Current Neurovascular Research - Volume 11, Issue 3, 2014

One of the novel observations reported by Paul Ehrlich who later was awarded the Nobel Prize in Medicine was the separate compartments that were present between the organs of the body and the brain. Ehrlich was able to show that aniline dye injected into the arterial system was able to color multiple organs, such as the heart and kidneys, but did not affect the brain. When such an experiment was performed in a reverse mode years later with injection of the dye into the cerebrospinal fluid of the brain by Ehrlich’s student, Edwin Goldmann, the peripheral organs were noted to be unstained and without evidence of dye. These early experiments identified the existence of the blood brain barrier (BBB) that may years later would be identified as a critical system that controls the flow of agents to and from the brain as well as leading to brain pathology during dysfunction of the BBB system. The BBB is composed of tight junctions that exist between endothelial cells in the central nervous system. Due to the highly restrictive transport across the BBB, few drugs are able to pass through this barrier. Dependent upon the drug composition, some estimates point to only 1% of drugs can pass the through the BBB and be active in the central nervous system. Agents that pass though the BBB through diffusion must be lipid soluble, small in size (<400 Da), and have low hydrogen binding. Other mechanisms that facilitate transport across the BBB include processes that involve carrier-mediated transport and receptor mediated transport, such as the transport of insulin across the BBB. Dynamics and functional stability of the BBB can become severely altered during disease states such as infection or ischemic injury. For example, following a stroke, vascular reperfusion can ensue that results in the initial disruption of the integrity of the BBB, subsequent leakage, and cerebral edema with concurrent neuronal cell loss. These processes also may lead to hemorrhagic transformation following an ischemic stroke. Although these initial insults to the BBB may eventually be repaired with restoration of endothelial tight junctions, it appears that activation of inflammatory pathways as well as other toxic processes may be the primary culprits that are responsible for injury to the brain during the vulnerable period that occurs with BBB dysfunction. However, even low-grade chronic BBB injury may result in neurodegenerative disease. In regards to chronic neurodegenerative disorders such as Alzheimer’s disease, the brains of patients with Alzheimer’s disease have been reported to show BBB leakage that was associated with the APOE4 allele, suggesting that BBB dysfunction may contribute to the onset and/or progression of Alzheimer’s disease. In this issue of Current Neurovascular Research, several novel studies are presented that focus upon the critical role that the BBB and vascular disease may have during both acute and chronic neurodegenerative disorders. In the paper by Ning et al., the authors examine the role of cerebral microemboli during β-amyloid deposition in an amyloid precursor protein/presenilin 1 (APP/PS1) double transgenic mouse model of Alzheimer’s disease. The authors demonstrate that internal carotid artery injection of microemboli can result in increased β-amyloid deposition that occurs during elevated matrix metalloproteinase-9 expression, suggesting that BBB injury may be a factor leading to the increased β-amyloid accumulation in the brain. Interestingly, Provias and Jeynes illustrate that in the brains of Alzheimer’s patients, capillary vascular endothelial growth factor is significantly reduced in the superior temporal, hippocampus and brainstem regions of the brain. Given the significant role of vascular endothelial growth factor for endothelial cell viability and BBB maintenance, their work provides further support for the premise that vascular disease in conjunction with BBB injury may be a contributory factor for the development or progression of Alzheimer's disease. In conjunction with vascular injury, inflammatory disease also may be a significant factor that severely impacts neurodegenerative cell injury. Barichello et al. use an experimental model of pneumococcal meningitis to examine inflammatory parameters and BBB disruption. Following treatment with daptomycin or ceftriaxone, interleukin-4 and neutrophil chemoattractant -1 were reduced with restoration of BBB integrity, suggesting that modulation of inflammatory mediators may be necessary for the maintenance of BBB function. In related work, Yang et al. also point to a vital role for inflammatory cell function. The authors show that in a murine model of chronic cerebral hypoperfusion that reduction in microglial activation and inhibition of autophagy reduces the presence of white matter lesions and improves working memory, suggesting that intact cognitive function may be closely tied to inflammatory cell activity. The role of BBB damage during central nervous system injury is further highlighted in our review articles in this issue of Current Neurovascular Research. Jin et al. discuss the role of early ischemic BBB damage during tissue plasminogen activatorassociated hemorrhagic transformation and the ability to develop new therapeutic strategies to prevent hemorrhagic complications during thrombolytic treatment for stroke. In the review article by Michels et al., the authors highlight the role of microglia during sepsis in the central nervous system and describe the ability of these cells to generate cytokines and chemokines, lead to oxidative stress, and result in BBB breakdown. Their work and analysis places a sharp focus on targeting neuroinflammatory mechanisms for the development of new therapies against disorders in the nervous and vascular systems. Our other review articles complement these papers by shedding light on the role of glycogen synthase kinase-3β (GSK- 3β) during ischemic neuronal injury and the mechanisms and therapeutic options for the development of depression in individuals who suffer a stroke. Pradeep et al. discuss a number of pathways in which GSK- 3β can mediate cell injury, such as through oxidative stress, mitochondrial dysfunction, and activation of inflammatory mediators. All of these pathways including GSK- 3β have been implicated in not only neuronal cell injury, but also BBB dysfunction and cognitive loss. Nabavi et al. examine the pathways that can lead to depression in stroke patients and provide insight into the potential anti-depressant role of reorganization of neural circuitry through brain derived neurotrophic factor (BDNF), an agent that can promote endothelial cell survival, vascular growth, and maintenance of BBB integrity. In this issue of Current Neurovascular Research, a number of novel and elegant studies highlight the critical role played by the BBB in both health and illness. Yet, the link of BBB injury to several acute and chronic neurodegenerative disorders is not always self-evident. Therefore, the ability to launch new therapeutic strategies that can either maintain or control BBB permeability during disease states requires meticulous experimental analysis to correctly “connect the dots” from disorders of the central nervous system to internal cellular pathways that govern the intricate system of cerebral endothelial cells tightly linked together that comprise a selective gatekeeper for the brain known as the BBB.

We investigated the effects of cerebral arterial microemboli on amyloid β protein (Aβ) deposition in the hippocampal region of amyloid precursor protein/presenilin 1 (APP/PS1) double transgenic mice and evaluated the role of cerebral arterial microemboli in Alzheimer's disease (AD) pathogenesis. The mice were divided into a wild-type sham surgery group (n = 15), a wild-type coupled with microemboli group (n =15), an APP/PS1 double transgenic sham surgery group (n =15) and an APP/PS1 double transgenic coupled with microemboli group (n =15). The microemboli mice were injected via the left internal carotid artery with 300 µL of a normal saline suspension containing 100 whole blood clot-derived microemboli (25-50 µm). The sham surgery mice were injected with equal volumes of saline. After the mouse model was established for 1, 2 or 4 weeks, the Aβ1-42 deposition in the left hippocampal region and the matrix metalloproteinase-9 (MMP-9) and glial fibrillary acidic protein (GFAP) expression levels were determined through immunohistochemical staining. The Aβ1-42 deposition level in the left hippocampi of transgenic microemboli group was significantly greater than in the transgenic sham group at week 1 and 2 (P<0.001) but not at week 4. No Aβ1-42 deposition was detected in the wild-type groups. Only sporadic MMP-9- and GFAP-positive cells were observed in the wild-type sham group. Significantly more MMP-9- and GFAP-positive cells were detected in the transgenic groups (P<0.001), particularly in the transgenic microemboli group. An intragroup analysis of the time factor for the microemboli groups showed significantly more MMP-9- and GFAP-positive cells at week 1 than at week 2 or 4 (P<0.001). No difference was detected between time points in the sham groups. Cerebral microemboli increased Aβ deposition in the hippocampal region of APP/PS1 double transgenic mice. MMP-9 and GFAP expression may play an important role in excess Aβ deposition, which is caused by an imbalance between the protein’s synthesis and removal.

Fifteen Alzheimer (AD) and fifteen normative (NM) age-matched autopsy brains were analyzed in superior temporal cortex, hippocampal and brainstem samples. Vascular endothelial growth factor (VEGF) positive capillaries were quantitatively analyzed in all three sites in the 30 cases. Amyloid β42 senile plaques and VEGF positive capillaries were counted and statistically analyzed using Mann-Whitney, Kruskal–Wallis and the non-parametric Spearman’s test. There is a significantly different expression of capillary VEGF between normative and Alzheimer brains. Within Alzheimer’s superior temporal, hippocampus and brainstem sites there was reduced VEGF expression, with the P value being less than 0.05 in all three sites (superior temporal less than 0.035, hippocampus less than 0.001, brainstem less than 0.006). As VEGF is an important endothelial growth factor involved in vascular remodeling, angiogenesis, and endothelial/blood brain barrier maintenance, its reduced expression in Alzheimer’s disease is evidence for altered capillary function in this disease, which may be contributory to its pathogenesis by altering beta amyloid handling and efflux.

Pneumococcal meningitis is associated with neurologic sequelae, such as learning and memory impairment. Most recently, a nonbacteriolytic antibiotic has been investigated to minimise the inflammatory host response and prevent cognitive damage. In this study, we compared daptomycin (DPTO) or ceftriaxone (CFX) treatment on the inflammatory parameters and on the blood-brain barrier (BBB) integrity in experimental pneumococcal meningitis. In the first experiment, the animals received 10 µl of a Streptococcus pneumoniae suspension or artificial cerebrospinal fluid by intracerebroventricular (i.c.v.) and were treated with CFX or DPTO at 18 h post-infection. The animals were euthanised at 18, 20, 24, 36 and 40 h post-infection. In the hippocampus, brain-derived neurotrophic factor (BDNF), tumour necrosis factor alpha (TNF-α), interleukin-6 (IL-6) and IL-10 levels were not different between treatment groups; however, IL-4 and cytokine-induced neutrophil chemoattractant 1 (CINC-1) levels decreased in the CFX group. In the frontal cortex, TNF-α, IL- 4, IL-6, IL-10 and BDNF levels were not different between treatment groups. Only CINC-1 levels decreased at 40 h postinfection with CFX treatment. In the second experiment, the animals received DPTO or CFX for 7 days and were euthanised 10 days after pneumococcal meningitis induction. TNF-α, IL-6, IL-10, CINC-1 and BDNF levels were not different between treatment groups in the hippocampus; however, IL-4 levels decreased in CFX group. In the third experiment, the animals received 10 µl of an S. pneumoniae suspension or artificial CSF by i.c.v. and were treated with a single dose of CFX or DTPO antibiotic; assessment of the BBB breakdown showed that both antibiotics prevented the BBB disruption. Both treatments equally protected the BBB integrity, and there were no significant difference in cytokine production.

Microglial activation plays a vital role in the pathogenesis of white matter lesions (WMLs) during chronic cerebral hypo perfusion. Autophagy has been associated with both microglia survival and cell death. Yet, the role of autophagy during microglial activation in chronic cerebral ischemia is still unknown. We used a chronic cerebral hypoperfusion model by permanent stenosis of bilateral common carotid artery in mice to study microglial activation and autophagy. However, the autophagy inhibitor (3-methyladenine) could attenuate microglial autophagic activation, decrease white matter lesions, and improve working memory during chronic cerebral hypoperfusion in mice. In conclusion, chronic cerebral hypoperfusion that leads to microglial activation and autophagy induction exacerbates white matter lesions and cognitive deficits in mice. Our findings represent a potential novel target for chronic cerebral hypoperfusion therapy.

Influence of undifferentiated bone marrow stromal cells (BMSCs) combined with vein graft on transected sciatic nerve repair was studied in diabetic rats. A nerve segment, 10 mm, was excised and a vein graft (VG) was used to bridge the gap. 10 microliter undifferentiated BMSCs (2x107 cells /mL) were administered into the graft in treatment group (VG/BMSC). Phosphate buffered saline was only administered into the graft in control group. The regenerated nerve fibers were studied in three time points of 4, 8 and 12 weeks after surgery. Evaluation of the repair process was based on behavioral, functional (Walking Track Analysis), electrophysiological, histomorphometrical and immunohistochemical criteria. The behavioral, functional and electrophysiological studies indicated that there was significant recovery in regeneration of axons in VG/BMSC group (P<0.05). Morphometric evaluations showed that the number and diameter of myelinated fibers in VG /BMSC group were significantly higher than those in the control group (P<0.05). This indicates the potential of using undifferentiated BMSCs combined with vein graft in peripheral nerve repair in diabetic rats with no restrictions of donor-site morbidity associated with isolation of Schwann cells. This technique is also cost saving because of decrease in interval from tissue harvest until administration of the cells and simplicity of laboratory techniques in comparison with undifferentiated BMSCs. It may have clinical implications for the surgical management of diabetic patients after nerve transection.

Elevated urinary albumin to creatinine ratio (ACR) and white matter hyperintensity (WMH) volume seen on brain MRI are measures of microvascular disease which may have shared susceptibility to metabolic and vascular insults. We hypothesized that elevated ACR may be useful as inexpensive biomarker to predict presence of cerebral microvascular disease. We assessed the association between ACR at study entry and subsequent WMH volume. We evaluated pulse pressure, mean arterial pressure, hypertension duration, waist circumference, fasting glucose, glomerular filtration rate (GFR) and C-reactive protein (CRP) as potential mediators and diabetes as a moderator of the association between ACR and WMH. Data were collected at study entry and at follow-up approximately 7 years later in a multiethnic population sample of 1281 participants (mean age=51, SD=9.5) from Dallas County. Overall, ACR differences were only marginally (p= 0.05) associated with subsequent WMH. In mediator analysis, however, ACR differences related specifically to arterial pulsatility(β=0.010, bootstrap 95% Confidence Interval (CI): 0.002 to 0.021) and waist circumference (β= -0.004, bootstrap 95% CI: -0.011 to -0.001) were significantly associated with WMH. ACR differences related to serum glucose and CRP were not associated with WMH. ACR evaluated at the same time as WMH had a higher level of significance (p< 0.001) indicating greater utility in predicting current cerebrovascular insults.

Iron overload may contribute to brain damage that involves delayed brain atrophy, edema, and neuronal cell death as well as unfavorable outcome following ischemic stroke and intracerebral hemorrhage (ICH). This prospective study was performed to determine the association of serum ferritin level, an iron storage protein, with perihematoma edema (PHE) growth as well as in-hospital mortality and long-term clinical outcome of patients with ICH. Data was collected from patients with ICH from February 2011 to April 2012. Demographic and clinical data were recorded and serum ferritin was measured on admission. Brain CT scan was performed on admission and 72 hours later. Volume of hematoma and PHE was calculated using ABC/2 formula. Functional outcome was assessed using modified Rankin Scale. A total of 63 patients were included in this study, of those 11 (17.5%) patients died during the first 72 hours of admission. There was a significant correlation between PHE growth during first 72 hours of hospitalization and serum ferritin (P<0.001) as well as history of diabetes mellitus (P<0.001). PHE growth during the first 72-hours of hospitalization and baseline hematoma volume were both predictors of in-hospital mortality and poor outcome (P=0.026 and P=0.035, respectively). These results indicate the role of iron overload in the development of PHE following ICH. However, it seems that serum ferritin level is not directly associated with in-hospital mortality and long-term functional outcome.

Tissue plasminogen activator (tPA) thrombolysis, remains to be the only United States Food and Drug Administration (FDA) approved treatment for acute ischemia stroke. However, the use of tPA has been profoundly constrained due to its narrow therapeutic time window and the increased risk of potentially deadly hemorrhagic complications. TPA-associated hemorrhagic transformation (HT) often occurs as a result of catastrophic failure of the blood brain barrier (BBB), wherein the affected cerebral capillaries can no longer hold blood constituents. Due to its direct contribution to edema and HT, reperfusion-associated BBB damage has been extensively studied, while BBB damage that occurs within the thrombolytic time window is largely neglected. Of note, ischemia-induced BBB damage in the early stroke stages is increasingly appreciated to negatively affect the safety and efficacy profiles of thrombolytic therapy for ischemic stroke. In this review, we discussed the recent findings of spatio-temporal evolution of BBB injury in the early stages of cerebral ischemia and its association with intracerebral hemorrhage following tPA thrombolysis. The increased understanding of early ischemic BBB damage and its close link to tPA-associated HT is of particular importance for developing new preventive and therapeutic strategies to reduce the hemorrhagic complications in stroke thrombolysis.

Neuroinflammation is presented in the acute phase brain damage as well as chronic diseases. Cells that are directly or indirectly involved in immune responses compose the central nervous system (CNS). Microglia are resident cells of the CNS and, as peripheral macrophages, are activated in presence of some cellular insult, producing a large number of cytokines and chemokines in order to remove toxins from the extracellular space. This activation can lead to a breakdown of the blood-brain barrier, production of reactive oxygen species that is involved in the progression of CNS damage as occurs in septic encephalopathy. Given the growing relevance of microglia in the area of neurotoxicology, we describe the role of microglia and the cellular mechanisms that activate these cells during sepsis. Thus, in this review we focused on the relationship between microglia and neuroinflammation associated with sepsis.

Glycogen synthase kinase 3β (GSK-3β) is implicated in diverse cellular processes such as cell signaling and survival. Accumulating lines of evidence indicate that increased GSK-3β activity contributes to neuronal death and pathogenesis of ischemic stroke. Considering predominant roles of GSK-3β in neuronal apoptosis, modulation of this protein kinase is a reliable strategy for ischemic neuroprotection. In this review, we survey and synthesize the current knowledge about the role of GSK-3β in neuroprotection following the ischemic stroke.

Post-stroke depression is an important psychological consequence of ischemic stroke, and affects around one third of stroke patients at any time post-stroke. It has a negative impact on patient morbidity and mortality, and as such development of effective post-stroke recognition and treatment strategies are very important. There are several therapeutic strategies for post-stroke depression, including both pharmacological and non-pharmacological approaches. In this review, we present evidence regarding the underlying biology of post-stroke depression, commonalities between post-stroke depression and Major Depressive Disorder and explore several treatment approaches, including antidepressant therapy, psychotherapy, surgical therapy, electroconvulsive therapy, acupuncture, music therapy and natural products. Further experimental and clinical studies are required, particularly in emerging fields such as the role of nutraceuticals in the treatment of stroke.