Bilirubin production and oxidation in CSF of patients with cerebral vasospasm after subarachnoid hemorrhage

Delayed cerebral vasospasm after subarachnoid hemorrhage (SAH) remains a significant cause of mortality and morbidity, however, the etiology is, as yet, unknown, despite intensive research efforts. Research in this laboratory indicates that bilirubin and oxidative stress may be responsible by leading to formation of bilirubin oxidation products (BOXes), so we investigated changes in bilirubin concentration and oxidative stress in vitro, and in cerebral spinal fluid (CSF) from SAH patients. Non-SAH CSF, a source of heme oxygenase I (HO-1), and blood were incubated, and in vitro bilirubin production measured. CSF from SAH patients was collected, categorized using stimulation of vascular smooth muscle metabolism in vitro (CSFv and CSFc for with vasospasm and without vasospasm respectively). CSF was analyzed for hemoglobin, total protein and bilirubin, BOXes, malonyldialdehyde and peroxidized lipids (indicators of an oxidizing environment), and HO-1 concentration. The study patients comprised 7 males and 5 females. The median age was 46, and the ages ranged from 32 to 72 years old at time of hemorrhage. The CSFc samples averaged 0.92 ± 0.31 g/dL total protein and 0.39 ± 0.06 g/dL hemoglobin, compared with 0.94 ± 0.37 g/dL and 0.41 ± 0.07 g/dL in the CSFv samples. HO-1 concentrations are significantly higher (p < 0.05) in CSFv, compared with CSFc. When bilirubin concentration is compared for patients with and without clinical vasospasm, there is significantly greater bilirubin in the vasospasm patients (5.6 ± 0.93 vs. 29.4 ± 3.9 μmols/L). We found that total peroxidized lipids (LOOH) is significantly elevated in vasospasm patients compared to control (18.24 ± 4.4 vs. 7.5 ± 0.53 umols/L respectively). We have found that the concentration of BOXes in CSFv of patients following SAH is significantly elevated compared to CSFc (1.33 ± 0.07 versus 0.02 ± 0.004 uM respectively). Bilirubin, BOXes, HO-1 and peroxidized lipid content were significantly higher in CSF from SAH patients with vasospasm, compared with non-vasospasm SAH CSF, and correlated with occurrence of vasospasm. We conclude that vasospasm may be more likely in patients with elevated BOXes. The conditions necessary for the formation of BOXes are indeed present in CSF from SAH patients with vasospasm, but not CSF from SAH patients without vasospasm.