bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mast cell responsiveness to IL-10 is altered by genetic background (151.14)
Authors: Kolawole Motunrayo , Ryan John 
Published: January 2023

Abstract
Mast cell activation has been shown to be antagonized by the inhibitory cytokine IL-10. We have found that there are strain differences in IL-10 responsiveness, as BALB/c mast cell progenitors were resistant to IL-10-mediated apoptosis, while C57BL/6 mast cell progenitors were killed by IL-10. We now report that IL-10-mediated effects on the high affinity IgE receptor vary with genetic background. IL-10 enhanced FceRI expression on mast cells derived from Th2-prone BALB/c or 129/Sv mice, but not on C57BL/6 mast cells. We also found that IL-10 surprisingly enhanced IgE-mediated production of inflammatory cytokines MIP-1 alpha and IL-13 even on C57BL/6 mast cells. These data illustrate that cytokine-mediated immunoregulation may be altered by genetic background as well as the inflammatory factors being studied.
Mast cell deficient mice display a pulmonary Th2 polarization with an increase in alternatively activated alveolar macrophages (151.17)
Authors: Wang Xiaojia , Hilderbrand Susana , Brown Jared 
Published: January 2023

Abstract
Macrophages are innate immune cells capable of coordination with the adaptive immune system during an immune response. The classical activation of macrophages is induced by IFN-γ, while the alternative activation of macrophages is triggered by IL-4 and IL-13. Mast cells are well recognized for their role in allergic inflammation, asthma and innate immunity. Due to a membrane tyrosine kinase mutation in c-Kit, B6.Cg-KitW-sh mice are unable to respond to stem cell factor and therefore do not develop mast cells. Thus, B6.Cg-KitW-sh mice do not develop IgE -dependent passive cutaneous anaphylaxis in the ear and joint as well as passive anaphylaxis. In the current study, we show that B6.Cg-KitW-sh mice display a Th2 phenotype with elevated levels of IL-4, IL-5 and IL-13 in serum and decreased IFN-γ levels in lung BAL fluid. This data led us to investigate whether alveolar macrophage phenotype is altered due to mast cell deficiency. We found an increase in the expression of genes associated with alternative activation of alveolar macrophages that included arginase-1 (Arg1), chitinases (Chi313), and FIZZ1 (Retnla) in BAL cells, when compared to background control mice (C57BL/6). In addition, these alternative activated macrophages display a &gt; 25 fold change in class B scavenger receptor (Scarb2) expression. These data begin to suggest that mast cells may be important in regulation of alveolar macrophage phenotype.
Developing a passive sensitization procedure for basophils to diagnose IgE-mediated drug hypersensitivity (151.19)
Authors: Gentinetta Thomas , Hausmann Oliver , Kopac Peter , Colagiovanni Amira , Pichler Werner J. 
Published: January 2023

Abstract
Introduction: The basophil activation test is quite well established for protein allergens. It is normally done with the patients’ basophils and requires fresh cells and rapid processing. It would be a particularly interesting test for drug allergy, as only very few drug specific in vitro IgE assays exist and it seems to be a highly relevant read out system. Objective: To set up a passive sensitization protocol to test IgE-mediated drug hypersensitivity. Methods: PMBCs of basophil donors are incubated with lactic acid (pH=3.9) to remove surface bound IgE. Afterwards, re-sensitization with patient serum and subsequent stimulation with serial dilutions of drugs. Degranulation can be measured by flow cytometry (anti-CCR3-APC/anti-CD63-PE). Results: We were able to establish a well functional procedure for protein allergens and for some drug allergens (cephalosporines, NMBAs). Passive sensitization assays using serum of patients with well-documented IgE-mediated amoxicillin allergy were reproducibly false negative. Conclusion: Using defined donor basophils for passive sensitizations with serum would have several advantages over direct assays, as it would facilitate diagnosis, as patient’s sera is easy to store and as it would provide a direct proof of an IgE-mediated hypersensitivity reaction. However, a better understanding of how small drugs are actually able to cross-link cell surface bound IgE seems to be a prerequisite to improve this assay and to widen its application.
Mast cells induce a novel pro-inflammatory function of myeloid-derived suppressor cells (151.2)
Authors: Morales Johanna , Faber Travis , Saunders Bryan , Graham Laura , Bear Harry , Lantz Chris , Ryan John 
Published: January 2023

Abstract
Mast cells and myeloid-derived suppressor cells (MDSC) are both increased at sites of growing tumors. Like mast cells, MDSC have recently been suggested to function in parasitic or bacterial infection. We have tested the effects of mast cell-MDSC interactions in vitro and in a mouse model of parasitic infection. Surprisingly, co-culture of these cells resulted in a drastic increase in pro-inflammatory mediator release, which was attributed entirely to MDSC. MDSC depletion from mice infected with Nippostrongylus brasiliensis resulted in delayed worm expulsion and a drastically increased intestinal worm burden reminiscent of mast cell-deficient mice. Mast cells can therefore reprogram MDSC to a pro-inflammatory phenotype, suggesting that the evolutionary purpose of these suppressive cells may be to potentiate inflammatory Th2 responses necessary for parasite expulsion.
MTORC1 and mTORC2 control normal and neoplastic mast cell homoeostasis through regulation of cell division rather than maintenance of survival (151.20)
Authors: Smrz Daniel , Kim Mi-Sun , Silhankova Sarka , Wilson Todd , Metcalfe Dean , Gilfillan Alasdair 
Published: January 2023

Abstract
Mature mast cells are terminally differentiated and non-dividing. However, a pathologic increase in mast cells is associated with the myeloproliferative disorder mastocytosis which has been linked to expression of an activating mutation (D816V) in the mast cell growth factor receptor, KIT. We found that in primary human mast cells developed from peripheral blood CD34+ progenitor cells, KIT activation led to activation of mTORC1 and mTORC2 pathways through phosphorylation of mTOR (mammalian target of rapamycin) at S2481 and that these pathways were constitutively activated in both D816V positive (HMC-1.2) and negative (HMC-1.1, LAD2) neoplastic human mast cells. A dual acting inhibitor of the mTORC1 and mTORC2 pathways, Torin1, and a more selective mTORC1 inhibitor, Rapamycin, abolished proliferation of the malignant mast cells regardless of the presence of the D816V mutation. However, little reduction in numbers of the non-dividing primary mast cells was observed. The effects of the inhibitors on cell proliferation correlated to downregulation of mTORC1 and mTORC2 pathways which was verified by shRNA. Furthermore, no evidence of apoptosis in response to the inhibitors was observed in either neoplastic or primary mast cells. Collectively, our data demonstrate that by targeting the mTORC1 and mTORC2 pathways, it may be possible to inhibit the proliferation of rapidly dividing mast cell populations associated with mastocytosis while leaving normal mast cells largely unaffected.
Prostaglandin E2 induces activation of mTORC1 and mTORC2 in mast cells: selective utilization of mTORC2 for the regulation of chemotaxis and mediator release (151.6)
Authors: Jung Mi-Yeon , Kuehn Hye Sun , Beaven Michael , Metcalfe Dean , Gilfillan Alasdair 
Published: January 2023

Abstract
PGE2 is a potent modifier/regulator of multiple mast cell responses including FcϵRI-mediated degraulation, chemokine production, adhesion to basement membrane proteins, and chemotaxis. Nevertheless, how PGE2 regulates these diverse responses remains unclear. In mouse bone marrow-derived mast cells, we observed that PGE2 induced activation of mTORC1 (mTOR-raptor complex), as indicated by increased p70S6K and 4E-BP1 phosphorylation; and mTORC2 (mTOR-rictor complex), as indicated by increased phosphorylation of AKT at position Ser473. We thus investigated the roles of these signaling complexes in PGE2-mediated mast cell responses. Inhibition of mTORC1, through the use of the selective mTORC1 inhibitor rapamycin or by raptor-targeted shRNA, had minimal impact on PGE2-mediated chemotaxis or chemokine production. In contrast Torin, which inhibited both mTORC1 and mTORC2, and rictor-targeted shRNA, markedly reduced PGE2-mediated chemotaxis and this was associated with down-regulation of PGE2-induced cytoskeletal reorganization. Downregulation of mTORC2 furthermore attenuated PGE2-induced production of the chemokine CCL2, which was linked to a significant reduction in ROS production. Mast cell degranulation and adhesion were unaffected by down-regulation of either mTOR complex. Taken together, these data illustrate that mTOR2, is an important signal for chemotaxis and chemokine production from PGE2-activated mast cells.
Establishing the kinetics of TFH cell development in vivo (152.11)
Authors: Baumjohann Dirk , Okada Takaharu , Ansel K. 
Published: January 2023

Abstract
T follicular helper (TFH) cells have recently emerged as the main regulators of T cell-dependent humoral immune responses. Yet both the developmental pathway and the maintenance of TFH cells are only poorly understood. In addition, it is not clear if TFH cells are a stable T helper cell lineage or if the TFH phenotype represents a transient activation state. The transcriptional repressor BCL6 has been shown to be crucial for the establishment of the TFH phenotype. However, the mechanism by which BCL6 accomplishes this task and the regulation of BCL6 in TFH cells at the pre- and post-transcriptional level remain largely unknown. In order to generate a comprehensive kinetic profile of BCL6 expression in TFH cells on the genomic and protein level, we are taking advantage of a novel murine BCL6-reporter strain and improved intracellular BCL6 staining reagents.
Secreted Frizzled-related protein-1 regulates human peripheral CD4 T cell responses (152.16)
Authors: Lee Yoon-Sook , Lee Kyoo-A , Kang Chang-Yuil 
Published: January 2023

Abstract
Secreted Frizzled-related protein-1 (sFRP1) has been described as a modulator for complex Wnt signaling pathway. In addition to its essential function in T cell development, recent studies have indicated a critical role for Wnt/β-catenin signaling pathway in the biology of mature T cells such as differentiation, polarization, and survival. Here, we investigated the effect of sFRP1 on TCR/CD28 stimulated memory and naive CD4 T cells derived from human peripheral blood in the context of cytokine response of Th subsets. sFRP1 treatment inhibited proliferation and early cytokine secretion of CD3/CD28 activated memory CD4(+)CD45RO(+) T cells, whereas it increased proliferation of CD3/CD28 activated naïve CD4(+)CD25(-)CD45RO(-) T cells. Moreover, sFRP1 influenced lineage choice of naïve CD4 T cells, resulting in enhanced Th1 and Th17 polarization with suppressed Th2 fate. Regulation of Th1/Th2 polarization by sFRP1 was expected from its antagonistic effect on Wnt/β-catenin signaling reported to favor Th2 over Th1 polarization in CD4 T cells. Interestingly, sFRP1 also positively regulated Th17 polarization and sFRP1 treatment of naïve CD4 T cells in Th17-polarizing conditions up-regulated IL-17 expression. Taken together, our observations suggest the function of sFRP1 in the control of activated human peripheral CD4 T cell subsets.
Acute-phase serum amyloid A causes Th1 differentiation in murine CD4+ T lymphocytes (152.18)
Authors: Lui Asona , He Rong 
Published: January 2023

Abstract
Serum Amyloid A (SAA) is an acute-phase protein, released predominantly from the liver during large scale inflammation. High SAA levels, up to 1000 times normal, are also associated with autoimmune conditions like Rheumatoid Arthritis (RA). SAA is found in very high levels in the arthritic joints of patients, along with a high concentration of certain subsets of T lymphocytes, notably helper T cell type 1 (Th1). The marker for Th1 cells is production of the cytokine interferon gamma (IFNγ). The purpose of this study is to determine if SAA stimulation of murine T cells causes Th1 differentiation. CD4+ T-cells isolated from the spleens of wild type mice have been stimulated in vitro with SAA protein. IFNγ production was measured and compared to that of the known inducer of Th1 cells, interleukin-12 (IL-12). Indeed, SAA stimulation induced IFNγ production with levels as high as 22,088 pg/mL, which is comparable to the 20,237 pg/mL induced by IL-12. Our data also shows that SAA and IL-12 can co-stimulate T cells and result in even higher levels of IFNγ secretion. Future experiments will confirm Th1 differentiation by detecting cell surface markers with flow cytometry and will also investigate the intracellular actions of SAA stimulation. These results suggest that SAA and Th1 cells are found together in arthritic joints because SAA induces this T-cell subset’s production, and is perhaps a missing link in our understanding of RA.
E3 ubiquitin ligase Cbl-b negatively regulates T helper 2 cell responses by targeting Stat-6 for ubiquitination (152.19)
Authors: Qiao Guilin , Ying Haiyan , Guo Hui , Chen Bohao , Zhu Xiangdong , Solway Julian , Penninger Josef , Langdon Wallace , Zhang Jian 
Published: January 2023

Abstract
Casitas-B-lineage lymphoma protein-b (Cbl-b), an E3 ubiquitin ligase, belongs to the Cbl family. Recent studies indicate that Cbl-b emerges as a gatekeeper that controls the activation threshold of the antigen receptor and maintains the balance between tolerance and autoimmunity. However, the role of Cbl-b in T helper (Th) cell differentiation remains unknown. In this study, we show that Cbl-b deficiency selectively results in a biased Th2 differentiation in vitro. Furthermore, using a mouse model of asthma, we find that the absence of Cbl-b results in severe airway inflammation which associates with stronger Th2 responses including elevated IL-4, IL-5, and IL-13, and accumulation of eosinophils in bronchoalveolar lavage (BAL) fluid and higher titers of IgE in the serum. At the molecular level, Cbl-b selectively associates with Stat-6 upon IL-4 ligation, and targets Stat-6 for K48-linked polyubiquitination and subsequently degradation in the proteasome. These processes are heightened in the presence of TCR/CD28 costimulation. IL-4-induced Cbl-b-Stat6 association requires Cbl-b TKB domain, whereas TCR/CD28-induced Cbl-b-Stat6 interaction is mediated by tyrosine residues of Cbl-b and SH2 domain of Stat6. Therefore, we have discovered a previously uncharacterized function of Cbl-b in Th2 cell differentiation that may serve as an important therapeutic target for allergic diseases including asthma.
IL-10 production by Th2 cells in vivo occurs by sub-differentiation into a novel, suppressive IL-10+CTLA-4hi Th2 subset (152.28)
Authors: Altin John , Goodnow Chris , Cook Matthew 
Published: January 2023

Abstract
IL-10 is a critical regulatory cytokine that can be produced by many cell-types of the immune system, including IL-4-producing Th2 cells where it was first identified. However, current knowledge of Th2 cell IL-10 production comes largely from cultures in vitro, and little is known about how IL-10 is regulated at the single cell level during Th2 cell differentiation in vivo. Here, by directly measuring and manipulating IL-4 and/or IL-10-producing cells formed during inflammatory responses in vivo, we show that Th2 cells acquire the ability to produce IL-10 as part of a regulated differentiation process. This process is characterised by the co-ordinate upregulation of a program of other ‘regulatory’ genes, particularly CTLA-4. Differentiation is exquisitely dependent on Card11-dependent external signals that condition IL-10-CTLA-4lo ‘early’ Th2 cells to become IL-2-responsive IL-10-CTLA-4int Th2 ‘intermediates’. Upon exposure to cytokines such as IL-2, IL-10 and IL-21, these intermediates progress into IL-10+CTLA-4hi ‘end state’ Th2 cells that have potent suppressive function. Our findings define a novel process of external signal-regulated Th2 cell sub-differentiation in vivo, culminating in a suppressive subset that may account for negative feedback during chronic Th2 responses.
Molecular mechanism for pre-B cell loss in mutant rabbits (153.17)
Authors: Robbins Gregory , Knight Katherine 
Published: January 2023

Abstract
VH4 is the most frequently rearranged VH gene during B cell development in mutant ali/ali rabbits; however, most VH4-utilizing cells do not survive the pre-B cell stage. The mechanism(s) by which these pre-B cells are lost remains an enigma. We hypothesize that VH4-utilzing pre-B cells are lost due to inefficient pre-BCR formation and inadequate pre-BCR signaling. Ex-vivo cultures of ali/ali bone marrow cells on OP9 stroma yield fewer blasting pre-B cells and reduced levels of phosphorylated-Syk than cultures of wildtype(wt) controls, suggesting that VH4-utilzing pre-BCRs have impaired signaling. We previously determined that pre-BCRs containing VH4-μ chains are less abundant on the cell surface compared to the pre-BCRs containing VH1-μ chains as are found in wt rabbits. A single amino acid substitution in FR2 of VH4(Y47W)restored the levels of cell surface pre-BCR expression to that of wt pre-BCR, and current studies are testing if this substitution is sufficient to restore pre-BCR cell signaling and pre-B cell development. Loss of VH4-utilizing pre-B cells is not likely due to gross structural defects in VH4-μ chains since the conventional BCRs utilizing VH4 are both functional and diverse. Therefore, studies utilizing the ali/ali rabbit offer a unique model system to identify necessary amino acids for pre-BCR formation and signaling.
Role of antigen in later stages of B cell development (153.20)
Authors: Taham-zadeh Dariush , Wu Yuan , Ratcliffe Michael 
Published: January 2023

Abstract
B cell development occurs in the gut associated lymphoid tissue (GALT) of many mammals and birds. In chickens, the early stages of B cell development, including colonization of bursal follicles, B cell proliferation within bursal follicles and repertoire diversification by gene conversion, are supported by surface Ig receptor related constructs that lack antigen binding capacity. Thus early B cell development requires Ig receptor expression but not sIg ligation. In contrast later stages of bursal development, including cortico-medullary redistribution of B cells and their maintenance after hatch, is not supported by those constructs. The later stages of bursal B cell development occur in the presence of gut derived antigens suggesting the possibility that later stage may require antigen mediated sIg receptor ligation as opposed to simply receptor expression. To address this directly, we have introduced Ig receptor related constructs with defined antigen specificity into developing chick embryos. Expression of such constructs is sufficient to support early stages of B cell development, and in the absence of cognate antigen, is not sufficient to support the later stages of B cell development. Preliminary evidence suggests, however, that introduction of cognate antigen results in extended maintenance of receptor expressing B cells after hatch suggesting that ligation of the BCR may act as a survival signal in the later stages of B cell development in the bursa.
Role of CD38 during ontogeny of murine B cells in bone marrow (153.23)
Authors: Romero-Ramirez Hector , Pelayo-Camacho Rosana , Lopez-Santiago Ruben , Rodriguez-Alba Juan Carlos , Santos-Argumedo Leopoldo 
Published: January 2023

Abstract
CD38 is a 45 KD transmembrane protein, expressed by immature and mature lymphocytes. Agonistic antibodies directed to this molecule induce several biological responses: cellular proliferation, protein phosphorylation, intracellular Ca2+ mobilization, rescue from and induction of apoptosis. CD38 ligation with a mAb anti-CD38 (NIM-R5) induces apoptosis of the murine pro-B cell line (Ba/F3); this phenomenon depends on tyrosine kinases activation and the MAP kinase ERK. The expression and function of CD38 during B-cell ontogeny has been poorly analyzed in mice. We analyzed the expression of CD38 during the ontogeny of B cells in murine bone marrow. CD38 is expressed from the preproB stage. ProB, preB and immature B cells from murine bone marrow were all positive for the expression of CD38, but immature B cells had the highest fluorescence intensity, and the proB stage shows a low and high expression of CD38 that suggest two probable populations. The results suggest that CD38 may play a role during the ontogeny of murine B cells. We are analysing whether CD38 ligation on these subpopulations, induces proliferation, maturation or apoptosis.
Molecular interactions of the oncogene NUP98-HOXA9 (153.31)
Authors: Sarma Nayan , Yaseen Nabeel 
Published: January 2023

Abstract
NUP98-HOXA9 is the prototype of the fusion oncoproteins that result from NUP98 gene rearrangements and are associated with acute myeloid leukemia. It consists of an N-terminal FG-rich portion of the nucleoporin NUP98 fused to a C-terminal portion of the homeobox protein HOXA9, and acts as an aberrant transcription factor. Using a yeast two-hybrid assay we have identified several interacting partners of NUP98-HOXA9 including Dynein Light Chain 1 (DYNLT1) and Amino-terminal Enhancer of Split (AES). We showed that the FG repeat region of NUP98 is critical for these interactions. In addition, we found that DYNLT1 localizes to the nuclear periphery, interacts with other nucleoporins and these interactions are dependent on the C-terminal of DYNLT1. Knockdown of DYNLT1 resulted in reduction of the ability of NUP98-HOXA9 to activate its target promoters. AES localizes primarily to the interior of the nucleus and showed a strong interaction with NUP98 but not with other FG repeat nucleoporins. AES augmented the transcriptional activity of NUP98-HOXA9. In the presence of NUP98-HOXA9, AES caused an increase in long-term proliferation of primary human CD34+ cells with an increase in the numbers of primitive cells. AES caused a shift away from the erythroid lineage in primary CD34+ cells, thus counteracting the effects of NUP98-HOXA9 on differentiation. These data establish that DYNLT1 and AES interact and cooperate with NUP98-HOXA9 in transcriptional regulation and cell transformation.
Estrogenic microenvironment influences myeloipoiesis to generate regulatory myeloid cells (153.34)
Authors: VanGundy Zachary , Muth Dillon , White Angela , Taylor Cassandra , Papenfuss Tracey 
Published: January 2023

Abstract
Myeloid cells include macrophages, dendritic cells (DCs), monocytes and their precursors. These cells can promote and regulate inflammation and immunity. Regulatory macrophages, tolerogenic DCs (Tol-DCs) and myeloid derived suppressor cells (MDSCs) are all examples of regulatory myeloid cells. We have found that estriol (E3), a pregnancy specific estrogen, generates Tol-DCs in vivo that protect against the inflammatory autoimmune disease experimental autoimmune encephalomyelitis. To understand the generation of E3-Tol-DCs, we examined E3 effects on myelopoiesis and DC differentiation. Bone marrow (BM) from mice was differentiated for 3-7 days + E3 and phenotypic and regulatory function assessed. Similar to in vivo E3-Tol-DCs, BM-DCs differentiated with E3 showed phenotypic increases in CD80, CD86, PD-L1 and PD-L2 expression and suppressed proliferation of responder cells. E3 exposure also affected myeloid progenitors where E3-BM cells (day 3) suppressed proliferation of responder cells in a dose-dependent manner. These effects were not due to altered differentiation (based on CD11b, Gr-1 or CD11c expression of BM cells). Interestingly, E3-BM cells increased FoxP3+ cells in vitro after 4-5 days in culture with FoxP3+GFP+ reporter splenocytes. These results suggest that E3 promotes Tol-DCs through actions on myelopoiesis and that hormonal modulation during myelopoiesis contributes to the generation of regulatory myeloid cells.