Abstract
Ferroptotic cell death is characterized by iron-dependent lipid peroxidation that is initiated by ferrous iron and H2O2 via Fenton reaction, in which the role of activating transcription factor 3 (ATF3) remains elusive. Brucine is a weak alkaline indole alkaloid extracted from the seeds of Strychnos nux-vomica, which has shown potent antitumor activity against various tumors, including glioma. In this study, we showed that brucine inhibited glioma cell growth in vitro and in vivo, which was paralleled by nuclear translocation of ATF3, lipid peroxidation, and increases of iron and H2O2. Furthermore, brucine-induced lipid peroxidation was inhibited or exacerbated when intracellular iron was chelated by deferoxamine (500 μM) or improved by ferric ammonium citrate (500 μM). Suppression of lipid peroxidation with lipophilic antioxidants ferrostatin-1 (50 μM) or liproxstatin-1 (30 μM) rescued brucine-induced glioma cell death. Moreover, knockdown of ATF3 prevented brucine-induced accumulation of iron and H2O2 and glioma cell death. We revealed that brucine induced ATF3 upregulation and translocation into nuclei via activation of ER stress. ATF3 promoted brucine-induced H2O2 accumulation via upregulating NOX4 and SOD1 to generate H2O2 on one hand, and downregulating catalase and xCT to prevent H2O2 degradation on the other hand. H2O2 then contributed to brucine-triggered iron increase and transferrin receptor upregulation, as well as lipid peroxidation. This was further verified by treating glioma cells with exogenous H2O2 alone. Moreover, H2O2 reversely exacerbated brucine-induced ER stress. Taken together, ATF3 contributes to brucine-induced glioma cell ferroptosis via increasing H2O2 and iron.
Highlights
Gliomas are the most common type of malignant brain tumors with higher mortality
Intracellular iron level is mainly regulated by transferrin receptor (TFR) which accounts for transporting extracellular iron–TF complex into cells via clathrin-mediated endocytosis, ferritin composed of light chain and heavy chain and responsible for storing iron, and ferroportin (FPN) in charge of iron exportation [5]
activating transcription factor 3 (ATF3) improved H2O2 by upregulating NADPH oxidase 4 (NOX4) and superoxide dismutase 1 (SOD1) and downregulating xCT and catalase To clarify the role of ATF3 in promotion of brucine-induced increase of H2O2, we examined brucine-induced changes in cysteine that could be converted from cystine and used as a material used for GSH synthesis
Summary
Gliomas are the most common type of malignant brain tumors with higher mortality. The median survival of the patients with high-grade glioma is not longer than 14.6 months, even if they accept surgery and are treated with chemotherapy and radiotherapy [1]. Ferroptosis is a newly established type of programmed necrosis and implicated in multiple pathological processes such as neurodegenerative diseases, carcinogenesis, ischemia reperfusion, brain trauma, and cerebral bleeding [3]. It is featured by presence of mitochondria with condensed mitochondrial membrane densities and decreased size [4]. Intracellular iron level is mainly regulated by transferrin receptor (TFR) which accounts for transporting extracellular iron–TF complex into cells via clathrin-mediated endocytosis, ferritin composed of light chain and heavy chain and responsible for storing iron, and ferroportin (FPN) in charge of iron exportation [5]
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