Investigation and modulation of interleukin-6 following subarachnoid hemorrhage: targeting inflammatory activation for cerebral vasospasm.

BACKGROUND: Cerebral vasospasm (CV) can contribute to significant morbidity in subarachnoid hemorrhage (SAH) patients. A key unknown is how CV induction is triggered following SAH. METHODS: Human aneurysmal blood and cerebral spinal fluid were collected for evaluation. To confirm mechanism, c57/bl6 wild type and c57/bl6 IL-6 female knockout (KO) mice were utilized with groups: saline injected, SAH, SAH + IL-6 blockade, SAH IL-6 KO, SAH IL-6 KO + IL-6 administration, SAH + p-STAT3 inhibition. Dual-labeled microglia/myeloid mice were used to show myeloid diapedesis. For SAH, 50 µm blood was collected from tail puncture and administered into basal cisterns. IL-6 blockade was given at various time points. Various markers of neuroinflammation were measured with western blot and immunohistochemistry. Cerebral blood flow was also measured. Vasospasm was measured via cardiac injection of India ink/gelatin. Turning test and Garcia's modified SAH score were utilized. P < 0.05 was considered significant. RESULTS: IL-6 expression peaked 3 days following SAH (p < 0.05). Human IL-6 was increased in aneurysmal blood (p < 0.05) and in cerebral spinal fluid (p < 0.01). Receptor upregulation was periventricular and perivascular. Microglia activation following SAH resulted in increased caveolin 3 and myeloid diapedesis. A significant increase in BBB markers endothelin 1 and occludin was noted following SAH, but reduced with IL-6 blockade (p < 0.01). CV occurred 5 days post-SAH, but was absent in IL-6 KO mice and mitigated with IL-6 blockade (p < 0.05). IL-6 blockade, and IL-6 KO mitigated effects of SAH on cerebral blood flow (p < 0.05). SAH mice had impaired performance on turn test and poor modified Garcia scores compared to saline and IL-6 blockade. A distinct microglia phenotype was noted day 5 in the SAH group (overlap coefficients r = 0.96 and r = 0.94) for Arg1 and iNOS, which was altered by IL-6 blockade. Day 7, a significant increase in toll-like receptor 4 and Stat3 was noted. This was mitigated by IL-6 blockade and IL-6 KO, which also reduced Caspase 3 (p < 0.05). To confirm the mechanism, we developed a p-STAT3 inhibitor that targets the IL-6 pathway and this reduced NFΚB, TLR4, and nitrotyrosine (p < 0.001). Ventricular dilation and increased Tunel positivity was noted day 9, but resolved by IL-6 blockade (p < 0.05). CONCLUSION: Correlation between IL-6 and CV has been well documented. We show that a mechanistic connection exists via the p-STAT3 pathway, and IL-6 blockade provides benefit in reducing CV and its consequences mediated by myeloid cell origin diapedesis.

Scavenging Free Iron Reduces Arteriolar Microvasospasms After Experimental Subarachnoid Hemorrhage.

BACKGROUND AND PURPOSE: Subarachnoid hemorrhage (SAH) is associated with acute and delayed cerebral ischemia resulting in high acute mortality and severe chronic neurological deficits. Spasms of the pial and intraparenchymal microcirculation (microvasospasms) contribute to acute cerebral ischemia after SAH; however, the underlying mechanisms remain unknown. We hypothesize that free iron (Fe3+) released from hemolytic red blood cells into the subarachnoid space may be involved in microvasospasms formation. METHODS: Male C57BL/6 mice (n=8/group) received 200 mg/kg of the iron scavenger deferoxamine or vehicle intravenously and were then subjected to SAH by filament perforation. Microvasospasms of pial and intraparenchymal vessels were imaged three hours after SAH by in vivo 2-photon microscopy. RESULTS: Microvasospasms occurred in all investigated vessel categories down to the capillary level. Deferoxamine significantly reduced the number of microvasospasms after experimental SAH. The effect was almost exclusively observed in larger pial arterioles (>30 µm) covered with blood. CONCLUSIONS: These results provide proof-of-principle evidence that Fe3+ is involved in the formation of arteriolar microvasospasms after SAH and that arteriolar and capillary microvasospasms are triggered by different mechanisms. Deciphering the mechanisms of Fe3+-induced microvasospasms may result in novel therapeutic strategies for SAH patients.

Peroxisome proliferator­activated receptor ß/δ regulates cerebral vasospasm after subarachnoid hemorrhage via modulating vascular smooth muscle cells phenotypic conversion.

Cerebral vasospasm (CVS) is a common complication of subarachnoid hemorrhage (SAH) with high deformity rates and cerebral vascular smooth muscle cells (VSMCs) phenotypic switch is considered to be involved in the regulation of CVS. However, to the best of the authors' knowledge, its underlying molecular mechanism remains to be elucidated. Peroxisome proliferator­activated receptor ß/δ (PPARß/δ) has been demonstrated to be involved in the modulation of vascular cells proliferation and maintains the autoregulation function of blood vessels. The present study investigated the potential effect of PPARß/δ on CVS following SAH. A model of SAH was established by endovascular perforation on male adult Sprague­Dawley rats, and the adenovirus PPARß/δ (Ad­PPARß/δ) was injected via intracerebroventricular administration prior to SAH. The expression levels of phenotypic markers α­smooth muscle actin and embryonic smooth muscle myosin heavy chain were measured via western blotting or immunofluorescence staining. The basilar artery diameter and vessel wall thickness were evaluated under fluorescence microscopy. SAH grade, neurological scores, brain water content and brain swelling were measured to study the mechanisms of PPARß/δ on vascular smooth muscle phenotypic transformation. It was revealed that the expression levels of synthetic proteins were upregulated in rats with SAH and this was accompanied by CVS. Activation of PPARß/δ using Ad­PPARß/δ markedly upregulated the contractile proteins elevation, restrained the synthetic proteins expression and attenuated SAH­induced CVS by regulating the phenotypic switch in VSMCs at 72 h following SAH. Furthermore, the preliminary study demonstrated that PPARß/δ downregulated ERK activity and decreased the expression of phosphorylated (p­)ETS domain­containing protein Elk­1 and p­p90 ribosomal S6 kinase, which have been demonstrated to serve an important role in VSMC phenotypic change. Additionally, it was revealed that Ad­PPARß/δ could positively improve CVS by ameliorating the diameter of the basilar artery and mitigating the thickness of the vascular wall. Furthermore, subsequent experiments demonstrated that Ad­PPARß/δ markedly reduced the brain water content and brain swelling and improved the neurological outcome. Taken together, the present study identified PPARß/δ as a useful regulator for the VSMCs phenotypic switch and attenuating CVS following SAH, thereby providing novel insights into the therapeutic strategies of delayed cerebral ischemia.

The calcimimetic R-568 attenuates subarachnoid hemorrhage-induced vasospasm through PI3K/Akt/eNOS signaling pathway in the rat model.

Cerebral vasospasm (CVS) causes mortality and morbidity in patients after subarachnoid hemorrhage (SAH). The mechanism and adequate treatment of CVS are still elusive. R-568 is a calcimimetic agent known to exert a vasodilating effect. However, there is no report on its vasodilator effect against SAH-induced vasospasm. In the present study, we investigated the therapeutic effect of R-568 on the SAH-induced CVS model in rats. Seventy-two adult male Sprague-Dawley rats were divided into 8 groups: sham surgery; SAH only; SAH + Vehicle, SAH + R-568; SAH + R-568 + Wortmannin (the PI3K inhibitor); SAH + Wortmannin; SAH + R-568 + Calhex-231 (a calcilytic agent); SAH + Calhex-231. SAH was induced by blood (0.3 mL) given by intracisternal injection. R-568 (20 µM) was administered intracisternal immediately prior to experimental SAH. Basilar arteries (BAs) were obtained to evaluate PI3K/Akt/eNOS pathway (immunoblotting) and morphological changes 48 h after SAH. Perimeters of BAs were decreased by 24.1% in the SAH group compared to the control group and the wall thickness was increased by 75.3%. With R-568 treatment, those percentages were 9.6% and 29.6%, respectively, indicating that vasospasm was considerably improved when compared with the SAH group (P < 0.001 in both). While p-PI3K/PI3K and p-Akt/Akt ratio and eNOS protein expression were markedly decreased in the SAH rats, treatment with R-568 resulted in a significant increase in these levels. The beneficial effects of R-568 were partially blocked in the presence of Calhex-231 and completely blocked in the presence of Wortmannin. Herein, we found that treatment with R-568 would attenuate SAH-induced CVS through the PI3K/Akt/eNOS pathway and demonstrate therapeutic promise in CVS treatment following SAH.

SIRT1 mediates hypoxic preconditioning induced attenuation of neurovascular dysfunction following subarachnoid hemorrhage.

BACKGROUND AND PURPOSE: Vasospasm and delayed cerebral ischemia (DCI) contribute significantly to the morbidity/mortality associated with aneurysmal subarachnoid hemorrhage (SAH). While considerable research effort has focused on preventing or reversing vasospasm, SAH-induced brain injury occurs in response to a multitude of concomitantly acting pathophysiologic mechanisms. In this regard, the pleiotropic epigenetic responses to conditioning-based therapeutics may provide an ideal SAH therapeutic strategy. We previously documented the ability of hypoxic preconditioning (PC) to attenuate vasospasm and neurological deficits after SAH, in a manner that depends on the activity of endothelial nitric oxide synthase. The present study was undertaken to elucidate whether the NAD-dependent protein deacetylase sirtuin isoform SIRT1 is an upstream mediator of hypoxic PC-induced protection, and to assess the efficacy of the SIRT1-activating polyphenol Resveratrol as a pharmacologic preconditioning therapy. METHODS: Wild-type C57BL/6J mice were utilized in the study and subjected to normoxia or hypoxic PC. Surgical procedures included induction of SAH via endovascular perforation or sham surgery. Multiple endpoints were assessed including cerebral vasospasm, neurobehavioral deficits, SIRT1 expression via quantitative real-time PCR for mRNA, and western blot for protein quantification. Pharmacological agents utilized in the study include EX-527 (SIRT1 inhibitor), and Resveratrol (SIRT1 activator). RESULTS: Hypoxic PC leads to rapid and sustained increase in cerebral SIRT1 mRNA and protein expression. SIRT1 inhibition blocks the protective effects of hypoxic PC on vasospasm and neurological deficits. Resveratrol pretreatment dose-dependently abrogates vasospasm and attenuates neurological deficits following SAH - beneficial effects that were similarly blocked by pharmacologic inhibition of SIRT1. CONCLUSION: SIRT1 mediates hypoxic preconditioning-induced protection against neurovascular dysfunction after SAH. Resveratrol mimics this neurovascular protection, at least in part, via SIRT1. Activation of SIRT1 is a promising, novel, pleiotropic therapeutic strategy to combat DCI after SAH.

Biospecimens and Molecular and Cellular Biomarkers in Aneurysmal Subarachnoid Hemorrhage Studies: Common Data Elements and Standard Reporting Recommendations.

INTRODUCTION: Development of clinical biomarkers to guide therapy is an important unmet need in aneurysmal subarachnoid hemorrhage (SAH). A wide spectrum of plausible biomarkers has been reported for SAH, but none have been validated due to significant variabilities in study design, methodology, laboratory techniques, and outcome endpoints. METHODS: A systematic review of SAH biomarkers was performed per the Preferred Reporting Items for Systematic Reviews and Meta-Analyses (PRISMA) guidelines. The panel's recommendations focused on harmonization of (1) target cellular and molecular biomarkers for future investigation in SAH, (2) standardization of best-practice procedures in biospecimen and biomarker studies, and (3) experimental method reporting requirements to facilitate meta-analyses and future validation of putative biomarkers. RESULTS: No cellular or molecular biomarker has been validated for inclusion as "core" recommendation. Fifty-four studies met inclusion criteria and generated 33 supplemental and emerging biomarker targets. Core recommendations include best-practice protocols for biospecimen collection and handling as well as standardized reporting guidelines to capture the heterogeneity and variabilities in experimental methodologies and biomarker analyses platforms. CONCLUSION: Significant variabilities in study design, methodology, laboratory techniques, and outcome endpoints exist in SAH biomarker studies and present significant barriers toward validation and translation of putative biomarkers to clinical use. Adaptation of common data elements, recommended biospecimen protocols, and reporting guidelines will reduce heterogeneity and facilitate future meta-analyses and development of validated clinical biomarkers in SAH.

Increased expression of thymic stromal lymphopoietin receptor in a rat model of middle cerebral artery occlusion.

Thymic stromal lymphopoietin (TSLP) is an epithelial cell-derived cytokine expressed in the skin, gut, lungs, and thymus. TSLP triggers dendritic cell-mediated T helper 2 inflammatory responses by formation of a ternary complex consisting of a heterodimer of interleukin-7 (IL-7) receptor α chain (IL-7Rα), TSLP, and the TSLP receptor chain (TSLPR). The present study aimed to investigate the expression of this ternary complex and its interaction with signal transducer and activator of transcription 5 (STAT5) in a ischemic stroke model using middle cerebral artery occlusion. Using immunofluorescence staining, we found that TSLPR was expressed widely in neurons and gliocytes. Using immunoprecipitation analysis, we detected an increased interaction between STAT5 and the ternary complex in the cortex of stroke rats. Moreover, using western blots, we found that expressions of the ternary complex and STAT5 were markedly increased in the cortex of stroke rats compared with the control and sham rats. These results suggest that the formation of the ternary TSLPR : TSLP : IL-7Rα complex may activate STAT5 or a STAT5-related signaling pathway to mediate neuroinflammation in ischemic stroke.

The relationship between IL-6, ET-1 and cerebral vasospasm, in experimental rabbit subarachnoid hemorrhage.

BACKGROUND: In subarachnoid hemorrhage (SAH), occurrence of cerebral vasospasm (CVS) mediated by endothelin (ET)-1 might be a result of a compartmental inflammatory response with interleukin (IL)-6 release. We aim to investigate the relationship between ET-1 and IL-6 in association of CVS. METHODS: A total of 24 New Zealand white rabbits where randomly allocated into 3 groups: SAH (N.=10), IL-6 (N.=10), and sham (N.=4). SAH was induced by a closed cranium extracranial-intracranial shunt model. In the IL-6 group, IL-6 was injected into the cisterna magna. CVS of the basilar artery was assessed by digital subtraction angiography. IL-6 and ET-1 concentrations were measured using enzyme-linked immunosorbent assay. Neuronal damage was assessed by terminal deoxynucleotidyl transferase dUTP nick end labeling. RESULTS: A significant increase between baseline (day 0) and follow-up (day 3) was found in CSF IL-6 levels of animals in the SAH and IL-6 group. There was a statistically significant correlation between IL-6 and ET-1 levels in the CSF (Pearson's r=0.454, P=0.003). CVS at day 3 was more pronounced in the SAH than in the IL-6 group: 26.0 ±7.2 % and 16.7 ±5.0 % respectively. TUNEL positive apoptotic neurons in the hippocampal formation were present in the SAH group and in a lesser degree in the IL-6 group. CONCLUSIONS: The results indicate that IL-6 triggered CVS after SAH is ET-1 dependent. IL-6 may be a target for new therapeutic approaches.

Therapeutically Targeting Platelet-Derived Growth Factor-Mediated Signaling Underlying the Pathogenesis of Subarachnoid Hemorrhage-Related Vasospasm.

INTRODUCTION: Vasospasm accounts for a large fraction of the morbidity and mortality burden in patients sustaining subarachnoid hemorrhage (SAH). Platelet-derived growth factor (PDGF)-ß levels rise following SAH and correlate with incidence and severity of vasospasm. METHODS: The literature was reviewed for studies investigating the role of PDGF in the pathogenesis of SAH-related vasospasm and efficacy of pharmacological interventions targeting the PDGF pathway in ameliorating the same and improving clinical outcomes. RESULTS: Release of blood under high pressure into the subarachnoid space activates the complement cascade, which results in release of PDGF. Abluminal contact of blood with cerebral vessels increases their contractile response to PDGF-ß and thrombin, with the latter upregulating PDGF-ß receptors and augmenting effects of PDGF-ß. PDGF-ß figures prominently in the early and late phases of post-SAH vasospasm. PDGF-ß binding to the PDGF receptor-ß results in receptor tyrosine kinase domain activation and consequent stimulation of intracellular signaling pathways, including p38 mitogen-activated protein kinase, phosphatidylinositol-3-kinase, Rho-associated protein kinase, and extracellular regulated kinase 1 and 2. Consequent increases in intracellular calcium and increased expression of genes mediating cellular growth and proliferation mediate PDGF-induced augmentation of vascular smooth muscle cell contractility, hypertrophy, and proliferation. CONCLUSION: Treatments with statins, serine protease inhibitors, and small molecular pathway inhibitors have demonstrated varying degrees of efficacy in prevention of cerebral vasospasm, which is improved with earlier institution.

Effects of Tenascin-C Knockout on Cerebral Vasospasm After Experimental Subarachnoid Hemorrhage in Mice.

A matricellular protein tenascin-C (TNC) has been suggested to play a role in the pathogenesis of cerebral vasospasm after subarachnoid hemorrhage (SAH), but the direct evidence remains lacking. In this study, we examined effects of TNC knockout (TNKO) on cerebral vasospasm after experimental SAH in mice. C57BL/6 wild-type (WT) or TNKO mice were subjected to SAH by endovascular puncture. Ten WT and ten TNKO mice were randomized to WT sham (n = 4), TNKO sham (n = 4), WT SAH (n = 6), and TNKO SAH (n = 6) groups. In addition to neurobehavioral impairments and severity of SAH, cerebral vasospasm was assessed by morphometric measurements of the left internal carotid artery (ICA). Infiltration of inflammatory cells in the subarachnoid periarterial space was also assessed, and expressions of TNC and mitogen-activated protein kinases (MAPKs) in the ICA were immunohistochemically evaluated at 24 h post-surgery. TNC was induced in the smooth muscle cell layers and the adventitia in the spastic ICAs as well as the periarterial inflammatory cells in WT SAH mice. Compared with WT SAH mice, TNKO SAH mice showed better neurological scores and less severe cerebral vasospasm, as well as fewer inflammatory cell infiltration in the periarterial space. Post-SAH activation of MAPKs in the smooth muscle cell layers of the ICAs was also prevented in TNKO SAH mice. The findings in the present study suggest that TNC causes the development of cerebral vasospasm via pro-inflammatory effects and activation of MAPKs.

Soluble Fms-Like Tyrosine Kinase 1 (sFlt-1) and Risk of Cerebral Vasospasm After Aneurysmal Subarachnoid Hemorrhage.

BACKGROUND: The molecular mechanisms underlying cerebral vasospasm and delayed cerebral ischemia (DCI) after aneurysmal subarachnoid hemorrhage (aSAH) are incompletely understood. We hypothesized that circulating antiangiogenic factors, such as soluble Fms-like tyrosine kinase 1 (sFlt-1) and soluble transforming growth factor ß coreceptor, soluble endoglin (sEng), are important markers of their pathophysiology. METHODS: We performed a prospective study in patients with aSAH and measured cerebrospinal fluid and serum levels of sFlt-1 and sEng on postbleed day 1 and 6 and correlated levels with incidence and severity of cerebral vasospasm and DCI. RESULTS: Twenty-seven patients with aSAH were enrolled in the study. Severe angiographic vasospasm was present in 14.8% of patients and DCI occurred in 33.3%. Serum sFlt1 levels were increased on postbleed day 6 in patients who developed vasospasm. However, on postbleed day 1, there were no differences in patients who developed vasospasm. Increased serum sFlt-1 levels on postbleed day 1 were found to predict the development of severe angiographic vasospasm with an area under the curve of 0.818 with an optimal cutoff value of 95 pg/mL. Alterations in sFlt1 were not associated with DCI. Serum and cerebrospinal fluid sEng levels did not correlate with vasospasm or DCI. CONCLUSIONS: Serum levels of sFlt-1 are increased in patients with aSAH who are at risk for severe vasospasm. Further studies with larger sample sizes are needed to evaluate whether sFlt-1 levels may predict onset of severe vasospasm and DCI.

Activation of Cytotoxic Natural Killer Cells After Aneurysmal Subarachnoid Hemorrhage.

OBJECTIVE: Cell-mediated inflammation is critical in the development of cerebrovascular complications after aneurysmal subarachnoid hemorrhage. We analyzed the course for activated CD16brightCD56dim cytotoxic natural killer (NK) cells in cerebrospinal fluid of 15 patients. METHODS: Patients were classified by occurrence of cerebral vasospasm (CV) and delayed cerebral ischemia. NK were monitored by flow cytometry between day 1 and 14 after hemorrhage. RESULTS: Twelve patients (80%) developed CV with a mean day of detection at 3.9 ± 1.6. In those patients, cell count for NK increased from 1.40 ± 1.42 cells/µL on day 1 to a peak of 11.66 ± 11.56 cells/µL on day 6.1 ± 2.9 (P = 0.001). An increase of mean cerebral blood flow velocity in transcranial Doppler from 71.33 ± 12.93 cm/second to 166.20 ± 20.19 cm/second (P < 0.01) and an increase in number of vascular axes affected by CV was detected (P < 0.01). In patients with grade 3 CV (n = 4, 33.3%), activated NK counts were significantly higher than in patients who did not have CV (23.18 ± 13.92 cells/µL vs. 0.02 ± 0.01 cells/µL; P = 0.029). NK counts were significantly different between patients with grade 1 and grade 3 CV (P = 0.04). Patients who did not have CV who showed low NK counts achieved better functional outcome (Glasgow Outcome Scale [GOS] score, 4.6 ± 0.6) at discharge than did patients with CV grade 2 (GOS score, 3.3 ± 0.5) and CV grade 3 (GOS score, 2.3 ± 0.5) who showed increased NK cell counts (CV grade 0 vs. CV grade 2, P = 0.048; CV grade 0 vs. CV grade 3, P = 0.001). Activated CD16brightCD56dim cytotoxic NKCSF cell counts showed a mean maximum (14.15 ± 12.21 cells/µL) when delayed cerebral ischemia occurred. CONCLUSIONS: The increase of activated CD16brightCD56dim cytotoxic NK cells in cerebrospinal fluid after aneurysmal subarachnoid hemorrhage suggests an increased risk of CV and delayed cerebral ischemia.

Pharmacokinetic Modeling of Intra-arterial Nimodipine Therapy for Subarachnoid Hemorrhage-Related Cerebral Vasospasm.

PURPOSE: Intra-arterial (IA) administration of nimodipine has been shown to be an effective treatment for subarachnoid hemorrhage-related cerebral vasospasm. The concentrations achieved in cerebral arteries during this procedure, though, are unknown. Therefore, there are no clinical studies investigating dose-dependent effects of nimodipine. We aimed at providing a pharmacokinetic model for IA nimodipine therapy for this purpose. METHODS: A two-compartment pharmacokinetic model for intravenous nimodipine therapy was modified and used to assess cerebral arterial nimodipine concentration during IA nimodipine infusion into the internal carotid artery (ICA). RESULTS: According to our simulations, continuous IA nimodipine infusion at 2 mg/h and 1 mg/h resulted in steady-state cerebral arterial concentrations of about 200 ng/ml and 100 ng/ml assuming an ICA blood flow of 200 ml/min and a clearance of 70 l/h. About 85 % of the maximal concentration is achieved within the first minute of IA infusion independent on the infusion dose. Within the range of physiological and pharmacokinetic data available in the literature, ICA blood flow has more impact on cerebral arterial concentration than nimodipine clearance. CONCLUSION: The presented pharmacokinetic model is suitable for estimations of cerebral arterial nimodipine concentration during IA infusion. It may, for instance, assist in dose-dependent analyses of angiographic results.

Epidermal growth factor-like repeats of tenascin-C-induced constriction of cerebral arteries via activation of epidermal growth factor receptors in rats.

Tenascin-C (TNC), one of matricellular proteins, has been suggested to be involved in cerebral vasospasm after aneurysmal subarachnoid hemorrhage. However, the mechanisms of how TNC constricts cerebral arteries remain unclear. The aim of this study was to examine if epidermal growth factor (EGF)-like repeats of TNC is involved in TNC-induced constriction of cerebral arteries in rats via EGF receptor (EGFR) activation. Two dosages of recombinant TNC (r-TNC) consisting of the EGF-like repeats was administered intracisternally to healthy rats, and its vasoconstrictor effects were evaluated by neurobehavioral tests and India-ink angiography at 24, 48, and 72 hours after the administration. Western blotting and immunohistochemistry were performed to explore the underlying mechanisms on constricted cerebral arteries after 24 hours. The effects of a selective EGFR tyrosine kinase inhibitor (AG1478) on r-TNC-induced vasoconstriction were evaluated by neurobehavioral tests, India-ink angiography and immunohistochemistry at 24 hours after the administration. A higher dosage of r-TNC induced cerebral arterial constriction more severely, which continued for 48 hours. The effects were associated with the activation of EGFR and extracellular signal-regulated kinase (ERK)1/2 in the smooth muscle cell layer of the constricted cerebral artery, while c-Jun N-terminal kinase and p38 were not activated. AG1478 blocked r-TNC-induced vasoconstrictive effects, as well as activation of EGFR and ERK1/2. These findings demonstrate that TNC induces constriction of cerebral arteries via activation of EGFR and ERK1/2.

Signaling Pathway in Cerebral Vasospasm After Subarachnoid Hemorrhage: News Update.

Subarachnoid hemorrhage (SAH) caused by ruptured intracranial aneurysms is a serious threat to human health and life. Although advances in surgical and anesthetic techniques have improved the prognosis of patients with aneurysms, the rate of death and disability caused by SAH remains high, predominantly due to cerebral vasospasm (CVS) after SAH and early brain damage. In particular, CVS is a common complication after SAH. However, its pathogenesis has not yet been fully elucidated, and clinically effective prevention and treatment measures are still lacking. Spasm of blood vessels can decrease cerebral blood flow, leading to ischemia and hypoxia in brain tissues and ultimately severe neurological dysfunction. Currently, there is no comprehensive theory that can fully explain the mechanisms underlying SAH-caused CVS. However, studies on signal transduction, apoptosis, and glial cell-mediated mechanisms in recent years have shed new light on the treatment of CVS.

Expression of Sphingosine-1-phosphate (S1P) on the cerebral vasospasm after subarachnoid hemorrhage in rabbits

PURPOSE:To demonstrate the relationship between of sphingosine-1-phosphate (S1P) expression and subarachnoid hemorrhage (SAH).METHODS:The basilar arteries from a "double-hemorrhage" rabbit model of SAH were used to investigate the relation between S1P expression and SAH. Various symptoms, including blood clots, basilar artery cross-sectional area, and S1P phosphatase expression were measured at day 3, 5, 7, 9.RESULTS: The expression of S1P was enhanced in the cerebral vasospasm after subarachnoid hemorrhage in the rabbits. And S1P expression was consistent with the basilar artery cross-sectional area changes at day 3, 5, 7, 9.CONCLUSION: Sphingosine-1-phosphate expression in the cerebral arterial may be a new indicator in the development of cerebral vasospasm after subarachnoid hemorrhage and provide a new therapeutic method for SAH.

Alterations of the thioredoxin system during subarachnoid hemorrhage-induced cerebral vasospasm.

BACKGROUND: The exact underlying pathogenic mechanisms and effective preventive or therapeutic interventions for cerebral vasospasm remain obscure. The thioredoxin (Trx) system performs important functions in the central nervous system including neurotrophic and neuroprotective actions. There is no study directly investigating the effects of subarachnoid hemorrhage (SAH) induced cerebral vasospasm on the Trx system in the literature. METHODS: Sixteen male New Zealand rabbits were randomly divided into two groups of eight rabbits each: a control group and a SAH group. The control group, (n = 8) was a sham surgery group in which SAH was not induced. In the SAH group, (n = 8), the SAH protocol was used to induce cerebral vasospasm. The brain and brainstem were removed and each brainstem was cut coronally into two pieces: an anterior part that contains basilar artery and a dorsal part that contains brainstem tissue. The brainstem tissue thioredoxin-1(Trx1), thioredoxin-2 (Trx2), thioredoxin reductase (TrxR), thioredoxin reductase-1 (TrxR1), thioredoxin-interacting protein (TXNIP) levels were investigated. Total oxidant status (TOS), total antioxidant status (TAS), malondialdehyde levels (MDA) and tumor necrosis factor alpha (TNF-alpha) levels were investigated for determining the oxidative-antioxidative status of the related brain tissues. Basilar artery segments were investigated for cross-sectional area and wall thickness measurements. RESULTS: SAH statistically significantly reduced the tissue levels of Trx1 (p < 0.01) and TrxR (p < 0.01). Trx2 levels were not significantly altered after SAH (p > 0.05). SAH significantly reduced the expression of TrxR1 (p < 0.01) and significantly increased the expression of TXNIP (p < 0.01) when compared with controls. TOS levels and MDA levels significantly increased after SAH (p < 0.01) and TAS levels significantly reduced after SAH (p < 0.01). TNF-alpha levels significantly increased after SAH (p < 0.01). SAH-induced cerebral vasospasm significantly (p < 0.05) increased the wall thickness and reduced the mean cross-sectional area of the basilar artery (p < 0.05). CONCLUSIONS: The Trx system seems to be negatively affected by the simultaneously interrelated enzymatic alterations during cerebral vasospasm.

Early diagnosis of cerebral ischemia in cerebral vasospasm by oxygen-pulse near-infrared optical topography.

PURPOSE: Early diagnosis of vasospasm is a key factor in the choice of treatment after subarachnoid hemorrhage (SAH). However, a noninvasive method of diagnosing delayed ischemic neurological deficit (DIND) has not been established. We therefore propose a new method of diagnosing cerebral ischemia using near-infrared optical topography (OT) with oxygen inhalation. MATERIALS AND METHODS: We used a 44-channel OT system that covers the bilateral front otemporoparietal areas to assess 29 patients who underwent surgery within 72 h of the onset of SAH. The patients inhaled room air followed by oxygen for 2 min, and then peripheral oxygen saturation (SpO2) was continuously monitored at the index fingertip. The patients were assessed by N-isopropyl-p-[¹²³I]iodoamphetamine (IMP)-SPECT and OT on the same day. Ischemic findings were confirmed using principal component analysis with reference to the systemic SpO2value. RESULTS: Seven of 29 patients developed DIND. Evidence of ischemia was identified by OT in all seven of these patients before the onset of DIND. The OT and SPECT findings agreed in 27 (93 %) of the 29 patients. DISCUSSION AND CONCLUSIONS: Our method might detect cerebral ischemia before the onset of DIND and thus be clinically useful for assessing cerebral ischemia with vasospasm.

Protective effect of stellate ganglion block on delayed cerebral vasospasm in an experimental rat model of subarachnoid hemorrhage.

Stellate ganglion block (SGB) is a blockade of sympathetic ganglia innervating the head and neck, and is known to function through vasodilation of the target region. However, the effectiveness of SGB in relieving cerebral vasospasm (CVS) through dilation of intracerebral vessels has not been evaluated. The aim of the present study is to investigate the therapeutic effects of SGB in a rat model of subarachnoid hemorrhage (SAH) complicated by delayed CVS, and explore the underlying mechanisms. The SAH model was established by double injection of autologous arterial blood into the cisterna magna. We simulated SGB by transection of the cervical sympathetic trunk (TCST), and measured changes in the diameter, perimeter and cross-sectional area of the basilar artery (BA) and middle cerebral artery (MCA) to evaluate its vasodilatory effect. To investigate the underlying mechanisms, we determined the expression level of vasoactive molecules endothelin-1 (ET-1) and calcitonin gene-related peptide (CGRP) in the plasma, and apoptotic modulators Bcl-2 and Bax in the hippocampus. We found a significant increase in the diameter, perimeter and cross-sectional area of the BA and right MCA in SAH rats subjected to TCST. Application of SGB significantly reduced the expression of ET-1 while increasing that of CGRP in SAH rats. We also found a significant increase in the expression of Bcl-2 and decrease in the expression of Bax in the hippocampus of SAH rats subjected to TCST, when compared to untreated SAH rats. The mechanism of action of SGB is likely mediated through alterations in the ratio of ET-1 and CGRP, and Bax and Bcl-2. These results suggest that SGB can alleviate the severity of delayed CVS by inducing dilation of intracerebral blood vessels, and promoting anti-apoptotic signaling. Our findings provide evidence supporting the use of SGB as an effective and well-tolerated approach to the treatment of CVS in various clinical settings.