Combination of cyanidin-3-O-glucoside and cisplatin induces oxidative stress and apoptosis in HeLa cells by reducing activity of endogenous antioxidants, increasing bax/bcl-2 mRNA expression ratio, and downregulating Nrf2 expression.

Abstract

Investigation on potentiation of existing drugs with natural compounds to enhance efficacy and reduce toxic effect of the drugs has been increasing in recent years. This paper reports cytotoxic effect (apoptosis-related and oxidative stress-related effect) of cyanidin-3-O-glucoside (C3G), cisplatin (DDP), and their combination (C3G-DDP) on cervical cancer HeLa cells. Concentration of intracellular reactive oxygen species (ROS) was determined by employing fluorescent marker 2',7'-dichlorodihydrofluorescein diacetate. On the other hand, malondialdehyde (MDA) and glutathione (GSH) concentration, and activity of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px) were quantitated by commercially available assay kits. C3G-DDP significantly inhibited the activity of SOD, CAT, and GSH-Px. Simultaneously, C3G-DDP reduced GSH concentration while increased the concentration of ROS and MDA. Moreover, Western blot analysis suggested that C3G-DDP significantly reduced the expression of nuclear factor erythroid 2-related factor-2 (Nrf2) and Nrf2 target proteins: heme oxygenase-1 (HO-1) and NAD(P)H quinone dehydrogenase 1 (NQO1). In contrast, C3G-DDP increased the expression of Keap1. Furthermore, C3G-DDP significantly upregulated and downregulated the mRNA expressions of bax and bcl-2, respectively, thereby increasing bax/bcl-2 mRNA expression ratio. Overall, our findings propose that potentiation of DDP with C3G improves cancer cell susceptibility, specifically cervical cancer cells, to DDP. PRACTICAL APPLICATIONS: Cisplatin is recommended by most medical oncologists worldwide to treat cancer. Despite its neoplastic efficacy, it has undesirable side effects including nausea, vomiting, nephrotoxicity, and hepatotoxicity. Natural biologically active food ingredients are suggested to be used as antioxidants along with DDP therapy to prevent cisplatin-induced toxicity. C3G-DDP protected HeLa cells from oxidative stress by reducing NQO1 and HO-1 levels and regulated the Nrf2 signaling pathway. In addition, C3G-DDP protected HeLa cells from oxidative stress-induced apoptosis by increasing bcl-2 levels and decreasing bax levels. These results expanded our understanding of the role of C3G in a cervical cancer cell model, and provided a potential new treatment strategy for this cancer, as well as a theoretical basis for the development of new drugs in the future.