RESUMO
Malignant melanoma is the most aggressive type of skin cancer with increasing incidence rates worldwide. On the other hand, watercress is a rich source of phenethyl isothiocyanate (PEITC), among others, which has been widely investigated for its anticancer properties against various cancers. In the present study, we evaluated the role of a watercress extract in modulating apoptotic induction in an in vitro model of human malignant melanoma consisting of melanoma (A375, COLO-679, COLO-800), non-melanoma epidermoid carcinoma (A431) and immortalized, non-tumorigenic keratinocyte (HaCaT) cells. Moreover, the chemical composition of the watercress extract was characterized through UPLC MS/MS and other analytical methodologies. In addition, cytotoxicity was assessed by the alamar blue assay whereas apoptosis was determined, initially, by a multiplex activity assay kit (measuring levels of activated caspases -3, -8 and -9) as well as by qRT-PCR for the identification of major genes regulating apoptosis. In addition, protein expression levels were evaluated by western immunoblotting. Our data indicate that the extract contains various phytochemicals (e.g. phenolics, flavonoids, pigments, etc.) while isothiocyanates (ITCs; especially PEITC) were the most abundant. In addition, the extract was shown to exert a significant time- and dose-dependent cytotoxicity against all malignant melanoma cell lines while non-melanoma and non-tumorigenic cells exhibited significant resistance. Finally, expression profiling revealed a number of genes (and corresponding proteins) being implicated in regulating apoptotic induction through activation of the intrinsic apoptotic cascade. Overall, our data indicate the potential of PEITC as a promising anti-cancer agent in the clinical management of human malignant melanoma.
RESUMO
Malignant melanoma is an aggressive type of skin cancer characterised by high metastatic capacity and mortality rate. On the other hand, Epilobium parviflorum is known for its medicinal properties, including its anticancer potency. In this context, we aimed to (i) isolate various extracts of E. parviflorum, (ii) characterize their phytochemical content, and (iii) determine their cytotoxic potential in an in vitro model of human malignant melanoma. To these ends, we utilized various spectrophotometric and chromatographic (UPLC-MS/MS) approaches to document the higher content of the methanolic extract in polyphenols, soluble sugars, proteins, condensed tannins, and chlorophylls -a and -b as opposed to those of dichloromethane and petroleum. In addition, the cytotoxicity profiling of all extracts was assessed through a colorimetric-based Alamar Blue assay in human malignant melanoma (A375 and COLO-679) as well as non-tumorigenic immortalized keratinocyte (HaCaT) cells. Overall, the methanolic extract was shown to exert significant cytotoxicity, in a time- and concentration-dependent manner, as opposed to the other extracts. The observed cytotoxicity was confined only to human malignant melanoma cells, whereas non-tumorigenic keratinocyte cells remained relatively unaffected. Finally, the expression levels of various apoptotic genes were assessed by qRT-PCR, indicating the activation of both intrinsic and extrinsic apoptotic cascades.
RESUMO
Watercress is an enriched source of phenethyl isothiocyanate (PEITC), among other phytochemicals, with an antioxidant capacity. The aim of this study was to (i) chemically characterize and (ii) biologically evaluate the profile of the main health-promoting compounds contained in edible (i.e., mixture of leaves and lateral buds) and non-edible (i.e., stems) parts of watercress in an in vitro model of malignant melanoma consisting of human malignant melanoma (A375), non-melanoma (A431) and keratinocyte (HaCaT) cells. The extraction of the main constituents of watercress was performed by subjecting the freeze-dried edible and non-edible samples through different extraction protocols, whereas their concentration was obtained utilizing analytical methodologies. In addition, cell viability was evaluated by the Alamar Blue assay, whereas levels of oxidative stress and apoptosis were determined by commercially available kits. The edible watercress sample contained a higher amount of various nutrients and phytochemicals in the hexane fraction compared to the non-edible one, as evidenced by the presence of PEITC, phenolics, flavonoids, pigments, ascorbic acid, etc. The cytotoxicity potential of the edible watercress sample in the hexane fraction was considerably higher than the non-edible one in A375 cells, whereas A431 and HaCaT cells appeared to be either more resistant or minimally affected, respectively. Finally, levels of oxidative stress and apoptotic induction were increased in both watercress samples, but the magnitude of the induction was much higher in the edible than the non-edible watercress samples. Herein, we provide further evidence documenting the potential development of watercress extracts (including watercress waste by-products) as promising anti-cancer agent(s) against malignant melanoma cells.
RESUMO
Malignant melanoma is one of the most aggressive types of skin cancer with an increasing incidence worldwide. Thus, the development of innovative therapeutic approaches is of great importance. Salvia fruticosa (SF) is known for its anticancer properties and in this context, we aimed to investigate its potential anti-melanoma activity in an in vitro model of human malignant melanoma. Cytotoxicity was assessed through a colorimetric-based sulforhodamine-B (SRB) assay in primary malignant melanoma (A375), non-malignant melanoma epidermoid carcinoma (A431) and non-tumorigenic melanocyte neighbouring keratinocyte (HaCaT) cells. Among eight (8) different fractions of S. fruticosa extracts (SF1-SF8) tested, SF3 was found to possess significant cytotoxic activity against A375 cells, while A431 and HaCaT cells remained relatively resistant or exerted no cytotoxicity, respectively. In addition, the total phenolic (Folin-Ciocalteu assay) and total flavonoid content of SF extracts was estimated, whereas the antioxidant capacity was measured via the inhibition of tert-butyl hydroperoxide-induced lipid peroxidation and protein oxidation levels. Finally, apoptotic cell death was assessed by utilizing a commercially available kit for the activation of caspases - 3, - 8 and - 9. In conclusion, the anti-melanoma properties of SF3 involve the induction of both extrinsic and intrinsic apoptotic pathway(s), as evidenced by the increased activity levels of caspases - 8, and - 9, respectively.
