bluuebunny/crossref_metadata_2025_split huggingface dataset:
Differential effects of cyclosporine A and TGFβ1 on IgE- versus IL-33-mediated mast cell activation (HYP3P.407)
Authors: Chernushevich Oksana , Ndaw Victor , Ryan John 
Published: January 2023

Abstract
Cyclosporine A (CsA), a potent calcineurin antagonist and immunosuppressant, is known to inhibit IgE-mediated mast cell activation. The present research investigates the role of this drug in suppressing IL-33-mediated mast cell activation. IL-33 is a novel cytokine of IL-1 family that elicits mast cells to produce inflammatory cytokines independently of IgE-FcεRI signals. We have found that low concentrations of CsA (10nM) are capable of suppressing IgE-mediated cytokine production by 50-80% in mouse bone marrow-derived mast cells. By comparison, IL-33-mediated secretion of TNF, IL-6, and MCP-1 was unaffected by CsA under these conditions. These results were consistent among mast cells derived from C57BL/6, 129Sv, C3H, and A/J mouse strains. We have previously reported that TGFβ1 inhibits IgE-mediated cytokine production. Our current experiments showed that TGFβ1 similarly antagonizes IL-33-induced cytokine secretion, and that CsA does not enhance this effect. These data demonstrate that mast cell activation by IL-33, thought to be a key regulator of innate immunity, is responsive to TGFβ1-mediated suppression, but relatively resistant to CsA effects. Understanding how mast cells are regulated by innate signals will shed light on many inflammatory diseases.
Dexamethasone suppresses IL-33-mediated mast cell activation (HYP3P.409)
Authors: Paranjape Anuya , Chernushevich Oksana , Ryan John 
Published: January 2023

Abstract
Dexamethasone, a potent synthetic member of the glucocorticoid class of steroid drugs is known for its immunosuppressant and anti-inflammatory effects. The present research investigates the role of this anti-allergic drug in suppressing IL-33-mediated mast cell activation. IL-33 is a novel cytokine of IL-1 family, which promotes mast cells to produce inflammatory cytokines independently of IgE-FcεRI signals. Initial studies have shown that micromolar quantities of dexamethasone suppress the production of TNF-α and IL-6 by IL-33-stimulated mast cells. While confirming these results by intracellular staining, it was observed that dexamethasone not only decreases the fraction of cytokine-positive cells but also causes a drop in cytokine production per cell. Although dexamethasone pretreatment resulted in a significant reduction in the IL-33 receptor subunit T1/ST2, this did not correlate with decreased cytokine production. These results demonstrate the ability of dexamethasone to suppress mast cell activation during innate immune responses, and support further work into the mechanisms by which this occurs.
Mir-34a regulates macrophage polarization by targeting IL-4Ralpa/STAT6 axis (INM2P.423)
Authors: Tundup Smanla , Srivastava Leena , Chandora Kapil , Harn Donald 
Published: January 2023

Abstract
Macrophages perform diverse functions during infection as well as metabolic diseases. M1 macrophages promote inflammation while M2 macrophages suppress inflammation and drive anti-inflammatory responses. Emerging evidence suggests that microRNAs control numerous cellular processes; however their role in controlling M2 macrophage maturation is not well understood. To test if microRNAs play roles in macrophage M2 polarization, we performed microarray analysis of microRNA expression in mouse M2 macrophages generated under both in-vivo (i.p injection of Schistosome eggs or LNFPIII glycan) and in-vitro (incubation with recombinant IL-4) conditions. Among the microRNAs differentially expressed, we found that expression of mir-34a was significantly upregulated in M2 macrophages and that over expression of miR-34a mimics led to decreased M2 maturation measured as Fizz1 expression. Using reporter assays and western blot analysis we found that miR-34a targets 3' UTR of stat6 gene thereby affecting the IL-4Rα-STAT6 signaling cascade. Our findings show a unique role of mir-34a in regulating macrophage polarization via targeting the IL-4Rα-STAT6 axis, suggesting that mir-34a plays an important role in regulation of inflammation and may have therapeutic function for inflammatory and metabolic diseases.
Enhanced phagocytosis of auto antibodies-opsonized blood cells by macrophages from virally infected mice. (INM3P.440)
Authors: Legrain Sarah , Su Dan , Detalle Laurent , Izui Shozo , Verbeek Sjef , Coutelier Jean-Paul 
Published: January 2023

Abstract
Autoimmune haemolytic anemia and idiopathic thrombocytopenic purpura are common immune disorders resulting from the phagocytosis of autoantibodies-opsonized blood cells. It has been reported that viral infection may be associated with autoimmune haemolytic anemia and idiopathic thrombocytopenic purpura in patients and may incriminate IgG and IgM antibody isotypes. Lactate Dehydrogenase-elevating Virus triggers the activation of immune cells although it does not induce usually any pathology in mice. Here we report that LDV infection increases the pathogenicity of IgM anti-platelet mAbs which is mediated by FcαμR. Interestingly, we show that this mechanism results from an enhanced Fc-dependent phagocytosis by macrophages rather than an increased expression of FcαμR. Our results also indicate that the pathogenicity of IgG anti-red blood cells is increased in virally infected mice. To date, three activating FcγR have been identified in mice, binding IgGs with different affinity : FcγRI, FcγRIII and FcγRIV. Since FcγRI has a high affinity for monomeric IgG2a, this receptor is poorly available for the binding of circulating-opsonized IgG2a. However we report an increased expression of each receptor by macrophages in contrast to FcamR whose expression is not regulated. Furthermore, we show the functional involment of these three receptors in the exacerbation of IgG-mediated anemia.
Inflammatory protease, caspase-1, is secreted from monocytes in a stably active form in response to inflammasome activation (INM3P.441)
Authors: Shamaa Obada , Ghonime Mohammed , Srabani Das , Gavrilin Mikhail , Wewers Mark 
Published: January 2023

Abstract
Caspase-1 is an inflammatory protease which is required for activation of the pro-inflammatory cytokines, interleukin-1β (IL-1β) and IL-18, in addition to a role in a form of cell death termed pyroptosis. Caspase-1 is synthesized as a zymogen which requires the assembly of a multi-protein complex, termed the inflammasome, for its activation. This assembly forms in response to pathogen or danger associated molecular patterns, but where and how this assembly happens in the cell is poorly understood. To address this question we compared the activity of the released caspase-1 to the cytosolic form in the human monocyte cell line, THP1. In response to endotoxin (LPS) and ATP, pro-caspase-1 is rapidly processed and released from monocytes in its mature, p20/p10, form. We found that the released form has activity which is stable for over 12h and this activity is inhibited by the caspase-1 specific inhibitor, YVAD-cmk. Interestingly, we were unable to completely immunodeplete the released caspase-1 activity or show its ability to cleave exogenously added proIL-1β. This contrasts to the in-vitro cell-extract model, where concentrating monocytic lysates spontaneously activates caspase-1. This cytosolic caspase-1 cleaves endogenous proIL-18, but rapidly loses function. These results suggest that caspase-1 activation requires multi-level regulation that links assembly of the inflammasome complex and enzymatic activity to its interaction with its target substrates during release.
Scaffold functions of 14-3-3 adaptors in B cell immunoglobulin class switch DNA recombination (IRC3P.472)
Authors: Lam Tonika , Thomas Lisa , White Clayton , Li Guideng , Pone Egest , Xu Zhenming , Casali Paolo 
Published: January 2023

Abstract
Immunoglobulin (Ig) class switch DNA recombination (CSR) crucially diversifies antibody biological effector functions. CSR involves the induction of activation-induced cytidine deaminase (AID) expression and targeting to switch (S) regions by 14-3-3 adaptors. We hypothesized that 14-3-3 adaptors function as scaffolds to stabilize CSR elements on S regions. Here, we demonstrate that all seven 14-3-3 adaptors (β, ε, γ, η, σ, τ and ζ), directly interacted with AID, PKA-Cα (catalytic subunit) and PKA-RIα (regulatory inhibitory subunit) and uracil DNA glycosylase (Ung). 14-3-3 adaptors, however, did not interact with AIDΔ(180-198) C-terminal truncation-mutant or AIDF193A and AIDL196A point-mutants (which have been shown not to bind to S region DNA and fail to mediate CSR). 14-3-3 adaptors colocalized with AID and replication protein A (RPA) in B cells undergoing CSR. 14-3-3 and AID binding to S region DNA was disrupted by HIV-1 viral protein R (Vpr), which inhibited CSR without altering AID expression. Accordingly, we demonstrate that 14-3-3 directly interacted with Vpr, which in turn, also interact with AID, PKA-Cα and Ung. Thus, we describe that 14-3-3 adaptors play important scaffold functions and nucleate the assembly of CSR factors on S regions. Such assembly can be disrupted by a viral protein, thereby allowing us to hypothesize that small molecule compounds that specifically block 14-3-3 interactions with AID, PKA and/or Ung can be used to inhibit unwanted CSR.
Mechanism of B cell membrane repair and the effects of repair on B cell receptor activation (IRC3P.460)
Authors: Miller Heather , Tam Christina , Corrotte Matthias , Castro-Gomes Thiago , Andrews Norma , Song Wenxia 
Published: January 2023

Abstract
The plasma membrane of B cells is at risk for injury by bacterial toxins, viruses, and possible mechanical damage induced by interactions with other cells during antigen presentation. Whether and how B cells repair membrane damage and its impact on B-cell functions are unknown. Here we show that upon injury by the pore forming toxin streptolysin O (SLO), primary mouse splenic B cells can repair their membrane in a Ca2+-dependent manner, like muscle and epithelial cells. B cells exocytose their lysosomes in response to Ca2+ influx and release acid sphingomyelinase from lysosomes, which induces endocytosis. In B cells activated by cross-linking B-cell receptors (BCRs), repairing membrane damage by SLO inhibits BCR clustering and internalization, consequently reducing BCR-induced tyrosine phosphorylation. Further analyses show that membrane repair inhibits the colocalization of BCRs with lipid rafts on the cell surface and during their internalization, indicating that membrane repair-induced endocytosis competes with BCRs for lipid rafts and interferes with the interaction of BCRs with lipid raft-associated kinases. Our results reveal the ability and the mechanism for B cells to repair membrane damage, and discover a negative regulatory role of membrane repair on B cell signaling and antigen uptake functions.
Integrin β8 is a marker of mouse regulatory T-cells (Treg) and is required for processing of latent TGF-β1 from the GARP/latent TGF-β1 complex. (IRC4P.483)
Authors: Edwards Justin , Thornton Angela , Shevach Ethan 
Published: January 2023

Abstract
Previous studies have demonstrated that activated Tregs express latent TGF-β1 on their cell surface bound to the tethering molecule GARP. Although integrins have been implicated in mediating the release of biologically active TGF-β1 from the complex of latent TGF-β1 and latent TGF-β1 binding protein, their role in processing latent TGF-β1 from the latent TGF-β1/GARP complex is unclear. We detected integrin β8 expression by CD4+Foxp3+ Treg, but not by CD4+Foxp3- T cells, by qRT-PCR. Furthermore, integrin β8 expression appeared to be a marker of thymically-derived Treg (tTreg), as its could not be detected on Foxp3+ T cells induced in vitro nor in vivo on peripherally induced Foxp3+Helios- Tregs. While Tregs from integrin β8 conditional knockouts suppressed T effector cell proliferation normally in vitro and in vivo in a model of colitis, integrin β8 deficient Tregs failed to provide a source of TGF-β1 to drive Th17 or iTreg differentiation in vitro. In addition, the knockout Tregs expressed higher levels of latent TGF-β1 on their cell surface consistent with defective processing. Taken together, integrin β8 is not only a marker of tTregs, but also functions in a cell intrinsic manner as the major pathway for processing latent TGF-β1 from the latent TGF-β1/GARP complex on the surface tTregs.
Type I IFNs modulate T effector (Teff) and T regulatory (Treg) cell function during experimental autoimmune encephalomyelitis (EAE) (IRC4P.485)
Authors: Metidji Amina , Shevach Ethan 
Published: January 2023

Abstract
Type I IFNs (IFNα and IFNβ) are a family of cytokines that mediate multiple roles in inflammation and autoimmunity. Previous studies have demonstrated that Type I IFNs primarily act on myeloid cells during the course of EAE. We have re-examined the role of Type I IFNs in EAE by generating mixed bone marrow chimeras between wild type (WT) mice and mice with a deficiency in the IFN receptor (IFNAR-/-). IFN plays a role in the T lymphocyte homeostasis as the ratio of WT to IFNAR-/- CD4+ T cells is skewed toward WT and the skewing is more marked in the Treg compartment. Surprisingly, after induction of EAE, the ratio of WT to IFNAR-/- T cells is reversed in the periphery and the mixed chimeras develop more severe disease than the WT/WT chimeras. Almost all of the CD4+Foxp3- and CD4+Foxp3+ T cells in the CNS of chimeric mice are derived from the IFNAR-/- donor, but the percentage of Foxp3+ T cells was similar in WT/WT and WT/IFNAR-/- chimeras. Both CD4+Foxp3- and CD4+Foxp3+ T cells from the IFNAR-/- donor expressed higher levels of α4 integrins, which may account for their increased numbers in the CNS during EAE. Mice with a conditional deletion of the IFNAR in Treg rapidly developed a very severe form of EAE. Taken together, these results demonstrate that signaling via the IFNAR is required for Treg suppressor function in EAE and that IFNAR signaling may also modulate homing of Teff cells to the CNS and/or their susceptibility to suppression by Treg.
PCAF promotes TGF-β-induced Foxp3+ regulatory T cell production (IRC4P.486)
Authors: Liu Yujie , Wang Liqing , Han Rongxiang , Beier Ulf , Dent Sharon , Hancock Wayne 
Published: January 2023

Abstract
We recently showed that 2 histone/protein acetyltransferases (HATs), p300 and CBP, are important for Foxp3+ Treg function, and that their targeting promotes anti-tumor immunity in immunocompetent mice. We now report on functions of another HAT, PCAF, in Tregs. Like CBPflflp300flflFoxp3YFP-cre mice, PCAF-/-GCN5flflFoxp3YFP-cre mice develop severe autoimmunity and die in early life, indicating their importance for Treg development. While mice with a single deletion (PCAF-/- or GCN5flflFoxp3YFP-cre) are born at expected Mendelian ratios and do not develop obvious autoimmunity, T cells from PCAF-/- mice are activated, with more CD4+CD44hiCD62Llow, CD4+CD69+ and CD8+IFN-γ+ cells than WT mice. PCAF-/- mice have normal CD4+Foxp3+ percentages in thymus and peripheral lymphoid tissues, but these undergo increased apoptosis upon CD3/CD28 stimulation. Furthermore, compared to control naïve CD4+ T cells (CD4+CD25-CD44lowCD62Lhigh), PCAF naïve CD4 T cells show impaired conversion into Foxp3+ Tregs when cultured with TGF-β in vitro. Adoptive transfer of naïve PCAF-/- vs. WT CD4 cells into Rag1-/- mice also resulted in fewer PCAF-/- CD4 T cells converting into Foxp3+ Tregs than with WT controls, and was accompanied by lower pSmad2 levels in PCAF-/- CD4 T cells. Hence, PCAF regulates iTreg production in vivo and in vitro. Our studies suggest that PCAF is a promising target for development of pharmacologic inhibitors to modulate iTreg production and promote anti-tumor responses.
R-Ras promotes autoimmunity through negative regulation of natural T regulatory cell numbers by inhibiting tolerogenic dendritic cell development (IRC4P.488)
Authors: Ray Avijit , Basu Sreemanti , Chan Andrew , Dittel Bonnie 
Published: January 2023

Abstract
R-Ras is a member of the Ras family of small GTPases, which regulate various cellular processes including adhesion, survival, proliferation, trafficking, and cytokine production. R-Ras is expressed by immune cells and has been shown to modulate dendritic cell (DC) function in vitro. Although, higher expression of R-Ras is associated with liver autoimmunity in humans, its pathogenic mechanism is unknown. We used Rras-/- mice to identify the mechanism whereby R-Ras contributes to autoimmunity using experimental autoimmune encephalomyelitis (EAE), a mouse model of the CNS autoimmune disease multiple sclerosis. We found that a lack of R-Ras in peripheral immune cells resulted in attenuated EAE disease. In accordance with reduced EAE disease, numbers of inflammatory immune cells in the CNS of Rras-/- mice at the peak of EAE were significantly reduced. Further investigation revealed that during EAE, absence of R-Ras promoted the formation of MHC IIlo tolerogenic DC concomitant with a significant increase in Neuropilin-1hiCD4+Foxp3+ natural T regulatory cell (nTreg) numbers in the periphery. Mechanistically, we found that Rras-/- tolerogenic DCs induced proliferation of nTregs thereby enhancing their numbers and maintaining them in vivo. Our study suggests a novel role for R-Ras in promoting autoimmunity through negative regulation of nTreg numbers by inhibiting tolerogenic DC development.
WRNIP1 is a key regulator of antigen-specific Treg cells. (IRC4P.480)
Authors: Du Weiting , Wang Ding , Shen Yueh-Wei , Lee Wen-Hui , Riggs Art , Liu Chih-Pin 
Published: January 2023

Abstract
It is well documented that, both Foxp3+ regulatory T cells (Treg) and antigen-expanded Foxp3lo/- Treg play critical roles in regulating immunity and autoimmunity, as well as in preventing autoimmune diseases. However, the molecular mechanisms and genes responsible for modulating function of these Treg still remain incompletely understood. WRNIP1 (Werner helicase-interacting protein 1) was identified as a protein that interacts with WRN, the Werner syndrome responsible gene product, and may participate in DNA synthesis, damage and repair. Here, we report an unexpected finding that WRNIP1 plays an important role in regulating the function of autoantigen-expanded Foxp3lo/- Treg. Down regulation of WRNIP1 in these Treg significantly increased their proliferation and reduced activation-induced cell death. More importantly, WRNIP1-downregulated Treg showed markedly enhanced suppressive function in vitro, partly due to an increased production of IL-10 and TGF-β. These Treg also significantly improved their ability in preventing diabetes development in NOD mice. To understand how WRNIP1 regulates Treg function, we found that elevated Foxo1 expression may contribute to the enhanced Treg function through promoting IL-10 and TGF-β production. These studies demonstrate that WRNIP1 negatively regulates Treg function and suggest previously undisclosed molecular pathways and targets for improving Treg-mediated immune tolerance to treat autoimmune diseases like type 1 diabetes.
Chronic viral infection: the perfidious role of IL-2 and IL-15 on CD8 T cell exhaustion (IRM4P.500)
Authors: Beltra Jean-Christophe , Charpentier Tania , Michaud Eva , Lamarre Alain , Decaluwe Hélène 
Published: January 2023

Abstract
CD8 T cell exhaustion is a major immuno-regulatory mechanism by which chronic viruses escape the immune response. However, little is known on the factors regulating the unresponsiveness of CD8 T cells is this context. Here, we investigated the combined role of IL-2 and IL-15, two common gamma chain-dependent cytokines, on CD8 T cell differentiation and exhaustion during a chronic viral infection. We performed adoptive transfer of WT or IL-2RβKO P14 CD8 T cells (unresponsive to both IL-2 and IL-15 signals) in mice challenged with LCMV clone 13. Initially, IL-2Rβ-signals were critical for short-lived effector cell differentiation. IL-2 and IL-15 conditioned optimal proliferation of early effector CD8 T cells and promoted their cytotoxic functions. Interestingly, over the chronic phase, IL-2Rβ-signals dramatically worsened CD8 T cell exhaustion. IL-2 and IL-15 sustained the expression of several inhibitory receptors and fostered the development of the highly exhausted PD-1hi progeny population. Finally, IL-2Rβ-signals precluded central memory CD8 T cell differentiation capable of homeostatic proliferation and robust secondary expansion. Altogether, these data demonstrated for the first time an important deleterious effect of IL-2 and IL-15 on the CD8 T cell response to a chronic viral infection. Reconsidering their role in this context may provide new insights for therapeutic regimens against chronic viruses.
Pregnane X receptor negatively regulates inflammation by directly modulating the Toll-like receptor 4 pathway (IRM5P.701)
Authors: Qiu Zhijuan , Cicek Basak , Cervantes Jorge , Salazar Juan , Mani Sridhar , Khanna Kamal 
Published: January 2023

Abstract
Pregnane X receptor (PXR) is a nuclear receptor that plays a critical role in regulating the metabolism of xenobiotics. Here we report a new role for PXR as a negative regulator of the inflammatory pathway. Quiescent Pxr-/- BMDMs exhibited a pro-inflammatory state as transcripts of inflammatory genes were significantly elevated. Upon stimulation with Listeria monocytogenes (Lm), Pxr-/- BMDMs produced significantly higher protein levels of inflammatory cytokines. Moreover, enhanced inflammation was also observed in Pxr-/- mice challenged with Lm. Furthermore, disruption of the toll-like receptor 4 (TLR4) gene reversed the pro-inflammatory state in Pxr-/- BMDMs suggesting that PXR negatively regulated inflammation through the TLR4 pathway. In fact, both Pxr-/-Tlr4-/- BMDMs and mice exhibited normal inflammation in response to Lm infection. Transfection of HEK-Blue™-mTLR4 cells with DNA vector expressing PXR inhibited the response to LPS in a dose-dependent manner, which suggested that PXR can directly modulate the TLR4 pathway. In conclusion, our study demonstrates that PXR negatively regulates the inflammatory pathway and the regulation is through directly modulating the TLR4 pathway. Thus, our results suggest that PXR can be targeted as a therapeutic strategy to treat inflammatory diseases.
Differential phosphorylation of T-bet in the presence and absence of mTORC1 signaling (IRM6P.722)
Authors: Chornoguz Olesya , Powell Jonathan 
Published: January 2023

Abstract
Previously we have demonstrated that T cells lacking the mTORC1 activator Rheb fail to differentiate in Th1 cells upon appropriate skewing conditions. T-box transcription factor T-bet plays a critical role in Th1 differentiation. We wondered whether mTORC1 regulates either T-bet expression or activity. Interestingly, T-bet contains several motifs of sites typically phosphorylated by AGC kinases. This raised the possibility that mTORC1 activation might play a role in the phosphorylation and control of T-bet function. To test this hypothesis we employed targeted mass-spectrometry proteomic analysis (MRM-MS) to identify novel T-bet phosphorylation sites that might be controlled by mTORC1. By analyzing activated WT and Rheb-/- T cells as well as T cells stimulated in the presence of rapamycin we were able to detect 6 T-bet phosphorylation sites. Specifically S52, T54, S224 are novel and Y77, Y118, S508 are previously published phosphorylations. T54, Y77 and S508 were either absent or downregulated in Rheb deficient T-cells and T-cells treated with Rapamycin suggesting that they are controlled by mTORC1 signaling. Studies are underway to determine the precise functional importance of these sites. Thus using MRM-MS we have identified a potentially novel mechanism by which mTORC1 signaling controls T-bet.