- Victoria, Australia
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The b-amyloid protein (Ab) is the major protein component of amyloid plaques found in the Alzheimer brain. Although there is a loss of acetylcholinesterase (AChE) from both cholinergic and non-cholinergic neurones in the brain of... more
The b-amyloid protein (Ab) is the major protein component of amyloid plaques found in the Alzheimer brain. Although there is a loss of acetylcholinesterase (AChE) from both cholinergic and non-cholinergic neurones in the brain of Alzheimer patients, the level of AChE is increased around amyloid plaques. Previous studies using P19 cells in culture and transgenic mice which overexpress human Ab
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With the advent of effective therapeutic agents for the treatment of Alzheimer's disease (AD), accurate diagnosis will become increasingly important. To date, clinical diagnosis based on neuropsychological assess-ment is time... more
With the advent of effective therapeutic agents for the treatment of Alzheimer's disease (AD), accurate diagnosis will become increasingly important. To date, clinical diagnosis based on neuropsychological assess-ment is time consuming and often inaccurate. Therefore, ...
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Pathogenic substitutions in leucine-rich repeat kinase 2 (LRRK2, Lrrk2) have been genetically linked to familial, late-onset Parkinsonism. End-stage disease is predominantly associated with nigral neuronal loss and Lewy body pathology,... more
Pathogenic substitutions in leucine-rich repeat kinase 2 (LRRK2, Lrrk2) have been genetically linked to familial, late-onset Parkinsonism. End-stage disease is predominantly associated with nigral neuronal loss and Lewy body pathology, but patients may have gliosis, tau or ubiquitin inclusions (pleomorphic pathology). The anatomical distribution of Lrrk2 protein may provide insight into its function in health and neurodegeneration, thus we performed a comparative study with 'in-house' and commercially available Lrrk2 antibodies using brain tissue from wild type and human Lrrk2 transgenic bacterial artificial chromosome (BAC) mice, and from diffuse Lewy body disease (DLBD) patients. Lrrk2 protein was ubiquitously expressed and relatively abundant in most brain regions, including the substantia nigra, thalamus and striatum. Lrrk2 was not a major component of Lewy body or neuritic pathology associated with Parkinson's disease. However, selective loss of dopaminergic neurons in Lrrk2-associated Parkinsonism argues the protein may have regional-specific interactions. Lrrk2 immunohistochemical staining was present in the subventricular zone, a region containing stem cells that give rise to both neurons and glia. A role for Lrrk2 in neurogenesis might provide further insight into the aberrant role of mutant protein in age-associated neurodegeneration with pleomorphic pathology.
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Parkinson&am... more
Parkinson's disease (PD) is a severe neurodegenerative disorder characterised by loss of dopaminergic neurons of the substantia nigra. The pathological hallmarks are cytoplasmic inclusions termed Lewy bodies consisting primarily of aggregated alpha-synuclein (alphaSN). Different lines of transgenic mice have been developed to model PD but have failed to recapitulate the hallmarks of this disease. Since treatment of rodents with the pesticide rotenone can reproduce nigrostriatal cell loss and other features of PD, we aimed to test chronic oral administration of rotenone to transgenic mice over-expressing human alphaSN with the A53T mutation. Initial assessment of this transgenic line for compensatory molecular changes indicated decreased brain beta-synuclein expression and significantly increased levels of the PD-associated oxidative stress response protein, DJ-1, and the E3 ubiquitin ligase enzyme, Parkin. Rotenone treatment of 30 mg/kg for 25 doses over a 35-day period was tolerated in the transgenic mice and resulted in decreased spontaneous locomotor movement and increased cytoplasmic alphaSN expression. The mitochondrial Parkinson's-associated PTEN-induced kinase 1 protein levels were also increased in transgenic mouse brain after rotenone treatment; there was no change in brain dopamine levels or nigrostriatal cell loss. These hA53T alphaSN transgenic mice provide a useful model for presymptomatic Parkinson's features and are valuable for study of associated compensatory changes in early Parkinson's disease stages.
Research Interests: Cognitive Science, Molecular Neuroscience, Transgenic Mice, Alpha Synuclein, Molecular, and 13 moreDopamine, Humans, Protein Kinases, Mice, Animals, Substantia nigra, Enzyme, Ubiquitin ligase, Transgenic Mouse Technology, Neurosciences, parkinsonian disorders, Rotenone, and Neurodegenerative disorder
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Research Interests: Genetics, Regulation, Biological Sciences, Humans, Sequence alignment, and 15 moreProtein Kinases, Mutation, Animals, Human Molecular Genetics, Phosphorylation, Cattle, Histones, Protein Kinase, Amino Acid Sequence, Recombinant Proteins, Parkinson Disease, Down-Regulation, Mitochondrial Proteins, Molecular Sequence Data, and Medical and Health Sciences
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Research Interests: Oxidative Stress, Gene expression, Amyotrophic Lateral Sclerosis, Biological Sciences, Copper, and 12 moreMovement, Mice, Animals, Spinal Cord, Zinc, Superoxide Dismutase, Mouse Model, Glial Fibrillary Acidic Protein, Protein Quaternary Structure, Cell Survival, Amyloid Beta Precursor Protein, and Gliosis
The primary constituent of the amyloid plaque, β-amyloid (Aβ), is thought to be the causal “toxic moiety” of Alzheimer's disease. However, despite much work focused on both Aβ and its parent protein, amyloid precursor protein (APP), the... more
The primary constituent of the amyloid plaque, β-amyloid (Aβ), is thought to be the causal “toxic moiety” of Alzheimer's disease. However, despite much work focused on both Aβ and its parent protein, amyloid precursor protein (APP), the functional roles of APP and its cleavage products remain to be fully elucidated. Protein–protein interaction networks can provide insight into protein function, however, high-throughput data often report false positives and are in frequent disagreement with low-throughput experiments. Moreover, the complexity of the CNS is likely to be under represented in such databases. Therefore, we curated the published work characterizing both APP and Aβ to create a protein interaction network of APP and its proteolytic cleavage products, with annotation, where possible, to the level of APP binding domain and isoform. This is the first time that an interactome has been refined to domain level, essential for the interpretation of APP due to the presence of multiple isoforms and processed fragments. Gene ontology and network analysis were used to identify potentially novel functional relationships among interacting proteins.
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The amyloid protein (A beta or beta A4) is the major constituent of amyloid plaques in the Alzheimer's disease brain. A beta is cleaved from the amyloid precursor protein (APP) by a mechanism which is poorly understood.... more
The amyloid protein (A beta or beta A4) is the major constituent of amyloid plaques in the Alzheimer's disease brain. A beta is cleaved from the amyloid precursor protein (APP) by a mechanism which is poorly understood. Cell culture studies suggest that APP may be cleaved by secretases within the late Golgi compartment. Studies performed so far have mainly used exogenous APP and synthetic peptides as substrates. For this study, a Golgi and plasma membrane-enriched fraction was isolated from rat brain and incubated at 37 degrees C at pH 7.2 to study the degradation of endogenous APP. The breakdown of APP was accompanied by the concomitant generation of A beta-containing C-terminal fragments, in a time-dependent fashion. The metal ion chelators EDTA and 1,10-phenanthroline inhibited this degradation. The inhibition by EDTA was reversed by 50 microM Zn2+ but not by other metal ions. The protease activity was not inhibited by cysteine, serine or aspartic protease inhibitors nor was it inhibited by compounds which are inhibitors of known metalloendopeptidases and matrix metalloproteinases (cFP, phosphoramidon and TIMP-2). Our data suggest that a novel Zn(2+)-dependent metalloprotease activity associated with a Golgi and plasma membrane-enriched fraction can degrade endogenous APP to generate A beta containing C-terminal fragments. This protease may generate amyloidogenic fragments of APP which may serve as precursors for A beta.
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Τhe main component of cerebral amyloid angiopathy (CAA) in Alzheimer's disease is the amyloid-β protein (Aβ), a 4-kDa polypeptide derived from the β-amyloid protein precursor (APP). The accumulation of Aβ in the basement membrane has been... more
Τhe main component of cerebral amyloid angiopathy (CAA) in Alzheimer's disease is the amyloid-β protein (Aβ), a 4-kDa polypeptide derived from the β-amyloid protein precursor (APP). The accumulation of Aβ in the basement membrane has been implicated in the degeneration of adjacent vascular smooth muscle cells (VSMC). However, the mechanism of Aβ toxicity is still unclear. In this study, we examined the effect of substrate-bound Aβ on VSMC in culture. The use of substrate-bound proteins in cell culture mimics presentation of the proteins to cells as if bound to the basement membrane. Substrate-bound Aβ peptides were found to be toxic to the cells and to increase the rate of cell death. This toxicity was dependent on the length of time the peptide was allowed to ‘age’, a process by which Aβ is induced to aggregate over several hours to days. Oxidative stress via hydrogen peroxide (H2O2) release was not involved in the toxic effect, as no decrease in toxicity was observed in the presence of catalase. However, substrate-bound Aβ significantly reduced cell adhesion compared to cells grown on plastic alone, indicating that cell–substrate adhesion may be important in maintaining cell viability. Aβ also caused an increase in the number of apoptotic cells. This increase in apoptosis was accompanied by activation of caspase-3. Homocysteine, a known risk factor for cerebrovascular disease, increased Aβ-induced toxicity and caspase-3 activation in a dose-dependent manner. These studies suggest that Aβ may activate apoptotic pathways to cause loss of VSMC in CAA by inhibiting cell–substrate interactions. Our studies also suggest that homocysteine, a known risk factor for other cardiovascular diseases, could also be a risk factor for hemorrhagic stroke associated with CAA.
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Research Interests: Psychology, Animal Behavior, Transgenic Mice, Anxiety, Alpha Synuclein, and 17 moreHumans, Mutation, Mice, Animals, Open Field, Young Adult, Clinical Sciences, Experimental, Analysis of Variance, Elevated Plus Maze, ANXIETY, Experimental Neurology, Exploratory Behavior, Maze Learning, Neurosciences, Knock Out Mice, and Motor activity
Mutations in PTEN-induced kinase 1 (PINK1) gene cause PARK6 familial Parkinsonism. To decipher the role of PINK1 in pathogenesis of Parkinson’s disease (PD), researchers need to identify protein substrates of PINK1 kinase activity that... more
Mutations in PTEN-induced kinase 1 (PINK1) gene cause PARK6 familial Parkinsonism. To decipher the role of PINK1 in pathogenesis of Parkinson’s disease (PD), researchers need to identify protein substrates of PINK1 kinase activity that govern neuronal survival, and establish whether aberrant regulation and inactivation of PINK1 contribute to both familial Parkinsonism and idiopathic PD. These studies should take into account the several unique structural and functional features of PINK1. First PINK1 is a rare example of a protein kinase with a predicted mitochondrial-targeting sequence and a possible resident mitochondrial function. Second, bioinformatic analysis reveals unique insert regions within the kinase domain that are potentially involved in regulation of kinase activity, substrate selectivity and stability of PINK1. Third, the C-terminal region contains functional motifs governing kinase activity and substrate selectivity. Fourth, accumulating evidence suggests that PINK1 interacts with other signaling proteins implicated in PD pathogenesis and mitochondrial dysfunction. The most prominent examples are the E3 ubiquitin ligase Parkin, the mitochondrial protease high temperature requirement serine protease 2 and the mitochondrial chaperone tumor necrosis factor receptor-associated protein 1. How PINK1 may regulate these proteins to maintain neuronal survival is unclear. This review describes the unique structural features of PINK1 and their possible roles in governing mitochondrial import, processing, kinase activity, substrate selectivity and stability of PINK1. Based upon the findings of previous studies of PINK1 function in cell lines and animal models, we propose a model on the neuroprotective mechanism of PINK1. This model may serve as a conceptual framework for future investigation into the molecular basis of PD pathogenesis.
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The major constituent of amyloid plaques in the Alzheimer disease (AD) brain is the amyloid protein (Aβ). Aβ has been shown to be neurotoxic to cells, but the exact mechanism of its effects are still not known. Most studies have focussed... more
The major constituent of amyloid plaques in the Alzheimer disease (AD) brain is the amyloid protein (Aβ). Aβ has been shown to be neurotoxic to cells, but the exact mechanism of its effects are still not known. Most studies have focussed on Aβ neurotoxicity, but little is known ...
Research Interests:
Parkinson's disease (PD) is a severe neurodegenerative disorder characterised by loss of dopaminergic neurons of the substantia nigra. The pathological hallmarks are cytoplasmic inclusions termed Lewy bodies consisting primarily of... more
Parkinson's disease (PD) is a severe neurodegenerative disorder characterised by loss of dopaminergic neurons of the substantia nigra. The pathological hallmarks are cytoplasmic inclusions termed Lewy bodies consisting primarily of aggregated α-synuclein (αSN). Different lines of transgenic mice have been developed to model PD but have failed to recapitulate the hallmarks of this disease. Since treatment of rodents with the pesticide rotenone can reproduce nigrostriatal cell loss and other features of PD, we aimed to test chronic oral administration of rotenone to transgenic mice over-expressing human αSN with the A53T mutation. Initial assessment of this transgenic line for compensatory molecular changes indicated decreased brain β-synuclein expression and significantly increased levels of the PD-associated oxidative stress response protein, DJ-1, and the E3 ubiquitin ligase enzyme, Parkin. Rotenone treatment of 30 mg/kg for 25 doses over a 35-day period was tolerated in the transgenic mice and resulted in decreased spontaneous locomotor movement and increased cytoplasmic αSN expression. The mitochondrial Parkinson's-associated PTEN-induced kinase 1 protein levels were also increased in transgenic mouse brain after rotenone treatment; there was no change in brain dopamine levels or nigrostriatal cell loss. These hA53T αSN transgenic mice provide a useful model for presymptomatic Parkinson's features and are valuable for study of associated compensatory changes in early Parkinson's disease stages.
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Cerebral amyloid angiopathy (CAA) is a major feature of Alzheimer's disease pathology. In CAA, degeneration of vascular smooth muscle cells (VSMCs) occurs close to regions of the basement membrane where the amyloid protein (Aβ) builds up.... more
Cerebral amyloid angiopathy (CAA) is a major feature of Alzheimer's disease pathology. In CAA, degeneration of vascular smooth muscle cells (VSMCs) occurs close to regions of the basement membrane where the amyloid protein (Aβ) builds up. In this study, the possibility that Aβ disrupts adhesive interactions between VSMCs and the basement membrane was examined. VSMCs were cultured on a commercial basement membrane substrate (Matrigel). The presence of Aβ in the Matrigel decreased cell-substrate adhesion and cell viability. Full-length oligomeric Aβ was required for the effect, as N- and C-terminally truncated peptide analogues did not inhibit adhesion. Aβ that was fluorescently labelled at the N-terminus (fluo-Aβ) bound to Matrigel as well as to the basement membrane heparan sulfate proteoglycan (HSPG) perlecan and laminin. Adhesion of VSMCs to perlecan or laminin was decreased by Aβ. As perlecan influences VSMC viability through the extracellular signal-regulated kinase (ERK)1/2 signalling pathway, the effect of Aβ1–40 on ERK1/2 phosphorylation was examined. The level of phospho-ERK1/2 was decreased in cells following Aβ treatment. An inhibitor of ERK1/2 phosphorylation enhanced the effect of Aβ on cell adhesion. The studies suggest that Aβ can decrease VSMC viability by disrupting VSMC–extracellular matrix (ECM) adhesion.
Research Interests: Neurochemistry, Immunohistochemistry, Atomic Force Microscopy, Extracellular Matrix, Signal Transduction, and 14 moreBasement Membrane, Female, Animals, NF-kappa B, Cell Viability, Neurons, Mitogen Activated Protein Kinase, Rats, Tumor necrosis factor-alpha, Transfection, Neurosciences, Ceramides, Down-Regulation, and Calcitonin Gene-Related Peptide
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Accumulation of beta amyloid (Aβ) in the brain is central to the pathogenesis of Alzheimer's disease. Aβ can bind to membrane lipids and this binding may have detrimental effects on cell function. In this study, surface plasmon resonance... more
Accumulation of beta amyloid (Aβ) in the brain is central to the pathogenesis of Alzheimer's disease. Aβ can bind to membrane lipids and this binding may have detrimental effects on cell function. In this study, surface plasmon resonance technology was used to study Aβ binding to membranes. Aβ peptides bound to synthetic lipid mixtures and to an intact plasma membrane preparation isolated from vascular smooth muscle cells. Aβ peptides were also toxic to vascular smooth muscle cells. There was a good correlation between the toxic effect of Aβ peptides and their membrane binding. ‘Ageing’ the Aβ peptides by incubation for 5 days increased the proportion of oligomeric species, and also increased toxicity and the amount of binding to lipids. The toxicities of various Aβ analogs correlated with their lipid binding. Significantly, binding was influenced by the concentration of cholesterol in the lipid mixture. Reduction of cholesterol in vascular smooth muscle cells not only reduced the binding of Aβ to purified plasma membrane preparations but also reduced Aβ toxicity. The results support the view that Aβ toxicity is a direct consequence of binding to lipids in the membrane. Reduction of membrane cholesterol using cholesterol-lowering drugs may be of therapeutic benefit because it reduces Aβ-membrane binding.
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The major constituent of amyloid plaques in the Alzheimer disease (AD) brain is the amyloid protein (Aβ). Aβ has been shown to be neurotoxic to cells, but the exact mechanism of its effects are still not known. Most studies have focussed... more
The major constituent of amyloid plaques in the Alzheimer disease (AD) brain is the amyloid protein (Aβ). Aβ has been shown to be neurotoxic to cells, but the exact mechanism of its effects are still not known. Most studies have focussed on Aβ neurotoxicity, but little is known ...
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Bovine and normal human articular chondrocytes in suspension culture were treated with misoprostol (an analog of prostaglandin E(1) [PGE(1)]), alone and in combination with interleukin-1 (IL-1) and/or diclofenac sodium, to study effects... more
Bovine and normal human articular chondrocytes in suspension culture were treated with misoprostol (an analog of prostaglandin E(1) [PGE(1)]), alone and in combination with interleukin-1 (IL-1) and/or diclofenac sodium, to study effects on proteoglycan metabolism. A concentrations of 50 ng ml(minus sign1) and above, misoprostol suppressed synthesis of proteoglycans but did not affect their rate of catabolism. The mild inhibitory effect of misoprostol on proteoglycan synthesis was additive to that of IL-1, especially in chondrocytes from the superficial zone of bovine articular cartilage. In IL-1-treated cultures, diclofenac NA caused a modest improvement in proteoglycan synthesis, but this beneficial effect was diminished by the simultaneous addition of misoprostol. Cartilage or chondrocyte cultures treated with IL-1, in which proteoglycan synthesis is suppressed, serve as model systems in which to study metabolic responses of chondrocytes to potential therapeutic agents, but in these experiments, no chondroprotective effects of misoprostol were observed in IL-1-activated chondrocytes.
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Reelin is a glycoprotein that modulates synaptic function and plasticity in the mature brain, thereby favouring memory formation. We recently reported altered cerebral Reelin expression in... more
Reelin is a glycoprotein that modulates synaptic function and plasticity in the mature brain, thereby favouring memory formation. We recently reported altered cerebral Reelin expression in Alzheimer's disease (AD). Here we demonstrate pronounced Reelin changes at protein and mRNA levels in the frontal cortex in adult Down's syndrome (DS), where the extra copy of chromosome 21 leads to overexpression of beta-amyloid. In cortical extracts of fetal DS samples we detected increased levels of the full-length Reelin and the 310-kDa fragment. Overexpression of mutant human amyloid precursor protein also led to an increase in levels of Reelin fragments in Tg2576 transgenic mice for human beta-amyloid. Finally, in vitro Abeta42 treatment of SH-SY5Y neuroblastoma cells led to increased Reelin levels. An altered pattern of Reelin glycosylation was detected in extracts from the frontal cortex of AD patients and in Abeta42-treated SH-SY5Y cells, supporting the notion that beta-amyloid triggers altered Reelin processing. These results provide evidence that Reelin expression and processing is altered in several amyloid conditions.
Research Interests: Neurobiology Of Disease, Neurobiology, Down Syndrome, Transgenic Mice, Amyloid, and 23 moreBrain, Humans, Cerebral Cortex, Mice, Female, Animals, Male, P-glycoprotein, Neuronal Plasticity, Frontal Cortex, Glycosylation, Clinical Sciences, Aged, Middle Aged, Adult, SYNAPSES, Fetus, Alzheimer Disease, Neurosciences, Full Length Movies, Gene Expression Regulation, Extracellular Matrix Proteins, and Amyloid Beta Precursor Protein
Research Interests: Kinetics, Western blotting, Amyloid, Humans, Animals, and 15 morePeptides, Trypsin, Sodium Dodecyl Sulfate-Polyacrylamide Gel Electrophoresis, Amyloid Beta, European, Time Factors, Sympathetic Nervous System, Protein isoforms, Recombinant Proteins, Protein Binding, Alzheimer Disease, Serine Protease, PLANT PROTEINS, Biochemistry and cell biology, and Amyloid Beta Precursor Protein
Research Interests: Gene expression, Western blotting, Humans, Heparin, Mutagenesis, and 15 moreCircular Dichroism, Escherichia coli, Pichia pastoris, Affinity chromatography, Molecular cloning, Protein Secondary Structure Prediction, Amino Acid Profile, Heparan Sulphate, Recombinant Protein, Polyacrylamide Gel Electrophoresis, Recombinant Proteins, Protein Binding, Alzheimer Disease, Biochemistry and cell biology, and Amyloid Beta Precursor Protein
Transgenic mice carrying mutant Cu/Zn superoxide dismutase (SOD1) recapitulate the motor impairment of human amyotrophic lateral sclerosis (ALS). The amyloid-β (Aβ) peptide associated with Alzheimer's disease is neurotoxic. To investigate... more
Transgenic mice carrying mutant Cu/Zn superoxide dismutase (SOD1) recapitulate the motor impairment of human amyotrophic lateral sclerosis (ALS). The amyloid-β (Aβ) peptide associated with Alzheimer's disease is neurotoxic. To investigate the potential role of Aβ in ALS development, we generated a double transgenic mouse line that overexpresses SOD1G93A and amyloid precursor protein (APP)-C100. The transgenic mouse C100.SOD1G93A overexpresses Aβ and shows earlier onset of motor impairment but has the same lifespan as the single transgenic SOD1G93A mouse. To determine the mechanism associated with this early-onset phenotype, we measured copper and zinc levels in brain and spinal cord and found both significantly elevated in the single and double transgenic mice compared with their littermate control mice. Increased glial fibrillary acidic protein and decreased APP levels in the spinal cord of C100.SOD1G93A mice compared with the SOD1G93A mice agree with the neuronal damage observed by immunohistochemical analysis. In the spinal cords of C100.SOD1G93A double transgenic mice, soluble Aβ was elevated in mice at end-stage disease compared with the pre-symptomatic stage. Buffer-insoluble SOD1 aggregates were significantly elevated in the pre-symptomatic mice of C100.SOD1G93A compared with the age-matched SOD1G93A mice, correlating with the earlier onset of motor impairment in the C100.SOD1G93A mice. This study supports abnormal SOD1 protein aggregation as the pathogenic mechanism in ALS, and implicates a potential role for Aβ in the development of ALS by exacerbating SOD1G93A aggregation.