bluuebunny/crossref_metadata_2025_split huggingface dataset:
Dynamic monitoring of pathological and immunological changes and treatment effect of killer artificial antigen presenting cells in a murine experimental autoimmune encephalomyelitis model
Authors: wan xin 
Published: January 2023

Abstract
Multiple sclerosis (MS) is a chronic, autoimmune disease of the central nervous system. Experimental autoimmune encephalomyelitis (EAE) are widely used to study the pathological features and mechanisms underlying MS. Here, a murine EAE model was induced. The disease degree was daily assessed; The local inflammatory infiltration was observed and the expressions of MBP and GFAP were detected. The frequencies and reactivity of MOG antigen-specific CD4+ T cell and CD8+ T cells were concurrently investigated. As a result, the dynamic changes of inflammation, myelin loss and astrocyte proliferation in CNS were highly consistent with clinical manifestations observed in EAE course. Data also emphasize the pathogenic roles of both CD4+ T cells and CD8+T cells in the development of EAE, while their reactivity in response to the antigen was different from their dynamic changes of frequencies.
Killer artificial antigen presenting cells (KaAPCs) are thought to be a powerful tool to inhibit autoimmune effect and kill autoreactive T cells avoiding systemic immune suppression. Here, a KaAPC was generated by covalently co-coupling targeted binding molecules (H-2Db/MOG40-54 dimmer, H-2I-Ab/MOG35-55 multimer), negative regulation molecules (anti-mouse Fas mAbs, PD-L1-Ig, TGF-β1) and antiphagecytic molecules (CD47-Ig) onto poly lactic-co-glycolic acid (PLGA) microspheres. We found decreased clinical score of EAE mice after several times treatment of KaAPCs.
Collectively, this study presented a clear histopathologic and immunologic pattern of mouse EAE model, which may contribute to better understanding of the EAE process. Also, KaAPCs are feasible strategy for treating EAE, which lead to a new approach for MS therapies.
NKG2D signaling between human NK cells enhances TACE-mediated TNF-α release
Authors: Markiewicz Mary A , Sharma Neekun 
Published: January 2023

Abstract
Natural killer group 2 member D (NKG2D) is a strong NK cell activating receptor, with engagement by ligands triggering granule release and cytokine production from NK cells. The function of NKG2D signaling in NK cells has largely been studied in the context of engagement of the receptor by ligands expressed on the surface of target cells, such as tumor cells. Here we report that upon activation with IL-12, IL-15 and IL-18, human NK cells expressed NKG2D ligands of the UL16 binding protein (ULBP) family on the cell surface. NKG2D-ULBP engagement during homotypic interaction between cytokine-stimulated NK cells increased the activity of the metalloprotease TNF-α-converting enzyme (TACE). This enhanced TACE activity significantly increased the release of TNF-α and ULBP proteins from the surface of the NK cells. These results demonstrate that NKG2D signaling is critical for maximal TNF-α release by NK cells. Further, they are the first to demonstrate a role for NKG2D-ligand interaction via homotypic NK cell contact in NK cell effector function.
Neutrophil Cathepsin G inhibits hapten-presenting dendritic cell production of IL-12 and skewing of hapten-reactive CD4 T cell development to IFN-γ- and IL-17-producing effector cells in allergic contact sensitivity
Authors: Kish Danielle , Min Susie , Fairchild Robert L 
Published: January 2023

Abstract
Contact hypersensitivity (CHS) is a CD8 T cell-mediated response to hapten skin sensitization and challenge. Sensitization of wild-type (WT) mice induces hapten-reactive CD8 T cells that produce IFN-γ and IL-17 and mediate CHS and hapten-reactive CD4 T cells producing IL-4 that cannot mediate CHS. Although CXCR2-dependent neutrophil recruitment into the hapten challenged skin is required to direct effector CD8 T cell infiltration into the site to elicit CHS in WT mice, CHS was elicited following challenge of hapten sensitized CXCR2−/− mice. Sensitization of CXCR2−/− mice or antibody-mediated depletion of neutrophils during sensitization of WT mice generated hapten-reactive CD4 and CD8 T cells producing IFN-γ and IL-17 and either T cell population could mediate CHS independently of the other. The hapten-reactive CD4 cells producing IFN-γ and IL-17 and their CHS responses were not generated during hapten sensitization of IL-12−/− mice depleted of neutrophils or WT mice depleted of neutrophils and treated with anti-IL-12 mAb. Neutrophil depletion during hapten sensitization of WT mice markedly increased numbers of IL-12-producing dendritic cells in the skin draining lymph nodes. Sensitization of mice lacking the neutrophil serine protease cathepsin G induced hapten-reactive CD4 and CD8 T cells producing IFN-γ and IL-17 with elevated and elongated CHS responses to challenge. Induction of CHS effector CD4 T cells in neutrophil-depleted WT mice was eliminated by subcutaneous injection of recombinant cathepsin G during sensitization. Thus, hapten skin sensitization induces neutrophil production of cathepsin G that regulates hapten-presenting dendritic cell production of IL-12 and the development of CHS effector CD4 T cells.
Skin-resident effector memory CD8+CD28− T cells exhibit a pro-fibrotic phenotype in patients with systemic sclerosis
Authors: Fuschiotti Patrizia , Larregina Adriana T , Domsic Robyn T , Stolz Donna B , Medsger Thomas A , Lafyatis Robert A 
Published: January 2023

Abstract
Loss of CD28 expression by CD8+ T cells occurs with age and during chronic inflammatory conditions. CD8+CD28− T cells are a heterogeneous cell subpopulation whose function ranges from immunosuppressive to effector. Here we analyzed the role of CD8+CD28− T cells in the pathogenesis of systemic sclerosis (SSc), a connective tissue disorder characterized by autoimmunity, vasculopathy and extensive cutaneous and visceral fibrosis. We show that the frequency of CD8+CD28− T cells is increased in the blood and affected skin of SSc patients, independent of patient age, and correlates with the extent of skin fibrosis. We found that the majority of skin-tropic CD8+CD28− T cells are resident in the skin lesions of patients in the early stage of the disease, exhibit an effector memory phenotype and present a strong cytolytic activity ex vivo. Skin-resident and circulating SSc CD8+CD28− T cells produce high levels of the pro-fibrotic cytokine IL-13, which induces collagen production by normal and SSc dermal fibroblasts. Thus, our findings indicate that CD8+CD28− T cells represent a pathogenic T-cell subset in SSc and likely play a critical role in the early stage of SSc skin disease.
Group 2 Innate Lymphoid Cells Recruit and Activate Airway Macrophages to facilitate Pulmonary Epithelial Regeneration
Authors: Copenhaver Alan M , Dagher Rania , Berlin Aaron A , Besnard Valerie , Maret Marielle , Hamidi Fatima , Aubier Michel , Kolbeck Roland , Pretolani Marina , Humbles Alison A 
Published: January 2023

Abstract
Group 2 innate lymphoid cells (ILC2s) are a subset of ILCs that are resident in many tissues including the lung. These cells respond to epithelial cytokines and in turn potently produce many type 2 associated cytokines, such as IL-4, IL-5, and IL-13. Beyond cytokine production, ILC2s are thought to aid tissue repair after damage by producing growth factors such as amphiregulin or by expressing the enzyme arginase-1. Alternatively activated macrophages and epithelial progenitor cells also play important roles in directing successful repair after damage, but it is not known whether ILC2s work in concert with either of these cells to mediate effective repair. Using a model of sterile epithelial damage, we investigated the role of ILC2s during the course of epithelial repair and regeneration. Lung ILC2s were significantly activated 4–6 days after epithelial damage and produced increased levels of GM-CSF, IL-5 and IL-13. Interestingly, mice lacking ILCs (Rag2−/−Il2rg−/−) had reduced numbers of macrophages in the lung upon injury and subsequently decreased levels of macrophage-derived growth factors, such as IGF-1 and BRP-39. Reconstitution of ILC2s into Rag2−/−Il2rg−/− mice restored macrophage numbers and growth factor expression within the lung. These data suggest that ILC2s are part of a larger, multicellular circuit required for both recruitment and stimulation of alternatively activated macrophages which initiate the epithelial repair process. Thus, ILC2s can mediate both inflammation and repair at barrier sites by coordinating the activation state of specific effector cells.
IL-33/ST2 macrophage crosstalk promotes lung epithelial repair
Authors: Dagher Rania , Copenhaver Alan M , Besnard Valerie , Maret Marielle , Hamidi Fatima , Berlin Aaron A , Aubier Michel , Kolbeck Roland , Humbles Alison A , Pretolani Marina 
Published: January 2023

Abstract
Increasing evidence points to a central role for macrophages in tissue regeneration, however, the molecular mechanisms underlying this function remain unknown. Here, using depletion and adoptive transfer approaches, we demonstrate a major role of alveolar macrophages in facilitating bronchial re-epithelialization. In addition, we identify a novel, and paradoxical, protective role for the IL-33/ST2 axis in epithelial repair following naphthalene (NA)-induced lung injury. We defined two subsets of ST2 expressing myeloid cells, namely recruited monocyte-derived cells and resident alternatively activated macrophages (AAMs), which infiltrate the airways during the regeneration of the bronchial epithelium. Further, ST2-deficient mice exhibited an incomplete epithelial repair which was associated with a dysfunctional AAM phenotype and a reduction in the proliferation of a self-renewing subset of Clara progenitor stem cells. Notably, reconstitution of ST2+ AAMs, post NA, completely restored the epithelium in ST2-deficient animals. In contrast, the monocyte/macrophage-dependent CCL2/CCR2 axis was redundant for repair, however, anti ST2-antibody treatment to CCR2−/− mice resulted in ineffective epithelial regeneration and confirmed the observations seen with ST2-deficiency after NA-induced injury. Thus, the IL-33/ST2 pathway regulates effective Clara cell regeneration and bronchial re-epithelialization via AAM polarization and may lead to new therapeutic insights in acute respiratory diseases.
Novel inhibitory receptor signatures revealed by 18 parameter flow cytometry and mapped via a combination of automated analysis algorithms
Authors: Belkina Anna C , Fleury Michelle , Olson Alex , Dooms Hans , Kukuruzinska Maria , Lin Nina , Snyder-Cappione Jennifer 
Published: January 2023

Abstract
Inhibitory Receptors (IRs) such as PD-1, TIM-3, CD160, and TIGIT are upregulated during activated, exhausted, and pathological immune cell states. How the combinational expression of multiple IRs on individual cells tracks with functional status and/or fate is largely unknown. Due to paucity of human samples, large panels for immune cell characterization are of a high value, and combining them with automated platforms of multi-dimensional single cell analysis would allow objective and comprehensive population characterization. We report an 18-parameter (16-color) fluorescent panel that allows IR signature analysis as well as cell sorting for functional studies, and adopt algorithmic methods previously used for mass cytometry into a novel platform to analyze fluorescent datasets in an unbiased analytical workflow. PBMC from diseased and healthy control subjects as well as tumor specimens were stained and analyzed on a FACSARIA cell sorter. For the automated analysis, the datasets were processed with t-SNE algorithm into a 2D map of phenotypic space that was subsequently clustered using SPADE. From the samples, a 32-plex combinational IR signature was determined for the CD4+ T, CD8+ T, NK, iNKT, and γδ T cell lineages. The results indicate novel combinational IR ‘hits’ for particular immune subsets and some immune subset IR signatures correlated with disease status. In conclusion, 18-parameter flow cytometry with automated analysis provides robust results from human blood and solid tissue samples. In combination with functional and other readouts, measurement of single immune cell IR signatures could provide new insight into activation, exhaustion, and pathogenic processes and potentially reveal novel therapeutic targets.
Recombination signal sequences shape antigen receptor repertoires and enforce allelic exclusion
Authors: Wu Glendon S , Yang-Iott Katherine S , Bassing Craig H 
Published: January 2023

Abstract
Antigen receptor loci are arrays of variable (V), diversity (D), and joining (J) gene segments that the RAG endonuclease assembles into complete AgR genes. RAG binds to and cleaves at recombination signal sequences (RSSs) that flank gene segments. V(D)J recombination of Tcrb, Igh, and Igk loci is regulated such that functional AgR genes are assembled on and expressed from one allele (allelic exclusion) in most cells. It is assumed that allelic exclusion is achieved by mono-allelic initiation and feedback inhibition of the V-to-(D)J recombination step, and intrinsic features of RSSs have been proposed to mediate mono-allelic V-to-DJ recombination of Tcrb and Igh loci. We established and analyzed mice with the Trbv2 or Trbv31 RSS replaced by an RSS that possesses greater intrinsic recombination activity. Each of these RSS substitutions causes a profound increase in the development of T cells expressing either Trbv2+ or Trbv31+ TCRβ chains, reflecting that the RSSs flanking Trbv segments are major determinants of Trbv recombination frequency. Each RSS replacement also promotes a modest increase in the fraction of cells expressing Trbv2+ or Trbv31+ TCRβ chains along with another type of TCRβ chain (allelic inclusion). Strikingly, mice with Trbv2 and Trbv31 RSS replacements on opposite alleles have a 30-fold higher than normal level of allelic inclusion for Trbv2+ and Trbv31+ TCRβ chains, demonstrating that RSSs flanking Trbv segments enforce TCRβ allelic exclusion. We conclude that the infrequency of Trbv recombination as determined by inherent inefficiencies of Trbv RSSs limits bi-allelic assembly of Tcrb genes within the time window before feedback inhibition permanently cements allelic exclusion.
Rhbdf1 is a novel target of TRAF3 inactivation in B lymphocytes
Authors: Zhu Sining , Luo Chang , Lalani Almin , Hart Ronald , Xie Ping 
Published: January 2023

Abstract
TRAF3 is a new tumor suppressor gene in B lymphocytes. Deletions and mutations of TRAF3 were detected in human non-Hodgkin lymphoma (NHL) and multiple myeloma (MM). We previously reported that specific deletion of TRAF3 in B lymphocytes causes prolonged survival of mature B cells, which eventually leads to B lymphoma development in mice. Here we identified Rhbdf1, a 7-transmembrane domain protein implicated in the trafficking of ER proteins, as a novel target of TRAF3 inactivation in B cells. Rhbdf1 expression was strikingly up-regulated in TRAF3-deficient premalignant B cells, B lymphomas and human MM at both the mRNA and protein levels. Normal physiological stimuli such as BAFF, CD40 or BCR could not induce the expression of Rhbdf1 in B cells regardless of the presence of TRAF3, suggesting that Rhbdf1 is specifically and aberrantly induced by TRAF3 inactivation in B cells. Biochemical fractionation studies revealed that Rhbdf1 was mainly localized at the ER in mouse splenic B cells and human MM. Interestingly, lentiviral shRNA vector-mediated knockdown of Rhbdf1 induced apoptosis and inhibited proliferation in human MM cells with TRAF3 deletions or mutations. Furthermore, bortezomib, an effective clinical drug for MM, drastically decreased Rhbdf1 protein levels in both TRAF3-deficient mouse B lymphoma and human MM. Taken together, our findings suggest that Rhbdf1 is a novel target of TRAF3 inactivation in B cells and may contribute to oncogenic B cell survival.
IL-7-dependent STAT1 activation limits homeostatic CD4 T cell expansion
Authors: Catalfamo Marta , Le Saout Cecile , Luckey Megan A. , Villarino Alejandro , Smith Mindy , Hasley Rebecca B. , Myers Timothy G. , Imamichi Hiromi , Park Jung-Hyun , O’Shea John J , Lane H. Clifford 
Published: January 2023

Abstract
IL-7 regulates homeostatic mechanisms that maintain the overall size of the T cell pool throughout life. We show that under steady state conditions, IL-7 signaling is principally mediated by activation of Signal Transducers and Activators of Transcription 5 (STAT5). In contrast, under lymphopenic conditions there is a modulation of STAT1 expression resulting in an IL-7-dependent STAT1 and STAT5 activation. Consequently, the IL-7-induced transcriptome is altered with enrichment of interferon (IFN) stimulated genes (ISGs). Moreover, STAT1 overexpression was associated with reduced survival in CD4 T cells undergoing Lymphopenia-Induced Proliferation (LIP). We propose a model in which T cells undergoing LIP upregulate STAT1 protein, “switching on” an alternate IL-7-dependent program. This mechanism could be a physiological process to “sense space” regulating the size of the CD4 T cell pool. During HIV infection the virus could exploit this pathway leading to the homeostatic dysregulation of the T cell pools observed in these patients.
Knockout validation of antibodies to Ki67: a marker for cellular proliferation
Authors: Dreja Hanna S , Ayton Julie , Bruce David , Lochead Julia , Renshaw Simon , Parton Laura , Hamilton Bruce J , Beer Samantha , Munro Melanie , Solache Alejandra 
Published: January 2023

AbstractAt Abcam, providing a high level of validation has always been a priority and our strategies are constantly evolving as new technologies are developed. The recent knockout (KO) validation initiative, made possible through a partnership with Horizon Discovery and the use of their CRISPR/Cas9 KO cell lines, addresses both antibody specificity and reproducibility on a large scale.Ki67 is a proliferation marker and determining its expression levels has prognostic and predictive values in cancer (1–5). Hence, it is critical that antibodies used to target Ki67 are highly specific. This study demonstrates the evolving strategies used in our laboratory to validate antibodies to Ki67. The use of a human haploid cell lines where Ki67 expression has been knocked out has unequivocally identified antibodies that bind specifically to Ki67.Utilising this technology has enabled KO validation of antibodies and, at the time of abstract submission, there are over 650 KO validated antibodies on our catalogue. It is our hope that by using this technique we can help to raise industry standards and contribute to the generation of reproducible research.
Immunology Antibody Validation: PTEN a target case in the use of human KO cell lines for antibody validation
Authors: Ayton Julie S , Dreja Hanna S , Bruce David , Lochead Julia , Koch Sofia , Parton Laura , Hamilton Bruce J , Munro Melanie , Solache Alejandra 
Published: January 2023

AbstractAt Abcam we are constantly improving our manufacturing processes and adopting more robust antibody validation technologies as they are developed. Recently we have implemented the use of human knockout (KO) cell lines to interrogate the specificity of our antibodies on a large scale. This has been possible through a partnership with Horizon Discovery and the use of their KO cell lines generated using CRISPR/Cas9 technology.PTEN, is a well-known tumour suppressor that plays a critical role in the host immune response. Recent research has highlighted the importance of this protein with regards to a) lineage stability and homeostasis in Treg cells (1) and b) negative regulation of the NK cell immune response (2, 3). In both cases PTEN acts by regulation of the PI3-kinase signalling pathway.We have identified several monoclonal antibodies to both PTEN and PI3K and confirmed their specificity by KO validation. We performed this analysis in either denatured (western blot) or native (immunocytochemistry) conditions. Our KO models represent true negative controls and provide confidence that these antibodies are binding to their intended target. This technology has allowed us to develop a growing range of over 650 KO-validated antibodies, including a number that are applicable to immunology/oncology research.
Role of innate immunity in neonates during a viral CNS infection
Authors: Ganesan Priya , Kuhn Alison M , Dunphy Abigail A , O’Donnell Lauren A 
Published: January 2023

Abstract
Viral infections in the central nervous system (CNS) are associated with devastating neurological consequences in newborns. This may be due, in part, to deficiencies in the immature innate and adaptive immune response. To study viral CNS infections in neonates, we use a transgenic mouse model of neuronally-restricted measles virus (MV) infection (NSE-CD46 mice), where the human isoform of CD46, a MV receptor, is expressed under the control of the neuron-specific enolase promoter. Adult CD46+ mice survive infection and control MV in an interferon gamma (IFNg)- and T cell-dependent manner. In contrast, neonatal CD46+ mice succumb to the infection despite T cell infiltration into the brain parenchyma. CD46+/RAG2-KO neonates, which lack T- and B-cells, experience less severe disease, prolonged survival, and lower viral load compared to CD46+ neonates. CD46+/RAG2-KO neonates also show greater infiltration of neutrophils and natural killer cells and higher expression of the pathogen recognition receptors melanoma differentiation-associated gene 5 (MDA5) and retinoic acid-inducible gene I (RIGI) during infection. Both CD46+ and CD46+/RAG2-KO neonates upregulate IFNg but CD46+/RAG2-KO neonates show higher expression of IFN-beta. CXCL10, a IFNg inducible chemokine, is also upregulated in CD46+ neonates (84-fold) and CD46+/RAG2-KO neonates (771-fold). Current studies aim to explore the role of IFN-beta in early viral control in the neonates and explore the contribution of CXCL10 in the infiltration of innate immune cells in the CNS.
Supported by 1R15NS087606-01A1.
Mechanisms of T follicular helper (Tfh) impairment in chronic HIV infection
Authors: Chakhtoura Marita , Cubas Rafael , Talla Aarthi , TARDIF Virginie , Haddad Elias 
Published: January 2023

Abstract
Human immunodeficiency virus (HIV) infects millions of people worldwide. In secondary lymphoid organs, site where the anti-HIV immune response is mounted, the virus has been shown to induce a predominant dysregulation and disturbance of the microvenvironment. Previous work in our laboratory has identified the lack of protective anti-HIV antibodies in patients to be due, at least in part, to an inadequate T follicular helper (Tfh) cell help to germinal center (GC) B cells in draining lymph nodes through impaired PD-1/PD-L1 interactions. Since the mechanism of this impairment is not yet fully elucidated, we have used our co-culture assay followed by a unique gene array analysis to screen for key molecular pathways that could be accountable for the difference between helper programs in Tfh cells of HIV positive and negative individuals. The major driver of gene profile clustering was observed within Tfh proliferation programs as well as infection status. Gene set enrichment analysis included repair mechanisms, cell cycle regulation, metabolism, signal transduction and co-stimulation in the top 10 enriched pathways. Careful examination of gene fold changes in proliferating vs non-proliferating Tfh cells between healthy and HIV individuals showed a decrease with HIV, in immune regulatory gene expression such as the transcription factor c-maf and its mediators, responsible for Tfh differentiation, development, survival and performance. Investigating this pathway would contribute to a better understanding of the HIV-mediated dysregulation.