metallothionein and Parkinson-Disease

Disbalance of zinc (Zn2+) and copper (Cu2+) ions in the central nervous system is involved in the pathogenesis of numerous neurodegenerative disorders such as multisystem atrophy, amyotrophic lateral sclerosis, Creutzfeldt-Jakob disease, Wilson-Konovalov disease, Alzheimer's disease, and Parkinson's disease. Among these, Alzheimer's disease (AD) and Parkinson's disease (PD) are the most frequent age-related neurodegenerative pathologies with disorders in Zn2+ and Cu2+ homeostasis playing a pivotal role in the mechanisms of pathogenesis. In this review we generalized and systematized current literature data concerning this problem. The interactions of Zn2+ and Cu2+ with amyloid precursor protein (APP), β-amyloid (Abeta), tau-protein, metallothioneins, and GSK3β are considered, as well as the role of these interactions in the generation of free radicals in AD and PD. Analysis of the literature suggests that the main factors of AD and PD pathogenesis (oxidative stress, structural disorders and aggregation of proteins, mitochondrial dysfunction, energy deficiency) that initiate a cascade of events resulting finally in the dysfunction of neuronal networks are mediated by the disbalance of Zn2+ and Cu2+.

Topics: Alzheimer Disease; Amyloid beta-Peptides; Amyloid beta-Protein Precursor; Copper; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Metallothionein; Oxidative Stress; Parkinson Disease; Reactive Oxygen Species; tau Proteins; Zinc

Clinically, Parkinson's disease (PD) is a neurodegenerative disorder characterized by the development of tremors and rigidity that is found primarily in patients over the age of 50. At the cellular level, it is clear that the pathology of PD results from the progressive loss of dopaminergic neurons in the substantia nigra. Several lines of evidence have implicated oxidative stress as a contributing factor to the depletion of dopaminergic neurons in PD. Under conditions of oxidative stress, the neurotransmitter dopamine can be oxidized to form neurotoxic quinone and semiquinone products. While dopaquinones are known to be extremely reactive towards sulfhydryl groups of many cellular substrates, mounting evidence suggests that their toxic effects can be quenched by intrinsic antioxidant mechanisms (e.g., glutathione). However, to respond appropriately to differing levels of oxidative stress, cells require a mechanism to regulate an appropriate response. This manuscript proposes metallothionein as a major cellular sensor of oxidized dopamine stress and metallothionein-mediated Zn2+ mobilization as an effecter signal that is used by the cell to manage oxidized dopamine as an intrinsic neurotoxin.

Topics: Amino Acid Sequence; Animals; Antioxidants; Dopamine; Humans; Metallothionein; Molecular Sequence Data; Oxidative Stress; Parkinson Disease; Protein Structure, Tertiary; Signal Transduction

Parkinson's disease is characterized by a progressive loss of dopaminergic neurons in the substantia nigra zona compacta, and in other sub-cortical nuclei associated with a widespread occurrence of Lewy bodies. The cause of cell death in Parkinson's disease is still poorly understood, but a defect in mitochondrial oxidative phosphorylation and enhanced oxidative and nitrative stresses have been proposed. We have studied control(wt) (C57B1/6), metallothionein transgenic (MTtrans), metallothionein double gene knock (MTdko), alpha-synuclein knock out (alpha-syn(ko)), alpha-synuclein-metallothionein triple knock out (alpha-syn-MTtko), weaver mutant (wv/wv) mice, and Ames dwarf mice to examine the role of peroxynitrite in the etiopathogenesis of Parkinson's disease and aging. Although MTdko mice were genetically susceptible to 1, methyl, 4-phenyl, 1,2,3,6-tetrahydropyridine (MPTP) Parkinsonism, they did not exhibit any overt clinical symptoms of neurodegeneration and gross neuropathological changes as observed in wv/wv mice. Progressive neurodegenerative changes were associated with typical Parkinsonism in wv/wv mice. Neurodegenerative changes in wv/wv mice were observed primarily in the striatum, hippocampus and cerebellum. Various hallmarks of apoptosis including caspase-3, TNFalpha, NFkappaB, metallothioneins (MT-1, 2) and complex-1 nitration were increased; whereas glutathione, complex-1, ATP, and Ser(40)-phosphorylation of tyrosine hydroxylase, and striatal 18F-DOPA uptake were reduced in wv/wv mice as compared to other experimental genotypes. Striatal neurons of wv/wv mice exhibited age-dependent increase in dense cored intra-neuronal inclusions, cellular aggregation, proto-oncogenes (c-fos, c-jun, caspase-3, and GAPDH) induction, inter-nucleosomal DNA fragmentation, and neuro-apoptosis. MTtrans and alpha-Syn(ko) mice were genetically resistant to MPTP-Parkinsonism and Ames dwarf mice possessed significantly higher concentrations of striatal coenzyme Q10 and metallothioneins (MT 1, 2) and lived almost 2.5 times longer as compared to control(wt) mice. A potent peroxynitrite ion generator, 3-morpholinosydnonimine (SIN-1)-induced apoptosis was significantly attenuated in MTtrans fetal stem cells. These data are interpreted to suggest that peroxynitrite ions are involved in the etiopathogenesis of Parkinson's disease, and metallothionein-mediated coenzyme Q10 synthesis may provide neuroprotection.

Topics: alpha-Synuclein; Animals; Apoptosis; Brain; Coenzymes; Disease Models, Animal; Dopamine; Gene Expression Regulation; Humans; Metallothionein; Mice; Mice, Neurologic Mutants; Mice, Transgenic; MPTP Poisoning; Nerve Tissue Proteins; Parkinson Disease; Synucleins; Ubiquinone

Topics: Animals; Brain; Humans; Metallothionein; Mitochondria; Neurodegenerative Diseases; Neuroprotective Agents; Oxidative Phosphorylation; Parkinson Disease; Ubiquinone

Topics: Animals; Humans; Male; Metallothionein; Oxidation-Reduction; Parkinson Disease

Parkinson's disease (PD) is the second most common neurodegenerative disease caused by selective degeneration of dopaminergic neurons in the substantia nigra. Metallothionein has been shown to act as a neuroprotectant in various brain injury. Thus, this study aims to identify the effects of full-length human metallothionein 2 peptide (hMT2) in paraquat-induced brain injury in the zebrafish.. A total of 80 adult zebrafish were divided into 4 groups namely control, paraquat-treated, pre-hMT2-treated, and post-hMT2-treated groups. Fish were treated with paraquat intraperitoneally every 3 days for 15 days. hMT2 were injected intracranially on day 0 (pre-treated group) and day 16 (post-treated group). Fish were sacrificed on day 22 and the brains were collected for qPCR, ELISA and immunohistochemistry analysis.. qPCR analysis showed that paraquat treatment down-regulated the expression of genes related to dopamine activity and biosynthesis (. Paraquat has been identified as one of the pesticides that can cause the death of dopaminergic neurons and affect dopamine biosynthesis. Treatment with exogenous hMT2 could reverse the effects of paraquat in the zebrafish brain.

Topics: Animals; Brain Injuries; Dopamine; Dopaminergic Neurons; Humans; Metallothionein; Mice; Mice, Inbred C57BL; Neurodegenerative Diseases; Neuroprotective Agents; Paraquat; Parkinson Disease; Substantia Nigra; Zebrafish

Neurodegenerative disorders (ND) like Alzheimer's (AD), Parkinson's (PD), Huntington's or Prion diseases share similar pathological features. They are all age dependent and are often associated with disruptions in analogous metabolic processes such as protein aggregation and oxidative stress, both of which involve metal ions like copper, manganese and iron. Bush and Tanzi proposed 2008 in the 'metal hypothesis of Alzheimer's disease' that a breakdown in metal homeostasis is the main cause of NDs, and drugs restoring metal homeostasis are promising novel therapeutic strategies. We report here that metallothionein (MT), an endogenous metal detoxifying protein, is increased in young amyloid ß (Aß) expressing Caenorhabditis elegans, whereas it is not in wild type strains. Further MT induction collapsed in 8 days old transgenic worms, indicating the age dependency of disease outbreak, and sharing intriguing parallels to diminished MT levels in human brains of AD. A medium throughput screening assay method was established to search for compounds increasing the MT level. Compounds known to induce MT release like progesterone, ZnSO

Topics: Aging; alpha-Synuclein; Alzheimer Disease; Amyloid beta-Peptides; Animals; Animals, Genetically Modified; Benzothiazoles; Caenorhabditis elegans; Caenorhabditis elegans Proteins; Clioquinol; Disease Models, Animal; Emodin; Gene Knockdown Techniques; Homeostasis; Metallothionein; Metals; Neuroprotective Agents; Parkinson Disease; Quercetin; Signal Transduction

Mirtazapine, a noradrenergic and specific serotonergic antidepressant (NaSSA), is known to activate serotonin (5-HT) 1A receptor. Our recent study demonstrated that stimulation of astrocytic 5-HT1A receptors promoted astrocyte proliferation and upregulated antioxidative property in astrocytes to protect dopaminergic neurons against oxidative stress. Here, we evaluated the neuroprotective effects of mirtazapine against dopaminergic neurodegeneration in models of Parkinson's disease (PD). Mirtazapine administration attenuated the loss of dopaminergic neurons in the substantia nigra and increased the expression of the antioxidative molecule metallothionein (MT) in the striatal astrocytes of 6-hydroxydopamine (6-OHDA)-injected parkinsonian mice via 5-HT1A receptors. Mirtazapine protected dopaminergic neurons against 6-OHDA-induced neurotoxicity in mesencephalic neuron and striatal astrocyte cocultures, but not in enriched neuronal cultures. Mirtazapine-treated neuron-conditioned medium (Mir-NCM) induced astrocyte proliferation and upregulated MT expression via 5-HT1A receptors on astrocytes. Furthermore, treatment with medium from Mir-NCM-treated astrocytes protected dopaminergic neurons against 6-OHDA neurotoxicity, and these effects were attenuated by treatment with a MT-1/2-specific antibody or 5-HT1A antagonist. Our study suggests that mirtazapine could be an effective disease-modifying drug for PD and highlights that astrocytic 5-HT1A receptors may be a novel target for the treatment of PD.

Topics: Animals; Antioxidants; Astrocytes; Cells, Cultured; Dopamine; Dopaminergic Neurons; Female; Male; Metallothionein; Mice; Mice, Inbred ICR; Mirtazapine; Neuroprotection; Neuroprotective Agents; Oxidative Stress; Oxidopamine; Parkinson Disease; Pregnancy; Rats, Sprague-Dawley; Receptor, Serotonin, 5-HT1A; Substantia Nigra

Intracellular aggregates of α-synuclein are the pathological hallmark of Parkinson's disease (PD) and dementia with Lewy bodies (DLB), being linked to neurotoxicity. Multiple triggers of α-synuclein aggregation have been implicated, including raised copper. The potential protective role of the endogenous copper-/zinc-binding proteins, metallothioneins (MT), has been explored in relation to copper-induced α-synuclein aggregation. Up-regulated endogenous expression of MT was induced in SHSY-5Y cells by the synthetic glucocorticoid analogue, dexamethasone. After treatment to induce endogenous MT expression, immunofluorescence confocal microscopy was used to quantify protein aggregates in cells with/without copper treatment. MT induction resulted in significant (p < 0.01), dose-dependent up-regulation of MT expression and significant reduction in Cu-dependent α-synuclein intracellular aggregates (p < 0.01) that could be suppressed by MT-specific siRNA. Ubiquitous (MT-2) and brain-specific (MT-3) isoforms were investigated by transient transfection of the GFP-fusion proteins, observing equivalent α-synuclein aggregate suppression by each. These studies indicate MT induction could have potential in PD/DLB neuroprotective therapy by suppressing α-synuclein aggregation.

Topics: alpha-Synuclein; Brain; Cell Line, Tumor; Copper; Dexamethasone; Humans; Lewy Body Disease; Metallothionein; Parkinson Disease

An excess of reactive oxygen species (ROS) relative to the antioxidant capacity causes oxidative stress, which plays a role in the development of Parkinson's disease (PD). Because mitochondria are both sites of ROS generation and targets of ROS damage, the delivery of antioxidants to mitochondria might prevent or alleviate PD. To transduce the antioxidant protein human metallothionein 1A (hMT1A) into mitochondria, we computationally designed a cell-penetrating artificial mitochondria-targeting peptide (CAMP). The recombinant CAMP-conjugated hMT1A fusion protein (CAMP-hMT1A) successfully localized to the mitochondria. Treating a cell culture model of PD with CAMP-hMT1A restored tyrosine hydroxylase expression and mitochondrial activity and reduced ROS production. Furthermore, injection of CAMP-hMT1A into the brain of a mouse model of PD rescued movement impairment and dopaminergic neuronal degeneration. CAMP-hMT1A delivery into mitochondria might be therapeutic against PD by alleviating mitochondrial damage, and we predict that CAMP could be used to deliver other cargo proteins to the mitochondria.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Amino Acid Sequence; Animals; Cell Death; Cell Line, Tumor; Cell-Penetrating Peptides; Computer Simulation; Disease Models, Animal; Green Fluorescent Proteins; Humans; Metallothionein; Mice; Mitochondria; Neurons; Parkinson Disease; Protein Transport; Recombinant Fusion Proteins; Substantia Nigra; Tyrosine 3-Monooxygenase

Disordered gambling is a moderately heritable trait, but the underlying genetic basis is largely unknown. We performed a genome-wide association study (GWAS) for disordered gambling using a quantitative factor score in 1312 twins from 894 Australian families. Association was conducted for 2 381 914 single-nucleotide polymorphisms (SNPs) using the family-based association test in Merlin followed by gene and pathway enrichment analyses. Although no SNP reached genome-wide significance, six achieved P-values < 1 × 10(-5) with variants in three genes (MT1X, ATXN1 and VLDLR) implicated in disordered gambling. Secondary case-control analyses found two SNPs on chromosome 9 (rs1106076 and rs12305135 near VLDLR) and rs10812227 near FZD10 on chromosome 12 to be significantly associated with lifetime Diagnostic and Statistical Manual of Mental Disorders, fourth edition pathological gambling and South Oaks Gambling Screen classified probable pathological gambling status. Furthermore, several addiction-related pathways were enriched for SNPs associated with disordered gambling. Finally, gene-based analysis of 24 candidate genes for dopamine agonist-induced gambling in individuals with Parkinson's disease suggested an enrichment of SNPs associated with disordered gambling. We report the first GWAS of disordered gambling. While further replication is required, the identification of susceptibility loci and biological pathways will be important in characterizing the biological mechanisms that underpin disordered gambling.

Topics: Adult; Ataxin-1; Ataxins; Case-Control Studies; Chromosomes, Human, Pair 12; Chromosomes, Human, Pair 9; Dopamine Agonists; Female; Gambling; Genetic Markers; Genome-Wide Association Study; Genotype; Humans; Male; Metallothionein; Nerve Tissue Proteins; Nuclear Proteins; Parkinson Disease; Polymorphism, Single Nucleotide; Receptors, LDL

The aggregation of α-synuclein (α-Syn), the major component of intracellular Lewy body inclusions in dopaminergic neurons of the substantia nigra, plays a critical role in the etiology of Parkinson disease (PD). Long-term effects of redox-active transition metals (Cu, Fe) and oxidative chemical imbalance underlie the disease progression and neuronal death. In this work, we provide evidence that a brain metalloprotein, Zn₇-metallothionein-3 (Zn₇MT-3), possesses a dynamic role in controlling aberrant protein-copper interactions in PD. We examined the properties of the α-Syn-Cu(II) complex with regard to molecular oxygen, the biological reducing agent ascorbate, and the neurotransmitter dopamine. The results revealed that under aerobic conditions α-Syn-Cu(II) possesses catalytic oxidase activity. The observed metal-centered redox chemistry significantly promotes the production of hydroxyl radicals and α-Syn oxidation and oligomerization, processes considered critical for cellular toxicity. Moreover, we show that Zn₇MT-3, through Cu(II) removal from the α-Syn-Cu(II) complex, efficiently prevents its deleterious redox activity. We demonstrate that the Cu(II) reduction by thiolate ligands of Zn₇MT-3 and the formation of Cu(I)₄Zn₄MT-3, in which an unusual oxygen-stable Cu(I)₄-thiolate cluster is present, comprise the underlying molecular mechanism by which α-Syn and dopamine oxidation, α-Syn oligomerization, and ROS production are abolished. These studies provide new insights into the bioinorganic chemistry of PD.

Topics: alpha-Synuclein; Animals; Cells, Cultured; Copper; Disease Models, Animal; Disease Progression; Dopamine; Homeostasis; Humans; Male; Metallothionein; Neurons; Oxidation-Reduction; Parkinson Disease; Rats; Substantia Nigra

The role of glial cells in Parkinson's disease (PD) is unclear. We have previously reported a striking up-regulation of DnaJB6 heat shock protein in PD substantia nigra astrocytes. Whole genome transcriptome analysis also indicated increased expression of metallothionein genes in substantia nigra and cortex of sporadic PD cases. Metallothioneins are metal-binding proteins in the CNS that are released by astrocytes and associated with neuroprotection. Metallothionein expression was investigated in 18 PD cases and 15 non-PD controls using quantitative real-time polymerase chain reaction (qRT-PCR), in situ hybridisation (ISH) and immunocytochemistry (ICC). We observed a strong increase in the expression of metallothioneins MT1E, MT1F, MT1G, MT1H, MT1M, MT1X and MT2A in both PD nigra and frontal cortex. Expression of LRP2 (megalin), the neuronal metallothionein receptor was also significantly increased. qRT-PCR confirmed metallothionein up-regulation. Astrocytes were found to be the main source of metallothioneins 1 and 2 based on ISH results, and this finding was confirmed by ICC. Our findings demonstrate metallothionein expression by reactive astrocytes in PD nigra and support a neuroprotective role for these cells. The traditional view that nigral astrocytes are non-reactive in PD is clearly incorrect. However, it is possible that astrocytes are themselves affected by the disease process which may explain their comparatively modest and previously overlooked response.

Topics: Aged; Aged, 80 and over; Amino Acid Sequence; Astrocytes; Cluster Analysis; Female; Humans; Male; Metallothionein; Microarray Analysis; Molecular Sequence Data; Parkinson Disease; Protein Isoforms; Sequence Alignment; Up-Regulation

The concentration of plasma copper, ceruloplasmin (CRP), non-ceruloplasmin-bound Cu (NCBC), and metallothioneins (MTs) were studied as putative biomarkers for neurodegenerative diseases in patients and in their first-degree relatives. We found increased levels of Cu in the plasma of Alzheimer's disease (AD), Parkinson's disease (PD), and vascular dementia (VD) patients, and the increase observed in VD group was linked to the evolution of the disease. CRP was also elevated in response to the inflammatory component of the diseases, however, a correlation with illness progression was only observed in VD patients. The level of MTs is proportional to the evolution of VD. The Cu/CRP and Cu/MTs ratios are both indicative of disease progression for AD patients but not for those with PD or VD. Moreover, there is a correlation between the NCBC levels and the cognitive impairment estimated through the Mini-mental State Examination (MMSE) scale. This dependence is linear for AD and PD patients and non-linear for the VD ones. The relative values of NCBC showed dependence on the disease duration, especially for AD. Copper measurement and the Cu/CRP ratio may be predictive markers of risk for the first-degree relatives of AD patients. We believe that these results are valuable as a reliable clinical tool.

Topics: Adult; Aged; Alzheimer Disease; Biomarkers; Ceruloplasmin; Copper; Dementia, Vascular; Disease Progression; Family; Female; Humans; Linear Models; Male; Metallothionein; Middle Aged; Parkinson Disease; Risk Factors; Time Factors

Dopamine agonists have neuroprotective properties in addition to their original pharmacologic function. We examined the effects of pergolide mesilate (PM) on the levels of metallothionein mRNA expression and lipid peroxidation in the corpus striata of 1-methyl 4-phenyl 1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinsonian mice. Mice were administered normal saline (vehicle as a control), PM, or MPTP. A consecutive 7-d administration of MPTP via a gastric tube at a dose of 30 mg/kg significantly decreased metallothionein (MT)-I mRNA expression but did not influence MT-III mRNA expression. Lipid peroxidation, measured as the production of malondialdehyde reactive substances, did not increase after MPTP treatment. Although PM administration alone did not effect MT-I expression, an additional consecutive 7-d administration of PM (30 mug/kg) following MPTP treatment recovered the decreased MT-I level and increased MT-III expression. Lipid peroxidation was significantly suppressed. These results suggest that PM exerts an antioxidative property through the induction of MT-I and MT-III mRNAs simultaneously in response to cellular and/or tissue injury.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Antioxidants; Brain; Disease Models, Animal; Dopamine Agonists; Gene Expression; Lipid Peroxidation; Malondialdehyde; Metallothionein; Mice; Mice, Inbred C57BL; Parkinson Disease; Pergolide; RNA, Messenger

The exact molecular mechanism of progressive loss of neuromelanin containing nigrostriatal dopaminergic neurons in Parkinson's disease (PD) remains unknown, yet evidence suggests that iron might play an important role in PD pathology. In this study we have determined the neuroprotective role of coenzyme Q(10) (CoQ(10)) in ironinduced apoptosis in cultured human dopaminergic (SK-N-SH) neurons, in metallothionein gene- manipulated mice, and in alpha-synuclein knockout (alpha-synko) mice with a primary objective to assess a possible therapeutic and anti-inflammatory potential for CoQ(10) in PD. Iron-induced mitochondrial damage and apoptosis were characterized by reactive oxygen species production, increased metallothionein and glutathione synthesis, caspase- 3 activation, NF-kappaB induction, and decreased Bcl-2 expression, without any significant change in Bax expression. Lower concentrations of FeSO4 (1-10 microM) induced perinuclear aggregation of mitochondria, whereas higher concentrations (100-250 microM) induced CoQ(10) depletion, plasma membrane perforations, mitochondrial damage, and nuclear DNA condensation and fragmentation. FeSO(4)-induced deleterious changes were attenuated by pretreatment with CoQ(10) and by deferoxamine, a potent iron chelator, in SK-N-SH cells. 1-Methyl, 4-phenyl, 1,2,3,6- tetrahydropyridine (MPTP)-induced striatal release of free iron, and NF-kappaB expression were significantly increased; whereas ferritin and melanin synthesis were significantly reduced in the substantia nigra pars compacta (SNpc) of MT(dko) mice as compared with control(wt) mice, MT(trans) mice, and alpha-synko mice. CoQ(10) treatment inhibited MPTP-induced NF-kappaB induction in all of the genotypes. These data suggest that glutathione and metallothionein synthesis might be induced as an attempt to combat iron-induced oxidative stress, whereas exogenous administration of CoQ(10) or of metallothionein induction might provide CoQ(10)-mediated neuroprotection in PD.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; Animals; Apoptosis; Cell Line; Coenzymes; Deferoxamine; Dopamine; Dopamine Agents; Glutathione; Humans; Iron; Lipid Peroxidation; Melanins; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Knockout; Mice, Transgenic; Neurons; Neuroprotective Agents; NF-kappa B; Parkinson Disease; Reactive Oxygen Species; Siderophores; Ubiquinone; Vitamins

Regional distribution of coenzyme Q10 and mitochondrial complex-1 activity were estimated in the brains of control-(C57BL/6), metallothionein knock out-, metallothionein transgenic-, and homozygous weaver mutant mice; and human dopaminergic (SK-N-SH) cells with a primary objective to determine the neuroprotective potential of coenzyme Q10 in Parkinson's disease. Complex-1 activity as well as coenzyme Q10 were significantly higher in the cerebral cortex as compared to the striatum in all the genotypes examined. Complex-1 activity and coenzyme Q10 were significantly reduced in weaver mutant mice and metallothionein knock out mice, but were significantly increased in metallothionein transgenic mice. The reduced complex-1 activity and 18F-DOPA uptake occurred concomitantly with negligible differences in the coenzyme Q10 between in the cerebral cortex and striatum of weaver mutant mice. Administration of coenzyme Q10 increased complex-1 activity and partially improved motoric performance in weaver mutant mice. Direct exposure of rotenone also reduced coenzyme Q10, complex-1 activity, and mitochondrial membrane potential in SK-N-SH cells. Rotenone-induced down-regulation of complex-1 activity was attenuated by coenzyme Q10 treatment, suggesting that complex-1 may be down regulated due to depletion of coenzyme Q10 in the brain. Therefore, metallothionein-induced coenzyme Q10 synthesis may provide neuroprotection by augmenting mitochondrial complex-1 activity in Parkinson's disease.

Topics: Analysis of Variance; Animals; Brain; Cell Line, Tumor; Chromatography, High Pressure Liquid; Coenzymes; Dihydroxyphenylalanine; Disease Models, Animal; Electron Transport Complex I; Fluorine Radioisotopes; Humans; Male; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Neurologic Mutants; Mice, Transgenic; Neuroblastoma; Neuroprotective Agents; Parkinson Disease; Positron-Emission Tomography; Statistics as Topic; Tissue Distribution; Ubiquinone

We have examined potent peroxynitrite ion (ONOO-) generator 3-morpholinosydnonimine (SIN-1)-induced neurotoxicity in control wild-type (control(wt)) mice, metallothionein double knockout (MT(dko)) mice, metallothionein-transgenic (MT(trans)) mice, and in cultured human dopaminergic (SK-N-SH) neurons to determine the neuroprotective potential of metallothionein against ONOO(-)-induced neurodegeneration in Parkinson disease (PD). SIN-1-induced lipid peroxidation, reactive oxygen species synthesis, caspase-3 activation, and apoptosis were attenuated by metallothionein gene overexpression and augmented by metallothionein gene down-regulation. A progressive nigrostriatal dopaminergic neurodegeneration in weaver mutant (wv/wv) mice was associated with enhanced nitrite ion synthesis, metallothionein down-regulation, and significantly reduced dopamine synthesis and 18F-DOPA uptake as determined by high-resolution micropositron emission tomography neuroimaging. The striatal (18)F-DOPA uptake was significantly higher in MT(trans) mice than in MT(dko) and alpha-synuclein knockout (alpha-Syn(ko)) mice. These observations provide further evidence that nitric oxide synthase activation and ONOO- synthesis may be involved in the etiopathogenesis of PD, and that metallothionein gene induction may provide neuroprotection.

Topics: Animals; Apoptosis; Caspase 3; Caspases; Coenzymes; Dopamine; Enzyme Activation; Lipid Peroxidation; Metallothionein; Mice; Mice, Inbred C57BL; Mice, Transgenic; Molsidomine; Oxidative Stress; Parkinson Disease; Reactive Oxygen Species; Ubiquinone

Parkinson's disease (PD) is characterized by a progressive loss of dopaminergic neurons in the substantia nigra zona compacta and in other subcortical nuclei associated with a widespread occurrence of Lewy bodies. The causes of cell death in Parkinson's disease are still poorly understood, but a defect in mitochondrial oxidative phosphorylation and enhanced oxidative stress has been proposed. We have examined 3-morpholinosydnonimine (SIN-1)-induced apoptosis in control and metallothionein-overexpressing dopaminergic neurons, with a primary objective to determine the neuroprotective potential of metallothionein (MT) against peroxynitrite-induced neurodegeneration in PD. SIN-1 induced lipid peroxidation and triggered plasma membrane blebbing. In addition, it caused DNA fragmentation, alpha-synuclein induction, and intramitochondrial accumulation of metal ions (copper, iron, zinc, and calcium), and it enhanced the synthesis of 8-hydroxy-2-deoxyguanosine. Furthermore, it downregulated the expression of Bcl-2 and poly(adenosine diphosphate-ribose) polymerase, but upregulated the expression of caspase-3 and Bax in dopaminergic (SK-N-SH) neurons. SIN-1 induced apoptosis in aging mitochondrial genome knockout cells, alpha-synuclein-transfected cells, metallothionein double-knockout cells, and caspase-3-overexpressed dopaminergic neurons. SIN-1-induced changes were attenuated with selegiline or in metallothionein-transgenic striatal fetal stem cells. SIN-1-induced oxidation of dopamine (DA) to dihydroxyphenylacetaldehyde (DopaL) was attenuated in metallothionein-transgenic fetal stem cells and in cells transfected with a mitochondrial genome, and was enhanced in aging mitochondrial genome knockout cells, in metallothionein double-knockout cells, and caspase-3 gene-overexpressing dopaminergic neurons. Selegiline, melatonin, ubiquinone, and metallothionein suppressed SIN-1-induced downregulation of a mitochondrial genome and upregulation of caspase-3 as determined by reverse transcription polymerase chain reaction. These studies provide evidence that nitric oxide synthase activation and peroxynitrite ion overproduction may be involved in the etiopathogenesis of PD, and that metallothionein gene induction may provide neuroprotection.

Topics: Animals; Cell Line; DNA, Complementary; Glutathione; Humans; Metallothionein; Mice; Microscopy, Fluorescence; Neuroprotective Agents; Oligonucleotide Array Sequence Analysis; Oxidative Stress; Parkinson Disease; Peroxynitrous Acid

The mechanism of copper (Cu) neurotoxicity was studied in the RCSN-3 neuronal dopaminergic cell line, derived from substantia nigra of an adult rat. The formation of a Cu-dopamine complex was accompanied by oxidation of dopamine to aminochrome. We found that the Cu-dopamine complex mediates the uptake of (64)CuSO(4) into the Raúl Caviedes substantia nigra-clone 3 (RCSN3) cells, and it is inhibited by the addition of excess dopamine (2 m M) (63%, p < 0.001) and nomifensine (2 microM) (77%, p < 0.001). Copper sulfate (1 m M) alone was not toxic to RCSN-3 cells, but was when combined with dopamine or with dicoumarol (95% toxicity; p < 0.001) which inhibits DPNH and TPNH (DT)-diaphorase. Electron spin resonance (ESR) spectrum of the 5,5-dimethylpyrroline-N-oxide (DMPO) spin trap adducts showed the presence of a C-centered radical when incubating cells with dopamine, CuSO(4) and dicoumarol. A decrease in the expression of CuZn-superoxide dismutase and glutathione peroxidase mRNA was observed when RCSN-3 cells were treated with CuSO(4), dopamine, or CuSO(4) and dopamine. However, the mRNA expression of glutathione peroxidase remained at control levels when the cells were treated with CuSO(4), dopamine and dicoumarol. The regulation of catalase was different since all the treatments with CuSO(4) increased the expression of catalase mRNA. Our results suggest that copper neurotoxicity is dependent on: (i) the formation of Cu-dopamine complexes with concomitant dopamine oxidation to aminochrome; (ii) dopamine-dependent Cu uptake; and (iii) one-electron reduction of aminochrome.

Topics: Animals; Catalase; Cell Line; Copper Sulfate; Dicumarol; Dopamine; Electron Spin Resonance Spectroscopy; Enzyme Induction; Glutathione Peroxidase; Indolequinones; Indoles; Ion Transport; Metallothionein; NAD(P)H Dehydrogenase (Quinone); Neurons; Nomifensine; Oxidation-Reduction; Oxidative Stress; Parkinson Disease; Rats; RNA, Messenger; Substantia Nigra; Superoxide Dismutase

Virtually any neurological disorder leads to activation of resident microglia and invasion of blood-borne macrophages, which are accompanied by an increase in number and change in phenotype of astrocytes, a phenomenon generally termed reactive astrocytosis. One of the functions attributed to activation of astrocytes is thought to involve restoration of tissue damage. Hitherto, the role of astrocytes in the inflammatory reaction occurring in Parkinson's disease has not received much attention. In the present study, we examined the inflammatory events in autopsies of the substantia nigra and putamen from Parkinson's disease patients using age-matched autopsies from normal patients as controls. In the substantia nigra, activation of microglia was consistently observed in all Parkinson's disease autopsies as verified from immunohistochemical detection of CR3/43 and ferritin. Activation of resident microglia was not observed in the putamen. No differences were observed between controls and Parkinson's disease autopsies from the substantia nigra and putamen, in terms of distribution, cellular density or cellular morphology of astrocytes stained for glial fibrillary acidic protein or metallothioneins I and II, the latter sharing high affinity for metal ions and known to be induced in reactive astrocytes, possibly to exert anti-oxidative effects. Together, these findings indicate that the inflammatory process in Parkinson's disease is characterized by activation of resident microglia without reactive astrocytosis, suggesting that the progressive loss of dopaminergic neurons in Parkinson's disease is an ongoing neurodegenerative process with a minimum of involvement of the surrounding nervous tissue. The absence of reactive astrocytosis in Parkinson's disease contrasts what follows in virtually any other neurological disorder and may indicate that the inflammatory process in Parkinson's disease is a unique phenomenon.

Topics: Aged; Aged, 80 and over; Astrocytes; Ferritins; Glial Fibrillary Acidic Protein; Gliosis; Humans; Iron; Metallothionein; Microglia; Nerve Degeneration; Neuritis; Oligodendroglia; Oxidative Stress; Parkinson Disease; Putamen; Receptors, Complement; Substantia Nigra