Genes and the Environment in Neurodegeneration

Neurodegenerative diseases are a heterogeneous group of pathologies which includes complex multifactorial diseases, monogenic disorders and disorders for which inherited, sporadic and transmissible forms are known. Factors associated with predisposition and vulnerability to neurodegenerative disorders may be described usefully within the context of gene–environment interplay. There are many identified genetic determinants for neurodegeneration, and it is possible to duplicate many elements of recognized human neurodegenerative disorders in animal models of the disease. However, there are similarly several identifiable environmental influences on outcomes of the genetic defects; and the course of a progressive neurodegenerative disorder can be greatly modified by environmental elements. In this review we highlight some of the major neurodegenerative disorders (Alzheimer’s disease, Parkinson’s disease, Amyotrophic lateral sclerosis, Huntington’s disease, and prion diseases.) and discuss possible links of gene–environment interplay including, where implicated, mitochondrial genes.     

Generation of Reactive Oxygen Species by Mitochondrial Complex I: Implications in Neurodegeneration

Mitochondrial Complex I [NADH Coenzyme Q (CoQ) oxidoreductase] is the least understood of respiratory complexes. In this review we emphasize some novel findings on this enzyme that are of relevance to the pathogenesis of neurodegenerative diseases. Besides CoQ, also oxygen may be an electron acceptor from the enzyme, with generation of superoxide radical in the mitochondrial matrix. The site of superoxide generation is debated: we present evidence based on the rational use of several inhibitors that the one-electron donor to oxygen is an iron-sulphur cluster, presumably N2. On this assumption we present a novel mechanism of electron transfer to the acceptor, CoQ. Complex I is deeply involved in pathological changes, including neurodegeneration. Complex I changes are involved in common neurological diseases of the adult and old ages. Mitochondrial cytopathies due to mutations of either nuclear or mitochondrial DNA may represent a useful model of neurodegeneration. In this review we discuss Parkinson’s disease, where the pathogenic involvement of Complex I is better understood; the accumulated evidence on the mode of action of Complex I inhibitors and their effect on oxygen radical generation is discussed in terms of the aetiology and pathogenesis of the disease. Special issue article in honor of Dr. Anna Maria Giuffrida-Stella.     

The cerebral proteopathies

The abnormal assembly and deposition of specific proteins in the brain is the probable cause of most neurodegenerative disease afflicting the elderly. These “cerebral proteopathies” include Alzheimer’s disease (AD), Parkinson’s disease (PD), Huntington’s disease (HD), prion diseases, and a variety of other disorders. Evidence is accumulating that the anomalous aggregation of the proteins, and not a loss of protein function, is central to the pathogenesis of these diseases. Thus, therapeutic strategies that reduce the production, accumulation, or polymerization of pathogenic proteins might be applicable to a wide range of some of the most devastating diseases of old age.     

Brain-Specific Aminopeptidase: From Enkephalinase to Protector Against Neurodegeneration

The major breakthrough discovery of enkephalins as endogenous opiates led our attempts to determine their inactivation mechanisms. Because the NH2-terminal tyrosine is absolutely necessary for the neuropeptides to exert analgesic effects, and aminopeptidase activities are extraordinarily high in the brain, a specific “amino-enkephalinase” should exist. Several aminopeptidases were identified in the central nervous system during the search. In fact, our laboratory found two novel neuron-specific aminopeptidases: NAP and NAP-2. NAP is the only functionally active brain-specific enzyme known. Its synaptic location coupled with its limited substrate specificity could constitute a “functional” specificity and contribute to enkephalin-specific functions. In addition, NAP was found to be essential for neuron growth, differentiation, and death. Thus, aminopeptidases are likely important for mental health and neurological diseases. Recently, puromycin-sensitive aminopeptidase (PSA) was identified as a modifier of tau-induced neurodegeneration. Because the enzymatic similarity between PSA and NAP, we believe that the depletion of NAP in Alzheimer’s disease (AD) brains plays a causal role in the development of AD pathology. Therefore, use of the puromycin-sensitive neuron-aminopeptidase NAP could provide neuroprotective mechanisms in AD and similar neurodegenerative diseases. Special issue in honor of Naren Banik.     

Dopaminergic Agonists: Possible Neurorescue Drugs Endowed with Independent and Synergistic Multisites of Action

Dopaminergic agonists have been usually used as adjunctive therapy for the cure of Parkinson’s disease (PD). It is generally believed that treatment with these drugs is symptomatic rather then curative and does not stop or delay the progression of neuronal degeneration. However, several DA agonists of the DA D2–receptor family (including D2, D3 and D4-subtypes) have recently been shown to possess neuroprotective properties in different in vitro and in vivo experimental PD models. Here we summarize some recent data from our and other groups underlining the wide pharmacological spectrum of DA agonists currently used for treating PD patients. In particular, the mechanism of action of different DA agonists does not appear to be restricted to the stimulation of selective DA receptor subtypes being these drugs endowed with intrinsic, independent, and peculiar antioxidant effects. This activity may represent an additional pharmacological property contributing to their clinical efficacy in PD. Special issue dedicated to Dr. Moussa Youdim.     

Free Copper,Ferroxidase and SOD1 Activities,Lipid Peroxidation and NO<Subscript><Emphasis Type="Italic">x</Emphasis></Subscript> Content in the CSF. A Different Marker Profile in Four Neurodegenerative Diseases

The understanding of oxidative damage in different neurodegenerative diseases could enhance therapeutic strategies. Our objective was to quantify lipoperoxidation and other oxidative products as well as the activity of antioxidant enzymes and cofactors in cerebrospinal fluid (CSF) samples. We recorded data from all new patients with a diagnosis of either one of the four most frequent neurodegenerative diseases: Parkinson’s disease (PD), Alzheimer’s disease (AD), Huntington’s disease (HD) and lateral amyotrophic sclerosis (ALS). The sum of nitrites and nitrates as end products of nitric oxide (NO) were increased in the four degenerative diseases and fluorescent lipoperoxidation products in three (excepting ALS). A decreased Cu/Zn-dependent superoxide dismutase (SOD) activity characterized the four diseases. A significantly decreased ferroxidase activity was found in PD, HD and AD, agreeing with findings of iron deposition in these entities, while free copper was found to be increased in CSF and appeared to be a good biomarker of PD.     

Voltage-gated Calcium Channels Provide an Alternate Route for Iron Uptake in Neuronal Cell Cultures

Recent studies suggest that iron enters cardiomyocytes via the L-type voltage-gated calcium channel (VGCC). The neuronal VGCC may also provide iron entry. As with calcium, extraneous iron is associated with the pathology and progression of neurodegenerative diseases such as Parkinson’s and Alzheimer’s disease. VGCCs, ubiquitously expressed, may be an important route of excessive entry for both iron and calcium, contributing to cell toxicity or death. We evaluated the uptake of ⁴⁵Ca²⁺ and ⁵⁵Fe²⁺ into NGF-treated rat PC12, and murine N-2α cells. Iron not only competed with calcium for entry into these cells, but iron uptake (similar to calcium uptake) was inhibited by nimodipine, a specific L-type VGCC blocker, and enhanced by FPL 64176, an L-VGCC activator, in a dose-dependent manner. Taken together, these data suggest that voltage-gated calcium channels are an alternate route for iron entry into neuronal cells under conditions that promote cellular iron overload toxicity. Special issue dedicated to Dr. Moussa Youdim.     

Applications of electron paramagnetic resonance to studies of neurological disease

Electron paramagnetic resonance spectroscopy (EPR) has the potential to give much detail on the structure of the paramagnetic transition ion coordination sites, principally of Cu²⁺, in a number of proteins associated with central nervous system diseases. Since these sites have been implicated in misfolding/mis-oligomerisation events associated with neurotoxic molecular species and/or the catalysis of damaging redox reactions in neurodegeneration, an understanding of their structure is important to the development of therapeutic agents. Nevertheless EPR, by its nature an in vitro technique, has its limitations in the study of such complex biochemical systems involving self-associating proteins that are sensitive to their chemical environment. These limitations are at the instrumental and theoretical level, which must be understood and the EPR data interpreted in the light of other biophysical and biochemical studies if useful conclusions are to be drawn. Australian Society for Biophysics Special Issue: Metals and Membranes in Neuroscience, held in Melbourne on 11 July 2007.     

Amyotrophic lateral sclerosis--the tools of the trait

The aim of this review is to analyze how our knowledge on the etiology, pathology, and treatment of amyotrophic lateral sclerosis (ALS) has profited from the application of biotechnology tools for the identification of disease markers, the development of animal disease models, and the design of innovative therapeutics. In humans, ALS-specific clinical, genetic or protein biomarkers, or panels of biomarkers stemming from genomics and proteomics analyses can be critical for early diagnosis, monitoring of disease progression, drug validation in clinical trials, and identification of therapeutic targets for subsequent drug development. At the same time, animal models representing a number of human superoxide dismutase 1 mutations, intermediate-filament disorganization or axonal-transport defects have been invaluable in unraveling aspects of the pathophysiology of the disease; in each case, these only represent a small proportion of all ALS patients. Preclinical and clinical trials, although at present heavily concentrating on pharmacological approaches, are embracing the emerging alternative strategies of stem-cell and gene therapy. In combination with a further subcategorization of patients and the development of corresponding model systems for functional analyses, they will significantly influence the already changing face of ALS therapy.     

Quantitative Proteomic Analysis of Mitochondria in Aging PS-1 Transgenic Mice

Accumulating evidence suggests mitochondrial alterations are intimately associated with the pathogenesis of Alzheimer’s disease (AD). In order to determine if mutations of presenilin-1 (PS-1) affect levels of mitochondrial proteins at different ages we enriched mitochondrial fractions from 3-, 6-, 12-month-old knock-in mice expressing the M146V PS-1 mutation and identified, and quantified proteins using cleavable isotope-coded affinity tag labeling and two-dimensional liquid chromatography/tandem mass spectrometry (2D-LC/MS/MS). Using this approach, 165 non-redundant proteins were identified with 80 of them present in all three age groups. Specifically, at young ages (3 and 6 months), Na⁺/K⁺ ATPase and several signal transduction proteins exhibited elevated levels, but dropped dramatically at 12 months. In contrast, components of the oxidative phosporylation pathway (OXPHOS), the mitochondrial permeability transition pore (MPTP), and energy metabolism proteins remained unchanged at 3 months but significantly increased with age. We propose that alterations in calcium homeostasis induced by the PS-1 mutation have a major impact in young animals by inhibiting the function of relevant proteins and inducing compensatory changes. However, in older mice combination of the PS-1 mutation and accumulated oxidative damage results in a functional suppression of OXPHOS and MPTP proteins requiring a compensatory increase in expression levels. In contrast, signal transduction proteins showed decreased levels due to a break down in the compensatory mechanisms. The dysfunction of Na⁺/K⁺ ATPase and signal transduction proteins may induce impaired cognition and memory before neurodegeneration occurs. Electronic supplementary material  The online version of this article (doi:) contains supplementary material, which is available to authorized users.     

Osteoporosis in neurodegeneration

Osteoporosis affects bone microarchitecture and reduces bone mass. There are more than 200 million people with osteoporosis worldwide, and the prevalence is slowly increasing. The highest prevalences are found in Scandinavia and USA, also slowly increasing. A parallel increase in neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, and multiple sclerosis has been noted since the middle of this century. Osteoporosis is more common in patients with each of these neurodegenerative conditions than in the general population. Several metals with neurotoxic properties accumulate in bone and can substitute for calcium in hydroxyapatite, the main mineral component of bone. Especially cadmium, but also lead, aluminum and arsenic affect bone mineral density negatively. Metals with neurotoxic properties have also been found in brain and cerebrospinal fluid from patients with Alzheimer's disease, Parkinson's disease, Amyotrophic lateral sclerosis, and multiple sclerosis, and markers for neurodegeneration such as amyloid beta peptide and amyloid precursor protein have been detected in bone tissue from patients with osteoporosis. A common mechanism contributing to the pathogenesis of both neurodegeneration and osteoporosis can be suspected. The hypothesis that neurodegenerative disorders are associated with osteoporosis is presented and discussed.     

Neurodegeneration in mnd2 mutant mice is not prevented by parkin transgene

Parkinson’s disease (PD) is a common neurodegenerative disorder. The motor neuron degeneration 2 mutant (mnd2) mouse is considered to be an animal model of PD, and exhibits striatal neuron loss, severe muscle wasting, weight loss and death before 40 days of age. We found for the first time that parkin expression was decreased in the mnd2 mouse brain. Since parkin is a crucial protein for PD, the neurodegenerative disorder in mnd2 mice may be caused by parkin protein loss. We therefore examined whether compensation of parkin protein prevents neurodegenerative disorders in mnd2 mice by generating parkin-transgenic (parkin-Tg) mnd2 mice. However, both parkin-Tg mnd2 mice and mnd2 mice were smaller than wild type mice. In muscle strength and survival rate, parkin-Tg mnd2 mice showed similar values to mnd2 mice. Our data suggest that repression of parkin protein does not play a major role in neurodegeneration of mnd2 mice and administration of parkin protein does not rescue mnd2 mice.     

BLOC1S2 interacts with the HIPPI protein and sensitizes NCH89 glioblastoma cells to apoptosis

The HIPPI (HIP-1 protein interactor) protein is a multifunctional protein that is involved in the regulation of apoptosis. The interaction partners of HIPPI include HIP-1 (Huntingtin-interacting protein-1), Apoptin, Homer1c, Rybp/DEDAF, and BAR (bifunctional apoptosis regulator). In search for other binding partners of HIPPI, we performed a yeast two hybrid screen and identified BLOC1S2 (Biogenesis of lysosome-related organelles complex-1 subunit 2) as a novel HIPPI-interacting protein. In co-immunoprecipitation assays, BLOC1S2 specifically associates with HIPPI, but not with HIP-1. To study the expression of BLOC1S2 on the protein level, we generated a mouse monoclonal antibody specific for BLOC1S2 and a multiple tissue array comprising 70 normal and cancer tissue samples of diverse origin. BLOC1S2 protein is widely expressed in normal tissue as well as in malignant tumors with a tendency towards lower expression levels in certain subtypes of tumors. On the subcellular level, BLOC1S2 is expressed in an organellar-like pattern and co-localizes with mitochondria. Over-expression of BLOC1S2 in the presence or absence of HIPPI does not induce apoptosis. However, BLOC1S2 and HIPPI sensitize NCH89 glioblastoma cells to the pro-apoptotic actions of staurosporine and the death ligand TRAIL by enhancing caspase activation, cytochrome c release, and disruption of the mitochondrial membrane potential. Given its interaction with HIPPI and its pro-apoptotic activity, BLOC1S2 might play an important functional role in cancer and neurodegenerative diseases.     

A comparison of the neuronal dysfunction caused by <Emphasis Type="Italic">Drosophila</Emphasis> tau and human tau in a <Emphasis Type="Italic">Drosophila</Emphasis> model of tauopathies

Hyperphosphorylation and aggregation of tau into tangles is a feature of disorders such as Alzheimer’s disease and other Tauopathies. To model these disorders in Drosophila melanogaster, human tau has been over-expressed and a variety of phenotypes have been observed including neurotoxicity, disrupted neuronal and synaptic function and locomotor impairments. Neuronal dysfunction has been seen prior to neuronal death and in the absence of tangle formation. The Drosophila tau protein shares a large degree of homology with human tau but differs in the crucial microtubule binding domains. Although like human tau Drosophila tau can induce neurotoxicity, little is known about its ability to disrupt neuronal function. In this study we demonstrate that like human tau, over-expression of Drosophila tau results in disrupted axonal transport, altered neuromuscular junction morphology and locomotor impairments. This indicates that like human tau, over-expression of Drosophila tau compromises neuronal function despite significant differences in microtubule binding regions.     

Plasma amino acids concentration in amyotrophic lateral sclerosis patients

Summary.  Previous investigations showed an impairment of amino acids (AA) metabolism in amyotrophic lateral sclerosis (ALS). It was hypothesized that excitatory AA may play an important role in the etiopathogenesis of this disease. The aim of the study was to determine plasma AA concentrations in ALS patients, and to examine the relationship between AA and the clinical state of ALS patients, the type of ALS onset and the duration of the disease. The study involved 20 ALS patients and 30 control group people. The AA analysis was performed by ion – exchange chromatography on an automatic AA analyser. The results showed significantly decreased concentrations of valine, isoleucine, leucine, tyrosine and aspartate in the plasma of the whole group of ALS patients compared to the control group, and a significantly decreased concentration of arginine in the patients with a long duration of ALS compared to the patients with a short duration. The clinical state of ALS patients significantly influenced only plasma alanine concentration. Other plasma AA concentrations were not significantly associated with clinical parameters of the disease. Our study confirms that metabolic abnormalities concerning AA exist in ALS patients. However, the normal plasma glutamate concentration observed in this study in the whole group of ALS patients compared to the controls does not exclude that this excitatory AA may play a role in neurodegeneration in ALS. Received June 22, 2002 Accepted October 3, 2002 Published online January 23, 2003 Correspondence: Joanna Iłżecka M.D., Department of Neurology, Medical University, Jaczewskiego 8, 20-954 Lublin, Poland, Fax: +48 81 742 55 34, E-mail: Ilzecka@medscape.com     

Key players in neurodegenerative disorders in focus—New insights into the proteomic profile of Alzheimer's disease,schizophrenia, ALS,and multiple sclerosis—24th HUPO BPP Workshop

The HUPO Brain Proteome Project (HUPO BPP) held its 24th workshop in Vancouver, Canada, September 29, 2015. The focus of the autumn workshop was on new insights into the proteomic profile of Alzheimer's disease, schizophrenia, ALS and multiple sclerosis.     

Mitochondrial Dysfunction in Neurodegenerative Diseases

Mitochondria play a pivotal role in mammalian cell metabolism, hosting a number of important biochemical pathways including oxidative phosphorylation. As might be expected from this fundamental contribution to cell function, abnormalities of mitochondrial metabolism are a common cause of human disease. Primary mutations of mitochondrial DNA result in a diverse group of disorders often collectively referred to as the mitochondrial encephalomyopathies. Perhaps more importantly in numerical terms are those neurodegenerative diseases caused by mutations of nuclear genes encoding mitochondrial proteins. Finally there are mitochondrial abnormalities induced by secondary events e.g. oxidative stress that may contribute to senescence, and environmental toxins that may cause disease either alone or in combination with a genetic predisposition. Special issue article in honor of Dr. Anna Maria Giuffrida-Stella.     

Applications of neutron activation analysis to the study of age-related neurological diseases

Although the etiology and pathogenesis of Alzheimer’s disease, Pick’s disease, and amyotrophic lateral sclerosis are still unknown, it has been suggested that perturbations in element metabolism may play a role. Even if not causative factors, these imbalances may prove to be markers that could aid in diagnosis. We have employed a sequential neutron activation analysis (NAA) procedure to determine elemental concentrations in brain, hair, fingernails, blood, and cerebrospinal fluid (CSF) of these patients and age-matched controls. Samples are first irradiated with accelerator-produced 14-MeV neutrons for determination of nitrogen and phosphorus, then with reactor thermal neutrons for the instrumental determination of 16–18 minor and trace elements, and, finally, reactor-irradiated again, followed by a rapid radiochemical separation procedure (RNAA) to determine four additional elements. Major advantages of NAA are: (1) its simultaneous multielement capability; (2) the relative freedom from reagent and laboratory contamination; (3) the absence of major matrix effects; and (4) an adequate sensitivity for most elements of interest. Ranges of concentrations by INAA and RNAA in selected control tissues and interelement correlations in control brain are presented to illustrate results obtained by the procedure. Longitudinal studies of tissues from Alzheimer’s disease (AD) and amyotrophic lateral sclerosis (ALS) patients are still in progress.     

Patients with Amyotrophic Lateral Sclerosis and Other Neurodegenerative Diseases Have Increased Levels of Neurofilament Protein in CSF

Abstract: In the present study we describe an ELISA to quantify the light subunit of the neurofilament triplet protein (NFL) in CSF. The method was validated by measuring CSF NFL concentrations in healthy individuals and in two well-characterized groups of patients with amyotrophic lateral sclerosis (ALS) and Alzheimer's disease (AD). The levels were increased in ALS (1,743 ± 1,661 ng/L; mean ± SD) and AD (346 ± 176 ng/L) compared with controls (138 ± 31 ng/L; p < 0.0001 for both). Within the ALS group, patients with lower motor neuron signs only had lower NFL levels (360 ± 237 ng/L) than those with signs of upper motor neuron disease (2,435 ± 1,633 ng/L) ( p < 0.05). In a second study patients with miscellaneous neurodegenerative diseases were investigated (vascular dementia, olivopontocerebellar atrophy, normal pressure hydrocephalus, cerebral infarctions, and multiple sclerosis), and the CSF NFL level was found to be increased (665 ± 385 ng/L; p < 0.0001). NFL is a main structural protein of axons, and we suggest that CSF NFL can be used to monitor neurodegeneration in general, but particularly in ALS with involvement of the pyramidal tract.