Cold-injury of the cerebral cortex: immunolocalization of cellular proteins and blood-brain barrier permeability studies

The occurrence of blood-brain barrier (BBB) permeability alterations and neovascularization are well documented in the cerebral cortical cold-injury model. This model was used to determine whether the glucose transporter (glutI) protein was present in endothelium of cerebral vessels with breakdown of BBB to protein and when regenerating endothelial cells become immunoreactive for glutI protein. Secondly, the protein products of c-fos and c-jun were localized to determine whether these early immediate genes are activated in this model. Observations were made over a period of 12 hours to 14 days after the cold-injury. Blood-brain barrier permeability was assessed using horseradish peroxidase (HRP) as a tracer. Since HRP may not be able to enter thrombosed vessels within the cold lesion, immunohistochemistry was used to detect extravasation of endogenous serum proteins using antisera to rat serum proteins. The proteins-glut1, GFAP, c-fos and c-jun-were localized by immunohistochemistry. Endothelium of vessels which were permeable to protein, whether in the cold-injury site or in the perilesional area, all contained glut1 protein; hence, the presence of glut1 did not appear to correlate with an intact BBB to protein. An interesting point is that in the process of neovascularization, regenerating endothelial cells become immunoreactive for glut1 at 5 days and this coincides with the presence of tight junctions in these cells. Immunoreactivity for c-fos was observed in regenerating endothelium within the lesion site, in astrocytes, and to a lesser extent in endothelial cells and neurons in the perilesional area. Few astrocytes showed immunoreactivity for c-jun at 4 and 5 days. Possibly, the growth factors generated to promote angiogenesis and repair led to activation of the c-fos gene with deposition of c-fos protein. The results suggest that during nervous system development or endothelial regeneration, the presence of glut1 in cerebral endothelium coincides with the presence of an intact BBB to protein and protein tracers. However, in pathological states presence of glut1 in cerebral endothelium does not appear to correlate with an intact BBB to protein. This model lends itself to the study of angiogenesis and repair processes in the cerebral cortex in an environment unaffected by ischemia and thus the findings may be relevant to traumatic injuries of the human cerebral cortex.     

The effect of photon irradiation on blood-brain barrier permeability to methotrexate in mice

Methotrexate was administered by intraperitoneal injection (100 mg/kg) to unirradiated mice, and to mice receiving varying doses of cranial irradiation. The animals were sacrificed 24 hours after injection, and methotrexate assays were performed on brain tissue. No methotrexate was detected in the brains of the unirradiated animals. Detectable levels of methotrexate were present after 2000 rad cranial irradiation, but not after 500 rad, 1000 rad, or 1500 rad. The implications of these findings are discussed.     

Insight into the regulation by second messenger molecules of the permeability of the blood-brain barrier

Recent advances in our knowledge of the blood-brain barrier have in part been made by studying the properties and function of cerebral endothelial cells in vitro. After an era of working with a fraction, enriched in cerebral microvessels by centrifugation, the next generation of in vitro blood-brain barrier model systems was introduced, when the conditions for routinely culturing the endothelial cells were established. This review summarizes the results obtained mainly from this in vitro approach. Different elements of the intracellular signaling messenger systems have been detected in the course of our studies in the cerebral endothelial cells. It has been shown that the synthesizing enzymes of and substrate proteins for the second messenger molecules are present in the cerebral endothelial cells, and their activity and/or amount can change in pathological circumstances, i.e., during the formation of brain oedema. Pharmacological treatments interfering with the second messenger systems proved to be effective in the prevention of brain oedema formation.     

Differential permeability of the blood-brain barrier to two pancreatic peptides: insulin and amylin

OBJECTIVE: To review the available evidence regarding efficacy, benefits, and risks of magnesium sulfate seizure prophylaxis in women with preeclampsia or eclampsia. DATA SOURCES: The English-language literature in MEDLINE was searched from 1966 through February 1998 using the terms "magnesium sulfate," "seizure," "preeclampsia," "eclampsia," and "hypertension in pregnancy." Reviews of bibliographies of retrieved articles and consultation with experts in the field provided additional references. METHODS OF STUDY SELECTION: All relevant English-language clinical research articles retrieved were reviewed. Randomized controlled trials, retrospective reviews, and observational studies specifically addressing efficacy, benefits, or side effects of magnesium sulfate therapy in preeclampsia or eclampsia were chosen. TABULATION, INTEGRATION, AND RESULTS: Nineteen randomized controlled trials, five retrospective studies, and eight observational reports were reviewed. The criteria used for inclusion were as follows: randomized controlled trials evaluating use of magnesium sulfate in eclampsia, preeclampsia, and hypertensive disorders of pregnancy; nonrandomized studies of historical interest; "classic" observational studies; and recent retrospective studies evaluating efficacy of magnesium sulfate therapy, using relative risk and 95% confidence intervals where applicable. Magnesium sulfate therapy has been associated with increased length of labor, increased cesarean delivery rate, increased postpartum bleeding, increased respiratory depression, decreased neuromuscular transmission, and maternal death from overdose. A summary of randomized, controlled trials in women with eclampsia reveals recurrent seizures in 216 (23.1%) of 935 women treated with phenytoin or diazepam, compared with recurrent seizures in only 88 (9.4%) of 932 magnesium-treated women. Randomized controlled trials in women with severe preeclampsia collectively revealed seizures in 22 (2.8%) of 793 women treated with antihypertensive agents, compared with seizures in only seven of 815 (0.9%) magnesium-treated women. CONCLUSION: The evidence to date confirms the efficacy of magnesium sulfate therapy for women with eclampsia and severe preeclampsia. However, there is a need for a randomized controlled trial to determine efficacy of magnesium sulfate therapy for women with mild preeclampsia and gestational hypertension.     

Reversible blood-brain barrier dysfunction to peroxidase after a small stab wound in the rat cerebral cortex

Small stab wounds were made in the rat frontal lobe. The animals were injected with horseradish peroxidase intravenously at different times after the injury in order to study the extravasation of this tracer. There was a leakage of peroxidase into the brain during the first 3 days after the injury. The route of passage from the vessel lumen into the brain was through disrupted blood vessels in the injured region. Endothelial pinocytosis and formation of thin, trans-endothelial channel-like structures with or without a content of peroxidase were two other possible routes of passage across the blood vessels. Occasionally, badly damaged endothelial cells displayed a diffuse cytoplasmic distribution of peroxidase, indication a diffusion into and possibly across these injured cells. No widened tight junctions were seen. Thus, this study indicated four possible routes of passage of horseradish peroxidase across the endothelial cells: cellular gross damage with disrupture of the cells, diffusion across badly injured endothelial cells, possibly pinocytosis and formation of trans-endothelial channel-like structures. The cellular uptake of the tracer was vesicular in most neurons, astrocytes, oligodendrocytes and hematogeneous phagocytes. However, a diffuse distribution of the tracer was seen in some "dark" neurons near leaking vessels in the vicinity of the stab wound.     

Mechanisms of edema formation after intracerebral hemorrhage: effects of thrombin on cerebral blood flow, blood-brain barrier permeability, and cell survival in a rat model

Recently, the authors showed that thrombin contributes to the formation of brain edema following intracerebral hemorrhage. The current study examines whether the action of thrombin is due to an effect on cerebral blood flow (CBF), vasoreactivity, blood-brain barrier (BBB) function, or cell viability. In vivo solutions of thrombin were infused stereotactically into the right basal ganglia of rats. The animals were sacrificed 24 hours later; CBF and BBB permeability were measured. The actions of thrombin on vasoreactivity were examined in vitro by superfusing thrombin on cortical brain slices while monitoring microvessel diameter with videomicroscopy. In separate experiments C6 glioma cells were exposed to various concentrations of thrombin, and lactate dehydrogenase release, a marker of cell death, was measured. The results indicate that thrombin induces BBB disruption as well as death of parenchymal cells, whereas CBF and vasoreactivity are not altered. The authors conclude that cell toxicity and BBB disruption by thrombin are triggering mechanisms for the edema formation that follows intracerebral hemorrhage.     

Intracellular delivery of lipopolysaccharide during DNA transfection activates a lipid A-dependent cell death response that can be prevented by polymyxin B

The presence of lipopolysaccharide (LPS) as a contaminant in plasmid DNA prepared from Escherichia coli is well documented, and we have previously demonstrated that LPS internalization during adenovirus-mediated gene transfer can generate a toxicity in some primary cell types. We demonstrate here that in addition to adenoviral systems, several commonly used nonviral methods of gene transfer also activate this cell death response in the presence of LPS. Subcomponents of LPS were analyzed and the toxicity was found to be due to the lipid A component of LPS. The LPS-chelating antibiotic polymyxin B, when present at concentration of 10-30 micrograms/ml, can block this toxicity.     

Vascular clogging, mononuclear cell margination, and enhanced vascular permeability in the pathogenesis of human cerebral malaria

To document histopathologic evidence on the pathogenic mechanism of human cerebral malaria, we used light microscopy to study brain specimens from 23 patients who died of central nervous system involvement with Plasmodium falciparum. Sequestration of parasitized red blood cells (PRBCs) leading to cerebral capillary clogging was seen. In a few specimens, vascular clogging by PRBCs was associated with margination of mononuclear cells. In others, capillaries were virtually empty and lymphocytes and monocytes were seen in apposition (marginated) to the capillary endothelial surface. The endothelial cells appeared plump, hypertrophied, and prominent. The capillary wall appeared thickened by fibrinous material. Massive intercellular brain edema along with extravasated red blood cells, mononuclear cells, and plasmatic fluid was also noticed. In addition to hypoxia induced by PRBC-mediated vascular clogging, marginating mononuclear cells may contribute to the pathogenesis of cerebral malaria. The precise role played by this phenomenon needs further evaluation.     

Importance of platelets in increased vascular permeability evoked by experimental haemarthrosis in synovium of the rat

Increased vascular permeability of synovium induced by experimental haemarthrosis was studied in the stifle joint of the rat. Abnormal permeability was detected by injecting animals intravenously with colloidal carbon and examining the synovial vessels for intramural deposits of carbon. Both fresh and heparinized whole blood injected into the joint induced a marked permeability response of synovial venules which persisted for 18 hours. Platelets suspended in heparinized Tyrode solution induced a similar response, but one lasting only 12 hours. The permeability effects of suspensions of leucocytes and erythrocytes on the synovial vasculature were relatively mild and were maximal 5 and 12 h respectively after intra-articular injection. Heparinized platelet-free plasma had no significant permeability effects. These results indicate that platelets may play an important role in the pathogenesis of synovial inflammation in haemarthrosis.     

Blood-brain barrier permeability during the development of experimental bacterial meningitis in the rat

A 60-year-old man presented with progressive large fiber sensory loss in the right first three fingers and, to a lesser extent, in both fourth and fifth fingers. Electrophysiologic studies were characteristic of chronic sensory demyelinating polyneuropathy, a variant of chronic inflammatory demyelinating polyneuropathy. Plasma exchange was unsuccessful, but intravenous immunoglobulin (IVIG) led to complete recovery of sensation for 2 months, although neurophysiologic abnormalities persisted. A battery of noninvasive tests to measure hand grip strength, tactile sensation at the fingertips, and motor control of prehension during precision grip revealed marked abnormalities in the right hand before IVIG. One month after IVIG, all test results had normalized, but they returned to pretreatment levels after 3 months. Functional evaluation of the hand may be a sensitive method to objectively quantify loss of and changes in cutaneous mechanoreceptor function of the fingers in large fiber sensory neuropathy.     

Latent inhibition in rats is abolished by NMDA-induced neuronal loss in the retrohippocampal region, but this lesion effect can be prevented by systemic haloperidol treatment.

Latent inhibition (LI) refers to the retardation in learning about the significance of a neutral stimulus that results from its nonreinforced preexposure. There is evidence that electrolytic or aspiration lesions of the hippocampal formation can disrupt LI (I. Weiner, see record 1991-06652-001). It has been suggested that this effect may stem from the interruption of a projection from the retrohippocampal region to the nucleus accumbens (A. J. M. Clark et al, 1992). The present experiment assessed this possibility by comparing LI in rats with retrohippocampal N-methyl-D-aspartate (NMDA) lesions extending from the entorhinal cortex to the ventral subiculum to that seen in vehicle controls and unoperated controls. LI was abolished by the retrohippocampal lesion. The effect of the lesion on LI was prevented by treatment with systemic haloperidol (0.2 mg/kg). The results are discussed with respect to an animal model of schizophrenia. (PsycINFO Database Record (c) 2010 APA, all rights reserved)     

Lack of autoregulatory response of the cerebral circulation after osmotic opening of the blood-brain barrier in dogs

The reversibility of osmotic opening of the blood-brain barrier was studied in dogs one hour after intracarotid 3 M urea injection. At that time the permeability of cerebral blood vessels to albumin is restored as evidenced by lack of Evans blue extravasation. Despite that, the response of the urea-perfused hemisphere to changes of perfusion pressure was abnormal. Blood flow in that hemisphere followed passively blood pressure changes in contrast to the contralateral hemisphere in which the blood flow remained independent of the perfusion pressure.     

Vulnerability to cerebral hypoxic-ischemic insult in neonatal but not in adult rats is in parallel with disruption of the blood-brain barrier

BACKGROUND AND PURPOSE: Vulnerability to cerebral hypoxic-ischemic (H-I) insult and its relation to disruption of the blood-brain barrier were investigated in postnatal rats. METHODS: Pups of postnatal day (P) 7, P14, and P21 underwent ligation of a unilateral carotid artery and were exposed to hypoxic conditions. For the detection of early-phase deterioration, brains were perfusion-fixed 24 hours after H-I insult and examined by argyrophil III method. For the detection of later infarction, animals were fixed at 72 hours after the H-I insult. RESULTS: In either case, tissue damage was detected in the striatum, parietal cortex, and hippocampus. The vulnerability of P7 and P21 rats was remarkable, as compared with P14 rats. Although the developmental status of the vasculature was not significantly different at each age, the permeability of IgG after H-I injury was prominent in P7 rats and to a lesser extent in P14 rats. In P21 rats, however, there was little IgG leakage even 24 hours after the insult. Dexamethasone pretreatment blocked the extravasation of IgG and reduced the damaged tissue in P7 and P14 rats but not in P21 rats. Percentages of reduction in infarcted areas by the dexamethasone became smaller in proportion to ages. CONCLUSIONS: The results suggest that in younger rats vulnerability to H-I insult was in parallel with permeability of the blood-brain barrier, whereas in adults in might be more dependent on cellular vulnerability.     

Cloned lines of Plasmodium berghei ANKA differ in their abilities to induce experimental cerebral malaria

Infection with Plasmodium berghei ANKA is usually lethal. The parasite causes in some mouse strains a neurovascular syndrome, experimental cerebral malaria (ECM), involving immunopathological reactions. The effects on the development of ECM of the mouse genetic background have been clearly demonstrated, but nothing is known about the effects of the clonal diversity of the parasite. We showed that various cloned lines derived from a polyclonal line of P. berghei ANKA caused ECM but that the extent of ECM induction was dependent on the amount of inoculum. Subtle differences in ECM characteristics (survival time and hypothermia) were also observed. We also confirmed, using the 1.49L cloned line, that the mouse genetic background strongly affects ECM.     

Levofloxacin versus ciprofloxacin, flucloxacillin, or vancomycin for treatment of experimental endocarditis due to methicillin-susceptible or -resistant Staphylococcus aureus

Levofloxacin is the L isomer of ofloxacin, a racemic mixture in which the L stereochemical form carries the antimicrobial activity. Levofloxacin is more active than former quinolones against gram-positive bacteria, making it potentially useful against such pathogens. In this study, levofloxacin was compared to ciprofloxacin, flucloxacillin, and vancomycin for the treatment of experimental endocarditis due to two methicillin-susceptible Staphylococcus aureus (MSSA) and two methicillin-resistant S. aureus (MRSA) isolates. The four test organisms were susceptible to ciprofloxacin, the levofloxacin MICs for the organisms were low (0.12 to 0.25 mg/liter), and the organisms were killed in vitro by drug concentrations simulating both the peak and trough levels achieved in human serum (5 and 0.5 mg/liter, respectively) during levofloxacin therapy. Rats with aortic endocarditis were treated for 3 days. Antibiotics were injected with a programmable pump to simulate the kinetics of either levofloxacin (350 mg orally once a day), ciprofloxacin (750 mg orally twice a day), flucloxacillin (2 g intravenously four times a day), or vancomycin (1 g intravenously twice a day). Levofloxacin tended to be superior to ciprofloxacin in therapeutic experiments (P = 0.08). More importantly, levofloxacin did not select for resistance in the animals, in contrast to ciprofloxacin. The lower propensity of levofloxacin than ciprofloxacin to select for quinolone resistance was also clearly demonstrated in vitro. Finally, the effectiveness of this simulation of oral levofloxacin therapy was at least equivalent to that of standard treatment for MSSA or MRSA endocarditis with either flucloxacillin or vancomycin. This is noteworthy, because oral antibiotics are not expected to succeed in the treatment of severe staphylococcal infections. These good results obtained with animals suggest that levofloxacin might deserve consideration for further study in the treatment of infections due to ciprofloxacin-susceptible staphylococci in humans.     

E- and P-selectin are not involved in the recruitment of inflammatory cells across the blood-brain barrier in experimental autoimmune encephalomyelitis

In experimental autoimmune encephalomyelitis (EAE) inflammatory cells cross the endothelial blood-brain barrier (BBB) and gain access to the central nervous system (CNS). Here we show that E- and P-selectin are not involved in the recruitment of inflammatory cells across the BBB. Neither expression of E- nor P-selectin is induced in BBB-forming endothelium at any time after initiation of EAE. Some of the inflammatory cells present in the CNS during EAE express ligands for E- or P-selectin. However, anti-E- and P-selectin antibodies influence neither immigration of inflammatory cells across the BBB nor the development of EAE. In general, suppression of E- and P-selectin expression on BBB endothelium is dependent on factors derived from the CNS microenvironment, eg, astrocytes. Our results suggest that during EAE suppression of E- and P-selectin expression on the BBB provides a CNS-specific mechanism to reduce leukocyte recruitment into the CNS.     

Phospholipid composition of the mammalian red cell membrane can be rationalized by a superlattice model

Although the phospholipid composition of the erythrocyte membrane has been studied extensively, it remains an enigma as to how the observed composition arises and is maintained. We show here that the phospholipid composition of the human erythrocyte membrane as a whole, as well as the composition of its individual leaflets, is closely predicted by a model proposing that phospholipid head groups tend to adopt regular, superlattice-like lateral distributions. The phospholipid composition of the erythrocyte membrane from most other mammalian species, as well as of the platelet plasma membrane, also agrees closely with the predictions of the superlattice model. Statistical analyses indicate that the agreement between the observed and predicted compositions is highly significant, thus suggesting that head group superlattices may indeed play a central role in the maintenance of the phospholipid composition of the erythrocyte membrane.