Use of ganoderic acid a in preparation of product for preventing and / or treating non-alcoholic fatty liver disease

Abstract

Use of ganoderic acid A in the preparation of a product for preventing and / or treating non-alcoholic fatty liver disease. Ganoderic acid A can regulate bile acid reabsorption in C57BL / 6J mice induced by a high-fat diet, and reduce the absorption of triglycerides and cholesterol. Ganoderic acid A can also enhance the expression of the gene of cholesterol 7α-hydroxylase, a key enzyme for liver cholesterol decomposition, thereby promoting the conversion of liver cholesterol into bile acid, and alleviating the accumulation of liver cholesterol. Ganoderic acid A has a significant preventive and ameliorative effect on non-alcoholic fatty liver disease induced by a high-fat diet by means of regulating bile acid metabolism, inhibiting lipid absorption, and reducing liver lipid accumulation. Therefore, ganoderic acid A can be used as a health care product, a functional product, or a drug for preventing and treating non-alcoholic fatty liver disease.

Description

Application of ganoderic acid A in preparation of products for preventing and / or treating non-alcoholic fatty liver disease TECHNICAL FIELD

[0001] The present application belongs to the technical field of medicine, and particularly relates to application of ganoderic acid A in preparation of products for preventing and / or treating non-alcoholic fatty liver disease. BACKGROUND

[0002] Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease worldwide and also the liver disease that causes the most disability and changes in life expectancy. It can cause a series of pathological changes such as liver inflammation and fibrosis, and develop into non-alcoholic steatohepatitis. Unhealthy eating habits, sedentary lifestyle and aging can increase the risk of NAFLD. According to literature reports, the global prevalence of NAFLD increased from 25.3% in 1990-2006 to 38.0% in 2016-2019. China is also a high-incidence area of NAFLD, and the incidence of NAFLD in China was between 23.8% and 29% as of 2018. NAFLD seriously threatens human health and quality of life, and has caused huge economic and medical burdens to China. Despite the high and increasing incidence of NAFLD, there is still a lack of safe and effective drugs for treating NAFLD so far. Targeting lipid metabolism is considered to be the main treatment strategy for NAFLD. Drugs that interfere with lipid metabolism are often used to improve NAFLD, such as weight loss drugs and lipid-lowering drugs, but long-term use of these drugs can cause serious adverse reactions. Therefore, it is of great significance to develop safe and effective, long-term use of drugs or health care products for preventing and treating NAFLD.

[0003] Ganoderma lucidum is widely used as a traditional Chinese medicine in Asian regions. Ganoderic acid compounds are an important active substance in G. lucidum extract, which are triterpenoids with a lanostane skeleton composed of 30 carbon atoms. According to the number and position of double bonds of the parent ring, ganoderic acid can be divided into different monomers. In the ganoderic acid extract, the content of ganoderic acid A is higher than that of other ganoderic acid monomers, and it has various pharmacological effects such as anti-inflammatory and antioxidant effects, and can also relieve diseases caused by lipid metabolism disorders. However, the effect and mechanism of ganoderic acid A in improving NAFLD have not been reported. SUMMARY

[0004] Non-alcoholic fatty liver disease (NAFLD) is the most common liver disease in the world, and is the most common liver disease that causes disability and changes life. The application provides application of ganoderic acid A in preparation of products for preventing and / or treating non-alcoholic fatty liver disease. The effect of ganoderic acid A on relieving non-alcoholic fatty liver disease is remarkable. Ganoderic acid A can prevent and treat the formation of non-alcoholic fatty liver by inhibiting the reabsorption of bile acid and the expression of liver cholesterol accumulation related genes, and can be used for preventing and treating non-alcoholic fatty liver disease in various ways.

[0005] Ganoderic acid A has the advantages of safety, effectiveness and suitability for long-term use, and has good application prospect. Technical solutions

[0006] In order to solve the above technical problems, the application provides application of ganoderic acid A in preparation of products for preventing and / or treating non-alcoholic fatty liver disease.

[0007] The chemical structural formula of the ganoderic acid A is shown as formula I:

[0008]

[0009] In the product, ganoderic acid A can be used as the only active ingredient, or as one of the active ingredients.

[0010] The application further provides application of a composition containing ganoderic acid A in preparation of products for preventing and / or treating non-alcoholic fatty liver disease.

[0011] In the above application, the composition further contains a pharmaceutically acceptable excipient and / or a drug compatible with ganoderic acid A.

[0012] In the application, the prevention and / or treatment of non-alcoholic fatty liver disease is reflected in at least one of the following aspects:

[0013] 1) inhibiting the increase of body weight, liver specific gravity and / or white fat specific gravity caused by non-alcoholic fatty liver disease;

[0014] 2) improving liver damage caused by non-alcoholic fatty liver disease;

[0015] 3) reducing liver lipid accumulation caused by non-alcoholic fatty liver disease;

[0016] 4) regulating bile acid reabsorption and inhibiting the absorption of lipids (triglycerides and cholesterol);

[0017] 5) enhancing the expression of key enzyme cholesterol 7 alpha-hydroxylase (CYP7A1) gene of liver cholesterol decomposition.

[0018] Further, the improvement of liver injury caused by non-alcoholic fatty liver disease is embodied in that ganoderic acid A can significantly reduce the increase of alanine aminotransferase and aspartate aminotransferase levels caused by non-alcoholic fatty liver disease.

[0019] In the present application, the non-alcoholic fatty liver disease is caused by high-fat diet.

[0020] In the present application, the product can be a drug, a health product and / or a functional food.

[0021] The drug contains an effective dose of ganoderic acid A. The effective dose is a unit administration dose form (such as the content in a piece of drug) or a unit dose for a patient (such as a unit weight dose). In the present application, the subject of drug treatment is a human or a mammal (such as a mouse, etc.).

[0022] The dosage form of the drug in the present application is not limited as long as it can effectively reach the active ingredient in the body, including tablets, sugar-coated tablets, film-coated tablets, enteric-coated tablets, capsules, hard capsules, soft capsules, buccal tablets, granules, instant tablets, pills, powders, pastes, pills, suspensions, powders, solutions, injections, suppositories, ointments, plasters, creams, sprays, drops, patches, etc.

[0023] The present application also provides a method for preventing and / or treating non-alcoholic fatty liver disease.

[0024] The method for preventing and / or treating non-alcoholic fatty liver disease provided by the present application comprises the following steps: administering ganoderic acid A or the composition containing ganoderic acid A to a recipient animal or a human to prevent and / or treat non-alcoholic fatty liver disease.

[0025] The present application uses a C57BL / 6J mouse model induced by high-fat diet to find that supplementing ganoderic acid A has a relieving effect on non-alcoholic fatty liver disease. Compared with the blank group, the body weight, liver index and white fat index of the mice in the high-fat group are significantly increased. Compared with the high-fat group, after the mice are orally administered with 20 mg / kg body weight of ganoderic acid A daily for 16 weeks, the body weight, liver index and white fat index of the mice are significantly decreased.

[0026] The present application shows that ganoderic acid A can significantly reduce the concentrations of alanine aminotransferase and aspartate aminotransferase in the serum of high-fat diet mice by determining the blood biochemical indicators of the mice in the blank group, the high-fat group and the high-fat+ganoderic acid A group, which indicates that ganoderic acid A can improve the liver injury induced by high-fat diet.

[0027] The present application observes that the hepatocytes of the high-fat group mice are obviously enlarged, and the cells are filled with lipids to form fat bubbles through HE staining of liver tissue sections. After intervention of ganoderic acid A, the volume of the fat bubbles in the hepatocytes is significantly reduced. The results of liver oil red O staining show that ganoderic acid A can significantly reduce the lipid deposition in the liver. The present application finds that ganoderic acid A can significantly reduce the levels of triglyceride and cholesterol in the liver by using a kit for detection, which indicates that ganoderic acid A can effectively alleviate the occurrence and development of non-alcoholic fatty liver.

[0028] Specifically, ganoderic acid A can reduce the lipid accumulation in the liver of mice caused by high-fat diet.

[0029] Specifically, ganoderic acid A can reduce the increase of the contents of triglyceride and cholesterol in the liver tissue of mice caused by high-fat diet.

[0030] The present application finds that ganoderic acid A significantly affects the cholesterol / bile acid metabolism of mice and reduces the levels of some long-chain fatty acids by detecting the metabolites at the end of the ileum of mice in the blank group, the high-fat group and the high-fat+ganoderic acid A group. Meanwhile, it is found that ganoderic acid A can significantly increase the content of bile acid in feces by using a kit for detection. And it is found that ganoderic acid A can significantly increase the lipid content in feces by using a kit for detection. It is indicated that ganoderic acid A can inhibit lipid absorption by regulating the bile acid / cholesterol metabolism and bile acid reabsorption of mice.

[0031] Specifically, ganoderic acid A can reduce the levels of harmful bile acids such as taurine cholic acid and glycochenodeoxycholic acid, and inhibit bile acid reabsorption. Ganoderic acid A inhibits the absorption of triglyceride and cholesterol.

[0032] The present application detects the cholesterol decomposition related genes in the liver of mice in the blank group, the high-fat group and the high-fat+ganoderic acid A group by using fluorescence quantitative PCR technology.

[0033] Specifically, ganoderic acid A can promote the decomposition of liver cholesterol by increasing the expression of cholesterol 7alpha-hydroxylase (CYP7A1) gene in the liver, and reduce the accumulation of liver cholesterol. Beneficial effects

[0034] (1) The present application discovers a new natural active substance for preventing and treating non-alcoholic fatty liver disease, which has the advantages of safety, effectiveness and suitability for long-term use, and has good application prospect.

[0035] (2) The effect of ganoderic acid A on alleviating non-alcoholic fatty liver disease is significant.

[0036] (3) Ganoderic acid A can prevent and treat the formation of non-alcoholic fatty liver by inhibiting the reabsorption of bile acid and the expression of liver cholesterol accumulation related genes, and can be used for preventing and treating non-alcoholic fatty liver disease in multiple ways. BRIEF DESCRIPTION OF DRAWINGS

[0037] Figure 1 is the effect of ganoderic acid A on the body weight of mice.

[0038] Figure 2 is the effect of ganoderic acid A on the liver index and fat index.

[0039] Figure 3 is the effect of ganoderic acid A on the concentration of alanine aminotransferase and aspartate aminotransferase in the blood of mice.

[0040] Figure 4 is the HE staining and oil red O staining of liver sections of mice.

[0041] Figure 5 is the effect of ganoderic acid A on the content of triglyceride and cholesterol in the liver tissue of mice.

[0042] Figure 6 is the differential bile acid metabolites in the ileum tissue of mice.

[0043] Figure 7 is the differential long-chain fatty acids in the ileum tissue of mice.

[0044] Figure 8 is the effect of ganoderic acid A on the triglyceride, cholesterol and total bile acid in the feces of mice.

[0045] Figure 9 is the effect of ganoderic acid A on the expression level of Cyp7a1 gene in the liver tissue of mice. Embodiments of the present application

[0046] The present application will be further described in conjunction with the specific embodiments. The examples given are only to illustrate the present application, and are not intended to limit the scope of the present application. The examples provided below can serve as a guide for further improvement by those of ordinary skill in the art, and do not in any way constitute a limitation on the present application.

[0047] In the following examples, the experimental methods are conventional methods, and are performed according to the techniques or conditions described in the literature in the art or according to the product instructions, unless otherwise specified. The materials, reagents, etc. used in the following examples can be obtained commercially, unless otherwise specified. In the following examples, quantitative tests were set up in triplicate, and the results were averaged, unless otherwise specified.

[0048] Experimental grouping and administration in the following examples:

[0049] After 18 eight-week-old C57BL / 6J mice were adaptively fed for one week, they were randomly divided into three groups, each group of six, namely a blank group, a high-fat group and a high-fat + ganoderic acid A group. The blank group was fed with normal feed, and the high-fat group and the high-fat + ganoderic acid A group were fed with high-fat feed. The blank group and the high-fat group were administered with 0.5% sodium carboxymethyl cellulose physiological saline solution each time, and the high-fat + ganoderic acid A group was administered with ganoderic acid A (20 mg / kg, dissolved in 0.5% sodium carboxymethyl cellulose physiological saline solution) each time. The administration frequency was once a day, and the administration lasted for 16 weeks. The normal and high-fat feed were purchased from Beijing Keao Hesuolil Foodstuff Co., Ltd.

[0050] Example 1: Effect of Ganoderic Acid A on alleviating non-alcoholic fatty liver disease

[0051] The mice were weighed every week and the body weight of each group of mice was recorded. After the last administration experiment, the mice were dissected to remove the liver, white fat. The liver, white fat weight was weighed, and the liver ratio and white fat ratio (formula: liver ratio = liver weight / body weight * 100%; white fat ratio = white fat weight / body weight * 100%) were calculated.

[0052] Experimental results: see Figures 1 and 2. Ganoderic acid A can significantly inhibit the increase of body weight, liver ratio and white fat ratio caused by high-fat diet.

[0053] Detection of serum glutamic-pyruvic transaminase and glutamic-oxalacetic transaminase: after the last administration experiment, the mice were fasted for 12 h, blood was taken, centrifuged at 3000 r / min for 10 min, serum was separated for detection, and the serum glutamic-pyruvic transaminase and glutamic-oxalacetic transaminase levels were determined.

[0054] Experimental results: see Figure 3. Ganoderic acid A significantly reduced the increase of glutamic-pyruvic transaminase and glutamic-oxalacetic transaminase levels caused by high-fat diet, indicating that ganoderic acid A significantly improved liver damage caused by high-fat diet.

[0055] HE staining, oil red O staining: after the last administration experiment, the mice were dissected, the mouse liver tissue was fixed with 4% paraformaldehyde, then embedded in paraffin, and the section thickness was 4 μm. Hematoxylin and eosin (H&E) and oil red O staining were used for pathological evaluation.

[0056] Experimental results: see Figure 4. Through HE staining of liver tissue sections, it can be observed that the hepatocytes of high-fat group mice are significantly enlarged, and the intracellular lipid is filled to form fat vacuoles. After intervention of ganoderic acid A, the volume of fat vacuoles in hepatocytes is significantly reduced. The results of liver oil red O staining show that ganoderic acid A can significantly reduce the lipid deposition in liver.

[0057] Triglyceride and cholesterol content detection: after the last administration experiment, the mouse liver tissue was taken, and the triglyceride and cholesterol detection kit of Proliferon Company was used to detect the triglyceride and cholesterol content of liver according to the instruction.

[0058] Experimental results: see Figure 5, ganoderic acid A can significantly reduce the increase of triglyceride and cholesterol content in liver tissue of mice caused by high-fat diet.

[0059] Example 2: Effect of Ganoderic Acid A on lipid absorption in high-fat diet mice

[0060] Non-targeted metabolome detection: After the end of the last administration experiment, the mice were dissected and the mouse ileum contents were collected, and the metabolome analysis was performed using liquid chromatography-tandem mass spectrometry technology to detect the components of the mouse ileum contents.

[0061] Experimental results: see Figures 6 and 7. Compared with the high-fat group and the high-fat + ganoderic acid A group, the results showed that ganoderic acid A significantly affected the cholesterol / bile acid metabolic pathway in mice, reducing the levels of harmful bile acids such as taurocholic acid (leading to excessive lipid absorption) and glycochenodeoxycholic acid (which can cause glycolipid metabolic disorders). At the same time, ganoderic acid A reduced the levels of various long-chain fatty acids in the ileum. The above results suggest that intragastric administration of ganoderic acid A can affect the bile acid / cholesterol metabolism of mice and affect lipid absorption by changing bile acid metabolism.

[0062] Triglyceride, cholesterol and bile acid content in feces. After the end of the last administration experiment, the mouse feces were collected. The triglyceride and cholesterol content was detected using the kit from Pu Li Lei Company, and the bile acid content was detected using the kit from Nanjing Jiancheng Company, and the operation was performed according to the instructions.

[0063] Experimental results: see Figure 8. Ganoderic acid A significantly increased the levels of triglycerides, cholesterol and bile acids in feces, indicating that ganoderic acid A promoted the excretion of lipids and bile acids, reducing lipid absorption.

[0064] Example 3: Mechanism of ganoderic acid A in relieving liver lipid accumulation

[0065] Fluorescent quantitative PCR method was used to detect the expression level of Cyp7a1 gene in liver tissue.

[0066] Experimental results: see Figure 9, ganoderic acid A significantly increased the expression level of Cyp7a1, which is responsible for converting excess liver cholesterol into bile acids and relieving liver lipid accumulation.

[0067] Cross-reference of related applications:

[0068] This application claims priority to Chinese patent application (application number: 202410887701.5) with a filing date of July 3, 2024, the entire contents of the above patent application are hereby incorporated by reference.

[0069] The application has been described in detail above. For those skilled in the art, the application can be implemented in a wider range under equivalent parameters, concentrations and conditions without departing from the spirit and scope of the application and without unnecessary experiments. Although the application gives a specific example, it should be understood that the application can be further improved. In summary, according to the principle of the application, the present application intends to include any change, use or improvement of the application, including changes made by conventional techniques known in the art, which deviates from the range disclosed in the present application. Some basic features can be applied within the scope of the following attached claims. Industrial applicability

[0070] The application provides application of ganoderic acid A in preparation of products for preventing and / or treating non-alcoholic fatty liver disease. The ganoderic acid A can prevent and treat formation of non-alcoholic fatty liver by inhibiting reabsorption of bile acid and expression of liver cholesterol accumulation related genes, and can be used in various ways for preventing and treating non-alcoholic fatty liver disease. The ganoderic acid A has the advantages of safety, effectiveness and suitability for long-term use, and has good application prospect.

Claims

1. Use of ganoderic acid A in the preparation of a product for preventing and / or treating non-alcoholic fatty liver disease.

2. Use of a composition comprising ganoderic acid A in the preparation of a product for preventing and / or treating non-alcoholic fatty liver disease.

3. Use according to claim 2, characterized in that: The composition further comprises pharmaceutically acceptable adjuvants and / or drugs compatible with ganoderic acid A.

4. Use according to any one of claims 1 to 3, characterized in that: The prevention and / or treatment of non-alcoholic fatty liver disease is manifested in at least one of the following aspects: 1) inhibiting the increase of body weight, liver specific gravity and / or white fat specific gravity caused by non-alcoholic fatty liver disease; 2) improving liver damage caused by non-alcoholic fatty liver disease; 3) reducing liver lipid accumulation caused by non-alcoholic fatty liver disease; 4) regulating bile acid reabsorption and inhibiting lipid absorption; 5) enhancing the expression of the key enzyme cholesterol 7a-hydroxylase (CYP7A1) gene for liver cholesterol decomposition.

5. Use according to claim 4, characterized in that: The improvement of liver damage caused by non-alcoholic fatty liver disease is specifically manifested in that ganoderic acid A can significantly reduce the increase of alanine aminotransferase and aspartate aminotransferase levels caused by non-alcoholic fatty liver disease.

6. Use according to any one of claims 1 to 5, characterized in that: The product can be a drug and / or health product and / or functional food.

7. Use of ganoderic acid A in the prevention and / or treatment of non-alcoholic fatty liver disease.

8. Use of a composition comprising ganoderic acid A in the prevention and / or treatment of non-alcoholic fatty liver disease.

9. A method for preventing and / or treating non-alcoholic fatty liver disease, comprising the following steps: administering ganoderic acid A or a composition comprising ganoderic acid A to a subject animal or human to prevent and / or treat non-alcoholic fatty liver disease.

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