Application of osmundone in medicine with neuroprotective effect on Parkinson's disease

Abstract

The invention belongs to the technical field of medicines, and particularly discloses application of osmundone in a medicine with a neuroprotective effect on Parkinson's disease, which is specifically characterized in that an alpha-Syn plasmid is transfected into a mouse dopaminergic neuron MES23.5 cell to establish an alpha-Syn overexpressed PD cell model, and the PD cell is intervened by adding the osmundone, so that the neuroprotective effect on Parkinson's disease is achieved. Results prove that osmundone can improve cell viability and reduce cytotoxicity by reducing mitochondrial damage of PD cells, and has a neuroprotective effect on PD damaged neurons; a PD mouse model is established by utilizing MPTP intraperitoneal injection, intervention is carried out through osmundone intragastric administration, and a result proves that osmundone can effectively improve the athletic ability of a PD mouse. The results show that osmundone can effectively play a neuroprotective role on PD, and has a good clinical application prospect.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, and particularly relates to the application of osmundacetone in drugs for neuroprotection against Parkinson's disease. Background Art

[0002] The main pathological feature of Parkinson's disease (PD) is the progressive loss of dopaminergic neurons in the substantia nigra (SN) of the midbrain, which leads to a decrease in dopamine levels in the striatum and subsequently triggers a series of movement disorders. In addition, Lewy body-like structures will form in the remaining neurons, and these structures are mainly composed of abnormal aggregation of α-synuclein (α-Syn). Traditional PD animal models mainly use various neurotoxins, such as 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), rotenone, paraquat, 6-hydroxydopamine, etc., to cause selective damage to dopaminergic neurons. Such animal models well simulate the damage of dopaminergic neurons in the substantia nigra of PD and the motor symptoms, but lack the progressive Lewy body-like pathological changes. In recent years, more and more evidence has shown that the abnormal increase of α-Syn and the resulting mitochondrial dysfunction play a key role in the pathogenesis of PD. The specific mechanisms include: the presence of α-Syn in mitochondria: part of α-Syn is endogenously or overexpressed in mitochondria, which may lead to the down-regulation of mitochondrial complex I activity and then cause mitochondrial damage. Mitochondrial morphological disruption: in α-Syn transgenic mice, the morphology of brain mitochondria is disrupted, which may affect the normal function of mitochondria. Mitochondrial fragmentation and dysfunction: α-Syn may exist in the form of small polymers, inducing mitochondrial fragmentation, which may lead to subsequent mitochondrial dysfunction and neuronal death.

[0003] Mitochondria are the main sites for generating ATP in animal and plant cells and are crucial for promoting cell energy conversion and participating in processes such as apoptosis. Mitochondria store energy in the form of electrochemical potential energy in the inner mitochondrial membrane through the respiratory oxidation process to form the mitochondrial membrane potential (MMP).

[0004] In view of the key role of the abnormal increase of α-Syn and the resulting mitochondrial dysfunction in the pathogenesis of PD, drugs that reduce the production of α-Syn, promote its clearance or prevent its abnormal aggregation may become a new direction for the treatment of PD, or improving the function of mitochondria through drugs or other means may help delay the progression of PD.

[0005] Osmundacetone (OAC) is a phenolic active ingredient in Phellinus linteus. According to reports at home and abroad, the pharmacological activity research of OAC is scarce and not in-depth. Shunyan Mo et al. detected that OAC has good cytotoxic effects on four tumor cell lines, MCF-7, Bel-7402, Ketr3, and HCT-8, by the MTT method. Myeong-Seok Lee et al. reported that OAC has strong scavenging activity against free radicals. The key role of OAC in NSCLC mitochondrial energy metabolism is to inhibit cell proliferation and tumor growth by downregulating GLUD1 to inhibit glutamate metabolism, thereby reducing the generation of energy metabolism substrates and inhibiting oxidative phosphorylation. The research results related to the nervous system show that OAC improves Alzheimer's disease-like pathology by inhibiting β-amyloid fibrillization, oxidative damage, and neuroinflammation in APP / PS1 transgenic mice. However, so far, there is no relevant research on whether OAC has a protective effect on PD nerve injury. We propose the hypothesis that OAC can reduce the cell damage of PD dopaminergic neurons and relieve the motor dysfunction of PD by inhibiting mitochondrial damage. For clarifying whether OAC can play a protective role in PD nerve injury by alleviating mitochondrial dysfunction, and then the application of OAC in the preparation of drugs with a protective effect on PD nerve injury has good market prospects. Summary of the Invention

[0006] The object of the present invention is to provide an application of osmundacetone in drugs for the neuroprotection of Parkinson's disease. Specifically, osmundacetone reduces the cell damage and cytotoxicity of dopaminergic neuron MES23.5 cells induced by overexpressed α-Syn protein, improves the mitochondrial dysfunction caused by overexpressed α-Syn protein in dopaminergic neuron MES23.5, relieves the motor disorder of Parkinson's disease mice, and then plays a protective role in Parkinson's disease nerve injury.

[0007] In order to achieve the above object, the technical solution adopted by the present invention is as follows:

[0008] One object of the present invention is to provide an application of osmundacetone in drugs for the neuroprotection of Parkinson's disease.

[0009] Further, the osmundacetone plays a protective role in neuron injury.

[0010] Further, the osmundacetone reduces the cell damage and cytotoxicity of dopaminergic neuron MES23.5 cells induced by overexpressed α-Syn protein, improves the mitochondrial dysfunction caused by overexpressed α-Syn protein in dopaminergic neuron MES23.5, relieves the motor disorder of Parkinson's disease mice, and then plays a protective role in Parkinson's disease nerve injury.

[0011] Another object of the present invention is to provide a drug with neuroprotective effect on Parkinson's disease, and the main component of the drug is osmundacetone.

[0012] Further, the effective concentration of osmundacetone for reducing the damage and cytotoxicity of dopaminergic neuron MES23.5 cells induced by overexpression of α-Syn protein in vitro is 1 μM - 4 μM.

[0013] Further, the effective concentration of osmundacetone for improving the motor ability of PD model mice induced by MPTP in vivo is 1 mg / kg - 8 mg / kg.

[0014] Further, the drug is prepared into a clinically acceptable pharmaceutical preparation with osmundacetone as the main component, plus pharmaceutically acceptable excipients or auxiliary components.

[0015] Further, the pharmaceutical preparation of the drug includes any one of tablets, pills, capsules, granules, syrups, powders, powders, suppositories, drops, emulsions, solutions, suspensions.

[0016] The third object of the present invention is to provide a pharmaceutical composition for treating and / or preventing Parkinson's disease, and the pharmaceutical composition is prepared into a clinically acceptable pharmaceutical preparation with osmundacetone as the main component, plus pharmaceutically acceptable excipients or auxiliary components.

[0017] Further, the content of osmundacetone in the pharmaceutical composition is 1 mg / kg - 8 mg / kg.

[0018] Further, the pharmaceutical preparation of the drug includes any one of tablets, pills, capsules, granules, syrups, powders, powders, suppositories, drops, emulsions, solutions, suspensions.

[0019] Generally speaking, as a drug, it is usually clinically applied after being prepared into a preparation. The pharmaceutical composition of the present invention can be prepared according to the methods well known in the art. The pharmaceutical composition of the present invention can be combined with one or more pharmaceutically acceptable solid or liquid excipients and / or adjuvants to form any dosage form suitable for human or animal use.

[0020] The pharmaceutical composition of the present invention or the pharmaceutical composition containing it can be administered in unit dosage form, and the administration route can be enteral or parenteral, such as oral, intravenous injection, intramuscular injection, subcutaneous injection, nasal cavity, oral mucosa, eye, lung and respiratory tract, skin, vagina, rectum, etc.

[0021] The dosage form for administration can be a liquid dosage form, a solid dosage form or a semi-solid dosage form. The liquid dosage form can be a solution (including true solution and colloidal solution), an emulsion (including o / w type, w / o type and multiple emulsion), a suspension, an injection (including aqueous injection, powder injection and infusion), an eye drop, a nasal drop, a lotion, a liniment, etc.; the solid dosage form can be a tablet (including ordinary tablet, enteric-coated tablet, buccal tablet, dispersible tablet, chewable tablet, effervescent tablet, orally disintegrating tablet), a capsule (including hard capsule, soft capsule, enteric-coated capsule), a granule, a powder, a pellet, a dropping pill, a suppository, a film, a patch, an aerosol (powder aerosol), a spray, etc.; the semi-solid dosage form can be an ointment, a gel, a paste, etc.

[0022] The pharmaceutical composition of the present invention can be made into an ordinary preparation, or can also be made into a sustained-release preparation, a controlled-release preparation, a targeted preparation and various particulate drug delivery systems. In order to make the pharmaceutical composition of the present invention into a tablet, various excipients well-known in the art can be widely used, including diluents, binders, wetting agents, disintegrants, lubricants, glidants. The diluent can be starch, dextrin, sucrose, glucose, lactose, mannitol, sorbitol, xylitol, microcrystalline cellulose, calcium sulfate, calcium hydrogen phosphate, calcium carbonate, etc.; the wetting agent can be water, ethanol, isopropanol, etc.; the binder can be starch paste, dextrin, syrup, honey, glucose solution, microcrystalline cellulose, gum arabic paste, gelatin paste, sodium carboxymethyl cellulose, methyl cellulose, hydroxypropyl methyl cellulose, ethyl cellulose, acrylic resin, carbomer, polyvinylpyrrolidone, polyethylene glycol, etc.; the disintegrant can be dry starch, microcrystalline cellulose, low-substituted hydroxypropyl cellulose, cross-linked polyvinylpyrrolidone, cross-linked sodium carboxymethyl cellulose, sodium carboxymethyl starch, sodium bicarbonate and citric acid, polyoxyethylene sorbitan fatty acid ester, sodium lauryl sulfate, etc.; the lubricant and glidant can be talc powder, silicon dioxide, stearate, tartaric acid, liquid paraffin, polyethylene glycol, etc.

[0023] The tablet can be further made into a coated tablet, such as a sugar-coated tablet, a film-coated tablet, an enteric-coated tablet, or a bilayer tablet and a multilayer tablet.

[0024] In order to make the dosage unit into a capsule, the active ingredient, the pharmaceutical composition of the present invention, can be mixed with a diluent and a glidant, and the mixture can be directly placed into a hard capsule or a soft capsule. Alternatively, the active ingredient, the pharmaceutical composition of the present invention, can be first made into granules or pellets with a diluent, a binder and a disintegrant, and then placed into a hard capsule or a soft capsule. The varieties of each diluent, binder, wetting agent, disintegrant, glidant used for preparing the tablet of the pharmaceutical composition of the present invention can also be used for preparing the capsule of the pharmaceutical composition of the present invention.

[0025] To prepare the pharmaceutical composition of the present invention into an injection, water, ethanol, isopropanol, propylene glycol or a mixture thereof can be used as a solvent, and appropriate solubilizers, cosolvents, pH adjusters, and osmotic pressure regulators commonly used in the art can be added. The solubilizer or cosolvent can be poloxamer, lecithin, hydroxypropyl-β-cyclodextrin, etc.; the pH adjuster can be phosphate, acetate, hydrochloric acid, sodium hydroxide, etc.; the osmotic pressure regulator can be sodium chloride, mannitol, glucose, phosphate, acetate, etc. When preparing a freeze-dried powder injection, mannitol, glucose, etc. can also be added as a bulking agent.

[0026] In addition, if necessary, coloring agents, preservatives, fragrances, flavoring agents or other additives can also be added to the pharmaceutical preparation. In summary, due to the adoption of the above technical solutions, the beneficial effects of the present invention are:

[0027] The application of the osmundacetone of the present invention in drugs for the neuroprotective effect of Parkinson's disease. Specifically, an α-Syn overexpressing PD cell model was established by transfecting α-Syn plasmid into mouse dopaminergic neuron MES23.5 cells, and Parkinson's disease (PD) cells were intervened by adding osmundacetone (OAC). The cell viability and cytotoxicity damage of PD cells were detected by cell viability experiments and LDH release experiments to evaluate the effect of OAC on the cell viability of PD cells; the mitochondrial membrane potential (MMP) of cells was detected by Mito-Tracker staining experiments to evaluate the effect of OAC on the mitochondrial function of cells. The experimental results showed that treatment with 2 μM OAC could effectively increase the cell viability of PD cells, reduce cytotoxicity, and effectively increase the mitochondrial membrane potential of PD cells, rescuing mitochondrial damage. The above results proved that OAC could increase cell viability and reduce cytotoxicity by reducing mitochondrial damage in PD cells, playing a neuroprotective role in PD damaged neurons. A PD mouse model was established by intraperitoneal injection of MPTP, and PD model mice were intervened by gavage with OAC. Further, the effect of OAC on the motor ability of mice was evaluated by open field experiments and rotarod experiments. The results of animal experiments found that 8 mg / kg OAC could significantly improve the average movement speed of PD model mice in open field experiments and increase the falling rod latency of PD mice in rotarod experiments, indicating that osmundacetone could effectively improve the motor function of Parkinson's disease mice. Therefore, osmundacetone has good clinical application prospects for the treatment and / or prevention of Parkinson's disease. Brief Description of the Drawings

[0028] Figure 1 It is a diagram for establishing an α-Syn overexpressing PD cell model of the present invention. Among them Figure 1Figure A shows the protein electrophoresis results of MES23.5 cells transfected with NC and SNCA plasmids for 48 h. Figure B shows the comparison results of the relative expression levels of α-Syn protein in the cell proteins of each group. In the figure, Con: blank cell group; NC: group transfected with NC plasmid; SNCA: group transfected with α-Syn plasmid; α-Syn: α-synuclein; β-actin: β-actin. n = 3, ***P < 0.001.

[0029] Figure 2 Figure A shows the statistical chart of the significant inhibition of cytotoxic damage of PD model cells by Osmundacetone (OAC) of the present invention. Among them, Figure A shows the results of the cell viability detection experiment, showing the relative cell viability of each group of cells. Figure B shows the results of the cytotoxicity detection experiment, showing the percentage of LDH release of each group of cells. In the figure, Con: blank cell group; NC: group transfected with NC plasmid; SNCA: each group transfected with α-Syn plasmid. n = 3, *P < 0.05, ***P < 0.001, ****P < 0.0001.

[0030] Figure 3 Figure and statistical chart show the significant inhibition of mitochondrial damage of PD model cells by Osmundacetone (OAC) of the present invention.

[0031] Among them, Figure A shows the staining results of MitoTracker and Hoechst33342 of each group of cells. Figure B shows the comparison of the relative fluorescence intensity values of MitoTracker staining of each group of cells. In the figure, Con: blank cell group; NC: group transfected with NC plasmid; SNCA: group transfected with α-Syn plasmid; SNCA + OAC: group transfected with α-Syn plasmid and intervened with OAC. n = 3, *P < 0.05, ****P < 0.0001.

[0032] Figure 4 Figure shows the results of the significant improvement of the motor ability of PD model mice by Osmundacetone (OAC) of the present invention. Among them, Figure A shows the open field movement trajectory diagrams of each group of mice. Figure B shows the statistical chart of the average movement speed of the open field experiment of each group of mice. Figure C shows the statistical chart of the falling rod latency of the rotarod test of each group of mice. In the figure, Con: control group; PD: Parkinson's disease group; OAC: Osmundacetone. n = 5, *P < 0.05, ***P < 0.001, ****P < 0.0001. Detailed implementation manners

[0033] To make the above objects, features, and advantages of the present invention more apparent and understandable, the following detailed description of the specific embodiments of the present invention will be provided in conjunction with the accompanying drawings. Many specific details are set forth in the following description to facilitate a full understanding of the present invention. However, the present invention can be implemented in many other ways different from those described herein, and those skilled in the art can make similar improvements without departing from the spirit of the present invention. Therefore, the present invention is not limited by the specific embodiments disclosed below.

[0034] Experiment:

[0035] 1. Materials and Methods

[0036] 1.1 Cell Culture and Treatment

[0037] Establishment of PD cell model: Mouse dopaminergic neurons (MES23.5 cells) were cultured in DMEM / F12 medium (Gibco, New York, USA) containing 10% fetal bovine serum and Sato's. 2 μg of α-Syn plasmid or empty vector (NC) plasmid was transfected into each well of a 6-well plate using Lipofectamine 3000 (Gibco, New York, USA). The cells were divided into 3 groups: Con group, NC group, and SNCA group. The cells were treated for 48 h, and subsequent Western blotting was performed for detection.

[0038] Drug intervention: 100 ng of α-Syn plasmid (SNCA) or empty vector (NC) plasmid was transfected into each well of a 96-well plate using Lipofectamine 3000. After 24 h of plasmid transfection, OAC dissolved in DMSO (Sigma, MO, USA) was added to the cell culture medium overexpressing α-Syn protein, and the final concentrations were 0 μM, 1 μM, 2 μM, and 4 μM, respectively. The cells were divided into 6 groups: Con group, NC group, SNCA-OAC 0 μM group, SNCA-OAC 1 μM group, SNCA-OAC 2 μM group, and SNCA-OAC 4 μM group. The cells were continuously treated for 24 h, and subsequent cell viability detection, cytotoxicity detection, and mitochondrial membrane potential detection were performed.

[0039] 1.2 Western blotting experiment

[0040] Collect and lyse the cells treated differently according to the instructions of the Cellular Protein Preparation Kit (Applygen, Beijing, China). The cells are separated and aggregated into different tubes. The protein concentration is evaluated using the BCA Protein Quantification Kit (Thermo Fisher Scientific, Massachusetts, USA). Then, 20 μg of protein from each fraction is separated by SDS-PAGE and transferred to a PVDF membrane (Sigma, MO, USA). Then, it is blocked with 5% non-fat milk powder, and anti-α-Syn and anti-β-actin antibodies (Abcam, Cambridge, UK) are added respectively and incubated overnight. The corresponding fluorescent secondary antibody is combined with the primary antibody at room temperature for 1 h. After four washes, the membrane is scanned and imaged with an ODYSSEY imaging system (LI-COR, Nebraska, USA).

[0041] 1.3 Cell viability detection

[0042] Use CellTiter AQueous One Solution Cell Proliferation Assay kit (Promega, WI, USA) to detect cell viability. Aspirate the old medium from the cells in the 96-well plate and replace it with 100 μL of fresh medium per well. Add 20 μL of CellTiter AQueous One Solution Reagent to the treated cells and incubate at 37 °C in a 5% CO 2 environment for 1 h. Read the absorbance value at a wavelength of 490 nm using an enzyme-linked immunosorbent assay (ELISA) reader.

[0043] 1.4 Cytotoxicity detection

[0044] Use CytoTox Non-Radioactive Cytotoxicity Assay kit (Promega, WI, USA) to detect the LDH release and evaluate cytotoxic damage. Take 50 μL of the old medium from all test wells and control wells and transfer it to a clean 96-well plate. Add 50 μL of CytoTox Reagent and incubate in the dark at room temperature for 30 min. Read the absorbance value at a wavelength of 490 nm using an ELISA reader.

[0045] 1.5 Mitochondrial membrane potential (MMP) detection

[0046] The mitochondrial membrane potential (MMP) of cells was detected using a mitochondrial membrane potential detection kit (Beyotime, Shanghai, China). The old culture medium in the 96-well plate was aspirated, and the cells were washed once with PBS. Then, 96.5 μL of binding buffer was added to each well, followed by 2 μL of Mito-Tracker Red CMXRos staining solution and 5 μL of Hoechst 33342 staining solution. The mixture was gently vortexed. After incubating at room temperature in the dark for 20 min, the cells were observed under a fluorescence microscope. MitoTracker Red CMXRos emits red fluorescence, and Hoechst 33342 emits blue fluorescence.

[0047] Establishment of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP)-induced Parkinson's disease (PD) model and intervention with oleanolic acid (OAC)

[0048] Twenty-five 8-week-old male C57 / BL6N mice were purchased from Cyagen Biosciences (Taicang) Co., Ltd. (Production License No.: SCXK (Jiangsu) 2018-0003) and housed in the SPF animal facility of Guilin Medical University. The mice had free access to food and water and were maintained on a 12-h light / dark cycle. The mice were randomly divided into five groups of five mice each. Four of the groups were intraperitoneally injected with MPTP for 7 days to establish a PD mouse model. Three of these groups were pre-treated with intragastric administration of OAC at doses of 1 mg / kg, 4 mg / kg, and 8 mg / kg for 7 days and continuously administered OAC by gavage for 7 days during the PD modeling process to establish PD models with different concentrations of OAC intervention, namely the PD + 1 mg / kg OAC group, the PD + 4 mg / kg OAC group, and the PD + 8 mg / kg OAC group. The fourth group was pre-treated with intragastric administration of the same volume of normal saline for 7 days and continuously administered normal saline by gavage for 7 days during the modeling process as the PD group. The fifth group was pre-treated with intragastric administration of normal saline for 14 days and received intraperitoneal injection of normal saline starting from day 8 as the control (Con) group. After the modeling, the motor ability of the mice in each group was detected by behavioral tests. The open-field test was used to evaluate the spontaneous motor ability of the mice, that is, the spontaneous movement trajectory and distance of the mice within 5 min were recorded, and the average movement speed was calculated based on the distance and time. The rotarod test was used to detect the coordinated motor ability of the mice. First, the mice were trained at a speed of 4 rpm / min for 3 times, 2 min each time. Then, the rotarod speed was adjusted to accelerate from 4 rpm / min to 40 rpm / min within 5 min, and the time experienced by each mouse before falling off the rotarod, that is, the fall latency of the rotarod, was recorded.

[0049] 1.7 Statistical analysis

[0050] Data are expressed as the mean ± standard deviation. Statistical analysis was performed using GraphPad Prism software version 9.0 (GraphPad, California, USA). One-way analysis of variance (ANOVA) was used to evaluate the differences between groups, followed by Tukey's multiple comparison test. A P value < 0.05 was considered statistically significant.

[0051] 2. Results

[0052] 2.1 Establishment of PD model cell model

[0053] The WB results showed that compared with the Con group (100%) and the NC group (99.03%), the expression of α-Syn in the α-Syn transfection group (SNCA group, 2565.77%) was significantly increased (P<0.001), indicating that the PD cell model with overexpression of α-Syn was successfully established ( Figure 1 ).

[0054] 2.2 OAC significantly inhibits the cytotoxic damage of PD model cells

[0055] The results of cell viability detection and LDH release experiment showed that compared with the Con group (cell viability 100%, LDH release 100%) and the NC group (cell viability 98.56%, LDH release 94.36%), the cell viability (47.30%) of the α-Syn transfection group (SNCA-OAC 0 μM group) was significantly decreased, and the LDH release (158.10%) was significantly increased (P<0.0001, P<0.001). Compared with the SNCA-OAC 0 μM group, after treatment with 1 μM OAC, the cell viability increased to 52.41%, and the LDH release decreased to 137.03%; after treatment with 2 μM OAC, the cell viability increased to 67.45%, and the LDH release decreased to 117.79%; after treatment with 4 μM OAC, the cell viability increased to 61.81%, and the LDH release decreased to 156.82%. Among them, the treatment effect of 2 μM OAC was the best (P<0.05, P<0.05)( Figure 2 A, B).

[0056] 2.3 OAC significantly inhibits the mitochondrial function damage of PD model cells

[0057] The results of mitochondrial membrane potential detection showed that compared with the Con group (100%) and the NC group (89.22%), the mitochondrial membrane potential (MitoTracker fluorescence intensity, 33.98%) of the α-Syn transfection group (SNCA group) was significantly decreased (P<0.0001). Compared with the SNCA group, after treatment with 2 μM OAC, the MitoTracker fluorescence intensity (64.49%) was significantly increased (P<0.05)( Figure 3 A, B).

[0058] 2.4 OAC significantly improves the motor ability of PD model mice

[0059] The results of the open field test showed that compared with the Con group, the movement trajectories of the mice in the PD group were significantly sparser and concentrated around the four sides, while the trajectories of the mice intervened with different concentrations of OAC gradually became denser, especially in the mice in the OAC intervention group at 8 mg / kg ( Figure 4 A). Compared with the Con group (64.71 mm / s), the open field movement speed of the mice in the PD group decreased significantly to 31.76 mm / s (P < 0.0001); while the average open field movement speed of the mice in the 1 mg / kg OAC intervention group increased to 36.00 mm / s, the average open field movement speed of the mice in the 4 mg / kg OAC intervention group increased to 43.58 mm / s, and the average open field movement speed of the mice in the 8 mg / kg OAC intervention group increased to 48.37 mm / s. Among them, the average open field movement speed of the mice in the 8 mg / kg OAC intervention group increased most significantly (P < 0.05). In addition, the results of the rotarod test showed that compared with the Con group (265.8 s), the falling rod latency of the mice in the PD group in the rotarod test was significantly shortened to 133.8 s (P < 0.001); while the falling rod latency of the mice in the 1 mg / kg OAC intervention group in the rotarod test increased to 143.8 s, the falling rod latency of the mice in the 4 mg / kg OAC intervention group in the rotarod test increased to 182.6 s, and the falling rod latency of the mice in the 8 mg / kg OAC intervention group in the rotarod test increased to 224.2 s. Among them, the falling rod latency of the mice in the 8 mg / kg OAC intervention group in the rotarod test increased most significantly (P < 0.05)( Figure 4 B, C).

[0060] In summary, the treatment with 2 μM osmundacetone can effectively increase the cell viability of PD cells, reduce the cytotoxicity, and effectively increase the mitochondrial membrane potential of PD cells, and rescue the mitochondrial damage; the treatment with 8 mg / kg osmundacetone can effectively increase the average open field movement speed of PD mice and prolong the falling rod latency of the rotarod test. The above results prove that osmundacetone can increase the cell viability, reduce the cytotoxicity, and improve the motor ability of PD mice by reducing the mitochondrial damage of PD cells, and has a neuroprotective effect on PD, with good clinical application prospects.

[0061] The above-described embodiments only represent several implementation manners of the present invention. The description is relatively specific and detailed, but it should not be construed as a limitation on the scope of the present invention. It should be noted that for those of ordinary skill in the art, without departing from the concept of the present invention, several deformations and improvements can still be made, and these all belong to the protection scope of the present invention. Therefore, the protection scope of the present invention should be subject to the appended claims.

Claims

1. Application of purpurogenol as a neuroprotective drug for Parkinson's disease.

2. The use of porphyrin in a neuroprotective drug for Parkinson's disease according to claim 1, characterized in that: The porphyrin plays a protective role in neuronal damage.

3. The use of porphyrin in a neuroprotective drug for Parkinson's disease according to claim 1, characterized in that: The purpurogenol reduces the damage and cytotoxicity of dopaminergic neuron MES23.5 cells induced by overexpression of α-Syn protein, improves the mitochondrial dysfunction caused by overexpression of α-Syn protein in dopaminergic neuron MES23.5, alleviates the movement disorder of Parkinson's disease mice, and thus plays a protective role in Parkinson's disease nerve damage.

4. A drug for protecting Parkinson's disease nerves, characterized in that: The main component of the medicine is porphyrone.

5. The drug for protecting Parkinson's disease nerves according to claim 4, characterized in that: The effective concentration of purpurogenol in reducing the damage and cytotoxicity of dopaminergic neuron MES23.5 cells induced by overexpression of α-Syn protein in vitro is 1 μM-4 μM.

6. The drug for protecting Parkinson's disease nerves according to claim 4, characterized in that: The medicine is prepared into a clinically acceptable medicine preparation with porphyrin as the main component and pharmaceutically acceptable auxiliary materials or auxiliary components.

7. The drug for protecting Parkinson's disease nerves according to claim 4, characterized in that: The preparation of the drug includes any one of tablets, pills, capsules, granules, syrups, powders, granules, suppositories, drops, emulsions, solutions and suspensions.

8. A pharmaceutical composition for treating and / or preventing Parkinson's disease, characterized in that: The pharmaceutical composition is prepared from porphyrin as a main component and pharmaceutically acceptable auxiliary materials or auxiliary components to form a clinically acceptable pharmaceutical preparation.

9. The pharmaceutical composition for treating and / or preventing Parkinson's disease according to claim 8, characterized in that: The content of porphyrone in the pharmaceutical composition is 1 mg / kg-8 mg / kg.

10. The pharmaceutical composition for treating and / or preventing Parkinson's disease according to claim 8, characterized in that: The preparation of the drug includes any one of tablets, pills, capsules, granules, syrups, powders, granules, suppositories, drops, emulsions, solutions and suspensions.

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