Phytochemical-induced reactive oxygen species and endoplasmic reticulum stress-mediated apoptosis and differentiation in malignant melanoma cells

Phytochemicals are derived from plants, vegetables and daily products and exert chemopreventive effects. Malignant melanoma is highly metastatic, and melanoma patients can develop chemotherapeutic resistance against conventional melanoma therapies. In the present study, we investigated the anti-cancer effect of the phytochemicals kaempferol (Kaem), genistein (Gen), and 3′3-diindolylmethane (DIM) on melanoma cell viability. We also evaluated the altered expression of cell cycle-related genes. We verified the production of intracellular reactive oxygen species (ROS) and endoplasmic reticulum (ER) stress at both the protein and cellular level using a western blot, TUNEL assay, and Dihydrodichlorofluorescein diacetate (DCF-DA) assay. Treatment of A375SM melanoma cells with phytochemicals resulted in inhibition of cell growth. Treatment with phytochemicals increased the gene expression of p21 and decreased the gene expression of cyclin E and/or cyclin B. The three phytochemicals activated the ROS-p38-p53 apoptotic pathway by increasing the level of phosphorylated p38 MAPK and p53, and they activated the ER stress-mediated apoptotic pathway by increasing the level of phosphorylated eIF2α and C/EBP homologous protein (CHOP). Both the ROS-p38-p53 and ER stress-mediated pathway induced the mitochondrial apoptotic pathway by attenuating Bcl-2 expression and upregulating BAX. Detection of morphological changes demonstrated that Kaem and Gen can induce differentiation in A375SM cell line. These results indicate that phytochemicals are potentially useful in treatments for melanoma due to their ability to inhibit melanoma cell growth and division via the ROS and ER stress pathway.