Human fallopian tube epithelial cells exhibit stemness features, self-renewal capacity, and Wnt-related organoid formation
Human fallopian tube epithelial cells exhibit stemness features, self-renewal capacity, and Wnt-related organoid formation
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DOI：
10.1186/s12929-019-0602-1
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发表时间：
2020-02-08
影响因子：
11
通讯作者：
Ding, Dah-Ching
中科院分区：
文献类型：
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作者：
Chang, Yu-Hsun;Chu, Tang-Yuan;Ding, Dah-Ching
输卵管上皮细胞（FTEC）被认为是高级别浆液性卵巢癌（HGSOC）的起源。对FTEC的干性或启动特征的认识是不够的。在此之前，我们已经对FTEC的干细胞标记物进行了表征，本研究旨在进一步表征FTEC的克隆原性和球形特征。方法成功地从人输卵管上皮层中获得输卵管内皮细胞。我们观察了它们的形态、增殖率、倍增时间和克隆生长情况。第3代观察明胶包被培养、悬浮培养和基质培养上的球形成情况，并检测LGR5、SSEA3、SSEA4等茎干标记的表达情况。此外，我们还检测了FTEC、输卵管间质细胞（FTMSC）和内皮细胞（HUVEC）共培养的组织重构类器官。结果FTEC细胞呈长方体形态，并能保持恒定增殖速率达9代（P9）。FTEC可以从单个细胞中增殖，克隆效率为4%。流式细胞术显示正常干细胞标志物（SSEA3、SSEA4和LGR5）和肿瘤干细胞标志物（CD24、CD44、CD117、ROR1和CD133）的表达。在低附着皿中培养时，FTEC形成球形和菌落。在Matrigel的作用下，茎干性和菌落形成活性明显增强。在与FTMSC和HUVEC共培养时，FTEC可以形成可被Wnt抑制剂DKK1阻断的类器官。添加DKK1后，LGR5和FOXJ1的表达也降低。结论在人类ftec中大量存在依赖于Wnt信号的干细胞，这些干细胞能够有效地形成集落、球体和类器官。我们也首次报道了从组织中重组细胞系生成类器官。这可能为研究输卵管上皮的再生和恶性转化提供一个新的模型。
Background Fallopian tube epithelial cells (FTEC) were thought to be the origin of high-grade serous ovarian carcinoma (HGSOC). Knowledge of the stemness or initiating characteristics of FTEC is insufficient. Previously, we have characterized the stemness cell marker of FTEC, this study aims to further characterize the clonogenicity and spheroid features of FTEC. Methods We successfully derived FTECs from the epithelial layer of the human fallopian tubes. We examined the morphology, proliferation rate, doubling time, and clonal growth of them. At passage 3, the sphere formations on gelatin-coated culture, suspension culture, and matrigel culture were observed, and the expression of LGR5, SSEA3, SSEA4, and other stemness markers was examined. Furthermore, tissue-reconstituted organoids from coculture of FTEC, fallopian stromal cells (FTMSC) and endothelial cells (HUVEC) were examined. Results FTEC exhibited cuboidal cell morphology and maintained at a constant proliferation rate for up to nine passages (P9). FTEC could proliferate from a single cell with a clonogenic efficiency of 4%. Flow cytometry revealed expressions of normal stem cell markers (SSEA3, SSEA4, and LGR5) and cancer stem cell markers (CD24, CD44, CD117, ROR1, and CD133). FTEC formed spheres and colonies when cultured on low attach dish. In the presence of Matrigel, the stemness and colony formation activity were much enhanced. In co-culturing with FTMSC and HUVEC, FTEC could form organoids that could be blocked by Wnt inhibitor DKK1. Expressions of LGR5 and FOXJ1 expression were also decreased by adding DKK1. Conclusion We demonstrated abundantly presence of stem cells in human FTECs which are efficient in forming colonies, spheres and organoids, relying on Wnt signaling. We also reported for the first time the generation of organoid from reconstitutied cell lineages in the tissue. This may provide a new model for studying the regneration and malignant transformation of the tubal epithelium.