Fig. 7: L-arginine supplementation protects the gut from 5-FU and TNF-α induced intestinal epithelial damage in a Wnt2b-dependent manner.

a–c Mice were treated with normal water or 7 mg/ml L-arginine in their drinking water for 14 days and then injected with 5-FU (300 mg/kg) or PBS as a control once a day for 5 days. In Wnt2b antibody neutralization experiment, every mouse were given 300 μg Wnt2b neutralizing antibody once on the first day of 5-FU injection. a Pathology of the small intestine by H&E staining. Scale bar, 200 μm. b FCM analysis of live Lgr5^hi ISC frequency in SI crypts, n = 3. c SI organoid frequency of crypts from mice in different groups. Representative images of crypt culture from each group are shown, n = 6. Scale bar, 50 μm. d–e SI organoids-CD90⁺ stromal cells co-culture model was cultured with ENR-medium or ENR-medium supplemented L-arginine (1 mM), TNF-α (100 ng/ml), WNT2B protein (100 ng/ml), Wnt2b neutralizing antibody (1 μg/ml) and their combinations for 3 days. d The light microscope observation of the co-culture model treated with L-arginine, TNF-α, and their combinations. Scale bar, 200 μm. The number of total organoids and disrupted organoids with altered morphology per well were counted, n = 6. e The light microscope observation of co-culture model treated with L-arginine, TNF-α, WNT2B protein, Wnt2b neutralizing antibody and their combinations. Scale bar, 200 μm. The number of total organoids and disrupted organoids with altered morphology per well were counted, n = 6. Data are the mean ± SD; comparisons performed with t-tests (two groups) or analysis of variance (ANOVA) (multiple groups). *P < 0.05, **P < 0.01, ***P < 0.001. Results are representative of two or three independent experiments.