Macrophages play a critical role in cerebral aneurysm formation and rupture. The purpose of this study is to demonstrate the feasibility and optimal parameters of imaging macrophages within human cerebral aneurysm wall using ferumoxytol-enhanced-MRI.
Methods and Results
19 unruptured aneurysms in 11 patients were imaged using T2*-GE-MRI sequence. Two protocols were utilized. Protocol A: infusion of 2.5mg/kg of ferumoxytol and imaging at day 0 and 1. Protocol B: infusion of 5mg/kg of ferumoxytol and imaging at day 0 and 3. All images were reviewed independently by two neuroradiologists to assess for ferumoxytol-associated loss of MRI signal intensity within aneurysm wall. Aneurysm tissue was harvested for histologic analysis.
Fifty percent(5/10) of aneurysms in protocol A showed ferumoxytol-associated signal changes in aneurysm walls compared to 78% (7/9) of aneurysms in protocol B. Aneurysm tissue harvested from patients infused with ferumoxytol stained positive for both CD68+, demonstrating macrophage infiltration, and Prussian-Blue, demonstrating uptake of iron particles. Tissue harvested from controls stained positive for CD68 but not Prussian-Blue.
Imaging with T2*-GE-MRI at 72 hours post-infusion of 5mg/kg of ferumoxytol establishes a valid and useful approximation of optimal dose and timing parameters for macrophages imaging within aneurysm wall. Further studies are needed to correlate these imaging findings with risk of intracranial aneurysm rupture.
Intracranial Aneurysm; Inflammation; Magnetic Resonance Imaging; Ferumoxytol; USPIO
Many significant microsurgical series of patients with giant aneurysms predate changes in practice during the endovascular era.
A contemporary surgical experience is presented to examine changes in management relative to earlier reports, to establish the role of open microsurgery in the management strategy, and to quantify results for comparison with evolving endovascular therapies.
During a 13-year period, 140 patients with 141 giant aneurysms were treated surgically. 100 aneurysms (71%) were located in the anterior circulation, and 41 aneurysms were located in the posterior circulation.
108 aneurysms (77%) were completely occluded, 14 aneurysms (10%) had minimal residual aneurysm, and 16 aneurysms (11%) were incompletely occluded with reversed or diminished flow. 3 patients with calcified aneurysms were coiled after unsuccessful clipping attempts. 18 patients died in the perioperative period (surgical mortality, 13%). Bypass-related complications resulted from bypass occlusion (7 patients), aneurysm hemorrhage due to incomplete aneurysm occlusion (4 patients), or aneurysm thrombosis with perforator or branch artery occlusion (4 patients). 13 patients were worse at late follow-up (permanent neurological morbidity, 9%; mean length of follow-up, 23±1.9 months). Overall, good outcomes (GOS 5 or 4) were observed in 114 patients (81%) and 109 patients (78%) were improved or unchanged after therapy.
A heavy reliance on bypass techniques plus indirect giant aneurysm occlusion distinguishes this contemporary surgical experience from earlier ones, and obviates the need for hypothermic circulatory arrest. Experienced neurosurgeons can achieve excellent results with surgery as the “first-line” management approach and endovascular techniques as adjuncts to surgery.
bypass; direct clipping; giant aneurysm; indirect aneurysm occlusion; microsurgery
Brain arteriovenous malformations (BAVMs) are an important cause of intracranial hemorrhage (ICH) in young adults. Gene expression profiling of blood has led to the identification of stroke biomarkers, and may help identify BAVM biomarkers and illuminate BAVM pathogenesis. It is unknown whether blood gene expression profiles differ between 1) BAVM patients and healthy controls, or 2) unruptured and ruptured BAVM patients at presentation. We characterized blood transcriptional profiles in 60 subjects (20 unruptured BAVM, 20 ruptured BAVM, and 20 healthy controls) using Affymetrix whole genome expression arrays. Expression differences between groups were tested by ANOVA, adjusting for potential confounders. Genes with absolute fold change ≥ 1.2 (false discovery rate corrected p ≤ 0.1) were selected as differentially expressed and evaluated for over-representation in KEGG biological pathways (p ≤ 0.05). Twenty-nine genes were differentially expressed between unruptured BAVM patients and controls, including 13 which may be predictive of BAVM. Patients with ruptured BAVM compared to unruptured BAVM differed in expression of 1490 genes, with over-representation of genes in 8 pathways including MAPK, VEGF, Wnt signaling and several inflammatory pathways. These results suggest clues to the pathogenesis of BAVM and/or BAVM rupture and point to potential biomarkers or new treatment targets.
arteriovenous malformation; blood; gene expression; intracranial hemorrhage; microarray analysis
Elevated levels of B-type natriuretic peptide (BNP) have been associated with cardiac dysfunction and adverse neurological outcomes after subarachnoid hemorrhage (SAH). We sought to determine whether elevated levels of BNP are independently associated with radiographic cerebral infarction after SAH.
Plasma BNP levels were measured after admission, a mean of 5.5 ± 3.0 days after SAH onset. Cerebral infarction was determined by retrospective review of computerized tomography (CT) scans. Cerebral vasospasm was confirmed by the presence of vascular narrowing on cerebral angiogram. The association between BNP and cerebral infarction was quantified using multivariable logistic regression and reverse stepwise elimination of clinical covariates. A stratified analysis was performed to quantify the association between BNP levels and infarction in patients with and without angiographic vasospasm.
BNP levels were measured from 119 subjects. The median BNP level was 105 pg/ml (interquartile range 37–275 pg/ml). In our multivariable model, the top quartile of BNP levels (≥276 pg/ml) were associated with an increased odds of cerebral infarction (OR 4.2, P = 0.009). The stratified analysis showed that the association between BNP and infarction was strongest in patients without angiographic vasospasm (OR 7.8, P = 0.006).
Elevated levels of BNP are strongly and independently associated with cerebral infarction, and the association is most pronounced in patients without angiographic vasospasm. These results provide further evidence that other mechanisms can contribute to infarction, and BNP may be a useful biomarker in detecting patients at risk for adverse outcomes without large vessel vasospasm.
Subarachnoid hemorrhage; Cerebral infarction; Cerebral vasospasm; B-type natriuretic peptide
Estimation of the stability of dysmorphic fusiform aneurysms of the intra-cranial internal carotid artery requires precise monitoring of their volumes. In this report we apply a method using MRI and 3D post-processing to study the evolution of these aneurysms on a prospective cohort of patients not immediately suitable for surgery or endovascular treatment.
Materials and Methods
Ten patients with fusiform aneurysms of the intra-cranial internal carotid artery underwent serial MRI studies. Five patients were studied at two time points and the remainder at multiple time points (mean delay between studies: 12.6 +/− 3.8 months). For each patient, studies from all time points were co-registered. Volumes of each vessel component were calculated.
Mean aneurysm volume was 833 +/− 878 mm3. Mean annual rate of volume progression was 1.37 +/− 2.09 % per year. All the aneurysms were thrombus-free.
This study indicates that, given the relatively low rate of progression of these dysplastic fusiform aneurysms and the complexity of their shape, 3D quantitative volumetric methods can be helpful in monitoring whether any growth has occurred.
Brain arteriovenous malformations (bAVM) are an important cause of hemorrhagic stroke. The underlying mechanisms are not clear. No animal model for adult bAVM is available for mechanistic exploration. Patients with Hereditary Hemorrhagic Telangiectasia Type2 (HHT2) with activin receptor-like kinase 1 (ALK1; ACVRL1) mutations have a higher incidence of bAVM than the general population. We tested the hypothesis that VEGF stimulation with regional homozygous deletion of Alk1 induces severe dysplasia in the adult mouse brain, akin to human bAVM.
Alk12f/2f (exons 4–6 flanked by loxP sites) and wild-type (WT) mice (8–10 weeks old) were injected with Ad-Cre (2×107 PFU, adenoviral vector expressing Cre recombinase) and AAV-VEGF (2×109 genome copies, adeno-associated viral vectors expressing VEGF) into the basal ganglia. At 8 weeks, blood vessels were analyzed.
Gross vascular irregularities were seen in Alk1 2f/2f mouse brain injected with Ad-Cre and AAV-VEGF. The vessels were markedly enlarged with abnormal patterning resembling aspects of the human bAVM phenotype, displayed altered expression of the arterial and venous markers (EphB4 and Jagged-1), and showed evidence of arteriovenous shunting. Vascular irregularities were not seen in similarly treated WT mice.
Our data indicate that post-natal, adult formation of the human disease bAVM is possible, and that both genetic mutation and angiogenic stimulation are necessary for lesion development. Our work not only provides a testable adult mouse bAVM model for the first time, but also suggests that specific medical therapy can be developed to slow bAVM growth and potentially stabilize the rupture-prone abnormal vasculature.
Brain arteriovenous malformations (BAVM) are high-flow vascular lesions prone to intracranial hemorrhage (ICH). Abnormal angiogenesis is a key characteristic of BAVM tissue. Angiopoietin-like 4 (ANGPTL4), a secreted glycoprotein, is thought to be involved in angiogenesis and required for proper postnatal blood vessel partitioning. We investigated whether common single nucleotide polymorphisms (SNPs) in ANGPTL4 were associated with risk of BAVM or ICH.
Methods and Results
We conducted a case-control study of 216 Caucasian BAVM cases and 246 healthy controls, and a secondary case-only analysis, comparing 83 ruptured (ICH) with 133 unruptured BAVM cases at presentation. Four tagSNPs in ANGPTL4 captured variation over a 10-kb region (rs2278236, rs1044250, rs11672433, and rs1808536) and were tested for association with BAVM or ICH. The minor allele (A) of rs11672433 (exon 6, Pro389Pro) was associated with an increased risk of BAVM (p = 0.006), which persisted after adjusting for multiple comparisons (p = 0.03). After adjustments for age and sex, carriers of the minor allele (A) remained at higher risk for BAVM compared to noncarriers (odds ratio, OR = 1.56; 95% confidence interval, CI = 1.01–2.41; p = 0.046) and risk of BAVM was increased with increasing copy of the minor A allele (OR = 1.49, 95% CI = 1.03–2.15; ptrend = 0.03). Five common haplotypes (frequency >1%) were inferred; overall haplotype distribution differed between BAVM cases and controls (χ2 = 12.2, d.f. = 4, p = 0.02). Neither SNPs (p > 0.05) nor haplotype distribution (χ2 = 1.1, d.f. = 4, p = 0.89) were associated with risk of ICH among BAVM cases.
A synonymous SNP in ANGPTL4 and haplotypes carrying it are associated with risk of BAVM but not with ICH presentation in BAVM cases.
Arteriovenous malformations; Cerebrovascular disorders; Epidemiology; Genetics; Intracranial hemorrhage
Background and Purpose
Abnormal vascular remodeling triggered by hemodynamic stresses and inflammation is believed to be a key process in the pathophysiology of intracranial aneurysms. Numerous studies have shown infiltration of inflammatory cells, especially macrophages, into intracranial aneurysmal walls in humans. Using a mouse model of intracranial aneurysms, we tested whether macrophages play critical roles in the formation of intracranial aneurysms.
Intracranial aneurysms were induced in adult male mice using a combination of a single injection of elastase into the cerebrospinal fluid and angiotensin-II-induced hypertension. Aneurysm formation was assessed three weeks later. Roles of macrophages were assessed utilizing clodronate liposome-induced macrophage depletion. In addition, the incidence of aneurysms was assessed in mice lacking monocyte chemotactic protein-1 (MCP-1, CCL2), and mice lacking matrix metalloproteinase-12 (MMP-12, macrophage elastase).
Intracranial aneurysms in this model showed leukocyte infiltration into the aneurysmal wall, the majority of leukocytes being macrophages. Mice with macrophage depletion had a significantly reduced incidence of aneurysms compared to control mice (1/10 vs. 6/10; P < 0.05). Similarly, there was a reduced incidence of aneurysms in mice lacking MCP-1, compared to incidence of aneurysms in wild-type mice (2/10 vs. 14/20, P < 0.05). There was no difference in the incidence of aneurysms between mice lacking MMP-12 and wild-type mice.
These data suggest critical roles of macrophages and proper macrophage functions in the formation of intracranial aneurysms in this model.
Intracranial aneurysm; stroke; inflammation; animal model; macrophage
Brain and spinal cord arteriovenous malformations (AVMs) are characterized by aberrant angiogenesis and vascular remodeling. Endothelial progenitor cells (EPCs) can be recruited by stromal cell-derived factor-1 (SDF-1), and participate in vascular remodeling in both physiological and pathological settings. We hypothesized that there was increased EPC levels in the brain and spinal cord AVM nidus.
Microsurgical specimens without endovascular embolization and radiosurgery from the brain (n=12) and spinal cord (n=5) AVMs were examined. Hemangioblastoma, meningioma, cerebral cortex obtained from epilepsy surgery, and the basilar artery (BA) from the autopsy were chosen for control comparisons. EPCs were identified as cells that were double-positive for the stem cell marker CD133 and the endothelial cell marker VEGFR-2 (vascular endothelial growth factor receptor-2 or KDR). In addition, SDF-1 was characterized by immunohistochemistry.
Both brain and spinal AVM tissues displayed more CD133, SDF-1, and CD68-positive signals than epilepsy and basilar artery control tissues. The level of EPCs was increased in the brain and spinal cord AVM nidus, mainly at the edge of the vessel wall. The expression of SDF-1 was co-localized with CD31-positive and α-smooth muscle cells, and was predominantly found within the vessel wall.
Our data demonstrate that EPCs are present in the nidus of the brain and spinal cord AVMs, which may mediate pathological vascular remodeling and impact the clinical course of AVMs.
Angiogenesis; Endothelial progenitor cell; Stromal cell-derived factor-1; Vascular malformation
Estimating risk of intracranial hemorrhage (ICH) for patients with unruptured brain arteriovenous malformations (AVMs) in the natural course is essential for assessing risks and benefits of treatment. Traditionally, the survival period starts at the time of diagnosis and ends at ICH, but most patients are quickly censored because of treatment. Alternatively, a survival period from birth to first ICH, censoring at the date of diagnosis, has been proposed. The authors quantitatively compared these 2 timelines using survival analysis in 1,581 Northern California brain AVM patients (2000–2007). Time-shift analysis of the birth-to-diagnosis timeline and maximum pseudolikelihood identified the point at which the 2 survival curves overlapped; the 95% confidence interval was determined using bootstrapping. Annual ICH rates per 100 patient-years were similar for both the birth-to-diagnosis (1.27, 95% confidence interval (CI): 1.18, 1.36) and the diagnosis-to-ICH (1.17, 95% CI: 0.89, 1.53) timelines, despite differences in curve morphology. Shifting the birth-to-diagnosis timeline an optimal amount (10.3 years, 95% CI: 3.3, 17.4) resulted in similar ICH survival curves (P = 0.979). These results suggest that the unconventional birth-to-diagnosis approach can be used to analyze risk factors for natural history risk in unruptured brain AVM patients, providing greater statistical power. The data also suggest a biologic change around age 10 years influencing ICH rate.
arteriovenous malformations; intracranial hemorrhages; risk factors; survival analysis
Patient age, hemorrhagic presentation, nidal diffuseness, and deep perforating artery supply are important factors when selecting patients with brain arteriovenous malformations for surgery. We hypothesized that these factors outside of the Spetzler-Martin grading system could be combined into a simple, supplementary grading system that would accurately predict neurological outcome and refine patient selection.
A consecutive, single-surgeon series of 300 patients with AVMs treated microsurgically was analyzed in terms of change between preoperative and final postoperative Modified Rankin Scale scores. Three different multivariable logistic models (full, Spetzler-Martin, and supplementary models) were constructed to test the association of combined predictor variables with the change in MRS score. A simplified supplementary grading system was developed from the data which combined age, hemorrhagic presentation, and diffuseness in a manner analogous to the Spetzler-Martin grading system, with points assigned according to each variable and added together for a supplementary AVM grade.
Predictive accuracy was highest for the full multivariable model (receiver operating characteristic curve area, 0.78), followed by the supplementary model (0.73), and least for the Spetzler-Martin model (0.66). Predictive accuracy of the simplified supplementary grade was significantly better than that of the Spetzler-Martin grade (P=0.042), with ROC curve areas of 0.73 and 0.65, respectively. The predictive accuracy of the supplementary grade was only slightly less than a full point score with all 7 weighted variables (P=0.364), with areas under the ROC curve of 0.73 and 0.75, respectively.
This new AVM grading system supplements rather than replaces the well established Spetzler-Martin grading system, and is a better predictor of neurological outcomes after AVM surgery. The supplementary grading scale has high predictive accuracy on its own and stratifies surgical risk more evenly. Supplementary grades can confirm risk predicted by the Spetzler-Martin grade, or in cases of mismatched grades, may alter management decisions. The supplementary grading system is easily applicable at the bedside, where it is intended to improve preoperative risk prediction and patient selection for surgery.
Arteriovenous malformation; Microsurgery; Patient selection; Prediction models; Spetzler-Martin grading system; Supplementary grading system
Evolution of intracranial aneurysms is known to be related to hemodynamic forces such as Wall Shear Stress (WSS) and Maximum Shear Stress (MSS). Estimation of these parameters can be performed using numerical simulations (computational fluid dynamics - CFD) but can also be directly measured with MRI using a time-dependent 3D phase-contrast sequence with encoding of each of the three components of the velocity vectors (7D-MRV). In order to study the accuracy of 7D-MRV in estimating these parameters in–vivo, in comparison with CFD, 7D-MRV and patient-specific CFD modeling was performed for three patients who had intracranial aneurysms. A visual and a quantitative analysis of the flow pattern and the distribution of velocities, MSS, and WSS were performed between the two techniques. Spearman's coefficients of correlation between the two techniques were 0.56 for the velocity field, 0.48 for MSS and 0.59 for WSS. Visual analysis and Bland-Altman plots showed a good agreement for flow pattern and velocities but large discrepancies for MSS and WSS. In conclusion, these results indicate that in-vivo 7D-MRV can be used to measure velocity flow fields and to estimate MSS and WSS but is not currently able to provide accurate quantification of these two last parameters.
Phase-contrast MRI; Computational Fluid Dynamics; Intracranial aneurysm; Wall Shear Stress
A role for the Notch signalling pathway in the formation of arteriovenous malformations during development has been suggested. However, whether Notch signalling is involved in brain arteriovenous malformations in humans remains unclear. Here, we performed immunohistochemistry on surgically resected brain arteriovenous malformations and found that, compared with control brain vascular tissue, Notch-1 signalling was activated in smooth muscle and endothelial cells of the lesional tissue. Western blotting showed an activated form of Notch-1 in brain arteriovenous malformations, irrespective of clinical presentation and with or without preoperative embolization, but not in normal cerebral vessels from controls. In addition, the Notch-1 ligands Jagged-1 and Delta-like-4 and the downstream Notch-1 target Hes-1 were increased in abundance and activated in human brain arteriovenous malformations. Finally, increased angiogenesis was found in adult rats treated with a Notch-1 activator. Our findings suggest that activation of Notch-1 signalling is a phenotypic feature of brain arteriovenous malformations, and that activation of Notch-1 in normal vasculature induces a pro-angiogenic state, which may contribute to the development of vascular malformations.
Notch-1; AVM; human; brain; signalling; angiogenesis
Brain arteriovenous malformations (BAVM) are a tangle of abnormal vessels directly shunting blood from the arterial to venous circulation and an important cause of intracranial hemorrhage (ICH). EphB4 is involved in arterial-venous determination during embryogenesis; altered signaling could lead to vascular instability resulting in ICH. We investigated the association of single-nucleotide polymorphisms (SNPs) and haplotypes in EPHB4 with risk of ICH at clinical presentation in BAVM patients.
Methods and Results
Eight haplotype-tagging SNPs spanning ∼29 kb were tested for association with ICH presentation in 146 Caucasian BAVM patients (phase I: 56 ICH, 90 non-ICH) using allelic, haplotypic, and principal components analysis. Associated SNPs were then genotyped in 102 additional cases (phase II: 37 ICH, 65 non-ICH) and data combined for multivariable logistic regression. Minor alleles of 2 SNPs were associated with reduced risk of ICH presentation (rs314313 C, P=0.005; rs314308 T, P=0.0004). Overall, haplotypes were also significantly associated with ICH presentation (χ2=17.24, 6 df, P=0.008); 2 haplotypes containing the rs314308 T allele (GCCTGGGT, P=0.003; GTCTGGGC, P=0.036) were associated with reduced risk. In principal components analysis, 2 components explained 91% of the variance, and complemented haplotype results by implicating 4 SNPs at the 5′ end, including rs314308 and rs314313. These 2 SNPs were replicated in the phase II cohort, and combined data resulted in greater significance (rs314313, P=0.0007; rs314308, P=0.00008). SNP association with ICH presentation persisted after adjusting for age, sex, BAVM size, and deep venous drainage.
EPHB4 polymorphisms are associated with risk of ICH presentation in BAVM patients, warranting further study.
cerebrovascular disorders; genetics; hemorrhage; receptors; risk factors
The elevation of serum alanine aminotransferase (ALT) is regarded as an indicator of liver damage based on the presumption that ALT protein is specifically and abundantly expressed in the liver. However, ALT elevation is also observed in non-liver injury conditions (e.g., muscle injury) and in apparently healthy people. Conversely, serum ALT activity is normal in many patients with confirmed liver diseases (e.g., cirrhosis and hepatitis C infection). To improve the diagnostic value of the ALT assay and to understand the molecular basis for serum ALT changes in various pathophysiological conditions, we have cloned rat ALT isoenzyme ALT1 and ALT2 cDNAs, examined their tissue expressions at the mRNA and protein levels, and determined ALT1 and ALT 2 serum levels in response to liver damage in rodents. Quantitative real-time PCR (qRT-PCR) analysis shows that ALT1 mRNA is widely distributed and mainly expressed in intestine, liver, fat tissues, colon, muscle and heart, in the order of high to low expression level, whereas ALT2 gene expression is more restricted, mainly in liver, muscle, brain, and white adipose tissue. The tissue distribution pattern of ALT1 and ALT2 proteins largely agrees with their mRNA expression. Interestingly, hepatic ALT2 protein is about four times higher in male rats than female rats. In addition, ALT isoenzymes distribute differentially at the subcellular level in that ALT1 is a cytoplasmic protein and ALT2 a mitochondrial protein, supporting bioinformatic prediction of mitochondrial localization of ALT2. Finally, using animal models of hepatoxicity induced by carbon tetrachloride and acetaminophen, we found that both serum ALT1 and ALT2 protein levels were significantly elevated and correlated with ALT activity, providing, for the first time, the molecular basis for the elevated total serum ALT activity.
Brain arteriovenous malformations (AVMs) are an important cause of neurological morbidity in young adults. The pathophysiology of these lesions is poorly understood. A soluble form of endoglin (sEng) has been shown to cause endothelial dysfunction and induce preeclampsia. We tested if sEng would be elevated in brain AVM tissues relative to epilepsy brain tissues, and also investigated whether sEng overexpression via gene transfer in the mouse brain would induce vascular dysplasia and associated changes in downstream signaling pathways.
Expression levels of sEng in surgical specimens were determined by Western blot assay and ELISA. Vascular dysplasia, levels of MMP and oxidative stress were determined by immunohistochemistry and gelatin zymography.
Brain AVMs (n=33) had higher mean sEng levels (245 ± 175 vs 100 ± 60, % of control, P=0.04) compared with controls (n=8), as determined by Western blot. In contrast, membrane-bound Eng was not significantly different (108 ± 79 vs 100 ± 63, % of control, P=0.95). sEng gene transduction in the mouse brain induced abnormal vascular structures. It also increased matrix metalloproteinase (MMP) activity by 490 ± 30% (MMP-9), 220 ± 30% (MMP-2), and oxidants by 260 ± 20% (4-hydroxy-2-nonenal) at 2 weeks after injection, suggesting that MMPs and oxidative radicals may mediate sEng-induced pathological vascular remodeling.
The results suggest that elevated sEng may play a role in the generation of sporadic brain AVMs. Our findings may provide new targets for therapeutic intervention for patients with brain AVMs.
Stereotactic radiosurgery makes brain arteriovenous malformations (AVM) more manageable during their microsurgical resection. To better characterize these effects, we compared results of microsurgical resection of radiated (RS+) and non-radiated (RS−) AVMs to demonstrate that prior radiosurgery facilitates surgery and decreases operative morbidity.
From of series of 344 patients who had AVM resections at the University of California, San Francisco (1997–2007), 21 RS+ patients were matched with 21 RS− patients based on pre-treatment clinical and AVM characteristics. Matching was blinded to outcomes, which were assessed with the modified Rankin Scale (mRS).
Mean AVM volume was reduced by 78% (P<0.01) and Spetzler-Martin grades were reduced in 52% of RS+ patients (P < 0.001). Preoperative embolization was used less in RS+ than in RS− patients (P< 0.001). Mean operative time (P< 0.01), blood loss (P < 0.05), and length of hospital stay (P< 0.05) were lower in the RS+ group. Surgical morbidity was 14% higher in RS− patients, and they had significant worsening in mRS scores after surgery while RS+ patients did not (P<0.01). RS+ patients deteriorated between AVM diagnosis and surgery due to hemorrhages during the latency period (P< 0.05).
Prior radiosurgery facilitates AVM microsurgery and decreases operative morbidity. Radiosurgery is recommended for unruptured AVMs that are not favorable for microsurgical resection. Microsurgical resection is recommended for radiated AVMs that are not completely obliterated after the 3-year latency period, but altered favorably for surgery, even in asymptomatic patients. Prompt resection of persistent AVMs should be considered to avoid the risk of post-latency hemorrhage and to optimize patient outcomes.
Arteriovenous malformation; microsurgical resection; stereotactic radiosurgery; endovascular embolization; multimodality treatment
Polymorphisms in the proinflammatory cytokine interleukin (IL)-1β gene have been associated with systemic atherogenesis, thrombosis and rupture. The aim of this study was to investigate associations between single nucleotide polymorphisms (SNPs) in IL-1β and intracranial hemorrhage (ICH) in the natural course of brain arteriovenous malformation (BAVM) patients.
Two IL-1β promoter SNPs (−511C→T, −31T→C) and 1 synonymous coding SNP in exon 5 at +3953C→T (Phe) were genotyped in 410 BAVM patients. We performed a survival analysis of time to subsequent ICH, censoring cases at first treatment, death or last follow-up. A Cox regression analysis was performed to obtain hazard ratios (HRs) for genotypes adjusted for age, sex, Caucasian race/ethnicity and hemorrhagic presentation.
Subjects with the −31 CC genotype (HR = 2.7; 95% CI 1.1–6.6; p = 0.029) or the −511 TT genotype (HR = 2.6; 95% CI 1.1–6.5; p = 0.039) had a greater risk of subsequent ICH compared with reference genotypes, adjusting for covariates. The +3953C→T SNP was not significantly associated with an increased ICH risk (p = 0.22). The IL-1β promoter polymorphisms were also associated with BAVM susceptibility among a subset of 235 BAVM cases and 255 healthy controls of Caucasian race/ethnicity (p < 0.001).
IL-1β promoter polymorphisms were associated with an increased risk of ICH in BAVM clinical course and with BAVM susceptibility. These results suggest that inflammatory pathways, including the IL-1β cytokine, may play an important role in ICH.
Arteriovenous malformations; Cerebrovascular disease; Inflammation; Single nucleotide polymorphisms
Background and Purpose:
Evolution of intracranial aneurysmal disease is known to be related to hemodynamic forces acting on the vessel wall. Low Wall Shear Stress (WSS) has been reported to have a negative effect on endothelial cells normal physiology and may be an important contributor to local remodeling of the arterial wall and to aneurysm growth and rupture.
Seven patient-specific models of intracranial aneurysms were constructed using MR angiography data acquired at two different time points (mean 16.4 +/− 7.4 months between the two time points). Numerical simulations of the flow in the base-line geometries were performed to compute WSS distributions. The lumenal geometries constructed from the two time points were manually co-registered and the radial displacement of the wall was calculated on a pixel-by-pixel basis. This displacement, corresponding to the local growth of the aneurysm, was compared to the time-averaged wall shear stress (WSSTA) through the cardiac cycle at that location. For statistical analysis, radial displacement was considered to be significant if it was larger than half of the MR pixel resolution (0.3 mm).
Mean WSSTA values obtained for the areas with a displacement smaller and greater than 0.3 mm were 2.55 +/− 3.6 and 0.76 +/− 1.5 Pa respectively (p<0.001). A linear correlation analysis demonstrated a significant relationship between WSSTA and surface displacement (p<0.001).
These results indicate that aneurysm growth is likely to occur in regions where the endothelial layer lining the vessel wall is exposed to abnormally low wall shear stress.
Magnetic Resonance; Aneurysm; Hemodynamics; Endothelium
To understand the molecular mechanisms underlying compound-induced hemangiosarcomas in mice, and therefore, their human relevance, a systems biology approach was undertaken using transcriptomics and Causal Network Modeling from mice treated with 2-butoxyethanol (2-BE). 2-BE is a hemolytic agent that induces hemangiosarcomas in mice. We hypothesized that the hemolysis induced by 2-BE would result in local tissue hypoxia, a well-documented trigger for endothelial cell proliferation leading to hemangiosarcoma. Gene expression data from bone marrow (BM), liver, and spleen of mice exposed to a single dose (4 h) or seven daily doses of 2-BE were used to develop a mechanistic model of hemangiosarcoma. The resulting mechanistic model confirms previous work proposing that 2-BE induces macrophage activation and inflammation in the liver. In addition, the model supports local tissue hypoxia in the liver and spleen, coupled with increased erythropoeitin signaling and erythropoiesis in the spleen and BM, and suppression of mechanisms that contribute to genomic stability, events that could be contributing factors to hemangiosarcoma formation. Finally, an immunohistochemistry method (Hypoxyprobe) demonstrated that tissue hypoxia was present in the spleen and BM. Together, the results of this study identify molecular mechanisms that initiate hemangiosarcoma, a key step in understanding safety concerns that can impact drug decision processes, and identified hypoxia as a possible contributing factor for 2-BE–induced hemangiosarcoma in mice.
hemangiosarcoma; hypoxia; angiogenesis; endothelial cells; endothelial precursor cells; mechanism of action; human relevance; 2-butoxyethanol
Background and Purpose
As part of the Carotid Occlusion Surgery Study (COSS), patients with chronic cervical carotid artery occlusive disease are selected for extracranial-intracranial bypass surgery based on the results of 15O2/H215O PET imaging. The purpose of this study was to compare the results of Perfusion-CT (PCT) with those of 15O2/H215O PET in a subset of COSS patients.
Materials & Methods
Six patients enrolled in COSS underwent a standard-of-care PCT in addition to the 15O2/H215O PET study used for determining study eligibility. PCT and PET studies were coregistered and then processed separately by different radiologists. Relative measurement of cerebral blood flow (CBF) and oxygen extraction fraction (OEF) without arterial sampling were calculated from two PET scans, one for O15O inhalation and one for H215O injection. PCT datasets were processed using different arterial input functions (AIF), on the side of the carotid occlusion (“ischemic” inputs) and on the contralateral side (“nonischemic” inputs). The same sets of symmetric regions of interests (anterior, middle and posterior cerebral artery distribution) were drawn on both hemispheres on matching slices from both imaging modalities (PCT and PET). Relative PCT and PET CBF values (“ischemic” side divided by “nonischemic” side) were compared using linear regression model, in order to determine the most appropriate arterial input function for PCT. As a secondary analysis, PCT values of relative CBF, cerebral blood volume (CBV) and mean transit time (MTT) using the most accurate arterial input function were evaluated for linear regression with respect to relative PET OEF values, which are used for determining study eligibility in COSS.
The most accurate PCT relative CBF maps with respect to the gold standard PET CBF, were obtained when CBF values for each arterial territory are calculated using a dedicated AIF for each territory (ACA AIF for ACA territory, right MCA AIF for right MCA territory, left MCA AIF for left MCA territory) (R2 = 0.796, p < 0.001). PCT MTT is the parameter that showed the best correlation with the count-based PET OEF ratios (slope = 0.124, intercept = 0.676, R2 = 0.590, p < 0.001).
PCT relative CBF compares favorably to PET relative CBF in patients with chronic carotid occlusion when processed using a dedicated AIF for each territory. The PCT MTT parameter correlated best with PET relative OEF. Prospective investigations are needed to assess whether PCT can be used as an alternative to PET OEF to identify patients with recently symptomatic carotid artery occlusion who are at high risk for subsequent stroke.
chronic carotid occlusion; PET; perfusion-CT; processing; arterial input function
Brain arteriovenous malformations (AVM) have high MMP-9, IL-6 and MPO expression, and polymorphic variations in inflammatory genes are associated with increased risk of hemorrhage. In this study, we characterized the presence of inflammatory cells in AVM lesional tissues.
Immunohistochemistry was used to identify and localize neutrophils (MPO as marker), macrophages/microglia (CD68 as marker), T lymphocytes (CD3 as marker), and B lymphocytes (CD20 as marker). Endothelial cell (EC) marker CD31 was used as an index to assess vascular mass (EC mass). Surgical specimens from 20 unruptured, non-embolized AVMs were examined; seven cortical samples from temporal lobectomy were used as controls. Positive signals for inflammatory cell markers were counted and analyzed by normalizing to the area of the tissue section and the amount of endothelial cells (cells/mm2/EC mass pixels). Levels of MPO and MMP-9 were determined by ELISA.
Neutrophils and macrophages are all frequently identified in the vascular wall of AVM tissues. In contrast, T and B lymphocytes are rarely observed in AVM tissues. AVM tissues displayed more neutrophil and macrophage/microglia markers than epilepsy control tissues (MPO: 434 ± 333 vs 5 ± 4, P=0.0001; CD68: 454 ± 404 vs 4 ± 2, P=0.0001; cells/mm2/EC mass pixels). In ex vivo studies, neutrophil quantity, MPO, and MMP-9 levels were all co-linear(R2=0.98–0.99).
Our study demonstrates that inflammatory cells are present in AVM tissues. Taken together with prior genetic and cytokine studies, these data are consistent with a novel view that inflammation is associated with AVM disease progression and rupture.
CD68; inflammation; myeloperoxidase; vascular malformations
Background and Purpose
Vasospasm following aneurysmal subarachnoid hemorrhage (SAH) remains a leading cause of death and disability after aneurysm rupture. Decreased availability of nitric oxide (NO) may be crucial in the pathogenesis. We hypothesize that endothelial NO synthase (eNOS) polymorphisms may determine susceptibility to vasospasm in SAH patients.
We conducted a prospective cohort study of SAH patients, and determined vasospasm by cerebral angiography. We genotyped three eNOS polymorphisms: intron 4 variable-number-tandem-repeat (VNTR), promoter single-nucleotide-polymorphism (-786T>C SNP), and coding SNP in exon 7 (894G>T encoding E298D). Using multivariable logistic regression, we quantified the association of eNOS polymorphisms in patients with vasospasm confirmed by cerebral angiogram.
For the eNOS promoter -786T>C SNP, the presence of the CC genotype compared to any T genotype (CT or TT) was associated with increased odds of vasospasm (OR 2.97, 95%CI 1.32-6.67, p=0.008). No association with vasospasm was observed for the eNOS 894G>T or VNTR polymorphisms.
These findings suggest that genetic variation influencing NO regulation contributes to risk of angiographic vasospasm in patients with SAH. The specific role of the promoter SNP (-786T>C) may determine the effect of NO regulated by this pathway distinct from other known eNOS polymorphisms.
Endothelial nitric oxide; Genetics; Subarachnoid hemorrhage; Vasospasm
Tetracyclines may be useful in preventing pathological vascular remodeling, thus decreasing the risk of spontaneous hemorrhage from brain vascular malformations.
Arteriovenous malformation (AVM) and intracranial aneurysm patients undergoing non-invasive management were treated with minocycline or doxycycline (200mg/day) up to 2 years in a prospective open-label safety pilot trial. The primary outcome was to compare dose limiting intolerance (DLI), defined as treatment-related dose-reduction and withdrawal between the agents.
26 patients with AVMs (n=12) or aneurysms (n=14) were recruited. Adverse event rates were similar to other reported trials of these agents; four of 13 (31%) minocyline and three of 13 (23%) doxycycline patients had DLI (hazard ratio=3.1, 95%CI=0.52–18.11, log rank p =0.70,).
It is feasible to propose a long-term trial to assess the potential benefit of tetracycline therapy to decrease hemorrhagic risk in brain vascular malformations.
brain vascular malformations; doxycycline; minocycline; tolerability; vasculostatic therapy
Circulating blood endothelial progenitor cells (EPCs) contribute to postnatal vasculogenesis, providing a novel therapeutic target for vascular diseases. However, the molecular mechanism of EPC-induced vasculogenesis is unknown. Interleukin-6 (IL-6) plays multiple functions in angiogenesis and vascular remodeling. Our previous study demonstrated that the polymorphism (174G>C) in IL-6 gene promoter was associated with brain vascular disease. In this study, we investigated if IL-6 receptor is expressed in human endothelial progenitor cells (EPCs) derived from circulating mononuclear cells, and if IL-6 stimulates EPC angiogenesis in vitro.
First, we isolated and cultured mononuclear cells from adult human circulating blood. We obtained EPC clones that were further cultured and expended for the angiogenesis study. We found that the EPCs possessed human mature endothelial cell phenotypes; however, they proliferated much faster than mature endothelial cells (p<0.05). We then found that IL-6 receptor (gp-80) was expressed in the EPCs, and that administration of IL-6 could activate receptor gp80/gp130 signaling pathways including downstream ERK1/2 and STAT-3 phosphorylation in EPCs. Furthermore, IL-6 stimulated EPC proliferation, migration and matrigel tube formation in a dose-dependent manner (p<0.05); anti-IL-6 antibodies or IL-6 receptor could abolish these effects (P<0.05). These results suggest that IL-6 plays a crucial role in the biological behavior of blood-derived EPCs, which may help clarify the mechanism of IL-6 inflammatory-related diseases.
angiogenesis; endothelial progenitor cell; ERK 1/2; interleukin-6; proliferation; migration; tube formation