IDDF2023-ABS-0286 Intestinal microbiota mediates redox biotransformation of pyrrolizidine alkaloids-induced acute hepatotoxicity in rats

BackgroundPyrrolizidine alkaloids (PA) of retronecine-type and otonecine-type are phytotoxins present in ~6000 plant species and known to act as human toxins via metabolic activation to form pyrrole-protein adducts (PPA) leading to hepatotoxicity. To investigate PA absorption and disposition in the body, our group recently conducted toxicokinetic (TK) of several PA in rats, and found that regardless of significantly lower oral bioavailability, hepatotoxicity via oral (PO) was significantly higher than that via intravenous (IV) administration for retronecine-type (Retrorsine (RTS)) but not otonecine-type (Clivorine (CLI)) PA (Table 1).Based on structures, only retronecine-type PA can metabolically form PA N-oxides (PANO). We hypothesize that this unusual phenomenon may be caused by metabolic redox-conversion between N-oxidation and N-reduction in the intestine mediated by intestinal microbiota. Therefore, the present study aims to further confirm orally retronecine-type PA-induced more potent hepatotoxicity and delineate the underlying mechanism.MethodsTo investigate the role of gut microbiota, male SD rats were orally administered with/without mixed non-absorbable antibiotics (ABX) once daily for 5 consecutive days to eradicate microbiota. One day after the last ABX administration, rats were treated orally/intravenously with/without RTS (40 mg/kg), followed by sacrificed at 48 h after RTS dosing.ResultsRTS exhibited severer hepatotoxicity via PO than IV administration, as indicated by remarkably elevated ALT activity (Figure 1), obvious histopathological changes indicated by H&E staining and immunostaining of sinusoidal endothelial cells, and significantly higher levels of PPA formed in plasma, red blood cells (RBCs) and liver (Figure 2). Further, ABX pre-treatment remarkably alleviated PA-ILI in rats as evidenced by significantly lowered ALT activity and PPA levels.Abstract IDDF2023-ABS-0286 Table 1TK parameters of plasma and RBC level of PPAs in rats after a single PO or IV administration of 20 mg/kg RTS (Retrorsine, retronecine-type PA) and CLI (Clivorine, otonecine-type PA) TK parameters Plasma AUC 0-720 (μM × min) RBC AUC 0-720 (μM × min) RTS IV267.03 ± 68.7513579.91 ± 1311.06RTS PO459.85 ± 37.02 **18918.88 ± 2473.79 **CLI IV154.02 ± 13.771585.94 ± 290.31CLI PO94.18 ± 13.09***1014.71 ± 178.72 *Data are presented as mean ± SD (n=4).*: p