Butyrate inhibits deoxycholic-acid-resistant colonic cell proliferation via cell cycle arrest and apoptosis

Authors: Zeng, Huawei; Safratowich, Bryan; Wang, Thomas - Tom; HAMLIN, STEPHANIE - University Of North Dakota; JOHNSON, LUANN - University Of North Dakota

Submitted to: Molecular Nutrition and Food Research
Publication Type: Peer Reviewed Journal
Publication Acceptance Date: 1/24/2020
Publication Date: 1/30/2020
Citation: Zeng, H., Safratowich, B.D., Wang, T.T., Hamlin, S., Johnson, L. 2020. Butyrate inhibits deoxycholic-acid-resistant colonic cell proliferation via cell cycle arrest and apoptosis. Molecular Nutrition and Food Research. https://doi.org/10.1002/mnfr.201901014.
DOI: https://doi.org/10.1002/mnfr.201901014

Interpretive Summary: Colon cancer accounts for approximately 140,000 new cancer cases and 50,000 deaths each year in the US, and it is predicted that half the Western population will develop at least one colon tumor by the age of 70 years. Recent studies have shown that consumption of high fat diets increases certain fecal bile acids which may cause DNA mutations in colonic cells and are implicated as promoters of colon cancer. In contrast, butyrate, an intestinal bacterial by-product of dietary fiber, may be preventive against colon cancer. We hypothesize that butyrate (at naturally occurring concentrations) eliminates bile acid-induced mutations in the colon. In the present study, we established a mutant colonic cell line generated in the presence of mutation causing (fecal) bile acids. Furthermore, we demonstrated that butyrate inhibits these mutant colonic cells at the cellular and molecular level. These findings suggest that butyrate may protect against cancer through a specific targeting of bile acid-induced mutations in the colon. The information will be useful for scientists and health-care professionals who are interested in dietary fiber intake and obesity-related colon cancer prevention.

Technical Abstract: Butyrate, an intestinal microbiota metabolite of dietary fiber, exhibits colon cancer preventive effects. In contrast, intake of a high dietary fat causes an increase in fecal secondary bile acids, such as deoxycholic acid (DCA), which are implicated as promoters of colon cancer. Human studies suggest that long term consumption of a high-fat (>40% fat as energy) diet may selectively enrich mutant epithelial cells with an abnormally high resistance to DCA-induced apoptosis in the colon. We hypothesize that physiological concentrations of butyrate inhibit DCA-resistant colonic cell proliferation. With the use of HCT-116 cells as parental colonic cells, we developed a DCA-resistant cell line (DCA-RCL) after 8 to 10 cell culture passages with increased DCA and subsequently maintained DCA-RCL cells in the presence of DCA (0.2 mM). Further characterization showed that DCA treatment (0.1 to 0.3 mM) increased intracellular reactive oxygen species (ROS, an apoptotic trigger) and cell apoptosis in HCT-116 cells, but to a 10-fold lesser extent in DCA-RCL cells. Consistent with this observation, we identified, using a PCR array analysis, 10 pro-apoptotic genes, 13 anti-apoptotic genes, and 18 cellular mediator genes with more than 1- fold increase (or decrease) in mRNA levels in DCA-RCL cells when compared with the parental HCT-116 cells. These genes are involved in the tumor necrosis factor alpha receptor, caspase recruitment domain-containing protein, and B-cell lymphoma-2 regulated pathways. Butyrate treatment (0.5 to 2 mM) inhibited DCA-RCL cell proliferation with similar efficacy when compared with HCT116 cells. At the cellular signaling level, butyrate reduced p38 mitogen-activated protein kinase (MAPK)/cellular myelocytomatosis oncogene (c-Myc) protein content, a cell proliferation promoting pathway. Collectively, we characterized DCA-RCL cells, and demonstrated that butyrate inhibited DCA-RCL cell proliferation at the cellular and molecular level. These data suggest that butyrate can protect against colon carcinogenesis through a specific targeting of DCA-resistant colonic cells.