bluuebunny/crossref_metadata_2025_split huggingface dataset:
EPOR expression by peritoneal macrophages and B1b cells (TUM6P.1010)
Authors: Riggs James , Goldman Naomi , Orlowski Michelle 
Published: January 2023

Abstract
Erythropoietin (EPO) is used in the treatment of chemotherapy-related anemia. A higher mortality rate among cancer patients treated with EPO has been documented. Research in this area has focused upon EPOR expression by cancer cells, either cell lines or ex vivo tumor tissue. We find that addition of exogenous EPO suppresses the activation of peritoneal cavity (PerC) T cells. Via FACs analysis of cultured C57BL/6J (wildtype,WT) PerC cells, we detected the EPO receptor (EPOR) on F4/80+, CD11b+ macrophages (Mfs). The specificity of the anti-EPOR pAb was validated by inhibition of staining with soluble EPOR. The percentage of Mfs expressing the EPOR increased with culture duration. EPOR+ Mfs also increased expression of the immunoinhibitory molecule PDL1/B7H1. Interestingly, EPOR expression also appeared on cultured B1b (IgMhi, CD11blo) cells. Although less evident in WT mice assessed ex vivo, EPOR+ PerC Mfs were found in IL10KO mice. Considerably more EPOR+ B1b cells were observed in the inflammatory ascites of mice with the ovarian cancer that develops following injection of the ID8 mouse epithelial carcinoma cell line. We are currently investigating how EPO contributes to T cell suppression in vitro and impacts Mf and B1b cell function and phenotype. Our data support a link between inflammation and EPOR expression by PerC Mfs and B1b cells.
Increasing Th1 phenotype in Th17 cells improves anti-tumor T cells function (VAC7P.1032)
Authors: Chatterjee Shilpak , Kesarwani Pravin , Soloshchenko Myroslawa , Fu Jianing , Paulos Chrystal , Yu Xue-Zhong , Mehrotra Shikhar 
Published: January 2023

Abstract
Immunotherapy of cancer is a promising approach for successful eradication of large and established tumor. While cytolytic CD8+ T cells have been extensively investigated and used for adoptive cell therapy (ACT), accumulating evidence indicates that CD4+ T helper (Th) cells including Th1 and Th17 subsets are also effective in tumor immunotherapy. Although Th1 cells possess key anti-tumor effector functions as IFNγhi, CD107ahi, T-bethi, Granzyme Bhi, but their poor persistence due to lack of stem-cell like characteristic effect long-term tumor control and reduce their potential in ACT. However, Th17 cells programmed ex vivo in TGFβ exhibit increased stem-cell like features and reduced effector function as compared to Th1 cells, which corelates to their increased persistence but reduced long-term tumor control. Herein we identified the ex vivo programming conditions that combine potent Th1 effector conditions with key stemness feature of Th17 to generate hybrid Th17/Th1 cells that exhibit improved control of murine melanoma (B16-F10) and human melanoma (624-MEL), as compared to Th1 or Th17 cells alone. The hybrid Th1/Th17 cells were less susceptible to activation induced cell death, exhibited increased persistence along with potent effector function, and have potential in future translational studies.
Improving anti-tumor immunity by engineering T cell receptors to prevent mispairing (VAC7P.1033)
Authors: Bethune Michael , Gee Marvin , Lee Mark , Kuhns Michael , Baltimore David 
Published: January 2023

Abstract
T cells transduced with the genes encoding the α and β chains of a tumor-specific T cell receptor (TCR) can mediate anti-tumor immunity in patients. Such TCR-transduced T cells co-express both the therapeutic and an endogenous TCR. As each receptor is an αβ heterodimer, the endogenous α chain can mispair with the therapeutic β chain and vice versa. Such mispairing - which is observed in transduced human T cells - reduces export of the therapeutic TCR and produces potentially self-reactive TCRs. The latter can precipitate graft-vs-host disease, a lethal complication reported in mice following TCR gene transfer. We have developed a general strategy for preventing TCR mispairing. Select constant domains are swapped between the α and β chains of a tumor-specific T cell receptor, creating a domain-swapped (ds)TCR. When correctly paired, these dsTCR chains retain all domains necessary to recruit CD3 proteins, to express on the T cell surface, and to mediate functional T cell responses upon engaging a target antigen. By contrast, mispaired TCRs containing one dsTCR chain and one wild-type TCR chain lack key domains necessary for CD3 recruitment, export, and signaling, and thus are unable to mediate deleterious autoimmunity. We validate the potential of this approach in cell-based assays, as well as in a mouse model of TCR gene transfer-induced graft-vs-host disease. Thus, this work will enable the design of safer, more effective TCR gene therapy candidates for cancer immunotherapy.
Preservation of Raboral V-RG® viability by vitrification (VAC9P.1100)
Authors: Tran Yen-Nhi , Kemp Jeffrey , Nash Paul 
Published: January 2023

Abstract
Raboral V-RG® is a recombinant vaccinia virus used to orally immunize wildlife such as raccoons and coyotes against rabies. Since it is a live virus vaccine that expresses the rabies virus glycoprotein, stabilizing the virus and allowing for more methods of incorporation into bait can both be advantageous. This may also be true for other live viruses or cells. Two methods of preserving the virus were tested: vitrification and centrifugal evaporation. Vitrification in a sugar or polymer matrix provides protection for the virus through the transition of the matrix into a glass state and eliminates the need for containment of liquid virus suspension. Centrifugal evaporation in a sugar matrix provides protection for the virus through drying out the sample and is an alternative method of eliminating the need for containment of liquid virus suspension. Vitrification in several different matrices provided the virus with a significant level of protection. Using vaccinia virus as a surrogate for Raboral V-RG, we also examined whether simple drying might also be effective.
Novel approach to simultaneously monitor local and systemic in vivo effects of varying doses of intradermally injected LPS (VET1P.1123)
Authors: Byrne Kristen , Erf Gisela , Falcon Daniel 
Published: January 2023

Abstract
We recently established the dermis (pulp) of growing feathers (GF) as a minimally invasive test-site to monitor in vivo tissue effects of test substances. The objective of this study was to use the GF test-system to simultaneously monitor local and systemic effects of varying doses of endotoxin (LPS) injected into GF on leukocyte profiles and activities. Growing feathers of 16 chickens were injected with either 10 µL of endotoxin-free PBS or 10, 100, or 1000 ng of LPS per feather (18 GF/chicken). Blood and three GF were collected at each time point: 0 (before injection), 2, 4, 8, and 24 h post-injection. The concentration of leukocytes in the blood and proportions of leukocytes within the feather pulp were determined for each time point. Both LPS 1000 and 100 exhibited similar patterns of leukocyte recruitment in the blood and GF. Specifically, GF injection of LPS resulted in an initial drop (2 h) in blood leukocytes followed by increasing levels at 4 and 8 h and returning to near control levels by 24 h. The drop in blood leukocytes at 2 h was not observed with the LPS 10 treatment. At the site of injection (GF), increased levels of heterophils were already observed at 2 h for LPS 10 and 100, whereas this increase was delayed for LPS 1000. At 8 h post LPS injection, leukocyte levels were greatly elevated for all LPS treatments. Overall, based on leukocyte recruitment patterns in blood and tissue, the lowest dose of LPS may serve as a model for the local inflammatory response.
Understanding manatee immunogenetics and the implications for conservation (VET1P.1121)
Authors: Breaux Breanna , Heard-Ganir Ashley , Sena Leo , Hunter Margaret , Bonde Robert , Criscitiello Michael 
Published: January 2023

Abstract
The completion of the genome draft of the Florida manatee (Trichechus manatus latirostris) in 2012 made immunogenetic exploration of the Sirenia order more feasible. The endangered manatees are sentinel species for coastal ecosystems and their evolutionary origins are complex. It is believed that they evolved from a wading terrestrial mammal that became fully aquatic, which suggests distinctive adaptations for transitioning to a new environment. Discovering the genetic details of their immune system will help understand manatee evolution and allow species-specific diagnostic tools to be developed. Due to bottleneck signatures at other loci, the genetic diversity of the major histocompatibility complex (MHC) is expected to be low. This could pose problems for conservation efforts because low diversity can increase the chance of extirpation. The primary focus of this project is to determine the immune robustness of manatees by looking at their antibody repertoire and MHC polymorphism. Two species will be compared: the West Indian manatee (T. manatus) and the Amazonian manatee (T. inunguis). Using next generation sequencing, we will robustly sample the IgH and IgL rearrangements contributing to the antibody repertoire, and identify dominant MHC alleles and estimate their frequencies. The allelic distribution between the species could reveal clues to their evolutionary relationship and aid conservation efforts to help this species become more stable in their environment.
IL-15 drives CD8 T cell cycling and differentiation in chronic HIV-1 infection (VIR6P.1162)
Authors: younes souheil , shive carey , Reynaldi Arnold , Freeman Michael , Panigrahi Soumya , Estes Jacob , Anderson Jodi , Schacker Timothy , Davenport Miles , McCune Joseph , Hunt Peter , Lee Sulggi , Serrano-Villar Sergio , Canaday David , Cameron Cheryl , Sekaly Rafick-Pierre , Rodriguez Benigno , Sieg Scott , Lederman Michael 
Published: January 2023

Abstract
HIV-1 infection is characterized by sustained CD8 T cell expansion that is linked to morbid outcomes even after HIV replication is controlled with antiretroviral therapy (ART). The drivers of this persistent CD8 expansion in HIV-1 infection are not well understood. Here we show that cycling frequency is increased in memory CD8 T cells in untreated HIV-1 infected patients and cycling CD8 T cells have a broad T cell receptor repertoire tightly linked to the repertoire of non-cycling cells suggesting that they are largely driven into cycle by bystander mechanisms. In HIV-1 infection, cycling and granzyme B expression were increased among CD8 T cells binding viral peptide/HLA tetramers and this enriched cycling/cytolytic phenotype among virus-reactive CD8 T cells could be generated in vitro by exposure to interleukin (IL)-15. Lymph nodes of HIV-1 infected untreated patients were enriched for interleukin-15 expression that correlated with circulating CD8 T cell counts and normalized with drug-induced control of HIV-1 replication. The determinants of persistent CD8 T cell expansion and the mechanisms whereby this expansion is linked to increased morbidity merit further examination.
Sensor function for butyrophilin 3A1 in prenyl pyrophosphate stimulation of human Vγ2Vδ2 T cells (APP5P.100)
Authors: Morita Craig , Wang Hong 
Published: January 2023

Abstract
Vγ2Vδ2 T cells play important roles in human immunity to pathogens and in cancer immunotherapy by responding to isoprenoid metabolites such as (E)-4-hydroxy-3-methyl-but-2-enyl pyrophosphate and isopentenyl pyrophosphate. The immunoglobulin superfamily protein, butyrophilin 3A1 (BTN3A1), has been shown to be required for prenyl pyrophosphate stimulation. We proposed that the intracellular B30.2 domain of BTN3A1 binds prenyl pyrophosphates resulting in a change in the extracellular BTN3A1 dimer that is detected by Vγ2Vδ2 TCRs. Such B30.2 binding has been recently demonstrated. However, others have reported that the extracellular BTN3A1 IgV domain binds prenyl pyrophosphates leading to the proposal that the Vγ2Vδ2 TCR recognizes the complex. To distinguish between these mechanisms, we mutagenized residues in the binding sites to alanine and tested the mutant BTN3A1 proteins for their ability to mediate prenyl pyrophosphate stimulation of Vγ2Vδ2 T cells to proliferate and secrete TNF-α. Mutagenesis of residues in the IgV site had no effect whereas mutagenesis of residues within the basic pocket and surrounding V regions of the B30.2 domain abrogated prenyl pyrophosphate-induced proliferation and reduced or abolished TNF-α production. Some mutations further from the B30.2 site also diminished stimulation suggesting that the B30.2 domain may interact with a second protein. These findings support intracellular sensing of prenyl pyrophosphates rather than extracellular presentation.
Direct interaction of recombinant TAPBPR with MHC-I molecules: stabilization of peptide-free MHC-I promotes high affinity peptide loading (APP5P.102)
Authors: Morozov Giora , Zhao Huaying , Mage Michael , Boyd Lisa , Venna Ramesh , Norcross Michael , McMurtrey Curtis , Hildebrand William , Schuck Peter , Jiang Jiansheng , Natarajan Kannan , Margulies David 
Published: January 2023

Abstract
The loading of MHC-I molecules with peptides for cell surface display is a crucial step in self-tolerance and activation of CD8 T cells. We studied TAPBPR (TAP binding protein-related) protein, a tapasin homolog, which is widely expressed and IFN-γ inducible, but is not part of the classical MHC-I peptide-loading complex. We produced recombinant soluble TAPBPR and evaluated its interactions with several recombinant MHC-I molecules in vitro, by gel-shift, size exclusion chromatography, ultracentrifugation, and surface plasmon resonance. We show that TAPBPR binds MHC-I after photolysis of a bound peptide, and that the TAPBPR/MHC-I complex is dissociated by exposure to peptides that bind the MHC-I molecule, indicating a role of TAPBPR in stabilizing a peptide-receptive form of the MHC-I/β2m complex. Peptide-dependent release of MHC-I from TAPBPR is directly proportional to the peptide’s affinity for MHC-I. Peptide binding experiments indicate a role for TAPBPR in selection of high affinity peptides. Mutagenesis of TAPBPR and MHC-I confirm the importance of amino acid residues conserved with the putative tapasin/MHC-I binding site and reveal additional residues important for the TAPBPR/MHC-I interaction. Molecular docking simulations suggest a detailed mechanism for the interaction of TAPBPR with peptide free MHC-I. These studies are consistent with the view that TAPBPR functions as a chaperone that stabilizes peptide-free MHC-I to permit binding of high affinity peptides.
ERAP1-ERAP2 dimers trim MHC I-bound precursor peptides: a molecular understanding of peptide editing (APP5P.103)
Authors: Bouvier Marlene , Chen Hanna , Li Lenong , Weimershaus Mirjana , Evnouchidou Irini , Van Endert Peter 
Published: January 2023

Abstract
The human endoplasmic reticulum aminopeptidase 1 (ERAP1) and ERAP2 are critically important in the final processing of MHC class I antigens. To date, the molecular context of peptide trimming by ERAPs, and how ERAPs shape antigen repertoires, remain open questions. Using a cell-free system composed of ERAP1 and ERAP2 heterodimers (ERAP1/2), MHC class I molecules, and N-terminally extended model and natural peptides, we characterized the function of ERAPs. We provide conclusive evidence that ERAP1/2 trims MHC I-bound precursor peptides to the final lengths, albeit more slowly than the corresponding free precursors. We show that trimming of MHC I-bound precursors by ERAP1/2 increases the conformational stability of MHC I/peptide complexes. We will discuss a molecular mechanistic model of peptide editing by ERAP1/2 that unifies and advances our understanding of these important enzymes.
Identification of self-specific CD4+ T cell subsets resistant to PD-1 blockade (BA11P.130)
Authors: Martinov Tijana , Pauken Kristen , Nelson Christine , Spanier Justin , Heffernan James , Sahli Nathanael , Osum Kevin , Jenkins Marc , Vezys Vaiva , Fife Brian 
Published: January 2023

Abstract
The inhibitory receptor programmed death (PD)-1 and its ligand PD-L1 regulate T cell function to limit autoimmunity. PD-1 is highly expressed on exhausted T cells, limiting their antiviral or antitumor activity. Even though PD-1 pathway blockade has gained momentum in cancer treatment, it is unclear how PD-1/PD-L1 blockade impacts different T cell subsets. We examined the effects of PD-1 blockade on self-reactive CD4+ T cells in mouse models of varying autoimmune susceptibilities. We used insulin-peptide/MHC Class II tetramers to track endogenous insulin-specific CD4+ T cells in diabetes-prone (NOD), and diabetes-resistant (NOR and B6.g7) mice. PD-1 was expressed at lower levels on insulin-specific cells in diabetes-resistant mice compared to cells from NOD mice. Moreover, PD-1 pathway blockade accelerated autoimmunity in all NOD mice and 30% of NOR mice, but B6.g7 mice remained protected from disease. By transferring congenically marked islet-specific CD4+ T cells into pre-diabetic NOD mice, we asked whether anti-PD-L1 reinvigorates anergic cells. Surprisingly, anergic cells remained functionally blunted compared to effector cells after anti-PD-L1, suggesting that established anergic cells were not dependent on PD-1 signaling to maintain their unresponsive state. This work highlights how the differentiation status of a T cell may predetermine its susceptibility to PD-1 blockade, and influence autoimmunity or antitumor responses that ensue.
The dynamics of T cell restimulation in the pancreatic islets indicate a period of T cell tolerance during autoimmune diabetes (BA11P.125)
Authors: Friedman Rachel , Lindsay Robin , Gebert Matthew , Bradley Brenda , Krummel Matthew , Haskins Kathryn , Jacobelli Jordan 
Published: January 2023

Abstract
In type 1 diabetes, the pancreatic islets are an important site for therapeutic intervention since immune infiltration and attack of the islets is well established at diagnosis. To understand how T cell stimulation and function evolve at different stages of islet infiltration, we initially analyzed the dynamics of T cell movement and T cell interactions with CD11c+ APCs within individual islets. Using 2-photon imaging, we defined a correlation between increased islet infiltration and increased T cell motility. Early T cell arrest was antigen-dependent and due, at least in part, to antigen recognition through stable interactions with CD11c+ APCs in the islets, which were lost with increased islet infiltration. Stable T cell-APC interactions resulted in TCR clustering, supported IFN-γ production, and CD8 T cell killing of islet APCs. As islet infiltration increased, T cell motility became antigen-independent, with a loss of sustained interactions with APCs resulting in reduced IFN-γ production and APC killing. These studies suggest that the autoimmune T cell response in the islets is temporarily dampened during the course of islet infiltration and disease progression. We suggest that an infiltrating population of CD11c+ APCs, which we can distinguish from islet-resident APCs via expression levels of CX3CR1, plays a role in establishing a period of T cell tolerance within the pancreatic islets following initial T cell infiltration.
Influence of cytokines and TLR agonists on CD49a, CD49b and CD103 expression by T cells (CAM4P.152)
Authors: Melssen Marit , Hutchison Ciara , Bullock Timothy , Slingluff Craig 
Published: January 2023

Abstract
Integrins CD49a (alpha1), CD49b (alpha2) and CD103 (alphaE) are highly expressed on tumor infiltrating lymphocytes (TILs) compared to circulating lymphocytes. Interactions of the integrins with known ligands support retention of lymphocytes in peripheral tissues; thus, they may be considered retention integrins (RI). T cells in peripheral tissues expressing RI have a cytotoxic phenotype and RI+ TIL are associated with improved patient survival. Despite their importance in retention of tumor reactive TILs, little is known about pathways inducing RI expression, except that TGFβ induces CD103. We hypothesized that other cytokines, TLR agonists, and T cell receptor (TCR) stimulation can induce RI expression by circulating T cells. We stimulated PBMC with cytokines or TLR agonists after TCR stimulation, and evaluated RI expression by flow cytometry after 2-7 days. Both CD103 and CD49a are induced by TGFβ, most strikingly on activated T cells. IL2 and IL10 also influence CD49a expression. CD49b expression is increased by IL2, IL4 and IL10, although this depends on T cell subset and activation state. TLR2, TLR6 and TLR7 agonists also appear to induce CD49a and CD49b. In conclusion, our results support our hypothesis by showing that RI expression is tightly regulated by cytokine and TCR stimulation. Furthermore these regulating factors have different effects among the RI, suggesting a different role for each.
A new technology for in vitro chemotaxis assays (CAM4P.159)
Authors: APPLEDORN DANIEL , O'Clair Lindy , Tilhonenko Maria , Roddy Meagan , Schroeder Kirk 
Published: January 2023

Abstract
Chemotactic-driven cell migration is an important biological process in many areas of life science research including immunology, cancer and developmental biology. The dominant high-throughput (96-well format) solution for measuring cellular chemotaxis in vitro is the Boyden-chamber. However, Boyden chambers have some inherent liabilities including: a) the inability to directly visualize the cell migration process, b) a requirement of tens of thousands of cells per well, c) the need to label cells for quantitation, d) a relative insensitivity to surface integrin signaling, and e) high random migration artifacts. Microfluidic solutions have sought to eliminate some of these deficiencies, specifically by providing the ability to visualize the cell migration process, but at the expense of relatively low throughput and cumbersome quantitation methods. We have developed a new technology which eliminates most of the deficiencies of existing Boyden chambers and most importantly, provides a direct visualization of the migration process as it occurs. Integrated metrics precisely quantify the chemotactic response using significantly fewer cells (500-5000 cells per well) in a 96-well format. This assay method does not require labeling for quantitation, has a kinetic read-out, and is sensitive to surface-integrin signaling. Pharmacological data examples from T cells, neutrophils, cancer cells, and endothelial cells will be shown with observations enabled by the real-time visualization.
Sphingosine 1-phosphate receptor-3 contributes to host defense against polymicrobial sepsis via modulating macrophage chemotaxis (CAM4P.161)
Authors: Shu Qiang , Hou JinChao , Chen QiXing , Fang XiangMing 
Published: January 2023

Abstract
Objective Sphingosine 1-phosphate receptor-3 (S1P3) is an important G protein-coupled receptor regulating host immune response. However, the role of S1P3 in polymicrobial sepsis remains unclear. Method Sepsis was induced in wild-type (WT) and S1PR3-deficient mice via cecal ligation and puncture (CLP). The mortality of mice, as well as organ injury and bacterial clearance during sepsis were assessed. The role of S1PR3 in sepsis was further confirmed by administration of an agonist peptide representing part of the intracellular domain of S1PR3 (GPS-725.017) in septic WT mice. Results S1PR3 deficient mice showed reduced survival rate, increased lung and liver damage, and more bacterial burden during sepsis. There was also significantly less macrophages chemotactic into the peritoneal cavity in the S1PR3 deficient mice. GPS-725.017 could active S1PR3, including internalization of the membrane protein S1PR3 into cytoplasm and activation of MAPKs signaling. In vivo, GPS-725.017 treatment increased recruitment of phagocyte into the peritoneal cavity, enhanced bacterial clearance and improved survival during sepsis. Conclusion The present findings uncover S1PR3 might be a critical modulator of macrophage chemotaxis in the pathogenesis of sepsis. The detailed mechanisms need to be further addressed.