Bulk RNA-seq of T cells from EMT6 tumors treated with aTGFb/aPD-L1
Ontology highlight
ABSTRACT: We performed bulk RNA sequencing on sorted T cells (7AAD- Calcein blue+ CD45+ THY1.1- TCRb+) in an orthotopic EMT6 tumor model 7 days after treatment initiation in four experimental groups: 1) Control 2) aPD-L1 3) aTGFb 4) aPD-L1 and aTGFb.
Project description:We performed 10x single cell RNA-seq on sorted populations: T cell (7AAD- Calcein blue+ CD45+ THY1.1- TCRb+), fibroblasts (7AAD- Calcein Blue+ CD45- THY1.1- CD31- PDPN+), myeloid cells (7AAD- Calcein Blue+ CD45+ THY1.1- CD11b+) and tumor cells (7AAD- Calcein Blue+ CD45- THY1.1+) from cells in an orthotopic EMT6 tumor model 7 days after treatment initiation in four experimental groups: 1) Control 2) aPD-L1 3) aTGFb 4) aPD-L1 and aTGFb.
Project description:To identify heterogeneity of stromal cells in human lymph nodes, we performed single-cell RNA-seq of stromal cells (CD45-, CD31- PDPN+) from 3 donors. Additionally, dendritic cells (DCs, CD45+, CD11C+) were isolated from the LNs of the same donors to predict potential interactions between these two types of cells. As potentially human identifiable data, he raw sequencing data files for this experiment will be deposited in the controlled-access EGA archive.
Project description:We performed 10x single cell RNA and VDJ sequencing on sorted T cells (7AAD- Calcein blue+ CD45+ THY1.1- TCRb+) in an orthotopic EMT6 tumor model 7 days after treatment initiation in four experimental groups: 1) Control 2) aPD-L1 3) aTGFb 4) aPD-L1 and aTGFb.
Project description:Lymphoma growth is facilitated by the well-balanced infrastructure of the microenvironment in lymph nodes (LN). LNs undergo rapid expansion during lymphoma progression, often accompanied by excessive blood vessel growth. Here we report that aggressive lymphoma cause severe high endothelial venule (HEV) regression which impairs lymphocyte recirculation. In this study, we used transferred (i.v.) Eµ-Myc tumor cells as a mouse model of aggressive lymphoma. Blood endothelial cells from tumor bearing and control LNs were isolated by FACS and processed for single-cell RNA sequencing. The results revealed a detrimental mechanisms causing HEV-dedifferentiation during lymphoma growth with potential implications on tumor-targeting immune surveillance and strategies of immune therapy in LNs
Project description:With the advent of cancer immunotherapy, intense investigation has been focused on tumor-infiltrating immune cells. With only a fraction of patients responding to these new therapies, a better understanding of all elements of the tumor microenvironment (TME) that may influence therapeutic outcome is needed. Stromal elements of the TME, chiefly fibroblasts, have emerged as potential contributors to tumor progression and most recently resistance to immunotherapy, but their precise composition and clinical relevance remain incompletely understood. Here we use single-cell transcriptomics to chart the fibroblastic landscape during pancreatic ductal adenocarcinoma (PDAC) progression in animal models, identifying two healthy tissue fibroblast subsets that co-evolve along individual trajectories into four subsets of carcinoma-associated fibroblasts (CAFs).
Project description:Bulk RNA-seq was performed on fibroblasts and mesothelial cells from 11 different healthy mouse tissues such as bone, epididymal, brown and inguinal fat pads, mesentery, omentum, pancreas, liver, lung, lymph node and spleen. This data was used to examine tissue specific gene expression programs of fibroblasts and mesothelial cells.
Project description:The tumor microenvironment (TME) is a complex mixture of tumor cells, immune cells, endothelial cells and fibroblastic stroma cells (FSC). While cancer-associated fibroblasts are generally seen as a tumor-promoting entity, it is conceivable that distinct FSC populations within the TME contribute to immune-mediated tumor control. Here, we show that intra-tumoral injection of a recombinant LCMV-based vaccine vector (r3LCMV) expressing the melanocyte differentiation antigen TRP2 results in T cell-dependent eradication of melanomas. Analysis of the TME revealed that viral vector transduction precipitates activation of particular FSC subsets. Using single-cell RNA-seq analysis, we identified a Cxcl13-expressing FSC population with a pronounced immune-stimulatory signature and increased expression of the inflammatory cytokine IL-33. Genetic ablation of Il33 in Cxcl13-Cre+ FSC impeded functionality of intratumoral T cells and consequently tumor control. Thus, reprogramming of distinct FSC subsets in the TME through LCMV-based vectors efficiently promotes tumor eradication by locally sustaining the activity of tumor-specific T cells.
Project description:Alterations in global mRNA decay can broadly impact multiple upstream and downstream stages of gene expression. For example, accelerated cytoplasmic mRNA degradation can trigger a reduction in mammalian RNA polymerase II (RNAPII) transcription, although signals that connect these seemingly distal processes remain largely unknown. Here, we used tandem mass tag labeling with mass spectrometry to chart how changes in Xrn1-dependent mRNA degradation impact nuclear-cytoplasmic protein distribution in human cells. Notably, accelerating mRNA decay through expression of a gammaherpesviral endonuclease known to coordinate with Xrn1 drove nuclear relocalization of many RNA binding proteins. Particularly enriched in the relocalized subset were factors linked to the poly(A) tail. Conversely, cells lacking Xrn1 exhibited changes in the localization and/or abundance of numerous factors linked to mRNA turnover. Based on these data, we uncovered a new role for cytoplasmic poly(A) binding protein in repressing RNAPII transcription upon its mRNA decay-induced translocation to the nucleus.
Project description:Myocarditis is a heart condition that causes inflammation and results in the loss of heart muscle cells, often leading to fibrosis (scarring) of the heart tissue and heart failure. However, the molecular mechanisms underlying immune cell control and maintenance of tissue integrity in the inflamed cardiac microenvironment remain elusive. Based on our finding that bone morphogenic protein-4 (BMP4) serum concentration was reduced in myocarditis patients in combination with comprehensive single cell and single nucleus RNA sequencing analyses of inflamed murine and human myocardial tissue indicated that BMP4 gradients maintain cardiac tissue homeostasis. Indeed, restoration of BMP signaling through antibody-mediated neutralization of the BMP-inhibitors GREM1 and GREM2 reduced CD4+ T cell-mediated myocardial inflammation and blocked disease progression by reduction of adverse fibrotic remodelling. These results unveil a key function of the BMP4-GREM1/2 axis as promising approach for treating myocardial inflammation and the serious complications of cardiac fibrosis and heart failure.
Project description:By generating a paired single cell RNA-sequencing database of the tumor niche from 10 newly diagnosed MM patients, we created a unique dataset allowing the in-depth analyses of stromal-immune interactions within the tumor microenvironment (see related accession number). Using this database, we identified the presence of inflammatory stromal fibroblasts in the bone marrow of Myeloma patients.The stromal inflammation was associated with NF-κB signaling, and sources of IL-1β or TNFα were specific immune subsets previously shown to be altered in MM, suggesting the presence of an immune cell-mediated feed-forward loop of bone marrow inflammation in MM. By tracking inflammatory signatures over time in individual patients undergoing first-line treatment using bulk RNA sequencing, we show that bone marrow inflammation is not reverted by successful anti-tumor therapy (this dataset), suggesting a role for stromal fibroblasts and bone marrow inflammation in disease persistence or relapse. Raw sequencing data files will be deposited to EGA.