Vasogenic Edema Formation Research Articles

Decrease in dystrophin expression prior to disruption of brain–blood barrier within the rat piriform cortex following status epilepticus

Increased permeability of the brain–blood barrier (BBB) in the piriform cortex (PC) has been reported in various animal models of temporal lobe epilepsy. Since BBB disruption induced by epileptogenic insult has not fully clarified, we attempted to determine whether changes in BBB-related molecules are associated with vasogenic edema in the PC. One day after status epilepticus (SE), PC neurons and astrocytes showed a pyknotic nucleus and shrunken cytoplasm accompanied by vasogenic edema. At this time point, SMI-71 (an endothelial barrier antigen) immunoreactivity had decreased in the PC. Prior to vasogenic edema formation (12h after SE), dystrophin immunoreactivity disappeared within astrocytes, while the change in glial fibrillary acidic protein immunoreactivity was negligible. However, glucose transporter-1 (an endothelial cell marker) had increased at 12h after SE. These findings indicate that dysfunction of dystrophin induced by SE may result in endothelial and astroglial damage with BBB breakdown and increase vascular permeability, leading to vasogenic edema that is involved in pathogenesis of epileptogenesis.

Cardiac arrest-induced regional blood–brain barrier breakdown, edema formation and brain pathology: a light and electron microscopic study on a new model for neurodegeneration and neuroprotection in porcine brain

Brief cardiac arrest and survival is often associated with marked neurological alterations related to cognitive and sensory motor functions. However, detail studies using selective vulnerability of brain after cardiac arrest in animal models are still lacking. We examined selective vulnerability of five brain regions in our well-established cardiac arrest model in pigs. Using light and electron microscopic techniques in combinations with immunohistochemistry, we observed that 5, 30, 60 and 180min after cardiac arrest results in progressive neuronal damage that was most marked in the thalamus followed by cortex, hippocampus, hypothalamus and the brain stem. The neuronal damages are largely evident in the areas showing leakage of serum albumin in the neuropil. Furthermore, a tight correlation was seen between neuronal damage and increase in brain water content and Na(+) indicating vasogenic edema formation after cardiac arrest. Damage to myelinated fibers and loss of myelin as seen using Luxol fast blue and myelin basic protein (MBP) immunoreactivity is clearly evident in the brain areas exhibiting neuronal damage. Upregulation of GFAP positive astrocytes closely corresponds with neuronal damages in different brain areas after cardiac arrest. At the ultrastructural level, perivascular edema together with neuronal, glial and endothelia cell damages is frequent in the brain areas showing albumin leakage. Damage to both pre- and post-synaptic membrane is also common. Treatment with methylene blue, an antioxidant markedly reduced neuronal damage, leakage of albumin, overexpression of GFAP and damage to myelin following cardiac arrest. Taken together, these observations suggest that (a) cardiac arrest is capable to induce selective neuronal, glial and myelin damage in different parts of the pig brain, and (b) antioxidant methylene blue is capable to induce neuroprotection by reducing BBB disruption. These observations strongly suggest that the model could be used to explore new therapeutic agents to enhance neurorepair following cardiac arrest-induced brain damage for therapeutic purposes.

Matrix Metalloproteinase-9 as a Marker for Acute Ischemic Stroke: A Systematic Review

Matrix metalloproteinase-9 (MMP-9) is a possible marker for acute ischemic stroke (AIS). In animal models of cerebral ischemia, MMP expression was significantly increased and was related to blood-brain barrier disruption, vasogenic edema formation, and hemorrhagic transformation. The definition of the exact role of MMPs after ischemic stroke will have important diagnostic implications for stroke and for the development of therapeutic strategies aimed at modulating MMPs. The objectives of the present study were to determine (1) whether MMP-9 is a possible marker for AIS; (2) whether MMP-9 levels correlate with infarct volume, stroke severity, or functional outcome; and (3) whether MMP-9 levels correlate with the development of hemorrhagic transformation after tissue plasminogen activator (t-PA) administration. The literature was searched using MEDLINE and EMBASE with no year restriction. All relevant reports were included. A total of 22 studies (3,289 patients) satisfied the inclusion criteria. Our review revealed that higher MMP-9 values were significantly correlated with larger infarct volume, severity of stroke, and worse functional outcome. There were significant differences in MMP-9 levels between patients with AIS and healthy control subjects. Moreover, MMP-9 was a predictor of the development of intracerebral hemorrhage in patients treated with thrombolytic therapy. MMP-9 level was significantly increased after stroke onset, with the level correlating with infarct volume, stroke severity, and functional outcome. MMP-9 is a possible marker for ongoing brain ischemia, as well as a predictor of hemorrhage in patients treated with t-PA.

Cerebral glucose metabolism: Influence on perihematomal edema formation after intracerebral hemorrhage in cat models

Most intracerebral hemorrhage (ICH) imaging studies focus on structural brain changes. Stereotactic neuroimaging techniques such as magnetic resonance imaging (MRI) and positron emission tomography/computed tomography (PET/CT) are helpful in the diagnosis of ICH, monitoring the subsequent recovery and investigating its functional mechanisms. To explore the influence of the changes in cerebral glucose metabolism on perihematomal edema formation in an experimental cat model of ICH. Forty-eight cats were divided into 1 sham operation group (6 cats) and 7 ICH model groups (42 cats)". The ICH model groups were injected with 1.0 ml autologous nonheparinized blood into their thalami using accurate stereotactic guidance apparatus. MRI and (18)F-fluorodeoxyglucose (FDG) PET/CT scans were acquired at 2, 6, 12, 24, 48, 72, and 120 h following the intervention. Pearson's correlation test was used to evaluate the association between T2-weighted signal intensity and the edema volume. Student's t test and q test were used to identify the times of significant temporal changes. The volume of perilesional edema did not significantly increase from 2 h to 12 h after ICH, but then increased by 229.4% at 24 h, peaked (by 273.5%), and steadily decreased by 72 h. The FDG intensity in perihematomal edema tissues was markedly reduced 2 h after ICH on PET images, reached its lowest level at 12 h, and then steadily increased at 24 h and 48 h. The changes of standard absorption value (SUV) in perihematomal edema were consistent with those of FDG intensity. Perihematomal glucose metabolism abnormalities have a close relationship with the formation of vasogenic edema. Furthermore, abnormal glucose metabolism may impair capillary integrity and increase blood-brain barrier permeability.

Cerebrolysin treatment attenuates heat shock protein overexpression in the brain following heat stress

The possibility that overexpression of heat shock proteins (HSPs) in the CNS represents a neurodestructive signal following hyperthermia was examined in a rat model using a potent neuroprotective drug, Cerebrolysin (Ebewe Pharma, Austria). Rats subjected to four hours of heat stress in a biological oxygen demand incubator at 38 degrees C developed profound hyperthermia (41.23 +/- 0.14 degrees C) and overexpressed HSP 72 kD in several brain regions: cerebral cortex, hippocampus, cerebellum, thalamus, hypothalamus, brain stem, and spinal cord compared to controls. This HSP overexpression closely correlated with the leakage of blood-brain barrier permeability and vasogenic edema formation in these brain areas. HSP positive cells are largely confined in the edematous brain regions showing Evans blue leakage. Pretreatment with Cerebrolysin (5 mL/kg, i.v.) 30 minutes before heat stress markedly attenuated hyperthermia (39.48 +/- 0.23 degrees C, P < 0.01) and the induction of HSP to all the brain regions examined. Leakage of Evans blue albumin and increase in brain water content in these brain areas are also markedly reduced with Cerebrolysin pretreatment. These results are the first to show that Cerebrolysin, if administered before heat stress, attenuates hyperthermia induced stress reaction and HSP 72 kD induction. Taken together, these novel observations suggest that upregulation of HSP 72 kD in brain represents neurodestructive signals and a reduction in cellular stress mechanisms leading to decline in HSP expression is neuroprotective in nature.

Effect of Acute Poly(ADP-Ribose) Polymerase Inhibition by 3-AB on Blood–Brain Barrier Permeability and Edema Formation after Focal Traumatic Brain Injury in Rats

Recent evidence supports a crucial role for matrix metalloproteinase-9 (MMP-9) in blood-brain barrier (BBB) disruption and vasogenic edema formation after traumatic brain injury (TBI). Although the exact causes of MMP-9 upregulation after TBI are not fully understood, several arguments suggest a contribution of the enzyme poly(ADP-ribose)polymerase (PARP) in the neuroinflammatory response leading to MMP-9 activation. The objectives of this study were to evaluate the effect of PARP inhibition by 3-aminobenzamide (3-AB) (1) on MMP-9 upregulation and BBB integrity, (2) on edema formation as assessed by magnetic resonance imaging (MRI), (3) on neuron survival as assessed by (1)H magnetic resonance spectroscopy ((1)H-MRS), and (4) on neurological deficits at the acute phase of TBI. Western blots and zymograms showed blunting of MMP-9 upregulation 6 h after TBI. BBB permeability was decreased at the same time point in 3-AB-treated rats compared to vehicle-treated rats. Cerebral MRI showed less "free" water in 3-AB-treated than in vehicle-treated rats 6 h after TBI. MRI findings 24 h after TBI indicated predominant cytotoxic edema, and at this time point no significant differences were found between 3-AB- and vehicle-treated rats with regard to MMP-9 upregulation, BBB permeability, or MRI changes. At both 6 and 24 h, neurological function was better in the 3-AB-treated than in the vehicle-treated rats. These data suggest that PARP inhibition by 3-AB protected the BBB against hyperpermeability induced by MMP-9 upregulation, thereby decreasing vasogenic edema formation 6 h after TBI. Furthermore, our data confirm the neuroprotective effect of 3-AB at the very acute phase of TBI.

Inhibition of myosin light chain kinase reduces brain edema formation after traumatic brain injury

The role of the endothelial contractile apparatus in the process of brain edema formation after brain trauma is not characterized. Phosphorylation of myosin light chains by myosin light chain kinases (MLCK) activates endothelial contractile elements and results in a rearrangement of the cytoskeleton. This may enhance post-traumatic blood-brain barrier dysfunction. In order to investigate the role of the MLCK on brain edema formation and blood-brain barrier permeability after brain injury, mice were anesthetized and subjected to a controlled cortical impact (CCI). MLCK expression is significantly up-regulated after CCI with a maximum 12 h post-injury. Specific inhibition of MLCK by ML-7 resulted in a reduction of phosphorylation of myosin light chains and improved blood-brain-barrier integrity. Accordingly, ML-7 attenuated post-traumatic brain edema formation and intracranial hypertension 24 h after CCI. Prevention of brain edema formation did not translate into improved neurological outcome or reduced brain lesion. In conclusion, the results confirm that the endothelial contractile apparatus is activated by CCI and opens the endothelial barrier leading to vasogenic brain edema formation. Lack of neurological and histological improvement suggests that specific targeting of vasogenic brain edema at the endothelial level is not sufficient to limit secondary brain damage and has, therefore, to be combined with other potential neuroprotective strategies.

Aggravation of infarct formation by brain swelling in a large territorial stroke: a target for neuroprotection?

In territorial stroke vasogenic edema formation leads to elevated intracranial pressure (ICP) and can cause herniation and death. Brain swelling further impairs collateral blood flow to the ischemic penumbra and causes mechanical damage to adjacent brain structures. In the present study the authors sought to quantify the impact of this space-occupying effect on ischemic lesion formation. Wistar rats were assigned to undergo bilateral craniectomy or a sham operation and then were subjected to temporary middle cerebral artery occlusion (MCAO) for 90 minutes. A clinical evaluation and 7-T MR imaging studies were performed 5 and 24 hours after MCAO. The absolute brain water content was determined at 24 hours by using the wet/dry method. Bilateral craniectomy before MCAO led to a drastic reduction in lesion volume at both imaging time points (p < 0.0001). Ischemic lesion volume was 2.7- and 2.3-fold larger in sham-operated animals after 5 and 24 hours, respectively. Clinical scores were likewise better in rats that had undergone craniectomy (p < 0.05). After 24 hours the midline shift differed significantly between the 2 groups (p < 0.001), but not after 5 hours. The relation between brain water content and ischemic lesion volume as well as the T2 relaxation time within the infarcted area was not different between the groups (p > 0.05). The data indicated that collateral damage caused by the space-occupying effect of a large MCA territory stroke contributes seriously to ischemic lesion formation. The elimination of increased ICP thus must be regarded as a highly neuroprotective measure, rather than only a life-saving procedure to prevent cerebral herniation. Further clinical trials should reveal the neuroprotective potential of surgical and pharmacological ICP-lowering therapeutic approaches.

Delayed intracerebral hemorrhage after superficial temporal artery–middle cerebral artery anastomosis in a patient with moyamoya disease: possible involvement of cerebral hyperperfusion and increased vascular permeability

Background Postoperative intracerebral hemorrhage is a rare complication after surgical revascularization for moyamoya disease, and its mechanism is totally undetermined. Case Description A 47-year-old woman with moyamoya disease, experiencing crescendo transient ischemic attack on her left hand, underwent STA-MCA anastomosis on the right hemisphere. Postoperative MR imaging 1 day after surgery demonstrated asymptomatic vasogenic edema without ischemic change at the subcortex under the site of the anastomosis that expanded the next day, and STA-MCA bypass was apparently patent with the strong high signal by MR angiography. N-isopropyl- p-[ 123I]iodo-amphetamine single-photon emission CT showed marked increase in the CBF on the hemisphere operated on. Four days after surgery, the patient complained of sudden headache and experienced severe monoparesis in her left hand due to the intracerebral hemorrhage at the corresponding lesion to the prior vasogenic edema. Edema around hematoma was prolonged for as long as a month; while intensive blood pressure control and the use of adrenocorticosteroid gradually relieved her symptoms. The patient completely recovered from her symptoms 2 months later, and she was discharged without neurologic deficit. Her transient ischemic attacks completely disappeared postoperatively. Conclusion Early increase in CBF associated with vasogenic edema formation at the site of the anastomosis could be the warning sign for subsequent hemorrhagic complication, and intensive blood pressure control is warranted in such patients. Alternatively, it would be necessary to elucidate the biochemical mechanism of the deleterious cascade during reperfusion in moyamoya disease to avoid this rare complication.

Midline-shift Corresponds to the Amount of Brain Edema Early After Hemispheric Stroke—An MRI Study in Rats

Vasogenic brain edema formation is a serious complication in hemispheric stroke. Its space-occupying effect can lead to midline-shift (MLS), cerebral herniation, and death. Clinical studies indicate that quantification of MLS can predict cerebral herniation and subsequent death at early time-points, even before clinical deterioration becomes apparent. The present experimental study was designed to determine the relation between MLS, absolute edema volume, lesion size, and clinical findings in a rat stroke model. Middle cerebral artery-occlusion was performed in 24 rats using the suture technique. Clinical evaluation and magnetic resonance imaging (MRI) (Bruker PharmaScan 7.0T) was performed 24 hours later. Lesion volume, the volume-increase within the affected hemisphere (%HEV), and MLS were quantified on T2-weighted images. The absolute increase of hemispheric water content (DeltaH2O) was determined in a subgroup using the wet-dry method (n=12). MLS correlated significantly with the total amount of brain edema (magnetic resonance imaging study: r=0.82; P<0.01; wet-dry analysis r=0.80; P<0.01). MLS correlated only moderately with T2-lesion volume (r=0.55; P<0.01). No significant correlation could be detected between MLS and clinical scores (r=0.26; P>0.05). MLS thus quantitatively reflects the amount of vasogenic brain edema within the affected hemisphere at early time-points. MLS quantification can be regarded as an easily assessable and valid global quantitative parameter for brain edema and thus might facilitate the surgical and nonsurgical management of edema in acute stroke patients.

Absence of the Chemokine Receptor CCR2 Protects Against Cerebral Ischemia/Reperfusion Injury in Mice

The chemokine, monocyte chemoattractant protein-1 (CCL2), is a major factor driving leukocyte infiltration into the brain parenchyma in a variety of neuropathologic conditions associated with inflammation, including stroke. In addition, recent studies indicate that CCL2 and its receptor (CCR2) could have an important role in regulating blood-brain barrier (BBB) permeability. This study evaluated the role of the CCL2/CCR2 axis in regulating postischemic inflammation, BBB breakdown, and vasogenic edema formation. CCR2(-/-) and CCR2(+/+) mice were subjected to focal transient cerebral ischemia. BBB permeability and brain edema formation were observed at days 1 and 5 of reperfusion by evaluating the product surface area for fluorescein isothiocyanate-albumin and measuring water and electrolyte contents. Immunohistochemistry was used to assess leukocyte infiltration. cDNA gene and protein arrays for inflammatory cytokines were used to assess inflammatory profiles in CCR2(+/+) and CCR2(-/-) mice. CCR2(-/-) mice had reduced infarct sizes and significantly reduced BBB permeability and brain edema formation in the affected ischemic hemisphere compared with CCR2(+/+) mice. This reduction in injury was closely associated with reduced infiltration of not only monocytes but also neutrophils (7- and 4-fold decreases, respectively). In addition, CCR2(-/-) mice had reduced expression/production of inflammatory cytokines during reperfusion. These data suggest that inhibiting the CCL2/CCR2 axis affects brain reperfusion outcome by reducing brain edema, leukocyte infiltration, and inflammatory mediator expression.

Characterization of microvascular basal lamina damage and blood–brain barrier dysfunction following subarachnoid hemorrhage in rats

Vasogenic brain edema is one of the major determinants for mortality following subarachnoid hemorrhage (SAH). Although the formation of vasogenic brain edema occurs on the microvascular level by opening of endothelial tight junctions and disruption of the basal lamina, microvascular changes following experimental SAH are poorly characterized. The aim of the present study was therefore to investigate the time course of blood–brain barrier (BBB) dysfunction and basal lamina damage following SAH as a basis for the better understanding of the pathophysiology of SAH. SAH was induced in Sprague–Dawley rats by an endovascular filament. Animals were sacrificed 6, 24, 48, and 72 h thereafter ( n = 9 per group). Microvascular basal lamina damage was quantified by collagen type IV immunostaining. Western blotting was used to quantify collagen IV protein content and bovine serum albumin (BSA) extravasation as a measure for basal lamina damage and blood–brain barrier disruption, respectively. BSA Western blot revealed significant ( p < 0.05) BBB opening in the cerebral cortex ipsilateral to the hemorrhage beginning 6 h and peaking 48 h after SAH. Significant ( p < 0.05) basal lamina damage occurred with gradual increase from 24 to 72 h. Basal lamina damage correlated significantly with BBB dysfunction ( r = − 0.63; p = 0.0001). Microvascular damage as documented by collagen IV degradation and albumin extravasation is a long lasting and ongoing process following SAH. Due to its delayed manner microvascular damage may be prone for therapeutic interventions. However, further investigations are needed to determine the molecular mechanisms responsible for basal lamina degradation and hence damage of the microvasculature following SAH.

Treatment of Cerebral Edema

Cerebral edema is a potentially devastating complication of various acute neurologic disorders. Its successful treatment may save lives and preserve neurologic function. Different pathophysiological mechanisms are responsible for the formation of cytotoxic and vasogenic edema. Yet, these 2 types of edema often coexist and their treatment tends to overlap, with the exception of corticosteroids, which should be only used to ameliorate vasogenic edema. Currently available to control brain swelling include osmotic agents (with emphasis on mannitol and hypertonic saline solutions), corticosteroids, hyperventilation, sedation (propofol, barbiturates), neuromuscular paralysis, hypothermia, and surgical interventions. This article discusses the indications, advantages, and limitations of each treatment modality following an evidence-based approach. The therapy for brain edema remains largely empirical. More research aimed at enhancing our understanding of the pathophysiology of cerebral edema is needed to identify new and more effective forms of treatment.

A study on blood–brain barrier ultrastructural changes induced by cerebral hypoperfusion of different stages

Objectives: To evaluate the ultrastructural changes in the blood–brain barrier (BBB) induced by cerebral hypoperfusion of different stages, which may predispose the brain to the formation of vasogenic edema and hemorrhage under cerebral luxury perfusion.Methods: Twenty cerebral steal models with left neck arteriovenous fistula (AVF) were surgically created in Wistar's rats, leading to a noninfarctional reduction in the cerebral blood flow (CBF) by between 25 and 50%, resulting in cerebral hypoperfusion in the AVF side for 3 days (acute stage), 3 weeks (subacute stage) and 3 months (chronic stage), respectively. Another six sham-operated models were made in age-matched rats as control. The BBB ultrastructural changes were assessed by transmission electron microscopy. Ridit analysis was conducted to compare the positive ratio of ultrastructural changes among multiple groups.Results: Electron microscopy demonstrated no ultrastructural change at the acute stage, however, at the subacute stage, slight vacuolar degeneration was found in the astrocytic foot process layer encircling the capillaries; furthermore, at the chronic stage, the astrocytic foot processes expressed marked vacuolization associated with the adjacent astrocytic degeneration. Meanwhile, in both capillary endothelium and basal lamina layers, no abnormal ultrastructures similar to those in the astrocytic foot processes layer were identified. After cerebral luxury perfusion took place, BBB was disrupted where astrocytic foot processes vacuolization was most distinguished.Conclusion: Astrocytes generate ultrastructural abnormality as a result of chronic cerebral hypoperfusion. Astrocytic foot process vacuolization, which constitutes the major ultrastructural change in the BBB, is the extension of the degeneration of astrocyte body. It is inferred that BBB is prone to structure weakness and function instability, which forms the morphological basis of cerebral luxury perfusion.

Pathophysiology of glioma cyst formation

Fluid filled cystic cavities are accompaniments of some cerebral gliomas. These tumoural cysts together with peritumoural vasogenic brain oedema add to the morbid effects of the gliomas in terms of mass effect and increased intracranial pressure. Although different mechanisms have been suggested as to the pathogenesis of glioma-associated cysts, it is still unclear why these cysts appear in only a limited number of cerebral gliomas while brain oedema, a probable precursor of glioma cysts, is a usual accompaniment of most gliomas. Here, the authors present a two-hit hypothesis of brain glioma cyst formation. We suggest that after the formation of vasogenic tumoural brain oedema, microvascular phenomena may lead to the formation of microcysts, which might later become confluent and grow to form macroscopic cysts. Progress in the understanding of pathogenesis of cerebral glioma cysts might set targets for treatment of brain edema and glioma cysts.

Pathophysiology of Blood-Spinal Cord Barrier in Traumatic Injury and Repair

Blood-spinal cord barrier (BSCB) plays an important role in the regulation of the fluid microenvironment of the spinal cord. Trauma to the spinal cord impairs the BSCB permeability to proteins leading to vasogenic edema formation. Several endogenous neurochemical mediators and growth factors contribute to trauma induced BSCB disruption. Studies carried out in our laboratory suggest that those drugs and neurotrophic factors capable to attenuate the BSCB dysfunction following trauma are neuroprotective in nature. Whereas, agents that do not exert any influence on the BSCB disruption failed to reduce cell injury. These observations are in line with the idea that BSCB disruption plays an important role in the pathophysiology of spinal cord injuries. The probable mechanism(s) of trauma induced BSCB dysfunction and its contribution to cell injuries are discussed.

The selective Na +–Ca 2+exchange inhibitor attenuates brain edema after radiofrequency lesion in rats

2-[4-[(2,5-Difluorophenyl)methoxy]phenoxy]-5-ethoxyaniline (SEA0400), a specific inhibitor of the Na +–Ca 2+ exchanger, exerts cytoprotective action and reduces brain infarct volume after cerebral ischemia. We examined the effect of SEA0400 on vasogenic brain edema in rats. Histological observations showed that radiofrequency current caused brain infarct and extravasation of endogenous albumin in the brain. SEA0400 (3 and 10 mg/kg, i.v.) significantly suppressed the increase in brain water content with attenuation of Evans blue dye and sodium fluorescein extravasation after radiofrequency lesion. The findings suggest that the Na +–Ca 2+ exchanger plays a role in vasogenic edema formation after radiofrequency lesion.

Transcellular transport as a mechanism of blood-brain barrier disruption during stroke.

It is well-known that ischemia causes disruption of the blood-brain barrier (BBB), which leads to the formation of vasogenic brain edema. One major mechanism of BBB opening is enhanced pinocytotic vesicle formation that may be induced after transient focal ischemia by several mechanisms, including nitric oxide production, release of neurotransmitters, inflammatory mediators and hemodynamic alterations. In the present study we sought to characterize the extent of pinocytosis in cerebral endothelium during both ischemia/reperfusion (I/R) and elevated intravascular pressure. Transient focal ischemia was induced for 1 hour with 24 hours of reperfusion using the filament occlusion model in male Wistar rats, after which occluded middle cerebral arteries (MCAs) were dissected and mounted on glass cannulas in an arteriograph chamber. This system allowed control over intravascular pressure, measurement of lumen diameter and perfusion with various tracers (Lucifer Yellow and horseradish peroxidase) for measurement of transcellular transport and quantification of pinocytosis using transmission electron microscopy. I/R was found to increase vesicle formation by 166% basolaterally without a change in vesicle formation apically compared to non-ischemic control MCAs at 75 mmHg (p less than 0.01). Similarly, an acute increase in pressure to 200 mmHg caused a 78% increase in apical pinocytosis (p less than 0.05) and a non-significant 42% increase basolaterally. These results were confirmed by permeability measurements using Lucifer Yellow and demonstrate that both I/R and acute elevations in intravascular pressure enhance cerebral endothelial cell pinocytosis. The increase in basolateral pinocytosis during ischemia suggests enhanced efflux mechanisms that may be transporting substances from brain to blood. In addition, since the enhanced pinocytosis after an increase in pressure occurred in isolated arteries in vitro without the influence of metabolic or neuronal factors, these findings demonstrate that elevated intravascular pressure is a primary stimulus for pinocytosis in cerebral endothelial cells.

Neutrophils both reduce and increase permeability in a cell culture model of the blood–brain barrier

This study was carried out to determine the effects that human neutrophils have on permeability across a model of the blood–brain barrier (BBB) formed by primary cultures of bovine brain microvessel endothelial cells (BBMEC). Transendothelial electrical resistance (TEER) was used to measure changes in permeability across BBMEC monolayers in a dual compartment system, during neutrophil interactions. When neutrophils (5×10 6/ml) were applied to monolayers, TEER increased (permeability decreased). Adenosine was implicated, since the TEER increase was blocked by adenosine deaminase (1 U/ml) and the adenosine A 2 receptor antagonist ZM 241385 (at 10 −6 M but not 10 −8 M, implicating A 2B receptors). Oxygen free radicals were implicated as the TEER increase was blocked by combined catalase (100 U/ml) and superoxide dismutase (60 U/ml). When a gradient of the bacterial chemoattractant peptide formyl methionyl leucine phenylalanine (fMLP, 10 −7 M) was applied to neutrophils, the TEER decreased (permeability increased), concurrent with migration. When fMLP (10 −7 M) was added to the neutrophils, without migration, no change occurred. The TEER decrease was blocked by loading endothelium with the calcium buffer BAPTA (10 μM) and partially blocked by the serine protease inhibitor aprotinin (20 μg/ml). Measures to block the potential extracellular triggers heparin binding protein, glutamate, oxygen free radicals and binding to intercellular cell adhesion molecule-1 (ICAM-1) were ineffective. These data indicate that neutrophils both reduce and increase permeability in a cell culture model of the BBB, correlated to their proximity and migration through the endothelium. They explore the role of neutrophils in BBB breakdown, and the formation or amelioration of vasogenic cerebral edema.

Aquaporin‐4 water channel expression in human vasogenic brain oedema

A number of brain lesions result in brain oedema formation. This oedema is considered to be vasogenic in nature, resulting from breakdown of the blood–brain barrier. Brain microvessels are surrounded by astrocyte foot processes that express the water channel protein aquaporin 4 (aqp4). Aqp4 has been implicated in the formation of vasogenic oedema in animal models, but little is known about its involvement in vasogenic oedema in man. Therefore formalin‐fixed samples of a variety of brain lesions that give rise to brain oedema were obtained from 17 patients who underwent craniotomy at atkinson morley’s hospital during 1993–9. Immunocytochemistry was then performed on the samples using a polyclonal aqp4 antibody (ab3068, chemicon) visualised with immunoperoxidase and diaminobenzidine hydrochloride. In histologically normal cerebral cortex (n = 1) aqp4 immunoreactivity was restricted to astrocyte foot processes lining the pial membrane and surrounding some microvessels. In contused cortex (n = 3) astrocyte aqp4 expression was up‐regulated in accordance with the degree of tissue damage and the severity of oedema, assessed by contrast enhancement on ct. Aqp4 immunoreactivity was not present in nonastrocytic tumour cells including adenocarcinoma (n = 4), meningioma (n = 4), oligodendroglioma (n = 1) and medulloblastoma (n = 1). However diffuse aqp4 immunoreactivity was present in cells with the morphology of reactive astrocytes at the periphery of these tumours and occasionally ‘trapped’ within them. In addition up‐regulation of aqp4 immunoreactivity was observed in astrocyte end feet surrounding the microvessels in the perimeningioma cortex. Diffuse aqp4 immunoreactivity was observed in a subependyal astrocytoma (n = 1). Strong aqp4 immunolabelling of astrocytes was observed in cerebral cortex affected by bacterial meningitis (n = 1). Up‐regulation of aqp4 immunoreactivity was also observed in the foot processes of astrocytes lining the pial membrane and surrounding microvessels in this tissue. Up‐regulation of aqp4 immunoreactivity was not observed in brain tissue affected by subarachnoid haemorrhage (n = 1), from a patient who did not show any evidence of oedema on ct. Therefore brain lesions with different aetiologies that result in vasogenic brain oedema appear to cause a similar up‐regulation of aqp4 expression in astrocytes associated with the lesions. These results suggest that aqp4 up‐regulation in astrocytes may be part of a ‘final common pathway’ leading to oedema formation.