bluuebunny/crossref_metadata_2025_split huggingface dataset:
Mobilization of Notch ligand to a poly-HEMA scaffold to promote thymopoiesis and model T cell development (58.19)
Authors: Falcon Jessica , Lelkes Peter , Conway Daniel 
Published: January 2023

Abstract
Notch signaling is critical for thymopoiesis. Notch ligand’s expression on thymic epithelial cells is crucial to the development naïve T lymphocytes and therefore characterized as one of the primary signaling pathways. We are developing a biomaterial-based model of thymopoiesis. We hypothesize that by binding the Notch ligand onto a biomaterial based scaffold, we can maintain the Notch ligand in its active form, and differentiate mouse embryonic stem cells into T-lymphocytes. The common protocol for differentiating mouse embryonic stem cells into T lymphocytes is using the OP9 system. In this system, the stem cells are supported by a monolayer of stromal cells that express the Notch ligand on its surface. Our project aims to bypass the monolayer of living cells and replace them with a biocompatible inorganic matrix that provides the cells with the necessary environment to differentiate into T lymphocytes. We are currently using Poly(2-hydroxyethyl methacrylate) which is both biocompatible and has a similar Young’s modulus of the lymph nodes, which also expresses Notch. We have confirmed activation of the ligand through ELISA and are in the process of differentiating the embryonic stem cells. This project holds significant implications for the field on immunology. If successful, this early prototype can become the basis of the first artificial thymus.
Modulation of the Inflammatory and Metabolic Profile of Pancreatic β-cells Following TLR Ligation. (61.5)
Authors: Mc Evoy Fiona , Holthofer Harry , Loscher Christine 
Published: January 2023

Abstract
Type 2 Diabetes is a multifactorial metabolic disorder characterised by chronic hyperglycemia, insulin resistance and an insulin secretion deficit due to β cell exhaustion. Evidence from animal models and humans has indicated that systemic inflammation plays a role in the pathophysiological processes of diabetes and is facilitated by innate immune responses. TLRs (Toll-like receptors) are key innate immune receptors that recognise both exogenous (pathogen-associated molecular patterns) and endogenous (molecules generated after stress and free fatty acids) ligands. Their activation leads to a proinflammatory reaction that is fundamental for the initiation of an innate immune response and subsequent adaptive responses, but can also damage tissues. Although TLR expression is increased in many inflammatory disorders the effects of metabolic aberrations on TLRs and their role in diabetes is still emerging. Our objective was to elucidate the role of TLRs in the insulin secreting β cell line MIN6. We have shown that MIN6 cells express surface markers vital to the immune response including TLR2, TLR4, TLR5, MHCII, CD14 and CD40. Specific TLR stimulation alters β-cell insulin secretion in response to glucose. It also results in the secretion of various chemokines (Mip-1, Mip-2, MCP and RANTES), and alters the secretion of cytokines (IL-4 & IL-27). These results suggest that engagement of TLRs on β-cells has the ability to affect both the metabolic and inflammatory profile of the cell.
Fluorescent transgenic reporter mice can be used for in vivo Imaging of lymphatic vessels and high endothelial venules in a Sjögren’s model (61.7)
Authors: Ruddle Nancy , Truman Lucy , Bentley Kevin , Alonso-Gonzalez Noelia 
Published: January 2023

Abstract
Dramatic changes occur in the lymph node vasculature after immunization that affect antigen priming. There is a temporary defect in lymph flow and high endothelial venules (HEV) revert to an immature phenotype. To visualize these dynamic changes, we have made transgenic mice with red-fluorescent lymphatic vessels and green-fluorescent HEV that are suitable for live imaging. The lymphatic vessels of ProxTomxHec6st/eGFP reporter mice were successfully visualized in vivo using 2-photon laser scanning microscopy. We are interested in tertiary lymphoid organs (TLO) that develop in the autoimmune disease Sjögren’s-syndrome. Changes in lymphatic vessels and HEV may precede lymphocyte infiltration and autoantibody production. The presence of germinal centers in the minor salivary glands of Sjögren’s patients predicts the onset of extra-nodal lymphoma. We show lymphatic vessels, lymphocyte infiltration, HEV, T and B cell compartmentalization, and spontaneous lymphoma in BL/6 mice over 15 months old. We have made transgenic mice that express LTα and LTβ under the control of salivary amylase to model Sjögren’s syndrome. We are crossing the ProxTomxHec6st/eGFP reporter mice with mice that over-express lymphotoxin in their salivary glands, to understand the temporal sequence of the vascular changes that occur in Sjögren’s syndrome.
Hodgkin and Reed-Sternberg cells modulate endothelial cell function in classical Hodgkin lymphoma (61.9)
Authors: Lim Yaw-Chyn , Fhu Chee-Wai , Yap Celestial , Graham Anne , Chong Siew-Meng 
Published: January 2023

Abstract
Growth and survival of neoplastic Hodgkin and Reed-Sternberg cells (H-RS) cells in classical Hodgkin lymphoma (cHL) is dependent on cell-cell contact and soluble mediator signals derived from the inflammatory infiltrate within the lesion. However, the mechanisms underlying recruitment of the inflammatory infiltrate, particularly T cells, into lesional lymph nodes remain unclarified. In this study, we test the hypothesis that H-RS cells secrete soluble mediators to induce endothelial cell (EC) activation to modulate T cell recruitment in cHL. We examined in-vitro interactions, under static and flow conditions, between freshly isolated T cell subsets and EC treated with conditioned medium (CM) harvested from the H-RS cell line, KM-H2. Both CM-stimulated EC and TNFα-stimulated EC support CD4+ naive and memory T-cell interactions under flow conditions. Both CD4+ naive and memory T cell demonstrated enhanced transmigration across CM-stimulated EC monolayers in response to CXCL12 as compared to TNFα-stimulated or unstimulated EC. Our data show that the upregulation of inducible adhesion molecules on CM-stimulated EC is dependent on the NFκB pathway; but the dominant mediator involved is not TNFα. However, blockade of cyclooxygenase activity in KM-H2 cells partially attenuated the efficacy of the CM in activating EC. These findings suggest that H-RS cells secrete soluble mediators that can modulate endothelial cell function and influence recruitment of T cells into the cHL lesion.
A novel immunoassay strategy to measure total serum lymphotoxin-α levels in cynomolgus monkeys dosed with an anti-LTα antibody (62.5)
Authors: Young Judy , Nguyen Allen , Qiu Zhihua Julia , Ying Yong , Gao Xiaoying , Reed Chae , Chuntharapai Anan , Deng Rong , Luttman Jeff , Grogan Jane , Wong Wai Lee , DeForge Laura 
Published: January 2023

Abstract
Lymphotoxin α (LTα) is a TNF superfamily cytokine secreted from activated lymphocytes as a homotrimer (LTα3) or complexed on the cell-surface with LTβ as heterotrimers. Compelling evidence indicates that LTα is involved in the pathophysiology of autoimmune diseases. Given that blocking the activity of LTα holds promise as a treatment option, a neutralizing monoclonal antibody (mAb) is under clinical development as a potential therapeutic. In preclinical studies of mAb therapeutics, measuring the level of the target antigen is often of interest as a pharmacodynamic marker. We found that the presence of anti-LTα caused a dose-dependent decrease in ability to detect LTα by immunoassay due to the inability of detection antibodies to bind in the presence of bound therapeutic mAb. To circumvent this problem, an alternate assay strategy was devised to accurately measure total LTα3 (free LTα and LTα/anti-LTα complexes). Specifically, excess anti-LTα was added to fully saturate all binding sites on LTα3. Bound anti-LTα was detected using an anti-idiotype mAb. Using this approach, total LTα was measured in serum of cynomolgus monkeys dosed with anti-LTα. Circulating LTα concentrations showed dose related increases up to 8,000-fold that correlated with anti-LTα PK. This assay strategy may be useful in other situations where low molecular weight and/or multimeric structure of a target antigen makes accurate detection in the presence of therapeutic mAb problematic. KEY WORDS LTα mAb PD
Cytokine profiles of patients with Staphylococcus aureus skin infections (70.12)
Authors: Bartman Caroline , Chen Luqiu , Chong Anita , Daum Robert , Alegre Marisa 
Published: January 2023

Abstract
Methicillin-resistant S. aureus (MRSA) causes skin infections in epidemic proportions among otherwise healthy individuals. Several lines of evidence suggest that Th17 cells may be crucial to prevent localized S. aureus infections, as they occur in patients deficient in STAT3 or mice lacking IL-17A and F. In this study, we tested the hypothesis that patients presenting to the emergency room (ER) with skin infections have a reduction in Th17 differentiation, either globally, or specifically to S. aureus antigens because of inhibitory factors produced by MRSA strains. Blood samples from 71 patients presenting to the ER with skin infections and 142 controls were stimulated with MRSA lysates, or with the T cell mitogen OKT3, and supernatants were collected at 24h for detection of cytokines by ELISA. Upon stimulation with MRSA lysates, PBMCs of infected patients displayed increased production of IL-1β and IL-6, innate cytokines that can drive Th17 differentiation. In contrast, production of IFNγ and IL-17 in response to OKT3 was similar in patients and controls. These data suggest that unlike immunodeficient patients with recurrent S. aureus infections, our otherwise healthy ER patients with skin infections do not have a global intrinsic defect in Th17 differentiation. The next set of experiments is addressing the specific T cell recall responses to S. aureus antigens in patients and controls to determine whether patients can mount an anti-S.aureus T cell response in vivo.
Colitis in mice with WASP deficiency in innate immune cells is associated with impairment in IL-10 production (71.10)
Authors: Nguyen Deanna , Goettel Jeremy , Wurbel Marc-Andre , Marin Romela , Ahmed Osub , Horwitz Bruce , Snapper Scott 
Published: January 2023

Abstract
Colitis occurs in mice and some patients deficient in the Wiskott-Aldrich Syndrome protein (WASP), a hematopoietic-specific molecule that regulates the actin cytoskeleton. This colitis is lymphocyte-dependent, but severe disease is observed in chimeric mice with WT T cells and WASP KO innate immune cells, generated by transfer of WT CD4+ T cells into WASP/RAG DKO mice. Given the immunomodulatory function of IL-10, coupled with our findings that mixed bone marrow chimeras are free of colitis, we investigated whether alterations in IL-10 play a role in our colitis model. We found reduced IL-10 transcript levels in mesenteric lymph nodes of chimeric mice. To tease out the cellular source of decreased IL-10 production, WASP/RAG DKO DCs were stimulated in vitro and observed to produce less IL-10. We then transferred IL-10GFP CD4+ cells and found a reduction in the IL-10-expressing proportion of T cells in WASP/RAG DKO compared to RAG KO mice. Lastly, exogenous administration of IL-10-Ig completely abrogated disease in chimeric mice. In summary, besides impaired Treg homeostasis and function, severe colitis in our chimeric mice is associated with reduced IL-10 production in MLNs and in vitro stimulated DCs. Taken together with prevention of disease with IL-10 treatment, colitis in this model may result from decreased IL-10 production by WASP KO innate immune cells.
Imprinting CD103+ LP-DCs and the acquisition of luminal antigen is dependent upon CCR6 (71.12)
Authors: Wheeler Leroy , McDole Jeremiah , McDonald Keely , Miller Mark , Newberry Rodney 
Published: January 2023

Abstract
Dendritic cells (DCs) play a crucial role initiating and guiding immune responses. DCs are found within the gut associated lymphoid tissue, within the lamina propria (LP), and closely associated with the small intestinal epithelium. The epithelia associated DCs (EADC) are positioned to be imprinted by epithelial cells and to acquire luminal antigens, however the factors recruiting LPDCs to the epithelium are not known. We observed that CCL20 is expressed by small intestine villous enterocytes, and that the receptor for CCL20, CCR6, is expressed by small intestine EADCs and LPDCs. CCR6-/- mice had a diminished population of EADCs, but not LPDCs when compared with wildtype mice. Mixed bone marrow chimeric mice revealed that CCR6-/- LPDCs were less efficient at associating with the epithelium, demonstrating a requirement for CCR6 recruiting EADCs. Consistent with our observation that goblet cells (GC) act as passages delivering luminal antigens and GC antigens, LPDCs from CCR6-/- mice were inefficient at presenting luminal antigens to T-cells and rarely stained positive for the goblet cell protein cytokeratin 18. Moreover CD103+ LPDCs from CCR6-/- mice had decreased aldehyde dehydrogenase activity, consistent with the requirement for DC-epithelia associations to imprint DCs with a mucosal phenotype. These observations demonstrate an essential role for CCR6 in recruiting LPDCs to the epithelium where these DCs are imprinted and acquire luminal and GC antigens.
Isolation of neutralizing monoclonal antibodies against HIV-1 with predetermined conformational epitope specificity (76.4)
Authors: Sholukh Anton , Mukhtar Muhammad , Humbert Michael , Essono Sosthene , Watkins Jennifer , Shanmuganathan Vivekanandan , Hemashettar Girish , Hu Shiu-Lok , Montefiori David , Polonis Victoria , Schur Peter , Ruprecht Ruth 
Published: January 2023

Abstract
Existing technologies allow isolating antigen-specific monoclonal antibodies (mAbs) from B cells. However, using the whole antigen for B-cell sorting implies a need for extensive screening of newly isolated mAbs, followed by epitope mapping. We devised a direct approach to isolate mAbs with predetermined conformational epitope specificity, using epitope mimetics (mimotopes) that reflect the three-dimensional structure of given antigen subdomains. We performed differential biopanning using bacteriophages encoding random peptide libraries and polyclonal antibodies (Abs) that had been affinity-purified with either native or denatured antigen. This strategy yielded conformational mimotopes. We then generated mimotope-fluorescent protein fusions, which were used as baits to isolate single memory B cells from rhesus monkeys (RMs). To amplify RM immunoglobulin variable regions, we developed RM-specific PCR primers and generated chimeric simian-human mAbs with predicted epitope specificity. Using our strategy we isolated mAbs targeting the conformational V3 loop crown of HIV Env. The new mAbs showed strong dependence on epitope conformation and cross-neutralized viruses of different clades. The novel technology allows isolating mAbs from RMs or other hosts given experimental immunogens or infectious agents.
Elevated IL-17A levels in human lung of fatal cases of influenza A (76.9)
Authors: Rosas-Taraco Adrian , Rodriguez-Ramirez Heidi , Salinas-Carmona Mario , Melo Americo , Barboza-Quintana Oralia 
Published: January 2023

Abstract
Influenza A is an infectious disease and world health problem due to high morbi-mortality. Hypercytokinemia has been detected in sera or plasma of severe cases. However, little is known about the microenvironment in lung of patients in fatal cases. The aim of this study was to analyze the cytokines environment in human lung tissue from fatal cases of influenza A and its relation with viral load. A total of 6 biological samples from fatal cases of influenza A were included in this study. RNA was isolated from formalin fixed paraffin-embedded lung tissue of patients and RT-qPCR was performance to detect and quantify the viral load and T cells and macrophages related cytokines (IL-2, IL-4, IL-6, IL-10, IL-12, IL-17A, IL-23, iNOS, Arg1, IFN-γ, TNF-α, TGF-β1 and RETNLB). Viral load was detected in all human lung samples (4-173 copies/mm3). Low levels of iNOS, IL-2, IL-6 and Arg1 gene expression were found in fatal cases of influenza A (p&lt;0.05). On the other hand, high levels of IL-17A mRNA were found in the same samples (p &lt;0.01). We also found a positive correlation between IL-6 expression and viral load. In conclusion, IL-17A could be associated with fatal outcome in influenza A infected patients.
The involvement of CD62L in the effect of IVIg on the cytotoxic activity of CD8 T cells. (P5031)
Authors: Chabot Dominique , Trépanier Patrick , Bazin Renée 
Published: January 2023

Abstract
Intravenous immunoglobulin (IVIg) is used for the treatment of an increasing number of autoimmune disorders. In the past decades, its uses increased constantly, creating a potential risk of shortage. In order to further understand the mechanisms by which IVIg exerts immunomodulatory effects, our group previously demonstrated that IVIg inhibits the in vitro CD8 T cell activation. The CD8 T cell response is known to contribute to the progression of several autoimmune diseases. Our results showed that antigen-mediated CD8 T cells activation was reduced by more than 50% (proliferation, CD69 expression) in the presence of IVIg (Trépanier et al., Blood 2012). In addition, recent results showed a decreased lytic activity of CD8 T cells in the presence of IVIg. In this work, we used an in vitro cross-presentation assay with bone marrow-derived dendritic cells (DC) from C57BL/6 mice and ovalbumin-specific CD8 T cells (OT-I) to study the mechanism by which IVIg affects cytotoxicity. Preliminary results suggest that IVIg prevents the shedding of CD62L/L-selectin from the surface of T cells. Shedding of CD62-L from the surface of cytotoxic T cells was recently shown to be linked to the acquisition of lytic activity. Therefore, the effect of IVIg observed on the expression of CD62L on the CD8 T cell surface may contribute to the decreased cytotoxicity of these cells. The elucidation of the mechanisms of immunomodulatory effects of IVIg could help to design a potent substitute.
Multiple defects in HLA class II-mediated immune recognition of Burkitt's lymphoma (P5055)
Authors: Haque Azizul , God Jason , Zhao Dan , Cameron Christine , Amria Shereen , Bethard Jennifer 
Published: January 2023

Abstract
While Burkitt Lymphoma (BL) has a well-known defect in HLA class I-mediated Ag presentation, the exact role of BL-associated HLA class II in generating a poor CD4+ T cell response remains unresolved. Here, we found that BL cells are deficient in their ability to optimally stimulate CD4+ T cells via the HLA class II pathway. This defect in CD4+ T cell recognition was not associated with low levels of costimulatory molecules on BL cells, as addition of external costimulation failed to elicit CD4+ T cell activation by BL. Further, the defect was not caused by faulty Ag/class II interaction, because antigenic peptides bound with measurable affinity to BL-associated class II molecules. Interestingly, functional class II-peptide complexes were formed at acidic pH 5.5, which restored immune recognition. Acidic buffer (pH 5.5) eluate from BL cells contained molecules which impaired class II-mediated Ag presentation and CD4+ T cell recognition. Biochemical analysis showed that these molecules were greater than 30 kDa in size, and proteinaceous in nature. In addition, BL was found to have decreased expression of a 47 kDa enolase-like molecule which enhances class II-mediated Ag presentation in B cells, macrophage and dendritic cells, but not in BL. These findings demonstrate that BL likely has multiple defects in HLA class II-mediated Ag presentation and immune recognition, which may be exploited for future immunotherapies.
P2X4 receptor: A mediator of alcohol-induced effects on microglia cells. (P4185)
Authors: Gofman Larisa , Cenna John , Potula Raghava 
Published: January 2023

Abstract
Activation of purinergic receptors on glial cells is associated with functional responses under normal and pathophysiological conditions. Modulation of purinergic receptors in neurodegenerative and neuroprotective processes have become important therapeutic targets in inflammation and immune-related diseases. Chronic alcohol use causes structural and functional abnormalities of the brain and has a profound impact on the neuroimmune system. As immune effectors in the central nervous system (CNS), microglia are activated in pathological conditions and alcohol induces inflammatory processes in the brain leading to neurodegeneration. Among the P2XR subtypes, P2X4R is copious in the CNS and data suggests them to be important mediators of alcohol-induced effect. Our previous unpublished studies have shown that alcohol significantly increases microglia P2X4 mRNA and protein expression. In this study, we investigated the consequences of P2X4R expression on microglia immune function. Fractalkine induced microglia migration was decreased by 75% following 48hrs of alcohol treatment compared to control (p&lt;0.001), which was attenuated 2.3 fold by pretreatment with 5-BDBD, a P2X4 antagonist. Similarly, 48hrs of alcohol treatment decreased phagocytosis by 15% compared to control (p&lt;0.05), this was significantly reversed when pre-treated with 5-BDBD (p&lt;0.001). Our findings suggest that P2X4 receptors play a role in mediating alcohol-induced effects on microglia immune effector functions.
Extracellular signal-regulated kinase 5, a central regulator of Toll-like receptor signaling in vivo and in vitro (P4193)
Authors: Wilhelmsen Kevin , Xu Fengyun , Tran Alphonso , Hellman Judith 
Published: January 2023

Abstract
Background: Sepsis is initiated by interactions between microbes and Toll-like receptors (TLRs), which leads to inflammation, coagulopathy and organ failure. We recently discovered that ERK5, a member of the mitogen activated protein kinase family, mediates TLR2-dependent activation of monocytes and endothelial cells, and promotes TLR2-dependent expression of the coagulation factor plasminogen activator inhibitor 1 (PAI-1). We tested the hypotheses that ERK5 also regulates TLR4-dependent endothelial activation, and that ERK5 mediates TLR2- and TLR4-dependent activation of inflammation and coagulopathy in vivo in mice. Methods: HUVEC were treated with FSL-1 or Pam3Cys (TLR2 agonists), or with LPS (TLR4 agonist), in the presence of the ERK5 inhibitor XMD8-92. Mice were pre-treated with ERK5 inhibitor (XMD-92) and then challenged IV with Pam3Cys or LPS. Cytokines were quantified in HUVEC supernatants and mouse plasmas. PAI-1 was quantified in mouse plasmas. Results: ERK5 inhibition resulted in the downregulation of TLR2- and TLR4-dependent cytokine production by HUVEC in vitro (p&lt;0.05). Similarly treatment with ERK5 inhibitor reduced the induction of cytokines and PAI-1 in plasmas of mice treated with Pam3Cys or LPS (p&lt;0.05). Conclusions: Our results highlight critical, previously overlooked functions for ERK5 in regulating TLR-dependent inflammatory signaling in sepsis.