Use of α-ketoglutaric acid in preparation of drug for preventing and treating demyelination-related diseases

Abstract

The use of α-ketoglutaric acid in the preparation of a drug for preventing and treating demyelination-related diseases. The α-ketoglutaric acid or a derivative thereof is used for preventing and / or treating myelin sheath defects in demyelination-related diseases, such as demyelinating diseases, amyotrophic lateral sclerosis, Huntington's disease, hypomyelinating leukodystrophy, Alzheimer's disease, Parkinson's syndrome and diabetes-related visual impairment, and can promote myelin sheath generation, regeneration or repair, and improve the immune microenvironment in the lesion area.

Description

Application of α-ketoglutaric acid in the preparation of drugs for preventing and treating demyelination-related diseases Technical Field

[0001] The present invention relates to the use of α-ketoglutaric acid in preparing drugs for preventing and treating demyelination-related diseases, and specifically relates to the use of α-ketoglutaric acid or its derivatives in preparing drugs for preventing and treating demyelination-related diseases, as well as drugs for preventing and treating demyelination-related diseases, belonging to the field of biomedicine technology. Background Art

[0002] Myelin defects occur in many neurological diseases, typically demyelinating diseases (such as multiple sclerosis), neurodegenerative diseases (such as Alzheimer's disease and Parkinson's disease), and hypomyelinated leukodystrophy. In addition to some genetic factors, other environmental factors, including trauma, smoking, drug abuse, and infection, can increase the risk of developing the above diseases. The main cause of hypomyelinated leukodystrophy is genetic defects, including PLP1, GJA1, GJC2, which are all high-risk genes for the disease. In multiple sclerosis, a large number of immune cells, including B cells and T cells, are enriched in the affected area, causing an excessive immune response and thus damaging the myelin sheath.

[0003] Currently, most drugs used to treat multiple sclerosis are anti-immune drugs, including CD20 antibodies targeting B cells such as ocrelizumab and ofatumumab, and lymphocyte blockers such as fingolimod. There is currently no specific treatment for hypomyelinated leukodystrophy. Multiple sclerosis, senile demyelination, and hypomyelinated leukodystrophy all involve impaired oligodendrocyte differentiation. Although anti-immune therapy can effectively alleviate the symptoms of multiple sclerosis, myelin damage is difficult to restore, and anti-immune therapy alone cannot effectively promote myelin regeneration. Therefore, the treatment of demyelinating diseases still needs to focus on how to promote remyelination. Drugs currently under development to promote myelin regeneration include liothyronine, bexarotene, and GSK239512, but their current effectiveness has been less than ideal.

[0004] In addition to multiple sclerosis, Alzheimer's disease also presents with a decrease in overall myelin levels. Studies have shown that enhancing myelin formation through genetic means can alleviate Alzheimer's-related cognitive impairment. Currently, new drug development for the treatment of Alzheimer's disease focuses on the following: eliminating Aβ deposits (Aducanumab, Lecanemab, etc.), inhibiting tau protein neurofibrillary tangles (Cambino, Semorinemab, etc.), replenishing neurotransmitters (L-serine, Atomoxetine, etc.), inhibiting neuroinflammation (dasatinib + quercetin, etanercept, etc.), and regulating lipid metabolism (efavirenz, etc.). However, drugs that enhance myelin formation have not yet been used in Alzheimer's disease.

[0005] Optic nerve / retinal diseases caused by conditions such as glaucoma and traumatic brain injury also severely impact patients' vision and can even cause irreversible blindness. However, the visual changes associated with these diseases remain understudied. Studies have found that the onset of glaucoma is accompanied by significant optic nerve demyelination, including decreased MBP markers and a reduction in oligodendrocyte numbers. Demyelination also precedes axonal degeneration, suggesting that demyelination is an early marker of optic nerve dysfunction in glaucomatous eyes. During the pathophysiology of glaucoma, abnormal activation of retinal microglia produces neurotoxic and inflammatory factors, further accelerating retinal neuronal apoptosis and optic nerve myelin damage. Current treatments for glaucomatous optic neuropathy include ocular hypotensive drugs (e.g., β-adrenergic receptor blockers, Rho kinase inhibitors), and optic nerve protective drugs (e.g., calcium channel blockers, neurotrophic factors). However, no currently available drugs address glaucomatous optic nerve myelin damage and immune regulation. Therefore, developing new treatments is necessary for the large number of patients with vision loss caused by glaucoma.

[0006] There is also a direct link between diabetic visual impairment and pathological changes in the optic nerve myelin sheath, and oligodendrocyte differentiation disorders during the course of the disease are the main cause of pathological changes in the myelin sheath. Impaired optic nerve neomyelination may be an important cause of early diabetic visual dysfunction, and drugs that promote oligodendrocyte differentiation can rescue visual dysfunction. Currently, drugs for diabetic optic neuropathy are still under research. Although clemastine used in some studies can promote oligodendrocyte differentiation, it can cause side effects such as drowsiness and loss of appetite in clinical use. Therefore, it is necessary to develop safer therapeutic drugs for diabetes-related visual impairment and myelin damage.

[0007] Furthermore, cancer patients also experience white matter lesions after chemotherapy and radiotherapy, manifesting as abnormal microglial activation and focal demyelination, clinically manifesting as significant cognitive impairment. For example, during the treatment of nasopharyngeal carcinoma, radiotherapy requires prophylactic irradiation of the middle cranial fossa within the radiation field. This radiation dose, combined with the inherent radiation dose exceeding the acceptable dose for normal brain tissue, inevitably causes damage to the white matter of normal brain tissue. Another example is methotrexate, oxaliplatin, and fluorouracil, which can also cause white matter lesions during cancer treatment. Currently, little research has been conducted on the mechanisms of white matter damage after chemoradiotherapy and radiotherapy for cancer. Furthermore, chemoradiotherapy and radiotherapy can induce persistent microglial activation in the brain, disrupting various mechanisms that promote neuroplasticity and leading to treatment-related cognitive impairment. The mechanisms underlying abnormal microglial activation in cancer patients after chemoradiotherapy and radiotherapy remain unclear and require further investigation. Therefore, promoting myelin regeneration and improving the immune microenvironment in the central nervous system after chemoradiotherapy and radiotherapy may help alleviate cognitive impairment in cancer patients after treatment. This could lead to the development of more effective and safer drugs that promote myelin regeneration and immunomodulation.

[0008] α-ketoglutaric acid is an intermediate metabolite of the tricarboxylic acid cycle in organisms and participates in the synthesis of amino acids, vitamins, and organic acids, as well as in energy metabolism. Therefore, it is widely used as an additive in dietary supplements. For example, Patent Publication No. CN107412216A describes the use of α-ketoglutaric acid in improving reproductive function impairment in animals caused by a high-fat diet. There are also reports of the use of α-ketoglutaric acid in the preparation of drugs for treating myocardial infarction, such as Patent Publication No. CN114028371A, and in drugs for preventing and treating skin tumors, such as Patent Publication No. CN115414343A. However, there are no studies on the use of α-ketoglutaric acid to promote remyelination.

[0009] Summary of the Invention

[0010] The present invention aims to provide the use of α-ketoglutaric acid in the preparation of drugs for preventing and treating demyelinating-related diseases. It is the first time that α-ketoglutaric acid or its derivatives are used to prevent and / or treat myelin defects in demyelinating-related diseases. The invention is a drug with effectiveness and safety.

[0011] The technical solutions provided by the present invention are as follows:

[0012] The present invention provides the use of α-ketoglutaric acid or its derivatives in the preparation of drugs for preventing and treating demyelination-related diseases. The α-ketoglutaric acid or its derivatives prevents and treats myelin defects in demyelination-related diseases by promoting myelin generation, regeneration or repair.

[0013] The demyelination-related disease is a demyelinating disease, which includes multiple sclerosis, acute disseminated encephalomyelitis, demyelinating diseases with myelin formation defects, and inflammatory demyelinating diseases of the central nervous system in children.

[0014] The demyelination-related disease is amyotrophic lateral sclerosis, Huntington's disease, hypomyelinating leukodystrophy, Alzheimer's disease, Parkinson's syndrome, diabetes-related visual impairment, or damage to retinal neurons and optic nerves caused by glaucoma or craniocerebral trauma.

[0015] The remyelination includes remyelination in demyelinated areas of the spinal cord, remyelination after demyelination, and remyelination after optic nerve injury.

[0016] Derivatives of α-ketoglutaric acid include α-ketoglutarate dimethanol, α-ketoglutarate dimethyl ester, α-ketoglutarate sodium, and α-ketoglutarate disodium.

[0017] The present invention also provides a drug for preventing and treating demyelinating diseases, comprising α-ketoglutaric acid or its derivatives. α-ketoglutaric acid or its derivatives prevent and treat demyelinating diseases by promoting myelin formation, regeneration or repair. The demyelinating diseases include multiple sclerosis, acute disseminated encephalomyelitis, demyelinating diseases with myelin formation defects, and inflammatory demyelinating diseases of the central nervous system in children.

[0018] The remyelination includes remyelination in demyelinated areas of the spinal cord, remyelination after demyelination, and remyelination after optic nerve injury.

[0019] The drugs for preventing and treating demyelinating diseases provided by the present invention all contain α-ketoglutaric acid or its derivatives as their effective substances, and may also contain medically acceptable excipients.

[0020] Furthermore, these drugs can also be prepared into any pharmaceutically acceptable dosage form, including but not limited to: decoctions, powders, pills, wine preparations, lozenges, gels, tea preparations, koji preparations, cakes, lotions, stick preparations, thread preparations, strip preparations, nail preparations, moxibustion preparations, ointments, pills, liposome preparations, aerosols, injections, mixtures, oral ampoules, tablets, capsules, pills, emulsions, films or sponges.

[0021] Compared with the prior art, the present invention has the following advantages and beneficial effects:

[0022] (1) α-ketoglutaric acid itself is a metabolite of the tricarboxylic acid cycle in the body and is used in dietary supplements and health products. It has good effects in anti-aging and muscle growth. As a dietary supplement, no obvious side effects have been reported in adults. It can successfully cross the blood-brain barrier and enter the brain parenchyma. Therefore, it has better safety and effectiveness.

[0023] (2) In existing studies on promoting myelin regeneration, many screened drugs are mainly used to treat protein targets that inhibit myelin regeneration. The present invention provides a new direction for promoting myelin regeneration. By injecting α-ketoglutaric acid supplements, a metabolite of the tricarboxylic acid cycle, ATP synthesis in cells is increased, thereby activating OPCs and promoting myelin regeneration.

[0024] (3) The present invention is the first to use α-ketoglutaric acid for demyelinating diseases of the nervous system, such as multiple sclerosis and senile demyelination, and provides a new treatment principle. The current treatment principle is mostly to use anti-immune drugs to reduce the damage of endogenous immune cells to the myelin sheath, while the present invention treats the disease by promoting myelin regeneration after demyelination through α-ketoglutaric acid or its derivatives.

[0025] (4) Since neurodegenerative diseases such as Alzheimer's disease can also lead to a decrease in the overall level of myelin, the present invention also proposes the potential therapeutic effect of α-ketoglutaric acid or its derivatives in the treatment of neurodegenerative diseases, and experimentally proves that α-ketoglutaric acid has the function of improving the memory ability of Alzheimer's disease.

[0026] (5) The present invention can be used to prevent and treat white matter lesions in the aging process, and can enhance the proliferation of oligodendrocytes and myelination in the aging brain, thereby improving the learning and memory ability related to aging. At present, for aging-related white matter lesions, in addition to the general selective nutritional intake treatment, most treatment plans require selective treatment based on the cause of the disease. For example, white matter lesions caused by diabetes use hypoglycemic drugs (such as gliclazide sustained-release tablets, metformin hydrochloride tablets, acarbose tablets, etc.), and white matter lesions caused by hypertension use antihypertensive drugs (such as captopril tablets, enalapril maleate tablets, felodipine sustained-release tablets, etc.). These drugs can only alleviate white matter lesions and have little significance for the regeneration and repair of myelin.

[0027] (6) For visual impairment caused by glaucoma, mechanical damage, diabetes, etc., mainstream treatment methods include surgery, drugs to reduce intraocular pressure, etc., while drugs for repairing optic neuropathy are mainly vitamins and neuroprotectants, such as nerve growth factor, gangliosides, etc. However, there is still a lack of treatment options for improving the immune microenvironment of optic nerve lesions and promoting myelin regeneration. This invention is the first to use α-ketoglutaric acid to improve the immune microenvironment of damaged optic nerves, which can promote axonal myelination and thus promote vision recovery.

[0028] (7) Currently, there are relatively few studies on the mechanism of white matter damage after chemotherapy and radiotherapy for cancer. Chemotherapy and radiotherapy can also induce the continuous activation of microglia in the brain, destroying the various mechanisms of neural plasticity and developing into treatment-related cognitive impairment. Therefore, promoting myelin regeneration after chemotherapy and radiotherapy and improving the immune microenvironment of the brain center are expected to improve the cognitive impairment of cancer patients after treatment. More effective and safe drugs can be developed to promote myelin regeneration and immune regulation. This invention is the first to use α-ketoglutaric acid to improve white matter damage after chemotherapy and radiotherapy for cancer, improve the immune microenvironment of the brain center, and thus improve cognitive impairment related to cancer treatment. BRIEF DESCRIPTION OF THE DRAWINGS

[0029] FIG1 is a schematic diagram showing how energy metabolism (ATP metabolism) promotes oligodendrocyte maturation and myelination.

[0030] Figure 2 is a schematic diagram of AKG participating in the tricarboxylic acid cycle and promoting ATP synthesis.

[0031] FIG3 is a schematic diagram of DMKG treatment in the LPC spinal cord demyelination model.

[0032] Figure 4 shows the immunofluorescence images (H3K27ac and MBP) of the DMKG-treated group and the Vec solvent control group (hereinafter referred to as DMKG and Vec) in the LPC spinal cord demyelination model.

[0033] Figure 5 shows the number of H3K27ac / DOR positive cells per unit area in the LPC spinal cord demyelination model DMKG and Vec.

[0034] FIG6 is an RNA in situ hybridization image of MBP and plp in the LPC spinal cord demyelination model DMKG and Vec.

[0035] Figure 7 is an electron microscopic image of the spinal cord cross-section of the LPC spinal cord demyelination model DMKG and Vec.

[0036] FIG8 is a graph showing the percentage of axons containing regenerated myelin in DMKG and Vec in the LPC spinal cord demyelination model.

[0037] FIG9 is a statistical diagram of the axonal g-ratio distribution of DMKG and Vec in the LPC spinal cord demyelination model.

[0038] Figure 10 shows the tamoxifen-induced marker (tdTomato red fluorescence) transgenic aging mice (Cspg4-Cre ERT :toTomato) oligodendrocyte precursors, and then tracking the proliferation and differentiation of oligodendrocytes in the hippocampus of DMKG and Vec fluorescence images.

[0039] FIG11 is a graph showing the number of GSTπ-positive cells in the hippocampus of aging mice DMKG and Vec.

[0040] FIG12 is an immunofluorescence image (Iba1 and BrdU) of microglia and their proliferation in the hippocampus of aging mice DMKG and Vec.

[0041] FIG13 is a diagram of the Morris water maze test of aging mice DMKG and Vec (time to reach the hidden platform on the water surface).

[0042] Figure 14 shows the Morris water maze test results of aged mice DMKG and Vec (total distance swum after the platform was removed, the number of times the original platform position was crossed, and the percentage of time swimming in the area where the original platform was located).

[0043] FIG15 is a schematic diagram of DMKG treatment of the LPC myelin injury model in aged mice.

[0044] FIG16 is a fluorescence immunofluorescence image of DMKG and Vec in the LPC myelin damage model of aging mice (DOR).

[0045] FIG17 is a fluorescence immunofluorescence image (H3K27ac and MBP) of DMKG and Vec in the LPC myelin damage model of aged mice.

[0046] FIG18 is a graph showing the number of H3K27ac and DOR positive cells per unit area in the DMKG and Vec models of LPC myelin damage in aged mice.

[0047] FIG19 is a fluorescence immunofluorescence image of DMKG and Vec (GSTπ) in the LPC myelin damage model of aged mice.

[0048] FIG20 is a graph showing the number of GSTπ-positive cells per unit area in the DMKG and Vec models of LPC myelin damage in aged mice.

[0049] Figure 21 is an electron microscopic image of DMKG and Vec in the LPC myelin damage model of aging mice.

[0050] Figure 22 shows the percentage of myelinated axons in the DMKG and Vec myelin sheath injury models of LPC in aged mice.

[0051] FIG23 is a Morris water maze test trajectory diagram of Alzheimer's disease mouse models DMKG and Veh.

[0052] FIG24 shows the Morris water maze test of Alzheimer's disease mouse models DMKG and Veh (the number of times the mouse crossed the original platform position after the platform was removed).

[0053] FIG25 shows the Morris water maze test of Alzheimer's disease mouse models DMKG and Veh (swimming distance in the original platform area after the platform was removed).

[0054] FIG26 shows the Morris water maze test of Alzheimer's disease mouse models DMKG and Veh (swimming time in the original platform area after the platform was removed).

[0055] FIG27 shows the Morris water maze test of Alzheimer's disease mouse models DMKG and Veh (average swimming time in the area where the platform was originally located after the platform was removed).

[0056] FIG28 shows the Morris water maze test of Alzheimer's disease mouse models DMKG and Veh (average swimming distance to the original platform position after the platform was removed).

[0057] Figure 29 shows the Morris water maze test for Alzheimer's disease mouse models DMKG and Veh (after the platform was removed, the ratio of swimming time in the limit where the original platform was located to the other limits)

[0058] FIG30 is a fluorescence immunofluorescence image of GSTπ-positive cells and their proliferation in the optic nerve injury mouse model DMKG and Vec, as well as a graph showing the number of proliferating GSTπ-positive cells per unit area.

[0059] Figure 31 shows electron micrographs of DMKG and Vec in the optic nerve injury mouse model and the percentage of myelinated axons. DETAILED DESCRIPTION

[0060] The objectives, technical solutions and beneficial effects of the present invention are further described in detail below.

[0061] It should be noted that the following detailed descriptions are exemplary and are intended to provide further explanation of the claimed invention. Unless otherwise defined, all technical and scientific terms used herein have the same meaning as commonly understood by one of ordinary skill in the art to which the invention belongs.

[0062] Currently, α-ketoglutarate (AKG) is primarily used as a dietary supplement, with potential benefits in anti-aging, wound healing, and muscle growth, but there are no reports on AKG's relevance to myelination. This study, through extensive experimental research, demonstrated that intraperitoneal injection of the AKG supplement dimethyl α-ketoglutarate (DMKG) into mice effectively altered the dense chromatin structure of oligodendrocyte precursor cells (OPCs) and promoted the expression of genes associated with oligodendrocyte differentiation and maturation in the treatment of myelin retardation and myelin damage repair.

[0063] Specifically, myelination originates from the differentiation and maturation of oligodendrocytes, and this process requires the joint action of a large number of transcription factors and ATP-dependent chromatin remodeling molecules, including Brg1, Chd7, etc. Therefore, the activation of oligodendrocyte precursors OPCs requires a large amount of energy to unwind the dense chromatin structure and initiate the expression of genes involved in oligodendrocyte differentiation and maturation, as shown in Figure 1. From the perspective of intracellular metabolic pathways, the production of ATP mainly depends on the tricarboxylic acid cycle. Among them, AKG is an intermediate product of the tricarboxylic acid cycle and is involved in various metabolic and cellular pathways, as shown in Figure 2. One tricarboxylic acid cycle can produce 10 molecules of ATP. Therefore, by introducing exogenous AKG, the tricarboxylic acid cycle metabolic pathway can be enhanced, providing a large amount of ATP for cell activities, thereby providing a large amount of energy for chromatin unwinding and gene expression.

[0064] Glossary:

[0065] ATP: adenosine triphosphate (ATP), chemical formula is C 10 H 16 N5O 13 P3, with a molecular weight of 507.18, is an unstable, high-energy compound composed of one adenine molecule, one ribose molecule, and three phosphate groups. Also known as adenosine triphosphate, it releases a significant amount of energy upon hydrolysis and is the most direct energy source in organisms.

[0066] Brg1: A SWI / SNF-related actin-dependent chromatin regulator with ATP hydrolase, DNA polymerase, and nucleic acid binding activities. It participates in nervous system development, positive regulation of the Wnt signaling pathway, and positive regulation of RNA polymerase II transcription.

[0067] Chd7: Chromatin helicase DNA-binding protein 7, a transcription factor with ATP-binding activity, involved in organ development, etc.

[0068] The following experiments verify the improvement of AKG supplement dimethyl α-ketoglutarate (DMKG) on myelination and Alzheimer's disease.

[0069] Application examples:

[0070] 1.DMKG formula: Mix DMKG and normal saline at a ratio of 1:10.

[0071] 2. Experimental groups:

[0072] Type of mice treated:

[0073] (1) The LPC (lysolecithin) demyelination mouse model was constructed by injecting LPC into the white matter region of the spinal cord of adult mice (5 months old), causing demyelination at the injection site, thereby establishing a mouse demyelination model.

[0074] (2) Aged mice (16-17 months old, with the genotype Cspg4-CreERT / Rosa26-STOP-tdTomato) were constructed by injecting tamoxifen to induce the expression of the red fluorescent protein tdTomato to track newborn oligodendrocytes.

[0075] (3) Alzheimer's disease mouse model (9 months old)*, using the conventional transgenic mouse model of this disease, APP / PS1.

[0076] (4) Optic nerve injury mouse model: The optic nerve of adult mice was injured by squeezing the optic nerve behind the mouse eyeball using sharp forceps.

[0077] Grouping: solvent administration group (control group) and DMKG administration group.

[0078] 3. Administration: Intraperitoneal injection, 10 mg / kg (mouse body weight) per day.

[0079] 4. Post-treatment effect detection:

[0080] (1) Immunofluorescence and RNA in situ hybridization to detect myelin regeneration

[0081] Tissue collection: After anesthetizing mice, the right atrium was opened and the left ventricle was perfused with 1X PBS and then 4% paraformaldehyde. (For electron microscopy, only the electron microscopy perfusion solution is needed: 4% paraformaldehyde, 40% glutaraldehyde, 0.02 mol / L sodium cacodylate, pH 7.2-7.4. Dissolve in ultrapure water and prepare freshly prepared samples.)

[0082] Tissue type: mouse brain (hippocampus region), spinal cord;

[0083] Tissue sectioning: vibrating section for mouse brain and frozen section for spinal cord;

[0084] Detection methods: immunofluorescence, RNA in situ hybridization, transmission electron microscopy;

[0085] Detection markers: Mbp, GSTπ, etc.

[0086] (2) Behavioral testing of the recovery status of drug-administered mice

[0087] Behavioral testing: Morris water maze test

[0088] Morris Water Maze Test: Day 1, Visible Platform Phase: Water is added to the maze, leaving the circular platform above the surface, allowing the mouse to just land. The mouse is placed in the water at a 45-degree angle facing the center of the circle and allowed to swim to the platform spontaneously or induced. The duration is set to 1 minute, with the mouse remaining on the platform for 10 seconds. Days 2-6, Hidden Platform Phase (Acquisition Training): More water is added, leaving the platform just submerged. The platform is placed in one of four random starting positions: east, west, south, or north. The time it takes for the mouse to find the submerged platform is recorded. During the first few training sessions, if this time exceeds 60 seconds, the animal is guided to the platform. The animal is allowed to remain on the platform for 10 seconds. Each animal is trained four times daily, with a 15-20 minute interval between training sessions, for 5 consecutive days. Day 7, Platform Removal Phase (Exploration Training): The day after the last acquisition training session, the platform is removed, and 60 seconds of exploration training begins. The animal is placed in the water from the quadrant opposite the original platform. The time the animal spent in the target quadrant (the quadrant where the platform was originally placed) and the number of times it entered the quadrant were recorded as indicators of spatial memory.

[0089] 5. Experimental results after drug treatment

[0090] (1) DMKG treatment helps enhance myelin regeneration in demyelinated areas of the spinal cord

[0091] In a mouse model of LPC demyelination, the DMKG group (DMKG) was injected with the DMKG formula, while the vehicle group (Vec) was injected with a control vehicle (normal saline). Injections were performed on days 14 and 21. Spinal cord tissue was collected the day after injection, and the therapeutic effect of DMKG on the LPC spinal cord demyelination model mice was observed by immunofluorescence, as shown in Figure 3.

[0092] The results of immunofluorescence and RNA in situ hybridization showed that in the lesion recovery area after 14 days, the protein levels of myelin protein MBP, transcription factor Trp53inp2 / DOR, and super enhancer marker H3K27ac in the DMKG treatment group were significantly higher than those in the control group, and the number of Plp-positive cells (immature oligodendrocytes) was also significantly increased, as shown in Figures 4 to 6. After 21 days of treatment, the number of GSTπ-positive cells (mature oligodendrocytes) in the DMKG group also increased significantly. Projection electron microscopy experiments also showed that axons of different calibers in the DMKG group showed an increase in the number of myelinated axons and thickening of the myelin sheath, as shown in Figure 7.

[0093] 8 and 9 , it can be seen that after the demyelination model mice were treated with DMKG, the percentage of remyelinated axons increased significantly, and the thickness of the myelin sheath increased significantly regardless of the thickness of the axons.

[0094] (2) DMKG treatment can promote myelination and enhance the learning and memory abilities of aged mice

[0095] In aged mice, the DMKG group (DMKG) was injected with a DMKG formula, while the vehicle group (Vec) was injected with a control vehicle (normal saline). In aged mice (16-17 months), Trp53inp2 expression levels decreased, and new myelin sheaths were also reduced. However, DMKG treatment enhanced MBP expression in aged mice, enhanced myelin formation in the aged brain, and promoted the maturation of OPCs.

[0096] By expressing the genotype Cspg4-Cre ERT :tdTomato aged mice (16 months old) were injected with Tamoxfen to activate tdTomato red fluorescent protein to mark oligodendrocytes. After one month of DMKG treatment, the number of tdTomato-positive cells (newborn oligodendrocyte cell line) in the CA1 region of the hippocampus of the aged mice increased significantly compared with the non-DMKG treatment group, as shown in Figure 10. Approximately 37% of the tdTomato-positive cells were also GSTπ-positive (mature oligodendrocytes), while the control group had only 8%, as shown in Figure 11. At the same time, microglia also proliferated, as shown in Figure 12.

[0097] Furthermore, in the Morris water maze test, DMKG-treated aging mice showed improved memory function, reaching the hidden platform on the water surface in a shorter time (see Figure 13). After the platform was removed, the number of crossings to the original platform position increased significantly, and the percentage of time spent swimming in the platform-remaining zone increased significantly (see Figure 14).

[0098] (3) DMKG treatment can promote remyelination after demyelination in aged mice

[0099] For the LPC aged mouse myelin injury model, the DMKG group (DMKG) was injected with the DMKG formula, and the solvent group (Vec) was injected with the control solvent (normal saline), as shown in Figure 15. The myelin regeneration ability of aged mice showed a significant downward trend, but after DMKG treatment, as shown in Figure 16, the expression of the Trp53inp2 gene in the lesion of aged mice was upregulated, MBP and H3K27ac immunofluorescence labeling increased, and the expression of GSTπ, a marker of mature oligodendrocytes, also increased, as shown in Figures 17 to 20. At the same time, the number of myelinated axons also increased significantly, as shown in Figures 21 and 22.

[0100] As shown in FIG22 , in the aged LPC myelin injury mouse model treated with DMKG, the percentage of remyelinated axons in the injury area increased to more than 10%, while in the untreated model, this value was only about 3%.

[0101] (4) DMKG treatment can improve the memory ability of Alzheimer's mice

[0102] For the Alzheimer's disease mouse model, the DMKG group (DMKG) was injected with DMKG formula, and the solvent group (Veh) was injected with control solvent (normal saline).

[0103] After DMKG treatment, the Morris water maze test showed that compared with mice not injected with DMKG, there was no difference in the visible platform period between AD mice that received continuous intraperitoneal drug injections; there was no significant difference in the hidden platform training period, but in the platform withdrawal period, the number of crossing the original platform increased significantly, and other indicators showed a certain increasing trend, indicating that continuous intraperitoneal injection of DMKG in AD mice can improve spatial memory, see Figures 23 to 29 for details.

[0104] (5) DMKG treatment can promote myelin regeneration after optic nerve injury

[0105] For the optic nerve injury mouse model, the DMKG group (DMKG) was injected with DMKG formula, and the solvent group (Vec) was injected with control solvent (normal saline).

[0106] After DMKG injection, the number of oligodendrocytes at the site of optic nerve injury increased, with a particularly significant increase in mature oligodendrocytes. Electron microscopy also showed enhanced axonal remyelination after DMKG treatment, indicating that DMKG treatment can promote remyelination after optic nerve injury (see Figures 30 and 31 for details).

[0107] It should be noted that no obvious side effects were found in all the above-mentioned DMKG treatment experiments.

[0108] The above description is only a preferred embodiment of the present invention and does not limit the present invention in any form. Any simple modification or equivalent change made to the above embodiment based on the technical essence of the present invention shall fall within the scope of protection of the present invention.

Claims

1. Use of α-ketoglutaric acid or its derivatives in the preparation of drugs for preventing and treating demyelination-related diseases, characterized in that: The α-ketoglutarate or its derivatives prevent and treat myelin deficiency in demyelination-related diseases by promoting myelin generation, regeneration or repair.

2. The use according to claim 1, characterized in that: The demyelination-related diseases are demyelinating diseases, which include multiple sclerosis, acute disseminated encephalomyelitis, demyelinating diseases with myelin formation defects, and inflammatory demyelinating diseases of the central nervous system in children.

3. The use according to claim 1, characterized in that: The demyelination-related disease is amyotrophic lateral sclerosis, Huntington's disease, hypomyelinating leukodystrophy, Alzheimer's disease, Parkinson's syndrome, diabetes-related visual impairment, or damage to retinal neurons and optic nerves caused by glaucoma or craniocerebral trauma.

4. The use according to claim 1, characterized in that: The remyelination includes remyelination in demyelinated areas of the spinal cord, remyelination after demyelination, and remyelination after optic nerve injury.

5. The use according to claim 1, characterized in that: Derivatives of α-ketoglutaric acid include α-ketoglutarate dimethanol, α-ketoglutarate dimethyl ester, α-ketoglutarate sodium, and α-ketoglutarate disodium.

6. A drug for preventing and treating demyelinating diseases, characterized in that: The invention comprises α-ketoglutaric acid or its derivatives, which can prevent and treat demyelinating diseases by promoting myelin formation, regeneration or repair. The demyelinating diseases include multiple sclerosis, acute disseminated encephalomyelitis, demyelinating diseases with myelin formation defects and inflammatory demyelinating diseases of the central nervous system in children.

7. The drug according to claim 6, characterized in that: The remyelination includes remyelination in demyelinated areas of the spinal cord, remyelination after demyelination, and remyelination after optic nerve injury.

8. The drug according to claim 6, characterized in that: Derivatives of α-ketoglutaric acid include α-ketoglutarate dimethanol, α-ketoglutarate dimethyl ester, α-ketoglutarate sodium, and α-ketoglutarate disodium.

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