Fig. 5
L-citrulline confers protection of intestinal epithelial cells against ionizing radiation. (A) Veen diagram showing unique and shared metabolites in feces and colon tissue. (B) Table showing common metabolites in feces and colon tissue. (C) Correlation network of microbes and metabolites from arginine biosynthesis, which included L-citrulline. (D) The STA Sankey Network for arginine biosynthesis. Aserisks (*) indicate statistically significant correlations with metabolites. The pink/blue color of nodes indicate up/downregulation. The red/green bands indicate the positive/negative correlations with metabolites. (E)-(H) The effects of L-citrulline treatment on cell viability and lactate dehydrogenase (LDH) release of intestinal cells (HIEC and IEC-6). *P < 0.05, **P < 0.01, compared with the control group. (I) A schematic timeline of treatment in intestinal cells, HIEC and IEC-6 cells were pretreated with L-citrulline, then irradiated, followed by ROS detecting, apoptosis detecting and clonogenic survival assay. (J)-(K) Effect of L-citrulline on ROS production after different dose of irradiation as determined by DCFH-DA staining in HIEC and IEC-6. (L)-(O) Effect of L-citrulline on radiation sensitivity as determined by a colony formation assay followed different dose of radiation. (P)-(Q) Effect of L-citrulline on cell apoptosis determined by AV/PI staining in HIEC and IEC-6 cells. (R) Schematic showing the protective effect of L-citrulline against radiation-induced intestinal injury. (S) The body weight of mice after different treatment. (T) Representative H&E-stained sections of the small intestines in different group. Scale bar = 200 μm. (U) The height of small intestine villus in different group. (V) Western blotting analysis of H3Cit17 in IEC-6 cells. (W) Immunofluorescence assay of H3Cit17 distribution in IEC-6 cells. Scale bar = 10 μm. The data are shown as the mean ± SEM for three independent experiments. (X) Mean fluorescence intensity of H3Cit17 in IEC-6 cells. The data are shown as the mean ± SEM for three independent experiments. *P < 0.05; **P < 0.01