bluuebunny/crossref_metadata_2025_split huggingface dataset:
Macrophages produce a soluble mediator which enables neutrophils to kill the parasitic nematode Strongyloides stercoralis (51.4)
Authors: O’Connell Amy E , Abraham David 
Published: January 2023

Abstract
Neutrophil recruitment to the parasite microenvironment via CXCR2 is required for killing of Strongyloides stercoralis infective larvae during the primary and secondary immune responses. Neutrophils isolated from other cell types within diffusion chambers are capable of killing L3 in vivo, however, isolated neutrophils are unable to kill larvae in vitro. Macrophages have been shown to be required for in vivo killing of other nematode species. We hypothesized that neutrophils require soluble mediators that can be supplied by macrophages in vivo, explaining their inability to kill in isolation in vitro. Combining neutrophils with macrophages at a 4:1 ratio in vitro led to a robust larvicidal response, while neither cell type was able to facilitate larval killing independently. The killing response occurred whether naïve or immune mouse serum was used, which suggests that neutrophil-macrophage cooperation is a component of the primary immune response. Furthermore, the killing process was complement-dependent, as killing was ablated when serum from C3−/− mice was used. Combining cell free supernatant from cultured macrophages with neutrophils also led to larval killing, indicating that the cooperation between neutrophils and macrophages is dependent on a soluble factor from macrophages inducing larval killing by neutrophils.
Defensin α-1 expression is up-regulated in human cells in response to early Trypanosoma cruzi infection as an innate immune mechanism to decrease cellular infection via membrane pore formation leading to apoptosis (51.6)
Authors: Madison Marisa Nia , Kleshchenko Yuliya , Nde Pius , Simmons Kaneatra , Lima Maria F , Villalta Fernando 
Published: January 2023

Abstract
Defensins play a fundamental role in the initiation of immune responses to selected human pathogens. Here, we show that early cellular infection by T. cruzi up-regulates the expression and secretion of defensin α-1, which displayed a trypanocidal role mediated by pore-formation resulting in apoptosis of the parasite. Analysis of the whole human epithelial cell transcriptome upon short infection of cells by T. cruzi indicates that the parasite up-regulates the levels of transcripts of defensin α-1. Defensin α-1, at concentrations not toxic for host cells, significantly reduced the viability of trypomastigote forms of T. cruzi. Defensin α-1 integrates into the trypanosome membrane to cause pore formation, cellular and mitochondrial membrane depolarization, membrane disorganization and blebbing, DNA fragmentation, and cytoplasmic vacuolization leading to damage and death. Pre-incubation of trypomastigotes with exogenous defensin α-1 followed by exposure to human epithelial cells significantly reduced trypanosome infection. Membrane depolarization of trypanosomes abolished the trypanocidal activity of defensin α-1, indicating that the mechanism of defensin α-1 mediated trypanosome killing is membrane-voltage dependent. We conclude that defensin α-1 gene expression and peptide secretion is an effective host innate immune response to control T. cruzi infection.
(Supported by NIH grants)
Heat shock protein gp96 is a critical chaperone for Toll-like receptor 9 (52.2)
Authors: Liu Bei , Yang Yi , Li Zihai 
Published: January 2023

Abstract
Toll-like receptors (TLR) are important pattern-recognition receptors and play significant roles in both innate and adaptive immunity. As a ubiquitous ER luminal protein, gp96 has been previously implicated in the folding and surface expression of TLR1, TLR2 and TLR4. However, whether gp96 chaperones other TLRs, especially intracellular TLR9, is unknown. We herein addressed this question using a genetic approach. We found that gp96 mutant cells, via insertional mutagenesis or homologous recombination, were unresponsive to TLR9 ligand, which was completely rescued by restoration of gp96. Furthermore, we demonstrated the direct interaction between gp96 and TLR9; such an interaction is dependent on N-linked glycosylation and normal ER calcium store. These findings illustrate that gp96 is an essential chaperone for the folding of TLR9, and provide a new potential target to regulate TLR function in the treatment of a variety of diseases.
Complement C5 regulates insulin-like growth factors binding proteins in chronic Experimental Autoimmune Encephalomyelitis (EAE) (53.8)
Authors: Cudrici Cornelia Diana , Zafranskaia Ekaterina , Weerth Susanna , Chen Hegang , Gherman Adrian , Ito Takahiro , Rus Violeta , Raine Cedric , Rus Horea 
Published: January 2023

Abstract
Complement activation plays a central role in autoimmune demyelination. We have analyzed the role of complement C5 in EAE using C5-deficient (C5-d) and C5-sufficient (C5-s) mice and found that the absence of C5 resulted in fiber loss and extensive scarring. In this study, we investigated thetranscriptional profile induced by C5 in chronic EAE using oligonucleotide array.During chronic EAE (day 90, post-immunization), we found that 462 genes were differentially regulated in C5-s when compared with C5-d mice. The majority of these genes were related to cell growth and signal transduction; agroup of genes belonging to the insulin-like growth factors binding proteins family was also significantly regulated by C5. This category include insulin like growth factors (IGFBP) 3, 4 and 6, connective tissue growth factor (CTGF), WNT1 inducible signaling pathway protein 2, cysteine rich protein 61 and endothelial cell specific molecule-1. Using Real-time PCR we confirmed the down-regulation of these genes in C5-d mice in chronic EAE. When IGFBP3 and CTGF levels in chronic EAE were compared with those in control mice (day 0, post-immunization), significantly lower levels of mRNA expression were found only in C5-d mice. The down-regulation of genes belonging to the insulin-like growth factor binding family suggests that they might be responsible for the axonal loss and gliosis seen in C5-d during chronic EAE.
Cellular interactions during GALT development (81.10)
Authors: Yeramilli Venkata , Knight Katherine L. 
Published: January 2023

Abstract
In rabbits, B cells develop in the bone marrow and migrate to the GALT where they proliferate, form organized follicles, and develop the primary antibody repertoire by Ig gene diversification. Our goal is to determine the mechanism by which these processes are induced in GALT. We hypothesize that B cells in GALT are polyclonally stimulated in an antigen and T cell independent manner. To test this idea, we injected newborn rabbits with adenoviruses (Ad) expressing soluble receptors, CTLA4Ig and CD40Ig, to inhibit activation of T cells and T-B cell interactions, respectively. We found that GALT developed normally in rabbits injected with AdCTLA4Ig, as evidenced by the presence of follicles with proliferating B cells. In contrast, we found no follicles with proliferating B cells in rabbits injected with AdCD40Ig. These results indicate that activated T cells are not required for GALT development, but that CD40:CD40L interactions are required. Using a similar approach, we found that B cells require signaling through the BAFF receptor and complement receptor 2 (CR2) for proliferation in GALT. The source of CD40L, BAFF and CR2 ligand, molecules required for B cell activation in GALT, are under investigation.
Supported by NIH AI50260
Functional deficiency in IL-7 caused by an ENU-induced point mutation (82.11)
Authors: Wang Hongsheng , Feng Jianxun , Morse Herbert C. 
Published: January 2023

Abstract
N-ethyl-N-nitrosourea (ENU)-induced mutagenesis provides a powerful approach to identifying genes involved in immune regulation and diseases. Here we describe a new mutant strain, HLB368, with hereditary leukopenia. At necropsy, the mutant mice had very small thymuses and spleens. All but the inguinal nodes were absent and there were no Peyer’s patches. By flow cytometry, the ratios of T cell subsets were normal, but B cell development was blocked at the pre-pro-B cell stage. The development of B1 and marginal zone B cells was relatively normal. The mutation was mapped to chromosome 3 between D3Mit221 and D3Mit224, a region that contains the Il7 gene. cDNA and genomic DNA sequences of Il7 revealed a T-to-C missense transition resulting in a change of Leu to Pro within the leader peptide that would be predicted to inhibit secretion. In keeping with this concept, we found that in vitro treatment of B cell progenitors from mutant mice with IL-7 induced them to differentiate into pre-BII cells. Phenotypic comparisons of HLB368 with genetically targeted Il7 null mice showed many similarities along with a few differences, indicating that this ENU-induced mutant carries a novel allele. This new strain thus provides a new model for studying the functions of IL-7 on a pure C57BL/6 background.
Exposure to bead immobilized antigen dramatically increases the survival and expansion of anti-CD40 stimulated HEL transgenic B cells. (83.9)
Authors: Marran-Nichol Kathleen 
Published: January 2023

Abstract
Since B cells primarily respond to conformational determinants produced by the three-dimensional assembly of proteins, the assumption has remained that B cells respond mainly to soluble antigens. Recently data clearly show that dendritic cells derived from inflammatory sites exhibit the capability of presenting non-processed and cell surface associated antigens to resting B cells. We show that HEL B cells stimulated with bead associated HEL plus anti-CD40 proliferate and survive to a much greater extant than similar B cells stimulated in culture with soluble HEL plus anti-CD40. ACS analysis of phenotypic characteristics and CSFE staining patterns demonstrate that although similar numbers of B cells initially enter cell cycle in both experimental groups, HEL B cells stimulated by immobilized antigen plus anti-CD40 are found to complete up to twice as many division cycles. In addition, a subset of these B cells appears to enter the plasmocyte differentiation pathway in the absence of exogenous cytokine supplementation. These findings indicate that presentation of bound antigen to resting HEL B cells impacts on the ability of the B cells to effectively respond to T helper cell-derived signals and support the possibility that direct physical presentation of intact antigen by APC emigrating from inflammatory sites represents an additional means for communication between the innate and adaptive immune systems.
Thymopoiesis is Enhanced by Inhibition of gp130 Signaling in Young and Aged Mice (85.15)
Authors: Kendall Shellie Marie , Ventevogel Melissa S. , Sempowski Gregory D. 
Published: January 2023

Abstract
Normal aging is characterized by attenuation of cellular immune function and progressive decline in ability to respond to pathogens. Understanding mechanisms of immunosenescence with aging will enable development of targeted therapies for enhancing immune response. We previously reported that both human and mouse thymus aging is characterized by increased expression of IL-6 family cytokines. Further we demonstrated that these cytokines are directly suppressive and involute the thymus. IL-6 family cytokines activate the gp130 cell surface receptor on thymocytes, thus we hypothesized inhibition of gp130 receptor signaling will stimulate thymopoiesis in aged mice. To test this hypothesis we infused aged BALB/c mice (20 mo) or young mice (2 mo) with neutralizing gp130 Ab and monitored thymus function over 8 weeks. Inhibition of gp130 signaling significantly elevated thymus weight and TCR gene rearrangement in the aged thymus. In young mice we also observed with gp130 inhibition significant increases in thymus weight, cellularity, TCR gene rearrangement, and naïve export to the periphery. Overall, these data demonstrate release of thymosuppressive cytokine signaling stimulates thymopoiesis by both young and aged thymus tissue. Furthermore, gp130 signaling is an important thymoregulatory pathway to explore in the pursuit of human T cell immune reconstitution therapies.
Supported by NIH HL67314, AG025150.
Aged mice exhibit increased frequencies of CD4+ T cells with a regulatory phenotype (85.20)
Authors: Maue Alexander C , Eaton Sheri , Haynes Laura 
Published: January 2023

Abstract
In vivo, age-related defects in CD4+ T cell function lead to reduced expansion, IL-2 production, NF-κB activation and B cell helper activity. These defects are significant as they result in reduced vaccine efficacy in the elderly. To date, relatively little is known about what factors contribute to these age-related defects in vivo. By using real time-PCR (RT-PCR) and intracellular flow cytometry, we have identified candidate cell subsets and molecules that may contribute to the observed immunosuppression. Spleens of young (4 months) and aged (24 months) mice displayed significantly higher expression of interleukin-10 (IL-10) as determined by RT-PCR. Analysis of cell subsets isolated by sorting revealed a significant increase in IL-10 expression in aged CD4+ T cells compared to young. Following intracellular cytokine staining, a statistically significant increase in CD4+IL-10+ cells (~15 fold) was observed in aged (26 months) compared to young (2 months) mice. Aged CD4+ T cells exhibited significantly higher expression of CD25 (~3 fold), GITR (~5 fold) and CTLA-4 (~4 fold) compared to young CD4+ T cells, as well as greater overall numbers of cells expressing each marker. Following intracellular staining, both intact and transgenic CD4+ T cell populations in aged mice displayed greater numbers of Foxp3-expressing cells compared to young populations. Our results demonstrate that in aged mice, several molecules and cells associated with immune regulation are present in significantly higher amounts than in young mice and that future experiments are required to determine the effects of these elements on naïve CD4+ T cell and aged immune responses.
This research was supported by NIH grants AG02160 and AG21054.
Inflammation regulates memory CD8 T cell development (85.3)
Authors: Cui Weiguo , Joshi Nikhil , Kaech Susan 
Published: January 2023

Abstract
During viral or bacterial infection, the distinct effects of inflammation and antigenic stimulation on effector and memory cell differentiation in CD8 T cells remain to be better defined. Our recent findings show that two subsets of effector CD8 T cells that have distinct cell fates are formed during acute infection -- the KLRG1hi IL-7Rlo short-lived effector cells (SLECs) and the KLRG1lo IL-7Rhi memory precursor cells (MPCs). SLECs will undergo apoptosis after infection subsides whereas MPCs survive and become memory CD8 T cells. Here we show that inflammation plays an important role in generating SLECs during infection. Specifically, we found that priming CD8 T cells in the absence of overt inflammation using peptide-coated dendritic cells (DCs) induced primarily MPCs, but SLECs did not form. However, if CD8 T cell activation occurred in the presence of CpG then SLECs formed, and this was TLR9-dependent. In addition, the two signals (antigen + inflammation) needed to act in concert to induce formation of the SLECs. Next, we found that IL-12 alone, but not IFNγ, was sufficient to induce SLECs. However, IFNγ appears to be permissive in this process since IFN γR−/− CD8 T cells failed to form SLECs with peptide-coated DCs + CpG. Together, these results provide new insight into how inflammatory signals control effector cell differentiation and longevity and thus, provides a novel link between inflammation and T cell homeostasis.
Impaired Peripheral B cell Development and Function in RasGRP3-deficient Mice (86.17)
Authors: Hoek Kristen L , Carlesso Gianluca , Castro Iris , Rapoport Rebecca , Bautch Victoria L , Khan Wasif N 
Published: January 2023

Abstract
Signaling through the B cell receptor (BCR) is critical for peripheral B cell maturation. We have shown that diacylglycerol (DAG) is preferentially produced in late (T2) compared to early (T1) transitional immature splenic B cells upon BCR engagement. Two major downstream effectors of DAG in B cells are PKCs and RasGRP3. BCR-mediated activation of RasGRP3 leads to activation of the Ras/ERK pathway, which is important for B cell proliferation and differentiation. To determine the role of RasGRP3 in B cell development, we analyzed the B cell compartment of RasGRP3-deficient mice obtained by gene trap (Mol Cell Biol 24:101515–28). Here we show that loss of RasGRP3 results in a decrease in splenic B cells compared to wt controls (25.6±16.6 vs 69.3±23.1, mean B cell number (x106) ± SD), with a block in peripheral B cell development between the T2 and mature follicular (FoB) stages. Additionally, peritoneal B2 B cells are severely reduced in RasGRP3−/− animals. However, development of marginal zone (MZ) and B1 B cells appears unaffected. In agreement with previous reports, we found that RasGRP3−/− B cells are impaired in BCR-mediated functions in vitro; B cell proliferation, induction of activation markers, and upregulation/maintenance of survival proteins are severely impaired in response to anti-IgM stimulation. These results reveal that RasGRP3 is necessary for BCR-mediated B cell differentiation/survival.
Zinc functions as ionic signaling molecule in primary T cell responses (87.30)
Authors: Lee Won-Woo , Pryshchep Sergey , Cui Dapeng , Weyand Cornelia M. , Goronzy Jorg J. 
Published: January 2023

Abstract
Zn is an important catalytic cofactor for enzymes and transcription factors, however, it is unknown whether these mechanisms account for the compromised immune competence with Zn deficiency or immune-enhancing function of Zn. To examine the effect of Zn on T cell priming, we stimulated naive CFSE-labeled CD4 T cells with DCs and the superantigen TSST. The system allows for separately examining high- (Vß2+) and low-avidity (Vß2-) T-cell responses. External Zn2+ concentrations inversely correlated with the activation threshold of low-avidity T-cells, pointing to an early signaling event. FACS showed that external Zn concentrations only influenced cytoplasmic Zn in activated and not in resting cells. Confocal microscopy was used to monitor Zn fluxes in FluoZin3-labeled T cells after activation with TSST-loaded DC. TCR synapse formation was visualized with anti-CD3. Zn2+ accumulated in the subsynaptic region of the T-cell activation platform within 5 min after activation, likely due to relocation and activation of zinc importers. The local Zn accumulation correlated with external Zn2+ concentration, peaked early, and started to decline again within ten min. Free Zn2+ did not show the global cytoplasmic spreading seen with Ca2+. Our data suggest that Zn functions as an ionic signal. TCR-induced Zn influx may modulate early signaling events which increases sensitivity of T-cell responses to subthreshold stimuli.
Galectin-3 translocates to the immunological synapse and inhibits cytokine production in CD4+ T cells activated by engagement of TCR (88.12)
Authors: Chen Huan-Yuan , Weng I-Chun , Maverakis Emanual , Yang Ri-Yao , Hsu Daniel K , Liu Fu-Tong 
Published: January 2023

Abstract
Galectin-3 is induced in CD4+ T cells upon activation by mitogens or TCR engagement, but its function in this cell type has not been determined. We have compared the cytokine responses of gal3−/− and gal3+/+ CD4+ T cells upon activation induced by TCR engagement and found that gal3−/− CD4+ T cells secreted more IFNγ and IL-4 compared to gal3+/+ cells. We also found that serum levels of IFNγ were significantly higher in gal3−/− than gal3+/+ mice after exposure to Staphylococcus enterotoxin B. In addition, gal3−/− CD4+ T cells exhibited higher levels of tyrosine-phosphorylated proteins, as well as enhanced phosphorylation of early signaling molecules, including LAT and Zap70, upon anti-CD3 stimulation. These results suggest that endogenous galectin-3 suppresses T cell responses. Immunofluorescence analysis revealed that galectin-3 was recruited to the immunological synapse in CD4+ T cells after TCR engagement. Our preliminary data suggest that galectin-3 expression in T cells correlates with an enhanced down-regulation of TCR when the cells are activated by engagement of the receptor, in a manner similar to that described for other intracellular proteins that negatively regulate T cell activation. We conclude that galectin-3 is an inhibitory regulator of T cell activation that down-regulates signal transduction, and functions by recruitment to the immunological synapse following activation by TCR engagement.
CD4+ T cells promoting humoral responses in mice (88.22)
Authors: Eddahri Fouad , Denanglaire Sébastien , Leo Oberdan , Andris Fabienne 
Published: January 2023

Abstract
Following antigen inoculation in naïve transgenic mice, we identified an antigen-specific CD4+ T cell subset expressing high level of both CD62L and CD44 that differ from the well characterized inflammatory cytokine producer CD62Llo CD44hi T cells. Antigen-primed CD62LhiCD44hi T cells induced extensive antibody secretion in B cell cooperation tests despite defective IL-4 and IFN-γ production, while CD62Llo CD44hi T cells secreted high amount of IL-4 and IFN-γ but did not induce optimal IgG secretion by cocultured B cells, suggesting that T cell-dependent antibody secretion by B lymphocytes (Th-B function) could be dissociated from elevated Th1 and Th2 cytokine production.
We therefore studied the stimulation requirements leading to humoral and inflammatory immune responses following a dendritic cell-based immunisation protocol. The data obtained suggested that injection of antigen-pulsed mature DC primed naïve T cells for IFN-γ secretion in vivo whereas injection of antigen-pulsed immature DC did not. However, immature DC were fully capable of promoting antigen specific antibody secretion, suggesting that activation protocols described as “suboptimal” for Th cell cytokine secretion induced optimal Th-B capacities.
Together, these studies should help in the development of novel vaccination strategies inducing optimal antibody responses while minimizing local inflammatory reactions.
FOXP3 is a homo-oligomer, and it interactions with histone acetyltransferase and class II histone deacetylases are required for repression (88.30)
Authors: Li Bin , Samanta Arabinda , Song Xiaomin , Iacono Kathryn T , Brennan Patrick , Riley James , Chatila Talal A , Banham Alison , Shen Yuan , Saouaf Sandra J , Greene Mark I 
Published: January 2023

Abstract
We have found that FOXP3 is an oligomeric component of a large supramolecular complex. The Zinc-Leuzip region of FOXP3 is sufficient to mediate its homotetramerization. Wild type FOXP3 directly binds to the human IL-2 promoter, but the E251 deletion in FOXP3 in XLAAD/IPEX patient’s T cells disrupts its homo-oligomerization, heteromerization with FOXP1, its association with the IL-2 promoter, and limits repression of IL-2 transcription after T cell activation. Additionally we report that transcriptional repression by FOXP3 involves a histone acetylase-deacetylase complex that includes TIP60 and HDAC7 and HDAC9. FOXP3 can be acetylated, and TIP60 promotes FOXP3 acetylation in vivo. Knockdown of endogenous TIP60 relieved FOXP3-mediated transcriptional repression. A minimum FOXP3 ensemble containing native TIP60 and HDAC7 is necessary for IL-2 production regulation in T cells. Moreover, FOXP3 association with HDAC9 is antagonized by T cell stimulation, and can be restored by the Trichostatin A indicating a complex dynamic aspect of T suppressor cell regulation. These findings identify a novel complex based mechanism by which FOXP3 actively mediates transcriptional repression.