Isolation, Culture, and Differentiation of Mammary Epithelial Stem/Progenitor Cells from Fresh or Ex Vivo Cultured Human Breast Tissue.
Isolation, Culture, and Differentiation of Mammary Epithelial Stem/Progenitor Cells from Fresh or Ex Vivo Cultured Human Breast Tissue.
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DOI：
10.1002/cpcb.65
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发表时间：
2019-03-01
影响因子：
--
通讯作者：
Sun, Lu-Zhe
中科院分区：
文献类型：
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作者：
Chen, Guang;Bouamar, Hakim;Sun, Lu-Zhe
体内移植是表征干/祖细胞自我更新、组织再生和肿瘤发生的金标准方法。该方法需要代表给定组织的一小部分的干细胞的富集群体。富集的干/祖细胞群增加了植入的可能性并减少了体内移植所需的受体动物的数量。乳腺上皮干细胞和祖细胞形成乳腺球的方法已被广泛采用，用于富集干细胞/祖细胞，使研究人员能够研究遗传和表观遗传特性，与其他细胞类型的相互作用，以及分化和致癌转化。然而，乳腺球的产生是复杂的，涉及许多步骤并需要特殊的技能。在这里，我们描述了一个详细的mammosphere协议，包括人原代乳腺上皮干/祖细胞的分离和培养，以及它们在3D类器官培养中的分化和传代。我们还描述了一种用于临床治疗测定的新鲜人乳腺组织的离体培养方案。分步说明详细介绍了通过干/祖细胞生成的3D类器官的传代进行的组织处理，可用于评估乳腺干/祖细胞的性质、功能和肿瘤转化。John Wiley & Sons，Inc.
In vivo transplantation is the gold standard method for characterization of stem/progenitor cell self-renewal, tissue regeneration, and tumorigenesis. The method requires an enriched population of stem cells that represent a small fraction of a given tissue. An enriched population of stem/progenitor cells increases the likelihood of engraftment and reduces the number of recipient animals needed for in vivo transplantation. Methods for mammosphere formation by mammary epithelial stem and progenitor cells have been widely adopted for enriching stem/progenitor cells, allowing researchers to study genetic and epigenetic properties, interaction with other cell types, and differentiation and oncogenic transformation. The generation of mammospheres is complex, however, involving many steps and requiring particular skill. Here we describe a detailed mammosphere protocol, including isolation and culture of human primary mammary epithelial stem/progenitor cells and their differentiation and passage in 3D organoid culture. We also describe a protocol for ex vivo culture of fresh human breast tissue for use in assays of clinical treatment. Step-by-step instructions detail tissue handling through passage of the stem/progenitor cell-generated 3D organoids, which can be used to assess the properties, function, and neoplastic transformation of mammary stem/progenitor cells. © 2018 by John Wiley & Sons, Inc.