Purpose: The goal was to provide a multifaceted insight into the impact of a continuous exposure of low doses of cereulide on metabolic responses of Caco-2 and and HepG2 cells as models for intestinal and liver toxicity.
Methods: Caco-2 and HepG2 cells were exposed to food-relevant low concentrations of CER to investigate the effect of a longer exposure. To explore the mechanisms involved in the cytotoxic response and mitochondrial function, the Seahorse Bioscience XFe24 analyzer (Massachusetts, USA) was used in combination with well-established assays for mitochondrial activity (MTT) and changes in protein content (SRB (sulforhodamine B)). The effects of cereulide on the mitochondrial oxygen consumption rate (OCR) were assessed using the Seahorse Bioscience XF Cell Mito Stress Test assay kit. In this assay, modulators of cellular respiration (oligomycin, FCCP, and a mix of rotenone and antimycin A) were serially injected providing insight into different aspects of mitochondrial function. High-resolution mass spectrometry was used to unravel the metabolic profile of CER.
Results: Both MTT and SRB assays showed toxicity on undifferentiated cells at 0.125 ng/ml CER after three days of exposure. The three-day treatment with low concentrations of CER on mitochondrial respiration in intact human intestinal epithelial cells showed perturbations in mitochondrial respiration at a concentration of 0.125 ng/ml.
Significance: These in vitro data suggest that repeated exposure of CER might injure intestinal cells even at relative low doses. Cereulide appear to be more toxic than other cyclodepsipeptide toxins with ionophoretic properties like valinomycin and beauvericin.