Haptoglobin administration into the subarachnoid space prevents hemoglobin-induced cerebral vasospasm
Michael Hugelshofer, … , Emanuela Keller, Dominik J. Schaer
Michael Hugelshofer, … , Emanuela Keller, Dominik J. Schaer
Published December 2, 2019; First published August 27, 2019
Citation Information: J Clin Invest. 2019;129(12):5219-5235. https://doi.org/10.1172/JCI130630.
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Research Article Neuroscience Vascular biology

Haptoglobin administration into the subarachnoid space prevents hemoglobin-induced cerebral vasospasm

Abstract

Delayed ischemic neurological deficit (DIND) is a major driver of adverse outcomes in patients with aneurysmal subarachnoid hemorrhage (aSAH), defining an unmet need for therapeutic development. Cell-free hemoglobin that is released from erythrocytes into the cerebrospinal fluid (CSF) is suggested to cause vasoconstriction and neuronal toxicity, and correlates with the occurrence of DIND. Cell-free hemoglobin in the CSF of patients with aSAH disrupted dilatory NO signaling ex vivo in cerebral arteries, which shifted vascular tone balance from dilation to constriction. We found that selective removal of hemoglobin from patient CSF with a haptoglobin-affinity column or its sequestration in a soluble hemoglobin-haptoglobin complex was sufficient to restore physiological vascular responses. In a sheep model, administration of haptoglobin into the CSF inhibited hemoglobin-induced cerebral vasospasm and preserved vascular NO signaling. We identified 2 pathways of hemoglobin delocalization from CSF into the brain parenchyma and into the NO-sensitive compartment of small cerebral arteries. Both pathways were critical for hemoglobin toxicity and were interrupted by the large hemoglobin-haptoglobin complex that inhibited spatial requirements for hemoglobin reactions with NO in tissues. Collectively, our data show that compartmentalization of hemoglobin by haptoglobin provides a novel framework for innovation aimed at reducing hemoglobin-driven neurological damage after subarachnoid bleeding.

Authors

Michael Hugelshofer, Raphael M. Buzzi, Christian A. Schaer, Henning Richter, Kevin Akeret, Vania Anagnostakou, Leila Mahmoudi, Raphael Vaccani, Florence Vallelian, Jeremy W. Deuel, Peter W. Kronen, Zsolt Kulcsar, Luca Regli, Jin Hyen Baek, Ivan S. Pires, Andre F. Palmer, Matthias Dennler, Rok Humar, Paul W. Buehler, Patrick R. Kircher, Emanuela Keller, Dominik J. Schaer

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Figure 3

Hb-induced vasospasm of cerebral arteries and protection by haptoglobin.

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Hb-induced vasospasm of cerebral arteries and protection by haptoglobin....
(A) SEC elution profiles of CSF collected from the subarachnoid space at baseline (blue), after intraventricular infusion of Hb at t = 0 minutes (red), and after infusion of haptoglobin at t = 90 minutes (green). Standard elution profiles are shown on the top. (B) Angiograms of the middle cerebral artery (MCA) at baseline, 45 minutes after Hb infusion (t = 45 minutes) and 45 minutes after haptoglobin infusion (t = 135 minutes). The square in the far-right image indicates the position of the zoom-images. DSA images were obtained with a ×1.5 magnification. (C) Relative change in diameter of cerebral arteries 60 minutes after infusion of Hb or Hb-haptoglobin. ACA, anterior cerebral artery; BA, basilar artery; ICA, internal carotid artery. Diamonds represent the mean and the 95% confidence interval (n = 4 sheep per group). (D) Cumulative analysis of the relative diameter changes of analyzed arterial segments 60 minutes after infusion of aCSF (n = 32), Hb (n = 16), or Hb-haptoglobin (n = 16). Colors represent vascular areas defined in C. Group means were compared by 1-way ANOVA, P < 0.001. The scheme depicts the experimental protocol. (E) NO-mediated relaxation profiles (n = 8 per condition) of porcine basilar arteries immersed in sheep CSF collected after the 60-minute posttreatment angiograms. Blue line: CSF samples after infusion of artificial CSF (CSF-heme 0 μM). Red line: CSF samples after infusion of Hb (CSF-heme 200–240 μM). Green line: CSF samples after infusion of Hb-haptoglobin (CSF-heme 200–240 μM). Thick lines represent the treatment group means. Dilatory responses were induced with a single bolus of MAHMA-NONOate. (F) Addition of equimolar haptoglobin to the Hb-containing sheep CSF restored the dilatative response to MAHMA-NONOate. Red line: before haptoglobin addition, same CSF as in E. Black line: after haptoglobin addition. Thick lines represent treatment group means of relaxation profiles (n = 8).