bluuebunny/crossref_metadata_2025_split huggingface dataset:
The Role of Chemokine-Like Receptor-1 in Recruitment and Activation of Myeloid Cells During WNV Encephalitis
Authors: Kwok Keven , Durrant Douglas 
Published: November 2024

Abstract
Recent increases in West Nile Neuroinvasive Disease (WNND) outbreaks in the US underscore the urgent need to understand CNS viral clearance mechanisms. In response to WNV, neurons secrete chemokines (CCL2, CCL5, and CXCL10), facilitating leukocyte entry into the CNS. CNS-infiltrating macrophages amplify this signal, recruiting and activating neuroprotective leukocytes, limiting harmful accumulation, and aiding repair. CMKLR1 promotes CNS inflammatory responses during infectious and autoimmune diseases via inducing macrophage and DC migration following chemerin activation. However, the specific role of chemerin/CMKLR1 in WNND remains unclear. WT and CMKLR1-/- mice were infected with WNV to investigate this. WNV-infected CMKLR1-/- mice exhibited a slight increase in mortality but elevated clinical scores and reduced recovery compared to WT. We hypothesize that CMKLR1 limits WNV pathogenesis by regulating CNS recruitment and activation of inflammatory myeloid cells during WNV encephalitis. Assessing key inflammatory mediators and activation markers in CNS-resident and infiltrating cells in WT and CMKLR1-/- animals will elucidate how CMKLR1 influences the recruitment/activation of these cells, clarifying processes resulting in infection resolution and neural repair.
Unveiling the immune surveillance role of extracellular vesicles
Authors: Kumari Puja , Vasudevan Swathy , Russo Ashley , Wright Skylar , Fraile-Ágreda Víctor , Krajewski Dylan , Jellison Evan , Rubio Ignacio , Bauer Michael , Shimoyama Atsushi , Fukase Koichi , Zhang Yuanpeng , Pachter Joel , Kailasan Vanaja Sivapriya , Rathinam Vijay 
Published: November 2024

Abstract
Intracellular immune surveillance for systemic microbial components during homeostasis and infections governs host physiology and immunity. However, a long-standing question is how circulating microbial ligands become accessible to intracellular receptors. Here, we show a role for host-derived extracellular vesicles (EVs) in this process; human and murine plasma- and cell culture-derived EVs have an intrinsic capacity to bind bacterial lipopolysaccharide (LPS). Remarkably, circulating host EVs capture blood-borne LPS in vivo, and the LPS-laden EVs confer cytosolic access for LPS, triggering noncanonical inflammasome activation of GSDMD and pyroptosis. Mechanistically, the interaction between EV lipid bilayer and LPS’ lipid A underlies EV capture of LPS. Furthermore, the intracellular transfer of LPS by EVs is mediated by CD14-dependent receptor-mediated endocytosis. Overall, this study demonstrates that EVs capture and escort systemic LPS to the cytosol licensing inflammasome responses, uncovering EVs as a previously unrecognized link between systemic pathogen-associated molecular pattern (PAMPs) and intracellular immune surveillance.
O-GlcNAc transferase plays dual antiviral roles by integrating innate immunity and lipid metabolism
Authors: Dong Hong , Liang Chenxi , Yount Jacob , Liu Shan-Lu , Wen Haitao 
Published: November 2024

Abstract
Challenge of host cells with viral pathogens induces robust metabolic reprogramming; however, whether viral component(s) is detected by metabolic enzyme remains completely unknown. Our previous study identified the O-GlcNAc transferase (OGT), an important glucose metabolic enzyme, as a crucial mediator of MAVS-dependent antiviral response. Here, by studying a mouse model with enzyme-inactive OGT, we demonstrate an enzyme-independent function of OGT in limiting influenza A virus (IAV) infection, in addition to its enzyme-dependent effect on MAVS. Biochemical studies reveal a critical antiviral effect of OGT binding with IAV genomic RNA via its N-terminal tetratricopeptide repeats-4 domain. Binding with viral RNA causes a robust translocation of nuclear OGT to cytosolic lipid droplets (LDs) and subsequently antagonizes LD accumulation via destabilizing perilipin 2. This LD-targeting effect of OGT limits IAV replication without altering MAVS signaling. Our findings reveal OGT as a multifaceted metabolic sensor by integrating MAVS-dependent innate immune signaling and lipid metabolism.
B cell-derived IL-10 is an autocrine survival factor and maintains immune homeostasis in the spleen
Authors: Chen Chuxiao , Ramaswami Bala , Mohib Kanishka , Rothstein David 
Published: November 2024

Abstract
IL-10 is a key anti-inflammatory cytokine and IL-10 KO mice develop colitis. Mice with a global IL-10 KO or constitutive B cell-specific IL-10 KO (CD19-Cre.IL-10fl/fl) have otherwise normal B cells. Surprisingly, we found that within 14-21 days, mice with a tamoxifen (Tam) -induced B cell specific deletion of IL-10R (hCD20-ERT2-Cre.IL-10Rfl/fl) had a 50% decrease in splenic B cell number vs. Cre-controls. B cell frequency was unchanged due to a concomitant fall in both T cells and innate cells resulting in a 58% fall in splenocyte number. Plasma cells, CD4+ and CD8+ T cells, and neutrophils were disproportionately decreased, resulting in a 20-40% reduction in frequency. A similar but somewhat less severe loss of B cells/splenocytes was seen in Tam-treated hCD20-ERT2-Cre.IL-10fl/fl mice. Adoptive transfer competition studies in immunized µMT hosts treated with Tam revealed a selective loss of hCD20-ERT2-Cre.IL-10fl/fl vs. CD19-Cre.IL-10fl/fl B cells. Furthermore, by day 21, acute loss of B cell IL-10 or IL-10R resulted in ~20% weight loss and colon length, consistent with colitis (pathology pending). In conclusion, IL-10 is an autocrine B cell survival factor and acute loss of B cell IL-10 or IL-10R is associated with a marked contraction of B cells and associated collapse in T cells and innate cells in the spleen, resulting in immune dysregulation and colitis. This requirement for B cell IL-10 has not been previously recognized because it is compensated for developmentally.
Loss of UCP2 in CD4+ T cell promotes Th1/2 Pathogenic Phenotype
Authors: Choi Seungho , Chakraborty Paramita , Hedley Zachariah , Oberholtzer Nathaniel , Mills Stephanie , Mehrotra Shihar 
Published: November 2024

Abstract
Uncoupling protein 2 (UCP2) is an anion carrier protein located in the mitochondrial inner membrane. Mitochondria are the major sites for energy metabolism which is tightly linked with immune cell fate. In the present study, we have investigated UCP2’s function in T cells against murine melanoma model and the experimental autoimmune encephalomyelitis (EAE) model. Loss of UCP2 in CD4+ T cells showed a significant rise in the Th2/Th1 ratio. These cells exhibited an improved Th2 phenotype with increased production of nitric oxide (NO), and Inducible nitric oxide synthase (iNOS). Therefore, CD4+ T cells were treated with NOC-18, a source of NO, or L-NMMA, inhibitor for NOS to demonstrate NO-mediated Th2 polarization. Furthermore, Treatment of Ucp2 siRNA or, Genipin (inhibitor for UCP2) mimicked the effect of UCP2-/- CD4+ T cell with more cytotoxic markers and Th2/Th1 double positive cells. Transcriptomic and metabolomic analysis revealed the lower response of calcium influx by TCR activation and increased NLRF3 activation by uric acid in UCP2 deficient CD4+ T cells. Adoptive transfer of Genipin-treated Tyrp CD4+ cells exhibited superior tumor control as compared to Th1 and Th2, according to the findings. However, in UCP2-/- mice, the severity of EAE worsened, and heterogenic Th2 cells were infiltrated into the spinal cord. Our findings imply that the loss of UCP2 in CD4 increases Th2 polarization with a cytotoxic phenotype and may be relevant to various illnesses.
The TGF-β -- Eomes Axis Maintains Stemness in TCF-1+ CD8+ T Cells by Restricting Tissue Egress From Secondary Lymphoid Tissues.
Authors: Im John , Ma Chaoyu , Srinivasan Saranya , Zhang Nu 
Published: November 2024

Abstract
T-cell exhaustion is characterized by dysfunctional cytotoxic T lymphocytes (CTLs) that express inhibitory and exhaustion markers, such as programmed cell death 1 (PD-1) and CD101, with a reduced capacity to clear antigens. Exhausted CTLs arise from ‘stem-like’ precursors that express T cell factor-1 (TCF-1) and have a unique gene signature similar to hematopoietic stem cells. These self-renewing precursors reside in secondary lymphoid tissues, requiring Transforming growth factor beta (TGF-β) to maintain their residency and mitigate their effector differentiation, partially via controlling the expression of a4 integrins. Downstream of TGF-β, expression of the transcription factor Eomes has been associated with greater levels of exhaustion and is highly upregulated in terminally exhausted cells. Using a chronic lymphocytic choriomeningitis virus (LCMV) infection model in wild-type B6 mice, we found that TGF-β inhibited Eomes expression in TCF-1+ stem-like CD8 T cells. And that deletion of Eomes largely corrected the defects in TCF-1+ stem-like T cells lacking TGF-β receptor signaling. Our data indicate that the TGF-β -- Eomes axis helps to maintain stem-like TCF1+ precursor cells within lymphoid tissues, and that this pathway could be targeted to reinvigorate exhausted CD8+ T cells.
Role of MNK/eIF4E pathway in CD4+ Th2 cytokine-induced lung fibrosis in chronic allergic asthma
Authors: Fattahi Fatemeh , Vallance Laura , Walker Natalie , Holden Julia , Huang Steven , Atasoy Ulus 
Published: November 2024

Abstract
Despite therapeutic advances in allergic asthma, the persistent challenge of airway remodeling still exists, necessitating further exploration. We aimed to investigate the influence of CD4+ Th2 cytokines on fibrosis associated with airway remodeling, with a focus on mesenchymal cell (MC) transformation.
Using both human and mouse lung MCs, we investigated the pivotal role of MNK/eIF4E pathway in regulating fibrous protein expression driven by CD4+ Th2 cytokines. To assess this, we employed western blot and flow cytometry studies, while Luminex assays measured cytokine and chemokine secretion from activated lung MCs responding to Th2 cytokines.
Treatment with Th2 cytokines significantly increased ATX expression in lung MCs, a response mitigated by the specific MNK1/2 inhibitor (eFT-508). Furthermore, this inhibitor not only abolished the phosphorylation of eIF4E but also reduced collagen levels within activated MCs induced by Th2 cytokines. Luminex assay results demonstrated a significant decrease in IL-6, CCL-2, and GM-CSF in the activated MCs upon inhibiting the MNK1/2 pathway. Additionally, eFT-508 exhibited a marked reduction in lymphocyte migration, utilizing the Boyden chamber transwell assay.
Our study reveals that targeting the MNK/eIF4E pathway holds promise in mitigating airway remodeling associated with MCs fibrosis and provides valuable molecular insights for future therapeutic interventions related to fibrosis in the airway remodeling of allergic asthma.
Harnessing temperature-dependent STING signaling to reprogram regulatory T cells and enhance effector T cell immunity.
Authors: Smith Rachael , Heintzman Darren , Fisher Emilie , Voss Kelsey , Ye Xiang , Toudji Yasmine , Chi Channing , Schulman Jacob , Rathmell Jeffrey , Wilson John 
Published: November 2024

Abstract
The cGAS-STING pathway plays a critical role in immune surveillance through its capacity to sense aberrantly localized DNA in the cytosol caused by infection, cancer cell death, or cell stress-induced self-DNA leakage. STING pathway activation ultimately results in the production of Type I Interferons (IFN-I) and other inflammatory cytokines that stimulate T cell immunity. However, the T cell-intrinsic effects of STING agonists have been underexplored, especially in regulatory T cells (Tregs), which have an opposing but crucial role in suppressing excessive inflammation. Unexpectedly, we have found that STING activation repolarizes Tregs to IFN-I producing Th1/Th9-like cells with enhanced antitumor function and reduced oxidative metabolism and suppressive capacity. Inflammation can also coincide with heat (i.e. fever-range temperature, FRT, 39°C), which can cause nuclear or mitochondrial self-DNA leakage, yet the effects of FRT on STING in Tregs are unknown. We were surprised to find that FRT strongly potentiates the effects of STING activation in Tregs and that FRT appears to prime cGAS/STING signaling. Together, our initial data suggest a novel axis linking FRT to STING signaling in inhibiting Treg immunosuppressive function and in promoting Treg transdifferentiation to inflammatory effector-like T cells. These exciting findings motivate further investigation to dissect the mechanisms intertwining STING activation and FRT and their effect on Treg function and metabolism.
BLIMP-1 and CEACAM1 cooperate to regulate human regulatory T cell proliferation and function
Authors: Ding Ying , Yu Aixin , Vujanac Milos , Copsel Sabrina , Moro Alejandro , Nivelo Luis , Tchitchek Nicolas , Rosenzwajg Michelle , Villarino Alejandro , Levy Robert , Koatzmann David , Malek Thomas 
Published: November 2024

Abstract
BLIMP-1, encoded by the PRDM1 gene, is a major transcriptional hub distinct from Foxp3 in human Tregs, but its mechanisms of action have not been fully determined. Here, using CRISPR/Cas9-mediated disruption of PRDM1, we show that BLIMP-1 limits Treg proliferation while supports Treg functional activity. PRDM1KO human Tregs are less suppressive and exhibit increased expansion in a humanized mouse model of xenogeneic GVHD. BLIMP-1 restrains Treg proliferation by lowering IL-2R signaling through downregulation of CD25 and by inhibiting responsiveness to TCR/CD28 signaling and mTOR activation through upregulation of CEACAM1, an immune checkpoint. Expression of CEACAM1 is induced by IL-2 in human, but not mouse, Tregs. Prolonged IL-2R signaling leads to optimal BLIMP-1 expression and chromatin opening of human CEACAM1, leading to high cell surface CEACAM1 through a 3’ regulatory region containing motifs for binding PRDM1 and STATs. Correspondingly, CEACAM1 is highly induced on Tregs from autoimmune patients undergoing low-dose IL-2 therapy, and these Tregs showed reduced proliferation. In vivo studies using xenogeneic GVHD mouse model show that CEACAM1KO human Tregs have higher suppressive activities, indicating a potential role of CEACAM1 in Treg function. Collectively, our findings uncover a novel IL-2-dependent regulatory loop whereby BLIMP-1 and CEACAM1 act to limit human Treg proliferation and contribute to Treg suppressive function.
Evaluation of a human peripheral blood NK humanized hIL-15 mouse model lacking murine Fc gamma receptors
Authors: Dubé Philip , Smith Nicholas , Buczek Monika , MacBride Megan , Andino Esther , Sack Emily , Freer Debra , Vedder Michelle , Bott Kathleen , Baskin Louise , Richardson Janell 
Published: November 2024

Abstract
Fc gamma receptors (FcγRs) on residual murine immune cells can interact with human IgG-based therapeutics and confound preclinical results. This is critical for therapeutics with human NK cell mediated antibody-dependent cellular cytotoxicity (ADCC) as their primary mode of action. We compared humanization, kinetics, and persistence of NK cell reconstitution in human IL-15-expressing mice with or without FcγRs using intraperitoneal (IP) or intravenous (IV) administration of cells from multiple donors.
Methods: Negatively selected human NK cells were engrafted into hIL-15 NOG or FcResolv™ hIL-15 NOG (FcγR knockout) with identical protocols. Human reconstitution, immune profile and persistence was evaluated via serial blood sampling and terminal collections for blood, spleen, and bone marrow.
Results: Human NK cell reconstitution and persistence was equivalent between strains. There was a significant difference in the overall chimerism in IV vs. IP administration (IV &gt; IP). There was no significant difference in % body weight, body condition scores, morbidity or mortality across study duration. NK cell persistence and kinetics were consistent between strains, administration, and donors, with a peak at 4-5 WPE and persistence to 12+ WPE in the blood. The dominant circulating NK phenotype was CD56+CD16+.
Conclusions: huNK-engrafted hIL-15 NOG and FcResolv hIL-15 NOG mice provide a 12+ week study window, with equivalent reconstitution, immune profiling and kinetics.
Evaluation of an IgG therapeutic efficacy in a humanized immune system mouse model lacking murine Fc gamma receptors
Authors: Baskin Louise , Smith Nicholas , Buczek Monika , MacBride Megan , Andino Esther , Sack Emily , Freer Debra , Vedder Michelle , Bott Kathleen , Richardson Janelle 
Published: November 2024

Abstract
Fc gamma receptors (FcγRs) on residual murine immune cells in humanized immune system (HIS) mice can interact with human IgG-based therapeutics and confound preclinical results. We assessed impact of murine FcγRs on anti-PD1 efficacy in HIS mice engrafted with lung adenocarcinoma cells treated with pembrolizumab or vehicle. We also present humanization results for the newly generated FcγR knockout NOG-EXL. Methods: HIS NOG (huNOG) or HIS FcγR knockout NOG mice (FcResolv™ huNOG) were made using identical protocols with CD34+ cells (3 shared donors). Reconstitution was evaluated in naïve animals and HCC827 cells were inoculated in remaining animals. From D7, mice were dosed twice weekly for 4 wks then euthanized for blood, spleen, and tumor analysis. Results: Humanization was equivalent between strains. Pembro treatment showed significant tumor growth inhibition in 1 donor in FcResolv huNOG, but not in donor-matched huNOG. Human TILs in pembro-treated mice were significantly different between the strains for all donors, with more CD8+ T cells and fewer TAMs in FcResolv huNOG compared to vehicle-treated mice, and no significant differences in huNOG. Murine TIL analysis showed differences in murine macrophage populations between strains. Conclusions: Anti-PD1-treated FcResolv huNOG mice show expected pharmacodynamic changes and donor-dependent efficacy, whereas pembro-treated huNOG mice showed neither, demonstrating the impact of murine FcγRs on antibody IgG-based therapeutics.
Assessment of SARS-CoV-2 Spike and NCAP antigen-specific memory B cell frequency and Ig class/subclass usage following natural infection and/or COVID-19 mRNA vaccination
Authors: Kirchenbaum Greg , Becza Noemi , Yao Lingling , Liu Zhigang , Chepke Jack , Lehman Paul 
Published: November 2024

Abstract
An adaptive humoral response can encompass multiple Ig classes/subclasses, each capable of contributing to host defense through different effector functions. Which Ig class/subclass are elicited in response to an antigen encounter is the consequence of context, such as site of antigen contact, presence of immunostimulatory motifs and the cytokine secretion signature of helper T cells. Moreover, the number of antigen encounters and duration of the ensuing immune responses can further diversify the memory B cell compartment’s epitope specificity, affinity and Ig class/subclass usage. Beyond characterization of Ig usage in blood, whether differences also exist in the Bmem compartment following multiple SARS-CoV-2 infections and/or COVID-19 vaccinations is still not well understood. Leveraging multiplexed B cell ImmunoSpot assays, the scope of this work is to characterize SARS-CoV-2 S- and N-specific Bmem and their respective Ig class/subclass usage in the context of complex immune exposure histories. In agreement with recent observations detailing the antibody response elicited following COVID-19 mRNA vaccination and/or breakthrough infection, alterations in the affinity, breadth and IgG subclass usage of S-specific Bmem was also observed. Collectively, our findings demonstrate the feasibility of performing in-depth characterization of Bmem in a streamlined pipeline that requires minimal cell material and serves to motivate usage of the multiplexed FluoroSpot platform.
Label-free morphometric characterization of T cells for cell and gene therapy research and development.
Authors: Teranishi Kazuki , Wagatsuma Keisuke , Uematsu Mika , Tsubouchi Asako , An Yuri , Tamoto Ryo , Yanagihashi Yuichi , Kawamura Yoko , Murata Yuri , Akai Satoru , Komoriya Kaoru , Schneider Greg , Nomaru Hiroko , Ota Sadao 
Published: November 2024

Abstract
Identification, characterization, and minimally invasive isolation of T cells is critical for developing cellular therapies. Here we used VisionSort, an artificial intelligence (AI)-driven cellular analysis and sorting platform, to characterize and isolate human T cell subsets with therapeutically relevant phenotypes, label-free. By capturing single-cell digital phenotypes, we characterized human T-cells and generated ‘ground truth’ functional profiles for 1) glycolysis level, 2) exhaustion state 3) activation or resting profiles and 4) viability. A set of machine-learning derived classifiers was generated to identify these phenotypic classes label-free. The classifiers showed area under the curve (AUC) performance ranges for detecting specific, phenotypically defined T-cell populations between 0.923 and 0.995. Using a combination of supervised and unsupervised machine learning approaches, we also show identification of follicular helper T (Tfh) cells from other CD4+ T cells based on morphology alone. When we analyzed the morphological profiles of T cell subsets by unsupervised machine learning using uniform maniform projection and approximation (UMAP), we found that both gamma delta (γδ) T cells were distinct from alpha beta (αβ) T cells and CD8+ T cells were distinct from CD4+ T cells. Here we report on the use of AI-based cytometry to characterize and isolate phenotypically defined T cell subsets label-free, with applications in basic life science and cell therapy R&D.
Advancements in Research Grade Biosimilars Enable the Study of Therapeutic Antibodies
Authors: Cantor Joseph , Gao Guo-Jian , Rigaud Stephanie , Wei Ai-Li , Bazirgan Omar , Verma Shilpi 
Published: November 2024

Abstract
Biosimilars for therapeutic antibodies contain identical variable sequences to FDA-approved
antibodies. Recombinant BD research-grade biosimilars can be used to model several important aspects of major antibody therapies in use today. Our biosimilars with the huIgG1 (N297A) Fc mutation exhibit greatly reduced background binding and clarify the identification/study of therapeutic antibody target cell subsets. These flow cytometry reagents are available with many fluorochrome options to facilitate deeper phenotyping using large panels. Secreted targets (e.g., TNF-α) of therapeutic antibodies can either be bound in solution or in a membrane/cell-associated state; biosimilars may also be useful in studying this context-dependent target engagement. Alternatively, wildtype huIgG1 Fc biosimilars in no-azide/endotoxin-free (NA/LE) formats permit comparative mechanism-of-action (MOA) studies. Finally, biosimilars can be used in pre-blocking experiments to identify non-competing and competing clones used to study membrane target regulation, or gate on cell subsets defined by these targets in models of pre-exposure to therapeutic antibodies. In sum, our research-grade biosimilar antibodies can empower multiple avenues of research on therapeutic antibodies, which in turn are having major clinical impacts across medicine.
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