Haptoglobin and Hemopexin Redirect Heme-Driven Oxidative Stress and Neurotoxicity in Organotypic Brain Slices
The mechanistic links between hemoglobin oxidation, heme release, and neuronal death after intracerebral hemorrhage are incompletely defined. Using organotypic brain-slice cultures (BSCs) that model the RBC-toxin cascade, we dissect how the scavenger proteins haptoglobin (Hp) and hemopexin (Hpx) interrupt this pathology. BSCs exposed to free hemoglobin, heme, or their complexes (HbHp, hemeHpx) reveal that both Hb and heme drive iron-dependent neuronal injury, while Hp and Hpx confer protection by distinct mechanisms. Multi-modal readouts – including iron quantification, lipid peroxidation, neuronal apoptosis, and RNA-seq – demonstrate that Hpx abolishes toxicity by sequestering heme, whereas Hp stabilizes Hb and enables Nrf2-dependent cytoprotective gene expression. These data quantitatively map how Hp and Hpx reroute hemolytic toxicity toward resilience. Overall design: RNA-seq profiling of wildtype C57BL/6J organotypic brain slice cultures exposed to free hemoglobin, free heme, or their respective scavenger complexes (haptoglobin for hemoglobin, hemopexin for heme).