Project description:Single cell RNA sequencing of the mouse colonic mesenchyme. Mesenchymal/lamina propria cells were isolated from the middle and distal colon of wild type mice and were pooled. The vast majority of intestinal epithelial cells were depleted by Ethylenediaminetetraacetic acid (EDTA) treatment of the tissue samples and mesenchymal/lamina propria cells were isolated after enzymatic treatment with collagenase XI and dispase. Single cell RNA sequencing was performed with the Drop-seq protocol.
Project description:Single cell RNA sequencing of the mouse colonic mesenchyme. Mesenchymal/lamina propria cells were isolated from the middle and distal colon of wild type mice in two biological replicates. For each biological replicate the colons of n = 2 mice were pooled. The vast majority of intestinal epithelial cells were depleted by Ethylenediaminetetraacetic acid (EDTA) treatment of the tissue samples and mesenchymal/lamina propria cells were isolated after enzymatic treatment with collagenase XI and dispase. Single cell RNA sequencing was performed with the Drop-seq protocol. N =5 Drop-seq collections (samples) were processed in total, two from the first biological replicate and three from the second.
Project description:10x Chromium single cell RNA-Seq of colonic mesenchyme cell populations in health and Ulcerative Colitis in human patients and health in DSS-induced colitis in murine colon
Project description:Our study reports a role for tumor necrosis factor receptor 1 (TNFR1) in maintaining colonic mesenchymal cell diversity. Through scRNA-seq profiling, we show that a TNFR1-deficient mesenchyme has a reduced transcriptional diversity of mesenchymal cell populations, with a near complete loss of a cluster enriched in TNF, IFN, and cytokine signaling genes, which include MMP9, CD200, TNFRSF9, and TRAF1. We show that the transcriptional clustering of mesenchymal cells is consistently distinct and unique to the TNFR1-/- genotype. TNFR1-/- mesenchymal cells cluster as a single population, compared to control mesenchymal cells that cluster into 3 sub-populations. Through in vivo immunostaining in control and TNFR1-/- mice we show that the TNF signaling cluster localizes to the sub-epithelial niche and therefore may play an important role in the crosstalk between epithelial stem cells and mesenchyme-driven immune signaling in health and disease.
Project description:A cell suspension was prepared from wild-type P14 mouse retinas, and single-cell mRNAseq libraries were generated with Drop-Seq. Drop-Seq was performed on four separate days using the same age (P14) and strain (C57BL/6). On day 1, replicate 1 was obtained. On day 2, replicates 2 and 3 were obtained. On day 3, replicates 4-6 were obtained. On day 4, replicate 7 was obtained.
Project description:Our study reports a role for mesenchymal TNFR1 in maintaining colonic mesenchymal cell diversity and facilitation of a function epithelial stem cell niche. Through RNA-seq profiling, we show that a TNFR1-deficient mesenchyme has altered anchoring and cell substrate junctions, focal adhesions, extracellular matrix, and actin binding processes. We find specific reduction in the key stem cell niche factor, RSPO3, crucial for Lgr5+ stem cell maintenance, downregulated 5.1-fold in the TNFR1 deficient mesenchyme. Pathways classically associated with proliferation and differentiation, including Phosphoinositide 3-kinase (P13k) and Mitogen-activated protein kinase (MAPK), and those involved in migration, namely Ras-proximate-1 or Ras-related protein 1 (RAP1), TGF-β and RAS, were significantly enriched in TNFR1-/- mesenchyme compared to controls. In particular, Itga6, representing the integrin alpha 6 subunit, was one of the most highly upregulated transcripts. Therefore, the transcriptome profiling of the TNFR1 deficient mesenchyme deletion suggests an important role for TNFR1 in the regulation of mesenchymal cell-matrix interactions and niche transcript expression.
Project description:Cell suspensions were prepared from mouse lungs dissociated at baseline or 14 days after intratracheal bleomycin injury, and single cell mRNA-seq libraries were generated with Drop-seq.