Microglia regulate blood clearance in subarachnoid hemorrhage by heme oxygenase-1
Nils Schallner, … , Leo E. Otterbein, Khalid A. Hanafy
Nils Schallner, … , Leo E. Otterbein, Khalid A. Hanafy
Published May 26, 2015
Citation Information: J Clin Invest. 2015;125(7):2609-2625. https://doi.org/10.1172/JCI78443.
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Microglia regulate blood clearance in subarachnoid hemorrhage by heme oxygenase-1

Abstract

Subarachnoid hemorrhage (SAH) carries a 50% mortality rate. The extravasated erythrocytes that surround the brain contain heme, which, when released from damaged red blood cells, functions as a potent danger molecule that induces sterile tissue injury and organ dysfunction. Free heme is metabolized by heme oxygenase (HO), resulting in the generation of carbon monoxide (CO), a bioactive gas with potent immunomodulatory capabilities. Here, using a murine model of SAH, we demonstrated that expression of the inducible HO isoform (HO-1, encoded by Hmox1) in microglia is necessary to attenuate neuronal cell death, vasospasm, impaired cognitive function, and clearance of cerebral blood burden. Initiation of CO inhalation after SAH rescued the absence of microglial HO-1 and reduced injury by enhancing erythrophagocytosis. Evaluation of correlative human data revealed that patients with SAH have markedly higher HO-1 activity in cerebrospinal fluid (CSF) compared with that in patients with unruptured cerebral aneurysms. Furthermore, cisternal hematoma volume correlated with HO-1 activity and cytokine expression in the CSF of these patients. Collectively, we found that microglial HO-1 and the generation of CO are essential for effective elimination of blood and heme after SAH that otherwise leads to neuronal injury and cognitive dysfunction. Administration of CO may have potential as a therapeutic modality in patients with ruptured cerebral aneurysms.

Authors

Nils Schallner, Rambhau Pandit, Robert LeBlanc III, Ajith J. Thomas, Christopher S. Ogilvy, Brian S. Zuckerbraun, David Gallo, Leo E. Otterbein, Khalid A. Hanafy

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Figure 11

CO rescues HO-1 deficiency and impaired erythrophagocytosis, leading to increased cell death in vitro.

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CO rescues HO-1 deficiency and impaired erythrophagocytosis, leading to ...
(A) Quantification of the percentage of microglia positive for fluorescence-labeled red blood cells by flow cytometric analysis. *P = 0.0013 for LyzM-Cre Hmox1fl/fl red blood cells versus LyzM-Cre Hmox1fl/fl red blood cells plus CO (2-tailed Student’s t test; n = 3). (B) Mean fluorescent signal in microglial cells after exposure to fluorescence-labeled red blood cells. *P = 0.0154 for LyzM-Cre Hmox1fl/fl red blood cells versus LyzM-Cre Hmox1fl/fl red blood cells plus CO (2-tailed Student’s t test; n = 3). (C) Representative light microscopic images (original magnification, ×40; n = 3) of LyzM-Cre Hmox1fl/fl microglia after exposure to red blood cells with or without CO. (D) Quantification of red blood cells per microglia, analyzed by light microscopy. *P < 0.0001 for LyzM-Cre Hmox1fl/fl red blood cells versus LyzM-Cre Hmox1fl/fl red blood cells plus CO (2-tailed Student’s t test; n = 3). (E) Quantification of the percentage of red blood cell–positive microglia by light microscopy. *P = 0.0107 for LyzM-Cre Hmox1fl/fl red blood cells versus LyzM-Cre Hmox1fl/fl red blood cells plus CO (2-tailed Student’s t test; n = 3). (F) Representative images (original magnification, ×20; n = 3) of TUNEL staining (red) of neuronal cells cocultured with LyzM-Cre Hmox1fl/fl microglia with or without red blood cells and CO. Cell nuclei were counterstained with Hoechst 33258 (blue). (G) Quantification of TUNEL-positive neuronal cells per microscopic view. *P < 0.0001 for LyzM-Cre Hmox1fl/fl red blood cells versus LyzM-Cre Hmox1fl/fl red blood cells plus CO (2-tailed Student’s t test; n = 3).