leupeptins and Parkinsonian-Disorders

Parkinson disease (PD) is the second most common neurodegenerative disease, and it cannot be completely cured by current medications. In this study, DJ-1 protein was administrated into medial forebrain bundle of PD model rats those had been microinjected with 6-hydroxydopamine (6-OHDA) or MG-132. We found that DJ-1 protein could reduce apomorphine-induced rotations, inhibit reduction of dopamine contents and tyrosine hydroxylase levels in the striatum, and decrease dopaminergic neuron death in the substantia nigra. In 6-OHDA lesioned rats, uncoupling protein-4, uncoupling protein-5 and superoxide dismutase-2 (SOD2) mRNA and SOD2 protein were increased when DJ-1 protein was co-injected. Simultaneously, administration of DJ-1 protein reduced α-synuclein and hypoxia-inducible factor 1α mRNA and α-synuclein protein in MG-132 lesioned rats. Therefore, DJ-1 protein protected dopaminergic neurons in two PD model rats by increasing antioxidant capacity and inhibiting α-synuclein expression.

Topics: alpha-Synuclein; Animals; Antiparkinson Agents; Apomorphine; Corpus Striatum; Dopamine; Dopaminergic Neurons; Drug Evaluation, Preclinical; Humans; Hypoxia-Inducible Factor 1, alpha Subunit; Intracellular Signaling Peptides and Proteins; Ion Channels; Leupeptins; Male; Microinjections; Mitochondrial Membrane Transport Proteins; Mitochondrial Proteins; Mitochondrial Uncoupling Proteins; Motor Activity; Nerve Tissue Proteins; Neuroprotective Agents; Oncogene Proteins; Oxidative Stress; Oxidopamine; Parkinsonian Disorders; Protein Deglycase DJ-1; Rats; Rats, Sprague-Dawley; Substantia Nigra; Superoxide Dismutase; Tyrosine 3-Monooxygenase

Daytime somnolence is common in patients with Parkinson's disease (PD); however there is a lack of understanding of the cellular mechanisms involved in mediating these effects. It has been hypothesized that microglial activation and the subsequent increase of pro-inflammatory cytokines play an important role in the pathogenesis of PD. Because some cytokines are involved in the regulation of sleep, this study was designed to determine if tumor necrosis factor (TNF) and interleukin-1beta (IL-1beta), mediate daytime somnolence in the proteasome inhibitor (MG-132)-induced hemiparkinsonian rat model. Our results indicated that microglial activation caused the loss of dopaminergic neurons in the substantia nigra, and the expression of TNF-alpha, but not IL-1beta, increased in the midbrain and hypothalamus in MG-132-induced hemiparkinsonian rats. Slow-wave sleep (SWS) increased after the induction of hemiparkinsonism, but rapid eye movement (REM) sleep was not consistently altered. Application of the TNF receptor fragment (TNFRF) blocked hemiparkinsonism-induced SWS alteration, whereas the IL-1 receptor antagonist (IL-1ra) exhibited no effect. Increased nuclear translocation of NF-kappaB in the midbrain, and the blockade of SWS enhancement in MG-132-induced hemiparkinsonian rats by an inhibitor of NF-kappaB activation indicate that the TNF-NF-kappaB cascade is a critical mediator of MG-132 hemiparkinsonian-induced sleep alteration. This observation suggests potential therapeutic interventions to target the excessive daytime somnolence in patients with PD.

Topics: Animals; CD11b Antigen; Cysteine Proteinase Inhibitors; Disease Models, Animal; Disorders of Excessive Somnolence; Electroencephalography; Enzyme Inhibitors; Enzyme-Linked Immunosorbent Assay; Functional Laterality; Gene Expression Regulation; Leupeptins; Male; Microglia; Microscopy, Electron, Transmission; Neurons; NF-kappa B; Parkinsonian Disorders; Rats; Rats, Sprague-Dawley; Signal Transduction; Statistics, Nonparametric; Substantia Nigra; Tumor Necrosis Factor-alpha; Tyrosine 3-Monooxygenase

Proteasomal dysfunction and apoptosis are major hallmarks in the pathophysiology of Parkinson's disease (PD). PARK6 which is caused by mutations in the mitochondrial protein kinase PINK1 is a rare autosomal-recessively inherited disorder mimicking the clinical picture of PD. To investigate the cytoprotective physiological function of PINK1, we used primary fibroblasts from three patients homozygous for G309D-PINK1 as well as SHEP neuroblastoma cells stably overexpressing GFP-tagged wild type (wt) PINK1. Here we demonstrate that overexpression of wt PINK1 inhibits activation of Bax and release of cytochrome c, thereby diminishing caspase 9 processing and effector caspase activity after induction of proteasomal stress with the proteasome inhibitor (PI) MG132 in SHEP cells. Conversely, effector caspase activation induced by PIs, but not by the unrelated apoptotic stimulus staurosporine was potently enhanced in primary fibroblasts from homozygous PARK6 patients in comparison to those of heterozygous carriers or unaffected siblings. SHEP cells overexpressing wt PINK1 showed an elevated expression of the cytoprotective gene parkin, whereas PARK6 fibroblasts displayed significantly decreased expression of parkin in comparison to wild type control cells. Interestingly, overexpressed GFP-PINK1 was exclusively localized in the mitochondria of SHEP cells, but was redistributed to the cytoplasm under conditions of proteasomal stress. Our data indicate that PINK1 plays an important and specific physiological role in protecting cells from proteasomal stress, and suggest that PINK1 might exert its cytoprotective effects upstream of mitochondria engagement.

Topics: Analysis of Variance; Apoptosis; Blotting, Western; Cell Fractionation; Cysteine Proteinase Inhibitors; Cytochromes c; Cytoplasm; Fibroblasts; Flow Cytometry; Humans; Leupeptins; Microscopy, Confocal; Mitochondria; Parkinsonian Disorders; Proteasome Endopeptidase Complex; Protein Kinases; Reverse Transcriptase Polymerase Chain Reaction; Skin

The accumulation of misfolded and unfolded proteins in endoplasmic reticulum (ER) induces ER stress, activating the unfolded protein response (UPR). Recent evidence has suggested the relationship between UPR and dopaminergic neuronal cell death in Parkinson's disease (PD); however, it remains unclear whether it makes sense to modulate UPR, to mitigate the progression of PD. In this study, we investigated a role of the IRE1 alpha-XBP1 pathway in the survival of dopaminergic cells, under stress induced by PD-related insults. The exogenous expression of the active-form XBP1 (XBP1s) protein had protective effects against cell death induced by 1-methyl-4-phenylpyridinium (MPP+) and proteasome inhibitors. Moreover, adenoviral XBP1s expression significantly suppressed the degeneration of dopaminergic neurons in the mouse model of PD, as induced by 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). These results demonstrate that the enhancement of XBP1 could be a novel PD therapeutic strategy.

Topics: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine; 1-Methyl-4-phenylpyridinium; Acetylcysteine; Animals; Cell Death; Cell Line; Cell Survival; Cysteine Proteinase Inhibitors; DNA-Binding Proteins; Dopamine; Endoplasmic Reticulum; Endoribonucleases; Humans; Leupeptins; Male; Membrane Proteins; Mice; Mice, Inbred C57BL; Nerve Degeneration; Neurons; Parkinsonian Disorders; Protein Serine-Threonine Kinases; Regulatory Factor X Transcription Factors; RNA, Messenger; Signal Transduction; Stress, Physiological; Transcription Factors; X-Box Binding Protein 1

Spinocerebellar ataxia type 8 (SCA8) involves bidirectional expression of CUG (ATXN8OS) and CAG (ATXN8) expansion transcripts. The pathogenesis of SCA8 is complex and the spectrum of clinical presentations is broad. In the present study, we assessed the SCA8 repeat size ranges in Taiwanese Parkinson's disease, Alzheimer's disease and atypical parkinsonism and investigated the genetic variation modulating ATXN8 expression. Thirteen large SCA8 alleles and a novel ATXN8 -62 G/A promoter SNP were found. There is a significant difference in the proportion of the individuals carrying SCA8 larger alleles in atypical parkinsonism (P = 0.044) as compared to that in the control subjects. In lymphoblastoid cells carrying SCA8 large alleles, treatment of MG-132 or staurosporine significantly increases the cell death or caspase 3 activity. Although expressed at low steady-state, ATXN8 expression level is significantly higher (P = 0.012) in cells with SCA8 large alleles than that of the control cells. The ATXN8 transcriptional activity was significantly higher in the luciferase reporter construct containing the -62G allele than that containing the -62A allele in both neuroblastoma and embryonic kidney cells. Therefore, our preliminary results suggest that ATXN8 gene -62 G/A polymorphism may be functional in modulating ATXN8 expression.

Topics: Adult; Aged; Aged, 80 and over; Alzheimer Disease; Base Sequence; Case-Control Studies; Cell Line; Central Nervous System Diseases; DNA Primers; Female; Gene Expression; Humans; Leupeptins; Lymphocytes; Male; Microfilament Proteins; Middle Aged; Nerve Tissue Proteins; Parkinson Disease; Parkinsonian Disorders; Polymorphism, Single Nucleotide; Pyrimidines; RNA, Long Noncoding; RNA, Untranslated; Spinocerebellar Ataxias; Staurosporine; Sulfonamides; Trinucleotide Repeat Expansion

In Parkinson's disease (PD), dopaminergic (DA) neurons in the substantia nigra (SN, A9) are particularly vulnerable, compared to adjacent DA neurons within the ventral tegmental area (VTA, A10). Here, we show that in rat and human, one RAB3 isoform, RAB3B, has higher expression levels in A10 compared to A9 neurons. RAB3 is a monomeric GTPase protein that is highly enriched in synaptic vesicles and is involved in synaptic vesicle trafficking and synaptic transmission, disturbances of which have been implicated in several neurodegenerative diseases, including PD. These findings prompted us to further investigate the biology and neuroprotective capacity of RAB3B both in vitro and in vivo. RAB3B overexpression in human dopaminergic BE (2)-M17 cells increased neurotransmitter content, [(3)H] dopamine uptake, and levels of presynaptic proteins. AAV-mediated RAB3B overexpression in A9 DA neurons of the rat SN increased striatal dopamine content, number and size of synaptic vesicles, and levels of the presynaptic proteins, confirming in vitro findings. Measurement of extracellular DOPAC, a dopamine metabolite, following l-DOPA injection supported a role for RAB3B in enhancing the dopamine storage capacity of synaptic terminals. RAB3B overexpression in BE (2)-M17 cells was protective against toxins that simulate aspects of PD in vitro, including an oxidative stressor 6-hydroxydopamine (6-OHDA) and a proteasome inhibitor MG-132. Furthermore, RAB3B overexpression in rat SN both protected A9 DA neurons and resulted in behavioral improvement in a 6-OHDA retrograde lesion model of PD. These results suggest that RAB3B improves dopamine handling and storage capacity at presynaptic terminals, and confers protection to vulnerable DA neurons.

Topics: 3,4-Dihydroxyphenylacetic Acid; Animals; Cell Line; Corpus Striatum; Dopamine; Female; Gene Expression; Gene Expression Profiling; Humans; In Vitro Techniques; Leupeptins; Levodopa; Models, Neurological; Oxidopamine; Parkinson Disease; Parkinsonian Disorders; Presynaptic Terminals; rab3 GTP-Binding Proteins; Rats; Rats, Sprague-Dawley; RNA, Messenger; Substantia Nigra; Synaptic Vesicles; Ventral Tegmental Area

Parkinson's disease is a common progressive neurodegenerative disorder caused by the loss of dopaminergic neurons in the substantia nigra. We investigated whether cell therapy with human mesenchymal stem cells (hMSCs) had a protective effect on progressive dopaminergic neuronal loss in vitro and in vivo. In primary mesencephalic cultures, hMSCs treatment significantly decreased MG-132-induced dopaminergic neuronal loss with a significant reduction of caspase-3 activity. In rats received systemic injection of MG-132, hMSCs treatment in MG-132-treated rats dramatically reduced the decline in the number of tyrosine hydroxylase (TH)-immunoreactive cells, showing an approximately 50% increase in the survival of TH-immunoreactive cells in the substantia nigra compared with the MG-132-treated group. Additionally, hMSC treatment significantly decreased OX-6 immunoreactivity and caspase-3 activity. Histological analysis showed that the number of NuMA-positive cells was 1.7% of total injected hMSCs and 35.7% of these cells were double-stained with NuMA and TH. Adhesive-removal test showed that hMSCs administration in MG-132-treated rats had a tendency to decrease in the mean removal time. This study demonstrates that hMSCs treatment had a protective effect on progressive loss of dopaminergic neurons induced by MG-132 in vitro and in vivo. Complex mechanisms mediated by trophic effects of hMSCs and differentiation of hMSCs into functional TH-immunoreactive neurons may work in the neuroprotective process.

Topics: Animals; Caspase 3; Cell Communication; Cell Count; Cell Death; Cell Differentiation; Cell Survival; Cysteine Proteinase Inhibitors; Cytoprotection; Disease Models, Animal; Dopamine; Humans; Leupeptins; Male; Mesenchymal Stem Cell Transplantation; Mesenchymal Stem Cells; Nerve Growth Factors; Neurons; Parkinsonian Disorders; Rats; Rats, Sprague-Dawley; Substantia Nigra; Treatment Outcome; Tyrosine 3-Monooxygenase