Traditional Chinese medicine composition for treating metabolism-related fatty liver disease and preparation method thereof

Abstract

The invention belongs to the technical field of biological medicines, and particularly relates to a traditional Chinese medicine composition for treating metabolism-related fatty liver diseases and a preparation method thereof. The technical problem to be solved by the invention is to provide a new choice for treating metabolism-related fatty liver diseases. According to the technical scheme, the traditional Chinese medicine composition for treating the metabolism-related fatty liver disease is prepared from the following components: radix paeoniae rubra, cyrtomium fortunei, curcuma zedoary, sculellaria barbata, herba epimedii, fingered citron, semen cassiae and herba artemisiae scopariae. The invention is mainly used for providing a new choice for treating the metabolism-related fatty liver disease, and has a wide application prospect.

Description

Technical Field

[0001] The present invention belongs to the technical field of biomedicine, and specifically relates to a traditional Chinese medicine composition for treating metabolism-related fatty liver disease and a preparation method thereof. Background Art

[0002] Metabolic-associated liver disease (MAFLD) refers to liver diseases caused by metabolic abnormalities, typically associated with disturbances in the body's metabolism, affecting liver structure and function. These diseases are often closely associated with metabolic syndromes such as obesity, diabetes, and hyperlipidemia, and their incidence is increasing in modern society. MAFLD has become the most common liver disease worldwide and the leading liver disease in my country. It encompasses the full spectrum of fatty liver diseases, from simple hepatic steatosis (NAFL) to nonaleptic steatohepatitis (NASH) and NASH cirrhosis. Simple hepatic steatosis is characterized by lipid accumulation in hepatocytes. Nearly 60% of patients with simple hepatic steatosis may progress to steatohepatitis, characterized by inflammation and hepatocellular damage. 41% of patients with steatohepatitis also develop fibrosis. Of these, 22% of patients with advanced fibrosis may progress to cirrhosis, and 2-3% of patients with cirrhosis may develop hepatocellular carcinoma (HCC) within three years. This chronic liver disease has been dubbed a "silent killer."

[0003] MAFLD is closely related to lipid metabolism disorders. In the state of insulin resistance, the interaction between the disordered metabolic environment and local mediators of liver inflammation and fibrosis leads to liver fat accumulation and fatty liver hepatitis. Insulin resistance and inflammation form a vicious cycle, accelerating the development of MAFLD. MAFLD is also closely related to type 2 diabetes mellitus (T2DM). The prevalence of the two diseases is growing at almost the same rate, and the two have a bidirectional synergistic effect. Type 2 diabetes is also a risk factor for MAFLD to develop faster into fatty liver hepatitis, cirrhosis or hepatocellular carcinoma. Obesity is also strongly associated with an increased risk of liver cancer associated with MAFLD. For every 5 kg / m increase in BMI, the risk of MAFLD increasing from 1.5 kg / m 2 , increasing the risk of HCC by 24%. MAFLD patients also commonly suffer from comorbid hyperlipidemia (69.2%), hypertension (39.3%), and type 2 diabetes mellitus (T2DM) (22.5%). MAFLD also significantly increases the risk of cardiovascular disease (CVD), with atherosclerosis having the highest prevalence (55.5%). MAFLD's harmful effects not only lead to liver disease, disability, and death, but are also closely associated with a higher incidence of metabolic syndrome, type 2 diabetes, arteriosclerotic cardiovascular disease, and colorectal cancer, leading to increased morbidity and mortality from diabetes and cardiovascular disease.

[0004] Data from consulting firm Frost & Sullivan shows that the global MAFLD population is steadily increasing and is expected to reach 2.43 billion by 2030. The number of NASH patients is expected to reach 490 million, and the global market size of NASH drugs is expected to exceed US$30 billion. Over the past decade, MASH has been a hot topic in drug research and development. However, due to the complex etiology of MASH and the unclear pathogenic mechanism, the development of innovative drugs is quite difficult. It is known as a "R&D black hole" in the industry. Many multinational pharmaceutical companies such as Gilead, Novartis, and Bristol-Myers Squibb have failed in this field. For example, obeticholic acid (OCA) was denied approval for the treatment of NASH due to adverse reactions (pruritus, hepatobiliary events, and increased LDL levels). The Phase II clinical trial of BelaPectin for NASH with advanced liver fibrosis failed to achieve the primary clinical endpoint. The Phase III clinical trial of Elafibranor failed to achieve the primary clinical endpoint of alleviating NASH fibrosis. Selonsertib failed its Phase III clinical trial, failing to improve NASH-related fibrosis or compensated cirrhosis, highlighting the significant unmet need in the global NASH patient population. In 2024, the US Food and Drug Administration (FDA) granted accelerated approval to resmetirom, developed by Madrigal Pharmaceuticals, for the treatment of patients with steatohepatitis with liver fibrosis (NASH). This is the first innovative FDA-approved treatment for NASH, and its efficacy and safety await further testing after its launch. Traditional Chinese medicine (TCM) offers advantages in treating such complex pathological conditions, but it is noteworthy that since the implementation of the new TCM registration classification in 2020, no Category 1 new TCM liver disease drugs have been approved for marketing in China. Several Category 1 new TCM liver disease drugs are in Phase II clinical trials, including Kangyuan Pharmaceutical's Qiwei Zhigan Granules, Tasly's Picrorhizon Total Glycosides Capsules, Shanghai Tianchan Bio's Xuechan Ruanjian Rougan Tablets, and Sichuan Academy of Traditional Chinese Medicine's Qinwei Capsules. It can be seen that there is a huge market for drugs targeting MAFLD, but there are still major challenges in finding highly effective and safe drugs for the treatment of NASH.

[0005] Therefore, finding a suitable, more effective and safe drug to treat MAFLD is an urgent problem to be solved in clinical practice. Summary of the Invention

[0006] The technical problem to be solved by the present invention is to provide a new option for treating metabolism-related fatty liver disease.

[0007] The technical solution of the present invention is a traditional Chinese medicine composition for treating metabolism-related fatty liver disease, comprising the following ingredients: red peony root, rhizoma cyrtomii, zedoaria, scutellaria barbata, epimedium, bergamot, cassia seed and capillaris.

[0008] Furthermore, the dosage ratio of each ingredient is as follows: 6-12 parts by weight of red peony root, 6-12 parts by weight of rhizoma cyrtomii, 6-18 parts by weight of zedoaria, 5-15 parts by weight of scutellaria barbata, 6-18 parts by weight of epimedium, 6-18 parts by weight of bergamot, 6-18 parts by weight of cassia seed and 5-15 parts by weight of capillaris.

[0009] Preferably, the dosage ratio of the above ingredients is as follows: 6 parts by weight of red peony root, 6 parts by weight of rhizoma cyrtomii, 6 parts by weight of zedoaria, 5 parts by weight of scutellaria barbata, 6 parts by weight of epimedium, 6 parts by weight of bergamot, 6 parts by weight of cassia seed and 5 parts by weight of capillaris.

[0010] Preferably, the dosage ratio of the above ingredients is as follows: 12 parts by weight of red peony root, 12 parts by weight of cyperus rotundus, 18 parts by weight of zedoary turmeric, 15 parts by weight of scutellaria barbata, 18 parts by weight of epimedium, 18 parts by weight of bergamot, 18 parts by weight of cassia seed and 15 parts by weight of capillaris.

[0011] The present invention also provides a method for preparing the Chinese medicine composition, comprising the following steps: a. Mix red peony root, zedoary turmeric and epimedium, extract twice with 70% ethanol as extract, combine the extracts, recover ethanol, and obtain filtrate 1; b. Mix the medicinal residue after extraction in step a with Guanzhong, Scutellaria barbata, Artemisia capillaris, Citron, and Cassia seed, add water and extract twice, combine the extracts, and obtain filtrate 2; c. Combine filtrate 1 and filtrate 2, concentrate, dry and crush.

[0012] Specifically, in step a, the mass-to-volume ratio (g / mL) of the material to the liquid is 1:6-1:15.

[0013] Preferably, in step a, the mass-to-volume ratio (g / mL) of the feed liquid is 1:8.

[0014] Specifically, in step a, the extraction time for each extraction is 0.5h-2h.

[0015] Preferably, in step a, the extraction time for each extraction is 1 hour.

[0016] Furthermore, in step a, ethanol is recovered using a decompression technique.

[0017] Furthermore, in step b, the extraction is carried out by a heating, condensation and reflux method.

[0018] Specifically, in step b, the mass-to-volume ratio (g / mL) of the material to the liquid during the first extraction is 1:6-1:15; the mass-to-volume ratio (g / mL) of the material to the liquid during the second extraction is 1:6-1:12.

[0019] Preferably, in step b, the mass-to-volume ratio (g / mL) of the material to the liquid during the first extraction is 1:10; and the mass-to-volume ratio (g / mL) of the material to the liquid during the second extraction is 1:8.

[0020] Specifically, in step b, the extraction time is 0.5h-2h each time.

[0021] Preferably, in step b, the extraction time for each time is 1 hour.

[0022] The present invention also provides a traditional Chinese medicine composition prepared by the above preparation method.

[0023] The present invention also provides use of the above-mentioned traditional Chinese medicine composition in preparing a medicine for treating or improving metabolism-related fatty liver disease.

[0024] The present invention also provides a medicine or health product for weight loss, lipid-lowering and blood sugar-lowering, and anti-oxidation, comprising the above-mentioned traditional Chinese medicine composition or the traditional Chinese medicine composition obtained by the above-mentioned preparation method.

[0025] Specifically, the drug preparation further includes pharmaceutically acceptable excipients, and the dosage form of the preparation is tablets, pills, capsules, granules, powders, pastes, sprays or mixtures.

[0026] The present invention also provides the use of the above-mentioned traditional Chinese medicine composition in the preparation of a drug for controlling body weight and food intake, reducing liver index and liver weight, reducing body fat percentage, improving glucose intolerance and insulin resistance, improving blood lipids, improving liver damage, improving liver lipid accumulation, improving liver ballooning, improving liver inflammatory cell infiltration, improving liver fatty degeneration or lipid metabolism disorder caused by a high-sugar and high-fat diet, improving inflammatory response, improving liver fibrosis level and / or improving liver tissue oxidative stress level.

[0027] Beneficial Effects of the Invention: The present invention provides a traditional Chinese medicine composition comprising: red peony root, rhizoma cyrtomii, zedoaria, scutellaria barbata, epimedium, bergamot, cassia seed, and capillaris; a preparation method thereof is also provided; and clinical data demonstrate the efficacy of the composition in treating or ameliorating metabolic-related fatty liver disease. Through the synergistic action of multiple Chinese medicinal herbs, the composition has the effects of clearing heat and dampness, promoting blood circulation and removing blood stasis, soothing the liver and regulating qi, and tonifying the kidney and spleen, thereby targeting various pathological aspects of non-alcoholic fatty liver disease. Animal models were established, and a series of experimental data demonstrated that the composition can control body weight and food intake, reduce liver index and liver weight, lower body fat percentage, improve glucose intolerance and insulin resistance, improve blood lipids, ameliorate liver damage, improve liver lipid accumulation, improve liver ballooning, improve liver inflammatory cell infiltration, improve liver steatosis or lipid metabolism disorders caused by a high-sugar, high-fat diet, ameliorate inflammatory responses, improve liver fibrosis, and improve oxidative stress in liver tissue. The present invention provides a new option for treating or improving metabolism-related fatty liver disease and has broad application prospects. BRIEF DESCRIPTION OF THE DRAWINGS

[0028] Figure 1 Weight statistics of each group (n=6, ±s), the vertical axis is body weight (g), and the horizontal axis is the number of weeks of animal administration; compared with the model group: *P<0.05, **P<0.01, ***P<0.001, ****P<0.0001.

[0029] Figure 2 Appearance of the liver of mice in each group.

[0030] Figure 3 Blood glucose statistics of each group (n=6, ±s), the ordinate represents the blood glucose content (mmol / L), and the abscissa represents the time (min).

[0031] Figure 4 Representative images of H&E-stained liver sections in each group (n=3, x±s). H&E staining: hematoxylin-eosin staining.

[0032] Figure 5 Representative images of Oil Red O staining of liver sections in each group (n=3, x±s).

[0033] Figure 6 Liver sections of each group were stained with Sirius red (n=3, x±s).

[0034] Figure 7 F4 / 80 immunohistochemical staining of liver sections in each group (n=3, x±s). DETAILED DESCRIPTION

[0035] The Traditional Chinese Medicine (TCM) name for MAFLD is based on analysis of its symptoms, etiology, and pathogenesis, and is generally categorized as "liver disease," "flank pain," and "accumulation." Based on years of clinical practice and combined with efficacy research, the formula has been continuously optimized to form a fixed prescription consisting of red peony root, cyperus rotundus, zedoaria, barbata, epimedium, bergamot, cassia seed, and capillaris. In this study, based on the Western medical pathological characteristics and Traditional Chinese Medicine pathogenesis of MAFLD, a MAFLD mouse model that meets clinical characteristics was established by inducing a high-fat, high-sugar diet. Lipid (TG, TC, HDL-C, LDL-C) and glucose homeostasis (OGTT, HOMA-IR) indicators, liver function damage indicators (ALT, AST) and oxidative stress indicators (MDA, SOD), inflammation (TNF-α, MCP-1, IL-6, IL-1β), and fibrosis-related indicators (HA, Col-IV, LN, α-SMA, CoI1α1, Tgf-β1, FN-1) were detected. Combined with pathological staining (H&E staining, Oil Red O staining, Sirius Red staining, and immunohistochemistry staining), the pharmacological effects of the compound preparation in improving MAFLD were objectively evaluated, providing a scientific basis for its development as a potential drug for the prevention and treatment of MAFLD.

[0036] The present invention will be further described in detail below by way of examples, but the present invention is not limited thereto in any way.

[0037] The experimental animals used in the following examples are: Six-week-old C57BL / 6J male mice weighing 18-20 g were selected (purchased from Zhuhai Baishitong Biotechnology Co., Ltd.).

[0038] The experimental materials used in the following examples are: Red peony root (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Curcuma zedoaria (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Epimedium (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Rhizoma Cyperi (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Scutellaria barbata (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Artemisia capillaris (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Citron (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Semen Cassiae (purchased from Guangdong Provincial Medicinal Materials and Traditional Chinese Medicine Slice Factory), Silymarin (purchased from German Dr. Ma Pharmaceutical Factory), high-fat feed (containing 60% fat, D12492, Deitz, USA), D-glucose (purchased from Guangdong Guangshi Reagent Technology Co., Ltd.), and D-fructose (Shanghai MacLean Biochemical Technology Co., Ltd.).

[0039] Example 1 Extraction and Drying of Drugs—Low-Dose Compound Preparation ① Prepare the ingredients: weigh 6 g of red peony root, 6 g of cyperus rotundus, 6 g of zedoaria, 5 g of scutellaria barbata, 6 g of epimedium, 6 g of bergamot, 6 g of cassia seed, and 5 g of capillaris. ② Take red peony root, zedoary turmeric, and epimedium, add 8 times the amount of 70% ethanol, heat and reflux and extract twice, each time for 1 hour, filter, combine the two extracts, and recover the ethanol under reduced pressure to obtain filtrate 1; take Guanzhong, Scutellaria barbata, Artemisia capillaris, bergamot, and Cassia seed, and add the residue obtained after the above ethanol extraction and filtration, add water and heat and reflux and extract twice, the first time with 10 times the amount and the second time with 8 times the amount, each time for 1 hour, filter, and obtain filtrate 2; ③ Combine filtrate 1 and filtrate 2, concentrate under reduced pressure to obtain an extract, and freeze-dry the extract powder.

[0040] Example 2 Extraction and Drying of Drugs—High-Dose Compound Preparation ① Prepare the ingredients: weigh 12 g of red peony root, 12 g of cyperus rotundus, 18 g of zedoaria, 15 g of scutellaria barbata, 18 g of epimedium, 18 g of bergamot, 18 g of cassia seed, and 15 g of capillaris. ② Take red peony root, zedoary turmeric, and epimedium, add 8 times the amount of 70% ethanol, heat and reflux and extract twice, each time for 1 hour, filter, combine the two extracts, and recover the ethanol under reduced pressure to obtain filtrate 1; take Guanzhong, Scutellaria barbata, Artemisia capillaris, bergamot, and Cassia seed, and add the residue obtained after the above ethanol extraction and filtration, add water and heat and reflux and extract twice, the first time with 10 times the amount and the second time with 8 times the amount, each time for 1 hour, filter, and obtain filtrate 2; ③ Combine filtrate 1 and filtrate 2, concentrate under reduced pressure to obtain an extract, and freeze-dry the extract powder.

[0041] Example 3 Drug Intervention and Collection Random grouping: All mice were divided into a blank control group (Con), a model group (Mod), a positive drug silymarin group (Sily, 100 mg / kg), a high-dose compound preparation group (Preparation-H, 12.0 g / kg), and a low-dose compound preparation group (Preparation-L, 4.0 g / kg) according to their body weight using a random number table method; wherein, 12.0 g / kg in the high-dose compound preparation group means that 12.0 g of the drug needs to be administered per 1 kg of body weight of the mouse, and 12.0 g of the drug means that the amount of the raw medicinal material in the prescription of Example 2 is 12.0 g; 4.0 g / kg in the low-dose compound preparation group means that 4.0 g of the drug needs to be administered per 1 kg of body weight of the mouse, and 4.0 g of the drug means that the amount of the raw medicinal material in the prescription of Example 1 is 4.0 g.

[0042] After one week of adaptive feeding, 6-week-old male C57BL / 6J mice were fed a standard diet and drinking water. The Con group received a high-fat diet and high-sugar drinking water (23.1 g / L d-fructose + 18.9 g / L d-glucose) for 12 weeks to establish the MAFLD mouse model. After 12 weeks, mice in the Sily, Formulation-L, and Formulation-H groups were simultaneously administered the corresponding drug doses (0.1 mL / 10 g) dissolved in 0.5% CMC-Na. The remaining groups received the same dose of 0.5% CMC-Na for 12 weeks. After 12 weeks of drug administration, all groups were fasted for 12 hours. After anesthesia with isoflurane, blood was collected by enucleation. Following blood collection, the mouse hearts were perfused with 30 mL of normal saline, and the livers were removed sequentially. The following tests were performed on each group: 1. Determination of body fat percentage and liver index of mice Before sampling, based on the principle of nuclear magnetic resonance, a small nuclear magnetic analyzer was used to obtain the body fat-related values ​​of the mice.

[0043] After the autopsy, six mice were randomly selected from each group. Their body weight and liver wet weight were measured using an electronic balance. The organ index of each organ was calculated using the organ index formula: organ index = mouse organ fresh weight / mouse body weight × 100%.

[0044] 2. Oral glucose tolerance test One week before sampling, six mice were randomly selected from each group and fasted for 12 hours before the experiment. During this fasting period, mice were deprived of food but not water. Based on their body weight, they were injected with a 20% glucose solution at a dose of 2.0 g / kg. After disinfecting the mouse tail with an alcohol swab, blood was drawn with a lancet. Blood glucose levels were measured and recorded at 0 min (fasting blood glucose), 15 min, 30 min, 60 min, 90 min, and 120 min. The area under the curve (AUC) was calculated using GraPhPad Prism 9.0 software.

[0045] 3. Serum fasting insulin After drug intervention in MAFLD model mice, serum fasting insulin (FINS) levels were detected by ELISA, and the insulin resistance index was calculated according to the homeostatic insulin resistance index (HOMA-IR): HOMA-IR=FINS×FBG / 22.5.

[0046] 4. Tests of four serum lipids, liver function indicators alanine aminotransferase and aspartate aminotransferase After drug intervention in MAFLD model mice, the levels of serum total cholesterol, triglycerides, low-density lipoprotein cholesterol, and high-density lipoprotein cholesterol were determined by testing four blood lipids, and the levels of serum alanine aminotransferase and aspartate aminotransferase were detected using kits.

[0047] 5. Liver histomorphological evaluation (NAS score) The HE-stained images of liver tissue sections of mice in each group were scored according to the NAS scoring system developed by the Pathology Working Group of the American Association for Clinical Research on Nonalcoholic Fatty Liver Disease. Histological parameters included liver ballooning, steatosis, inflammation, and fibrosis, and the sum of the scores was summarized as the NAS score.

[0048] 6. Liver tissue TG, TC, MDA, SOD detection After drug intervention in MAFLD model mice, the levels of total cholesterol (TC), triglyceride (TG), malondialdehyde (MDA) and superoxide dismutase (SOD) in the liver tissue homogenate of mice were determined according to the kit instructions.

[0049] 7. Histopathological examination Liver tissue was collected and fixed in 4% paraformaldehyde for 24 hours. The tissue was then dehydrated using graded ethanol and xylene, embedded in paraffin, and sectioned at 4 μm thickness. Hematoxylin and eosin (HE) and Sirius red staining were performed according to the manufacturer's instructions. Frozen liver sections were dehydrated in sucrose solution and then stained with Oil Red O by incubation for 30 minutes. The sections were then observed and photographed under a microscope.

[0050] 8. Fibrosis index detection After drug intervention in MAFLD model mice, the serum levels of fibrosis indicators HA, Col-IV, and LN were detected by ELISA.

[0051] 9. Immunohistochemical staining Liver tissue was fixed in 4% paraformaldehyde and embedded in paraffin blocks. 4-μm liver sections were prepared according to standard immunohistochemistry protocols. Liver sections were dehydrated and incubated with primary and secondary antibodies. The area of ​​positive staining was measured in high-power fields on each slide and quantified using Image J software.

[0052] 10. Quantitative Real-Time PCR Total RNA was extracted from liver tissue and subjected to reverse transcription and quantitative real-time PCR (q-PCR) analysis using a real-time PCR system (LightCycler 480 Instrument II, Roche Diagnostics, Basel, BS, Switzerland) according to standard protocols. GAPDH was used as an internal control. Primer sequences are shown in Table 1.

[0053] Table 1 Primer sequences 11. Statistical analysis GraPhPad Prism 9.4.1 software was used for data analysis and plotting. Two-group comparisons were performed using a two-tailed unpaired t-test, and multiple group comparisons were performed using one-way ANOVA with Tukey's multiple comparison test. Data are presented as mean ± standard error ( ±s), and P<0.05 indicated that the difference was statistically significant.

[0054] Experimental results: 1. Body weight and food intake of mice The body weight of mice in each group Figure 1 , food intake are shown in Table 2. Compared with the blank control group, the body weight of the model group increased significantly, and the food intake decreased significantly; compared with the model group, the body weight of the high-dose and low-dose compound preparation groups decreased significantly.

[0055] Table 2 Statistics of food intake of each group (n=6, ±s) Note: Compared with the blank group: #### P<0.0001.

[0056] 2. Liver appearance, liver index and body fat percentage The appearance of the liver of mice in each group showed ( Figure 2 ), the livers of mice in the blank control group were dark red and soft in texture; the liver volume of the model group was significantly increased, the liver surface was earthy white and the toughness was enhanced; the liver volume of the compound preparation group was significantly reduced compared with the model group, the liver surface color turned dark brown and the toughness was weakened; the liver volume of the Sily-positive control group was significantly reduced compared with the model group, and the liver surface color did not change much.

[0057] The results of liver index and body fat percentage measurement of mice in each group showed (Table 3) that compared with mice in the Con group, the liver index of mice in the Mod group was significantly increased, the fat content was significantly increased, and the muscle content was significantly decreased; compared with the Mod group, the liver index of each drug-treated group was significantly decreased, the liver weight was significantly decreased, the fat percentage was significantly decreased, and the muscle percentage was significantly increased.

[0058] Table 3 Statistics of liver index and body fat percentage in each group (n=6, ±s) Note: Compared with the blank group: ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0059] 3. Oral glucose tolerance and insulin resistance The results of OGTT and insulin resistance of mice in each group showed that ( Figure 3 (Table 4). Compared with the Con group, the Mod group showed significantly decreased oral glucose tolerance and increased insulin resistance. All other treatment groups showed significantly improved oral glucose tolerance and decreased insulin resistance compared with the Mod group. This suggests that the compound preparation or Sily treatment effectively improves glucose tolerance and insulin resistance in MAFLD mice.

[0060] Table 4 Statistics of glucose area under the curve, serum insulin content, and insulin resistance in each group (n=6, ±s) Note: Compared with the blank group: ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0061] 4. Serum lipid levels and liver function damage Results of the four blood lipid profiles (Table 5) showed that compared with the Con group, the Mod group had significantly increased serum TC, TG, and LDL-C levels, and significantly decreased HDL-C levels. After intervention with Sily and different doses of the compound preparation, the mice's serum TC, TG, and LDL-C levels were significantly decreased, while their HDL-C levels were significantly increased. This suggests that the compound preparation can effectively improve the blood lipid profile of MAFLD mice, reducing serum TC, TG, and LDL-C levels and increasing HDL-C levels.

[0062] Table 5 Statistics of four blood lipids in each group (n=6, ±s) Note: Compared with the blank group: ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0063] The results of liver function index measurement showed (Table 6) that the serum AST and ALT levels of mice in the Mod group were significantly higher than those in the Con group, indicating that the liver function of mice in the Mod group was damaged. Sily and different doses of the compound preparation group could significantly reduce the serum AST and ALT levels of MAFLD mice induced by high fat and high sugar, and improve liver damage. Among them, the compound preparation was better than silymarin.

[0064] Table 6 Statistics of liver function indicators in each group (n=6, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0065] 5. Pathological staining and NAS scoring The results of H&E staining of liver tissue sections of mice in each group showed that ( Figure 4 ), the hepatocytes of the mice in the Mod group were swollen and rounded compared with those in the normal control group, with lipid droplet infiltration, ballooning and more intralobular inflammation; the liver plates of the mice in the Sily and different doses of the compound preparation groups were arranged relatively neatly, the hepatic lobule structure was basically normal, with a small amount of lipid droplet infiltration and ballooning, and the hepatic lobular inflammation was improved, with no obvious inflammatory cell accumulation foci, especially in the high-dose compound preparation group.

[0066] H&E-stained liver sections from each group of mice were observed microscopically, and five randomly selected fields of view were scored for NAS pathology. The results, as shown in Table 7, showed significant increases in the liver steatosis index, ballooning index, and inflammatory cell infiltration index in the Mod mice, demonstrating typical MAFLD characteristics. NAS pathology scores in all treatment groups were significantly decreased compared to those in the Mod group, demonstrating significant improvement. This suggests that Sily and the compound preparation can improve NAFLD pathological features such as hepatic lipid accumulation, ballooning, and inflammatory cell infiltration.

[0067] Table 7 NAS scores of each group (n=6, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001 6. Improvement of oxidative stress indicators and liver lipids The results of the oxidative stress levels in the liver tissues of each group showed (see Table 8) that compared with the Con group, the MDA level in the liver of the mice in the Mod group was significantly increased, the SOD level was significantly decreased, and the oxidative stress in the liver of the mice in the Mod group was enhanced; after treatment with Sily and different doses of the compound preparation, the MDA level in the liver of the MAFLD mice induced by high sugar and high fat was significantly reduced, the SOD level was increased, and the oxidative stress was improved.

[0068] Table 8 Statistics of oxidative stress levels in liver tissues of each group (n=6, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0069] like Figure 5 As shown, mice in the Mod group had extensive liver steatosis and lipid deposition (Table 9), with significantly elevated liver TG and TC levels (Table 10), indicating severe liver lipid accumulation. Administration of Sily and different doses of the compound preparation significantly improved liver lipid deposition and decreased liver TG and TC levels, indicating that the compound preparation effectively ameliorates hepatic steatosis in high-glucose and high-fat-induced MAFLD mice.

[0070] Table 9 Statistics of lipid droplet area in each group (n=3, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0071] Table 10 Statistics of TG and TC contents in liver tissues of each group (n=3, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0072] The results of gene expression related to lipid synthesis (Fasn, SrebP-1c, Scd1, PPar-γ, ACC), transport (Cd36) and fatty acid oxidation (CPt1a, PPar-α, Atgl) showed (Table 11). The compound preparation and Sily treatment could significantly reduce the mRNA levels of Fasn, SrebP-1c, Scd1, Cd36, ACC and PPar-γ in MAFLD mice, and reverse the abnormal downregulation of mRNA levels of Atgl, CPt1α and PPar-α, indicating that the compound preparation can effectively improve lipid metabolism disorders in MAFLD mice induced by high glucose and high fat.

[0073] Table 11 Statistics of mRNA levels of genes related to liver lipid metabolism in each group (n=6, ±s) Note: Compared with the blank group: ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001 7. Improvement of fibrosis indicators like Figure 6 As shown in Table 12, the livers of mice in the Mod group showed significant fibrotic lesions compared to the Con group, with significantly more collagen fibers concentrated around the portal areas and around degenerated hepatocytes. Treatment with Sily and different doses of the compound preparation significantly reduced collagen fiber deposition in MAFLD mice. Furthermore, the compound preparation significantly reduced serum levels of fibrosis markers LN, HA, and ColIV (Table 13). Compared with the Con group, the mRNA expression of pro-fibrotic markers was significantly upregulated in the Mod group. Administration of the compound preparation significantly inhibited the gene expression of α-SMA, Col1a1, FN-1, and Tgf-β1 in the livers of MAFLD mice (Table 14). This indicates that the compound preparation significantly improves liver fibrosis in MAFLD mice.

[0074] Table 12 Statistics of collagen fibers in liver sections of each group (n=3, ±s) Note: Compared with the blank group: ### P<0.001; compared with the model group: **P<0.01.

[0075] Table 13 Statistics of serum fibrosis factor levels in each group (n=6, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ###P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0076] Table 14 Statistics of mRNA levels of liver fibrosis-related genes in each group (n=6, ±s) Note: Compared with the blank group: ### P<0.001; compared with the model group: **P<0.01, ***P<0.001.

[0077] 8. Improvement of inflammation indicators F4 / 80 immunohistochemistry results showed that ( Figure 7 (Table 15) Compared with the Con group, the number of F4 / 80-positive cells in the livers of mice in the Mod group increased significantly, and the level of hepatic macrophage infiltration was significantly enhanced. Compared with the Mod group, the number of F4 / 80-positive cells in the livers of mice in the compound preparation group decreased significantly, indicating that the compound preparation significantly inhibited macrophage infiltration in the livers of mice. Real-time quantitative PCR was used to examine the gene expression of inflammatory factors in the livers of mice in each group. The results are shown in Table 16. Compared with the Mod group, long-term oral administration of the compound preparation significantly inhibited the gene expression of TNF-α, IL-1β, IL-6, and MCP-1 in MAFLD mice, and also showed a trend of activating the gene expression of the anti-inflammatory factor IL-10 (Table 16). This suggests that the compound preparation has an ameliorative effect on the inflammatory response in MAFLD mice.

[0078] Table 15 Statistics of positive areas of F4 / 80 immunohistochemical staining in liver sections of each group (n=3, ±s) Note: Compared with the blank group: # P<0.05, ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

[0079] Table 16 Statistics of mRNA levels of genes related to liver inflammatory factors in each group (n=6, ±s) Note: Compared with the blank group: ## P<0.01, ### P<0.001; compared with the model group: *P<0.05, **P<0.01, ***P<0.001.

Claims

1. A Chinese medicine composition for treating metabolic-related fatty liver disease, characterized in that: Includes the following ingredients: red peony root, cyperus rotundus, zedoaria, barbata scabra, epimedium, bergamot, cassia seed and capillaris.

2. The Chinese medicine composition according to claim 1, characterized in that The dosage ratio of the above ingredients is as follows: 6-12 parts by weight of red peony root, 6-12 parts by weight of rhizoma cyrtomii, 6-18 parts by weight of zedoaria, 5-15 parts by weight of scutellaria barbata, 6-18 parts by weight of epimedium, 6-18 parts by weight of bergamot, 6-18 parts by weight of cassia seed and 5-15 parts by weight of capillaris.

3. The Chinese medicine composition according to claim 2, characterized in that The dosage ratio of the above ingredients is as follows: 6 parts by weight of red peony root, 6 parts by weight of rhizoma cyrtomii, 6 parts by weight of zedoaria, 5 parts by weight of scutellaria barbata, 6 parts by weight of epimedium, 6 parts by weight of bergamot, 6 parts by weight of cassia seed and 5 parts by weight of capillaris; and / or, 12 parts by weight of red peony root, 12 parts by weight of rhizoma cyrtomii, 18 parts by weight of zedoaria, 15 parts by weight of scutellaria barbata, 18 parts by weight of epimedium, 18 parts by weight of citron, 18 parts by weight of cassia seed and 15 parts by weight of capillaris.

4. The method for preparing the Chinese medicine composition according to claim 1, characterized in that: The steps include: a. Mix red peony root, zedoary turmeric and epimedium, extract twice with 70% ethanol as extract, combine the extracts, recover ethanol, and obtain filtrate 1; b. Mix the medicinal residue after extraction in step a with Guanzhong, Scutellaria barbata, Artemisia capillaris, Citron, and Cassia seed, add water and extract twice, combine the extracts, and obtain filtrate 2; c. Combine filtrate 1 and filtrate 2, concentrate, dry and crush.

5. The preparation method according to claim 4, characterized in that: In step a, the mass-to-volume ratio (g / mL) of the feed and liquid is 1:6-1:15, and the time for each extraction is 0.5h-2h; in step b, the mass-to-volume ratio (g / mL) of the feed and liquid during the first extraction is 1:6-1:15; the mass-to-volume ratio (g / mL) of the feed and liquid during the second extraction is 1:6-1:12; the time for each extraction is 0.5h-2h.

6. The preparation method according to claim 5, characterized in that: In step a, the mass-to-volume ratio of the feed and liquid (g / mL) is 1:8, and the time for each extraction is 1 hour; in step b, the mass-to-volume ratio of the feed and liquid (g / mL) during the first extraction is 1:10; the mass-to-volume ratio of the feed and liquid (g / mL) during the second extraction is 1:8; the time for each extraction is 1 hour.

7. The preparation method according to claim 4, characterized in that: In step a, ethanol is recovered by using a decompression technique; and in step b, extraction is performed by using a heating, condensation, and reflux method.

8. A medicine or health product for weight loss, lipid-lowering and blood sugar-lowering, and anti-oxidation, characterized in that: The invention comprises the Chinese medicine composition according to any one of claims 1 to 3.

9. The drug according to claim 8, characterized in that The preparation of the medicine also includes pharmaceutically acceptable excipients, and the dosage form of the preparation is tablets, pills, capsules, granules, powders, ointments, sprays or mixtures.

10. Use of the Chinese medicine composition according to any one of claims 1 to 3 in the preparation of a medicament for treating metabolism-related fatty liver disease, improving metabolism-related fatty liver disease, controlling body weight and food intake, reducing liver index and liver weight, reducing body fat percentage, improving glucose intolerance and insulin resistance, improving blood lipids, improving liver damage, improving liver lipid accumulation, improving liver ballooning, improving liver inflammatory cell infiltration, improving liver steatosis or lipid metabolism disorders caused by a high-sugar and high-fat diet, improving inflammatory response, improving liver fibrosis levels and / or improving liver tissue oxidative stress levels.

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