KLF9-Mediated Transcriptional Promotion of HMGB2 Accelerates Cardiomyocyte Apoptosis, Inflammation, and Ferroptosis in Myocardial Ischemia/Reperfusion Injury

Cardiovasc Toxicol. 2025 Jun 19. doi: 10.1007/s12012-025-10028-0. Online ahead of print.

Abstract

High-mobility group protein B2 (HMGB2) has been confirmed to participate in regulating the process of myocardial ischemia/reperfusion (I/R) injury. However, the more roles and mechanisms of HMGB2 in myocardial I/R injury need to be further revealed. Cardiomyocytes (HL-1) were cultured under hypoxia/reoxygenation (H/R) conditions, and myocardial I/R injury mouse model was established by ligation of the left anterior descending coronary artery. The protein levels of HMGB2 and kruppel-like factor 9 (KLF9) were determined by western blot. Cell viability and apoptosis were examined by CCK8 assay and flow cytometry. The levels of inflammatory factors and ferroptosis-related markers were tested to assess cell inflammation and ferroptosis. The interaction between KLF9 and HMGB2 promoter was evaluated by ChIP assay and dual-luciferase reporter assay. HMGB2 was higher expressed in H/R-induced HL-1 cells and its silencing could suppress H/R-induced HL-1 cell apoptosis, inflammation, and ferroptosis. KLF9 had binding sites in HMGB2 promoter region, which could increase HMGB2 expression by enhancing its transcription. Silencing of KLF9 inhibited H/R-induced HL-1 cell apoptosis, inflammation, and ferroptosis, while these effects were reversed by overexpressing HMGB2. In addition, animal study revealed that interference of KLF9 alleviated myocardial tissues damage and fibrosis in I/R injury mice models by reducing HMGB2 expression. Collectively, our study indicated that KLF9 promoted myocardial I/R injury by aggravating cardiomyocyte apoptosis, inflammation, and ferroptosis through promoting HMGB2 transcription.

Keywords: HMGB2; KLF9; Myocardial ischemia/reperfusion injury.