| In vivo Mouse Mammary Gland Formation
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Author:
Date:
2020-07-05
[Abstract] For years, the mammary gland serves as a perfect example to study the self-renew and differentiation of adult stem cells, and the regulatory mechanisms of these processes as well. To assess the function of given genes and/or other factors on stemness of mammary cells, several In vitro assays were developed, such as mammospheres formation assay, detection of stem cell markers by mRNA expression or flow cytometry and so on. However, the capacity of reconstruction of whole mount in the cleared fat pad of recipient female mice is a golden standard to estimate the stemness of the cells. Here we described a step-by-step protocol for in vivo mammary gland formation assay, including preparation of “cleared” recipients and mammary cells for implantation, the surgery process and ...
[摘要] [摘要 ] 多年来，乳腺一直是研究成体干细胞的自我更新和分化以及这些过程的调控机制的完美例证。为了评估给定的基因和/或其他因素对乳腺细胞的干性，有几个函数体外测定法开发出来，如微球体由mRNA表达形成试验，干细胞标志物的检测或流式细胞术等。然而，在雌性小鼠清除的脂肪垫中整个坐骨的重建能力是估计细胞干性的黄金标准。在这里，我们描述了体内的分步操作方案 乳腺形成测定，包括准备“清除”的受体和用于植入的乳腺细胞，手术过程以及如何评估实验结果。结合通过基因编辑和/或药物处理对乳腺细胞的操作，该方案在乳腺干细胞和乳腺发育的研究中可能非常有用。
[背景 ] 作为哺乳动物最典型的器官之一，乳腺（MG）是外分泌腺，负责泌乳。MG的发育受某些性激素的控制，这些激素的水平精确地调节了MG在不同发育阶段的结构，细胞组成和功能变化（Henigighausen and Robinson，2005）。许多遗传和环境因素都参与了乳腺干细胞的调控和MG的发育。为了研究这些因素的功能和机理，已经开发了几种方法，特别是用于评估乳腺细胞的干性。先前的研究表明，只有MG的基底细胞而非管腔细胞能够在受体雌性小鼠清除的脂肪垫中重建上皮树，这表明乳腺干细胞仅存在于基底谱系中（Van Keymeulen 等，2011）。 ）。后来，包括我们在内的许多研究发现了乳腺干细胞的几种标志物（Prater ...
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| Isolation and Primary Culture of Adult Human Adipose-derived Stromal/Stem Cells
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Author:
Date:
2017-03-05
[Abstract] Adipose-derived stromal/stem cells (ASCs) are multipotent cells that can be isolated from adipose tissue. Studies have shown that cells have the capacity to self-renew and differentiate into adipocyte, chondrocyte, myocyte, and osteoblast lineages. Thus, significant interest regarding their use for regenerative purposes to restore aging or damaged tissue has grown in recent decades. These cells have also been shown to immunomodulate the microenvironment and secrete abundant growth factors, which minimize inflammation and aid repair and regeneration. ASCs can be readily isolated from the stromal vascular fraction (SVF) of lipoaspirates. Given their ease of accessibility, bountiful source, and potential application in regenerative medicine and tissue engineering, there is growing interest ...
[摘要] 脂肪来源的基质/干细胞（ASCs）是可以从脂肪组织分离的多能细胞。研究表明，细胞具有自我更新和分化成脂肪细胞，软骨细胞，肌细胞和成骨细胞谱系的能力。因此，近几十年来，对再生用途恢复老化或损伤组织的兴趣越来越大。这些细胞也被证明可以免疫微环境并分泌丰富的生长因子，从而使炎症最小化并辅助修复和再生。 ASCs可以容易地从脂质体的基质血管分数（SVF）中分离出来。鉴于其易于获取，丰富的来源和在再生医学和组织工程中的潜在应用，对于ASC的表征和利用越来越感兴趣。该方案描述了从成人人类脂肪组织中分离的ASC以及用于培养维持的方法，包括扩增和低温保存。
背景 脂肪来源的基质/干细胞（ASCs）表现出干细胞领域的巨大潜力。根据造血干细胞移植的治疗奇迹，ASCs代表干细胞的未来，因为它们更容易获得源 - 脂肪组织。 ASCs自我更新和分化成各种组织谱系（包括脂肪细胞，软骨细胞，肌细胞和成骨细胞谱系）的能力允许恢复损伤的组织。另外，推测ASCs有可能在体外复制组织。器官将允许更容易获得新颖药物的评估，从而显着降低药物生产成本。然而，隔离，维护和冷冻保存过程中的不一致，禁止集体分析世界各地不同实验室的结果。用于分离和培养ASC的标准方案对于确保一致的数据分析是必要的。
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| Isolation and Culture of Human Adipose-derived Stem Cells from Subcutaneous and Visceral White Adipose Tissue Compartments
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Author:
Date:
2016-11-20
[Abstract] Human Adipose-derived Stem/Stromal Cells (ASCs) have been widely used in stem cell and obesity research, as well as clinical applications including cell-based therapies, tissue engineering and reconstruction. Compared with mesenchymal stem cells (MSCs) derived from other tissues such as umbilical cord and bone marrow, isolation of ASCs from human white adipose tissue (WAT) has great advantages due to its rich tissue source and simple surgical procedure. In this detailed protocol we describe a protocol to isolate and characterize ASCs from human WAT. Molecular characterization of isolated ASCs was performed through surface marker expression profiling using flow cytometry. Adipogenic capacity of the isolated ASCs was confirmed through inducing adipogenic differentiation and Oil Red O ...
[摘要] Human Adipose-derived Stem/Stromal Cells (ASCs) have been widely used in stem cell and obesity research, as well as clinical applications including cell-based therapies, tissue engineering and reconstruction. Compared with mesenchymal stem cells (MSCs) derived from other tissues such as umbilical cord and bone marrow, isolation of ASCs from human white adipose tissue (WAT) has great advantages due to its rich tissue source and simple surgical procedure. In this detailed protocol we describe a protocol to isolate and characterize ASCs from human WAT. Molecular characterization of isolated ASCs ...
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