Browsing by Author "Alvarez, A"

Now showing 1 - 10 of 10
  • No Thumbnail Available
    Item
    Acetylcholinesterase accelerates assembly of amyloid-beta-peptides into Alzheimer's fibrils: Possible role of the peripheral site of the enzyme
    (1996) Inestrosa, N.C.; Alvarez, A; Perez, CA; Moreno, RD; Vicente, M; Linker, C; Casanueva, OI; Soto, C; Garrido, J
    Acetylcholinesterase (AChE), an important component of cholinergic synapses, colocalizes with amyloid-beta peptide (A beta) deposits of Alzheimer's brain. We report here that bovine brain AChE, as well as the human and mouse recombinant enzyme, accelerates amyloid formation from wild-type A beta and a mutant A beta peptide, which alone produces few amyloid-like fibrils. The action of AChE was independent of the subunit array of the enzyme, was not affected by edrophonium, an active site inhibitor, but it was affected by propidium, a peripheral anionic binding site ligand. Butyrylcholinesterase, an enzyme that lacks the peripheral site, did not affect amyloid formation. Furthermore, AChE is a potent amyloid-promoting factor when compared with other A beta-associated proteins. Thus, in addition to its role in cholinergic synapses, AChE may function by accelerating A beta formation and could play a role during amyloid deposition in Alzheimer's brain.
  • No Thumbnail Available
    Item
  • No Thumbnail Available
    Item
    Acetylcholinesterase promotes the aggregation of amyloid-beta-peptide fragments by forming a complex with the growing fibrils
    (1997) Alvarez, A; Opazo, C; Alarcon, R; Garrido, J; Inestrosa, NC
    Acetylcholinesterase (AChE), an enzyme involved in the hydrolysis of the neurotransmitter acetylcholine, consistently colocalizes with the amyloid deposits characteristic of Alzheimer's disease and may contribute to the generation of amyloid proteins and/or physically affect fibril assembly. Ln order to identify the structural domains of the amyloid-beta-peptide (A beta) involved in the aggregation induced by AChE, we have studied the effect of this cholinergic enzyme on A beta peptide fragments of different sizes. AChE enhanced the aggregation of the A beta(12-28) and A beta(25-35) peptides but not of the A beta(1-16) fragment. The inductive effect of AChE on the aggregation of A beta(12-28) was abolished by the presence of either A beta(1-16) or A beta(9-21). The effect of the enzyme was also analysed using two different mutant fragments, possessing a low and the other a high capacity for fibrillogenesis. The fragments used were A beta(12-28)(Val18-->Ala) and A beta(12-28)(Glu22-->Gln), respectively. AChE was able to promote the aggregation of these fragments in a very specific way and both mutant peptides were able to form amyloid fibrils, as revealed by negative staining under the electron microscope. Binding assays indicated that AChE was bound to A beta(12-28), as well as to the A beta(1-16) peptide. AChE was seen to form strong complexes with the A beta(12-28) fibrils as such complexes stained positively for both thioflavine-T and AChE activity, were resistant to high ionic strength treatment, and were partially sensitive to detergents, suggesting that hydrophobic interactions may play a role in the stabilization of the AChE-A beta complex. Our results suggest that such amyloid-AChE complexes are formed when AChE interacts with the growing amyloid fibrils and accelerates the assembly of A beta peptides. This is consistent with the fact that AChE is known to be present within A beta deposits including the pre-amyloid diffuse and mature senile plaques found in Alzheimer's brain. (C) 1997 Academic Press Limited.
  • No Thumbnail Available
    Item
    Acetylcholinesterase, a senile plaque component, affects the fibrillogenesis of amyloid-beta-peptides
    (1995) Alvarez, A; Bronfman, F; Perez, CA; Vicente, M; Garrido, J; Inestrosa, NC
    Acetylcholinesterase (AChE) colocalizes with amyloid-beta peptide (A beta) deposits present in the brain of Alzheimer's patients. Recent studies showed that A beta(1-40) Can adopt two different conformational states in solution (an amyloidogenic conformer, A beta ac, and a non-amyloidogenic conformer, A beta nac) which have distinct abilities to form amyloid fibrils. We report here that AChE binds A beta nac and accelerates amyloid formation by the same peptide. No such effect was observed with A beta ac, the amyloidogenic conformer, suggesting that AChE acts as a 'pathological chaperone' inducing a conformational transition from A beta nac into A beta ac in vitro.
  • No Thumbnail Available
    Item
    Laminin blocks the assembly of wild type Aβ and the Dutch variant peptide into Alzheimer's fibrils
    (1998) Bronfman, FC; Alvarez, A; Morgan, C; Inestrosa, NC
    Amyloid fibril formation is believed to be a nucleation-dependent polymerization process which may be influenced by various other factors with important consequences for the development, prevention or treatment of amyloidosis. We have previously shown that laminin inhibits A beta peptide fibril formation in vitro. Here we present a kinetic study that indicates laminin to be a potent anti-amyloidosis factor, as it not only inhibited alpha beta(1-40) fibril aggregation, but also inhibited the aggregation of the Dutch A beta(1-40) variant, a peptide with a higher capacity to aggregate than the wild-type A beta(1-40). The inhibitory effect of laminin on amyloid fibril formation was not overcome by the addition of pre-formed A beta fibrils, suggesting that laminin inhibits the fibril elongation process. At the present time, however, we cannot rule out the possibility that laminin also affects the initial nucleation process of A beta fibril formation. On other hand, laminin was not able to counteract the amyloid fibril formation promoted by acetylcholinesterase (AChE), another component of the amyloid deposits found in AD brains. The effect of laminin may bet important as an inhibitor of A beta amyloidogenesis in vivo, specifically at the level of cerebral blood vessels.
  • No Thumbnail Available
    Item
    Laminin inhibits amyloid-beta-peptide fibrillation
    (1996) Bronfman, FC; Garrido, J; Alvarez, A; Morgan, C; Inestrosa, NC
    Laminin, an important extracellular matrix component is induced by brain injury and colocalizes with amyloid-beta-peptide (A beta) deposits in Alzheimer brains. We report here that laminin inhibits amyloid fibril formation as determined by thioflavin T fluorescence spectroscopy and electron microscopic examination. The inhibition of amyloid formation by laminin was concentration dependent and was observed at a laminin concentration of 300 nM, corresponding to a laminin/A beta protein molar ratio of 1:800. The potential effect of laminin, may prove important to inhibit AP fibrillogenesis in vivo, specifically at the level of cerebral blood vessels.
  • No Thumbnail Available
    Item
    Molecular modeling of the amyloid-β-peptide using the homology to a fragment of triosephosphate isomerase that forms amyloid in vitro
    (1999) Contreras, CF; Canales, MA; Alvarez, A; De Ferrari, GV; Inestrosa, NC
    The main component of the amyloid senile plaques found in Alzheimer's brain is the amyloid-beta-peptide (A beta), a proteolytic product of a membrane precursor protein. Previous structural studies have found different conformations for the A beta peptide depending on the solvent and pH used. In general, they have suggested an alpha-helix conformation at the N-terminal domain and a beta-sheet conformation for the C-terminal domain. The structure of the complete A beta peptide (residues 1-40) solved by NMR has revealed that only helical structure is present in A beta. However, this result cannot explain the large beta-sheet A beta aggregates known to form amyloid under physiological conditions. Therefore, we investigated the structure of A beta by molecular modeling based on extensive homology using the Smith and Waterman algorithm implemented in the MPsrch program (Blitz server). The results showed a mean value of 23 % identity with selected sequences. Since these values do not allow a clear homology to be established with a reference structure in order to perform molecular modeling studies, we searched for detailed homology, A 28% identity with an alpha/beta segment of a triosephosphate isomerase (TIM) from Culex tarralis with an unsolved three-dimensional structure was obtained. Then, multiple sequence alignment was performed considering A beta, TIM from C.tarralis and another five TIM sequences with known three-dimensional structures. We found a TIM segment with secondary structure elements in agreement with previous experimental data for A beta. Moreover, when a synthetic peptide from this TIM segment was studied in vitro, it was able to aggregate and to form amyloid fibrils, as established by Congo red binding and electron microscopy, The A beta model obtained was optimized by molecular dynamics considering ionizable side chains in order to simulate A beta in a neutral pH environment. We report here the structural implications of this study.
  • No Thumbnail Available
    Item
    Protein kinase C inhibits amyloid β-peptide neurotoxicity by acting on members of the Wnt pathway
    (2002) Garrido, JL; Godoy, J; Alvarez, A; Bronfman, M; Inestrosa, NC
    Current evidence supports the notion that the amyloid beta-peptide (Abeta) plays a major role in the neurotoxicity observed in the brain in Alzheimer's disease. However, the signal transduction mechanisms involved still remain unknown. In the present work, we analyzed the effect of protein kinase C (PKC) on some members of the Writ signaling pathway and its implications for Abeta neurotoxicity. Activation of PKC by phorbol 12-myristate 13-acetate protected rat hippocampal neurons from A(3 toxicity. This effect was accomplished by inhibition of glycogen synthase kinase-3beta (GSK-3beta) activity, which led to the accumulation of cytoplasmic beta-catenin and transcriptional activation via beta-catenin/T-cell factor/lymphoid enhancer factor-1 (TCF/LEF-1) of Writ target genes, which in the present study were engrailed-1 (en-1) and cyclin D1 (cycD1). In contrast, inhibition of Ca2+-dependent PKC isoforms activated GSK-3beta and offered no protection from Abeta neurotoxicity. Wnt-3a and lithium salts, classical activators of the Writ pathway, mimicked PKC activation. Our results suggest that regulation of members of the Writ signaling pathway by Ca2+-dependent PKC isoforms may be important in controlling the neurotoxic process induced by Abeta.
  • No Thumbnail Available
    Item
    Stable complexes involving acetylcholinesterase and amyloid-β peptide change the biochemical properties of the enzyme and increase the neurotoxicity of Alzheimer's fibrils
    (1998) Alvarez, A; Alarcón, R; Opazo, C; Campos, EO; Muñoz, FJ; Calderón, FH; Dajas, F; Gentry, MK; Doctor, BP; De Mello, FG; Inestrosa, NC
    Brain acetylcholinesterase (AChE) forms stable complexes with amyloid-beta peptide (A beta) during its assembly into filaments, in agreement with its colocalization with the A beta deposits of Alzheimer's brain. The association of the enzyme with nascent A beta aggregates occurs as early as after 30 min of incubation. Analysis of the catalytic activity of the AChE incorporated into these complexes shows an anomalous behavior reminiscent of the AChE associated with senile plaques, which includes a resistance to low pH, high substrate concentrations, and lower sensitivity to AChE inhibitors. Furthermore, the toxicity of the AChE-amyloid complexes is higher than that of the A beta aggregates alone. Thus, in addition to its possible role as a heterogeneous nucleator during amyloid formation, AChE, by forming such stable complexes, may increase the neurotoxicity of A beta fibrils and thus may determine the selective neuronal loss observed in Alzheimer's brain.
  • No Thumbnail Available
    Item
    Wnt signaling involvement in β-amyloid-dependent neurodegeneration
    (2002) Inestrosa, NC; De Ferrari, GV; Garrido, JL; Alvarez, A; Olivares, GH; Barría, MI; Bronfman, M; Chacón, MA
    Alzheimer's disease (AD) is a progressive dementia paralleled by selective neuronal death, which is probably caused by the cytotoxic effects of the amyloid-p peptide (Abeta). We have observed that Abeta-dependent neurotoxicity induces a loss of function of Wnt signaling components and that activation of this signaling cascade prevent such cytotoxic effects. Therefore we propose that compounds which mimic this signaling cascade may be candidates for therapeutic intervention in Alzheimer's patients. (C) 2002 Elsevier Science Ltd. All rights reserved.