Abstract. Ursolic acid (UA) is a pentacyclic triterpenoid compound which exists widely in nature and is known to have a pleitropic biological activity profile. For the last few decades, extensive work has been carried out to establish its biological activities and pharmacological actions. It is described as a promising chemopreventive agent with an antiproliferative effect on cancer cells that stems from its ability to induce apoptosis. We investigated and compared the role played by mitochondria during the apoptotic process induced by UA in human HaCaT-derived keratinotic cells and M4Beu human melanoma cells. In both cell lines, UA induced significant caspase-3 activation, the downstream central effector of apoptosis. Subsequent JC-1/TOTO-3 double staining clearly demonstrated that UA induces strong mitochondrial-transmembrane potential collapse in M4Beu cells, while mitochondria from HaCaT-treated cells remain largely unstimulated. This was confirmed by Western blot analysis, which revealed a Bax/Bcl-2-balance change in favor of Bax, the proapoptotic member, in UA-treated M4Beu cells. It can be concluded that UA induces apoptosis in M4Beu through the mitochondrial pathway, while other mechanisms are activated in the case of HaCaT cells.
IntroductionAmong natural compounds, ursolic acid (UA), the most frequently studied natural triterpenic acid (1), seems to be a promising chemical entity for the protection of human skin. Indeed, in cosmetology UA is often used for photoaging protection (2). It prevents and improves the appearance of wrinkles and age spots by restoring skin collagen structure and elasticity, stimulates collagen production in cultured fibroblasts and increases the production of ceramides in human epidermal keratinocytes and skin (3). UA may also be an effective inhibitor of the UVA-modulated signal transduction pathway in human HaCaT cells, in which it significantly suppresses UVA-induced reactive oxygen species production and lipid peroxidation (4). Moreover, it possesses anti-tumor properties including the inhibition of skin tumorigenesis (5) and tumor promotion (6). UA also induces apoptosis in several cancer cell lines (7-13).Previously, we studied the effect of UA on human HaCaTderived keratinotic cells and M4Beu human melanoma cells with the intention of confirming its role as a promising candidate for skin cancer prevention. In those studies, we demonstrated for the first time that UA has a significant antiproliferative effect associated with the induction of the apoptotic process (14,15). In the literature, it is well established that the activation of the apoptotic transduction pathway is dependent on cell type and subcellular targets. The two main apoptotic pathways are the extrinsic and intrinsic pathways (16). The extrinsic pathway is characterized by death ligand attachment on extracellular receptors (e.g. TNF/TNF-R) with a subsequent caspase-8 activation, which in turn cleaves and activates caspase-3 (17). The intrinsic pathway is triggered by different mitochondrial stresses and i...