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Reconstruction of endometrium from human endometrial side population cell lines.

ABSTRACT: Endometrial regeneration is mediated, at least in part, by the existence of a specialized somatic stem cell (SSC) population recently identified by several groups using the side population (SP) technique. We previously demonstrated that endometrial SP displays genotypic, phenotypic and the functional capability to develop human endometrium after subcutaneous injection in NOD-SCID mice. We have now established seven human endometrial SP (hESP) cell lines (ICE 1-7): four from the epithelial and three from the stromal fraction, respectively. SP cell lines were generated under hypoxic conditions based on their cloning efficiency ability, cultured for 12-15 passages (20 weeks) and cryopreserved. Cell lines displayed normal 46XX karyotype, intermediate telomerase activity pattern and expressed mRNAs encoding proteins that are considered characteristic of undifferentiated cells (Oct-4, GDF3, DNMT3B, Nanog, GABR3) and those of mesodermal origin (WT1, Cardiac Actin, Enolase, Globin, REN). Phenotype analysis corroborated their epithelial (CD9+) or stromal (vimentin+) cell origin and mesenchymal (CD90+, CD73+ and CD45?) attributes. Markers considered characteristic of ectoderm or endoderm were not detected. Cells did not express either estrogen receptor alpha (ER?) or progesterone receptor (PR). The hESP cell lines were able to differentiate in vitro into adipocytes and osteocytes, which confirmed their mesenchymal origin. Finally, we demonstrated their ability to generate human endometrium when transplanted beneath the renal capsule of NOD-SCID mice. These findings confirm that SP cells exhibit key features of human endometrial SSC and open up new possibilities for the understanding of gynecological disorders such as endometriosis or Asherman syndrome. Our cell lines can be a valuable model to investigate new targets for endometrium proliferation in endometriosis.

SUBMITTER: Cervello I

PROVIDER: S-EPMC3119688 | biostudies-literature | 2011

REPOSITORIES: biostudies-literature

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Reconstruction of endometrium from human endometrial side population cell lines.

Cervelló Irene I Mas Aymara A Gil-Sanchis Claudia C Peris Laura L Faus Amparo A Saunders Philippa T K PT Critchley Hilary O D HO Simón Carlos C

PloS one 20110621 6

Endometrial regeneration is mediated, at least in part, by the existence of a specialized somatic stem cell (SSC) population recently identified by several groups using the side population (SP) technique. We previously demonstrated that endometrial SP displays genotypic, phenotypic and the functional capability to develop human endometrium after subcutaneous injection in NOD-SCID mice. We have now established seven human endometrial SP (hESP) cell lines (ICE 1-7): four from the epithelial and th ...[more]

PMID: 21712999

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Project description:BackgroundThe human endometrium undergoes cyclical regeneration throughout a woman's reproductive life. Ectopic implantation of endometrial cells through retrograde menstruation gives rise to endometriotic lesions which affect approximately 10% of reproductive-aged women. The high regenerative capacity of the human endometrium at eutopic and ectopic sites suggests the existence of stem/progenitor cells and a unique angiogenic system. The objective of this study was to isolate and characterize putative endometrial stem/progenitor cells and to address how they might be involved in the physiology of endometrium.Methodology/principal findingsWe found that approximately 2% of the total cells obtained from human endometrium displayed a side population (SP) phenotype, as determined by flow cytometric analysis of Hoechst-stained cells. The endometrial SP (ESP) cells exhibited preferential expression of several endothelial cell markers compared to endometrial main population (EMP) cells. A medium specific for endothelial cell culture enabled ESP cells to proliferate and differentiate into various types of endometrial cells, including glandular epithelial, stromal and endothelial cells in vitro, whereas in the same medium, EMP cells differentiated only into stromal cells. Furthermore, ESP cells, but not EMP cells, reconstituted organized endometrial tissue with well-delineated glandular structures when transplanted under the kidney capsule of severely immunodeficient mice. Notably, ESP cells generated endothelial cells that migrated into the mouse kidney parenchyma and formed mature blood vessels. This potential for in vivo angiogenesis and endometrial cell regeneration was more prominent in the ESP fraction than in the EMP fraction, as the latter mainly gave rise to stromal cells in vivo.Conclusions/significanceThese results indicate that putative endometrial stem cells are highly enriched in the ESP cells. These unique characteristics suggest that ESP cells might drive physiological endometrial regeneration and be involved in the pathogenesis of endometriosis.

Project description:Intratumoral heterogeneity is a major barrier against effective cancer therapy. Human malignant mesothelioma (HMM) that is closely associated with asbestos exposure is extremely heterogeneous in morphology and molecular phenotype. Contrast to genetic mutations, the role of epigenetic modifications in the generation and maintenance of heterogeneous populations in cancers remains largely undetermined. The present study was performed to investigate underlying molecular mechanisms for the emergence of intratumoral heterogeneity by identifying the global microRNA expression profile of distinct subpopulations of MS1 cell line, a HMM cell line. More aggressive cancer cells could be enriched by side population (SP) assay in HMM [20]. The sorted SP and NSP subpopulations were subjected to the microarray analysis of miRNA expression to investigate differentially altered miRNA genes defining tumor heterogeneity in HMM. Total RNAs were isolated from the sorted subpopulations of HMM cells, SP and non-SP fractions. The expression profile of miRNAs was evaluated using Affymetrix GeneChip miRNA Arrays. After data extraction and normalization, the microRNAs defining the cell subpopulations were determined using bioinformatics softwares. A total of 95 miRNAs including 42 up-regulated and 53 down-regulated were identified based on the criteria of 2 fold difference and a p-value < 0.05. Functional ontology of the dysregulated miRNAs revealed that a large number of target genes were categorized into the regulation of various cellular processes, including cell proliferation, programmed cell death, cell migration, cellular response to stress, and stem cell maintenance. The data show that microRNAs are significantly involved in the generation and maintenance of intratumoral heterogeneity and their regulation could be an effective strategy to eradicate a more aggressive cancer cell subpopulation. This is the first to report the profile of miRNA expression in CSCs in HMM by using side population assay assisted with flow cytometry. It will be valuable to understand the regulatory function of HMM CSC miRNAs in generation and maintenance of intratumoral heterogeneity.

Dataset Information

Reconstruction of endometrium from human endometrial side population cell lines.

Publications

Reconstruction of endometrium from human endometrial side population cell lines.

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