Figure 6.

HDAC9 knockdown alleviates ferroptotic neuronal death by blocking the increase of HIF-1 binding on TfR1 gene and by enhancing Sp1 binding on GPX4 gene in the periischemic temporoparietal cortex of tMCAO mice. A-C, Representative Western blot and quantification of HDAC9 (A), HIF-1alpha (B), and Sp1 (C) protein levels in temporoparietal cortex of mice after tMCAO or sham operated animals (Sham) (n = 5/6); *, p ≤ 0.05 versus Sham. D, G, Representative Western blot and quantification of HIF-1alpha (D) and Sp1 (G) protein levels in temporoparietal cortex of sham operated mice or mice subjected to 24h of tMCAO and treated with siHDAC9 or scrambled siRNA (siCTL) (n=5); *, p ≤ 0.05 versus Sham; #, p ≤ 0.05 versus siCTL after 24h of tMCAO. E, H, Chromatin immunoprecipitation (ChIP) of HIF-1 alpha on TfR1 promoter (E) and Sp1 on GPX4 promoter (H) performed in temporoparietal cortex of sham operated mice or mice subjected to 24h of tMCAO and treated with siHDAC9 or siCTL. Anti-IgG antibody has been used as negative control (n = 3); *, p ≤ 0.05 versus Sham, #, p ≤ 0.05 versus siCTL after 24h of tMCAO. F, I, Representative Western blot and quantification of TfR1 (F) and GPX4 (I) protein levels in temporoparietal cortex of sham operated mice or mice subjected to 24h of tMCAO and treated with siHDAC9 or siCTL (n = 5); *, p ≤ 0.05 versus Sham; #, p ≤ 0.05 versus siCTL after 24h of tMCAO. J, K, quantification of relative Fe²⁺ (J) and 4-HNE (K) levels in temporoparietal cortex of sham operated mice or mice subjected to 24h of tMCAO and treated with siHDAC9 or scrambled siRNA (n = 5); *, p ≤ 0.05 versus Sham; #, p ≤ 0.05 versus siCTL after 24h of tMCAO. Bars represent mean ± SD.