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Introduction: CD8+ cytotoxic T lymphocytes (CTLs) are highly effective in defending against viral infections and tumours. They are activated through the recognition of peptide-MHC-I complex by the T-cell receptor (TCR) and co-stimulation. This cognate interaction promotes the organisation of intimate cell-cell connections that involve cytoskeleton rearrangement to enable effector function and clearance of the target cell. This is key for the asymmetric transport and mobilisation of lytic granules to the cell-cell contact, promoting directed secretion of lytic mediators such as granzymes and perforin. Mitochondria play a role in regulating CTL function by controlling processes such as calcium flux, providing the necessary energy through oxidative phosphorylation, and its own protein translation on 70S ribosomes. However, the effect of acute inhibition of cytosolic translation in the rapid response after TCR has not been studied in mature CTLs. Methods: Here, we investigated the importance of cytosolic protein synthesis in human CTLs after early TCR activation and CD28 co-stimulation for the dynamic reorganisation of the cytoskeleton, mitochondria, and lytic granules through short-term chemical inhibition of 80S ribosomes by cycloheximide and 80S and 70S by puromycin. Results: We observed that eukaryotic ribosome function is required to allow proper asymmetric reorganisation of the tubulin cytoskeleton and mitochondria and mTOR pathway activation early upon TCR activation in human primary CTLs. Discussion: Cytosolic protein translation is required to increase glucose metabolism and degranulation capacity upon TCR activation and thus to regulate the full effector function of human CTLs.
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Linfocitos T CD8-positivos , Citosol , Activación de Linfocitos , Mitocondrias , Biosíntesis de Proteínas , Receptores de Antígenos de Linfocitos T , Humanos , Receptores de Antígenos de Linfocitos T/metabolismo , Receptores de Antígenos de Linfocitos T/inmunología , Activación de Linfocitos/inmunología , Citosol/metabolismo , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Mitocondrias/metabolismo , Mitocondrias/inmunología , Citoesqueleto/metabolismo , Linfocitos T Citotóxicos/inmunología , Linfocitos T Citotóxicos/metabolismo , Ribosomas/metabolismo , Transducción de Señal , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
A 10-year-old spayed female Dachshund presented with abdominal pain and generalized severe ileus. An exploratory laparotomy was performed, confirming a severe ileus of undetermined origin. Multiple intestinal biopsy results confirmed acute intestinal leiomyositis. Immunohistochemistry (IHC) stains confirmed a T-cell predominant inflammatory infiltrate. Intravenous immunoglobulin (hIVIG) was administered prior to immunosuppressive therapy. Within 10 days of hIVIG treatment, functional peristaltic activity returned, and symptoms resolved. Long-term management, including the use of mycophenolate, resulted in sustained functional peristaltic recovery. Further studies are needed to explore the potential benefits of hIVIG treatment in the stabilization phase of this commonly fatal, treatment-refractory disease.
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BACKGROUND: Relapsed medulloblastoma (MB) poses a significant therapeutic challenge due to its highly immunosuppressive tumor microenvironment. Immune checkpoint inhibitors (ICIs) have struggled to mitigate this challenge, largely due to low T-cell infiltration and minimal PD-L1 expression. Identifying the mechanisms driving low T-cell infiltration is crucial for developing more effective immunotherapies. METHODS: We utilize a syngeneic mouse model to investigate the tumor immune microenvironment of MB and compare our findings to transcriptomic and proteomic data from human MB. RESULTS: Flow cytometry reveals a notable presence of CD45hi/CD11bhi macrophage-like and CD45int/CD11bint microglia-like tumor-associated macrophages (TAMs), alongside regulatory T-cells (Tregs), expressing high levels of the inhibitory checkpoint molecule VISTA. Compared to sham control mice, the CD45hi/CD11bhi compartment significantly expands in tumor-bearing mice and exhibits a myeloid-specific signature composed of VISTA, CD80, PD-L1, CTLA-4, MHCII, CD40, and CD68. These findings are corroborated by proteomic and transcriptomic analyses of human MB samples. Immunohistochemistry highlights an abundance of VISTA-expressing myeloid cells clustering at the tumor-cerebellar border, while T-cells are scarce and express FOXP3. Additionally, tumor cells exhibit immunosuppressive properties, inhibiting CD4 T-cell proliferation in vitro. Identification of VISTA's binding partner, VSIG8, on tumor cells, and its correlation with increased VISTA expression in human transcriptomic analyses suggests a potential therapeutic target. CONCLUSIONS: This study underscores the multifaceted mechanisms of immune evasion in MB and highlights the therapeutic potential of targeting the VISTA-VSIG axis to enhance anti-tumor responses.
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BACKGROUND AND 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 MHC class II (HLA-DQ2.5), which facilitates gluten antigen recognition by CD4+ T cells in CeD. METHODS: Epithelial MHC class II (MHCII) was determined in active and treated CeD, and in non-immunized and gluten-immunized DR3-DQ2.5 transgenic mice, lacking mouse MHCII molecules. Organoid monolayers from DR3-DQ2.5 mice were treated with or without IFN-γ, and MHCII expression was evaluated by flow cytometry. Organoid monolayers and CD4+ T cell co-cultures were incubated with gluten, pre-digested, 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: Active CeD patients 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 IL-2, IFN-γ, and IL-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 pre-digestion with microbial elastase. Therapeutics directed at IECs may offer a novel approach for modulating both adaptive and innate immunity in CeD patients.
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We developed a highly sensitive assay for detecting protein-protein interaction using chimeric receptors comprising two molecules of interest in the extracellular domain and interferon alpha and beta receptor subunit 1 or 2 (IFNAR1/2) in the intracellular domain. This intracellular IFNAR1/2 reconstitution system (IFNARRS) proved markedly more sensitive than the NanoBiT system, currently considered one of the best detection systems for protein interaction. Employing chimeric receptors with extracellular domains from the IFNγ or IL-2 receptor and the intracellular domains of IFNAR1/2, the IFNARRS system effectively identifies low IFNγ or IL-2 levels. Cells stably expressing these chimeric receptors responded to IFNγ secreted by activated T cells following various stimuli, including a specific peptide-antigen. The activation signals were further enhanced by the expression of relevant genes, such as costimulators, via IFN-stimulated response elements in the promoters. Besides IFNγ or IL-2, the IFNARRS system demonstrated the capability to detect other cytokines by using the corresponding extracellular domains from these target cytokine receptors.
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Interferón gamma , Interleucina-2 , Receptor de Interferón alfa y beta , Linfocitos T , Humanos , Receptor de Interferón alfa y beta/metabolismo , Receptor de Interferón alfa y beta/genética , Linfocitos T/metabolismo , Linfocitos T/inmunología , Interleucina-2/metabolismo , Interferón gamma/metabolismo , Receptores de Interleucina-2/metabolismo , Receptores de Interleucina-2/genética , Unión Proteica , Activación de Linfocitos , Células HEK293RESUMEN
Recent in vitro studies of human sex chromosome aneuploidy showed that the Xi ("inactive" X) and Y chromosomes broadly modulate autosomal and Xa ("active" X) gene expression. We tested these findings in vivo. Linear modeling of CD4+ T cells and monocytes from individuals with one to three X chromosomes and zero to two Y chromosomes revealed 82 sex-chromosomal and 344 autosomal genes whose expression changed significantly with Xi and/or Y dosage in vivo. Changes in sex-chromosomal expression were remarkably constant in vivo and in vitro; autosomal responses to Xi and/or Y dosage were largely cell-type specific (â¼2.6-fold more variation than sex-chromosomal responses). Targets of the sex-chromosomal transcription factors ZFX and ZFY accounted for a significant fraction of these autosomal responses both in vivo and in vitro. We conclude that the human Xi and Y transcriptomes are surprisingly robust and stable, yet they modulate autosomal and Xa genes in a cell-type-specific fashion.
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One of the biggest challenges for in vivo gene therapy are vectors mediating highly selective gene transfer into a defined population of therapy-relevant cells. Here we present DARPin-targeted AAVs (DART-AAVs) displaying DARPins specific for human and murine CD8. Insertion of DARPins into the GH2/GH3 loop of the capsid protein 1 (VP1) of AAV2 and AAV6 resulted in high selectivity for CD8-positive T cells with unimpaired gene delivery activity. Remarkably, the capsid core structure was unaltered with protruding DARPins detectable. In complex primary cell mixtures, including donor blood or systemic injections into mice, the CD8-targeted AAVs were by far superior to unmodified AAV2 and AAV6 in terms of selectivity, target cell viability, and gene transfer rates. In vivo, up to 80% of activated CD8+ T cells were hit upon a single vector injection into conditioned humanized or immunocompetent mice. While gene transfer rates decreased significantly under non-activated conditions, genomic modification selectively in CD8+ T cells was still detectable upon Cre delivery into indicator mice. In both mouse models, selectivity for CD8+ T cells was close to absolute with exceptional detargeting from liver. The CD8-AAVs described here expand strategies for immunological research and in vivo gene therapy options.
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Methionine, an indispensable amino acid crucial for dietary balance, intricately governs metabolic pathways. Disruption in its equilibrium has the potential to heighten homocysteine levels in both plasma and tissues, posing a conceivable risk of inducing inflammation and detriment to the integrity of vascular endothelial cells. The intricate interplay between methionine metabolism, with a specific focus on S-adenosyl-L-methionine (SAM), and the onset of thoracic aortic dissection (TAD) remains enigmatic despite acknowledging the pivotal role of inflammation in this vascular condition. In an established murine model induced by ß-aminopropionitrile monofumarate (BAPN), we delved into the repercussions of supplementing with S-adenosyl-L-methionine (SAM) on the progression of TAD. Our observations uncovered a noteworthy improvement in aortic dissection and rupture rates, accompanied by a marked reduction in mortality upon SAM supplementation. Notably, SAM supplementation exhibited a considerable protective effect against BAPN-induced degradation of elastin and the extracellular matrix. Furthermore, SAM supplementation demonstrated a robust inhibitory influence on the infiltration of immune cells, particularly neutrophils and macrophages. It also manifested a notable reduction in the inflammatory polarization of macrophages, evident through diminished accumulation of MHC-IIhigh macrophages and reduced expression of inflammatory cytokines such as IL1ß and TNFα in macrophages. Simultaneously, SAM supplementation exerted a suppressive effect on the activation of CD4 + and CD8 + T cells within the aorta. This was evidenced by an elevated proportion of CD44- CD62L + naïve T cells and a concurrent decrease in CD44 + CD62L- effector T cells. In summary, our findings strongly suggest that the supplementation of SAM exhibits remarkable efficacy in alleviating BAPN-induced aortic inflammation, consequently impeding the progression of thoracic aortic dissection.
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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.
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The development of effective vaccines against Hepatitis C Virus (HCV) remains a global health priority and challenge. In this study, we employed an integrative approach combining computational epitope prediction with experimental validation to identify immunogenic peptides targeting the E1 glycoprotein of HCV. In the present report, computational data from various epitope prediction algorithms such as IEDB and SYFPEITHI, followed by molecular dynamics (MD) simulations and immuno-informatics analysis is presented. Through computational screening, we identified potential epitope candidates, with QVRNSSGLY (P3) and QLFTFSPRH (P7) emerging as promising candidates. MD simulations revealed stable interactions between these epitopes and MHC molecule, further validated by free energy estimations using MMPBSA method. Immuno-informatics analysis supported these findings, showing high binding potential and immunogenicity scores for the selected peptides. Subsequent synthesis and characterization of epitope peptides confirmed their structural integrity and purity required for conducting immune activation assays. Experimental immunological assays carried out in this study involved epitope peptide induced activation of CD8 + and CD4 + T cells from healthy human subjects and HCV- recovered patients. Data from experimental validation revealed significant cytokine release upon exposure to epitope peptides, particularly TNF-a, IL-6, and GM-CSF, indicative of robust immune responses. Notably, peptides P3 and P7 exhibited the most pronounced cytokine induction profiles, underscoring their potential as vaccine candidates. Further investigations addressing the mechanism of action of these epitope peptides under preclinical and clinical settings may help in developing effective vaccine against HCV.
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T-cell engaging (TCE) bispecific antibodies are potent drugs that trigger the immune system to eliminate cancer cells, but administration can be accompanied by toxic side effects that limit dosing. TCEs function by binding to cell surface receptors on T cells, frequently CD3, with one arm of the bispecific antibody while the other arm binds to cell surface antigens on cancer cells. On-target, off-tumor toxicity can arise when the target antigen is also present on healthy cells. The toxicity of TCEs may be ameliorated through the use of pro-drug forms of the TCE, which are not fully functional until recruited to the tumor microenvironment. This can be accomplished by masking the anti-CD3 arm of the TCE with an autoinhibitory motif that is released by tumor-enriched proteases. Here, we solve the crystal structure of the antigen-binding fragment of a novel anti-CD3 antibody, E10, in complex with its epitope from CD3 and use this information to engineer a masked form of the antibody that can activate by the tumor-enriched protease matrix metalloproteinase 2 (MMP-2). We demonstrate with binding experiments and in vitro T-cell activation and killing assays that our designed prodrug TCE is capable of tumor-selective T-cell activity that is dependent upon MMP-2. Furthermore, we demonstrate that a similar masking strategy can be used to create a pro-drug form of the frequently used anti-CD3 antibody SP34. This study showcases an approach to developing immune-modulating therapeutics that prioritizes safety and has the potential to advance cancer immunotherapy treatment strategies.
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Anticuerpos Biespecíficos , Complejo CD3 , Inmunoterapia , Profármacos , Linfocitos T , Anticuerpos Biespecíficos/inmunología , Anticuerpos Biespecíficos/farmacología , Humanos , Complejo CD3/inmunología , Inmunoterapia/métodos , Linfocitos T/inmunología , Profármacos/farmacología , Profármacos/química , Neoplasias/inmunología , Neoplasias/terapia , Neoplasias/tratamiento farmacológico , Ingeniería de Proteínas/métodos , Metaloproteinasa 2 de la Matriz/inmunologíaRESUMEN
Sialic acids are found as terminal sugars on glycan structures on cellular surfaces. T cells carry these sialoglycans abundantly, and they are thought to serve multiple functions in cell adhesion, cell migration, and protection from complement attack. We studied the role of sialoglycans on T cells in a mouse model with a T cell-specific deletion of cytidine monophosphate-sialic acid synthase (CMAS), the enzyme that is crucial for the synthesis of sialoglycans. These mice showed a T-cell deficiency in peripheral lymphoid organs. Many T cells with an undeleted Cmas allele were found in the periphery, suggesting that they escaped the Cre-mediated deletion. The remaining peripheral T cells of T cell-specific Cmas KO mice had a memory-like phenotype. Additional depletion of the complement factor C3 could not rescue the phenotype, showing that the T-cell defect was not caused by a host complement activity. Cmas-deficient T cells showed a high level of activated caspase 3, indicating an ongoing apoptosis. In bone marrow chimeric cellular transfer experiments, we observed a strong competitive disadvantage of Cmas-deficient T cells compared to wild-type T cells. These results show that sialoglycans on the surface of T cells are crucial for T-cell survival and maintenance. This function has not been recognized before and is similar to the function of sialoglycans on B cells.
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Ratones Noqueados , Ácidos Siálicos , Linfocitos T , Animales , Ratones , Linfocitos T/inmunología , Linfocitos T/metabolismo , Ácidos Siálicos/metabolismo , Supervivencia Celular , Ratones Endogámicos C57BL , Apoptosis , Complemento C3/metabolismo , Complemento C3/inmunología , Complemento C3/genética , Oxigenasas de Función MixtaRESUMEN
Inadequate antigen-specific T cells activation hampers immunotherapy due to complex antigen presentation. In addition, therapeutic in vivo T cell expansion is constrained by slow expansion rates and limited functionality. Herein, we introduce a model fusion protein termed antigen-presenting cell-mimic fusion protein (APC-mimic), designed to greatly mimicking the natural antigen presentation pattern of antigen-presenting cells and directly expand T cells both in vitro and in vivo. The APC-mimic comprises the cognate peptide-human leukocyte antigen (pHLA) complex and the co-stimulatory marker CD80, which are natural ligands on APCs. Following a single stimulation, APC-mimic leads to an approximately 400-fold increase in the polyclonal expansion of antigen-specific T cells compared with the untreated group in vitro without the requirement for specialized antigen-presenting cells. Through the combination of single-cell TCR sequencing (scTCR-seq) and single-cell RNA sequencing (scRNA-seq), we identify an approximately 600-fold monoclonal expansion clonotype among these polyclonal clonotypes. It also exhibits suitability for in vivo applications confirmed in the OT-1 mouse model. Furthermore, T cells expanded by APC-mimic effectively inhibits tumor growth in adoptive cell transfer (ACT) murine models. These findings pave the way for the versatile APC-mimic platform for personalized therapeutics, enabling direct expansion of polyfunctional antigen-specific T cell subsets in vitro and in vivo.
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Systemic lupus erythematosus (SLE) is a classic autoimmune disease characterized by abnormal autoantibodies, immune complex deposition, and tissue inflammation. Despite extensive research, the exact etiology and progression of SLE remain elusive. Cytidine/uridine monophosphate kinase 2 (CMPK2), a mitochondrial nucleoside monophosphate kinase, has garnered attention for its potential involvement in the development of various diseases, including SLE, where it has been observed to be dysregulated in affected individuals. However, the specific involvement of CMPK2 in the pathogenesis of SLE remains unclear. This study aims to clarify the expression level of CMPK2 in SLE CD4+ T cells and explore its impact on CD4+ T cells. The expression levels of the CMPK2 gene and the corresponding CMPK2 protein in CD4+ T cells of SLE patients were quantified using RT-qPCR and Western blot, respectively. Immunofluorescence and RT-qPCR were used to assess the mitochondrial function of SLE CD4+ T cells. Flow cytometry was used to assess CD4+ T cell activation and apoptosis levels. The impact of CMPK2 on CD4+ T cells was investigated by gene transfection experiment. We found that CMPK2 was significantly upregulated in SLE CD4+ T cells at both gene and protein levels. These cells demonstrated aberrant mitochondrial function, as evidenced by elevated mitochondrial reactive oxygen species (mtROS) levels, mitochondrial membrane potential, and mitochondrial DNA (mtDNA) copy number. Flow cytometry revealed a notable increase in both apoptosis and activation levels of CD4+ T cells in SLE patients. Gene transfection experiments showed that suppressing CMPK2 led to a significant improvement in these conditions. These findings suggest that CMPK2 may be involved in the pathogenesis of SLE by regulating mitochondrial dysfunction in CD4+ T cells and thus affecting CD4+ T cell activation and apoptosis. Our study may provide a new target for the treatment of SLE.
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High-grade serous ovarian cancer (HGSOC) is an aggressive disease known to develop resistance to chemotherapy. We investigated the prognostic significance of tumor cell states and potential mechanisms underlying chemotherapy resistance in HGSOC. Transcriptome deconvolution was performed to address cellular heterogeneity. Kaplan-Meier survival curves were plotted to illustrate the outcomes of patients with varying cellular abundances. The association between gene expression and chemotherapy response was tested. After adjusting for surgery status and grading, several cell states exhibited a significant correlation with patient survival. Cell states can organize into carcinoma ecotypes (CE). CE9 and CE10 were proinflammatory, characterized by higher immunoreactivity, and were associated with favorable survival outcomes. Ratios of cell states and ecotypes had better prognostic abilities than a single cell state or ecotype. A total of 1265 differentially expressed genes were identified between samples with high and low levels of C9 or CE10. These genes were partitioned into three co-expressed modules, which were associated with tumor cells and immune cells. Pogz was identified to be linked with immune cell genes and the chemotherapy response of paclitaxel. Collectively, the survival of HGSOC patients is correlated with specific cell states and ecotypes.
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Cistadenocarcinoma Seroso , Neoplasias Ováricas , Femenino , Humanos , Neoplasias Ováricas/mortalidad , Neoplasias Ováricas/genética , Neoplasias Ováricas/patología , Neoplasias Ováricas/inmunología , Neoplasias Ováricas/tratamiento farmacológico , Cistadenocarcinoma Seroso/genética , Cistadenocarcinoma Seroso/mortalidad , Cistadenocarcinoma Seroso/patología , Cistadenocarcinoma Seroso/inmunología , Pronóstico , Regulación Neoplásica de la Expresión Génica , Persona de Mediana Edad , Clasificación del Tumor , Transcriptoma , Estimación de Kaplan-Meier , Anciano , Resistencia a Antineoplásicos/genéticaRESUMEN
While existing synthetic technologies for ex vivo T-cell activation face challenges like suboptimal expansion rates and low effectiveness, artificial antigen-presenting cells (aAPCs) hold great promise for enhanced T-cell based therapies. In particular, gold nanoparticles (AuNPs), known for their biocompatibility, ease of synthesis, and versatile surface chemistry, are strong candidates for use as nanoscale aAPCs. In this study, we developed spiky AuNPs with branched geometries to present activating ligands to primary human T-cells. The special structure of spiky AuNPs enhances biomolecule loading capacity and significantly improves T-cell activation through multivalent binding of costimulatory ligands and receptors. Our spiky AuNPs outperform existing systems including Dynabeads and soluble activators by promoting greater polyclonal expansion of T-cells, boosting sustained cytokine production, and generating highly functional T-cells with reduced exhaustion. In addition, spiky AuNPs effectively activate and expand CD19 CAR-T cells while demonstrating increased in vitro cytotoxicity against target cells using fewer effector cells than Dynabeads. This study underscores the potential of spiky AuNPs as a powerful tool, bringing new opportunities to adoptive cell therapy applications.
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Oro , Activación de Linfocitos , Nanopartículas del Metal , Linfocitos T , Oro/química , Oro/farmacología , Humanos , Nanopartículas del Metal/química , Linfocitos T/inmunología , Linfocitos T/efectos de los fármacos , Activación de Linfocitos/efectos de los fármacosRESUMEN
T cell activation is critical for an effective immune response against pathogens. However, dysregulation contributes to the pathogenesis of autoimmune diseases, including Juvenile Idiopathic Arthritis (JIA). The molecular mechanisms underlying T cell activation are still incompletely understood. T cell activation promotes the acetylation of histone 3 at Lysine 27 (H3K27ac) at enhancer and promoter regions of proinflammatory cytokines, thereby increasing the expression of these genes which is essential for T cell function. Co-activators E1A binding protein P300 (P300) and CREB binding protein (CBP), collectively known as P300/CBP, are essential to facilitate H3K27 acetylation. Presently, the role of P300/CBP in human CD4+ T cells activation remains incompletely understood. To assess the function of P300/CBP in T cell activation and autoimmune disease, we utilized iCBP112, a selective inhibitor of P300/CBP, in T cells obtained from healthy controls and JIA patients. Treatment with iCBP112 suppressed T cell activation and cytokine signaling pathways, leading to reduced expression of many proinflammatory cytokines, including IL-2, IFN-γ, IL-4, and IL-17A. Moreover, P300/CBP inhibition in T cells derived from the inflamed synovium of JIA patients resulted in decreased expression of similar pathways and preferentially suppressed the expression of disease-associated genes. This study underscores the regulatory role of P300/CBP in regulating gene expression during T cell activation while offering potential insights into the pathogenesis of autoimmune diseases. Our findings indicate that P300/CBP inhibition could potentially be leveraged for the treatment of autoimmune diseases such as JIA in the future.
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Reactive oxygen species (ROS) are central to inter- and intracellular signaling. Their localized and transient effects are due to their short half-life, especially when generated in controlled amounts. Upon T cell receptor (TCR) activation, regulated ROS signaling is primarily initiated by complexes I and III of the electron transport chain (ETC). Subsequent ROS production triggers the activation of nicotinamide adenine dinucleotide phosphate oxidase 2 (NADPH oxidase 2), prolonging the oxidative signal. This signal then engages kinase signaling cascades such as the mitogen-activated protein kinase (MAPK) pathway and increases the activity of REDOX-sensitive transcription factors such as nuclear factor-kappa B (NF-κB) and activator protein-1 (AP-1). To limit ROS overproduction and prevent oxidative stress, nuclear factor erythroid 2-related factor 2 (Nrf2) and antioxidant proteins such as superoxide dismutases (SODs) finely regulate signal intensity and are capable of terminating the oxidative signal when needed. Thus, oxidative signals, such as T cell activation, are well-controlled and critical for cellular communication.
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Especies Reactivas de Oxígeno , Transducción de Señal , Linfocitos T , Especies Reactivas de Oxígeno/metabolismo , Humanos , Linfocitos T/metabolismo , Linfocitos T/inmunología , Animales , Activación de Linfocitos , Estrés Oxidativo , Oxidación-Reducción , Receptores de Antígenos de Linfocitos T/metabolismo , Factor 2 Relacionado con NF-E2/metabolismoRESUMEN
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.
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Experiencias Adversas de la Infancia , Trastorno Depresivo Mayor , Activación de Linfocitos , Recurrencia , Linfocitos T Reguladores , Humanos , Trastorno Depresivo Mayor/inmunología , Masculino , Femenino , Adulto , Experiencias Adversas de la Infancia/estadística & datos numéricos , Linfocitos T Reguladores/inmunología , Persona de Mediana Edad , Linfocitos B/inmunología , Estudios de Casos y ControlesRESUMEN
T cell activation is a complex biological process of naive cells maturing into effector cells. Proteomic and phospho-proteomic approaches have provided critical insights into this process, yet it is not always clear how changes in individual proteins or phosphorylation sites have functional significance. Here, we developed the Phosphorylation Integrated Thermal Shift Assay (PITSA) that combines the measurement of protein or phosphorylation site abundance and thermal stability into a single tandem mass tags experiment and apply this method to study T cell activation. We quantified the abundance and thermal stability of over 7500 proteins and 5000 phosphorylation sites and identified significant differences in chromatin-related, TCR signaling, DNA repair, and proliferative phosphoproteins. PITSA may be applied to a wide range of biological contexts to generate hypotheses as to which proteins or phosphorylation sites are functionally regulated in a given system as well as the mechanisms by which this regulation may occur.