bluuebunny/crossref_metadata_2025_split huggingface dataset:
AN2728, a new boron-based topical anti-inflammatory agent, inhibits phosphodiesterase 4 (IRC4P.611)
Authors: Freund Yvonne , Dong Chen , Virtucio-Frates Charlotte , Rock Fernando , Mak Yvonne , Zhou Yasheen , Zane Lee , Jarnagin Kurt 
Published: January 2023

Abstract
The novel boron-containing compound, AN2728, is being investigated in phase 3 trials for atopic dermatitis treatment. It exerts its anti-inflammatory effect by inhibiting phosphodiesterase-4 (PDE4), which catalyzes the breakdown of cAMP to AMP. AN2728 competes with cAMP to inhibit the PDE4B1-catalytic domain with Ki of 173±26 nM; thus AN2728 interacts at the enzyme-active site. The X-ray structure of PDE4B-catalytic domain with AN2728 and its structural relative AN2898 reveals that the boron atom interacts with the bimetal center and occupies a position in the catalytic site similar to that of the cAMP phosphate. AN2728 has good affinity across PDE4 gene products A-D. Its selective affinity for PDE4 is 4-10-fold greater than its affinity for PDE1, 2, 3A, 6, or 7B. It is inactive on PDE3B, 5, 7A1, and 8-11. AN2728 activity increases intracellular cAMP and activates PKA, followed by phosphorylation and negative regulation of various cytokine transcription factors. AN2728 inhibits production of specific cytokines with a pattern similar to that of other established PDE4 inhibitors and distinct from those of a glucocorticoid and calcineurin inhibitor. To investigate reactivity against other targets, AN2728 was tested against a set of 50 receptors and ligand-gated ion channels for inhibition at 10 µM. Inhibition was &lt;25% for all receptors; thus, AN2728 is specific for PDE4 and appears to exert an anti-inflammatory effect through inhibition of inflammatory cytokine production.
Genetic control of isotype specific antibody responses to flagellin in mice (IRC4P.450)
Authors: LU CHUNCHI , Lopez-Yglesias Americo , CHEN HSINHUI , Zhao Xiaodan , VandenBos Tim , Strong Roland , Smith Kelly 
Published: January 2023

Abstract
Adjuvants can improve the efficacy of vaccines by inducing class-switched immune responses. Host factors controlling the magnitude of class-switched antibody responses are poorly understood. We have observed flagellin-immunized mouse strains differ in the magnitude of isotype specific antibody responses. Some strains are high responders (A/J, Swiss Webster) and other strains are low responders (C57BL6, Balb/c). All strains had comparable primary anti-flagellin IgM responses, suggesting common genetic variations influence isotype class switching. We hypothesized different genetic differences between A/J and C57BL/6 mice determine magnitude of responder phenotype. We bred C57BL/6 and A/J mice and tested F1 and F2 offspring for isotype specific antibody responses. Mice were immunized with a FliC protein mutated in the Naip5/6 recognition domain to overcome known differences in Naip5-dependent inflammasome activation between these mouse strains. We demonstrate anti-flagellin IgG1, IgG2a/c and IgA responses are inherited as a quantitative trait. Using consomic mice, we show the high responder phenotype is not controlled by the IgH locus on A/J chromosome 12. Using mutant flagellin proteins, we demonstrate the high responder phenotype is primarily controlled by the FliC D2/D3 domain. Understanding the host factors contributing to FliC D2/D3 induced antibody responses aid in the rationale design of vaccines, and help establish principles for future adjuvant design and development.
Effects of short chain fatty acids and GPR43 stimulation on human Treg function (IRC5P.631)
Authors: Mamontov Polina , Neiman Eric , Cao Tinghua , Perrigoue Jacqueline , Friedman Joshua , Das Anuk , Mora J. 
Published: January 2023

Abstract
SCFA (acetate, propionate, butyrate) are produced when dietary fiber is metabolized by commensal microbiota. In the mouse, SCFA increase colonic Foxp3+ regulatory T cell (Treg) function, promote immune tolerance, and ameliorate intestinal inflammation through the G protein-coupled receptor GPR43 (FFAR2). However, despite the compelling literature in the mouse, it has not been reported whether SCFA can modulate human Treg function. Moreover, whether GPR43 stimulation is sufficient to mediate any effects of SCFA on human Treg remains unknown. Here we show that acetate or butyrate did not increase Foxp3 expression in ex vivo-differentiated human Treg. Nonetheless, consistent with a proposed effect of SCFA on Treg tolerogenic function, butyrate increased IL-10 production by ex vivo-differentiated Treg. Of note, we show that synthetic GPR43-agonists increased the in vitro suppressive capacity of human peripheral blood CD4+/CD25+ Treg in a T cell/DC co-culture system, an effect that correlated with a decreased production of pro-inflammatory cytokines. Thus, pharmacological GPR43-agonists may partially emulate SCFA effects on Treg. Our results show for the first time that SCFA can modulate human Treg function, and that pharmacological GPR43-agonists are sufficient to increase Treg suppressive capacity, with potential therapeutic implications for inflammatory/autoimmune diseases.
Inhibition of phosphorylation of 4E-BP1 by resveratrol suppresses monosodium urate crystal-induced production of interleukin-1β in human monocytes (IRM5P.646)
Authors: Chung Yeon-Ho , Chung Doo Hyun , Lee Won-Woo 
Published: January 2023

Abstract
Monosodium urate (MSU) crystal-induced production of interleukin-1β(IL-1β) from innate immune cells has a critical role for the pathogenesis of gouty inflammation. Recently, it has been reported that resveratrol has a protective effect on gouty arthritis in mouse model. However, the underlying mechanism remains unknown. In this study, we investigated the immunoregulatory role of resveratrol affecting MSU crystal-induced IL-1β production in human CD14+ monocytes. Our data showed that resveratrol inhibited the release of IL-1β by monocyte in response to MSU crystal stimulation and it was closely associated with a decreased level of production of intracellular pro-IL-1β. To investigate the mechanism involved with the negative effect of resveratrol on pro-IL-1β synthesis, we evaluated the phosphorylation status of cap-dependent translation inhibitor 4E-BP1 in MSU crystal-treated monocytes. Resveratrol markedly diminished the phosphorylation of 4E-BP1 in MSU crystal-treated monocytes. Moreover, the autoproteolytic activation of caspase-1 induced by MSU crystal-treatment was also greatly prevented by resveratrol. These results reveal the mechanism underlying how resveratrol prevents MSU crystal-induced production of IL-1β in monocytes and suggest its novel function in inhibition of gouty inflammation.
Rhbdd3 controls innate inflammatory diseases via negatively regulating TLR signaling (IRM5P.647)
Authors: Liu Juan 
Published: January 2023

Abstract
Rhomboid domain-containing protein 3 (Rhbdd3), which belongs to a family of proteins with rhomboid domain, is widely expressed in immune cells; however, the roles of Rhbdd3 in immunity remain unknown. Rhbdd3-deficient mice spontaneously developed autoimmune diseases due to increased IL-6 production, and developed more severe inducible autoinflammatory colitis and acute hepatic injury. Using Rhbdd3-deficient mice, we reveal that Rhbdd3 negatively regulates Toll like receptor ( TLR)-mediated NK cell and DC activation. Rhbdd3 respectively interacts with DAP12 and NEMO to inhibit MAPK and NF-kB signaling activation in TLR-triggered NK cells and DC. Furthermore, Rhbdd3 directly binds to K27-linked polyubiquitin chains on NEMO via its UBA domain, and recruits the deubiquitinase A20 to induce K63-linked deubiquitination of NEMO and thus inhibits activation of the transcription factor NF-kB in DCs. Our study outlines a mechanism for the negative regulation of TLR signaling and autoimmune diseases.
Calcium-dependent PKC is required for chromatin decondensation during the activation of peripheral T cells (IRM6P.651)
Authors: Rawlings Jason , Eikenbusch Sarah , Shields Molly , Bingham Kellie , Lee Megan , Meredith Jenna , Mitchell Taylor 
Published: January 2023

Abstract
Activation of the T cell receptor (TCR) through antigen binding results in chromatin decondensation required for the clonotypic proliferation observed during a proper immune response. Activation of the TCR results in the cleavage of phospholipase biphosphate (PIP2) into diacylglyercol (DAG) and inositol trisphosphate (IP3) by phospholipase C-γ (PLC-γ). Previous studies have shown that both the DAG and IP3 pathways can decondense chromatin, and that intracellular calcium is required for proper TCR-mediated chromatin decondensation. In this study, we used flow cytometry to analyze histone accessibility of chromatin in T cells and found that PKC activation is also required for proper activation-induced chromatin decondensation. Furthermore, we show that calcium is required for proper PKC-mediated chromatin decondensation. Our studies suggest that the PKC isoform(s) required for decondensation may be calcium-dependent. Finally, we show that activation of PKC and not IP3 is required for T cells to gain competence to respond to cytokine stimulation. Taken together, our data suggest that IP3 and PKC signaling work cooperatively to decondense chromatin and regulate clonotypic expansion of T cells during an immune response.
H3K9 trimethylation silences Fas expression to confer colon carcinoma immune escape and chemoresistance (IRM6P.654)
Authors: Paschall Amy , Yang Dafeng , Li Xia , Choi Jeong-Hyeon , Liu Feiyan , Figueroa Mario , Oberlies Nicholas , Pearce Cedric , Bollag Wendy , Nayak-Kapoor Asha , Liu Kebin 
Published: January 2023

Abstract
The Fas-FasL effector mechanism plays a key role in cancer immune surveillance by host T cells, but metastatic human colorectal carcinoma (CRC) often uses silencing of Fas expression as a mechanism of immune evasion. The molecular mechanism of FAS transcriptional silencing in human CRC is unknown. We performed genome-wide ChIP-Sequencing analysis and determined that the level of H3K9me3 in the FAS promoter region is significantly higher in metastatic than in primary human CRC cells, and is inversely correlated with the level of Fas expression. We discovered that verticillin A is a selective inhibitor of histone methyltransferases and inhibits H3K9 trimethylation in human CRC cells. Fas expression is abolished in metastatic human CRC cells and verticillin A treatment effectively restored Fas expression in these metastatic CRC cells. Furthermore, verticillin A exhibited greater efficacy than Decitabine and Vorinostat in overcoming metastatic human CRC resistance to FasL-induced apoptosis. Interestingly, verticillin A also effectively overcomes metastatic human CRC resistance to 5-Fluorouracil in vitro and in vivo. Using an orthotopic CRC mouse model, we demonstrated that tumor-infiltrating cytotoxic T lymphocytes are FasL+ and FasL-mediated cancer immune surveillance is essential for CRC inhibition in vivo. Our findings determine that H3K9 trimethylation of the FAS promoter is a dominant mechanism underlying FAS silencing and resultant CRC immune evasion and metastasis.
Nucleosomal remodeling defects at gene promoters underlie cross-phenotype effects in X-linked thrombocytopenia/Wiskott-Aldrich syndrome (IRM6P.655)
Authors: Sarkar Koustav , Sadhukhan Sanjoy , Han Seong-Su , Vyas Yatin 
Published: January 2023

Abstract
Loss-of-function mutations in Wiskott-Aldrich-Syndrome (WAS) gene associate with either X-linked thrombocytopenia (XLT) or XLT that progresses to immunodeficiency, yet the biological basis for gene-pleiotropy remains unknown. WASp, the protein deficient in WAS, generates F-actin in the cytosol via VCA-domain and supports transcription in the nucleus, the latter by an unknown mechanism. We report that nuclear-WASp, independently of its VCA-domain, is required for the recruitment of BAF47, BAF170, and EP400 to IFNG and TBX21 promoters during TH1-differentiation, but not to IL4 or GATA3 during TH2-differentiation. XLT-causing Thr45Met mutation that progresses to immunodeficiency, impairs recruitment of the above chromatin-remodelers, NF-κB(p65), and STAT1 to TH1-gene promoters, which show paucity of H2A.Z containing nucleosomes, and are therefore sub-optimally transcribed. Dissimilarly, Ala236Gly and Arg477Lys mutations that manifest in XLT without progressing to immunodeficiency do not impair TH1-transcriptional reprogramming. We identify a functional link between nuclear-WASp and SWI/SNF-complexes in T lymphocytes, perturbation of which provides a mechanism for the development of clinical crossphenotypes in human XLT/WAS.
Regulation and function of absent in melanoma 2 in human CD4 T cells (IRM7P.710)
Authors: Komori Heather 
Published: January 2023

Abstract
Recognition of dsDNA by absent in melanoma 2 (AIM2) leads to inflammasome formation, activation and secretion of IL-1β and IL-18 in innate cells. While AIM2 inflammasome function has been well characterized in macrophages, it has not been explored in adaptive cells. Recently, we have shown that human CD4 T cells robustly upregulate expression of AIM2 following T cell activation. Interestingly, memory CD4 T cells have a higher resting level of expression of AIM2 and more rapid upregulation of AIM2 mRNA following activation. High-throughput targeted bisulfite sequencing and ChIPseq show that AIM2 expression in CD4 T cells is epigenetically regulated by DNA methylation and histone modifications. The AIM2 promoter exhibits both hypomethylated CpGs and increased H3K4me3 and H3K4me2 in memory CD4 T cells compared to naïve. Such epigenetic marks allow for both a higher basal level of AIM2 expression and stronger induction of AIM2 expression following T cell activation. Robust protein expression of AIM2 is also found following stimulation of these cells. While AIM2 is known to form inflammasomes and drive IL-1β and IL-18 expression in macrophages, CD4 T cells do not express either of these cytokines, suggesting a non-canonical role for AIM2 function in T cells. Our current studies are focused on determining the function of AIM2 in CD4 T cells, and preliminary work suggests that AIM2 plays a role in the regulation of apoptosis following T cell receptor crosslinking
SLAM-associated protein deficiency in T cells from patients with systemic lupus erythematosus contributes to early signaling abnormalities and reduced IL-2 production (IRM7P.704)
Authors: Karampetsou Maria , Kis-Toth Katalin , Comte Denis , Tsokos George 
Published: January 2023

Abstract
T cells are key players in the pathogenesis of systemic lupus erythematosus (SLE), a chronic autoimmune disease. Following TCR stimulation, SLE T cells display increased levels of tyrosine phosphorylated proteins (pTyr) and augmented calcium (Ca++) responses compared to healthy donors. Impaired IL-2 and excessive IFNγ production also characterize SLE T cells. X-linked lymphoproliferative syndrome (XLP) is an immunodeficiency caused by loss-of-function mutations of the SLAM-associated protein (SAP), a SH2-domain-containing cytoplasmic adaptor molecule. XLP T cells display TCR-mediated early signaling aberrations and a cytokine profile similar to SLE. We determined SAP protein and mRNA levels in SLE T cells and found them significantly decreased. Silencing of SAP in normal T cells resulted in increased pTyr levels and Ca++ flux following aCD3 stimulation. Interestingly, silencing of SAP resulted in decreased IL-2 production. Forced expression of SAP in SLE T cells restored pTyr levels and Ca++ responses to normal levels while the production of IL-2 was corrected to normal levels. In conclusion, SLE T cells display reduced levels of the protein adaptor SAP and that restoration of SAP protein levels corrects the early signaling transduction defects and restores IL-2 production. Approaches to restore SAP levels in SLE T cells may prove of therapeutic value.
TRAF3 regulates CSK and PTPN22 to enhance T cell receptor signaling (IRM7P.705)
Authors: Wallis Alicia , Yi Zuoan , Bishop Gail 
Published: January 2023

Abstract
TCR signaling is critical for T cell activation, so further elucidation of this process is necessary to understanding T cell biology, and to manipulate T cell activation in clinical settings. Extensive research on TCR signaling has revealed many details, but additional information on the molecular mechanisms continues to be revealed. Our group previously produced a T cell-conditional (CD4-Cre) TRAF3-/- mouse, which demonstrated that TRAF3 plays an important positive role in TCR signaling and T cell functions. After TCR engagement, TRAF3 associates with the TCR complex, and TRAF3-/- cells display a marked reduction in the early phosphorylation of Lck and CD3-ζ activation sites. This impact of TRAF3 on proximal TCR signaling events led us to hypothesize that TRAF3 regulates CSK, a T cell membrane-localized protein that binds to and inhibits Src family kinases, such as Lck. Through immunoprecipitation, TRAF3 was shown to interact with CSK upon TCR stimulation. The interaction between TRAF3 and CSK occurs in both the membrane and cytoplasmic fractions. In the absence of TRAF3, elevated amounts of CSK remain in the plasma membrane of stimulated T cells. Another TCR inhibitor, the protein tyrosine phosphatase PTPN22, also interacts with the TRAF3:CSK complex. Similar to its impact on CSK, loss of TRAF3 results in elevated levels of PTPN22 in the plasma membrane. These data identify a new role for TRAF3 in regulating the localization of the TCR signaling inhibitors CSK and PTPN22.
A single amino acid residue of CDR3β alters the antigenicity and autoreactivity of human invariant natural killer T cell receptors (IRM7P.707)
Authors: Guo Tingxi , Chamoto Kenji , Hirano Naoto 
Published: January 2023

Abstract
Detection of human invariant natural killer T (iNKT) cells relies on staining for their T cell receptor (TCR), invariant Vα24 TCRα chain paired with semi-variant Vβ11 TCRβ chain, or α-GalCer-CD1d tetramer. It has been assumed that all Vα24+ iNKT cells are stained with similar intensity by anti-Vα24 mAbs, clones C15 and 6B11, regardless of the hypervariable Vβ11 CDR3β sequences. By analyzing a large panel of autoreactive human iNKT TCRs, we found that TCRs encoding CASR at the start of CDR3β were stained with significantly lower intensity by C15 than those encoding CASS. Substitution of S to R at the fourth CDR3β residue was sufficient to diminish staining intensity by anti-Vα24 mAbs. Interestingly, the autoreactivity was also reduced in the TCRs encoding CASR. Importantly, α-GalCer tetramer positive population of human primary T cells presented low and high intensity subsets when stained with anti-Vα24 mAbs. Vβ11 TCRβ chains were cloned from each sorted population, and the Vα24 low population contained a significantly higher frequency of Vβ11 TCRs encoding CASR compared to the Vα24 high population. Therefore, human iNKT TCRs encoding CASR may falsely be negative when stained with anti-Vα24 mAbs, which underestimate the frequency of bona fide Vα24+ iNKT cells. Our data demonstrate that the fourth CDR3β amino acid residue affects iNKT TCR autoreactivity and binding of anti-Vα24 mAbs, suggesting that the CDR3β sequence alters the structural conformation of the Vα24 TCRα chain.
Dendritic cells coordinate the development and homeostasis of organ-specific regulatory T cells (LYM2P.730)
Authors: Leventhal Daniel , Nishi Saki , Gilmore Dana , Malchow Sven , Kline Douglas , Kline Justin , Savage Peter 
Published: January 2023

Abstract
Foxp3+ regulatory T cells (Tregs) are critical for the maintenance of tolerance to self. However, the antigen presenting cells (APCs) controlling the thymic development and peripheral homeostasis of these cells have yet to be clearly defined. Previously, we identified a monoclonal population of prostate-specific Tregs, termed MJ23 Tregs, that require Autoimmune Regulator (Aire) for their thymic development. Using a variety of mixed bone marrow chimera experiments, we sought to determine the contributions of various thymic APC subsets towards MJ23 Treg development. We demonstrate that the thymic development of MJ23 Tregs requires both MHC-II and B7.1/.2 on bone-marrow derived cells. Moreover, development is abolished by the depletion of CD11c-expressing cells. Together, these data indicate that the antigen recognized by MJ23 Tregs in the thymus must be transferred from Aire-expressing medullary thymic epithelial cells to CD11c-expressing dendritic cells to coordinate MJ23 Treg development. In the periphery, we demonstrate that the depletion of CD11c-expressing cells eliminates the enrichment of MJ23 Tregs in the prostate-draining lymph nodes, demonstrating that dendritic cells also coordinate the anatomical distribution of organ-specific Tregs in the secondary lymphoid organs. In all, our results reveal a critical role for dendritic cells in orchestrating the development and homeostasis of Aire-dependent, organ-specific Treg cells.
Heterochronic parabiosis: allowing the dissection of the aged immune system (LYM2P.723)
Authors: Davies John , Pulko Vesna , Thompson Heather , Nikolich-Zugich Janko 
Published: January 2023

Abstract
Parabiosis is the surgical union of two organisms resulting in the development of a single, shared circulatory system. When animals of different ages are conjoined (i.e. heterochronic parabiosis), blood-borne factors from the younger animal can often beneficially affect the older animal and recapitulate the youthful phenotype & function in target tissues. However, the effects of heterochronic parabiosis on the aged immune system remain unexplored. An important question to be answered is whether the cellular defects involved in the aged immune system are due to intrinsic deficiencies or if they can be rescued by extrinsic factors. Also, T cell ontogenesis is known to diminish with age due to thymic involution as well as defects in lymphopoiesis that affect pre-T cells. The heterochronic parabiosis model allows testing of contributions of the aged thymus, pre-T cells and of systemic environment in immune aging. The preliminary data demonstrate differential migration patterns of T cells, B cells & dendritic cells as a function of age. Of interest, pre-T cells from either animal adequately seed the opposing thymus and undergo thymopoiesis and other data suggest that there are no intrinsic defects in old thymocytes, but rather of the old thymic environment. That latter defect may not be corrected by heterochronic parabiosis.
Homeostasis and function of Tregs at the site of chronic inflammation (LYM2P.728)
Authors: Spence Allyson , Kim Youmin , Tang Qizhi 
Published: January 2023

Abstract
This study is aimed at characterizing Treg dynamics at the site of chronic inflammation in the islets of NOD mice. Tregs expressing CD103 were present primarily in the inflamed islets and low in the pancreatic LN and spleen. Both CD103+ and CD103- Tregs from inflamed NOD islets could prevent diabetes when transferred to Treg-deficient CD28KO mice. CD103 was selectively induced and maintained in inflamed islets after transfer. However, fewer CD103+ Tregs were recovered after transfer compared with CD103- Tregs, suggesting that CD103+ Tregs have a survival defect. Indeed, CD103+ Tregs had lower Bcl2 expression despite higher CD25 expression and higher responsiveness to IL-2. CD103+ islet Tregs have reduced TCRb repertoire diversity and higher BrdU incorporation, indicating they clonally expand in the islets. Lastly, in NOD mice expressing a TCR signaling reporter, the Nur77-GFP transgene, CD103+ islet Tregs were found to be uniformly Nur77hi suggesting they were more recently activated by antigens. TCRb sequences used by high frequency clones of CD103+ Tregs could be found in Nur77hiCD103- but not Nur77loCD103- Tregs, consistent with sequential TCR activation and CD103 expression. Together, our results demonstrate that chronically inflamed islets in NOD mice contain functional Tregs, but fail to maintain the highly antigen-specific activated subset. Maintenance of this subset may have important implications for the treatment of chronic inflammatory disease.