CircDLGAP4 overexpression ameliorates neuronal injury in Parkinson's disease by binding to EIF4A3 and increasing HMGA2 expression.

Parkinson's disease (PD) is a prevalent neurodegenerative disease, and its prevalence increases steadily with age. Circular RNAs (circRNAs) are involved in various neurodegenerative diseases. Here, we aimed to explore the role of circRNA DLG-associated protein 4 (circDLGAP4) in 1-methyl-4-phenylpyridinium ion (MPP+ )-induced neuronal injury in PD. SH-SY5Y cells were treated with MPP+ to establish PD cell models. The levels of circDLGAP4 and high mobility group AT-hook 2 (HMGA2) in SH-SY5Y cells were detected. SH-SY5Y cell viability and apoptosis were detected. The levels of inflammatory damage (IL-1ß, IL-6, TNF-α) and oxidative stress (reactive oxygen species, lactate dehydrogenase, superoxide dismutase, and malondialdehyde)-related factors were measured. The binding of eukaryotic initiation factor 4A3 (EIF4A3) to circDLGAP4 and HMGA2 was analyzed using RNA pull-down or RNA immunoprecipitation. The stability of HMGA2 was detected after actinomycin D treatment, and its effects on neuronal injury were tested. CircDLGAP4 expression was decreased in MPP+ -induced SH-SY5Y cells. CircDLGAP4 upregulation restored cell activity, decreased apoptosis, and reduced inflammatory damage and oxidative stress in PD cell models. CircDLGAP4 bound to EIF4A3 to increase HMGA2 expression and stability. Silencing HMGA2 attenuated the protective effect of circDLGAP4 overexpression. Overall, circDLGAP4 upregulated HMGA2 by recruiting EIF4A3, thus increasing the mRNA stability of HMGA2 and alleviating neuronal injury in PD.

Icariside II suppresses ferroptosis to protect against MPP+-Induced Parkinson's disease through Keap1/Nrf2/GPX4 signaling.

Parkinson's disease (PD) is recognized as a degenerative and debilitating neurodegenerative disorder. The novel protective role of icariside II (ICS II) as a plant-derived flavonoid compound in neurodegenerative diseases has aroused much attention. Herein, the definite impacts of ICS II on the process of PD and the relevant action mechanism were studied. Human neuroblastoma SK-N-SH cells were challenged with 1-methyl-4-phenylpyridinium ion (MPP+) to construct the PD cell model. MTT assay and flow cytometry analysis, respectively, appraised cell viability and apoptosis. Caspase 3 Activity Assay examined caspase 3 activity. Corresponding kits examined oxidative stress levels. BODIPY 581/591 C11 assay evaluated lipid reactive oxygen species. Iron Assay Kit assessed iron content. Western blot tested the expression of apoptosis-, ferroptosis- and Kelch-like ECH-associated protein 1 (Keap1)/nuclear factor erythroid 2-related factor 2 (Nrf2)/glutathione peroxidase 4 (GPX4) signaling-associated proteins. Molecular docking verified the binding of ICS II with Keap1. The existing experimental results unveiled that ICS II elevated the viability whereas reduced the apoptosis, oxidative stress, and ferroptosis in MPP+-treated SK-N-SH cells in a concentration-dependent manner. Furthermore, ICS II declined Keap1 expression while raised Nrf2, heme oxygenase 1, and GPX4 expression. In addition, ICS II had a strong binding with Keap1 and Nrf2 inhibitor ML385 partially abolished the suppressive role of ICS II in MPP+-triggered apoptosis, oxidative stress, and ferroptosis in SK-N-SH cells. To summarize, ICS II might inhibit apoptosis, oxidative stress, and ferroptosis in the MPP+-stimulated PD cell model, which might be due to the activation of Keap1/Nrf2/GPX4 signaling.

Azoramide prevents MPP+-induced dopaminergic neuronal death via upregulating ER chaperone BiP expression.

Progressive death of dopaminergic (DA) neurons is the main cause of Parkinson's disease (PD). The discovery of drug candidates to prevent DA neuronal death is required to address the pathological aspects and alter the process of PD. Azoramide is a new small molecule compound targeting ER stress, which was originally developed for the treatment of diabetes. In this study, pre-treatment with Azoramide was found to suppress mitochondria-targeting neurotoxin MPP+-induced DA neuronal death and locomotor defects in zebrafish larvae. Further study showed that pre-treatment with Azoramide significantly attenuated MPP+-induced SH-SY5Y cell death by reducing aberrant changes in nuclear morphology, mitochondrial membrane potential, intracellular reactive oxygen species, and apoptotic biomarkers. The mechanistic study revealed that Azoramide was able to up-regulate the expression of ER chaperone BiP and thereby prevented MPP+-induced BiP decrease. Furthermore, pre-treatment with Azoramide failed to suppress MPP+-induced cytotoxicity in the presence of the BiP inhibitor HA15. Taken together, these results suggested that Azoramide is a potential neuroprotectant with pro-survival effects against MPP+-induced cell death through up-regulating BiP expression.

Integrated insight into the molecular mechanisms of selenium-modulated, MPP+-induced cytotoxicity in a Parkinson's disease model.

OBJECTIVE: Parkinson's disease (PD) is a neurodegenerative disease that is associated with oxidative stress. Due to the anti-inflammatory and antioxidant functions of Selenium (Se), this molecule may have neuroprotective functions in PD; however, the involvement of Se in such a protective function is unclear. METHODS: 1-methyl-4-phenylpyridinium (MPP+), which inhibits mitochondrial respiration, is generally used to produce a reliable cellular model of PD. In this study, a MPP+-induced PD model was used to test if Se could modulate cytotoxicity, and we further capture gene expression profiles following PC12 cell treatment with MPP+ with or without Se by genome wide high-throughput sequencing. RESULTS: We identified 351 differentially expressed genes (DEGs) and 14 differentially expressed long non-coding RNAs (DELs) in MPP+-treated cells when compared to controls. We further document 244 DEGs and 27 DELs in cells treated with MPP+ and Se vs. cells treated with MPP+ only. Functional annotation analysis of DEGs and DELs revealed that these groups were enriched in genes that respond to reactive oxygen species (ROS), metabolic processes, and mitochondrial control of apoptosis. Thioredoxin reductase 1 (Txnrd1) was also identified as a biomarker of Se treatment. CONCLUSIONS: Our data suggests that the DEGs Txnrd1, Siglec1 and Klf2, and the DEL AABR07044454.1 which we hypothesize to function in cis on the target gene Cdkn1a, may modulate the underlying neurodegenerative process, and act a protective function in the PC12 cell PD model. This study further systematically demonstrated that mRNAs and lncRNAs induced by Se are involved in neuroprotection in PD, and provides novel insight into how Se modulates cytotoxicity in the MPP+-induced PD model.

Neuroprotective microRNA-381 Binds to Repressed Early Growth Response 1 (EGR1) and Alleviates Oxidative Stress Injury in Parkinson's Disease.

As a common and disabling disease of the elderly, the standard therapies of Parkinson's disease (PD) fail to curb the ongoing neurodegeneration, thus calling for newer strategies. This work was conducted to examine the effect of microRNA-381 (miR-381) on oxidative stress injury to dopaminergic neurons in PD in vivo and in vitro. We established an in vivo mouse model of PD using 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) and an in vitro cell model of PD by treating dopaminergic neuron MN9D cells with 1-methyl-4-phenylpyridinium (MPP+). It was established that miR-381 was poorly expressed in the substantia nigra pars compacta (SNc) of MPTP-lesioned mice. The motor function of MPTP-lesioned mice was evaluated in the presence of ectopic miR-381 expression, and oxidative stress and dopaminergic neuron injury were also characterized. Restoration of miR-381 was demonstrated to diminish oxidative stress and damage in dopaminergic neurons, accompanied by enhanced motor functions. Mechanistically, the putative binding sites of miR-381 were retrieved through the starBase database, and the luciferase activity assay confirmed that it bound to EGR1 and repressed its expression, which then upregulated the expression of PTEN and p53. The neuroprotective effects of miR-381 on the motor function and dopaminergic neuronal damage were counteracted by ectopic EGR1 expression. Together, this study indicates that the binding of miR-381 to EGR1 upregulates PTEN/p53 to alleviate PD, which provides novel insights for a neuroprotective mechanism in PD.

Neuroprotection of Andrographolide against Neurotoxin MPP+-Induced Apoptosis in SH-SY5Y Cells via Activating Mitophagy, Autophagy, and Antioxidant Activities.

Parkinson's disease (PD) is associated with dopaminergic neuron loss and alpha-synuclein aggregation caused by ROS overproduction, leading to mitochondrial dysfunction and autophagy impairment. Recently, andrographolide (Andro) has been extensively studied for various pharmacological properties, such as anti-diabetic, anti-cancer, anti-inflammatory, and anti-atherosclerosis. However, its potential neuroprotective effects on neurotoxin MPP+-induced SH-SY5Y cells, a cellular PD model, remain uninvestigated. In this study, we hypothesized that Andro has neuroprotective effects against MPP+-induced apoptosis, which may be mediated through the clearance of dysfunctional mitochondria by mitophagy and ROS by antioxidant activities. Herein, Andro pretreatment could attenuate MPP+-induced neuronal cell death that was reflected by reducing mitochondrial membrane potential (MMP) depolarization, alpha-synuclein, and pro-apoptotic proteins expressions. Concomitantly, Andro attenuated MPP+-induced oxidative stress through mitophagy, as indicated by increasing colocalization of MitoTracker Red with LC3, upregulations of the PINK1-Parkin pathway, and autophagy-related proteins. On the contrary, Andro-activated autophagy was compromised when pretreated with 3-MA. Furthermore, Andro activated the Nrf2/KEAP1 pathway, leading to increasing genes encoding antioxidant enzymes and activities. This study elucidated that Andro exhibited significant neuroprotective effects against MPP+-induced SH-SY5Y cell death in vitro by enhancing mitophagy and clearance of alpha-synuclein through autophagy, as well as increasing antioxidant capacity. Our results provide evidence that Andro could be considered a potential supplement for PD prevention.

Protective Effects of Querectin against MPP+-Induced Dopaminergic Neurons Injury via the Nrf2 Signaling Pathway.

BACKGROUND: Parkinson's disease (PD) is a common selective and progressive neurodegenerative disorder of nigrostriatal dopaminergic (DA) neurons. Quercetin is a bioflavonoid with antioxidant, anti-inflammatory, anti-aging and anti-cancer properties. However, the exact mechanism by which quercetin exerts its protective effect on DAergic neurons remains unclear. PURPOSE: To investigate the underlying molecular mechanism of quercetin's protective effect on DA neurons using 1-methyl-4-phenylpyridinium (MPP+)-induced PD ferroptosis model in vitro. METHODS: MPP+ was used to induce cytotoxicity in SH-SY5Y/primary neurons. Cell viability and apoptosis were assessed by CCK-8 assay and flow cytometry. The expression levels of ferroptosis-related proteins (NCOA4, SLC7A11, Nrf2, and GPX4) were determined by Western blotting. Malondialdehyde (MDA), iron, and GPX4 levels were assesed using corresponding assay kits. Lipid peroxidation was assessed by C11-BODIPY staining. RESULTS: In the MPP+-induced ferroptosis model of SH-SY5Y cells, the expressions of SLC7A11 and GPX4 were inhibited, and the expression of NCOA4 protein was increased, causing the overproduction of MDA and lipid peroxidation. Quercetin can reduce the above changes caused by MPP+, that is, reduce the protein expression of NCOA4 in SH-SY5Y cells, increase SLC7A11 and GPX4 partially inhibited by MPP+, and reduce MDA overproduction and lipid peroxidation to protect DA neurons. Nrf2 inhibitor ML385 could inhibit quercetin-induced increase of GPX4 and SLC7A11 protein expression, indicating that the protective effect of quercetin was mediated through Nrf2. CONCLUSIONS: The results of this study suggest that quercetin regulates ferroptosis through Nrf2-dependent signaling pathways, thereby inhibiting MPP+-induced neurotoxicity in SH-SY5Y/primary neurons.

eEF1A2 siRNA Suppresses MPP+-Induced Activation of Akt and mTOR and Potentiates Caspase-3 Activation in a Parkinson's Disease Model.

The tissue-specific protein eEF1A2 has been linked to the development of neurological disorders. The role of eEF1A2 in the pathogenesis of Parkinson's disease (PD) has yet to be investigated. The aim of this study was to determine the potential neuroprotective effects of eEF1A2 in an MPP+ model of PD. Differentiated SH-SY5Y cells were transfected with eEF1A2 siRNA, followed by MPP+ exposure. The expression of p-Akt1 and p-mTORC1 was determined using Western blotting. The expression of p53, Bax, Bcl-2, and caspase-3 was evaluated using qRT-PCR. Cleaved caspase-3 levels and Annexin V/propidium iodide flow cytometry were used to determine apoptosis. The effects of PI3K inhibition were examined. The results showed that eEF1A2 siRNA significantly reduced the eEF1A2 expression induced by MPP+. MPP+ treatment activated Akt1 and mTORC1; however, eEF1A2 knockdown suppressed this activation. In eEF1A2-knockdown cells, MPP+ treatment increased the expression of p53 and caspase-3 mRNA levels as well as increased apoptotic cell death when compared to MPP+ treatment alone. In cells exposed to MPP+, upstream inhibition of the Akt/mTOR pathway, by either LY294002 or wortmannin, inhibited the phosphorylation of Akt1 and mTORC1. Both PI3K inhibitors increased eEF1A2 expression in cells, whether or not they were also treated with MPP+. In conclusion, eEF1A2 may function as a neuroprotective factor against MPP+, in part by regulating the Akt/mTOR pathway upstream.

MicroRNA-593-5p contributes to cell death following exposure to 1-methyl-4-phenylpyridinium by targeting PTEN-induced putative kinase 1.

Neurodegenerative diseases are characterized by a decline in neuronal function and structure, leading to neuronal death. Understanding the molecular mechanisms of neuronal death is crucial for developing therapeutics. MiRs are small noncoding RNAs that regulate gene expression by degrading target mRNAs or inhibiting translation. MiR dysregulation has been linked to many neurodegenerative diseases, but the underlying mechanisms are not well understood. As mitochondrial dysfunction is one of the common molecular mechanisms leading to neuronal death in many neurodegenerative diseases, here we studied miRs that modulate neuronal death caused by 1-methyl-4-phenylpyridinium (MPP+), an inhibitor of complex I in mitochondria. We identified miR-593-5p, levels of which were increased in SH-SY5Y human neuronal cells, after exposure to MPP+. We found that intracellular Ca2+, but not of reactive oxygen species, mediated this miR-593-5p increase. Furthermore, we found the increase in miR-593-5p was due to enhanced stability, not increased transcription or miR processing. Importantly, we show the increase in miR-593-5p contributed to MPP+-induced cell death. Our data revealed that miR-593-5p inhibits a signaling pathway involving PTEN-induced putative kinase 1 (PINK1) and Parkin, two proteins responsible for the removal of damaged mitochondria from cells, by targeting the coding sequence of PINK1 mRNA. Our findings suggest that miR-593-5p contributes to neuronal death resulting from MPP+ toxicity, in part, by impeding the PINK1/Parkin-mediated pathway that facilitates the clearance of damaged mitochondria. Taken together, our observations highlight the potential significance of inhibiting miR-593-5p as a therapeutic approach for neurodegenerative diseases.

Mitochondrial Complex I Inhibition in Dopaminergic Neurons Causes Altered Protein Profile and Protein Oxidation: Implications for Parkinson's disease.

Mitochondrial dysfunction and oxidative stress are critical to neurodegeneration in Parkinson's disease (PD). Mitochondrial dysfunction in PD entails inhibition of the mitochondrial complex I (CI) in the dopaminergic neurons of substantia nigra. The events contributing to CI inhibition and downstream pathways are not completely elucidated. We conducted proteomic analysis in a dopaminergic neuronal cell line exposed individually to neurotoxic CI inhibitors: rotenone (Rot), paraquat (Pq) and 1-methyl-4-phenylpyridinium (MPP+). Mass spectrometry (MS) revealed the involvement of biological processes including cell death pathways, structural changes and metabolic processes among others, most of which were common across all models. The proteomic changes induced by Pq were significantly higher than those induced by Rot and MPP+. Altered metabolic processes included downregulated mitochondrial proteins such as CI subunits. MS of CI isolated from the models revealed oxidative post-translational modifications with Tryptophan (Trp) oxidation as the predominant modification. Further, 62 peptides in 22 subunits of CI revealed Trp oxidation with 16 subunits common across toxins. NDUFV1 subunit had the greatest number of oxidized Trp and Rot model displayed the highest number of Trp oxidation events compared to the other models. Molecular dynamics simulation (MDS) of NDUFV1 revealed that oxidized Trp 433 altered the local conformation thereby changing the distance between the Fe-S clusters, Fe-S 301(N1a) to Fe-S 502 (N3) and Fe-S 802 (N4) to Fe-S 801 (N5), potentially affecting the efficiency of electron transfer. The events triggered by the neurotoxins represent CI damage, mitochondrial dysfunction and neurodegeneration in PD.

Inhibition of mitochondrial transcription by the neurotoxin MPP.

The neurotoxin MPP+ triggers cell death of dopamine neurons and induces Parkinson's disease symptoms in mice and men, but the immediate transcriptional response to this neurotoxin has not been studied. We therefore treated human SH-SY5Y cells with a low dose (0.1 mM) of MPP+ and measured the effect on nascent transcription by precision run-on sequencing (PRO-seq). We found that transcription of the mitochondrial genome was significantly reduced already after 30 min, whereas nuclear gene transcription was unaffected. Inhibition of respiratory complex I by MPP+ led to reduced ATP production, that may explain the diminished activity of mitochondrial RNA polymerase. Our results show that MPP+ has a direct effect on mitochondrial function and transcription, and that other gene expression or epigenetic changes induced by this neurotoxin are secondary effects that reflect a cellular adaptation program.

Rutin Attenuates Oxidative Stress Via PHB2-Mediated Mitophagy in MPP+-Induced SH-SY5Y Cells.

Oxidative stress plays a crucial role in the occurrence and development of Parkinson's disease (PD). Rutin, a natural botanical ingredient, has been shown to have antioxidant properties. Therefore, the aim of this study was to investigate the neuroprotective effects of rutin on PD and the underlying mechanisms. MPP+(1-methyl-4-phenylpyridinium ions)-treated SH-SY5Y cells were used as an in vitro model of PD. Human PHB2-shRNA lentiviral particles were transfected into SH-SY5Y cells to interfere with the expression of Prohibitin2 (PHB2). The oxidative damage of cells was analyzed by detecting intracellular reactive oxygen species (ROS), malondialdehyde (MDA), and mitochondrial membrane potential (MMP). Western blotting was used to detect the protein expression of antioxidant factors such as nuclear factor E2-related factor 2 (Nrf2), heme oxygenase-1 (HO-1), NADPH quinone oxidoreductase-1 (NQO-1), and mitophagy factors PHB2, translocase of outer mitochondrial membrane 20 (TOM20), and LC3II/LC3I (microtubule-associated protein II light chain 3 (LC3II) to microtubule-associated protein I light chain 3 (LC3I)). In addition, we also examined the expression of PHB2 and LC3II/LC3I by immunofluorescence staining. MPP+ treatment significantly increased the generation of ROS and MDA and the level of MMP depolarization and decreased the protein expression of Nrf2, HO-1, NQO1, TOM20, PHB2, and LC3II/LC3I. In MPP+-treated SH-SY5Y cells, rutin significantly decreased the generation of ROS and MDA and the level of MMP depolarization and increased the protein expression of Nrf2, HO-1, NQO-1, TOM20, PHB2, and LC3II/LC3I. However, the protective role of rutin was inhibited in PHB2-silenced cells. Rutin attenuates oxidative damage which may be associated with PHB2-mediated mitophagy in MPP+-induced SH-SY5Y cells. Rutin might be used as a potential drug for the prevention and treatment of PD.

The Study of Overexpression of Peroxiredoxin-2 Reduces MPP+-Induced Toxicity in the Cell Model of Parkinson's Disease.

Parkinson's disease (PD) is a chronic neurodegenerative disorder characterized by dopaminergic neuron loss, which is related to excessive reactive oxygen species (ROS) accumulation. Endogenous peroxiredoxin-2 (Prdx-2) has potent anti-oxidative and anti-apoptotic effects. Proteomics studies revealed plasma levels of Prdx-2 were significantly lower in PD patients than in healthy individuals. For further study of the activation of Prdx-2 and its role in vitro, SH-SY5Y cells and the neurotoxin 1-methyl-4-phenylpyridinium (MPP+) were used to model PD. ROS content, mitochondrial membrane potential, and cell viability were used to assess the effect of MPP+ in SH-SY5Y cells. JC-1 staining was used to determine mitochondrial membrane potential. ROS content was detected using a DCFH-DA kit. Cell viability was measured using the Cell Counting Kit-8 assay. Western blot detected the protein levels of tyrosine hydroxylase (TH), Prdx-2, silent information regulator of transcription 1 (SIRT1), Bax, and Bcl-2. The results showed that MPP+-induced accumulation of ROS, depolarization of mitochondrial membrane potential, and reduction of cell viability occurred in SH-SY5Y cells. In addition, the levels of TH, Prdx-2, and SIRT1 decreased, while the ratios of Bax and Bcl-2 increased. Then, Prdx-2 overexpression in SH-SY5Y cells showed significant protection against MPP+ -induced neuronal toxicity, as evidenced by the decrease in ROS content, increase in cell viability, increase in the level of TH, and decrease in the ratios of Bax and Bcl-2. Meanwhile, SIRT1 levels increase with the level of Prdx-2. This suggests that the protection of Prdx-2 may be related to SIRT1. In conclusion, this study indicated that overexpression of Prdx-2 reduces MPP+-induced toxicity in SH-SY5Y cells and may be mediated by SIRT1.

Doxycycline inhibits dopaminergic neurodegeneration through upregulation of axonal and synaptic proteins.

Doxycycline (DOX) is a widely used antibiotic that is able to cross the blood-brain barrier. Several studies have shown its neuroprotective effect against neurodegeneration and have associated it with antioxidant, anti-apoptotic, and anti-inflammatory mechanisms. We have recently demonstrated that DOX mimics nerve growth factor (NGF) signaling in PC12 cells. However, the involvement of this mechanism in the neuroprotective effect of DOX is unknown. Axonal degeneration and synaptic loss are key events at the early stages of neurodegeneration, and precede the neuronal death in neurodegenerative diseases, including Parkinson's disease (PD). Therefore, the regeneration of the axonal and synaptic network might be beneficial in PD. The effect of DOX in PC12 cells treated with the Parkinsonian neurotoxin 1-methyl-4-phenylpyridinium (MPP+) was addressed. Doxycycline reduced the inhibition of neuritogenesis induced by MPP+, even in cells deprived of NGF. The mechanism involved the upregulation of GAP-43, synapsin I, ß-III-tubulin, F-actin, and neurofilament-200, proteins that are associated with axonal and synaptic plasticity. Considering the role of axonal degeneration and synaptic loss at the initial stages of PD, the recent advances in early diagnosis of neurodegeneration, and the advantages of drug repurposing, doxycycline is a promising candidate to treat PD.

Hispidin in the Medicinal Fungus Protects Dopaminergic Neurons from JNK Activation-Regulated Mitochondrial-Dependent Apoptosis in an MPP+-Induced In Vitro Model of Parkinson's Disease.

Degenerative diseases of the brain include Parkinson's disease (PD), which is associated with moveable signs and is still incurable. Hispidin belongs to polyphenol and originates primarily from the medicinal fungi Inonotus and Phellinus, with distinct biological effects. In the study, MES23.5 cells were induced by 1-methyl-4-phenylpyridinium (MPP+) to build a cell model of PD in order to detect the protective effect of hispdin and to specify the underlying mechanism. Pretreatment of MES23.5 cells with 1 h of hispdin at appropriate concentrations, followed by incubation of 24 h with 2 µmol/L MPP+ to induce cell damage. MPP+ resulted in reactive oxygen species production that diminished cell viability and dopamine content. Mitochondrial dysfunction in MS23.5 cells exposed to MPP+ was observed, indicated by inhibition of activity in the mitochondrial respiratory chain complex I, the collapse of potential in mitochondrial transmembrane, and the liberation of mitochondrial cytochrome c. Enabling C-Jun N-terminal kinase (JNK), reducing Bcl-2/Bax, and enhancing caspase-9/caspase-3/PARP cleavage were also seen by MPP+ induction associated with increased DNA fragmentation. All of the events mentioned above associated with MPP+-mediated mitochondrial-dependent caspases cascades were attenuated under cells pretreatment with hispidin (20 µmol/L); similar results were obtained during cell pretreatment with pan-JNK inhibitor JNK-IN-8 (1 µmol/L) or JNK3 inhibitor SR3576 (25 µmol/L). The findings show that hispidin has neuroprotection against MPP+-induced mitochondrial dysfunction and cellular apoptosis and suggest that hispidin can be seen as an assist in preventing PD.

Investigating Therapeutic Effects of Indole Derivatives Targeting Inflammation and Oxidative Stress in Neurotoxin-Induced Cell and Mouse Models of Parkinson's Disease.

Neuroinflammation and oxidative stress have been emerging as important pathways contributing to Parkinson's disease (PD) pathogenesis. In PD brains, the activated microglia release inflammatory factors such as interleukin (IL)-ß, IL-6, tumor necrosis factor (TNF)-α, and nitric oxide (NO), which increase oxidative stress and mediate neurodegeneration. Using 1-methyl-4-phenylpyridinium (MPP+)-activated human microglial HMC3 cells and the sub-chronic 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced mouse model of PD, we found the potential of indole derivative NC009-1 against neuroinflammation, oxidative stress, and neurodegeneration for PD. In vitro, NC009-1 alleviated MPP+-induced cytotoxicity, reduced NO, IL-1ß, IL-6, and TNF-α production, and suppressed NLR family pyrin domain containing 3 (NLRP3) inflammasome activation in MPP+-activated HMC3 cells. In vivo, NC009-1 ameliorated motor deficits and non-motor depression, increased dopamine and dopamine transporter levels in the striatum, and reduced oxidative stress as well as microglia and astrocyte reactivity in the ventral midbrain of MPTP-treated mice. These protective effects were achieved by down-regulating NLRP3, CASP1, iNOS, IL-1ß, IL-6, and TNF-α, and up-regulating SOD2, NRF2, and NQO1. These results strengthen the involvement of neuroinflammation and oxidative stress in PD pathogenic mechanism, and indicate NC009-1 as a potential drug candidate for PD treatment.

LncRNA SNHG15 mediates 1-methyl-4-phenylpyridinium (MPP+)-induced neuronal damage through targeting miR-29c-3p/SNCA axis.

BACKGROUND: Parkinson's disease (PD) is the most prevalent neurodegenerative disease in the elderly people. Long non-coding ribose nucleic acids (LncRNAs) can serve as molecular sponges for micro RNA (miRNA) and regulate gene expression, which is implicated in the occurrence and progression of PD. In this work, we investigated the functional role of lncRNA SNHG15 in a neuronal damage cell model and its potential mechanism. METHODS: SK-N-SH cells treated with 1-methyl-4-phenylpyridinium (MPP+) were employed as the in vitro cellular model to mimic neuronal degeneration. The expression levels of SNHG15, miR-29c-3p, and SNCA were determined by qRT-PCR. ELISA, CCK-8 proliferation assay, and flow cytometry were conducted to explore the effects of SNHG15 and miR-29c-3p on the production of inflammatory factors, cell proliferation, and apoptosis, respectively. Dual-luciferase reporter assay was utilized to validate the functional interactions among SNHG15, miR-29c-3p, and SNCA. SNCA protein levels were examined by Western blot. RESULTS: SNHG15 was highly induced in the cell model of MPP+-induced neuronal damage. SNHG15 knockdown significantly mitigated MPP+-induced damages in SK-N-SH cells. SNHG15 served as a sponge to down-regulate miR-29c-3p, thereby releasing the inhibition of miR-29c-3p on SNCA expression, which promoted neuronal damages upon MPP+ challenge. CONCLUSION: The upregulation of SNHG15 upon MPP+ challenge mediates neuronal damages in SK-N-SH cells by regulating miR-29c-3p/SNCA axis. Future work is required to validate these findings in PD patients and animal models, which could provide insights into the diagnosis and therapy of PD.

Salsolinol Protects SH-SY5Y Cells Against MPP+ Damage and Increases Enteric S100-Immunoreactivity in Wistar Rats.

A dopamine derivative, 1-methyl-6,7-dihydroxy-1,2,3,4-tetrahydroisoquinoline, known as salsolinol (SAL), has increasingly gained attention since its first detection in the urine of Parkinson's disease patients treated with levodopa, and has been proposed as a possible neurotoxic contributor to the disease. Yet, so far, the neurobiological role of SAL remains unclear. Thus, the main aims of our study were to compare the neurotoxic potential of SAL with MPP+ (1-methyl-4-phenylpyridinium ion) in vitro, and to examine intestinal and metabolic alterations following intraperitoneal SAL administration in vivo. In vitro, SH-SY5Y neuroblastoma cell line was monitored following MPP+ and SAL treatment. In vivo, Wistar rats were subjected to SAL administration by either osmotic intraperitoneal mini-pumps or a single intraperitoneal injection, and after two weeks, biochemical and morphological parameters were assessed. SH-SY5Y cells treated with MPP+ (1000 µM) and SAL (50 µM) showed increase in cell viability and fluorescence intensity in comparison with the cells treated with MPP+ alone. In vivo, we predominantly observed decreased collagen content in the submucosal layer, decreased neuronal density with comparable ganglionic area in the jejunal myenteric plexus, and increased glial S100 expression in both enteric plexuses, yet with no obvious signs of inflammation. Besides, glucose and triglycerides levels were lower after single SAL-treatment (200 mg/kg), and low- to high-density lipoprotein (LDL/HDL) ratio and aspartate to alanine aminotransferases (AST/ALT) ratio levels were higher after continuous SAL-treatment (200 mg/kg in total over 2 weeks). Low doses of SAL were non-toxic and exhibited pronounced neuroprotective properties against MPP+ in SH-SY5Y cell line, which supports the use of SAL as a reference compound for in vitro studies. In vivo results give insight into our understanding of gastrointestinal remodeling following intraperitoneal SAL administration, and might represent morphological correlates of a microglial-related enteric neurodegeneration and dopaminergic dysregulation.

Ketone Body ß-Hydroxybutyric Acid Ameliorates Dopaminergic Neuron Injury Through Modulating Zinc Finger Protein 36/Acyl-CoA Synthetase Long-Chain Family Member Four Signaling Axis-Mediated Ferroptosis.

ß-hydroxybutyrate (BHB) is one of main component of ketone body, which plays an important protective role in various tissues and organs. Whereas, its exact regulatory roles and mechanisms in Parkinson's disease (PD) have not been full elucidated. In this study, SN4741 cells and C57BL/6 mice were treated with 1-methyl-4-phenylpyridinium ion (MPP+)/1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) to establish the PD model in vitro and in vivo. Cell viability and damage to dopaminergic neurons were measured by cell counting kit 8, Calcein-AM/PI staining, terminal dexynucleotidyl transferase (TdT)-mediated dUTP nick end labeling and hematoxylin & eosin staining. Corresponding assay kits and BODIPY 581/591 C11 probe evaluated oxidative stress and intracellular iron levels. Western blot examined the ferroptosis-related proteins. MPTP/MPP+-treatment reduced cell viability but triggered oxidative stress and ferroptosis in SNA4741 cells and brain tissues of mice. However, these effects were dramatically reversed by BHB and Fer-1 treatment. Mechanistically, Zinc finger protein 36 (ZFP36) was a target of BHB, and its depletion could reverse the anti-oxidative stress and anti-ferroptosis roles of BHB. Moreover, ZFP36 could directly bound to acyl-CoA synthetase long-chain family member 4 (ACSL4) mRNA to decay its expression, thus negatively modulating ACSL4-mediated oxidative stress and ferroptosis. Summary, BHB alleviated oxidative stress and ferroptosis of dopaminergic neurons in PD via modulating ZFP36/ACSL4 axis, which provided some new understanding for PD prevention and treatment.

A Novel Mitochondria-Targeting Iron Chelator Neuroprotects Multimodally via HIF-1 Modulation Against a Mitochondrial Toxin in a Dopaminergic Cell Model of Parkinson's Disease.

Coumarins are plant-derived polyphenolic compounds belonging to the benzopyrones family, possessing wide-ranging pharmaceutical applications including cytoprotection, which may translate into therapeutic potential for multiple diseases, including Parkinson's disease (PD). Here we demonstrate the neuroprotective potential of a new polyhydroxyl coumarin, N-(1,3-dihydroxy-2-(hydroxymethyl)propan-2-yl)-2-(7-hydroxy-2-oxo-2H-chromen-4-yl)acetamide (CT51), against the mitochondrial toxin 1-methyl-4-phenylpyridinium (MPP+). MPP+'s mechanism of toxicity relates to its ability to inhibit complex I of the mitochondrial electron transport chain (METC), leading to adenosine triphosphate (ATP) depletion, increased reactive oxygen species (ROS) production, and apoptotic cell death, hence mimicking PD-related neuropathology. Dopaminergic differentiated human neuroblastoma cells were briefly pretreated with CT51, followed by toxin exposure. CT51 significantly restored somatic cell viability and neurite processes; hence, the drug targets cell bodies and axons thereby preserving neural function and circuitry against PD-related damage. Moreover, MPP+ emulates the iron dyshomeostasis affecting dopaminergic neurons in PD-affected brains, whilst CT51 was previously revealed as an effective iron chelator that preferentially partitions to mitochondria. We extend these findings by characterising the drug's interactive effects at the METC level. CT51 did not improve mitochondrial coupling efficiency. However, voltammetric measurements and high-resolution respirometry analysis revealed that CT51 acts as an antioxidant agent. Also, the neuronal protection afforded by CT51 associated with downregulating MPP+-induced upregulated expression of hypoxia-inducible factor 1 alpha (HIF-1α), a protein which regulates iron homeostasis and protects against certain forms of oxidative stress after translocating to mitochondria. Our findings support the further development of CT51 as a dual functioning iron chelator and antioxidant antiparkinsonian agent.