Cerebrospinal fluid hemoglobin drives subarachnoid hemorrhage-related secondary brain injury

Secondary brain injury after aneurysmal subarachnoid hemorrhage (SAH-SBI) is a significant contributor to poor outcomes in patients after rupture of an intracranial aneurysm. The lack of diagnostic biomarkers and novel drug targets represent an unmet need. Prior experimental evidence has suggested cell-free hemoglobin in the cerebrospinal fluid (CSF-Hb) as a pathophysiological driver of SAH-SBI. The aim of this study was to investigate the clinical and pathophysiological association between CSF-Hb and SAH-SBI. We prospectively enrolled 47 consecutive patients and collected daily CSF samples within 14 days after aneurysm rupture. There was very strong evidence for a positive association between CSF-Hb and SAH-SBI. The diagnostic accuracy of CSF-Hb for SAH-SBI markedly exceeded that of established methods (area under the curve: 0.89 [0.85-0.92]). Temporal LC-MS/MS CSF proteomics demonstrated that erythrolysis accompanied by an adaptive macrophage response are the two dominant biological processes occurring in the CSF space after aneurysm rupture. To further investigate the pathophysiology between CSF-Hb and SAH-SBI, we explored the vasoconstrictive and lipid peroxidation activities of Hb ex-vivo. These experiments revealed critical inflection points overlapping CSF-Hb concentration thresholds in patients with SAH-SBI. Selective Hb depletion and in-solution neutralization by the Hb-scavenger haptoglobin or the heme-scavenger hemopexin efficiently attenuated the vasoconstrictive and lipid peroxidation activities of CSF-Hb in patient CSF. Collectively, the clinical association between high CSF-Hb levels and SAH-SBI, the underlying pathophysiological rationale, and the favorable effects of haptoglobin and hemopexin in ex-vivo experiments position CSF-Hb as a highly attractive biomarker and potential drug target.