bluuebunny/crossref_metadata_2025_split huggingface dataset:
B cell selection sites in the nurse shark spleen may represent evolutionary precursors of mammalian germinal centers
Authors: Matz Hanover C , Dooley Helen 
Published: January 2023

Abstract
Sharks are members of the oldest extant taxonomic group (Chondrichthyes) to have immunoglobulin (Ig)-based adaptive immunity. While research has demonstrated that sharks can produce antigen-specific antibody responses, affinity mature these antibodies, and establish immunological memory, histological examination has so far failed to find evidence of germinal centers (GCs). To explore this apparent discrepancy, we immunized several nurse sharks (Ginglymostoma cirratum) with phycoerythrin and visualized the subsequent splenic response using a combination of single nuclei (sn)RNA sequencing and fluorescent microscopy. Our snRNA data reveal the presence of both centrocyte-like and centroblast-like B cells in sharks, as well as a T cell subset expressing several markers found in mammalian T follicular helper cells. RNA fluorescent in situ hybridization experiments show distinct regions of AID expression, polarization of CXCR4 and CXCR5 expression, and association of T cell aggregates with splenic B cell follicles. Fluorescent microscopy clearly demonstrates presentation of nondegraded antigen in the center of B cell follicles. Finally, analysis of Ig variable regions amplified from such B cell follicles reveals evidence of positive selection of clonotypes. Together, our data supports the presence of B cell selection sites in the cartilaginous fish spleen that functionally support the selection of antigen-specific B cells and affinity maturation of the shark Ig repertoire. Further, these sites potentially represent the primordial structures that gave rise to mammalian GCs.
Supported by grants from NIH T32 AI095190, F31 AI147532
Cross-protection by SARS vaccines is improved by booster vaccination
Authors: Dangi Tanushree , Palacio Nicole , Sanchez Sarah , Park Mincheol , Class Jacob , Visvabharathy Lavanya , Ciucci Thomas , Koralnik Igor J , Richner Justin , Macmaster Pablo Penaloza 
Published: January 2023

Abstract
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) vaccines have shown extraordinary efficacy against SARS-CoV-2. Recently, we demonstrated that prime-boost coronavirus vaccine regimens can protect against heterologous coronaviruses (Dangi, The Journal of Clinical Investigation, 2021). However, it remains unclear whether boosters are required for such cross-protection. In this study, we show that booster immunizations are critical to elicit cross-protection against heterologous coronaviruses. We first vaccinated BALB/c mice intramuscularly with a poxvirus-based SARS-CoV-1 vaccine developed in 2004 (MVA-SARS-CoV-1) and compared cross-protection following an intranasal SARS-CoV-2 challenge, evaluating cross-protection after a prime-only regimen versus a prime-boost regimen. Interestingly, we show cross-protection only in mice that received boosters. We are currently testing the durability of cross-protection elicited by prime-boost vaccine regimens, and we are also extending these results to humans. Overall, our findings provide a rationale for universal coronavirus vaccines, and highlights the importance of boosters as a strategy to broaden cross-protection to other coronaviruses different than SARS-CoV-2.
Supported by NIDA DP2 Avenir (1DP2DA051912-01) EREEP Grant Northwestern University
Heterogeneity in NK memory stem cells after Zika virus infection using single cell approaches
Authors: Kujur Weshely , Murillo Oscar , Adduri SitaRamaRaju , Mulik Sachin 
Published: January 2023

Abstract
Zika virus infection during pregnancy causes congenital defects such as microcephaly and ocular pathologies. Our lab previously reported that Zika virus infection in mice induces memory like NK cells that express CD27, had higher TCF-1 levels and possessed superior antiviral ability than naïve CD27+ NK cells. Moreover, these cells revealed stem like properties. We termed those NK cells with memory and stem cell like features as “NK memory stem cells (NKSCM). We hypothesized that NKSCM may contain subpopulations that differ in stemness program and antiviral capacity. To determine this, we performed single cell RNA-seq on NKSCM cells. Our analysis revealed 8 distinct clusters in NKSCM. Of these, Cluster 0 showed upregulation of Dusp1, Dusp2, Junb, Fosb while cluster 4 showed downregulation of Dusp1, Dusp2, Junb, Fosb genes. Cluster 5 had upregulation of proliferation associated genes (Mki67, Cdk1 and Pcna). Cluster 3 revealed downregulation of cytotoxic genes (Prf1, Gzma, Gzmb) while gene upregulation of Id3, Myb, Sell, Cd226 and Cxcr3 was evident in this subset. Similarly, other clusters showed differential gene expression profiles. Currently, our analysis is ongoing for single cell ATAC-seq datasets to determine epigenome at single cell level in these NKSCM clusters. Finally, the experiments are underway to functionally characterize these NKSCM subpopulations and to evaluate their role in Zika viral defense.
This work was supported by Department of Pulmonary Immunology, University of Texas Health Science Center at Tyler, TX, USA and American Association of Immunology Careers in Immunology Fellowship
Highly multiplexed, single-cell spatial phenotyping of Epstein-Barr virus infected tissues.
Authors: Nikulina Nadezhda , Ben Cheikh Bassem , Braubach Oliver , Fennell Eanna , Leahy Ciara , Pugh Matthew , Taylor Graham , Murray Paul 
Published: January 2023

Abstract
The Epstein-Barr virus (EBV), a group 1 carcinogen, is a causative factor in nine different cancers that cause ~165,000 deaths globally each year. Almost all adults carry EBV asymptomatically; the virus persisting primarily in rare memory B cells for the lifetime of the host. However, in some infected individuals, for reasons that are unknown, EBV may contribute to the development of cancer. The immune microenvironment in which infected cells reside may influence or trigger cancer development, but we know very little about how this occurs. We have begun to address these critical questions by deploying a spatial biology assay for comprehensive study of the immune microenvironment of EBV-infected tissues. Using AKOYA’s immunofluorescence based Phenocycler assay, we developed an antibody panel that can detect 40 or more different biomarkers in situ at single cell resolution. The panel contains antibodies directed at EBV infected cells at various stages of viral infection, as well as biomarkers for different immune cell lineages, immune cell activation states and tissue structures. This design allows simultaneous identification of infected immune cells within their microenvironment, while, crucially, maintaining spatial relationships between them. We have now set out to produce unbiased and comprehensive spatial phenotypic maps of normal and EBV infected human FFPE tissues. Our data will provide an initial understanding of how the immune microenvironment of virus-infected cells influences tissue pathology and, ultimately, the development of cancer.
Neurological chikungunya virus infection induces corpus callosum degeneration associated with resident immune cell activation and peripheral immune cell infiltration
Authors: Knight Audrey Claire , Nagel Jonathan , Anderson Elizabeth J , Atkins Hannah M , Montgomery Stephanie , Baxter Victoria K 
Published: January 2023

Abstract
Chikungunya virus (CHIKV) is an Old World alphavirus that typically induces arthralgia and rash. However, CHIKV is also capable of infecting the central nervous system (CNS), resulting in encephalitis, myelitis, and peripheral neuropathy. Patients who survive acute infection typically have long lasting neurological effects. To date, an immunocompetent small animal model that recapitulates CHIKV CNS disease has not been established. To develop a mouse model of CNS CHIKV infection, we intracranially inoculated 4–6 week old C57BL/6J mice with 1 of 3 clinical CHIKV isolates: Asian lineage strains SM2013 and AF15561, and ECSA lineage strain SL15649. CHIKV CNS disease is most commonly reported in regions where ECSA lineage strains circulate; in agreement with observations in humans, SL15649 was associated with more severe neurological disease, while SM2013 induced minimal to no disease, and AF15561 induced an intermediate clinical phenotype. At 7dpi when mice demonstrated signs of CNS disease but infectious virus was cleared from the brain, significant corpus callosum degeneration was observed in mice infected with SL15649, and to a lesser extent AF15561. Recruited CD4+ T cells and B cells were significantly elevated in the corpus callosum of mice infected with SL15649 compared to mice infected with SM2013. SL1549 infected mice also had significant micro- and astrogliosis in this area. At 3dpi, prior to neurological signs, the corpus callosum in SL1549 infected mice did not show significant degeneration but did have an influx of peripheral immune cells and increased gliosis. These results suggest that the CNS manifestations of CHIKV are associated with corpus callosum degeneration driven by both peripheral and CNS resident immune cells.
Supported by the following grants: K01 OD026529 (VKB) T32AI0075151 (ACK) AAI Careers in Immunology Fellowship (VKB, ACK) UNC SOM Junior Investigator Development Award (VKB) UNC Department of Pathology and Laboratory Medicine (VKB)
Combined blockade of TNF-a and IFN-I receptors in experimental RSV infection: analysis of STAT protein modulation
Authors: Morris Dorothea R , Qu Yue , de Mello Aline Haas , Ivanciuc Teodora , Ansar Maria , Casola Antonella , Garofalo Roberto P 
Published: January 2023

Abstract
Respiratory syncytial virus (RSV) is the leading cause of acute lower respiratory illness in young children. Two cytokines key to RSV pathogenesis are tumor necrosis factor (TNF)-a and type-I interferon (IFN-I). Using a dual receptor antibody blockade of IFNAR1 with either TNFR1 or TNFR2, we previously reported that IFN-I and TNF-a synergistically contribute to clinical disease while also controlling peak lung viral replication in RSV-infected Balb/c mice. The objective of the current study was to investigate the possible effect of TNF-a/IFN-I on prolonged viral shedding and to dissect canonical signaling pathways that would likely mediate the above-described activity. We found that viral titer in the lung of all treatment groups was comparable to the RSV control mice at days six and seven post-infection, indicating that lack of these cytokine receptor signaling was not involved in prolonged viral shedding. We then determined expression levels of total lung STAT1, STAT2, STAT3 and their phosphorylated forms as well as IRF1, pIRF3, IRF7, and IRF9 by western blot. Though RSV/IFNAR1, RSV/TNFR1, and RSV/TNFR2 demonstrated variations in protein expression for several of these signaling molecules, none of these proteins were uniquely modulated by the dual receptor blockade. These findings collectively demonstrate that while TNF-a/IFN-I receptor signaling controls peak viral replication in the lung, they have no effect on extended viral shedding and that STAT1, STAT2, and STAT3 signaling appears to be dispensable for the enhanced viral replication observed in the dual receptor blockade treated mice.
Supported by grants from AI062885, AI25434, Clinical and Translational Science Award [(NRSA (TL1) Training Core TL1TR001440)],Supported by UTMB McLaughlin Fellowship
Understanding innate immune cell-mediated inflammation in UV-induced skin injury in lupus-prone mice.
Authors: Maz Mitra Padideh , Shi Hong , Wolf Sonya , Estadt Shannon Nicole , Reddy Alayka , Kahlenberg J Michelle 
Published: January 2023

Abstract
Systemic lupus erythematosus (SLE) is a heterogeneous autoimmune disorder characterized by sensitivity to ultraviolet (UV) light and overproduction of type I interferons (IFNs), in which skin is affected in up to 70% of total SLE patients. While UV light is one of few well-known triggers of SLE, precise mechanisms governing UV-mediated inflammation are incompletely understood. Our lab recently performed single-cell analysis of cutaneous lupus lesions, showing an increase in activated monocyte-derived CD16+ dendritic cells (DCs) in both lesional and non-lesional skin of lupus patients compared with healthy controls. With the goal of understanding the functionality of this subset, we have turned to lupus-prone murine models. RNA sequencing of UV-exposed skin samples from lupus-prone NZM2328 (NZM), iNZM (lupus-prone mice lacking the type I IFN receptor) and wildtype Balb/c mice demonstrated upregulation of monocyte chemoattractants like CCL2 in NZM greater than iNZM and Balb/c, which was verified at the transcript and protein level of UV-irradiated human lupus keratinocytes compared with healthy controls. We further verified recruitment and differentiation of monocytes into monocyte-derived DCs (moDC) in UV-irradiated skin of NZM, iNZM, and Balb/c mice, as well as increased MHC class II expression in skin-recruited moDCs of chronically UV-exposed NZM mice greater than iNZM and Balb/c, suggesting an importance of type I IFN signaling in this process. Future studies will be directed at understanding mechanisms of monocyte recruitment and differentiation at sites of UV-mediated inflammation, as well as type I IFN effects on moDC antigen uptake and presentation.
Supported by grants from NIH (T32-GM007863, R01-AR071384, and K24-AR076975)
Pathogenic Tissue-Resident Memory T Cells in Exocrine Gland Chronic Graft-Versus-Host Disease
Authors: Costa da Silva Ana C , Sharma Rubina , Kim Clara H , Javaid Ayesha , Martin Daniel , Mays Jacqueline W 
Published: January 2023

Abstract
Salivary glands are frequently affected in chronic graft-versus-host disease (cGVHD), an autoimmune-like disease that occurs after allogeneic hematopoietic stem cell transplant resulting in tissue sclerosis and gland dysfunction. Alloreactive T cells have been identified as disease drivers, yet the function/pathogenicity of tissue-resident memory cells (Trm) in target organs remain unclear. In this study, we examined glandular Trm populations and tissue-specific signals that regulate Trm differentiation and functionality. scRNAseq analysis of human labial minor salivary glands (MSG) revealed enrichment of immune cell clusters in MSG of oral cGVHD patients. Focusing on T CD8 cells, we identified 4 clusters, one of them unique to affected patients which had low expression of CD69, but expressed other markers of Trm, including CD103, BLIMP1 and PD1. Additionally, this cluster was enriched for genes associated with cytotoxicity (IFNG, PRF1 and GZMB) and exhaustion (TIM3, LAG3 and TIGIT). Interestingly, this population showed high expression of CXCR6 and ligand-receptor analysis predicted CXCL16 an important signaling ligand that targets T CD8 cells. Further analysis implicates macrophages as the main source of CXCL16, suggesting their role in local accumulation of CXCR6+ Trm in the gland. Flow cytometry confirmed increased number of CD69+CD103±CD8+ Trm cells in cGVHD patients. Lymphocytic infiltration in MSG using IHC, spatially related the elevated frequency of CD8+IFNg+ T cells within interacinar stroma and damaged acini. The importance of Trm in site-specific cGVHD still needs further investigation. Based on current data, site-specific Trm cells may represent a druggable target in the therapy of refractory local cGVHD.
Supported by NIDCR (1ZIADE000747-05)
Responses of injured nerve in CD137L knockout mice following sciatic nerve crush-induced neuropathic pain
Authors: Cao Ling , Di Nucci Bruno Souza , Wakley Alexa , Ohara Kinuyo 
Published: January 2023

Abstract
Previously, we showed that CD137L, a costimulatory molecule is involved in the development of neuropathic pain following sciatic nerve crush (SNC). CD137L knockout (KO) mice displayed reduced SNC-induced pain-like behaviors and faster recovery following SNC. To determine the responses at the nerve crush site, we further examined the phenotypic and inflammatory changes of injured sciatic nerve in both CD137L KO mice and wild type (WT) C57BL/6 mice. Both CD137L KO and WT mice underwent either SNC or sham surgery. Sample collection occurred at days 0 (naïve), 3, 7, 14, 28, and 56 post-surgery. First, the femur-level segment of sciatic nerve containing the injury site was analyzed via whole mount immunohistochemistry. At the baseline, CD137L KO mice showed higher staining intensity for IB4 (non-peptidergic c-fibers) compared to WT mice. In WT mice but not CD137L KO mice, SNC significantly increased staining intensity for CGRP (all times; peptidergic c-fibers) and IB4 (Days 3 and 28). These increases were not observed in the corresponding CD137L KO mice. Second, selected inflammatory markers were measured via real time qRT-PCR. At baseline, CD137L KO mice expressed significantly lower levels of TNFα, yet higher levels of IL-10. SNC induced higher levels of elevation of IL-6, CCL3, CCL4, and Arg-1 (all peaked at day 3) in CD137L KO mice compared to WT mice. WT but not CD137L KO mice, showed significant up-regulation of CCL5 (throughout the time course) and CXCL1 (peaked at day 14). Therefore, we propose that CD137L-mediated local inflammatory responses may contribute to SNC induced pain-like behaviors in part by affecting the phenotype of injured nerve.
Supported by NIH/NINDS R01NS098426 (Cao), COBRE Histology and Imaging Core funded by NIH/NIGMS P20GM103643 (Meng), and the Kahn Family Foundation through the Carmen Pettapiece Student Research Fellowship (Souza Di Nucci).
MicroRNA control of inflammatory and regulatory T cells in CNS autoimmunity
Authors: Gopal Murugaiyan , Fujiwara Mai , Raheja Radhika , Garo Lucien , Ajay Amrendra , Chitnis Tanuja , Weiner Howard L , Gandhi Roopali 
Published: January 2023

Abstract
A disequilibrium between immunosuppressive regulatory T cells (Tregs) and inflammatory interleukin (IL)-17-producing Th17 cells is a hallmark of autoimmune diseases, including multiple sclerosis (MS). However, molecular mechanisms underlying Treg and Th17 imbalance in the central nervous system (CNS) autoimmunity remain largely unclear and thus identifying factors which drive this imbalance is of high clinical interest. Recently, we found a major disease-promoting role for microRNA-92a (miR-92a) in CNS autoimmunity. MiR-92a is elevated in experimental autoimmune encephalomyelitis (EAE), and its loss attenuates EAE. Mechanistically, miR-92a mediates EAE susceptibility in a T cell-intrinsic manner by restricting Treg induction and suppressive capacity, while supporting Th17 responses by directly repressing the transcription factor, Foxo1. Correspondingly, miR-92a inhibitor therapy ameliorates EAE by modulating the balance between Tregs and Th17 cells. Analogous to mice, miR-92a is elevated in MS patient CD4+ T cells, and miR-92a silencing in patient T cells promotes Treg development while limiting Th17 differentiation. Together, our results identify a previously unknown function by which miR-92a drives CNS autoimmunity via sustaining the Treg/Th17 imbalance and implicate miR-92a as a potential therapeutic target for MS.
Supported by NIH R01 R01AI127853 NMSS RG-1507-05164
Secretome of adipose-derived mesenchymal stem cells establish anti-inflammatory milieu by developing M2b/c macrophage
Authors: Yoon Taejun , Ko Eunhee , Park Yong-Beom 
Published: January 2023

Abstract
Mesenchymal stem cells (MSCs) express immunomodulatory and anti-inflammatory effect toward immune system with several mechanisms such as secreting bioactive molecules. Distinct mechanisms of their immune actions, however, are not fully understood. As one of the important cells in immune system, macrophages play a vital role in innate immunity with two functional phenotypes, classically activated (M1) and alternatively activated (M2) macrophages. In this study, immunomodulatory ability of adipose-derived MSCs (AD-MSCs) secretome toward peritoneal macrophage was studied. Secretome was concentrated from human AD-MSCs using 3K TFF membrane. Murine peritoneal macrophages were cultured for 24 hours with secretome of AD-MSCs and 4 hours with LPS. Macrophages were analyzed by flow cytometry, ELISA, and real-time PCR. LC-MS/MS proteomics analysis of secretome was performed and acquired protein sets were analyzed by DAVID Bioinformatics. Secretome of AD-MSCs increased cell viability of peritoneal macrophages. Secretome treated peritoneal macrophages resulted in reduced M1 markers such as CD86, iNOS, and TNFα and increased M2 markers such as CD206, Arginase-1, and IL-10. Also, peritoneal macrophage expressed upregulation of mRNA level of LIGHT and MerTK, M2b and M2c markers, respectively. 83 proteins were identified in secretome according to the proteomics analysis. Several proteins among them were selected as candidates after confirmation of inhibiting the effect of secretome with neutralizing antibodies. In conclusion, secretome of AD-MSCs induced polarization of peritoneal macrophages from M1 macrophages to M2 macrophages and made them create anti-inflammatory milieu by several key secreted proteins.
Supported by the National Research Foundation of Korea(NRF) grant funded by the Korea government(MSIT) (2021R1F1A1049630) and the Ministry of Health and Welfare, Republic of Korea (HI14C1324)
The long noncoding RNA LUCAT1 promotes immune gene expression in human macrophages
Authors: Vergara Kevin E , Lal Sneha , Cattley Richard , Hawse William F , Atianand Maninjay 
Published: January 2023

Abstract
Long noncoding RNAs (lncRNA) are transcripts larger than 200 nucleotides that mediate their molecular function in an RNA-dependent manner by regulating gene expression at either the transcriptional or post-transcriptional level. lncRNAs play vital roles in many biological processes; however, their functions in immune cells are largely unexplored. Here we describe the expression, function and the mechanism of action of LUCAT1 (Lung cancer associated transcript 1; SCAL1) in human macrophages. While elevated LUCAT1 expression is observed across many cancers, its cellular function in physiological contexts remains unknown. We show that Toll-like receptor 4 (TLR4) and PMA upregulates LUCAT1 expression in human monocyte-derived macrophages and THP1 cells through ERK1/2 signaling. Using 3’-RACE, long-range PCR and RNA FISH, we demonstrate that the LUCAT1 gene transcribes an abundant, 2562-nucleotides long nuclear-localized RNA. Transient perturbation of the nuclear LUCAT1 using antisense oligonucleotides followed by RNA sequencing and protein assays revealed that LUCAT1 promotes the expression of cytokines and chemokines (CSF3, IL24 and CXCL5), transcription factors (NR4A1 and NR4A3), matrix metalloproteinases (MMP1, MMP3 and MMP19) and others in human macrophages. Using siRNA-mediated knockdown of LUCAT1-interacting nuclear proteins, we show that RBMX contributes to the molecular function of LUCAT1, whereas SFPQ controls the biogenesis of the mature LUCAT1. Further, the nuclear LUCAT1 is chromatin-associated, and acts at the level of gene transcription. Collectively, in this study we have uncovered the function of LUCAT1 in macrophages, underscoring the critical roles of lncRNAs in the immune system.