bluuebunny/crossref_metadata_2025_split huggingface dataset:
Efficient Antibody Selection and Quantitation Using Microcapillary Cytometry (B209)
Authors: Mergia Alemayehu , Fishwild Dianne , Kenney John , Tyagarajan Kamala 
Published: January 2023

Abstract
Optimal selection of antibodies for therapy or research utilizes information from relative affinity measurements, specificity, functional studies and antibody isotype. All of the above are further dependant on accurate quantitative levels of antibodies utilized in the experiments so that a proper comparison of clones may be performed. Guava® RapidQuant™ IgG assays are novel bead-based assays that use microcapillary cytometry to provide quantitation of mouse or human IgG. In this study, mice were immunized with human CD80 and supernatant from each resulting hybridoma was subjected to multiple analyses on the Guava EasyCyte™ platform. The antigen specificity was obtained using a cell-based Guava Express™ assay and quantitative levels of IgG in supernatant were determined using the Guava RapidQuant Mouse IgG Assay. Antibody isotype and relative affinity were also ascertained. From these analyses, top ranking clones from 455 tested were identified based on specificity and productivity data. This study demonstrates the utility of microcapillary cytometry in combination with bead-based and cell-based assays to obtain multiple layers of information in the ranking and selection of the best clones.
A study on the screening and classification of herb medicines that have an influence on interleukin-4 (B216)
Authors: Min Ji-young , Lee Youngseop , Lee Heekyung , Jeon Sung Bae , Shin Min-Kyu , Kim Yangseok , Bae Hyunsu , Hong Moo-Chang 
Published: January 2023

Abstract
This study has examined the effects of 204 herbs, which are used in Traditional Korean Medicine, on the immune system. After the separation of splenocytes of BALB/c mice, two concentrations (1 §¶/§¢, 10 §¶/§¢) of herb medicine were given, and then the inducement of IL-4 was examined and measured using a sandwich ELISA. The amount of IL-4 each herb induced was analyzed, and the herb medicines were classified using two methods. One was the classification method of Traditional Korean Medicine, and the other was according to botanical phylogenetic systematics.
There were significant results for the analysis according to the botanical phylogenetic classification (Order). When there was a low concentration of the herb medicines, IL-4 was decreased by Ranunculales and Rosales, and increased by Malvales. When there was a high concentration of the herb medicines, IL-4 was decreased by Liliales, Ranunculales, Asterales, Fabales, Lamiales, Rosales, and Rutlaes, and increased by Malvales. The results showed the characteristics of individual herb medicines that have an affect on the inducement of IL-4 in splenocytes of a BALB/c mouse. In relation to the immune system, there was significant meaning to the functions of drugs for dispersing actions of wind and cold, drugs for dispersing actions of wind and heat, drugs for reducing intense internal heat, and drugs for eliminating heat and dampness, and also classifying the herb medicines according to botanical phylogenetics (Order).
This work was supported by the Brain Korea 21 project in 2007
The role of PKC-Theta in CD8 T cell activation (B28)
Authors: Valenzuela Javier O. , Wang Xingyu , Hussain Sofia , Beato Francisca , Liang Yaming , Yu Xue-Zhong , Beg Amer A. 
Published: January 2023

Abstract
Protein kinase C- Theta (PKC𝛉) is known to be a key mediator of NF-κB activation downstream of TCR signaling. However, recent in vivo studies with PKC𝛉−/− mice have generated conflicting results regarding its role in T cell activation. To elucidate the role of PKC𝛉 in the activation of CD8 T cells, we generated ovalbumin (ova)-specific PKC𝛉−/− OT1 mice. Stimulation of PKC𝛉−/− OT1 CD8 T cells with microspheres coupled to H2Kb-ova demonstrated a critical function for PKC𝛉 in TCR-induced NF-κB activation and proliferation in vitro. In contrast, ova-induced proliferation and CTL generation in vivo was virtually the same as WT cells. In addition, PKC𝛉−/− CD8 T cells stimulated in vitro with ova-pulsed DC proliferated vigorously and to a similar extent as WT cells. Remarkably, comparable levels of NF-κB were activated in WT and PKC𝛉−/− T cells after stimulation with DC. Proliferation of PKC𝛉−/− cells is severely reduced when DC deficient for B7-1/2 are used to stimulate the cells. Costimulation provided by CD27L also plays a role, as demonstrated by the use of a specific blocking antibody. In contrast to the response against antigen, GVHD responses are severely impaired in the absence of PKC𝛉. These results suggest that impaired TCR/antigen-induced NF-κB activation in PKC𝛉−/− CD8 T cells does not have a significant impact on T cell proliferation when other non-TCR signaling pathways are able to induce NF-κB and T cell activation.
Genetic variation in PTPN22 corresponds to altered signaling in B lymphocytes (B38)
Authors: Arechiga Adrian Fernely , Buckner Jane Hoyt 
Published: January 2023

Abstract
PTPN22 is a gene encoding the protein tyrosine phosphatase Lyp. A missense mutation in the gene, changing residue 1858 from a Cytosine to a Thymidine (1858C/T), is associated with the development of multiple autoimmune disorders. Although studies have demonstrated that Lyp has an inhibitory effect on T cell receptor mediated signaling pathways, the mechanism by which the 1858T variant contributes to the development of autoimmunity remains elusive. In this study, we examine the impact of the 1858T variant on B lymphocyte function. B lymphocytes from healthy subjects expressing the variant allele were compared to those of healthy subjects without the variant. When B cells were stimulated via the B-cell receptor, we observed impaired BCR signal transduction among subjects possessing the variant allele. This was characterized by diminished Calcium mobilization and altered tyrosine phosphorylation in key signaling proteins. These findings demonstrate that the PTPN22 1858T variant alters B cell as well as T cell responses and implicates B cells in the mechanism by which the PTPN221858T variant contributes to autoimmunity.
This work is supported by a JDRF grant to The Center for Translational Research at Benaroya Research Institute.
Cytokines Maintain Lymphocyte Homeostasis through the Activity of Cdc25A (B56)
Authors: Kittipatarin Christina , Khaled Annette 
Published: January 2023

Abstract
Homeostasis of the peripheral immune system is maintained through signals transduced by cytokines like interleukin-7 (IL-7). In the absence of IL-7 cells undergo apoptosis, while excess IL-7 leads to proliferation. The mechanisms by which IL-7 regulates these processes are poorly understood. We observed that expressing an anti-apoptotic protein (BCL-2) or inhibiting a pro-apoptotic protein (BAX) prevented cell death but did not support growth. Using cytokine-dependent lymphocytes, we found that the G1-to-S phase progression was not solely dependent on the synthesis of cyclin dependent kinases (cdks) and cyclins or the inactivation of inhibitors. Rather it was the removal of inhibitory phosphorylations on cdks by a phosphatase, Cdc25A, which primarily transduced cytokine-mediated growth signals. Cytokine deprivation lead to cell cycle arrest by inducing the activity of p38 MAPK, which destabilized Cdc25A. Inhibiting p38 or expressing a mutant Cdc25A, in which p38 target sites were mutated, promoted cell cycling independent of a cytokine signal. Cells expressing the Cdc25A mutant, in the absence of cytokines, actively synthesized DNA, expressed hyperphosphorylated Rb protein, maintained glucose uptake and did not undergo apoptosis, even in the presence of p27. Cdc25A is a novel transducer of cytokine-mediated growth signals and regulation of its activity is a means by which IL-7 controls proliferation.
Aberrant Regulation of Hematopoiesis by T cells in BAZF-Deficient Mice (B7)
Authors: Dent Alexander L , Broxmeyer Hal E. , Cooper Scott H , Sehra Sarita 
Published: January 2023

Abstract
The BAZF (BCL6b) protein is highly similar to the BCL6 transcriptional repressor. While BCL6 has been characterized extensively, relatively little is known about the normal function of BAZF. In order to understand the physiological role of BAZF, we created BAZF-deficient mice. Unlike BCL6-deficient mice, BAZF-deficient mice are healthy and normal in size. However BAZF-deficient mice have a hematopoietic progenitor phenotype that is almost identical to that of BCL6-deficient mice. Compared to wild-type mice, both BAZF-deficient and BCL6-deficient mice have greatly reduced numbers of cycling hematopoietic progenitor cells (HPC) in the bone marrow and greatly increased numbers of cycling HPC in the spleen. In contrast to HPC from wild-type mice, HPC from BAZF-deficient and BCL6-deficient mice are resistant to chemokine-induced myelo-suppression and do not show a synergistic growth response to GM-CSF plus Stem Cell Factor. Depletion of CD8 T cells in BAZF-deficient mice reverses several of the hematopoietic defects in these mice. Since both BAZF- and BCL6-deficient mice have defects in CD8 T cell differentiation, we hypothesize that both BCL6 and BAZF regulate HPC homeostasis by an indirect pathway involving CD8 T cells.
Distinct Processing Patterns Involving Flanking Residues of a GAD65 Peptide Can Be Accountable for the Difference in Diabetes Susceptibility Between NOD and NOR Mice (B77)
Authors: Dai Yang D. , Jensen Kent P. , Marrero Idania , Li Ningli , Quinn Anthony , Sercarz Eli 
Published: January 2023

Abstract
An immunogenic peptide in the GAD65 molecule, p524–543, activates two distinct populations of T cells, which apparently play opposite roles in the development of diabetes in NOD mice. In this study, we compared the fine specificity of these two T-cell repertoires using a nested set of truncated peptides that cover the p524–543 region. Surprisingly, we found that all clones apparently required the same minimum core sequence, around p530–539, for optimal activation. It was the flanking residues surrounding this core that played the crucial role in selecting T cells to activate. Flanking residues on the N-terminal side of the core led to the selection of regulatory cells and a largely Vbeta 12 T-cell repertoire, whereas flanking residues on the C-terminal side led to selection of effector cells and a largely Vbeta 4 T-cell repertoire. We then compared the repertoire in NOD with that in MHC-matched but diabetes-resistant NOR mice. NOD mice exhibited an enriched response to a C-terminal peptide, whereas NOR mice favored a response requiring N-terminal flanking residues. These data suggest that processing or trimming of the flanking residues of diabetes-associated determinants is a decisive event in the selection of islet-infiltrating T cells in NOD mice.
Development of Altered Peptide Ligands for Immunotherapy of Experimental Collagen-induced Arthritis (B96)
Authors: Marshall Dana Robin , Zhou Fang , He Xiaowen , Kayes Tim , Stuart John M 
Published: January 2023

Abstract
Collagen-induced arthritis is an autoimmune disease that results from the immunization of susceptible mouse strains with type II collagen. Amino acid residues 260–274 in the bovine collagen II alpha chain (bCIIá) represent the major MHC Class II epitope for the human DR1 haplotype. Amino acid residues 267Q and 270K are predicted TCR contact residues. Replacement of either of these amino acids with a glycine does not alter the ability of the peptides to bind to soluble DR1 but does substantially decrease their ability to induce proliferation of lymphocytes from bCIIá-immunized B10.M.DR1 mice while still allowing them to interfere with bCIIá-induced proliferation. The altered peptides elicit moderate decreases in disease incidence when co-immunized with bCIIá or injected intraperitoneally prior to the onset of clinical signs of disease.
The immune function of aging T cells (35.6)
Authors: Mirza Noweeda N , Hoelzinger Dominique B , Dominguez Ana L , Lustgarten Joseph 
Published: January 2023

Abstract
Age related changes compromise T cell mediated immunity. Previous findings suggest that the aged immune T cell repertoire can be exploited to mount effective anti-tumor responses to immunogenic tumors. However, antitumor immune responses against tumor self-antigens are significantly dysregulated in the old; hence, the salient differences in the biology and function of T cells between young and old, should be elucidated. These differences fall into three categories: (i) there are lower surface levels of T cell receptor (TCR) on old CD4 and CD8 T cells, (ii) the TCR complex is cycled differently in aged T cells and (iii) the TCR complex may be disassociated in older animals. Therefore these preliminary findings support the hypothesis that aging causes a decline in T cell function. Data analyses from a T cell activation time-course microarray study show that there are large differences in the gene expression profiles of old and young naive cells in CD4 (2022 genes) and CD8 (3666 genes) subsets. These differing genetic signatures point to potentially dysregulated proteins in the old immune system. Further investigation of selected gene products using appropriate mouse models, may provide valuable preclinical information with significant therapeutic potential which could have the opportunity of being translated into clinical trials.
This work was sponsored in part by a grant from AFAR.
Gads regulates the signaling threshold for CD8+ T cell activation (35.9)
Authors: Zhang Elizabeth Yan , Yankee Thomas  M. 
Published: January 2023

Abstract
Gads is an important regulator in T cell receptor (TCR)-mediated Ca2+ mobilization. We investigated the role of Gads in peripheral CD8+ T cell proliferation, using the MHC Class I-restricted ovalbumin-specific TCR transgenic mouse line (OT-I) on either the Gads+/+ or Gads-/- genetic background. We found that the Gads deficiency impaired TCR-mediated proliferation both in vitro and in vivo. This defect was most evident when low doses of cognate antigen (SIINFEKL) or altered peptide ligands (APLs) with lower affinity for the OT-I TCR were used as stimuli. However, the requirement for Gads in the proliferation of CD8+ T cells could be overcome by stimulating cells with high doses of SIINFEKL or APLs. The defect of Gads-/- CD8+ T cells in proliferation was caused by both impaired survival and by delayed entry into the cell cycle. We conclude that Gads regulates the signaling threshold through the TCR by promoting survival and cell cycle entry of CD8+ T cells. This research is supported by NIH grant P20 RR016443.
Modulation of autoimmune diabetes by antibodies specific for N-acetyl-D-glucosamine (167.10)
Authors: Dizon Brian , Greenspan Neil , Kearney John 
Published: January 2023

Abstract
Type I diabetes (T1D) is an autoimmune disease in which β cells are destroyed by the immune system. Childhood exposure to Group A Streptococci (GAS) is negatively correlated with T1D. Additionally, mice and humans produce N-acetyl-D-glucosamine (GlcNAc) specific antibodies in response to GAS infection. Since GlcNAc-specific antibodies exhibit autoreactivity to β cells, we hypothesized that GlcNAc-specific B cells and their secreted antibodies modulate the development of autoimmune diabetes. When neonatal NOD mice were immunized with GAS or control bacteria and monitored for hyperglycemia and glycosuria, NOD mice immunized with GAS were protected from spontaneous autoimmune diabetes. Moreover, passive transfer of sera from NOD mice immunized with GAS protected WT NOD mice from diabetes, demonstrating a protective role for GlcNAc-specific antibodies in vivo. Purified GlcNAc-specific IgM prevented activation and production of TNFα, IL-2, and IFNγ by diabetogenic T cells in vitro. Finally, immunization with GAS inhibited diabetogenic CD4 T cell activation in pancreatic lymph nodes in vivo. These findings indicate that B cells specific for O-GlcNAc, when activated by microorganism-associated antigens during neonatal development, protect against autoimmune diabetes by a mechanism dependent on secretory autoantibodies. These data identify a potential vaccine-based therapy for T1D in which protective autoantibodies are generated upon exposure to GlcNAc-expressing microorganisms.
HMGB1-PAMP complex promote the transformation of SFs to RASFs (171.19)
Authors: Qin Yanghua , Chen Yan , Wang Weiwei , Wang Zhiwei , Tang Gusheng , Dai Shengming , Shen Qian 
Published: January 2023

Abstract
The specific pathogenesis mechanism of rheumatoid arthritis (RA) remains unclear, current views believe the transformation of normal synovial fibroblasts (SFs) to RASFs is crucial for the pathogenesis of RA. Our previous studies showed that HMGB1-PAMP complex directly induced experimental arthritis in DBA/1 mice; pathology showed synovial thickening, and SFs excessive proliferation, suggesting that HMGB1-PAMP complex might induce the formation of RASFs. In current study, osteoarthritis synovial fibroblasts (OASFs) isolated from tissue obtained during reconstructive surgery and cocultured with HMGB1-PAMP complex in vitro for 5-8 generations to induce the transformation of normal SFs to RA-like SF (tOASFs). Then the pathogenicity of tOASFs was evaluated both in vitro and in vivo. Our results showed that tOASFs attached to, invaded into and degraded co-implanted cartilage in SCID mouse model. Histochemitry showed tOASFs excessive proliferation, expression MMPs. In vitro, cell cycle analysis showed more tOASFs pass through the G1/S checkpoint and move to S or G2 phage. Flow cytometry and confocal microscope showed significant reduced apoptosis and enhanced autophagy in tOASFs compared to OASFs, respectively. We further showed up-regulated expression of certain receptors (TLR2, TLR4, and RAGE) and adhesion molecules (ICAM-1 and VCAM-1) in tOASFs. In conclusion, we proved that HMGB1-PAMP complex promoted the formation of RASFs.
Regulation of dendritic cell function by Notch receptors and ligands (172.40)
Authors: Krawczyk Connie , Pearce Edward , Borjian Alborz , Young Stephanie , Mahbuba Raya 
Published: January 2023

Abstract
Cells of the innate immune system, such as Dendritic Cells (DCs), are the first responders and play a key role in creating pathogen-specific microenvironments through the generation of secreted mediators including cytokine, chemokines and lipids, and through modification of cell phenotype. T cells respond to the different activation phenotypes of DCs and in turn differentiate accordingly into distinct effector types, designated as lineages. Differentiation down different lineages is driven by “signal 3” or differentiation cues provided by the microenvironment. We have previously shown that signal 3 provided by DCs is a powerful regulator of T cell function not only at the time of activation, but during differentiation and recall responses. Notch ligand expression on DCs has been shown to be differentially regulated in response to various stimuli such as bacteria, worms, and allergens. Because Notch ligands show distinct and correlative expression patters in response to stimulation with various activators it is enticing to speculate they play a role in inducing or propagating differential immune responses. However their role remains controversial. Notch receptors are also expressed on DCs and their role in regulating DC function is just emerging. We will present evidence that Notch receptors participate in the activation of DCs. Furthermore, we propose that Notch ligands may indirectly influence Th differentiation through direct modification of DC function.
Identification, characterization and inhibition of protein interactions in immune responses (172.42)
Authors: Hindie Valerie , Ruggiero Barbara , Tafelmeyer Petra , Rain Jean-Christophe , Formstecher Etienne 
Published: January 2023

Abstract
Protein interaction mapping provides molecular basis to protein functions and signaling pathways. We have developed a high-throughput yeast two-hybrid technology that allows the exhaustive screening of highly complex, domain-based cDNA libraries relevant to the immune system (e.g. leukocytes and activated mononuclear cells, thymocytes CD4+/CD8+ and T-cells). These libraries are screened to saturation thanks to an optimized mating procedure, allowing to test on average 100 million interactions per screen. This corresponds to a 10-fold coverage of the libraries, and ensures exhaustive and reproducible results. Moreover, multiple independent fragments are isolated for each interactant, enabling the immediate delineation of a minimal interacting domain and the computation of a confidence score. A database containing over five thousand cDNA screens realized with this technique permits a rigorous calculation of relative prey connectivity and understanding of true/false positives and sticky proteins. We have developed additional methods to probe and characterize protein interactions in vitro with a medium to high throughput, using Surface Plasmon Resonance and a miniaturized time-resolved FRET assay which has been used for drug discovery projects targeting protein-protein interactions. Here, we will present results that illustrate the strength and benefits of our protein interaction screening and validation technologies to decipher the molecular components of immune responses.
Hybrid Vγ-Cβ TCR chains support T cell development and an immunocompetent T cell repertoire (LYM7P.713)
Authors: Bowen Steven , Sharrow Susan , Hodes Richard 
Published: January 2023

Abstract
Developing T cells undergo VDJ recombination at four T cell receptor (TCR) loci (γ, δ, β and α) to generate a highly diverse γδ or αβ TCR. During the CD4-CD8- (DN) stage of development, simultaneous recombination at the TCRγ, TCRδ and TCRβ loci can result in unusual interchromosomal trans rearrangements between these loci, particularly between TCRβ and TCRγ, creating hybrid TCR genes encoded by two different chromosomes. We have found that in-frame trans rearrangements between the Vγ2 gene and a variety of Dβ and Jβ genes are transcribed, spliced and expressed on the surface of a small population of peripheral T cells. Vγ2-Cβ hybrid TCR chains are capable of mediating β-selection and require the expression of TCRα for subsequent developmental transitions. However, the relatively low frequency of these cells has made analysis of their function difficult. Here we report that transgenic expression of a Vγ2-Jβ2.4 trans rearrangement is capable of driving T cell development in the thymus through the CD4/CD8 double positive (DP) and single positive (SP) stages as well as maintaining a robust population of CD4+ and CD8+ T cells in the periphery. These mice are also capable of mounting a T-dependent IgG1 response to peptide immunization. These data suggest that non-conventional Vγ-Cβ TCR chains are sufficient to support the development of an immunocompetent T cell repertoire.
Tumoral expression of IL-33 promotes Anti-tumor Immune Responses (TUM2P.912)
Authors: Wang Xuefeng , Gao Xin , Zhao Xin , Yang Qianting , Qi Yuan , Zang Shayang , Zhang Xueguang , Lu Binfeng 
Published: January 2023

Abstract
One major approach of cancer immune therapy is to convert the tumor immunosuppressive microenvironment to one that favors antitumor immune responses. We have recently demonstrated that Interleukin 33 (IL-33) promotes effector functions of CD8 T cells, suggesting a potential function in antitumor immunity. Here we showed that overexpression of IL-33 in two tumor cell lines, 4T1 and B16, potently inhibited tumor growth in vivo. CD45+, CD8+ T cells, NK cells, IFN-γ+ CD8+ T cells and IFN-γ+ NK cells were greatly increased in the IL-33-expressing tumors when compared with those in the control tumors. We further demonstrated that the antitumor effect of IL-33 was dependent on NK cells and CD8 T cells. In contrast, MDSC were greatly decreased in the IL-33-expressing tumors when compared to those in the control tumors. Our results indicate that the intratumoral delivery of IL-33 can be a new strategy of tumor immunotherapy by promoting an immunogenic microenvironment.