Modulation of amyloid beta peptide(1-42) cytotoxicity and aggregation in vitro by glucose and chondroitin sulfate.

One mechanism leading to neurodegeneration during Alzheimer's disease (AD) is amyloid beta peptide (Abeta)-induced neurotoxicity. Among the factors proposed to potentiate Abeta toxicity is its covalent modification through carbohydrate-derived advanced glycation endproducts (AGEs). Other experimental evidence, though, indicates that certain polymeric carbohydrates like the glycosaminoglycan (GAG) chains found in proteoglycan molecules attenuate the neurotoxic effect of Abeta in primary neuronal cultures. Pretreatment of the 42-residue Abeta fragment (Abeta1-42) with the ubiquitous brain carbohydrates, glucose, fructose, and the GAG chondroitin sulfate B (CSB) inhibits Abeta1-42-induced apoptosis and reduces the peptide neurotoxicity on neuroblastoma cells, a cytoprotective effect that is partially reverted by AGE inhibitors such as pyridoxamine and L-carnosine. Thioflavin T fluorescence measurements indicate that at concentrations close to physiological, only CSB promotes the formation of Abeta amyloid fibril structure. Atomic force microscopy imaging and Western blot analysis suggest that glucose favours the formation of globular oligomeric structures derived from aggregated species. Our data suggest that at short times carbohydrates reduce Abeta1-42 toxicity through different mechanisms both dependent and independent of AGE formation.

Effect of the nucleotides CMP and UMP on exhaustion in exercise rats.

The aetiology of muscle fatigue has yet not been clearly established. Administration of two nucleotides, cytosine monophosphate (CMP) and uridine monophosphate (UMP), has been prescribed for the treatment of neuromuscular affections in humans. Patients treated with CMP/UMP recover from altered neurological functions and experience pain relief, thus the interest to investigate the possible effect of the drug on exhausting exercise. With such aim, we have determined, in exercised rats treated with CMP/UMP, exercise endurance, levels of lactate, glucose and glycogen, and the activity of several metabolic enzymes such as, creatine kinase (CK), lactate dehydrogenase (LDH), and aspartate aminotransferase (AST). Our results show that rats treated with CMP/UMP are able to endure longer periods of exercise (treadmill-run). Before exercise, muscle glucose level is significantly higher in treated rats, suggesting that the administration of CMP/UMP favours the entry of glucose in the muscle. Liver glycogen levels remains unaltered during exercise, suggesting that CMP/UMP may be implicated in maintaining the level of hepatic glycogen constant during exercise. Lactate dehydrogenase and aspartate aminotransferase activity is significantly lower in the liver of treated rats. These results suggest that administration of CMP/UMP enable rats to endure exercise by altering some metabolic parameters.

Effect of the nucleotides CMP and UMP on exhaustion in exercise rats

The aetiology of muscle fatigue has yet not been clearly established. Administrationof two nucleotides, cytosine monophosphate (CMP) and uridine monophosphate(UMP), has been prescribed for the treatment of neuromuscular affections inhumans. Patients treated with CMP/UMP recover from altered neurological functionsand experience pain relief, thus the interest to investigate the possible effect ofthe drug on exhausting exercise. With such aim, we have determined, in exercised ratstreated with CMP/UMP, exercise endurance, levels of lactate, glucose and glycogen,and the activity of several metabolic enzymes such as, creatine kinase (CK), lactatedehydrogenase (LDH), and aspartate aminotransferase (AST). Our results show thatrats treated with CMP/UMP are able to endure longer periods of exercise (treadmillrun).Before exercise, muscle glucose level is significantly higher in treated rats, suggestingthat the administration of CMP/UMP favours the entry of glucose in themuscle. Liver glycogen levels remains unaltered during exercise, suggesting thatCMP/UMP may be implicated in maintaining the level of hepatic glycogen constantduring exercise. Lactate dehydrogenase and aspartate aminotransferase activity is significantlylower in the liver of treated rats. These results suggest that administrationof CMP/UMP enable rats to endure exercise by altering some metabolic parameters (AU)

No dipsonible

Inactivation of phosphoglycerate mutase and creatine kinase isoenzymes in human serum.

AIMS/BACKGROUND: Total phosphoglycerate mutase (PGM) activity in serum has been shown to be increased in acute myocardial infarction with the same time course as creatine kinase (CK) activity. However, the increase in the muscle (MM) and in the cardiac (MB) PGM isoenzymes was not as high as expected. The present study was undertaken to characterise PGM inactivation by serum and to compare it with serum CK inactivation. METHODS: The PGM and the CK activities of extracts of human heart, skeletal muscle, and brain were determined spectrophotometrically after incubation with different media, namely: plasma, whole serum, dialysed serum, heated serum, serum ultrafiltrate, urate solution, and buffer solution. RESULTS: Type MM PGM was inactivated by plasma, whole serum, heated serum, dialysed serum, and serum ultrafiltrate. Inactivation in dialysed serum was reduced by EDTA and largely reversed by thiol agents. Inactivation in serum ultrafiltrate was not prevented by EDTA and only partially reversed by dithiothreitol. The muscle and type BB CK isoenzymes were inactivated in all the tested media. The incubation of human and rabbit skeletal muscle PGM and CK in urate solution showed that urate does not affect mutase activity under conditions that inactivate CK. CONCLUSIONS: These results confirm the mechanisms of CK inactivation proposed by others and show that the type M PGM subunit is inactivated by two different mechanisms, which appear to involve the thiol groups of the enzyme. One mechanism is caused by either a protein component or a protein bound serum component and involves calcium ions and/or another chelatable metal ion. The other mechanism is caused by a lower molecular weight serum component and is metal ion independent.

Association analysis of HTR6 and HTR2A polymorphisms in sporadic Alzheimer's disease.

In order to identify gene variants related to the serotonergic neurotransmitter system that possibly represent a hereditary risk factor for sporadic Alzheimer's disease (AD), patients suffering from AD and non-demented psychiatric inpatients without symptoms of dementia were genotyped for polymorphisms of HTR6 (267C/T) and HTR2A (-1438G/A). Although there was a tendency toward an increased number of the genotype TT of the 5-HT6 receptor polymorphism in AD patients when compared to controls (2.8% vs. 1.3%), neither this nor the 5-HT2A promoter polymorphism showed significant differences in their genotypic or allelic distribution among patients and controls. These polymorphisms probably do not represent major genetic risk factors of AD. However, further studies including other genetic variants of the serotonergic neurotransmitter system are needed in order to elucidate their role in AD.

Potential genetic markers of sporadic Alzheimer's dementia.

Polymorphisms within the genes encoding apolipoprotein E (ApoE), apolipoprotein CI (ApoCI), alpha1-antichymotrypsin (ACT), the low-density lipoprotein (LDL) receptor and lipoprotein lipase were investigated in patients suffering from Alzheimer's dementia and non-demented psychiatric patients as control subjects. The ApoE allele 4, well known as a risk factor in Alzheimer's disease, and the ApoCI allele A2, which is closely linked to the ApoE allele 4, were found elevated in the index group. Concerning the polymorphism within exon 8 of the LDL receptor (alanin/threonin), there was also a predominance of the allele carrying threonin in the index group, which barely missed significance. Distribution of the polymorphisms of ACT and lipoprotein lipase were similar in both groups investigated. We conclude that, apart from the ApoE allele 4, other genetically regulated factors like ApoCI and the LDL receptor modulate the individual risk for Alzheimer's disease.

Brain-derived neurotrophic factor and neurotrophin 3 in schizophrenic psychoses.

Disturbed neural development has been postulated as a crucial factor in the pathophysiology of schizophrenic psychoses. The neurobiochemical basis for such changes of cytoarchitecture and changed neural plasticity could involve an alteration in the regulation of neurotrophic factors. In order to test this hypothesis, BDNF and NT-3 levels in post-mortem brain tissue from schizophrenic patients were determined by ELISA. There was a significant increase in BDNF concentrations in cortical areas and a significant decrease of this neurotrophin in hippocampus of patients when compared with controls. NT-3 concentrations of frontal and parietal cortical areas were significantly lower in patients than in controls. These findings lend further evidence to the neurotrophin hypothesis of schizophrenic psychoses which proposes that alterations in expression of neurotrophic factors could be responsible for neural maldevelopment and disturbed neural plasticity, thus being an important event in the etiopathogenesis of schizophrenic psychoses.

Increased levels of calcium-sensitive adenylyl cyclase subtypes in the limbic system of alcoholics: evidence for a specific role of cAMP signaling in the human addictive brain.

We examined the amounts of several adenylyl cyclase (AC) isoforms and of cAMP-response element binding protein (CREB) in alcoholic and control brains. Immunoreactivity of type I AC was significantly increased in alcoholic nucleus accumbens and corpus amygdaloideum. Immunoreactivity of type VIII AC was also increased in alcoholic corpus amygdaloideum and hippocampus. CREB immunoreactivities were unchanged. These findings indicate that the brain-region specific increase of Ca(2+)-sensitive AC may contribute to the pathophysiology of alcoholism.

Immunoreactivity of cAMP response element binding protein is not altered in the post-mortem cerebral cortex or cerebellum of alcoholics.

We examined amounts of cAMP response element binding protein (CREB) and its phosphorylated form in post-mortem frontal and temporal cortices and cerebella from alcoholics and controls by immunoblotting. No significant differences were observed in the levels of these proteins in each brain region, suggesting that the assumed neuroadaptations to chronic ethanol intake may not be reflected by quantitative alterations of CREB in alcoholic brain.

Apolipoprotein E genotype in Spanish schizophrenic patients.

Apolipoprotein E (ApoE) is a lipoprotein that, in the central nervous system, is thought to play a role in neuronal growth and repair. ApoE has three isoforms (ApoE2, ApoE3 and ApoE4) coded by three different alleles (epsilon2, epsilon3 and epsilon4). Evidence from family, twin and adoption studies suggest that there is an important genetic contribution to the etiology of schizophrenia. Schizophrenia is in some cases associated with cognitive impairment similar to that of Alzheimer patients; therefore, one may postulate that the ApoE gene, whose role in the dementia of Alzheimer's type has been clearly demonstrated, may also be involved in schizophrenia. In the present study, we have genotyped 114 schizophrenic Spanish patients and 94 healthy matched controls, and found no association between the ApoE genotype and schizophrenia. Subdivision of patients in clinical subgroups showed a slight increase of ApoE4 in early-age onset of the disease and a slight decrease in positive family history for psychiatric diseases; the group with a poor response to neuroleptic drugs had a lower ApoE2 allele frequency. However, as the differences did not reach statistical significance, we cannot draw evidence of an association. Our negative data do not support an involvement of ApoE in schizophrenia, and suggest that the underlying mechanism for the cognitive impairment found in schizophrenic patients is not related to that of Alzheimer's patients nor to a higher prevalence of the ApoE allele 4.

Human post-mortem striatal alpha4beta2 nicotinic acetylcholine receptor density in schizophrenia and Parkinson's syndrome.

The density of nicotinic alpha4beta2 receptors, which are believed to largely mediate nicotine's effects, has been reported to be decreased in post-mortem hippocampus of patients with schizophrenia. In the present study, using [(3)H]cytisine as a radioligand, we observed a significant 30% decrease in post-mortem striatum of patients with schizophrenia (n=12) as compared to controls (n=12). A 25% decrease of striatal alpha4beta2 receptor density in patients with Parkinson's syndrome (n=12) was not significant. As an upregulation of alpha4beta2 receptors has been observed due to nicotine consumption, the beneficial effects of nicotine described in patients with schizophrenia may be partly due to a compensation for a decrease in alpha4beta2 nicotinic acetylcholine receptors.

Phosphoglycerate mutase, 2,3-bisphosphoglycerate phosphatase, creatine kinase and enolase activity and isoenzymes in breast carcinoma.

We have compared the levels of phosphoglycerate mutase (EC 5.4.2.1), 2,3-bisphosphoglycerate phosphatase (EC 3.1.3.13), creatine kinase (EC 2.7.3.2) and enolase (EC 4.2.1.11) activities and the distribution of their isoenzymes in normal breast tissue and in breast carcinoma. Tumour tissue had higher phosphoglycerate mutase and enolase activity than normal tissue. Creatine kinase activity was higher in seven out of 12 tumours. In contrast 2,3-bisphosphoglycerate phosphatase activity was lower. Phosphoglycerate mutase, enolase and 2,3-bisphosphoglycerate phosphatase presented greater changes in the oestrogen receptor-negative/progesterone receptor-negative breast carcinomas than in the steroid receptor-positive tumours. Determined by electrophoresis, type BB phosphoglycerate mutase, type BB creatine kinase and alpha alpha-enolase were the major isoenzymes detected in normal breast tissue. Types alpha gamma and gamma gamma enolase, types MB and MM phosphoglycerate mutase were detected in much lower proportions. In tumours a decrease of phosphoglycerate mutase isoenzymes possessing M-type subunit and some increase of enolase isoenzymes possessing gamma-type subunit was observed. No detectable change was observed in the creatine kinase phenotype.

Brain-derived neurotrophic factor and neurotrophin-3 levels in Alzheimer's disease brains.

In the present study, we investigated the levels of brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) in post-mortem brain tissue of Alzheimer's disease (AD) patients, and we observed a significant increase of BDNF concentration in hippocampus and parietal cortex of AD patients, as well as a negative correlation between NT-3 levels and age in hippocampus and putamen of control subjects, and for BDNF in frontal cortex. A defining feature of AD is the post-mortem identification of neuritic plaques and neurofibrillary tangles in the brain, however, a more significant neuropathological finding is the degeneration of cholinergic neurones of the basal forebrain, critically involved in memory and cognition. Neurotrophic factors are partly responsible for the maintenance of neuronal function and structural integrity in the adult brain. Our results provide, therefore, evidence that, under conditions of progressive neurodegeneration the brain stimulates the over expression of certain neurotrophic factors as a possible mechanisms of compensation, and that during senescence the expression of these molecules is regulated.

Increased frequency of the alpha-1-antichymotrypsin T allele in cerebral amyloid angiopathy.

Cerebral amyloid angiopathy (CAA) is a process of unknown etiology characterized by amyloid deposition in the wall of small cerebral and meningeal blood vessels. CAA is also a feature of Alzheimer's disease (AD) and of a subgroup of elderly people. Alpha-1-Antichymotrypsin (ACT) is a serum glycoprotein frequently associated with vascular and senile plaque amyloid. The ACT gene is known to have a bi-allele polymorphism that causes a simple amino acid substitution. In an attempt to clarify the possible role of ACT polymorphism in AD and in cases of CAA, the ACT genotype was investigated in AD, CAA, and intellectually intact controls. Representative brain areas (cerebral cortex, hippocampus, putamen, white matter, and gyrus cinguli) from all cases were studied using classical histologic staining techniques (hematoxylin-eosin (HE), Mallory's thrichromic or alkaline congo red stain), and immunohistochemistry for tau and beta-amyloid proteins. There was a significantly increased T allele and TT genotype frequency in the CAA group, but not in the AD group, suggesting a role for the ACT genotype in the development of vascular lesions. The presence of the apolipoprotein E4 allele (ApoE4) did not correlate with the ACT-A allele, as previously reported, and appeared to be independent of the risk for developing AD.

Brain-derived neurotrophic factor and neurotrophin-3 levels in Alzheimer's disease brains.

In the present study, we investigated the levels of brain-derived neurotrophic factor (BDNF) and neurotrophin-3 (NT-3) in post-mortem brain tissue of Alzheimer's disease (AD) patients, and we observed a significant increase of BDNF concentration in hippocampus and parietal cortex of AD patients, as well as a negative correlation between NT-3 levels and age in hippocampus and putamen of control subjects, and for BDNF in frontal cortex. A defining feature of AD is the post-mortem identification of neuritic plaques and neurofibrillary tangles in the brain, however, a more significant neuropathological finding is the degeneration of cholinergic neurones of the basal forebrain, critically involved in memory and cognition. Neurotrophic factors are partly responsible for the maintenance of neuronal function and structural integrity in the adult brain. Our results provide, therefore, evidence that, under conditions of progressive neurodegeneration the brain stimulates the over expression of certain neurotrophic factors as a possible mechanisms of compensation, and that during senescence the expression of these molecules is regulated.

Correlation between Apolipoprotein-E polymorphism and Alzheimer's disease pathology.

Alzheimer's disease (AD) and small vessel disease dementia (SVDD) are common causes of dementia. The ApoE genotype has been proposed as a risk factor for AD. The frequency of the three ApoE alleles was correlated with the neuropathological changes of AD (senile plaques, neurofibrillary tangles and amyloid angiopathy) and SVDD (status lacunaris, status cribosus, leucoencephalopathy, micronecrosis and vascular fibrohyalinosis) in order to validate previous ApoE genotyping results in AD and to identify pre-clinical AD. Representative cerebral regions (cortex, gyrus cinguli, putamen, hippocampus, white matter) from 28 AD cases, 7 SVDD and 38 non-neurological controls were studied using classical histological techniques and immunohistochemistry for tau protein and amyloid-beta. The frequency of the ApoE allele 4 was significantly increased not only in AD patients but also in aged controls. However, following a detailed histopathological examination was found 62% of this group to exhibit histological changes associated with AD in limited brain areas (entorhinal region, hippocampus and adjacent temporal cortex or entorhinal region and hippocampus, or only in the entorhinal region), but 87% of these cases were found to be ApoE4 positive. The significant differences found in the distribution of ApoE allele frequencies were more marked when these cases were excluded from the control group and included as AD cases. In contrast, the frequency of the ApoE allele 2 is significantly increased in SVDD patients. Using histological techniques we confirmed the clinical diagnoses of all cases and classified the AD patients according to the severity of cortical pathology related to AD, while re-grouping from the control group those cases which had no clinical history of the disease but exhibited typical AD and SVDD histological lessions which could be considered as "pre-clinical" forms of these diseases.

Investigations on oxidative stress and therapeutical implications in dementia.

Alzheimer's disease (AD) is a progressive dementia affecting a large proportion of the aging population. The histopathological changes in AD include neuronal cell death and formation of amyloid plaques and neurofibrillary tangles (NFTs) NFTs are composed of hyperphosphorylated tau protein, and senile plaques contain aggregates of the beta-peptide. There is also evidence that brain tissue in patients with AD is exposed to oxidative stress during the course of the disease. Advanced glycation endproducts (AGEs), which are formed by a non-enzymatic reaction of glucose with long-lived protein deposits, are potentially toxic to the cell, are present in brain plaques in AD, and its extracellular accumulation in AD may be caused by an accelerated oxidation of glycated proteins. The microtubuli-associated protein tau is also subject to intracellular AGE formation. AGEs participate in neuronal death causing direct (chemical) radical production: Glycated proteins produce nearly 50-fold more radicals than non-glycated proteins, and indirect (cellular) radical production: Interaction of AGEs with cells increases oxidative stress. During aging cellular defence mechanisms weaken and the damages to cell constituents accumulate leading to loss of function and finally cell death. The development of drugs for the treatment of AD remains at a very unsatisfying state. However, pharmacological approaches which break the vicious cycles of oxidative stress and neurodegeneration offer new opportunities for the treatment of AD. Theses approaches include AGE-inhibitors, antioxidants, and anti-inflammatory substances, which prevent radical production. AGE inhibitors might be able to stop formation of AGE-modified beta-amyloid deposits, antioxidants are likely to scavenge intracellular and extracellular superoxide radicals and hydrogen peroxide before these radicals damage cell constituents or activate microglia, and anti-inflammatory drugs attenuating microglial radical and cytokine production.

Alzheimer's disease--synergistic effects of glucose deficit, oxidative stress and advanced glycation endproducts.

Many approaches have been undertaken to understand Alzheimer's disease (AD) but the heterogeneity of the etiologic factors makes it difficult to define the clinically most important factor determining the onset and progression of the disease. However, there is increasing evidence that the previously so-called "secondary factors" such as a disturbed glucose metabolism, oxidative stress and formation of "advanced glycation endproducts" (AGEs) and their interaction in a vicious cycle are also important for the onset and progression of AD. AGEs are protein modifications that contribute to the formation of the histopathological and biochemical hallmarks of AD: amyloid plaques, neurofibrillary tangles and activated microglia. Oxidative modifications are formed by a complex cascade of dehydration, oxidation and cyclisation reactions, subsequent to a non-enzymatic reaction of sugars with amino groups of proteins. Accumulation of AGE-crosslinked proteins throughout life is a general phenomenon of ageing. However, AGEs are more than just markers of ageing since they can also exert adverse biologic effects on tissues and cells, including the activation of intracellular signal transduction pathways, leading to the upregulation of cytokine and free radical production (oxidative stress). Oxidative stress is involved in various divergent events leading to cell damage, including an increase in membrane rigidity, DNA strand breaks and an impairment in glucose uptake. In addition, other age-related metabolic changes such as depletion of antioxidants or decreased energy production by a disturbed glucose metabolism diminish the ability of the cell to cope with the effects of radical-induced membrane, protein and DNA damage. With our improving understanding of the molecular basis for the clinical symptoms of dementia, it is hoped that the elucidation of the etiologic causes, particularly the positive feedback loops involving radical damage and a reduced glucose metabolism, will help to develop novel "neuroprotective" treatment strategies able to interrupt this vicious cycle of oxidative stress and energy shortage in AD.

Variants in neurotrophic factor genes and schizophrenic psychoses: no associations in a Spanish population.

Possible associations between schizophrenic psychoses, a ciliary neurotrophic factor (CNTF) null mutation and a neurotrophin in 3 (NT3) bi-allele polymorphism were investigated in a Spanish population. The sample consisted of 107 patients suffering from schizophrenic psychoses and 100 healthy volunteers. There was no statistical difference in the frequencies of the mutant CNTF allele in the index and control groups (0.125 vs. 0.121). The frequency of the rare NT3 allele (Glu) was very low and similar in both groups (0.005). Previous findings supporting the involvement of these genetic markers in the pathogenesis of schizophrenic psychoses were thus not confirmed. In light of neurodevelopmental hypotheses of schizophrenia, however, genes coding for neurotrophic factors remain interesting susceptibility loci in schizophrenia research. Subsequent studies should not focus exclusively on genetic alterations but also take into account secondary changes of the neurotrophic factor system at the mRNA and protein levels. Further, the current unsatisfying operationalized classification of the heterogeneous syndrome 'schizophrenia' remains a crucial problem that could be partially resolved by introducing more differentiated diagnoses defined on the basis of neurobiological criteria.

Phosphoglycerate mutase, 2,3-bisphosphoglycerate phosphatase and creatine kinase activity and isoenzymes in human brain tumours.

The distribution of phosphoglycerate mutase (EC 5.4.2.1, PGM), 2,3-bisphosphoglycerate phosphatase (EC 3.1.3.13, BPGP) and creatine kinase (EC 2.7.3.2, CK) activity and isoenzymes in various regions of adult human brain and in brain tumours (astrocytomas, anaplastic astrocytomas, glioblastomas and meningiomas) has been determined using electrophoresis. PGM and cytosolic CK exist in mammalian tissues as three isoenzymes that result from the homodimeric and heterodimeric combinations of two subunits [types M (muscle) and B (brain)] coded by separated genes. In addition, a dimeric form and an octameric form of mitochondrial CK exist in mammals. Type BB-PGM was the major PGM isoenzyme found in normal brain, although type MB-PGM and type MM-PGM were also detected. All brain tumours possessed lower PGM activity than normal brain, and meningiomas showed higher BPGP activity. In astrocytic tumours, the proportion of type MB- and type MM-PGM decreased, and in meningiomas these isoenzymes were not detected. Type BB-CK and mitochondrial CK were the only CK isoenzymes detected in normal brain. Astrocytomas possessed lower CK activity than anaplastic astrocytomas and glioblastomas and, in addition, tended to possess lower CK content than normal brain. No qualitative changes of the normal CK isoenzyme pattern were observed in the tumours.