Hepatotoxic pyrrolizidine alkaloids—Bioavailability and cellular effects on human HEPG2 cells

Purpose: 1,2-Unsaturated pyrrolizidine alkaloids (PA) belong to the most toxic compounds. These substances are found in several plants such as Asteraceae and Boraginaceae families. Acute PA poisoning via food contamination causes severe damage to the liver; long-term, sub-lethal doses may cause cumulative damage or cancer. Methods: In this study, we analyzed the toxicological effects (cytotoxicity, ATP-content, and oxidative stress) of selected PA (retrorsine, senecionine, and seneciphylline) on the hepatocarcinoma cell line HepG2 by using a bioactivating system (S9-fraction). Furthermore, we characterized the uptake of senecionine and its N-oxidated form across the intestinal barrier by using the well-characterized human Caco-2 cell Transwell-model as an in vitro system for the human gut barrier. The PA content was analyzed by LC-MS/MS. Results: The metabolic bioactivation of PA with the S9-fraction of rat liver homogenate resulted in a weak increase of cellular cytotoxicity including a slight reduction of cellular ATP-content, as well as in a weak increase of the apoptosis rate and of oxidative stress. The Caco-2 cell absorption studies revealed a fast passage of senecionine from the apical into the basolateral compartment suggesting that active transport mechanisms are involved in the uptake and excretion of the substance. In contrast, the oxidized metabolite senecionine-N-oxide did not cross the differentiated Caco-2 monolayer from the apical to the basolateral side. In addition, we observed the conversion of senecionine into the oxidated form, as well as the reduction of senecionine-N-oxide. This indicates that Caco-2 cells are able to metabolize this compound.