bluuebunny/crossref_metadata_2025_split huggingface dataset:
Restoring T cell function to promote resolution of chronic HBV infection
Authors: Rexhouse Catherine , Bailey Jacob T , Moshkani Safiehkhatoon , Cimino Jesse L , Robek Michael D 
Published: August 2023

Abstract
An estimated 296 million people worldwide suffer from chronic hepatitis B virus (HBV) infection, with 1.5 million new infections occurring each year despite the availability of an effective vaccine. Chronic HBV infection persists in a dysfunctional immune environment, manifested by an ineffective T cell response to the virus. Patient HBV-specific CD8+ T cells have diminished proliferative capacity and cytokine production, and they exhibit increased and sustained expression of inhibitory immune checkpoint receptors. In mice with persistent HBV replication, reduction of viral antigens can be achieved by therapeutic immunization with alphavirus- and vesiculovirus-based vaccine platforms expressing HBV antigens. However, blocking PD-1 and CTLA-4 signaling may further enhance antiviral effector T cell function and therapeutic efficacy. Using a mouse model of HBV replication in which mice are transduced with adeno-associated virus encoding the HBV genome (AAV-HBV), we investigated the role of these inhibitory receptors in the T cell response to HBV. Antibody-mediated blockade of either PD-1 or CTLA-4 independently in AAV-HBV mice did not promote viral clearance. Likewise, PD-1 and CTLA-4 inhibition combined with therapeutic immunization with VSV expressing an HBV antigen did not improve anti-HBV T cell responses or virus clearance in high-antigen mice with persistent viral replication. However, in mice with lower initial antigen loads, blockade of these receptors along with VSV therapeutic immunization resulted in an enhanced decrease of circulating HBV antigen. These results suggest new methodologies for improving HBV-specific immune responses to promote a functional cure of the infection.
Supported by the National Institute of Allergy and Infectious Diseases and the National Institute of Diabetes and Digestive and Kidney Diseases of the National Institutes of Health under award numbers R01AI148354 and R44DK113858. The content is solely the responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.
Developing a Robust and High Throughput Assay for Screening Innate Immune Training Molecules
Authors: Forbes Greta E , James Heera , Henderson Mark , John Sinu P , Fraser Iain D C 
Published: August 2023

Abstract
Innate immune training molecules have several potential applications in the biomedical field. We developed an assay to screen for new training molecules by generating HiBiT tagged TNF cells in the THP1 monocytic cell line through CRISPR knock-in. The HiBiT tag, an 11 amino acid long peptide, was fused to the TNF C-terminus to produce luminescence upon addition of the LgBiT substrate. To optimize the assay, we first titrated different concentrations of multiple TLR ligands including pIC, p(dAdT), PGN, FLGN, LPS, R848, and P3C. LPS, PGN, and P3C induced the strongest signal to noise (S/N) ratio in the pool of TNF-HiBiT cells. From the TNF-HiBiT pool, we then single cell cloned and characterized them for optimum TNF response. The clones D3 and D6 had significantly higher S/N ratio compared to the pools, especially in response to P3C and PGN. To optimize the TNF-HiBiT cells for a training screen, we used previously known training molecules such as Syk kinase inhibitor, beta-Glucan and Rutaecarpine, to train multiple TNF-HiBiT cell clones. We found that the clones that induced a lower TNF signal upon acute TLR activation exhibited a more robust training phenotype compared to the clones that induced a more robust acute TNF signal. We found therefore that the optimal protocol to assess trained immunity is to use lower concentrations of activating TLR ligands (0.1, 0.1, and 0.0001 ng/mL of P3C, LPS and PGN) in the trained cells. This may be due to saturation of the TNF-HiBit reporter signal when cells are challenged with higher TLR ligand concentrations (10, 100, LPS, and 10 ng/mL of P3C, LPS and PGN). In brief, we have developed and optimized a robust assay for screening novel training molecules.
This work was supported by the Intramural Research Program of NIAID, NIH
Supported by grants from Intramural Research Program of NIAID, NIH and the Office of AIDS Research at NIH.
Tpl2−/− macrophages exhibit impaired mitochondrial respiration and function in response to LPS stimulation
Authors: Fahey Denise Lyn , Watford Wendy Tharpe 
Published: August 2023

Abstract
Macrophages are instrumental in maintaining immune homeostasis, and their dysfunction or depletion is tied to many human diseases, including autoimmunity, obesity, and cancer. Activated macrophages modify their cellular metabolic pathways, such as glycolysis and oxidative phosphorylation, to drive their diverse functions. Many metabolic processes occur in the mitochondria, including oxidative phosphorylation, metabolite production, and reactive oxygen species (ROS) generation. Previous research demonstrated that tumor progression locus 2 (Tpl2, MAP3K8, or COT) is required for the induction of oxidative phosphorylation and the alternatively activated phenotype in macrophages during Schistosome infection; however, Tpl2 regulation of cellular metabolism in classically activated macrophages remains unexplored. We hypothesize that Tpl2 normally promotes oxidative phosphorylation, and its absence causes mitochondrial dysfunction. LPS-stimulated Tpl2−/− bone marrow-derived macrophages (BMDMs) had decreased oxygen consumption, basal respiration, ATP production, and maximal respiration compared to LPS-stimulated wild type BMDMs. Additionally, Tpl2−/− BMDMs exhibited higher basal and LPS-stimulated ROS production than wild type BMDMs. These data suggest that Tpl2−/− mitochondria are functionally defective, and the absence of TPL2 increases mitochondrial oxidative stress. Together, this may reveal the underlying mechanism for defective Tpl2−/− macrophage behavior. Understanding the role of Tpl2 in classically activated macrophages will provide a foundation for developing targeted therapies to treat cancer and autoimmune diseases.
Supported by grant from NIH (R21 AI147003).
ZIP8 impacts protease activity in macrophages in response to Streptococcus pneumoniae
Authors: Haq Sabah , Smith Deandra R , von Pein Jessica B , Sweet Matthew J , Knoell Daren L 
Published: August 2023

Abstract
Zinc (Zn) is essential for immune function and Zn deficiency increases the incidence of community acquired pneumonia. The Zn transporter ZIP8 is required by myeloid cells to defend against pathogens. Previously, we reported that S. pneumoniae infected ZIP8 knockout mice (Zip8KO) have increased bacterial burden and mortality and that Zip8KO bone marrow derived macrophages (BMDMs) have reduced phagocytic activity. Others have shown that Zn induces cathepsin expression via the TFEB pathway. However, it is unknown whether ZIP8 impacts TFEB-dependent phagolysosomal processing in macrophages. Based on this, we hypothesized that ZIP8 facilitates phagolysosomal processing and eradication of bacteria via TFEB.
BMDMs were derived from wild type and Zip8KO mice and then exposed to S. pneumoniae (MOI=10) for 1 hour. At 6–18 hours post challenge we assessed cathepsin B and L mRNA and protein levels as well as total protease, cathepsin B and L activity. In conjunction, we measured p-TFEB and total TFEB proteins levels.
Active TFEB and cathepsin B protein levels were significantly lower in Zip8KO BMDMs with no difference in cathepsin L. Strikingly, Zip8KO BMDMs had significantly reduced total protease, cathepsin B and cathepsin L activity which also corresponded with decreased mRNA levels.
Based on these findings we predict that ZIP8 loss in macrophages impairs phagolysosomal processing and bacterial eradication in a TFEB-dependent manner. Overall, this warrants future studies given the high incidence of Zn deficiency and the rs13107325 defective ZIP8 allele in humans.
National Heart, Lung and Blood Institute #1R01HL156952-01-A1 (DLK)
Tryptophan diet promotes myeloid-derived suppressor cell response via aryl hydrocarbon receptor in Sjogren’s syndrome
Authors: Lin Xiang 
Published: August 2023

Abstract
Myeloid-derived suppressor cells (MDSCs) are heterogeneous myeloid cell populations with immunosuppressive capacity, which contribute to immune regulation and tolerance induction. We previously reported impaired MDSC function in patients with primary Sjögren’s syndrome (pSS) and mice with experimental SS (ESS). In this study, we first found that aryl hydrocarbon receptor (AhR) was highly expressed by human and murine polymorphonuclear MDSCs (PMN-MDSCs). Indole-3-propionic acid (IPA), a natural AhR ligand produced from dietary tryptophan, significantly promoted PMN-MDSC differentiation and their suppressive function on CD4+ T cells. In contrast, feeding tryptophan-free diet resulted in decreased PMN-MDSC response, a phenotype that could be reversed by IPA supplementation. Notably, AhR expression was reduced in PMN-MDSCs during ESS development, while AhR antagonism resulted in exacerbated ESS pathology and dysregulated T effector cells, which could be phenocopied by tryptophan-free diet. Interferon regulatory factor 4 (IRF4), a repressive transcription factor, was upregulated in PMN-MDSCs during ESS progression. ChIP analysis revealed the binding capacity of IRF4 to the promoter region of AhR, while IRF4 deficiency markedly enhanced AhR-mediated PMN-MDSC responses. Furthermore, tryptophan-enriched diet or dietary supplementation of IPA markedly ameliorated salivary glandular pathology in ESS mice with restored MDSC immunosuppressive function. Together, our results identify a novel function of AhR in modulating PMN-MDSC response and demonstrate the therapeutic potential of targeting AhR for the treatment of pSS.
CD4+ Treg expansion arrests disease progression in a pre-clinical model of alopecia areata
Authors: Lensing Maddison , Connell Samuel J , Kahl Payton , Zhu Zhaowen , Jabbari Ali 
Published: August 2023

Abstract
Alopecia Areata (AA) is a prevalent autoimmune disease that results in nonscarring hair loss. Disease is associated with the collapse of an immune privileged state of the hair follicle, characterized by increased MHC expression, immune cell infiltration and the presence of pro-inflammatory cytokines in the microenvironment. CD8 T cells have been identified as the main contributors to disease and the activating receptor NKG2D can identify this pathogenic population. There is currently one FDA-approved drug for AA, a JAK1/2 inhibitor, which is thought to act by dampening inflammatory mediators during disease. However, the potential of adverse effects and refractory patients necessitates the development of alternative therapeutics. Emerging data from our lab suggests that the observed expansion of CD4 T regulatory cells (Tregs), relative to effector CD8 T cells during AA, is inadequate to prevent disease onset. We hypothesized that pharmacologic enhancement of Treg numbers may restrain development of AA. To address this, we investigated the protective potential of Treg expansion using a murine model of AA. In vivo expansion of CD4+ CD25+ FoxP3+ Tregs was achieved using an IL-2 cytokine/antibody complex (IL-2C), which is known to selectively expand and enhance Tregs. We found that 6 weeks of treatment with IL-2C halted the progression of hair loss, while control mice exhibited robust hair loss. Additionally, IL-2C treatment led to a reduction in NKG2D+ CD8 T cells and IFNγ-producing CD8 T cells found infiltrating the skin, suggesting that increasing Tregs in the system is an effective strategy for restraining disease. Our findings provide translational insight for the use of Treg enrichment as a therapeutic strategy for patients with AA.
Supported by grants from NIH (R01 AR077194) and Dept. of VA (I01 BX004907)
Characterization of the immunosuppressant activity of the novel biogenic amine 3HKA and its analog derivatives
Authors: Martinez Aitziber Buque , Clement Cristina , Shetty Madhur , Santambrogio Laura 
Published: August 2023

Abstract
Tryptophan (Trp) is an essential amino acid which in mammalian cells 90% is processed through the kynurenine pathway. Kynurenine metabolites are among the most potent regulators of immune responses. Herein we report the characterization of a novel immunosuppressant biogenic amine, 3-hydroxy-L-kynurenamine (3HKA), generated through an uncharacterized lateral pathway of Trp catabolism, involving its decarboxylation by the aromatic amino acid decarboxylase. 3HKA is generated by both professional (Dendritic Cells, DC) and non-professional (Lymphatic Endothelial Cells) APC. Quantitative phosphoproteomic analysis of human-monocytes (CD14+) derived DCs, identified key signaling transduction receptors, IFNR1, and signaling molecules from the JAK/STAT and cAMP mediated PKA pathways, regulated by 3HKA. Cell-based assays coupled with ELISA platforms showed that 3-HKA inhibited the IFN-g-mediated STAT1/NF-kB pathway in human DCs, and consequent decreased the release of pro-inflammatory chemokines and cytokines IL-1, TNFa, IL-6, and IL12p70. In vivo, 3-HKA reduced the intestinal inflammation in a model of dextran sodium sulfate-induced colitis. Furthermore, flow cytometry analysis revealed that 3-HKA increased the number of T-cells expressing 2, 3 or 4 exhaustion markers (PD-1, CTLA4, LAG3 or TIM3), exclusively in CD8+CD4− cells. We used, in silico approaches for ligand and pharmacophore-based drug discovery and developed 30 novel 3-HKA analogues with ADMET predicted drug-like properties. Structure-activity relationships studies, leaded to identify compounds with improved anti-inflammatory profiles. The novel anti-inflammatory 3-HKA analogues showed promising activity for the treatment of autoimmune diseases
Targeting of GPR65 with Small Molecules alters Th17 cell differentiation
Authors: Hernandez Jeniffer B , Cave Si , Patel Dhruv , Liang Yuqiong , Yang Qiyuan , Wiater Ezra , Zheng Ye , Montminy Marc 
Published: August 2023

Abstract
GPR65 has been shown to be a critical regulator of Th17 pathogenicity. Loss of GPR65 in mice results in a decrease in Th17 cells and reduced susceptibility to experimental autoimmune encephalomyelitis. The CREB/CRTC2 pathway has emerged as an important regulator of immune function. We have previously shown that the CREB/CRTC2 pathway modulates autoimmune disease by promoting differentiation of Th17 cells. In this study we performed RNA-seq to identify Th17 genes regulated by the CREB/CRTC2 pathway. Our RNA-seq analysis led us to uncover the first mechanism of regulation of the orphan receptor GPR65 by the CREB/CRTC2 pathway. We show that GPR65 is a target of the CREB/CRTC2 pathway through expression studies and chromatin immunoprecipitation. In addition, we show that targeting of GPR65 with small molecules alters Th17 cell differentiation. Understanding the regulation of GPR65 will be crucial in developing small molecules to treat patients with Th17-mediated diseases.
B Cell Targeting mAb–siRNA Conjugates Reduced Autoantibody in Experimental Myasthenia
Authors: Huda Ruksana 
Published: August 2023

Abstract
Conjugates of monoclonal antibodies (mAbs) and siRNA have been used for target cell-specific gene repression. B cells are integrally involved in the anti-Acetylcholine receptor autoantibody-mediated pathogenic development of autoimmune myasthenia gravis (MG). Here we show that simultaneous targeting of the critical receptors expressed on pathogenic B-cell subsets with the optimized dose of mAb-siRNA conjugates markedly depleted target B cells and substantially reduced pathogenic autoantibody in a mouse model of MG. The results show the potential of mAb-conjugates in the treatment of myasthenia gravis.
Supported by grants from AFM-Telethon, France
14-3-3ζ Pseudogenes: Physiologically and powerful evolutionary forces in the genome
Authors: Lughmani Haroon Yahya , Chakravarti Ritu , Chattopadhyay Srestha , Patel Hrushil 
Published: August 2023

Abstract
Contrary to the rigid dogma ruling out the phenotypic relevance of pseudogenes, recent RNA-seq and meta-analyses indicate that 10% of all pseudogenes are transcribed, and 40% of those pseudogenes are translated into protein. This investigation unpackages the 14-3-3ζ (YWHAZ) gene, which has been shown to regulate cytokine signaling, induce antibody response, and reduce inflammation in rats with rheumatoid arthritis. The original YWHAZ gene is located on the 8th human chromosome. Our detailed investigation in the human genome shows that there are ten pseudogenes of YWHAZ. Located across the human genome, these pseudogenes display unique properties, such as having allelic SNPs for Body Mass Index, Prostatic Neoplasms, and Macular Degeneration. Diverse mRNA expression profiles skewed towards reproductive organs and an unusually high transcription rate (9 of 10 being actively transcribed) suggest a strong physiological relevance. Additionally, contrary to the prevailing pseudogene formation theory where all exons are transcribed, YWHAZ pseudogenes have omitted the sixth exon’s UTR region, resulting in a more compact, equally functional gene structure. The evolution of the YWHAZ pseudogene from the parent gene’s framework, combined with some pseudogenes having allelic properties, demonstrates how pseudogenes may be the body’s way to repackage and repurpose parent genes for a more diverse genome and proteome.
The CD16 and CD32b Fc-gamma receptors regulate antibody-mediated responses in mouse natural killer cells
Authors: Aguilar Oscar A , Gonzalez-Hinojosa Maria D R , Arakawa-Hoyt Janice S , Millan Alberto J , Gotthardt Dagmar , Nabekura Tsukasa , Lanier Lewis L 
Published: August 2023

Abstract
Natural killer (NK) cells are innate lymphocytes capable of mediating immune responses without prior sensitization. NK cells express Fc-gamma receptors (FcγR) that engage the Fc region of IgG. Studies investigating the role of FcγR on mouse NK cells have been limited due to lack specific reagents. In this study, we characterize the expression and biological consequences of activating mouse NK cells through their FcγR. We demonstrate that most NK cells express the activating CD16 receptor, and a subset of NK cells also express the inhibitory CD32b receptor. Critically, these FcγRs are functional on mouse NK cells and can modulate antibody-mediated responses. We also characterized mice with conditional knock-out alleles of Fcgr3 (CD16) or Fcgr2b (CD32b) in the NK and innate lymphoid cell (ILC) lineage. NK cells in these mice did not reveal any developmental defects and were responsive to cross-linking activating NK receptors, cytokine stimulation, and killing of YAC-1 targets. Importantly, CD16-deficient NK cells failed to induce antibody-directed cellular cytotoxicity (ADCC) of antibody-coated B cell lymphomas in in vitro assays. In addition, we demonstrate the important role of CD16 on NK cells using an in vivo model of cancer immunotherapy using anti-CD20 antibody treatment of B cell lymphomas.
This project was funded by fellowships from Burroughs Wellcome Fund (BWF) and the Cancer Research Institute to O.A.A., and grants from the Parker Institute for Cancer Immunotherapy (PICI), and the NIH (AI068129 and AI146581) to L.L.L.
BPA modulates the neutrophil response and cytokine environment in a model of tail fin regeneration
Authors: Walser-Kuntz Debby , Wang Hongshen , Harmon Bailey 
Published: August 2023

Abstract
The ER-α and β ligand bisphenol A (BPA) is used in the production of polycarbonate plastic and epoxy resins. BPA presents a potential environmental challenge to both human and wildlife immunity due to its widespread use and its resulting presence in surface water and wastewater runoff. We observed that BPA exposure downregulates the expression of a neutrophil-recruiting chemokine in both mice (cxcl1) and zebrafish (cxcl8) and that BPA exposure decreases neutrophil recruitment to a tail injury in zebrafish. Neutrophils are essential for the removal of damaged tissue and invading microbes, thus facilitating tissue regeneration of the tail fin. To investigate BPA’s effects on larval zebrafish tail regeneration, we induced a tail injury at day 3 post fertilization (dpf) and found that fish exposed to 100 ng/ml BPA regenerate a smaller tail fin area at 3 dpi (p-value &lt; 0.01). We hypothesized that neutrophil recruitment could be tied to regeneration by altering the expression of inflammation- and regeneration-related genes. 3 dpf embryos exposed to BPA show modulated levels of background wnt10a and il-10 expression compared to controls. BPA exposure reduces the 3 hour post-injury inflammatory response by decreasing both il-1 and il-6 expression. By modifying the cytokine environment both prior to and following infection and injury, BPA impacts both the inflammatory response and tissue regeneration. Multiple cytokine and chemokine genes are regulated by the transcription factor NF-kB and Wnt plays a key role in tissue regeneration; these two proteins also cross regulate one another. The impact of BPA on wnt10a and nfkb expression following tissue injury and LPS exposure will be discussed.
Supported by Carleton College.
Regulation of CCR6 Expression on Epidermal γδ T Cells in Response to Damage or Stress in the Skin
Authors: Castellanos Tanya , Lawler William Keele 
Published: August 2023

Abstract
Chronic conditions, including type 2 diabetes and psoriasis, interfere with the timing and location of necessary inflammatory cellular functions. Inflammation in the skin is at least partly mediated by epidermal γδ T cells in humans and mice, which secrete growth factors, cytokines and chemokines. However, less is known about which chemokines regulate epidermal γδ T cells and whether functional epidermal T cell subsets exist. One signaling pathway of interest is the C-C Motif Chemokine Receptor 6 (CCR6)/C-C Motif Chemokine ligand 20 (CCL20) axis, which recruits T cells to the epidermis for wound repair. To investigate how this signaling pathway initiates and regulates epidermal γδ T cell functions, we utilized flow cytometry, publicly available scRNA-seq data. Luminex and immunofluorescence microscopy were utilized to evaluate the location and timing of CCR6 expression by epidermal γδ T cells. Initial findings indicate that CCR6 expression is upregulated in a subset of activated epidermal γδ T cells and that CCL20 induces pro-inflammatory IL-17-biased gene expression. Analysis of publicly available single cell RNA sequencing data from mice with psoriasis show that predominantly proinflammatory genes are upregulated in the CCR6+ epidermal γδ T cell subset. Some proinflammatory genes that are expressed include IL-22, IL-17F and IL-17A. Further, upregulation of CCR6 expression by a subset of epidermal γδ T cells begins within the first hours post wounding and remains elevated until a decline is observed 72 hours post wounding.Further understanding of the role of CCR6 in inflammation can lead to the development of novel treatments for conditions such as chronic non healing wounds or psoriasis.