Fig. 5 | Nature Communications

Fig. 5

From: Anti-LRP5/6 VHHs promote differentiation of Wnt-hypersensitive intestinal stem cells

Fig. 5

Anti-LRP5/6 VHHs drive collective differentiation of R/Z-mutant tumor organoids. a Anti-LRP5/6P3 VHHs block tumorigenic R/Z mutant organoid growth. Organoids were cultured in EN and treated with 10 μM of the indicated anti-LRP5/6P3 VHH for 4 d (Scale bar, 400 μm). Red asterisks indicate cell death; green arrows indicate organoids showing villi and crypts; orange arrows indicate organoids showing a mixed phenotype of cell death and villi crypts structures. b Anti-LRP5/6P3 VHHs strongly diminish cell viability of tumorigenic R/Z mutant organoids. Organoids were cultured in EN and treated with 10 μM of the indicated anti-LRP5/6P3 VHHs for 4 d. Graph represents relative cell viability normalized to PBS treatment. Mean ± s.d. (n = 3) are plotted. c Schematic representation of the cell types present in the intestine and the role of Wnt signaling in maintaining intestinal physiology. d qRT-PCR showing anti-LRP5/6P3 VHH treatment (3 d) strongly inhibits the expression of Wnt target genes, stem cell-associated genes and Paneth cell markers while inducing a transcriptional program for differentiation in tumorigenic R/Z mutant organoids. Graph represents fold change in gene expression as compared to PBS treatment ( = 1). Mean ± s.d. (n = 3) are plotted. e Anti-LRP5/6P3 VHH treatment (3 d) attenuates proliferation as shown by a decrease in Ki67 staining (left). Anti-LRP5/6P3 VHH treatment decreases the number of Paneth cells (Lys) while increasing the number of differentiated enteroendocrine cells (ChgA) (right). Phalloidin staining was used to mark the organoids (Scale bar, 10 μm)

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