Project description:To demonstrate the efficacy of our developed Automated Live-imaging and cell Picking System for downstream RNA sequencing (ALPS&RNA-seq), we isolated single cells of three cell lines (murine T cell, murine leukemia cell, and murine hematopoietic progenitor cell) and of murine peripheral blood mononuclear cells (PBMCs). The detemined profiles of the three cell lines were different, and the profiles of the PBMSc showed three subpopulations which were determined as CD4+ T cells, CD8+ T cells, and B cells based on the marker genes.
Project description:Here we report bulk TCR sequencing data associated with open repertoire murine CD4+, CD4+CD8+, and CD8+ T cells isolated from B16 melanoma tumor resections
Project description:We report here that persistent activation of signal transducer and activator of transcription 5 (STAT5) in tumor-specific CD4+ T cells drives the development of polyfunctional T cells with superior antitumor activities. We showed that ectopic expression of a constitutively active form of murine STAT5A (CASTAT5) in tumor-specific CD4+ T cells established a distinct epigenetic and transcriptional landscape, endowing CD4+ T cells polyfunctionality, exhaustion-resistance and tumor-infiltrating capability. Single cell RNA sequencing analysis (scRNAseq) identified a subset of cells with the molecular signature indicative of their role as progenitor polyfunctional T cells. Importantly, T cells engineered to co-express CD19-targeting chimeric antigen receptors (CD19CAR) and CASTAT5 gave rise to polyfunctional CD4+ CAR T cells capable of providing optimal help to CD8+ T cells to achieve durable and curative outcomes in mice with advanced B-cell lymphoma. Evidence of CASTAT5 functional activities in primary human CD4+ T cells underscores its potential clinical relevance.
Project description:Conventional CD4 and CD8 single positive T cell lineages constitute the main differentiation pathway in the thymus. In human thymus, a minor TCRαβ differentiation pathway diverges from the early double positive stage, consisting of CD10+ PD-1+ cells. These cells are phenotypically and functionally similar to murine agonist-selected intraepithelial T lymphocyte precursors (IELps) which home to the small intestine. Here, the progeny of the human agonist-selected IEL lineage was identified in antigen-inexperienced cord blood (CB) with a polyclonal T cell receptor (TCR) repertoire exhibiting a bias towards early TCR alpha chain rearrangements and elevated autoreactive indices. Single-cell RNA sequencing allowed further delineation of this unconventional lineage in CB. Trajectory analysis, along with TCR repertoire analysis, transcriptomics and proteomics, suggests a precursor-progeny relationship with the thymic IELps. The distinct, heterogeneous CB population can now be defined as CD3+ TCRαβ+ CD4- CCR7- CD26-. Besides recent thymic emigrants, this population also consists of newly identified effector clusters and previously described populations: the suppressive NK receptor expressing CD8+ Treg population, the KIR/NKG2A+ EOMES+ virtual memory population and the CD8αα+ T cell populations. The population shows a discriminating stable HELIOS expression and is exclusively able to downregulate CD8β expression, resulting in double negative T cells. The functional properties of this population suggest that the cells expand on inflammatory cues and exert cytotoxic and proinflammatory activity.
Project description:Depletion of CD4+ cells by anti-CD4 monoclonal antibody (anti-CD4 mAb) induces expansion of tumor-reactive CD8+ T cells and exhibits strong antitumor effects in several murine tumor models. However, whether the anti-CD4 mAb treatment activates particular or a broad variety of tumor-reactive CD8+ T cell clones is not answered. To investigate the changes of TCR repertoire induced by the anti-CD4 mAb treatment, we performed unbiased high throughput TCR sequencing in a B16F10 mouse subcutaneous melanoma model.
