bluuebunny/crossref_metadata_2025_split huggingface dataset:
The PGI2 signaling pathway decreases glycolysis and mitochondria respiration in mouse Th2 cells
Authors: Zhou Weisong , Zhang Jian , Chowdhury Nowrin U , Norlander Allison E , Toki Shinji , Newcomb Dawn C , Peebles R Stokes 
Published: January 2023

Abstract
Prostaglandin I2 (PGI2) is a lipid molecule produced in the cyclooxygenase (COX) metabolic pathway and regulates T cell function and inflammation. We have previously shown that PGI2 inhibited type 2 cytokine production by Th2 cells in vitro and in vivo. To further investigate the mechanism of the inhibitory effect of PGI2 on Th2 differentiation, we determined the effect of the PGI2 analog cicaprost on T cell metabolism under Th2 conditions. Mouse naïve CD4 T cells from the spleen were activated with antibodies against CD3 and CD28 under Th2 polarization conditions and treated with cicaprost or vehicle for 4 days. Seahorse assays measured cellular glycolysis and mitochondria respiration. We found that cicaprost significantly decreased basal and maximum glycolysis in Th2 cells. Furthermore, when we performed Seahorse assays on the cells rested for 2 days after the initial 4 day T cell activation, we discovered that cicaprost not only suppressed basal glycolysis and glycolytic reserve but also dose-dependently inhibited basal and maximum mitochondria respiration. These results suggest that the PGI2 signaling pathway inhibits Th2 cell metabolism, providing a possible mechanism for PGI2-mediated inhibition of Th2 differentiation and Th2 cell-dependent immune disorders.
The Department of Veterans Affairs BX004299 (RSP), NIH AI095227 (RSP), NIH AI111820 (RSP), NIH AI145265 (RSP), NIH AI124456 (RSP), NIH AI145397 (RSP).
Contributions of the PD-1/PD-L1 pathway to Borrelia persistence and inflammation
Authors: Helble Jennifer D , McCarthy Julie E , Starnbach Michael N , Hu Linden T 
Published: January 2023

Abstract
Borrelia burgdorferi, the causative agent of Lyme disease, establishes a long-term infection inside its mammalian hosts. Despite the continued presence of the bacteria in animal models of disease, inflammation is transitory and resolves spontaneously. Exhausted T cells are present in many long-term infections and mediate a balance between pathogen clearance and preventing tissue damage resulting from excess inflammation. Indeed, we have found that CD4+ T cells in the draining inguinal and popliteal lymph nodes, as well as the heart and ankle joint, significantly upregulate the immunoinhibitory molecule PD-1 following B. burgdorferi infection. While PD-1 expression on T cells is a strong indication of CD4+ T cell exhaustion, we have extended our findings by assessing expression of a ligand for PD-1, PD-L1, on antigen presenting cells in vitro and in vivo following B. burgdorferi infection. We have shown that not only is PD-L1 is significantly upregulated on both bone marrow derived macrophages (BMDMs) and dendritic cells (BMDCs) following B. burgdorferi stimulation, but is also upregulated on macrophages and a subset of conventional dendritic cells (cDC2s) in the draining lymph nodes following in vivo infection. We are currently testing if mice deficient in either PD-1 or PD-L1 exhibit changes in ankle joint or heart inflammation compared to wild-type mice, which would suggest that the PD-1/PD-L1 axis is critical for regulating inflammation during B. burgdorferi infection. Together, these data will allow us to determine how this immune checkpoint regulates inflammation during B. burgdorferi infection.
Supported by grants from the NIH (R01 AI150157), the Harvard Medical School Fairbairn Family Lyme Research Initiative, and the Tufts University School of Medicine Natalie Zucker Research Center for Women’s Scholars Award.
Cutaneous CD4+ T cell subsets promote distinct outcomes in human keratinocytes and fibroblasts
Authors: Morawski Peter A , DeBerg Hannah , Fahning Mitch , Stefani Caroline , Lacy-Hulbert Adam , Gratz Iris , Campbell Daniel J 
Published: January 2023

Abstract
Epithelial, stromal, and immune cells in the skin cooperate to ensure appropriate response to pathogens and environmental damage. Recent single cell transcriptional analyses of healthy and fibrotic human skin revealed a wide range of phenotypes that arise in response to distinct cytokine milieu. Cutaneous T cells promote host defense and the subsequent resolution of inflammation and repair of damaged skin through the production of cytokines, but the precise mechanisms T cells use to impact the differentiation and function of skin cells involved in inflammatory and tissue-repair responses remain understudied. To assess the functional capacity of cutaneous lymphocytes we isolated skin-tropic and skin-resident T helper cells including a novel population of pro-fibrotic migratory tissue resident memory cells we recently discovered. We measured both quantitative T cell cytokine production following TCR stimulation, and the impact of T cell derived cytokines on human keratinocyte and fibroblast transcriptional profile and morphology. We find that phenotypically and functionally distinct circulating and skin-resident T cell subsets have the capacity to promote discrete cytokine-dependent transcriptional and cellular outcomes in the skin including the induction of metabolic, proliferative, and inflammatory gene profiles. Thus, cutaneous T cells support host-protective and tissue-repair responses through direct activity on keratinocytes and dermal fibroblasts.
Lymph node-targeted long-acting butyrate micelles induce regulatory immune modulation
Authors: Cao Shijie , Wang Ruyi , Bashir Mohamed Elfatih H , Su Yanlin , Sabados Matthew , Hesser Lauren A , Nagler Cathryn R , Hubbell Jeffrey A 
Published: January 2023

Abstract
Regulatory T cells (Tregs) play essential roles in maintaining immune homeostasis and preventing autoimmunity and allergy. Peripherally derived Tregs are induced most efficiently in the lymph nodes (LNs), yet current therapies are not effectively delivered to these sites. The short-chain fatty acid butyrate facilitates extrathymic differentiation of Tregs and has been considered a promising therapeutic candidate. However, there are challenges to its use, including its foul odor, quick metabolism, and low efficiency with traditional oral delivery. Herein, we designed butyrate-prodrug micelles that deliver butyrate to peripheral LNs through subcutaneous (SC) administration. We have developed two polymeric micelles, one with a neutral charge (termed NtL-ButM) and one with a negative charge (termed Neg-ButM), which both contain 28 wt% butyrate and have similar sizes of ~40 nm. In a biodistribution study, we observed that Neg-ButM, not NtL-ButM, accumulated in draining LNs for over one month. At the cellular level, the Neg-ButM were taken up mostly by the macrophages and dendritic cells in the LNs and inhibited LPS-induced activation of those cells. In antibiotic-treated mice, three weekly doses of Neg-ButM significantly increased the numbers of CD25+ Foxp3+ CD4+ Tregs in draining LNs compared to either PBS or NtL-ButM treatment. Furthermore, we demonstrated that, when combined with intragastrically administered peanut proteins, the SC-injected Neg-ButM protected from anaphylactic responses in a mouse model of peanut allergy. This approach provides a potential Treg-based therapy against food allergies.
Supported by a seed grant from the Chicago Immunoengineering Innovation Center of the University of Chicago.
Practical Solutions to Improve Animal Study Reproducibility
Authors: Ibsen Eric 
Published: January 2023

Abstract
A 2012 study published by Amgen researchers found that 90% of the in vivo oncology studies were not reproducible. Historically, 95% of post-phase III oncology clinical trial therapies failed to receive regulatory approval due to a lack of demonstrated efficacy, not toxicity. The overwhelming lack of reproducible data underscores the dire need to critically examine our current standard animal study conduct processes for areas of potential improvement. In today’s competitive climate, researchers must identify the causes of poor animal study irreproducibility and implement meaningful and effective solutions. Recently, questions and debates concerning the validity of preclinical science data have resulted in multiple published reviews on the problem. Fortunately, as covered in this presentation, cost-effective technology exists to mitigate this “Crisis of Animal Study Irreproducibility”. This presentation covers:
The factors contributing to poor animal study reproducibility.
Publicly-available and online resources and practically-applicable concepts, such as the ARRIVE guidelines, PHISPS protocols, and the N3R Design Assistant.
Attendees receive first-hand experiences with a variety of animal study conduct approaches and commercially-available animal study workflow software applications, along with a summary of each tool’s direct and indirect impacts on the causes of animal study irreproducibility.
Exploration of how these concepts are applied in one Animal Study Workflow Software to solve problems, improve animal study reproducibility and faithfully preserve detailed study results for future access.
Development of a serum-free, chemically-defined workflow solution for T cell culture
Authors: Lopes Vanda , Quintero Zayda , Ni Jessie 
Published: January 2023

Abstract
Cancer immunotherapy has become one of the most promising fields in the treatment of oncologic related disorders, by harnessing the power of T-cells. Expansion and/or differentiation of cells with the right therapeutic profiles, in a consistent and safe manner, requires the use of defined cell culture reagents. One major cause of variability is serum, with its undefined composition. The presence of undefined reagents can lead to the inability to reproduce results between labs, and stop the progression from discoveries to clinical applications. Here we introduce a chemically-defined serum substitute and a low DMSO cryopreservation solution specially designed for T cells. After identifying the essential cell property parameters, and establishing appropriate cell culture assays, we initiated formulation optimization. We were able to identify the critical components required for appropriate cell expansion and cryopreservation. We were able to develop a chemically-defined solution, comparable in performance to serum, while bringing the benefits of being fully defined. We were also able to develop a low DMSO cryopreservation solution, tailored for T cell applications, comparable in performance to home brew or other commercially available solutions containing 10% DMSO. Our development work was allowed to gain in depth knowledge of the impact of each component in T cell biology, allowing us at BioLegend to provide custom tailored solutions. The development of defined reagents provides a smooth transition and reproducibility between research and clinical development.
Alloantigen-specific Chimeric Antigen Receptor Regulatory T cell therapy in non-human primate islet transplantation
Authors: Ellis Gavin I , Coker Kimberly E , Winn Delaine W , Deng Mosha Z , Shukla Divanshu , Bhoj Vijay , Milone Michael C , Wang Wei , Liu Chengyang , Naji Ali , Duran-Struuck Raimon , Riley James L 
Published: January 2023

Abstract
Though islet transplantation has emerged as a promising therapeutic for type 1 diabetes, the immunosuppressive agents that prevent rejection carry significant risk of morbidity. Adoptive transfer of chimeric antigen receptor (CAR) Tregs has the potential to protect allografts and transform transplantation medicine in the way that CAR T cells have revolutionized the treatment of cancer. However, there lacks a large animal model in which to optimize the safety and efficacy of CAR Treg therapy. Therefore, we developed methods to sort, transduce, and expand Cynomolgus macaque CAR Tregs recognizing the human/non-human primate cross-reactive alloantigen Bw6. Our novel pan-primate artificial antigen presenting cells expanded Tregs to clinical sized doses of &gt;500 million cells from as few as 25k sorted cells. Expanded Tregs expressed protein and epigenetic markers indicative of stable CAR Tregs and displayed suppressor function without cytotoxicity. CAR Tregs transferred into an antigen negative recipient could be detected for 7 days in peripheral blood and 1 month in bone marrow. When transferred to a diabetic recipient simultaneously with allogeneic Bw6+ islets, islets were partially protected from rejection as measured by increased control of blood glucose with insulin treatment. Transferred Tregs persisted in peripheral blood for &gt;34 days while displaying markers of stable, activated Tregs including CAR, FoxP3, and Helios. Importantly, we could not detect CAR+ FoxP3− effector T cells which would target the graft and expedite graft rejection. Our results provide a proof of concept for CAR Treg adoptive therapy in a large animal model and establish a framework for optimization of therapy before a clinical trial in humans.
Paired Immunoglobulin-Like Receptors Impact The Differentiation of Reparative Macrophages Following Allogeneic Challenge
Authors: Warunek Jordan , Lucas Anna , Mathews Lisa , Ossart Jason , Oberbarnscheidt Martin H , Lakkis Fadi G , Turnquist Heth R 
Published: January 2023

Abstract
Monocytes differentiate into CD206+ CD301+ reparative macrophages (MΦs) that orchestrate tissue repair in response to damage signals and cytokines. Yet, transplanting allogeneic materials produces a novel situation where innate allorecognition of mismatched MHCI may disrupt normal MΦ responses. B6 monocyte paired immunoglobulin-like receptor (PIR)-A3 recognizes BALB/c H2-Dd to generate pro-inflammatory dendritic cells that promote kidney and heart transplant rejection. Conversely, monocyte PIR-B binding to self MHCI negatively regulates PIR-A signals. How PIRs shape MΦ differentiation after Tx was not understood, so we tested the hypothesis that recognition of self-versus-allogeneic MHCI by PIRs divergently modulates MΦ differentiation. Monocyte-derived MΦ differentiation in wild type (WT), Rag2−/− γc−/−, Pira−/− and Pirb−/− B6 mice was assessed by flow cytometry 3 days after exposure to 20×106 radiation-damaged BALB/c allogeneic (allo) or B6 syngeneic (syn) splenocytes administered intraperitoneally. Syn materials predominantly stimulated monocyte differentiation into phenotypically reparative MΦs. Allo cells instead transformed monocytes exclusively into pro-inflammatory Ly6chi CD86hi MΦs. Rag2−/− γc−/− mice behaved similarly. IL-33 released from injured cells metabolically reprograms infiltrating monocytes and MΦs to support their differentiation into CD206+ CD301+ MΦs. Interestingly, Pirb−/− mice display greatly reduced C206+ CD301+ MΦs, which correlated with their decreased expression of the IL-33 receptor. Our data provide important insights into how innate allorecognition by PIR-A disrupts the typical generation of reparative MΦs in response to injured self that is supported by PIR-B.
Supported by 5T32AI074490-14
Vista blockade reprograms tumor myeloid cells and relieves adaptive resistance
Authors: Lines J Louise , Schaafsma Evelien , Croteau Walburga , Nowak Elizabeth C , ElTanbouly Mohamed , Sarde Aurélien , Cheng Chao , Noelle Randolph J 
Published: January 2023

Abstract
treatment. Significantly, VISTA shows broad expression in many cancer types, with enhanced expression upon development of adaptive immune checkpoint resistance. In the CT26 colon cancer model, growth of small tumors (40mm3), are retarded by anti-VISTA monotherapy, and are readily rejected following anti-CTLA4/anti-PD-L1 (CP)therapy. In contrast, very large tumors (&gt;600mm3), are refractory to CP therapy. However, combinatorial therapy of CP with anti-VISTA leads to rejection of about half the tumors. Both in monotherapy and combination therapy the mechanisms of enhanced anti-tumor immunity were investigated using scRNAseq, multiplex image analysis and flow cytometry of the tumor immune infiltrate. In both treatment models, anti-VISTA upregulated stimulated myeloid antigen-presentation pathways and reduced myeloid mediated suppression. Multi-spectral imaging revealed an anti-VISTA stimulated increase in contacts between CD8+ T cells and myeloid cells, further supporting the notion of increased antigen presentation upon anti-VISTA treatment. Transcriptional analysis of antigen-specific CD8 T cells showed that anti-VISTA therapy induced T cell pathways highly distinct from the anti-exhaustion effects of CP. These data suggest the non-redundancy of CP and anti-VISTA treatment, and support the use of these treatments in combination to overcome the adaptive resistance seen in contemporary treatments involving PD-1 and/or CTLA-4.
Research was supported by NIH grants R01AR070760 (R.J.N.), R01CA214062 (R.J.N.), and 1R21CA227996 (C.C.) and the Cancer Prevention Research Institute of Texas (CPRIT) (RR180061 to C.C.). C.C. is a CPRIT Scholar in Cancer Research.
Glycogen synthase kinase (GSK-3) synergizes with PD-1/PDL1 blockade to generate super-armed CD8 killers against tumors
Authors: Issa Mark E , Krueger Janna , Kazanova Alexandra , Taylor Alison , Rudd Christopher E 
Published: January 2023

Abstract
Immune checkpoint blockade (ICB) of negative co-receptors on T-cells such as programmed cell death-1 (PD-1) is promising for the treatment of cancer. Despite success, the poor prognosis for most patients highlights the need for novel clinical interventions. We have shown that the kinase, glycogen synthase kinase-3 (GSK-3) negatively regulates T-cell activation due to altered PD-1 and LAG-3 expression (Taylor et al., 2016 Immunity; Rudd et al., 2019 Cell Reports). GSK-3 inhibition (GSK-3i) is as effective as anti-PD-1 in controlling the growth of melanoma (Taylor et al., 2017 Can Res; Krueger and Rudd, Immunity 2017; Stelle et al., 2021 iScience). GSK-3i increases Tbet (Tbx21) transcription, which inhibits PD-1/LAG-3 transcription, while increasing granzyme B (GZMB) and interferon gamma (IFNγ). Here, we show that Gsk3−/− mice and small molecule inhibitors (SMIs) synergize with anti-PD-1 to eliminate melanomas that are resistant to anti-PD-1 monotherapy. Transcriptomic profiling showed that GSK-3 × PD-1 cooperativity was characterized by a specific increase in a family of different granzymes (7/12 GZM genes out of a data base of 20,500 potential genes). Some GZMs have been characterized and others not, but as a family, this increased armory of GZMs in CD8+ T-cells is expected to greatly enhance tumor killing. Overall, our data shows the PD-1 × GSK-3 synergy in limiting tumor growth is due to a specific set of cytolysis mediators needed for tumor killing.
Pro-tumoral role of Granzyme B to aid in invasion and metastasis.
Authors: Tibbs Ellis J , Cao Xuefang 
Published: January 2023

Abstract
The granule exocytosis pathway has long been known to be involved in potent responses to pathogens and cancers. This is executed by the action of Perforin and a family of granule-associated serine proteases known as Granzymes (Gzm). Many studies have shown that these proteases exert their effects by killing affected cells, executing the major function by Cytotoxic T cells (Tc) and Natural Killer cells expressing GzmB. Studies have found that the tumor cells ability to create a suppressive environment is dependent on Gzm expression by Regulatory T lymphocytes (Treg). Bird et al published data showing GzmB aids in the transmigration of Tcs, however the ability of GzmB to aid in the migration of non-T cells has not been investigated. We hypothesize that Granzyme B secretion may cause extracellular matrix remodeling in the tumor microenvironment, aiding in the outgrowth of the tumor cells via promoting invasion or metastasis.
We test our hypothesis by measuring tumor invasion in vitro utilizing the xCelligence transmigration system. We found that culturing B16 cells with recombinant GzmB from Biolegend or Peprotech on Matrigel resulted in a 31% and 47% increase in cellular invasion, respectively. Also, the rate of invasion resulted in a 35% and 61% increase, respectively. We also tested our hypothesis in vivo with a preliminary study using Treg-specific GzmB KO mice (Foxp3cre GzmBfl/fl). We depleted CD8+ T cells, injected B16 cells intravenously, and activated Tregs. We found a 48% decrease in metastatic nests in mice deficient of GzmB in Treg cells after 8 days and a 37% decrease after 16 days compared to WT. These findings shed light on the pro-migratory ability of secretory GzmB and helps us understand an unassuming role of GzmB as a pro-tumoral factor.
Supported by funds through NIH (RO1 HL135325, T32 AI095190, P30 CA134274) and Maryland Department of Health’s Cigarette Restitution Fund Program.
Transcriptomic and epigenetic profiling of tumor-associated monocyte function
Authors: Contreras Cristina F , Kaczanowska Sabina , Kaplan Rosandra N 
Published: January 2023

Abstract
Monocytes are innate immune cells recognized for their ability to play both tumor permissive and surveillant roles in cancer. Circulating classical monocytes (CD14+ CD16−) can home to the tumor and suppress other immune cells through various mechanisms, including the production of arginase and the release of reactive oxygen species (ROS). Conversely, patrolling nonclassical monocytes (CD14− CD16+) have been shown to employ processes such as phagocytosis and presentation of tumor antigens to prevent metastasis. This heterogeneous monocyte function is influenced by tumor-derived factors that are released during cancer development and progression. Phenotypic and transcriptional alterations in circulating monocytes and other myeloid cells in patients with solid tumors have been reported and associated with poor clinical outcomes. However, perturbations of specific monocyte functions in the setting of solid tumors have not been well explored. Here we present a characterization of monocytes by coupling flow cytometry-based functional assays with sequencing (Func-seq). Healthy donor primary monocytes and monocytic cell lines were used to examine the production of ROS and arginase in response to osteosarcoma-conditioned media and monocyte-mediated phagocytosis of osteosarcoma cells. Bulk RNA-seq and ATAC-seq were performed on FACS-sorted populations to compare differentially expressed genes and establish transcriptomic and epigenetic signatures associated with monocyte-mediated immunosuppression and tumor-cell phagocytosis. The incorporation of functional selection into -omic characterization provides insights into monocyte behavior and potential therapeutic targets to alter their activity in solid tumors.
Funding: US National Institutes of Health grants ZIA BC 011332 and ZIA BC 011855 and NCI cancer moonshot.