Ingestion of aristolochic acid (AA), especially its major constituent aristolochic acid I (AAI), results in severe kidney injury known as aristolochic acid nephropathy (AAN). Although hepatic cytochrome P450s metabolize AAI to reduce its kidney toxicity in mice, the mechanism by which AAI is uptaken by renal cells to induce renal toxicity is largely unknown. In this study, we found that organic anion transporters (OATs) 1 and 3, proteins known to transport drugs from the blood into the tubular epithelium, are responsible for the transportation of AAI into renal tubular cells and the subsequent nephrotoxicity. AAI uptake in HEK 293 cells stably transfected with human OAT1 or OAT3 was greatly increased compared to that in the control cells, and this uptake was dependent on the AAI concentration. Administration of probenecid, a well-known OAT inhibitor, to the mice reduced AAI renal accumulation and its urinary excretion and protected mice from AAI-induced acute tubular necrosis. Further, AAI renal accumulation and severe kidney lesions induced by AAl in Oat1 and Oat3 gene knockout mice all were markedly suppressed compared to those in the wild-type mice. Together, our results suggest that OAT1 and OAT3 have a critical role in AAl renal accumulation and toxicity. These transporters may serve as a potential therapeutic target against AAN.

Comfrey has been consumed by humans as a vegetable and a tea and used as an herbal medicine for more than 2000 years. Comfrey, however, produces hepatotoxicity in livestock and humans and carcinogenicity in experimental animals. Comfrey contains as many as 14 pyrrolizidine alkaloids (PA), including 7-acetylintermedine, 7-acetyllycopsamine, echimidine, intermedine, lasiocarpine, lycopsamine, myoscorpine, symlandine, symphytine, and symviridine. The mechanisms underlying comfrey-induced genotoxicity and carcinogenicity are still not fully understood. The available evidence suggests that the active metabolites of PA in comfrey interact with DNA in liver endothelial cells and hepatocytes, resulting in DNA damage, mutation induction, and cancer development. Genotoxicities attributed to comfrey and riddelliine (a representative genotoxic PA and a proven rodent mutagen and carcinogen) are discussed in this review. Both of these compounds induced similar profiles of 6,7-dihydro-7-hydroxy-1-hydroxymethyl-5H-pyrrolizine (DHP)-derived DNA adducts and similar mutation spectra. Further, the two agents share common mechanisms of drug metabolism and carcinogenesis. Overall, comfrey is mutagenic in liver, and PA contained in comfrey appear to be responsible for comfrey-induced toxicity and tumor induction.

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Emodin, an anthraquinone derived from a plant and fungi, has been reported to possess potential genotoxicity, but the mechanism is not entirely clear. Here, we report that emodin causes DNA double-strand breaks (DSBs) through stabilization of topoisomerase (Topo) II-DNA cleavage complexes and inhibition of ATP hydrolysis. In our study, emodin did not induce mutagenecity in the salmonella mutation assay but caused genotoxicity in the thymidine kinase gene mutation assay and in the micronucleus test. Moreover, emodin induced DNA DSBs demonstrated by induction of comet tails, the expression of phosphorylated histone H2AX, and phosphorylation of ataxia telangiectasia mutated. Our studies also revealed that emodin exerted strong inhibitory activity against Topo II in the supercoiled pBR322 relaxation assay and in Topo II-mediated kinetoplast DNA decatenation, similar to the previous report. We also showed that the inhibitory effect of emodin on Topo II was because of its ability to stabilize Topo II-DNA complexes and to inhibit the ATP hydrolysis of Topo II. Furthermore, emodin was found to trigger DNA DSBs in a Topo II-dependent manner using the Topo II catalytic inhibitor aclarubicin and in Topo II-deficient mitoxantrone-resistant variant HL-60/MX2 cells. Together, these results suggest that in emodin-induced DNA DSBs and genotoxicity, stabilization of Topo II-DNA cleavage complexes and inhibition of ATP hydrolysis play an important role.