Dihydrotanshinone I inhibits human glioma cell proliferation via the activation of ferroptosis

Abstract

The aim of the present study was to investigate the effect of dihydrotanshinone I (DHI) on the survival of human glioma cells and the expression levels of ferroptosis-associated proteins. Human U251 and U87 glioma cells were cultured in vitro and treated with different concentrations of DHI and/or the ferroptosis inhibitor ferrostatin-1. A Cell Counting Kit-8 assay was used to determine the cell survival rate. The cells were further analyzed to determine their 5-, 12- and 15-hydroxyeicosatetraenoic acid (HETE), lactate dehydroge- nase (LDH) and malondialdehyde (MDA) levels, and reduced glutathione (GSH)/oxidized glutathione (GSSG) ratios. Western blotting was used to detect ferroptosis-associated glutathione peroxidase 4 (GPX4) and long-chain acyl-CoA synthetase 4 (ACSL-4). Changes in the mitochondrial membrane potential (MMP) were also observed using tetramethylrhodamine methyl ester staining and confocal fluo- rescence microscopy. The results revealed that DHI inhibited the proliferation of human glioma cells. Following treatment of the U251 and U87 cells with DHI, changes in the expres- sion levels of ferroptosis-associated proteins were observed; the expression level of GPX4 decreased and that of ACSL-4 increased. DHI also increased the levels of LDH and MDA in the human glioma cells and reduced the GSH/GSSG ratio. The DHI-treated cells also exhibited a marked reduction in MMP. Furthermore, ferrostatin-1 blocked the DHI-induced effects in human glioma cells. From these results, it may be concluded that DHI inhibits the proliferation of human glioma cells via the induction of ferroptosis.