Introduction: Colorectal cancer (CRC) is the third most commonly diagnosed cancer worldwide. Its incidence is influenced by genetics, age, lifestyle, and diet factors. Indeed, a balanced diet rich in fiber and regular physical activity can significantly reduce the risk of developing CRC. Functional Foods have aroused considerable interest due to their antiproliferative and anti-cancer effects. Interestingly, bergapten, the main furanocoumarin in bergamot, exhibiting antioxidant properties, has been particularly studied for its ability to inhibit proliferation and induce apoptosis in different tumour cell lines, including CRC. Paradoxically, several reports have highlighted the pro-oxidant effects of natural antioxidant compounds, which increase free radicals and exacerbate oxidative stress. Based on this evidence, we investigated the molecular mechanism by which bergapten may affect survival and ROS production in CRC cells.
Methods: CaCo-2 colorectal adenocarcinoma cells were used as a model to evaluate bergapten anti-cancer effects. ROS production and mitochondrial membrane potential were evaluated through flow cytometry. mRNA levels of stemness, the epithelial-to-mesenchymal transition (EMT), the cell cycle and apoptosis markers were quantified by qRT-PCR. Western blotting was carried out to detect protein levels.
Results: Low doses of bergapten induced a reduction in intracellular ROS levels in CaCo-2 cells, as is generally reported. Unexpectedly, high doses of the furanocumarin promoted a time-dependent significant increase in ROS levels. In CaCo-2 bergapten-treated cells, a reduction in mitochondrial membrane potential, a down-regulation of anti-apoptotic markers, and an increase in pro-apoptotic markers were observed, leading to cytochome C release. In agreement with this, bergapten induced a lowered expression of Cyclin D1, Notch family members, and Survivin compared to untreated cells. Interestingly, the bergapten-induced inhibition of Notch signalling also led to the down-regulation of the EMT, reducing CRC growth and progression.
Conclusions: These results suggest that bergapten, at high doses, may act as a promoter of oxidative stress, favoring antiproliferative effects through the activation of intrinsic apoptosis.