Expression of Bone Morphogenetic Proteins in Multiple Sclerosis Lesions.

Bone morphogenetic proteins (BMPs) are secreted proteins that belong to the transforming growth factor-ß superfamily. In the adult brain, they modulate neurogenesis, favor astrogliogenesis, and inhibit oligodendrogenesis. Because BMPs may be involved in the failure of remyelination in multiple sclerosis (MS), we characterized the expression of BMP-2, BMP-4, BMP-5, and BMP-7; BMP type II receptor (BMPRII); and phosphorylated SMAD (pSMAD) 1/5/8 in lesions of MS and other demyelinating diseases. A total of 42 MS lesions, 12 acute ischemic lesions, 8 progressive multifocal leukoencephalopathy lesions, and 10 central nervous system areas from four nonneuropathological patients were included. Lesions were histologically classified according to the inflammatory activity. The expression of BMP-2, BMP-4, BMP-5, BMP-7, BMPRII, and pSMAD1/5/8 was quantified by immunostaining, and colocalization studies were performed. In MS lesions, astrocytes, microglia/macrophages, and neurons expressed BMP-2, BMP-4, BMP-5, and BMP-7; BMPRII; and pSMAD1/5/8. Oligodendrocytes expressed BMP-2 and BMP-7 and pSMAD1/5/8. The percentage of cells that expressed BMPs, BMPRII, and pSMAD1/5/8 correlated with the inflammatory activity of MS lesions, and changes in the percentage of positive cells were more relevant in MS than in other white matter-damaging diseases. These data indicate that BMPs are increased in active MS lesions, suggesting a possible role in MS pathogenesis.

Proinsulin protects against age-related cognitive loss through anti-inflammatory convergent pathways.

Brain inflammaging is increasingly considered as contributing to age-related cognitive loss and neurodegeneration. Despite intensive research in multiple models, no clinically effective pharmacological treatment has been found yet. Here, in the mouse model of brain senescence SAMP8, we tested the effects of proinsulin, a promising neuroprotective agent that was previously proven to be effective in mouse models of retinal neurodegeneration. Proinsulin is the precursor of the hormone insulin but also upholds developmental physiological effects, particularly as a survival factor for neural cells. Adeno-associated viral vectors of serotype 1 bearing the human proinsulin gene were administered intramuscularly to obtain a sustained release of proinsulin into the blood stream, which was able to reach the target area of the hippocampus. SAMP8 mice and the control strain SAMR1 were treated at 1 month of age. At 6 months, behavioral testing exhibited cognitive loss in SAMP8 mice treated with the null vector. Remarkably, the cognitive performance achieved in spatial and recognition tasks by SAMP8 mice treated with proinsulin was similar to that of SAMR1 mice. In the hippocampus, proinsulin induced the activation of neuroprotective pathways and the downstream signaling cascade, leading to the decrease of neuroinflammatory markers. Furthermore, the decrease of astrocyte reactivity was a central effect, as demonstrated in the connectome network of changes induced by proinsulin. Therefore, the neuroprotective effects of human proinsulin unveil a new pharmacological potential therapy in the fight against cognitive loss in the elderly.

peIF4E as an independent prognostic factor and a potential therapeutic target in diffuse infiltrating astrocytomas.

Malignant transformation in tumors is a complex process requiring accumulation of numerous oncogenic abnormalities. Brain tumors show considerable phenotypic and genetic heterogeneity. In a series comprising diffuse infiltrating astrocytomas (DIA) and reactive gliosis, we investigated the main factors associated with signaling pathways. We assessed expression levels and their association with tumor progression and survival. We studied 19 grade II astrocytomas, 25 anaplastic astrocytomas (grade III), 60 glioblastomas (grade IV), and 15 cases of reactive gliosis. Epidermal growth factor receptor (EGFR), pMAPK, 4E-BP1, p4E-BP1, pS6, eIF4E, and peIF4E expression levels were evaluated using immunohistochemistry. Expression levels were semiquantitatively evaluated using a histoscore. Immunohistochemistry and PCR were used for IDH1 mutations. Statistical analysis was based on the following tests: chi-square, Student's t, Pearson correlation, Spearman's rho, and Mann-Whitney; ROC and Kaplan-Meier curves were constructed. A significant increase was observed between grades for expression of total and phosphorylated 4E-BP1 and for eIF4E, Ki67, EGFR, and cyclin D1. Although expression of EGFR, eIF4E, and Ki67 correlated with survival, only peIF4E was an independent predictor of survival in the multivariate analysis. Combining the evaluation of different proteins enables us to generate helpful diagnostic nomograms. In conclusion, cell signaling pathways are activated in DIAs; peIF4E is an independent prognostic factor and a promising therapeutic target. Joint analysis of the expression of 4E-BP1 and peIF4E could be helpful in the diagnosis of glioblastoma multiforme in small biopsy samples.

In vivo evaluation of amyloid deposition and brain glucose metabolism of 5XFAD mice using positron emission tomography.

Positron emission tomography (PET) has been used extensively to evaluate the neuropathology of Alzheimer's disease (AD) in vivo. Radiotracers directed toward the amyloid deposition such as [(18)F]-FDDNP (2-(1-{6-[(2-[F]Fluoroethyl)(methyl)amino]-2-naphthyl}ethylidene)malononitrile) and [(11)C]-PIB (Pittsburg compound B) have shown exceptional value in animal models and AD patients. Previously, the glucose analogue [(18)F]-FDG (2-[(18)F]fluorodeoxyglucose) allowed researchers and clinicians to evaluate the brain glucose consumption and proved its utility for the early diagnosis and the monitoring of the progression of AD. Animal models of AD are based on the transgenic expression of different human mutant genes linked to familial AD. The novel transgenic 5XFAD mouse containing 5 mutated genes in its genome has been proposed as an AD model with rapid and massive cerebral amyloid deposition. PET studies performed with animal-dedicated scanners indicate that PET with amyloid-targeted radiotracers can detect the pathological amyloid deposition in transgenic mice and rats. However, in other studies no differences were found between transgenic mice and their wild type littermates. We sought to investigate in 5XFAD mice if the radiotracers [(11)C]-PIB, and [(18)F]-Florbetapir could quantify the amyloid deposition in vivo and if [(18)F]-FDG could do so with regard to glucose consumption. We found that 5XFAD animals presented higher cerebral binding of [(18)F]-Florbetapir, [(11)C]-PIB, and [(18)F]-FDG. These results support the use of amyloid PET radiotracers for the evaluation of AD animal models. Probably, the increased uptake observed with [(18)F]-FDG is a consequence of glial activation that occurs in 5XFAD mice.

[Clinical variability of polymicrogiria: report of 35 new cases and review of the literature]. / Variabilidad clínica de la polimicrogiria: a propósito de 35 nuevos casos y revisión de la bibliografía.

INTRODUCTION: The study of polymicrogyria with magnetic resonance imaging (MRI) has made possible the report of several series of patients in which the main clinical manifestations differ considerably. The aims of the study were to review the literature and to know the clinical variability of the patients attended in a neuropediatric service. PATIENTS AND METHODS: A retrospective study was conducted between 1989-2011 for the patients attended in our neuro-pediatric service and diagnosed of polymicrogyria by MRI. RESULTS: On the totality of 44 patients having polymicrogyria, 9 did not satisfy de inclusion criteria (Barkovich's radiological criteria). The polymicrogyria was bilateral in 22/35 patients (1 frontal, 22 perisylvian) and unilateral in 13/35 (2 frontal, the rest perisylvian). All patients with bilateral polymicrogyria had intellectual disability, 71% had global development delay, 75% had oromotor disorder and 40% had epilepsy. Patients with unilateral polymicrogyria had the following symptoms: 65% intellectual disability, 55% global development delay, 55% oromotor disorder, 55% epilepsy and 2 patients where free of symptoms (the oldest 2 year old). The initial symptoms were depending upon the age: the oromotor disorder was the most common in the newborn period, global development delay if the symptoms started before 2 years old and after 2 years epilepsy was the initial most common symptom. CONCLUSION: In our study the most common symptom was intellectual disability (independently of the type of poly-microgyria), followed by oromotor disorder and, with fewer proportion, epilepsy (in contrast with other series).

Variabilidad clínica de la polimicrogiria: a propósito de 35 nuevos casos y revisión de la bibliografía / Clinical variability of polymicrogiria: report of 35 new cases and review of the literatureepi

Introducción. El estudio diagnóstico de polimicrogiria mediante resonancia magnética ha facilitado la publicación de varias series de pacientes en las que las manifestaciones clínicas predominantes varían considerablemente. Objetivo. Conocer la variabilidad fenotípica de la polimicrogiria basándose en la serie de pacientes atendidos en nuestro servicio y la revisión de la bibliografía. Pacientes y métodos. Estudio retrospectivo de los pacientes diagnosticados de polimicrogiria mediante resonancia magnética y seguidos en consultas durante los años 1989-2011.Resultados. De un total de 44 pacientes, nueve fueron excluidos por no cumplir los criterios diagnósticos radiológicospropuestos por Barkovich. La polimicrogiria fue bilateral en 22/35 pacientes (una frontal, 21 perisilvianas) y unilateral en13/35 (dos frontales, el resto perisilvianas). Todos los pacientes con polimicrogiria bilateral tenían algún tipo de discapacidadintelectual, un 71% tenía retraso global del desarrollo, un 75% tenía trastorno oromotor y un 40% tenía epilepsia. Los pacientes con polimicrogiria unilateral presentaron discapacidad intelectual (65%), retraso global del desarrollo (55%), trastorno oromotor (55%) y epilepsia (55%), estando asintomáticos dos pacientes (2 años de edad). La presentación clínica de los pacientes dependía de la edad: en el período neonatal, el síntoma guía fue el trastorno oromotor; antes de los 2 años, el retraso global del desarrollo; y partir de los 2 años, la epilepsia. Conclusión. En este estudio, a diferencia de otras series, el síntoma más prevalente fue la discapacidad intelectual (independientemente del tipo de polimicrogiria), seguido del trastorno oromotor y, en menor medida, la epilepsia (AU)

Introduction. The study of polymicrogyria with magnetic resonance imaging (MRI) has made possible the report of severalseries of patients in which the main clinical manifestations differ considerably. The aims of the study were to review the literature and to know the clinical variability of the patients attended in a neuropediatric service. Patients and methods. A retrospective study was conducted between 1989-2011 for the patients attended in our neuropediatric service and diagnosed of polymicrogyria by MRI.Results. On the totality of 44 patients having polymicrogyria, 9 did not satisfy de inclusion criteria (Barkovich’s radiological criteria). The polymicrogyria was bilateral in 22/35 patients (1 frontal, 22 perisylvian) and unilateral in 13/35 (2 frontal, the rest perisylvian). All patients with bilateral polymicrogyria had intellectual disability, 71% had global development delay, 75% had oromotor disorder and 40% had epilepsy. Patients with unilateral polymicrogyria had the following symptoms: 65% intellectual disability, 55% global development delay, 55% oromotor disorder, 55% epilepsy and 2 patients wherefree of symptoms (the oldest 2 year old). The initial symptoms were depending upon the age: the oromotor disorder was the most common in the newborn period, global development delay if the symptoms started before 2 years old and after 2 years epilepsy was the initial most common symptom.Conclusion. In our study the most common symptom was intellectual disability (independently of the type of polymicrogyria), followed by oromotor disorder and, with fewer proportion, epilepsy (in contrast with other series) (AU)

VAP-1/SSAO plasma activity and brain expression in human hemorrhagic stroke.

BACKGROUND: Vascular adhesion protein-1 (VAP-1) is a cell surface and circulating enzyme that belongs to the semicarbazide-sensitive amine oxidase (SSAO) family, which oxidatively deaminates primary amines and is implicated in leukocyte extravasation. Our aim was to investigate the alteration of soluble VAP-1/SSAO activity in plasma samples after acute intracerebral hemorrhage (ICH) and its presence in human ICH brain tissue. METHODS: VAP-1/SSAO activity was determined in plasma of 66 ICH patients and 58 healthy controls. In addition, we assessed the expression of VAP-1/SSAO in postmortem brain tissue from hemorrhagic stroke patients by Western blot and immunohistochemistry. RESULTS: We observed significantly higher levels of plasma VAP-1/SSAO activity in patients with ICH compared to matched elderly controls (p = 0.001). Plasma VAP-1/SSAO activity <2.7 pmol/min·mg and baseline ICH volume <17 ml were independent predictors of neurological improvement after 48 h (OR 6.8, 95% CI 1.14-41.67, p = 0.035, and OR 10.64, 95% CI 1.1-100, p = 0.041, respectively), after adjustment for baseline stroke severity. We also found that membrane-bound VAP-1/SSAO levels were lower in the perihematoma region than in the corresponding contralateral brain areas of patients deceased due to ICH (p = 0.024). CONCLUSIONS: Our data demonstrate that plasma VAP-1/SSAO activity is increased in ICH and predicts neurological outcome, suggesting a possible contribution of the soluble protein in secondary brain damage. Furthermore, anti-VAP-1/SSAO strategies might be a promising approach to prevent neurological worsening following ICH.

MMP-2/MMP-9 plasma level and brain expression in cerebral amyloid angiopathy-associated hemorrhagic stroke.

Cerebral amyloid angiopathy (CAA) is one of the main causes of intracerebral hemorrhage (ICH) in the elderly. Matrix metalloproteinases (MMPs) have been implicated in blood-brain barrier disruption and ICH pathogenesis. In this study, we determined the levels MMP-2 and MMP-9 in plasma and their brain expression in CAA-associated hemorrhagic stroke. Although MMP-2 and MMP-9 plasma levels did not differ among patients and controls, their brain expression was increased in perihematoma areas of CAA-related hemorrhagic strokes compared with contralateral areas and nonhemorrhagic brains. In addition, MMP-2 reactivity was found in ß-amyloid (Aß)-damaged vessels located far from the acute ICH and in chronic microbleeds. MMP-2 expression was associated to endothelial cells, histiocytes and reactive astrocytes, whereas MMP-9 expression was restricted to inflammatory cells. In summary, MMP-2 expression within and around Aß-compromised vessels might contribute to the vasculature fatal fate, triggering an eventual bleeding.

Brain perihematoma genomic profile following spontaneous human intracerebral hemorrhage.

BACKGROUND: Spontaneous intracerebral hemorrhage (ICH) represents about 15% of all strokes and is associated with high mortality rates. Our aim was to identify the gene expression changes and biological pathways altered in the brain following ICH. METHODOLOGY/PRINCIPAL FINDINGS: Twelve brain samples were obtained from four deceased patients who suffered an ICH including perihematomal tissue (PH) and the corresponding contralateral white (CW) and grey (CG) matter. Affymetrix GeneChip platform for analysis of over 47,000 transcripts was conducted. Microarray Analysis Suite 5.0 was used to process array images and the Ingenuity Pathway Analysis System was used to analyze biological mechanisms and functions of the genes. We identified 468 genes in the PH areas displaying a different expression pattern with a fold change between -3.74 and +5.16 when compared to the contralateral areas (291 overexpressed and 177 underexpressed). The top genes which appeared most significantly overexpressed in the PH areas codify for cytokines, chemokines, coagulation factors, cell growth and proliferation factors while the underexpressed codify for proteins involved in cell cycle or neurotrophins. Validation and replication studies at gene and protein level in brain samples confirmed microarray results. CONCLUSIONS: The genomic responses identified in this study provide valuable information about potential biomarkers and target molecules altered in the perihematomal regions.

The proteome of human brain after ischemic stroke.

Although stroke is among the most common causes of death and chronic disability worldwide, the proteome of the ischemic human brain remains unknown. Only a few studies have investigated the ischemic brain proteome in rodent stroke models. We performed a proteomic study of the human brain after ischemic stroke using a 2-dimensional differential gel electrophoresis-based proteomic approach. In brain samples from 6 deceased stroke patients and 3 control subjects, there was an average of 1,442 ± 231 protein spots in the gels. Changes of at least 1.5-fold in the relative expression of 132 protein spots between different cerebral areas (infarct core, peri-infarct, and contralateral tissue) were identified (p < 0.05); 39 of these were successfully identified by matrix-assisted laser desorption/ionization-time of flight mass spectrometry. Among the identified protein spots, we validated the results of 10 proteins by Western blot and determined the cellular localization in brain parenchyma for 3 of the identified proteins: dihydropyrimidinase-related protein 2, vesicle-fusing ATPase, and Rho dissociation inhibitor 1. These results contribute to understanding the processes that follow cerebral ischemia; moreover, some of the identified proteins may be therapeutic targets or biologic markers for determining the diagnosis and prognosis of stroke.

Plasma VAP-1/SSAO activity predicts intracranial hemorrhages and adverse neurological outcome after tissue plasminogen activator treatment in stroke.

BACKGROUND AND PURPOSE: Vascular adhesion protein-1 (VAP-1) is a cell surface and circulating enzyme involved in recruitment of lymphocytes and neutrophils through its semicarbazide-sensitive amine oxidase (SSAO) activity. We aimed to study plasma VAP-1/SSAO activity in relation to the risk for intracranial bleeding complications in patients with stroke treated with tissue plasminogen activator (tPA), the greatest safety concern with this treatment. METHODS: In 141 patients with ischemic stroke, we measured VAP-1/SSAO activity in plasma taken before tPA administration. Hemorrhagic events were classified according to brain CT criteria and functional outcomes evaluated using the National Institutes of Health Stroke Scale. We also assessed the potential therapeutic effect of blocking VAP-1/SSAO activity in a rat embolic stroke model treated with tPA. RESULTS: We saw significantly higher levels of plasma VAP-1/SSAO activity in patients who subsequently experienced hemorrhagic transformation. Elevated plasma VAP-1/SSAO activity also predicted worse neurological outcome in these patients. In the rat model, we confirmed that use of the inhibitor semicarbazide prevented adverse effects caused by delayed tPA administration, leading to a smaller infarct volume. CONCLUSIONS: Our data demonstrate that baseline VAP-1/SSAO activity predicts parenchymal hemorrhage after tPA, suggesting the safety of thrombolytic agents could be improved by considering VAP-1/SSAO activity. Furthermore, anti-VAP-1/SSAO drugs given with tPA may prevent neurological worsening in patients with ischemic stroke.

Brainstem dysgenesis in an infant prenatally exposed to cocaine.

Many authors described the effects on the fetus of maternal cocaine abuse during pregnancy. Vasoconstriction appears to be the common mechanism of action leading to a wide range of fetal anomalies. We report on an infant with multiple cranial-nerve involvement attributable to brainstem dysgenesis, born to a cocaine-addicted mother.

Proteomic study of neuron and astrocyte cultures from senescence-accelerated mouse SAMP8 reveals degenerative changes.

Senescence-accelerated prone (SAMP) strain 8 mice suffer an earlier development of cognitive age-related pathologies and a shorter life span than conventional mice. Protein alterations in astrocytes, in addition to those in neurons, may contribute to neurodegenerative damage. We applied proteomics techniques to study cell-specific early markers of brain aging-related degeneration in SAMP8. The two-dimensional protein expression patterns of the SAMP8 neuron and astrocyte cultures were compared with those obtained from senescence-accelerated resistant mouse strain 1 cultures. Differentially expressed spots were identified by matrix-assisted laser desorption/ionization-time of flight peptide map fingerprinting and database search. Proteins belonged to cell pathways of energy metabolism, biosynthesis, cell transduction and signaling, stress response, and the maintenance of cytoskeletal functions. Most of the changes were cell type specific. However, there was a general increase in cell transduction, signaling, and stress-related proteins and a decrease in cytoskeletal proteins. In addition, neurons showed an increased expression of proteins involved in biosynthetic pathways. A number of the protein alterations have been previously reported in the brain tissue proteome of SAMP8, aged brain or Alzheimer's disease brain. Alterations in neuron and astrocyte proteoma indicated that both cell types are involved in the brain degenerative changes of SAMP8 mice. However, network analysis suggests that neuronal changes are more complex and have a greater influence.

Vascular MMP-9/TIMP-2 and neuronal MMP-10 up-regulation in human brain after stroke: a combined laser microdissection and protein array study.

Matrix Metalloproteinases (MMPs) play an important role in brain injury after ischemic stroke. In the present study, we aimed to assess the global expression of MMP-Family proteins in the human brain after stroke by using a combination of Searchlight Protein Array and Laser Microdissection to determine their cellular origin. This study demonstrated that MMP-1, MMP-2, MMP-3, MMP-8, MMP-9, MMP-10, MMP-13, and TIMP-1 were upregulated in the infarcted tissue compared to healthy control areas. Using laser microdissection we obtained specific neuronal and vascular populations from both infarcted and control areas. From these fractions, we showed that MMP-9 and TIMP-2 were highly produced in brain microvessels while MMP-10 was notably increased in neurons of the ischemic brain but not in healthy areas. These findings demonstrate a selective cell-dependent MMP secretion, opening the possibility of selectively targeting specific MMPs for neuroprotection or vasculoprotection following stroke.

Matrix metalloproteinase-13 is activated and is found in the nucleus of neural cells after cerebral ischemia.

Matrix metalloproteinases (MMPs) have been implicated in the pathophysiology of ischemic stroke. In this study, we investigated the time course of gelatinolytic activation in a rat model of permanent ischemia. We observed an activation of MMPs as early as 30 mins after the ischemic insult, mainly in the nuclei of brain cells. Besides, we explored MMP-13 expression in brain samples of the animal model and stroke deceased patients. We observed an upregulation of active MMP-13 in rat brains (P<0.05) after 90 mins of cerebral ischemia. Human infarct/periinfarct samples also showed higher levels of active MMP-13 (P<0.05) compared with contralateral ones. Interestingly, we found that MMP-13 colocalized with 46-diamidino-2-phenyl indole signal by immunohistochemistry in both humans and rats, suggesting an intranuclear localization for MMP-13. Immunohistochemistry also revealed that MMP-13 was mainly produced by neurons, in both species, but also by oligodendrocytes in rats, and by astrocytes in humans. Finally we subjected a rat primary neuronal culture to oxygen and glucose deprivation (OGD) and we reproduced the nuclear translocation of MMP-13 in vitro. Nuclear extracts from cells confirmed upregulation of active MMP-13 after OGD (P<0.05). These results suggest that MMP-13 activation and its nuclear translocation is an early consequence of an ischemic stimulus.

Stroke and multi-infarct dementia as presenting symptoms of giant cell arteritis: report of 7 cases and review of the literature.

Cerebrovascular accidents (CVAs) and multi-infarct dementia have rarely been reported as presenting symptoms of giant cell arteritis (GCA), although 3%-4% of patients with GCA may present with CVAs during the course of the disease. We describe 7 patients with biopsy-proven GCA who presented with stroke or multi-infarct dementia. Most of them had other symptoms of GCA when the disease began that were misdiagnosed or not noticed. The internal carotid arteries were involved in 4 patients and the vertebrobasilar arteries in 3, with bilateral vertebral artery occlusion in 1. Small cerebral infarction foci on cranial computed tomography (CT) scan and magnetic resonance imaging (MRI) were found in 5 cases, and cerebellar infarction, in 2. MR angiography showed intracranial arteritis in 4 cases. Treatment with glucocorticoids and adjunctive antiplatelet or anticoagulant therapy was given in all cases, with neurologic improvement in 5. Two patients died. Necropsy demonstrated generalized GCA involving the medium and small cerebral vessels in 1 case. Central nervous system involvement is a rare complication in GCA but is important to recognize, as it can be reversible if diagnosed and treated promptly. Suspicion should arise in elderly patients suffering from strokes with a quickly progressing stepwise course and associated headache, fever, or inflammatory syndrome. In these cases, temporal artery biopsy should be performed without delay. Early diagnosis of GCA and immediate initiation of corticosteroid treatment may prevent progressive deterioration and death. Additional antiplatelet or anticoagulant therapy should be evaluated according to the individual risk and benefit to the patient under care.

Fas system activation in perihematomal areas after spontaneous intracerebral hemorrhage.

BACKGROUND AND PURPOSE: Apoptosis has been implicated as the prominent form of cell death in the brain perihematomal region in animal models and in autopsy or postsurgical human studies. Both the Fas system and caspase activation play a central role in apoptotic pathways. The aims of this study were to investigate soluble Fas (s-Fas) plasma levels after acute intracerebral hemorrhage (ICH), to determine its influence on clinical and radiologic features, and to assess Fas receptor and Fas ligand (Fas-L) protein expression in human ICH brain tissue. METHODS: s-Fas plasma levels were determined on admission in 78 consecutive ICH patients and serially in a subgroup of 21 of them, at the time of neurologic assessment, by means of ELISA. ICH and perihematomal edema volumes were determined at baseline and on follow-up computed tomography scans, and ICH and perihematomal edema growth was calculated. The presence of Fas receptor and Fas-L was assessed in different brain tissue samples by immunoblotting from 6 deceased ICH patients and from 2 control subjects. RESULTS: Mortality reached 20.5% of patients at the third month, and 48% of survivors had an unfavorable outcome (modified Rankin Scale score >/=3). The baseline s-Fas level in ICH patients was significantly lower than in healthy controls [160 (160-245) vs 269 (230-332) pg/mL, P<0.001], returning to normal values by 24 hours (P<0.05 for all determinations). Regarding radiologic features, the baseline s-Fas value was found to be inversely correlated to perihematomal edema growth at follow-up (r=-0.33, P=0.041). Finally, Fas-L content was highest in the perihematomal area compared with contralateral and remote ipsilateral areas in ICH patient and control samples. CONCLUSIONS: A decreased plasma s-Fas level together with an increased Fas-L amount in perihematomal brain tissue suggest Fas-mediated apoptosis involvement in this disease.

MMP-9-positive neutrophil infiltration is associated to blood-brain barrier breakdown and basal lamina type IV collagen degradation during hemorrhagic transformation after human ischemic stroke.

BACKGROUND AND PURPOSE: An abnormal expression of some matrix metalloproteinases (MMPs) is related with hemorrhagic transformation events after stroke. Our aim was to investigate MMP-2 and MMP-9 in the ischemic brain and its relation with blood-brain barrier breakdown after hemorrhagic transformation in human stroke. METHODS: We assessed 5 cases of fatal ischemic strokes with hemorrhagic complications; brain samples were obtained from infarct, hemorrhagic, and contralateral tissue. MMP-9 and MMP-2 content was analyzed by zymography and immunohistochemistry was performed to localize MMP-9 and to assess collagen IV integrity in the basal lamina. Laser capture microdissection was performed to isolate blood-brain barrier vessels to study these MMPs. RESULTS: Overall, MMP-9 levels were higher both in hemorrhagic and nonhemorrhagic infarcted tissue compared to contralateral areas (P<0.0001 and P<0.05). Moreover, levels of the cleaved MMP-9 85kDa-form were significantly elevated in the hemorrhagic compared to nonhemorrhagic and contralateral areas (P=0.033 and P<0.0001). No changes were found for MMP-2 content. Immunostaining revealed a strong MMP-9-positive neutrophil infiltration surrounding brain microvessels associated with severe basal lamina type IV collagen degradation and blood extravasation. Microdissection confirmed that content of MMP-9 was similarly high in microvessel endothelium from hemorrhagic and infarcted areas compared to contralateral hemisphere vessels (P<0.05), pointing to neutrophils surrounding dissected microvessels as the main source of MMP-9 in hemorrhagic areas. CONCLUSIONS: Our results show a strong neutrophil infiltration in the infarcted and hemorrhagic areas with local high MMP-9 content closely related to basal lamina collagen IV degradation and blood-brain barrier breakdown. Microvessel and inflammatory MMP-9 response are associated with hemorrhagic complications after stroke.

Expression of metallothionein-I, -II, and -III in Alzheimer disease and animal models of neuroinflammation.

In recent years it has become increasingly clear that the metallothionein (MT) family of proteins is important in neurobiology. MT-I and MT-II are normally dramatically up-regulated by neuroinflammation. Results for MT-III are less clear. MTs could also be relevant in human neuropathology. In Alzheimer disease (AD), a major neurodegenerative disease, clear signs of inflammation and oxidative stress were detected associated with amyloid plaques. Furthermore, the number of cells expressing apoptotic markers was also significantly increased in these plaques. As expected, MT-I and MT-II immunostaining was dramatically increased in cells surrounding the plaques, consistent with astrocytosis and microgliosis, as well as the increased oxidative stress elicited by the amyloid deposits. MT-III, in contrast, remained essentially unaltered, which agrees with some but not all studies, of AD. In situ hybridization results in a transgenic mouse model of AD amyloid deposits, the Tg2576 mouse, which expresses human Abeta precursor protein harboring the Swedish K670N/M671L mutations, are in accordance with results in human brains. Overall, these and other studies strongly suggest specific roles for MT-I, MT-II, and MT-III in brain physiology.

Increased brain expression of matrix metalloproteinase-9 after ischemic and hemorrhagic human stroke.

BACKGROUND AND PURPOSE: Abnormal expression of some matrix metalloproteinases (MMP) has shown to play a deleterious role in brain injury in experimental models of cerebral ischemia. We aimed to investigate MMP-2 (gelatinase A) and MMP-9 (gelatinase B) in brain parenchyma in both ischemic and hemorrhagic strokes. METHODS: Postmortem fresh brain tissue from 6 ischemic and 8 hemorrhagic stroke patients was obtained within the first 6 hours after death. Finally, 78 brain tissue samples from different areas (infarct, peri-infarct, perihematoma and contralateral hemisphere) were studied. To quantify gelatinase content we performed gelatin zymograms that were confirmed by Western Blot Analysis, immunohistochemistry to localize MMP source, and in situ zymography to detect gelatinase activity. RESULTS: Among ischemic cases, gelatin zymography showed increased MMP-9 content in infarct core although peri-infarct tissue presented also higher levels than contralateral hemisphere (P<0.0001 and P=0.042, respectively). Within infarct core, MMP-9 was mainly located around blood vessels, associated to neutrophil infiltration and activated microglial cells. In peri-infarct areas the major source of MMP-9 were microglial cells. Tissue around intracranial hemorrhage also displayed higher MMP-9 levels than contralateral hemisphere (P=0.008) in close relationship with glial cells. MMP-2 was constitutively expressed and remained invariable in different brain areas. CONCLUSIONS: Our results demonstrate in situ higher levels of MMP-9 in human brain tissue after ischemic and hemorrhagic stroke, suggesting a contribution of MMP-9 to ischemic brain injury and perihematoma edema.