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Title: Reversible toxic effects of the dietary supplement Indole-3-Carbinol in an immune compromised rodent model: intestine as the main target

Author
item FLETCHER, ARNETTA - University Of Maryland
item HUANG, HAIQIU - University Of Maryland
item YU, LU - University Of Maryland
item PHAM, QUYNHCHI - University Of Maryland
item YU, LIANGLI - University Of Maryland
item Wang, Thomas - Tom

Submitted to: Journal of Dietary Supplement
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 7/17/2016
Publication Date: 5/1/2017
Citation: Fletcher, A.R., Huang, H., Yu, L., Pham, Q., Yu, L., Wang, T.T. 2017. Reversible toxic effects of the dietary supplement Indole-3-Carbinol in an immune compromised rodent model: Intestine as the main target. Journal of Dietary Supplement. 14:(3):303-322. doi: 10.1080/19390211.

Interpretive Summary: Indole-3-carbinol (I3C), a bioactive compound commonly found in cruciferous vegetables, is known for its protective effects against chronic diseases. These benefits promoted its use as a commercially available dietary supplement. However, the safety of I3C, especially in high amounts, is unresolved. A nude mouse model commonly used in preclinical cancer studies were fed 0- 100 µmoles I3C/ g diet for 5 weeks. Results from this study indicated negative results when fed the highest concentration, and mice in this group were not viable after 7 days of treatment. Conversely, switching mice to the control diet without I3C upon detecting stress resulted in a 75% recovery rate. Mice fed 10 and 50 µmol I3C/ g diets survived through the entire study. Although, significant weight loss occurred in those fed 50 µmol I3C/ g diet. Physical signs of distress included skin flush and the presence of fecal blood within these mice. Mice fed 10 µmol I3C/ g diet had a similar healthy appearance as controls, yet fecal blood discharge was observed. The gastrointestinal tract as the primary affected tissue and significant changes in the size, shape, and viability of small intestine and colon region. The results from this study is the first report of hazardous effects specific to the gastrointestinal tract and should serve as a caution for excessive use of I3C.

Technical Abstract: Indole-3-carbinol (I3C), a bioactive compound generated by the hydrolysis of glucobrassicin from cruciferous vegetables, is known for its potential protective effects against chronic diseases. These benefits fostered its availability as a commercially available dietary supplement. However, the safety of orally consumed I3C, especially in high amounts remains largely unresolved. Immune-deficient BALB/C nu/nu male mice (6 to 8 weeks old), a model commonly used in tumorigenesis studies, were fed 0- 100 µmoles I3C/ g diet for 5 weeks. Immunohistochemistry and real-time RT PCR was employed to elucidate pathophysiological changes underlying intestinal damage. Results of one-way ANOVA were reported as means ± SE. Mice were biologically responsive to 10-100 µmoles I3C/ g diet, concentrations used in previous animal studies. Significant induction of CYP1A1 expression by 200 fold, and other Phase 1 and 2 xenobiotic enzymes were observed livers of mice treated with 50 µmol I3C/ g diet and in human HepG2 liver carcinoma cells. This supports involvement of aryl hydrocarbon receptor-mediated pathway in I3C metabolism. I3C also altered the expression of Phase 2 xenobiotic metabolizing enzymes in HepG2. Mice fed an AIN-93 diet supplemented with 100 µmoles I3C/ g diet were not viable after seven days. Conversely, switching mice to the control diet without I3C upon detecting stress resulted in a 75% recovery rate. Mice fed 10 and 50 µmol I3C/ g diets survived through the entire study. Although, significant weight loss occurred in those fed 50 µmol I3C/ g diet. Symptoms included skin flush and fecal blood with these mice. Mice fed 10 µmol I3C/ g diet had a similar appearance as controls, yet fecal blood discharge was observed. Hematoxylin and eosin staining of intestine, spleen, kidney, and liver revealed the gastrointestinal tract as the primary affected tissue. Number and width of intestinal villi were significantly altered by all I3C concentrations. Intestinal analysis using Ki67 proliferation marker and TUNEL assay indicated a dose-dependent reduction in cell proliferation and increase in apoptosis, respectively. This study is the first report of hazardous effects specific to the gastrointestinal tract and should serve as a caution for excessive use of I3C.