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Impact of 12/15-Lipoxygenase on Brain Injury After Subarachnoid Hemorrhage.

ABSTRACT: Background and Purpose- Subarachnoid hemorrhage (SAH) is a devastating form of stroke. Oxidative stress contributes to brain injury, but the mechanisms have been poorly studied. Here, we evaluated the role of 12/15-lipoxygenase (12/15-LOX), an enzyme known to cause cell death in ischemic stroke, on brain injury in a mouse model of SAH. Methods- C57Bl6 wild-type mice and Alox15 knockout mice were subjected to SAH using a direct blood injection technique. In SAH wild-type mice, half received the 12/15-LOX inhibitor ML351 and half received vehicle. Immunohistochemistry, brain edema, blood-brain barrier leakage and functional outcomes were assessed 1 and 3 days after SAH induction. Results- SAH led to increased 12/15-LOX in macrophages of the brain parenchyma, adjacent to the subarachnoid blood. Neuronal cell death after SAH was reduced by ML351 and in Alox15 knockout mice. Similarly, SAH induced brain edema, which was 12/15-LOX dependent. Finally, Alox15 gene knockout and inhibitor treatment in wild-type mice with SAH led to an improved behavioral outcome. Conclusions- 12/15-LOX is overexpressed in macrophages after SAH in mice, and inhibition of the 12/15-LOX pathway decreases brain injury and improves neurological outcome. This study suggests 12/15-LOX as a novel therapeutic target to limit brain injury after SAH.

SUBMITTER: Gaberel T

PROVIDER: S-EPMC6349484 | biostudies-literature | 2019 Feb

REPOSITORIES: biostudies-literature

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Impact of 12/15-Lipoxygenase on Brain Injury After Subarachnoid Hemorrhage.

Gaberel Thomas T Gakuba Clément C Zheng Yi Y Lépine Matthieu M Lo Eng H EH van Leyen Klaus K

Stroke 20190201 2

Background and Purpose- Subarachnoid hemorrhage (SAH) is a devastating form of stroke. Oxidative stress contributes to brain injury, but the mechanisms have been poorly studied. Here, we evaluated the role of 12/15-lipoxygenase (12/15-LOX), an enzyme known to cause cell death in ischemic stroke, on brain injury in a mouse model of SAH. Methods- C57Bl6 wild-type mice and Alox15 knockout mice were subjected to SAH using a direct blood injection technique. In SAH wild-type mice, half received the 1 ...[more]

PMID: 30602353

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Project description:BackgroundInnate immunity can facilitate early brain injury (EBI) following subarachnoid hemorrhage (SAH). Numerous studies suggest that pyroptosis could exacerbate extracellular immune responses by promoting secretion of inflammatory cytokines. Transforming growth factor-β-activated kinase 1 (TAK1) is a quintessential kinase that positively regulates inflammation through NF-κB and MAPK signaling cascades. However, the effects of TAK1 on neuroinflammation in EBI following SAH are largely unknown.MethodsTwo hundred and forty-six male C57BL/6J mice were subjected to the endovascular perforation model of SAH. A selective TAK1 inhibitor, 5Z-7-oxozeaenol (OZ) was administered by intracerebroventricular (i.c.v) injection at 30 min after SAH induction. To genetic knockdown of TAK1, small interfering RNA (siRNA) was i.c.v injected at 48 h before SAH induction. SAH grade, brain water content, BBB permeability, neurological score, western blot, real-time PCR, ELISA, transmission electron microscope, and immunofluorescence staining were performed. Long-term behavioral sequelae were evaluated by the rotarod and Morris water maze tests. Furthermore, OZ was added to the culture medium with oxyhemoglobin (OxyHb) to mimic SAH in vitro. The reactive oxygen species level was detected by DCFH-DA staining. Lysosomal integrity was assessed by Lyso-Tracker Red staining and Acridine Orange staining.ResultsThe neuronal phosphorylated TAK1 expression was upregulated following SAH. Pharmacologic inhibition of TAK1 with OZ could alleviate neurological deficits, brain edema, and brain-blood barrier (BBB) disruption at 24 h after SAH. In addition, OZ administration restored long-term neurobehavioral function. Furthermore, blockade of TAK1 dampened neuronal pyroptosis by downregulating the N-terminal fragment of GSDMD (GSDMD-N) expression and IL-1β/IL-18 production. Mechanistically, both in vivo and in vitro, we demonstrated that TAK1 can induce neuronal pyroptosis through promoting nuclear translocation of NF-κB p65 and activating nucleotide-binding oligomerization domain (NOD)-like receptor pyrin domain containing 3 (NLRP3) inflammasome. TAK1 siRNA treatment mitigated SAH-induced neurobehavioral deficits and restrained phosphorylated NF-κB p65 expression and NLRP3 inflammasome activation. TAK1 blockade also ameliorated reactive oxygen species (ROS) production and prevented lysosomal cathepsin B releasing into the cytoplasm.ConclusionsOur findings demonstrate that TAK1 modulates NLRP3-mediated neuronal pyroptosis in EBI following SAH. Inhibition of TAK1 may serve as a potential candidate to relieve neuroinflammatory responses triggered by SAH.

Project description:BACKGROUND:Proteomic analysis of cerebrospinal fluid (CSF) has shown great promise in identifying potential markers of injury in neurodegenerative diseases [1-13]. Here we compared CSF proteomes in healthy individuals, with patients diagnosed with traumatic brain injury (TBI) and subarachnoid hemorrhage (SAH) in order to characterize molecular biomarkers which might identify these different clinical states and describe different molecular mechanisms active in each disease state. METHODS:Patients presenting to the Neurosurgery service at the Louisiana State University Hospital-Shreveport with an admitting diagnosis of TBI or SAH were prospectively enrolled. Patients undergoing CSF sampling for diagnostic procedures were also enrolled as controls. CSF aliquots were subjected to 2-dimensional gel electrophoresis (2D GE) and spot percentage densities analyzed. Increased or decreased spot expression (compared to controls) was defined in terms of in spot percentages, with spots showing consistent expression change across TBI or SAH specimens being followed up by Matrix-Assisted Laser Desorption/Ionization mass spectrometry (MALDI-MS). Polypeptide masses generated were matched to known standards using a search of the NCBI and/or GenPept databases for protein matches. Eight hundred fifteen separately identifiable polypeptide migration spots were identified on 2D GE gels. MALDI-MS successfully identified 13 of 22 selected 2D GE spots as recognizable polypeptides. RESULTS:Statistically significant changes were noted in the expression of fibrinogen, carbonic anhydrase-I (CA-I), peroxiredoxin-2 (Prx-2), both ? and ? chains of hemoglobin, serotransferrin (Tf) and N-terminal haptoglobin (Hp) in TBI and SAH specimens, as compared to controls. The greatest mean fold change among all specimens was seen in CA-I and Hp at 30.7 and -25.7, respectively. TBI specimens trended toward greater mean increases in CA-I and Prx-2 and greater mean decreases in Hp and Tf. CONCLUSIONS:Consistent CSF elevation of CA-I and Prx-2 with concurrent depletion of Hp and Tf may represent a useful combination of biomarkers for the prediction of severity and prognosis following brain injury.

Dataset Information

Impact of 12/15-Lipoxygenase on Brain Injury After Subarachnoid Hemorrhage.

Publications

Impact of 12/15-Lipoxygenase on Brain Injury After Subarachnoid Hemorrhage.

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