Gentiopicroside and swertiamarin induce non-selective oxidative stress-mediated cytotoxic effects in human peripheral blood mononuclear cells

Abstract

Gentiopicroside (Gp) and swertiamarin (Sm), secoiridoid glycosides commonly found in plants of the Gentianaceae family, differ in one functional group. They exhibit promising cytotoxic effects in cancer cell lines and overall protective outcomes, marking them as promising molecules for developing novel pharmaceuticals. To investigate potential variations in cellular sensitivity to compounds of similar molecular structures, we analyzed the mode of Gp and Sm induced cell death in human peripheral blood mononuclear cells (PBMCs) after 48 h of treatment. The lowest tested concentration that significantly reduces cell viability, 50 μM, was applied. Oxidative stress parameters were estimated by measuring the levels of prooxidative/antioxidative balance, lipid peroxidation products, and 8-oxo-7,8-dihydro-2-deoxyguanosine, while gene expression of DNA repair enzymes was evaluated by employing quantitative real-time PCR. Cellular morphology was analyzed by fluorescent microscopy, and immunoblot analysis of apoptosis and necroptosis-related proteins was used to assess the type of cell death induced by the treatments. The discriminatory impact of Gp/Sm treatments on apoptosis and necroptosis-induced cell death was evaluated by monitoring the cell survival in co-treatment with specific cell death inhibitors. Obtained results show greater cytotoxicity of Gp than Sm suggesting that variations in the molecular structures of the tested compounds can substantially affect their biological effects. Gp/Sm co-treatment with apoptosis and necroptosis inhibitors revealed a distinct, albeit non-specific mechanism of PBMCs cell death. Although the therapeutic may not directly cause a specific type of cell death, its extent can be pivotal in assessing the safety of therapeutic application and developing phytopharmaceuticals with improved features. Since phytopharmaceuticals affect all exposed cells, identification of cytotoxic mechanisms on PBMCs after Gp and Sm treatment is important for addressing the formulation and dosage of potential phytopharmaceuticals.