Figure 8

Acetylation-dependent release of HMGB1 via its dissociation from SIRT1 is correlated with endotoxin toxicity.

(A,B,D) Tissues were prepared from BALB/c mice infected with Ad-Myc-SIRT1, Ad-Flag-HMGB1, and/or Ad-Flag-HMGB1^K282930R at a multiplicity of infection of 0.5 × 10¹⁰ via the tail vein, followed by the infusion of LPS or vehicle (5 mg/kg, i.p.) 3 days later. Interaction of HMGB1 and SIRT1 (A) were analyzed by Western blot and immunoprecipitation using whole-tissue lysates. Circulating levels of Flag-HMGB1 (B) and cytokines (D) were detected by Western blotting or ELISA, respectively, using sera prepared from samples collected at 16 h post-injection. Results are expressed as means ± standard error (n = 3 or 7). (C) BALB/c mice (n = 10–11 per group) were infected with Ad-Myc-SIRT1, Ad-Flag-HMGB1, and/or Ad-Flag-HMGB1^K282930R at a multiplicity of infection of 0.5 × 10¹⁰ via the tail vein, followed by a lethal infusion of endotoxin (LPS, 1 mg/kg, i.p.) 3 days later. Survival was monitored daily for up to 2 weeks. (E) Schematic representation of acetylation-dependent interaction of HMGB1 and SIRT1. ^*p < 0.01 compared with the Ad-LacZ-infected group; ^#p < 0.01 and ^##p < 0.05 compared with Ad-LacZ + LPS-treated group; ^†p < 0.01 and ^††p < 0.05 compared with the Ad-Flag-HMGB1 + LPS-treated group; ^‡p < 0.01 compared with the Ad-Flag-HMGB1 + Ad-Myc-SIRT1 + LPS-treated group; ^∮p < 0.001 compared with the Ad-Flag-HMGB1 + Ad-Myc-SIRT1 + LPS-treated group; and ^∬p < 0.001 compared with the Ad-Flag-HMGB1^K282930R + LPS-treated group.