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      Unravelling the Mystery of Stem/Progenitor Cells in Human Breast Milk

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          Abstract

          Background

          Mammary stem cells have been extensively studied as a system to delineate the pathogenesis and treatment of breast cancer. However, research on mammary stem cells requires tissue biopsies which limit the quantity of samples available. We have previously identified putative mammary stem cells in human breast milk, and here, we further characterised the cellular component of human breast milk.

          Methodology/Principal Findings

          We identified markers associated with haemopoietic, mesenchymal and neuro-epithelial lineages in the cellular component of human breast milk. We found 2.6±0.8% (mean±SEM) and 0.7±0.2% of the whole cell population (WCP) were found to be CD133+ and CD34+ respectively, 27.8±9.1% of the WCP to be positive for Stro-1 through flow-cytometry. Expressions of neuro-ectodermal stem cell markers such as nestin and cytokeratin 5 were found through reverse-transcription polymerase chain reaction (RT-PCR), and in 4.17±0.2% and 0.9±0.2% of the WCP on flow-cytometry. We also established the presence of a side-population (SP) (1.8±0.4% of WCP) as well as CD133+ cells (1.7±0.5% of the WCP). Characterisation of the sorted SP and non-SP, CD133+ and CD133- cells carried out showed enrichment of CD326 (EPCAM) in the SP cells (50.6±8.6 vs 18.1±6.0, P-value  = 0.02). However, culture in a wide range of in vitro conditions revealed the atypical behaviour of stem/progenitor cells in human breast milk; in that if they are present, they do not respond to established culture protocols of stem/progenitor cells.

          Conclusions/Significance

          The identification of primitive cell types within human breast milk may provide a non-invasive source of relevant mammary cells for a wide-range of applications; even the possibility of banking one's own stem cell for every breastfeeding woman.

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          Most cited references27

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          The prognostic role of a gene signature from tumorigenic breast-cancer cells.

          Breast cancers contain a minority population of cancer cells characterized by CD44 expression but low or undetectable levels of CD24 (CD44+CD24-/low) that have higher tumorigenic capacity than other subtypes of cancer cells. We compared the gene-expression profile of CD44+CD24-/low tumorigenic breast-cancer cells with that of normal breast epithelium. Differentially expressed genes were used to generate a 186-gene "invasiveness" gene signature (IGS), which was evaluated for its association with overall survival and metastasis-free survival in patients with breast cancer or other types of cancer. There was a significant association between the IGS and both overall and metastasis-free survival (P<0.001, for both) in patients with breast cancer, which was independent of established clinical and pathological variables. When combined with the prognostic criteria of the National Institutes of Health, the IGS was used to stratify patients with high-risk early breast cancer into prognostic categories (good or poor); among patients with a good prognosis, the 10-year rate of metastasis-free survival was 81%, and among those with a poor prognosis, it was 57%. The IGS was also associated with the prognosis in medulloblastoma (P=0.004), lung cancer (P=0.03), and prostate cancer (P=0.01). The prognostic power of the IGS was increased when combined with the wound-response (WR) signature. The IGS is strongly associated with metastasis-free survival and overall survival for four different types of tumors. This genetic signature of tumorigenic breast-cancer cells was even more strongly associated with clinical outcomes when combined with the WR signature in breast cancer. Copyright 2007 Massachusetts Medical Society.
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            Urine derived cells are a potential source for urological tissue reconstruction.

            Contemporary approaches to tissue engineering and cell therapy for urinary tract reconstruction require invasive tissue biopsies to obtain autologous cells. However, these procedures are associated with potential complications. We determined whether the cells present in urine have characteristics of normal bladder cells and investigated their potential uses for urological reconstructive procedures. A total of 55 urine samples were collected from 15 healthy individuals and 8 patients with vesicoureteral reflux. Urine derived cells were isolated, expanded and tested for progenitor and differentiated cell specific markers using flow cytometry, immunofluorescence and Western immunoblotting. The chromosomal stability of cultured urine derived cells was determined by karyotype analysis. Clones were successfully established from primary cultures of urine derived cells. Isolated cells showed 3 phenotypes, including fully differentiated, differentiating and progenitor-like cells. Some urine derived cells stained positive for the surface markers c-Kit, SSEA4, CD105, CD73, CD91, CD133 and CD44. Two to 7 cells per 100 ml urine were multipoint progenitors that could expand extensively in culture. Single progenitor cells had the ability to differentiate into the cell lineages expressing urothelial, smooth muscle, endothelial and interstitial cell markers. The expression of lineage markers was characterized by Western blot and immunofluorescence analysis. Urine derived cells also maintained a normal karyotype after serial culture. A subpopulation of cells isolated from urine had progenitor cell features and the potential to differentiate into several bladder cell lineages. Urine derived cells could serve as an alternative cell source for urinary tract tissue engineering and reconstruction.
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              A method for quantifying normal human mammary epithelial stem cells with in vivo regenerative ability.

              Previous studies have demonstrated that normal mouse mammary tissue contains a rare subset of mammary stem cells. We now describe a method for detecting an analogous subpopulation in normal human mammary tissue. Dissociated cells are suspended with fibroblasts in collagen gels, which are then implanted under the kidney capsule of hormone-treated immunodeficient mice. After 2-8 weeks, the gels contain bilayered mammary epithelial structures, including luminal and myoepithelial cells, their in vitro clonogenic progenitors and cells that produce similar structures in secondary transplants. The regenerated clonogenic progenitors provide an objective indicator of input mammary stem cell activity and allow the frequency and phenotype of these human mammary stem cells to be determined by limiting-dilution analysis. This new assay procedure sets the stage for investigations of mechanisms regulating normal human mammary stem cells (and possibly stem cells in other tissues) and their relationship to human cancer stem cell populations.
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                Author and article information

                Contributors
                Role: Editor
                Journal
                PLoS One
                plos
                plosone
                PLoS ONE
                Public Library of Science (San Francisco, USA )
                1932-6203
                2010
                28 December 2010
                : 5
                : 12
                Affiliations
                [1 ]Experimental Fetal Medicine Group, Department of Obstetrics and Gynaecology, Yong Loo Lin School of Medicine, National University Health System, Singapore, Singapore
                [2 ]School of Biomedical, Biomolecular and Chemical Sciences, The University of Western Australia, Crawley, Perth, Australia
                [3 ]Department of Reproductive Medicine, KK Women's and Children's Hospital, Singapore, Singapore
                [4 ]Cancer and Stem Cell Biology Program, Duke-NUS Graduate Medical School, Singapore, Singapore
                Istituto Dermopatico dell'Immacolata, Italy
                Author notes

                Conceived and designed the experiments: YF MC MDC JKYC. Performed the experiments: YF. Analyzed the data: YF YSC MC MDC JKYC. Contributed reagents/materials/analysis tools: YF YSC MC MDC JKYC. Wrote the paper: YF YSC MC MDC JKYC.

                [¤]

                Current address: Medela AG, Baar, Switzerland

                Article
                10-PONE-RA-21055R1
                10.1371/journal.pone.0014421
                3010984
                21203434
                7add6aa4-397e-4676-ae17-cda54b7a1793
                Fan et al. This is an open-access article distributed under the terms of the Creative Commons Attribution License, which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited.
                Page count
                Pages: 11
                Categories
                Research Article
                Cell Biology
                Developmental Biology/Stem Cells
                Obstetrics/Breast Feeding
                Oncology/Breast Cancer

                Uncategorized
                Uncategorized

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