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. 2023 Dec 1;133(23):e168146. doi: 10.1172/JCI168146

Figure 5. Yap inhibition hinders effect of matrix stiffness in EAC PDOs in vivo xenograft model.

Figure 5

(A) Schematic of in vivo transplantation experiment of EAC PDOs within NorHA hydrogels into mouse subcutaneous pockets and treatment with intraperitoneal injections of verteporfin or DMSO. Created with BioRender.com. (B) Histological (H&E) microcopy images and quantification of PDO (C) size (area) and (D) density within NorHA hydrogels at 28 days after encapsulation, in vivo transplantation, and treatment with verteporfin or DMSO. Data are represented as mean ± SEM. (C) n = at least 17 organoids analyzed across 10 samples per group. (D) n = at least 55 organoids analyzed across 10 samples per group. Kruskal-Wallis test with Dunn’s multiple-comparisons test showed significant differences between 1000 Pa+DMSO and 1000 Pa +verteporfin, 1000 Pa+DMSO and 100 Pa+DMSO and no significant differences between 100 Pa+DMSO and 100 Pa+verteporfin (P > 0.05). **P < 0.01; ***P < 0.001; ****P < 0.0001. Scale bar: 100 μm. (E) Fluorescence microcopy images of PDOs within NorHA hydrogels at 28 days after encapsulation, in vivo transplantation, and treatment with verteporfin or DMSO stained for Sox9, Yap, Ki67, and CD44. Scale bar: 100 μm. Original magnification, ×5 (insets). (F–I) Quantification of (F) percentage of nuclear Yap+ cells (%Yap+), (G) percentage of Sox9+ cells (%Sox9+), (H) percentage of Ki67+ cells (%Ki67+), and (I) CD44 fluorescence intensity per EAC PDO within NorHA hydrogels at 28 days after encapsulation, in vivo transplantation, and treatment with verteporfin or DMSO. Data are represented as mean ± SEM. (F–I) n = at least 6 organoids analyzed per group. Kruskal-Wallis test with Dunn’s multiple-comparisons test showed significant differences between 1000 Pa+DMSO and 1000 Pa+verteporfin, 1000 Pa+DMSO and 100 Pa+DMSO and no significant differences between 100 Pa+DMSO and 100 Pa+verteporfin (P > 0.05). *P < 0.05; ***P < 0.001; ****P < 0.0001. (A–G) Two independent experiments were performed, and data are presented for 1 of the experiments. Every independent experiment was performed with 2 gels per mouse and 5 mice per experimental group.