WRN conditioned media is sufficient for in vitro propagation of intestinal organoids from large farm and small companion animals.
WRN conditioned media is sufficient for in vitro propagation of intestinal organoids from large farm and small companion animals.
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DOI：
10.1242/bio.021717
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发表时间：
2017-05-15
期刊：
影响因子：
2.4
通讯作者：
Behnke MS
中科院分区：
文献类型：
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作者：
Powell RH;Behnke MS
近年来，在连续繁殖源自肠隐窝的类器官的能力方面已经有了显著的发展。这些进展已应用于小鼠和人类样本，为胃肠道组织发育和疾病提供模型。我们采用这些方法从各种大型农场和小型伴侣（LF/SC）动物，包括猫，狗，牛，马，猪，羊和鸡的肠道类器官（肠）的繁殖。我们发现，LF/SC类肠细胞在含有信号传导因子Wnt 3a、R-spondin-3和头蛋白（WRN）的L-WRN条件培养基中繁殖和扩增。每个种属均实现了多次成功分离，LF/SC类肠体的生长保持在高传代次数。LF/SC肠样组织表达隐窝干细胞标记物LGR 5和低水平的间充质标记物Vim。用EdU标记还显示了在标记隐窝样区域的肠内的细胞增殖的不同区域。生长和维持LF/SC肠样细胞系的能力为研究胃肠道发育生物学提供了额外的模型，并为研究肠道细胞与具有医学重要性的人畜共患病肠道病原体之间的宿主-病原体相互作用提供了平台。总结：用于繁殖隐窝衍生的小鼠和人肠上皮细胞的培养方法适用于大型农场和小型伴侣动物，证明了隐窝信号微环境的保护。
Recent years have seen significant developments in the ability to continuously propagate organoids derived from intestinal crypts. These advancements have been applied to mouse and human samples providing models for gastrointestinal tissue development and disease. We adapt these methods for the propagation of intestinal organoids (enteroids) from various large farm and small companion (LF/SC) animals, including cat, dog, cow, horse, pig, sheep and chicken. We show that LF/SC enteroids propagate and expand in L-WRN conditioned media containing signaling factors Wnt3a, R-spondin-3, and Noggin (WRN). Multiple successful isolations were achieved for each species, and the growth of LF/SC enteroids was maintained to high passage number. LF/SC enteroids expressed crypt stem cell marker LGR5 and low levels of mesenchymal marker VIM. Labeling with EdU also showed distinct regions of cell proliferation within the enteroids marking crypt-like regions. The ability to grow and maintain LF/SC enteroid cell lines provides additional models for the study of gastrointestinal developmental biology as well as platforms for the study of host-pathogen interactions between intestinal cells and zoonotic enteric pathogens of medical importance. Summary: Culturing methods for the propagation of crypt-derived mouse and human enteroids are applicable to large farm and small companion animals, demonstrating conservation of the crypt signaling microenvironment.