To validate a novel "Pre-Reperfusion HBOC Intervention" strategy, a cross-disciplinary initiative bridging hematology's oxygen carriers with neurology's penumbra salvage, for mitigating cerebral ischemia-reperfusion injury, and to investigate its underlying mechanism focused on ferroptosis regulation.

Effective neuroprotection in acute ischemic stroke is challenged by the narrow therapeutic window of traditional agents and their single-target nature. We posited that a hemoglobin-based oxygen carrier (HBOC), administered prior to recanalization, could offer a unique cross-disciplinary solution by sustaining the ischemic penumbra through oxygen delivery, potentially via a multi-target mechanism involving ferroptosis.

Sprague-Dawley rats were randomly assigned to HBOC or saline control groups (n=6/group) in a blinded tMCAO model. Treatment was administered intravenously 5 minutes post-ischemia, followed by reperfusion at 1 hour. We assessed safety (serum biochemistry, multi-organ histopathology, hemorrhagic transformation), infarct volume (MRI, TTC), and ultrastructural integrity (mitochondrial, axonal, and myelin integrity via TEM/immunofluorescence). Mechanisms were investigated by quantifying ferroptosis markers (e.g., MDA, GSH, GPX4, Fe²⁺), neuroinflammation (cytokines, microglial polarization), and behavioral outcomes.

HBOC administration proved safe and did not increase hemorrhagic transformation. This ultra-early intervention significantly reduced infarct volume by 23% (P=0.047) and robustly improved motor function. It also preserved neuronal density, blood-brain barrier integrity, and ameliorated damage to myelin and mitochondria. Mechanistically, HBOC enhanced antioxidant capacity and suppressed lipid peroxidation and iron dysmetabolism, indicating ferroptosis inhibition. Concomitant immunomodulation was observed, characterized by decreased pro-inflammatory and increased anti-inflammatory cytokines, alongside a shift in microglial polarization toward the M2 phenotype.

The pre-reperfusion HBOC strategy protects against cerebral IRI by inhibiting lipid peroxidation, mitigating ferroptosis, and modulating neuroinflammation. This work proposes a novel "Pre-hospital HBOC first-aid + Recanalization" paradigm, offering a promising translatable solution to critical time constraints in acute stroke care.