搜索结果: 'gmp t cell activator'

AbstractBackgroundParkinson's disease (PD) is a chronic neurodegenerative disorder characterized by the loss of dopaminergic neurons in the substantia nigra,which promotes a sustained inflammatory environment in the central nervous system. Regulatory T cells (Tregs) play an important role in the control of inflammation and might play a neuroprotective role. Indeed,a decrease in Treg number and function has been reported in PD. In this context,pramipexole,a dopaminergic receptor agonist used to treat PD symptoms,has been shown to increase peripheral levels of Treg cells and improve their suppressive function. The aim of this work was to determine the effect of pramipexole on immunoregulatory Treg cells and its possible neuroprotective effect on human dopaminergic neurons differentiated from human embryonic stem cells.MethodsTreg cells were sorted from white blood cells of healthy human donors. Assays were performed with CD3/CD28?activated and non?activated Treg cells treated with pramipexole at concentrations of 2 or 200 ng/mL. These regulatory cells were co?cultured with in vitro?differentiated human dopaminergic neurons in a cytotoxicity assay with 6?hydroxydopamine (6?OHDA). The role of interleukin?10 (IL?10) was investigated by co?culturing activated IL?10?producing Treg cells with neurons. To further investigate the effect of treatment on Tregs,gene expression in pramipexole?treated,CD3/CD28?activated Treg cells was determined by Fluidigm analysis.ResultsPramipexole?treated CD3/CD28?activated Treg cells showed significant protective effects on dopaminergic neurons when challenged with 6?OHDA. Pramipexole?treated activated Treg cells showed neuroprotective capacity through mechanisms involving IL?10 release and the activation of genes associated with regulation and neuroprotection.ConclusionAnti?CD3/CD28?activated Treg cells protect dopaminergic neurons against 6?OHDA?induced damage. In addition,activated,IL?10?producing,pramipexole?treated Tregs also induced a neuroprotective effect,and the supernatants of these co?cultures promoted axonal growth. Pramipexole?treated,activated Tregs altered their gene expression in a concentration?dependent manner,and enhanced TGF??related dopamine receptor regulation and immune?related pathways. These findings open new perspectives for the development of immunomodulatory therapies for the treatment of PD. Pramipexole?treated,activated regulatory T cells protect dopaminergic neurons against 6 OHDA damage and promote primary neurite length. This could be due to the production of the regulatory cytokine IL?10 and an increased expression of genes related to regulation and neuroprotection. View Publication

T cell dysfunction is a pivotal driving factor in autoimmune diseases,yet its underlying regulatory mechanisms remain incompletely understood. The role of long non-coding RNAs (lncRNAs) in immune regulation has gradually been recognized,although their functional mechanisms in T cells remain elusive. This study focuses on lncBADR (LncRNA Branched-chain Amino acids Degradation Regulator),elucidating its mechanism by which it regulates branched-chain amino acids (BCAAs) metabolism to influence T cell effector functions. Mice with specific knockout of lncBADR (T celllncBADR−/−) exhibited markedly ameliorated experimental autoimmune encephalomyelitis (EAE) symptoms. Mechanistic investigations revealed that lncBADR inhibits BCAAs degradation by binding to the enzymes Mccc1 and Pcca,leading to the accumulation of BCAAs within T-cells. This,in turn,activates the mTOR-Stat1 signaling pathway,promoting IFN-γ secretion and exacerbating EAE pathology. In contrast,knockout of lncBADR restored BCAAs degradation,significantly reducing IFN-γ secretion in T cells and suppressing their pathogenic functions. Further studies demonstrated that high-BCAAs feeding partially reversed the protective effects of lncBADR knockout,indicating that lncBADR plays a crucial role in autoimmune inflammation by regulating BCAAs metabolism. This study offers new insights into targeting lncBADR or modulating BCAAs metabolism as potential therapeutic strategies for autoimmune diseases. View Publication

Plasminogen activator inhibitor-1 (PAI-1) has a pro-tumorigenic function via its pro-angiogenic and anti-apoptotic activities. Here,we demonstrate that PAI-1 promotes the recruitment and M2 polarization of monocytes/macrophages through different structural domains. Its LRP1 interacting domain regulated macrophage migration,while its C-terminal uPA interacting domain promoted M2 macrophage polarization through activation of p38MAPK and nuclear factor $\kappa$B (NF-$\kappa$B) and induction of an autocrine interleukin (IL)-6/STAT3 activation pathway. We then show in several experiments in mice that expression of PAI-1 is associated with increased tumorigenicity,increased presence of M2 macrophages,higher levels of IL-6,and increased STAT3 phosphorylation in macrophages. Strong positive correlations between PAI-1,IL-6,and CD163 (M2 marker) expression were also found by meta-analysis of transcriptome data in many human cancers. Altogether,these data provide evidence for a mechanism explaining the paradoxical pro-tumorigenic function of PAI-1 in cancer. View Publication

Despite the growing knowledge of T cell responses in COVID-19 patients,there is a lack of detailed characterizations for T cell-antigen interactions and T cell functions. Here,with a predicted peptide library from SARS-CoV-2 S and N proteins,we found that specific CD8+ T cell responses were identified in over 75% of COVID-19 convalescent patients (15/20) and an epitope from the N protein,N361-369 (KTFPPTEPK),was the most dominant epitope from our selected peptide library. Importantly,we discovered 2 N361-369-specific T cell receptors (TCRs) with high functional avidity that were independent of the CD8 co-receptor. These TCRs exhibited complementary cross-reactivity to several presently reported N361-369 mutant variants,as to the wild-type epitope. Further,the natural functions of these TCRs in the cytotoxic immunity against SARS-CoV-2 were determined with dendritic cells (DCs) and the lung organoid model. We found that the N361-369 epitope could be normally processed and endogenously presented by these different types of antigen presenting cells,to elicit successful activation and effective cytotoxicity of CD8+ T cells ex vivo. Our study evidenced potential mechanisms of cellular immunity to SARS-CoV-2,and illuminated potential ways of viral clearance in COVID-19 patients. These results indicate that utilizing CD8-independent TCRs against SARS-CoV-2-associated antigens may provide functional superiority that is beneficial for the adoptive cell immunotherapies based on natural or genetically engineered T cells. Additionally,this information is highly relevant for the development of the next-generation vaccines with protections against continuously emerged SARS-CoV-2 mutant strains. View Publication

To date,studies on T cells in acute myeloid leukemia (AML) have been limited to flow cytometric analysis of whole peripheral blood mononuclear cell (PBMC) specimens or functional work looking at the impact of AML myeloblasts on normal or remission T cells. This lack of information on T cells at the time of presentation with disease is due in part to the difficulty in isolating sufficiently pure T cells from these specimens for further study. Negative immunomagnetic selection has been the method of choice for isolating immune cells for functional studies due to concerns that binding antibodies to the cell surface may induce cellular activation,block ligand-receptor interactions or result in immune clearance. In order specifically to study T cells in presentation AML specimens,we set out to develop a method of isolating highly pure CD4 and CD8 T cells by negative selection from the peripheral blood (PB) of newly diagnosed AML patients. This technique,unlike T cell selection from PB from normal individuals or from patients with chronic lymphocytic leukaemia,was extremely problematic due to properties of the leukaemic myeloblasts. A successful method was eventually optimized requiring the use of a custom antibody cocktail consisting of CD33,CD34,CD123,CD11c and CD36,to deplete myeloblasts. View Publication

Interleukin 17 (IL-17)-producing CD4(+) helper T cells (T(H)-17 cells) share a developmental relationship with Foxp3(+) regulatory T cells (T(reg) cells). Here we show that a T(H)-17 population differentiates in the thymus in a manner influenced by recognition of self antigen and by the cytokines IL-6 and transforming growth factor-beta (TGF-beta). Like previously described T(H)-17 cells,the T(H)-17 cells that developed in the thymus expressed the transcription factor RORgamma t and the IL-23 receptor. These cells also expressed alpha(4)beta(1) integrins and the chemokine receptor CCR6 and were recruited to the lung,gut and liver. In the liver,these cells secreted IL-22 in response to self antigen and mediated host protection during inflammation. Thus,T(H)-17 cells,like T(reg) cells,can be selected by self antigens in the thymus. View Publication

Regulation of cytokine production in stimulated T cells can be disrupted in autoimmunity,immunodeficiencies,and cancer. Systematic discovery of stimulation-dependent cytokine regulators requires both loss-of-function and gain-of-function studies,which have been challenging in primary human cells. We now report genome-wide CRISPR activation (CRISPRa) and interference (CRISPRi) screens in primary human T cells to identify gene networks controlling interleukin-2 (IL-2) and interferon-$\gamma$ (IFN-$\gamma$) production. Arrayed CRISPRa confirmed key hits and enabled multiplexed secretome characterization,revealing reshaped cytokine responses. Coupling CRISPRa screening with single-cell RNA sequencing enabled deep molecular characterization of screen hits,revealing how perturbations tuned T cell activation and promoted cell states characterized by distinct cytokine expression profiles. These screens reveal genes that reprogram critical immune cell functions,which could inform the design of immunotherapies. View Publication

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 . View Publication

Severe lupus often includes psychiatric and neurological sequelae,although the cellular contributors to CNS disease remain poorly defined. Using intravascular staining to discriminate tissue-localized from blood-borne cells,we find substantial accumulation of CD8+ T cells relative to other lymphocytes in brain tissue,which correlates with lupus disease and limited neuropathology. This is in contrast to all other affected organs,where infiltrating CD4+ cells are predominant. Brain-infiltrating CD8+ T cells represent an activated subset of those found in the periphery,having a resident-memory phenotype (CD69+CD122-PD1+CD44+CD62L-) and expressing adhesion molecules (VLA-4+LFA-1+) complementary to activated brain endothelium. Remarkably,infiltrating CD8+ T cells do not cause tissue damage in lupus-prone mice,as genetic ablation of these cells via $\beta$2 m deficiency does not reverse neuropathology,but exacerbates disease both in the brain and globally despite decreased serum IgG levels. Thus,lupus-associated inflammation disrupts the blood-brain barrier in a discriminating way biased in favor of non-pathogenic CD8+ T cells relative to other infiltrating leukocytes,perhaps preventing further tissue damage in such a sensitive organ. View Publication

Regulatory T cell (Treg) therapy has the potential to induce transplantation tolerance so that immunosuppression and associated morbidity can be minimized. Alloantigen-reactive Tregs (arTregs) are more effective at preventing graft rejection than polyclonally expanded Tregs (PolyTregs) in murine models. We have developed a manufacturing process to expand human arTregs in short-term cultures using good manufacturing practice-compliant reagents. This process uses CD40L-activated allogeneic B cells to selectively expand arTregs followed by polyclonal restimulation to increase yield. Tregs expanded 100- to 1600-fold were highly alloantigen reactive and expressed the phenotype of stable Tregs. The alloantigen-expanded Tregs had a diverse TCR repertoire. They were more potent than PolyTregs in vitro and more effective at controlling allograft injuries in vivo in a humanized mouse model. View Publication

How HIV controllers (HICs) maintain undetectable viremia without therapy is unknown. The strong CD8(+) T-cell HIV suppressive capacity found in many,but not all,HICs may contribute to long-lasting viral control. However,other earlier defense mechanisms may be involved. Here,we examined intrinsic HIC cell resistance to HIV-1 infection. After in vitro challenge,monocyte-derived macrophages and anti-CD3-activated CD4(+) T cells from HICs showed low HIV-1 susceptibility. CD4 T-cell resistance was independent of HIV-1 coreceptors and affected also SIVmac infection. CD4(+) T cells from HICs expressed ex vivo higher levels of p21(Waf1/Cip1),which has been involved in the control of HIV-1 replication,than cells from control subjects. However,HIV restriction in anti-CD3-activated CD4(+) T cells and macrophages was not associated with p21 expression. Restriction inhibited accumulation of reverse transcripts,leading to reduction of HIV-1 integrated proviruses. The block could be overcome by high viral inocula,suggesting the action of a saturable mechanism. Importantly,cell-associated HIV-1 DNA load was extremely low in HICs and correlated with CD4(+) T-cell permissiveness to infection. These results point to a contribution of intrinsic cell resistance to the control of infection and the containment of viral reservoir in HICs. View Publication

Developing efficacious vaccines against enteric diseases is a global challenge that requires a better understanding of cellular recruitment dynamics at the mucosal surfaces. The current paradigm of T cell homing to the gastrointestinal (GI) tract involves the induction of $$4$$7 and CCR9 by Peyer's patch and mesenteric lymph node (MLN) dendritic cells (DCs) in a retinoic acid-dependent manner. This paradigm,however,cannot be reconciled with reports of GI T cell responses after intranasal (i.n.) delivery of antigens that do not directly target the GI lymphoid tissue. To explore alternative pathways of cellular migration,we have investigated the ability of DCs from mucosal and nonmucosal tissues to recruit lymphocytes to the GI tract. Unexpectedly,we found that lung DCs,like CD103(+) MLN DCs,up-regulate the gut-homing integrin $$4$$7 in vitro and in vivo,and induce T cell migration to the GI tract in vivo. Consistent with a role for this pathway in generating mucosal immune responses,lung DC targeting by i.n. immunization induced protective immunity against enteric challenge with a highly pathogenic strain of Salmonella. The present report demonstrates novel functional evidence of mucosal cross talk mediated by DCs,which has the potential to inform the design of novel vaccines against mucosal pathogens. View Publication