bluuebunny/crossref_metadata_2025_split huggingface dataset:
Aiolos restrains the acquisition of cytotoxic features by CD4+ T cells
Authors: Jones Devin M , Read Kaitlin A , Pokhrel Srijana , Hales Emily , Eisele Caprice , Warren Robert T , Collins Patrick , Freud Aharon , Oestreich Kenneth J 
Published: January 2023

Abstract
Classically, CD4+ T cells have been defined as “helper” type cells, providing aid to other immune cell populations via the secretion of cytokines and direct cell-cell interactions. More recently, a subset of CD4+ T cells with cytotoxic capabilities, termed CD4+ cytotoxic T lymphocytes (CD4-CTLs), have been observed in both mice and humans, performing protective functions in the settings of infection and cancer while conversely contributing to the pathogenesis of autoimmunity. Despite their well-documented importance in several disease contexts, the complete mechanisms that underlie their differentiation and function remain unknown. Here, we identify the Ikaros family member, Aiolos, as a novel regulator of CD4+ CTL differentiation and function. We find that Aiolos deficiency results in increased expression of key CD4+ CTL transcription factors and effector molecules both in vitro and in an in vivo murine model of influenza infection. Mechanistically, we find that Aiolos deficiency results in increased IL-2/STAT5 signaling, supporting a repressive role for Aiolos in CD4+ CTL differentiation via negative regulation of the IL-2/STAT5 pathway. Collectively, this work identifies Aiolos as a novel negative regulator of CD4+ cytotoxic gene programming, and thus may represent a therapeutic target for the treatment of autoimmune diseases and enhanced anti-tumor immunity.
Sponsored by a grant from NIAID (NIH-RO1 AI134972) and funds through The Ohio State University College of Medicine
The role of ERK2 in regulating germinal center B cell fate decisions
Authors: Negron Austin Joseph , Abdul-Baki Nawal Samer , Perez Rebecca , Perez Christina , Bartsch Brenden L , Forsthuber Thomas G 
Published: January 2023

Abstract
The regulation of B cell activation, differentiation, and survival is a crucial control point for the maintenance of B cell tolerance in the germinal center (GC) and involves the precise coordination of several complex intracellular signaling pathways. Dysregulation of these pathways can lead to the generation and persistence of autoreactive memory B cells and antibody-secreting cells (ASCs), resulting in autoimmune disease. The mitogen-activated protein kinase (MAPK) pathway is central to signals required for GC B cell selection and differentiation, such as those received through the B cell receptor (BCR) and C-X-C chemokine receptor type 4 (CXCR4), respectively. Extracellular signal-regulated kinases 1 and 2 (ERK1/2), important effectors of this pathway, play a major role in B cell activation and differentiation. Though ERK2 specifically regulates the pro-apoptotic protein, Bmf, its role in regulating GC B cell fate is unknown. Our results show that peripheral Erk2-deficient B cells express DEC205, which is associated with follicular T helper cell interactions and CD80, which is involved in co-stimulation and immunoglobulin class switching. Thus, we hypothesize that ERK2 regulates several aspects of GC B cell selection. Our results elucidate key molecular mechanisms underlying the role of ERK2 in B cell differentiation, potentially offering new approaches to treating B cell-driven autoimmune diseases.
Supported by grant R21AI44731 from the NIH/NIAID (TGF), and partially funded by the UTSA president’s distinguished researcher fellowship and NIH/NIGMS RISE grant GM60655 (A.N.).
BCL6-dependent TCF-1+ progenitor cells maintain effector and helper CD4 T cell responses to persistent antigen
Authors: Xia Yu , Sandor Katalin , Pai Joy A , Daniel Bence , Raju Saravanan , Wu Renee , Hsiung Sunnie , Qi Yanyan , Yangdon Tenzin , Okamoto Mariko , Schreiber Robert D , Murphy Kenneth M , Satpathy Ansuman , Egawa Takeshi 
Published: January 2023

Abstract
Shortly after priming, the fate of activated CD4 T cells is segregated into BCL6+ follicular helper T (Tfh) and BCL6− effector (Teff) cells. However, it remains unknown how these subsets are sustained in the presence of chronic antigen stimulation. Using a combination of single cell- and population-based approaches, we show that in chronic viral infection, activated CD4 T cells differentiate into BCL6-dependent TCF-1+ progenitor cells with superior capacity to expand and give rise to both Teff and Tfh. They share properties with progenitor-exhausted CD8 T cells and are required for the continued generation of Teff cells as antigen persists. In response to tumors, an analogous CD4 T cell population develops in draining lymph nodes. Our study reveals the heterogeneity and plasticity of CD4 T cells upon encountering persistent antigen and highlights their population dynamics through a stable bipotent intermediate state.
Effect of Tritrichomonas foetus, Tf-31 (a cattle isolate) and Tff6 (a feline isolate) on TLR signaling pathway in cervical epithelial cells.
Authors: Kucknoor Ashwini S , Lavergne Randall 
Published: January 2023

Abstract
Bovine trichomonosis is caused by Tritrichomonas foetus, which is widely prevalent in 33 US states, and is a reportable disease due to potential huge economic losses to cattle industry. T. foetus also causes chronic large-bowel diarrhoea in cats, complicated not only by its chronicity but for its difficulty to treat, with few registered or effective products available. T. foetus persistently colonize the lower genital tract of bulls but usually do not cause either major clinical signs or inflammation while vaginal inflammation is a common disease of the dairy cows’ reproductive tract. Mechanisms of immune response to this pathogen are poorly studied. This study for the first time, reports the effect of T. foetus; Tf-31-cattle isolate and Tff6-cat isolate, on activation of toll-like receptors (TLRs) and inflammatory response in vaginal epithelial cells. The expression levels of serial effectors related to the TLRs-MyD88/NF-κB signaling pathway (TLR2, TLR4, MyD88, NF-κB, IL-1β, IL-6, TNF-α, and IL-10) were measured with real-time polymerase chain reaction (RT-PCR), ELISA, and Western blot analyses. Of the 6 cell-membrane TLRs tested, TLR2 and TLR4 were activated by cells, as noted by increased mRNA expression levels of TLR2 and TLR4 in both Tf31 and Tff6 treated cervical epithelial cells relative to control cells. ELISA assays showed up-regulation on IL-10, both in Tf-31 and Tff6 treated epithelial cells. Treatment with inhibitors of MyD88 signaling pathway abolished the upregulation of downstream cytokines. These data indicate that T. foetus isolates activate TLR-MyD88 signaling pathway to induce innate immune response. However, further detailed in vitro and in vivo studies are required to confirm these findings.
Supported by Biology department
Breakdown of the blood brain barrier in Experimental Cerebral Malaria is caused by CD8+ T cells with low affinity TCRs
Authors: Cornwall Douglas , Jacobs Jesica , Kolawole Elizabeth M , Evavold Brian D , Lamb Tracey J 
Published: January 2023

Abstract
Malaria is a worldwide health concern killing over 400,000 people each year, many of them children under 5 years old. One severe outcome of Plasmodium infection is cerebral malaria caused by a dysregulation of the immune response in the central nervous system. Using an established mouse model of cerebral malaria called experimental cerebral malaria(ECM) using Plasmodium berghei ANKA (PbA) infections of C57BL/6J mice have shown that breakdown of the blood brain barrier is mediated by IFN-g producing CD8 T cells. Here, using novel techniques to determine the affinity of the MHC:peptide:TCR interaction we have discovered that although there are high affinity cells in the population trafficking to the CNS, surprisingly a large majority of these PbA-reactive pathogenic CD8 T cells have low affinity TCRs, and a large percentage of them cannot be detected by tetramer. Importantly we show that these low affinity cells are also producing IFN-g suggesting they are pathogenic. Through adoptive transfer experiments we show that high and low affinity T cells remain stable in their affinity profile. Furthermore we show here that low affinity cells are needed for breakdown of the blood brain barrier – CD8 T cells bearing high affinity TCRs for the GAP50 peptide of Plasmodium have an impaired ability to cause ECM compared to low affinity CD8 T cells. This work demonstrates that low affinity cells are the major component of the immunopathology causing ECM when infected with PbA and contrasts with current dogma which suggests that high affinity cells are the main cells responding in these types if infections.
Supported by grants from NIH (R01 AI167422)
Epithelial HNF4A shapes the intraepithelial lymphocyte compartment via direct regulation of immune signaling molecules
Authors: Lei Xuqiu , Carneiro Natalia Ketelut , Galia Liraz , Wilson Ruth , Vierbuchen Tim , Chen Yongzhi , Ward Doyle , Fitzgerald Katherine A 
Published: January 2023

Abstract
Hepatocyte Nuclear Factor 4A (HNF4A) is a highly conserved orphan nuclear receptor that has been associated with ulcerative colitis. Mice lacking Hnf4a specifically in the intestinal epithelial cells (IECs) are more susceptible to DSS-induced colitis and develop spontaneous colonic inflammation with aging. However, the mechanisms how HNF4A maintains intestinal homeostasis and protects against colitis remain unclear. Here we demonstrate that epithelial HNF4A shapes the intraepithelial lymphocyte (IEL) compartment by selectively promoting the development of natural IEL subsets that are TCRgd+ or TCRab+ CD8aa+. Mechanistically, HNF4A serves as a transcription factor that directly regulates epithelial expression of immune signaling molecules involved in the IEC-IEL crosstalk. Those molecules include the butyrophilin-like (Btnl) molecules Btnl1 and Btnl6. Further, while HNF4A is stably expressed along the intestinal tract, these molecules are more abundant in the small intestine, which correlates with the distribution of natural IELs. We propose HNF4A paralog, HNF4G, which is restricted to the small intestine, cooperates with HNF4A in driving expression of signaling molecules and accounts for the geographic specificity. Moreover, HNF4A-BTNL regulatory axis is conserved in Caco2 cells of human colorectal origin. Collectively, our findings underscore the importance of HNF4A as a conserved transcription factor controlling IEC-IEL crosstalk and suggest that HNF4A maintains intestinal homeostasis through regulation of the IEL compartment.
Spatially mapping T cell receptors and transcriptomes
Authors: Liu Sophia , Iorgulescu Julian Bryan , Li Shuqiang , Barrera-Lopez Irving A , Borji Mehdi , Shanmugam Vignesh , Morriss Julia W , Garcia Zoe N , Murray Evan , Braun David A , Livak Kenneth J , Wu Catherine J , Chen Fei 
Published: January 2023

Abstract
T cells mediate antigen-specific immune responses to disease through the specificity and diversity of their T-cell receptors (TCRs). Although determining the spatial distributions of T cell clonotypes in tissues is essential to understanding T cell maturation and behavior, spatial sequencing methods remain unable to profile the TCR repertoire. We have developed Slide-TCR-seq, a 10-μm-resolution method to sequence whole transcriptomes and TCRs within the spatial context of intact tissues. We first confirmed the ability of Slide-TCR-seq to map the characteristic architecture of T cells and their receptors in mouse spleen. In human reactive lymph node and tonsil, we identified spatially distinct TCR repertoires both between germinal center (GC) and non-GC regions as well as between GCs. We then spatially profiled T cell clonotypes and their infiltration in renal cell carcinoma specimens before and following immune checkpoint blockade, which identified uniquely distributed T cell clonotypes and transcriptional states, highlighting the infiltration and expression heterogeneity both between clonotypes and within a T cell clonotype. Our method is anticipated to facilitate dissection of the immune microenvironment, yielding insights into the complex spatial relationships between T cell clonotypes, neighboring cell types, and gene expression that drive T cell development and response to disease.
This work was supported in part by the NIH U19AI082630, the Dana-Farber/Harvard Cancer Center Kidney Cancer SPORE (NCI P50CA101942), and the NCI IMAT grant.
A rapid and fully automated in vitro micronucleus assay using imaging flow cytometry and convolutional neural network analysis
Authors: Mendoza Maria Gracia Garcia , Sutton Alex , Kong Raymond , Rodrigues Matthew , Pugsley Haley 
Published: January 2023

Abstract
Micronuclei (MN) originate from whole chromosomes or chromosome fragments that lag behind during cell division and fail to be incorporated into one of the two main nuclei. As a result, scoring MN using the well-established in vitro micronucleus assay evaluates the ability of chemicals or other agents to induce DNA damage. This technique is typically performed by manual microscopy, which can be time-consuming and prone to variability. Additionally, automated methods lack cytoplasmic visualization when using slide-scanning microscopy, and conventional flow cytometry doesn’t provide visual confirmation of MN.
The ImageStream®X Mk II (ISX) imaging flow cytometer combines the high-resolution imagery of microscopy with conventional flow cytometry’s speed and statistical robustness in a single system. Previously, we developed a rapid and automated MN assay based on high-throughput image capture and feature-based image analysis using IDEAS® Software. However, the feature-based analysis was not readily applicable to multiple cell lines and chemicals, so we developed a deep learning method based on convolutional neural networks to score imaging flow cytometry data in both the cytokinesis-blocked and unblocked versions of the MN assay using Amnis® AI Software.
Our current study validates our previously established assay and analyses using three different chemicals (Mitomycin C, Cyclophosphamide, and Eugenol) and three different cell lines (TK6, L5178Y, CHO-K1). Here, we demonstrate how using Amnis AI to score imagery acquired on the ISX provides a rapid and fully automated in vitro MN assay with improved accuracy, reproducibility, and time-to-results in toxicity and biodosimetry applications across multiple cell lines.
Derivation of a parsimonious Tuberculosis gene signature using the digital NanoString nCounter platform.
Authors: Kaipilyawar Vaishnavi , Zhao Yue , Wang Xutao , Joseph Noyal M , Babu Senbagavalli Prakash , Hochberg Natasha S , Sarkar Sonali , Horsburgh Charles , Ellner Jerrold J , Johnson William Evan , Salgame Padmini 
Published: January 2023

Abstract

Rationale
Currently, there are several host immune response-based biomarkers to triage individuals suspected to have Tuberculosis (TB), to predict TB treatment failure and to predict risk of progression from latent TB infection (LTBI) to TB disease. However, validation of these biomarkers in diverse populations, with well-defined clinical endpoints, has been scarce. Moreover, a robust platform to validate TB biomarkers is essential to the development of a universal diagnostic or prognostic clinical test.

Objectives
NanoString nCounter technology is an amplification-free, highly sensitive digital detection platform that directly measures gene expression of up to 800 targets. Here, we investigated whether NanoString technology could serve as a platform to extensively validate existing TB biomarkers, in a new cohort from South India.

Methods
A NanoString Codeset that probes 107 genes (NS-TB107), comprising 12 published TB gene signatures and 6 house-keeping genes was developed to quantify RNA from whole blood samples of 80 pulmonary TB patients and 102 individuals with LTBI. The TBSignatureProfiler software was used to score samples for each signature. An ensemble of machine-learning algorithms were used to derive a parsimonious gene signature.

Results
Gene signatures present in NS-TB107 had statistically significant power to distinguish TB from LTBI in our cohort. A six-gene set (NANO6) derived from this cumulative data, when tested on 10 publicly-available transcriptomic datasets, was highly diagnostic for active TB.

Conclusions
The NanoString nCounter system provides an efficient platform to validate a panel of existing TB biomarkers and to derive a highly-diagnostic parsimonious candidate TB biomarker.
This project has been funded by Award No. USB-31150-XX-13 of the US Civilian Research & Development Foundation (CRDF Global) and by the National Science Foundation under Cooperative Agreement No. OISE-9531011 with Federal funds from the Government of India’s (GOI) Department of Biotechnology (DBT), the Indian Council of Medical Research (ICMR), the United States National Institutes of Health (NIH), National Institute of Allergy and Infectious Diseases (NIAID), Office of AIDS Research (OAR), and distributed in part by CRDF Global.
StarBright Dyes:-Build bigger better panels with superior dyes excitable by the violet and ultraviolet lasers.
Authors: Blundell Michael , Sanderson Sharon , Huang Leon , Cichewicz Magdalena , Li Lei 
Published: January 2023

Abstract
StarBright UltraViolet and StarBright Violet Dyes are the brightest ultraviolet and violet laser excitable fluorescent dyes specifically developed for flow cytometry. Their narrow excitation and emission mean that despite the increased brightness there is no loss of resolution due to excessive spillover and spreading. StarBright Dyes can be pre-mixed, fixed and have no requirement for a special buffer when multiplexing, providing researchers with the flexibility to build bigger, better panels.
Here we demonstrate this flexibility by comparing freshly made and premixed antibody cocktails from Bio-Rad Antibodies to immunophenotype human peripheral blood. Cells were stained using common staining buffers with acquisition on the ZE5 Cell Analyzer to show how bigger multiplex panels can be built with ease.
Expanding the size of multicolor panels with ease with new StarBright Blue and StarBright Yellow Dyes.
Authors: Blundell Michael , Sanderson Sharon , Huang Leon , Cichewicz Magdalena , Li Lei 
Published: January 2023

Abstract
Bio-Rad is committed to improving multicolor flow cytometry and is excited to announce a novel series of StarBright Dyes, exceptionally bright fluorescent nanoparticles developed for use with flow cytometry antibodies. These bright dyes have the additional benefits of narrow excitation and emission, high stability, minimal lot-to-lot variation, no requirement for a special buffer and can be fixed making them ideal for inclusion in multicolor immunophenotyping panels containing multiple antibodies conjugated to StarBright Dyes and traditional fluorescent dyes.
Here we show how new blue (488 nm) and yellow/green (561 nm) laser excitable StarBright Dyes can be incorporated into multicolor panels in both conventional and full spectrum flow cytometry to give unrivalled resolution, flexibility and choice. Available by the end of 2022 these new dyes will be available conjugated to our highly cited, validated antibodies.
DEFINING THE ROLE FOR THE GUT MICROBIOME IN THE CLINICAL EFFICACY OF SULFASALAZINE THERAPY FOR IBD ASSOCIATED SPONDYLOARTHRITIS
Authors: Lima Svetlana Ferreira , Rupert Amanda , Viladomiu Monica , Marderstein Andrew , Pires Silvia , Putzel Gregory , Woo Viola , Funez-dePagnier Gabriela , Jin Wen-Bing , Guo Chun-Jun , Scherl Ellen , Longman Randy S 
Published: January 2023

Abstract
Joint inflammation (spondyloarthritis, SpA), is the most common extra-intestinal manifestation of inflammatory bowel disease (IBD), but the specific role for therapies targeting SpA is not well defined. Sulfasalazine (SAS) is a prodrug composed of two chemical moieties, 5-aminosalicylate and the anti-folate antibiotic sulfapyridine, with efficacy in peripheral arthritis. Our study aims to evaluate the role for the gut microbiome in clinical response of SpA to SAS and to define microbial mechanisms targeted by SAS.
We longitudinally follow IBD patients with SpA who have a medical indication for SAS therapy. Clinical data and fecal samples from 22 patients were collected before initiation of SAS and at week 12 after initiation of SAS. The fecal microbiome of SAS-responders was distinct from that observed in non-responders and 6 pre-treatment microbial markers (including the short chain fatty acid (SCFA) producer Faecalibacterium prausnitzii) predicted SAS-response (AUC=0.9). Fecal metabolome of SAS responders had lower thymine and higher deoxyuridine compared to non-responders consistent with evidence of a folate trap in response to SAS treatment. SAS therapy in SPF mouse-model of chemically-induced colitis alleviated colitis in GPR 109a- and 43-dependent fashion consistent with a synergistic role for SCFA. In vitro and in vivo models revealed SAS direct regulation of F. prausnitzii transcription and metabolic function and its impact on host immune response.
Collectively, these findings highlight the potential role for microbial diagnostics to improve SAS efficacy, and drug modulation of microbial markers to potentiate therapy for IBD patients with SpA.
Supported by New York Crohn’s Foundation
Bispecific CD8 Treg modulators regulate a novel regulatory CD8 T cell network and eliminate pathogenic CD4 T cells in live cell co-culture system
Authors: Gardell Jennifer L , Crane Courtney , Bowser Justin , Fasnacht Rachael , Julien Susan , Maurer Meghan E , Templeton Megan , Therriault Jon , Yang Su Jung , Swiderek Kristine M 
Published: January 2023

Abstract

INTRODUCTION
Others have described a subset of CD8 T cells (CD8 Treg) with immunosuppressive characteristics in inflammatory disease settings. CD8 Treg activation through canonical T cell receptors results in their oligoclonal expansion and perforin dependent elimination of pathogenic CD4 T cells. We have described the CD8 Treg network in Celiac patients and its potential to eliminate pathogenic CD4 T cells. Here we describe bispecific CD8 Treg modulators that activate CD8 Tregs, resulting in pathogenic CD4 T cell death.

METHODS
Novel bispecific CD8 Treg modulators were tested for target specificity by Octet and cell binding. The functional impact of CD8 Treg modulators was evaluated using flow and Luminex assays, and in a live cell co-culture system using Celiac patient derived CD8 Tregs and gliadin activated CD4 T cell targets. CD8 Treg modulators with specific molecular formats were then ranked for functional efficacy.

RESULTS
We tested a panel of bispecific CD8 Treg modulators with monovalent binding affinities in the low nanomolar range that selectively bind the CD8 Treg subset. Within 48 hours of CD8 Treg modulator addition to T cell co-cultures, CD8 Tregs had a rapid increase of cytolytic markers and degranulation in response to CD4 T cell targets. We observed a concomitant increase of activated CD4 T cell death and decrease of pro-inflammatory cytokines.

CONCLUSIONS
Our results indicate that a targeted approach to engage and activate CD8 Treg reduces pathogenic CD4 T cells. As this newly described CD8 Treg cell network is observed in many autoimmune diseases, we postulate that bispecific CD8 T cell modulators may represent a novel, selective and broadly applicable therapeutic approach for the treatment of human autoimmune diseases.