P18-50 In vitro impact of hesperidin on human melanoma cells

The skin is an organ that undergoes numerous changes over time because it is in direct contact with the external environment, is vulnerable to microbial, physical, and chemical stress, and can lead to superficial lesions and even cancerous pathologies. A skin pathology where therapeutic intervention must be immediately implemented is malignant cutaneous melanoma, with a high mortality rate worldwide, being the most aggressive form of skin cancer. Due to the complexity of the skin organ, compounds that possess benefits at this level are of interest in both the medical and cosmetic spheres, mainly due to the emergence of drug resistance. Hesperidin (HES), a bioflavonoid found in citrus fruits, may be a possible anti-tumor candidate due to its multiple therapeutic properties. Considering these aspects, the present study aims to investigate the in vitro effect of HES on the A375 human melanoma cell line. The in vitro biological impact of HES on melanoma cells was investigated by several methods, including the evaluation of cell viability (through MTT colorimetric assay), determination of cellular morphological changes through bright-field microscopy, and cytotoxic potential (using LDH test). The effect of HES on the nuclear and mitochondrial aspects of A375 cells was evaluated using immunofluorescence techniques, the Hoechst 33342 and MitoTracker™ Red CMXRos method. The results indicated that treatment of A375 cells with HES (50- 100 µM) induced a dose-dependent decrease in cell viability, and the percentages of released LDH increased slightly with rising concentrations. Morphological analysis suggested reduced confluency and signs of cell detachment from the plaque, with the most pronounced dysmorphologies occurring at 100 µM HES treatment. Additionally, Hoechst 33342 staining for cell nuclei showed the presence of shrunken and condensed nuclei at 100 µM. At the same time, mitochondrial evaluation pointed out that treatment with 100 µM HES did not produce consistent alterations compared to the appearance of mitochondria in untreated cells. These findings revealed the in vitro anticancer potential of HES, which caused a cytotoxic effect in A375 cells. The current study was conducted within the Research Centre for Pharmaco-Toxicological Evaluations, “Victor Babes” University of Medicine and Pharmacy, Eftimie Murgu Square, no 2 RO-300041, Timisoara, Romania.