TIM-4⁺ skeletal muscle Resident Tissue Macrophages Ferroptosis mediated Rhabdomyolysis in Exertional Heatstroke

Skeletal muscle resident tissue macrophages (smRTMs) are strategically positioned to sense myofiber injury and coordinate inflammatory responses, but their mechanistic contribution to exertional heatstroke (EHS)-associated rhabdomyolysis (RM) remains poorly defined. Here, we delineate a ferroptosis-dependent pathway in smRTMs that drives RM during EHS. Using mouse model of EHS, in combination with single-cell RNA sequencing, smRTM-targeted Hmox1 deletion and pharmacological modulation of ferroptosis and inflammasome activity, we identify a T cell membrane protein 4-positive (TIM-4⁺) smRTM subset as selectively vulnerable to ferroptosis. EHS robustly induces heme oxygenase-1 (HMOX1), iron-dependent lipid peroxidation and ferroptotic death in TIM-4⁺ smRTMs, accompanied by accumulation of the lipid peroxidation-derived aldehyde octanal. Octanal engages olfactory receptor 2 (Olfr2) and provides a proximal signal for activation of the NOD-like receptor family pyrin domain containing 3 (NLRP3) inflammasome, caspase-1 cleavage and interleukin-1β (IL-1β) release. Chromatin and functional assays further establish JunD as a transcription factor that binds the Olfr2 promoter and is required for Olfr2 upregulation downstream of HMOX1-driven ferroptosis. Genetic or pharmacological inhibition of HMOX1, or blockade of ferroptosis, reduces TIM-4⁺ smRTM ferroptosis, dampens the JunD-Olfr2-NLRP3-IL-1β axis and significantly attenuates RM in both species. These data identify HMOX1-dependent ferroptosis in TIM-4⁺ smRTMs as a central immunometabolic mechanism of EHS-associated RM and nominate smRTM ferroptosis and the Olfr2-NLRP3 pathway as rational therapeutic targets.