OBJECTIVE: Mitophagy is known to contribute towards progression of Parkinson's disease. Korean red ginseng (KRG) is a widely used medicinal herb in East Asia, and recent studies have reported that KRG prevents 1-methyl-4-phenylpyridinium ion (MPP METHODS: SH-SY5Y cells were incubated with KRG for 24 h, and subsequently exposed to MPP RESULTS: MPP CONCLUSION KRG effectively prevents MPP
1-Methyl-4-phenylpyridinium/toxicity* ; Apoptosis ; Cell Line, Tumor ; Mitochondria ; Mitophagy ; Panax ; Reactive Oxygen Species

We previously demonstrated that 1-methyl-4-phenylpyridinium (MPP+) causes caspase-independent, non-apoptotic death of dopaminergic (DA) neuronal cells. Here, we specifically examined whether change of glucose concentration in culture medium may play a role for determining cell death modes of DA neurons following MPP+ treatment. By incubating MN9D cells in medium containing varying concentrations of glucose (5~35 mM), we found that cells underwent a distinct cell death as determined by morphological and biochemical criteria. At 5~10 mM glucose concentration (low glucose levels), MPP+ induced typical of the apoptotic dell death accompanied with caspase activation and DNA fragmentation as well as cell shrinkage. In contrast, MN9D cells cultivated in medium containing more than 17.5 mM (high glucose levels) did not demonstrate any of these changes. Subsequently, we observed that MPP+ at low glucose levels but not high glucose levels led to ROS generation and subsequent JNK activation. Therefore, MPP+-induced cell death only at low glucose levels was significantly ameliorated following co-treatment with ROS scavenger, caspase inhibitor or JNK inhibitor. We basically confirmed the quite similar pattern of cell death in primary cultures of DA neurons. Taken together, our results suggest that a biochemically distinct cell death mode is recruited by MPP+ depending on extracellular glucose levels.
1-Methyl-4-phenylpyridinium ; Cell Death* ; DNA Fragmentation ; Dopaminergic Neurons* ; Glucose* ; Neurons ; Parkinson Disease ; Reactive Oxygen Species

BACKGROUND: 1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) produces an irreversible and severe parkinsonian-like syndrome. A licorice compound glycyrrhizin exerts a cytoprotective or anti-oxidant effect in various disease conditions, but its effect against the MPTP-induced brain tissue damage remains uncertain. The present study elucidates the protective effects of glycyrrhizin against brain tissue damage in the MPTP mouse model of Parkinson's disease. METHODS: We measured the activities of antioxidant enzymes and formation of tissue peroxidation products in the brains of MPTP-treated mice. We also performed an in vitro assay to examine the effects of 1-methyl-4-phenylpyridinium (MPP+) on the mitochondrial respiratory electron flow, membrane potential and cytochrome c release and measured the scavenging action of glycyrrhizin against reactive oxygen species. RESULTS: The MPTP treatment increased activities of total superoxide dismutase, catalase, and glutathione peroxidase and levels of malondialdehyde and carbonyls in the basal ganglia, diencephalon plus midbrain compared to the control mouse brain. Co-administration of glycyrrhizin (16.8 mg/kg = 20 micrometer) attenuated the MPTP effect on the enzyme activities and formation of tissue peroxidation products. Glycyrrhizin attenuated the 500 micrometer MPP+ -induced inhibition of electron flow, changes in the membrane potential and cytochrome c release in isolated brain mitochondria. Glycyrrhizin (1-50 micrometer) showed a scavenging action against superoxide radicals, hydrogen peroxide and hydroxyl radicals. CONCLUSIONS: Glycyrrhizin may prevent the toxicity of MPTP against brain tissue by suppressing mitochondrial damage and oxidative tissue damage. Glycyrrhizin seems to attenuate oxidative brain tissue damage occurring in Parkinson's disease through antioxidant action and prevention of mitochondrial dysfunction.
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine* ; 1-Methyl-4-phenylpyridinium ; Animals ; Antioxidants ; Basal Ganglia ; Brain ; Catalase ; Cytochromes c ; Diencephalon ; Glutathione Peroxidase ; Glycyrrhiza ; Glycyrrhizic Acid* ; Hydrogen Peroxide ; Malondialdehyde ; Membrane Potentials ; Mesencephalon ; Mice* ; Mitochondria ; Parkinson Disease* ; Reactive Oxygen Species ; Superoxide Dismutase ; Superoxides

The present study examined the inhibitory effect of licorice compounds glycyrrhizin and a metabolite 18 beta-lycyrrhetinic acid on the neurotoxicity of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) in the mouse and on the 1-methyl-4-phenylpyridinium (MPP+-induced cell death in differentiated PC12 cells. MPTP treatment increased the activities of total superoxide dismutase, catalase and glutathione peroxidase and the levels of malondialdehyde and carbonyls in the brain compared to control mouse brain. Co-administration of glycyrrhizin (16.8 mg/kg) attenuated the MPTP effect on the enzyme activities and formation of tissue peroxidation products. In vitro assay, licorice compounds attenuated the MPP+induced cell death and caspase-3 activation in PC12 cells. Glycyrrhizin up to 100 micrometer significantly attenuated the toxicity of MPP+ Meanwhile, 18beta-lycyrrhetinic acid showed a maximum inhibitory effect at 10 micrometer; beyond this concentration the inhibitory effect declined. Glycyrrhizin and 18beta-lycyrrhetinic acid attenuated the hydrogen peroxide- or nitrogen species-induced cell death. Results from this study indicate that glycyrrhizin may attenuate brain tissue damage in mice treated with MPTP through inhibitory effect on oxidative tissue damage. Glycyrrhizin and 18 beta-lycyrrhetinic acid may reduce the MPP+toxicity in PC12 cells by suppressing caspase-3 activation. The effect seems to be ascribed to the antioxidant effect.
1-Methyl-4-phenyl-1,2,3,6-tetrahydropyridine ; 1-Methyl-4-phenylpyridinium ; Animals ; Antioxidants ; Brain ; Caspase 3 ; Catalase ; Cell Death ; Glutathione Peroxidase ; Glycyrrhiza ; Glycyrrhizic Acid ; Hydrogen ; Malondialdehyde ; Mice ; Nitrogen ; PC12 Cells ; Superoxide Dismutase

BACKGROUND: The extract and hemiterpene glycosides of Ilex Rotunda Thunb exert antioxidant and anti-inflammatory effects. The effect of rotundarpene on apoptosis in neuronal cells caused by the 1-methyl-4-phenylpyridinium (MPP⁺) has not been reported previously. METHODS: Using differentiated PC12 cells and human neuroblastoma SH-SY5Y cells, we investigated the effect of rotundarpene on MPP⁺-caused apoptosis in relation to the cell death process. RESULTS: MPP⁺-induced cell death was identified using the MTT and neutral red uptake tests. Apoptosis was induced by eliciting decreases in the cytosolic levels of Bid and Bcl-2 proteins, increases in the cytosolic levels of Bax and p53, disruption of the mitochondrial transmembrane potential, and the release of cytochrome c and the activation of caspase-8, -9, and -3 in differentiated PC12 cells and SH-SY5Y cells. Treatment with rotundarpene reduced the MPP⁺-induced changes in the levels of apoptosis-regulated proteins, formation of reactive oxygen species, depletion and oxidation of glutathione, and cell death in both PC12 and SH-SY5Y cells. CONCLUSIONS: Rotundarpene may reduce MPP⁺-induced apoptosis in neuronal cells by suppressing the activation of the mitochondria-mediated pathway and the caspase-8 and Bid pathways. Rotundarpene appears to act by inhibiting the production of reactive oxygen species and by the depletion and oxidation of glutathione.
1-Methyl-4-phenylpyridinium ; Animals ; Apoptosis ; Caspase 8 ; Cell Death* ; Cytochromes c ; Cytosol ; Glutathione ; Glycosides ; Humans ; Ilex ; Membrane Potentials ; Neuroblastoma ; Neurons ; Neutral Red ; PC12 Cells ; Reactive Oxygen Species

BACKGROUND: 1-Methyl-4-phenylpyridinium (MPP+) causes a neuronal cell injury that is similar to the findings observed in Parkinson's disease. Caffeoylquinic acid derivatives have demonstrated anti-oxidant and anti-inflammatory effects. Nevertheless, the effect of 3,4,5-tricaffeoylquinic acid (3,4,5-triCQA) on the neuronal cell death due to exposure of parkinsonian toxin MPP+ remains unclear. METHODS: Using differentiated PC12 cells, the preventive effect of 3,4,5-triCQA on the MPP+-induced cell death in relation to apoptotic process was examined. RESULTS: MPP+ induced a decrease in Bid, Bcl-2 and survivin protein levels, increase in Bax levels, loss of the mitochondrial transmembrane potential, cytochrome c release, activation of caspases (-8, -9 and -3), cleavage of PARP-1, and an increase in the tumor suppressor p53 levels. 3,4,5-Tricaffeoylquinic acid attenuated the MPP+-induced changes in the apoptosis-related protein levels, formation of reactive oxygen species, depletion of GSH, nuclear damage and cell death. 3,4,5-Tricaffeoylquinic acid attenuated another parkinsonian neurotoxin rotenone-induced cell death. CONCLUSIONS: 3,4,5-Tricaffeoylquinic acid may attenuate the MPP+-induced apoptosis in PC12 cells by suppressing the activation of the mitochondrial pathway and the caspase-8- and Bid-dependent pathways. The preventive effect seems to be ascribed to its inhibitory effect on the formation of reactive oxygen species and depletion of GSH.
1-Methyl-4-phenylpyridinium ; Animals ; Apoptosis* ; Caspases ; Cell Death ; Cytochromes c ; Membrane Potentials ; Neurons ; Parkinson Disease ; PC12 Cells ; Reactive Oxygen Species

BACKGROUND: Parkinson disease (PD) is caused by selective cell death of dopaminergic neurons in the substantia nigra. An early onset form of PD, autosomal recessive juvenile parkinsonism has been associated with a mutation in the parkin gene. The function of parkin is known to remove misfolding proteins and protect cell death. We aimed to investigate the role of parkin against oxidative stress in neuronal cells. METHODS: Parkin knockout embryonic stem cells (PKO ES cells) were differentiated into neurons by adherent monolayer culture method. Oxidative stress was induced by the treatment of 1-methyl-4-phenylpyridinium (MPP+) in neurons derived from wild type and PKO ES cells, and cell viability was examined by MTT assay. After exposure to MPP+, Tuj1-positive cell population was compared between PKO and wild type cells by fluorescence activated cell sorter (FACS) analysis. The activated caspase3 protein level was also measured by Western blot analysis, FACS and immunocytochemistry. RESULTS: There was no difference in the efficiency of neuronal differentiation between wild type and PKO ES cells. After exposure to MPP+, no significant differences were found in cell viability and Tuj1-positive cell population between the two groups determined by MTT assay and FACS analysis, respectively. The activated caspase3 protein levels examined by Western blot analysis, FACS and immunocytochemistry were not changed in PKO cells compared with those of wild type cells after MPP+ treatment. CONCLUSION: These results suggest that PKO neuronal cells including dopaminergic neurons are not sensitive to caspase3-dependent cell death pathway during the response against MPP+-induced oxidative stress.
1-Methyl-4-phenylpyridinium ; Blotting, Western ; Cell Death ; Cell Survival ; Dopaminergic Neurons ; Embryonic Stem Cells ; Fluorescence ; Immunohistochemistry ; Neurons ; Oxidative Stress* ; Parkinson Disease ; Parkinsonian Disorders ; Substantia Nigra

BACKGROUND: Protein casein kinase 2 is involved in signal transduction, cell growth, and apoptosis. However, it has not been elucidated whether parkinsonian toxin 1-methyl-4-phenylpyridinium (MPP+)-induced neuronal cell death is mediated by a casein-kinase-2-mediated pathway. METHODS: We monitored apoptosis-related protein activation, changes in the level of casein kinase 2, nuclear damage, and apoptosis in differentiated PC12 cells exposed to MPP+ in combination with casein kinase 2 inhibitor. RESULTS: Casein kinase 2 inhibitors [4,5,6,7-tetrabromobenzotriazole (TBB), 5,6-dichloro-1-beta-D-ribofuranosylbenzimidazole, and apigenin] reduced MPP+- and rotenone-induced cell death in differentiated PC12 cells. TBB inhibited the MPP+-induced activation of apoptosis-related proteins (decreases in Bid and Bcl-2 levels, increase in Bax levels, cytochrome c release, and caspase-3 activation), increase in casein kinase 2 levels, and nuclear damage. CONCLUSIONS: Administering casein kinase 2 inhibitor TBB at concentrations that do not induce toxic effects may reduce MPP+-induced cell death in differentiated PC12 cells by suppressing the apoptosis-related protein activation that leads to cytochrome c release and subsequent activation of caspase-3. The results suggest that MPP+-induced cell death process is mediated by a casein kinase 2 pathway.
1-Methyl-4-phenylpyridinium ; Animals ; Apoptosis ; Casein Kinase II ; Casein Kinases ; Caseins ; Caspase 3 ; Cell Death ; Cytochromes c ; Neurons ; PC12 Cells ; Proteins ; Signal Transduction ; Triazoles

The aim of the present study was to investigate the protective effects of crude polysaccharides from Chroogomphus rutilus on dopaminergic neurons impaired by MPP(+). SH-SY5Y cells were pretreated with crude polysaccharides (200, 400 and 800 μg/mL), and then MPP(+) was added to cell medium. After 48 h of incubation, MTT method was used to detect the survival rate of SH-SY5Y cells damaged by MPP(+). Annexin V-FITC staining and flow cytometry were used to detect apoptotic rate. The results showed that pretreating SH-SY5Y cells with crude polysaccharides (400 and 800 μg/mL) increased the survival rates, and reduced the apoptotic rates of SH-SY5Y cells. To rule out the possibility that crude polysaccharides may decrease actual concentration of MPP(+) by direct binding, we washed off crude polysaccharides before MPP(+) addition. Under this experimental condition, MTT results showed the survival rates of the SH-SY5Y cells were still significantly increased by 800 μg/mL crude polysaccharides pretreatment. These results suggest a protective effect of polysaccharides on the SH-SY5Y cells. Most of this protection is contributed by direct action of polysaccharides on the cells, not by binding with MPP(+). It is indicated that the crude polysaccharides from Chroogomphus rutilus can be developed as a potential drug for Parkinson's disease prevention and treatment in the future.
1-Methyl-4-phenylpyridinium ; adverse effects ; Apoptosis ; drug effects ; Basidiomycota ; chemistry ; Cell Line, Tumor ; Dopaminergic Neurons ; drug effects ; Humans ; Polysaccharides ; pharmacology

The aim of this study was to investigate the effect of edaravone (Eda) on the balance of mitochondrial fusion and fission in Parkinson's disease (PD) cell model. A cell model of PD was established by treating PC12 cells with 500 μmol/L 1-methyl-4-phenylpyridinium (MPP). Thiazole blue colorimetry (MTT) was used to detect the effect of different concentrations of Eda on the survival rate of PC12 cells exposed to MPP. The mitochondrial morphology was determined by laser confocal microscope. Western blot was used to measure the protein expression levels of mitochondrial fusion- and fission-related proteins, including OPA1, MFN2, DRP1 and Fis1. The results showed that pretreatment with different concentrations of Eda antagonized MPP-induced PC12 cell damage in a dose-dependent manner. The PC12 cells treated with MPP showed mitochondrial fragmentation, up-regulated DRP1 and Fis1 protein expression levels, and down-regulated MFN2 and OPA1 protein expression levels. Eda could reverse the above changes in the MPP-treated PC12 cells, but did not affect Fis1 protein expression. These results suggest that Eda has a protective effect on the mitochondrial fusion disruption induced by MPP in PC12 cells. The mechanism may be related to the up-regulation of OPA1/MFN2 and down-regulation of DRP1.
1-Methyl-4-phenylpyridinium ; Animals ; Dynamins ; Edaravone ; pharmacology ; GTP Phosphohydrolases ; Mitochondria ; drug effects ; Mitochondrial Dynamics ; Mitochondrial Proteins ; PC12 Cells ; Parkinson Disease ; Rats ; Up-Regulation