bluuebunny/crossref_metadata_2025_split huggingface dataset:
Are commercial soy-sauce products naturally gluten-free? Preliminary evaluation of selected products sold in the USA
Authors: Shivaramaiah Shambhavi Devarahalli , Shah Yash , Jorgensen Rick Dean , Gangur Venugopal 
Published: August 2023

Abstract
Gluten present in wheat, rye, and barley has been linked to 3 types of immune mediated disorders: Celiac disease, non-celiac gluten sensitivity and wheat hypersensitivity (or wheat allergy). Consequently, detection of gluten in food products is routinely used to validate gluten-free products meant for gluten-sensitive subjects. Fermentation processing has been shown to be effective in degrading food allergens including gluten. Most commercial soy sauce products (SSPs) use wheat in addition to soybean as a major ingredient in their production, thus presence of wheat gluten is expected in such products. Here we tested the hypothesis that commercial SSPs sold in the retail market in the USA will test positive for gluten independent of the country of origin. Commercial SSPs were tested for gluten presence using gluten test kit (LOD, 5 ppm). The test kit was validated for positivity using purified gluten extracted from durum wheat flour and for negativity by screening 2 commercial gluten-free SSPs. The following products have been tested: 4 products from China, 3 from USA, 1 from Canada and Taiwan respectively. Commercial SSPs when used without dilution in the food extract buffer rendered the test kit system invalid for testing. Therefore, a dilution of 1/10 was used in testing. Despite identifying wheat as an ingredient on the product label, all SSPs from USA, Canada, and Taiwan tested negative for gluten. Among these products, only 1 from china tested slightly positive for gluten. These results demonstrate that most commercial SSPs sold in the united states, independent of country of origin, used in this study test negative for the gluten content despite identifying wheat as a major ingredient on the labels. Further product screening is in progress.
USDA/NIFA
The ESCRT protein CHMP5 controls skeletal muscle homeostasis and coordinates myeloid cell-mediated tissue repair
Authors: Trichka Josephine , Adoro Stanley 
Published: August 2023

Abstract
Macrophages promote skeletal muscle homeostasis by participating in crosstalk with satellite cells, fibroadipogenic precursors, and myofibers that drives regeneration after injury. Dysregulation of this crosstalk contributes to conditions like myopathy, inefficient wound healing, and sarcopenia. Despite their central role in muscle homeostasis and regeneration, the tissue-intrinsic mechanisms that coordinate the recruitment and activity of these macrophages are poorly understood. In this study, we investigated how skeletal muscle homeostasis is regulated by the ESCRT protein CHMP5. CHMP5 was initially characterized as a member of the ESCRT family of proteins that coordinate membrane scission events in eukaryotic cells. However, recent studies by us and others have revealed non-canonical roles for CHMP5 wherein CHMP5 promotes the stability of client proteins required for cellular differentiation and cell fate decisions in hematopoietic and non-hematopoietic tissues. Using an in vitro model of myogenesis, we found that CHMP5 knockdown impaired the differentiation of C2C12 myoblasts into myotubes. Furthermore, when wild-type BMDMs were cocultured with CHMP5-knockdown C2C12 myoblasts, they failed to polarize into M2-like macrophages. In vivo, mice with muscle-specific CHMP5 heterozygous deletion (CHMP5hetmice) displayed diminished myeloid cell presence in muscle following cardiotoxin-induced injury. 12 days post-injury, histologic and flow cytometric analyses of muscles from CHMP5hetmice revealed less regenerative progress than littermate controls. These data suggest a critical function for CHMP5 in promoting muscle homeostasis both in the steady state and in the context of injury and inflammation.
Supported in part by intramural funding from the National Cancer Institute, T32 GM007250, T32 NS077888-08, and NIH R01AI143992-03.
NF kappa B regulator Bcl3 controls development and function of classical dendritic cells required for resistance to Toxoplasma gondii
Authors: Guha June 
Published: August 2023

Abstract
The atypical IκB family member Bcl3 associates with p50/NF-κB1 or p52/NF-κB2 homodimers in the nucleus, and positively or negatively modulates transcription in a context-dependent manner. Since Bcl3 expression in dendritic cells (DC) is pivotal for antigen presentation and since classical DCs (cDC) are major antigen presenting cells, we investigated the role of Bcl3 specifically in cDCs in vivo by crossing Zbtb46 cre mice with Bcl3flx/flx mice. Bcl3flx/flx Zbtb46 cre mice were as susceptible to lethal T. gondii infection as total Bcl3−/− mice and generated poor Th1 immune responses. Consistent with this, compared to wildtype controls, splenic Xcr1+ Bcl3-deficient cDC1 cells were defective in presenting Ova antigen to OT-I cells both for Ova257–264 peptide and after infection with Ovalbumin-expressing T. gondii. Moreover, splenic CD4+ and CD8+ T cells from infected Bcl3flx/flx Zbtb46 cre mice exhibited decreased T. gondii-specific priming as revealed by both reduced cytokine production and reduced T. gondii-specific tetramer staining. Consistent with this, splenocyte single cell RNA seq (scRNAseq) in infected mice revealed Bcl3-dependent expression of genes involved in antigen processing in cDCs. We also identified by scRNAseq, a unique Bcl3-dependent hybrid subpopulation of Zbtb46+ DCs co-expressing the monocyte/macrophage transcription factor Lysozyme M. Likewise, by flow cytometry we identified two T. gondii-induced hybrid subpopulations of Bcl3-dependent cDC1 and cDC2 cells both expressing monocyte/macrophage markers, designated as icDC1 and icDC2. Together, our results indicate that Bcl3 in classical DCs is a major determinant of protective T cell responses and survival in T. gondii-infection.
Complement Turning Violent in Ultra-Rare Genetic Disorder
Authors: Pandey Manoj K , Trivedi Vyoma Snehal , Magnusen Albert Frank , Rani Reena , Woods Christopher K , DiPasquale Betsy Ann 
Published: August 2023

Abstract
Sandhoff disease (SD) is an ultra-rare lysosomal storage disorder (LSD), which affects ~ 1/1000, 000 live birth. SD is caused by genetic deficiency of beta (β) Hexosaminidase and resulting excess central nervous system (CNS) synthesis of GM2 ganglioside (GM2) and its impact on neuron death. The exact mechanisms underlying such GM2-driven neuron death are unknown in SD. Glucosylceramide (GC) induced complement 5a (C5a) and its C5aR1 activation causes tissue inflammation in experimental and clinical Gaucher disease. Additionally, C5a-C5aR1 axis was found to sparks CNS inflammation and neurodegeneration in several brain diseases including, intracerebral hemorrhage, traumatic brain injury, myasthenia gravis, amyotrophic lateral sclerosis, neuromyelitis optica spectrum, Alzheimer, and Huntington’s diseases. Here, we determined the increased brain levels of C5a and C5aR1 in a beta (β) hexosaminidase inhibitor (HABI)-induced experimental mouse model of SD, when compared to vehicle (PBS) injected WT control mice. Also, HABI-induced experimental mouse model of SD showed increased microglial cells activation, massive brain generation of pro-inflammatory cytokines and loss of neurons, when compared to vehicle injected WT mice. To assess the relevance of C5a-C5aR1 axis activation for brain inflammation in SD, we targeted β hexosaminidase with HAB-I in WT and C5aR1−/− mice. Strikingly, as compared to HABI-injected WT mice, HABI-injected C5aR1−/− mice caused marked reduction in microglial cell activation, brain production of pro-inflammatory cytokines, and the loss of neurons. These data suggest that the C5a-C5aR1 axis is a critical driver of neuroinflammation and neurodegeneration in SD.
No Funding
The role of complement C3 in the outcome of regional myocardial infarction
Authors: Fang Zhou 
Published: August 2023

Abstract
Coronary heart disease leading to myocardial ischemia is a major cause of heart failure. A hallmark of heart failure is myocardial fibrosis. Using a murine model of myocardial ischemia/reperfusion injury (IRI), we showed that, following IRI, in mice genetically deficient in the central factor of complement system, C3, myocardial necrosis was reduced compared with WT mice. Four weeks after the ischemic period, the C3−/−mice had significantly less cardiac fibrosis and better cardiac function than the WT controls. Overall, our results suggest that innate immune response through complement C3 plays an important role in necrotic cell death, which contributes to the cardiac fibrosis that underlies post-infarction heart failure.
CRISPR screening for cytotoxic granule degranulation pathway in CD8 T cell
Authors: Shibata Hirofumi , Golec Dominic P , Phelan James , Johansen Kristoffer , Huang Bonnie , Schwartzberg Pamela L 
Published: August 2023

Abstract
Degranulation of perforin-containing cytotoxic granules is important for NK cells and CD8 cytotoxic T lymphocytes (CTLs) to kill infected cells or tumors. Defects in this pathway cause the primary form of hemophagocytic lymphohistiocytosis (HLH), which is known as familial HLH (FHL) syndrome. Most of reported FHL causative genes encode molecules that are involved in the intracellular trafficking of cytotoxic granules and their fusion to the plasma membrane, followed by the delivery of their contents into the target cell through immunological synapses. However, other components essential for cytotoxic granule degranulation for CTLs are not well understood.
To identify indispensable molecules for cytotoxic granule degranulation, we established an efficient in vitro CD8 amplification and degranulation system to perform a CRISPR-screening. To test this system, we transduced OT-I CD8 cells with a gRNA library targeting PIP3 binding proteins, which are important for cell adhesion, subcellular trafficking and T cell activation. Cells were harvested at short (Day 8) and medium-timepoints (Day 15) to evaluate the frequency of gRNAs in the degranulated (CD107+) and non-degranulated fractions.
Of the top 10 hits of genes whose deletion suppressed degranulation, five were already listed in the 2022 IUIS classification of inborn error of immunity. We will use individual knockouts to evaluate how these genes affect degranulation. To expand to genome-wide screening, we have also subcloned a full library of sgRNAs into our high efficiency retroviral vector. Thus, we established an efficient assay system to perform a CRISPR-screening for CD8 T cells and can detect genes affected by primary immunodeficiencies.
This work was supported by the Intramural Research Program of NIAID, NIH.
A20 is a negative regulator of both NF-κB and JNK signaling following T cell receptor stimulation in humans
Authors: Dabbah-Krancher Gina L , Ruchinskas Allison , Snow Andrew L 
Published: August 2023

Abstract
A20 is an NF-κB-induced, dual function ubiquitin editing enzyme that negatively regulates NF-κB signaling induced by TNF and other innate immune receptors, constraining inflammation. Indeed, A20 haploinsufficiency drives a severe autoinflammatory disease in humans. Although A20 serves a similar negative feedback function for T cell receptor (TCR) signaling, the molecular mechanisms utilized and their ultimate impact on human T cell function remain unclear. TCR engagement triggers the assembly of the CARD11-BCL10-MALT1 (CBM) protein complex, a signaling platform that governs the activation of downstream transcription factors including NF-κB and c-Jun. We employed an extensive set of mutant expression constructs to better elucidate how A20 regulates TCR signaling output in Jurkat T cells. Intriguingly, A20 knockout cells showed enhanced, sustained NF-κB and JNK signaling after stimulation, which was reversed upon wild-type (WT) A20 complementation. A20 deubiquitinase function was entirely dispensable for NF-κB and JNK regulation, whereas zinc finger domains that confer E3 ligase activity (ZnF4) and linear ubiquitin-binding capacity (ZnF7) were required. Abolishing MALT1-dependent proteolytic cleavage of A20 did not affect suppressive function, although neither N- nor C-terminal A20 cleavage fragments retained regulatory capacity. We also found that CRISPR-mediated deletion of A20 adversely affects the survival and expansion of primary human T cells. Ongoing studies will determine how A20 deletion affects human T cell effector responses after stimulation, and decipher how dysregulated NF-κB and/or JNK signaling contributes to T cell-intrinsic abnormalities that perturb adaptive immune homeostasis.
An old dog with ‘new’ tricks: the MDR1 transporter in T cell-mediated immunity
Authors: Balasubramanian Akshaya , Chen Mei Lan , Hegner Courtney Lynn , Sundrud Mark S 
Published: August 2023

Abstract
Multidrug Resistance Protein 1 (MDR1) is an orphan ATP-binding cassette transporter, originally discovered for removing cytostatic drugs from resistant tumor cells. However, MDR1 is also functionally expressed in many normal cell types. Work from our group has shown that MDR1 is constitutively expressed in cytotoxic T cells (CTLs), where it promotes CTL-mediated immunity to infection. Adoptive transfer of Lymphocytic choriomeningitis virus (LCMV)-specific MDR1 (Abcb1a/b)-deficient naïve CTLs into wild-type recipients, followed by LCMV infection, has revealed that MDR1-deficient CTLs fail to accumulate to wild-type levels in response to either acute (Armstrong) or chronic (Clone-13) LCMV infection. This defective accumulation is observed as early as 5-days post-infection, suggesting that endogenous MDR1 transport activity may preferentially regulate early T cell receptor (TCR)-elicited CTL functions. New single cell (sc)RNA-seq experiments, coupled with RNA velocity and lineage trajectory analyses, suggest that MDR1 expression is induced rapidly during TCR activation, coinciding with the induction of other genes involved in mitochondrial and redox metabolism. In vitro-activated CTLs lacking MDR1 also experience oxidative stress, mitochondrial dysfunction and display mitochondrial localization of MDR1. Together, these data support a new model of MDR1 function, in which MDR1-dependent transport of endogenous substrates suppresses oxidative stress and supports redox homeostasis during periods of metabolic activation. In turn, these concepts provide a novel framework for improving the efficacy and durability of vaccines, and for elucidating metabolic vulnerabilities of multidrug-resistant cancers.
An inter-species amino acid variation modulates signaling strength and function of CD28 in vitro and in vivo
Authors: Brady Alexander , McGeachy Mandy 
Published: August 2023

Abstract
Animal models do not always reflect the human condition. One such example was a phase one clinical trial of a CD28 super agonist antibody (CD28SA). In mice, CD28SA treatment expanded Tregs and reduced autoimmune disease; yet when human volunteers received CD28SA, they experienced life-threatening cytokine storm and multi-organ failure. CD28 is constitutively expressed on T cells and provides the canonical co-stimulatory “signal 2” needed for T cell activation. The proline residues in the CD28 PYAP domain are required for downstream signaling and this motif is conserved across species. However, the human PYAP domain is adjacent to an additional proline, while murine PYAP is followed by an inactive alanine. Previous in vitro studies have suggested this single amino acid variation modulates downstream signaling. We have developed a “humanized” mouse with a CD28 A to P substitution at residue 210 adjacent to the PYAP domain (CD28A210P). In vitro, CD28A210P T cells had increased p-Akt, JunB abundance, and produced more IL-2 compared to WT T cells when stimulated in vitro. In vivo, immunization with antigen/CFA emulsion, CD28A210P mice display increased antibody production, suggestive of enhanced CD28-mediated Tfh activity. In addition, we assessed the response of the “humanized” CD28 mice to the CD28 super agonist antibody. After injection with CD28 super agonist, CD28A210P mice had significantly increased weight loss, elevated and sustained pro-inflammatory cytokine production, and less Treg expansion compared to WT mice. This study suggests interspecies variation in CD28 signaling due to differences outside of the well-studied signaling domains can alter outcomes of CD28 ligation and promote divergent outcomes when targeted in vivo.
Single-Cell RNA Sequencing Reveals Disease Stage-Dependent Transcriptomic Profiles of Regulatory B Cells in Systemic Lupus Erythematosus
Authors: Alajoleen Razan M , Daamen Andrea , Timilesena Prakash R , Li Song , Lipsky Peter E , Luo Xin 
Published: August 2023

Abstract
Systemic lupus erythematosus (SLE) is an autoimmune disease associated with abnormal activation of immune cells. Regulatory B (Breg) cells are a B cell subset that negatively regulate immune responses via secretion of immunoregulatory cytokines interleukin (IL)-10, IL-35, and transforming growth factor (TGF)-β. We had previously shown that in lupus-prone MRL/lpr mice, pre-disease Breg cells were more potent in suppressing autoimmunity than active-disease Breg cells. In this study, using single-cell RNA (scRNA) sequencing, we profiled ~10,000 Breg cells from female MRL/lpr mice at the pre-disease (6–8 weeks of age) vs. active-disease (10–12 weeks of age) stages. Based on known markers of Breg subsets, scRNA analysis identified respective clusters as transtional-2 marginal zone precursor B cells (T2-MZP), marginal zone B cells (MZB), transitional 1 B cells (T1), germinal center B cells (GC), and B1 B cells. Our data showed that the pre-disease Breg cells were predominantly T2-MZP and MZB cells. Follicular B cells outside the marginal zone, on the other hand, are significantly increased in the active-disease stage. Two long noncoding RNAs (lncRNAs), Malat1 and Xist, were significantly increased in active-disease Breg cells, where the expression of Lglas3, Slpi, and Spp1 was also increased. Notably, active-disease Breg cells exhibited an exhausted phenotype with increased expression of T-bet (Tbx21). Collectively, our findings suggest that the SLE disease stage-dependent functions of Breg cells are regulated at the transcriptional level. In future experiments, we will determine whether the immunosuppressive functions of active-disease Breg cells can be restored by knocking down the differentially overexpressed genes.
CSF1R blockade improves clinical parameters in prediabetic rhesus macaques
Authors: Stahr Natalie Ann , Phou Amy , Hattler Julian , McGuire David , Kim Woong-Ki 
Published: August 2023

Abstract
It is unknown whether dysregulated CSF1Rhi monocytes/macrophages drive the progression to diabetes and if their removal and endogenous replacement with fresh macrophages resolves disease. Therefore, we hypothesize that depletion of activated CSF1R+ tissue macrophages using a monoclonal antibody against CSF1R and replacement with freshly differentiated macrophages will alleviate inflammation in tissues and ameliorate diabetes-associated disease in prediabetic rhesus macaques.
We identified 6 aged Indian rhesus macaques with heightened fasting blood glucose as well as higher area-under-the-curve in an intravenous glucose tolerance test (IVGTT), indicating prediabetes. Three animals were treated with a rhesus IgG1 recombinantanti-CSF1R antibody administered i.v. (15 mg/kg) once every two weeks for 6 weeks. An IgG1 control was administered to the remaining 3 animals. Clinical parameters such as blood pressure, plasma levels of total cholesterol, glucose, insulin, adiponectin, colony stimulating factor 1 (CSF1), IL1β, monocyte chemoattractant protein 1 (MCP-1) were monitored throughout treatment and up to 8 weeks after the 3rd injection. A 27-color flow cytometry panel were used to monitor changes in immune cell populations throughout the study.
Treatment with anti-CSF1R resulted in decreased total cholesterol, increased adiponectin, increased CSF1, and increased IL1β levels in the blood throughout the study. Flow cytometry experiments confirmed the depletion of CD16+ monocytes (known to be CSF1R+) in treated animals.
Treatment with anti-CSF1R resulted in improvement in multiple clinical parameters associated with the progression to type-2 diabetes mellitus.
Complete tolerogenic response adjuvant stimulates Treg response to immunization
Authors: Janjam Sri Sushma Santhi , Geng Yajun , Yan Zhaoqi , Shin Soo Min , Joshi Kanak , Panicker Anjali Jacob , Shankar Archana , Elangovan Ramya , Koehler Jason , Gnanasekar Varun , Williams Amanda , Feng Addysen , Ellis Christian , Vattikota Anirudh , Chen Aoshuang , Zheng Guoxing 
Published: August 2023

Abstract
We identified the lowest composition necessary to generate a vaccination adjuvant that promotes a Treg response following immunization in mice and named it “complete tolerogenic adjuvant.” This novel adjuvant may allow us to utilize the well-established “antigen plus adjuvant” method of vaccination to induce Treg cell-mediated antigen-specific immunosuppression. The minimal composition is dexamethasone, rapamycin, and monophosphoryl lipid A with the following mass ratios: 8:20:3. We have demonstrated why immunosuppressive and immunogenic substances are both required for the formation of genuine adjuvants for Treg cells by dissecting the roles of each of these components during vaccination. Currently, research is ongoing in our group to begin to assess the usefulness of this adjuvant for antigen immunization in murine models of human autoimmune/inflammatory diseases. Specifically, we are evaluating the tolerogenic efficacy of this adjuvant for vaccination regimen targeting the apolipoprotein B antigen, using the ApoE−/− mouse model of atherosclerosis. These studies are expected to help drive the development of further, perhaps more effective, complete tolerogenic adjuvants that could be used to create a plethora of new, novel vaccines for treating immunological illnesses.
Master program and NIH grant