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. 2023 Jan 11;14:156. doi: 10.1038/s41467-022-35487-9

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© The Author(s) 2023

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Fig. 1 — a Schematic showing induction of quiescence and senescence in fibroblasts, conditioned media collection, and organoid culture in conditioned media. The figure was created with BioRender.com. b Representative images of organoids after 5 d of culture in conditioned media. Enlarged images (boxed area) are shown at the right. Scale bar, 200 μm. c Quantification of data from (b). The percentage of cystic organoids in organoids cultured in conditioned media is shown (mean ± s.d.; two-tailed t-test). Each dot represents a result from organoids established from a mouse, 3 biological replicates per experiment (n = 3). d H&E staining (left column) and immunostaining of paraffin-embedded organoids. Immunostaining for Ki67 (green) and phospho-Histone H3 (red) in intestinal organoids cultured in conditioned media and stained with Hoechst (blue). Scale bar, 50 μm. e Gene expression in organoids exposed to SCM determined by qPCR and represented as a ratio relative to organoids exposed to QCM (mean ± s.d.; n = 4; two-tailed t-test). f–m Numbers of cells expressing the listed markers in organoids exposed to QCM or SCM (mean ± SEM; n = 10; two-tailed t-test). n UMAP of pooled snRNA-seq data of nuclei from QCM and SCM exposed organoids colored by cell-type annotation (right). o UMAP showing contribution of nuclei from QCM and SCM exposed organoids labeled in blue and orange, respectively. p Relative contribution of cells from QCM and SCM exposed organoids to each cluster. Cells from QCM and SCM exposed organoids were labeled in blue and orange, respectively. q Representative images of organoids taken every 2 days starting on day 4. From day 0 to 4, organoids were cultured in quiescent conditioned media (left panel) and senescent conditioned media (right panel). On day 4, quiescent-conditioned media was changed to senescent-conditioned media (left bottom panel), whereas senescent-conditioned media was changed to quiescent-conditioned media (right bottom panel). In all conditions, media was changed every other day. Scale bars, 100 μm. See also Fig. S1. Source data are provided as a Source Data file.