搜索结果: 'gmp t cell activator'

BackgroundAbnormalities in T cell activation play an important role in the pathogenesis of myocarditis,and persistent T cell responses can lead to autoimmunity and chronic cardiac inflammation,as well as even dilated cardiomyopathy. Although previous work has examined the role of T cells in myocarditis in animal models,the specific mechanism for human cardiomyocytes has not been investigated.MethodsIn this study,we constructed the human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) and established the T cell-mediated cardiac injury model by co-culturing with activated CD4 + T or CD8 + T cells that were isolated from peripheral mononuclear blood to elucidate the pathogenesis of myocardial cell injury caused by inflammation.ResultsBy combination of quantitative proteomics with tissue and cell immunofluorescence examination,we established a proteome profile of inflammatory myocardia from hiPSC-CMs with obvious cardiomyocyte injury and increased levels of lactate dehydrogenase content,creatine kinase isoenzyme MB and cardiac troponin. A series of molecular dysfunctions of hiPSC-CMs was observed and indicated that CD4 + cells could produce direct cardiomyocyte injury by activating the NOD-like receptor signals pathway.ConclusionsThe data presented in our study established a proteome map of inflammatory myocardial based on hiPSC-CMs injury model. These results can provide guidance in the discovery of improved clinical treatments for myocarditis.Supplementary InformationThe online version contains supplementary material available at 10.1186/s13287-024-03791-4. View Publication

Recent studies demonstrate that brain infiltration of peripheral immune cells and their interaction with brain-resident cells contribute to Parkinson’s disease (PD). However,mechanisms of T cell-brain cell communication are not fully elucidated and models allowing investigation of interaction between T cells and brain-resident cells are required. In this study,we developed a three-dimensional (3D) model composed of stem cell-derived human midbrain organoids (hMO) and peripheral blood T cells. We demonstrated that organoids consist of multiple midbrain-specific cell types,allowing to study T cell motility and interactions with midbrain tissue in a spatially organized microenvironment. We optimized co-culture conditions and demonstrated that T cells infiltrate hMO tissue,leading to neural cell loss. Our work establishes a novel 3D cell co-culture model as a promising tool to investigate the effect of the adaptive immune system on the midbrain and can be used in future studies to address these processes in the context of PD. View Publication

Checkpoint blockade therapies targeting PD-1/PD-L1 and CTLA-4 are clinically successful but also evoke adverse events due to systemic T-cell activation. We engineered a bispecific,mAb targeting CD28 homolog (CD28H),a newly identified B7 family receptor that is constitutively expressed on T and natural killer (NK) cells,with a PD-L1 antibody to potentiate tumor-specific immune responses. The bispecific antibody led to T-cell costimulation,induced NK-cell cytotoxicity of PD-L1-expressing tumor cells,and activated tissue-resident memory CD8+ T cells. Mechanistically,the CD28H agonistic arm of the bispecific antibody reduced PD-L1/PD-1-induced SHP2 phosphorylation while simultaneously augmenting T-cell receptor signaling by activating the MAPK and AKT pathways. This bispecific approach could be used to target multiple immune cells,including CD8+ T cells,tissue-resident memory T cells,and NK cells,in a tumor-specific manner that may lead to induction of durable,therapeutic antitumor responses. View Publication

Pulmonary metastasis of breast cancer requires recruitment and expansion of T-regulatory cells (Treg) that promote escape from host protective immune cells. However,it remains unclear precisely how tumors recruit Tregs to support metastatic growth. Here we report the mechanistic involvement of a unique and previously undescribed subset of regulatory B cells. These cells,designated tumor-evoked Bregs (tBreg),phenotypically resemble activated but poorly proliferative mature B2 cells (CD19(+) CD25(High) CD69(High)) that express constitutively active Stat3 and B7-H1(High) CD81(High) CD86(High) CD62L(Low) IgM(Int). Our studies with the mouse 4T1 model of breast cancer indicate that the primary role of tBregs in lung metastases is to induce TGF-$\beta$-dependent conversion of FoxP3(+) Tregs from resting CD4(+) T cells. In the absence of tBregs,4T1 tumors cannot metastasize into the lungs efficiently due to poor Treg conversion. Our findings have important clinical implications,as they suggest that tBregs must be controlled to interrupt the initiation of a key cancer-induced immunosuppressive event that is critical to support cancer metastasis. View Publication

Associations between chronic antigen stimulation,T cell dysfunction,and the expression of various inhibitory receptors are well characterized in several mouse and human systems. During chronic hepatitis B virus (HBV) infection (CHB),T cell responses are blunted with low frequencies of virus-specific T cells observed,making these parameters difficult to study. Here,using mass cytometry and a highly multiplexed combinatorial peptide-major histocompatibility complex (pMHC) tetramer strategy that allows for the detection of rare antigen-specific T cells,we simultaneously probed 484 unique HLA-A*1101-restricted epitopes spanning the entire HBV genome on T cells from patients at various stages of CHB. Numerous HBV-specific T cell populations were detected,validated,and profiled. T cells specific for two epitopes (HBVpol387 and HBVcore169) displayed differing and complex heterogeneities that were associated with the disease progression,and the expression of inhibitory receptors on these cells was not linearly related with their extent of T cell dysfunction. For HBVcore169-specific CD8+ T cells,we found cellular markers associated with long-term memory,polyfunctionality,and the presence of several previously unidentified public TCR clones that correlated with viral control. Using high-dimensional trajectory analysis of these cellular phenotypes,a pseudo-time metric was constructed that fit with the status of viral infection in corresponding patients. This was validated in a longitudinal cohort of patients undergoing antiviral therapy. Our study uncovers complex relationships of inhibitory receptors between the profiles of antigen-specific T cells and the status of CHB with implications for new strategies of therapeutic intervention. View Publication

BACKGROUND Radiotherapy enhances antitumor immunity. However,it also induces immunosuppressive responses,which are major hurdles for an effective treatment. Thus,targeting the immunosuppressive tumor microenvironment is essential for enhancing the antitumor immunity after radiotherapy. Retrospective studies show that a blockade of PI3K$\delta$ and/or $\gamma$,which are abundant in leukocytes,exhibits antitumor immune response by attenuating activity of immune suppressive cells,however,the single blockade of PI3K$\delta$ or $\gamma$ is not sufficient to completely eliminate solid tumor. METHODS We used BR101801,PI3K$\delta$/$\gamma$ inhibitor in the CT-26 syngeneic mouse model with a subcutaneously implanted tumor. BR101801 was administered daily,and the target tumor site was locally irradiated. We monitored the tumor growth regularly and evaluated the immunological changes using flow cytometry,ELISpot,and transcriptional analysis. RESULTS This study showed that BR101801 combined with irradiation promotes systemic antitumor immunity and abscopal response by attenuating the activity of immune suppressive cells in the CT-26 tumor model. BR101801 combined with irradiation systemically reduced the proliferation of regulatory T cells (Tregs) and enhanced the number of tumor-specific CD8$\alpha$+ T cells in the tumor microenvironment,thereby leading to tumor regression. Furthermore,the high ratio of CD8$\alpha$+ T cells to Tregs was maintained for 14 days after irradiation,resulting in remote tumor regression in metastatic lesions,the so-called abscopal effect. Moreover,our transcriptomic analysis showed that BR101801 combined with irradiation promoted the immune-stimulatory tumor microenvironment,suggesting that the combined therapy converts immunologically cold tumors into hot one. CONCLUSIONS Our data suggest the first evidence that PI3K$\delta$/$\gamma$ inhibition combined with irradiation promotes systemic antitumor immunity against solid tumors,providing the preclinical result of the potential use of PI3K$\delta$/$\gamma$ inhibitor as an immune-regulatory radiosensitizer. View Publication

Although obesity is known to be critical for cancer development,how obesity negatively impacts antitumor immune responses remains largely unknown. Here,we show that increased fatty acid oxidation (FAO) driven by activated STAT3 in CD8+ T effector cells is critical for obesity-associated breast tumor progression. Ablating T cell Stat3 or treatment with an FAO inhibitor in obese mice spontaneously developing breast tumor reduces FAO,increases glycolysis and CD8+ T effector cell functions,leading to inhibition of breast tumor development. Moreover,PD-1 ligation in CD8+ T cells activates STAT3 to increase FAO,inhibiting CD8+ T effector cell glycolysis and functions. Finally,leptin enriched in mammary adipocytes and fat tissues downregulates CD8+ T cell effector functions through activating STAT3-FAO and inhibiting glycolysis. We identify a critical role of increased oxidation of fatty acids driven by leptin and PD-1 through STAT3 in inhibiting CD8+ T effector cell glycolysis and in promoting obesity-associated breast tumorigenesis. View Publication

The adaptive immune response involves T cell differentiation and migration to sites of inflammation. T cell trafficking is initiated by rolling on inflamed endothelium. Tethers and slings,discovered in neutrophils,facilitate cell rolling at high shear stress. Here,we demonstrate that the ability to form tethers and slings during rolling is highly inducible in T helper 1 (Th1),Th17,and regulatory T (Treg) cells but less in Th2 cells. In vivo,endogenous Treg cells rolled stably in cremaster venules at physiological shear stress. Quantitative dynamic footprinting nanoscopy of Th1,Th17,and Treg cells uncovered the formation of multiple tethers per cell. Human Th1 cells also showed tethers and slings. RNA sequencing (RNA-seq) revealed the induction of cell migration and cytoskeletal genes in sling-forming cells. We conclude that differentiated CD4 T cells stabilize rolling by inducible tether and sling formation. These phenotypic changes approximate the adhesion phenotype of neutrophils and support CD4 T cell access to sites of inflammation. View Publication

A key event in the successful induction of adaptive immune responses is the antigen-specific activation of T cells by dendritic cells (DCs). Although LFA-1 (lymphocyte function-associated antigen 1) on T cells is considered to be important for antigen-specific T-cell activation,the role for LFA-1 on DCs remains elusive. Using 2 different approaches to activate LFA-1 on DCs,either by deletion of the αL-integrin cytoplasmic GFFKR sequence or by silencing cytohesin-1-interacting protein,we now provide evidence that DCs are able to make use of active LFA-1 and can thereby control the contact duration with naive T cells. Enhanced duration of DC/T-cell interaction correlates inversely with antigen-specific T-cell proliferation,generation of T-helper 1 cells,and immune responses leading to delayed-type hypersensitivity. We could revert normal interaction time and T-cell proliferation to wild-type levels by inhibition of active LFA-1 on DCs. Our data further suggest that cytohesin-1-interacting protein might be responsible for controlling LFA-1 deactivation on mature DCs. In summary,our findings indicate that LFA-1 on DCs needs to be in an inactive state to ensure optimal T-cell activation and suggest that regulation of LFA-1 activity allows DCs to actively control antigen-driven T-cell proliferation and effective immune responses. View Publication

BACKGROUND Hypersensitivity pneumonitis (HP) is an interstitial lung disease characterized by antigen-triggered neutrophilic exacerbations. Although CD4+ T cells are sufficient for HP pathogenesis,this never translated into efficient T cell-specific therapies. Increasing evidence shows that B cells also play decisive roles in HP. Here,we aimed to further define the respective contributions of B and T cells in subacute experimental HP. METHODS Mice were subjected to a protocol of subacute exposure to the archaeon Methanosphaera stadmanae to induce experimental HP. Using models of adoptive transfers of B cells and T cells in Rag1-deficient mice and of B cell-specific S1P1 deletion,we assessed the importance of B cells in the development of HP by evaluating inflammation in bronchoalveolar lavage fluid. We also aimed to determine if injected antibodies targeting B and/or T cells could alleviate HP exacerbations using a therapeutic course of intervention. RESULTS Even though B cells are not sufficient to induce HP,they strongly potentiate CD4+ T cell-induced HP?‘associated neutrophilic inflammation in the airways. However,the reduction of 85% of lung B cells in mice with a CD19-driven S1P1 deletion does not dampen HP inflammation,suggesting that lung B cells are not necessary in large numbers to sustain local inflammation. Finally,we found that injecting antibodies targeting B cells after experimental HP was induced does not dampen neutrophilic exacerbation. Yet,injection of antibodies directed against B cells and T cells yielded a potent 76% inhibition of neutrophilic accumulation in the lungs. This inhibition occurred despite partial,sometimes mild,depletion of B cells and T cells subsets. CONCLUSIONS Although B cells are required for maximal inflammation in subacute experimental HP,partial reduction of B cells fails to reduce HP-associated inflammation by itself. However,co-modulation of T cells and B cells yields enhanced inhibition of HP exacerbation caused by an antigenic rechallenge. View Publication

BACKGROUND Chimeric antigen receptor (CAR) T cell therapy targeting B cell maturation antigen (BCMA) on multiple myeloma (MM) produces fast but not long-lasting responses. Reasons for treatment failure are poorly understood. CARs simultaneously targeting two antigens may represent an alternative. Here,we (1) designed and characterized novel A proliferation inducing ligand (APRIL) based dual-antigen targeting CARs,and (2) investigated mechanisms of resistance to CAR T cells with three different BCMA-binding moieties (APRIL,single-chain-variable-fragment,heavy-chain-only). METHODS Three new APRIL-CARs were designed and characterized. Human APRIL-CAR T cells were evaluated for their cytotoxic function in vitro and in vivo,for their polyfunctionality,immune synapse formation,memory,exhaustion phenotype and tonic signaling activity. To investigate resistance mechanisms,we analyzed BCMA levels and cellular localization and quantified CAR T cell-target cell interactions by live microscopy. Impact on pathway activation and tumor cell proliferation was assessed in vitro and in vivo. RESULTS APRIL-CAR T cells in a trimeric ligand binding conformation conferred fast but not sustained antitumor responses in vivo in mouse xenograft models. In vitro trimer-BB$\zeta$ CAR T cells were more polyfunctional and formed stronger immune synapses than monomer-BB$\zeta$ CAR T cells. After CAR T cell-myeloma cell contact,BCMA was rapidly downmodulated on target cells with all evaluated binding moieties. CAR T cells acquired BCMA by trogocytosis,and BCMA on MM cells was rapidly internalized. Since BCMA can be re-expressed during progression and persisting CAR T cells may not protect patients from relapse,we investigated whether non-functional CAR T cells play a role in tumor progression. While CAR T cell-MM cell interactions activated BCMA pathway,we did not find enhanced tumor growth in vitro or in vivo. CONCLUSION Antitumor responses with APRIL-CAR T cells were fast but not sustained. Rapid BCMA downmodulation occurred independently of whether an APRIL or antibody-based binding moiety was used. BCMA internalization mostly contributed to this effect,but trogocytosis by CAR T cells was also observed. Our study sheds light on the mechanisms underlying CAR T cell failure in MM when targeting BCMA and can inform the development of improved treatment strategies. View Publication

OBJECTIVES We sought to develop medium throughput standard operating procedures for screening cryopreserved human peripheral blood mononuclear cells (PBMCs) for CD4+ and CD8+ T cell responses to potential autoantigens. METHODS Dendritic cells were loaded with a peptide cocktail from ubiquitous viruses or full-length viral protein antigens and cocultured with autologous T cells. We measured expression of surface activation markers on T cells by flow cytometry and cytometry by time of flight 24-72 h later. We tested responses among T cells freshly isolated from healthy control PBMCs,cryopreserved T cells,and T cells derived from a variety of T cell expansion protocols. We also compared the transcriptional profile of CD8+ T cells rested with interleukin (IL)7 for 48 h after 1) initial thawing,2) expansion,and 3) secondary cryopreservation/thawing of expanded cells. To generate competent antigen presenting cells from PBMCs,we promoted differentiation of PBMCs into dendritic cells with granulocyte macrophage colony stimulating factor and IL-4. RESULTS We observed robust dendritic cell differentiation from human PBMCs treated with 50 ng/mL GM-CSF and 20 ng/mL IL-4 in as little as 3 days. Dendritic cell purity was substantially increased by magnetically enriching for CD14+ monocytes prior to differentiation. We also measured antigen-dependent T cell activation in DC-T cell cocultures. However,polyclonal expansion of T cells with anti-CD3/antiCD28 abolished antigen-dependent upregulation of CD69 in our assay despite minimal transcriptional differences between rested CD8+ T cells before and after expansion. Furthermore,resting these expanded T cells in IL-2,IL-7 or IL-15 did not restore the antigen dependent responses. In contrast,T cells that were initially expanded with IL-2 + IL-7 rather than plate bound anti-CD3 + anti-CD28 retained responsiveness to antigen stimulation and these responses strongly correlated with responses measured at initial thawing. SIGNIFICANCE While screening techniques for potential pathological autoantibodies have come a long way,comparable full-length protein target assays for screening patient T cells at medium throughput are noticeably lacking due to technical hurdles. Here we advance techniques that should have broad applicability to translational studies investigating cell mediated immunity in infectious or autoimmune diseases. Future studies are aimed at investigating possible CD8+ T cell autoantigens in MS and other CNS autoimmune diseases. View Publication