bluuebunny/crossref_metadata_2025_split huggingface dataset:
A novel murine α4 integrin splicing variant is a specific receptor for VCAM-1 (61.2)
Authors: Kon Shigeyuki , Kouro Hitomi , Matsuda Tadashi 
Published: January 2023

Abstract
Integrins affect multiple cell motility in control of cell survival, growth, or differentiation mediated by cell-cell and cell-extracellular matrix interaction. We previously reported that α9 integrin splicing variant, SFα9, promotes wild type α9 integrin-dependent adhesion. In this report, we introduce a new murine α4 integrin splicing variant mSFα4, which have new short cytoplasmic tail. In inflamed tissues including rheumatoid joint, the expression of mSFa4, as well as wild type α4 integrin, is upregulated. Interestingly, cells expressing mSFα4 bind to VCAM-1 but not other α4 integrin ligand such as fibronectin CS1 or osteopontin. Binding of cells coexpressing mSFα4 and wild type α4 integrin to α4 integrin ligands is promoted than cells expressing wild type α4 integrin. Thus, mSFα4 functions modulation of α4 integrin. Now, we pursue the metastatic function of mSFα4 by using mSFα4-specific knockdown tumor cells.
Migratory properties of pulmonary dendritic cells are developmentally programmed (61.4)
Authors: Cook Donald , Burgents Joseph , Nakano Hideki 
Published: January 2023

Abstract
Pulmonary dendritic cells (DCs) take up inhaled allergens and migrate to draining lymph nodes (LN) to present antigens to naïve T cells. However, pulmonary DCs are also required to elicit inflammation after allergen challenge, suggesting the presence of both migratory and non-migratory DCs. There are two phenotypically distinct major lung DC subsets, defined by their expression of CD103 and CD11b, respectively. We used a novel Ccr7-gfp knock-in reporter mouse to study Ccr7 expression in lung DCs. Ccr7-gfp was expressed in many CD103 DCs, but in few CD11bhi DCs. Using the dye PKH26 to track lung DCs, we found that many CD103+ DCs migrate to draining LNs, whereas relatively few CD11bhi DCs do so. Thus, CD11bhi lung DCs are comprised of both migratory and non-migratory cells. Further analysis revealed that monocyte-derived (mo)CD14hi CD11bhi DCs, including LPS-elicited Ly-6Chi inflammatory DCs, do not express Ccr7 or migrate to LNs. Flt3l-/- mice, which lack DCs derived from the common DC precursor (CDP) but retain moDCs, possessed almost no migratory Ccr7-expressing DCs in the lung. Conversely, mice lacking both Ccr2 and Cx3cr1 lacked moDCs, but retained migratory, CDP-derived DCs. These findings show that the migratory activity of CD11bhi DCs is developmentally programmed; CDP-derived CD103+ and CD11bhi pulmonary DCs are migratory, whereas monocyte-derived CD11bhi DCs do not express Ccr7 and therefore remain in the lung, even during acute pulmonary inflammation.
Differentiation and detection of mouse CD4+ Th cells by measuring intracellular cytokine production (62.6)
Authors: Santos Mark , Williams Yuko , Weldon Don , Olcott Angelica , Hsu Matthew 
Published: January 2023

Abstract
Naïve CD4+ T cells can give rise to a variety of subsets of Th cells depending on the nature of the immune response, and subsequently release a distinct subset of cytokines. Conventionally these cytokines can be measured in a secreted format (e.g. Luminex or ELISAs), but here we describe a novel method to reliably differentiate mouse CD4+ T cells, and further characterize the differentiated lineages by measuring intracellular cytokines using flow cytometry. In addition, we also describe the use of a proprietary fixable viability dye in order to eliminate the false positives associated with non-specific staining. Based on an optimized protocol for differentiating CD4+ T cells, we have developed differentiation tools designed to obtain the desired Th cell lineages. Following naïve T cell differentiation toward specific Th lineages, we further developed a flow cytometry assay to measure intracellular cytokine production by blocking the secretion of the cytokine with Brefeldin A. We further eliminate non-specific staining due to dead or dying cells using a proprietary fixable viability dye and accurately determine the percentage of Th lineages using guava benchtop flow cytometer and InCyte analysis software.
Thymic Selection of T cells expressing CD8 independent high affinity human TCR αβ in TCR transgenic mice (63.7)
Authors: Thyagarajan Krishnamurthy , Al-Khami Amir , Naga Osama , Fang Quan , Nishimura Michael , Mehrotra Shikhar 
Published: January 2023

Abstract
Thymocytes undergo a number of selection events before egressing the thymus and moving to lymphoid organs as mature T cells. The chronology of these molecular and cellular events is crucial for determining the function of mature T cells. We have established a transgenic mouse model using a CD8 independent human TIL1383I αβTCR that was isolated from MHC I restricted CD4+ T cells obtained from tumor infiltrating lymphocytes of a metastatic melanoma patient and recognizes tyrosinase epitope presented by HLA-A2. We analyzed the cell fate of thymocytes in these mice in the presence of different selecting ligands. Surprisingly, this HLA-A2 restricted TCR was expressed on peripheral CD4+ and CD8+ T cells in HLA-A2- C57BL/6 mice. When bred into MHC I and MHC II deficient mice, CD8+ and CD4+ T cells respectively were absent from the periphery indicating that transgenic T cells are selected by MHC I and MHC II ligands in the mouse. Development of T cells in murine MHC deficient thymus directed majority of the transgenic T cells to CD3+CD4-CD8- phenotype. The presence of HLA-A2 also resulted in &gt;90% of the transgenic T cells to egress the thymus with a CD3+CD4-CD8- double-negative phenotype. While peripheral transgenic T cells in MHC deficient or MHC sufficient HLA-A2+ background were CD4 and CD8 coreceptor deficient, they retained antigen recognition function. The above data suggests that CD8 independent TIL1383I αβTCR does not strictly follow rules of MHC-specific thymic selection.
IL-9 is Required for TSLP-mediated Allergic Airway Inflammation (64.3)
Authors: Yao Weiguo , Zhang Yanlu , Tepper Robert , Kaplan Mark , Zhou Baohua 
Published: January 2023

Abstract
Thymic stromal lymphopoietin (TSLP) is an epithelial cell derived cytokine important for the initiation and development of Th2-mediated allergic inflammation. In this study, we identified a significant association between IL-9 and TSLP concentration in the serum of infants with atopic dermatitis. Th9 cultures increased IL-9 production in a TSLP dose-dependent manner, parallel with an increase in pSTAT5 binding to Il9, Irf4 and Gata3 loci. Lung-specific transgenic expression of TSLP stimulated IL-9 production in the lung, and anti-IL-9 treatment attenuated TSLP-induced airway inflammation. Using an adoptive transfer model, we observed that TSLP promoted Th9-induced allergic inflammation in an IL-9-dependent manner, with affects directly on T cells. Together, our results demonstrate that TSLP promotes Th9 differentiation, and define a requirement for IL-9 in TSLP-induced allergic inflammation.
Transglutaminase 2 expressed in mast cell activation induces IgE production in B cells, airway inflammation and remodeling via up-regulating CD40L and cytokine expression in mouse allergic asthma (64.4)
Authors: Ro Jai , Lee Yun Song , Kim Soo Youl , Hong Gwan 
Published: January 2023

Abstract
TGase 2 is over-expressed in a variety of inflammatory diseases including allergic asthma. This study aimed to investigate the role of TGase 2 in signaling pathways in mast cells related to OVA-induced allergic asthma in vitro and in vivo. BMMCs isolated from C57BL/6 WT or TGase 2-/- mice were activated with Ag/Ab. Mice were sensitized and challenged with OVA to induce asthma. [Ca2+]i level were determined by confocal, mast cells by May-Giemsa, LT, IgE and cytokine levels by ELISA, expression of TGase 2, PLA2, EMBP, Muc5ac, collagen, cytokines, MMPs and TIMPs by Western blotting or RT-PCR, NF-kB activity by EMSA, the recruitment of inflammatory cells into BAL fluid or lung tissues by Diff-Quik and H&E, AHR by Fexivent system. Act-WT-BMMCs expressed TGase 2, and increased the [Ca2+]i level, activities of I-kB, NF-kB and PLA2, LTs level, expression of CD40/CD40L, cytokines and MMPs, collagen deposition, and IgE production in act-B cells, whereas act-KO-BMMCs reversed all of them. TGase 2-/- mice protected against OVA-specific IgE production as well as AHR, expression of signaling molecules and mediator release in BAL cells and lung tissues versus those in WT mice. Our data suggest that TGase 2 expression and Ca2+ influx required by bidirectional events in mast cell activation facilitate IgE production in B cells via regulating mast cell CD40L expression, and induce the expression of numerous molecules associated with airway inflammation and remodeling in allergic asthma.
Antigen-specific B cells are not required for soluble OVA protein induced T-cell tolerance through i.v. route (65.8)
Authors: Zhang Ai-Hong , Scott David 
Published: January 2023

Abstract
Intravenous exposure to soluble proteins can lead to antigen-specific immune tolerance. Moreover, resting B cells have been shown to be tolerogenic antigen-presenting cells (APC) in vivo. However, it is still not clear whether this APC function is fulfilled by the rare antigen-specific B cells or the abundant antigen non-specific B cells. To address this, we used soluble chicken ovalbumin (OVA) as a model antigen and employed B-cell receptor (BCR) transgenic mice (8.18C5 Tg) of C57BL6 (B6) background. In the homozygous 8.18C5 Tg mice, ~97% of peripheral B cells contain recombined immunoglobulin heavy chain specific for non-related myelin oligodendrocyte glycoprotein. As expected, a single i.v. dose of soluble OVA in wt B6 mice induced significant B+T cell tolerance. The 8.18C5 Tg mice immunized with OVA+CFA failed to produce any detectible anti-OVA antibody response, confirming the lack of OVA-specific endogenous B cells in these mice. Importantly, pretreatment of the 8.18C5 Tg mice with 25µg soluble OVA i.v. also resulted in profound T-cell tolerance upon OVA+CFA challenge a week later. Since antigen non-specific B cells can present processed antigen after soluble protein exposure in vivo, our results support the notion that antigen non-specific resting B cells in the steady state may participate in the peripheral tolerance to self proteins.
ROR-alpha is required for natural helper cell development in all tissues from a bone-marrow derived progenitor and the induction of allergic inflammation (67.11)
Authors: Halim Timotheus , MacLaren Aric , Romanish Mark , Gold Matthew , McNagny Kelly , Takei Fumio 
Published: January 2023

Abstract
Natural Helper (NH) cells are Th2 cytokine-producing innate lymphoid cells (ILCs) in, or associated with the mucosal tissues of the lung and gut. Currently, the lineage relationship between NH cells in different tissues and between NH cells and interleukin-22 (IL-22)-producing retinoid receptor-related orphan receptor (ROR)γt-positive ILCs is unclear. Here, we report that NH cells express RORα but not RORγt. RORα-deficient, but not RORγt-deficient, mice lack NH cells in the lung, small and large intestines while all other lymphocytes including RORγt+ ILCs are unaffected. We have also identified RORα-dependent Lin-Sca-1hic-kit-CD127+CD25+ NH cell progenitors different from common lymphoid progenitors in the bone marrow. The putative NH cell progenitors transplanted into NOD/SCID/Il2rg-/- mice develop into NH cells but not other lymphocytes including RORγt+ ILCs. Thus, all NH cells belong to a distinct cell lineage separate from other lymphoid cell lineages. NH cell-deficient mice generated by RORα-deficient bone marrow transplantation into lymphodeficient Rag2-/-Il2rg-/- mice have normal Th2 cell response but fail to develop acute allergic lung inflammation in response to protease allergens. NH cell-deficient mice further reveal the role of NH cells in allergic lung inflammation.
Efficient capture of infected neutrophils by dendritic cells in the skin inhibits the early anti-leishmanial response (67.13)
Authors: Ribeiro-Gomes Flavia , Peters Nathan , Sacks David 
Published: January 2023

Abstract
Neutrophils and dendritic cells (DC) converge at sites of acute inflammation in the skin following pathogen deposition by the bites of arthropod vectors or by needle injection. How neutrophils might interact with and modulate the function of DCs in these inflammatory settings has not been previously addressed. Neutrophils are the predominant recruited and infected cells during the earliest stage of Leishmania major infection in the skin. The infected neutrophils recovered from the skin expressed elevated levels of apoptotic markers and were preferentially captured by dermal DCs, the majority of which acquired their parasites via uptake of infected neutrophils. Neutrophil depletion augmented the expression of activation markers on infected DCs and their ability to present Leishmania antigen ex vivo, and markedly enhanced CD4+ T cell priming against parasite-derived antigen in vivo. The findings suggest that L. major exploits the apoptotic cell clearance function of DCs to suppress the development of acquired resistance until the acute neutrophilic response is resolved.
The impact of type I interferon on the development of the adaptive immune response (67.14)
Authors: Pinto Amelia , Daffis Stephane , Brien James , Sheehan Kathleen , Schreiber Robert , Diamond Michael 
Published: January 2023

Abstract
Determining the impact of type I interferon on immune responses using mice missing the common IFN-α/β signaling receptor (IFNAR- /-) is limited to studying the functions of type I IFN at all stages of viral infection. To define the temporal functions of type I IFN signaling in the context of infection by West Nile virus (WNV), we treated mice with MAR1-5A3, a neutralizing, non cell-depleting anti-IFNAR antibody at later stages of infection. Inhibition of type I IFN signaling at or before day 2 after infection was associated with markedly enhanced viral burden, whereas treatment at day 4 altered WNV dissemination less dramatically. Blockade of type I IFN signaling starting at day 4 induced dysfunctional CD8+ T cells with depressed cytokine responses and expression of phenotypic markers suggesting exhaustion. The later maturation phase (days 4-8) of anti-WNV CD8+ T cell development requires type I IFN signaling. Collectively, our results suggest that cell non-autonomous type I IFN signaling shapes maturation of antiviral CD8+ T cell response at a stage distinct from the initial priming event.
Genome-wide RNAi screen reveals a novel role of WNT/CTNNB1 signaling pathway in regulation of innate antiviral responses (67.7)
Authors: Lamarre Daniel , Baril Martin , Es-Saad Salwa , Chatel-Chaix Laurent , Karin Fink Karin Fink , Audette Karine , Guenier1 Anne-Sophie , Germain Marie-Anne , Duchaine Jean , Servant Marc , Grandvaux Nathalie 
Published: January 2023

Abstract
To identify new regulators of innate antiviral immunity, we completed the first genome-wide gene silencing screen assessing the transcriptional response at the interferon-β gene (IFNB1) promoter following Sendai virus (SeV) infection. We identified 237 potential modulator genes for which negative or positive actions of gene products were mapped to the different steps of the antiviral responses from virus sensing, signal propagation/amplification up to the feedback regulation. In the present study, we will report on specific proteins that promote IFNB1 expression and innate antiviral response. The functional genomics screen uncovers a novel link between WNT family members and innate antiviral immunity. We show that virus-induced secretion of WNT2B and WNT9B down regulates IFNB1 and ISG56 expression in a β-catenin (CTNNB1)-dependent mechanism. The antiviral response is drastically reduced by GSK3 inhibitors but completely restored in CTNNB1 knockdown cells. The findings confirm a novel regulation of the innate antiviral response by a canonical WNT/GSK3/CTNNB1 pathway in a negative feedback mechanism. The study identifies novel avenues for therapeutically regulating innate immunity for effective treatment of viral infection and prevention of excessive response in autoimmune diseases.
Molecular Mechanisms Responsible for the Selective and Low-grade Induction of Pro-inflammatory Mediators by Lipopolysaccharide (68.10)
Authors: Li Liwu , Maitra Urmila , Deng Hui , Baker Bianca 
Published: January 2023

Abstract
Low dose endotoxemia is prevalent in humans with adverse health conditions, and correlates with the pathogenesis of chronic inflammatory diseases such as atherosclerosis, diabetes, and neurological inflammation. However, the underlying molecular mechanisms are poorly understood. Here, we demonstrate that subclinical low dose lipopolysaccharide (LPS) skews macrophages into a mild pro-inflammatory state, through cell surface toll-like-receptor 4 (TLR4), interleukin-1 receptor associated kinase-1 (IRAK-1), and the toll-interacting-protein (Tollip). Unlike high dose LPS, low dose LPS does not induce robust activation of nuclear factor kappa-B (NFκB), mitogen-activated protein kinases (MAPK), phosphoinositide 3-kinase (PI3K), or anti-inflammatory mediators. Instead, low dose LPS induces activating transcription factor 2 (ATF2) through Tollip-mediated generation of mitochondrial reactive oxygen species (ROS), allowing mild induction of pro-inflammatory mediators. Our data reveal novel mechanisms responsible for skewed and persistent low grade inflammation, a cardinal feature of chronic inflammatory diseases.
Intrinsic defect in the phosphoinositide 3-kinase - Akt pathway plays a role in cytokine dysregulation in aged mouse macrophages stimulated with TLR ligands or heat killed Streptococcus pneumoniae (68.11)
Authors: Fallah Mosoka , Rogier Eric , Garvy Beth , Bondada Subbarao 
Published: January 2023

Abstract
Age-associated defects in innate immunity have a role in reduced immunity and vaccine efficacy to many Gram positive bacteria like Streptococcus pneumoniae . We have previously shown that splenic macrophages from aged mice exhibited a cytokine dysregulation; impaired production of pro-inflammatory cytokines but enhanced production of IL-10 upon signaling via TLR-2 and TLR-4 or when stimulated with IL-1 or Heat killed S. pneumoniae (HKSP). We showed in this study that aged BMDM demonstrated a cytokine dysregulation phenotype similar to splenic macrophages upon stimulation with TLR2, 4 and 9 ligands, or HKSP. We hypothesized that an intrinsic age-associated increase in the activity of phosphatidyl inositol 3-kinase (PI3K)-Akt- signaling pathway may be playing a causal role in the age-associated cytokine dysregulation. We showed a heightened activity of PI3K in the aged BMDM compared to the young. Inhibition of PI3K in BMDM increased translocation of p65Nf-κB and induction of pro-inflammatory cytokines when stimulated with these ligands. The increase in activity of PI3K-Akt pathway appears to be responsible for cytokine dysregulation in the aged macrophages independent of the splenic microenvironment and could thus be modulated to produce cytokines needed to support B-cell activation and differentiation.
Acidic residues in the juxta-transmembrane region of toll-like receptor 9 are crucial for binding UNC93B1 (68.12)
Authors: Kim Jihee , Brinkmann Melanie , Ploegh Hidde , Kim You-Me 
Published: January 2023

Abstract
The nucleotide-sensing toll-like receptors (TLRs) such as TLR3, 7, 9 traffic from the ER to endolysosomes where they recognize their cognate ligands and initiate signaling. A polytopic membrane protein, UNC93B1 delivers the intracellular TLRs to endolysosomes by binding to the transmembrane region of the receptors. In the cells expressing a mutant UNC93B1 incapable of binding the nucleotide-sensing TLRs, the receptors fail to exit the ER and cannot signal. However, the detailed molecular mechanism underlining the UNC93B1-mediated TLR trafficking is not known. Here we show that acidic residues in the juxta-transmembrane region of TLR9 are critical for the interaction with UNC93B1. Mutation of those acidic residues in TLR9 inhibits its binding to UNC93B1 and prevents it from trafficking to endolysosomes without affecting the localization of UNC93B1. Consequently, the TLR9 mutants are unable to rescue CpG-mediated signlaing in the TLR9 knock-out dendritic cells and macrophages. Based on these observations, we are currently trying to develop a method to pharmacologically disrupt the interaction of endogenous TLR9 and Unc93B1. The specific inhibition of the nucleotide-sensing TLRs by means of preventing the interaction with UNC93B1, without affecting other TLRs, may be beneficial for the treatment of autoimmune diseases.
Toll-like receptor 2 ligands enhance monocyte Fcγ receptor function (68.16)
Authors: Shah Prexy , Justiniano Steven , Dunn Robert , Jarjoura David , Byrd John , Butchar Jon , Tridandapani Susheela 
Published: January 2023

Abstract
Fcγ receptors (FcγR) are major effectors of antibody therapy so it is critical to seek new ways to enhance their effectivness. Toll-like receptors (TLR) recognize a wide range of microbial components, eliciting cellular responses such as cytokine production and generation of antimicrobial factors. Among these receptors, TLR2 recognizes both bacterial and fungal components. Given the strong cellular proinflammatory responses to TLR2 activation, we explored the possibility that TLR2 agonists could strengthen FcγR activity. Human peripheral blood monocytes (PBM) were treated with or without the TLR2 agonist PAM2CSK4 and tested for FcγR expression and function. Results showed that overnight treatment with PAM2CSK4 led to an increase in the expression of both FcγRIIa and the common γ-subunit, while expression of the inhibitory receptor FcγRIIb remained unchanged. Along with this, PAM2CSK4 significantly increased IgG-mediated TNFα production in plate-bound assays. We then tested its effect on phagocytosis, using fluorescently-labeled opsonized sheep red blood cells as targets. PAM2CSK4 significantly enhanced the phagocytic ability of PBM. To test the effect of PAM2CSK4 in vivo, we injected it in combination with anti-CD20 antibody and measured B-cell depletion. Results showed that PAM2CSK4 led to greater depletion of splenic B cells. These findings suggest that TLR2 agonists modulate FcγR expression and function and should be investigated as possible adjuvants for antibody therapy.
Cryptococcus neoformans activates the inflammasome and its capsule plays an important role in this process (P1357) (68.9)
Authors: Saavedra Pedro , Capparelli Manuella , Matos Larissa , Zamboni Dario , Casadevall Arturo , Bocca Anamelia , Magalhaes Kelly 
Published: January 2023

Abstract
Cryptococcus neoformans is an encapsulated fungus widely distributed around the globe and capable of causing pulmonary infection and meningitis in immunocompromised individuals. Its capsule is well known for conferring C. neoformans the ability to evade host immune system, leading to its proliferation. Regarding the importance of initial innate immune response to cryptococcal infections, we decided to investigate if C. neoformans is capable of activating the inflammasome as well as the role of its capsule in this pathway. In order to verify whether C. neoformans can trigger inflammasome activation, peritoneal (PMs), bone marrow-derived dendritic cells (BMDCs) or the human monocyte cell line (THP-1) were pretreated or not with caspase-1 and potassium efflux inhibitors and infected with wild type C. neoformans (B3501) or its acapsular mutant (Cap67). Our results showed that acapsular mutant of C. neoformans, but not the wild type, induced IL-1β secretion in both murine and human infected cells. Besides, IL-1β secretion seemed to be dependent of fungal viability as heat-killed fungi elicited lower levels of this cytokine compared the living microorganism. In addition, acapsular C. neoformans triggered caspase-1-dependent and potassium efflux independent inflammasome activation. Taken together, these data suggest that C. neoformans activates the inflammasome and its capsule plays an important role in this process.