BBB Disruption Research Articles

Increase in TNFα Transport after SCI Is Specific for Time, Region, and Type of Lesion

The dynamic changes of the blood–brain barrier and blood–spinal cord barrier (BBB) are an important part of the CNS response to injury. This study addresses the permeability of the BBB in the acute phase of spinal cord injury (SCI) to the thoracic region. SCI by compression or by complete transection was generated in mice. BBB disruption was evaluated by spinal cord uptake of radiolabeled albumin. The BBB of the thoracic spinal cord was disrupted immediately after compression injury, lasting for 2 days. This was followed by a delayed permeability increase in the cervical spinal cord beginning 3 days after injury. After transection, BBB disruption was limited to the thoracic spinal cord and was present only immediately postinjury. The entry of TNFα not only was increased at the time of BBB disruption, following the same pattern, but also had secondary changes after the BBB permeability to albumin had returned to normal. The increase of TNFα entry, best explained by upregulation of the specific transport system for TNFα, was pronounced in the lumbar spinal cord as well as the thoracic region, and followed a different time course after the two types of injury. Integrating our results with those of the literature regarding the roles of inflammatory responses and the effects of TNFα in spinal cord regeneration, we conclude that the time-, region-, and lesion-specificity of the upregulation of TNFα transport is part of the regulatory changes at the BBB in response to SCI.

Hyperglycemic Transient Ischemia Induces Massive Neutrophil Deposition in Brain.

P80 Acute hyperglycemia worsens neurological signs and accentuates neuropathology after ischemia, but the mechanisms are poorly understood. In this study, we tested whether polymorphonuclear leukocytes (neutrophils) might be contributory. Anesthetized, physiologically monitored Wistar rats underwent global forebrain ischemia for 12.5 min by bilateral carotid artery occlusions plus hypotension (45 mm Hg). To induce hyperglycemia, rats received 2.5 ml of 25% dextrose i.p. 30 min prior to ischemia. Normoglycemic rats received saline. Plasma glucose levels were 340±66 and 133 ±21 mg/dl, respectively (mean±SD). Animals were sacrificed at either 24 h or 3 days by perfusion-fixation with FAM. Brain sections were reacted for the immunohistochemical visualization of myeloperoxidase (MPO), a specific and quantitative marker of neutrophil activity in the brain. In sham rats and in normoglycemic-ischemic animals, almost no MPO-positive cells were identified. In marked contrast, brains of hyperglycemic-ischemic rats studied at 24 h contained dramatic accumulations of MPO-positive cells within pial and parenchymal blood vessels as well as within cortical and subcortical parenchyma. MPO-positivity was most robust in areas of severe injury (on H&E sections), but MPO cells were also observed in areas without overt injury. Intravascular MPO-positive cells commonly obstructed the vascular lumen. Following 3-d survival, MPO-positivity of hyperglycemic-ischemic brains had significantly decreased. In hyperglycemic brains studied at 24 h, median numbers of MPO-positive cells were increased by 130-fold in cortex and 110-fold in striatum above values in normoglycemic-ischemic rats. In summary, this study shows that preischemic hyperglycemia triggers the early and dramatic deposition of polymorphonuclear leukocytes in the postischemic brain, with neutrophil adherence to cerebral blood vessels and their migration into brain parenchyma following brief forebrain ischemia. These events may contribute to the enhanced tissue destruction, extension of infarction, and BBB disruption observed in hyperglycemic ischemia. Supported by NIH Grant NS05820.

Differential Upregulation of Aquaporin-4 mRNA Expression in Reactive Astrocytes after Brain Injury: Potential Role in Brain Edema

Astrocytes and aquaporin-4 (AQP4) play a significant role in brain ion homeostasis. Consequently the regulation of AQP4 mRNA in the CNS after different neurological insults was of interest. A single intrastriatal injection of ringer or quinolinic acid strongly induced AQP4 mRNA in the striatum, specially at the core of the lesion. Colocalization studies demonstrated that AQP4 mRNA induction was restricted to hypertrophic astrocytes. The extent of striatal AQP4 mRNA induction did not correlate with neuronal degeneration, but it did with extravasation of Evans blue dye as a marker of BBB disruption. Distant lesions were additionally induced by either 6-OHDA or a knife cut in the medial forebrain bundle (MFB). The former, but not the latter, induced a high AQP4 mRNA expression in the lesioned substantia nigra. However, axotomy of the MFB induced a high AQP4 mRNA expression at the lesion site. We conclude that the induction of AQP4 mRNA expression is related to disruption of the blood–brain barrier and under brain edema conditions this water channel plays a key role in the reestablishment of the brain osmotic equilibrium.

Simultaneous assessment of cerebral hemodynamics and contrast agent uptake in lesions with disrupted blood-brain-barrier.

The purpose of this study was to develop a method that eliminates the influence of the T1 relaxation time upon the signal-time course in perfusion-weighted imaging of cerebral lesions with blood-brain-barrier (BBB) disruption. On a 1.5 T whole body clinical magnetic resonance (MR) imager, we implemented a dual-echo RF-spoiled FLASH sequence (TE=6/23.6 ms). We developed a postprocessing routine that allowed to calculate a signal-time course representing only the change in T2* and another one representing only the change in T1. Using this method, we examined 7 patients with various brain lesions showing evidence of BBB disruption. In the signal-time-curves obtained from the early echo we found a distinct signal drop due to the T2* effect. These effects could be eliminated by the correction algorithm yielding a 67% higher signal increase. Correction of the signal-time curve of the late echo yielded a more pronounced maximum signal drop and a decrease in postcontrast signal intensity. We found that without this correction the relative regional cerebral blood volume and the first moment of the concentration-time curve were underestimated by 72% and 22%, respectively. The dual echo-sequence combined with the postprocessing algorithm separates T1 and T2* effects and thus allows to assess cerebral hemodynamics and contrast agent kinetics simultaneously. This method may be a useful tool for characterizing, staging, and therapy monitoring of brain tumors.

Elevated Cerebrospinal Fluid Glutamate in Patients With HIV-related Dementia-Reply

In Reply. —Drs Gurwitz and Kloog raise some points regarding possible alternative interpretations of our data. Human immunodeficiency virus—related encephalitis is present in all patients with AIDS-related dementia, and its severity is correlated with the extent of viral infection and degree of dementia. 1 For this reason, we believe that an increased glutamate level in CSF can be directly explained by interaction of viral proteins or factors from virus-infected cells (cytokines, free radicals, and nitric oxide) with glutamate transporters within the central nervous system. The alternative explanation of increased CSF glutamate levels as a reflection of elevated plasma levels appears less likely, for several reasons. Plasma glutamate levels were similarly elevated in AIDS patients with and without dementia, 2 and BBB disruption also may be present in nondemented HIV-positive patients who have normal CSF glutamate levels. 3 Moreover, although glutamate levels are normally 10 times higher in blood than in CSF, in

Gene therapy for neurodegenerative disorders and malignant brain tumors

AbstractGene therapy approaches have great promise in the treatment of neurodegenerative disorders and malignant brain tumors. Neuwelt et al. review available viral-mediated gene therapy methods and their blood-brain-barrier (BBB) disruption delivery technique, briefly mentioning nonviral mediated gene therapy methods. This commentary discussed the BBB disruption delivery technique, viral and nonviral mediated gene therapy approaches to Parkinson's disease, and the potential use of antisense oligo to suppress malignant brain tumors.

Long term exposure to heat reduces edema formation after closed head injury in the rat.

Cerebral edema is one of the major consequences of head trauma (HT); its evolution may cause secondary ischemia and neuronal damage. In a closed head injury model in rats, we have shown BBB disruption and edema formation during the post traumatic period. We have previously shown that chronic exposure to moderate heat improves clinical outcome of rats subjected to HT. Long term exposure to heat results in the achievement of a stable acclimated state, characterized by a lower metabolic rate and improved heat tolerance. In the present study, we investigated the effect of chronic exposure to heat on edema formation after HT. Rats were held at 24 degrees C (CON) or 34 degrees C (ACC) for one month. Injury was then induced under ether anesthesia by a weight drop device. Four or 48 hours later, they were sacrificed for evaluation of BBB integrity (Evans blue, EB, extravasation) or edema formation (specific gravity, SG, or percent water). We found that EB uptake by the contused hemisphere was 6 fold lower in the ACC rats as compared to CON (p < 0.001). Furthermore, edema measured at 48 h by both SG and percent water methods was significantly lower in the acclimated rats (p < 0.01). We suggest that heat acclimation offers protection to rats subjected to head injury, possibly by reduction of plasma proteins extravasation.

Changes in brain polyamine levels following head injury

The changes in polyamines levels in the brain after closed head injury were studied in rats. At 1 and 15 min, 24 and 48 h after closed head injury cortical tissue from the site of injury, from the contralateral region, and from remote areas were taken. The levels of the diamine putrescine and the polyamines spermine and spermidine were assayed by thin layer liquid chromatography of their dansyl derivatives. Head injury induced a significant increase in putrescine at 48 h at the site of injury and in the frontal lobe of the injured hemisphere, respectively. In the contralateral hemisphere only minor changes in putrescine were found. Spermine and spermidine showed minor changes at that time course. We have previously shown that at 24–48 h after injury, severe edema is found at the site injury. In order to study the role of putrescine in edema formation in this model we treated the traumatized rats with α-difluoromethyl-ornithine (DFMO), an inhibitor of ornithine-decarboxylase, the rate limiting enzyme in putrescine biosynthesis. This drug did not affect the level of edema 4 or 48 h after injury although it abolished the increase in putrescine. The effect of DFMO on blood-brain barrier function was studied, using Evans blue extravasation, at the early post-traumatic period (15 min-4 h), where a massive amount of dye is taken up by traumatized brain. No changes in the amount of dye extracted was found after DFMO treatment. On the other hand, DFMO had a beneficial effect on the neurological outcome, as evaluated by a set of clinical criteria. These results demonstrate the activation of the biosynthetic pathway of polyamines as a result of primary mechanical injury to the brain. Their role in early BBB disruption and in the development of edema at the delayed (48 h) phase can be ruled out based on the present results, since DFMO, despite its effective inhibition of putrescine, was ineffective in protecting BBB or reducing edema. However, the improvement of the clinical outcome points to a role these compounds might play in the development of delayed neuronal damage.

Dextran-magnetite particles: contrast-enhanced MRI of blood-brain barrier disruption in a rat model.

Dextran-magnetite particles (DMP) were studied for their use as a MR contrast agent to visualize lesions with a blood-brain barrier (bbb) disruption. A freezing injury to the rat cerebral cortex was used as a model of bbb disruption. The biodistribution of iv-injected DMP was studied using atomic absorption spectrophotometry, electron microscopy, and MRI. One hour after injection, focal accumulation of the particles in capillary endothelial cells could be demonstrated in the freezing lesion. Despite the observation that the relaxivity of DMP in vivo appears to be less well pronounced than that in vitro, the MR imaging studies show that DMP can be used to visualize bbb disruption with adequate contrast.

The fallacy of the localized supratentorial malignant glioma.

Supratentorial glioblastoma multiforme (GBM) is a lethal malignancy. Subtotal surgical resection and post operative external beam radiotherapy palliate symptoms and prolong survival, but very few patients survive more than 3 years following diagnosis.‘,3,20*32,43,46,48*5’*58,66 Whereas the probability of survival of supratentorial anaplastic astrocytoma (AA) is slightly better, most patients will still succumb to the tumor.58-60*66 The resistance of these neoplasms to conventional therapies has prompted many physicians to undertake new clinical and laboratory investigations. These have included the use of chemotherapy,44 immunotherapy,40 multiple-fraction-perday radiotherapy,2’,34,49 radiotherapy with chemical radiosensitizers,6’ and neutron radiotherapy.45 As interest in the biology and treatment of malignant gliomas has increased, many conceptual verities have been proposed to guide therapeutic investigations, These include the significance of the cellular heterogeneity of AA and GBM,‘2,24 the presence of the blood-brain-barrier (BBB) as a functional obstruction to chemotherapy, and the local nature of these tumors. In this editorial, we focus on the last of these premises. We have selected this issue because of its major implications for any treatment that is directed at a specific portion of the brain rather than the whole brain: BBB disruption, intracarotid drug administration, and brain brachytherapy. Interstitial brachytherapy is used for primary and recurrent GBM and AA. The technique consists of the stereotactic computed tomography (CT) or magnetic resonance imaging (MRI) guided placement of coaxial catheters within and around the tumor. Radioactive sources are after-loaded into the catheters to deliver continuous low-dose rate irradiation. After a specified dose is delivered, the system is removed.22~27~28~39~45 Interstitial hyperthermia generated by implanted microwave antennae may be administered in conjunction with the radiation.52 One of the justifications invoked for brachytherapy is the concept of malignant gliomas as localizedlesions. Allusions to this concept abound in the literature. “Most malignant gliomas are localized to a single area of the brain, central nervous system (CNS) metastases from these tumors are uncommon and systemic metastases are rare.“29 “Since an astrocytoma is almost invariably a nonmetastatizing single mass, ideally it should lend itself best to regional treatment.“** “Because a primary brain tumor is most often a localized disease, hyperthermia, intratumoral chemotherapy, and interstitial irradiation, the stereotactic placement of radioactive sources directly into brain tumors, have been suggested as possible local treatments for brain tumors.“*’

Ischemic brain edema and the osmotic gradient between blood and brain.

The relationship of the osmotic pressure gradient between blood and brain, and the development of ischemic brain edema was studied. Focal cerebral ischemia was produced by left middle cerebral artery occlusion in rats. Brain osmolality was determined with a vapor pressure osmometer, brain water content by wet-dry weight, and tissue sodium and potassium contents by flame photometry. Permeability of the BBB was tested by Evans blue. Measurements were made from the ischemic cortex within 14 days of occlusion. Brain osmolality increased from 311 +/- 2 to 329 +/- 2 mOsm/kg by 6 h after occlusion. Serum osmolality did not change significantly. The osmotic gradient between blood and brain peaked at approximately 26 mOsm/kg. Brain osmolality then decreased to 310 +/- 2 mOsm/kg by 12 h after occlusion and remained at about that same level. Water content increased progressively within 1 day of occlusion, then gradually decreased by 14 days. Brain tissue sodium plus potassium content did not increase within 6 h of occlusion, and Evans blue extravasation was not seen within that time. These findings indicate that an osmotic pressure gradient contributes to the formation of edema only during the early stage of cerebral ischemia. Furthermore, the increase in brain osmolality is not related to tissue electrolyte change or BBB disruption to protein.

The effect of systemic arterial hypertension on blood-to-tissue transport in experimental gliomas.

Systemic arterial hypertension was induced with epinephrine in 15 rats with 39 transplanted RG-2 brain tumors in an attempt to increase blood-to-tissue transport of a water-soluble compound. In 4 rats, hypertension was induced acutely (less than 5 sec), and in 11 hypertension was induced more slowly (over 5 min). Regional values of the unidirectional blood-to-tissue transfer constant (K) of alpha aminoisobutyric acid were measured with quantitative autoradiography. Mean arterial blood pressure (BP) over the experimental period increased from 117 +/- 17 mmHg (SD) to 168 +/- 18 mmHg in the rats with slowly induced hypertension, and from 124 +/- 4 to 142 +/- 5 mmHg in the acute hypertension group. Peak BP was 208 +/- 16 in the first group and 216 +/- 13 mmHg in the second. Intracerebral hemorrhage occurred in 10/15 animals, and there was disruption of BBB in tumor-free brain in 10/15 animals. Averaged mean whole tumor K of AIB in all hypertensive rats was 0.052 +/- 0.022 ml/g/min, compared to 0.037 +/- 0.015 ml/g/min in normotensive controls; there was no difference in mean tumor K between the two hypertensive groups. However, in intraparenchymal tumors without hemorrhage, K was only 0.039 +/- 0.013 ml/g/min. Although the mean K of AIB was higher in brain tumors of the hypertensive rats, the increase is unlikely to be meaningful in terms of augmented delivery of water-soluble drugs to brain tumors, and the high incidence of intracerebral hemorrhage countermands any clinical use of this approach.

CSF-plasma relationships for DSIP and some other neuropeptides

The relationships between CSF and plasma hormonal levels of several peptides were studied in the same samples of simultaneously obtained plasma and CSF. A significant correlation existed between CSF and plasma levels of DSIP as well as gastrin. Preliminary results also showed a correlation between CSF and plasma levels of NT, but not VIP or calcitonin. CSF/plasma ratios and the effect of BBB disruption also varied from peptide to peptide. These diverse CSF/plasma relationships are not easily explained by models of nonspecific passage but may indicate individual systems or axes that could be involved in the central effects of peripherally administered peptides.