Department of Internal Medicine, Division of Gastroenterology, University of Michigan Medical School, Ann Arbor, MI 48109, USA.
Department of Pathology, University of Michigan, Ann Arbor, MI 48109, USA.
Development. 2018 Mar 14;145(6):dev153049. doi: 10.1242/dev.153049.
The intestine is maintained by stem cells located at the base of crypts and distinguished by the expression of LGR5. Genetically engineered mouse models have provided a wealth of information about intestinal stem cells, whereas less is known about human intestinal stem cells owing to difficulty detecting and isolating these cells. We established an organoid repository from patient-derived adenomas, adenocarcinomas and normal colon, which we analyzed for variants in 71 colorectal cancer (CRC)-associated genes. Normal and neoplastic colon tissue organoids were analyzed by immunohistochemistry and fluorescent-activated cell sorting for LGR5. LGR5-positive cells were isolated from four adenoma organoid lines and were subjected to RNA sequencing. We found that LGR5 expression in the epithelium and stroma was associated with tumor stage, and by integrating functional experiments with LGR5-sorted cell RNA sequencing data from adenoma and normal organoids, we found correlations between LGR5 and CRC-specific genes, including dickkopf WNT signaling pathway inhibitor 4 () and SPARC-related modular calcium binding 2 (). Collectively, this work provides resources, methods and new markers to isolate and study stem cells in human tissue homeostasis and carcinogenesis.
肠道由位于隐窝底部的干细胞维持，这些干细胞的特征是表达 LGR5。基因工程小鼠模型为肠道干细胞提供了丰富的信息，而由于难以检测和分离这些细胞，人们对人类肠道干细胞的了解较少。我们从患者来源的腺瘤、腺癌和正常结肠建立了类器官库，并对 71 个结直肠癌（CRC）相关基因的变体进行了分析。对正常和肿瘤结肠组织类器官进行了 LGR5 的免疫组织化学和荧光激活细胞分选分析。从四个腺瘤类器官系中分离出 LGR5 阳性细胞，并进行 RNA 测序。我们发现上皮和基质中的 LGR5 表达与肿瘤分期相关，通过将功能实验与腺瘤和正常类器官的 LGR5 分选细胞 RNA 测序数据进行整合，我们发现 LGR5 与 CRC 特异性基因之间存在相关性，包括 dickkopf WNT 信号通路抑制剂 4（）和 SPARC 相关模块化钙结合蛋白 2（）。总之，这项工作提供了资源、方法和新的标记物，以分离和研究人类组织稳态和癌变中的干细胞。
