Single Lgr5- or Lgr6-expressing taste stem/progenitor cells generate taste bud cells ex vivo
Single Lgr5- or Lgr6-expressing taste stem/progenitor cells generate taste bud cells ex vivo
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DOI：
10.1073/pnas.1409064111
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发表时间：
2014-11-18
影响因子：
11.1
通讯作者：
Jiang, Peihua
中科院分区：
文献类型：
--
作者：
Ren, Wenwen;Lewandowski, Brian C.;Jiang, Peihua
富含亮氨酸重复的G蛋白偶联受体5（Lgr 5）及其同源物（例如，lgr 6）标记多种组织中的成体干细胞。最近，我们和其他人已经表明，Lgr 5标志着舌后部的成年味觉干/祖细胞。然而，表达Lgr 5（Lgr 5（+））的细胞的再生潜力和在舌前部再生味觉组织的成体味觉干/祖细胞的身份仍然难以捉摸。在目前的工作中，我们描述了一种培养系统，其中来自味觉组织的单个分离的Lgr 5（+）或Lgr 6（+）细胞可以产生连续扩展的3D结构（“类器官”）。这些味觉类器官内的许多细胞是周期性的，并且对增殖细胞标志物细胞角蛋白K5和Sox 2呈阳性，并且掺入5-溴-2 '-脱氧尿苷。重要的是，成熟的味觉感受器细胞，表达味觉蛋白，碳酸酐酶4，味觉感受器类型1成员3，核苷三磷酸二磷酸水解酶-2，或细胞角蛋白K8存在于味觉类器官。使用钙成像测定，我们发现从分离的Lgr 5（+）细胞衍生的味觉类器官中生长出来的细胞是功能性的，并且以剂量依赖性方式对促味剂产生反应。遗传谱系追踪显示Lgr 6（+）细胞在舌前、后区的味乳头中均能分化为味蕾细胞。RT-PCR结果表明Lgr 5和Lgr 6可能在味觉干/祖细胞的前后方向上标记同一个亚群。总之，我们的数据表明，功能性味觉细胞可以从单个Lgr 5（+）或Lgr 6（+）细胞离体产生，验证了该模型用于味觉细胞产生研究的用途。
Leucine-rich repeat-containing G protein-coupled receptor 5 (Lgr5) and its homologs (e.g., Lgr6) mark adult stem cells in multiple tissues. Recently, we and others have shown that Lgr5 marks adult taste stem/progenitor cells in posterior tongue. However, the regenerative potential of Lgr5-expressing (Lgr5(+)) cells and the identity of adult taste stem/progenitor cells that regenerate taste tissue in anterior tongue remain elusive. In the present work, we describe a culture system in which single isolated Lgr5(+) or Lgr6(+) cells from taste tissue can generate continuously expanding 3D structures ("organoids"). Many cells within these taste organoids were cycling and positive for proliferative cell markers, cytokeratin K5 and Sox2, and incorporated 5-bromo-2'-deoxyuridine. Importantly, mature taste receptor cells that express gustducin, carbonic anhydrase 4, taste receptor type 1 member 3, nucleoside triphosphate diphosphohydrolase-2, or cytokeratin K8 were present in the taste organoids. Using calcium imaging assays, we found that cells grown out from taste organoids derived from isolated Lgr5(+) cells were functional and responded to tastants in a dose-dependent manner. Genetic lineage tracing showed that Lgr6(+) cells gave rise to taste bud cells in taste papillae in both anterior and posterior tongue. RT-PCR data demonstrated that Lgr5 and Lgr6 may mark the same subset of taste stem/progenitor cells both anteriorly and posteriorly. Together, our data demonstrate that functional taste cells can be generated ex vivo from single Lgr5(+) or Lgr6(+) cells, validating the use of this model for the study of taste cell generation.