bluuebunny/crossref_metadata_2025_split huggingface dataset:
Intracellular signaling as measured by single cell network profiling (SCNP) is stable in longtitudinal samples of peripheral blood mononuclear cells (PBMC) from healthy donors (HD) across multiple immune cell subsets (TECH1P.839)
Authors: Ptacek Jason , Hawtin Rachael , Louie Brent , Evensen Erik , Cordeiro James , Mittleman Barbara , Atallah Michelle , Cesano Alessandra , Gregersen Peter , Bridges S. 
Published: January 2023

Abstract
Biomarkers of immune function from viable cells would be of high value for many clinical applications. The immune system responds rapidly to a variety of stimuli, so identifying immune measures that are stable, robust, and relevant to clinical features/outcomes can be challenging. SCNP is a multiparametric flow cytometry assay measuring induced changes in intracellular signaling, providing a functional measure of pathway capacity in multiple cell subsets without physical separation. To assess the longitudinal stability of signaling capacity, PBMCs from 11 healthy donors (7 female, age 29 - 60) were collected at 2 time points and characterized by SCNP of 32 nodes (combinations of a modulator and an intracellular signaling readout) in 21 cell subsets. Signaling stability was assessed for each node:subset combination, calculating the ratio of the signaling variability between time points to the mean signaling level. Of 258 node:subset combinations measured: 160 (62%) had a ratio &lt;0.1; 227 (88%) were &lt;0.2 (i.e. the variability between time points is &lt;20% of the average signaling). Memory B cells had the greatest, though modest, variability in signaling between sample times (median ratio 0.15, range 0.04 - 0.43); naive CD4- T cells had the least variation (median ratio 0.066, range 0.03 - 0.13). This study serves as reference for biomarker development for immune monitoring, demonstrating that signaling capacity is a stable, robust phenotype over time and within an individual.
Self-education via Ly49 receptor engagement of donor and host iNKT cells following in utero hematopoietic cellular transplantation (TRAN1P.874)
Authors: Strong Beverly , Rotzler Tess , Wadhwani Ram , Turner Lucas , Shaaban Aimen 
Published: January 2023

Abstract
Invariant NKT (iNKT) cells recognize glycolipid-CD1d complexes via their TCR but also express inhibitory Ly49 receptors (iLy49s) specific for MHC I ligands. However, little is known regarding the importance of iLy49s in the self-recognition of MHC I by iNKT cells during development. We hypothesized that self-engagement of iLy49s during development alters the phenotype and function of self-responsive iNKT cells. We examined this hypothesis in prenatal Balb/c to B6 and B6 to Balb/c allogeneic chimeras that were established by in utero hematopoietic cellular transplantation (IUHCT) of E14 fetal liver light density cells into age-matched fetal recipients. After birth, we compared iLy49 expression and the functional response of iNKT cells in the allogeneic chimeras to age-matched controls. The levels of allospecific iLy49s on host iNKT and donor iNKT were significantly reduced in all organs tested suggesting developmental recognition of both donor and host MHC class I had occurred. In contrast, the expression of irrelevant receptors was not altered. Lastly, iNKT cells from chimeric mice that expressed allospecific iLy49s exhibited significantly increased production of IL-4 in response to TCR stimulation suggesting that a shift toward a regulatory phenotype had occurred. Future studies will directly evaluate the impact of self-recognition by iNKT cells in the developmental pathogenesis of autoimmunity, allergy and allospecific tolerance.
Lectin-like transcript-1 (LLT1) and exosomal PCNA inhibit NK cell effector function against prostate cancer cells (TUM2P.892)
Authors: Mathew Porunelloor , Horton Nathan , Bowen Kelly , Mathew Stephen 
Published: January 2023

Abstract
NK cell functions are regulated by inhibitory and activating receptors binding corresponding ligands on the surface of target cells, providing vital first line defenses against infections and cancer. In order to establish and grow, cancer cells must escape NK cell-mediated killing. By altering the expression of ligands for NK cell receptors (down-regulating activating ligands and up-regulating inhibitory ligands) cancer cells may evade NK mediated killing. Here we have examined the expression of LLT1 (Lectin-like transcript-1 or CLEC2D), a ligand for the NK cell inhibitory receptor NKRP1A (CD161) as well as proliferating cell nuclear antigen (PCNA), a ligand for NKp44 on surface of prostate cancer cell DU145. Expression of LLT1 on DU145 cell inhibited NK cell cytolytic activity though NKRP1A (CD161). Similarly, expression of exosomal PCNA by DU145 inhibited NK cell cytotoxicity through NKp44. The association of HLA I and PCNA on DU145 forms an inhibitory ligand for NKp44. Blocking the inhibitory signals by anti-LLT1 and anti-NKp44 monoclonal antibodies enhanced the killing of prostate cancer cells by NK cells. Thus tumor cells may use multiple pathways to inhibit NK effector function and combinatorial targeting of these pathways could lead to efficient elimination of cancer cells.
Biomarker: long term study of breast cancer specific CDR3s in peripheral blood (TUM2P.894)
Authors: Alleyne Jessica , Wang Chunlin , Yang Qunying , Byrne-Steele Miranda , Ng Enders , Kwong Ava , Shin Vivian , Lu Chiung-Mei , Han Jian 
Published: January 2023

Abstract
We have identified a group of T cell receptors, represented by their unique CDR3 sequences, that are closely associated with breast cancer. Multiplex PCR was used to amplify the T cell receptor beta (TCRβ) inclusively and semi-quantitatively; the amplicons were sequenced with the Illumina HiSeq platform. Here, we report a longitudinal study where one patient’s peripheral blood samples were evaluated for repertoire changes before and after the treatment. At regular intervals over four years, we collected five peripheral blood samples from the patient. The blood samples were sorted into Tc, Tr, Th, and NKT cell subsets, and the TCRβ genes were amplified and sequenced. We have also sequenced the TCRβ from the patient’s surgically removed breast cancer tissue. We have identified two kinds of biomarkers for breast cancer: public and private biomarkers. The public biomarkers are those CDR3s that are shared with other patients who were also diagnosed with breast cancer; the private biomarkers are expressed exclusively within a particular patient and are also found in the disease tissue. The study has shown that a group of CDR3s can be used as breast cancer-specific biomarkers because they are tightly associated with breast cancer patients, and their expression level can be quantitatively measured from peripheral blood. The discovery of this new class of cancer specific biomarkers will facilitate the development of new drugs and also improve the utility of existing therapies.
Natural killer (NK) cell high-mobility group box 1 (HMGB1) is required for anti-tumor function (TUM2P.895)
Authors: LI Guanqiao , Liang Xiaoyan , Kaus James , Basse Per , Zeh Herbert , Lotze Michael 
Published: January 2023

Abstract
HMGB1 is an evolutionarily ancient protein, serving as genome guardian, autophagy promoter and DAMP molecule, essential in immunity and cancer development. Its role in NK cell activitity(lysis, DC maturation and cytokine secretion) remains unknown. To study the role of HMGB1 in NK cell functionality, we generated transgenic NKH mice with targeted genomic ablation of HMGB1 in NK cells. HMGB1- NK cells are less responsive to IL-2/IL-18 in vivo and vitro(p=.0001). Consistent with a less mature phenotype(CD27+/CD11b-), they expand more effectively with Flt3 ligand(p=.03). Reserve capacities of OXPHOS and glycolysis decrease with HMGB1 deletion(p&lt;.001). Activated NK cells secrete HMGB1 via vesicular exocytosis and up-regulate cognate receptors such as RAGE. This fails to occur in NKH model. Deletion of HMGB1 impairs NK ability to mature DCs. This can be rescued with addition of exogenous HMGB1. Serum IFN-γ levels and cytotoxicity are reduced in NKH mice receiving high dose IL-2(p&lt;.001). Additionally, tumors engraft more readily in NKH mice(p &lt;.0001), but grow at comparable rates with respect to WT, suggesting NK cells are crucial in early tumor surveillance, but not overall growth kinetics. Thus, accelerated tumor engraftment results in diminished survival in NKH cohorts (p &lt;.05). In conclusion, endogenous HMGB1 promotes NK cell development and IL-2-induced proliferation. Besides, HMGB1 sustains NK cell bioenergetics and promotes diverse NK functions including early tumor control.
Functional consequences of TCR transgenic β-chain expression (TUM2P.908)
Authors: Kazansky Dmitry , Silaeva Yulia , Vagida Murad , Kalinina Anastasia , Khromykh Ludmila 
Published: January 2023

Abstract
We have generated mouse strain expressing transgenic β-chain of TCR 1D1 (belonging to the Vβ6 family) on the genetic background B10.D2(R101) (KdIdDb). The repertoire of peripheral T lymphocytes in these mice contains 70-80% of T cells expressing transgenic β-chain and 20-30% of T cells expressing endogenous β-chains. Expression of transgenic β-chain resulted in T cells with phenotype CD44-CD62L+ characteristic of “naïve” T lymphocytes. On the contrary, CD8+ T lymphocytes expressing endogenous β-chains had surface phenotype of activated effectors CD44+CD62L-. In wild-type B10.D2(R101) mice, immunization with lymphoma EL4 (H-2b) cells induced a vigorous CTL response targeted to the H-2Kb molecule and resulted in full rejection of the tumor cells. In contrast, transgenic mice developed a compromised proliferative response in MLR and were unable to reject transplanted allogeneic EL4 cells. During the immune response to EL4 cells, mostly CD8+ T-lymphocytes with endogenous β-chains became CD44+CD62L- effectors in transgenic mice, whereas only a small part of the T-lymphocytes expressing transgenic β-chains became effectors. Effectors CD8+CD44+CD62L- expressing endogenous β-chains showed enhanced expression of PD-1 and contained increased percent of KLRG1-positive cells. The acquisition of these characteristic features was associated with immune selection of H-2Kb-negative EL4 cells and subsequent tumor escape.
A common gut commensal as a cross-reactive candidate in antiphospholipid syndrome (HUM3P.250)
Authors: Dehner Carina , Ruff William , VIEIRA SILVIO , Goodman Andrew , Kriegel Martin 
Published: January 2023

Abstract
Antiphospholipid syndrome (APS) is an autoimmune clotting disease characterized by autoantibodies interfering with coagulation. Given the vast antigenic potential of the gut microbiota, we hypothesized that adaptive immune responses to gut commensals may sustain chronic autoreactivity via cross-reactivity. We identified in in silico searches the common, gram-positive gut commensal Roseburia intestinalis as a prime candidate for molecular mimicry, containing peptide sequences highly homologous to both T and B cell immunodominant epitopes of the major autoantigen in APS, the plasma protein b2-glycoprotein I (b2GPI). PBMCs from APS patients proliferated significantly more to R. intestinalis protein extracts compared to phylogenetically closely related Eubacterium rectale, which lacks homologous peptides. We therefore cloned b2GPI-specific CD4 memory T cells and tested cross-reactivity with peptides in vitro. Preliminary data supports that IL-2 but not IL-4, IL-10 or IL-17 secretion can be elicited from an autoantigen-specific Th1 clone in response to a R. intestinalis peptide. Furthermore, Western blotting using patient plasma samples revealed specific bands for R. intestinalis including a potential cross-reactive protein at 24 kDa. Pre-adsorption using recombinant b2GPI prior to Western blotting showed a decrease in this candidate band. These data support a working model whereby autoreactive T and B cell responses are sustained by cross-reactivity with a common gut commensal.
A novel subset of fibrocyte-like cells in synovial fluid communicate with CD8 T cells and shape the local inflammatory milieu in Juvenile Idiopathic Arthritis (JIA). (HUM3P.253)
Authors: Ferguson Ian , Yano Hiroshi , Griffin Patricia , Michel Joshua , Kietz Daniel , de Vallejo Abbe 
Published: January 2023

Abstract
We reported previously that children with JIA carry senescent CD31+CD8+ T cells disproportionate with age. Such cells are inflammatory effectors via CD31-driven TCR-independent activation pathway. Here, we hypothesized that these T cells interact with non-hematopoietic cells in the synovial space to perpetuate joint inflammation. JIA patients medically indicated to undergo arthrocentesis were recruited. Synovial aspirates were screened by flow cytometry, and short-term cultures were established. Results showed two subsets of plastic-adherent fibrocyte-like cells (FLC). About 75% were procollagen-CD45-CD14- cells that variably expressed CD34. The other ~25% were procollagen+CD45+CD14+IL17RA+ CD34- FLC; a finding unlike similar cells reported for adult-onset rheumatoid arthritis. FLC exposed to rIL-17 and/or rTNFα induced production of MMPs that were reversed with corticosteroid, TNFi, anti-IL6R, or experimental NFkB inhibitors. Co-culture of FLC with autologous CD8 T cells resulted in production of the same array of MMPs and inflammatory cytokines that were partially blocked by neutralizing anti-CD31 antibody. These results suggest a FLC-CD8 T cell communication circuit in the maintenance of joint inflammation in JIA. Further understanding of the regulation and/or the consequences of this local cell-cell interaction could shed light into the systemic complications of this childhood disease that impose lifelong health burden. [Supported by Nancy E Taylor Foundation and NIH].
Functional evaluation of PTPN22 modulation in human T cells (HUM3P.260)
Authors: Perry Daniel , Saikumar Lakshmi Priya , Zhang Lin , Brusko Todd 
Published: January 2023

Abstract
A major event in type 1 diabetes (T1D) pathogenesis occurs when T cells aberrantly target and destroy insulin-producing beta cells. A SNP variant of protein tyrosine phosphatase, non-receptor type 22 (PTPN22) has been associated with T1D by GWAS. This gene encodes for lymphoid phosphatase (LYP), which functions to dephosphorylate proximal TCR signaling molecules, and is thus a major determinant for T cell function. The susceptible variant produces an R620W amino acid change in the LYP protein. As yet, there is no consensus as to how this variant affects T cell signaling and function, and ultimately, contributes to T1D pathogenesis. We hypothesized that the utilization of lentiviruses to stably transduce primary human T cells would elucidate the mechanisms of the LYP-620W variant in T1D pathogenesis. We found that TCR signaling and proliferation were blunted by overexpressing the LYP-620R variant (-25.39±4.15% Ca++ flux, 0.616±0.041 fold expansion), but were only modestly affected by the LYP620W variant (-5.66±2.11% Ca++ flux, 0.823±0.041 fold expansion). In addition, LYP modulation only affected proliferation when APCs were used for activation, as opposed to antibody-coated beads. Finally, endogenous PTPN22 expression was found to be dynamic throughout an activation time course. Our results implicate the T1D-associated LYP-620W variant to be a hypomorph, and have important implications when considering interventions to restore defective pathways for proper immune regulation.
Forced miR-146a expression causes autoimmune lymphoproliferative syndrome in mice via downregulation of Fas in germinal center B cells (HUM3P.249)
Authors: Li Jingyi , Guo Qiuye , Wan Ying , Wu Yuzhang 
Published: January 2023

Abstract
MicroRNAs, a class of small non-coding RNAs, play a major role in various cellular activities during psychological and pathological processes through inhibiting the direct targets expression. MiR-146a is remarkable induced after LPS stimulation and virus infection and highly expressed in patients with immune disorders such as rheumatoid arthritis, Sjögren's syndrome and psoriasis. To date, whether the high level of miR-146a contributes the pathogenesis of immune disorder disease s remains unknown. In order to elucidate precious functions of miR-146a in vivo, we generated a miR-146a over-expression transgenic mouse. The level of miR-146a has been increased 3 to 8 folds higher than WT mice, which is similar to the levels of patients with immune disorders. miR-146a transgenic mice spontaneously developed a ALPS feathered disease with the distinct manifests including elevated double negative T cells, enlarged spleens and lymph nodes, higher serum IgG, and inflammatory infiltrations in livers and lungs at early age. Further analysis revealed that Fas was the direct target of miR-146a and was specifically down-regulated in germinal center B cells. In addition, B cells of miR-146a TG mice showed hyper-homeostasis proliferation after adoptive transferred into SCID mice. Collectively, our results clearly establish the notion that miR-146a may get involved in the pathogenesis of ALPS through targeting Fas in GC B cells and might be a potential therapeutic target.
Myeloid-derived cells reside in subcutaneous adipose tissue but are not found in circulation: a DC-expressing BDCA-1 CD206 and hematopoetic progenitor cells (HUM4P.273)
Authors: Gonzalez Juana , Walker Jeanne , Stanwix Jenny , Correa da Rosa Joel , Krueger James 
Published: January 2023

Abstract
Obesity is an insulin resistance-associated metabolic disorder which is a hallmark of and risk factor for type 2 diabetes and the metabolic syndrome. It is also often linked to cardiovascular disease, certain cancers and inflammatory diseases. Different immune cells have been identified in visceral adipose tissue (VAT) and subcutaneous adipose tissue (SAT) which are indicators of inflammation associated with insulin resistance and systemic endothelial dysfunction in the peripheral vasculature. However, detailed studies of infiltrating leukocytes and associated cytokines are largely absent in human adipose tissue. One hypothesis that may link obesity with inflammatory immune response is that Th17 T-cells may be involved with pathological processes. Since past studies have shown increased production of CCL20 by adipocytes in obese humans , this chemokine would be expected to chemoattract DCs and Th17 T-cells into adipose tissue; associated Th17 activation could be the nidus for inflammatory diseases. In this study, we have found that obese SAT samples have a higher percentage of DCs than in the lean SAT samples of a DC which is BDCA-1 positive. Half of these BDCA-1 cells express CD206, this population is not found in blood. We also found a population unique to SAT, a hematopoietic progenitor cell. We additionally found an increase of Th17 cells in blood of obese subjects in comparison to lean subjects.
FcRγ- and IL-33-dependent house dust mite extract antigen-uptake in lung antigen presenting cells (HYP4P.311)
Authors: Camacho Daniel , Tjota Melissa , Sperling Anne 
Published: January 2023

Abstract
Allergic sensitization can be generated against various allergens, leading to Th2-mediated inflammation and subsequent asthmatic symptoms. However, the exact mechanisms behind allergic sensitization are not precisely understood. Our previous work demonstrated that house dust mite (HDM) signals through the FcRγ-associated receptor Dectin-2 and induces expression of IL-33 by DCs, leading to the differentiation of Th2 cells and asthma pathogenesis. Here, we sought to determine whether changes in antigen uptake by immune cells in the lung interstitium may be responsible for the diminished inflammation in response to HDM previously demonstrated in FcRγ- and IL-33-knockout mice. To this end, we evaluated antigen uptake in lung APC subsets 18 hours after intratracheal instillation of fluorescently labeled HDM in FcRγ- and IL-33-knockout mice. Our findings demonstrate the uptake of HDM by fewer lung-interstitial Ly6C+ monocytes in FcRγ-deficient mice, but equal uptake by DC subsets. Surprisingly, a defect in IL-33 alone resulted in reduced numbers of Ly6C+ monocytes, moDCs, and CD11b+ cDCs containing HDM in the lung interstitium. These data suggest a mechanism by which an FcRγ-dependent receptor is required for uptake of HDM by lung-interstitial Ly6C+ monocytes in order to upregulate IL-33 production in the lung, which may promote uptake by other APC subsets and induce Th2-mediated allergic airway inflammation.
Statins alter IL-33-mediated cytokine production in mouse mast cell (HYP4P.315)
Authors: Taruselli Marcela , Kolawole Elizabeth , Ryan John 
Published: January 2023

Abstract
Statins are HMG-CoA reductase inhibitors widely used to reduce hypercholesterolemia. Recent publications have shown that statins can act as immune suppressors by decreasing pro-inflammatory functions of APCs and T cells. We have found that the statin drug Fluvastatin suppresses IgE-mediated pro-inflammatory cytokine release from mast cells, likely by the decreasing prenylation of small G proteins. We extended these studies to the mast cell activator IL-33. Similar to previous studies, Atorvastatin decreased IL-33-mediated pro-inflammatory cytokine production. Unexpectedly, we have found that Fluvastatin increased TNF and IL-6 production upon IL-33-activation. This augmentation is dependent on stem cell factor (SCF), since in the absence of SCF Fluvastatin does not increase cytokine production. Suppressing G protein prenylation by treating mast cells with farnesyl protein transferase and geranylgeranyl transferase inhibitors did not mimic the effects of Fluvastatin. Furthermore, the product of HMG-CoA reductase, melvalonic acid, was unable to reverse Fluvastatin-mediated inhibition. These data suggest that the pro-inflammatory effects of Fluvastatin are an “off target” effect. Examining the effect of statins in IL-33-activated mast cells will elucidate how innate immunity may be altered by commonly prescribed medications.
Didox suppresses IgE-mediated mast cell response (HYP4P.317)
Authors: McLeod Jamie , Elford Howard , Ryan John 
Published: January 2023

Abstract
Mast cells play a central role in allergic reactions through high-affinity IgE receptor (FcεRI)-mediated responses. FcεRI cross-linking on mast cells activates multiple signaling pathways that lead to sustained inflammation through degranulation, prolonged generation of reactive oxygen species, and production of various cytokines and chemokines. These cellular responses are involved in inflammatory diseases including asthma, rheumatoid arthritis, multiple sclerosis and heart disease. As a synthetic anti-oxidant and a potent ribonucleotide reductase inhibitor capable of suppressing gene expression, Didox (3,4-Dihyroxybenzohydroxamic acid) has become an attractive therapeutic for treating inflammatory diseases. Here we investigated the anti-inflammatory abilities of Didox as a therapeutic approach to attenuate the allergic response. Our results show that Didox treatment yields a dose-dependent reduction in IgE-stimulated cytokine secretion among bone marrow-derived mast cell (BMMC). These effects are not directly linked to reduced receptor expression, or an increase in cell death and suggest a decrease in signaling associated with cytokine secretion. Together these data could indicate a role for Didox in the treatment of MC-associated inflammatory and autoimmune diseases.