bluuebunny/crossref_metadata_2025_split huggingface dataset:
CyTOF enables unprecedented resolution of intracellular readouts to reveal novel functional diversity of human T Cells
Authors: Mcconney Christina , Smith-Mahoney Erika , Cohen Michael , Belkina Anna , Selvanantham Thirumahal , King David , Loh Christina , Cappione Amedeo , Snyder-Cappione Jennifer 
Published: November 2024

Abstract
Measuring functional signatures at the single-cell level in a cross-lineage manner provides key insights into several facets of immunology, including the plasticity of innate T cells, T/B memory cell diversity following infection/vaccination, and the panoply of activities associated with cell therapy success/failure. To better understand the dynamics of the immune system, flow cytometry phenotyping panels often include intracellular markers of functional potential, such as cytokines, phosphorylation events, and transcription factors. Many of these readouts, including cytokines of the Th2/Treg lineages, have been historically difficult to detect in human cells using flow cytometry. We compared the signal resolution of three 13-parameter panels on a Cytek™ Aurora spectral cytometer and the CyTOF™ XT mass cytometer. Each panel measured surface and intracellular analytes; fluorescent spectral panels were designed with little/no spectral overlap. PBMC-derived data was analyzed using PhenoGraph clustering and visualized by opt-SNE. Data collected on CyTOF XT™ demonstrated superior signal resolution for a range of intracellular readouts, including cytokines IL-5, IL-10, and IL-13. Also, when better signal/noise enabled more accurate clustering, a higher number of immune cell subsets possessing unique signatures resulted from CyTOF datasets. Overall, cytometry by CyTOF provided clearer understanding of the immuno-diversity present in a sample in the context of unbiased analysis.
Chromatin remodeler ACTL6A in CD8+ T cells promotes antitumor immunity
Authors: Wu Bogang , Thant Win , Bitman Elena , Liu Ting , Ellisen Leif 
Published: November 2024

Abstract
Chromatin remodeling plays a pivotal role in dictating epigenetic and transcriptomic regulation governing cytotoxic T lymphocyte differentiation and effector function. However, the regulatory mechanisms underlying chromatin remodeler assembly and actions remains unclear. Actin-like gene family members have been implicated in the organization of chromatin landscapes, however their role in T cell functional regulation remains unknown. We identified Actin-like protein 6A/Actl6a as the most upregulated gene within actin-like family members upon CD8+ T cell activation across multiple species. Additionally, ACTL6A is highly expressed in human tumor-infiltrating CD8+ mitotic tissue-resident memory T cells, suggesting a functional importance of ACTL6A in this subset. Deletion of mouse Actl6a in mature T lymphocytes impairs immune responses to transplanted syngeneic mouse tumors. T cell-specific Actl6a deletion mice exhibit a reduced effector memory population and attenuated cytotoxicity in their tumor infiltrating CD8+ T cells. T cell adoptive transfer further corroborates a CD8+ T cell-intrinsic role of Actl6a on antitumor adaptive immunity. Transcriptomic/epigenetic investigations provide mechanistic insights into the action of ACTL6 in the CD8+ T cells regulation. Furthermore, our study suggests that pharmacological or genetic approaches to increase Actl6a expression in CD8+ T cells may have the potential to boost immune based anticancer therapies including CAR-T therapies.
introduction of expanded tools for spatial biology applications
Authors: Serafin Jen , Montoya Leticia , Harvey Austin , Voeun Mae , Clark Scott 
Published: November 2024

Abstract
Fluorescence labelling is a highly versatile approach to immunohistochemistry (IHC), offering the ability to detect numerous targets simultaneously with excellent sensitivity and specificity. Currently, the fluorescent labelling method utilized for IHC is the use of secondary antibodies. This multi-step process is restricted by targets that share the same species isotype, limiting the number of targets possible in one sample. Another method used for multiplex staining is cyclic labeling, the process of staining and stripping a tissue sample with different antibodies to detect multiple targets. This process is time-consuming and increases the risk of tissue damage or loss of antigenicity. Our new reagents will enable researchers to stain in fewer steps with preserved antigenicity, ensuring accurate and reliable detection of target antigens in tissue samples. One of the key benefits of these products is the ability to achieve higher-plex labeling, allowing researchers to multiplex with 8-10 biological targets on a single sample in one staining cocktail. This reduces the need for cyclic labeling, streamlines the workflow, and reduces overall time required for analysis. We provide a process that enables successful detection of multiple protein markers across diverse tissue organs while simultaneously detecting transcriptomic targets, providing comprehensive insights into complex biological processes and disease mechanisms. Intended for research use only.
Using single cell CITE-seq and high parameter flow cytometry to characterize human regulatory T cells
Authors: D'Cruz Louise , Shi Xiaoshan , Rezvan Ali , Sen Moen , Sakofsky Cynthia , Genero Gracia , Widmann Syephanie , Tyznik Aaron 
Published: November 2024

Abstract
Regulatory T cells (Tregs) are an essential population of T cells that are critical for maintenance of peripheral tolerance. Imbalanced Treg frequency or dysregulation of Treg function can lead to the development of autoimmune diseases. Thus, the modulation of Treg frequency or function has become an important therapeutical approach to treat autoimmune disorders. Despite increased study, whether distinct populations of Tregs with unique functions exist is unclear. Here, we used a combination of single-cell CITE-Seq (scCITE-Seq) and high parameter flow cytometry to resolve the heterogeneity of Tregs in humans. Using magnetic cell enrichment in conjunction with fluorescence-activated cell sorting (FACS), we isolated Tregs and performed downstream scCITE-Seq. By using more than a hundred antibody-oligo conjugates, we identified distinct populations of Tregs, uncovered the proteomics signature associated with these populations, and revealed donor-to-donor variability in the peripheral Treg compartment. The protein expression pattern and antigen density on Tregs revealed by scCITE-Seq correlated to that of high dimensional flow cytometry. Trajectory analysis segregated Treg subsets into distinct differentiation pathways with unique proteomic signatures ranging from naïve, activated to memory Tregs. These findings improve our understanding of the heterogeneity of Tregs and provide a single-cell proteomic atlas for dissecting the complex roles of Tregs in health and disease.
Ce3D-IBEX: Achieving Multiplex 3-dimensional Imaging for Deep Phenotyping of Cells in Tissues
Authors: Arroyo Armando , Ichise Hiroshi , Hor Jyh Liang , Chu Colin , Croteau Joshua , Radtke Andrea , Germain Ronald 
Published: November 2024

Abstract
2D multiplexed antibody-based imaging provides a framework to study cell-cell interactions in thin tissue sections but lacks the ability to interrogate spatial relationships in larger 3D micro-anatomical structures. Several methods for 3D imaging of optically cleared tissue exist, but current approaches do not provide a deep phenotyping capability as afforded by high content 2D imaging. Here,we developed a method to overcome this constraint by combining a fast hydrophilic tissue clearing technique, clearing-enhanced 3D (Ce3D), and the recently developed Iterative Bleaching Extends Multiplexity (IBEX) technique together with a robust image registration algorithm suite to create a highly multiplex 3D imaging method: Ce3D-IBEX. It has been used to analyze murine lung, lymph nodes, small intestine, retina, tumor, and cornea as well as human retina, kidney, liver, tonsil, and jejunum, yielding seamless views of large-scale tissue structures while enabling visualization of high dimensional phenotypic markers at single-cell resolution, permitting the identification of discrete cell subsets, quantification of protein expression level and their mutual spatial association within tissue structures. To date, 25+ markers have been visualized in a single 3D sample. This represents a major advance in the emerging field of highly multiplexed 3D imaging. With aims to open new avenues for characterization of spatial dynamics of immune cells in healthy and diseased tissue.
A comprehensive analytical validation framework for immunofluorescence assay design using CellScape precise spatial multiplexing
Authors: Braubach Oliver , Jackson Charle , Christians Arne , Boog Jannik , Ingalls Matthew , Meshik Xenia , Northcutt Adam , Cashion Kayla , Schwarz John 
Published: November 2024

Abstract
Multiplex immunofluorescence (mIF) is a powerful immunology research tool that enables deep phenotyping of cells within the tissue microenvironment. Careful optimization of multiplex antibody panels for specificity, sensitivity, and reproducibility, is crucial to achieving robust data and is a requirement for assay repeatability and standardization. Here, we describe our validation procedure for multiplex assay panels.
Experiments were performed using the CellScape™ platform for precise spatial multiplexing. To design a ready-to-use panel for immune cell phenotyping, individual antibodies were first screened for specificity and then optimized for labeling sensitivity. Signal-to-noise, inter-assay reproducibility, and intra-assay precision were evaluated in a multi-stage testing matrix to validate each antibody in isolation and assembled into a panel. Measurements of both signal and quantitative immune phenotyping across technical replicates showed a high degree of intra-assay precision and inter-assay reproducibility.
Pairing the CellScape platform with comprehensively validated antibody panels streamlined deep immune profiling. The 15-plex, ready-to-use VistaPlex™ immune profiling panel for human FFPE tissues was validated on human tonsil, lung, breast, and colon tissues demonstrating the ability to positively identify epithelial cells and 22 cellular immune phenotypes most critical to understanding tumor immune responses.
High-throughput and high-dimensional single-cell analysis of antigen-specific CD4+ T cells in COVID-19 and influenza
Authors: Guo Yuwan , Huang Chao , Jiang Ning 
Published: November 2024

Abstract
This study aims to elucidate the tissue-specific dynamics of antigen-specific CD4+ T cells in response to SARS-CoV-2 and influenza. We have developed TetTCRII-SeqHD, a novel high-throughput, high-dimensional single-cell analysis platform that utilizes barcoded tetramers for antigen-specific TCR sequencing. It enables simultaneous profiling of CD4+ T cells' antigen specificities, TCR sequences, gene and surface-protein expression in human tissues. Using this technology, we have profiled &gt;50,000 SARS-CoV-2-specific CD4+ T cells from lung, lung-associated lymph node, spleen, and blood.
Our results show distinct TCR repertoires and responses of CD4+ T cells in COVID-19 and influenza. Tissue-isolated CD4+ T cells display overlapping core signatures, yet distinct tissue cues influence their phenotype, pointing to site-specific functions. In COVID-19, unique transcriptional profiles of CD4+ T cells is characterized by a pronounced cytotoxic signature and a reduction in Treg, in contrast to flu-specific cells. We also observed that tissue context influences the Treg TCR repertoire, with clonal diversity tending to be more restricted in lung tissues compared to the spleen.
This study provides a comprehensive understanding of CD4+ T cell heterogeneity and function in tissues during viral infections. The novel TetTCRII-SeqHD platform presents an advancement in analyzing tissue-localized immune responses, offering potential implications for future immune research and vaccine strategies.
Streamlined 96-well plate method for simultaneous single-cell TCR immune receptor repertoire analysis and immunophenotyping
Authors: Chenchik Alex , Liu Tianbing , Makhanov Mikhail , Hu Dongfang , Kobzik Lester , Ghias Khadija , Diehl Paul 
Published: November 2024

Abstract
This study describes an efficient immune profiling method by FACS-sorting single cells in a 96-well plate to assess clonotype repertoire diversity, paired-chain information, and cell phenotypes in a single, multiplex RT-PCR-based assay without the use of any specialized equipment. It is a cost-effective alternative to conventional single-cell technologies such as microwell arrays or droplet microfluidics. The 96-well plate contains primers for either T-cell receptor (TCR) / or TCR / chains, together with primers for 30 key T-cell markers. T cells from PBMCs are sorted in a single-cell-per-well format. We conduct RT-PCR library prep using the DriverMapTM Adaptive Immune Receptor (AIR) Repertoire Profiling Assay and sequence the CDR3 region. The results show clonotype frequencies, chain pairing details for / and / chains, and T-cell subtype classifications based on gene expression profiling. This enables in-depth analysis of TCR gene rearrangements at the single-cell level, enhancing understanding of T cell development, proliferation, and clonality, which are important in studying cancer, immunodeficiencies, and autoimmune disorders. Additionally, we can effectively identify and characterize γδ T cells by integrating single-cell TCR sequencing with RNA sequencing data, potentially useful in the development of γδ cancer immunotherapies. Overall, this method provides an efficient way to simultaneously analyze clonotypes and immunophenotypes in a single assay.
Lactobacillus gasseri KBL697 ameliorates psoriatic inflammation through the modulation of gut-skin axis
Authors: Kim Jun-Hyeong , Lee Kiuk , Jang Sung Jae , Jo Eun Jung , Kim Daehwan , Cho Bo-Ram , Lim Sang Kyun , Ko GwangPyo 
Published: November 2024

Abstract
Psoriasis is a chronic inflammatory disease arising from aberrant interactions between various immune cells and cytokines. The current standard therapy includes topical steroids, methotrexate or anti-IL-17A/IL-23 antibodies. However, due to the limited efficacy and associated side effects, there is still an unmet medical need for the development of novel therapeutic agents for this disease.
KBL697 is a strain of Lactobacillus gasseri, isolated from the vaginal samples of healthy donors. We previously reported that KBL697 ameliorated the symptoms of an inflammatory bowel disease mouse model. At the molecular level, KBL697 decreased pro-inflammatory cytokines including IFN-gamma, IL-6, and IL-17A, while an anti-inflammatory cytokine IL-10 was increased.
In the current study, we tested KBL697 in pre-clinical models for psoriasis. Oral administration of KBL697 showed a significant beneficial effect in imiquimod-induced mouse models, judged by the skin inflammation, PASI score and immune profile change. Further studies revealed that IL-10 is induced by lipoteichoic acid (LTA) of KBL697, indicating the critical role of LTA/IL-10 in the anti-inflammatory function of KBL697. In the skin analysis, key cytokines in psoriasis, such as IL-17A, IL-23, and IL-36, are downregulated while genes for skin regeneration are upregulated, supporting the model for KBL697 working in the gut-skin axis. Therefore, KBL697 has great potential as a novel therapeutic agent for the treatment of psoriasis.
Siglec-E is a novel co-inhibitory receptor in the modulation of innate activation after transplantation
Authors: Rodrigues Lima Karina , Gassen Rodrigo , Liu Kaifeng , Ganchiku Yoshikazu , T. Ribas Guilherme , J. Borges Thiago , Riella Leonardo 
Published: November 2024

Abstract
Early after transplantation, inflammation and tissue injury are associated with significant activation of the innate immune cells, such as DCs. This activation drives adaptive immune responses, triggering acute rejection while hindering the development of transplant tolerance. Current immunosuppressive drugs primarily target T cells, while innate immunity is not efficiently inhibited in transplantation. The sialic acid–binding immunoglobulin-like lectin E (Siglec-E, or SigE) is an innate receptor that binds to ligands carrying sialic acids and inhibits TLRs-triggered inflammatory responses. Here, we demonstrate the role of SigE in experimental solid organ transplantation mouse models. Upon heart transplantation in mice, SigE is upregulated in graft-infiltrating APCs, including DCs. In humans, Siglec-7 and -9, the human homologs of SigE, are also upregulated in human heart biopsies following transplantation. Genetic ablation of recipient SigE leads to accelerated rejection of heart, kidney, and skin allografts. Mechanistically, SigE-deficient DCs are more susceptible to activation by DAMPS with an enhanced NF-kB activation and production of TNF-a. Moreover, DCs lacking SigE have a higher ability to induce allo-T cell responses in vitro and in vivo. In contrast, overexpression of SigE on DCs decreases their activation by DAMPs and reduces their T cell allostimulatory capacity. Thus, Siglec-E is a major inhibitory receptor in transplantation by controlling APC activation.
Respiratory mucosal immunity against SARS-CoV-2 after vaccination and infection
Authors: Tang Jinyi , Chaudhuri Arka , Qu Panke , Wu Yue , Wavell Kristin , Spano Marthajoy , Taylor Justin , Liu Shan-lu , Teague William , Sun Jie 
Published: November 2024

Abstract
The current COVID-19 mRNA vaccine is effective in systemic immune memory but lacks in generating mucosal anti-SARS-CoV-2 immunity. While prior SARS-CoV-2 infection establishes respiratory immunity, its strain-specific nature and limited duration pose challenges for sustained protection. Recent research has shown that hybrid immunity, a combination of vaccination and infection, results in a more robust, durable, and broadly reactive immune response in circulation compared to vaccination or infection alone. However, the characteristics and protective mechanisms of hybrid immunity in the respiratory mucosa are not fully understood. Moreover, it remains to be determined whether mucosal booster vaccines could potentially induce responses that are even stronger and more broadly reactive than hybrid immunity. Here, we collect nasal wash, bronchoalveolar lavage, and blood from controls, vaccinee, convalescents, or hybrid individuals. We found that hybrid individuals displayed higher levels of specific Ab, nAb, and B cells than other groups in the respiratory mucosa. In our mouse models, although hybrid immunity showed significant mucosal humoral and cellular immune responses, an adenovirus-based mucosal vaccine booster induced more robust protective respiratory immunity against SARS-CoV-2 variants. Our findings reveal the mechanisms of protection of hybrid immunity and underscore the importance of developing mucosal vaccine boosters that can emulate heightened mucosal immunity.
Persistence of Bordetella pertussis as aggregates within nasal tissues of aPV immunized mice prevents recruitment of Siglec-F+ neutrophils and bacterial clearance
Authors: Hall Jesse , Gutiérrez-Ferman Jessica , Shamseldin Mohamed , Guo Myra , Gupta Yash , Deora Rajendar , Dubey Purnima 
Published: November 2024

Abstract
Despite global vaccination, pertussis caused by Bordetella pertussis (Bp) is resurging. One explanation for the increased incidence of infections in immunized individuals is the inability of acellular pertussis vaccines (aPV) to clear Bp from the nose. To understand the host and bacterial mechanisms that contribute to Bp persistence, we evaluated bacterial localization and the immune response in the nasal associated tissues (NT) of naïve and immunized mice following Bp challenge. Bp resided in the NT of naïve and aPV-immunized mice as aggregates, while aggregate formation was not observed in mice immunized with a whole cell pertussis vaccine (wPV). CXCL1 was detected in the NT of wPV immunized mice. Siglec-F+ neutrophils, critical for eliminating Bp from the nose, were elicited at higher levels in wPV immunized mice compared to aPV immunized mice. Together, our data suggest that the TH2 polarized immune response generated by aPV vaccination permitted Bp to persist in the NT by establishing a niche in epithelial tissues that formed a protective barrier to infiltration of immune effectors. In contrast, immune responses generated by wPV, including the neutrophil chemoattractant CXCL1, recruited neutrophils that rapidly eliminated the bacterial burden and prevented establishment of a nasal reservoir. Overall, development of a nasal Bp reservoir and subsequent transmission may be prevented by a modified aPV that elicits a TH1/TH17 phenotype generated by wPV.
Rehabilitation, the disciplining of the body, and disability identity: Reflections from psychotherapy with disabled people
Authors: Watermeyer Brian P. 
Published: November 2024

Background: Recently, attention has been paid to how rehabilitation not only provides medical treatment and instrumental skills but also impacts psychological well-being and identity. We all have psychological structures that discipline the self, enforcing norms internalised during early life and exacting judgments when we fail to ‘make the grade’. In cases of congenital disabilities, rehabilitation interventions may span many years, involving strict programmes of therapy, exercise and self-discipline. These regimes may align with internalised rules in harmful ways, as striving for functional improvements takes on a moral dimension, affecting psychological health and empowered disability identities.Objectives: This study explores rehabilitation by examining the experiences of adults with congenital disabilities, who have undergone childhood medical and rehabilitative interventions.Method: This study was based on the experience of a psychoanalytic psychotherapist working with adults with disability, and presents composite case material to illustrate how interactions with medical authority figures, such as rehabilitation professionals, can have a formative influence on self-identity and entitlement to inclusion.Results: The findings vividly reflected how ‘medical socialisation’ created meanings of disability that were enacted and repeated well into adulthood.Conclusion: The discipline of rehabilitation still has much to do in examining its value-laden assumptions and practices, and how these may shape the internal and relational worlds of people with disability.Contribution: This article contributes to the debate in critical rehabilitation studies, focussing on the issue of constructions of disability which may be communicated to people with disability, with implications for self-advocacy and the growth of the disability movement.