bluuebunny/crossref_metadata_2025_split huggingface dataset:
Extensive characterization of islet-reactive CD8 T cells in type 1 diabetes by mass cytometry (CyTOF)
Authors: Wiedeman Alice E , James Eddie A , Greenbaum Carla , Long Sarah Alice 
Published: January 2023

Abstract
In type 1 diabetes (T1D), insulin-secreting beta cells of the pancreatic islets are targets of autoimmune destruction by T lymphocytes, resulting in insulin deficiency and dysregulation of glucose metabolism. CD8 T cells reactive to islet antigens have been implicated in this process, and phenotypic changes in CD8 T cells have been observed in the transition to clinical diabetes. While efforts have led to identification of peptide antigens targeted by autoreactive CD8 T cells, less has been done to characterize the phenotype and function of this population. We developed a 25+ marker mass cytometry (CyTOF) panel to identify and extensively characterize CD8 T cells reactive to pooled class I (HLA-A2) tetramer loaded with peptides of viral or islet antigens in the same sample. Results were confirmed with several smaller flow panels. In PBMCs of three healthy controls and four T1D patients, we found virus-specific CD8 T cells had a more memory- and exhausted-like phenotype compared to total CD8 T cells, including increased T effector memory (CD45RO+CCR7−), KLRG1, CD57, TIGIT, and PD1, and decreased naïve T cells (CD45RO−CCR7+). In contrast, islet-specific CD8 T cells (detectable in three T1D patients) did not vary significantly from total CD8 T cells. Given the differences in phenotype between viral and total CD8 T cells, we successfully used unsupervised clustering analysis software to reveal a distinct virus-specific CD8 T cell population. This approach of using pooled tetramer for detection of total viral and autoreactive CD8 T cells in the same sample, in combination with extensive phenotyping by mass cytometry will be useful in monitoring important phenotypic changes in this population during disease progression and in response to therapy.
AMPK-dependent and independent effects of AICAR and Compound C on T-cell responses
Authors: Rao Enyu , Zhang Yuwen , Li Qiang , Hao Jiaqing , Egilmez Nejat , Li Bing 
Published: January 2023

Abstract
As a master metabolic sensor, AMP-activated protein kinase (AMPK) is involved in different fundamental cellular processes. Regulation of AMPK activity either by agonists (e.g., AICAR) or by antagonists (e.g., Compound C) has been widely employed to study the physiological functions of AMPK. However, mounting evidence indicates AMPK-independent effects for these chemicals. Herein, using T cells from AMPK conditional knockout mice and their wild type littermates, we demonstrate that AICAR and Compound C can, indeed, activate or inhibit AMPK activity in T cells, respectively. Specifically, AICAR inhibits, but Compound C promotes, Ca2+-induced T cell death in an AMPK-dependent manner. In contrast, our data also demonstrate that AICAR and Compound C inhibit T cell activation and cytokine production in an AMPK-independent manner. Moreover, we find that the AMPK-independent activity of AICAR and Compound C is mediated via the mTOR signaling pathway in activated T cells. Our results not only reveal the critical role of AMPK in regulating T cell survival and function, but also demonstrate AMPK-dependent and independent roles of AICAR/Compound C in regulating T cell responses, thus suggesting a context-dependent effect of these “AMPK regulators”.
A checkpoint requiring cognate antigen recognition at the effector stage is necessary for protective memory CD4 T cell generation.
Authors: Bautista Bianca L , Devarajan Priyadharshini , McKinstry K. Kai , Strutt Tara M , Swain Susan L 
Published: January 2023

Abstract
Effective vaccination depends on the generation of memory T cells, B cells, and long-lived plasma cells. However, the mechanisms that govern memory CD4 T cell formation are not well understood. Antigen (Ag) recognition during priming drives effector generation, but it is unclear how Ag recognition at the effector stage impacts T cell fate. Using an influenza infection model, we find that effector CD4 T cells must re-encounter Ag at the effector phase to avoid default apoptosis and become memory. Cognate interaction at this defined “memory checkpoint” drives continued proliferation, enhances survival, upregulates memory markers CD25, CD127, CXCR3, and Bcl-6, and drives IL-2 production, resulting in the formation of long-lived memory cells with enhanced production of multiple cytokines and increased protective function. A cold-adapted influenza virus, like those used in live-attenuated influenza vaccines, fails to present Ag at this critical effector time point and fails to induce substantial CD4 memory. However, memory CD4 T cell formation was rescued by adding short-lived Ag-pulsed antigen presenting cells during the checkpoint. Importantly, the Ag alone without infection-induced inflammation, was sufficient to restore memory formation. Our findings imply that in order for vaccines to promote optimal memory CD4 T cell formation, they must provide Ag presentation long enough that effector CD4 T cells are effectively restimulated during the “memory checkpoint”.
Induction of B10 function on ligature-induced experimental periodontitis
Authors: Hu Yang , Yu Pei , Yu Xinbo , Hu Xingxue , Han Xiaozhe 
Published: January 2023

Abstract

Objective
IL-10 producing B cells (B10 cells) play an essential role in immune system balance by suppressing excessive inflammatory responses. This study is to investigate induction of B10 function in vitro and their effect on ligature-induced experimental periodontitis in vivo.

Methods
Spleen B cells were isolated from C57BL/6J mice and cultured for 48 hours under the following conditions: control, CD40L (1 μg/ml), IL21 (25, 50, 100, 1000ng/ml), anti-Tim1 (2.5, 5, 10, 20 μg/ml), CD40L + IL21, CD40L + anti-Tim1, CD40L + IL21 + anti-Tim1. The mRNA level and secreted protein level of IL-10 expression were detected by RT-qPCR and ELISA respectively. Silk ligatures were tied around both maxillary second molars of C57BL/6J mice for two weeks. Optimized combination of CD40L, IL21 and anti-Tim1 and vehicle were injected into contralateral side of palatal gingiva on days 3, 6 and 9. The palatal gingival tissues and maxillary bone were collected on day 14 to determine expressions of IL10 and RANKL and periodontal bone resorption respectively.

Results
IL-10 expressions of cultured spleen B cells were significantly increased in the presence of CD40L, IL21 and anti-Tim1 combination when compared with control groups. Gingival IL-10 expression were significantly increased after injection of CD40L, IL21 and anti-Tim1 combination, when compared to the control side (p&lt;0.05). The gingival RANKL expression and periodontal bone loss were significantly decreased on the side with combined injection of CD40L, IL21 and anti-Tim1, as compared to the control side (p&lt;0.05).

Conclusion
CD40L, IL21 and anti-Tim1 combination induced IL-10 production by B cells and inhibited periodontal bone loss in ligature-induced experimental periodontitis.
Mechanism of central and peripheral T cell tolerance to an antigen of the central nervous system
Authors: Wang Lei , Hinterberger Maria , Winnewisser Julia , Federle Christine , Klein Ludger 
Published: January 2023

Abstract
Autoreactive CD4+ T cells are censored by two mechanisms, the so-called clonal deletion and the deviation into regulatory T cells in the thymus both requiring the same stimulus, namely the interaction with self-peptide. In order to study the mechanisms of central and peripheral tolerance that operate to shape the CD4 T-cell repertoire, we have generated a TCR transgenic mouse specific for myelin proteolipid protein (PLP). We found that PLP-specific T cells undergo clonal deletion and Treg differentiation concomitantly upon encounter of the cognate self-antigen PLP. Medullary thymic epithelial cells (mTECs) express and present the endogenous antigen PLP and mediated tolerance in an autonomous manner, whereas thymic dendritic cells (DCs) were dispensable for central tolerance induction to PLP. Furthermore, the transcriptional regulator AIRE expressed by mature mTEC is essential for tolerance induction of PLP-specific CD4 T cells. Although central tolerance induction to PLP is very potent it is not complete as a proportion of autoreactive T cells also escape to the periphery. However despite the presence of potentially dangerous cells in the periphery, the mice do not develop autoimmunity. Thus, additional tolerogenic mechanisms promote tolerance in the periphery exist. We could further show that autoreactive PLP-specific T cells are deleted or become functionally inactivation (anergy) by recognition of self-peptide on DCs in the periphery. In a set of bone marrow transplantation experiments we found that PLP is expressed by radio resistant stromal cells and subsequently cross-presented by dendritic cells. Taken together our results indicate a complementing role of the thymus and the periphery for tolerance induction to PLP.
Etv5 promotes IL-10 production in Th2 cells
Authors: Koh Byunghee , Kaplan Mark H 
Published: January 2023

Abstract
The anti-inflammatory cytokine, IL-10, suppresses inflammatory responses at mucosal surfaces. Among T helper cell subsets, Th2 cells produce the highest concentrations. However, the transcription factors that regulate Il10 gene expression have not been completely defined. Etv5 is a member of the ETS transcription factor family that promotes production of IL-17 in Th17 cells and IL-9 in Th9 cells. To define the effects of Etv5 deficiency on cytokine expression in Th2 cells, T cells from CD4 specific Etv5 knockout (Etv5fl/flCD4-Cre+, termed Etv5 KO below) and littermate control (Etv5fl/fl CD4-Cre-, termed WT below) mice were differentiated under Th2 conditions. Etv5 was required for maximal IL-10 production and modestly repressed the Th2 cytokines IL-4, IL-5 and IL-13. In mice sensitized with Aspergilus fumigatus extract, IL-10 production in both BAL and lung was significantly reduced in Etv5 KO mice, consistent with in vitro data. Using ChIP and reporter assays, we showed that Etv5 directly binds to the Il10 locus, specifically at the CNS3 region. Etv5 promotes IL-10 production by recruiting IL-10 inducing transcription factors including Jun family members, IRF4 and E4BP4, and the binding of these factors to the Il10 locus, but not the expression of these factors, was decreased in the absence of Etv5. Ectopic GATA3 and E4BP4 expression recovers IL-10 production to wild type amounts, suggesting that Etv5 enhances the efficiency of function of these factors. Importantly, ectopic Etv5 expression in Etv5 KO Th2 cells restored binding of the factors to the Il10 locus. Thus, Etv5 promotes IL-10 production by enhancing the recruitment of IL-10 inducing transcription factors.
Regulation of β1 Integrin recycling, cell migration and focal adhesion turnover by CD13
Authors: Ghosh Mallika , Thangada Shobha , Devarakonda Charan , Shapiro Linda H 
Published: January 2023

Abstract
CD13 is a multifunctional cell surface peptidase that is expressed on a variety of cells where we have shown that it modulates receptor-mediated endocytosis and ligand internalization to control downstream signaling pathways. In fibroblasts and epithelial cells, CD13 is localized with β1-integrin at focal adhesions (FAs), the sites of communication between the extracellular matrix and the actin cytoskeleton, prompting our exploration of potential contributions of CD13 focal adhesion turnover, integrin endocytosis and trafficking. Phenotypically, FAs in CD13KO fibroblasts are elongated and irregular with displaced FA accessory proteins, markedly reduced actin stress fibers and fewer microtubule extensions, consistent with a link between CD13 and the control of FA dynamics. In wild type cells, CD13 and β1-integrin co-internalize, traffic to EEA1+ Rab5+ early endosomes and recycle to the plasma membrane via Rab11a+ recycling endosomes. Pulse-chase assays with wild type and CD13 phospho tyrosine mutant confirmed that tyrosine phosphorylated CD13 must associate with the ARF6 GTPase for β1-integrin recycling to the membrane to occur. Conversely, the absence of CD13 results in trafficking of internalized β1-integrin from early endosomes to Rab7+ late endosomes and lysosomes and a failure to return to the cell surface. Functionally, in migrating cells CD13 accumulates at the leading edge and co-localizes with IQGAP1 and focal adhesion protein, Paxillin to regulate cell migration and adhesion. In conclusion, CD13 is responsible for proper localization of focal adhesion proteins, coordination of integrin recycling and focal adhesion dynamics, thereby regulating integrin signal transduction and cell motility.
Insights into MHC-I peptide loading obtained from the structure of a TAPBPR/MHC-I complex
Authors: Natarajan Kannan , Jiang Jiansheng , Boyd Lisa F , Morozov Giora I , Mage Michael G , Margulies David H 
Published: January 2023

Abstract
The cell surface expression of MHC-I molecules displaying peptides derived from intracellular proteolytic processing is a critical requirement for T cell-mediated immunity. A complex series of steps, guided by the peptide loading machinery in the endoplasmic reticulum, ensures that only stably folded MHC-I molecules carrying peptides of proper length and high affinity are transported for display at the cell surface. TAP-binding protein, related (TAPBPR), has been recently shown to directly bind to certain MHC-I alleles and to facilitate peptide loading. To elucidate mechanistic details, we have examined the binding interaction between purified recombinant TAPBPR and MHC-I molecules loaded with truncated peptides that leave a portion of the peptide binding groove unoccupied. Such MHC-I molecules interact with TAPBPR with nanomolar affinities as measured by surface plasmon resonance and these TAPBPR/MHC-I complexes dissociate when offered a high affinity peptide. An extensive crystallization screen with purified TAPBPR/MHC-I complexes yielded crystals that diffracted to 3.5 Å. The structure of the TAPBPR/MHC-I complex, solved by molecular replacement, and refined at this resolution readily reveals the binding footprint and further structural details of the interaction. Data will be presented on the binding interaction and structural characterization of the TAPBPR/MHC-I complex that reveal insights into the mechanism of TAPBPR- and tapasin-mediated loading of peptides onto MHC-I molecules. (KN and JJ contributed equally to this work).
Cytokine-mediated germinal center reprogramming enhances vaccine efficacy
Authors: Pravetoni Marco , Laudenbach Megan , Baruffaldi Federico , Robinson Christine , Carter Philipp 
Published: January 2023

Abstract
Candidate vaccines against infectious agents and non-communicable diseases consist of purified proteins, proteins conjugated to synthetic peptides or haptens, and use adjuvants or novel delivery platforms. Despite pre-clinical proof of concept, vaccine clinical efficacy is often limited to a small subset of immunized subjects. Additionally, very few adjuvants meet regulatory approval. Thus, more effective immunization strategies are needed. This study tested the hypothesis that vaccine efficacy is enhanced by cytokine-based immunomodulators that guide B and T cells toward germinal center (GC) activation and generation of effective antibody (Ab) responses. To this end, a series of cytokines and monoclonal Ab against cytokines or cytokine receptors was co-administered with a candidate therapeutic vaccine for treatment of opioid abuse. Immunization with a drug-hapten conjugated to immunogenic carriers generates anti-drug Ab that block drug distribution to the brain, and reduce drug-induced behavioral effects. In this study, targeting IL-4 signaling enhanced vaccine efficacy in blocking opioid distribution to the brain in mice. Vaccine efficacy was associated with increased vaccine-specific GC B cell and GC-TFH cell differentiation, serum anti-opioid IgG Ab levels and IgG subclasses shift, and a balanced Th1/Th2 environment. No side effects were found. This strategy generalized to a model peptide-protein conjugate immunogen and an approved tetanus-diphtheria-pertussis vaccine. These data support targeting cytokine signaling to safely program vaccine-induced GC formation and achieve clinically effective Ab. This strategy provides a blueprint for rational development of more effective next-generation vaccines.
A novel vaccine vector: herpes simplex virus type-2 deleted in glycoprotein D (HSV-2 ΔgD) and expressing modified influenza A (IAV) hemagglutinin (HA) antigens
Authors: Dardick Joseph , Weiss Kayla , Lukose Regy , Weinrick Brian , Herold Betsy , Jacobs William R 
Published: January 2023

Abstract
Herpes simplex virus types 1 and 2 (HSV-1 and HSV-2) cause significant global morbidity and mortality. In collaboration, the Jacobs and Herold laboratories generated an attenuated herpes simplex virus type 2 (HSV-2) strain deleted in glycoprotein D (HSV-2 ΔgD) and demonstrated that it triggers an FcγRIV activating response that elicits both antibody-dependent cell-mediated cytotoxicity and phagocytosis. The antibodies rapidly clear virus and prevent the establishment of latency in mice and guinea pigs following challenge with several clinical isolates of both HSV-1 and HSV-2. We therefore propose that HSV-2 ΔgD is an ideal vaccine vector to induce immunity against a wide array of pathogens.
The HSV genome is difficult to work with due to high GC content, repetitive regions, and tendency toward attenuation. Using Gibson cloning and homologous recombination we introduce exogenous genes such as red fluorescent protein (rfp) and influenza A (IAV) hemagglutinin (HA) into the HSV genome. IAV HA is a great model antigen to study the immunogenicity of proteins inserted into the HSV-2 ΔgD backbone. Moreover, cloning both full-length and stalk HAs into the HSV-2 ΔgD genome will test the influence of immunodominant antigens on the character of our antibody response.
Our expression vectors, verified by digestion and sequencing, encode a chimeric gD/HA protein that contains the signal sequence and cytosolic and transmembrane domains of HSV-2 gD. These modifications will promote trafficking of HA to the cell membrane and viral envelope. Our recombination system has produced an RFP+ HSV-2 ΔgD virus that we verified by fluorescence, digestion and sequencing. The kinetics of infection with the RFP+ virus match that its parental HSV-2 ΔgD strain.
Induction of Virus-specific Immune Tolerance in A Chronic Hepatitis B Virus Mouse Model
Authors: Lan Yu Hua , Lo Yu-Chieh , Wu Ping-Yi , Tao Mi-Hua 
Published: January 2023

Abstract
Hepatitis B virus (HBV) infection causes severe public health problem. There are estimated 250 million people being chronically infected by HBV and are at an increased risk of developing end-stage liver diseases, such as cirrhosis and hepatocellular carcinoma. It is well recognized that the chronicity of HBV infection is due to the impaired immune responses to HBV and thus not able to eliminate HBV from the infected hepatocytes. However, the immune mechanisms that lead to immune tolerance and HBV persistence have not been understood. A challenging question for HBV studies is lack of appropriate animal models due to the narrow host range of HBV. To address this challenge, we created a chronic HBV infection model in immunocompetent mice by transduction of adeno-associated virus serotype 8 (AAV8) vector which carries and these mice was transduced with carrying a replication-defective HBV genome with two point mutations in the polymerase(AAV/HBVp−). We showed that mice transduced with AAV/HBVp− produced HBV RNA and proteins, including serum HBV surface antigen and HBV e antigen as well as HBV core protein. The high levels of HBV RNA and proteins sustained for at least 1 year post AAV transduction. Viral transcripts and proteins were detected in the livers of the AAV-injected mice but not in other organs. DNA immunization of these AAV/HBVp− transduced mice failed to induce HBV-specific humoral and cellular immune responses, suggesting these mice developed immune tolerance toward HBV. Thus, the AAV/HBVp− transduced mouse model recapitulates many virological and immunological characteristics of chronic HBV in humans. This animal model could be useful for the development of new treatments and immune-based therapies for chronic HBV infections.
Zika virus infects human Sertoli cells and trespass the blood-testes barrier to migrate across the seminiferous epithelium
Authors: Siemann David N , Strange Daniel , Maharaj Payal , Verma Saguna 
Published: January 2023

Abstract
The 2015–2016 epidemic of Zika virus (ZIKV) has identified sexual transmission as a new route of disease spread. Presence of ZIKV in the semen when viremia is undetectable in humans suggests the ability of ZIKV to establish infection in the testes. Lack of specific therapeutic drugs to clear testicular infection warrants understanding of the associated mechanisms. Seminiferous tubules, the site of spermatogenesis, is an immune privileged site with an effective blood-testes barrier (BTB) constituted exclusively by specialized tight junctions proteins (TJP) between the adjacent Sertoli cells (SC) that protects germ cells from systemic immune attack. Increased inflammatory mediators such as TNF-α, IL-1β, matrix metalloproteinases and cell-adhesion molecules (CAMs) can disrupt BTB and facilitate testicular infections. We determined the infection kinetics of ZIKV in SC and its association with antiviral defense and BTB integrity. We demonstrate that SC were susceptible to ZIKV and induced cytokines (IFN-α, IFN-γ, TNF-α) and CAMs (VCAM), which correlated with peak virus titers. Interestingly, while ZIKV transmigrated efficiently across the in vitro BTB model, it did not affect barrier integrity, also supported by no difference observed in the levels of TJP such as ZO-1 and claudin between control and infected SC. However, the integrity of the BTB model decreased significantly following exposure to the supernatant from ZIKV-infected macrophages. Collectively, our data suggests infection of SC as one of the routes of virus transmigration into the seminiferous tubules and that inflammatory mediators from infected human macrophages may further compromise integrity of BTB thereby allowing persistent infection of ZIKV in the testes.
HVEM expression by intestinal epithelial cells modulates the microbiome and epithelial immunity
Authors: Seo Goo-Young , Shui Jr-Wen , Mikulski Zbigniew , Cheroutre Hilde , Kim Pyeung-Hyeun , Kronenberg Mitchell 
Published: January 2023

Abstract
The herpes virus entry mediator (HVEM), a member of the tumor necrosis factor receptor superfamily (TNFRSF), is highly expressed by intestinal epithelial cells (IEC). Previously, we reported that HVEM expression by epithelial cells was required for innate immune defense from acute infections in the lung and the intestine (Nature 488:222,2012). Here, we demonstrate a novel, constitutive role of HVEM expression by IEC in microbial homeostasis and epithelial immunity. Mice with an epithelial-specific deletion of the gene encoding HVEM (HvemΔIEC) had significantly increased segmented filamentous bacteria (SFB). This caused an increase in Th17 cells and IL-22+ILC3s in the intestine lamina propria. HvemΔIEC mice also exhibited increased goblet cell hypertrophy and hyperplasia, crypt hyperplasia, villous atrophy as they aged, and they showed a severe defect in fucosylation of cell surface proteins, even at younger ages. Treatment with the antibiotic vancomycin eliminated SFB and decreased Th17 cells and ILC3, but did not reverse the defect in fucosylation. Our findings suggest that epithelial cell expression of HVEM is important at steady state both for shaping the intestinal microbiota and for the homeostasis of IEC in the intestine.