bluuebunny/crossref_metadata_2025_split huggingface dataset:
Inhibition of E protein activity is required for iNKT cell development and function (P1458)
Authors: Hu Taishan , Wang Hongcheng , Simmons Amie , Zhao Ying , Sun Xiao-hong , Alberola-Ila Jose 
Published: January 2023

Abstract
iNKT cells are a distinct subset of αβ T cells with diverse immunoregulatory functions. Like conventional T cells, iNKT cells arise in thymus from a common CD4+CD8+ double positive (DP) precursor, but these cells are positively selected by glycolipid-loaded CD1d. Our previous result showed that the RAS/MAPK cascade (RAS-Erk-Egr-ID3), a central player during conventional T cell positive selection, is also required for iNKT cell development. Since RAS/MAPK signaling cascade has been reported to promote T cell positive selection by antagonizing E2A activity, we hypothesize that the same mechanism might be applied to iNKT cell development. To test this hypothesis, we utilized a conditional knock-in mouse model expressing ET-2, a chimeric protein composed of the TAD domain of E47 and the bHLH domain of SCL/Tal. Functioning as a dominant negative mutant of ID, ET-2 dramatically impaired iNKT cell development and function. iNKT cells in ET-2 mice were blocked during transition of stage 2 to stage 3, and these cells were also defective in IFN-γ production. Further mechanistic dissection showed that ET-2 might block iNKT cell development through inhibition of T-bet, a transcriptional regulator required for iNKT cell terminal maturation and homeostasis by controlling CD122. Forced expression of T-bet rescued the defects in CD122 expression and IFN-γ production. These results further confirm the role of the RAS/MAPK cascade in iNKT cell development and function.
Using Zinc-finger technology to study transcriptional regulation of ThPOK identifies potential cis-acting elements and transcription factors' binding sites (P1460)
Authors: Zha Jikun , He Xi , Dai Zhongping , Hua Xiang , Kappes Dietmar 
Published: January 2023

Abstract
T cell maturation involves key branch points at which cells choose between commitment to different lineages, i.e. gd versus ab T cell lineages and CD4+ helper versus CD8+ killer lineages. We focussed on the characterization of a unique line of spontaneous mutant mice, HD-/- mice, which are blocked in the generation of CD4+ helper T cells. Eventually, we discovered of the Zn-finger transcription factor Th-POK. We and others further established that the transcription factor ThPOK is necessary and sufficient to trigger adoption of the CD4 lymphocyte fate. We start to investigate the regulation of ThPOK expression and its subsequent control of CD4+T cell commitment. We identified six separate DHS sites, of which two coincide with putative promoters and the others could represent potential enhancers and silencers. Functional analysis of these putative regulatory elements using a series of reporter transgenics revealed that the DHS site A functions as a lineage specific silencer (referred to as DRE, distal regulatory element).To further dissect the contribution of these DHS sites to the regulation of ThPOK expression, we employed the technology of Zinc finger nucleases (ZFN) mediated genome mutation. Analyzing these mutant mice lines indicates that these both these TF binding motifs work in concert to regulate ThPOK expression, which further affect the determination of thymocytes fate.
CD8aa TCRab intraepithelial lymphocytes MHC restriction (P1462)
Authors: Lambolez Florence , Mayans Sofia , Stepniak Dariusz , Palida Sakina , Larange Alexandre , Cheroutre Hilde 
Published: January 2023

Abstract
T lymphocytes develop in the thymus into two major lineages characterized by expression of ab and gd TCR. Each lineage can be further subdivided into distinct subsets that differ in TCR specificity, phenotype and function. CD8aa TCRab intraepithelial lymphocytes (T-IEL) located in the epithelium of the small intestine is phenotypically different from conventional T cells. CD8aa T-IEL have been studied for years however the question of their MHC restriction has not been elucidated. Therefore, we decided to clone TCR isolated from naturally arising CD8aa T-IEL and retrovirally express these TCRs in BM chimera. First, we cloned four TCRab that were expressed retrovirally in RagKO BM chimera. Analysis of the chimera showed that all TCR clones gave rise to T cells. The T cell that developed were CD4-CD8b- but CD8a+ and phenotypically identical to CD8aa T-IEL isolated from wild type mice. In addition, they were preferentially found in the gut. In order to define the MHC restriction of these particular TCRs, BM chimeras in various MHC-deficient backgrounds were generated. Subsequent analysis of the chimera for the presence of CD8aaTCRab IEL demonstrated that they are dependent either on Kb or Db MHC I or b2m-dependent MHC I aside from KbDb. In conclusion, our results indicate that CD8aa T-IEL develops from precursors expressing particular TCRs that allow their engagement in this unique T cell lineage. In addition, CD8aa T-IEL can harbor clones selected on different MHC molecules.
Regulation of NKT cell development by SLAP2 (P1470)
Authors: Tuttle Kathryn , Peterson Lisa , Bedel Romain , Dragone Leonard , Gapin Laurent 
Published: January 2023

Abstract
NKT cell function in animal models is highly variable. This variation is likely due to functionally distinct subpopulations of NKT cells that are present to different degrees in each mouse strain. NKT precursors proceed through several developmental stages marked by the sequential expression of activation markers and NK cell receptors. At each step of development, NKT precursors receive signals that promote progression to the next stage. The nature of these signals is not completely understood, however T cell receptor (TCR), SLAM receptor and cytokine signaling are important. Src-Like adaptor protein 2 (SLAP2) is an adaptor protein that has been implicated in regulation of TCR and cytokine signaling in cell lines. While TCR signaling and development of conventional αβ T cells are normal in SLAP2-/- thymocytes, NKT cell development is altered. SLAP2-deficient thymocytes are enriched for immature NKT cells, a population known to be biased towards IL-4 secretion. Consequently, a large increase in the proportion of IL-4-dependent "innate CD8 lymphocytes" is detected in the thymus of SLAP2-/- mice. We hypothesize that SLAP2 modulates currently undefined signaling pathways that are important for NKT cell development.
Innate lymphoid cells are critical regulators of host-microbiota interactions and intestinal homeostasis (P3152)
Authors: Sonnenberg Gregory , Hepworth Matthew , Fung Thomas , Monticelli Laurel , Artis David 
Published: January 2023

Abstract
Mammals are colonized with trillions of commensal bacteria that are essential for normal intestinal physiology and immune cell development. However, dysregulated host-microbiota interactions have been causally linked with the pathogenesis and progression of numerous chronic infectious, inflammatory and metabolic diseases, such as HIV, cancer, obesity, viral hepatitis and inflammatory bowel disease (IBD). Innate lymphoid cells (ILCs) are an emerging family of immune cells that have recently been identified to play critical roles in regulation of immunity, inflammation and homeostasis in the intestine of mice and furthermore alterations in ILC populations have been observed in several chronic human diseases. However, whether ILCs are involved in the regulation of host-microbiota interactions is poorly understood. Here we show that ILCs are critical regulators of host immune responses to the microbiota through several distinct pathways. Surprisingly, abrogation of ILCs or their effector pathways resulted in pathologic host adaptive immune cell responses to resident intestinal microbiota and induction of spontaneous inflammation in the intestine and peripheral tissues. Collectively, these results identify an essential role for ILCs in regulating interactions between the mammalian adaptive immune system and commensal microbiota that dictate the state of intestinal health, and may represent a novel therapeutic target in chronic human disease.
Effect of the green tea catechin, epigallocatechin gallate, on innate immune responses in HIV-1 infection as a microbicide (P3170)
Authors: Nance Christina , Mata Melinda , Shearer Willliam 
Published: January 2023

Abstract
Our preclinical data showed binding of green tea catechin, epigallocatechin gallate(EGCG), to CD4 with inhibition of HIV-1-gp120 binding to human CD4+Tcells throughout a broad spectrum of virus strains lead to implementation of our Phase I/IIa clinical trial in HIV-1-infected indivduals. As mucosal dendritic cells(DC), Langerhans cells(LC) and monoctye-derived dendritic cells(MDDC), with CD4 receptor binding to HIV-1-gp120, act as first line defense in mucosa against invading pathogens via innate immune responses,EGCG in the context of an HIV-1 microbicide was studied. Human CD34+ hematopoietic stem cells and CD14+ cells differentiatied to LC and MDDC,respectively were assessed for HIV-1-gp120 binding, Annexin V-binding apopotosis and receptor profiling by flow cytometry, cytokine profiling by Luminex, toll-like receptor(TLR) profiling by RT-PCR, antigen uptake/presentation(APC) by RIA, cytotoxicity by MTT and statistical significance by Student’s t test. EGCG(1-50uM) downregulated TLR-6,-8 mRNA expresion (25%;p&lt;0.05;43%;p&lt;0.01, respectively),not TLR-2 or-3. TLR9 ligation downregulated CD86 expression and was reversed by EGCG (50uM) (62%;p&lt;0.05).EGCG (1-50uM) did not significantly increase DC cytotoxicity, apoptosis, APC nor promote proinflammatory ctyokine secretiion of TNFa,IL1a,IL1B,orIL6. R5 chemokines, MIP1a,MIP1BandRANTES were upregulated by EGCG (25uM) (29%,33%,37%,respectively;p&lt;0.01). EGCG has potential as an HIV-1 microbicide.
Inducible bronchus-associated lymphoid tissue reduces Th2-driven pathology by redistributing leukocytes within the lung (P6214)
Authors: Hwang Ji Young , Rangel-Moreno Javier , Garcia-Hernandez Maria de la Luz , Randall Troy 
Published: January 2023

Abstract
Inducible bronchus-associated lymphoid tissue (iBALT) is an ectopic lymphoid tissue formed in the lung after pulmonary infection or inflammation. This local lymphoid tissue is structurally similar to conventional secondary lymphoid organs, with separated B and T cell areas, specialized stromal cells and lymphoid dendritic cells (DCs). Here, we tested whether the presence of iBALT affects pulmonary immune responses to allergens. We induced iBALT formation in neonatal mice by pulmonary administration of LPS, and when the mice were adults, challenged them intranasally with OVA. We found that although mice with iBALT had more lymphocytes in their lungs, they had fewer eosinophils, less goblet cells and had reduced Th2 cytokine production in bronchoalveolar (BAL) fluid and lung tissue after OVA challenge. Surprisingly, using IL-4 reporter and by transferring OVA-specific OT-II cells, we found that the presence of iBALT did not reduce the number of Th2 cells that accumulated in the lung. Despite similar numbers of Th2 effectors in the lungs of mice with and without iBALT, we found that Th2 cells were differentially distributed in these two groups of mice. Those in the lungs of mice without iBALT were more widely dispersed, whereas the cells in the lungs of mice with iBALT were predominantly concentrated in the lymphoid tissue. These results suggest that the reduction in Th2-driven pathology with iBALT is due to the redistribution of leukocytes within the lung after iBALT formation.
Molecular mechanisms of TRAF6 regulation and intranuclear localization (P1305)
Authors: Su An-Jey , Adamik Juraj , Galson Deborah , Auron Philip 
Published: January 2023

Abstract
Tumor-necrosis factor receptor associated factors (TRAFs) are widely regarded as cytoplasmic adaptor proteins, that are important for the assembly of signaling complexes necessary for cell-receptor mediated immunological response. Among the TRAFs, TRAF6 is uniquely able to couple TLR/IL-1 and TNFR activation of NF-κB. Insomuch, the regulation of TRAF6 is critical for normal cellular physiology and responses to activation events. Accumulated evidence suggests that TRAF6 is regulated by both 1) interactions with other molecules as well as 2) an intramolecular interaction with itself. We have previously reported autoinhibition by the intramolecular interaction of TRAF6’s N-terminal Ring-zinc (RZ) and C-terminal MATH domains in trans. We now provide evidence that IRAK1, which contains a TRAF-Interaction Motif (TIM), can counteract TRAF6 autoinhibition. Further, we newly identify that the transcription factor Sox4 interacts with TRAF6 and that overexpression of either Sox4 or Syntenin, a dual PDZ domain containing protein, alters TRAF6 activation of NF-κB and translocates the normally cytoplasmic TRAF6 into the nucleus. While many studies have focused on cytoplasmic TRAF6 and its activation, there are few reports on the inhibition of TRAF6 and none have provided a mechanism by which it enters the nucleus.
Platelet CXCL7 and CXCL4 inhibit chemokine scavenging and improve innate immunity to bacterial infection (P1317)
Authors: Mei Junjie , Kowalska M. Anna , Dai Ning , Liu Yuhong , Hudock Kristin , Jeyaseelan Samthamby , Lee Janet , Guttentag Susan , Poncz Mortimer , Worthen G. Scott 
Published: January 2023

Abstract
We recently demonstrated the critical role of CXCL5 in chemokine scavenging, neutrophil homeostasis and host defense to bacterial pneumonia. Here we generated Cxcl7-/- mice and found that Cxcl7 deletion also disrupted expression of CXCL4 and CXCL5, but did not affect expression of two other neutrophil chemokines, CXCL1 and CXCL2. Cxcl4, 7, 5 form a conserved genomic locus (14.5 kb in length) in both human and mice, suggesting their importance during evolution. Heparin treatment of blood led to over 10-fold more plasma CXCL4, but comparable plasma CXCL7 as compared to PBS treatment. In addition, the inactive PPBP form of murine CXCL7 showed considerable binding affinity with red cell DARC. We further delineated the role of CXCL7 and CXCL4 by comparing the phenotypes of Cxcl7-/- and Cxcl5-/- mice. CXCL7 and CXCL4 inhibit chemokine scavenging not only in blood, but also in tissues, and platelet-derived CXCL7 is present in alveolar space and peritoneum. In CXCL1- and nebulized LPS-induced acute lung inflammation models, K. pneumoniae pneumonia and bacteremia models, as compared to Cxcl5-/- mice, Cxcl7-/- mice showed attenuated neutrophil chemokine levels in tissues and blood, decreased neutrophil influx to the lung, and impaired innate immunity. This study is the first to reveal the central role of homeostatic CXCL7 and CXCL4 in increasing chemokine levels in blood and tissues, and potentiating neutrophil transmigration during inflammation and infection.
Mechanisms and intervention strategies for chronic low-grade inflammation. (P1361)
Authors: Morris Matt , Li Liwu 
Published: January 2023

Abstract
Low-grade inflammation contributes to diseases such as atherosclerosis, arthritis, and diabetes. Contributors to low-grade inflammation include circulating microbial products such as lipopolysaccharide (LPS). High plasma levels of LPS cause acute shock, but low levels of LPS predispose individuals to mild inflammatory conditions. This predisposition results from mild expression of pro-inflammatory cytokines without compensating anti-inflammatory modulators, leading to persistent, unresolved inflammation. Despite the clinical importance of chronic inflammation, its underlying mechanisms are poorly understood. Glycogen synthase kinase 3 (GSK3) is implicated in the pro-inflammatory conditioning of innate immunity. We tested whether inhibition of GSK3 and upstream kinases can prevent inflammatory skewing by low dose LPS. We measured the expression of pro-inflammatory IL-6 and anti-inflammatory IL-33 in pre-monocytic THP-1 cells treated with low dose LPS (50 pg/mL) and a GSK3 inhibitor. We observed that while low dose LPS alone induced mild expression of IL-6, GSK3 inhibition skews THP-1 cells into an anti-inflammatory state by augmenting the induction of IL-33 by low dose LPS while diminishing the induction of IL-6. We found that broad inhibition of tyrosine kinases also diminishes IL-6 induction. These observations suggest a complex intracellular circuit that dynamically modulates the expression of pro- and anti-inflammatory mediators upon treatment with low dose LPS.
MiR-155 regulates NK cell activation (P1366)
Authors: Sullivan Ryan , Leong Jeffrey , Schneider Stephanie , Keppel Catherine , Romee Rizwan , Fehniger Todd 
Published: January 2023

Abstract
MiR-155 is a well-documented small RNA regulator of a variety of immune cells, including adaptive T and B lymphocytes. However, its role in regulating the innate NK cell response is unclear. We evaluated the expression of miR-155 using real-time qPCR and found that miR-155 is expressed at modest levels in resting mouse and human NK cells, but is upregulated upon stimulation, especially with IL-12+15+18 (30-fold). Therefore, we hypothesized that miR-155 regulates NK cell function following activation. We utilized miR-155-/- mice (155KO), NK-specific miR-155 overexpression mice (155OE), and miR-155 lentiviral transduction to examine the role of miR-155 in NK cell functional responses. miR-155KO, miR-155OE, and miR-155-transduced NK cells were all found to secrete more IFN-γ (by ELISA) upon ligation-mediated or cytokine stimulation, with no apparent defects in maturation or development. When further dissected by flow cytometry, we found that 155OE and transduced NK cells had increased IFN-γ MFI, indicating more “per-cell” production. In contrast, the 155KO NK cells had a higher percentage of IFN-γ+ NK cells, with similar MFI, indicating a potential activation threshold defect. Though both 155KO and 155OE NK cells produce more IFN-γ, the cellular basis for each, i.e. miR-155 target, likely differs. Therefore, we used unbiased RISC-IP to identify relevant miR-155 targets, which included previously validated miR-155 targets (e.g., SHIP-1) as well as novel miR-155 NK cell targets.
DDX41 recognizes bacterial secondary messengers cyclic di-GMP and cyclic di-AMP to activate a type I interferon immune response (P1375)
Authors: Parvatiyar Kislay , Zhang Zhiqiang , Teles Rosane , Ouyang Songying , Jiang Yan , Liu Zhi-Jie , Iyer Shankar , Zaver Shivam , Schenk Mirjam , Zeng Shang , Zhong Wenwan , Modlin Robert , Liu Yongjun , Cheng Genhong 
Published: January 2023

Abstract
Cytosolic detection of bacterially derived secondary messengers cyclic-di-GMP (c-di-GMP) or cyclic -di-AMP (c-di-AMP) by the host immune system activates an innate immune response characterized by the induction of type I interferons (IFNs). Induction of IFN by c-di-GMP or c-di-AMP has been shown to be dependent on a stimulator of IFN genes-TANK binding kinase 1-IFN regulatory factor 3 (STING-TBK1-IRF3) signaling axis. Although STING has been shown to interact with c-di-GMP, an upstream sensor of these cyclic dinucleotides is unknown. Here we identify the helicase, DEAD (Asp-Glu-Ala-Asp) box polypeptide 41 (DDX41) as the pattern recognition receptor (PRR) that senses both c-di-GMP and c-di-AMP. DDX41 specifically and directly interacts with c-di-GMP. Knockdown of DDX41 via shRNA in murine or human immune cells inhibits the induction of innate immune genes and results in defective STING, TBK1 and IRF3 activation in response to c-di-GMP or c-di-AMP. Our findings suggest a mechanism whereby c-di-GMP and c-di-AMP molecules are detected by the DDX41 PRR, which complexes with the STING adaptor to signal to TBK1-IRF3 and activate the IFN response.
Multiple pools of dynein and their roles in T cell effector functions (P1028)
Authors: Nath Shubhankar , Christian Laura , Lee Siu , Artz Jonathan , Yuan Denley , Poenie Martin 
Published: January 2023

Abstract
Dramatic cytoskeletal activities and protein movements are seen when T cells engage cognate targets or antigen presenting cells. These include formation of a specialized contact site known as the immunological synapse (IS), clustering of vesicles around the microtubule organizing center (MTOC) and translocation of the MTOC up to the IS. Ultimately, this leads to focused secretion towards the cognate target. In an effort to understand how MTOC translocation takes place, we have investigated the roles of dynein, p150Glued, Lis1,NDE1 and NDEL1 using Jurkat cells as a model system. Using expressed molecular traps aimed at dynein intermediate chain (DIC) or LC8 (a dynein light chain), and Jurkat cells expressing fluorescent CTLA-4 constructs targeting secretory vesicles, we have shown that dynein is required for both vesicle clustering and MTOC translocation. When p150Glued expression was reduced, vesicles became dispersed but MTOC translocation was unaffected suggesting that dynactin was needed for vesicle movements but not MTOC translocation. MTOC translocation was potently blocked when Lis1 expression was reduced. We also find that NDE1 accumulates at the IS and pulls down with DIC and Lis1. Overexpression of GFP-NDE1 blocks MTOC polarization. The combination of localization and functional data suggests that there are two separate dynein complexes, a dynein-Lis1-NDE1/NDEL complex that mediates MTOC translocation and a dynein-dynactin complex that mediates vesicle clustering.
Th2 cytokines IL-4 and IL-13 negatively regulate lymphangiogenesis (P1038)
Authors: Sin Ki-Hyuk , Kataru Raghu , Lee Seung-Hyo 
Published: January 2023

Abstract
The vascular system consists of two major components, blood vessels (BVs) and lymphatic vessels (LVs). Both vascular systems are essential for homeostasis in healthy organism and related to the pathogenesis of many diseases. Other than fluid homeostasis LVs are also important for immune surveillance. In the present study, we show novel anti-lymphangiogenic factors IL-4 and IL-13 which is major effector cytokine produced by type 2 helper T cells (Th2 cells). By using murine lymphatic endothelial cells (mLECs) and human LECs (hLECs), we demonstrate that IL-4 and IL-13 down-regulate Prox-1 which is essential transcription factor of LECs. These cytokines inhibited tube formation of LECs also. Surprisingly, co-culture LECs with Th2 cells inhibited tube formation, but this effect was fully restored by blocking IL-4 and IL-13. Furthermore, in vivo blockade of IL-4 and IL-13 in a murine asthma model increase not only the density of LVs in the lung but also the function of LVs. Taken together our findings demonstrate a novel anti-lymphangiogenic function for Th2/IL-4 and IL-13, suggest that these effects could be involved in pathogenesis of asthma.