Powell Robin H, Behnke Michael S
Department of Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA 70803, USA.
Department of Pathobiological Sciences, School of Veterinary Medicine, Louisiana State University, Baton Rouge, LA 70803, USA
Biol Open. 2017 May 15;6(5):698-705. doi: 10.1242/bio.021717.
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.
近年来，源自肠隐窝的类器官连续传代培养能力取得了重大进展。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了模型。我们将这些方法应用于从各种大型农场动物和小型伴侣动物（LF/SC），包括猫、狗、牛、马、猪、羊和鸡中培养肠道类器官（肠类器官）。我们发现，LF/SC肠类器官在含有信号因子Wnt3a、R-spondin-3和Noggin（WRN）的L-WRN条件培养基中能够增殖和扩增。每个物种都实现了多次成功分离，并且LF/SC肠类器官的生长维持到了高传代次数。LF/SC肠类器官表达隐窝干细胞标志物LGR5和低水平的间充质标志物VIM。用EdU标记也显示出肠类器官内标记隐窝样区域的不同细胞增殖区域。培养和维持LF/SC肠类细胞系的能力为胃肠道发育生物学研究提供了额外的模型，也为研究肠道细胞与具有医学重要性的人畜共患肠道病原体之间的宿主-病原体相互作用提供了平台。
