Evaluation of In Vitro Models for Pyrrolizidine Alkaloid‐Induced Hepatocellular Injury

Abstract

ObjectivePyrrolizidine alkaloids (PAs) are toxic chemicals found in many herbal medicines, dietary supplements and even food sources such as honey, bread, and tea. Cases of drug‐induced liver injury due to herbal and dietary supplements are relatively common in Asia and are increasing in the United States. The literature suggests that PAs containing macrocyclic structure are more toxic than those with smaller sidechains. The goal of this project was to evaluate two PAs with macrocyclic structure (monocrotaline and retrorsine) and one PA will a smaller sidechain (retronecine) for their potential to elicit toxicity across two in vitro model systems.MethodsPA toxicity was measured by determining the concentration at which 50% of the cell population was killed (LC50). PAs were dissolved in DMSO and PA‐induced toxicity was measured after 48 hours in two in vitro models: TAMH (transforming growth factor‐α transgenic mouse hepatocytes) and THLE‐2 (transformed human liver epithelial‐2 cells). Statistical analysis of cell viability data was performed using SPSS (Probit regression) to obtain LC50 values.ResultsA decrease in cell viability was observed as PA concentration increased. Interestingly, both cell lines were relatively resistant to PA‐induced toxicity. In many cases only predicted LC50 values for each compound could be generated and are indicated by an asterisk (7.0mM*, 11mM*, and 30.mM* in TAMH for monocrotaline, retrorsine and retronecine, respectively; and 2.2mM, 3.2mM, and >100mM* in THLE‐2 for monocrotaline, retrorsine and retronecine, respectively. Based on the toxicologic parameters, monocrotaline and retrorsine were more toxic (lower LC50 values) than retronecine.ConclusionsWhile PAs have been associated with hepatotoxicity, limited toxicity was seen in both the TAMH and THLE‐2 cell lines. Predicted LC50 values were higher than expected, suggesting TAMH and THLE‐2 cells are not good models for studying PA‐induced liver toxicity. Retronecine, which contains a smaller sidechain, was found to be less toxic than the larger compounds. Future work is currently underway to identify biomarkers responsible for the observed resistance to PA‐induced toxicity as potential targets for preventing hepatocellular injury.Support or Funding InformationThis work was supported by the Pacific University Research Incentive Grant Program and an ASPET Summer Undergraduate Research Fellowship (SURF) award at Pacific University.