Puerarin triggers sensitivity to ferroptosis in glioblastoma cells by activating SIRT3/NCOA4-dependent autophagy

Abstract

SIRT3 has been found to involve in the tumorigenesis and progression of glioblastoma, and it is reported that puerarin can inhibit the growth of glioblastoma cells. Therefore, we aimed to investigate the biological function of SIRT3 in autophagy and ferroptosis in glioblastoma cells and study the effects of puerarin on ferroptosis and SIRT3/NCOA4-dependent autophagy in cancer cells. The results showed that overexpression of SIRT3 significantly promoted ferroptosis sensitization by reducing cell viability and GSH/GSSH ratio and increasing ROS levels, whereas knockout of SIRT3 significantly triggered cell viability and GSH/GSSH ratio and decreasing ROS levels in U87MG cells (P < 0.05). Moreover, overexpression of SIRT3 significantly also promoted the ratio of LC3-Ⅱ/Ⅰ and upregulated NCOA4 and Fe²⁺ levels, but downregulated the expression of p62 and FTH (P < 0.05); while knockout of SIRT3 has the opposite effects. Besides, autophagy inhibitor antagonized the effects of SIRT3 overexpression on the expression of autophagy-associated proteins and Fe²⁺ levels. Additionally, knockout of NCOA4 significantly increased cell viability and GSH/GSSH ratio and reduced ROS levels in RSL3-treated cells overexpressing SIRT3. Puerarin treatment significantly upregulated the levels of ROS, Fe²⁺, SIRT3, NCOA4, and the ratio of LC3-Ⅱ/Ⅰ, but downregulated the levels of cell viability, GSH/GSSH ratio, p62, and FTH in U87MG cells (all P < 0.05). Autophagy inhibitor, SIRT3 or NCOA4 deletion significantly reduced the effects of puerarin on the autophagy-dependent ferroptosis in U87MG cells (all P < 0.05). SIRT3 drives sensitivity to ferroptosis by activating NCOA4-mediated autophagy, and we proposed puerarin, a promising therapeutic drug for glioblastoma.