Application of colchicine in the preparation of drugs for the prevention and treatment of cardiovascular complications caused by Kawasaki disease

Abstract

This invention discloses the application of colchicine in the preparation of drugs for the prevention and treatment of cardiovascular complications caused by Kawasaki disease. Colchicine can significantly protect the integrity of the coronary artery endothelium, reduce inflammatory cell infiltration in the blood vessels and surrounding myocardial tissue, and effectively maintain the structural regularity of elastic fibers in the blood vessel wall, preventing them from loosening and breaking. Simultaneously, colchicine significantly reduces the infiltration of neutrophils (Ly6G⁺) and macrophages (CD86⁺) around the heart vessels in model mice. The application described in this invention provides a new candidate drug for Kawasaki disease-related cardiac injury, and is expected to be used to prevent or treat coronary artery damage, myocarditis, and other lesions caused by Kawasaki disease, thereby reducing the risk of serious cardiovascular events in children with Kawasaki disease.

Description

Technical Field

[0001] This invention belongs to the field of medicine, specifically relating to the application of colchicine in the preparation of drugs for the prevention and treatment of cardiovascular complications caused by Kawasaki disease. Background Technology

[0002] Kawasaki disease (KD), also known as mucocutaneous lymph node syndrome, is an acute, self-limiting disease characterized primarily by systemic vasculitis, most commonly affecting infants and young children under 5 years old. The most significant and serious complications of Kawasaki disease are cardiovascular complications, primarily including coronary arteritis, coronary artery dilation (CAD), and coronary artery aneurysm (CAA). These conditions can lead to myocardial ischemia, myocardial infarction, and even sudden death, and have become a leading cause of acquired heart disease in children.

[0003] Currently, the first-line treatment for Kawasaki disease is intravenous immunoglobulin (IVIG) combined with high-dose oral aspirin. However, approximately 15-20% of children with Kawasaki disease do not respond to IVIG treatment and face a higher risk of coronary artery damage. Furthermore, existing treatments lack effective methods to reverse or repair existing coronary artery damage. Therefore, developing novel drugs that can effectively prevent and treat Kawasaki disease-related cardiac damage, particularly by protecting the vascular endothelium and inhibiting perivascular inflammation, has significant clinical and social value.

[0004] Colchicine is a classic microtubule inhibitor with broad-spectrum anti-inflammatory effects, and it has been used clinically to treat gouty arthritis, pericarditis, and other diseases. In recent years, its anti-inflammatory applications in cardiovascular diseases, such as stable coronary artery disease and acute coronary syndrome, have also attracted attention. However, the specific pharmacological effects and related mechanisms of colchicine in Kawasaki disease-induced cardiac injury, especially coronary artery injury, have not yet been reported. Summary of the Invention

[0005] Purpose of the invention: This invention provides colchicine for the preparation of drugs to prevent and treat cardiovascular complications caused by Kawasaki disease, aiming to solve the technical problem of the lack of effective drugs in the prior art for repairing or alleviating cardiac and coronary artery damage caused by Kawasaki disease.

[0006] Specifically, this invention proposes the use of colchicine in the preparation of a medicament for the prevention and treatment of cardiovascular complications caused by Kawasaki disease.

[0007] The cardiovascular complications caused by Kawasaki disease include coronary artery injury, coronary arteritis, coronary artery dilation, or coronary artery aneurysm.

[0008] In some embodiments, the drug is used to protect the integrity of the cardiac vascular endothelium caused by Kawasaki disease.

[0009] In some embodiments, the drug is used to reduce the loosening and breakage of elastic fibers in the walls of the heart vessels caused by Kawasaki disease.

[0010] In some embodiments, the drug is used to reduce inflammatory cell infiltration of the cardiovascular vessels and surrounding myocardial tissue caused by Kawasaki disease.

[0011] In some embodiments, the inflammatory cells include neutrophils and / or macrophages.

[0012] The drug is administered orally or by injection. Preferably, it is administered orally.

[0013] Based on a classic animal model, this invention found that colchicine can significantly improve cardiac damage in mice with Kawasaki disease induced by Candida albicans cell wall extract (CAWS).

[0014] This invention describes a CAWS-induced Kawasaki disease mouse model treated with colchicine via gavage. Results showed that colchicine significantly protected the integrity of the coronary artery endothelium, reduced inflammatory cell infiltration in the blood vessels and surrounding myocardium, and effectively maintained the structural regularity of elastic fibers in the vascular wall, preventing their loosening and breakage. Simultaneously, colchicine significantly reduced the infiltration of neutrophils (Ly6G⁺) and macrophages (CD86⁺) around the heart vessels in the model mice.

[0015] Therefore, the application described in this invention provides a new candidate drug for Kawasaki disease-related cardiac injury, which is expected to be used to prevent or treat coronary artery damage, myocarditis and other lesions caused by Kawasaki disease, thereby reducing the risk of serious cardiovascular events in children with Kawasaki disease.

[0016] Beneficial effects: Compared with the prior art, this invention reveals for the first time a novel use of colchicine in the prevention and treatment of Kawasaki disease-related cardiac injury, with the following outstanding advantages: (1) Protecting the vascular endothelium: Colchicine can effectively protect the integrity of the vascular endothelium of the coronary arteries, which is the key to preventing coronary artery damage in Kawasaki disease; (2) Protecting elastic fibers: It can prevent the elastic fibers of the blood vessel wall from loosening and breaking, and maintain the normal structure and function of the blood vessel wall; (3) Inhibits inflammatory infiltration: It significantly reduces inflammatory cell infiltration in blood vessels and surrounding myocardial tissue, especially reducing the aggregation of Ly6G⁺ neutrophils and CD86⁺ macrophages, thus playing a cardioprotective role from an anti-inflammatory perspective. Attached Figure Description

[0017] Figure 1 The study showed that colchicine improved cardiovascular pathological damage in a mouse model of Kawasaki disease. Figure 2 Colchicine was shown to reduce inflammatory cell infiltration around the heart and blood vessels in a Kawasaki disease model mouse. Detailed Implementation

[0018] The present invention will be further described in detail below with reference to the accompanying drawings and specific embodiments, and the advantages of the present invention in the above and / or other aspects will become clearer.

[0019] Example 1: Colchicine improved cardiovascular pathological damage in a mouse model of Kawasaki disease.

[0020] 1. Experimental Materials Drug: Colchicine (purity ≥99%) was purchased from MCE (product number HY16569).

[0021] Animals: Four-week-old SPF-grade male C57BL / 6 mice, with an initial weight of 12-13 g, were purchased from Hangzhou Ziyuan Laboratory Animal Technology Co., Ltd. They were housed at the Barrier Environment Animal Experiment Center of the Children's Hospital Affiliated to Soochow University, at a temperature of 24±2℃ and a relative humidity of 55±5%, with 12h / 12h light-dark cycles. All experimental procedures were approved by the Laboratory Animal Ethics Committee of Soochow University.

[0022] Main reagent: Candida albicans cell wall extract (CAWS) (NBRC Collection Center, Japan, NBRC1385), prepared by the laboratory according to standard methods.

[0023] 2. Experimental Methods CAWS preparation: (1) Prepare YM broth medium, sterilize at 113°C for 20 min, add Candida albicans solution and shake at 26°C and 160 rpm for 72 h. (2) Prepare C-limiting medium, sterilize at 125°C for 30 min, add the bacterial solution from (1) and shake at 26°C and 250 rpm for 72 h. (3) Add an equal volume of anhydrous ethanol and incubate at 4°C overnight. (4) Centrifuge at 9000 rpm for 15 min, take the precipitate, add ddH2O to dissolve, and stir for 2 h. (5) Centrifuge at 9000 rpm for 15 min, take the supernatant, add an equal volume of anhydrous ethanol, and incubate at 4°C overnight. (6) Centrifuge at 9000 rpm for 15 min, take the precipitate, wash with acetone, centrifuge at 3000 rpm for 10 min, discard the supernatant, and air dry. (7) Weigh the dried product, dissolve it in PBS to a concentration of 20 mg / mL, and aliquot and store.

[0024] Model Replication and Drug Administration: C57BL / 6 mice were randomly divided into three groups (at least six mice per group): a normal control group (Control), a model group (CAWS), and a colchicine group. Mice in the model and colchicine groups were intraperitoneally injected with 0.2 mL of CAWS solution (20 mg / mL) once daily for 5 consecutive days to replicate the Kawasaki disease mouse model; mice in the normal control group were injected with an equal volume of PBS. Simultaneously with model establishment, mice in the colchicine group were administered 100 µL of colchicine (0.2 mg / kg / d) by gavage daily for 7 consecutive days; mice in the normal and model groups were administered an equal volume of PBS by gavage. After the last CAWS injection, mice were fed for 14 days. At the end of the experiment, mice were anesthetized, and heart tissue was collected for subsequent analysis.

[0025] 3. Detection Method HE staining: Heart tissue was fixed with 4% paraformaldehyde, dehydrated with graded ethanol, cleared with xylene, embedded in paraffin, and 4 μm sections were prepared. After dewaxing and hydration, the sections were stained with hematoxylin for 5 min, rinsed with running water, treated with blue solution for 1 min, stained with 0.5% eosin for 5 min, dehydrated and cleared, mounted, and observed under an optical microscope for vascular endothelium and inflammatory infiltration.

[0026] EVG staining: After dewaxing and hydration, the sections were immersed in pre-prepared EVG staining solution for 5 min until the elastic fibers turned purplish-black. After counterstaining, the sections were dehydrated, cleared, and mounted to observe the morphology of the elastic fibers.

[0027] 4. Experimental Results like Figure 1 HE staining showed that the cardiac vascular endothelium of mice in the normal control group was intact and smooth, with no inflammatory cell infiltration. In the model group, the cardiac vascular endothelium of mice was rough, structurally incomplete, with disordered cell arrangement, and extensive inflammatory cell infiltration was observed in the blood vessels and surrounding myocardial tissue. In contrast, the cardiac vascular endothelium of mice in the colchicine intervention group was more intact, and inflammatory cell infiltration was significantly reduced.

[0028] like Figure 1 EVG staining showed that the elastic fibers in the heart and blood vessel walls of normal control mice were arranged regularly and coiled in an orderly manner. In the model group, the elastic fibers were disordered, loose, and broken. In the colchicine intervention group, the elastic fibers were more regularly arranged, with no obvious breakage observed.

[0029] The above results indicate that colchicine can significantly improve cardiovascular pathological damage in Kawasaki disease model mice.

[0030] Example 2: Colchicine reduces inflammatory cell infiltration around the heart and blood vessels in a Kawasaki disease model mouse.

[0031] 1. Experimental Materials and Methods The mouse model replication and drug administration regimen were the same as in Example 1. The preparation of heart tissue sections was the same as in Example 1.

[0032] 2. Detection Method Multiplex fluorescence immunohistochemistry: After dewaxing and antigen retrieval of sections, endogenous peroxidase was inactivated and the sections were blocked. Primary antibodies CD31 (YM8207, ImmunoWay Biotechnology Co., USA), Ly6G (YM8307, ImmunoWay Biotechnology Co., USA), CD86 (91882, Cell Signaling Technology, USA), and their corresponding secondary antibodies and fluorescence signal amplification solutions were incubated sequentially. After each incubation, the sections were peeled off using antibody stripping buffer. Finally, the sections were mounted with a DAPI-containing anti-quenching mounting solution, and fluorescence images were acquired by full-slide scanning.

[0033] 3. Experimental Results like Figure 2 As shown, multiplex immunohistochemical staining results revealed a significant increase in Ly6G⁺ (neutrophils) and CD86⁺ (macrophages) cells around the heart vessels in the CAWS model group mice compared to the normal control group. However, colchicine treatment significantly reduced the abundance of both of these pro-inflammatory immune cells.

[0034] This result further confirms that colchicine exerts a cardioprotective effect by inhibiting the infiltration of perivascular neutrophils and macrophages, thereby reducing the local inflammatory response induced by Kawasaki disease.

[0035] In summary, this invention experimentally demonstrates the novel use of colchicine in the preparation of drugs for the prevention and treatment of Kawasaki disease-related cardiac injury. Colchicine effectively improves cardiac injury in CAWS-induced Kawasaki disease model mice through mechanisms such as protecting the integrity of coronary artery endothelium, maintaining elastic fiber structure, and inhibiting perivascular inflammatory infiltration. Therefore, colchicine shows promise as a candidate drug for the prevention and treatment of Kawasaki disease-related cardiac injury (especially coronary artery injury).

[0036] This invention provides a new indication for colchicine. Many methods and approaches exist for implementing this technical solution; the above description is merely a preferred embodiment of the invention. It should be noted that those skilled in the art can make various improvements and modifications without departing from the principles of this invention, and these improvements and modifications should also be considered within the scope of protection of this invention. All components not explicitly stated in this embodiment can be implemented using existing technologies.

Claims

1. The application of colchicine in the preparation of drugs for the prevention and treatment of cardiovascular complications caused by Kawasaki disease.

2. The application according to claim 1, characterized in that, Cardiovascular complications caused by Kawasaki disease include coronary artery injury, coronary arteritis, coronary artery dilation, or coronary aneurysm formation.

3. The application according to claim 1, characterized in that, The drug is used to protect the integrity of the cardiac vascular endothelium caused by Kawasaki disease.

4. The application according to claim 1, characterized in that, The drug is used to reduce the loosening and breakage of elastic fibers in the heart and blood vessel walls caused by Kawasaki disease.

5. The application according to claim 1, characterized in that, The drug is used to reduce inflammatory cell infiltration in the cardiovascular system and surrounding myocardial tissue caused by Kawasaki disease.

6. The application according to claim 5, characterized in that, The inflammatory cells include neutrophils and / or macrophages.

7. The application according to claim 1, characterized in that, The drug can be administered orally or by injection.

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