MicroRNA expression profiles distinguish the carcinogenic effects of riddelliine in rat liver

Pyrrolizidine alkaloids (PAs) are the most common plant constituents that poison livestock, wildlife and humans. Riddelliine is a prototype genotoxic PA and has been nominated to be classified as a reasonably anticipated human carcinogen by the US National Toxicology Program (NTP) in the 12th Report on Carcinogens. Riddelliine's nomination is due to the high incidence of liver tumours that were observed in both mice and rats in the NTP tumourigenicity bioassay study. In this current study, we explored whether riddelliine treatment could alter microRNA (miRNA) expression in rat liver and whether the possible deregulation of miRNA was related to mutagenicity and carcinogenicity of riddelliine. Groups of six rats were administered riddelliine at a mutagenic dose of 1 mg/kg body weight or with control vehicle 5 days a week for 12 weeks. A group of six rats treated with aristolochic acid, a renal carcinogen, was used as a tissue-specific negative control. The animals were sacrificed 1 day after the last treatment and the livers were isolated for miRNA expression analysis using miRNA microarrays. miRNA expression was significantly altered by riddelliine treatment. Principal component analysis and hierarchical clustering analysis showed that the miRNA expression profiles were clearly classified into two groups, riddelliine treatment versus other samples. Forty-seven miRNAs were significantly dysregulated by riddelliine treatment, among which 38 were up-regulated and 9 were down-regulated. Functional analysis of these differentially expressed miRNAs by riddelliine revealed that these miRNAs were involved in liver carcinogenicity and toxicity, such as liver proliferation, liver necrosis/cell death, hepatocellular carcinoma, liver hepatomegaly, liver inflammation and liver fibrosis. These results suggest that miRNAs actively respond to a mutagenic dose of riddelliine and the pattern of miRNA expression has the potential to be used as a biomarker of genotoxicity and carcinogenicity for riddelliine and possibly other PAs.