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Manipulating the Avian Epiblast and Epiblast-Derived Stem Cells

Compared to eutherian mammals, birds retain a primitive form of epiblast development. Molecular studies of the avian epiblast can provide valuable insight for mammalian epiblast research. Here, we introduce several basic techniques in handling epiblast-stage embryos of the chick, ...

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Chicken Embryonic Stem Cells: Establishment and Characterization

Embryonic stem (ES) cells are unique models for investigating early development and cell differentiation. First identified in mouse and later in other mammals, these cells have also been isolated in avian species. Here, using chicken as a model, we describe a set of protocols allowing the isola ...

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Analysis of Molecular Markers for Staging Peri-gastrulating Bovine Embryos

Whole-mount in situ hybridization (WISH) is a method to visualize gene expression through hybridization of in vitro synthesized riboprobes to cellular mRNAs. WISH has been used in developmental biology for decades and was adapted to many species, especially for model organisms of devel ...

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Detection In Vitro and Quantitative Estimation of Artificial Microterritories Which Promote Osteogenic Differentiation and Maturation of Stromal Stem

Extracellular matrix can regulate multipotent mesenchymal stromal cells (MMSC) differentiation, with potential applications for tissue engineering. A relief of mineralized bone takes essential effect on MMSC fate. Nevertheless, delicate structure and a hierarchy of nich ...

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Primary Marrow-Derived Stromal Cells: Isolation and Manipulation

Marrow stromal cells (MSCs) are relatively rare cells difficult to visualize in marrow biopsies or detect in aspirated marrow. Under specific conditions MSC can be expanded in vitro and the population can give rise to several mesenchymal lineages. “MSC” also refers to mesenchymal stem cells ...

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Isolation and Enrichment of Stro-1 Immunoselected Mesenchymal Stem Cells from Adult Human Bone Marrow

The availability of mesenchymal stem cells (MSCs) or skeletal stem cells (SSCs) is vital to many of the tissue engineering strategies currently being developed for repairing bone and cartilage. One difficulty with using this cell population is that SSCs represent only a small fraction of the ...

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Isolation of Embryonic Hematopoietic Niche Cells by Flow Cytometry and Laser Capture Microdissection

Hematopoietic stem cells (HSCs) can differentiate into several types of hematopoietic cells, such as erythrocytes, megakaryocytes, lymphocytes, neutrophils, or macrophages, and also undergo self-renewal to sustain hematopoiesis throughout an organism’s lifetime. HSCs e ...

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In Vitro Construction of 2D and 3D Simulations of the Murine Hematopoietic Niche

Hematopoietic stem cells (HSC) undergo multilineage differentiation or self-renewal to maintain normal hematopoiesis and to sustain the size of the HSC pool throughout life. These processes are determined by a complex interplay of molecular signals between HSC and other cellular c ...

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Morphometric Evaluation of the Spermatogonial Stem Cell Distribution and Niche in Vertebrates

Morphometry is a classical quantitative method often used in biology to provide a data basis for functional interpretations/interactions of a particular organ or system. Herein we took advantage of this valuable approach to evaluate the spermatogonial stem cell niche using the horse t ...

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Visualization of Adult Stem Cells Within Their Niches Using the Drosophila Germline as a Model System

The germaria of the fruit fly Drosophila melanogaster present an excellent model to study germline stem cell–niche interactions. Two to three adult stem cells are surrounded by a number of somatic cells that form the niche. Here we describe how Drosophilae germaria can be dissected and specif ...

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Genetic, Immunofluorescence Labeling, and In Situ Hybridization Techniques in Identification of Stem Cells in Male and Female Germline Niches

Stem cells have an enormous capacity of self-renewal, as well as the ability to differentiate into specialized cell types. Proper control of these two properties of stem cells is crucial for animal development, growth control, and reproduction. Germline stem cells (GSCs) are a self-renewing ...

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Isolation and Propagation of Colon Cancer Stem Cells

The design of tissue culture conditions that faithfully reproduce the characteristics of cells in their native environment remains one of the main challenges of cancer stem cell (CSC) biology. Here we describe a detailed methodology for the isolation and expansion of both human colon CSCs a ...

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Serial Orthotopic Transplantation of Epithelial Tumors in Single-Cell Suspension

Orthotopic transplantation of tumor tissue into recipient mice has long been established to study the role of the microenvironment in tumorigenesis and metastasis. Many of these transplantation assays involve the surgical implantation of an undissociated piece of tumor tissue. H ...

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Immunostaining of Germline Stem Cells and the Niche in Drosophila Ovaries

Stem cells have the ability to switch between proliferative (self-renewal) and differentiation modes. The Drosophila germarium is a well-established in vivo model for the study of communication between stem cells and their niche. One commonly used technique for such study is immunost ...

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Identification and Separation of Normal Hematopoietic Stem Cells and Leukemia Stem Cells from Patients with Acute Myeloid Leukemia

Mounting evidences indicate that leukemic cells in patients with acute myeloid leukemia (AML) are derived from leukemia stem cells (LSC). In analogy to normal hematopoietic stem cells (HSC), LSC remain mostly dormant and are hence resistant to conventional chemotherapy. Residual, ph ...

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Detection and Isolation of Human Disseminated Tumor Cells in the Murine Bone Marrow Stem Cell Niche

The presence of disseminated tumor cells (DTCs) in the bone marrow is associated with poor prognosis of cancer patients. However, little is known about the biology of DTCs due to lack of relevant animal models. Here, we describe the methods for detecting and isolating human DTCs from the murine bone m ...

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Hypoxia and Visualization of the Stem Cell Niche

It is widely accepted that mammalian stem cells reside in a specialized cellular and a cellular microenvironment called the niche. The niche contrary to other tissues is characterized by a low partial Oxygen pressure (ppO2). This microenvironment protects stem cells from deleterious e ...

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Isolation of c-Kit+ Human Amniotic Fluid Stem Cells from Second Trimester

Amniotic fluid-derived stem (AFS) cells have been described as an appealing source of stem cells because of their (1) fetal, non-embryonic origin, (2) easy access during pregnancy overcoming the ethical issues related both to the use of human embryonic cells and to the postnatal tissue biopsy w ...

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Modulation of the Host Skeletal Muscle Niche for Donor Satellite Cell Grafting

Skeletal muscle tissue has a remarkable capability of regenerating in pathological conditions or after injury. The principal muscle stem cells, satellite cells, are responsible for this prompt and efficient process. Normally quiescent in their niches underneath the basal lamina of ...

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Human and Murine Skeletal Muscle Reserve Cells

Study of stem cell phenotype and functions requires their proper isolation. Stem cells isolated from skeletal muscle are a useful tool to explore molecular pathways involved in the regulation of myogenesis. Among progenitor cells, a subset of cells, called reserve cells, has been identif ...

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