bluuebunny/crossref_metadata_2025_split huggingface dataset:
Lipid raft-based membrane order is important for antigen-specific clonal expansion of CD4+ T cells (IRC2P.451)
Authors: Bamezai Anil , Schieffer Daniel , Naware Sanya 
Published: January 2023

Abstract
Lipid rafts contribute to the plasma membrane order and to its spatial asymmetry. Lipid rafts are cholesterol and saturated lipid-rich, nanometer sized membrane domains that are hypothesized to play an important role in spatiotemporal regulation of cell signaling. The raft nanodomains on the surface of CD4+ T lymphocytes coalesce during their interaction with antigen presenting cells (APCs). Sensing of foreign antigen by the antigen receptor on CD4+ T cells occurs during these cell-cell interactions. In response to foreign antigen the CD4+ T cells proliferate, allowing the expansion of few antigen-specific primary CD4+ T cell clones. Proliferating CD4+ T cells undergo differentiation into appropriate effectors tailored to mount an effective immunity to the pathogen. To investigate the role of lipid raft-based membrane order in the clonal expansion phase of primary CD4+ T cells, we have disrupted membrane order by incorporating an oxysterol, 7-ketocholesterol (7-KC) into the plasma membrane of primary CD4+ T cells expressing a T cell receptor specific to chicken ovalbumin323-339 peptide sequence and tested their antigen-specific response. We report that 7-KC, at appropriate concentrations, significantly diminishes the c-Ovalbumin323-339 peptide-specific clonal expansion of primary CD4+ T cells. Our findings suggest that lipid raft-based membrane order and membrane asymmetry is important for clonal expansion of CD4+ T cells in response to a model antigen.
α-galactosylceramide may lead to an enhanced affinity immune response (IRC3P.459)
Authors: Hawksworth David 
Published: January 2023

Abstract
α-galactosylceramide (αGalCer) has been shown to stimulate B cell proliferation in mice, leading to enhanced antibody titers. We sought to discover whether, in addition to antibody titer, we could develop a higher affinity immune response in mice administered αGalCer in the context of our standard, adjuvant based immunization methodologies. Mice were immunized three times, every 4-6 weeks, with various antigens emulsified in Freund’s adjuvant. Test animals were administered αGalCer at the time of primary immunization, in addition to the antigen. Relative affinity was determined by testing serum antibody for reactivity to limiting concentrations of solution phase antigen. Results show that, for some antigen systems, administration of αGalCer does result in a trend toward a higher relative affinity vs. control animals. Pooled data from 3 antigen systems shows an average relative affinity of 2.105 nM for αGalCer mice vs. 3.134 nM for control mice. It was also observed, for one antigen system, that only those animals administered αGalCer developed a significant immune response. Our results indicate that administration of αGalCer at the time of primary immunization may lead to a slightly higher average affinity response in mice and can also be the difference between the development of a significant immune response vs. a very weak response.
B cells potently amplify T cell IL-4 responses following non-BCR mediated cysteine protease uptake (IRC3P.463)
Authors: Dwyer Daniel , Woodruff Matthew , Carroll Michael , Austen K. , Gurish Michael 
Published: January 2023

Abstract
Many common allergens such as house dust mite antigen and grass pollen are complex mixes of proteins and other compounds, both of which may have innate signaling capacity. Understanding the role of each of these signals is essential to determining what separates allergens from other innocuous proteins. Papain is a cysteine protease structurally similar to proteases found in many common allergens, including grass pollen and house dust mite antigen. Papain induces a potent IL4 response directed by T cells in the popliteal lymph nodes (PLN) of mice following footpad immunization directed against the protease, but the mechanisms behind this response and all of the relevant antigen presenting cells driving this polarization have not yet been fully elucidated. We have identified a novel capacity for B cells to rapidly bind and internalize papain to endosomes in an innate manner not mediated by the specificity of the B cell receptor. B cells subsequently regulate the adaptive immune response by enhancing CD4+ T cells Inducible T cell Costimulator (ICOS) expression on CD4+ T cells and amplifying both Th2 cell and T follicular helper cell induction. Antibody blockade of ICOS ligand, expressed by PLN B cells but not DC at the peak of the immune response, inhibits this IL-4 induction in WT but not B cell-deficient μMT mice. Thus, B cells play a critical role in amplifying adjuvant-dependent Th2 polarization following non-canonical acquisition and internalization of cysteine protease.
Cellular mechanism for immunogenicity of PEGylated “stealth” liposomes (IRC3P.466)
Authors: Watanabe Akiko , Cain Derek , Hanson Melissa , Kim Mikyung , Reinherz Ellis , Irvine Darrell , Kelsoe Garnett 
Published: January 2023

Abstract
Liposomes have been used as a vehicle to deliver pharmaceutical agents and as a carrier of vaccine antigens for malaria, cancer and HIV. The potency of a liposome-based vaccination is affected by the lipid formulation and inclusion of adjuvants, in addition to the immunogenicity of target antigens. While liposomal vaccines elicit robust humoral responses against target antigens, recent studies have shown that lipid components, such as monophosphoryl lipid A (MPLA), or liposome adducts, such as polyethylene glycol (PEG), are also capable of being targets of induced antibody. To understand the cellular interactions and molecular pathways responsible for the anti-liposome responses, we immunized mice with PEGylated “stealth” liposomes containing lipid-arrayed membrane-proximal external region (MPER) of HIV-1, and followed the kinetics of the humoral responses against liposome components as well as MPER. Immunized mice mounted robust IgM and IgG against PEG and, to a lesser extent, MPLA. PEG IgG was dependent on CD154, yet the requirement for T cells was less stringent. Moreover, PEG IgG was generated independent of MHCII, CD1d, and TCRδ, suggesting a novel form of T-cell help. Anti-MPER IgG, in contrast, was dependent on CD154, MHCII, and LAT. These results suggest that anti-liposome IgG responses are induced by atypical T-B cognate interaction, which is independent of CD4, NKT and γδ T cells and are distinct from MPER IgG responses, which require classical T-B collaboration.
TRIF signaling is essential for TLR4-driven IgE class switching (IRM5P.707)
Authors: Janssen Erin , Ozcan Esra , Liadaki Kyriaki , Jabara Haifa , Manis John , Ullas Sumana , Akira Shizuo , Fitzgerald Katherine , Golenbock Douglas , Geha Raif 
Published: January 2023

Abstract
The TLR4 ligand LPS causes mouse B cells to undergo IgE and IgG1 isotype switching in the presence of IL-4. TLR4 activates two signaling pathways mediated by the adaptor molecules MyD88 and TRAM, which recruits TRIF. Following stimulation with LPS+IL-4, Tram-/- and Trif-/- B cells completely failed to express Cε germ line transcripts (GLT) and secrete IgE. In contrast, Myd88-/- B cells had normal expression of Cγ1 GLT, but reduced IgE secretion in response to LPS+IL-4. Following LPS+IL-4 stimulation, Cγ1 GLT expression was modestly reduced in Tram-/- and Trif-/- B cells, whereas Aicda expression and IgG1 secretion were reduced in Tram-/-,Trif-/-, and Myd88-/- B cells. B cells from all strains secreted normal amounts of IgE and IgG1 in response to anti-CD40+IL-4. Following stimulation with LPS+IL-4, Trif-/- B cells failed to sustain NFκB p65 nuclear translocation beyond 3 hours and had reduced binding of p65 to the Iε promoter. Addition of the NFκB inhibitor, JSH-23, to wild-type B cells 15 hours after LPS+IL-4 stimulation selectively blocked Cε GLT expression and IgE secretion, but had little effect on Cγ1 GLT expression and IgG secretion. These results indicate that sustained activation of NFκB driven by TRIF is essential for LPS+IL-4 driven activation of the Cε locus and class switching to IgE.
RGC-32 is essential for TGF-β mediated Th17 cell differentiation (LYM3P.728)
Authors: Rus Violeta , Tegla Cosmin , Nguyen Vinh , Cudrici Cornelia , Badea Tudor , Rus Horea 
Published: January 2023

Abstract
Our group has cloned and characterized the Response Gene to Complement (RGC)-32 which is induced by C5b-9 and exerts an important role in cell cycle activation via cyclin B1-CDC2 activation and Akt phosphorylation. RGC-32 is also a downstream target of TGF-β and plays a role in fibrogenesis. As Th17 and induced T regulatory cells (iTreg) differentiate under the control of TGF-β, we determined whether RGC-32 plays a role in this process using RGC-32 KO mice generated in our lab. RGC-32 was not required for peripheral T cell homeostasis as we observed normal cellularity, T cell subset composition and phenotypic markers. However, RGC-32 KO CD4+ T cells failed to differentiate normally to Th17 lineage as they exhibited a significant reduction in IL-17 and RORγt mRNA and protein expression in vitro and a decreased proportion of IL-17+ cells in lamina propria lymphocytes in vivo. The defect in IL-17 expression appeared to be mediated by decreased expression of IL-6R and TGF-βRI at baseline and under Th17 conditions and was not restored by IL-2 inhibition. A trend for increased iTreg differentiation was noted in RGC-32 KO CD4+ T cells under Treg skewing conditions but the difference was not statistically significant. Differentiation of CD4+ T cells into Th1 cells was decreased in RGC-32 KO mice while Th2 cells developed normally. These results suggest that RGC-32 is a novel mediator of Th17 differentiation in vitro and in vivo through modulation of IL-6R and TGF-βRI expression.
Type I IFN regulates non-canonical inflammasome activation and impairs protective immunity during fatal ehrlichia-induced toxic shock (MPF1P.762)
Authors: Yang Qin , Stevenson Heather , Scott Melanie , Ismail Nahed 
Published: January 2023

Abstract
Inflammasome complexes such as NLRP3 play a major role in antimicrobial defense to infection. Canonical and non-canonical pathways activate caspase 1 and caspase 11 to secrete IL-1β and induce inflammatory cell death. Ehrlichia, obligate intracellular bacteria that cause fatal toxic shock in humans and mice, activate inflammasomes and induce expression of type 1 interferon (IFN1). The contribution of the NLRP3 and IFN1 to Ehrlichia-induced toxic shock is not known. We show that NLRP3-/- and Caspase 1-/- mice succumb to lethal Ehrlichia infection similar to wild type (WT) mice. Unlike caspase 1-/- and WT, NLRP3-/- mice have lower bacterial burdens with higher IFNγ, suggesting that NLRP3 impairs bacterial clearance in a caspase 1-independent manner. Unexpectedly, infected IFNαR-/- mice were protected against a normally lethal dose of Ehrlichia with effective bacterial clearance, altered pathology with less apoptosis, increased IFNγ, decreased IL-10 and IL-10 producing Treg cells, and increased number of protective CD4+ Th1 and NKT cells. Lack of IFN1 signaling abrogated caspase 11 activation and IL-1β secretion, suggesting that IFN1 activates the non-canonical inflammasome. Our data also show that WT, but not IFNαR-/-, cells exhibit defective autophagy. Together, these data demonstrate that IFN 1 signaling suppresses autophagy and activates the non-canonical inflammasome pathway contributing to impaired protective immunity and development of fatal Ehrlichia-induced toxic shock.
Deleterious role for type I interferon during acute blood stage malaria infection (MPF1P.767)
Authors: Spaulding Emily , Lauvau Gregoire 
Published: January 2023

Abstract
Malaria is a severe and life-threatening disease that affects over 219 million people worldwide every year. In patients, a robust induction of the type I IFN pathway is observed, yet studies in mouse models have failed to determine whether type I IFN contributes to host protection or pathogenesis. We thus hypothesized that type I IFN is essential during blood stage malaria infection. Using Plasmodium yoelii (Py) YM which causes an acute lethal blood infection in C57BL/6 WT mice, we found that while WT mice rapidly succumbed, mice lacking the type I IFN receptor (IFNαβR-/-) mostly survived (&gt;80%). Innate immune cells, particularly monocytes and NK cells, underwent early and robust activation and overwhelming production of inflammatory mediators in WT mice, whereas this was prevented in IFNαβR-/- mice. We are currently investigating precisely the cellular source of type I IFN and the mechanisms that regulate the activation of type I IFN-producing cells during this infection. Our results favor a model in which type I IFN promotes over-exuberant activation of innate monocytes and NK cells, leading to overwhelming inflammation and death of Py YM-infected mice. Overall, our work highlights a novel and unsuspected role for type I IFN in this significant human parasitic disease. We believe these results contribute to our understanding of the immunological cues leading to severe disease in malaria patients.
Iron acts as a second signal during coinfection with M. tuberculosis and N. brasiliensis leading to exacerbated M. tuberculosis disease outcome (MPF1P.769)
Authors: Dietzold Jillian , Gopalakrishnan Archana , Konowich Jill , Salgame Padmini 
Published: January 2023

Abstract
Previously, our laboratory has demonstrated that N. brasiliensis (Nb), an intestinal helminth, is able to exacerbate Mtb burden in coinfected Balb/c mice through generation of alternatively activated macrophages (AAMs). To examine the role of host genetic background in infection we performed coinfection experiments in C57Bl/6 mice. In contrast to Balb/c, coinfected C57Bl/6 mice were able to control Mtb burden comparable to mice infected with Mtb alone, despite mounting a Th2 response. This led us to hypothesize that a factor in addition to the generation of AAMs was responsible for the increased Mtb burden seen in the coinfection model. Concurrent experiments designed to understand Mtb survival within in vitro derived AAMs showed that IL-4 and IL-13 treatment was insufficient to cause differences in Mtb growth, compared to untreated macrophages. However the addition of iron significantly increased Mtb growth in AAMs. Iron, besides being important for Mtb growth; is also present in the lungs following tissue damage caused by Nb larval migration. To determine the role of iron during Nb infection we evaluated the expression of iron responsive genes in the murine lungs at 3, 5, and 7 days post infection. Balb/c and C57Bl/6 mice exhibited distinct profiles in response to Nb infection consistent with iron recycling and iron sequestration, respectively. Future studies aim to further understand the role of iron as important cofactor in regulating bacterial burden during coinfection.
IgG anti nocardia brasiliensis antibodies block IgM mediated protective effect in BALB/c mice. (MPF2P.814)
Authors: Salinas-Carmona Mario , Perez Rivera Luz , Vazquez Anna , Welsh Oliverio 
Published: January 2023

Abstract
Intracellular bacteria such as, Nocardia brasiliensis, produce a chronic human disease named actinomycetoma. Diagnosis of mycetoma is made on clinical findings as swelling and deformation of infected tissue and confirmed by culturing the offending microbe or by an ELISA test. Pathogenesis of mycetoma is little known and role of antibodies is controversial but it has been published that that cell mediated immunity is responsible of host protection in other intracellular bacterial infections. We used Nocardia brasiliensis to infect BALB/c mice and studied the role of both monoclonal and polyclonal antigen-specific IgM and IgG antibodies. We investigated antibodies role in host protection using passive immunization. Mice recipients of anti N. brasiliensis antibodies were then infected to assess actinomycetoma development. We found that IgM but not IgG anti N. brasiliensis antibodies from sera of infected animals and IgM, but not IgG anti N. brasiliensis monoclonal antibodies were able to confer protection and prevented actinomycetoma infection in experimental animals. IgG antibodies blocked the IgM protective effect in vivo. Implications of these finding in vaccine development are presented.
Phenotypic and functional analysis of the immune response to chronic T.cruzi infection in C3H/HePas mice (MPF2P.805)
Authors: Fonseca Raíssa , Borges da Silva Henrique , Salgado Rafael , do Nascimento Rogério , D'Império Lima Maria , Álvarez José 
Published: January 2023

Abstract
C3H/HePAS mice infected with T. cruzi Sylvio X10/4 develop a chronic cardiomyopathy similar to observed in humans, associated with presence of inflammatory infiltrates in the heart, suggesting that immune responses mounted during chronic infection play an important role in disease progression. However, these immune responses are not well understood. Thus, we analyzed the activation profile of blood and spleen immune cells at 390 days post infection, compared to age matched controls. We observed, at 390dpi, an increase in spleen and blood CD8+ T cell, and a decrease in CD11b+CD11c+ blood cell frequencies. We also observed increased PDL1 expression on both CD4+ and CD8+ spleen T cells, and of PDL1 on heart of chronic mice, as assessed by qRT-PCR, indicating that PD1/PDL1 inhibitory pathway may be activated. Chronic mice had also more CD4+IFNγ+ spleen cells, which had better proliferation ability when stimulated with T. cruzi antigen. There was increased CD69 expression in splenic CD4+ and blood CD8+ T cells. Importantly, there was a positive correlation between the expression of PDL1, PD1 and CD69 in CD4+ and CD8+ spleen T cells. In blood, correlations of PD1 and PDL1 expression were found on B, CD4+ and CD8+ T cells. Our results suggest that continued activation in chronic mice may induce higher levels of PD1/PDL1, which can be important to inhibit heart infiltrating cells, favoring parasite persistence. New experiments are needed to evaluate the importance of this pathway.
Intestinal epithelial cells contribute to Citrobacter rodentium induced infectious colitis through an NF-κB-dependent mechanism (MUC2P.826)
Authors: Mackos Amy , Galley Jeff , Parry Nicola , Boyaka Prosper , Bailey Michael 
Published: January 2023

Abstract
Exposing mice to a psychological stressor during challenge with the enteric pathogen Citrobacter rodentium (Cr) exacerbates Cr-induced infectious colitis. Enhanced recruitment of inflammatory monocytes to the colon, via a CCL2-dependent mechanism, is essential for the exacerbation of infectious colitis by stressor exposure, however the cell type responsible for the increase of colonic CCL2 is unknown. Previous evidence demonstrates that Cr-induced CCL2 is through an NF-κB mediated mechanism, thus this study was designed to test the hypothesis that NF-κB-derived gene expression from intestinal epithelial cells is necessary for the exacerbation of colitis via the recruitment of inflammatory monocytes, as well as the corollary hypothesis that inflammatory monocytes contribute to infectious colitis also via NF-κB activation. To test these hypotheses, two different strains of mice were challenged with Cr in which classical NF-κB activation is defective via the deletion of IKKβ in either intestinal epithelial cells (IKKβΔIEC) or myeloid-derived cells (IKKβΔMY). There was a reduction in Cr-induced colon mass, colonic histopathology, and colonic inflammatory mediator gene expression in IKKβΔIEC mice as compared to wild type controls. In addition, Cr-induced colon mass, histopathology, and colonic gene expression is further reduced in IKKβΔMY mice. Together, these data illustrate the deleterious effects of pathogen induced NF-κB activation has during enteric pathogen challenge.
TGFβ converts Th1 cell into Th17 cells through stimulation of Runx1 expression under inflammatory conditions in intestines (MUC2P.819)
Authors: Liu Houpu , Feng Ting , Li Qingjie , Zhang Wenbo , Yao Suxia , Elson Charles , Cong Yingzi 
Published: January 2023

Abstract
Although accumulating evidence demonstrates that differentiated CD4 cells preserve plasticity to alter phenotypes under various conditions, it is still unclear how stable Th1 cells are and whether Th1 cells can convert into Th17 cells. The high stability of Th1 is supported by some epigenetic studies and numerous reports of Treg and Th17 cells conversion into Th1 cells but not vice versa. However, recent reports of Th1 cell conversion to Treg, Th2 and Tfh cells argue the absolute stability of Th1 lineage. By using IFNγThy1.1 CBir1 TCR transgenic reporter mice, whose TCR is specific for an immunodominant microbiota antigen, we investigate the stability of Th1 cells under intestinal inflammatory conditions. Transfer of purified CBir1 specific IFNγ+Th1 cells induces colitis in RAG-/- mice and IFNγ+Th1 cells convert into IL-17+ Th17 but not Foxp3+ Treg cells in the inflamed intestines. TGFβ, IL-6 and IL-2, but not hypoxia factors, differentially regulate Th1 to Th17 conversion. TGFβ induction of transcriptional factors, Runx1 and RORγt, is crucial for the conversion, in that silencing Runx1 by siRNA inhibits Th1 conversion into Th17 cells. Furthermore, using ChIP assay, we show that TGFβ enhances histone acetylation but inhibits trimethylation of Runx1 and RORγt binding sites on il-17 or rorc gene in Th1 cells. In conclusion, our data demonstrate that Th1 cells convert into Th17 cells under inflammatory conditions in intestines, which is mediated by TGFβ-induction of Runx1.
SLP76 N-terminal tyrosine residues regulate a dynamic signaling equilibrium involving feedback of proximal TCR signaling (TECH1P.848)
Authors: Ji Qinqin , Ding Yiyuan , Salomon Arthur 
Published: January 2023

Abstract
SLP76 is a cytosolic adaptor protein that plays an important role in TCR mediated T-cell signaling pathway. SLP76 consists of three functional domains: an amino-terminal acidic domain with three tyrosine phosphorylation sites, a central proline-rich domain, and a carboxy-terminal SH2 domain. The direct influence of each domain of SLP76 in T-cell signaling pathway is not well understood. In our work, phosphoproteomic methods in mass spectrometry were used to do a phosphoproteomic analysis of cells containing a mutant form of SLP76 where three tyrosine residues are replaced with phenylalanines. Thousands of unique phosphorylation sites upon a time course of T-cell receptor stimulation in these cells are identified. Mutation of the three N-terminal tyrosine residues of SLP76 phenocopied SLP76 deficient cells for the majority of tyrosine phosphorylation sites observed including feedback on proximal TCR signaling proteins. Meanwhile, reversed phosphorylation changes were observed on Tyr192 of Lck when comparing Y3F mutant cells to SLP76 deficient cells. In addition, the N-terminal tyrosine sites of SLP76 also perturbed phosphorylation of Tyr440 of Fyn, Tyr702 of PLCγ1, Tyr204, Tyr397 and Tyr69 of ZAP-70, revealing new modes of regulation on these sites. All these findings confirmed the central role of N-terminal tyrosine residues of SLP76 in T cell signaling and shed light on novel signaling networks that are uniquely regulated by SLP76 N terminal tyrosine residues.