CD4+ T-cell-induced inflammatory killing controls immune evasive tumors
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ABSTRACT:
PROVIDER: PRJNA961118 | ENA |
REPOSITORIES: ENA
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Project description:Most clinically applied cancer immunotherapies rely on the ability of CD8+ cytolytic T-cells to directly recognise and kill tumor cells. These strategies are limited by the emergence of MHC-deficient tumor cells and the development of an immunosuppressive tumor microenvironment. The ability of CD4+ effector cells to contribute to anti-tumor immunity independently of CD8+ T-cells is increasingly recognised, but strategies to unleash their full potential remain to be identified. Here, we describe a mechanism through which a small number of CD4+ T-cells is sufficient to eradicate MHC-deficient tumors that escape direct CD8+ T-cell targeting. The CD4+ effector T-cells preferentially cluster at tumor invasive margins where they interact with MHC-II+CD11c+ antigen-presenting cells. We show that Th1-directed CD4+ T-cells and innate immune stimulation reprogram the tumor-associated myeloid cell network towards IFN-activated antigen-presenting and iNOS-expressing tumoricidal effector phenotypes. Together, CD4+ T-cells and tumoricidal myeloid cells orchestrate a remote inflammatory cell death process that indirectly eradicates IFN-unresponsive and MHC-deficient tumors. These results warrant to clinically exploit the ability of CD4+ T-cells and innate immune stimulators as a strategy to complement the direct cytolytic activity of CD8+ T- and NK-cells and advance cancer immunotherapies.
2023-05-24 | GSE230427 | GEO
Project description:Mycobacterium smegmatis is a model non-pathogenic mycobacterium that is efficiently killed by macrophages. Here, we explore the role of NF-?B in the innate immune response, focusing in detail on the mechanisms of the first killing period (1-4h) of M. smegmatis which coincides with phagosome-lysosome fusion. We show that infection of macrophages with M. smegmatis induces an activation of NF-?B and this activation is required for killing since treatment of macrophages with NF-?B inhibitors or siRNA silencing of the NF-?B subunit p65 increases bacterial survival. NF-?B induced proteins were thus hypothesized to be essential during the first phase of M. smegmatis killing. We therefore identified, using RNA microarray, the genes that were regulated during infection in the absence and presence of NF-?B inhibitors. By subtraction this provided a list of pro-inflammatory proteins that were under the control of NF-?B and putatively involved in the killing response. Among these category of genes were those for lysosomal enzymes and membrane trafficking regulators, including Cathepsins, LAMP-2 and Rab34, are regulated by NF-?B. Moreover, inhibition of NF-?B signaling retarded the delivery of v-ATPase, LAMP-2, CtsZ and CtsH thereby impairing the maturation of mycobacterial phagosomes. Collectively; our data provide the first compelling evidence that the innate immune response via NF-?B activation is linked to phagosome fusion with lysosomes that is essential for killing of mycobacteria. Keywords: NFkB inhibitor treated vs untreated The study includes J774 macrophage cell lines which are infected with Mycobacterium smegmatis in the presence and absense of NFkB inhibitor SC-514.Total RNA was isolated from individual samples and each sample was hybridised to CodeLink Mouse whole genome bioarray slides.This study was intended to know the role of NFkB regulated genes in killing non-pathogenic mycobacteria during 4 hour post infection of macrophages.
2007-10-23 | E-GEOD-8999 | biostudies-arrayexpress
Project description:Mouse macrophages J774A.1 were pre-treated with the anti-inflammatory lipid eicosapenaenoic acid (EPA) or the pro-inflammatory lipid ceramide (Cer) for 3h. Macrophages were then infected with Mycobacterium smegmatis for 1h, and total RNA was collected. In a parallel experiment, infected macrophages were infected for 1h, 4h and 24h. At these time points, macrophages were lysed, bacteria was collected and quantified by the Colony Forming Units (CFU) Assay. This provided the kinetics of the killing of Mycobacteria inside mouse macrophages. CFU experiments revealed that cells pre-treated with EPA showed an increased number of bacteria inside macrophages, in contrast to cells pre-treated with Cer. To dissect the molecular mechanisms involved in the survival and killing of mycobacteria infected macrophages, mediated by lipids, gene expression studies were performed. Cultures of Mycobacterium smegmatis mc2155 harbouring a p19-(long lived) EGFP plasmid were grown to exponential growth phase. Bacteria were pelleted, washed in PBS and resuspended in medium DMEM with a multiplicity of infection (MOI) of 10 (10 bacteria per macrophage). Clumps of bacteria were removed by ultrasonic treatment of bacterial suspensions in an ultrasonic water bath for 15 minutes, followed by a low speed centrifugation for 2 minutes. Mouse macrophages J774A.1 were pre-.treated with the anti-inflammatory lipid eicosapenaenoic acid (EPA) or the pro-inflammatory lipid ceramide (Cer), 3h before infection. Mouse macrophages J774A.1 were infected with Mycobacterium smegmatis mc2155 for 1h, at 37M-BM-:C and 5% CO2. After 1h of infection, cells were washed with PBS. After washing, 1 ml Trizol was added per well, to collect total RNA. The experimental condition were: samples 1.1, 1.2, 1.3: Untreated macrophages; samples 2.1, 2.2, 2.3: Mock treated macrophages (ethanol 1ul/ml); samples 3.1, 3.2, 3.3: EPA (15uM) treated macrophages (ethanol 1ul/ml); samples 4.1, 4.2, 4.3: Cer (5ug/ml) treated macrophages (ethanol 1ul/ml); samples 5.1, 5.2, 5.3: Untreated macrophages, infected with Mycobacterium smegmatis mc2155 for 1h; samples 6.1, 6.2, 6.3: Mock treated macrophages (ethanol 1ul/ml) infected with Mycobacterium smegmatis mc2155 for 1h; samples 7.1, 7.2, 7.3: EPA (15uM) treated macrophages (ethanol 1ul/ml) infected with Mycobacterium smegmatis mc2155 for 1h; samples 8.1, 8.2, 8.3: Cer (5ug/ml) treated macrophages (ethanol 1ul/ml) infected with Mycobacterium smegmatis mc2155 for 1h; In a parallel experiment, infected macrophages remained with the bacteria for 1h, 4h and 24h after infection. After these time points, the macrophages were lysed, the bacteria was collected and quantified by the Colony Forming Units (CFU) Assay. This provided the kinetics of the killing of non-pathogenic intracellular bacteria inside mouse macrophages. For the CFU assay, after 1h of infection, cells were washed with PBS and Gentamicin (10ug/ml) in DMEM to kill the extracellular bacteria. At discrete time points, cells were washed with PBS and lysed with sterilized water. Quantitative cultures of bacteria were performed in a 10-fold serial dilutions, inoculated on 7H10 agar plates. 5ul were plated in triplicate and the number of colonies were counted after 48h.
2015-01-20 | E-GEOD-27277 | biostudies-arrayexpress
Project description:Mouse macrophages J774A.1 were pre-treated with the anti-inflammatory lipid eicosapenaenoic acid (EPA) or the pro-inflammatory lipid ceramide (Cer) for 3h. Macrophages were then infected with Mycobacterium smegmatis for 1h, and total RNA was collected. In a parallel experiment, infected macrophages were infected for 1h, 4h and 24h. At these time points, macrophages were lysed, bacteria was collected and quantified by the Colony Forming Units (CFU) Assay. This provided the kinetics of the killing of Mycobacteria inside mouse macrophages. CFU experiments revealed that cells pre-treated with EPA showed an increased number of bacteria inside macrophages, in contrast to cells pre-treated with Cer. To dissect the molecular mechanisms involved in the survival and killing of mycobacteria infected macrophages, mediated by lipids, gene expression studies were performed.
2015-01-20 | GSE27277 | GEO
Project description:Mycobacterium smegmatis is a model non-pathogenic mycobacterium that is efficiently killed by macrophages. Here, we explore the role of NF-κB in the innate immune response, focusing in detail on the mechanisms of the first killing period (1-4h) of M. smegmatis which coincides with phagosome-lysosome fusion. We show that infection of macrophages with M. smegmatis induces an activation of NF-κB and this activation is required for killing since treatment of macrophages with NF-κB inhibitors or siRNA silencing of the NF-κB subunit p65 increases bacterial survival. NF-κB induced proteins were thus hypothesized to be essential during the first phase of M. smegmatis killing. We therefore identified, using RNA microarray, the genes that were regulated during infection in the absence and presence of NF-κB inhibitors. By subtraction this provided a list of pro-inflammatory proteins that were under the control of NF-κB and putatively involved in the killing response. Among these category of genes were those for lysosomal enzymes and membrane trafficking regulators, including Cathepsins, LAMP-2 and Rab34, are regulated by NF-κB. Moreover, inhibition of NF-κB signaling retarded the delivery of v-ATPase, LAMP-2, CtsZ and CtsH thereby impairing the maturation of mycobacterial phagosomes. Collectively; our data provide the first compelling evidence that the innate immune response via NF-κB activation is linked to phagosome fusion with lysosomes that is essential for killing of mycobacteria. Keywords: NFkB inhibitor treated vs untreated
2007-10-23 | GSE8999 | GEO
Project description:T cells are important for the control of acute myeloid leukemia (AML), a common and often deadly malignancy. We observed that some AML patient samples are resistant to killing by human engineered cytotoxic CD4+ T cells. Single-cell RNA-seq of primary AML samples and CD4+ T cells before and after their interaction uncovered transcriptional programs that correlate with AML sensitivity or resistance to CD4+ T cell killing. Resistance-associated AML programs were enriched in AML patients with poor survival, and killing-resistant AML cells did not engage T cells in vitro. Killing-sensitive AML potently activated T cells before being killed, and upregulated ICAM1, a key component of the immune synapse with T cells. Without ICAM1, killing-sensitive AML became resistant to killing to primary ex vivo-isolated CD8+ T cells in vitro, and engineered CD4+ T cells in vitro and in vivo. Thus, ICAM1 on AML acts as an immune trigger, allowing T cell killing, and could affect AML patient survival in vivo.
2024-05-09 | GSE254282 | GEO
Project description:Functionally distinct CD4+ helper T (Th) cell subsets, such as Th1, Th2, Th17, and regulatory T cells (Treg), play a pivotal role in the host-defense against pathogen invasion and the pathogenesis of inflammatory disorders. In this project, DIA-MS-based proteome analysis was performed on naïve CD4+ T, Th0, Th1, Th2, Th17 and iTreg cells using Q Exactive HF-X (Thermo Fisher Scientific) to search for proteins that differ among the cell subsets.
2023-01-04 | PXD036065 | JPOST Repository
Project description:Upon antigen-specific T Cell Receptor (TCR) engagement, human CD4+ T cells proliferate and differentiate, a process associated with rapid transcriptional changes and metabolic reprogramming utilizing aerobic glycolysis together with maintenance of oxidative phosphorylation1,2. However, the role of glycolytic-reprogramming during T-cell activation remains largely unclear3,4,5. Here, we show that maintenance of cytosolic pyruvate production is an essential requirement for remodeling of the CD4+ T cell epigenome after TCR-engagement. Furthermore, we provide evidence that the local inflammatory environment sustains metabolic reprogramming of CD4+ T-cells and impacts histone modification in a pyruvate-dependent manner. Mechanistically, we demonstrate that rapid and sustained generation of cytosolic, but not mitochondrial, pyruvate is an essential step for acetyl-coA production and subsequent H3K27ac epigenome remodeling. TCR-activation was found to induce nuclear import of pyruvate dehydrogenase (PDH) and its association with both the p300 acetyltransferase and histone H3K27ac. Disrupting PDH nuclear import impacted expression of activation-induced genes. These results reveal a direct connection between CD4+ T cell metabolic reprogramming and transcriptional regulation, with the generation of cytosolic pyruvate being an essential step in T cell activation. These data support tight integration of metabolic enzymes and histone modifying enzymes, allowing metabolic reprogramming to fuel CD4+ T cell activation.
2023-10-27 | PXD018132 | Pride
Project description:The crosstalk between tumor and immune cells has important implications for cancer treatment, in particular for immunotherapies. Pancreatic ductal adenocarcinoma (PDAC) generally displays a chemoresistant phenotype and is refractory to immune-based therapies. Highly tumorigenic and stem-like cells known as cancer stem cells (CSCs) within PDAC tumors have been demonstrated to contribute to its aggressiveness, metastatic capacity and chemoresistance. Here, we show that CSCs could be also relevant to establish an immunosuppresive environment. Using genetically engineered mouse models (GEMMs) of PDAC we isolated a triple positive (EpCAM+, Sca-1+ and CD133+) population of tumor cells, functionally validated these cells as bona fide CSCs and probed their transcriptional profile by microarray analysis. In addition to stem pathways, signatures related to tumor immune evasion, immune modulation, invasiveness, and innate immune response were enriched in this population. Peptidoglycan recognition protein 1 (Pglyrp1), an innate immune response anti-bacterial protein expressed primarily by neutrophils, was highly expressed in this triple-positive population and in CSC-enriched spheres from human cell lines. Interestingly, modulation of Pglyrp1 expression in murine PDAC primary cultures affected their immune evasive properties, specifically their ability to escape macrophage- and T-cell-mediated killing in vitro and to establish tumors in immunocompetent hosts. Pglyrp1 expression in tumor cells seemed to interfere with TNFα-mediated killing by T cells and to promote activated T-cell cytotoxicity. Importantly, these results could be validated in CSCs from human patient-derived xenografts, and PGLYRP1 protein levels were also significantly higher in PDAC patient samples versus healthy controls. Thus, our study demonstrates a hierarchical organization of cancer cells in a murine model of PDAC, validating the CSC concept, and identifies Pglyrp1 as a putative PDAC biomarker that confers immune privilege properties to CSCs which could have important therapeutic implications.
2024-09-30 | GSE222985 | GEO
Project description:The crosstalk between tumor and immune cells has important implications for cancer treatment, in particular for immunotherapies. Pancreatic ductal adenocarcinoma (PDAC) generally displays a chemoresistant phenotype and is refractory to immune-based therapies. Highly tumorigenic and stem-like cells known as cancer stem cells (CSCs) within PDAC tumors have been demonstrated to contribute to its aggressiveness, metastatic capacity and chemoresistance. Here, we show that CSCs could be also relevant to establish an immunosuppresive environment. Using genetically engineered mouse models (GEMMs) of PDAC we isolated a triple positive (EpCAM+, Sca-1+ and CD133+) population of tumor cells, functionally validated these cells as bona fide CSCs and probed their transcriptional profile by microarray analysis. In addition to stem pathways, signatures related to tumor immune evasion, immune modulation, invasiveness, and innate immune response were enriched in this population. Peptidoglycan recognition protein 1 (Pglyrp1), an innate immune response anti-bacterial protein expressed primarily by neutrophils, was highly expressed in this triple-positive population and in CSC-enriched spheres from human cell lines. Interestingly, modulation of Pglyrp1 expression in murine PDAC primary cultures affected their immune evasive properties, specifically their ability to escape macrophage- and T-cell-mediated killing in vitro and to establish tumors in immunocompetent hosts. Pglyrp1 expression in tumor cells seemed to interfere with TNFα-mediated killing by T cells and to promote activated T-cell cytotoxicity. Importantly, these results could be validated in CSCs from human patient-derived xenografts, and PGLYRP1 protein levels were also significantly higher in PDAC patient samples versus healthy controls. Thus, our study demonstrates a hierarchical organization of cancer cells in a murine model of PDAC, validating the CSC concept, and identifies Pglyrp1 as a putative PDAC biomarker that confers immune privilege properties to CSCs which could have important therapeutic implications.
2024-09-30 | GSE222983 | GEO