MRNA-seq analysis of mouse cytotoxic T lymphocytes (CTLs) over-expressing Akt isoforms
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ABSTRACT: PI3K/Akt pathway plays a central role in T-cell activation and is impaired in Ag-specific CTLs during chronic viral infections. In this study, we genetically engineered HBV-specific CTLs and adoptively transferred these Akt-expressing viral Ag-specific CTLs into HBV-infected mice and monitored the virological and immunological responses. In the meantime, we also analyzed the mRNA profiles of control-CTLs and CTLs-overexpressing Akt1 and Akt2, respectively before adoptive transfer and after in vitro restimulation. We expect to elucidate the potential mechanisms about how the Akt signaling in CTLs prevents T-cell exhaustion and enhance T-cell expansion in the liver during persistent HBV infection from this mRNA-seq analysis.
Project description:Transcriptional profiling of mouse tumor-specific cytotoxic T lymphocytes (CTL) comparing activation-induced gene expression profiles induced by tumor-specific antigen (Ag) and CD3 mAb. CTLs were stimulated with either Ag or CD3 mAb for 3 and 24 h, respectively. The stimulated CTLs were compared to un-stimulated CTLs. The objective was to identify differential gene expression profiles induced by the two activation (Ag vs CD3 mAb) conditions. Two-condition experiments, un-stimulated vs stimulated CTLs. Biological replicates: 3 replicates for each comparison. Unstimulated CTLs were compared to stimulated CTLs (Ag and CD3 mAb, respectively).
Project description:We conducted immune- and RNA-sequencing of HLA-A24-restricted CMVpp65-specific CTLs to better understand the immune reconstitution of CMV-CTLs after allo-HCT. To the best of our knowledge, this is the first report on the features of TCRβ-CDR3, diversity, and GEP of HLA-A24 CMV-CTLs according to the CMV-reactivation pattern among recipients after allo-HCT. In addition, we further sought to demonstrate homogeneity or heterogeneity according to individual CTL clones using single-cell RNA-sequencing technology.
Project description:Transcriptional profiling of mouse tumor-specific cytotoxic T lymphocytes (CTL) comparing activation-induced gene expression profiles induced by tumor-specific antigen (Ag) and CD3 mAb. CTLs were stimulated with either Ag or CD3 mAb for 3 and 24 h, respectively. The stimulated CTLs were compared to un-stimulated CTLs. The objective was to identify differential gene expression profiles induced by the two activation (Ag vs CD3 mAb) conditions.
Project description:SARS-CoV-2 CTLs can be generated from convalescent donors by viral peptide stimulation and enrichment via the CliniMACS® Cytokine Capture System.
Project description:Celiac disease is an intestinal inflammatory disorder induced by dietary gluten in genetically susceptible individuals. The mechanisms underlying the massive expansion of interferon g–producing intraepithelial cytotoxic T lymphocytes (CTLs) and the destruction of the epithelial cells lining the small intestine of celiac patients have remained elusive. We report massive oligoclonal expansions of intraepithelial CTLs that exhibit a profound genetic reprogramming of natural killer (NK) functions. These CTLs aberrantly expressed cytolytic NK lineage receptors, such as NKG2C, NKp44, and NKp46, which associate with adaptor molecules bearing immunoreceptor tyrosine-based activation motifs and induce ZAP-70 phosphorylation, cytokine secretion, and proliferation independently of T cell receptor signaling. This NK transformation of CTLs may underlie both the self-perpetuating, gluten-independent tissue damage and the uncontrolled CTL expansion leading to malignant lymphomas in severe forms of celiac disease. Because similar changes were detected in a subset of CTLs from cytomegalovirus-seropositive patients, we suggest that a stepwise transformation of CTLs into NK-like cells may underlie immunopathology in various chronic infectious and inflammatory diseases. Keywords: NKG2C; LAK; CTL; NK receptor; IEL; Mucosal Immunity; Celiac Disease
Project description:Transcriptom-wide RNA expression profilng in human CAR-T cells treated with lenalidomide in the presense or absense of Ag-specific restimulation
Project description:Genome-wide DNA accessibility maps of chromatin state in human CAR-T cells treated with lenalidomide in the presense or absense of Ag-specific restimulation
Project description:Bach2 codes for a transcriptional regulator exerting major influences on T cell mediated immune regulation. Effector CTLs derived from in vitro activation of murine CD8+ T cells showed increased proliferative and cytolytic capacity in the absence of BACH2. Before activation, BACH2-deficient CD8+ T cells had a higher abundance of memory and reduced abundance of naïve cells compared to wild-type. CTLs derived from central memory T cells were more potently cytotoxic than those derived from naïve T cells, but even within separated subsets, BACH2-deficiency conferred a cytotoxic advantage. Immunofluorescence and electron microscopy revealed larger granules in BACH2-deficient compared to wild-type CTLs, and proteomic analysis showed an increase in granule content, including perforin and granzymes. Thus, the enhanced cytotoxicity observed in effector CTLs lacking BACH2 arises not only from differences in their initial differentiation state but also inherent production of enlarged cytolytic granules. These results demonstrate how a single gene deletion can produce a CTL super-killer.