Heparin attenuates cytotoxic and inflammatory activity of Alzheimer amyloid-β in vitro (original) (raw)
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Heparin attenuates cytotoxic and inflammatory activity of Alzheimer amyloid-� in vitro
Neurobiol Aging, 2002
Amyloid- protein (A) is implicated in the pathogenesis of Alzheimer's disease because of its neurotoxicity and the ability to trigger local inflammation. Compounds that interact with the amino acids of the N-terminal region or interfere with aggregation can reduce the A biologic activity. We evaluated the effect of heparin on A (1-42) neurotoxicity and on its ability to activate complement and contact system. On differentiated PC12 cells, a reliable model of neuronal cells, heparin at the doses of 10 and 20 g/ml significantly counteracted A cytotoxicity as assessed by measuring MTT conversion. We then explored the effect of heparin on A (1-42)-induced complement and contact system activation. A (1-42) was incubated with heparin in presence of normal plasma as the source of complement and contact system factors. Heparin reduced, in a dose-dependent manner, complement and contact system activation, assessed by measuring the degree of C4 and high molecular weight kininogen cleavage. The present data show that heparin can attenuate neurotoxic and pro-inflammatory activity of A and suggest that this drug could represent a new strategy to reduce the progressive neurodegeneration in AD.
Journal of Neuroscience, 2004
We investigated the effect of long-term, peripheral treatment with enoxaparin, a low molecular weight heparin, in transgenic mice overexpressing human amyloid precursor protein 751. Enoxaparin (6 IU per mouse intraperitoneally, three times a week for 6 months) significantly lowered the number and the area occupied by cortical -amyloid deposits and the total -amyloid (1-40) cortical concentration. Immunocytochemical analysis of glial fibrillary acid protein-positive cells showed that enoxaparin markedly reduced the number of activated astrocytes surrounding -amyloid deposits. In vitro, the drug dose-dependently attenuated the toxic effect of -amyloid on neuronal cells. Enoxaparin dose-dependently reduced the ability of -amyloid to activate complement and contact systems, two powerful effectors of inflammatory response in AD brain. By reducing the -amyloid load and cytotoxicity and proinflammatory activity, enoxaparin offers promise as a tool for slowing the progression of Alzheimer's disease.
Journal of Biological Chemistry, 1997
Aggregation and deposition of the 40-42-residue amyloid -protein (A) are early and necessary neuropathological events in Alzheimer's disease. An understanding of the molecular interactions that trigger these events is important for therapeutic strategies aimed at blocking A plaque formation at the earliest stages. Heparan sulfate proteoglycans may play a fundamental role since they are invariably associated with A and other amyloid deposits at all stages. However, the nature of the A-heparan sulfate proteoglycan binding has been difficult to elucidate because of the strong tendency of A to self-aggregate. Affinity co-electrophoresis can measure the binding of proteoglycans or glycosaminoglycans to proteins without altering the physical state of the protein during the assay. We used affinity co-electrophoresis to study the interaction between A and the glycosaminoglycan heparin and found that the aggregation state of A governs its heparin-binding properties: heparin binds to fibrillar but not nonfibrillar A. The amyloid binding dye, Congo red, inhibited the interaction in a specific and dose-dependent manner. The "Dutch" mutant A E22Q peptide formed fibrils more readily than wild type A and it also attained a heparin-binding state more readily, but, once formed, mutant and wild type fibrils bound heparin with similar affinities. The heparin-binding ability of aggregated A E22Q was reversible with incubation in a solvent that promotes ␣-helical conformation, further suggesting that conformation of the peptide is important. Studies with another human amyloidogenic protein, amylin, suggested that its heparinbinding properties were also dependent on aggregation state. These results demonstrate the dependence of the A-heparin interaction on the conformation and aggregation state of A rather than primary sequence alone, and suggest methods of interfering with this association.
Journal of Biological Chemistry, 1999
Serum amyloid A isoforms, apoSAA1 and apoSAA2, are apolipoproteins of unknown function that become major components of high density lipoprotein (HDL) during the acute phase of an inflammatory response. ApoSAA is also the precursor of inflammation-associated amyloid, and there is strong evidence that the formation of inflammation-associated and other types of amyloid is promoted by heparan sulfate (HS). Data presented herein demonstrate that both mouse and human apoSAA contain binding sites that are specific for heparin and HS, with no binding for the other major glycosaminoglycans detected. Cyanogen bromide-generated peptides of mouse apoSAA1 and apoSAA2 were screened for heparin binding activity. Two peptides, an apoSAA1derived 80-mer (residues 24 -103) and a smaller carboxyl-terminal 27-mer peptide of apoSAA2 (residues 77-103), were retained by a heparin column. A synthetic peptide corresponding to the CNBr-generated 27-mer also bound heparin, and by substituting or deleting one or more of its six basic residues (Arg-83, His-84, Arg-86, Lys-89, Arg-95, and Lys-102), their relative importance for heparin and HS binding was determined. The Lys-102 residue appeared to be required only for HS binding. The residues Arg-86, Lys-89, Arg-95, and Lys-102 are phylogenetically conserved suggesting that the heparin/HS binding activity may be an important aspect of the function of apoSAA. HS linked by its carboxyl groups to an Affi-Gel column or treated with carbodiimide to block its carboxyl groups lost the ability to bind apoSAA. HDL-apoSAA did not bind to heparin; however, it did bind to HS, an interaction to which apoA-I contributed. Results from binding experiments with Congo Red-Sepharose 4B columns support the conclusions of a recent structural study which found that heparin binding domains have a common spatial distance of about 20 Å between their two outer basic residues. Our present work provides direct evidence that apoSAA can associate with HS (and heparin) and that the occupation of its binding site by HS, and HS analogs, likely caused the previously reported increase in amyloidogenic conformation sheet) of apoSAA2 (McCubbin, W. D., Kay, C. M., Narindrasorasak, S., and Kisilevsky, R. (1988) Biochem. J. 256, 775-783) and their amyloid-suppressing effects in vivo
Heparin promotes ß-secretase cleavage of the Alzheimer's amyloid precursor protein
Neurochemistry International, 1997
In Alzheimer's disease, abnormal processing of the amyloid precursor protein (APP) is thought to play an important role in amyloid deposition. We investigated the effect of heparin, a highly sulfated glycosaminoglycan related to heparan sulfate, on the secretion of the ß-secretase cleavage product of APP (sAPPß) in a human neuroblastoma cell line. Heparin induced an increase in the secretion of
Biochemistry, 2006
BACE1 is an aspartic protease that generates the N-terminus of the -amyloid protein (Α ) from the -amyloid precursor protein (APP). BACE1 is a key target for Alzheimer drug development. However, little is known about the physiological regulation of the enzyme. Heparin can promote -secretase cleavage of APP in neuroblastoma cells. However, heparin has also been reported to directly inhibit BACE1 activity in vitro. To clarify the role of heparin in regulating BACE1, we examined the effect of heparin on the activity of recombinant human BACE1 (rBACE1) in vitro. Low concentrations (1 µg/mL) of heparin were found to stimulate rBACE1, increasing enzyme V max and decreasing the K M . In contrast, higher concentrations of heparin (10 or 100 µg/mL) were inhibitory. Heparin affinity chromatography demonstrated that heparin interacted strongly with the zymogen form of rBACE1 and bound to a peptide homologous to the N-terminal pro sequence of BACE1. Mature (pro sequence cleaved) enzyme lacked the capacity to be stimulated by heparin, indicating that the pro domain was necessary for the stimulation by heparin. Furthermore, in the presence of stimulatory concentrations of heparin, there was an increase in autocatalytic cleavage of the protease domain and a subsequent loss of enzyme activity in vitro. Our results strongly suggest that heparin stimulates the partially active BACE1 zymogen, and we propose that the activation is mediated by high-affinity binding of heparin to the pro domain. Our study provides evidence that heparan sulfate proteoglycans could regulate the rate of Α production in vivo.
Febs Letters, 1997
Deletion mutagenesis studies have suggested that there are two domains within APP which bind heparan sulphate. These domains have been cloned and expressed in the yeast Pichia pastoris. Both recombinant proteins bound to heparin. One domain (APP316-447) was further characterised by binding studies with peptides encompassing this region. Peptides homologous to APP316-346 and APP416-447 were found to bind heparin. Circular dichroism studies show that APP416-447 shifted towards an a-helical conformation in the presence of heparin. This study suggests that heparin-binding domains may lie within regions high in a-helical structure.
Heparin Oligosaccharides as Potential Therapeutic Agents in Senile Dementia
Current Pharmaceutical Design, 2007
Heparin is a glycosaminoglycan mixture currently used in prophylaxis and treatment of thrombosis. Heparin possesses non-anticoagulant properties, including modulation of various proteases, interactions with fibroblast growth factors, and anti-inflammatory actions. Senile dementia of Alzheimer's type is accompanied by inflammatory responses contributing to irreversible changes in neuronal viability and brain function. Vascular factors are also involved in the pathogenesis of senile dementia. Inflammation, endogenous proteoglycans, and assembly of senile plagues and neurofibrillary tangles contribute directly and indirectly to further neuronal damage. Neuron salvage can be achieved by antiinflammation and the competitive inhibition of proteoglycans accumulation. The complexity of the pathology of senile dementia provides numerous potential targets for therapeutic interventions designed to modulate inflammation and proteoglycan assembly. Heparin and related oligosaccharides are known to exhibit anti-inflammatory effects as well as inhibitory effects on proteoglycan assembly and may prove useful as neuroprotective agents.