Jean-Luc Olivier

Soluble oligomers of amyloid-β peptide induce neuronal apoptosis by activating a cPLA2-dependent sphingomyelinase-ceramide pathway

Recent data have revealed that soluble oligomeric amyloid-beta peptide (Abeta) may be the proximate effectors of neuronal injuries and death in Alzheimers disease (AD) by unknown mechanisms. Consistently, we recently demonstrated the critical role of a redox-sensitive cytosolic calcium-dependent phospholipase A2 (cPLA2)-arachidonic acid (AA) pathway in Abeta oligomer-induced cell death. According to the involvement of oxidative stress and polyunsaturated fatty acids like AA in the regulation of sphingomyelinase (SMase) activity, the present study underlines the role of SMases in soluble Abeta-induced apoptosis. Soluble Abeta oligomers induced the activation of both neutral and acidic SMases, as demonstrated by the direct measurement of their enzymatic activities, by the inhibitory effects of both specific neutral and acidic SMase inhibitors, and by gene knockdown using antisense oligonucleotides. Furthermore, soluble Abeta-mediated activation of SMases and subsequent cell death were found to be inhibited by antioxidant molecules and a cPLA2-specific inhibitor or antisense oligonucleotide. We also demonstrate that sphingosine-1-phosphate is a potent neuroprotective factor against soluble Abeta oligomer-induced cell death and apoptosis by inhibiting soluble Abeta-induced activation of acidic sphingomyelinase. These results suggest that Abeta oligomers induce neuronal death by activating neutral and acidic SMases in a redox-sensitive cPLA2-AA pathway.

Cytosolic phospholipase A2 mediates neuronal apoptosis induced by soluble oligomers of the amyloid-β peptide

Recent data have revealed that soluble oligomeric forms of amyloid peptide (Aβ) may be the proximate effectors of the neuronal injury and death occurring in Alzheimers disease (AD). However, the molecular mechanisms associated with the neuronal cell death induced by the nonfibrillar Aβ 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 Aβ. The treatment of rat cortical neurons with low concentrations of soluble Aβ(1‐40) or Aβ(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 Aβ. Furthermore, inhibitors of the PKC, p38, and MEK/ERK pathways that are involved in cPLA2 phosphorylation and activation reduced Aβ‐induced cell death. Finally, we demonstrate that inhibitors of cyclooxygenase‐2 reduced the Aβ‐induced cell death by 55%. Our studies suggest a novel neuronal response of soluble oligomers of Aβ, 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.

Docosahexaenoic acid prevents neuronal apoptosis induced by soluble amyloid-β oligomers

A growing body of evidence supports the notion that soluble oligomers of amyloid‐β (Aβ) peptide interact with the neuronal plasma membrane, leading to cell injury and inducing death‐signalling pathways that could account for the increased neurodegeneration occurring in Alzheimers 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 Aβ oligomers. DHA pre‐treatment was observed to significantly increase neuronal survival upon Aβ 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 Aβ oligomers. Such neuroprotective effects could be of major interest in the prevention of AD and other neurodegenerative diseases.

Microtubule-associated Protein MAP1A, MAP1B, and MAP2 Proteolysis during Soluble Amyloid β-Peptide-induced Neuronal Apoptosis SYNERGISTIC INVOLVEMENT OF CALPAIN AND CASPASE-3

A growing body of evidence supports the notion that soluble oligomeric forms of the amyloid β-peptide (Aβ) may be the proximate effectors of neuronal injuries and death in the early stages of Alzheimer disease. However, the molecular mechanisms associated with neuronal apoptosis induced by soluble Aβ remain to be elucidated. We recently demonstrated the involvement of an early reactive oxygen species-dependent perturbation of the microtubule network (Sponne, I., Fifre, A., Drouet, B., Klein, C., Koziel, V., Pincon-Raymond, M., Olivier, J.-L., Chambaz, J., and Pillot, T. (2003) J. Biol. Chem. 278, 3437–3445). Because microtubule-associated proteins (MAPs) are responsible for the polymerization, stabilization, and dynamics of the microtubule network, we investigated whether MAPs might represent the intracellular targets that would enable us to explain the microtubule perturbation involved in soluble Aβ-mediated neuronal apoptosis. The data presented here show that soluble Aβ oligomers induce a time-dependent degradation of MAP1A, MAP1B, and MAP2 involving a perturbation of Ca2+ homeostasis with subsequent calpain activation that, on its own, is sufficient to induce the proteolysis of isoforms MAP2a, MAP2b, and MAP2c. In contrast, MAP1A and MAP1B sequential proteolysis results from the Aβ-mediated activation of caspase-3 and calpain. The prevention of MAP1A, MAP1B, and MAP2 proteolysis by antioxidants highlights the early reactive oxygen species generation in the perturbation of the microtubule network induced by soluble Aβ. These data clearly demonstrate the impact of cytoskeletal perturbations on soluble Aβ-mediated cell death and support the notion of microtubule-stabilizing agents as effective Alzheimer disease drugs.

Ciliary Neurotrophic Factor Cell-Based Delivery Prevents Synaptic Impairment and Improves Memory in Mouse Models of Alzheimer's Disease

The development of novel therapeutic strategies for Alzheimers disease (AD) represents one of the biggest unmet medical needs today. Application of neurotrophic factors able to modulate neuronal survival and synaptic connectivity is a promising therapeutic approach for AD. We aimed to determine whether the loco-regional delivery of ciliary neurotrophic factor (CNTF) could prevent amyloid-β (Aβ) oligomer-induced synaptic damages and associated cognitive impairments that typify AD. To ensure long-term administration of CNTF in the brain, we used recombinant cells secreting CNTF encapsulated in alginate polymers. The implantation of these bioreactors in the brain of Aβ oligomer-infused mice led to a continuous secretion of recombinant CNTF and was associated with the robust improvement of cognitive performances. Most importantly, CNTF led to full recovery of cognitive functions associated with the stabilization of synaptic protein levels in the Tg2576 AD mouse model. In vitro as well as in vivo, CNTF activated a Janus kinase/signal transducer and activator of transcription-mediated survival pathway that prevented synaptic and neuronal degeneration. These preclinical studies suggest that CNTF and/or CNTF receptor-associated pathways may have AD-modifying activity through protection against progressive Aβ-related memory deficits. Our data also encourage additional exploration of ex vivo gene transfer for the prevention and/or treatment of AD.

Membrane cholesterol interferes with neuronal apoptosis induced by soluble oligomers but not fibrils of amyloid-β peptide

Neuronal cell death in Alzheimers disease (AD) is partly induced by the interaction of the amyloid‐β peptide (Aβ) with the plasma membrane of target cells. Accordingly, recent studies have suggested that cholesterol, an important component of membranes that controls their physical properties and functions, plays a critical role in neurodegenerative diseases. We report here that the enrichment of the neuronal plasma membrane with cholesterol protects cortical neurons from apoptosis induced by soluble oligomers of the Aβ(1,40) peptide. Conversely, cholesterol depletion using cyclodextrin renders cells more vulnerable to the cytotoxic effects of the Aβ‐soluble oligomers. Increasing the cholesterol content of small unilamellar liposomes also decreases Aβ‐dependent liposome fusion. We clearly demonstrate that cholesterol protection is specific to the soluble conformation of Aβ, because we observed no protective effects on cortical neurons treated by amyloid fibrils of the Aβ(1‐40) peptide. This may provide a new opportunity for the development of an effective AD therapy as well as elucidate the impact of the cholesterol level during AD development.

N-truncated amyloid-β oligomers induce learning impairment and neuronal apoptosis

N-terminal-truncated forms of amyloid-beta (A beta) peptide have been recently suggested to play a pivotal role early in Alzheimers disease (AD). Among them, A beta 3(pE)-42 peptide, starting with pyroglutamyl at residue Glu-3, is considered as the predominant A beta species in AD plaques and pre-amyloid lesions. Its abundance is reported to be directly proportional to the severity of the clinical phenotype. The present study investigates the effects of soluble oligomeric A beta 3(pE)-42 after intracerebroventricular injection on mice learning ability and the molecular mechanisms of its in vitro neurotoxicity. Mice injected with soluble A beta 3(pE)-42 or A beta(l-42) displayed impaired spatial working memory and delayed memory acquisition in Y-maze and Morris water maze tests, while those injected with soluble A beta(42-1) showed no effect. These cognitive alterations were associated with free radical overproduction in the hippocampus and olfactory bulbs, but not in the cerebral cortex or cerebellum. In vitro, A beta 3(pE)-42 oligomers induced a redox-sensitive neuronal apoptosis involving caspase activation and an arachidonic acid-dependent pro-inflammatory pathway. These data suggest that A beta 3(pE)-42 could mediate the neurodegenerative process and subsequent cognitive alteration occurring in preclinical AD stages.

Towards a nutritional approach for prevention of Alzheimer's disease : Biochemical and cellular aspects

Alzheimers disease (AD) is a major public health concern in all countries. Although the precise cause of AD is still unknown, a growing body of evidence supports the notion that soluble amyloid beta-peptide (Abeta) may be the proximate cause of synaptic injuries and neuronal death early in the disease. AD patients display lower levels of docosahexaenoic acid (DHA, C22:6 ; n-3) in plasma and brain tissues as compared to age-matched controls. Furthermore, epidemiological studies suggest that high DHA intake might have protective properties against neurodegenerative diseases. These observations are supported by in vivo studies showing that DHA-rich diets limits the synaptic loss and cognitive defects induced by Abeta peptide. Although the molecular basis of these neuroprotective effects remains unknown, several mechanisms have been proposed such as (i) regulation of the expression of potentially protective genes, (ii) activation of anti-inflammatory pathways, (iii) modulation of functional properties of the synaptic membranes along with changes in their physicochemical and structural features.

Additional Use of Aβ42/Aβ40 Ratio with Cerebrospinal Fluid Biomarkers P-Tau and Aβ42 Increases the Level of Evidence of Alzheimer's Disease Pathophysiological Process in Routine Practice

BACKGROUND Cerebrospinal fluid (CSF) biomarkers have recently been included in the criteria for the diagnosis of Alzheimers disease (AD). Since interpretation of CSF profile requires the combination of three parameters, biological data are not always conclusive and isolated elevation of phosphorylated tau (P-tau) or reduction of amyloid-β (Aβ)42 alone can be observed. In these cases, Aβ42/Aβ40 ratio could be more relevant than Aβ42 absolute values by considering inter-individual variations in the total amyloid load. OBJECTIVE The objective of this study was to assess the use of Aβ42/Aβ40 ratio to improve the accuracy of biological conclusions in the diagnosis of patients with ambiguous CSF Aβ42 or tau results. METHODS Among 386 lumbar punctures analyzed in the lab in 2 years, 122 showed ambiguous biological data that were completed by CSF Aβ40 quantification and Aβ42/Aβ40 ratio calculation. A biological conclusion was then made using 0.05 as the Aβ42/Aβ40 ratio cut-off. RESULTS Our results showed that one-third of the biological profiles of patients with atypical dementia were ambiguous. The addition of Aβ42/Aβ40 ratio increased the proportion of interpretable biological profiles from 69% to 87%, without changing the conclusion when usual biomarkers (Aβ42 and P-tau) were concordant. CONCLUSION Our results support the use of the Aβ42/Aβ40 ratio in addition to the usual CSF AD biomarkers for patients with ambiguous biological profiles. This method could be specifically directed to this population in order to improve the level of certainty for clinical routine practice.

Brain region‐specific immunolocalization of the lipolysis‐stimulated lipoprotein receptor (LSR) and altered cholesterol distribution in aged LSR+/− mice

Brain lipid homeostasis is important for maintenance of brain cell function and synaptic communications, and is intimately linked to age‐related cognitive decline. Because of the blood–brain barriers limiting nature, this tissue relies on a complex system for the synthesis and receptor‐mediated uptake of lipids between the different networks of neurons and glial cells. Using immunofluorescence, we describe the region‐specific expression of the lipolysis‐stimulated lipoprotein receptor (LSR), in the mouse hippocampus, cerebellum Purkinje cells, the ependymal cell interface between brain parenchyma and cerebrospinal fluid, and the choroid plexus. Colocalization with cell‐specific markers revealed that LSR was expressed in neurons, but not astrocytes. Latency in arms of the Y‐maze exhibited by young heterozygote LSR+/− mice was significantly different as compared to control LSR+/+, and increased in older LSR+/− mice. Filipin and Nile red staining revealed membrane cholesterol content accumulation accompanied by significantly altered distribution of LSR in the membrane, and decreased intracellular lipid droplets in the cerebellum and hippocampus of old LSR+/− mice, as compared to control littermates as well as young LSR+/− animals. These data therefore suggest a potential role of LSR in brain cholesterol distribution, which is particularly important in preserving neuronal integrity and thereby cognitive functions during aging.