Isofurans and isoprostanes as markers of delayed brain injury and mitochondrial dysfunction following aneurysmal subarachnoid hemorrhage. A prospective observational study

IntroductionF2-Isoprostanes (F2-IsoPs) and Isofurans (IsoF), specific markers of lipid peroxidation in vivo, have been reported to be elevated and have prognostic implications following subarachnoid hemorrhage (SAH). Platelet activation and vasoconstriction are attributed to these compounds. Elevated IsoF: F2-IsoPs ratios have been previously suggested as indicative of mitochondrial dysfunction. In this small study we examined their performance as specific biomarkers for delayed brain injury (DBI) development following SAH. We also explored if evidence of mitochondrial dysfunction could be found in a cohort of SAH patients.MethodsEighteen patients with SAH and 7 controls with normal neuroimaging and CSF analysis results underwent CSF sampling and abstraction of clinical, demographic, and laboratory data. Samples (two) of CSF were collected on day 1 and once on days 5-8 post-bleed. F2-IsoP and IsoF assays were performed at Vanderbilt Eicosanoid Core Lab by gas chromatography/mass spectroscopy. Levels are expressed in median (IQR) for non-parametric data. Repeated sample measurement were compared using the Wilcoxon signed-rank test, whereas the Mann Whitney test was used for other non-parametric data.ResultsMean age was 61.2 + 15.7 (SAH cases) vs. 47.6 + 10 (controls) years, and 80% of SAH patients were female. Median Hunt-Hess score was 3 (2-4) and modified Fisher scale 3 (3-4). Thirty nine percent of patients developed DBI. F2-IsoP were significantly higher in SAH cases than in controls [47.5 (30.2-53.5) vs. 26.0 (21.2-34.5) pg/mL]. No significant differences were observed in patients with or without DBI [41 (33.5-52) vs. 44 (28.5-55.5) pg/mL]. IsoF were elevated in the second CSF sample in 9 patients, but undetectable in the remainder cases and all controls. Patients who developed DBI had significantly higher IsoF than cases who did not [(57 (34-72) vs. 0 (0-34) pg/mL]. Patients who met criteria for delayed injury had a significantly higher IsoF: F2IsoPs ratio on the late CSF sample [1.03 (1-1.38) vs. 0 (0-0.52)].ConclusionsPreliminary findings from this study suggest that IsoF may represent a specific biomarker predicting DBI following SAH and provide possible evidence of CNS mitochondrial dysfunction in SAH. Future studies to further explore the value of IsoF as biomarkers of secondary brain injury and the contribution of mitochondrial dysfunction and ferroptosis to clinical outcomes following SAH seem warranted.

Melatonin affects hypoxia-inducible factor 1α and ameliorates delayed brain injury following subarachnoid hemorrhage via H19/miR-675/HIF1A/TLR4

Background: This study was aimed to investigate the molecular mechanism of how melatonin interferes with hypoxia-inducible factor 1α (HIF1A) and TLR4 expression, which is implicated in the management of delayed brain injury (DBI) after subarachnoid hemorrhage (SAH). Method: Luciferase assay, real-time PCR, Western-blot analysis and immunohistochemistry (IHC) assays were utilized to explore the interaction among H19, miR-675, HIF1A and TLR4, and to evaluate the effect of MT on the expression of above transcripts in different groups. Results: MT enhanced H19 expression by promoting the transcription efficiency of H19 promoter, and HIF1A was identified as a target of miR-675. HIF1A enhanced TLR4 expression via promoting the transcription efficiency of TLR4 promoter. Furthermore, administration of MT up-regulated miR-675 but suppressed the expressions of HIF1A and TLR4. Treatment with MT alleviated neurobehavioral deficits and apoptosis induced by SAH. According to the result of IHC, HIF1A and TLR4 protein levels in the SAH group were much higher than those in the SAH+MT group. Therefore, the administration of MT increased the levels of H19 and miR-675 which have been inhibited by SAH. In a similar way, treatment with MT decreased the levels of HIF1A and TLR4 which have been enhanced by SAH. Conclusion: MT could down-regulate the expression of HIF1A and TLR4 via the H19/miR-675/HIF1A/TLR4 signaling pathway, while TLR4 is crucial to the release of pro-inflammatory cytokines. Therefore, the treatment with MT could ameliorate post-SAH DBI.

New endovascular perforation subarachnoid hemorrhage model for investigating the mechanisms of delayed brain injury

OBJECTIVEDelayed brain injury (DBI) is considered one of the most important causes of mortality and morbidity after subarachnoid hemorrhage (SAH). However, no suitable experimental rat endovascular perforation (EVP) SAH model was available for investigating DBI. The authors added early cerebral hypoperfusion to a mild EVP SAH model by unilateral common carotid artery occlusion (UCCAO) 24 hours after induction of SAH to mimic the clinical course of early cerebral hypoperfusion after SAH.METHODSA total of 109 adult male Sprague-Dawley rats were randomly divided into 2 groups: no SAH and SAH. Next, no-SAH rats were randomly divided on day 1 into 2 groups: sham and UCCAO. SAH rats with a neurological score of 15 or greater were randomly divided into 2 groups: SAH − UCCAO and SAH + UCCAO group.RESULTSThe mild SAH model had a lower mortality rate of 5.4% within the first 24 hours. No rat died in the SAH + UCCAO group until day 7. DBI as well as early brain injury (EBI), reactive astrogliosis, and cerebral vasospasm significantly worsened in the SAH + UCCAO group.CONCLUSIONSThe present SAH + UCCAO model can simulate EBI with aggravation of reactive astrogliosis, cerebral vasospasm, and DBI but without high mortality.

Abstract TP259: Ischemic Postconditioning Protects Against Focal Ischemia in Mouse

Ischemic Postconditioning Protects Against Focal Ischemia in Mouse Background: Ischemic postconditioning(IPost) has been shown to attenuate cerebral ischemia/reperfusion injury in a few rat models. Here we further established a IPost model in mice, which offers oppotunities to study protective mechanism of IPost in mice with various genetic backgrounds. Material and Methods: Focal ischemia was induced by middle cerebral artery (MCA) suture occlusion for 45min. IPost was conducted by a series of repetitive, brief occlusion/reperfusion via withdrawl and insertion of the suture in the internal carotid artery. Infarction sizes were measured 3 days or 2 weeks after stroke. Functional neurological scores were also assessed. Results: we found that rapid IPost with three cycles of occlusion/reperfusion (15sec/30sec), or with two cycles of occlusion/reperfusion ( 60sec/60sec), which were performed 2 min after reperfusion, significantly reduced infarct sizes as measured 3 days post-stroke. Delayed IPost performed 3 hours after reperfusion aslo significantly reduced infarct sizes( n=9, P<0.01). Rapid IPost with 3 cycles of occlusion/reperfusion(15/30s) initiated 2 min after reperfusion resulted in the smallest infarct size. The spared infarct area were seen in border zones between the cortical territories of the anterior cerebral artery(ACA) and those of the MCA, as well as in the ventromedial and dorsolateral striatum. Delayed brain injury measured by silver staining 2 weeks after stroke and mortality were also reduced by rapid IPost. Conclusions: IPost reduces cerebral infarction and improve functional neurological status in mice, which offers a model to investigate the underlying protective mechanism of postconditioning using mice with various genetic backgrounds in the future.