Gene expression profile of zebrafish kidney side population (SP) cells
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ABSTRACT:
ORGANISM(S): Danio rerio
PROVIDER: GSE12955 | GEO | 2008/10/10
SECONDARY ACCESSION(S): PRJNA110937
REPOSITORIES: GEO
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Project description:Side population (SP) cells are identified based on their capacity to efflux of the fluorescent dye Hoechst 33342, and are enriched for hematopoietic stem cells (HSCs) in mammalian bone marrow. We recently demonstrated that SP cells were present in the teleost kidney, the main hematopoietic organ in teleosts, and were enriched for HSCs. In this analysis, to identify the regulated genes in teleost HSCs, gene expression analysis of zebrafish kidney SP cells were performed using the GeneChip Zebrafish Genome Array. Experiment Overall Design: Based on their Hoechst fluorescence intensity, lymphoid cells (FS-low, SS-low) from zebrafish kidney were subdivided into SP and MP populations (referred to as âzSPâ and âzMPâ). To minimize the biological variability, 20 zebrafish were used for 3 independent cell sorting experiments and lysates from these 3 experiments were pooled by cell type. Each RNA sample was split into two aliquots and used for amplification, labeling, and hybridization to independent arrays.
2008-10-26 | E-GEOD-12955 | biostudies-arrayexpress
Project description:Analysis of Hoechst dye 33342-effluxing side population (SP) cells from B-CLL peripheral blood mononuclear cells.
2011-09-27 | GSE30171 | GEO
Project description:The traditional view of hematopoiesis has been that all the cells of the peripheral blood are the progeny of a unitary homogeneous pool of hematopoietic stem cells (HSCs). Recent evidence suggests that the hematopoietic system is actually maintained by a consortium of HSC subtypes with distinct functional characteristics. We show here that myeloid-biased HSCs (My-HSCs) and lymphoid-biased (Ly-HSCs) can be purified according to their capacity for Hoechst dye efflux in combination with canonical HSC markers. We used microarray expression profiling to determine the transcriptional profiles of myeloid-biased lower-SP HSCs and lymphoid-biased upper-SP HSCs
2010-03-01 | GSE16475 | GEO
Project description:The traditional view of hematopoiesis has been that all the cells of the peripheral blood are the progeny of a unitary homogeneous pool of hematopoietic stem cells (HSCs). Recent evidence suggests that the hematopoietic system is actually maintained by a consortium of HSC subtypes with distinct functional characteristics. We show here that myeloid-biased HSCs (My-HSCs) and lymphoid-biased (Ly-HSCs) can be purified according to their capacity for Hoechst dye efflux in combination with canonical HSC markers. We used microarray expression profiling to determine the transcriptional profiles of myeloid-biased lower-SP HSCs and lymphoid-biased upper-SP HSCs Three biological replicates were analyzed for each HSC subpopulation. Lower-SP and upper-SP HSCs were purified from three pools of mice on separate days. HSCs were further purified with the addition of canonical HSC makers; Sca-1+ c-Kit+ Lineage-
2010-03-06 | E-GEOD-16475 | biostudies-arrayexpress
Project description:Analysis of Hoechst dye 33342-effluxing side population (SP) cells from B-CLL peripheral blood mononuclear cells. 9 biological replicates from B-CLL patients sorted into CD5+CD19+ SP and non-SP subsets. Two color comparative gene expression using Agilent microarrays.
2011-09-26 | E-GEOD-30171 | biostudies-arrayexpress
Project description:The kidney is the main hematopoietic organ in teleost fish. Various stages of hematopoietic cells were observed in the interstitial tissue of the kidney. We recently demonstrated that zebrafish hematopoietic stem cells (HSCs) expressing jam1a were specifically localized along the renal collecting ducts in the adult kidney. Interestingly, most of HSCs invaded into the intracellular spaces of the epithelium in collecting ducts, suggesting that collecting ducts provide a specific microenvironment for HSCs. In order to identify niche factors in collecting ducts, we performed microarray analysis in collecting ducts isolated from the Tg(jam1a:EGFP) zebrafish kidney. Collecting ducts were isolated from Tg(jam1a:EGFP) zebrafish under a fluorescent microscope. To examine the effect of X-ray irradiation on the niche, collecting ducts were isolated from the fish irradiated with 25Gy. The whole kidney tissues were also used for a comparison analysis. Two independent replicates consisting of five zebrafish were prepared for each sample.
2012-08-24 | E-GEOD-32363 | biostudies-arrayexpress
Project description:The kidney is the main hematopoietic organ in teleost fish. Various stages of hematopoietic cells were observed in the interstitial tissue of the kidney. We recently demonstrated that zebrafish hematopoietic stem cells (HSCs) expressing jam1a were specifically localized along the renal collecting ducts in the adult kidney. Interestingly, most of HSCs invaded into the intracellular spaces of the epithelium in collecting ducts, suggesting that collecting ducts provide a specific microenvironment for HSCs. In order to identify niche factors in collecting ducts, we performed microarray analysis in collecting ducts isolated from the Tg(jam1a:EGFP) zebrafish kidney.
2012-08-24 | GSE32363 | GEO
Project description:During reproductive life, the human endometrium undergoes around 480 cycles of growth, breakdown and regeneration should pregnancy not be achieved. This outstanding regenerative capacity is the basis for women’s cycling and its dysfunction may be involved in the etiology of pathological disorders. Therefore, the human endometrial tissue must rely on a remarkable endometrial somatic stem cells (SSC) population. Here we explore the hypothesis that human endometrial side population (SP) cells correspond to somatic stem cells. We isolated, identified and characterized the SP corresponding to the stromal and epithelial compartments using endometrial SP genes signature, immunophenotyping and characteristic telomerase pattern. We analyzed the clonogenic activity of SP cells under hypoxic conditions and the differentiation capacity in vitro to adipogenic and osteogenic lineages. Finally, we demonstrated the functional capability of endometrial SP to develop human endometrium after subcutaneous injection in NOD-SCID mice. Briefly, SP cells of human endometrium from epithelial and stromal compartments display genotypic, phenotypic and functional features of SSC. Human tissue collection: Human endometrial biopsies were taken using a Pipelle catheter under sterile conditions (from 18 to 48 years) throughout the menstrual cycle. Epithelial and stromal separation: Epithelial and stromal fractions were isolated using an established protocol with modifications. Briefly, samples were carefully dissected and minced into 1-2mm fragments, and enzymatically digested in DMEM containing 10 mg/ml collagenase type IA. Stromal cells (single cells or small aggregates) and epithelial glands were separated on a size basis using gravity sedimentation and membrane filtration. Cell suspensions were treated with erythrocyte lysis solution and evaluation of cell viability was performed with Propidium Iodide (PI; 5 μg/ml). Hoechst 33342 labeling: Cells isolated from the endometrial epithelial and stromal fractions were resuspended in DMEM prewarmed at 37ºC and supplemented with 2% FBS and 10mM HEPES. The cell suspension was labeled in the same media with 5 μg/ml of Hoechst 33342 dye (Ho-33342), either alone or in combination with 100 μM verapamil (Vp), in a water bath at 37ºC for 90-120 minutes. Then, cells were centrifuged for 6 minutes at 4ºC, and were resuspended in cold HBSS supplemented with 2%FBS and 10mM HEPES. PI permits to exclude dead cells prior to the flow cytometric analysis and sorting. Isolation of human endometrial SP cells: Cells were analyzed and sorted by a MoFlo® jet-in-air high speed sorter. Excitation was performed with a water cooled Enterprise II ion laser which operated at the 351 nm and 488 nm wavelengths, and worked at 30 mW. Hoechst 33342 blue and red fluorescences were detected through 405/30 and 670/20 nm band-pass filters respectively, by measuring the signals on a linear scale. PI fluorescence was detected through a band-pass filter of 613/20 on a logarithmic scale. The gates for cell sorting were defined to collect live cells with a low Hoechst 33342 fluorescence (SP fraction), as well as live cells with a high Hoechst 33342 fluorescence (NSP fraction). Microarrays experiments: Endometrial tissue consisted in a total of 8 endometrial biopsies (epithelial (n=8) and stromal (n=8) endometrial cell suspensions, epithelial SP fraction (n=8) and stromal SP fractions (n=8) separately) which were pooled in pairs at the RNA levels. Four microarrays were analyzed per group.
2010-08-15 | E-GEOD-21633 | biostudies-arrayexpress
Project description:Immature cell populations, including stem cells and progenitor cells, can be found in “side-population (SP)” cells. Although SP cells isolated from some adult tissues have been reported elsewhere, isolation and characterization of human trophoblast SP cells remained to be reported. We used microarrays to detail the global program of gene expression underlying cell differentiation and identified up-regulated genes of SP cells. HTR-8/SVneo SP cells or NSP cells were isolated using a cell sorter for RNA extraction and hybridization on Affymetrix microarrays. To obtain many SP or NSP cells, we cultured a large amount of HTR-8/SVneo cells (1×10^7 cells). The cells were labeled with 2.5µg/ml Hoechst 33342 dye (Molecular Probes, Eugene, OR) for 120 minutes at 37 ℃, either alone or in combination with 50 µmol/L verapamil (Sigma-Aldrich, ST Louis, MO). Finally, the cells were then passed through EPICS ALTRA HyPerSort (Beckman Coulter, Fullerton, CA) using dual wavelength analysis (blue, 424-444 nm; red, 675 nm) after excitation with 350 nm UV light.
2011-03-12 | E-GEOD-27909 | biostudies-arrayexpress
Project description:Side population (SP) cells isolated from limbal and conjunctival epithelia derive from cells that are slow cycling in vivo, a known feature of tissue stem cells. The purpose of this study was to define the molecular signature of the conjunctival side population cells by global differential gene expression to identify markers and signaling pathways associated with this cell phenotype. Four overnight cultures of freshly isolated human conjunctival epithelial cells stained with Hoechst 3342 were cytometrically sorted into SP and non-SP cohorts. RNA was isolated and processed for microarray analysis using a commercial oligonucleotide array representing more than 55,000 transcripts derived from about 30,000 different genes. Selected microarray results were validated at the gene and protein levels by quantitative PCR, and immunological methods. Data mining methods were used to identify cellular processes relevant for stem cell function. Comparative analysis of transcripts expression based on expression levels and present/absent software calls across 4 replicate experiments identified 16993 expressed conjunctival epithelial transcripts including 10,266 unique known genes. Of those genes, 1254 and 363 were over expressed (> 2-fold ) or under expressed (< 0.5-fold), respectively, in the SP. The overexpressed set included genes coding for non-epithelial genes (e.g., CD62E/E-selectin and CD93), genes that have been associated with stem cell function in other cellular systems, including several homeodomain genes and genes whose over- or under-expression may underpin the stem cell slow cycling phenotype ( e.g., dual specificity phosphatases and cyclin kinases). Experiment Overall Design: Whole human conjunctivae from unidentifiable cadaver donors, aged 55 to 65, were obtained from the National Disease Research Interchange (NDRI, Philadelphia, PA) within 48 hours of collection. No donor details apart from age, sex, and cause of death were released. The use of human tissue in the study was in accordance with the provisions of the Declaration of Helsinki and sanctioned by the Institutional Review Board. Fresh rabbit tissue was obtained from local abattoirs within one hour of sacrifice. Unless stated otherwise all reagents were procured from Sigma (St. Louis, Mo). Experiment Overall Design: Eight microarray experiments using side population (Hoechst 33342-transporting cells; SP) or non side population (Hoechst 33342-stained cells; nSP) cells from four cadavers.
2009-11-11 | E-GEOD-12631 | biostudies-arrayexpress