are acutely activated in the presence of the ADRB2 natural ligand, nor-epinephrine (NE), secrete significantly less IFN-γ and TNF-α. Similarly, the cytolytic capacity of these cells is reduced when NE is present. This downregulation by NE was reversed in the presence of a ADRB2-specific antagonist, but not a pan α-adrenergic or β1-adrenergic receptor specific antagonists. Additionally, mouse CD8+ T cells behaved similarly to human cells to these antagonists, suggesting species conservation of this neural-immune network. Moreover, ADRB2-KO CD8+ T cells were completely resistant to the effects of NE. Current experiments using an adoptive transfer model will elucidate how ADRB2 signaling modulates CD8+ T cell function in vivo. These experiments will provide insight into how the nervous system controls CD8+ T cell function during the course of an infection.
TCR strength of signal is upstream of and dominates over cytokine effects in controlling CD4+ T cell polarization in vivo (IRC2P.449)
Authors: van Panhuys Nicholas , Klauschen Frederick , Germain Ronald 
Published: January 2023

Abstract
Both quantitative antigenic signal strength and the qualitative effects of cytokine signaling have been reported to direct the polarization of CD4+ T cells into effectors with distinct patterns of cytokine secretion. Most current models of the polarization process attribute a dominant role to the effects of cytokines, with polarizing adjuvants presumed to work mainly through elicitation of the relevant cytokines. Here we have varied both the adjuvants used for DC activation and the strength of TCR signaling to examine whether quantitative or qualitative aspects of CD4+ T cell activation play the primary role in effector choice in vivo. Using two-photon intravital microscopy we compared DC stimulated with TH1 or TH2 promoting adjuvants and found that it is the duration of interaction and the strength of signal that dominates in controlling CD4+ T cell fate. Adjustments to antigen level that affect interaction duration can reverse the fate outcome irrespective of the adjuvant used to treat the DC, clearly pointing to the quantitative aspects of signaling as dominant over qualitative effects arising from adjuvant-induced cytokine production. Indicating that cytokines operate as a secondary step in a developmental system in which the strength of initial CD4+ T cell signaling regulates the capacity of the cells to respond to cytokine mediators.
Interplay of phosphorylation kinetics and methylation dynamics leads to complex behavior of Foxp3 and PTEN in computational model of T cell differentiation (IRM6P.709)
Authors: Miskov-Zivanov Natasa , Hawse WIlliam , Morel Penelope , Faeder James 
Published: January 2023

Abstract
We have recently published a mathematical model that demonstrates that differential activation of signaling pathways downstream of TCR by high and low concentrations of antigen can be modeled by varying the duration of antigen stimulation, which results in a mixed population of regulatory (Treg), helper (Th) and inactive cells, a prediction we confirmed experimentally. Analysis of the dynamics of this scenario demonstrated an important role for a feedback loop involving Foxp3, PTEN, and Akt-mTOR signaling, which affects the timing of transient Foxp3 expression and PTEN activity, and the ratio of different phenotypes in the mixed population. We hypothesized that the correlation in the dynamics of Foxp3 and PTEN, which we previously modeled as a direct link between Foxp3 and PTEN, stems from the regulation of both Foxp3 and PTEN expression by the transcription factor Foxo1. We updated the model to test this hypothesis, and preliminary results suggest that the interplay of rapid modulation of Foxo1 activity by Akt with the slower kinetics of methylation/demethylation at the Foxp3 promoter mediated by mTOR can produce dramatic changes in the resulting distribution of phenotypes when the timing of these events is varied. To calibrate this timing in the model, we are currently conducting experiments that will help us determine the time scale for Foxo1 inactivation by Akt and for the effect of Foxo1 on Foxp3 and PTEN expression.
IL-17-mediated regulation of C/EBPβ alternative translation: Implications for signaling and response to opportunistic fungal infections (IRM6P.721)
Authors: Simpson-Abelson Michelle , Bishu Shrinivas , Childs Erin , Conti Heather , Gaffen Sarah 
Published: January 2023

Abstract
Th17 cells and IL-17 play essential host-defensive roles in immunity to pathogens, particularly fungi such as Candida albicans. However, the specific signaling pathways used by IL-17 and its receptor are still poorly defined. We have shown that IL-17 activates the CCAAT enhancer binding protein β (C/EBPβ) transcription factor, which is required for induction of many IL-17 target genes. C/EBPβ is translated into 3 distinct protein isoforms whose expression is regulated by the differential use of different translational initiation start sites, known as LAP* (38kDa), LAP (35kDa) and LIP (22kDa). We have shown that IL-17 induces the alternative translation of C/EBPβ, inducing a marked increase in LAP* and LIP. However, the functional significance of differential C/EBPβ isoform expression remains enigmatic. To determine the biological significance of C/EBPβ alternative translation, we used C/EBPβ-/- mice and a knockin that cannot generate the LAP isoform (C/EBPβM20A). We then assessed susceptibility of these mice to oral C. albicans infection, immunity to which is a strongly IL-17-dependent event. We found that C/EBPβ-/- but not C/EBPβM20 mice were highly susceptible to disease, indicating that LAP is dispensable but C/EBPβ is required. In evaluating a panel of downstream genes to elucidate mechanisms of C/EBPβ-dependent immunity, β-defensin 3 (BD3) was strongly downregulated in C/EBPβ-/- mice. These results highlight a key role for C/EBP in antifungal immunity mediated by IL-17.
MicroRNAs, T follicular helper cells, and inflammatory status during aging (IRM6P.714)
Authors: Hu Ruozhen , Kagele Dominique , Huffaker Thomas , Runtsch Marah , Alexander Margaret , Bake Erin , Su Wei , Rao Dinesh , Möller Thomas , Garden Gwenn , Round June , O'Connell Ryan 
Published: January 2023

Abstract
Chronic, low-grade inflammation is associated with many life-shortening human diseases that afflict the elderly. By understanding the molecular mechanisms underlying chronic inflammation effective treatments can be developed to limit such conditions as metabolic disorders, autoimmunity and cancer. Using a mouse model of age-dependent inflammation, we demonstrate that miR-155 is a promoter of disease symptoms in aging mice. Upon analyzing lymphocytes from inflamed versus healthy mice, we observed increased numbers of T follicular helper (Tfh) cells and germinal center (GC) B cells, correlating with increased production of autoantibodies as mice reach middle age. Interestingly, this inappropriate humoral response occurred in a miR-155-dependent manner. Next, we generated T cell-specific miR-155-/- mice and discovered a T-cell intrinsic role for miR-155 in promoting Tfh cell formation, GC B cell development, and production of antigen-specific IgG antibodies. To begin unraveling the mechanistic basis for this phenotype, we employed a pipeline involving RNA-seq, bioinformatics and experimental validation. This approach identified a subset of miR-155 target genes that are relevant to Tfh cell formation. Taken together, our study demonstrates that miR-155 promotes Tfh cells development during age-dependent chronic inflammation, and identifies miR-155 as a potential therapeutic target to diminish age-related inflammatory conditions.
PTEN regulates CD4+ T cell differentiation (IRM6P.716)
Authors: Hawse WIlliam , Miskov-Zivanov Natasa , Faeder James , Morel Penelope 
Published: January 2023

Abstract
Naïve CD4+ T cells differentiate into either helper (Th) or regulatory (Treg) cells following encounter with antigen. T cell fate is in part determined by signaling strength where lower TCR and AKT/mTOR signaling favors Treg differentiation and higher TCR and AKT/mTOR signaling drives Th differentiation. AKT/mTOR signaling is regulated by PTEN, a phosphatase that is highly expressed in Tregs. We recently published a computational model that suggested a critical role for PTEN in T cell differentiation. Our current work focuses on validating the model predictions and explaining how PTEN is regulated during T cell differentiation. We demonstrate that PTEN inhibition increases Akt/mTOR signaling leading to increased Th induction. When the TCR signal strength is high, PTEN is ubiquitinated and phosphorylated and PTEN protein levels are dramatically reduced. Conversely, PTEN levels only decrease transiently when TCR signal strength is low and the resulting Treg express higher PTEN levels. We are exploring the transcriptional regulation of PTEN. One potential regulator is FOXO1, which has been shown by ChIP-Seq to bind the PTEN promoter. High Akt/mTOR signaling results in phosphorylation of FOXO1 and exclusion of FOXO1 from the nucleus. Low Akt/mTOR signaling results in minimal phosphorylation and sustained FOXO1 nuclear localization. Thus, PTEN protein levels are tightly controlled by TCR signal strength and regulation of PTEN plays an important role in determining T cell fate.
Mechanisms of CD4+ T Cell Exhaustion by persisting antigen (LYM3P.737)
Authors: Lober Benedikt , Obst Reinhard 
Published: January 2023

Abstract
When faced with chronic antigen presentation T cells are functionally impaired over time. Since detailed studies of antigen-exhausted CD4+ T cells are scarce, we followed AND TCR transgenic T cells transferred into antigen-transgenic recipients where they received chronic TCR signals of varying strength. In one of the recipient lines, transient antigen expression regulated by doxycycline treatment allowed for the efficient differentiation of memory T cells, with IL7R, Ly6C and swift cytokine production as their hallmarks, while chronic TCR signals exhausted the cells. We found two kinds of exhaustion: Strong stimulation was accompanied by reversible TCR downregulation, signaling impairment and an increase of PD-1 and the anergy markers CD73 and FR4. When stimulated with PMA/Ionomycin, which circumvent TCR-proximal signaling components, these cells produced effector cytokines almost like memory T cells. In contrast, T cells that experienced a weaker TCR stimulus chronically showed no cytokine production following PMA/Ionomycin restimulation. Since in such T cells TCR signaling via the AKT, MAPK and Ca2+ pathways was found only partially impaired, we conclude that strong signals impair TCR-proximal signaling components while weaker signals affect more distal elements. We suggest that the degree of T cell exhaustion correlates with antigen strength, leading either to a fast TCR and signaling downmodulation or to an irreversible functional impairment.
Ndfip1 limits TH17 differentiation and proinflammatory cytokine production. (LYM3P.741)
Authors: Layman Awo A. , Oliver Paula 
Published: January 2023

Abstract
Ubiquitination and subsequent degradation of pathway proteins is an important way of terminating an effector cell response. Three classes of enzymes: E1 ubiquitin activating enzymes, E2 ubiquitin conjugating enzymes, and E3 ubiquitin ligases, work sequentially to ubiquitinate selected substrates. Ndfip1 (Nedd4-family interacting protein 1) is a protein that activates the catalytic function of several related E3 ubiquitin ligases. Previous work from our lab showed that mice lacking Ndfip1 have increased numbers of TH17 cells and that Ndfip1 is expressed in T cells. Thus, we hypothesized that Ndfip1 has a T cell-intrinsic role in limiting TH17 cell differentiation. Using a mixed chimera model, we show that T cells that lack Ndfip1 are much more likely to produce IL-17A and GM-CSF, two proinflammatory cytokines produced by TH17 cells. Additionally, Ndfip1-deficient T cells produce more IL-17A and GM-CSF when cultured under TH17 polarizing conditions. These data may help to explain why mice that lack Ndfip1 are more susceptible to IL-17-mediated pathology in a DSS colitis model, compared to their Ndfip1-sufficient counterparts. This work reveals a biologically relevant role for Ndfip1 in limiting TH17 differentiation and GM-CSF production, as well as associated inflammatory pathology. We are now working to identify components of Ndfip1-mediated ubiquitin complexes and determine the underlying mechanism by which Ndfip1 limits proinflammatory cytokine production by TH17 cells.