Skeletal muscle ischaemia-reperfusion (I/R) injury involves complex redox dysregulation with limited treatments. Although ferroptosis contributes to other organ I/R injuries, its role and regulation in skeletal muscle remain unclear. This study aimed to investigate the role and regulatory mechanism of ferroptosis in skeletal muscle I/R injury, specifically focusing on whether hypoxia-inducible factor 1 alpha (HIF1A) transcriptionally activates cyclin-dependent kinase inhibitor 1a (CDKN1A/p21) to drive this process.

We employed integrative transcriptomics and Cleavage Under Targets and Tagmentation (CUT&Tag, a chromatin mapping technique) sequencing in murine I/R models. Genetic inhibition (Hif1a siRNA) and pharmacological inhibition (LW6) were utilized in vitro and in vivo. Cdkn1a overexpression was performed for rescue experiments. Ferroptosis was assessed by examining mitochondrial ultrastructure, quantifying lipid peroxidation, and evaluating the expression of key proteins: glutathione peroxidase 4 (GPX4), acyl-CoA synthetase long-chain family member 4 (ACSL4), and prostaglandin-endoperoxide synthase 2 (PTGS2). Clinical relevance was evaluated by co-expression analysis of HIF1A and CDKN1A in human I/R-affected muscle biopsies.

HIF1A directly bound to the Cdkn1a promoter during nuclear translocation, upregulating its expression. Both HIF1A inhibition (genetic or pharmacological) significantly attenuated ferroptosis, evidenced by preserved mitochondria, reduced lipid peroxidation, and normalized ferroptosis-related protein levels. Crucially, Cdkn1a overexpression reversed the anti-ferroptotic effects of Hif1a knockdown, confirming CDKN1A as a key downstream effector. Strong positive co-expression of HIF1A and CDKN1A was observed in human I/R biopsies (Spearman's r = 0.543, p = 0.006). Mechanistically, the HIF1A-CDKN1A axis exacerbated redox stress via glutathione depletion and intracellular free iron accumulation.

Our findings establish HIF1A as a context-dependent ferroptosis amplifier in skeletal muscle I/R injury, acting through direct transcriptional activation of Cdkn1a. This HIF1A-CDKN1A axis drives ferroptosis by disrupting redox homeostasis. Targeting HIF1A and CDKN1A within this pathway provides two complementary molecular entry points for mitigating skeletal muscle I/R injury.

Targeting the HIF1A-CDKN1A axis offers a promising therapeutic approach to reduce skeletal muscle damage and improve clinical outcomes after ischaemia-reperfusion injury.

CDKN1A

Ferroptosis

HIF1A

Ischaemia-reperfusion injury

Skeletal muscle

Transcriptional regulation

© 2026 The Authors. Published by Elsevier B.V. on behalf of Chinese Speaking Orthopaedic Society.