Use of kynurenic acid in preparing medicament for treating cerebral stroke

Abstract

Provided is use of kynurenic acid in preparing a medicament for preventing and / or alleviating and / or treating cerebral stroke or a symptom thereof. The medicament is used for preventing and / or treating cerebral infarction, alleviating the severity of cerebral infarction, improving neurological functions, improving systemic inflammatory phenotype, ameliorating intestinal flora dysbiosis, ameliorating intestinal barrier dysfunction, and improving the level of antimicrobial peptides in the intestinal tract in a patient with cerebral stroke.

Description

Use of kynurenic acid in the preparation of drugs for treating stroke Technical Field

[0001] This application belongs to the field of biomedicine, and in particular relates to the use of quinoline acid in the preparation of drugs for the treatment of stroke. Background Technology

[0002] Stroke is an acute cerebrovascular disease caused by the sudden rupture or blockage of blood vessels in the brain, resulting in brain tissue damage. It includes ischemic and hemorrhagic strokes. It is one of the most prevalent diseases in the world today, with its incidence increasing year by year. Stroke patients experience a decline in quality of life, and in severe cases, it can lead to death.

[0003] Currently, the focus of stroke treatment is intravenous thrombolysis or endovascular therapy, such as intravenous injection of recombinant tissue plasminogen activator (r-tPA). This method is still considered the most important treatment, but it has strict time window requirements. In addition, depending on each patient's specific condition, there are antiplatelet therapies, anticoagulants, neuroprotective agents, and other symptomatic treatments, including the treatment and prevention of hyperglycemia, hypertension, and acute stroke complications. However, effective treatment methods are still lacking, and prevention is currently considered the best approach. Given the increasingly serious nature of stroke, there is an urgent need to provide safe, effective, and low-toxicity new treatment methods, targets, and drugs. Summary of the Invention

[0004] To address the deficiencies of the prior art, this application discovers that quinoline acid has the effects of preventing, alleviating, and / or treating stroke or its symptoms. It can prevent and / or treat cerebral infarction in stroke patients, improve the degree of cerebral infarction in stroke patients, improve the neurological function of stroke patients, improve the systemic inflammatory phenotype in stroke patients, improve intestinal flora imbalance in stroke patients, improve intestinal barrier function abnormalities in stroke patients, and improve the level of antimicrobial peptides in the intestines of stroke patients.

[0005] The first aspect of this application provides the use of quinolinic acid in the preparation of medicaments for the prevention and / or relief and / or treatment of stroke or its symptoms.

[0006] A second aspect of this application provides the use of kynurenic acid in the preparation of a medicament, said medicament being used in any one or more of the following:

[0007] 1) Prevention and / or treatment of cerebral infarction in stroke patients;

[0008] 2) Improves the severity of cerebral infarction in stroke patients;

[0009] 3) Improves neurological function in stroke patients;

[0010] 4) Improves the systemic inflammatory phenotype in stroke patients;

[0011] 5) Improves gut microbiota dysbiosis in stroke patients;

[0012] 6) Improves abnormal intestinal barrier function in stroke patients;

[0013] 7) Improves the level of antimicrobial peptides in the gut of stroke patients.

[0014] A third aspect of this application provides a medicament for the prevention and / or relief and / or treatment of stroke or its symptoms, comprising an effective amount of canine quinolinic acid.

[0015] The fourth aspect of this application provides a method for preventing and / or alleviating and / or treating stroke or its symptoms, comprising administering to a patient the drug used in the first aspect, or the drug used in the second aspect, or the drug used in the third aspect.

[0016] The beneficial effects of this application are as follows:

[0017] This application is the first to discover that kynurenic acid has the effects of preventing, alleviating and / or treating stroke or its symptoms. It can prevent and / or treat cerebral infarction in stroke patients, improve the degree of cerebral infarction in stroke patients, improve the neurological function of stroke patients, improve the systemic inflammatory phenotype in stroke patients, improve intestinal flora disorder in stroke patients, improve intestinal barrier function abnormality in stroke patients, and improve the level of antimicrobial peptides in the intestine of stroke patients. Moreover, only a low concentration of 40 mg / Kg of kynurenic acid is needed to achieve the effects of prevention, alleviation and treatment, providing a new, feasible and effective solution for the prevention, alleviation and treatment of stroke. Attached Figure Description

[0018] Figure 1 shows that kynurenic acid can improve the degree of cerebral infarction in mice after stroke in Example 2 of this application.

[0019] Figure 2 shows that kynurenic acid can improve the neurological function of mice after stroke in Example 3 of this application.

[0020] Figure 3 shows that kynurenic acid can improve the systemic inflammatory phenotype in mice after stroke in Example 4 of this application.

[0021] Figure 4 shows that quinolinic acid can improve intestinal flora disorder in mice after stroke in Example 5 of this application.

[0022] Figure 5 shows that kynurenic acid can improve abnormal intestinal barrier function in mice after stroke, as demonstrated in Example 6 of this application.

[0023] Figure 6 shows that quinolinic acid can improve the level of antimicrobial peptides in the intestines of mice after stroke in Example 7 of this application. Detailed Implementation

[0024] The following specific examples illustrate the implementation of this application. Those skilled in the art can easily understand other advantages and effects of this application from the content disclosed in this specification. This application can also be implemented or applied through other different specific embodiments, and various details in this specification can also be modified or changed based on different viewpoints and applications without departing from the spirit of this application.

[0025] Before further describing the specific embodiments of this application, it should be understood that the scope of protection of this application is not limited to the specific embodiments described below; it should also be understood that the terminology used in the embodiments of this application is for describing specific embodiments and not for limiting the scope of protection of this application; in the specification and claims of this application, unless otherwise expressly stated in the text, the singular forms "a", "an" and "this" include the plural forms.

[0026] When numerical ranges are given in the embodiments, it should be understood that, unless otherwise stated in this application, both endpoints of each numerical range and any value between the two endpoints may be selected. Unless otherwise defined, all technical and scientific terms used in this application have the same meaning as commonly understood by one of ordinary skill in the art. In addition to the specific methods, apparatus, and materials used in the embodiments, based on the knowledge of the prior art possessed by one of ordinary skill in the art and the description in this application, any prior art methods, apparatus, and materials similar to or equivalent to those described, apparatus, and materials in the embodiments of this application may be used to implement this application.

[0027] In this application, "prevention" refers to the preventive administration of the combination to healthy subjects to prevent the occurrence of a disease or its symptoms, and may also include the preventive administration of the combination to patients in the pre-treatment stage of an allergic disease to be treated. "Prevention" does not require the complete elimination of the possibility of the disease or its symptoms occurring; prevention may also mean that the possibility or severity of the disease or its symptoms is reduced in the presence of the applied combination.

[0028] In this application, relief refers to reducing, shortening, or delaying a symptom, disease, condition, or phenotype. The symptom, disease, condition, or phenotype may include subjective perceptions of the subject, such as pain, dizziness, or other physiological disturbances, or medically detectable indicators, such as lesions detected through medical testing.

[0029] In this application, the treatment generally refers to a clinical intervention that alters the natural processes of the treated individual or cells in a clinicopathological process, and may include improving the disease state, eliminating lesions, or improving prognosis.

[0030] Kynurenic acid is a product of tryptophan metabolism, formed from kynurenic acid via aminotransfer and excreted in the urine. It has biological activity, is an antagonist of N-methyl-D-aspartic acid, and is closely related to the regulation of leukocyte-endothelial cell binding.

[0031] The chemical formula of quinolinic acid is C 10 The structure of H7NO3, with a molecular weight of 189.17, is as follows:

[0032] Stroke, also known as cerebrovascular accident (CVA), is an acute cerebrovascular disease caused by the sudden rupture or blockage of blood vessels in the brain, resulting in brain tissue damage. It includes ischemic stroke and hemorrhagic stroke. Due to the lack of effective treatments, prevention is considered the best approach.

[0033] Ischemic stroke generally refers to a group of diseases caused by insufficient blood supply to a specific or widespread area of ​​the brain, leading to dysfunction of the neurological tissue in that area. Insufficient blood supply to the brain can be caused by a variety of diseases or abnormalities, such as sickle cell anemia, vascular compression, ventricular tachycardia, arterial plaque buildup, thrombosis, severe hypotension, and congenital heart defects. Sickle cells are more prone to clotting than normal blood cells, obstructing blood flow to the brain; vascular compression can lead to cerebral ischemia by blocking arteries carrying oxygen to the brain, and causes of vascular compression include tumors; ventricular tachycardia can cause complete cardiac arrest, leading to cessation of blood flow, while arrhythmias can also cause blood clots, resulting in cerebral ischemia; arterial blockage due to plaque buildup can also cause cerebral ischemia, and even small plaque buildups can narrow the channels, leading to thrombus formation and thus cerebral ischemia; coagulation disorders, such as large blood clots, can also cause cerebral ischemia by obstructing blood flow; heart attacks can also cause cerebral ischemia, as they can slow blood flow and cause blood to clot, preventing blood from reaching the brain; there is a correlation between heart attacks and hypotension, and improper use and response to medications can also lead to excessively low blood pressure, which usually indicates insufficient tissue oxygenation. In this application, ischemic stroke can also be achieved by surgically occluding blood flow to the middle cerebral artery in mice. The neurological dysfunction may include neuronal death. Due to the dysfunction of different areas of nerve tissue, the main symptoms may include: changes in breath / taste / hearing or vision, swallowing / pupil light reflex disorders, body movement disorders, aphasia, changes in respiration and heart rate, ischemia of other tissues or organs, loss of consciousness, etc.

[0034] This application first provides the use of quinolinic acid in the preparation of medicaments for the prevention and / or relief and / or treatment of stroke or its symptoms.

[0035] In specific embodiments of this application, the stroke includes ischemic stroke and hemorrhagic stroke.

[0036] As some embodiments of this application, the stroke described is ischemic stroke.

[0037] In a specific embodiment of this application, the drug has one or more of the following effects:

[0038] 1) To prevent and / or alleviate and / or treat cerebral infarction in stroke patients;

[0039] 2) Improves the severity of cerebral infarction in stroke patients;

[0040] 3) Improves neurological function in stroke patients;

[0041] 4) Improves the systemic inflammatory phenotype in stroke patients;

[0042] 5) Improves gut microbiota dysbiosis in stroke patients;

[0043] 6) Improves abnormal intestinal barrier function in stroke patients;

[0044] 7) Improves the level of antimicrobial peptides in the gut of stroke patients.

[0045] Cerebral infarction refers to ischemic necrosis or softening of localized brain tissue caused by impaired blood supply to the brain, resulting in ischemia and hypoxia.

[0046] The degree of cerebral infarction refers to the extent of damage to brain tissue caused by cerebral infarction, which can be assessed through infarct volume, infarct proportion, and NIHSS score. Infarct volume refers to the actual size of the infarcted area, usually measured in milliliters (ml). This volume can be determined through imaging examinations (such as CT and MRI). The infarct proportion refers to the ratio of the infarcted area to the entire brain; this proportion reflects the extent of the infarct, i.e., what percentage of brain tissue is covered by the infarct.

[0047] In a specific embodiment of this application, the improvement in the degree of cerebral infarction in stroke patients is manifested in the reduction of the infarct volume and the proportion of cerebral infarction. That is, after the stroke patients are given the drug, compared with those who have not been given the drug, the infarct volume and the proportion of cerebral infarction are reduced.

[0048] The neurological function refers to the various physiological activities and functions performed by the nervous system (including the brain, spinal cord, and peripheral nerves), which can be evaluated by statistically scoring using neurological function scoring criteria. These criteria can include, for example, NIHSS, GCS, MESSS, and NSS. Specifically, when scoring neurological function in mice, the following criteria can be used: tail tuck test (3 points), forelimb flexion (1 point), hindlimb flexion (1 point), deviation from vertical axis >100° (1 point), placing the rat on the floor (normal value = 0; maximum value = 3) (3 points), normal walking (0 points), inability to walk in a straight line (1 point), circling towards the side of mild paralysis (2 points), tilting towards the side of mild paralysis (3 points), balance beam test (normal value = 0; maximum value = 6) (6 points), stable balance posture (0 points), gripping the edge of the balance beam (0 points). 1 point, clinging tightly to the balance beam, one limb dangling from the beam; 2 points, clinging tightly to the balance beam, two limbs dangling from the beam or rotating on the beam (>60 seconds); 3 points, attempting to balance on the beam but falling (>40 seconds); 4 points, attempting to balance on the beam but falling (>20 seconds); 5 points, falling, not attempting to balance on the beam; 6 points, auricular reflex (shaking head when touched in the external auditory canal); 1 point, corneal reflex (blinking when the cornea is lightly touched with a cotton thread); maximum score 14 points.

[0049] In a specific embodiment of this application, the improvement in the neurological function of stroke patients is a reduction in the patient's neurological function score, that is, after the stroke patient is given the drug, the patient's neurological function score is lower than that without the drug.

[0050] The systemic inflammatory phenotype refers to the characteristics and patterns of systemic inflammatory responses exhibited by the body in response to various internal and external stimuli (such as infection, trauma, metabolic disorders, etc.). This phenotype typically involves the activation of multiple cytokines and inflammatory mediators, leading to heterogeneity in inflammatory responses under different disease states, which can be evaluated through the levels of inflammatory factors.

[0051] In a specific embodiment of this application, the improvement in the systemic inflammatory phenotype of stroke patients is manifested by a decrease in the levels of inflammatory factors IFN-γ and / or TNF-α in the serum of stroke patients. Specifically, after administration of the drug to stroke patients, compared to the absence of drug administration, the levels of inflammatory factors IFN-γ and / or TNF-α in the patient's serum are reduced. Here, IFN-γ is interferon-gamma, and TNF-α is tumor necrosis factor-α.

[0052] The aforementioned gut microbiota dysbiosis, also known as gut microecological imbalance, refers to an imbalanced state caused by changes in the composition, activity, or distribution of the gut microbiota. Changes in the gut microbiota can be assessed by extracting and sequencing the DNA of the microbiota and analyzing the microbiota data.

[0053] Abnormal intestinal barrier function refers to the disruption of the intestinal mucosal barrier function, leading to the entry of intestinal endotoxins, bacteria, and other harmful substances into the bloodstream, triggering a series of inflammatory responses and related diseases. The intestinal barrier function mainly includes the mucosal barrier, immune barrier, and biological barrier, which work together to prevent the invasion of enterogenic bacteria. This can be assessed by the levels of intestinal barrier-related genes Muc1, Tjp1, Ocln, and Cldn4 in intestinal tissue. The encoded proteins are Muc1 (polymorphic epithelial mucin), Tjp1 (tight junction protein 1), Ocln (Occludin), and Cldn4 (Claudin-4).

[0054] In a specific embodiment of this application, the improvement of intestinal barrier function abnormality in stroke patients is manifested by an increase in the expression levels of Muc1, and / or Tjp1, and / or Ocln, and / or Cldn4 genes in the intestines of stroke patients. That is, after the medication is administered to stroke patients, compared with no medication, the expression levels of Muc1, Tjp1, Ocln, and Cldn4 genes in the patient's intestines are increased.

[0055] The level of antimicrobial peptides in the gut refers to the concentration or activity of antimicrobial peptides (AMPs) secreted by the gut. Antimicrobial peptides are polypeptides with antimicrobial activity, exhibiting broad-spectrum and highly effective bactericidal activity against bacteria.

[0056] In a specific embodiment of this application, the improvement in antimicrobial peptide levels in the intestines of stroke patients is manifested by increased expression levels of Reg3g, and / or Defa, and / or Lyz, and / or Ang4 genes in the intestines of stroke patients. That is, after drug administration to stroke patients, compared to no drug administration, the expression levels of Reg3g, Defa, Lyz, and Ang4 genes in the patient's intestines are increased. The encoded Reg3g is regenerating islet-derived protein 3γ, Defa is α-defensin, Lyz is lysozyme, and Ang4 is angiopoietin 4.

[0057] In a specific embodiment of this application, the content of quinoline acid in the drug can be from 0.00001% w / w to 99.99% w / w.

[0058] The kynurenic acid content in the medicine can be selected from 0.00001% w / w, 0.0001% w / w, 0.001% w / w, 0.01% w / w, 0.1% w / w, 1% w / w, 2% w / w, 3% w / w, 4% w / w, 5% w / w, 6% w / w, 7% w / w, 8% w / w, 9% w / w, 10% w / w, 15% w / w, 20% w / w, 25% w / w, 30% w / w, 35% w / w, 40% w / w, 45% w / w, 50% w / w, 55% w / w, 60% w / w, 65% w / w, 70% w / w, 75% w / w, 80% w / w, 85% w / w, 90% w / w, 9 1% w / w, 92% w / w, 93% w / w, 94% w / w, 95% w / w, 96% w / w, 97% w / w, 98% w / w, 99% w / w, 99.9% w / w, 99.99% w / w.

[0059] In a specific embodiment of this application, when the drug is used, the dosage of canine quinolinic acid is 0.01 to 200 mg / kg; preferably, the dosage of canine quinolinic acid is 4.4 to 13.5 mg / kg.

[0060] The dosage of kynurenic acid is selected from 0.01 mg / kg, 0.1 mg / kg, 1 mg / Kg, 2 mg / Kg, 3 mg / Kg, 4 mg / Kg, 4.4 mg / Kg, 4.5 mg / Kg, 4.6 mg / Kg, 4.7 mg / Kg, 4.8 mg / Kg, 4.9 mg / Kg, 5.0 mg / Kg, 5.1 mg / Kg, 5.2 mg / Kg, 5.3 mg / Kg, 5.4 mg / Kg, 5.5 mg / Kg, 5.6 mg / Kg, 5.7 mg / Kg, 5.8 mg / Kg, 5.9 mg / Kg, 6.0 mg / Kg, 6.1 mg / Kg, 6.2 mg / Kg, 6.3 mg / Kg, 6.4 mg / Kg, 6.5 mg / Kg, 6.6 mg / Kg, 6.7 mg / Kg, 6.8 mg / Kg, 6.9 mg / Kg, 7.0 mg / Kg, 7.1 mg / Kg, 7.2 mg / Kg, 7.3 mg / Kg, 7.4 mg / Kg, 7.5 mg / Kg, 7.6 mg / Kg, 7.7 mg / Kg, 7.8 mg / Kg, 7.9 mg / Kg, 8.0 mg / Kg, 8.1 mg / Kg, 8.2 mg / Kg, 8.3 mg / Kg, 8.4 mg / Kg, 8.5 mg / Kg, 8.8 mg / Kg, 8.7 mg / Kg, 8.8 mg / Kg, 8.9 mg / Kg, 9.0 mg / Kg, 9.1 mg / Kg, 9.2 mg / Kg, 9.3 mg / Kg, 9.4 mg / Kg, 9.5 mg / Kg, 9.6 mg / Kg, 9.7 mg / Kg, 9.8 mg / Kg, 9.9 mg / Kg, 10.0 mg / Kg, 10.1 mg / Kg, 10.2 mg / Kg, 10.3 mg / Kg, 10.4 mg / Kg, 10.5 mg / Kg, 10.6 mg / Kg, 10.7 mg / Kg, 10.8 mg / Kg, 10.9 mg / Kg, 11.0 mg / Kg, 11.1 mg / Kg, 11.2 mg / Kg, 11.3 mg / Kg, 11.4 mg / Kg, 11.5 mg / Kg, 11.6 mg / Kg, 11.7 mg / Kg, 11.8 mg / Kg, 11.9 mg / Kg, 12 mg / Kg, 12.1 mg / Kg, 12.2 mg / Kg, 12.3 mg / Kg, 12.4 mg / Kg, 12.5 mg / Kg, 12.6 mg / Kg, 12.7 mg / Kg, 12.8 mg / Kg, 12.9 mg / Kg, 13 mg / Kg, 13.1 mg / Kg, 13.2 mg / Kg, 13.3 mg / Kg, 13.4 mg / Kg, 13.5mg / Kg, 15mg / Kg, 20mg / Kg, 25mg / Kg, 30mg / Kg, 35mg / Kg, 40mg / Kg, 45mg / Kg, 50mg / Kg, 55mg / Kg, 60mg / Kg, 65mg / Kg, 70mg / Kg, 75mg / Kg, 80mg / Kg, 85 mg / Kg, 90mg / Kg, 95mg / Kg, 100mg / Kg, 110mg / Kg, 120mg / Kg, 130mg / Kg, 140mg / Kg, 150mg / Kg, 160mg / Kg, 170mg / Kg, 180mg / Kg, 190mg / Kg, 200mg / Kg. .

[0061] This application also provides the use of kynurenic acid in the preparation of a medicament, said medicament being used in any one or more of the following:

[0062] 1) Prevention and / or treatment of cerebral infarction in stroke patients;

[0063] 2) Improves the severity of cerebral infarction in stroke patients;

[0064] 3) Improves neurological function in stroke patients;

[0065] 4) Improves the systemic inflammatory phenotype in stroke patients;

[0066] 5) Improves gut microbiota dysbiosis in stroke patients;

[0067] 6) Improves abnormal intestinal barrier function in stroke patients;

[0068] 7) Improves the level of antimicrobial peptides in the gut of stroke patients.

[0069] In a specific embodiment of this application, the improvement in the degree of cerebral infarction in stroke patients is manifested as a reduction in the volume and proportion of cerebral infarction, as described above.

[0070] In a specific embodiment of this application, the improvement in the neurological function of stroke patients is defined as a decrease in the patient's neurological function score, as described above.

[0071] In a specific embodiment of this application, the improvement of the systemic inflammatory phenotype in stroke patients is manifested by a decrease in the levels of inflammatory factors IFN-γ and TNF-α in the serum of stroke patients, as described above.

[0072] In a specific embodiment of this application, the improvement of abnormal intestinal barrier function in stroke patients is manifested by increased expression levels of Muc1, and / or Tjp1, and / or Ocln, and / or Cldn4 genes in the intestine of stroke patients, as described above.

[0073] In a specific embodiment of this application, the improvement of antimicrobial peptide levels in the intestines of stroke patients is manifested by increased expression levels of Reg3g, and / or Defa, and / or Lyz, and / or Ang4 genes in the intestines of stroke patients, as described above.

[0074] In a specific embodiment of this application, the stroke is selected from ischemic stroke or hemorrhagic stroke.

[0075] As some embodiments of this application, the stroke described is ischemic stroke.

[0076] In a specific embodiment of this application, the content of quinoline acid in the drug can be from 0.00001% w / w to 99.99% w / w.

[0077] The kynurenic acid content in the medicine can be selected from 0.00001% w / w, 0.0001% w / w, 0.001% w / w, 0.01% w / w, 0.1% w / w, 1% w / w, 2% w / w, 3% w / w, 4% w / w, 5% w / w, 6% w / w, 7% w / w, 8% w / w, 9% w / w, 10% w / w, 15% w / w, 20% w / w, 25% w / w, 30% w / w, 35% w / w, 40% w / w, 45% w / w, 50% w / w, 55% w / w, 60% w / w, 65% w / w, 70% w / w, 75% w / w, 80% w / w, 85% w / w, 90% w / w, 9 1% w / w, 92% w / w, 93% w / w, 94% w / w, 95% w / w, 96% w / w, 97% w / w, 98% w / w, 99% w / w, 99.9% w / w, 99.99% w / w.

[0078] In a specific embodiment of this application, when the drug is used, the dosage of canine quinolinic acid is 0.01 to 200 mg / kg; preferably, the dosage of canine quinolinic acid is 4.4 to 13.5 mg / kg.

[0079] The dosage of kynurenic acid is selected from 0.01 mg / kg, 0.1 mg / kg, 1 mg / Kg, 2 mg / Kg, 3 mg / Kg, 4 mg / Kg, 4.4 mg / Kg, 4.5 mg / Kg, 4.6 mg / Kg, 4.7 mg / Kg, 4.8 mg / Kg, 4.9 mg / Kg, 5.0 mg / Kg, 5.1 mg / Kg, 5.2 mg / Kg, 5.3 mg / Kg, 5.4 mg / Kg, 5.5 mg / Kg, 5.6 mg / Kg, 5.7 mg / Kg, 5.8 mg / Kg, 5.9 mg / Kg, 6.0 mg / Kg, 6.1 mg / Kg, 6.2 mg / Kg, 6.3 mg / Kg, 6.4 mg / Kg, 6.5 mg / Kg, 6.6 mg / Kg, 6.7 mg / Kg, 6.8 mg / Kg, 6.9 mg / Kg, 7.0 mg / Kg, 7.1 mg / Kg, 7.2 mg / Kg, 7.3 mg / Kg, 7.4 mg / Kg, 7.5 mg / Kg, 7.6 mg / Kg, 7.7 mg / Kg, 7.8 mg / Kg, 7.9 mg / Kg, 8.0 mg / Kg, 8.1 mg / Kg, 8.2 mg / Kg, 8.3 mg / Kg, 8.4 mg / Kg, 8.5 mg / Kg, 8.8 mg / Kg, 8.7 mg / Kg, 8.8 mg / Kg, 8.9 mg / Kg, 9.0 mg / Kg, 9.1 mg / Kg, 9.2 mg / Kg, 9.3 mg / Kg, 9.4 mg / Kg, 9.5 mg / Kg, 9.6 mg / Kg, 9.7 mg / Kg, 9.8 mg / Kg, 9.9 mg / Kg, 10.0 mg / Kg, 10.1 mg / Kg, 10.2 mg / Kg, 10.3 mg / Kg, 10.4 mg / Kg, 10.5 mg / Kg, 10.6 mg / Kg, 10.7 mg / Kg, 10.8 mg / Kg, 10.9 mg / Kg, 11.0 mg / Kg, 11.1 mg / Kg, 11.2 mg / Kg, 11.3 mg / Kg, 11.4 mg / Kg, 11.5 mg / Kg, 11.6 mg / Kg, 11.7 mg / Kg, 11.8 mg / Kg, 11.9 mg / Kg, 12 mg / Kg, 12.1 mg / Kg, 12.2 mg / Kg, 12.3 mg / Kg, 12.4 mg / Kg, 12.5 mg / Kg, 12.6 mg / Kg, 12.7 mg / Kg, 12.8 mg / Kg, 12.9 mg / Kg, 13 mg / Kg, 13.1 mg / Kg, 13.2 mg / Kg, 13.3 mg / Kg, 13.4 mg / Kg, 13.5mg / Kg, 15mg / Kg, 20mg / Kg, 25mg / Kg, 30mg / Kg, 35mg / Kg, 40mg / Kg, 45mg / Kg, 50mg / Kg, 55mg / Kg, 60mg / Kg, 65mg / Kg, 70mg / Kg, 75mg / Kg, 80mg / Kg, 85 mg / Kg, 90mg / Kg, 95mg / Kg, 100mg / Kg, 110mg / Kg, 120mg / Kg, 130mg / Kg, 140mg / Kg, 150mg / Kg, 160mg / Kg, 170mg / Kg, 180mg / Kg, 190mg / Kg, 200mg / Kg. .

[0080] This application also provides a medicine for the prevention and / or relief and / or treatment of stroke or its symptoms, including an effective amount of canine quinolinic acid.

[0081] The drug is used to treat stroke or its symptoms, wherein the stroke is selected from ischemic stroke or hemorrhagic stroke; preferably, the stroke is ischemic stroke.

[0082] The drug can prevent and / or alleviate and / or treat cerebral infarction in stroke patients, improve the degree of cerebral infarction in stroke patients, improve neurological function in stroke patients, improve systemic inflammatory phenotype in stroke patients, improve gut microbiota dysbiosis in stroke patients, improve intestinal barrier dysfunction in stroke patients, and improve antimicrobial peptide levels in the gut of stroke patients, as specifically described above.

[0083] The drug in this application can be used on mammals, such as but not limited to humans, primates, livestock (e.g., sheep, cattle, horses, donkeys, pigs), pets (e.g., dogs, cats), laboratory test animals (e.g., mice, rabbits, rats, guinea pigs, hamsters) or captured wild animals (e.g., foxes, deer); preferably, the drug can be used on mice or humans.

[0084] The drug described in this application is intended for patients who have had, are currently having, or are at risk of having a stroke.

[0085] The dosage form or state of the drug in this application is not limited. It may be in solid form, such as capsules, tablets, pills, granules, small capsules or lozenges; or in liquid form, such as solutions, suspensions, emulsions or syrups.

[0086] The drug in this application is administered via the gastrointestinal tract. Research conducted in this application has shown that administration of the drug via the gastrointestinal tract has better therapeutic effects than other methods.

[0087] The gastrointestinal administration method is selected from one or more of oral administration, gastric tube administration, and enteral administration.

[0088] The drug in this application may also contain pharmaceutically acceptable excipients or carriers.

[0089] Pharmaceutically acceptable excipients or carriers include one or more components selected from the following: binders, fillers, lubricants, flow aids, disintegrants, and humectants.

[0090] The adhesive is selected from one or more of the following: gum arabic, alginate, calcium carboxymethyl cellulose, sodium carboxymethyl cellulose, glucose binder, dextrin, dextran, ethyl cellulose, gelatin, liquid glucose, guar gum, hydroxyethyl cellulose, hydroxypropyl cellulose, hydroxypropyl methyl cellulose, magnesium aluminum silicate, maltodextrin, methyl cellulose, polymethyl methacrylate, polyvinylpyrrolidone, pregelatinized starch, sodium alginate, sorbitol, starch, and syrup.

[0091] The filler is selected from one or more of the following: calcium carbonate, calcium phosphate, calcium sulfate, calcium carboxymethyl cellulose, sodium carboxymethyl cellulose, concentrated sugar, sugar paste, glucose binder, dextrin, dextrose, calcium hydrogen phosphate dihydrate, calcium hydrogen phosphate, fructose, palmitate stearate, glycerol, hydrogenated vegetable oil type 1, kaolin, lactose, corn starch, magnesium carbonate, magnesium oxide, maltodextrin, mannitol, microcrystalline cellulose, polymethyl methacrylate, potassium chloride, powdered cellulose, pregelatinized starch, sodium chloride, sorbitol, starch, sucrose, sugar spheres, talc, calcium phosphate, and xylitol.

[0092] The lubricant is selected from one or more of the following: calcium stearate, glyceryl monostearate, glyceryl palmitate, magnesium stearate, microcrystalline cellulose, sodium benzoate, sodium chloride, sodium lauryl sulfate, stearic acid, sodium stearyl fumarate, talc, and zinc stearate.

[0093] The flow aid is selected from one or more of colloidal silica, powdered cellulose, magnesium trisilicate, silica, and talc.

[0094] The disintegrant is selected from one or more of the following: alginate, calcium carboxymethyl cellulose, sodium carboxymethyl cellulose, colloidal silica, cross-linked sodium carboxymethyl cellulose, polyvinylpyrrolidone, guar gum, magnesium aluminum silicate, microcrystalline cellulose, methyl cellulose, polyvinylpyrrolidone, potassium polacrine, pregelatinized starch, sodium alginate, sodium dodecyl sulfate, and sodium starch glycolate.

[0095] The medicines mentioned in this application may include health care products.

[0096] Health products generally refer to products that share the common characteristics of ordinary foods, can regulate bodily functions, and are suitable for consumption by specific groups of people, but are not intended to treat diseases; they are also known as dietary supplements. These include tea, wine, bee products, beverages, soups, fresh juices, and medicinal cuisine.

[0097] The drugs mentioned in this application may also include biological products.

[0098] Biological products generally refer to products prepared using biological materials such as microorganisms, cells, and various animal and human tissues and fluids obtained through common or genetic engineering, cell engineering, protein engineering, fermentation engineering, and other biotechnologies, for the prevention, treatment, and diagnosis of human diseases. These include bacterial vaccines, toxins, toxoids, blood products, immunoglobulins, antigens, allergens, cytokines, hormones, enzymes, fermentation products, monoclonal antibodies, and recombinant DNA products.

[0099] In one embodiment of this application, the drug is formulated to be administered orally, for example, orally.

[0100] For example, oral dosage forms may include capsules, tablets, pills, granules, or syrups.

[0101] For example, oral dosages can be in the range of about 0.01 to 200 mg / kg body weight, about 0.01 to 180 mg / kg, about 0.01 to 160 mg / kg, about 0.01 to 140 mg / kg, about 0.01 to 120 mg / kg, about 0.01 to 100 mg / kg, about 0.1 to 200 mg / kg, about 0.1 to 150 mg / kg, and about 0.1 to 100 mg / kg. Dosages of approximately 0.1 to 80 mg / kg, approximately 1 to 60 mg / kg, approximately 0.1 to 40 mg / kg, approximately 0.1 to 20 mg / kg, 25-200 mg / kg, approximately 50-200 mg / kg, approximately 100-200 mg / kg, approximately 25-50 mg / kg, approximately 25-100 mg / kg, approximately 50-100 mg / kg, and approximately 50-200 mg / kg are administered.

[0102] For example, the oral administration method may include taking it once or more daily, daily, every other day, weekly, every two weeks, monthly, or every two months.

[0103] In this application, the kynurenic acid is substantially not broken down and / or inactivated by digestive juices. These digestive juices may include saliva, gastric juice, intestinal juice, pancreatic juice, and bile.

[0104] For example, saliva, with a pH of 6.6 to 7.1, mainly consists of salivary amylase, lysozyme, and small amounts of inorganic substances (such as inorganic salts containing sodium, potassium, and calcium). The kynuroline acid is essentially not broken down and / or inactivated by the saliva.

[0105] For example, gastric juice, with a pH of 0.9 to 1.5, mainly consists of pepsin, gastric acid (i.e., hydrochloric acid), mucus, and inorganic substances such as sodium and potassium salts. The kynuroline acid is essentially not broken down and / or inactivated by the gastric juice.

[0106] For example, pancreatic juice, with a pH of 7.8 to 8.4, mainly consists of sodium bicarbonate, pancreatic amylase, pancreatic lipase, trypsinogen, and chymotrypsinogen, etc., and the kynurenic acid is essentially not decomposed and / or inactivated by the pancreatic juice.

[0107] For example, bile, with a pH of approximately 6.8 to 7.4, is mainly composed of bile salts and bile pigments, and the kynurenic acid is essentially not broken down and / or inactivated by the bile.

[0108] For example, intestinal fluid, with a pH of approximately 7.6, contains various digestive enzymes such as amylase, maltase, sucrase, lactase, peptidase, and lipase. The kynurenic acid is essentially not broken down and / or inactivated by the intestinal fluid.

[0109] For example, the quinolinic acid is substantially not broken down and / or inactivated by saliva, gastric juice, pancreatic juice, bile, or small intestinal juice.

[0110] For example, the kynurenic acid is substantially not decomposed and / or inactivated at pH values ​​of 6.6–7.1, 0.9–1.5, 7.8–8.4, 6.8–7.4, and 7.6.

[0111] This application also provides a method for preventing and / or alleviating and / or treating stroke, which involves administering the drug used for the above purposes, or the drug described above, to a patient.

[0112] In a specific embodiment of this application, the stroke is selected from ischemic stroke or hemorrhagic stroke; preferably, the stroke is ischemic stroke.

[0113] In a specific embodiment of this application, when the drug is used, the dosage of quinoline acid is 0.01 to 200 mg / kg; preferably, when the drug is used, the dosage of quinoline acid is 4.4 mg / kg to 13.5 mg / kg.

[0114] The dosage of kynurenic acid is selected from 0.01 mg / kg, 0.1 mg / kg, 1 mg / Kg, 2 mg / Kg, 3 mg / Kg, 4 mg / Kg, 4.4 mg / Kg, 4.5 mg / Kg, 4.6 mg / Kg, 4.7 mg / Kg, 4.8 mg / Kg, 4.9 mg / Kg, 5.0 mg / Kg, 5.1 mg / Kg, 5.2 mg / Kg, 5.3 mg / Kg, 5.4 mg / Kg, 5.5 mg / Kg, 5.6 mg / Kg, 5.7 mg / Kg, 5.8 mg / Kg, 5.9 mg / Kg, 6.0 mg / Kg, 6.1 mg / Kg, 6.2 mg / Kg, 6.3 mg / Kg, 6.4 mg / Kg, 6.5 mg / Kg, 6.6 mg / Kg, 6.7 mg / Kg, 6.8 mg / Kg, 6.9 mg / Kg, 7.0 mg / Kg, 7.1 mg / Kg, 7.2 mg / Kg, 7.3 mg / Kg, 7.4 mg / Kg, 7.5 mg / Kg, 7.6 mg / Kg, 7.7 mg / Kg, 7.8 mg / Kg, 7.9 mg / Kg, 8.0 mg / Kg, 8.1 mg / Kg, 8.2 mg / Kg, 8.3 mg / Kg, 8.4 mg / Kg, 8.5 mg / Kg, 8.8 mg / Kg, 8.7 mg / Kg, 8.8 mg / Kg, 8.9 mg / Kg, 9.0 mg / Kg, 9.1 mg / Kg, 9.2 mg / Kg, 9.3 mg / Kg, 9.4 mg / Kg, 9.5 mg / Kg, 9.6 mg / Kg, 9.7 mg / Kg, 9.8 mg / Kg, 9.9 mg / Kg, 10.0 mg / Kg, 10.1 mg / Kg, 10.2 mg / Kg, 10.3 mg / Kg, 10.4 mg / Kg, 10.5 mg / Kg, 10.6 mg / Kg, 10.7 mg / Kg, 10.8 mg / Kg, 10.9 mg / Kg, 11.0 mg / Kg, 11.1 mg / Kg, 11.2 mg / Kg, 11.3 mg / Kg, 11.4 mg / Kg, 11.5 mg / Kg, 11.6 mg / Kg, 11.7 mg / Kg, 11.8 mg / Kg, 11.9 mg / Kg, 12 mg / Kg, 12.1 mg / Kg, 12.2 mg / Kg, 12.3 mg / Kg, 12.4 mg / Kg, 12.5 mg / Kg, 12.6 mg / Kg, 9mg / Kg, 12.7 mg / Kg, 12.8 mg / Kg, 12.9 mg / Kg, 13 mg / Kg, 13.1 mg / Kg, 13.2 mg / Kg, 13.3 mg / Kg, 13.4 mg / Kg, 13.5mg / Kg, 15mg / Kg, 20mg / Kg, 25mg / Kg, 30mg / Kg, 35mg / Kg, 40mg / Kg, 45mg / Kg, 50mg / Kg, 55mg / Kg, 60mg / Kg, 65mg / Kg, 70mg / Kg, 75mg / Kg, 80mg / Kg, 85 mg / Kg, 90mg / Kg, 95mg / Kg, 100mg / Kg, 110mg / Kg, 120mg / Kg, 130mg / Kg, 140mg / Kg, 150mg / Kg, 160mg / Kg, 170mg / Kg, 180mg / Kg, 190mg / Kg, 200mg / Kg. .

[0115] In a specific embodiment of this application, the patient is a patient who has had, is currently having, or is at risk of having a stroke.

[0116] In specific embodiments of this application, the patient is a mammal, such as, but not limited to, a human, a primate, livestock (e.g., sheep, cattle, horses, donkeys, pigs), a pet (e.g., dogs, cats), a laboratory test animal (e.g., mice, rabbits, rats, guinea pigs, hamsters), or a captured wild animal (e.g., foxes, deer); preferably, the patient is a mouse or a human.

[0117] In a specific embodiment of this application, the drug is administered via the gastrointestinal tract; preferably, the gastrointestinal administration is selected from one or more of oral administration, gastric tube administration, and enteral administration. Research conducted in this application has found that administering the drug via the gastrointestinal tract provides better therapeutic effects than other methods.

[0118] The present application will be described below through specific embodiments. Unless otherwise stated, the experimental methods, detection methods, and preparation methods disclosed in this invention all employ conventional techniques in molecular biology, biochemistry, chromatin structure and analysis, analytical chemistry, cell culture, recombinant DNA technology, and related fields. These techniques have been well described in existing literature; see Sambrook et al., *MOLECULAR CLONING: A LABORATORY MANUAL*, Second edition, Cold Spring Harbor Laboratory Press, 1989 and Third edition, 2001; Ausubel et al., *CURRENT PROTOCOLS IN MOLECULAR BIOLOGY*, John Wiley & Sons, New York, 1987 and periodic updates; the series *METHODS IN ENZYMOLOGY*, Academic Press, San Diego; Wolffe, *CHROMATIN STRUCTURE AND FUNCTION*, Third edition, Academic Press, San Diego, 1998; *METHODS IN ENZYMOLOGY*, Vol. 304, Chromatin (PM Wassarman and AP Wolffe, eds.), Academic Press, San Diego, 1999; and *METHODS IN MOLECULAR*. BIOLOGY, Vol. 119, Chromatin Protocols (PB Becker, ed.) Humana Press, Totowa, 1999, etc.

[0119] Example 1: Construction and Drug Administration of a Mouse Model

[0120] In this embodiment, mice with the MCAO model were continuously treated with quinolinic acid in their urine. Male C57BL / 6 mice aged 8-10 weeks, weighing 25-30g, were used in this embodiment. After one week of acclimatization, they were randomly divided into three groups: (1) MCAO group: Mice with normal feeding underwent MCAO modeling, and the phenotype of the mice was observed 3 days after MCAO modeling; (2) Low-KYNA group: Mice were treated with low-concentration quinolinic acid (40mg / Kg) via gavage for 2 weeks before undergoing MCAO modeling surgery, followed by 3 days of treatment with the same concentration of quinolinic acid after surgery; (3) High-KYNA group: Mice were treated with high-concentration quinolinic acid (120mg / Kg) via gavage for 2 weeks before undergoing MCAO modeling surgery, followed by 3 days of treatment with the same concentration of quinolinic acid after surgery. All gavage interventions were performed between 8-10 am daily.

[0121] Example 2: Kynurenic acid can improve the degree of cerebral infarction in mice after stroke.

[0122] This embodiment observes the effect of kynurenic acid on the proportion of infarct volume after stroke in mice.

[0123] Experimental methods:

[0124] The method for preparing brain slice staining is as follows:

[0125] (1) The MCAO model obtained in the manner described above;

[0126] (2) Prepare TTC powder using PBS. The concentration of TTC staining solution is 2%. Store in the dark.

[0127] (3) After anesthetizing the mice (0.2 ml / 10 g tribromoethanol), open the thoracic cavity, cut open the right atrial appendage, and perfuse the heart with frozen PBS for about 2 min;

[0128] (4) The brain was removed after decapitation. The brain tissue was placed in the brain groove, and then the brain groove was placed in a -80℃ freezer for 7-8 minutes before being removed and a 1.5mm thick coronal section was prepared.

[0129] (5) Place the cut brain tissue in TTC staining solution and stain for 10 minutes in the dark. Normal brain tissue is dark red, while the ischemic stroke area is unstained (grayish-white). After fixing the stained sections with formaldehyde, remove them and take pictures;

[0130] (6) ImagePlus Soft image software was used to analyze the brain injury volume. The brain edema factor was excluded from the analysis results, and the infarct volume was calculated using the formula for edema correction: brain injury area = direct injury volume - (ipsilateral hemisphere of the body - contralateral hemisphere of the body), and finally the proportion of brain injury volume to the total cerebral hemisphere was obtained.

[0131] Experimental results:

[0132] The results showed that, compared with the MCAO group, the proportion of cerebral infarction volume in both the Low-KYNA and High-KYNA groups was significantly reduced, suggesting that canine quinolinic acid intervention significantly restored post-stroke damage in mice (see Figure 1).

[0133] Example 3: Quinolinic acid can improve neurological function in mice after stroke.

[0134] In this embodiment, mice with MCAO model were continuously treated with canine quinolinic acid (Low-KYNA group, High-KYNA group, intervention procedure and dosage were the same as in Example 1) to observe its effect on neurological function after stroke in mice.

[0135] Experimental methods:

[0136] Neurological function was assessed in mice of the MCAO, Low-KYNA, and High-KYNA groups at specified time points after MCAO modeling. The specific scoring criteria are shown in Table 1 below.

[0137] Table 1. Detailed Rules for Neurological Function Scoring

[0138] The results showed that, compared with the MCAO group, the neurological function scores of mice in both the Low-KYNA and High-KYNA groups were significantly reduced, indicating that the neurological function of mice was significantly restored after quinolinic acid intervention (see Figure 2).

[0139] Example 4: Quinolinic acid can improve the systemic inflammatory phenotype in mice after stroke.

[0140] In this embodiment, kynurenic acid was continuously administered to mice with the MCAO model (Low-KYNA group, High-KYNA group, intervention procedure and dosage were the same as in Example 1) to observe the effect of kynurenic acid intervention on the serum levels of inflammatory factors IFN-γ and TNF-α in mice after stroke.

[0141] Experimental methods:

[0142] Blood samples were collected from mice after neurological function assessment. Blood collection methods included orbital sampling and posterior vena cava sampling. Mouse serum samples were stored at -80°C before testing. The levels of inflammatory factors such as IFN-γ and TNF-α in mouse serum were measured strictly according to the instructions of the ELISA kit (GeneMed, ELISA Lab). A standard curve was constructed, and the expression levels of each factor were calculated based on the standard curve.

[0143] The results showed that, compared with the MCAO group, the serum inflammatory factor IFN-γ levels in mice in the Low-KYNA and High-KYNA groups were significantly reduced. The serum inflammatory factor TNF-α levels in mice in both the Low-KYNA and High-KYNA groups were significantly reduced, suggesting that quinolinic acid intervention can significantly alleviate the phenotype of systemic inflammation after stroke in mice (see Figure 3).

[0144] Example 5: Quinolinic acid can improve gut microbiota dysbiosis in mice after stroke.

[0145] In this embodiment, kynurenic acid was continuously administered to mice with the MCAO model (Low-KYNA group, High-KYNA group, intervention procedure and dosage were the same as in Example 1) to observe the effect of kynurenic acid intervention on the gut microbiota structure after stroke in mice.

[0146] Experimental methods:

[0147] Extraction of total DNA from intestinal contents (total DNA of gut microbiota)

[0148] In mice from the MCAO group, Low-KYNA group, High-KYNA group, and blank control group, total bacterial DNA was extracted from samples of jejunal, ileal, cecal, and colonic contents using the MinkaGene Stool DNA Kit. The extraction procedure was strictly performed according to the kit's instructions. The extracted total bacterial DNA was stored at -80°C for subsequent PCR amplification of the target gene.

[0149] PCR amplification of the gene signature sequence of bacterial 16S rRNA V4 region

[0150] The bacterial total DNA sample was amplified using universal primers with a barcode on the 16S rRNA V4 variable region. The PCR cycle parameters were set as shown in Table 2 below.

[0151] Table 2 PCR Cycling Steps

[0152] The PCR system setup was performed under aseptic conditions using a biosafety cabinet. The PCR reaction volume was 25 μl. The PCR reaction system is shown in Table 3 below:

[0153] Table 3 PCR reaction system:

[0154] PCR amplification was performed using universal primers with barcodes in the V4 region of 16S rRNA. The universal primers were V4F: GTGYCAGCMGCCGCGGTAA (SEQ ID No. 19); V4R: GGACTACNVGGGTWTCTAAT (SEQ ID No. 20).

[0155] After amplification, the PCR products were stored in a -20°C freezer, awaiting mixing and sequencing.

[0156] Sequencing and microbial community data analysis

[0157] The PCR products of the successfully amplified target band were quantitatively mixed at a concentration of 100 μg / L and the bacterial community genome was sequenced in our laboratory using Illumina Iseq100 (PE 150) sequencing technology. All raw sequences were preprocessed using BIPES sequencing technology (see Zhou HW, Li DF, Tam NF, et al. BIPES, a cost-effective high-throughput method for assessing microbial diversity. ISME J, 2011, 5:741-749).

[0158] Sequencing results were processed using bioinformatics tools such as Mothur, QiIME, and BIPES. Sequences underwent quality control, chimera removal, and assembly to obtain target tag sequences, which were then sorted into individual samples. Further, OTU species classification was performed using Usearch or UPARSE clustering (see He Y, Caporaso JG, Jiang XT, et al. Stability of operational taxonomic units: an important but neglected property for analyzing microbial diversity[J]. Microbiome, 2015, 3:20). The final results were then used to obtain corresponding alpha and beta diversity parameters, and multivariate statistical analyses such as PCoA and PCA were performed. Finally, tools such as LEfSe were used to identify differences in bacterial community structure among different groups.

[0159] OTUs (Operational Taxonomic Units): In bioinformatics analysis, each sequence obtained from sequencing represents a bacterium. Sequences are clustered based on their similarity (specifying similarity as 96%, 97%, or 98%) into groups (each group being an OTU). After OTU classification, bioinformatics statistical analysis is performed to determine the number of bacterial species, genera, etc., in a given sequencing sample. The PyNAST algorithm is used to align and calibrate the sequences of each OTU, with the representative sequence of each OTU determined by its frequency. Chimeras are then removed, and the representative sequences of the OTUs are inserted into FastTree software for processing, generating a phylogenetic tree containing the metabolic sequences of all OTUs. Microbial community analysis is then performed based on this phylogenetic tree.

[0160] Principal component analysis (PCA) has been widely used to compare the community structures of different gut microbiota. PCA uses dimensionality reduction methods based on the correlation between samples to transform multiple indicators into a few comprehensive indicators for analysis. In this embodiment, a principal coordinate analysis (PCoA) method, similar to PCA, is used to analyze the β-diversity between groups using both unweighted and weighted methods. The PCoA analysis method starts from the clustering matrix between samples and, through rotation of the three-dimensional coordinate axes, re-calibrates the samples in a new coordinate system based on the cluster distance.

[0161] PCoA results showed that the β-diversity of gut microbiota in mice after stroke modeling was significantly altered. The Unweighted UniFrac results are shown in Figure 4. Compared with the blank control group, the gut microbiota in the jejunum, ileum, cecum, and colon of mice after MCAO modeling (MCAO group) was significantly dysregulated.

[0162] The results showed that, compared with the MCAO group, mice in the Low-KYNA and High-KYNA groups exhibited overall recovery of gut microbiota in the jejunum, ileum, cecum, and colon, suggesting that canine quinolinic acid intervention can significantly alleviate the disordered gut microbiota in mice after stroke (see Figure 4).

[0163] Example 6: Quinolinic acid can improve intestinal barrier dysfunction in mice after stroke.

[0164] In this embodiment, MCAO model mice were continuously treated with canine quinolinic acid (Low-KYNA group, High-KYNA group, intervention procedure and dosage were the same as in Example 1). The effects of canine quinolinic acid intervention on the levels of intestinal barrier-related genes Muc1, Tjp1, Ocln, and Cldn4 in intestinal tissue after stroke in mice were observed using upstream and downstream primers as shown in SEQ ID NO: 1-10.

[0165] At specified time points after stroke modeling, ileum and colon tissues of mice were rapidly collected and frozen at -80°C to determine the relative expression levels of genes related to the intestinal barrier pathway, including Muc1, Tjp1, Ocln, and Cldn4. The primer sequences are shown in Table 4.

[0166] Table 4 Primer sequences for gene PCR amplification

[0167] The specific steps are as follows:

[0168] (1) Use Minibeadbeater (Biospec Products, Bartlesville) to homogenize colon tissue;

[0169] (2) RNA extraction using the Trizol reagent method (Invitrogen): Add 1 ml of TRIZOL to every 50-100 mg of homogenized tissue sample for homogenization. Then add 0.2 ml of chloroform, cover, and shake vigorously for 15 seconds. Incubate at 15-30°C for 2-3 minutes, then centrifuge at 12,000 g for 15 minutes at 2-8°C. Transfer the upper aqueous phase, add 0.5 ml of isopropanol, incubate for 10 minutes at 15-30°C, then centrifuge at 12,000 g for 10 minutes at 2-8°C. Wash the RNA precipitate with 1 ml of 75% ethanol for 5 minutes. Dissolve the RNA and determine its concentration.

[0170] (3) cDNA was obtained from each RNA sample using TaqMan reverse transcription reagent (Takara Bio);

[0171] (4) Add 2 μl of cDNA as template (concentration of 50-100 ng / μl) to a 20 μl volume, and add the upstream and downstream primers as shown in SEQ ID NO: 1-10 at a final concentration of 250 nM to perform SYBR Green (TakaraBio) real-time PCR. The reaction system is the same as in Table 3.

[0172] (5) Data were extracted using the ViiA 7 real-time PCR system (Applied Biosystems) and analyzed using the comparative Ct method. The expression levels of each Gapdh gene were used as internal controls, and their primer sequences are shown in Table 4.

[0173] Experimental results:

[0174] The results showed that, compared with the MCAO group, the relative expression levels of Muc1, Tjp1, Ocln, and Cldn4 genes in the ileum of mice in both the Low-KYNA and High-KYNA groups were significantly increased. The relative expression levels of Tjp1, Ocln, and Cldn4 genes in the colon of mice in the Low-KYNA group were significantly increased. The relative expression levels of Muc1, Tjp1, Ocln, and Cldn4 genes in the colon of mice in the High-KYNA group were also significantly increased.

[0175] The above results suggest that canine quinolinic acid intervention can significantly improve intestinal barrier damage in mice after stroke (see Figure 5).

[0176] Example 7: Quinolinic acid can improve the level of antimicrobial peptides in the intestines of mice after stroke.

[0177] In this embodiment, mice with MCAO model were continuously treated with canine quinolinic acid (Low-KYNA group, High-KYNA group, intervention procedure and dosage were the same as in Example 1). The effects of canine quinolinic acid intervention on the levels of intestinal antimicrobial peptide-related genes Reg3g, Defa, Lyz, and Ang4 in intestinal tissue after stroke were observed using upstream and downstream primers as shown in SEQ ID NO: 9-18.

[0178] Experimental method: The primer sequences are shown in Table 4, and the detection method is the same as in Example 6.

[0179] The results showed that compared with the MCAO group, the relative expression levels of Reg3g, Defa, Lyz, and Ang4 genes in the ileum and colon tissues of mice in the Low-KYNA and High-KYNA groups were significantly increased, suggesting that canine quinolinic acid intervention can significantly improve the level of intestinal antimicrobial peptides in mice after stroke (see Figure 6).

[0180] The above embodiments are merely illustrative of the principles and effects of this application and are not intended to limit this application. Any person skilled in the art can modify or alter the above embodiments without departing from the spirit and scope of this application. Therefore, all equivalent modifications or alterations made by those skilled in the art without departing from the spirit and technical concept disclosed in this application should still be covered by the claims of this application.

Claims

1. Use of quinolinic acid in the preparation of medicines for the prevention and / or relief and / or treatment of stroke or its symptoms.

2. The use according to claim 1, characterized in that, The drug has one or more of the following effects: 1) To prevent and / or alleviate and / or treat cerebral infarction in stroke patients; 2) Improves the severity of cerebral infarction in stroke patients; 3) Improves neurological function in stroke patients; 4) Improves the systemic inflammatory phenotype in stroke patients; 5) Improves gut microbiota dysbiosis in stroke patients; 6) Improves abnormal intestinal barrier function in stroke patients; 7) Improves the level of antimicrobial peptides in the gut of stroke patients.

3. Use of kynurenic acid in the preparation of a medicament, wherein the medicament is used in any one or more of the following: 1) Prevention and / or treatment of cerebral infarction in stroke patients; 2) Improves the severity of cerebral infarction in stroke patients; 3) Improves neurological function in stroke patients; 4) Improves the systemic inflammatory phenotype in stroke patients; 5) Improves gut microbiota dysbiosis in stroke patients; 6) Improves abnormal intestinal barrier function in stroke patients; 7) Improves the level of antimicrobial peptides in the gut of stroke patients.

4. The use according to claim 2 or 3, characterized in that, The improvement in the degree of cerebral infarction in stroke patients is manifested by a reduction in the volume and proportion of cerebral infarction. And / or, the improvement in neurological function in stroke patients is a decrease in the patient's neurological function score; And / or, the improvement in the systemic inflammatory phenotype of stroke patients is manifested by a decrease in the serum inflammatory factors IFN-γ and / or TNF-α in stroke patients; And / or, the improvement of intestinal barrier function abnormality in stroke patients is manifested by increased expression levels of Muc1, and / or Tjp1, and / or Ocln, and / or Cldn4 genes in the intestine of stroke patients; And / or, the improvement in antimicrobial peptide levels in the gut of stroke patients is manifested by increased expression levels of Reg3g, and / or Defa, and / or Lyz, and / or Ang4 genes in the gut of stroke patients.

5. The use according to any one of claims 1 to 4, characterized in that, The stroke is selected from ischemic stroke or hemorrhagic stroke; preferably, the stroke is ischemic stroke. And / or, the kynurenic acid is the active ingredient in the drug, and when the drug is used, the dosage of kynurenic acid is 0.01 to 200 mg / kg; preferably, when the drug is used, the dosage of kynurenic acid is 4.4 to 13.5 mg / kg. And / or, the drug is used for patients who have had, are having, or are at risk of having a stroke; And / or, the drug is used in mammals; preferably, the drug is used in mice or humans; And / or, the drug contains pharmaceutically acceptable excipients or carriers; And / or, the drug is administered via the gastrointestinal tract; preferably, the gastrointestinal administration is selected from one or more of oral administration, gastric tube administration, and enteral administration; 6. Medications used to prevent and / or alleviate and / or treat stroke or its symptoms, including effective amounts of canine quinolinic acid.

7. The drug according to claim 6, characterized in that, The drug has one or more of the following effects: 1) To prevent and / or alleviate and / or treat cerebral infarction in stroke patients; 2) Improves the severity of cerebral infarction in stroke patients; 3) Improves neurological function in stroke patients; 4) Improves the systemic inflammatory phenotype in stroke patients; 5) Improves gut microbiota dysbiosis in stroke patients; 6) Improves abnormal intestinal barrier function in stroke patients; 7) Improves the level of antimicrobial peptides in the gut of stroke patients.

8. The medicament according to claim 7, characterized in that, The improvement in the degree of cerebral infarction in stroke patients is manifested by a reduction in the volume and proportion of cerebral infarction. And / or, the improvement in neurological function in stroke patients is a decrease in the patient's neurological function score; And / or, the improvement in the systemic inflammatory phenotype of stroke patients is manifested by a decrease in the serum inflammatory factors IFN-γ and / or TNF-α in stroke patients; And / or, the improvement of intestinal barrier function abnormality in stroke patients is manifested by increased expression levels of Muc1, and / or Tjp1, and / or Ocln, and / or Cldn4 genes in the intestine of stroke patients; And / or, the improvement in antimicrobial peptide levels in the gut of stroke patients is manifested by increased expression levels of Reg3g, and / or Defa, and / or Lyz, and / or Ang4 genes in the gut of stroke patients.

9. The medicament according to claim 8, characterized in that, The stroke is selected from ischemic stroke or hemorrhagic stroke; preferably, the stroke is ischemic stroke. And / or, when the drug is used, the dosage of quinoline acid is 0.01 to 200 mg / kg; preferably, when the drug is used, the dosage of quinoline acid is 4.4 to 13.5 mg / kg. And / or, the drug is used for patients who have had, are having, or are at risk of having a stroke; And / or, the drug is used in mammals; preferably, the drug is used in mice or humans; And / or, the drug contains pharmaceutically acceptable excipients or carriers; And / or, the drug is administered via the gastrointestinal tract; preferably, the gastrointestinal administration is selected from one or more of oral administration, gastric tube administration, and enteral administration.

10. A method for preventing and / or alleviating and / or treating stroke or its symptoms, comprising administering to a patient a drug for use as described in any one of claims 1 to 2, or a drug for use as described in any one of claims 3 to 5, or a drug for use as described in any one of claims 6 to 9.

11. The method according to claim 10, characterized in that, The stroke is selected from ischemic stroke or hemorrhagic stroke; preferably, the stroke is ischemic stroke. And / or, when the drug is used, the dosage of quinoline acid is 0.01 to 200 mg / kg; preferably, when the drug is used, the dosage of quinoline acid is 4.4 to 13.5 mg / kg. And / or, the patient is a patient who has had, is currently having, or is at risk of having a stroke; And / or, the patient is a mammal; preferably, the patient is a mouse or a human; And / or, the drug is administered via the gastrointestinal tract; preferably, the gastrointestinal administration is selected from one or more of oral administration, gastric tube administration, and enteral administration.

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