Some markers of neuronal damage in cerebrospinal fluid of multiple sclerosis patients in relapse.

In this paper the performance of cerebrospinal fluid (CSF) protein biomarkers important for monitoring damage of brain astrocytes and neurons for MS is reviewed. We estimated neurofilament, tau and phospho-tau proteins, ß-APP, Aß, S-100B and neuron-specific enolase in CSF of MS patients during relapse. We noted elevation of neurofilament, tau and phospho-tau proteins, S-100B, neuron-specific enolase and c-terminal epitopes of ß-APP; concomitantly decrease of Aß was observed. These CSF biomarkers for MS relapse should reflect the central pathogenic processes in the brain, i.e., axonal and neuronal degeneration.

Free radical peroxidation products in cerebrospinal fluid and serum of patients with multiple sclerosis after glucocorticoid therapy.

Multiple sclerosis (MS) patients were found to have elevated thiobarbituric acid reactive material levels, increased soluble sulfhydryl groups and reduced protein sulfhydryl groups in cerebrospinal fluid and serum, and slightly reduced superoxide dismutase in serum, which suggested disease activating free radical peroxidation. Moreover, levels of these varied across methylprednisolone (MP) therapy. We observed significant differences in the levels of peroxidation products between MS patients and controls. These changes were most evident in relapse. After MP therapy, levels of these indicators approached control values, especially in the remission period. Our findings suggest that MP protects against free radical attack.

Homocysteine-induced acute excitotoxicity in cerebellar granule cells in vitro is accompanied by PP2A-mediated dephosphorylation of tau.

Our results demonstrate that acute exposure of primary rat cerebellar granule cell cultures to homocysteine at millimolar concentrations induces a glutamate receptor-mediated decrease in tau protein phosphorylation, which is accompanied by excitotoxic neuronal damage. This contrasts with the previously reported hyperphosphorylation of tau in homocysteine-treated neurons, and with the assumption that hyperhomocysteinemia is one of the risk factors in Alzheimer disease, in which abnormal hyperphosphorylation of tau protein leads to neurofibrillary degeneration. The mechanisms of homocysteine neurotoxicity have been explained mainly either by disturbances in methylation processes, that may also lead to the accumulation of phosphorylated tau due to reduced activity of tau-selective protein phosphatase 2A, or by excitotoxicity. Since the relationships between homocysteine excitotoxicity and tau phosphorylation are unclear, the aim of this study was to characterize these processes in neurons acutely treated with homocysteine at neurotoxic concentrations, and to link them to the activities of glutamate receptors and protein phosphatase 2A. Within 24h following a 30 min exposure of neuronal cultures to 20mM d,l-homocysteine, significant neurotoxicity was induced. This could be reduced by treatment with an uncompetitive NMDA receptor antagonist, MK-801 (0.5 microM), or by mGlu1 and mGlu5 receptor antagonists, LY367385 and MPEP, respectively (both at 25 microM). Western blot analysis showed a rapid decrease in immunostaining of phospho-tau, 2h after incubation of cell cultures with 15 mM D,L-homocysteine, which persisted for 6h after the insult. Application of MK-801, LY367385 or okadaic acid (100 nM), an inhibitor of protein phosphatases 1 and 2A, significantly prevented dephosphorylation of tau, implying a role for the activation of glutamate receptors and protein phosphatase 2A. The phosphorylation of tau may be increased or reduced by treatment with homocysteine, and the nature of the cellular response to this sulfur-containing amino acid depends on the neuronal phenotype.

The immature endothelial cell in new vessel formation following surgical injury in rat brain.

OBJECTIVES: We investigated neovasculatization in the cerebral cortex of the adult rat after surgical brain injury by ultrastructural, immunocytochemical and immunochemical means. Previously we described endothelial-like cell that participates in new vessel formation on plasma proteins that served as a provisional matrix in the region immediately adjacent to the traumatic injury. In the present study we describe new vessel formation in the multistep process with the alterations in endothelial-like cell immunophenotype. METHOD: The observations were conducted from 2 to 7 days after induction of cortical trauma. Traumatic injury was induced in the frontotemporal region of cerebral cortex. RESULTS: We show that endothelial-like cell could not successfully terminate its development without the presence of pericyte and astrocyte. New formed blood vessels were accompanied by fibroblast and lipofibroblast cells differentiated probably from common progenitor. MAIN FINDINGS: Our result suggests that endothelial-like cell is committed endothelial cell between progenitor endothelial cell and terminally differentiated endothelium stage. Therefore, we propose that our of endothelial stem cells present in blood at different stages of morphogenetic differentiation and specifically arrested in "check points" of development trauma mobilize the group of the most differentiated progenitors. These may contribute to new vessel formation. CONCLUSION: Our model should be useful for the characterization of endothelial commitment and endothelial cell differentiation after brain injury.

2-deoxyglucose induces beta-APP overexpression, tau hyperphosphorylation and expansion of the trans-part of the Golgi complex in rat cerebral cortex.

The effects of a single intraperitoneal injection of a non-metabolizable glucose analog 2-deoxyglucose (2-DG, 500 mg/kg) on the levels of beta-APP expression, and phosphorylated and unphosphorylated tau protein in the rat cerebral cortex were investigated. The effects of 2-DG on the ultrastructure of cortical neurons with particular emphasis on the morphology of the Golgi apparatus, and on brain bioenergetics assessed by in vivo 31P-MRS technique were also evaluated. Seven and a half hours after injection of 2-deoxyglucose a significant increase in brain cortex beta-APP expression, increased tau phosphorylation, and a marked relative expansion of the trans- part of the Golgi intracellular secretory pathway in cortical neurons has been found. The changes of beta-APP expression and tau phosphorylation appeared within 1 h after 2-DG application and continued for at least 24 h. However, brain 31P resonance spectra remained unchanged for up to 7.5 h after 2-DG. It is suggested that the increase of beta-APP expression represents a response of brain tissues to 2-DG-evoked biochemical stress, while tau hyperphosphorylation and the change in Golgi morphology may be secondary phenomena.

Cerebrospinal fluid free-radical peroxidation products and cognitive functioning patterns differentiate varieties of normal pressure hydrocephalus.

The aims of the study were as follows: first, to verify the hypothesis that free radical peroxidation may be one of the factors implicated in pathophysiology of normal pressure hydrocephalus (NPH) and, second, to find out whether these biochemical characteristics together with neuropsychological cognitive deficits can differentiate between various types of NPH. This provides prognostic criteria for selection of patients for shunt surgery. Lipid peroxidation was measured in terms of thiobarbituric acid-reactive material (TBAR) and protein sulphydryl (SH) groups were measured as CSF content. Cognitive deficits were assessed using a number of neuropsychological tests. In the sample of NPH patients (n = 24), three categories were distinguished using these criteria: idiopathic active hydrocephalus (A), arrested hydrocephalus (AH), and post-traumatic hydrocephalus (PT). TBAR levels for NPH patients were higher than that of controls without CNS pathology (n = 2). Moreover, NPH patients had increased levels of total and soluble protein groups, and decreased levels of protein SH groups, which suggests the occurrence of processes that activate peroxidation of free radicals in normal pressure hydrocephalus. Levels of these indicators varied across NPH types. Two categories of NPH patients, with active (A) or posttraumatic (PT) hydrocephalus differed significantly from the controls (C)--their TBAR levels were 0.58, 0.56 and 0.28 nmol/mg protein, respectively; soluble SH levels: 41.5; 58.15 and 11.3 nmol/mg protein, and protein SH levels: 34.3, 21.8 and 57.5 nmol/mg protein. In PT group, many individual differences were noticed. These findings seem promising because the studied biochemical indicators may serve as additional diagnostic criteria for selection of NPH patients for shunting.

N-methyl-d-aspartate receptor-mediated processing of beta-amyloid precursor protein in rat hippocampal slices: in vitro--superfusion study.

Abnormal proteolytic degradation of the beta amyloid precursor protein (beta-APP) may result in accumulation of potentially neurotoxic beta amyloid (betaA). The role of various receptors in the regulation of beta-APP processing has been suggested. This study aimed to determine how NMDA receptors and Ca2+ ions regulate proteolysis of beta-APP in rat hippocampus in vitro. Adult rat hippocampal slices were superfused with NMDA-containing media, and immunoreactivity of soluble beta-APP derivatives was detected in dialysates. Application of 100 microM and 250 microM NMDA for 20 min in Ca2+-containing medium induced dose-dependent release of aminoterminal beta-APP derivatives, and a fragment of Abeta sequence, whereas carboxy-terminal fragments of beta-APP were only slightly detected. This indicates activation of beta-APP processing, and release of its soluble cleavage products. This effect was inhibited by NMDA receptor antagonist 1 microM MK-801 and 100 microM CPP in Ca2+-free medium, thus indicating that NMDA receptors and calcium ions mediate proteolytic non-amyloidogenic degradation of the beta-APP.