Acceleration of benzo(a)pyrene-induced colon carcinogenesis by Western diet in a rat model of colon cancer

Abstract

Colorectal cancer (CRC) is the third leading cause of cancer-related mortalities in the USA and around 52,550 people were expected to die from this disease by December 2023. The objective of this study was to investigate the effect of diet type on benzo(a)pyrene [B(a)P]-induced colon cancer in an adult male rat model, the Polyposis In the Rat Colon (PIRC) kindred type. Groups of PIRC rats (n = 10) were fed with AIN-76A regular diet (RD) or Western diet (WD) and received 25, 50 and 100 µg B(a)P/kg body wt. via oral gavage for 60 days. Rats fed diets alone, but no B(a)P, served as controls. After exposure, rats were euthanized; colon and liver samples were analyzed for activation of drug metabolizing enzymes (DMEs) CYP1A1, CYP1B1, SULT and GST. Plasma and tissue samples were analyzed by reverse phase-HPLC for B(a)P metabolites. In addition to these studies, DNA isolated from colon and liver tissues was analyzed for B(a)P-induced DNA adducts by the ³²P-postlabeling method using a thin-layer chromatography system. Western diet consumption resulted in a marked increase in DME expression and B(a)P metabolite concentrations in rats that were administered 100 µg/kg B(a)P + WD (p < 0.05) compared to other treatment groups. Our findings demonstrate that WD accelerates the development of colon tumors induced by B(a)P through enhanced biotransformation, and the products of this process (metabolites) were found to bind with DNA and form B(a)P-DNA adducts, which may have given rise to colon polyps characterized by gain in tumor number, sizes, and dysplasia.

Keywords: Benzo(a)pyrene; Colon cancer; DNA adducts; Lipid peroxidation; Polycyclic aromatic hydrocarbons; Western diet.

Graphical abstract

Fig. 1

A. RT-PCR analysis of liver CYP1A1 mRNA levels in PIRC rat liver samples following treatment with B(a)P and consumption of RD or WD. b. CYP1A1 protein expression in liver samples of PIRC rats following treatment with B(a)P and consumption of RD or WD. c. RT-PCR analysis of liver CYP1B1 mRNA levels in PIRC rat liver samples following treatment with B(a)P and consumption of RD or WD. d CYP1B1 protein expression in liver samples of PIRC rats. Annotations denote statistical significance (*p < 0.05, ^Φp < 0.0001 compared to RD Diet Only, ^Øp < 0.05 compared to WD Only, and ^∞p < 0.001 compared to WD Only) among diets and B(a)P concentrations used.

Fig. 2

A. SULT1A1 protein expression in liver samples of PIRC rats following treatment with B(a)P and consumption of RD or WD. b. GST protein expression in liver samples of PIRC rats. Annotations denote statistical significance (^±p < 0.001, ^tp < 0.0001, ^≠p < 0.05 compared to RD Only, ⁺p < 0.01 compared to RD Only, ^Φp < 0.0001 compared to RD Only, ^Øp < 0.05 compared to WD Only, and ^∼p < 0.0001 compared to WD Only) among diets and B(a)P concentrations used.

Fig. 3

A. RT-PCR analysis of liver CYP1A1 mRNA levels in PIRC rat liver samples following treatment with B(a)P and consumption of RD or WD. b. CYP1A1 protein expression in colon samples of PIRC rats following treatment with B(a)P and consumption of RD or WD. c. RT-PCR analysis of liver CYP1B1 mRNA levels in PIRC rat liver samples following treatment with B(a)P and consumption of RD or WD. d. CYP1B1 protein expression in colon samples of PIRC rats following treatment with B(a)P and consumption of RD or WD. Annotations denote statistical significance (*p < 0.05, ^±p < 0.001, ⁺p < 0.01 compared to RD Only, ⊥p < 0.001 compared to RD Only) among diets and B(a)P doses used.

Fig. 4

A. SULT1A1 protein expression in colon samples of PIRC rats following treatment with B(a)P and consumption of RD or WD. Annotations denote statistical significance (^xp < 0.01, and ^Φp < 0.0001 compared to RD Only, and ^∼p < 0.0001 compared to WD Only) among diets and B(a)P doses used. B. GST protein expression in colon samples of PIRC rats following treatment with B(a)P and consumption of RD or WD. Annotations denote statistical significance (^xp < 0.01 and ^Øp < 0.05 compared to Western Diet Only) among diets and B(a)P doses used.

Fig. 5

A–C B(a)P metabolite concentrations in plasma, liver and colon samples from PIRC rats following treatment with 25, 50, and 100 µg B(a)P/kg bw for 60 days via oral gavage and consumption of RD diet or WD. d. Representative analysis of Benzo(a)pyrene metabolism in colon sample of PIRC rat exposed to 50 µg B(a)P/kg bw and provided RD or WD.

Fig. 6

A–C. DNA strand breaks in liver (A) and colon (B) samples of PIRC rats following treatment with 25, 50, and 100 µg B(a)P/kg bw for 60 days via oral gavage and consumption of RD or WD. c,d. Oxidative stress marker concentrations in serum (C) and colon (D) samples from PIRC rats following treatment with 25, 50, and 100 µg B(a)P/kg bw for 60 days via oral gavage and consumption of RD or WD. Annotations denote statistical significance (^xp < 0.01, ⁺p < 0.01 compared to RD Only, ^#p < 0.01 compared to WD Only) among diets and B(a)P concentrations used.

Fig. 7

A-B. Benzo(a)pyrene-DNA adduct total concentrations in colon (A) and liver (B) tissues of PIRC rats that were fed either RD or WD and exposed to 25-,50- and 100 µg B(a)P/kg bw. The bars represent mean ± S.E. for three independent experiments. *p < 0.05 in adduct concentrations for the respective B(a)P + RD treatment group compared to B(a)P + WD treatment group.