bluuebunny/crossref_metadata_2025_split huggingface dataset:
Multiplex imaging to reveal the dynamics of MAVS-mediated control of Ebola virus replication in tissues.
Authors: Bettis Natalia , Speranza Emily , Sturdevant Gail , Robertson Shelly , Germain Ronald , Best Sonja 
Published: January 2023

Abstract
The RIG-I like receptor pathway that signals through the mitochondrial antiviral signaling protein (MAVS) is important in sensing RNA viruses, driving production of interferons, and inhibiting viral spread. How this signaling coordinates immune responses in tissues is incompletely understood. To understand the mechanisms of MAVS-mediated control, we examined the spatial relationship between EBOV antigen and cells in livers. C57BL/6J (B6) or MAVS−/− mice were inoculated intraperitoneally with 100 FFU of either wild-type (WT-EBOV) or mouse-adapted EBOV (MA-EBOV) with samples collected at 3 and 5 days post infection (dpi). This resulted in a spectrum of disease outcomes: complete survival (B6 WT-EBOV), weight loss and survival (B6 MA-EBOV and MAVS−/− WTEBOV) and 100% lethality (MAVS−/− MA-EBOV). Macrophage (MF), dendritic cell (DC) and T-Cell (TC) responses were examined by Opal-Plex, a multiplex staining method developed in house, combined with quantitative image analysis through object-based segmentation and spatial statistics. In B6 mice at 3 dpi, there was an increase in cellular recruitment of monocytes and CD8+ T-cells. An association between EBOV nucleoprotein (NP) and F4/80+ cells was observed in the survival phenotype and was especially prominent at 3 dpi. Further development of LysMCre MAVSfl/fl mice will determine if MAVS signaling in the macrophages is required for viral control to the macrophage compartment or if the phenotypes observed could have resulted from a bystander effect. This work will help to define dynamics of host control of viral dissemination and further define the relationship between tissue architecture and disease outcome.
All studies were done with approval from the ICUC of the NIH/NIAID.
Supported by The Intramural Program of the NIH/NIAID
Elucidating neuronal innate immune mechanisms in the central nervous system.
Authors: Negatu Seble G , Jurado Kellie A 
Published: January 2023

Abstract
Neurotropic viruses are the most common cause of infectious encephalitis. Published studies have increased our understanding of how resident glial cells and infiltrating leukocytes contribute to neuroimmune responses during pathogen invasion. However, neuron-specific immune responses remain poorly described. Neurons are readily infected by neurotropic viruses, but studies show neuronal stimulation by viral components poorly induces pathways that are critical for viral control, such as the type I interferon (IFN) response. This leads our central question of how do neurons orchestrate effective antiviral immunity during CNS infection? To probe type I IFN responses in the CNS, I have developed an intracranial neonatal infection model using La Crosse Virus (LACV), an emerging RNA virus. Preliminary data show that infections of C57Bl/6 wild-type, Type I IFN receptor (IFNAR) receptor knock-out (KO), and non-canonical type I IFN pathway (STING) KO neonates with both wild-type LACV and LACV-ΔNSs (lacks the known type I IFN antagonist, NSs) viruses causes severe disease and reduced survival of neonates. IFNAR KO and STING KO mice succumb to disease earlier than C57Bl/6 mice. Additionally, in all genetic backgrounds, viral titers have an inverse relationship with survival. Future work will assess cell-specific infectivity of LACV and type I IFN responses in the CNS. I have also developed an in vitro infection pipeline where primary neurons isolated and cultured from mice are infected and further characterized using fluorescence in situ hybridization and RNA-sequencing. I plan to infect neurons of differential maturation states with LACV-ΔNSs and evaluate changes in gene expression compared to uninfected, bystander cells.
Supported by Penn Presidential Fellowship.
A granular view of X-linked chronic granulomatous disease exploiting single-cell transcriptomics
Authors: Muzumdar Sukalp , Ballouz Sara , Lam Fung , Degrange Maureen , Kreuzburg Samantha , Chong Hey , Zerbe Christa , Jongco Artemio , Gillis Jesse 
Published: January 2023

Abstract
X-linked chronic granulomatous disease (X-CGD) is a rare monogenetic immunodeficiency primarily affecting phagocytes. Precipitated by mutations in the CYBB gene, patients exhibit a compromised oxidative burst, leading to recurrent infections which can be life-threatening. Curiously, autoimmune manifestations are also common in patients and carriers. Here, exploiting the cell type-specific nature of this disorder, we characterize X-CGD on a transcriptional level using single-cell sequencing. Peripheral blood from 14 X-CGD probands and 10 carriers signed onto IRB approved protocol NCT00404560, as well as from 15 controls was sampled, and PBMCs and isolated monocytes were subjected to single-cell sequencing. Probands exhibited a strong differential expression signal relative to controls. This was composed of not only genes previously described to be up-regulated in X-CGD such as IFI27, and indeed an autoimmunity-associated broader type I interferon response, but also previously undescribed genes involved in monocyte function (ARG1), antimicrobial proteins (CAMP, SLPI), and inflammasome components (AIM2). Surprisingly, expression variability was not greater in carriers relative to probands or controls, indicating a lack of cell autonomous effects from the deletion of CYBB. Interestingly, aggregate expression of differentially expressed genes in the probands was able to classify carriers from sex-matched controls with high accuracy (AUROC = 0.92), indicating the presence of an X-CGD-specific gene signature. This gene signature was also strongly co-expressed across 17 chordate species, pointing towards the disruption of ancestral pathways important in antimicrobial immunity in X-CGD probands and carriers.
This work was partially supported by a Swiss National Science Foundation fellowship to S.M.
Phenotypic Heterogeneity in Patients and Mice with Immunodeficiency Caused by SASH3 Mutations
Authors: Majumdar Shamik , McDermott David , Jaiswal Hemant , Cho Kyoungin , Khillan Jaspal , Murphy Philip 
Published: January 2023

Abstract
SASH3 encodes a putative adaptor protein designated SAM and SH3 domain-containing 3 (SASH3), also known as SH3-containing Lymphocyte Protein 1 (SLY1), that is expressed in the cytoplasm and nucleus of lymphocytes. Recently we reported mutations in SASH3 in 4 unrelated male patients as the cause of X-linked recessive immunodeficiency presenting with B, NK and CD4+ T cell lymphopenia; recurrent sinopulmonary, cutaneous and mucosal infections; and refractory autoimmune cytopenias. Patient P1 was hemizygous for a cysteine substitution at the highly conserved Arg347 located in the C-terminus resulting in a protein-positive phenotype, while the other three patients were hemizygous for nonsense protein-negative variants. P1’s immunophenotype was confounded by concomitant large granular lymphocyte leukemia (LGL) and was clinically associated with severe neutropenia, low IgM, significant wart burden on all four extremities, recurrent aphthous stomatitis, and a history of pneumonia. To study the effects p.Arg347Cys, we developed a knock in (KI) mouse model in the C57BL/6 strain designated SASH3KI by CRISPR-Cas9 gene editing. Similar to the patient, SASH3KI mice displayed significant T cell lymphopenia in blood, whereas B cell numbers were normal. CD8+ T cells in SASH3KI mice were reduced in frequency and absolute number in bone marrow, spleen and both mesenteric and inguinal lymph nodes, but were normal in thymus. After infection with LCMV, T cells in SASH3KI mice proliferated normally and produced normal levels of effector molecules. These findings contrast with those reported previously in Sly1−/− and Sly1d/d mutant mice, which both exhibit thymic and splenic hypocellularity and B and T cell lymphopenia.
Human G Protein-Coupled Receptor 15 mutations associated with inflammatory bowel disease.
Authors: Cheng Yan , Cui Jing , Ravell Juan C , Lenardo Michael J 
Published: January 2023

Abstract
G Protein-Coupled Receptor 15 (GPR15) is a chemokine receptor, primarily expressed in T cells based on human single-cell RNA sequencing data. Its natural ligand, GPR15L, is found in the gastrointestinal basal epithelial region and presumably guide GPR15+ T cells to that location. Previous studies showed that Gpr15−/− mice were more susceptible to C. rodentium-induced colitis but the function of GPR15 and whether its deficiency plays a role in inflammatory bowel disease (IBD) in humans are unclear. We recruited three pediatric patients who developed severe early onset IBD and found novel GPR15 mutations using whole-exome sequencing. A reduction in T cells was observed in patient colon biopsies. Surface staining of patient T cells after TCR stimulation further revealed low expression of the mutant GPR15 alleles compared to the wildtype. Characterizing the GPR15 mutants in transduced stable cell lines, we found that the mutations led to lower GPR15L-dependent signaling, causing decreased cell chemotaxis towards the ligand. Accordingly, Gpr15−/− mouse colon also exhibited reduced T cell phenotype observed in patients. Our results indicate that GPR15 is critical for T cell and migration to the human colon and its absence may lead to immunodeficiency in the region, predisposing individuals to IBD induced by specific pathogenic microbiota. This research was supported NIAID by the Intramural Research Program of the NIH.
This research was supported NIAID by the Intramural Research Program of the NIH.
From genetic variants to diseases: the role of LACC1 in T cells
Authors: Li Yingcong , Chandra Vivek , Vijayanand Pandurangan , Kronenberg Mitchell 
Published: January 2023

Abstract
Expression quantitative trait loci (eQTLs) studies provide associations of genetic variants with the expression of particular genes. But how these eQTLs directly affect gene expression and in what cell types of the immune system they act remain largely unknown. One hypothesis is that functional eQTLs in the cis-regulatory elements may affect cell type-specific transcription factor binding, thus reducing gene expression and even contributing to the cause of autoimmune and inflammatory diseases. Previously, we identified T cell-specific eQTLs of a gene known as laccase domain containing 1 (LACC1), which are GWAS hits for Crohn’s disease. Four of them are located in the LACC1 promoter region in the same haploblock. Direct association of disease-risk variants with lower LACC1 expression was confirmed by comparing pre-mRNA quantity of the different alleles in LACC1 heterozygous human CD4 T cells. We further validated the functional role of the eQTLs using CRISPR Cas9-mediated base-pair editing. LACC1 function has been analyzed primarily in macrophages and shown to affect their function, but the GWAS hits do not affect expression in macrophages. Moreover, in our in vitro studies, human CD4 T cells with LACC1 knockdown showed altered metabolic programming and proliferation, which may contribute to inflammatory diseases. Overall, our study provides an excellent model for analyzing and explaining how genetic variants contribute to diseases by influencing gene expression in specific cell types and changing cell functions.
Targeting the NLRP3 inflammasome in Rare hereditary blood disorder Fanconi Anemia
Authors: Beesetti Swarna , Sumpter Rhea , Green Douglas R 
Published: January 2023

Abstract
Fanconi anemia (FA) patients are at higher risk of developing bone marrow failure (BMF) or cancer upon exposure to potential threats like viral infections, bacterial infections, carcinogens, and mutagens. Deleterious mutations in FA genes cause multiple cellular defects which include impaired ability to repair DNA interstrand crosslink lesions in the nucleus and impaired removal of damaged mitochondria via mitophagy in the cytoplasm. Fancc-deficient mice exhibit enhanced inflammatory response and are hypersensitive to lipopolysaccharide-induced septic shock as a result of hematopoietic suppression. Considering the role of inflammation in pathogenesis of Fanconi anemia and bone marrow failure diseases in general, we asked whether inflammasome inhibition could be a promising therapeutic for the treatment of FA. Our studies demonstrate that MCC950, a small molecule inhibitor of the NLRP3 inflammasome, can protect Fancc null mice from LPS-induced hemopoietic suppression and septic shock as evidenced by increased survival rate of Fancc−/− mice. MCC950 partially rescues the thrombocytopenia in peripheral blood and partially restores the loss of megakaryocyte-erythroid progenitor (MEP) and granulocyte-macrophage progenitor (GMP) pools upon LPS challenge. Further, Treatment with MCC950 significantly improved the bone marrow colony-forming capacity of Fancc−/− bone marrow cells in vitro. Our studies demonstrate that MCC950, a small molecule inhibitor of the NLRP3 inflammasome, can protect Fancc null mice from LPS-induced hemopoietic suppression and septic shock and warrants further evaluation as a potential therapeutic strategy in FA.
Augmentation of lymphoid tissues in the human immune system mouse to advance host directed therapies for HIV
Authors: Holloway Alex , Saito Tais B , Naqvi Kubra F , Huante Matthew , Fan Xiuzhen , Lisinicchia Joshua , Gelman Benjamin B , Endsley Janice , Endsley Mark 
Published: January 2023

Abstract
The study of HIV infection and pathogenicity in physical reservoirs requires a biologically relevant model which resembles the human immune system and physiology. The human immune system (HIS) mouse is well established as a model of HIV, but defects in immune tissue reconstitution remain a challenge for examining pathology in tissues. Herein we show that exogenous injection of the human cytokine FLT-3L into the HSC cord blood HIS mouse model significantly expanded the total area of axial lymph nodes and the circulating percentage of human T cells, thus enabling us to visualize and quantify HIV infectivity in the secondary lymphoid tissues of the spleen and axial node. Further, we detected cell death and T cell depletion in tissues, consistent with HIV pathogenesis. Treatment with the caspase-1 inhibitor VX-765 significantly decreased viral load as measured by gag qPCR, and apoptosis as measured by TUNEL assay, in the spleen. A significant restoration of CD4+ T cells in the spleen due to VX-765 treatment was observed via flow cytometry. In situ hybridization further demonstrated a significant decrease in viral RNA in both the spleen and axial lymph nodes. Transcriptomic analysis further revealed an upregulation in host HIV restriction factors such as APOBR, SAMHD1 and APOBEC3A as a result of VX-765 administration. These findings demonstrate FLT-3L as a mechanism whereby the human immune environment in HIS mice can be enhanced to support investigations of HIV pathogenesis and immune outcomes in tissue compartments. Preliminary results indicate that targeting inflammasome pathways with VX-765 in HIS mice treated with FLT-3L, and infected with HIV, preserved T cell populations and decreased viral load.
Supported by R01 A1HL129881, NIH/NHLBI
The effect of miRNAs targeting TGFβ-Signaling in Multiple Sclerosis
Authors: Rau Christina N , Severin Mary E , Lovett-Racke Amy E 
Published: January 2023

Abstract
Multiple sclerosis (MS) is a demyelinating disease of the central nervous system (CNS) that is mediated by a dysregulated immune system. It can lead to severe neurological issues and is one of the major causes for disability in young adults. The cause of the disease is unknown, the immune and neurodegenerative mechanisms underlying the pathophysiology of the disease are poorly understood. To this date there is no cure and current therapies are not able to completely slow down disease progression.
In a large miRNA profiling study on naïve and effector/memory CD4 T cell in untreated MS patients, our lab found a number of miRNAs differently expressed. One of the main pathways affected by this dysregulation of miRNAs is the TGFβ signaling pathway. This impairment limits the differentiation of regulatory T cells. To study the effect of TGFβ-targeting miRNAs in a murine model of MS, neonatal mice were injected with miRNAs that were found to be upregulated in MS patients at day 0 and 21 of life and experimental autoimmune encephalomyelitis (EAE) was induced using conditions under which the controls would only get a mild form of the disease.
Mice, injected with these miRNAs had an earlier onset of EAE and disease severity was significantly increased compared to control mice. Not only did an injection of TGFβ-targeting miRNAs caused mice to have reduced numbers of Tregs in the spleen and thymus compared to control mice, but these Tregs also showed a dramatic loss of TCR diversity in most Vβ genes as TCR Vβ deep sequencing of sorted Tregs revealed. Our findings contribute to the understanding of Treg dysfunction in MS patients and determining candidate markers for disease susceptibility and treatment efficacy.
Supported by grants from NIH (R01 AI152435-01)
Single-cell RNA sequence (scRNAseq) analysis of T regulatory cells in relapsing remitting multiple sclerosis (RRMS)
Authors: Wang Yan , Seyedsadr Maryamsadat , Markovic-Plese Silva 
Published: January 2023

Abstract
Patients with RRMS have decreased numbers and function of Tregs, which may play a causative role in the pathogenesis of the disease. Here, we used scRNAseq to define Tregs populations in peripheral blood of 3 RRMS patients and 3 matched healthy controls (HCs) and compare their transcriptomes and function. We clustered sorted CD4+ CD25+ CD127− Tregs into active (a) and resting (r)Tregs. Our initial scRNAseq data analysis of the active Tregs identified 324 up-regulated differentially expressed genes (DEGs) and 208 down-regulated genes in RRMS in comparison to HCs. Upregulated genes include major histocompatibility complex (MHC) class I (HLAA, HLAB) and class II (HLADRB5) and TRCA gene, supportive of their activated status; ICAM2, involved in the migration to the CNS; ID3, involved in maintaining Treg pool and suppressive function; LAIR2, an indicator for exhaustive Tregs; NEAT1, whose silencing promotes Treg/Th17 balance, and TCF7, that limits Treg thymic generation. Genes with decreased expression include CD7, required for Treg homeostasis; costimulatory CD81; CTLA4 and JUNB required for optimal Treg function; IFTM1 and IF16 that may regulate IFN type I induction of Tregs; CXCR4; LGALS1 and LGALS3 coding for galectin 1 and 3 which inhibit Treg function; and NFKBIA and TNFAIP3, all suggestive of decreased Treg function in RRMS. Pathway enrichment analysis for DEGs in RRMS Tregs identified the strongest enrichment for type I IFN signaling pathway. Understanding the transcriptional changes involved in deficient Tregs functions in RRMS is crucial for better understanding of the disease pathogenesis and for a development of effective treatment for this disabling disease.
State of Pennsylvania grant: Deficient T regulatory Cell (Treg) Function in Relapsing Remitting Multiple Sclerosis (RRMS)
Spatial transcriptomics and single cell sequencing identifies a unique injury associated resident macrophage subpopulation and reveals differential response to injury
Authors: Erman Elise , Cheung Matthew D , La Fontaine Jennifer , Yang Zhengqin , Agarwal Anupam , George James 
Published: January 2023

Abstract
Acute kidney injury (AKI) is a growing concern in the U.S. and globally. Despite recovery of kidney function, patients remain at increased risk for developing chronic kidney disease (CKD). The cause of this increased risk is unknown and biomarkers to determine kidney injury are severely lacking. A potential candidate is the network of kidney resident macrophages interwoven throughout the tissue, and specifically a subpopulation of kidney injury associated macrophages (KIA) we first identified to appear after AKI and lack MHC II expression. KIA cells were localized within the tissue using Visium Spatial Transcriptomics and flow cytometry. Comparison of KIA cell kinetics between AKI and CKD indicated a correlation between the presence of KIA cells and the persistence of injury. Additionally, KIA cells have localized to the sight of greatest injury following an ischemic event. As kidney function recovers, KIA cells decrease in number and return to quiescent levels by 14 days post injury. In contrast, in the severe CKD model, KIA cells persist for up to six weeks, composing approximately 20% of the total kidney resident macrophage population. A less severe version of CKD results in fewer KIA cells but similar persistence, indicating appearance of KIA cells directly correlates with injury severity. Targeting KIA cells to either enhance or negate their function is a possible therapeutic target to reduce the risk of developing CKD following AKI. As we have identified KIA cells in multiple models of kidney injury, in the future they should be investigated as a potential injury biomarker. We hypothesize that KIA cells are uniquely involved in the pathogenesis of injury and may be a key to an AKI to CKD transition.
Supported by grants from the American Heart Association: (AHA 827257)
P66 is a bacterial “don’t eat me signal” that mimics mammalian CD47 and facilitates immune evasion by Borrelia burgdorferi.
Authors: Tal Michal Caspi , Hansen Paige , Ramadoss Nitya , Volk Regan , Zaro Balyn , Weissman Irving L 
Published: January 2023

Abstract
Innate immunity, the first line of defense against pathogens, relies on efficient elimination of the invading agents by phagocytes. Thus in the co-evolution of host and pathogen, pathogens developed mechanisms to dampen and evade phagocytic clearance. Here, we report that bacterial pathogens can evade clearance by macrophages through molecular mimicry of a mammalian anti-phagocytic “don’t eat me” signal. Using a high affinity structural probe for human CD47, a dominant “don’t eat me” signal, we discovered a bacterial protein that mimics CD47’s structure on the surface of Borrelia burgdorferi (Bb), a bacterial spirochete that can establish infection in mammals including Lyme Disease (LD). Blockade of the mimic promotes clearance of the infection in vivo. We identified P66, a known virulence factor, as the bacterial mimic of CD47. Finally, we determined that patients who return to health following LD infection are more likely to generate antibodies to p66 compared to patients who do not. This study demonstrates molecular mimicry as a means used by Bb to inhibit macrophages and evade phagocytic clearance; this mechanism may have broad implications for understanding host-pathogen interactions and the development of therapeutic strategies to combat bacterial infection.
Research reported in this publication was supported by the Fairbairn family foundation; Bay Area Lyme Foundation; the Younger family foundation; the Robert J. Kleberg, Jr., and Helen C. Kleberg Foundation; the Virginia and D. K. Ludwig Fund for Cancer Research; AML grant R01CA086017; the PCBC from NIHLB U01HL099999; as well as grant U19AI109662. M.C.T. was supported by Stanford Immunology training grant 5T32AI007290, and the NIH NRSA 1 F32 AI124558-01 award. L.B.T.D. was supported by a Stanford Diversifying Academia Recruiting Excellence fellowship. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.