Department of Medicine, Division of Digestive Diseases, Emory University, Atlanta, GA 30322, USA.
Graduate Program in Genetics and Molecular Biology, Emory University, Atlanta, GA 30322, USA.
Development. 2024 Dec 1;151(23). doi: 10.1242/dev.202941. Epub 2024 Nov 29.
Progenitors and mature cells can maintain the intestinal epithelium by dedifferentiation and facultative intestinal stem cell (fISC) function when active ISCs (aISCs) are lost to damage. Here, we modeled fISC activation in mouse intestinal organoids with doxorubicin (DXR) treatment, a chemotherapeutic known to ablate Lgr5+ aISCs in vivo. Similar fISC gene activation was observed between organoids treated with low versus high DXR, despite significantly decreased survival at the higher dose. aISCs exhibited dose-dependent loss after DXR treatment but survived at doses compatible with organoid survival. We ablated residual aISCs after DXR treatment using a Lgr52A-DTR allele and observed that aISC survival of the initial genotoxic insult is required for organoid survival following DXR treatment. These results suggest that although typical fISC genes are activated by DXR-induced injury in organoids, functional stemness remains dependent on the aISC pool. Finally, we show that human intestinal organoids require higher doses of DXR to induce loss of survival and downregulation of LGR5. Our data establish a reproducible model of DXR-induced injury in intestinal organoids and reveal differences in in vitro responses to an established in vivo damage modality.
祖细胞和成熟细胞可以通过去分化和兼性肠干细胞（fISC）功能来维持肠道上皮，当活跃的 ISC（aISC）因损伤而丢失时。在这里，我们使用多柔比星（DXR）处理来模拟小鼠肠道类器官中的 fISC 激活，这种化疗药物已知可在体内消除 Lgr5+aISC。尽管在高剂量下存活率显著降低，但在低剂量和高剂量 DXR 处理的类器官中观察到类似的 fISC 基因激活。在 DXR 处理后，aISC 呈剂量依赖性丢失，但在与类器官存活相容的剂量下存活。我们使用 Lgr52A-DTR 等位基因消除 DXR 处理后的残留 aISC，并观察到 DXR 处理后，初始遗传毒性损伤中 aISC 的存活对于类器官存活是必需的。这些结果表明，尽管典型的 fISC 基因在类器官的 DXR 诱导损伤中被激活，但功能干性仍然依赖于 aISC 池。最后，我们表明人类肠道类器官需要更高剂量的 DXR 才能诱导存活丧失和 LGR5 的下调。我们的数据建立了一个可重现的 DXR 诱导的肠道类器官损伤模型，并揭示了对已建立的体内损伤模式的体外反应的差异。
