Manon Oliero1,2, Annie Calvé1, Gabriela Fragoso1, Thibault Cuisiniere1, Roy Hajjar1,3, Ulrich Dobrindt4, Manuela M Santos5,6. 1. Nutrition and Microbiome Laboratory, Institut du cancer de Montréal, Centre de recherche du Centre hospitalier de l'Université de Montréal (CRCHUM), 900 Rue Saint Denis, Montreal, QC, H2X 0A9, Canada. 2. Department of Medicine, Faculty of Medicine, Université de Montréal, 2900 boulevard Édouard-Montpetit, Montréal, QC, H3T 1J4, Canada. 3. Department of Surgery, Faculty of Medicine, Université de Montréal, 2900 boulevard Édouard-Montpetit, Montréal, QC, H3T 1J4, Canada. 4. Institute of Hygiene, Section Microbial Genome Plasticity, University of Münster, Mendelstraße 7, 48149, Münster, Germany. 5. Nutrition and Microbiome Laboratory, Institut du cancer de Montréal, Centre de recherche du Centre hospitalier de l'Université de Montréal (CRCHUM), 900 Rue Saint Denis, Montreal, QC, H2X 0A9, Canada. manuela.santos@umontreal.ca. 6. Department of Medicine, Faculty of Medicine, Université de Montréal, 2900 boulevard Édouard-Montpetit, Montréal, QC, H3T 1J4, Canada. manuela.santos@umontreal.ca.
Abstract
BACKGROUND: Colibactin is a genotoxin that induces DNA double-strand breaks that may lead to carcinogenesis and is produced by Escherichia coli strains harboring the pks island. Human and animal studies have shown that colibactin-producing gut bacteria promote carcinogenesis and enhance the progression of colorectal cancer through cellular senescence and chromosomal abnormalities. In this study, we investigated the impact of prebiotics on the genotoxicity of colibactin-producing E. coli strains Nissle 1917 and NC101. METHODS: Bacteria were grown in medium supplemented with 20, 30 and 40 mg/mL of prebiotics inulin or galacto-oligosaccharide, and with or without 5 μM, 25 μM and 125 μM of ferrous sulfate. Colibactin expression was assessed by luciferase reporter assay for the clbA gene, essential for colibactin production, in E. coli Nissle 1917 and by RT-PCR in E. coli NC101. The human epithelial colorectal adenocarcinoma cell line, Caco-2, was used to assess colibactin-induced megalocytosis by methylene blue binding assay and genotoxicity by γ-H2AX immunofluorescence analysis. RESULTS: Inulin and galacto-oligosaccharide enhanced the expression of clbA in pks+ E. coli. However, the addition of 125 μM of ferrous sulfate inhibited the expression of clbA triggered by oligosaccharides. In the presence of either oligosaccharide, E. coli NC101 increased dysplasia and DNA double-strand breaks in Caco-2 cells compared to untreated cells. CONCLUSION: Our results suggest that, in vitro, prebiotic oligosaccharides exacerbate DNA damage induced by colibactin-producing bacteria. Further studies are necessary to establish whether oligosaccharide supplementation may lead to increased colorectal tumorigenesis in animal models colonized with pks+ E. coli.
BACKGROUND:Colibactin is a genotoxin that induces DNA double-strand breaks that may lead to carcinogenesis and is produced by Escherichia coli strains harboring the pks island. Human and animal studies have shown that colibactin-producing gut bacteria promote carcinogenesis and enhance the progression of colorectal cancer through cellular senescence and chromosomal abnormalities. In this study, we investigated the impact of prebiotics on the genotoxicity of colibactin-producing E. coli strains Nissle 1917 and NC101. METHODS: Bacteria were grown in medium supplemented with 20, 30 and 40 mg/mL of prebiotics inulin or galacto-oligosaccharide, and with or without 5 μM, 25 μM and 125 μM of ferrous sulfate. Colibactin expression was assessed by luciferase reporter assay for the clbA gene, essential for colibactin production, in E. coli Nissle 1917 and by RT-PCR in E. coli NC101. The human epithelial colorectal adenocarcinoma cell line, Caco-2, was used to assess colibactin-induced megalocytosis by methylene blue binding assay and genotoxicity by γ-H2AX immunofluorescence analysis. RESULTS: Inulin and galacto-oligosaccharide enhanced the expression of clbA in pks+ E. coli. However, the addition of 125 μM of ferrous sulfate inhibited the expression of clbA triggered by oligosaccharides. In the presence of either oligosaccharide, E. coli NC101 increased dysplasia and DNA double-strand breaks in Caco-2 cells compared to untreated cells. CONCLUSION: Our results suggest that, in vitro, prebiotic oligosaccharides exacerbate DNA damage induced by colibactin-producing bacteria. Further studies are necessary to establish whether oligosaccharide supplementation may lead to increased colorectal tumorigenesis in animal models colonized with pks+ E. coli.
Entities:
Keywords:
Colibactin; Colorectal cancer; E. coli; Galacto-oligosaccharides; Genotoxin; Inulin; Iron
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