Use of a combination of *Lysimachia christinae* and *Brain-Clearing Agent* in the preparation of a medicament for the prevention and / or treatment of non-alcoholic fatty liver disease.

Abstract

This invention belongs to the field of traditional Chinese medicine and discloses the application of a *Dendrobium nobile* and *Ginkgo biloba* composition in the preparation of drugs for the prevention and / or treatment of non-alcoholic fatty liver disease. The *Dendrobium nobile* and *Ginkgo biloba* composition includes *Dendrobium nobile*, *Ginkgo biloba* leaves, *Panax notoginseng*, and *Houttuynia cordata*. The *Dendrobium nobile* and *Ginkgo biloba* composition can significantly reduce steatosis and ballooning degeneration in liver tissue and reduce lobular inflammation; simultaneously, it can inhibit macrophage infiltration in liver tissue, significantly reducing blood lipids and lipid levels in the liver. This invention expands the clinical application of the *Dendrobium nobile* and *Ginkrobium nobile* composition, such as in capsule form, and provides experimental and theoretical basis for the treatment of non-alcoholic fatty liver disease.

Description

Technical Field

[0001] This invention belongs to the field of traditional Chinese medicine, specifically, it relates to the use of a Dengyin Naotong composition in the preparation of a medicament for the prevention and / or treatment of non-alcoholic fatty liver disease. Background Technology

[0002] Nonalcoholic fatty liver disease (NAFLD) is the most common chronic liver disease worldwide, characterized primarily by excess fat accumulation and degeneration in the liver, promoting the progression of inflammation. In recent years, the incidence of NAFLD has been rising, affecting approximately one-quarter of adults globally, and its prevalence continues to increase. NAFLD is a collective term for a range of chronic liver diseases, starting with simple fatty liver. 20% of NAFLD patients will progress to nonalcoholic steatohepatitis (NASH) during disease progression, and some may also develop cirrhosis and hepatocellular carcinoma. Furthermore, NAFLD patients often experience cardiovascular complications such as hypertension and atherosclerosis, possibly related to their dyslipidemia and pro-inflammatory state throughout the body and in the liver.

[0003] The liver plays a crucial role in lipid metabolism, including lipid uptake, synthesis, oxidation, and distribution to peripheral tissues. The pathogenesis of NAFLD is not fully understood, but in general, the main cause is the imbalance between fatty acids transported to the liver (including from diet, endoadrenal lipid production (DNL), and lipolysis of adipose tissue) and lipids synthesized in the liver, and triglycerides (TG) exported from the liver as very low-density lipoprotein.

[0004] NAFLD has a slow onset, with most patients experiencing no obvious symptoms. Occasionally, nonspecific manifestations such as discomfort in the liver area and hepatosplenomegaly may occur. Liver biopsy revealing histological changes can definitively diagnose the disease. However, since performing liver biopsy on every patient is difficult to achieve, current clinical diagnosis primarily relies on relevant biochemical indicators such as blood lipids and liver function tests, combined with auxiliary examinations such as imaging changes. Alcoholic fatty liver and other liver diseases must also be ruled out. Currently, clinical management mainly focuses on controlling the disease through lifestyle interventions such as diet and exercise. Although studies have shown that good lifestyle habits can significantly improve NAFLD, most patients are unable to maintain these habits consistently due to various factors, and there is a lack of specific drugs for NAFLD. Therefore, the development of NAFLD-related treatments is particularly important.

[0005] Currently, there is no specific drug for treating NAFLD in clinical practice. Treatment often involves single or combined medications such as insulin sensitizers, lipid-lowering drugs, and hepatoprotective drugs. However, these drugs have significant side effects, and their effectiveness and safety require further confirmation. Traditional Chinese medicine's (TCM) approach of syndrome differentiation and treatment has unique advantages in the clinical treatment of NAFLD, offering benefits such as high efficacy and fewer side effects. Therefore, TCM, with its unique advantages, demonstrates extremely high research value in the treatment of NAFLD.

[0006] Traditional Chinese medicine (TCM) considers fatty liver to be mainly referred to as "accumulation," "abdominal masses," and "fatness." The "Expert Consensus on the Diagnosis and Treatment of Nonalcoholic Fatty Liver Disease with Traditional Chinese Medicine (2017)" classifies the disease into five syndrome types: dampness retention syndrome, liver stagnation and spleen deficiency syndrome, damp-heat accumulation syndrome, phlegm and blood stasis syndrome, and spleen and kidney deficiency syndrome. Among these, "phlegm" and "blood stasis" are the most important pathological factors.

[0007] The Dengyin Naotong drug composition, such as Dengyin Naotong capsules, is a traditional Chinese medicine compound preparation composed of four Chinese medicinal herbs: *Dendrobium nobile*, *Ginkgo biloba*, *Panax notoginseng*, and *Houttuynia cordata*. Its high-content active ingredients include dinoflagellin, ginkgolide A, ginkgolide B, ginsenoside Rg1, and notoginsenoside R1. Dengyin Naotong capsules are used clinically to treat stroke with blood stasis obstructing the meridians, possessing the functions of promoting qi and blood circulation, dispersing blood stasis, and clearing the meridians. Laboratory and clinical studies suggest that it has pharmacological effects such as promoting cognitive function, protecting against cerebral ischemia, and anticoagulation. Clinically, it is widely used for cerebrovascular diseases such as cerebral infarction and dementia. However, its mechanism of action requires further research, and more research data is needed to support its potential for wider clinical application and promotion.

[0008] The pharmacological effects and clinical applications of Dengyin Naotong capsules require further exploration. Currently, there are no literature reports on whether the Dengyin Naotong drug composition has an interventional effect on non-alcoholic fatty liver disease. Therefore, this invention is proposed. Summary of the Invention

[0009] The technical problem to be solved by the present invention is to overcome the shortcomings of the prior art and provide an application of the Dengyin Naotong composition in the preparation of drugs for the prevention and / or treatment of non-alcoholic fatty liver disease, thereby expanding the application scope of Dengyin Naotong capsules and providing experimental and theoretical basis for the clinical treatment of non-alcoholic fatty liver-related diseases.

[0010] To solve the above-mentioned technical problems, the basic concept of the technical solution adopted by the present invention is as follows:

[0011] The first objective of this invention is to provide the use of the Dengyin Naotong composition in the preparation of a medicament for the prevention and / or treatment of non-alcoholic fatty liver disease.

[0012] The Dengyin Naotong composition is composed of four traditional Chinese medicinal herbs: *Erigeron breviscapus*, *Ginkgo biloba*, *Panax notoginseng*, and *Houttuynia cordata*. Based on this composition, the existing product, Dengyin Naotong capsules, is listed in the National Drug Standard WS-10843(ZD-0843)-2002-2012Z. Currently, it is mainly used to treat stroke. Experiments have shown that the Dengyin Naotong composition has a significant inhibitory effect on non-alcoholic fatty liver disease (NAFLD) and can be used to prepare drugs that inhibit NAFLD.

[0013] A further approach is to use the Dengyin Naotong composition in the preparation of a drug that reduces steatosis in liver tissue.

[0014] A further approach is to use the Dengyin Naotong composition in the preparation of a drug that reduces ballooning degeneration in liver tissue.

[0015] A further proposed approach is the application of the Dengyin Naotong composition in the preparation of drugs that reduce inflammation in liver tissue.

[0016] Preferably, the inflammation is lobular inflammation of the liver tissue.

[0017] A further approach is to use the Dengyin Naotong composition in the preparation of a drug that reduces lipid deposition in the liver.

[0018] A second objective of this invention is to provide the use of the Dengyin Naotong composition in the preparation of a medicament for inhibiting macrophage infiltration in liver tissue.

[0019] A third objective of this invention is to provide the use of the Dengyin Naotong composition in the preparation of a medicament for reducing blood lipids and / or lipid levels in the liver.

[0020] A further approach is to use the Dengyin Naotong composition to reduce the levels of total cholesterol and triglycerides in serum and / or liver tissue.

[0021] In a further embodiment, the Dengyinnaotong composition comprises Dengzhanxixin, Ginkgo biloba leaf, Panax notoginseng, and Manshanxiang;

[0022] In a further embodiment, the Dengyinnaotong composition comprises: 300-600 parts by weight of Dengzhanxixin, 300-600 parts by weight of Manshanxiang, 300-600 parts by weight of Ginkgo biloba leaves, and 100-300 parts by weight of Panax notoginseng.

[0023] Preferably, the Dengyinnaotong composition comprises: 400-550 parts by weight of Dengzhanxixin, 400-550 parts by weight of Manshanxiang, 400-550 parts by weight of Ginkgo biloba leaves, and 150-250 parts by weight of Panax notoginseng.

[0024] More preferably, the Dengyinnaotong composition comprises: 500 parts by weight of Dengzhan Xixin, 500 parts by weight of Manshanxiang, 500 parts by weight of Ginkgo biloba leaves, and 200 parts by weight of Panax notoginseng.

[0025] Preferably, the lamp-silver brain-tonifying composition further includes pharmaceutically acceptable excipients;

[0026] Preferably, the dosage form of the Dengyin Naotong composition is selected from capsules, tablets, injections, powders, pills, granules, or oral liquids.

[0027] In a preferred embodiment, the Dengyin Naotong composition is Dengyin Naotong capsules, and more preferably, the approval number of the Dengyin Naotong capsules is: National Drug Approval Number Z20026228.

[0028] By adopting the above technical solution, the present invention has the following beneficial effects compared with the prior art:

[0029] 1. This invention has discovered that Dengyin Naotong capsules have a significant inhibitory effect on non-alcoholic fatty liver disease and a significant effect on reducing blood lipids and lipid levels in the liver. Specifically:

[0030] (1) H&E and Oil Red O staining of liver tissue showed that APOE- / - mice fed a high-fat diet developed obvious non-alcoholic fatty liver disease, and Dengyin Naotong capsules could significantly alleviate these pathological changes.

[0031] (2) Immunofluorescence staining of CD68, a macrophage marker in liver tissue, was used to further observe the inflammatory status of the livers in each group of mice. The results showed that compared with the normal group (C57 group), the non-alcoholic fatty liver disease model group (HFD group) had a large number of macrophage infiltrations in the liver tissue, which was consistent with the multiple inflammatory lesions observed by liver H&E staining. The mice in the Dengyin Naotong capsule administration group showed that this change was alleviated to varying degrees.

[0032] (3) The effects of Dengyin Naotong capsules on blood lipids and liver lipids in mice with non-alcoholic fatty liver disease were observed by biochemical detection. The results showed that Dengyin Naotong capsules could significantly reduce blood lipids and liver lipids in mice with non-alcoholic fatty liver disease.

[0033] The new uses of the Dengyin Naotong composition discovered in this invention expand the scope of clinical application of Dengyin Naotong composition (such as Dengyin Naotong capsules) and have broad application prospects. At the same time, it also provides experimental and theoretical basis for the treatment of non-alcoholic fatty liver disease.

[0034] The specific embodiments of the present invention will now be described in further detail with reference to the accompanying drawings. Attached Figure Description

[0035] The accompanying drawings, as part of this invention, are used to provide a further understanding of the invention. The illustrative embodiments and descriptions of the invention are used to explain the invention, but do not constitute an undue limitation of the invention. Obviously, the drawings described below are merely some embodiments, and those skilled in the art can obtain other drawings based on these drawings without creative effort. In the drawings:

[0036] Figure 1 A shows H&E staining of liver tissue sections from mice in each group. Mouse hepatocyte steatosis is indicated by a "red arrow", ballooning degeneration by a "white arrow", and hepatic lobule inflammation by a "black arrow". Figure 1 In the HFD group staining diagram of A, the arrows from top to bottom are white, red, and black. Figure 1 B represents the histological scores of hepatocyte steatosis, ballooning degeneration, lobular inflammation, and NAS score for each group of mice. Figure 1 C represents Oil Red O staining of liver tissue sections from mice in each group; Figure 1 D represents the statistical analysis of the positive area of ​​Oil Red O staining in each group of mice (Note: **P < 0.01);

[0037] Figure 2 A shows the immunofluorescence staining of CD68 in each group of mice. Figure 2 B represents the statistical analysis of CD68-positive area in each group of mice (Note: *P<0.05; ***P<0.001; nsP>0.05);

[0038] Figure 3 A represents the serum TC content of mice in each group. Figure 3 B represents the serum TG content of mice in each group. Figure 3 C represents the TC content in the liver tissue of mice in each group. Figure 3 D represents the TG content in the liver tissue of mice in each group (Note: *P<0.05; **P<0.01; ***P<0.001; ***P<0.0001; nsP>0.05).

[0039] It should be noted that these accompanying drawings and textual descriptions are not intended to limit the scope of the invention in any way, but rather to illustrate the concept of the invention to those skilled in the art by referring to specific embodiments. Detailed Implementation

[0040] To make the objectives, technical solutions, and advantages of the embodiments of the present invention clearer, the technical solutions in the embodiments will be clearly and completely described below with reference to the accompanying drawings. The following embodiments are used to illustrate the present invention, but are not intended to limit the scope of the present invention.

[0041] It should be noted that the Dengyin Naotong composition described in this invention includes any traditional Chinese medicine compound preparation composed of four medicinal herbs: *Erigeron breviscapus*, *Ginkgo biloba*, *Panax notoginseng*, and *Houttuynia cordata*. The dosage form is not limited to any particular herbal medicine; it can be a capsule, a tablet, or other dosage form. Specifically, Dengyin Naotong capsules were used in the experiments conducted in this invention.

[0042] I. Dengyin Naotong Composition (Dengyin Naotong Capsules)

[0043] Prescription: 500g of Asarum sieboldii, 500g of Clematis armandii, 500g of Ginkgo biloba leaves, and 200g of Panax notoginseng.

[0044] Preparation: Soak the above four ingredients in water for 4 hours, decoct three times for 2 hours each time, combine the decoctions, filter, add ethanol to the filtrate to make the alcohol content reach 70%, mix well, let stand for 24 hours, filter, recover the ethanol from the filtrate until there is no alcohol taste, dry under reduced pressure, grind into powder, add 100g starch, 30g microcrystalline cellulose, 3g polysorbate 80, 0.5g hydroxypropyl methylcellulose, 2g magnesium stearate, and 17.5g sodium carboxymethyl cellulose, granulate, fill into capsules, make 1000 capsules, and obtain the product.

[0045] The Dengyin Naotong capsules used in the following experiments of this invention were produced by Kunming Pharmaceutical Group Co., Ltd.

[0046] II. Experimental Methods

[0047] 1. Animal models and grouping for drug administration:

[0048] Select 8-week-old SPF-grade ApoE - / - Twenty-four mice, including eight C57 mice (half male and half female), were sourced from Shanghai Slack Animal Co., Ltd. They were housed at the Animal Experiment Center of Yueyang Hospital of Integrated Traditional Chinese and Western Medicine, affiliated with Shanghai University of Traditional Chinese Medicine. The housing temperature was 25±2℃; humidity was 40%-55%; and the light cycle was 12h light / 12h dark. All procedures complied with the Ethics Guidelines of the Experimental Animal Management Committee of Shanghai University of Traditional Chinese Medicine (18867). After one week of free access to water and food for acclimatization, the 24 ApoE mice were transferred to... - / -Mice were randomly divided into three groups: a model group (high-fat diet, 0.5% sodium carboxymethyl cellulose (CMCNA), designated HFD), a low-dose Dengyinnaotong administration group (high-fat diet, 0.2 g / kg / d Dengyinnaotong capsules dissolved in 0.5% CMCNA, designated DYNT-L), and a high-dose Dengyinnaotong administration group (high-fat diet, 1 g / kg / d Dengyinnaotong capsules dissolved in 0.5% CMCNA, designated DYNT-H). C57 mice were given a normal diet as a control group, designated C57. Each mouse in the administration groups was administered 200 μl of different concentrations of Dengyinnaotong capsules by gavage. The normal control group and the high-fat diet group were given an equal volume of 0.5% CMCNA for 8 consecutive weeks. The grouping and administration are shown in Table 1 below.

[0049] Table 1

[0050] Group Animal numbers diet dose Dosage time C57 8 normal diet -- Lasting 8 weeks HFD 8 High-fat diet -- Lasting 8 weeks DYNT-L 8 High-fat diet 0.2g / kg / d Lasting 8 weeks DYNT-H 8 High-fat diet 1g / kg / d Lasting 8 weeks

[0051] 2. Animal handling:

[0052] Following the final drug intervention, blood was collected from the orbital fossa the day after the intervention. The blood was allowed to stand for 20 minutes, then centrifuged at 4°C and 3500 rpm for 10 minutes to separate the serum. Mice were euthanized, their hearts were fixed and exposed, and the apex of the heart was perfused with 0.9% saline until clear fluid flowed out. The largest lobe of the left liver was harvested and fixed in 4% paraformaldehyde for 24 hours. Half of the lobe was dehydrated and embedded in paraffin to prepare 4μm paraffin sections; the other half was dehydrated with sucrose and embedded in OCT to prepare 8μm frozen sections for histopathological examination and analysis. The largest lobe of the right liver was cryopreserved at -80°C for subsequent molecular mechanism studies.

[0053] 3. Liver histopathological examination and analysis (H&E and Oil Red O staining)

[0054] a. Obtain liver paraffin sections for H&E staining to observe pathological changes in liver tissue.

[0055] b. Frozen sections of the liver were used for Oil Red O staining to observe lipid deposition in the aortic tissue.

[0056] c. Obtain liver paraffin sections for CD68 immunofluorescence staining to observe macrophage infiltration in liver tissue.

[0057] The methods are as follows:

[0058] H&E staining

[0059] (1) Drying the sheet: Dry the sheet in a 60℃ oven for 1 hour.

[0060] (2) Dewaxing: Place the baked paraffin slices in xylene I, II and III in sequence and soak for 30 min each.

[0061] (3) Hydration: The dewaxed paraffin slices were placed in anhydrous ethanol → anhydrous ethanol → 90% ethanol → 80% ethanol in sequence, and each soaked for 2 minutes. Then they were rinsed with running water for 10 minutes.

[0062] (4) Staining: Place the sections in hematoxylin staining solution for 4-7 minutes.

[0063] (5) Rinse: Rinse quickly with tap water and then rinse with running water for 10 minutes.

[0064] (6) Staining: Soak the sections in eosin staining solution for 2 minutes.

[0065] (5) Dehydration: Place the stained sections in 95% ethanol I and II for 30s each, then in anhydrous ethanol I and II for 2min each.

[0066] (6) Transparency: Place the dehydrated slices in xylene I and II in sequence and soak for 2 minutes each.

[0067] (7) Sealing: Neutral resin sealing.

[0068] (8) Image acquisition: Images were acquired using a regular optical microscope at 200x magnification.

[0069] (9) Data processing: The liver tissue sections of mice in each group were semi-quantitatively analyzed by the non-alcoholic fatty liver scoring (NAS) system [1].

[0070] ([1]Kleiner DE,Brunt EM,Van Natta M,et al.Design and validation of ahistological scoring system for nonalcoholic fatty liverdisease.Hepatology.2005;41(6):1313-1321.doi:10.1002 / hep.20701)

[0071] Oil Red O staining

[0072] (1) Preparation of dye solution: 2.5g of oil red O and 500mL of 70% ethanol are mixed and shaken several times at intervals. After 24 hours, a saturated solution is formed and it is ready for use.

[0073] (2) Air-dry the frozen tissue sections at room temperature for 15 minutes, rinse with running water for 5 minutes, and soak in 70% ethanol for 5 seconds.

[0074] (3) Immerse in Oil Red O staining solution for 15 min.

[0075] (4) Immerse in 70% ethanol for 30 seconds, then rinse twice with distilled water.

[0076] (5) Glycerin gelatin sealing sheet.

[0077] (6) Image acquisition: Images were acquired using a regular optical microscope at 200x magnification.

[0078] (7) Data processing: Use Image Pro to calculate the positive area.

[0079] Immunohistofluorescence staining

[0080] (1) Drying the sheet: Dry the sheet in a 60℃ oven for 1 hour.

[0081] (2) Dewaxing: Place the baked paraffin slices in xylene I, II and III in sequence and soak for 30 min each.

[0082] (3) Hydration: The dewaxed paraffin sections were placed in anhydrous ethanol, anhydrous ethanol, 90% ethanol and 80% ethanol in sequence, and each was soaked for 2 minutes. Then, they were rinsed with running water for 10 minutes, soaked in pure water for 5 minutes, and washed twice with PBST for 5 minutes each time.

[0083] (4) Antigen retrieval: Heat a 10% sodium citrate solution to boiling in a microwave oven → stop boiling and place the slice in the microwave oven → heat for 15 seconds in the microwave oven → stop for 105 seconds (avoid boiling the liquid as much as possible) → repeat heating for 30 minutes → cool naturally to room temperature.

[0084] (5) Membrane permeation: Wash 3 times with PBST for 10 min each time, and soak the sections in 0.1% Triton X-100 for 10 min to increase membrane permeability.

[0085] (6) Blocking: Wash 3 times with PBST for 10 min each time, wipe the tissue around the section dry, and incubate with 10% goat serum (consistent with the source of the secondary antibody) at room temperature for 60 min. The primary antibody is CD68: Anti-CD68 antibody (ab125212) abcam rabbit source, and the secondary antibody is sheep-rabbit: SIGMA C2306.

[0086] (7) Incubation with primary antibody: Wipe the tissue section dry, add primary antibody (CD68) to cover the specimen, and incubate overnight at 4°C.

[0087] (8) Incubation with secondary antibody: After taking the slides out of the 4℃ refrigerator, wash them 3 times with PBST for 10 min each time, wipe the slides dry, add fluorescent secondary antibody to cover the specimens, and incubate at 37℃ for 30 min.

[0088] (9) Nuclear counterstaining: Wash 3 times with PBST for 10 min each time, and add 1 drop of DAPI (final concentration of 1 ug / ml) to each tissue section.

[0089] (10) Mounting: Glycerin gelatin is used for mounting.

[0090] (11) Image acquisition: Images were acquired using an inverted fluorescence microscope at 200x magnification.

[0091] (12) Data processing: Image Pro was used to count the positive area of ​​CD68.

[0092] 4. Serum biochemical detection and analysis

[0093] After centrifuging mouse blood at 3500 rpm for 10 min to extract serum, the biochemical indicators TC and TG in the serum were detected using a biochemical kit from Nanjing Jiancheng Biotechnology Institute, following the instructions.

[0094] 5. Biochemical detection and analysis of liver tissue

[0095] Mouse liver tissue was homogenized using a grinder at a weight (g) to anhydrous ethanol (ml) ratio. The tissue homogenate was centrifuged at 2500 rpm for 10 min, and the supernatant was extracted. Biochemical reagent kits (Nanjing Jiancheng A110-1-1 and Nanjing Jiancheng A111-1-1) from Nanjing Jiancheng Bioengineering Institute were used to detect the biochemical indicators TC (total cholesterol) and TG (triglycerides) in the liver tissue supernatant according to the instructions.

[0096] 6. Statistical Analysis

[0097] GraphPad Prism 9.0 was used to perform statistical analysis and plotting of the experimental data. Data are expressed as mean ± standard error.

[0098] (Mean±SEM) indicates that one-way ANOVA was used for multiple groups of data that conform to normal distribution and homogeneity of variance, and Kruskal-Wallis test was used for comparison between groups for those that do not conform to normal distribution. P<0.05 indicates statistical difference.

[0099] IV. Experimental Results

[0100] 1. Dengyin Naotong capsules can significantly improve non-alcoholic fatty liver disease in APOE- / - mice fed a high-fat diet.

[0101] Figure 1 A, Figure 1C represents representative images of mouse liver tissue sections stained with H&E and Oil Red O, respectively. As shown in the figure, compared with the C57 group, the HFD group mice showed a large number of fatty degeneration, ballooning degeneration, and scattered inflammatory lesions, while the Dengyin Naotong capsule administration group showed a dose-dependent reduction in these pathological changes. Semi-quantitative analysis of the liver tissue sections of each group of mice was performed using the Non-Alcoholic Fatty Liver Scoring (NAS) system. Figure 1 B) The results showed that the high-dose group of Dengyin Naotong capsules significantly reduced hepatic steatosis (P<0.01), ballooning degeneration (P<0.01), hepatic lobular inflammation (P<0.05), and NAS score (P<0.01). Furthermore, statistical analysis of the Oil Red O staining-positive area of ​​liver tissue sections revealed that the high-dose group of Dengyin Naotong capsules significantly reduced lipid deposition in the liver (P<0.05), indicating that Dengyin Naotong capsules have a significant effect in improving non-alcoholic fatty liver disease.

[0102] 2. Dengyin Naotong capsules can significantly inhibit APOE in patients fed a high-fat diet. - / - The macrophage infiltration assay in mouse liver tissue showed that the macrophage infiltration score was 5.625±0.375 in the HFD group, while it was 4.75±0.366 and 3.625±0.375 in the DYNT-L and DYNT-H groups, respectively. Figure 1 According to the NAS scoring system, a score >3 indicates non-alcoholic fatty liver disease (NAFLD), and a score >5 indicates non-alcoholic steatohepatitis (NASH). Therefore, the experimental results suggest that the HFD group mice may be in a state of non-alcoholic steatohepatitis. To further assess the inflammatory status of the livers in NAFLD mice, we detected the infiltration of macrophages in the liver tissues of each group of mice using CD68 immunofluorescence staining. Figure 2 As shown, compared with the C57 group, the positive area of ​​CD68 in the liver tissue of mice in the HFD group was significantly increased, and the high-dose administration of Dengyin Naotong capsules could significantly inhibit macrophage infiltration (P<0.05).

[0103] 3. Dengyin Naotong capsules can significantly reduce APOE in patients fed a high-fat diet. - / - Blood lipids and lipid levels in the liver of mice

[0104] like Figure 3 As shown, the levels of TC and TG in the serum and liver tissue of mice in each group were detected using biochemical reagent kits. Compared with the C57 group, the levels of TC and TG in the serum and liver tissue of mice in the HFD group were significantly increased. However, compared with the HFD group, the high-dose administration of Dengyin Naotong significantly reduced the levels of TC and TG in the serum and liver of mice (P < 0.05).

[0105] V. Summary

[0106] This experiment used a high-fat diet to feed ApoE- / - mice to establish a non-alcoholic fatty liver disease model, and observed the intervention effect of Dengyin Naotong capsules on non-alcoholic fatty liver disease.

[0107] (1) H&E and Oil Red O staining of liver tissue showed that APOE- / - mice fed a high-fat diet developed obvious non-alcoholic fatty liver disease, and Dengyin Naotong capsules could significantly alleviate these pathological changes.

[0108] (2) Immunofluorescence staining of CD68, a macrophage marker in liver tissue, was used to further observe the inflammatory status of the livers in each group of mice. The results showed that compared with the normal group (C57 group), the non-alcoholic fatty liver disease model group (HFD group) had a large number of macrophage infiltrations in the liver tissue, which was consistent with the multiple inflammatory lesions observed by liver H&E staining. The mice in the Dengyin Naotong capsule administration group showed that this change was alleviated to varying degrees.

[0109] (3) The effects of Dengyin Naotong capsules on blood lipids and liver lipids in mice with non-alcoholic fatty liver disease were observed by biochemical detection. The results showed that Dengyin Naotong capsules could significantly reduce blood lipids and liver lipids in mice with non-alcoholic fatty liver disease.

[0110] Current experimental results suggest that Dengyin Naotong capsules have a significant inhibitory effect on non-alcoholic fatty liver disease and a significant reduction in blood lipids and liver lipid levels. The relevant mechanisms still need further in-depth research.

[0111] The above description is merely a preferred embodiment of the present invention and is not intended to limit the present invention in any way. Although the present invention has been disclosed above with reference to preferred embodiments, it is not intended to limit the present invention. Any person skilled in the art can make some modifications or alterations to the above-described technical content to create equivalent embodiments without departing from the scope of the present invention. Any simple modifications, equivalent changes, and alterations made to the above embodiments based on the technical essence of the present invention without departing from the scope of the present invention shall still fall within the scope of the present invention.

Claims

1. The use of a *Dendrobium nobile* composition in the preparation of a medicament for the prevention and / or treatment of non-alcoholic fatty liver disease, said *Dendrobium nobile* composition comprising the following components: 300-600 parts by weight of Asarum sieboldii, 300-600 parts by weight of Scutellaria baicalensis, 300-600 parts by weight of Ginkgo biloba leaves, and 100-300 parts by weight of Panax notoginseng.

2. The application according to claim 1, characterized in that, The combination of Dengyin Naotong is used in the preparation of drugs that reduce steatosis in liver tissue.

3. The application according to claim 1, characterized in that, The combination of Dengyin Naotong is used in the preparation of drugs that reduce ballooning degeneration in liver tissue.

4. The application according to claim 1, characterized in that, The combination of Dengyin Naotong is used in the preparation of drugs that reduce lipid deposition in the liver.

5. The application according to any one of claims 1-4, characterized in that, The Dengyin Naotong composition also includes pharmaceutically acceptable excipients.

6. The application according to any one of claims 1-4, characterized in that, The dosage form of the Dengyin Naotong composition is selected from capsules, tablets, injections, powders, pills, granules, or oral liquids.

7. The use of a *Dendrobium nobile* composition in the preparation of a medicament for lowering blood lipids and / or lipid levels in the liver, said *Dendrobium nobile* composition comprising the following components: 300-600 parts by weight of Asarum sieboldii, 300-600 parts by weight of Scutellaria baicalensis, 300-600 parts by weight of Ginkgo biloba leaves, and 100-300 parts by weight of Panax notoginseng.

8. The application according to claim 7, characterized in that, The combination of Dengyin Naotong reduces the levels of total cholesterol and triglycerides in serum and / or liver tissue.

9. The application according to claim 7, characterized in that, The Dengyin Naotong composition also includes pharmaceutically acceptable excipients.

10. The application according to any one of claims 7-9, characterized in that, The dosage form of the Dengyin Naotong composition is selected from capsules, tablets, injections, powders, pills, granules, or oral liquids.

Images