Browse
Submit Data
Databases
API
Help

Dataset Information

24 Views

0 Connections

0 Citations

0 Reanalyses

0 Downloads

Omics score: 0

Isolation and functional characterization of murine prostate stem cells.

ABSTRACT: The ability to isolate prostate stem cells is essential to explore their role in prostate development and disease. In vitro prostate colony- and sphere-forming assays were used to quantitatively measure murine prostate stem/progenitor cell enrichment and self-renewal. Cell surface markers were screened for their ability to positively or negatively enrich for cells with enhanced growth potential in these assays. Immunohistochemical and FACS analyses demonstrate that specific cell surface markers can be used to discriminate prostate stromal (CD34(+)), luminal epithelial (CD24(+)CD49f(-)), basal epithelial (CD24(+)CD49f(+)), hematopoietic (CD45(+), Ter119(+)), and endothelial (CD31(+)) lineages. Sorting for cells with a CD45(-)CD31(-)Ter119(-)Sca-1(+)CD49f(+) antigenic profile results in a 60-fold enrichment for colony- and sphere-forming cells. These cells can self-renew and expand to form spheres for many generations and can differentiate to produce prostatic tubule structures containing both basal and luminal cells in vivo. These cells also localize to the basal cell layer within the region of the gland that is proximal to the urethra, which has been identified as the prostate stem cell niche. Prostate stem cells can be isolated to a purity of up to 1 in 35 by using this antigenic profile. The remarkable similarity in cell surface profile between prostate and mammary gland stem cells suggests these markers may be conserved among epithelial stem cell populations.

SUBMITTER: Lawson DA

PROVIDER: S-EPMC1716155 | biostudies-literature |

REPOSITORIES: biostudies-literature

ACCESS DATA

Json Xml

Similar Datasets

Isolation, cultivation and characterization of adult murine prostate stem cells.

Project description:The successful isolation and cultivation of prostate stem cells will allow us to study their unique biological properties and their application in therapeutic approaches. Here we describe step-by-step procedures on the basis of previous work in our laboratory for the harvesting of primary prostate cells from adolescent male mice by a modified enzymatic procedure; the isolation of an enriched population of prostate stem cells through cell sorting; and the cultivation of prostate stem cells in vitro and characterization of these cells and their stem-like activity, including in vivo tubule regeneration. Normally, it will take approximately 8 h to harvest prostate cells, isolate the stem cell-enriched population and set up the in vitro sphere assay. It will take up to 8 weeks to analyze the unique properties of the stem cells, including their regenerative capacity in vivo.

| S-EPMC2943378 | biostudies-literature

High aldehyde dehydrogenase activity: a novel functional marker of murine prostate stem/progenitor cells.

Project description:We have shown previously that prostatic stem/progenitor cells can be purified from isolated prostate ducts, based on their high expression of the Sca-1 surface antigen. We now report that high levels of aldehyde dehydrogenase (ALDH) activity are present in a subset of prostate epithelial cells that coexpress a number of antigens found on stem/progenitor cells of other origins (CD9, Bcl-2, CD200, CD24, prominin, Oct 3/4, ABCG2, and nestin). Almost all of these cells expressing high levels of ALDH activity also express Sca-1 and a third of them express high levels of this antigen. The cells with high levels of ALDH activity have greater in vitro proliferative potential than cells with low ALDH activity. Importantly, in an in vivo prostate reconstitution assay, the cells expressing high levels of ALDH activity were much more effective in generating prostatic tissue than a population of cells with low enzymatic activity. Thus, a high level of ALDH activity can be considered a functional marker of prostate stem/progenitor cells and allows for simple, efficient isolation of cells with primitive features. The elucidation of the role of ALDH in prostate stem/progenitor cells may lead to the development of rational therapies for treating prostate cancer and benign prostatic hyperplasia.

| S-EPMC2887284 | biostudies-literature

Isolation and functional interrogation of adult human prostate epithelial stem cells at single cell resolution.

Project description:Using primary cultures of normal human prostate epithelial cells, we developed a novel prostasphere-based, label-retention assay that permits identification and isolation of stem cells at a single cell level. Their bona fide stem cell nature was corroborated using in vitro and in vivo regenerative assays and documentation of symmetric/asymmetric division. Robust WNT10B and KRT13 levels without E-cadherin or KRT14 staining distinguished individual stem cells from daughter progenitors in spheroids. Following FACS to isolate label-retaining stem cells from label-free progenitors, RNA-seq identified unique gene signatures for the separate populations which may serve as useful biomarkers. Knockdown of KRT13 or PRAC1 reduced sphere formation and symmetric self-renewal highlighting their role in stem cell maintenance. Pathways analysis identified ribosome biogenesis and membrane estrogen-receptor signaling enriched in stem cells with NF-ĸB signaling enriched in progenitors; activities that were biologically confirmed. Further, bioassays identified heightened autophagy flux and reduced metabolism in stem cells relative to progenitors. These approaches similarly identified stem-like cells from prostate cancer specimens and prostate, breast and colon cancer cell lines suggesting wide applicability. Together, the present studies isolate and identify unique characteristics of normal human prostate stem cells and uncover processes that maintain stem cell homeostasis in the prostate gland.

| S-EPMC5581994 | biostudies-literature

Isolation and long-term expansion of murine epidermal stem-like cells.

Project description:Epidermis is the most outer layer of the skin and a physical barrier protecting the internal tissues from mechanical and environmental insults. The basal keratinocytes, which, through proliferation and differentiation, supply diverse cell types for epidermal homeostasis and injury repair. Sustainable culture of murine keratinocyte, however, is a major obstacle. Here we developed murine keratinocyte lines using low-Ca2+ (0.06 mM) keratinocyte serum-free medium (KSFM-Ca2+) without feeder cells. Cells derived in this condition could be subcultured for >70 passages. They displayed basal epithelial cell morphology and expressed keratin (Krt) 14, but lacked the epithelial-characteristic intercellular junctions. Moreover, these cells could be adapted to grow in the Defined-KSFM (DKSFM) media containing 0.15 mM Ca2+, and the adapted cells established tight- and adherens-junctions and exhibited increased Krt1/10 expression while retained subculture capacity. Global gene expression studies showed cells derived in KSFM-Ca2+ media had enriched stem/proliferation markers and cells adapted in DKSFM media had epithelial progenitor signatures. Correspondingly, KSFM-Ca2+-derived cells exhibited a remarkable capacity of clonal expansion, whereas DKSFM-adapted cells could differentiate to suprabasal epithelial cell types in 3-dimentional (3D) organoids. The generation of stem-like murine keratinocyte lines and the conversion of these cells to epithelial progenitors capable of terminal differentiation provide the critically needed resources for skin research.

| S-EPMC8284819 | biostudies-literature

Characterization of Dicer-deficient murine embryonic stem cells.

Project description:Dicer is an RNase III-family nuclease that initiates RNA interference (RNAi) and related phenomena by generation of the small RNAs that determine the specificity of these gene silencing pathways. We have previously shown that Dicer is essential for mammalian development, with Dicer-deficient mice dying at embryonic day 7.5 with a lack of detectable multipotent stem cells. To permit a more detailed investigation of the biological roles of Dicer, we have generated embryonic stem cell lines in which their single Dicer gene can be conditionally inactivated. As expected, Dicer loss compromises maturation of microRNAs and leads to a defect in gene silencing triggered by long dsRNAs. However, the absence of Dicer does not affect the ability of small interfering RNAs to repress gene expression. Of interest, Dicer loss does compromise the proliferation of ES cells, possibly rationalizing the phenotype previously observed in Dicer-null animals. Dicer loss also affects the abundance of transcripts from mammalian centromeres but does so without a pronounced affect on histone modification status at pericentric repeats or methylation of centromeric DNA. These studies provide a conditional model of RNAi deficiency in mammals that will permit the dissection of the biological roles of the RNAi machinery in cultured mammalian cells.

| S-EPMC1185572 | biostudies-literature

Isolation and functional characterization of the arteriole hematopoietic stem cell niche

Project description:Hematopoietic stem cells (HSCs) inhabit distinct microenvironments within the adult bone marrow (BM) that govern the delicate balance between HSC quiescence, self-renewal, and differentiation. It has been suggested that quiescent HSCs localize adjacent to BM arteriole endothelial cells in a significant and non-random distribution. This data suggests that the arteriole BM vascular niche may be the primary HSC niche. Because the BM arteriole niche is composed of tightly-associated pericytes, including smooth muscle actin+, LepR+, Nestin+, NG2+, and nonmyelinating Schwann cells, we sought to begin to uncouple the arteriole BM EC niche by examining its capacity to support the maintenance and expansion of HSCs ex vivo and in vivo. We developed a method to isolate and culture BM arteriole endothelial cells in serum-/growth factor-free conditions, allowing for a non-biased approach to examining their instructive function. Utilizing our protocol, we demonstrate that BM endothelial cells, but not BM stromal cells, have the capacity to expand long-term repopulating, multi-lineage HSCs in lieu of complex serum and cytokine supplementation. In addition, transplantation of arteriole endothelial cells promoted rapid hematopoietic recovery and protected HSCs following an LD50 dose of myeloablative irradiation. These data demonstrate that arteriole-derived BM endothelial cells are endowed with the necessary signals to support the self-renewal and regenerative capacity of LT-HSCs and that transplantation of arteriole BM endothelial cells could be used as a therapeutic means to decrease pancytopenias associated with myeloablative treatments to treat a wide array of disease states. Transcriptome sequencing of bone marrow endothelial cells and bone marrow stroma, in vitro and in vivo, with and without HSC co-culture.

2015-11-04 | E-GEOD-61636 | biostudies-arrayexpress

Isolation and functional characterization of the arteriole hematopoietic stem cell niche

2015-11-04 | GSE61636 | GEO

Isolation and Characterization of Multipotent Turkey Tendon-Derived Stem Cells.

Project description:Tendon injuries are among the most common and severe hand injuries with a high demand for functional recovery. Stem cells have been identified and isolated from different species and a variety of tissues for the sake of regenerative medicine. Recently, turkey has been suggested as a potential new large animal model for flexor tendon-related research. However, turkey tissue-specific stem cells have not been investigated. Here, we presented the isolation and verification of tendon-derived stem cells (TDSCs) from 6- to 8-month-old heritage-breed turkey. TDSCs were isolated from turkey flexor tendon by plating nucleated cells at the determined optimal density. Approximately 4% of the nucleated cells demonstrated clonogenicity, high proliferation rate, and trilineage differentiation potential after induction culturing. These cells expressed surface antigens CD90, CD105, and CD44, but did not express CD45. There was a high level of gene expression of tenogenic markers in TDSCs, including mohawk, collagen type I, tenascin C, and elastin. Turkey TDSCs also expressed transcription factors PouV, Nanog, and Sox2, which are critically involved in the regulation of stemness. The successful isolation of tendon-derived stem cells from turkey was beneficial for future studies in tendon tissue engineering and would help in the development of new treatment for tendon diseases using this novel animal model.

| S-EPMC6011053 | biostudies-literature

Facile isolation and the characterization of human retinal stem cells.

Project description:This study identifies and characterizes retinal stem cells (RSCs) in early postnatal to seventh-decade human eyes. Different subregions of human eyes were dissociated and cultured by using a clonal sphere-forming assay. The stem cells were derived only from the pars plicata and pars plana of the retinal ciliary margin, at a frequency of approximately 1:500. To test for long-term self-renewal, both the sphere assay and monolayer passaging were used. By using the single sphere passaging assay, primary spheres were dissociated and replated, and individual spheres demonstrated 100% self-renewal, with single spheres giving rise to one or more new spheres in each subsequent passage. The clonal retinal spheres were plated under differentiation conditions to assay the differentiation potential of their progeny. The spheres were produced all of the different retinal cell types, demonstrating multipotentiality. Therefore, the human eye contains a small population of cells (approximately equal to 10,000 cells per eye) that have retinal stem-cell characteristics (proliferation, self-renewal, and multipotentiality). To test the in vivo potential of the stem cells and their progeny, we transplanted dissociated human retinal sphere cells, containing both stem cells and progenitors, into the eyes of postnatal day 1 NOD/SCID mice and embryonic chick eyes. The progeny of the RSCs were able to survive, migrate, integrate, and differentiate into the neural retina, especially as photoreceptors. Their facile isolation, integration, and differentiation suggest that human RSCs eventually may be valuable in treating human retinal diseases.

| S-EPMC524825 | biostudies-literature

Isolation and molecular characterization of cancer stem cells in MMTVWnt-1 murine breast tumors

Project description:In human breast cancers, a phenotypically distinct minority population of tumorigenic cancer (TG) cells (sometimes referred to as cancer stem cells) drives tumor growth when transplanted into immunodeficient mice. Our objective was to identify a mouse model of breast cancer stem cells that could have relevance to studying human breast cancer. To do so, we utilized breast tumors of the MMTVWnt-1 mice. MMTV-Wnt-1 breast tumors were harvested, dissociated into single cell suspensions, and FACS sorted on Thy1, CD24, and CD45. FACS sorted cells were then injected into recipient background FBV/NJ female mice. Thy1+CD24+ cancer cells, which constitute approximately 1-4% of tumor cells were highly enriched for cells capable of regenerating new tumors when compared to cells of the tumor that did not fit this profile (“Not Thy1+CD24+”). Resultant tumors were of the same phenotypic diversity as the original tumor and behaved in a similar manner when passaged. Microarray analysis comparing Thy1+CD24+ tumor cells to “Not Thy1+CD24+” cells identified a list of differentially expressed genes. Orthologs of these differentially expressed genes predicted survival of human breast cancer patients from two different study groups. These studies suggest that there is a cancer stem cell compartment in the MMTV-Wnt-1 murine breast tumor and that there is a clinical utility of this model for the study of cancer stem cells. Keywords: cell type comparison

2007-11-19 | GSE8828 | GEO

OmicsDI is part of the ELIXIR infrastructure

OmicsDI is an Elixir interoperability service. Learn more ›

Tweets

OmicsDI Databases

PRIDE
PeptideAtlas
MassIVE
JPOST Repository
Physiome Model Repository

EGA
EVA
ENA
LINCS
PAXDB
Cell Collective

MetaboLights
Metabolomics Workbench
MetabolomeExpress
GNPS
BioModels
FAIRDOMHub

ArrayExpress
dbGaP
ExpressionAtlas
GEO
NODE

Information

Databases
Help
API
Contact us
Code on GitHub
Terms of use
Submit Data