Fig. 2: Inhibition of LDH activity suppressed hemin-induced neuronal ferroptosis in vitro.

A After the indicated treatment of PCNs for 12 h, the levels of H3K14la, H3K9me3, and H4K5la were examined using Western blotting. H3 serves as a loading control. (H3K14la: n = 5 cultures; H3K9me3: n = 4 cultures; H4K5la: n = 3 cultures). BODIPY 581/591 C11 reagent was used to detect the changes of intracellular lipid ROS after 24 h treatment with 50 µM hemin and 50 µM GSK, and the cell death was detected by PI and Hoechst staining (B). The quantifications for cell death and the intracellular lipid ROS are shown (C). (lipid ROS: n = 3 cultures; PI: n = 6 cultures). D After LDHA and LDHB were silenced by siRNA, the expression of H3K14la was detected by Western blotting at 12 h after 50 µM hemin treatment in PCNs. n = 10 cultures. E After transfection with siRNA of LDHA and LDHB, the lipid ROS of 50 µM hemin-treated PCNs were detected at 24 h by BODIPY 581/ 591 C11 reagent. n = 7 cultures. F After transfection with siRNA of LDHA and LDHB, the cell viability of hemin-treated PCNs was detected at 24 h by PI and Hoechst staining. n = 6 cultures. Results are shown as scatter plots (Mean ± SD). One-way ANOVA followed by Tukey’s multiple comparison tests (A, C), or two-way ANOVA followed by Sidak (D–F) multiple comparison tests was used. *p < 0.05, **p < 0.01, ***p < 0.001 vs Veh; #p < 0.05, ##p < 0.01 vs Hemin; NS not significant.