No abstract

Recent data have revealed that soluble oligomeric forms of amyloid peptide (Abeta) may be the proximate effectors of the neuronal injury and death occurring in Alzheimer's disease (AD). However, the molecular mechanisms associated with the neuronal cell death induced by the nonfibrillar Abeta remain to be elucidated. In this study, we investigated the role of the cytosolic Ca2+-dependent phospholipase A2 (cPLA2), and its associated metabolic pathway, i.e., the arachidonic acid (AA) cascade, in the apoptotic cell death induced by soluble oligomers of Abeta. The treatment of rat cortical neurons with low concentrations of soluble Abeta(1-40) or Abeta(1-42) peptide resulted in an early calcium-dependent release of AA associated with a transient relocalization of cPLA2. Both cPLA2 antisense oligonucleotides and a selective inhibitor of cPLA2 activity abolished the release of AA from neurons and also protected cells against apoptosis induced by Abeta. Furthermore, inhibitors of the PKC, p38, and MEK/ERK pathways that are involved in cPLA2 phosphorylation and activation reduced Abeta-induced cell death. Finally, we demonstrate that inhibitors of cyclooxygenase-2 reduced the Abeta-induced cell death by 55%. Our studies suggest a novel neuronal response of soluble oligomers of Abeta, which occurs through a cPLA2 signaling cascade and an AA-dependent death pathway. This may prove to be crucial in AD processes and could provide important targets for drug development.

Transcobalamin (TC) is the plasma transporter that delivers vitamin B(12) to cells. We have already reported that HT-29 and Caco-2 cells secrete different TC variants. HT-29 secretes 2 TC isoproteins (codon 259-Pro/Arg [259-P/R]), exhibiting unequal concentrations (TC 259-P > TC 259-R), and Caco-2 cells only secrete the phenotype 259-R. We investigated the relation between phenotypic and genetic TC polymorphism in HT-29 cells transfected with Caco-2 TC complementary DNA and in 159 healthy Caucasians. We found that codon 259-R is buried and, thus, the genetic polymorphism provides no explanation why the TCs from HT-29 and Caco-2 cells have different isoelectric points in nondenaturing isoelectric focusing (IEF). The newly translated TC in HT-29 cells from the Caco-2 complementary DNA recombinant plasmid had the same isoelectric point as the TC constitutively expressed in HT-29 cells, suggesting that TC phenotypic variability involves a specific cell folding of the protein. The codon 259 polymorphism was found to have a biallelic distribution: homozygotes P = 34.6%, heterozygotes R/P = 47.8%, and homozygotes R = 17.6%. In heterozygous samples, the IEF showed that the TC 259-P/TC 259-R ratio = 1.6. The blood apo-TC concentration of 259-P homozygous Caucasians was significantly higher than that of homozygous 259-R (P <.0001) and heterozygous (P <.0006) Caucasians. The heterozygotes 259-R/P had homocysteine concentration significantly higher than the homozygotes 259-R and 259-P (P =.02 and P =.01, respectively). In conclusion, TC codon-259 polymorphism affects TC plasma concentration and may interfere in vitamin B(12) cellular availability and homocysteine metabolism.

In the present study, we have determined the nature and the kinetics of the cellular events triggered by the exposure of cells to non-fibrillar amyloid-␤ peptide (A␤). When cortical neurons were treated with low concentrations of soluble A␤ (1-40), an early reactive oxygen species (ROS)-dependent cytoskeleton disruption precedes caspase activation. Indeed, caspase activation and neuronal cell death were prevented by the microtubulestabilizing drug taxol. A perturbation of the microtubule network was noticeable after being exposed to A␤ for 1 h, as revealed by electron microscopy and immunocytochemistry. Microtubule disruption and neuronal cell death induced by A␤ were inhibited in the presence of antioxidant molecules, such as probucol. These data highlight the critical role of ROS production in A␤-mediated cytoskeleton disruption and neuronal cell death. Finally, using FRAP (fluorescence recovery after photo bleaching) analysis, we observed a time-dependent biphasic modification of plasma membrane fluidity, as early as microtubule disorganization. Interestingly, molecules that inhibited neurotubule perturbation and cell death did not affect the membrane destabilizing properties of A␤, suggesting that the lipid phase of the plasma membrane might represent the earliest target for A␤. Altogether our results convey the idea that upon interaction with the plasma membrane, the non-fibrillar A␤ induces a rapid ROS-dependent disorganization of the cytoskeleton, which results in apoptosis.A common feature of Alzheimer's disease (AD), 1 the most common form of dementia, is the accumulation and the aggregation of the amyloid-␤ peptide (A␤), a 39-to 43-amino acid peptide derived from the proteolytic cleavage of the amyloid precursor protein (1, 2). Although A␤ represents a key factor in AD (3), the nature of the toxic form of A␤ early involved in AD pathology remains unclear. Whether it is the fibrillar or the non-fibrillar peptides that are the more deleterious remains a controversial issue (4). The amyloid cascade hypothesis causally links AD clinico-pathological process and neuronal cell death to the aggregation and deposition of A␤ (5-7). However, this hypothesis has been challenged by recent evidences indicating that the non-fibrillar A␤ also plays a major role in AD (8, 9). A recent elegant study has demonstrated that the fibrils from AD brain are composed of amyloid peptide moieties arranged at right angles to the backbone of the amyloid P protein wrapped in glycosaminoglycans (10). Thus, the fibrils are not simply made of chains of self-aggregated A␤ and do not comprise long chains of multimeric A␤, similar to those used to evaluate the neurotoxicity of the fibrillar A␤ in vitro and in vivo. Moreover, the synaptic loss in AD brain has been correlated with the soluble pool of A␤ peptides rather than the fibrillar one, implying that the non-fibrillar A␤ may be a crucial pathological factor in AD (11-13). Several studies, based on the use of transgenic mice, have demonstrated that neurodegeneration and specific spatia...

A growing body of evidence supports the notion that soluble oligomers of amyloid-b (Ab) peptide interact with the neuronal plasma membrane, leading to cell injury and inducing deathsignalling pathways that could account for the increased neurodegeneration occurring in Alzheimer's disease (AD). Docosahexaenoic acid (DHA, C22:6, n-3) is an essential polyunsaturated fatty acid in the CNS and has been shown in several epidemiological and in vivo studies to have protective effects against AD and cognitive alterations. However, the molecular mechanisms involved remain unknown. We hypothesized that DHA enrichment of plasma membranes could protect neurones from apoptosis induced by soluble Ab oligomers. DHA pre-treatment was observed to significantly increase neuronal survival upon Ab treatment by preventing cytoskeleton perturbations, caspase activation and apoptosis, as well as by promoting extracellular signal-related kinase (ERK)-related survival pathways. These data suggest that DHA enrichment probably induces changes in neuronal membrane properties with functional outcomes, thereby increasing protection from soluble Ab oligomers. Such neuroprotective effects could be of major interest in the prevention of AD and other neurodegenerative diseases.