Genotypic and phenotypic characterization of the side population of gastric cancer cell lines
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ABSTRACT:
ORGANISM(S): Homo sapiens
PROVIDER: GSE20058 | GEO | 2010/12/01
SECONDARY ACCESSION(S): PRJNA124329
REPOSITORIES: GEO
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Project description:To sum up our results, we managed to isolate a reproducible SP cell fraction of human gastric carcinoma cell lines for the first time. Those cells differed from the rest of the population regarding morphology and biological behaviour. They had the ability to build a population resembling the basic tumor population and therefore performed differenziation and asymmetric cell division. SP cells expressed potential stem cell markers like musashi 1 and Cd133 and also showed a high expression of genes that encode for stem cell properties. The so identified phenotypic and genetic factors showed a variable expression in tumor samples of patient series and might be applicable as prognostic factors in the diagnosis of gastric carcinoma. Aims: The side population (SP) of tumor cell lines shares characteristics with tumor stem cells. In this study we phenotypically and genotypically characterized the SP of gastric cancer cell lines. Methods: The SP was obtained from MKN45- and AGS-gastric cancer cells using Hoechst 33342 staining and fluorescence-activated cell sorting (FACS). SP cells were subsequently studied morphologically (cytology, immunocytochemistry), on the transcriptional level (gene array) and in cell culture (recultivation assays). Genes differentially expressed in the SP cells were finally searched by immunohistochemistry in neoplastic and non-neoplastic gastric tissue from gastric cancer patients. Results: The SP was reproducibly obtained from gastric cancer cell lines. The SP cells were smaller and rounder then non-SP cells. SP cells self-renewed in re-cultivation experiments and differentiated into SP- and non-SP cells. Re-cultivated SP- and non-SP cells showed distinct phenotypes in culture regarding cell shape and colony-formation. SP cells had increased levels of the stem cell markers CD133 and Musashi 1. Transcriptional analyses demonstrated that SP cells express genes that encode for stem cell properties like FZD7, HEY1, SMO and ADAM17. Finally we detected the transcripts of these genes in tissue samples from patients with gastric cancer. Conclusions: Human gastric cancer cell lines enclose a phenotypically and genotypically distinct cell population with tumor stem cell features. Phenotypical characteristics of this distinct cell population are also present in gastric cancer tissue.
2010-12-01 | E-GEOD-20058 | biostudies-arrayexpress
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.
2010-08-15 | GSE21633 | GEO
Project description:Human solid tumors contain rare cancer side population (SP) cells, which expel the fluorescencent dye H33342 and display cancer stem cell characteristics. Transcriptional profiling of cancer SP cells isolated by H33342 fluorescence analysis is a newly emerging approach to discover cancer stem cell markers and aberrant differentiation pathways. Using Affymetrix expression microarrays this study investigated differential gene expression between SP and non-SP (NSP) cells isolated from the CAL-51 human mammary carcinoma cell line. Keywords: cell type comparison
2009-12-31 | GSE18773 | GEO
Project description:Background The side population (SP) phenotype, a subset of cells that extrude the nucleic acid dye Hoechst 33342, has been reported to be enriched for stem cells in several human normal tissues, cancers and cell lines, and thus may be useful for the identification and isolation of cancer stem cells. Methods We demonstrated the presence of SP fractions in all analyzed tumor cell lines ranging between 7- 20% of cells. To identify gene expression patterns that contribute to SP phenotype, microarray analysis of SP and non-SP cells was performed. We additionally confirmed regulation of some genes by qRT-PCR. Results Surprisingly, only a subset of few genes in SP cells showed altered gene expression. A total of 11 genes in A2C12, 103 genes in cRAF_cMYC and 101 genes in beta5 SP cells were regulated. Most regulated genes are involved in transcription / transcriptional regulation and transport. In addition, we found no enrichment of previously described stem cell marker like CD24a, CD90 or CD133 and also the ABC transporter ABCG2 was only slightly increased in side population fraction of two cell lines. But despite the few differences between SP and non-SP cells, the beta5 tumor cells were highly tumorigenic due to their capacity to form an original murine tumor when injected in NOD/SCID mice. Conclusion These findings stand in contrast to other observations but indicate that there are further factors responsible for the SP phenotype and that SP cells alone are not suitable as a universal stem cell marker.
2016-12-31 | GSE22275 | 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:Human solid tumors contain rare cancer side population (SP) cells, which expel the fluorescencent dye H33342 and display cancer stem cell characteristics. Transcriptional profiling of cancer SP cells isolated by H33342 fluorescence analysis is a newly emerging approach to discover cancer stem cell markers and aberrant differentiation pathways. Using Affymetrix expression microarrays this study investigated differential gene expression between SP and non-SP (NSP) cells isolated from the CAL-51 human mammary carcinoma cell line. Keywords: cell type comparison To characterize differential gene expression between CAL-51 breast cancer SP and NSP cells, three consecutive cell culture passages of CAL-51 were independently subjected to H33342 labeling and dual wavelength fluorescence analysis and were then flow cytometrically sorted into SP and NSP cell fractions. Subsequently, each of the six cell preparations was subjected to global transcriptional profiling using Affymetrix HG U133 Plus 2.0 expression microarrays.
2009-12-31 | E-GEOD-18773 | biostudies-arrayexpress
Project description:To identify the gene expression signature associated with CD133, the well-known stem cell markers, three gastric cancer cell lines were obtained (KATO-III, SNU201 and SNU601). Cultured gastric cancer celllines were sorted into CD133+ and CD133- population by FACS sorting and microarray-based gene expression profiling was performed.
2018-04-04 | GSE112631 | GEO
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.
2011-03-12 | GSE27909 | GEO
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
2014-09-18 | GSE61547 | GEO
Project description:We report the application of single-cell RNA sequencing data on Side Population (SP) cells and their Non-Side Population (NSP) counterparts in a mouse model of undifferentiated pleomorphic sarcoma (UPS). SP cells exhibit tumor propagting cell properties characterized by enhanced tumorigenicity and self-renewal potential. The purpose of this experiment is to investigate cellular heterogeneity at the gene expression level in UPS tumors and lineage relationships between different subpopulation of tumor cells. We generated the gene expression profiles of individual cells in the SP and NSP compartments of mouse UPS.
2022-01-07 | GSE162847 | GEO