Mus musculus
Ontology highlight
ABSTRACT:
PROVIDER: PRJNA198709 | ENA |
REPOSITORIES: ENA
Similar Datasets
Project description:The identification of Lgr5 as an intestinal stem cell marker has made it possible to isolate and study primary stem cells from small intestine and skins. Applying genome-wide resequencing of bisulfite-treated genomic DNA, we profiled the DNA methylation changes of FACS-sorted Lgr5+ve stem cells and their immediate undifferentiated daughter cells from small intestine and skin. In addition to this, we also analyzed terminally differentiated villus cells. We find that terminal differentiation of an adult solid-tissue stem cell does not require DNA methylation dynamics at transcriptional start sites, but is characterized by hypo-methylation of enhancer-like domains. We used cell fractions of intestines from Lgr5-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Lgr5 promoter. RNA was isolated from several FACS-sorted cell populations, one expressing GFP at high levels (GFPhi) and the other expressing GFP at low levels (GFPlow), both from small intestine and skin. We also isolated RNA from intestinal epithelial villus cells. Differentially labelled cRNA from GFPhi, GFPlow and villus cells from three different sorts (each combining three different mice) were hybridized on Affymetrix HT MG-430 PM arrays.
2013-04-25 | E-GEOD-46303 | biostudies-arrayexpress
Project description:The identification of Lgr5 as an intestinal stem cell marker has made it possible to isolate and study primary stem cells from small intestine and skins. Applying genome-wide resequencing of bisulfite-treated genomic DNA, we profiled the DNA methylation changes of FACS-sorted Lgr5+ve stem cells and their immediate undifferentiated daughter cells from small intestine and skin. In addition to this, we also analyzed terminally differentiated villus cells. We find that terminal differentiation of an adult solid-tissue stem cell does not require DNA methylation dynamics at transcriptional start sites, but is characterized by hypo-methylation of enhancer-like domains.
2013-04-25 | GSE46303 | GEO
Project description:The endodermal lining of the adult gastro-intestinal tract harbors stem cells that are responsible for the day-to-day regeneration of the epithelium. Stem cells residing in the pyloric glands of the stomach and in the small intestinal crypts differ in their differentiation program and in the gene repertoire that they express. Both types of stem cells have been shown to grow from single cells into 3D structures (organoids) in vitro. We show that single adult Lgr5-positive stem cells, isolated from small intestinal organoids, require Cdx2 to maintain their intestinal identity and are converted cell-autonomously into pyloric stem cells in the absence of this transcription factor. Clonal descendants of Cdx2null small intestinal stem cells enter the gastric differentiation program instead of producing intestinal derivatives. Conversely, forced expression of Cdx2 in gastric organoids results in their intestinalization. The intestinal genetic program is thus critically dependent on the single transcription factor encoding gene Cdx2. Small intestinal crypts and stomach glands were isolated from Cdx2-/fl / Lgr5-EGFP-CreERT2 mice and cultured for a week in order to generate small intestinal (SI) and stomach (Sto) in vitro organoids. The Lgr5-CreERT2 enzyme activity has been induced by overnight 4-hydroxytamoxifen induction. Tamoxifen treated and untreated Lgr5-EGFPhi SI and Sto stem cells were FACS sorted and seeded back into ENRWfg (Sto med) culture conditions in order to generate Cdx2-/fl small intestinal (Control SI), Cdx2null small intestinal (Cdx2null SI) and Cdx2-/fl stomach (Control Sto) clonal organoids. Cdx2-/fl SI organoids and Cdx2-/fl Sto organoids have been also cultured in ENR (SI med) to induce differentiation. After some passages of clonal organoid expansion, RNA was isolated from Control SI, Cdx2null SI and Control Sto Lgr5-EGFPhi FACS sorted stem cell populations and from smal intestinal and stomach organoids cultured in different conditions and hybridized on Affymetrix Mouse Gene ST 1.1 arrays.
2014-12-04 | E-GEOD-51751 | biostudies-arrayexpress
Project description:We used unbiased whole genome bisulfite sequencing (WGBS) to identify DNA methylation changes in the intestinal stem cells (ISCs) or their progeny during the suckling period of mouse colon development. Lgr5-EGFP mice were used to identify ISC populations in the colons. WGBS were performed using EGFP labeled Lgr5+ ISCs and epithelial cell adhesion molecule (EpCAM) labeled epithelial cells isolated at the beginning and end of the suckling period (postnatal day 0-P0 and P21).
2015-08-25 | E-GEOD-58532 | biostudies-arrayexpress
Project description:The small intestine is a rapidly proliferating organ that is maintained by a small population of Lgr5-expressing intestinal stem cells (ISCs). However, several Lgr5-negative ISC populations have been identified, and this remarkable plasticity allows the intestine to rapidly respond to both the local environment and to damage. The mediators of such plasticity are still largely unknown. Using intestinal organoids and mouse models, we show that upon ribosome impairment (driven by Rptor deletion, amino acid starvation, or low dose cyclohexamide treatment) ISCs gain an Lgr5-negative, fetal-like identity. This is accompanied by a rewiring of metabolism. Our findings suggest that the ribosome can act as a sensor of nutrient availability, allowing ISCs to respond to the local nutrient environment. Mechanistically, we show that this phenotype requires the activation of ZAKɑ, which in turn activates YAP, via SRC. Together, our data reveals a central role for ribosome dynamics in intestinal stem cells, and identify the activation of ZAKɑ as a critical mediator of stem cell identity.
2022-06-13 | PXD033122 | Pride
Project description:The identification of Lgr5 as an intestinal stem cell marker has made it possible to isolate and study primary intestinal stem cells. Applying quantitative mass spectrometry as well as transcriptomic analysis, we profiled the protein and gene changes between FACS-sorted Lgr5+ve stem cells and their immediate undifferentiated daughter cells. The overall comparison of mRNA and protein levels revealed a high level of correlation, implying that the initial control of intestinal stem cell biology occurs largely at the mRNA level. Taken together, our study presents a valuable resource for the study of intestinal stem cell biology. We used cell fractions of intestines from Lgr5-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Lgr5 promoter. RNA was isolated from two FACS sorted cell populations, one expressing GFP at high levels (GFPbright) and the other expressing GFP at low levels (GFPdim). The gates set to sort cells for the expression profiling were the same as for the cells used for the mass spectometry analysis. cRNA from GFPbright and GFPdim cells from three different sorts (each combining three to four mice) were hybridized on Affymetrix Mouse HT MG-430 PM plate arrays.
2012-08-12 | E-GEOD-33948 | biostudies-arrayexpress
Project description:To investigate the influence of Interleukin-22 (IL-22) on colonic intestinal stem cells, we assessed gene expression in these cells during homeostasis and after induction of DNA damage. IL-22 is a lymphocyte-derived cytokine that targets exclusively non-hematopoietic cells. The receptor is expressed on intestinal epithelial cells, including Lgr5+ stem cells. The colonic Lgr5+ epithelial stem cells were highly purified as DAPI-EpCam+CD45-CD24MedLgr5+. DNA damage was induced by whole body irradiation with 8 Gy and cells were isolated 24h after exposure. The following populations were analyzed: Wildtype, unirradiated (Ctrl) Il22-/-, unirradiated (Ctrl) Wildtype, 24h after 8Gy Il22-/-, 24h after 8Gy
2019-01-01 | E-MTAB-6723 | biostudies-arrayexpress
Project description:The mammalian intestinal epithelium has a unique organization where crypts harboring stem cells produce progenitors and finally clonal populations of differentiated cells. Remarkably, the epithelium is replaced every three to five days throughout adult life. Disrupted maintenance of the intricate balance of proliferation and differentiation leads to loss of epithelial integrity, barrier function, or cancer. There is a tight correlation between epigenetic status of genes and expression changes during differentiation; however, the mechanism of how changes in DNA methylation direct gene expression and the progression from stem cells to their differentiated descendants is unclear. Using conditional gene ablation of the maintenance methyltransferase Dnmt1, we demonstrate that reducing DNA methylation causes intestinal crypt expansion in vivo. Determination of the base-resolution DNA methylome in stem cells and their differentiated descendants shows that DNA methylation is dynamic at enhancers, which are often associated with genes important for both stem cell maintenance and differentiation. We establish that the loss of DNA methylation at intestinal stem cell gene enhancers causes inappropriate gene expression and delayed differentiation.
2014-03-13 | E-MTAB-2350 | biostudies-arrayexpress
Project description:The identification of Lgr5 as an intestinal stem cell marker has made it possible to isolate and study primary stem cells from small intestine. Using the cell cycle specific expression og the mKi67 gene, we generated a novel Ki67-RFP knock-in allele which identifies dividing cells. Using Lgr5-GFP;Ki67-RFP mice, we isolated CBCs with distinct Wnt signaling levels and cell cycle features, and analyzed their global gene expression pattern using microarrays. We concluded that the cycling Lgr5hi stem cells exit the cell cycle in transition into the secretory lineage. Lgr5med Ki67low intermediate precursors reside in the zone of differentiation, resemble quiescent stem cells and generate the Dll1+ secretory precursors and the label retaining cells. Our findings support the cycling stem cell hypothesis and highlight the heterogeneity of early progenitors during lineage commitment. We used cell fractions of intestines from Lgr5-EGFP-ires-CreERT2 mice, expressing GFP under the control of the Lgr5 promoter, and Ki67-TagRFP mice where the RFP is fused to the C-terminus of the endogenous Ki67 gene. RNA was isolated from several FACS sorted cell populations of combinations expressing different levels of GFP and RFP: GFP high RFP high (Lgr5hi Ki67hi), GFP high RFP low (Lgr5hi Ki67low), GFP medium RFP high (Lgr5med Ki67high) and GFP medium RFP low (Lgr5med Ki67low). Purified RNA was processed, hybridized, and scanned according to the manufacturerM-bM-^@M-^Ys protocol and were hybridized on Affymetrix Mouse Gene ST 1.1 arrays).
2014-09-08 | E-GEOD-52813 | biostudies-arrayexpress
Project description:Stomach and intestinal adult epithelia harbor stem cells that are responsible for their continuous regeneration. Stomach and intestinal stem cells differ in their differentiation program and in the gene repertoire that they express. We show that single adult Lgr5-positive stem cells, isolated from 3D cultured small intestinal organoids, require Cdx2 to maintain their intestinal identity and are converted cell-autonomously into stomach-pyloric stem cells in the absence of this transcription factor. In order to obtain Cdx2null intestinal stem cells carrying the Lgr5-EGFP marker, 5-6 days old small intestinal organoids generated from Cdx2-/fl/Lgr5-EGFP-Ires-CreERT2 mice were incubated with 1 µM of 4-hydroxytamoxifen in intestinal culture medium for 16h to activate the Cre recombinase. Controls were 4-hydroxytamoxifen-untreated small intestinal (Control SI) and stomach (Control Sto) organoids issued from mice with the same genotype. The organoids were dissociated and sorted for EGFPhi. Cdx2null, Control SI and Control Sto clonal organoids were generated and expanded from Lgr5-EGFPhi single cells in stomach specific culture medium (ENRWfg) and RNA was isolated for RNA-Seq analysis. Cdx2+ Stomach (Sto) organoids were generated by infection of the wild type stomach organoids with lentiviral stock expressing Cdx2. They were cultured in stomach medium (ENRWfg) and RNA was isolated for RNA-Seq analysis
2014-12-05 | E-GEOD-62784 | biostudies-arrayexpress