The study aimed to investigate the effect of the CD200R1 gene deletion on microglia activation and nigrostriatal dopamine neuron loss in the Parkinson's disease (PD) process. The CRISPR-Cas9 technology was applied to construct the CD200R1^-/- mice. The primary microglia cells of wild-type and CD200R1^-/- mice were cultured and treated with bacterial lipopolysaccharide (LPS). Microglia phagocytosis level was assessed by a fluorescent microsphere phagocytosis assay. PD mouse model was prepared by nigral stereotaxic injection of recombinant adeno-associated virus vector carrying human α-synuclein (α-syn). The changes in the motor behavior of the mice with both genotypes were evaluated by cylinder test, open field test, and rotarod test. Immunohistochemical staining was used to assess the loss of dopamine neurons in substantia nigra. Immunofluorescence staining was used to detect the expression level of CD68 (a key molecule involved in phagocytosis) in microglia. The results showed that CD200R1 deletion markedly enhanced LPS-induced phagocytosis in vitro by the microglial cells. In the mouse model of PD, CD200R1 deletion exacerbated motor behavior impairment and dopamine neuron loss in substantia nigra. Fluorescence intensity analysis results revealed a significant increase in CD68 expression in microglia located in the substantia nigra of CD200R1^-/- mice. The above results suggest that CD200R1 deletion may further activates microglia by promoting microglial phagocytosis, leading to increased loss of the nigrostriatal dopamine neurons in the PD model mice. Therefore, targeting CD200R1 could potentially serve as a novel therapeutic target for the treatment of early-stage PD.
Animals ; Microglia/physiology* ; Mice ; Phagocytosis ; Parkinson Disease/genetics* ; Disease Models, Animal ; Receptors, Cell Surface/physiology* ; Dopaminergic Neurons/pathology* ; Antigens, CD/metabolism* ; Gene Deletion ; Substantia Nigra ; Mice, Inbred C57BL ; Mice, Knockout ; Cells, Cultured ; Male ; alpha-Synuclein ; CD68 Molecule ; Orexin Receptors

Objective: To observe the dynamic changes of brainstem locus coeruleus (LC) damage in Parkinson' s disease (PD) -like mice by paraquat (PQ) . Methods: In October 2019, 36 male C57BL/6 mice were randomly divided into the exposure group and the control group, with 18 mice in each group. The mice in the exposure group were given intraperitoneal injection of 15 mg/kg PQ, and the mice in the control group were given intraperitoneal injection of 0.9% saline, twice a week for 8 weeks. Neurobehavioral changes (pole climbing test, swimming test, open field test, tail hanging test, high plus maze test and water maze test) were observed at 4 weeks, 6 weeks and 8 weeks, respectively, and the changes of motor ability, emotion and cognitive function were evaluated. The brain tissue of mice were taken and stained with Hematoxylin-Eosin (HE) to observe the pathological changes of LC. Nissl staining was used to detect the changes of neuronal Nissl bodies in LC. Immunohistochemistry (IHC) staining was used to detect the expression of neuron nuclear antigen (NeuN) , dopamine (DA) neurons and norepinephrine (NE) neuron markers tyrosine hydroxylase (TH) , α-synuclein (α-syn) in substantia nigra (SN) and LC. The expression levels of NeuN, TH and α-syn in the midbrain and brainstem were detected by Western blotting. TUNEL staining was used to detect neuronal apoptosis in LC. Results: Compared with the 4th week of PQ exposure group, the time of pole climbing and swimming immobility were gradually increased, the ratio of open arm residence time of high plus maze test and the number of times of the platform and the residence time of platform quadrant in water maze test were gradually decreased (P<0.05) in the exposure group with the progress of exposure time. The results of HE and Nissl staining showed that the neurons in LC gradually arranged loosely, the nucleus were deeply stained, the cytoplasm was pyknosis, and the number of Nissl bodies gradually decreased (P<0.05) in the exposure group with the progress of exposure time. IHC results showed that the number of NeuN and TH positive cells in SN and LC of mice were gradually decreased, and the positive expression of α-syn was gradually increased (P<0.05) in the exposure group with the progress of exposure time. Western blotting results showed that the expression levels of NeuN and TH in the midbrain and brainstem were gradually decreased, and the expression level of α-syn was gradually increased (P<0.05) in the exposure group with the progress of exposure time. TUNEL staining showed that the apoptosis rates of neurons in LC were gradually increased (P<0.05) in the exposure group with the progress of exposure time. Conclusion: PQ induces progressive damage in the LC area of PD-like mice, which may be caused by the abnormal accumulation of pathological α-syn in the LC area.
Animals ; Dopaminergic Neurons ; Locus Coeruleus/pathology* ; Male ; Mice ; Mice, Inbred C57BL ; Paraquat/toxicity* ; Parkinson Disease/metabolism* ; Substantia Nigra ; Tyrosine 3-Monooxygenase/metabolism*

The traditionally used oriental herbal medicine Moutan Cortex Radicis [MCR; Paeonia Suffruticosa Andrews (Paeoniaceae)] exerts anti-inflammatory, anti-spasmodic, and analgesic effects. In the present study, we investigated the therapeutic effects of differently fractioned MCR extracts in a 6-hydroxydopamine (OHDA)-induced Parkinson's disease model and neuro-blastoma B65 cells. Ethanol-extracted MCR was fractionated by n-hexane, butanol, and distilled water. Adult Sprague-Dawley rats were treated first with 20 μg of 6-OHDA, followed by three MCR extract fractions (100 or 200 mg·kg) for 14 consecutive days. In the behavioral rotation experiment, the MCR extract-treated groups showed significantly decreased number of net turns compared with the 6-OHDA control group. The three fractions also significantly inhibited the reduction in tyrosine hydroxylase-positive cells in the substantia nigra pars compacta following 6-OHDA neurotoxicity. Western blotting analysis revealed significantly reduced tyrosine hydroxylase expression in the substantia nigra pars compacta in the 6-OHDA-treated group, which was significantly inhibited by the n-hexane or distilled water fractions of MCR. B65 cells were exposed to the extract fractions for 24 h prior to addition of 6-OHDA for 30 min; treatment with n-hexane or distilled water fractions of MCR reduced apoptotic cell death induced by 6-OHDA neurotoxicity and inhibited nitric oxide production and neuronal nitric oxide synthase expression. These results showed that n-hexane- and distilled water-fractioned MCR extracts inhibited 6-OHDA-induced neurotoxicity by suppressing nitric oxide production and neuronal nitric oxide synthase activity, suggesting that MCR extracts could serve as a novel candidate treatment for the patients with Parkinson's disease.
Animals ; Anti-Inflammatory Agents ; pharmacology ; therapeutic use ; Antiparkinson Agents ; pharmacology ; therapeutic use ; Cell Death ; drug effects ; Cell Line ; Disease Models, Animal ; Drugs, Chinese Herbal ; chemistry ; Neurons ; pathology ; Nitric Oxide ; analysis ; Nitric Oxide Synthase Type I ; biosynthesis ; Oxidopamine ; toxicity ; Paeonia ; chemistry ; Parkinsonian Disorders ; chemically induced ; drug therapy ; Phytotherapy ; Plant Extracts ; pharmacology ; therapeutic use ; Plants, Medicinal ; Rats ; Rats, Sprague-Dawley ; Substantia Nigra ; drug effects ; enzymology ; Tyrosine 3-Monooxygenase ; genetics ; metabolism

Parkinson's disease (PD) is characterized by the selective loss of dopaminergic neurons of the substantia nigra pars compacta (SNc) with motor and nonmotor symptoms. Defective mitochondrial function and increased oxidative stress (OS) have been demonstrated as having an important role in PD pathogenesis, although the underlying mechanism is not clear. The etiopathogenesis of sporadic PD is complex with variable contributions of environmental factors and genetic susceptibility. Both these factors influence various mitochondrial aspects, including their life cycle, bioenergetic capacity, quality control, dynamic changes of morphology and connectivity (fusion, fission), subcellular distribution (transport), and the regulation of cell death pathways. Mitochondrial dysfunction has mainly been reported in various non-dopaminergic cells and tissue samples from human patients as well as transgenic mouse and fruit fly models of PD. Thus, the mitochondria represent a highly promising target for the development of PD biomarkers. However, the limited amount of dopaminergic neurons prevented investigation of their detailed study. For the first time, we established human telomerase reverse transcriptase (hTERT)-immortalized wild type, idiopathic and Parkin deficient mesenchymal stromal cells (MSCs) isolated from the adipose tissues of PD patients, which could be used as a good cellular model to evaluate mitochondrial dysfunction for the better understanding of PD pathology and for the development of early diagnostic markers and effective therapy targets of PD. In this review, we examine evidence for the roles of mitochondrial dysfunction and increased OS in the neuronal loss that leads to PD and discuss how this knowledge further improve the treatment for patients with PD.
Animals ; Cell Death ; Diptera ; Dopaminergic Neurons ; Energy Metabolism ; Fruit ; Genetic Predisposition to Disease ; Humans ; Life Cycle Stages ; Mesenchymal Stromal Cells ; Mice ; Mice, Transgenic ; Mitochondria ; Neurons ; Oxidative Stress ; Parkinson Disease* ; Pathology ; Quality Control ; Substantia Nigra ; Telomerase ; Biomarkers

Parkinson's disease (PD) is an age-related progressive neurodegenerative disease associated with selective loss of dopaminergic neurons. The characteristic hallmark of the disease is intracytoplasmic proteinacious inclusion bodies called Lewy bodies, primarily consisting of a presynaptic protein alpha-synuclein. Oxidative stress-mediated damage to macromolecules have been shown to occur frequently in PD. Oxidative damage to DNA in the form of oxidized guanine (8-oxodG) accumulates in both the mitochondrial and nuclear DNA of dopaminergic neurons of the substantia nigra in PD. 8-oxodG-mediated transcriptional mutagenesis has been shown to have the potential to alter phenotype of cells through production of mutant pool of proteins. This review comprehensively summarizes the role of oxidative stress-mediated damage incurred during neurodegeneration, and highlights the scope of transcriptional mutagenesis event in leading to alpha-synuclein aggregation as seen in PD.
Amino Acid Sequence ; Animals ; Deoxyguanosine/*analogs & derivatives/metabolism ; Humans ; Molecular Sequence Data ; Mutagenesis ; *Oxidative Stress ; Parkinson Disease/*genetics/metabolism/pathology ; Protein Aggregation, Pathological/*genetics/metabolism/pathology ; Substantia Nigra/metabolism/*pathology ; Transcription, Genetic ; alpha-Synuclein/chemistry/*genetics

OBJECTIVETo discuss the protective effect of alkaloids from Piper longum (PLA) in rat dopaminergic neuron injury of 6-OHDA-induced Parkinson's disease and its possible mechanism.

METHODThe rat PD model was established by injecting 6-OHDA into the unilateral striatum with a brain solid positioner. The PD rats were divided into the PLA group (50 mg x kg(-1) x d(-1)), the madorpa group (50 mg x kg(-1) x d(-1)) and the model group, with 15 rats in each group. All of the rats were orally given drugs once a day for 6 weeks. Meanwhile, other 15 rats were randomly selected as the sham operation group, and only injected with normal saline in the unilateral striatum. The behavioral changes were observed with the apomorphine (APO)-induced rotation and rotary rod tests. The number of tyrosine hydroxylase (TH)-positive cells in rat substantia nigra and the density of TH-positive fibers in striatum were detected by tyrosine hydroxylase immunohistochemistry. The content of superoxide dismutase (SOD), glutathione peroxidase (GSH-Px), glutathione (GSH), catalase (CAT), malondialdehyde (MDA), nitric oxide (NO) and nitric oxide synthase (NOS) in rat substantia nigra and striatum were measured by the spectrophotometric method.

RESULTAfter being induced by APO, PD rats showed obvious rotation behaviors, with decreased time stay on rotary rod and significant reduction in the number of TH-positive cells in sustantia nigra and the density of TH-positive fibers in striatum. The activities of SOD, GSH-Px, CAT, the content of GSH and the total antioxidant capacity significantly decreased, whereas the activities of NOS and the content of MDA, NO significantly increased. PLA could significantly improve the behavioral abnormality of PD rats and increase the number of TH-positive cells in sustantia nigra and the density of TH-positive fibers in striatum. It could up-regulate the activities of SOD, GSH-Px, CAT, the content of GSH and the total antioxidant capacity, and decrease the content of NOS and the content of MDA, NO.

CONCLUSIONAlkaloids from P. longum shows the protective effect in substantia nigra cells of 6-OHDA-induced PD model rats. Its mechanism may be related with their antioxidant activity.

Administration, Oral ; Alkaloids ; administration & dosage ; pharmacology ; Animals ; Apomorphine ; pharmacology ; Catalase ; metabolism ; Dopamine Agonists ; pharmacology ; Dopaminergic Neurons ; drug effects ; metabolism ; pathology ; Glutathione ; metabolism ; Glutathione Peroxidase ; metabolism ; Male ; Malondialdehyde ; metabolism ; Motor Activity ; drug effects ; Neostriatum ; drug effects ; metabolism ; Nitric Oxide ; metabolism ; Nitric Oxide Synthase ; metabolism ; Oxidopamine ; Parkinson Disease, Secondary ; chemically induced ; physiopathology ; prevention & control ; Piper ; chemistry ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Substantia Nigra ; drug effects ; metabolism ; Superoxide Dismutase ; metabolism ; Tyrosine 3-Monooxygenase ; metabolism

BACKGROUNDParkinson's disease (PD) is a common neurodegenerative disease, which occurs mainly in the elderly. Recent studies have demonstrated that apoptosis plays an important role in the occurrence and development of PD. Subthalamic nucleus deep brain stimulation (STN-DBS) has been recognized as an effective treatment for PD. Recent clinical observations have shown that STN-DBS was able to delay early PD progression, and experiments in animal models have also demonstrated a protective effect of STN-DBS on neurons. However, the correlation between the neuron-protective effect of STN-DBS and the progression of substantia nigra pars compacta (SNc) neuronal apoptosis is still unknown. The aim of this study was to investigate the protective effect and potential mechanism of STN-DBS on SNc neurons in PD rats.

METHODSAfter the establishment of a PD rat model by unilateral/2-point injection of 6-hydroxydopamine in the medial forebrain bundle of the brain, DBS by implanting electrodes in the STN was administered. Behavioral changes were observed, and morphological changes of SNc neurons were analyzed by Nissl staining and DNA in situ end-labeling. Through extracellular recording of single neuron discharges and microelectrophoresis, the causes of and changes in SNc excitability during STN-DBS were analyzed, and the protective effect and potential mechanism of action of STN-DBS on SNc neurons in PD rats was investigated.

RESULTSSNc neuron apoptosis was significantly decreased (P < 0.05) in the stimulation group, compared with the sham stimulation PD group. Spontaneous discharges of SNc neurons were observed in normal rats and PD model rats, and the mean frequency of spontaneous discharges of SNc neurons in normal rats ((40.65 ± 11.08) Hz) was higher than that of residual SNc neurons in PD rats ((36.71 ± 9.23) Hz). Electrical stimulation of the STN in rats was associated with elevated excitation in unilateral SNc neurons. However, administering the gamma-aminobutyric acid receptor blocker, bicuculline significantly reduced SNc neuron excitation, but the change in SNc neuron excitation was not present when MK801, a glutamate receptor blocker, was administered.

CONCLUSIONSHigh-frequency stimulation of the STN has a protective effect on SNc neurons in PD rats. The possible molecular mechanism may be related to changes in the distribution and metabolism of neurotransmitters in the SNc region.

Animals ; Deep Brain Stimulation ; methods ; Disease Models, Animal ; Male ; Neurons ; cytology ; metabolism ; Parkinson Disease ; pathology ; therapy ; Rats ; Rats, Sprague-Dawley ; Substantia Nigra ; cytology

Parkinson's disease (PD) is characterized by selective and progressive degeneration of dopamine (DA)-producing neurons in the substantia nigra pars compacta (SNpc) and by abnormal aggregation of alpha-synuclein. Previous studies have suggested that DA can interact with alpha-synuclein, thus modulating the aggregation process of this protein; this interaction may account for the selective vulnerability of DA neurons in patients with PD. However, the relationship between DA and alpha-synuclein, and the role in progressive degeneration of DA neurons remains elusive. We have shown that in the presence of DA, recombinant human alpha-synuclein produces non-fibrillar, SDS-resistant oligomers, while beta-sheet-rich fibril formation is inhibited. Pharmacologic elevation of the cytoplasmic DA level increased the formation of SDS-resistant oligomers in DA-producing neuronal cells. DA promoted alpha-synuclein oligomerization in intracellular vesicles, but not in the cytosol. Furthermore, elevation of DA levels increased secretion of alpha-synuclein oligomers to the extracellular space, but the secretion of monomers was not changed. DA-induced secretion of alpha-synuclein oligomers may contribute to the progressive loss of the dopaminergic neuronal population and the pronounced neuroinflammation observed in the SNpc in patients with PD.
Blotting, Western ; Cell Line, Tumor ; Dopamine/*metabolism ; Humans ; Levodopa/pharmacology ; Neurons/*metabolism/pathology/*secretion ; Parkinson Disease/metabolism/pathology ; Substantia Nigra/metabolism/pathology ; alpha-Synuclein/*biosynthesis/*secretion

Oxidative stress is a hallmark in the pathogenesis of Parkinson disease (PD), which involves the selective loss of nigral dopaminergic neurons in PD. Resveratrol (3,5,4'-trihydroxy-trans-stilbene) is well known for its powerful antioxidant property and a wide range of other biological effects. In this study, we investigated the protective effect of resveratrol derived from Rhizoma Et Radix Polygoni Cuspidati and its liposomal form on the nigral cells of PD rats induced by unilateral microinjection of 6-hydroxy dopamine in the striatum. The results showed that after 14 days gavage of resveratrol and resveratrol liposome respectively (20 mg x kg(-1) WB per day), the abnormal rotational behavior of PD rats were deceased evidently, the numbers of total nigral cells, total nigral neurons and TH immuno-positive neurons were more than that of PD rats without given resveratrol or resveratrol liposome, simultaneously, the number of apoptotic nigral cells were decreased obviously. The results also showed that resveratrol and resveratrol liposome could decrease the total ROS activity, increase the total antioxidant capability of the nigral tissues. All the data indicated that resveratrol liposome performed stronger effects than resveratrol except for behavioral improvement. Our study confirmed that resveratrol derived from Rhizoma Et Radix Polygoni Cuspidati and its liposomal form could inhibit the loss of dopaminergic neurons of PD rats, the underlying mechanism may be attributed to their radical scavenging effect and antioxidant property. Due to presumably increased bioavailability, resveratrol liposome possesses the stronger therapeutic effect and may become a better clinical agent for the treatment of PD than free resveratrol.
Animals ; Antioxidants ; metabolism ; Behavior ; drug effects ; Cell Death ; drug effects ; Disease Models, Animal ; Drugs, Chinese Herbal ; pharmacology ; Liposomes ; pharmacology ; Neurons ; drug effects ; metabolism ; Neuroprotective Agents ; pharmacology ; Oxidative Stress ; drug effects ; Parkinson Disease ; drug therapy ; pathology ; Rats ; Rats, Wistar ; Reactive Oxygen Species ; metabolism ; Stilbenes ; pharmacology ; Substantia Nigra ; drug effects ; metabolism ; pathology

OBJECTIVETo explore the effect of Bushen Huoxue Decoction (BHD) on the orphan receptor (Nurr1) and tyrosine hydroxylase (TH) in the brain of rats with Parkinson's disease (PD).

METHODSOne hundred and twenty SD rats were divided into 100 in the model group and 20 in the normal control group, fifty-eight SD rats from the model group, established into PD model successfully by injuring their substantia nigra (SSN) with 6-hydroxydopamine, were divided equally into the model group and the test group, and they were treated with saline and BHD, respectively, for eight successive weeks. The change in the rats' behavior before and after treatment was observed by counting the cycles of rotation induced by apomorphine injection; the pathology of neurons, level of Nurr1 mRNA expression, and amount of TH positive cells in SSN were observed after treatment.

RESULTSThe rats' behavior was improved in the tested group significantly, the rotation cycle after treatment being 84.0 ± 20.0 cycles/40 min, which was significantly lower than that in the model group (377.0 ± 62.3 cycles/40 min, P<0.01). Besides, the Nurr1 mRNA expression and TH positive cell in the test group were 0.97 ± 0.15 and 49.40 ± 14.72, respectively, which were significantly higher than those in the model group, 0.22 ± 0.03 and 5.45 ± 2.58, respectively (all P<0.01).

CONCLUSIONBHD could treat PD by enhancing the Nurr1 mRNA expression, increasing the TH content in brain, and promoting the repairing of injured neuron in cerebral SSN.

Animals ; Behavior, Animal ; drug effects ; Brain ; drug effects ; enzymology ; pathology ; Drugs, Chinese Herbal ; pharmacology ; therapeutic use ; Female ; Gene Expression Regulation ; drug effects ; Neurons ; drug effects ; enzymology ; pathology ; Nuclear Receptor Subfamily 4, Group A, Member 2 ; genetics ; metabolism ; Parkinson Disease ; drug therapy ; enzymology ; pathology ; RNA, Messenger ; genetics ; metabolism ; Rats ; Rats, Sprague-Dawley ; Substantia Nigra ; drug effects ; metabolism ; pathology ; Tyrosine 3-Monooxygenase ; metabolism