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CD45 is a cell surface phosphatase that shapes the T cell receptor signaling threshold but does not have a known ligand. A family of adenovirus proteins, including E3/49K, exploits CD45 to evade immunity by binding to the extracellular domain of CD45, resulting in the suppression of T cell signaling. We determined the cryo-EM structure of this complex and found that the E3/49K protein is composed of three immunoglobulin domains assembled as beads on a string that compel CD45 into a closely abutted dimer by cross-linking the CD45 D3 domain, leading to steric inhibition of its intracellular phosphatase activity. Inspired by the E3/49K mechanism, we engineered CD45 surrogate ligands that can fine-tune T cell activation by dimerizing CD45 into different orientations and proximities. The adenovirus E3/49K protein has taught us that, despite a lack of a known ligand, CD45 activity can be modulated by extracellular dimerizing ligands that perturb its phosphatase activity and alter T cell responses.

Determining the label to target ratio, also known as the degree of labeling (DOL), is crucial for quantitative fluorescence microscopy and a high DOL with minimal unspecific labeling is beneficial for fluorescence microscopy in general. Yet robust, versatile and easy-to-use tools for measuring cell-specific labeling efficiencies are not available. Here we present a DOL determination technique named protein-tag DOL (ProDOL), which enables fast quantification and optimization of protein-tag labeling. With ProDOL various factors affecting labeling efficiency, including substrate type, incubation time and concentration, as well as sample fixation and cell type can be easily assessed. We applied ProDOL to investigate how human immunodeficiency virus-1 pathogenesis factor Nef modulates CD4 T cell activation measuring total and activated copy numbers of the adapter protein SLP-76 in signaling microclusters. ProDOL proved to be a versatile and robust tool for labeling calibration, enabling determination of labeling efficiencies, optimization of strategies and quantification of protein stoichiometry.

Innate lymphoid cells (ILCs) are a heterogeneous population that play diverse roles in airway inflammation after exposure to allergens and infections. However, how ILCs respond after exposure to environmental toxins is not well understood. Here we show a novel method for studying the heterogeneity of rare lung ILC populations by magnetic enrichment for lung ILCs followed by particle-templated instant partition sequencing (PIP-seq). Using this method, we were able to identify novel group 1 and group 2 ILC subsets that exist after exposure to both fungal allergen and burn pit-related constituents (BPC) that include dioxin, aromatic hydrocarbon, and particulate matter. Toxin exposure in combination with fungal allergen induced activation of specific ILC1/NK and ILC2 populations as well as promoted neutrophilic lung inflammation. Oxidative stress pathways and downregulation of specific ribosomal protein genes (Rpl41 and Rps19) implicated in anti-inflammatory responses were present after BPC exposure. Increased IFNγ expression and other pro-neutrophilic mediator transcripts were increased in BPC-stimulated lung innate lymphoid cells. Further, the addition of BPC induced Hspa8 (encodes HSC70) and aryl hydrocarbon transcription factor activity across multiple lung ILC subsets. Overall, using an airway disease model that develops after occupational and environmental exposures, we demonstrate an effective method to better understand heterogenous ILC subset activation.

Drugs that inhibit HIV transcription and/or reactivation of latent HIV have been proposed as a strategy to reduce HIV-associated immune activation or to achieve a functional cure, yet comparative studies are lacking. We evaluated 26 drugs, including drugs previously reported to inhibit HIV transcription (inhibitors of Tat-dependent HIV transcription, Rev, HSF-1/PTEF-b, HSP90, Jak/Stat, or SIRT1/Tat deacetylation) and other agents that were not tested before (inhibitors of PKC, NF-κB, SP-1, or histone acetyltransferase; NR2F1 agonists), elongation (inhibitors of CDK9/ PTEF-b), completion (inhibitors of PolyA-polymerase), or splicing (inhibitors of human splice factors). To investigate if those drugs would vary in their ability to affect different blocks to HIV transcription, we measured levels of initiated, elongated, midtranscribed, completed, and multiply spliced HIV RNA in PBMCs from antiretroviral therapy-suppressed individuals following ex vivo treatment with each drug and subsequent T cell activation. We identified new drugs that prevent HIV reactivation, including CDK and splicing inhibitors. While some drugs inhibited 1 or 2 steps, other drugs (CDK inhibitors, splicing inhibitors, tanespimycin, and triptolide) inhibited multiple stages of HIV transcription and blocked the production of supernatant viral RNA. These drugs and targets deserve further study in strategies aimed at reducing HIV-associated immune activation or achieving a functional cure.

The mechanisms of postacute medical conditions and unexplained symptoms after SARS-CoV-2 infection [Long Covid (LC)] are incompletely understood. There is growing evidence that viral persistence, immune dysregulation, and T cell dysfunction may play major roles. We performed whole-body positron emission tomography imaging in a well-characterized cohort of 24 participants at time points ranging from 27 to 910 days after acute SARS-CoV-2 infection using the radiopharmaceutical agent [18F]F-AraG, a selective tracer that allows for anatomical quantitation of activated T lymphocytes. Tracer uptake in the postacute COVID-19 group, which included those with and without continuing symptoms, was higher compared with prepandemic controls in many regions, including the brain stem, spinal cord, bone marrow, nasopharyngeal and hilar lymphoid tissue, cardiopulmonary tissues, and gut wall. T cell activation in the spinal cord and gut wall was associated with the presence of LC symptoms. In addition, tracer uptake in lung tissue was higher in those with persistent pulmonary symptoms specifically. Increased T cell activation in these tissues was also observed in many individuals without LC. Given the high [18F]F-AraG uptake detected in the gut, we obtained colorectal tissue for in situ hybridization of SARS-CoV-2 RNA and immunohistochemical studies in a subset of five participants with LC symptoms. We identified intracellular SARS-CoV-2 single-stranded spike protein-encoding RNA in rectosigmoid lamina propria tissue in all five participants and double-stranded spike protein-encoding RNA in three participants up to 676 days after initial COVID-19, suggesting that tissue viral persistence could be associated with long-term immunologic perturbations.

The B cell receptor (BCR) signals together with a multi-component co-receptor complex to initiate B cell activation in response to antigen binding. Here, we take advantage of peroxidase-catalyzed proximity labeling combined with quantitative mass spectrometry to track co-receptor signaling dynamics in Raji cells from 10 s to 2 h after BCR stimulation. This approach enables tracking of 2,814 proximity-labeled proteins and 1,394 phosphosites and provides an unbiased and quantitative molecular map of proteins recruited to the vicinity of CD19, the signaling subunit of the co-receptor complex. We detail the recruitment kinetics of signaling effectors to CD19 and identify previously uncharacterized mediators of B cell activation. We show that the glutamate transporter SLC1A1 is responsible for mediating rapid metabolic reprogramming and for maintaining redox homeostasis during B cell activation. This study provides a comprehensive map of BCR signaling and a rich resource for uncovering the complex signaling networks that regulate activation.

NK cells in the peripheral blood of severe COVID-19 patients exhibit a unique profile characterized by activation and dysfunction. Previous studies have identified soluble factors, including type I IFN and TGF-β, that underlie this dysregulation. However, the role of cell-cell interactions in modulating NK cell function during COVID-19 remains unclear. To address this question, we combined cell-cell communication analysis on existing single-cell RNA sequencing data with in vitro primary cell coculture experiments to dissect the mechanisms underlying NK cell dysfunction in COVID-19. We found that NK cells are predicted to interact most strongly with monocytes and that this occurs via both soluble factors and direct interactions. To validate these findings, we performed in vitro cocultures in which NK cells from healthy human donors were incubated with monocytes from COVID-19+ or healthy donors. Coculture of healthy NK cells with monocytes from COVID-19 patients recapitulated aspects of the NK cell phenotype observed in severe COVID-19, including decreased expression of NKG2D, increased expression of activation markers, and increased proliferation. When these experiments were performed in a Transwell setting, we found that only CD56bright CD16- NK cells were activated in the presence of severe COVID-19 patient monocytes. O-link analysis of supernatants from Transwell cocultures revealed that cultures containing severe COVID-19 patient monocytes had significantly elevated levels of proinflammatory cytokines and chemokines, as well as TGF-β. Collectively, these results demonstrate that interactions between NK cells and monocytes in the peripheral blood of COVID-19 patients contribute to NK cell activation and dysfunction in severe COVID-19.

Adenosine to inosine (A-to-I) RNA editing by ADAR1 has been implicated in maintaining self-tolerance, preventing autoimmunity, and mediating antiviral immunity. Foreign viral double-stranded RNA triggers rapid interferon response and activates ADAR1 in the host immune system. Emerging data points to a role of ADAR1 A-to-I editing in the inflammatory response associated with severe COVID-19 disease. We identify A-to-I editing events within human whole transcriptome data from SARS-CoV-2 infected individuals, non-infected individuals, and individuals with other viral illnesses from nasopharyngeal swabs. High levels of RNA editing in host cells are associated with low SARS-CoV-2 viral load (p = 9.27 E-06), suggesting an inhibitory effect of ADAR1 on viral infection. Additionally, we find differentially expressed genes associated with RNA-modifications and interferon response. Single cell RNA-sequencing analysis of SARS-CoV-2 infected nasopharyngeal swabs reveals that cytotoxic CD8 T cells upregulate ADAR1 in COVID-19 positive samples (p = 0.0269). We further reveal ADAR1 expression increases with CD4 and CD8 T cell activation, and knockdown of ADAR1 leads to apoptosis and aberrant IL-2 secretion. Together, our data suggests A-to-I RNA editing is required to maintain healthy homeostasis of activated T cells to combat SARS-CoV-2 infection.

Abstract Background The pathogenesis of hypertension (HTN) in people living with HIV/AIDS (PLHIV) is complex and remains not fully understood. Chronic immune activation (IA) is postulated to be one of the culprits. This notion is derived from studies in HIV-uninfected populations and/or animals while data on HTN and how it relates to IA in PLHIV remains scarce. We determined the relationship between HTN and IA among antiretroviral therapy (ART) naïve PLHIV. Methods We analysed baseline data of 365 out of 430 clinical trial participants whose main aim was to investigate the effect of low-dose aspirin on HIV disease progression in PLHIV starting ART. Soluble CD14 (sCD14), T cells co-expressing CD38 and HLA-DR, and PD-1 were the IA and exhaustion markers, respectively studied and were analysed by flow cytometry. Mann-Whitney U-test was used for comparison of the markers by HTN status. A robust Poisson regression model was used to determine the predictors for HTN. Results A quarter of the 365 were hypertensive (25.3%, 95% CI 20.9–29.8%), and, had higher median (IQR) body mass index (kg/m2) (23.4 (19.6, 28.0) versus 21.9 (19.3, 25.1)) and lower median (IQR) estimated glomerular filtration rate (mL/min/1.73m2) (101.2 (79.4, 126.9) versus 113.6 (92.7, 138.8)) than normotensive participants (p 0.05). None of these markers significantly predicted the occurrence of HTN. Conclusion Studied markers of IA and exhaustion were higher in PLHIV with HTN than those without but were unpredictive of HTN. Larger multicentre studies with a wider range of markers are needed to confirm the role of IA in HIV-associated HTN.

Background: The pathogenesis of hypertension (HTN) in people living with HIV/AIDS (PLHIV) is complex and remains not fully understood. Chronic immune activation (IA) is postulated to be one of the culprits. This notion is derived from studies in HIV-uninfected populations and/or animals while data on HTN and how it relates to IA in PLHIV remains scarce. We determined the relationship between HTN and IA among antiretroviral therapy (ART) naïve PLHIV. Methods: We analysed baseline data of 365 out of 430 clinical trial participants whose main aim was to investigate the effect of low-dose aspirin on HIV disease progression in PLHIV starting ART. Soluble CD14 (sCD14), T cells co-expressing CD38 and HLA-DR, and PD-1 were the IA and exhaustion markers, respectively studied and were analysed by flow cytometry. Mann-Whitney U-test was used for comparison of the markers by HTN status. A robust Poisson regression model was used to determine the predictors for HTN. Results: A quarter of the 365 were hypertensive (25.3%, 95% CI 20.9–29.8%), and, had higher median (IQR) body mass index (kg/m2) (23.4 (19.6, 28.0) versus 21.9 (19.3, 25.1)) and lower median (IQR) estimated glomerular filtration rate (mL/min/1.73m2) (101.2 (79.4, 126.9) versus 113.6 (92.7, 138.8)) than normotensive participants (p < 0.05). Participants with HTN had higher median frequencies of all markers of IA and exhaustion but lower sCD14 (p > 0.05). None of these markers significantly predicted the occurrence of HTN. Conclusion: Studied markers of IA and exhaustion were higher in PLHIV with HTN than those without but were unpredictive of HTN. Larger multicentre studies with a wider range of markers are needed to confirm the role of IA in HIV-associated HTN. [ABSTRACT FROM AUTHOR]

Lymphocyte phosphatase-associated phosphoprotein (LPAP) is a binding partner of the phosphatase CD45, but its function remains poorly understood. Its close interaction with CD45 suggests that LPAP may potentially regulate CD45, but direct biochemical evidence for this has not yet been obtained. We found that in the Jurkat lymphoid cells the levels of LPAP and CD45 proteins are interrelated and well correlated with each other. Knockout of LPAP leads to the decrease in the surface expression of CD45, while its overexpression, on the contrary, caused its increase. No such correlation was found in the non-lymphoid K562 cells. We hypothesize that LPAP regulates expression level of CD45 and thus can affect lymphocyte activation. [ABSTRACT FROM AUTHOR]

LAG-3 is a member of the immunoglobulin superfamily expressed on activated T cells, but also on other immune cells. It has significant homology to CD4. Both molecules have four extracellular Ig-like domains with similar structural motifs but the sequence identity between LAG-3 and CD4 is low. Furthermore, unlike CD4 LAG-3 restrains T cell responses and antibodies targeting this receptor are emerging drugs in cancer immunotherapy. A combination of LAG-3 and PD-1 antibodies has already been approved for the treatment of metastatic melanoma. Despite this success, its biology is still not well understood. Here we summarize the current knowledge on expression, ligands, and function of LAG-3. We point to the differences between LAG-3 and other inhibitory immune checkpoints and describe obstacles to study the role of this receptor in T cell activation processes. Finally, we discuss future directions for scientific efforts to come to a more complete understanding of the biology of this eminent immune checkpoint., (© 2024 The Author(s). The FASEB Journal published by Wiley Periodicals LLC on behalf of Federation of American Societies for Experimental Biology.)

Objective: The aim of this study was to characterize T-cell activation, exhaustion, maturation and Treg frequencies in individuals who acquire perinatal HIV (PHIV), in individuals who acquired HIV as adult (AHIV), and in healthy controls., Design: This cross-sectional study included people with HIV at least 14 and younger than 40 years, HIV-RNA less than 50 copies/ml on antiretroviral therapy for at least 6 months, and HC., Methods: We assessed the expression of PD-1, TIM-3, EOMES, CD38 + DR+, maturation status by CD4 + and CD8 + T cells and the frequency of CD4 + and CD8 + Treg cells. Principal component analysis (PCA) and k-means cluster analysis investigated which combination of immunological parameters better associated with each group., Results: Twenty-six PHIV and 18 AHIV with median ages of 26 (8.0) and 28 (6.8) years were consecutively enrolled. PHIV showed significant higher frequency of naive and lower frequency of terminal effector memory CD4 + and CD8 + T cells than AHIV. AHIV exhibited higher expression of exhaustion and activation markers. The statistical analysis returned two clusters with 94% of specificity and 88% of sensitivity identifying PHIV vs. AHIV. The nine healthy controls had a lower expression of exhaustion markers on both CD4 + and CD8 + T lymphocytes than PHIV and AHIV., Conclusion: These data may exclude major alterations of lymphopoiesis in PHIV, with even lower state of immune-activation and exhaustion compared with AHIV. This suggests that recent lack of virological control, may affect immune activation and exhaustion of CD4 + and CD8 + T cells., (Copyright © 2024 Wolters Kluwer Health, Inc. All rights reserved.)

Protein-protein interactions (PPIs) play pivotal roles in directing T cell fate. One key player is the non-receptor tyrosine protein kinase Lck that helps to transduce T cell activation signals. Lck is mediated by other proteins via interactions that are inadequately understood. Here, we use the deep learning method AF2Complex to predict PPIs involving Lck, by screening it against ∼1,000 proteins implicated in immune responses, followed by extensive structural modeling for selected interactions. Remarkably, we describe how Lck may be specifically targeted by a palmitoyltransferase using a phosphotyrosine motif. We uncover "hotspot" interactions between Lck and the tyrosine phosphatase CD45, leading to a significant conformational shift of Lck for activation. Lastly, we present intriguing interactions between the phosphotyrosine-binding domain of Lck and the cytoplasmic tail of the immune checkpoint LAG3 and propose a molecular mechanism for its inhibitory role. Together, this multifaceted study provides valuable insights into T cell regulation and signaling., Competing Interests: Declaration of interests M.G. is a stockholder at AgnistaBio, Inc., (Copyright © 2024 Elsevier Inc. All rights reserved.)

Objective: To investigate and analyze the correlation between the expression levels of CD38, HLA-DR and programmed cell death-1 (PD-1) on peripheral blood CD8 + T cells and HIV-1 RNA viral load, immune activation and exhaustion in patients with human immunodeficiency virus (HIV)/acquired immunodeficiency syndrome (AIDS). Methods: A total of 81 HIV/AIDS patients (64 without antiretroviral therapy and 17 with therapy) and 40 healthy donors in the same period were enrolled as the control group. Flow cytometry was used to analyze the CD4 + T lymphocyte count and the expression levels of activation markers CD38 and HLA-DR and apoptosis marker PD-1 on CD8 + T cells. HIV-1 RNA in the plasma of HIV-1 infected patients was quantitatively detected by real-time fluorescence quantitative polymerase chain reaction. Variance analysis was used to compare the expression levels of CD38, HLA-DR and PD-1 on CD8 + T cells between HIV/AIDS patients and healthy controls. Spearman correlation analysis was used to analyze the correlation between different T lymphocyte counts and HIV RNA viral load, and the correlation between HIV RNA viral load and peripheral blood CD8 + T cell CD38, HLA-DR and PD-1. Results: Among the 81 HIV/AIDS patients, 69 (85.19%) were males and 12 (14.81%) were females, with an age M ( Q 1 , Q 3 ) of 58 (36.5, 65.0) years. There were 60 HIV/AIDS patients over 55 years old (74.07%) and 21 HIV/AIDS patients between 18 and 55 years old (25.93%). The results of variance analysis showed that compared with the healthy control group, the expression levels of CD38, HLA-DR and PD-1 on CD8 + T cells in HIV/AIDS patients increased, and the differences were statistically significant (all P <0.05). In addition, the expression of CD38, HLA-DR and PD-1 increased significantly in patients with CD4 + T cell count less than 350 cells/μl, and the differences were statistically significant (all P <0.05). Spearman correlation analysis showed that CD4 + and CD4 + /CD8 + were negatively correlated with viral load in HIV/AIDS patients ( r =-0.407 and -0.378, respectively, both P <0.05), and CD8 + was positively correlated with viral load ( r =0.356, P <0.05). When the HIV RNA level was≤10 5 CPs/ml, there was no correlation between the HIV RNA level and the expression levels of CD38, HLA-DR and PD-1 on CD8 + T cells (all P >0.05). However, when the level of HIV RNA was>10 5 CPs/ml, the level of HIV RNA was positively correlated with the expression levels of CD38, HLA-DR and PD-1 on CD8 + T cells ( r =0.412, 0.387, 0.395, respectively, all P <0.05). Conclusions: The activation levels of CD38 and HLA-DR and the expression of PD-1 on CD8 + T cells in the peripheral blood of HIV/AIDS patients are increased. When the viral load is high, the HIV RNA viral load is positively correlated with the activation and exhaustion levels of CD8 + T cells.

Background: . CD8 + Cytotoxic T lymphocytes play a key role in the pathogenesis of autoimmune diseases and clinical conditions such as graft versus host disease and graft rejection. Mesenchymal Stromal Cells (MSCs) are multipotent cells with tissue repair and immunomodulatory capabilities. Since they are able to suppress multiple pathogenic immune responses, MSCs have been proposed as a cellular therapy for the treatment of immune-mediated diseases. However, the mechanisms underlying their immunosuppressive properties are not yet fully understood. MSCs have the remarkable ability to sense tissue injury and inflammation and respond by donating their own mitochondria to neighboring cells. Whether mitochondrial transfer has any role in the repression of CD8 + responses is unknown., Methods and Results: . We have utilized CD8 + T cells from Clone 4 TCR transgenic mice that differentiate into effector cells upon activation in vitro and in vivo to address this question. Allogeneic bone marrow derived MSCs, co-cultured with activated Clone 4 CD8 + T cells, decreased their expansion, the production of the effector cytokine IFNγ and their diabetogenic potential in vivo. Notably, we found that during this interaction leading to suppression, MSCs transferred mitochondria to CD8 + T cells as evidenced by FACS and confocal microscopy. Transfer of MSC mitochondria to Clone 4 CD8 + T cells also resulted in decreased expansion and production of IFNγ upon activation. These effects overlapped and were additive with those of prostaglandin E2 secreted by MSCs. Furthermore, preventing mitochondrial transfer in co-cultures diminished the ability of MSCs to inhibit IFNγ production. Finally, we demonstrated that both MSCs and MSC mitochondria downregulated T-bet and Eomes expression, key transcription factors for CTL differentiation, on activated CD8 + T cells., Conclusion: . In this report we showed that MSCs are able to interact with CD8 + T cells and transfer them their mitochondria. Mitochondrial transfer contributed to the global suppressive effect of MSCs on CD8 + T cell activation by downregulating T-bet and Eomes expression resulting in impaired IFNγ production of activated CD8 + T cells., (© 2024. The Author(s).)

Extracellular vesicles (EVs) are nano-sized membrane particles secreted by various cell types that are involved in many important cellular processes. Recently, EVs originating from immune cells, such as dendritic cells, chimeric antigen receptor T cells, and natural killer cells, have attracted much attention because of their known direct and indirect antitumor activity. Here, we report the EVs released by cytokine-activated CD8+ T (caCD8) cells and its cytotoxicity against cancer cells. CaCD8 cells can release EVs following stimulation of CD8+ T cells with an anti-CD3 antibody and a cytokine cocktail ex vivo. The isolated vesicles have typical EV characteristics, such as an oval shape and a size distribution between 30 and 200 nm, as well as CD81 expression. Notably, caCD8-EVs displayed cytotoxicity against various cancer cells in vitro. Furthermore, mechanism analysis demonstrates that caCD8-EVs not only contain typical cytotoxic proteins (i.e. granzyme B and perforin), but also significantly enrich interferon γ (IFNγ) compared with caCD8 cells. EV-derived IFNγ participates in EV-induced apoptosis in cancer cells. Therefore, our data reveal antitumor effects of EVs secreted from caCD8 cells and the potential role of the EV-derived IFNγ., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Leukocyte Biology. All rights reserved. For commercial re-use, please contact reprints@oup.com for reprints and translation rights for reprints. All other permissions can be obtained through our RightsLink service via the Permissions link on the article page on our site—for further information please contact journals.permissions@oup.com.)

Aim: Rheumatoid arthritis (RA) is a prevalent autoimmune disease characterized by immune dysegulation, including an immune imbalance due to abnormal activation of non-classical Th1 cells (CD161 + Th1). This study investigated the effects of CCR5 on the activation and proliferation of CD161 + Th1 and their pathogenicity in patients with RA., Methods: The study was conducted on 53 patients with RA and 32 age- and sex-matched healthy controls (HC). The cell phenotype was assessed by flow cytometry and the cytokine levels in the supernatant were detected by ELISA., Results: We demonstrate a marked increase in CD161 + Th1 cells in the synovial fluid of RA patients. These cells exhibit a hyperactivated and hyperproliferative state alongside elevated CCR5 expression. Furthermore, the levels of CD161 + Th1 cells, CD25, and CCR5 in RA synovial fluid show a positive correlation with the disease activity. Additionally, our study reveals that CCR5 facilitates the activation, proliferation, and cytokine production of CD161 + Th1 cells through the pZAP70/NFAT signaling pathway., Conclusion: These findings contribute to a deeper understanding of RA pathogenesis and uncover a novel mechanism that regulates non-classical CD161 + Th1 responses in RA, which may provide a potential therapeutic target., (© 2024 Asia Pacific League of Associations for Rheumatology and John Wiley & Sons Australia, Ltd.)

Natural killer (NK) cells play a pivotal role in the immune response against viral infections, including SARS-CoV-2. However, our understanding of memory NK cell responses in the context of SARS-CoV-2 remains limited. To address this, we investigated the memory-like response of NK cells to SARS-CoV-2 peptides, presented by autologous cells. Blood samples from 45 donors underwent analysis for SARS-CoV-2 IgG antibodies, categorizing them into four groups based on the antibody kind and level. NK cells from SARS-CoV-2-experienced donors demonstrated enhanced degranulation and activation levels, IFNγ production and proliferative potential in response to SARS-CoV-2 peptides. Investigation of highly proliferating NK cells demonstrated the formation of distinct clusters depending on the SARS-CoV-2 peptide supplementation and the donor group. RNA sequencing revealed differential gene expression patterns, highlighting metabolism, protein transport, and immune response genes. Notably, KIR2DS4 expression correlated with enhanced IFNγ production, degranulation and proliferation levels, suggesting a role in SARS-CoV-2 recognition. Collectively, these findings provide detailed insights into antigen-specific NK cell responses to SARS-CoV-2 peptides, indicating potential mechanisms underlying NK cell activation in antiviral immunity., (© 2024 Wiley Periodicals LLC.)

Colorectal cancer (CRC) is a prevalent malignant tumor of the gastrointestinal system, with the third and second highest incidence and mortality rates globally in 2020, respectively. Immunotherapy has developed rapidly in recent years. Natural killer (NK) cells have received increasing attention in the field of tumor immunotherapy due to their recognition and killing tumor cells without the limitations of major histocompatibility complexes. However, constraints within the tumor microenvironment that impede the infiltration and proliferation of NK cells result in poor efficacy of NK cell therapy for solid tumors. Oncolytic viral therapy is an immunogenic treatment with the potential to enhance anti-tumour immune responses and promote immune cell infiltration. In this study, we synergistically combine NK cells with an oncolytic adenovirus carrying Decorin (rAd.DCN) for the treatment of colorectal cancer (CRC) in a xenograft mouse model. By using Flow cytometry, real-time quantitative PCR and Calcein-AM release assay, we found that rAd.DCN could effectively promote proliferation, activation and degranulation of NK cells, up-regulate expression and secretion of NK cell killing activity-related factors, and enhance their killing activity. The efficacy is better than that of the blank control oncolytic virus rAd.Null. Combined treatment significantly inhibited tumor growth, increased the number of NK cells in peripheral blood, promoted the killing function of NK cells, and increased the expression levels of perforin and IFN-γ. At the same time, more NK cells were recruited to infiltrate tumor tissue. Our study established the feasibility of combination NK cells and oncolytic adenovirus application, thus expanding the scope of potentially curative treatments for NK cells in CRC., (© 2024. The Author(s).)

In vitro diagnostics for drug hypersensitivity reactions distinguish between serological and cellular-based tests. A serological test used for the diagnosis of immediate type reactions is the detection of specific IgE antibodies. The cellular tests include the basophil activation test for immediate type reactions and the lymphocyte transformation test, which is mainly used to detect delayed type hypersensitivity reactions. Further cellular-based tests are the CAST-ELISA and the mast cell activation test. None of the above-mentioned tests can definitively exclude an allergy if the result is negative. In addition, it is important to note that even a positive test result is not necessarily associated with an allergy but has to be interpreted in the clinical context., (© 2024 The Authors. Journal der Deutschen Dermatologischen Gesellschaft published by Wiley‐VCH GmbH on behalf of Deutsche Dermatologische Gesellschaft.)

Objective: Rheumatoid arthritis (RA) is a chronic systemic autoimmune disease in which activated CD4 + T cells participate in the disease process by inducing inflammation. We aimed to investigate the role of Toll-like receptor 2 (TLR2) on CD4 + T cells in RA patients, and to elucidate the underlying mechanisms by which TLR2 contributes to the pathogenesis of RA., Methods: Serum samples were collected from RA patients and healthy controls. Soluble TLR2 levels were quantified using an enzyme-linked immunosorbent assay (ELISA). Flow cytometry was employed to assess the TLR2 expression level, activation status, cytokine production, reactive oxygen species (ROS) levels, and glucose uptake capacity of CD4 + T cells. Quantitative polymerase chain reaction (qPCR) was used to measure the expression of enzymes associated with glucose and lipid metabolism. The concentration of lactic acid in the culture supernatant was determined using a dedicated detection kit., Results: RA patients had higher levels of TLR2 in their serum, which positively correlated with C-reactive protein and rheumatoid factor. The expression level of TLR2 in CD4 + T cells of RA patients was increased, and TLR2 + cells showed higher activation levels than TLR2- cells. Activation of TLR2 in CD4 + T cells of RA patients promoted their activation, TNF-α secretion, and increased production of ROS. Furthermore, TLR2 activation led to changes in enzymes related to glucose metabolism, causing a shift in glucose metabolism towards the pentose phosphate pathway. Blocking oxidative phosphorylation and the pentose phosphate pathway had varying effects on CD4 + T cell function., Conclusion: TLR2 reprograms the glucose metabolism of CD4 + T cells in RA patients, contributing to the development of RA through ROS-mediated cell hyperactivation and TNF-α secretion. Key Points • TLR2 is upregulated in CD4 + T cells of RA patients and correlates with disease severity markers such as CRP and RF. • Activation of TLR2 in CD4 + T cells promotes cell activation, TNF-α secretion, and increased ROS production, contributing to the pathogenesis of RA. • TLR2 activates glucose metabolism in CD4 + T cells, shifting towards the pentose phosphate pathway, which may be a novel therapeutic target for RA treatment. • Blocking glucose metabolism and ROS production can reduce CD4 + T cell hyperactivation and TNF-α secretion, indicating potential therapeutic strategies for RA management., (© 2024. The Author(s), under exclusive licence to International League of Associations for Rheumatology (ILAR).)

Substantial reduction of the intact proviral reservoir is essential towards HIV-1 cure. In vivo administration of broadly neutralizing antibodies (bNAbs) targeting the HIV-1 envelope glycoprotein (Env) trimer can decrease the viral reservoir, through Fc-mediated killing of infected cells. In this study, we compared three commonly used antibody engineering strategies to enhance Fc-mediated effector functions: (i) glyco-engineering, (ii) protein engineering, and (iii) subclass/hinge modifications in a panel of anti-HIV-1 antibodies. We found that antibody-dependent cellular phagocytosis (ADCP) was improved by elongating the hinge domain and switching to an IgG3 constant domain. In addition, potent NK cell activation and ADCC activity was observed for afucosylated antibodies and antibodies bearing the GASDALIE mutations. The combination of these engineering strategies further increased NK cell activation and induced antibody dependent cytotoxicity (ADCC) of infected cells at low antibody concentrations. The bNAb N6 was most effective at killing HIV-1 infected cells, likely due to its high affinity and optimal angle of approach. Overall, the findings of this study are applicable to other antibody formats, and can aid the development of effective immunotherapies and antibody-based treatments for HIV-1 cure strategies., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier B.V. All rights reserved.)

Foot-and-mouth disease (FMD) is a highly contagious and economically devastating viral disease of ruminants and swine, badly affecting the livestock industry worldwide. In clinical practice, vaccination is a frequently employed strategy to prevent foot-and-mouth disease (FMDV). However, commercial inactivated vaccines for FMD mainly rely on humoral immunity, exhibiting poor cellular immune responses and causing adverse reactions. Here, we use the double emulsion method to prepare poly (lactic-co-glycolic acid) nanoparticles (PLGA-NP) encapsulated with IL-2 cytokines, wrap the dendritic cell (DC) membrane carrying FMDV antigen information on the surface of the nanoparticles, obtaining a biomimetic nanoparticle vaccine Biom@DC with uniform size. This vaccine can effortlessly move through lymph nodes due to its nanoscale size advantage. It also possesses DC ability to present antigens, and antigen presentation can be made more effective with high biocompatibility. The sustained release of IL-2 encapsulated in the core of PLGA-NP in vivo can effectively promote the body's cellular immune response. Immune tests on mice have shown that Biom@DC may greatly increase T cell activation and proliferation both in vivo and in vitro, while also significantly reducing the fraction of inhibitory Treg cells. Furthermore, in the micro serum neutralization assay for FMDV, it has been demonstrated that the group vaccinated with Biom@DC exhibits a clear neutralizing effect. Given its strong immunogenicity, Biom@DC has the potential to develop into a novel, potent anti-FMDV vaccination., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Hepatocellular carcinoma (HCC) is a highly malignant tumor with significant global health implications. The role of CD4 + T cells, particularly conventional CD4 + T cells (Tconvs), in HCC progression remains unexplored. Furthermore, epigenetic factors are crucial in immune regulation, yet their specific role in HCC-infiltrating Tconv cells remains elusive. This study elucidates the role of MATR3, an epigenetic regulator, in modulating Tconv activity and immune evasion within the HCC microenvironment. Reanalysis of the scRNA-seq data revealed that early activation of CD4 + T cells is crucial for establishing an antitumor immune response. In vivo and in vitro experiments revealed that Tconv enhances cDC1-induced CD8 + T-cell activation. Screening identified MATR3 as a critical regulator of Tconv function, which is necessary for antitumour activity but harmful when overexpressed. Excessive MATR3 expression exacerbates Tconv exhaustion and impairs function by recruiting the SWI/SNF complex to relax chromatin in the TOX promoter region, leading to aberrant transcriptional changes. In summary, MATR3 is an HCC-specific epigenetic checkpoint that bidirectionally regulates Tconv antitumour immunity, suggesting new therapeutic strategies targeting epigenetic regulators to enhance antitumour immunity in HCC., (© 2024. The Author(s), under exclusive licence to CSI and USTC.)

The onset and progression of colorectal cancer is intricately linked to a multitude of factors. Among these, immune cells present within the tumor microenvironment play a pivotal role, particularly natural killer (NK) cells, which are essential for mediating anti-tumor immunity. This study aims to elucidate the mechanism by which the VWA2 protein facilitates the invasion and migration of colorectal cancer cells through the inhibition of NK cell activation. Understanding this molecular mechanism is crucial for deciphering the underlying processes involved in colorectal cancer. To achieve the study's objectives, various methodologies were employed, including cell culture techniques, transgenic technology, and assessments of NK cell functionality. The "limma" bioinformatics tool was utilised to identify differentially expressed genes (DEGs) between samples of colon cancer or polyps and normal tissue through transcriptome sequencing. Subsequent Wien analysis was conducted to pinpoint overlapping genes of interest. The impact of VWA2 on both the invasion and migration of colorectal cancer cell lines was assessed through experiments designed for the overexpression and knockout of VWA2.In addition, flow cytometry was employed to evaluate the activation status of NK cells, enabling an analysis of how VWA2 modulates relevant signaling pathways. The findings revealed that overexpression of VWA2 led to a marked inhibition of NK cell activation, which corresponded with reduced cytotoxic activity against tumor cells. Further examination indicated that VWA2 significantly amplified the migration and invasion capabilities of colorectal cancer cells by upregulating immunosuppressive factors while simultaneously downregulating pro-inflammatory factors. Conversely, the reduction of VWA2 expression was shown to markedly enhance NK cell functionality and decrease the invasive potential of colorectal cancer cells. Thus, the evidence suggests that the VWA2 protein actively promotes the migration and invasion of colorectal cancer cells primarily by suppressing NK cell activation, highlighting its potential role as a significant contributor to tumor progression in colorectal cancer., Competing Interests: Declaration of competing interest The authors declare no competing interests., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Tuberculosis (TB) is still an urgent global public health problem. Notably, mucosal-associated invariant T (MAIT) cells play an important role in early anti-TB immune response. Targeted control of them may be an effective method to improve vaccine efficacy and TB treatment. However, the biology and signal regulation mechanisms of MAIT cells in TB patients are still poorly understood. Previous studies have been limited by the lack of reagents to specifically identify MAIT cells. In addition, the use of alternative markers may subsume non-MAIT cell into MAIT cell populations. In this study, the human MR1 tetramer which can specifically identify MAIT cells was used to further explore the effect and mechanism of MAIT cells in anti-TB immune response. Our results showed that the tetramer + MAIT cells in peripheral blood of TB patients were mainly CD8 + or CD4 - CD8 - cells, and very few were CD4 + cells. After BCG infecting autologous antigen-presenting cells, MAIT cells in patients produced significantly higher levels of cytokines, lysis and proliferation compared with healthy controls. After suppression of mTORC1 by the mTORC1-specific inhibitor rapamycin, the immune response of MAIT cells in patients was significantly reduced. This study demonstrates that peripheral blood tetramer + MAIT cells from TB patients have significant anti-TB immune effect, which is regulated by mTORC1. This could provide ideas and potential therapeutic targets for the development of novel anti-TB immunotherapy., (© 2024 The Author(s). Immunology published by John Wiley & Sons Ltd.)

Background & Aims: Intestinal epithelial cell (IEC) damage is a hallmark of celiac disease (CeD); however, its role in gluten-dependent T-cell activation is unknown. We investigated IEC-gluten-T-cell interactions in organoid monolayers expressing human major histocompatibility complex class II (HLA-DQ2.5), which facilitates gluten antigen recognition by CD4 + T cells in CeD., Methods: Epithelial major histocompatibility complex class II (MHCII) was determined in active and treated CeD, and in nonimmunized and gluten-immunized DR3-DQ2.5 transgenic mice, lacking mouse MHCII molecules. Organoid monolayers from DR3-DQ2.5 mice were treated with or without interferon (IFN)-γ, and MHCII expression was evaluated by flow cytometry. Organoid monolayers and CD4 + T-cell co-cultures were incubated with gluten, predigested, or not by elastase-producing Pseudomonas aeruginosa or its lasB mutant. T-cell function was assessed based on proliferation, expression of activation markers, and cytokine release in the co-culture supernatants., Results: Patients with active CeD and gluten-immunized DR3-DQ2.5 mice demonstrated epithelial MHCII expression. Organoid monolayers derived from gluten-immunized DR3-DQ2.5 mice expressed MHCII, which was upregulated by IFN-γ. In organoid monolayer T-cell co-cultures, gluten increased the proliferation of CD4 + T cells, expression of T-cell activation markers, and the release of interleukin-2, IFN-γ, and interleukin-15 in co-culture supernatants. Gluten metabolized by P aeruginosa, but not the lasB mutant, enhanced CD4 + T-cell proliferation and activation., Conclusions: Gluten antigens are efficiently presented by MHCII-expressing IECs, resulting in the activation of gluten-specific CD4 + T cells, which is enhanced by gluten predigestion with microbial elastase. Therapeutics directed at IECs may offer a novel approach for modulating both adaptive and innate immunity in patients with CeD., (Copyright © 2024 AGA Institute. Published by Elsevier Inc. All rights reserved.)

Rheumatoid arthritis (RA) is a systemic autoimmune disease driven by highly active autoantibody-producing B cells. Activation of B cells is maintained within ectopic germinal centres found in affected joints. Fibroblast-like synoviocytes (FLS) present in inflamed joints support B-cell survival, activation, and differentiation. CD27+ memory B cells and naive B cells show very different responses to activation, particularly by CD40 ligand (CD40L). We show that FLS-dependent activation of human B cells is dependent on interleukin-6 (IL-6) and CD40L. FLS have been shown to activate both naive and memory B cells. Whether the activating potential of FLS is different for naive and memory B cells has not been investigated. Our results suggest that FLS-induced activation of B cells is dependent on IL-6 and CD40L. While FLS are able to induce plasma cell differentiation, isotype switching, and antibody production in memory B cells, the ability of FLS to activate naive B cells is significantly lower., (© 2024 The Author(s). Immunology published by John Wiley & Sons Ltd.)

Purpose: Immune cells are capable of eliminating leukemic cells, as evidenced by outcomes in hematopoietic cell transplantation (HCT). However, patients who fail induction therapy will not benefit from HCT due to their minimal residual disease (MRD) status. Thus, we aimed to develop an immunomodulatory agent to reduce MRD by activating immune effector cells in the presence of leukaemia cells via a novel fusion protein that chimerises two clinically tolerated biologics: a CD33 antibody and the IL15Ra/IL15 complex (CD33xIL15)., Methods: We generated a set of CD33xIL15 fusion protein constructs with varying configurations and identified those with the best in vitro AML-binding, T cell activation, and NK cell potentiation. Using 89 Zr-immunoPET imaging we then evaluated the biodistribution and in vivo tumour retention of the most favourable CD33xIL15 constructs in an AML xenograft model. Ex vivo biodistribution studies were used to confirm the pharmacokinetics of the constructs., Results: Two of the generated fusion proteins, CD33xIL15 (N72D) and CD33xIL15wt, demonstrated optimal in vitro behaviour and were further evaluated in vivo. These studies revealed that the CD33xIL15wt candidate was capable of being retained in the tumour for as long as its parental CD33 antibody, Lintuzumab (13.9 ± 3.1%ID/g vs 18.6 ± 1.1%ID/g at 120 h)., Conclusion: This work demonstrates that CD33xIL15 fusion proteins are capable of targeting leukemic cells and stimulating local T cells in vitro and of concentrating in the tumour in AML xenografts. It also highlights the importance of 89 Zr-immunoPET to guide the development and selection of tumour-targeted antibody-cytokine fusion proteins., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

T cells utilized in adoptive T cell immunotherapy are typically activated in vitro. Although these cells demonstrate proliferation and anti-tumor activity following activation, they often face difficulties in sustaining long-term survival post-reinfusion. This issue is attributed to the induction of T cells into a terminal differentiation state upon activation, whereas early-stage differentiated T cells exhibit enhanced proliferation potential and survival capabilities. In previous study, we delineated four T cell subsets at varying stages of differentiation: T N , T SCM , T CM , and T EM , and acquired their miRNA expression profiles via high-throughput sequencing. In the current study, we performed a differential analysis of miRNA across these subsets, identifying a distinct miRNA, hsa-miR-744-5p, characterized by progressively increasing expression levels upon T cell activation. This miRNA is not expressed in T SCM but is notably present in T EM . Target genes of miR-744-5p were predicted, followed by Gene Ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) pathway enrichment analyses, revealing that these genes predominantly associate with pathways related to the 'Wnt signaling pathway'. We established that miR-744-5p directly targets STK11, influencing its expression. Further, we investigated the implications of miR-744-5p on T cell differentiation and functionality. Overexpression of miR-744-5p in T cells resulted in heightened apoptosis, reduced proliferation, an increased proportion of late-stage differentiated T cells, and elevated secretion of the cytokine TNF-α. Moreover, post-overexpression of miR-744-5p led to a marked decline in the expression of early-stage differentiation-associated genes in T cells (CCR7, CD62L, LEF1, BCL2) and a significant rise in late-stage differentiation-associated genes (KLRG1, PDCD1, GZMB). In conclusion, our findings affirm that miR-744-5p contributes to the progressive differentiation of T cells by downregulating the STK11 gene expression., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

T cell receptor (TCR) engagement causes a global cellular response that entrains signaling pathways, cell cycle regulation, and cell death. The molecular regulation of mRNA translation in these processes is poorly understood. Using a whole-genome CRISPR screen for regulators of CD95 (FAS/APO-1)-mediated T cell death, we identified AMBRA1, a protein previously studied for its roles in autophagy, E3 ubiquitin ligase activity, and cyclin regulation. T cells lacking AMBRA1 resisted FAS-mediated cell death by down-regulating FAS expression at the translational level. We show that AMBRA1 is a vital regulator of ribosome protein biosynthesis and ribosome loading on select mRNAs, whereby it plays a key role in balancing TCR signaling with cell cycle regulation pathways. We also found that AMBRA1 itself is translationally controlled by TCR stimulation via the CD28-PI3K-mTORC1-EIF4F pathway. Together, these findings shed light on the molecular control of translation after T cell activation and implicate AMBRA1 as a translational regulator governing TCR signaling, cell cycle progression, and T cell death., Competing Interests: Competing interests statement:The authors declare no competing interest.

Inhibitory receptors are critical for regulating immune cell function. In cancer, these receptors are often over-expressed on the cell surface of T and NK cells, leading to reduced anti-tumor activity. Here, through the analysis of 11 commonly studied checkpoint and inhibitory receptors, we discern that only HAVCR2 (TIM3) and ENTPD1 (CD39) display significantly greater gene expression in glioblastoma compared to normal brain and lower grade glioma. Cell surface TIM-3, but not ENTPD1, was also elevated on activated CD4 + and CD8 + T cells, as well as on NK cells from glioblastoma patients compared to healthy donor T and NK cells. A subsequent analysis of molecules known to co-ordinate TIM-3 function and regulation was performed, which revealed that BAT3 expression was significantly reduced in CD4 + and CD8 + T cells, as well as NK cells from glioblastoma patients compared to counterparts from healthy donors. These pro-inhibitory changes are also correlated with reduced levels of the activation marker CD69 and the pro-inflammatory cytokine IFNγ in CD4 + and CD8 + T cells, as well as NK cells from glioblastoma patients. Collectively, these data reveal that glioblastoma-mediated CD4 + and CD8 + T cell and NK cell suppression is due, at least in part, to dysregulated TIM-3 and BAT3 expression and the associated downstream immunoregulatory and dysfunctional effects.

Ribosomal biosynthesis in nucleoli is an energy-demanding process driven by all RNA polymerases and hundreds of auxiliary proteins. We investigated how this process is regulated in activated T lymphocytes by T cell receptor (TCR) signals and the multiprotein complexes mTORC1 and mTORC2, both of which contain the kinase mTOR. Deficiency in mTORC1 slowed the proliferation of T cells, with further delays in each consecutive division, an effect not seen with deficiency in mTORC2. mTORC1 signaling was stimulated by components of conventional TCR signaling, and, reciprocally, TCR sensitivity was decreased by mTORC1 inhibition. The substantial increase in the amount of RNA per cell induced by TCR activation was reduced by 50% by deficiency in mTORC1, but not in mTORC2 or in S6 kinases 1 and 2, which are activated downstream of mTORC1. RNA-seq data showed that mTORC1 deficiency reduced the abundance of all RNA biotypes, although rRNA processing was largely intact in activated T cells. Imaging cytometry with FISH probes for nascent pre-rRNA revealed that deletion of mTORC1, but not that of mTORC2, reduced the number and expansion of nucleolar sites of active transcription. Protein translation was consequently decreased by 50% in the absence of mTORC1. Inhibiting RNA polymerase I blocked not only proliferation but also mTORC1 signaling. Our data show that TCR signaling, mTORC1 activity, and ribosomal biosynthesis in the nucleolus regulate each other during biomass production in clonally expanding T cells.

Identification and isolation of antigen-specific T cells for downstream transcriptomic analysis is key for various immunological studies. Traditional methods using major histocompatibility complex (MHC) multimers are limited by the number of predefined immunodominant epitopes and MHC matching of the study subjects. Activation-induced markers (AIM) enable highly sensitive detection of rare antigen-specific T cells irrespective of the availability of MHC multimers. Herein, we have developed an AIM assay for the detection, sorting and subsequent single-cell RNA sequencing (scRNA-seq) analysis of hepatitis C virus (HCV)-specific T cells. We examined different combinations of the activation markers CD69, CD40L, OX40, and 4-1BB at 6, 9, 18 and 24 h post stimulation with HCV peptide pools. AIM + CD4 T cells exhibited upregulation of CD69 and CD40L as early as 6 h post-stimulation, while OX40 and 4-1BB expression was delayed until 18 h. AIM + CD8 T cells were characterized by the coexpression of CD69 and 4-1BB at 18 h, while the expression of CD40L and OX40 remained low throughout the stimulation period. AIM + CD4 and CD8 T cells were successfully sorted and processed for scRNA-seq analysis examining gene expression and T cell receptor (TCR) usage. scRNA-seq analysis from this one subject revealed that AIM + CD4 T (CD69 + CD40L + ) cells predominantly represented Tfh, Th1, and Th17 profiles, whereas AIM + CD8 T (CD69 + 4-1BB + ) cells primarily exhibited effector and effector memory profiles. TCR analysis identified 1023 and 160 unique clonotypes within AIM + CD4 and CD8 T cells, respectively. In conclusion, this approach offers highly sensitive detection of HCV-specific T cells that can be applied for cohort studies, thus facilitating the identification of specific gene signatures associated with infection outcome and vaccination.

Limited knowledge is available on the differences in severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) specific antibody breadth and T cell differentiation among different COVID-19 sequential vaccination strategies. In this study, we compared the immunogenicity of the third different dose of COVID-19 vaccines, such as mRNA (I-I-M), adenoviral vector (I-I-A), and recombinant protein (I-I-R) vaccines, in terms of the magnitude and breadth of antibody response and differentiation of SARS-CoV-2-specific T and B cells. These studies were performed in the same clinical trial, and the samples were assessed in the same laboratory. IGHV1-69, IGHV3-9, and IGHV4-34 were the dominant B cell receptor (BCR) usages of the I-I-M, I-I-A, and I-I-R groups, respectively; the RBD + B cell activation capacities were comparable. Additionally, the I-I-R group was characterized by higher numbers of regulatory T cells, circulating T follicular helper cells (cTFH) - cTFH1 (CXRC3 + CCR6 - ), cTFH1-17 (CXRC3 + CCR6 + ), cTFH17 (CXRC3 - CCR6 + ), and cTFH-CM (CD45RA - CCR7 + ), and lower SMNE + T cell proliferative capacity than the other two groups, whereas I-I-A showed a higher proportion and number of virus-specific CD4 + T cells than I-I-R, as determined in ex vivo experiments. Our data confirmed different SARS-CoV-2-specific antibody profiles among the three different vaccination strategies and also provided insights regarding BCR usage and T/B cell activation and differentiation, which will guide a better selection of vaccination strategies in the future., Importance: Using the same laboratory test to avoid unnecessary interference due to cohort ethnicity, and experimental and statistical errors, we have compared the T/B cell immune response in the same cohort sequential vaccinated by different types of COVID-19 vaccine. We found that different sequential vaccinations can induce different dominant BCR usage with no significant neutralizing titers and RBD + B-cell phenotype. Recombinant protein vaccine can induce higher numbers of regulatory T cells, circulating TFH (CTFH)1, CTFH17, and CTFH-CM, and lower SMNE + T-cell proliferative capacity than the other two groups, whereas I-I-A showed higher proportion and number of virus-specific CD4 + T cells than I-I-R. Overall, our study provides a deep insight about the source of differences in immune protection of different types of COVID-19 vaccines, which further improves our understanding of the mechanisms underlying the immune response to SARS-CoV-2., Competing Interests: The authors declare no conflict of interest.

SATB1, a key regulator of T cell development, governs lineage-specific transcriptional programs upon T cell activation. The absence of SATB1 has been linked to the initiation and progression of autoimmunity. However, its precise roles in this process remain incompletely understood. Here we show that conditional knockout of Satb1 in CD4 + T cells in mice led to T cell hyperactivation and inflammatory cell infiltration across multiple organs. Transcriptional profiling on activated T cells revealed that the loss of SATB1 led to aberrant upregulation of CC chemokines. Treating Satb1 conditional knockout mice with CC chemokine receptor inhibitor alleviated inflammatory cell infiltration. Intriguingly, SATB1's transcriptional regulation of chemokine genes could not be attributed to its direct binding to chemokine promoters. Instead, SATB1 exerted its regulatory effects by controlling higher-order chromatin organization at a CC chemokine locus. The loss of SATB1 led to the emergence of a new chromatin domain encompassing the Ccl3, Ccl4, Ccl5, Ccl6, and Ccl9 genes and a distal enhancer, resulting in increased contacts between the enhancer and all five chemokine genes, thus inducing their upregulation. Collectively, these results demonstrate that SATB1 protects organs from immune cell infiltration by regulating chemokine expression, providing valuable insights into the development of autoimmunity-related phenotypes., (© 2024. The Author(s).)

The molecular mechanisms responsible for the heightened reactivity of quiescent T cells in human early life remain largely elusive. Our previous research identified that quiescent adult naïve CD4 + T cells express LINE1 (long interspersed nuclear elements 1) spliced in previously unknown isoforms, and their down-regulation marks the transition to activation. Here, we unveil that neonatal naïve T cell quiescence is characterized by enhanced energy production and protein synthesis. This phenotype is associated with the absence of LINE1 expression attributed to tonic T cell receptor/mTOR complex 1 (mTORC1) signaling and (polypyrimidine tract-binding protein 1 (PTBP1)-mediated LINE1 splicing suppression. The absence of LINE1 expression primes these cells for rapid execution of the activation program by directly regulating protein synthesis. LINE1 expression progressively increases in childhood and adults, peaking in elderly individuals, and, by decreasing protein synthesis, contributes to immune senescence in aging. Our study proposes LINE1 as a critical player of human T cell function across the human life span.

Immune cells are highly dynamic and able to migrate through environments with diverse biochemical and mechanical compositions. Their migration has classically been defined as amoeboid under the assumption that it is integrin independent. Here, we show that activated primary Th1 T cells require both confinement and extracellular matrix proteins to migrate efficiently. This migration is mediated through small and dynamic focal adhesions that are composed of the same proteins associated with canonical mesenchymal cell focal adhesions, such as integrins, talin, and vinculin. These focal adhesions, furthermore, localize to sites of contractile traction stresses, enabling T cells to pull themselves through confined spaces. Finally, we show that Th1 T cells preferentially follow tracks of other T cells, suggesting that these adhesions modify the extracellular matrix to provide additional environmental guidance cues. These results demonstrate not only that the boundaries between amoeboid and mesenchymal migration modes are ambiguous, but that integrin-mediated focal adhesions play a key role in T cell motility., (© 2024 Caillier et al.)

There is considerable evidence that the immune system plays a role in hypertension, however this role is not fully characterized. Our previous studies demonstrated that mesenteric perivascular adipose tissue (mPVAT) harbors a large T cell population, which is a cell type identified as contributing to hypertension. In the present study, we tested the hypothesis that soluble mediators in mPVAT influence T cell function just prior to the development of hypertension. Toward this end, we utilized a unique model of hypertension in which Dahl S rats on a high fat (HF) diet develop hypertension. We found that conditioned media (CM) from mPVAT from healthy Dahl S rats on control diet buffers T cell activation, however, mPVAT-CM from Dahl S rats on a HF diet markedly increased inflammatory cytokine induction (IFNγ, GM-CSF and IL-17a) by activated T cells. These cytokines are known to promote activation of macrophages and neutrophils, among other effects. Conversely, the anti-inflammatory cytokine, IL-10, was not different between the groups, suggesting the effect is selective for inflammatory cytokines. Furthermore, we conducted bulk RNA-seq on activated T cells cultured in mPVAT-CM from Dahl S rats on either control (CTL) or HF diet for 10 weeks. In accordance with the cytokine analysis, mPVAT-CM from HF diet-fed rats significantly upregulated many genes associated with IFNγ/IL-17 induction, whereas Th2/Treg-associated genes were downregulated. Taken together, these data strongly suggest soluble mediators from mPVAT influence T cell inflammatory status and may promote Th1/Th17 differentiation preceding the development of hypertension triggered by HF diet., Competing Interests: The authors have declared no competing interests., (Copyright: © 2024 Jin et al. This is an open access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.)

Chronic obstructive pulmonary disease (COPD) is regarded as an accelerated-age disease in which chronic inflammation, maladaptive immune responses, and senescence cell burden coexist. Accordingly, cellular senescence has emerged as a potential mechanism involved in COPD pathophysiology. In this study, 25 stable patients with COPD underwent a daily physical activity promotion program for 6 mo. We reported that increase of physical activity was related to a reduction of the senescent cell burden in circulating lymphocytes of patients with COPD. Senescent T-lymphocyte population, characterized by absence of surface expression of CD28, was reduced after physical activity intervention, and the reduction was associated to the increase of physical activity level. In addition, the mRNA expression of cyclin-dependent kinase inhibitors, a hallmark of cell senescence, was reduced and, in accordance, the proliferative capacity of lymphocytes was improved postintervention. Moreover, we observed an increase in functionality in T cells from patients after intervention, including improved markers of activation, enhanced cytotoxicity, and altered cytokine secretions in response to viral challenge. Lastly, physical activity intervention reduced the potential of lymphocytes' secretome to induce senescence in human primary fibroblasts. In conclusion, our study provides, for the first time, evidence of the potential of physical activity intervention in patients with COPD to reduce the senescent burden in circulating immune cells. NEW & NOTEWORTHY For the first time, we identified in patients with COPD a relation between physical activity intervention with respiratory function improvement and cellular senescence burden in lymphocytes that improved the T cell functionality and proliferative capacity of patients. In addition, our experiments highlight the possible impact of T-cell senescence in other cell types which could be related to some of the clinical lung complications observed in COPD.

N6-methyladenosine (m6A) is a RNA modification that can regulate post-transcriptional processes including RNA stability, translation, splicing, and nuclear export. In CD4+ lymphocytes, m6A modifications have been demonstrated to play a role in early differentiation processes. The role of m6A in CD4+ T cell activation and effector function remains incompletely understood. To assess the role of m6A in CD4+ T lymphocyte activation and function, we assessed the transcriptome-wide m6A landscape of human primary CD4+ T cells by methylated RNA immunoprecipitation sequencing. Stimulation of the T cells impacted the m6A pattern of hundreds of transcripts including tumor necrosis factor (TNF). m6A methylation was increased on TNF messenger RNA (mRNA) after activation, predominantly in the 3' untranslated region of the transcript. Manipulation of m6A levels in primary human T cells, the directly affected the expression of TNF. Furthermore, we identified that the m6A reader protein YTHDF2 binds m6A-methylated TNF mRNA, and promotes its degradation. Taken together, this study demonstrates that TNF expression in CD4+ T lymphocytes is regulated via m6A and YTHDF2, thereby providing novel insight into the regulation of T cell effector functions., Competing Interests: Conflict of interest statement. None declared., (© The Author(s) 2024. Published by Oxford University Press on behalf of Society for Leukocyte Biology.)

Expansion of CD4 + CD28 null T-lymphocytes is common in chronic heart failure (CHF) patients. Its ability to produce high levels of proinflammatory cytokines is probably the key role of these cells in CHF. IL-10 is a candidate for limiting CD4 + CD28 null T-lymphocyte responses, whereas tumour necrosis factor (TNF) is the cytokine most closely involved in the loss of CD28 expression. Serum levels of TNF and IL-10 were measured in 65 CHF patients (mean age, 65.2 ± 13.84 years). Patients with an IL-10/TNF ratio ≥1 had significantly lower levels of CD4 + CD28 null T-lymphocytes than those with a ratio <1. In vitro, IL-10 reduced the frequency of proliferative CD4 + CD28 null T-lymphocytes stimulated with anti-CD3. Pre-treatment with IL-10 before anti-CD3 stimulation was required for the cytokine to inhibit TNF production by CD4 + CD28 null T-lymphocytes. In addition to the previously described effect of IL-10 on HLA-DR and ICAM-1 expression, LFA-3 protein and mRNA levels were reduced in the presence of the cytokine in monocytes. IL-10 inhibition on CD4 + CD28 null T-lymphocytes may be mediated by a reduction in HLA class II and LFA-3 expression because blocking interactions with these costimulators has similar effects to those of IL-10 treatment. Moreover, costimulation through CD2/LFA-3 interaction is enough to induce proliferation and cytokine production in CD4 + CD28 null T-lymphocytes., (© 2024 John Wiley & Sons Ltd.)

As one of subunits for interleukin-2 receptor (IL-2R), CD122 can bind to IL-2 and then activate downstream signal transduction to participate in adaptive immune response. Although CD122 has been identified and investigated from several teleost species, studies on its function at T-cell level are still scarce for lack of specific antibodies. In this study, a typical CD122 in Nile tilapia (Oreochromis niloticus) was characterized by bioinformatics analysis, cloned to produce retrovirus infected NIH/3T3 cells for mouse immunization. After cell fusion and screening, we successfully developed a mouse anti-tilapia CD122 monoclonal antibody (mAb), which could specifically recognize CD122 and identify CD122-producing T cells of tilapia. Using the mAb to detect, CD122 was found to widely distribute in immune-related tissues, and significantly elevate post Edwardsiella piscicida infection or T-cell activation. More importantly, the expansion of CD122 + T cells and up-regulation of CD122 occurred both in total T cells and T-cell subsets during T-cell activation upon in vitro stimulation or in vivo infection. These results indicate that CD122 can be used as a T-cell activation marker in tilapia. Notably, CD122 mAb blocking blunted the activation of MAPK/Erk and mTORC1 pathways, and inhibited T-cell proliferation, suggesting a critical role of CD122 in ensuring proper proliferation of tilapia T cells. Therefore, this study enriches the knowledge of T-cell responses in fish and provides new evidence for understanding the evolution of lymphocyte-mediated adaptive immunity., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Atherosclerosis is a complex pathological process that results from the chronic inflammatory reaction of the blood vessel wall and involves various immune cells and cytokines. An imbalance in the proportion and function of the effector CD4 + T-cell (Teff) and regulatory T-cell (Treg) subsets is an important cause of the occurrence and development of atherosclerotic plaques. Teff cells depend on glycolytic metabolism and glutamine catabolic metabolism for energy, while Treg cells mainly rely on fatty acid oxidation (FAO), which is crucial for determining the fate of CD4 + T cells during differentiation and maintaining their respective immune functions. Here, we review recent research achievements in the field of immunometabolism related to CD4 + T cells, focusing on the cellular metabolic pathways and metabolic reprogramming involved in the activation, proliferation, and differentiation of CD4 + T cells. Subsequently, we discuss the important roles of mTOR and AMPK signaling in regulating CD4 + T-cell differentiation. Finally, we evaluated the links between CD4 + T-cell metabolism and atherosclerosis, highlighting the potential of targeted modulation of CD4 + T-cell metabolism in the prevention and treatment of atherosclerosis in the future., (© 2023. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

The commitment to specific T lymphocytes (T cell) lineages is governed by distinct transcription factors, whose expression is modulated through epigenetic mechanisms. Unravelling these epigenetic mechanisms that regulate T cell differentiation and function holds significant importance for understanding T cells. Menin, a multifunctional scaffolding protein, is implicated in various cellular processes, such as cell proliferation, cell cycle control, DNA repair and transcriptional regulation, primarily through epigenetic mechanisms. Existing research indicates Menin's impact on T cell differentiation and function, while a comprehensive and systematic review is currently lacking to consolidate these findings. In the current review, we have highlighted recent studies on the role of Menin in T cell differentiation and function, focusing mainly on its impact on the memory Th2 maintenance, Th17 differentiation and maintenance, CD4 + T cell senescence, and effector CD8 + T cell survival. Considering Menin's crucial function in maintaining effector T cell function, the potential of inhibiting Menin activity in mitigating inflammatory diseases associated with excessive T cell activation has also been emphasised., (© 2024 John Wiley & Sons Ltd.)

Polymorphisms in the IRGM gene are associated with susceptibility to tuberculosis in humans. A murine ortholog of Irgm, Irgm1, is also essential for controlling Mycobacterium tuberculosis (Mtb) infection in mice. Multiple processes have been associated with IRGM1 activity that could impact the host response to Mtb infection, including roles in autophagy-mediated pathogen clearance and expansion of activated T cells. However, what IRGM1-mediated pathway is necessary to control Mtb infection in vivo and the mechanistic basis for this control remains unknown. We dissected the contribution of IRGM1 to immune control of Mtb pathogenesis in vivo and found that Irgm1 deletion leads to higher levels of IRGM3-dependent type I interferon signaling. The increased type I interferon signaling precludes T cell expansion during Mtb infection. The absence of Mtb-specific T cell expansion in Irgm1 -/- mice results in uncontrolled Mtb infection in neutrophils and alveolar macrophages, which directly contributes to susceptibility to infection. Together, our studies reveal that IRGM1 is required to promote T cell-mediated control of Mtb infection in neutrophils, which is essential for the survival of Mtb-infected mice. These studies also uncover new ways type I interferon signaling can impact T H 1 immune responses., Competing Interests: Declaration of Competing Interest The authors declare the following financial interests/personal relationships which may be considered as potential competing interests: Christina Stallings reports financial support was provided by National Institute of Health. Michael McNehlan reports financial support was provided by National Institute of Health. Rachel Kinsella reports financial support was provided by Potts Memorial Foundation. Christina Stallings reports financial support was provided by Burroughs Wellcome Fund. If there are other authors, they declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 The Authors. Published by Elsevier Inc. All rights reserved.)

Background: Major depressive disorder (MDD) is characterized by increased T helper (Th)1 polarization, T cell activation (e.g., CD71+ and CD40L+), and cannabinoid receptor type 2 bearing CD20+ B cells; and lower T regulatory (Treg) numbers., Aims: To delineate the effects of adverse childhood experiences (ACEs) and recurrence of illness (ROI) on activated T and CB2-bearing B populations, and Tregs, including FoxP3 + CD152+, FoxP3 + GARP+, and FoxP3 + CB1+ cells., Methods: We measured ROI, ACEs, the number of activated T cells, Tregs, and CD20 + CB2+ B cells, in 30 MDD patients and 20 healthy controls., Results: A larger part of the variance in the depression phenome (40.8 %) was explained by increased CD20 + CB2+ and activated T cells, and lowered Tregs. ROI and lifetime suicidal behaviors were significantly and positively associated with CD20 + CB2+, CD3 + CD71+, CD3 + CD40L+, CD4 + CD71+, CD4 + CD40L+, and CD4HLADR+ numbers. ROI was significantly correlated with CD8 + CD40L+ numbers. The sum of ACEs was significantly associated with CD20 + CB2+, CD3 + CD40L+, CD4 + 40 L+ numbers, T cell activation (positively) and Treg (inversely) indices. One replicable latent vector could be extracted from activated T cells, lifetime and current suicidal behaviors, number of depressive episodes, and severity of depression, and 48.8 % of its variance was explained by ACEs., Conclusions: ACE-induced activation of T effector and cytotoxic cells and B cells with autoimmune potential, coupled with lowered Treg numbers are a key component of depression. The findings indicate that increasing ROI, the phenome of depression and suicidal behaviors, are caused by autoimmune processes, which are the consequence of ACEs and increasing sensitization of immune responses., Competing Interests: Declaration of competing interest The authors have no conflict of interest with any commercial or other association in connection with the submitted article., (Copyright © 2024 Elsevier B.V. All rights reserved.)