bluuebunny/crossref_metadata_2025_split huggingface dataset:
The actin-bundling protein L-plastin regulates alveolar macrophage pro-inflammatory cascade (INM2P.426)
Authors: Morley Sharon Celeste , Deady Lauren , Todd Elizabeth , Zhou Julie 
Published: January 2023

Abstract
Regulation of the actin cytoskeleton is essential to multiple immune cell functions, including cell motility, adhesion, and activation. In lymphocytes, the actin-bundling protein L-plastin (LPL) plays critical roles in sustaining cell polarization and formation of the immunological synapse. We now show defects in the innate immune system that lead to profound susceptibility to pulmonary pneumococcal infection. LPL is required for generation of alveolar macrophages: there are fewer alveolar macrophages in the bronchoalveolar lavage fluid of LPL-/- mice, and regeneration of alveolar macrophages following sublethal irradiation is delayed when bone marrow is donated by LPL-/- mice. Activation of LPL-/- alveolar macrophages is also defective, with reduced upregulation of IL-1-beta and IL-6 following LPS stimulation. Reduced numbers and activation of alveolar macrophages in LPL-/- mice correlate with disrupted podosome formation. Reduced numbers and activation of alveolar macrophages would be expected to result in an inability to clear pulmonary pathogens. When challenged with pulmonary infection with pneumococcus, LPL-/- mice show a clearance defect within 3 hours, with a ten-fold increase in recoverable pneumococci compared to wild-type mice. LPL-/- mice thus offer a model system in which to detail how actin cytoskeletal integrity regulates macrophage development and activation.
Carma1 and MALT1 are required for stimulation-dependent mTORC1 activation in T cells (IRC2P.455)
Authors: Hamilton Kristia , Shiva Sruti , Kane Lawrence 
Published: January 2023

Abstract
Stimulation of T cells through the T cell receptor (TCR) and co-stimulatory molecules induces a diverse set of signaling events leading to T cell activation. Some characteristics of T cell activation are cellular proliferation, increase in cell size and changes in metabolic pathways. Our work has primarily focused on understanding the roles that Carma1, MALT1 and Bcl10 (the CBM complex) play in T cell activation, specifically, elucidating their requirement in distinct T cell signaling pathways. Here, we show a requirement for Carma1 and MALT1 in activation of mTORC1. In the absence of Carma1 or MALT1, we noted impaired TCR/CD28-induced stimulation of ribosomal protein S6, p70S6K and 4E-BP1 phosphorylation. Further investigation into the specific molecular mechanism revealed constitutive association of MALT1 and p70S6K. Surprisingly, Bcl10 was not required for S6 or S6K phosphorylation in T cells. Pharmacological inhibition of this pathway with a MALT1 protease inhibitor (z-VRPR-fmk) impaired T cell proliferation upon TCR and co-stimulatory molecule stimulation, though this did not affect T cell “blasting” or early activation. Additionally, inhibition of MALT1 activity impaired T cell activation-induced glycolysis, as seen by a decrease in oxygen consumption rate (OCR) and extracellular acidification rate (ECAR). Thus, these findings present a novel mechanism for mTORC1 activation in T cells, through Carma1 and MALT1, leading to cellular proliferation and increased glycolysis.
Negative signalling in T cells with aging (IRC2P.446)
Authors: Fulop Tamas , Dupuis Gilles , Le Page Aurélie , Larbi Anis 
Published: January 2023

Abstract
Immune responses are generally impaired in aged humans. There is impaired production of IL-2 and T lymphocyte proliferation in response to antigenic stimulaton. These decreased responses involve altered signaling events associated with the early steps of T cell activation. The underlying causes of these changes are not fully understood but point to alterations in assembly of the machinery of T cell activation. We have tested the hypothesis that the T cell pool in elderly subjects displayed reduced functional capacities due to altered negative feedback mechanisms. We used tyrosine-specific mAbs and Western blot analysis to show that a dysregulation of the Csk/PAG loop in activated T cells from elderly individuals favored the inactive form of tyrosine-phosphorylated Lck (Tyr505). Confocal microscopy analysis revealed that the dynamic movements of these regulatory proteins in lipid raft microdomains was altered in T cells of aged individuals. Enzymic assays showed that SHP-1 activity was upregulated in T cells of aged donors, in contrast to young subjects. Pharmacological inhibition of SHP-1 resulted in recovery of TCR/CD28-dependent lymphocyte proliferation and IL-2 production of aged individuals to levels not significantly different than those of young donors. Our data suggest that the role of SHP-1 in T cell activation extends to its increased effect in negative feedback in aging. Modulation of SHP-1 activity could be a target to restore altered T cell functions in aging.
Nedd4-family interacting proteins regulate T cell homeostatic processes via activation of catalytic E3 ubiquitin ligases (IRC2P.447)
Authors: O'Leary Claire , Riling Christopher , Oliver Paula 
Published: January 2023

Abstract
Ubiquitylation tunes signaling pathways in stimulated T cells to regulate activation and function. Catalytic E3 ubiquitin ligases, like Itch, have known roles in these processes. We have found that Nedd4-family interacting protein 1 (Ndfip1) and Ndfip2 promote ubiquitin charging of Itch, a requisite step in protein ubiquitylation. In vitro, Ndfip1 and Ndfip2 have overlapping function. In vivo, Ndfip1 negatively regulates T cell activation and Th2 polarization. The in vivo role of Ndfip2 is unknown. To investigate this, we generated Ndfip2-/- mice. We found that, unlike Ndfip1, Ndfip2 is not a prominent negative regulator of T cell activation or Th2 polarization. However, loss of Ndfip2 exacerbates the inflammatory Ndfip1-/- phenotype, suggesting that, like Ndfip1, Ndfip2 dampens inflammatory processes. Our data indicate a T cell intrinsic role for Ndfips in limiting T cell activation and function, as Ndfip1/Ndfip2 DKO CD4+ T cells in mixed fetal liver chimeras are more activated and produce more cytokine than WT CD4+ T cells in the same host. In vitro coculture of naïve WT and DKO CD4+ T cells supports that Ndfip deficient T cells have increased viability and proliferative capacity. These results suggest that Ndfip1 and Ndfip2 together limit T cell survival and proliferation downstream of TCR engagement. We are taking a quantitative proteomic approach to identify ubiquitylation pathways engaged during TCR stimulation that are nucleated by Ndfip-E3 ligase interactions.
Activation of the aryl hydrocarbon receptor during development leads to an altered CD4+ T cell response to influenza A virus infection (IRC2P.448)
Authors: Boule Lisbeth , Lawrence B. Paige 
Published: January 2023

Abstract
Epidemiological evidence suggests developmental exposures to certain pollutants correlate with lower antibody responses to childhood immunizations, but the mechanism by which this occurs is unknown. The transcription factor aryl hydrocarbon receptor (AhR) is activated by a variety of chemicals, expressed by immune cells, and has been shown to influence immune function, such as CD4+ T cell subset specification and function. The effect of maternal exposure to AhR ligands on the function of CD4+ T cells has never been directly examined. We report here the consequences of developmental triggering of the AhR on the CD4+ T cell response to influenza virus infection. Mice developmentally exposed to 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), the prototypical AhR ligand, have fewer activated, virus-specific CD4+ T cells, Th1, and T follicular helper cells, yet an increase in regulatory T cells in their lung draining lymph nodes. Consistent with this, developmentally exposed mice have a reduction in their class-switched, virus-specific antibody response. By using reciprocal adoptive transfers, we demonstrate that the effects of developmental activation of the AhR on CD4+ T cells occur through both direct and indirect pathways. These findings suggest changes in the functional capacity of CD4+ T cells may be a key contributor to the decreased antibody response to immunizations observed in children of mothers exposed to AhR-binding chemicals.
Pivotal role of STAT3 signaling in germinal center B cell and T follicular helper cell differentiation (IRC3P.458)
Authors: Ding Chuanlin , Yan Jun 
Published: January 2023

Abstract
Germinal center (GC) B cells and T follicular helper (Tfh) cells are critical for long-lived plasma cell differentiation and high-affinity antibody production in response to T cell-dependent antigens. Previous studies demonstrated that GC formation was intact in the absence of STAT3 in B cells at day 12. However, emerging evidence suggests that IL-21/STAT3 signaling is a potent regulator for B cell differentiation. To better understand the role of STAT3 in the kinetics of GC formation and GC B cell response, we assessed GC B cells at consecutive time points following immunization with antigen OVA. In B cell conditional STAT3-dificient mice, fewer GC and GC B cells were generated upon immunization in the late phase (day 21) but not on day 14 compared with that of control mice. Increased cell death and decreased cell proliferation were found in the STAT3 deficient GC B cells. Consequently, plasma cell differentiation and antibody production were impaired in B cell STAT3-deficient mice. In addition, the Tfh cell response was correlated with the magnitude of the GC B cell response and significantly decreased in B cell STAT3-deficient mice. Furthermore, GC B cells and autoantibody production were also significantly decreased in autoreactive B cell STAT3-deficient mice after apoptotic cell immunization. These data provide new insights into the GC and Tfh differentiation and the function of STAT3 in humoral immune responses.
TIM-4 expression by B cells identifies an inflammatory B effector 1 subset that promotes allograft rejection and inhibits tumor metastases (IRC3P.471)
Authors: Mohib Kanishka , Ding Qing , Camirand Geoffrey , Rothstein David 
Published: January 2023

Abstract
B cells, like T cells, give rise to polarized subsets, termed Be1, Be2 and Breg that produce IFNγ, IL-4 and IL-10, respectively. We previously reported that TIM-1 is an inclusive marker for Be2 cells and Breg that can transfer allograft tolerance. We now show that TIM-4, a TIM-1 ligand, is expressed by a distinct subpopulation (~6-10%) of B cells. TIM-4 B cells are enriched for IFNγ (2.5%) and express relatively less IL-4 (1%) and IL-10 (4%) while TIM-1 B cells are enriched for IL-4 (6.5%) and IL-10 (14%) and express less IFNγ (1%). Accordingly, transfer of TIM-4 B cells to B cell deficient (JhD) islet allograft recipients accelerated rejection (MST 11d) vs. controls (no B cells; MST 15d) or TIM-1 B cell transfer (MST 70d), p&lt;0.05. TIM-4 B cell-mediated rejection required IFNγ as transfer of IFNγ-/- TIM-4 B cells modestly prolonged MST (15d to 25d, p&lt;0.05). In this model, transfer of TIM-4 B cells enhanced IFN-γ+ T cells (1.5X) while decreasing IL-4+, IL-10+ and Foxp3+ T cells (1.2-1.8X) whereas TIM-1 B cells increased IL-4+, IL-10+ and Foxp3+ T cells (1.2-1.7X) while decreasing IFNγ+ T cells (1.4X) vs. no B cell controls (p&lt;0.05 for each). Moreover, transfer of TIM-4 B cells into μMT mice decreased B16 melanoma lung metastases ~ 2-fold, while TIM-1 B cells increased lung metastases ~2-fold vs. controls (p &lt; 0.0001). Thus, we provide strong evidence that TIM-4 expression by B cells identifies a distinct Be1 inflammatory subset that can directly impact T cells responses.
Aging of the B-1a cell pool (IRC3P.473)
Authors: Holodick Nichol , Vizconde Teresa , Rothstein Thomas 
Published: January 2023

Abstract
B-1a cells arise predominately during fetal development, and their persistence throughout adult life is maintained through self-renewal. These cells are essential for immediate protection from S. pneumoniae infection. The unique ability of fetal derived B-1a cells to provide protection against S. pneumoniae is attributed to their spontaneously secreted natural IgM, which is germline-like due to minimal insertion of N-region additions and little somatic hypermutation. The B-1a cell pool is also maintained by B-1 cell progenitors found in the adult bone marrow. We reported these bone marrow derived (BMD) B-1a cells are functionally and phenotypically similar to native B-1a cells; however, BMD B-1a cells contain an increased number of N-region additions in their natural IgM as compared to native B-1a cells. Natural IgM derived from native B-1a cells in aged mice also contains more N-region additions. We show an increase in N-region addition in mice 6 months to 24 months of age. This suggests the repertoire of B-1a derived natural IgM changes with age, and the adult B-1a cell pool is not a closed system. We hypothesized the level of natural IgM protection changes with age as a result of repertoire alterations to the B-1a cell pool over time. We found the natural IgM from aged mice is less effective at protection from S. pneumoniae infection. Data presented here suggest age related changes of natural IgM can lead to less effective initial protection against infection.
B cells promote obesity-associated periodontitis and oral pathogen-associated inflammation (IRC3P.461)
Authors: Zhu Min , DeFuria Jason , Carr Jordan , Belkina Anna , Van Dyke Thomas , Gyurko Robert , Nikolajczyk Barbara 
Published: January 2023

Abstract
Chronic inflammation likely underlies the predisposition of type 2 diabetes (T2D) patients to periodontitis (PD), and the exacerbation of each disease when both are present. Several lines of evidence indicate that pro-inflammatory B cells promote T2D and PD individually, but the possibility that B cells are a key link between T2D to PD is untested. We compared outcomes from oral P. gingivalis challenge of lean WT or B cell-null mice to outcomes from mice that were obese and insulin resistance prior to oral challenge. Obese WT mice responded to oral P. gingivalis challenge with significant periodontal bone loss, while obese B cell-null mice were completely protected from PD. By contrast, lean WT and B cell-null mice suffer similar periodontal bone loss in response to oral pathogen. B cells from obese/insulin resistant hosts also support oral osteoclastogenesis, and both oral and systemic production of inflammatory cytokines, including the PD-associated cytokines TNF-α and MIP-2. B cells furthermore impact adipose tissue inflammation in obese P. gingivalis-infected hosts. Taken together, these data show that fundamentally different mechanisms regulate PD in lean and obese hosts, with B cells promoting PD only if the hosts are “primed” by obesity. These results justify more intense analysis of obesity-associated changes in B cells that predispose PD in human T2D, and may indicate the use of B cell depletion to alleviate PD and T2D simultaneously.
Tissue-specific tolerance of endogenous self antigen-specific CD4+ T cells (IRC4P.484)
Authors: Moon James , Lim Jong-Baeck , Legoux Francois 
Published: January 2023

Abstract
CD4+ T cells specific for self antigens are involved in autoimmunity, but little is known about the extent to which these potentially harmful T cells develop within a normal T cell repertoire or how they are maintained in a tolerant state. We modeled Cre recombinase as a neo-self antigen that is expressed by various different tissue-specific promoters in several strains of Cre transgenic mice. We mapped a CD4+ T cell epitope within the Cre protein and generated a corresponding peptide:MHCII tetramer, which we used in a magnetic enrichment strategy to directly detect rare populations of endogenous Cre-specific CD4+ T cells that naturally develop in the unmanipulated immune systems of these mice. We found that deletion of Cre-specific T cells was efficient in mice ubiquitously expressing Cre, but almost nonexistent in mice expressing Cre in tissue-restricted patterns. In mice expressing Cre exclusively in the pancreas, immunization with Cre antigen induced strong Cre-specific T cell expansion and cytokine production, suggesting immune ignorance to the pancreatic self antigen. In contrast, tolerance to Cre self antigen expressed in mucosal tissues such as the gut and lung relied on elevated steady state frequencies of Cre-specific regulatory T cells. While these Tregs efficiently controlled primary autoimmune responses, their ability to maintain tolerance was lost over successive antigenic challenges, indicating limits of peripheral tolerance to tissue-restricted self antigens.
An endogenous aryl hydrocarbon receptor (AhR) ligand, ITE induces regulatory T cells (Tregs) and ameliorates experimental colitis (IRC4P.490)
Authors: Singh Udai , Abron Jessicca , Singh Narendra , Guan Honbing , Hegde Venkatesh , Mishra Manoj , Scissum-Gunn Karyn , Price Robert , Nagarkatti Mitzi , Nagarkatti Prakash 
Published: January 2023

Abstract
Inflammatory bowel disease (IBD) is a chronic intestinal inflammatory condition that affects millions of people with high morbidity and health-care cost. Recent studies have shown that activation of the aryl hydrocarbon receptor (AhR) with the nontoxic ligand 2-(1’H-indole-3’-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE) induces regulatory T cells (Tregs) and suppresses autoimmune diseases. However, the effects of ITE on experimental colitis have not been investigated. We hypothesized that ITE induces in-vitro and in-vivo Tregs through activation of AhR and suppresses experimental colitis. Our data showed that ITE increases the frequency of Tregs in spleen, MLN and LPL as compared to vehicle. This induction of Tregs was reversed by AhR antagonist treatment in-vitro. Next, in-vivo colitis pathogenesis including a decrease in body weight was significantly reversed and systemic inflammatory cytokines were reduced in mice that received ITE treatments as compared to vehicle. ITE treatment also increased CD11c+, decreased activated T and CD11b+ cells from the spleen, MLN and LP as compared to vehicle alone. These findings clearly suggest that ITE diminishes colitis pathology through induction of Tregs, reduces inflammatory cytokines, decreases the inflammation score, and induces dendritic cells resulting in amelioration of colitis. Thus, our work demonstrates that the nontoxic endogenous AhR ligand, ITE, may serve as unique treatment modality against IBD.
TNFR2-deficiency contributes to sex-biased spontaneous experimental autoimmune encephalomyelitis (IRC4P.481)
Authors: McKarns Susan , Miller Patrick 
Published: January 2023

Abstract
Tumor necrosis factor-α (TNF) antagonism has improved the treatment of autoimmune disorders including rheumatoid arthritis and inflammatory bowel disease but is contraindicated in multiple sclerosis (MS) due to disease exacerbation. The mechanisms underlying these contrasting responses remain elusive. The objective of this study was to investigate the effect of selective TNFR2 ablation on the autoreactivity of myelin oligodendrocyte glycoprotein (MOG)-specific T cells. We report that 92% of female C57BL/6 TNFR2-/- MOG-specific (2D2) TCR transgenic mice spontaneously develop experimental autoimmune encephalomyelitis (EAE); whereas, only 8% of male TNFR2-/- 2D2 littermates developed an autoimmune phenotype. Demyelination accompanied B cell and CD4 T cell infiltration in the spinal cords and optic nerves of TNFR2-/- 2D2 mice with EAE. CNS inflammation in these mice correlated with increased production of IFN-γ and IL-17 and elevated sera titers of MOG35-55-specific IgG2b. Ex vivo co-culture studies demonstrated autonomous TNFR2-/- 2D2 CD4 FoxP3+ regulatory T cell (Treg) dysfunction as well as increased resilience by FoxP3-/- 2D2 CD4 T cells to respond to Treg suppression. These results suggest that TNFR2 plays a key role in the functional tolerance of MOG-specific CD4 T and B cells in a sex-related manner. This novel spontaneous autoimmune model provides a unique paradigm to study T and B cell interactions as well as sex-related factors in multiple sclerosis susceptibility.
Different roles for Smad4-dependent and independent TGFβ signaling pathways during the differentiation of virus-specific effector CD8 T cells (IRM4P.494)
Authors: Hu Yinghong , Cauley Linda 
Published: January 2023

Abstract
Transforming growth factor-β (TGFβ) is a regulatory cytokine that modulates the activities of other cells via multiple signaling pathways. The predominant pathways in hematopoietic cells are mediated by Sma and Mad-related family members (SMAD) and depend on a ubiquitous activating protein SMAD4. Other Smad4-independent pathways can also be activated by TGFβ however their role in immune regulation is less clearly defined. We examined the role of Smad4-dependent and -independent TGFβ signaling during the development of the CD8 T cell response to influenza virus infection. Our studies show that a complete blockade of all TGFβ-signaling accelerates the development of virus-specific CD8+ effector T cells (Teff) which express KLRG-1 and produce large quantities of cytokines in the lungs. In contrast, selective inhibition of only Smad4-dependent signaling prevents the development of a robust Teff response. These data point to a previously unexamined role of Smad4-independent TGFβ signaling in the regulation of virus-specific Teff cells. We are using this model to determine whether TGFβ regulation plays an major role in protective immunity in the lungs.
Simultaneous downmodulation of mTORC1 and PD-1 in activated CD8 T-cells generates expanded population of exhaustion-resistant memory T-cells (IRM4P.495)
Authors: Berezhnoy Alexey , Levay Agata , Puplampu-Dove Yvonne , Rajagopal Anugraha , Brenneman Randall , Gilboa Eli 
Published: January 2023

Abstract
It is possible to redirect recently activated T-cells toward memory differentiation by altering intracellular signaling of the mTORC1 pathway. As previously reported, treatment with rapamycin increased formation of memory cells; however, animals treated with the drug exhibited signs of immune suppression and the rapamycin-generated memory T-cells failed to protect animals against a tumor challenge. We introduced highly efficient and specific method to downmodulate mTORC1 in CD8 T cells, whereby siRNA against raptor, was targeted in vivo to activated CD8 T-cells by conjugating to a 4-1BB-binding RNA aptamer, leading to the generation of an enhanced memory response and protective antitumor immunity. Memory T-cells are resistant to immune suppression; yet, in vivo the population is experiencing attrition mediated in part via PD-1 signalling. We developed the conjugate with two siRNAs against raptor and PD-1 linked to a 4-1BB aptamer. In vitro, both siRNAs efficiently dissociated from the conjugate inside target cells and downregulated their respective mRNAs. In vivo, we demonstrated that treatment with 4-1BB-raptor-PD1 conjugates increased formation of memory CD8+ T-cells in a rate similar to the 4-1BB-raptor conjugate. Notably, CD8+ memory T-cells expanded with 4-1BB-raptor-PD1 conjugates expressed significantly less PD-1 on cell surface. Thus, we introduced a new method of cell-targeted delivery of two siRNA to generate CD8 memory cells with enhanced functional potential.