OCT1-dependent uptake of structurally diverse pyrrolizidine alkaloids in human liver cells is crucial for their genotoxic and cytotoxic effects

Archives of Toxicology · 6 authors, 4 centres

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This study provides the first evidence that open-chained PA diesters and PA monoesters are OCT1 substrates.

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This study provides the first evidence that open-chained PA diesters and PA monoesters are OCT1 substrates. In PHH, d-THP almost completely inhibited lasiocarpine cytotoxicity. A limitation is that both inhibitors undergo CYP-mediated metabolism, potentially competing with PAs for CYP3A4. The findings suggest OCT1-dependent uptake is crucial for PA-induced hepatotoxicity and may inform protective strategies in acute intoxication scenarios.

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other

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preclinical research

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ORIGINAL PAPER

PMC10567918

OCT1-dependent uptake of structurally diverse pyrrolizidine alkaloids in human liver cells is crucial for their genotoxic and cytotoxic effects

Archives of Toxicology

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ABSTRACT

Abstract Pyrrolizidine alkaloids (PAs) are important plant hepatotoxins, which occur as contaminants in plant-based foods, feeds and phytomedicines. Numerous studies demonstrated that the genotoxicity and cytotoxicity of PAs depend on their chemical structure, allowing for potency ranking and grouping. Organic cation transporter-1 (OCT1) was previously shown to be involved in the cellular uptake of the cyclic PA diesters monocrotaline, retrorsine and senescionine. However, little is known about the structure-dependent transport of PAs. Therefore, we investigated the impact of OCT1 on the uptake and toxicity of three structurally diverse PAs (heliotrine, lasiocarpine and riddelliine) differing in their degree and type of esterification in metabolically competent human liver cell models and hamster fibroblasts. Human HepG2-CYP3A4 liver cells were exposed to the respective PA in the presence or absence of the OCT1-inhibitors d -THP and quinidine, revealing a strongly attenuated cytotoxicity upon OCT1 inhibition. The same experiments were repeated in V79-CYP3A4 hamster fibroblasts, confirming that OCT1 inhibition prevents the cytotoxic effects of all tested PAs. Interestingly, OCT1 protein levels were much lower in V79-CYP3A4 than in HepG2-CYP3A4 cells, which correlated with their lower susceptibility to PA-induced cytotoxicity. The cytoprotective effect of OCT1 inhibiton was also demonstrated in primary human hepatocytes following PA exposure. Our experiments further showed that the genotoxic effects triggered by the three PAs are blocked by OCT1 inhibition as evidenced by strongly reduced γH2AX and p53 levels. Consistently, inhibition of OCT1-mediated uptake suppressed the activation of the DNA damage response (DDR) as revealed by decreased phosphorylation of checkpoint kinases upon PA treatment. In conclusion, we demonstrated that PAs, independent of their degree of esterification, are substrates for OCT1-mediated uptake into human liver cells. We further provided evidence that OCT1 inhibition prevents PA-triggered genotoxicity, DDR activation and subsequent cytotoxicity. These findings highlight the crucial role of OCT1 together with CYP3A4-dependent metabolic activation for PA toxicity. Supplementary Information The online version contains supplementary material available at 10.1007/s00204-023-03591-4. Keywords: Pyrrolizidine alkaloids, Cytotoxicity, Genotoxicity, OCT1, Transport, Primary human hepatocytes, γH2AX, p53

CITATION EXPORT

Manuel Haas, Gabriel Ackermann, Jan-Heiner Küpper et al.. OCT1-dependent uptake of structurally diverse pyrrolizidine alkaloids in human liver cells is crucial for their genotoxic and cytotoxic effects. Archives of Toxicology. (2023). https://doi.org/10.1007/s00204-023-03591-4

AUTHORS

6 · 4 CENTRES

Manuel Haas, Gabriel Ackermann, Jan-Heiner Küpper, Hansruedi Glatt, Dieter Schrenk, Jörg Fahrer

1Division of Food Chemistry and Toxicology, Department of Chemistry, RPTU Kaiserslautern-Landau, Erwin-Schroedinger-Str. 52, 67663 Kaiserslautern, Germany
2Division of Molecular Cell Biology, Department of Environment and Nature Science, Brandenburg University of Technology Cottbus-Senftenberg, 01968 Senftenberg, Germany
3Department Food Safety, German Federal Institute for Risk Assessment (BfR), Max-Dohrn-Strasse 8-10, 10589 Berlin, Germany
4Department of Nutritional Toxicology, German Institute of Human Nutrition (DIfE), Potsdam-Rehbrücke, Arthur-Scheunert-Allee 114-116, 14558 Nuthetal, Germany