Fig. 5: BMP signaling has a strong impact on gland turnover through the feed-forward loop and regulates R-spondin3 expression in myofibroblasts.

a qPCR for Bmp2 from organoids cultured in indicated conditions (n = 3). b qPCR for indicated genes and forming capacity from organoids exposed to different concentrations of rBMP2 (n = 3). c Representative images of organoids that re-grew in the full medium after exposure to the indicated rBMP2 concentrations. d qPCR for indicated genes and forming capacity from organoids grown without noggin, exposed to different concentrations of rBMP2 (n = 3). e Schematic representation: organoids were grown without noggin and treated with 5 ng/ml rBMP2 (blue) for 4 days at passage 0. Organoids were washed and passaged and either kept in full medium with noggin (dark blue) or without noggin (blue). f Organoid forming capacity at different passages from organoids treated (blue) or untreated (yellow) with rBMP. Organoids exposed to full medium with noggin (dark blue) recovered their forming capacity, while organoids without noggin progressively lost forming capacity (n = 2). g qPCR from the organoids from e at passage 2: Bmp2 expression triggered by rBMP is not observed in organoids that were re-grown in full medium, while in organoids grown without noggin, endogenous Bmp2 expression is strongly upregulated (n = 4). h Immunofluorescence image of myofibroblasts from Myh11CreErt2/Rosa26-tdTomato mice cultured in 2D. Scale bar: 100 µm. i qPCR for Id1 and Rspo3 from myofibroblasts cultured with noggin and BMP2 (n = 3). j qPCR for Id1 and Rspo3 from myofibroblasts cultured with different BMP2 concentrations (n = 2). k Immunofluorescence images of EdU proliferation assay performed with myofibroblasts untreated and treated with BMP2. Scale bar: 100 µm. l Quantification of the proportion of EdU positive cells (n = 3). Images are representative of at least three biological replicates. Scale bar: 250 µm, except where indicated. Data are presented as mean ± SEM. Statistical analyses were performed using Student’s t-test (two-tailed) for (g, i, l); using one-way ANOVA, followed by Tukey’s multiple comparisons test for (a). Source data are provided as a Source Data file.