bluuebunny/crossref_metadata_2025_split huggingface dataset:
Disease-associated dominant negative CARD11 mutations perturb the CARD11 Opening step during T cell receptor-induced activation of NF-κB and JNK2
Authors: Bedsaul Jacquelyn Renee , Pomerantz Joel 
Published: February 2023

Abstract
CARD11 is an oligomeric, multidomain scaffold that mediates lymphocyte activation. Upon antigen receptor engagement, CARD11 becomes active and signals through NF-κB, JNK, and mTOR to elicit the adaptive immune response. The CARD11 signaling cycle consists of at least four stages beginning with the Opening step, followed by Cofactor Recruitment, Enzyme Activation, and Complex Disassembly. Primary Immunodeficiency Diseases (PIDDs) have been attributed to both autosomal dominant and recessive CARD11 variants. Combined Immunodeficiency with atopy is caused by the inheritance of a single dominant negative CARD11 allele, but the mechanistic basis for the dominant negative effects is not well understood. In this study, we investigated the mechanistic basis for dominant negative CARD11 mutations in T cells by assessing the activity of loss-of-function variants in the presence and absence of co-expressed wild-type CARD11. In the absence of wild-type CARD11, the variants perturbed the Opening step and downstream signaling steps, including recruitment of Bcl10, HOIP, and MALT1 to CARD11, Bcl10 polyubiquitinylation, MALT1 protease activity, NF-κB and JNK2 activation, and IL-2 production. When the variants were co-expressed with wild-type CARD11, a range of dominant negative effects on NF-κB activation was observed, which correlated with the ability of the CARD11 oligomer to open, recruit cofactors, and mediate IL-2 production. We conclude that dominant negative mutations interfere with the CARD11 Opening step and provide insights into the manifestation of PIDDs caused by the inheritance of dominant and recessive CARD11 alleles.
The deficiency of Cbl family in human elicits heightened T-dependent immune activation with augmentation of TFH responses
Authors: Lee Kyungwoo , Kim Sun jung 
Published: February 2023

Abstract
T follicular helper (TFH) cells are functional subset CD4+ T cells in providing help to B cells and aberrant expansion of TFH cells is observed in systemic lupus erythematosus (SLE), which suggests a global defect in the maintenance of T and B cell tolerance. The mechanisms underlying uncontrolled TFH cells development and differentiation in SLE pathogenesis is fragmentary understand. Casitas B-lineage lymphoma (Cbl) family is an E3 ubiquitin ligase and has been implicated in SLE pathogenesis as a potent regulator in activating the immune cells and maintaining peripheral tolerance. The deficiency of cbl-b mice spontaneously develops autoimmunity and is highly susceptible to develop pathogenic auto-antibodies of autoimmune diseases. Through CRISPR KO system, we identified the deficiency of cbl/cbl-b in human naïve CD4+ T cells induces the TFH expansion in T cell stimulation with Activin A & IL12. Indeed, higher levels of chemokine (C-X-C motif) ligand 13 (CXCL13) which is constitutively produced in germinal center (GC) TFH cells were detected. Functionally, cbl/cbl-b KO TFH cells are more potent B cell activators compared to cbl/cbl-b competent TFH cells. There was increased plasmablast differentiation which secrete IgG. These data suggest that the deficiency of Cbl family in human elicits heightened T-dependent immune activation with augmentation of TFH responses, thus regulating humoral immune responses and the susceptibility to SLE in human. This project will advance our understanding of the mechanism of activation and maintenance of TFH cells in lupus and dissect possible avenues of therapeutic intervention in disease.
Lymphocyte phenotype in a mouse model of 22q11.2 deletion syndrome
Authors: Crockett Alexis Mary , Benallegue Nail , Adelman Philippa , Gur Raquel , Sullivan Kathleen E , Anderson Stewart A , Alvarez Jorge Ivan 
Published: February 2023

Abstract
Mice modeling the hemizygous deletion of chromosome 22q11.2 (22qMc) have been utilized to address various clinical phenotypes associated with the disease, including cardiac malformations, altered neural circuitry and behavioral deficits. Yet, the status of the immune system, an important clinical concern among 22q11.2 deletion syndrome (22qDS) patients, has not yet been addressed in this model despite the well-known effects of the immune system in regulating body homeostasis. While infancy and early childhood in 22qDS are associated with deficient T cell numbers due to thymic hypoplasia, which can be severe in a small subset of patients, clinical studies suggest a normalization of the T cell counts by adulthood. For the first time, we have studied the immune system in 22qMc and find that adult 22qMc do not exhibit thymic hypoplasia, immunodeficiencies of lymphoid organs, or altered T cell activation. Furthermore, deletion of the 22q11.2 homologous region does not affect T cell differentiation. Thus, this model accurately reflects the normalization of the T cell compartment in adulthood, as has been reported in human 22qDS. Altogether, our data suggest that 22qMc may serve as novel model to address experimental and translational aspects of immunity in adult 22qDS.
Borrelia burgdorferi peptidoglycan triggers inflammatory responses in Lyme arthritis
Authors: Rouse Joe , Wahhab Amanda , Danner Rebecca , Jutras Brandon , Lochhead Robert 
Published: February 2023

Abstract

Background:
Lyme arthritis (LA) is the predominant late-stage manifestation of Lyme disease, caused by the tick-borne spirochete Borrelia burgdorferi (Bb). Bacterial peptidoglycan (PG) is abundant in inflamed joints of LA patients, even months to years after antibiotic therapy, but the role of PG in LA pathogenesis is unclear.

Methods:
Primary murine macrophages and synovial fibroblasts, two cell types abundant in LA synovial tissue, were stimulated for 24–72 hours with IFNγ, PG, or both. Inflammatory responses were determined by flow cytometric analysis of CD80, CD86, MHC-II, CD206, ICAM1, and IL-15R.

Results:
Macrophage stimulation with PG+IFNγ for 24 hours led to up-regulation of costimulatory molecules CD80/CD86, compared with PG or IFNγ alone, consistent with classical (M1) activation. In contrast, stimulation for 72 hours resulted in marked suppression of MHC-II and upregulation of CD206, consistent with M2 macrophage polarization. When stimulated with PG+IFNγ for 24 hours, synovial fibroblasts upregulated activation markers ICAM1 and IL-15R, whereas these activation markers were only partially up-regulated when stimulated with PG or IFNγ alone. Current experiments include analyzing cytokine secretion by these cells at different time points.

Conclusion:
Both innate immune and stromal cells abundant in joint tissue are responsive to stimulation with PG, but these responses may be either pro-inflammatory or anti-inflammatory, depending on cell type and duration of stimulation. This highlights the complex relationship between PG and the cellular environment of infected joints. Future experiments will identify relevant immune response pathways, such as TLR2 and NOD2 signaling, in shaping these responses.
PI3Kd coordinates transcriptional, epigenetic and metabolic changes to promote effector CD8 T cells at the expense of memory
Authors: Cannons Jennifer Leigh , Villarino Alejandro V , Kapnick Senta M , Preite Silvia , Shih Han-Yu , Gomez-Rodriguez Julio Washington , Reilley Julie , Huang Bonnie , McBain Ian , Wu Tuoqi , Su Helen C , McGavern Dorian B , O’Shea John J , McGuire Peter J , Uzel Gulbu , Schwartzberg Pamela L 
Published: February 2023

Abstract
Patients with Activated-PI3Kd Syndrome (APDS) present with sinopulmonary infections, lymphadenopathy and CMV and/or EBV viremia, yet why patients fail to clear certain viral infections remains poorly understood. Using APDS patient samples and a mouse model (Pik3cdE1020K/+ mice), we demonstrate that, upon activation, Pik3cdE1020K/+ CD8+ T cells exhibit exaggerated features of short-lived effectors both in vitro and post-viral infection, associated with increased Fas-mediated apoptosis due to sustained phosphorylation of FoxO1 and derepression of FasL. In addition, Pik3cdE1020K/+ CD8+ T cells exhibit enhanced mTORC1 and c-Myc signatures; metabolic perturbations; and reorientation of their chromatin landscape. Conversely, Pik3cdE1020K/+ CD8+ T cells failed to sustain expression of proteins critical for maintenance of long-lived memory cells, including TCF1. Strikingly, activated Pik3cdE1020K/+ CD8+ T cells exhibit altered transcriptional and epigenetic circuits characterized by a pronounced IL-2/STAT5 signature associated with heightened IL-2 responses that prevented differentiation to memory-like cells in the presence of IL-15. Our data position PI3Kd as a central driver integrating multiple signaling circuits that promote terminal CD8+ T cell effector differentiation at the expense of memory and long-lived T cell responses.
This work was funded in part by the Intramural Research Program of NIAID, NIH.
Dendritic Cells Supply CD4+ T Cells With L-arginine
Authors: Crowther Rebecca R , Schmidt Stephanie M , Zhao Junfang , McKell Melanie C , Setchell Kenneth D , Qualls Joseph E 
Published: February 2023

Abstract
Tuberculosis (TB), caused by Mycobacterium tuberculosis, is responsible for over 1 million deaths each year. Mycobacteria-infected dendritic cells (DCs) migrate to the lymph node to initiate adaptive immune priming, which is vital to antimycobacterial immunity. This response is intimately tied to nutrient availability – especially the amino acid L-arginine (L-ARG), metabolism of which is altered in TB patients. We have characterized a pathway utilized by immune cells to synthesize L-ARG. Loss of L-ARG synthesis in CD11c+ cells, which includes DCs, results in increased mycobacterial burden following infection in mice. To characterize the role of this pathway in DCs, we developed a co-culture system with mycobacterial-specific CD4+ T cells and bone marrow derived DCs. Using CD4+ T cells and DCs with differing capabilities of L-ARG synthesis, we found 1) DC L-ARG synthesis supports CD4+ T cell proliferation and 2) activated T cells contain DC-derived L-ARG. We hypothesize DCs “share” synthesized L-ARG to support CD4+ T cell activation when L-ARG is limiting. Our data suggest nutrient availability as a 4th signal – following antigen presentation, co-stimulation, and cytokine receptor ligation – required for T cell activation. This work expands the current model of DC-T cell interactions and provides insight into the effects of nutrient availability in immune cells.
CD40-CD40L interaction is critical in mounting host immunity against m-CoV Mouse Hepatitis Virus induced neuroinflammatory demyelination
Authors: Sarma Jayasri Das , Saadi Fareeha , Chakravarty Debanjana , Kumar Saurav , Kamble Mithila , Khan Reas , Shindler Kenneth S 
Published: February 2023

Abstract
Chronic progressive neuroinflammatory disease multiple sclerosis (MS) is characterized by loss of neuronal functions resulting from demyelination with or without axonal degeneration. Infiltration of T lymphocytes and activation of microglia and their interplay are the major pathophysiological events leading to neurodegeneration in MS. Our studies in Mouse Hepatitis Virus (MHV) induced neuroinflammatory model demonstrated a protective role of CNS infiltrating CD4+ T cells. In the absence of CD4+ T cells, microglial activation fails to resolve, and mice are more susceptible to acute poliomyelitis and chronic demyelination with axonal bulbar vacuolation. Our studies also revealed that CD40L (expressed on CD4+ T cells) upregulates upon MHV infection but is downregulated in CD4−/− mice CNS. This led to a further delineation of the CD4-microglia nexus at the molecular level using CD40L−/− mice. Results showed that the absence of CD40L renders mice highly susceptible to MHV infection due to reduced microglia/macrophage activation and significantly dampened effector CD4+ T recruitment to the CNS at the acute-adaptive bridging phase (day 7–10 p.i.) of inflammation. Moreover, CD40L−/− mice exhibited severe demyelination, axonal loss, and persistent poliomyelitis at the chronic phase of infection, highlighting the protective role of CD40-CD40L in MHV induced neuroinflammatory demyelination. Together, these studies highlight that migration of peripheral T cells and their interaction with microglia via CD40-CD40L is essential to eliminate the virus and provide long-term neuroprotection. These findings can lead to designing potential therapeutic interventions against MS, targeting CD40-CD40L interaction.
Eos differentially regulates Th1 versus Tfh differentiation through feed-forward propagation of the IL-2/STAT5 signaling pathway
Authors: Read Kaitlin , Sreekumar Bharath , Powell Michael Duane , Jones Devin M , Pokhrel Srijana , Saadey Abbey , Collins Patrick , Ghoneim Hazem , Oestreich Kenneth J 
Published: February 2023

Abstract
The Ikaros zinc finger (IkZF) protein Eos is a transcription factor with known roles in driving regulatory T cell (Treg) suppressive function. However, whether it regulates the differentiation of effector CD4+ T cell subsets is currently unknown. Here, we find that Eos deficiency compromises both expression of the T helper 1 (Th1) differentiation program and effector function in vitro and in vivo, including production of IFN-gamma and TNF-alpha. Despite historical associations between IkZF factors and changes in gene expression via the alteration of chromatin structure, our ATAC-seq analysis indicated that there were minimal changes in chromatin accessibility between wildtype and Eos-deficient cells cultured under Th1-polarizing conditions. Rather, we find that IL-2 signaling promotes the STAT5-dependent induction of Eos expression, and that Eos both interacts with STAT5 and supports tyrosine phosphorylation-mediated STAT5 activation. Thus, STAT5 enrichment is reduced at Th1 target genes in the absence of Eos, including Prdm1, which encodes Blimp-1, an antagonist of the T follicular helper (Tfh) cell transcriptional program. Consequently, we find that Eos-deficient T cells also exhibit increased expression of signature Tfh genes. Together, our findings indicate that modulation of IL-2/STAT5 signaling represents a novel mechanism by which Eos regulates the Th1 and Tfh gene programs. Given the important role for IL-2/STAT5 signaling in both the positive and negative regulation of other T helper cell subsets, it will be of interest to determine whether this mechanism is conserved across additional CD4+ T cell populations.
TET-mediated regulation of effector CD4 T cells
Authors: Johnson Jordan T , Issuree Priya , Butler Noah 
Published: February 2023

Abstract
Functionally distinct subsets of pathogen-specific CD4 helper T cells, including T follicular helper (Tfh) and T helper type 1 (Th1) subsets, are critical for orchestrating the induction and maturation of innate and adaptive immune responses. Proper genetic programming of Tfh and Th1 differentiation is required for regulating immune responses to clear primary infections and promote long-lived memory CD4 T cell-mediated protection against pathogen re-exposures. Genetic approaches to study CD4 T cell differentiation have been largely focused on transcriptional programming, but gene regulation is also critically tied to epigenetic modifications, which represents a major knowledge gap in our understanding of CD4 T cell biology. Ten-eleven-translocation (TET) family enzymes facilitate DNA demethylation to influence chromatin accessibility and promote gene expression. Prior work has shown that TET2 activity has been linked to optimal Th1 responses in vitro. TET1 and TET3 proteins mediate Cd4 locus demethylation and are required for stable CD4 expression during thymocyte development. However, the precise role of either TET1 or TET3 activity to mediate CD4 T cell fate and function in vivo has not been directly investigated. Thus, we hypothesized that TET1 and TET3 activity in peripheral, mature CD4 T cells is required for regulating T helper cell differentiation and function. We used conditional genetic and chimeric approaches, coupled with robust systems of infectious disease immunology, to directly test our hypothesis. Our data show substantial CD4 T cell developmental skewing towards Tfh fates in the absence of specific TET family members. This suggests that TET1 and TET3 enzymes play a role in governing Tfh and Th1 CD4 cell fate.
Inhibition of GATA3 and Th2 cytokines expression in type 2 high asthmatics by a small molecule targeting RNA-binding protein HuR
Authors: Fattahi Fatemeh , Ellis Jason S , Sylvester Michael , Bahelda Kristin , Hietanen Sam , Correa Luis O , Lugogo Njira , Atasoy Ulus 
Published: February 2023

Abstract
RNA-binding protein human antigen R (HuR) is a master regulator of gene expression in human pathophysiology. Our animal studies in the past showed multiple critical roles for HuR in regulating proinflammatory markers in CD4+ T cells. We hypothesized HuR plays an important role in expression of GATA3 and Th2 cytokines in type 2 high asthma and its inhibition impairs these Th2 inflammatory responses. Peripheral CD4+ T cells were isolated from 26 healthy controls and 32 type 2 high asthmatics (blood eosinophil ≥300 cells/μl or FeNO levels ≥25ppb) and stimulated with anti-CD3/CD28 for 4 days, followed by determination of GATA3 and cytokines expression by flow cytometry and LEGENDplex. HuR levels were quantitated by Western blot, flow cytometry, and qPCR. Inhibition of inflammatory responses was evaluated using a cumarin-derived small molecule named CMLD-2 (a HuR-specific inhibitor). Actinomycin D treatment was used to measure mRNA decay rates of targeted HuR mRNAs. HuR protein levels in CD4+ T cells was significantly higher in type 2 high asthmatics compared to healthy controls. The expression and secretion of Th2 cytokines were significantly higher in asthmatics compared to controls. Our further mRNA and protein studies in type 2 high asthmatics showed reduced expression of GATA3 and Th2 cytokines after CMLD-2 treatment. There was also a significant reduction in gata3 mRNA stability with CMLD-2 treatment, compared with vehicle in the asthmatics. Our study is the first to demonstrate the role of RNA-binding protein HuR in type 2 high asthmatics and that interfering with HuR action using CMLD-2 ameliorates GATA3 and Th2 cytokines production. This data suggests HuR may serve as a novel and powerful therapeutic target to treat type 2 high asthmatics.
Student Application of Immunology Concepts and Mechanisms Through Test Based Learning on Canvas
Authors: Swanson-Mungerson Michelle Anne , Laddaga Richard , Szul Martin J. , Volin Michael V 
Published: February 2023

Abstract
Application of immunological concepts is a major challenge for students, especially during a pandemic when most students are learning remotely. Many students memorize the details, but often fail to critically process the information to form a comprehensive understanding of how the immune system works. To address this concern, we developed a “test-based learning” approach to engage students and reinforce important concepts. We converted our immunology lecture-based cases to a quiz format that was presented through our Learning Management Software (Canvas). This approach provides students the opportunity to remotely apply the information presented in lecture either independently or in a small group format through Teams. These cases engaged students to discuss the important aspects of the case and the immunological mechanisms involved. If students struggled, Faculty Members were available via Teams video chat or email to answer questions. We realized that this approach allowed for increased student participation, more active learning, and increased questions to faculty members in comparison to asking questions during lecture. A final advantage of this method is that faculty were able to quickly assess if students mastered key concepts by analyzing students’ scores. If faculty identified deficiencies, there was ample time to address misconceptions before an exam. This approach can be easily modified and used in any course that uses Learning Management Software (e.g. Canvas or Blackboard). Furthermore, this approach is not only applicable to medical education, but could be used at both the undergraduate and graduate levels.
Using the H5P digital platform as an active learning tool to build content-based critical thinking skills in an undergraduate immunology course
Authors: Carr William H 
Published: February 2023

Abstract
Recent studies have demonstrated that active learning or learning by doing has a more significant positive impact on learning outcomes than reading or watching videos. However the availability of free and open source tools for creating these active learning exercises has been limited by the requirement of computer programing skills. With the development of H5P technology, an open source, on-line tool, faculty and even undergraduate students can create interactive exercises that facilitate the learning process and promote formative learning. H5P is a plugin for existing publishing systems that enables the system to create interactive content, such as multiple-choice quizzes that provide feedback in real-time on quiz responses. Here are provided scaffolded assignments to enable implementation of this approach in an undergraduate immunology course that is taught online asynchronously. This activity was implemented at Medgar Evers College, City University of New York (Brooklyn, NY) in 2020–2021. These assignments enabled students to design interactive formative assessments, i.e., multiple choice questions, using the H5P digital platform (www.h5p.org). Results of undergraduate student surveys on their subjective impressions of using this tool are presented. Specifically, the learning outcome that this activity addressed was to increase understanding of the regulation of V(D)J recombination, which is often a difficult learning concept in undergraduate immunology courses. This is teaching innovation will also be presented as a workshop in the Education Session of Immunology 2021.
Engagement with In-class Activities Contributes to Student Success in an Accelerated, Flipped Classroom, Medical School Immunology Course
Authors: Steinel Natalie C , Corliss Stephanie , Lee Michael 
Published: February 2023

Abstract
In an era of decreasing basic science curriculum at medical schools, we sought to re-imagine how to optimally deliver three core basic science disciplines (immunology, microbiology, and pharmacology) together with infectious disease in a 5-week course. This course was developed as part of a new 1-year pre-clinical basic science curriculum at the recently established Dell Medical School (DMS) at the University of Texas at Austin. In line with the goals and missions of the DMS, course material was covered using primarily self-directed and flipped classroom pedagogies that incorporated approaches from the educational and cognitive psychology literature. Attendance and participation in flipped classroom activities was optional, and the materials and activities from these sessions were made available to all students. Analysis of student data of our integrated course shows a correlation between attendance in flipped classroom sessions and summative exam performance. We also observed a strong correlation between student performance on no-stakes formative assessments (weekly competency quizzes that did not count towards their course grade) and the summative exam as well. This suggests that, despite the abbreviated course time, engaging with the course as designed impacts achievement and may lead to superior outcomes. This offers valuable information that can be used to advise students on how to be successful in this type of high-impact course. It also may afford us the ability to predict which students may be at academic risk prior to the final summative exam.