Current Neurovascular Research - Volume 11, Issue 1, 2014

The level of global DNA methylation may be related to the cerebrovascular disease. The methylation level of Long Interspersed Nucleotide Element 1 (LINE-1) can represent the global methylation level. We investigated the association between the methylation levels of LINE-1 and ischemic stroke in Chinese. Two hundred and eighty patients of ischemic stroke and 280 age- and sex-matched controls were enrolled. The mean percentage of three CpG sites of LINE-1 was calculated for each subject and was used for analysis. Twenty four samples were re-checked for reproducibility of the methylation assay. Multivariate regression model was used to estimate the odds ratio of stroke risk for one percent change of methylation. Sex-specific analysis was also conducted. Thirty-two cases and 11 controls did not pass the methylation assay criteria, and were excluded from further analysis. The intra-class correlation has a coefficient of 0.97 for the methylation assay. The stroke cases had a lower methylation level than controls (p=0.002), especially male subjects (p=0.001). Sex-specific analysis showed that a decrease of 1% methylation level in men could increase a stroke risk by 1.2-fold after adjusting for other risk factors. LINE-1 methylation levels did not have a significant association with stroke in women. The present study shows that a lower level of LINE-1 methylation was associated with a higher risk for ischemic stroke in men, but methylation level in women did not affect the stroke risk. Our finding in Chinese is consistent with a previous result based on elderly white men.

Intracranial hemorrhage is the third most common cause of cerebrovascular disease. Some polymorphisms that affect clotting factors increase the risk of thrombosis. However, few reports have analyzed the effect of polymorphisms on the hemostatic state in bleeding disorders. The low-density lipoprotein receptor (LDLR) has been shown to contribute to factor VIII (FVIII) homeostasis, which represents a link between LDLR and hemostasis. FVIII plays a pivotal role in the coagulation cascade. Patients with high levels of FVIII are at an increased risk of arterial and venous thrombosis. On the other hand, patients with insufficient FVIII tend to bleed excessively, such as in hemophilia A. In a previous study, analysis of the genetic LDLR variant rs688 provided evidence suggesting that genetic polymorphisms of rs688 are associated with thrombotic cardiovascular diseases. The current study aimed to investigate the potential role of rs688 in primary intracerebral hemorrhage (PICH). This genetic association study was conducted within an isolated Taiwanese population (447 PICH patients and 430 controls). Genotypes C/C and C/T were used as the reference genotypes, and the genotype T/T was found to be associated with a 73% decreased risk of PICH. The preliminary evidence suggests that genetic polymorphisms of LDLR are associated with PICH.

Timing of treatment of aneurysmal subarachnoid hemorrhage has been controversial. This retrospective study was designed to access the safety and efficacy among cohorts of different timing of endovascular treatment patients with aneurysmal subarachnoid hemorrhage. A database of patients with aneurysmal subarachnoid hemorrhage was analyzed who were confirmed by CT, and underwent endovascular treatment between January 2005 and January 2012,. The patients were grouped into four cohorts according to the timing of treatment: ultra-early cohort (within 24 hours of onset which was confirmed by CT), early cohort (between 24 and 72 hours of onset which was confirmed by CT), intermediate cohort (between 4 and 10 days of onset which was confirmed by CT) and delayed cohort (after 11 days of onset which was confirmed by CT). Patient demographics, aneurysms features and clinical outcomes were analyzed to evaluate safety and efficacy for timing of endovascular treatment among four cohorts. In our series of 664 patients, 269 patients were grouped into ultra-early cohort, 62 patients in early cohort, 218 patients in intermediate cohort, and 115 patients in delayed cohort. The patient demographics, aneurysm characteristics and neurological conditions on admission among groups showed no statistical significance. As a result of the 9-month follow-up with 513 patients, good outcome (mRS< 2) was achieved in 78% patients in “ultra-early” cohort compared with that of 57% in the “intermediate” group(p=0.000), whereas other comparisons showed no statistical significance(p> 0.05) among the four groups. Dividing the patients with dichotomized mRS into “good outcome” group and “poor outcome” group (mRS> 2) at the 9-month follow-up, the results showed lower Hunt-Hess scores (p=0.000) and smaller size of aneurysms (p=.001) which were correlated with the good outcome. Hypertension (p=0.776), age (p=0.327), sex (p=0.551) and location (p=0.901) showed no statistical significance between groups. Endovascular treatment of aneurysmal subarachnoid hemorrhage which was confirmed by CT within 72 hours achieved better outcomes than that confirmed after 72 hours, especially in those patients treated within 24 hours of onset in comparison with patients treated between 4 and 10 days.

The aim of our study was to analyze morphological and functional aspects of cerebral veins by means of ecocolor- Doppler in young (i.e., ≤30 years old) and older (i.e., >30 years old) patients suffering from multiple sclerosis. 552 multiple sclerosis patients were evaluated by means of a dedicated Echo-Color-Doppler support (MyLab Vinco echocolor Doppler System, Esaote), in both supine and sitting positions. 458 (83%) showed alterations in their morphological and functional structures of cerebral veins and were divided in two different groups: 1) ≤30 (110 patients) and 2) >30 years old (348 patients). Young patients showed a statistically significant higher number of both hemodynamically (44% vs. 35%, p<0.01) and non-hemodynamically (51% vs. 45%, p<0.05) significant stenosis in the internal jugular veins as compared to older patients. A lower percentage of young patients showed blocked outflow in the cervical veins (50% vs. 65%, p<0.01) as compared to older ones. Patients >30 years old outlined a significantly higher disability degree (Expanded Disability Status Scale score: 5 vs. 3, p<0.01) as well as higher disease duration (12 vs. 5 months, p<0.01) than younger. No differences could be outlined about multiple sclerosis clinical form of the disease. It was evidenced that young and adult groups are different kind of patients, the former showing much more cerebral veins stenosis and blocked flow in internal jugular veins and vertebral veins than the latter. Duration of disease could explain such differences: the higher the diseases duration, the higher the degree of vascular alterations and, therefore, the disability degree. This could be due to the complex venous hemodynamic impairments induced by alterations in vascular walls: the blocked or difficult blood flow through stenosis could increase the hydrostatic pressure in the skull and this could induce damages to cerebral cells leading to the genesis of more advanced morphological abnormalities. Furthermore, the vessels’ alterations could impair venous endothelial functions which could turn in a possible alteration of the controls of cerebral vein return which could worsen the cerebral vascular outflow. It may be possible that early clinical, pharmacological and/or invasive vascular interventions could exert a possible role in the natural history of multiple sclerosis. Nevertheless, further trials are needed in order to confirm such considerations.

Fluoro-Gold (F-G) has been used extensively as a fluorescent retrograde neuronal-track tracer in the past. We now report that intraperitoneal administration of 10 to 30 mg/ kg of F-G from 30 min to 7 days prior to sacrifice labels vascular endothelial cells of the brain, choroid plexus and meninges and can be used to assess vascular integrity and damage. F-G vascular labeling co-localized with rat endothelial cell antigen (RECA-1) in the membrane. F-G also intensely labeled the nuclei of the endothelial cells, and co-localized with propidium iodide staining of these nuclei. As well, the administration of F-G during neurotoxic insults produced by amphetamine, kainic acid or “penetrating” wound to the brain can detect where vascular leakage/ hemorrhage has occurred. Histological methods to detect F-G labeled brain vasculature were performed in the same manner as that used for fluorescent visualization of neuronal elements labeled with F-G after perfusion fixation and coronal sectioning (15 to 40 μm) of the brain. This in vivo F-G labeling of endothelial cells and their nuclei yields a clear picture of the integrity of the vasculature and can be used to detect changes in structure. Vascular leaks after “penetrating” wounds through the cortex and striatum, hyperthermic amphetamine exposure or excitotoxic kainate exposure were detected by F-G in the extracellular space and via parenchymal F-G subsequently labeling the terminals and neurons adjacent to the lesioned or damaged vasculature. Further studies are necessary to determine the extent of the leakage necessary to detect vasculature damage. Visualization of the F-G labeling of vasculature structure and leakage is compatible with standard fluorescent immuno-labeling methods used to detect the presence and distribution of a protein in histological sections. This method should be directly applicable to studying brain vascular damage that occurs in the progression of Alzheimer’s disease, diabetes and for monitoring the brain vascular changes during development.

Although selective neurodegeneration of nigro-striatal dopaminergic neurons is widely accepted as a cause of Parkinson’s disease (PD), the role of vascular components in the brain in PD pathology is not well understood. However, the neurodegeneration seen in PD is known to be associated with neuroinflammatory-like changes that can affect or be associated with brain vascular function. Thus, dysfunction of the capillary endothelial cell component of neurovascular units present in the brain may contribute to the damage to dopaminergic neurons that occurs in PD. An animal model of PD employing acute, sub-acute and chronic exposures of mice to methyl-phenyl-tetrahydropyridine (MPTP) was used to determine the extent to which brain vasculature may be damaged in PD. Fluoro-Turquoise gelatin labeling of microvessels and endothelial cells was used to determine the extent of vascular damage produced by MPTP. In addition, tyrosine hydroxylase (TH) and NeuN were employed to detect and quantify dopaminergic neuron damage in the striatum (CPu) and substantia nigra (SNc). Gliosis was evaluated through GFAP immunohistochemistry. MPTP treatment drastically reduced TH immunoreactive neurons in the SNc (20.68±2.83 in acute; 22.98±2.14 in sub-acute; 10.20 ±2.24 in chronic vs 34.88 ±2.91in controls; p<0.001). Similarly, TH immunoreactive terminals were dramatically reduced in the CPu of MPTP treated mice. Additionally, all three MPTP exposures resulted in a decrease in the intensity, length, and number of vessels in both CPu and SNc. Degenerative vascular changes such as endothelial cell ‘clusters’ were also observed after MPTP suggesting that vasculature damage may be modifying the availability of nutrients and exposing blood cells and/or toxic substances to neurons and glia. In summary, vascular damage and degeneration could be an additional exacerbating factor in the progression of PD, and therapeutics that protect and insure vascular integrity may be novel treatments for PD.

We compared the efficacy of intravenous (IV) combination of low-dose tissue plasminogen activator (tPA) and urokinase (UK) versus either classical IV tPA or UK alone for acute ischemic stroke (AIS) within 4.5 h of symptom onset. One-hundred fifty-three AIS patients were treated with 1 of 3 different IV thrombolytic therapies within a 4.5-h time window. Clinical data included age, gender, type of therapy, NIHSS score, time from onset to needle, ASPECTS, mRS at 90 days, and medical history. The outcomes were δNIHSS-a (the difference between NIHSS scores at admission and 24 h); δNIHSS-b (difference between NIHSS scores at admission and 7 days), and mRS at 90 days. Multivariate logistic regression (MLR) was used to determine if treatments or other variables could predict these outcomes. Of 153 patients, 60.1% had a good outcome and 39.9% had a poor outcome. The most important predictors of 90-day mRS were AF history (p < 0.001) and NIHSS score at admission (p = 0.001). Age (p = 0.004) and treatment type (p = 0.043) that were also significantly associated with 90-day mRS. IV tPA yielded the best outcome, compared to low-dose tPA/UK (OR = 1.17) and UK alone (OR = 1.42). Low-dose tPA/UK also resulted in better outcome than UK alone did (OR = 1.12). We conclude that low-dose IV tPA with UK administered within a 4.5-h time window was effective and likely comparable to classical IV tPA thrombolysis.

The purpose of the study was to evaluate the expression of CD45, CD34, Sca-1 and CD117 in mouse bone marrow, Lin-ve and Lin+ve population. Bone marrow cells were isolated from C57/BL6J mouse and mononuclear population was separated from rest of the cell population. With the help of Magnetic associated cell sorter (MACS), Linve and Lin+ve cells were separated from the bone marrow. The expression of CD45, CD34, Sca1 and CD117 was evaluated in bone marrow, Lin-ve and Lin+ve population by flow cytometry. We found a significant increase in the expression of CD34 and CD117 in Lin-ve as compared to the bone marrow and Lin+ve population. These findings suggest that Lin-ve population has higher expression of stem cell progenitor markers and could be useful for tissue repair and regeneration.

The aim of this study was to investigate the flow volumes of the internal jugular veins (IJVs) in patients with cerebral venous sinus thrombosis (CVST) using Doppler ultrasonography (DUS) and to compare the findings with the control group. Forty patients diagnosed with CVST between 2008 and 2010 were included in the study. The patients diagnosed with a thrombosis via MRV and MRI underwent a bilateral examination of the IJVs by DUS. The patients were divided into three groups: Group I (n=29) unilateral total thrombosis; Group II (n= 6) bilateral diffuse thrombosis; and Group III (n=5) unilateral partial thrombosis. The IJV flow volumes of each group were compared to that of the control group (n=20). In Group I, the average flow volume was 53 ml/min on the side of the thrombosis. In Group II, the mean volume of the right and left IJV was 265 ml/min, and in Group III, the mean volume on the side of the partial thrombosis was 160 ml/min. The flow volume on the thrombosed side in Group I and Group III and the mean of the total bilateral flow volume in Group II were significantly lower than that of the control group. IJV flow volumes in the CVST group were significantly lower compared to the control group. Reduced flow volumes of the IJV may be diagnostic for CVST or an additional parameter to be considered with the use of MRI.

Sepsis is the complex syndrome characterized by an imbalance between proinflammatory and antiinflammatory response to infection. The brain may be affected during the sepsis, and acute and long-term brain dysfunctions have been observed in both animal models and septic patients. Oxidative stress and antioxidant systems may prove the basis underling brain dysfunction in sepsis. The antioxidant therapy may be theoretically achieved by the following strategies: restoring endogenous antioxidants and nutrients and supplementation with exogenous trace elements, vitamins, and nutrients with antioxidant proprieties; or administering drugs that reduce oxidative stress, such as NAcetylcysteine (NAC), vitamins and statins. In the review, we described below the involvement of oxidative stress and antioxidants defenses and potential utility of these strategies and present data regarding their use in sepsis.