Enhanced Utilization of Induced Pluripotent Stem Cell-Derived Human Intestinal Organoids Using Microengineered Chips.
Enhanced Utilization of Induced Pluripotent Stem Cell-Derived Human Intestinal Organoids Using Microengineered Chips.
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DOI：
10.1016/j.jcmgh.2017.12.008
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发表时间：
2018
影响因子：
7.2
通讯作者：
Barrett RJ
中科院分区：
文献类型：
--
作者：
Workman MJ;Gleeson JP;Troisi EJ;Estrada HQ;Kerns SJ;Hinojosa CD;Hamilton GA;Targan SR;Svendsen CN;Barrett RJ
来源于诱导多能干细胞的人类肠道类器官具有阐明肠道上皮在健康和疾病中的作用的巨大潜力，但难以直接测定这些复杂的结构。这项研究试图通过从诱导多能干细胞衍生的人类肠道类器官中获得上皮细胞并将其纳入小型微工程芯片中，使这项技术更适合研究。然后，我们研究了芯片内的这些细胞是否被极化，是否具有4种主要的肠上皮亚型，以及是否对外源性刺激有生物学反应。从人肠类器官中阳性选择上皮细胞，并将其掺入芯片中。连续介质流的影响进行了检查。免疫细胞化学和原位杂交结果表明，上皮细胞呈极化状态，具有肠上皮细胞的主要亚型。为了评估掺入的细胞是否具有生物学响应性，使用Western印迹分析和定量聚合酶链反应来评估干扰素（IFN）-γ的作用，并使用异硫氰酸荧光素-葡聚糖4 kDa渗透来评估IFN-γ和肿瘤坏死因子-α对屏障功能的作用。确定了最佳细胞接种密度和流速。连续给予血流导致形成极化的肠皱襞，其含有潘氏细胞、杯状细胞、肠上皮细胞和肠内分泌细胞沿着以及转运扩增干细胞和LGR 5+干细胞。IFN-γ给药1小时导致STAT 1磷酸化，而暴露3天导致IFN-γ相关基因显著上调。给予IFN-γ和肿瘤坏死因子-α 3天导致肠通透性增加。我们证明了肠芯片是极化的，包含所有的肠上皮亚型，并对外源性刺激有生物反应。这代表了一个更适合使用类器官技术的平台，并将高度适用于个性化医疗和广泛的胃肠道疾病。
Human intestinal organoids derived from induced pluripotent stem cells have tremendous potential to elucidate the intestinal epithelium’s role in health and disease, but it is difficult to directly assay these complex structures. This study sought to make this technology more amenable for study by obtaining epithelial cells from induced pluripotent stem cell–derived human intestinal organoids and incorporating them into small microengineered Chips. We then investigated if these cells within the Chip were polarized, had the 4 major intestinal epithelial subtypes, and were biologically responsive to exogenous stimuli. Epithelial cells were positively selected from human intestinal organoids and were incorporated into the Chip. The effect of continuous media flow was examined. Immunocytochemistry and in situ hybridization were used to demonstrate that the epithelial cells were polarized and possessed the major intestinal epithelial subtypes. To assess if the incorporated cells were biologically responsive, Western blot analysis and quantitative polymerase chain reaction were used to assess the effects of interferon (IFN)-γ, and fluorescein isothiocyanate–dextran 4 kDa permeation was used to assess the effects of IFN-γ and tumor necrosis factor-α on barrier function. The optimal cell seeding density and flow rate were established. The continuous administration of flow resulted in the formation of polarized intestinal folds that contained Paneth cells, goblet cells, enterocytes, and enteroendocrine cells along with transit-amplifying and LGR5+ stem cells. Administration of IFN-γ for 1 hour resulted in the phosphorylation of STAT1, whereas exposure for 3 days resulted in a significant upregulation of IFN-γ related genes. Administration of IFN-γ and tumor necrosis factor-α for 3 days resulted in an increase in intestinal permeability. We demonstrate that the Intestine-Chip is polarized, contains all the intestinal epithelial subtypes, and is biologically responsive to exogenous stimuli. This represents a more amenable platform to use organoid technology and will be highly applicable to personalized medicine and a wide range of gastrointestinal conditions.
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影响因子：
24.5
作者：
VanDussen KL;Marinshaw JM;Shaikh N;Miyoshi H;Moon C;Tarr PI;Ciorba MA;Stappenbeck TS
通讯作者：
Stappenbeck TS
影响因子：
2.5
作者：
Ciorba MA
通讯作者：
Ciorba MA
DOI：
10.1126/science.1188302
发表时间：
2010-06-25
期刊：
Science (New York, N.Y.)
影响因子：
--
作者：
Huh D;Matthews BD;Mammoto A;Montoya-Zavala M;Hsin HY;Ingber DE
通讯作者：
Ingber DE
影响因子：
14.8
作者：
Huh, Dongeun;Kim, Hyun Jung;Ingber, Donald E.
通讯作者：
Ingber, Donald E.
影响因子：
46.9
作者：
Bhatia, Sangeeta N.;Ingber, Donald E.
通讯作者：
Ingber, Donald E.