Acute ischemic stroke (AIS) is a major cerebrovascular disorder caused by the abrupt interruption of cerebral blood flow, resulting in acute focal neurological deficits [1]. Despite advances in acute stroke care, AIS remains a leading cause of death and long-term disability worldwide [2,3]. Current evidence-based reperfusion strategies, including intravenous thrombolysis with recombinant tissue plasminogen activator (r-tPA) and endovascular mechanical thrombectomy, can effectively restore cerebral perfusion [4,5]. However, a substantial proportion of patients still fail to achieve favorable recovery after reperfusion [6], and clinical benefit is often limited by cerebral ischemia–reperfusion (I/R)-associated neurovascular injury [7]. These observations highlight that AIS pathophysiology is not solely determined by vascular occlusion itself, but also by a complex multicellular network of secondary injury responses [8].

A central hub within this network is the blood–brain barrier (BBB), which serves as a critical interface between the circulation and the brain parenchyma and is among the earliest targets of injury after AIS. Cerebral I/R disrupts endothelial tight junctions (TJs), allowing plasma components and water to extravasate into the parenchyma, thereby promoting vasogenic edema and secondary tissue injury [9,10]. Barrier breakdown also rapidly activates innate neuroimmune responses, as microglia, astrocytes, and pericytes release pro-inflammatory cytokines and chemokines that amplify inflammatory cascades and aggravate neuronal damage [11,12]. Thus, BBB disruption is not only an early pathological hallmark of cerebral I/R, but also an upstream event that drives downstream neurovascular injury programs [13,14]. Importantly, endothelial cells, the principal cellular component of the BBB, also exert angiocrine functions by regulating post-stroke inflammation, tissue repair, and neuroregeneration through paracrine and contact-dependent signaling [15,16].

Consistent with this multicellular stress response, clinical metabolomics studies have revealed broad remodeling of the circulating metabolome in AIS, including recurrent alterations in glycerophospholipid and sphingolipid metabolism, amino acid pathways such as branched-chain amino acid and tryptophan–kynurenine metabolism, and energy/redox-related modules including lactate–pyruvate balance and glutathione/NADPH-associated signatures [17]. Within this metabolic context, membrane phospholipid–derived lipid mediators are increasingly recognized as rapid effector systems that translate ischemic stress into neurovascular dysfunction [18]. Under pathological conditions, endothelial secretory outputs shift toward the release of bioactive lipid mediators, coupling barrier destabilization to inflammatory amplification and redox imbalance, thereby forming a feed-forward loop that accelerates neuronal injury [19,20].

Among these lipid-mediator pathways, arachidonic acid (AA) is a major ω-6 polyunsaturated fatty acid esterified in membrane phospholipids and a key substrate mobilized during cellular stress [21]. Upon ischemic or inflammatory stimulation, AA is released and metabolized through the lipoxygenase (LOX) pathway to generate bioactive eicosanoids that regulate inflammation, immune signaling, and vascular homeostasis [22,23]. ALOX12 (12-LOX) catalyzes the conversion of AA to 12(S)-hydroperoxyeicosatetraenoic acid [12(S)-HpETE], which is subsequently reduced by glutathione peroxidases (GPxs) to 12(S)-hydroxyeicosatetraenoic acid [12(S)-HETE; 12-HETE] [24]. In particular, endothelial stress may engage the ALOX12–12-HETE pathway as an angiocrine signaling route, promoting ROS-related redox imbalance, disruption of tight-junction integrity, and amplification of secondary inflammatory injury within the neurovascular unit [25,26].

Although ALOX12 has been implicated in multiple pathological conditions, including myocardial, hepatic, and renal I/R injury, as well as diabetes [27], its cell-type-specific induction, temporal dynamics, and functional relevance in AIS remain incompletely understood. In the present study, we sought to define the endothelial ALOX12–12-HETE axis as a mechanistically relevant mediator of neurovascular injury after cerebral I/R by linking its activation to BBB disruption, inflammatory amplification, and redox imbalance. We further tested whether ALOX12 inhibition mitigates neurovascular injury and improves functional outcomes. In parallel, we evaluated whether circulating 12-HETE is associated with disease severity and may serve as a peripheral marker candidate in AIS.