Apiaceous Vegetable Intake Modulates Expression of DNA Damage Signaling Pathway Genes and miRNA in Wistar Rats

Abstract

2‐Amino‐1‐methyl‐6‐phenylimidazo[4,5‐b]pyridine (PhIP) is the most mass abundant heterocyclic aromatic amine in foods and is a colon‐specific carcinogen. We recently showed that PhIP‐DNA adducts in rat colon were reduced by 20% when the diet was supplemented with apiaceous vegetables (i.e., celery and parsnips) at a modest dose (21%, wt/wt) equivalent to a human intake of 1 cup/day (based on a 2,000 kcal diet). The mechanism of action, however, was not through a commonly hypothesized effect, inhibition of pro‐carcinogen activation. Thus, here we examine the effects of apiaceous vegetable intake on promising alternative mechanisms of dietary chemoprevention: 1) N‐methyltransferase (NMT) protein expression, 2) gene expression of PhIP transporters, 3) expression of genes related to DNA damage signaling pathways, and 4) changes in the global microRNA (miRNA) profile in rat colon. Male Wistar rats fed either the American Institute for Nutrition (AIN)‐93G diet (n=6), or the AIN‐93G supplemented with apiaceous vegetables (API; n=7), received a single intraperitoneal injection of PhIP (10 mg PhIP/kg body weight) after 6 days of feeding. A negative control group was fed AIN‐93G and injected with DMSO (n=5). No difference in hepatic NMT protein expression was noted amongst the groups. There was a trend for increased colonic mRNA expression of p‐glycoprotein, a minor PhIP efflux transporter (1.5‐fold; P=0.075); mRNA expression of other colonic PhIP efflux transporters, BCRP and MRP2, was not changed. PCR array analysis revealed that mRNA expression of Pole and Rad18, involved in DNA repair pathways and induced in response to DNA damage, was reduced by API (1.3‐fold for both; P<0.05). Also, in the global miRNA profiling analysis, 11 out of 421 miRNAs were either elevated or suppressed in response to apiaceous vegetable supplementation. In particular, miR‐19b‐3p and miR‐29b‐3p were increased by API diet (2.8‐ and 2.2‐fold increase, respectively, P<0.05). Predictions by Ingenuity Pathway Analysis suggest that these two miRNAs target BMPR2 and CCND1, among others. Some evidence suggests BMPR2 may be oncogenic, and there are reports of a positive correlation between CCND1 overexpression and cancer onset. In conclusion, apiaceous vegetable intake had negligible effects on NMT protein and PhIP transporter gene expression, but did modify genes and miRNA with potential chemopreventive roles. Further investigations are warranted regarding the impact of apiaceous‐mediated modulation of Pole and Rad18, and the net downstream effects of miRNA modulation as pertains to colon carcinogenesis.Support or Funding InformationUniversity of Minnesota Masonic Cancer Center