Use of the compound extract of radix paeoniae alba and astragalus membranaceus in preparation of a drug for targeted regulation of apo a4 target

Abstract

This invention relates to the application of a compound extract of Sophora flavescens and Astragalus membranaceus in the preparation of drugs targeting and regulating APOA4, belonging to the field of prevention and treatment technology for liver inflammation and cancer progression-related diseases. The compound extract of Sophora flavescens and Astragalus membranaceus proposed in this invention enhances lipid metabolism regulation by dose-dependently regulating the expression or activity of APOA4, thereby achieving effective intervention in liver inflammation and cancer progression-related diseases. The intervention scheme provided by this invention has clear targeting and dose dependence, and can be used to elucidate the function and mechanism of APOA4 in the process of liver inflammation and tumor development, and provide new experimental evidence and clinical application ideas for targeted intervention strategies for liver inflammation and cancer progression-related diseases based on APOA4.

Description

Technical Field

[0001] This invention relates to the application of a compound extract of bitter herbs and astragalus in the preparation of drugs that target and regulate APOA4, belonging to the field of prevention and treatment technology for liver inflammation and cancer progression-related diseases. Background Technology

[0002] Liver neoplastic diseases, especially hepatocellular carcinoma (HCC), are common and highly aggressive malignant diseases in clinical practice. Their occurrence and development are closely related to lipid metabolism reprogramming and persistent chronic inflammation. Abnormal lipid metabolism not only provides the energy and structural basis for abnormally proliferating cells, but also plays a crucial role in key biological processes such as inflammatory response remodeling, alteration of the immune microenvironment, and regulation of cell signal transduction. This continuously propels inflammatory diseases such as chronic hepatitis, liver fibrosis, cirrhosis, and metabolic-related fatty liver disease towards neoplasticity. In this pathological process, systemic imbalance in lipid metabolism is considered a crucial molecular basis connecting chronic inflammation and tumorigenesis.

[0003] The fine regulation of lipid metabolism largely depends on apolipoproteins (APOs)-mediated lipid transport and distribution. Existing research indicates that various APO isoforms are abnormally expressed in liver inflammation and neoplastic diseases. Among them, APOA4, as an important factor regulating triglyceride metabolism and lipid homeostasis, occupies a key position in the hepatic lipid metabolism network. Previous studies have found that APOA4 exhibits a continuous downregulation trend during the development and progression of liver tumors and is closely related to various lipid metabolism disorders; its decreased expression can further exacerbate lipid deposition and inflammatory microenvironment imbalance, thereby creating favorable metabolic conditions for tumor formation and progression. Therefore, APOA4 holds promise as an important regulatory node and potential intervention target connecting lipid metabolism abnormalities, chronic inflammation, and liver tumor development.

[0004] The "Sophora flavescens-Astragalus membranaceus" herb pair has been proven to have a clear basis for clinical application in previous studies. Its related preparations (such as Astragalus membranaceus injection and Kangai injection) are widely used as adjunctive therapy for viral hepatitis, chronic liver disease, and liver tumors due to their anti-inflammatory, anti-fibrotic, immunomodulatory, and anti-tumor biological activities. However, current research mainly focuses on the overall pharmacodynamic characteristics of this herb pair and its regulatory effects on downstream signaling pathways. Whether it plays a key role through specific lipid metabolism regulators remains unclear and requires a systematic and well-defined mechanistic explanation.

[0005] Currently, intervention strategies for liver inflammation-related cancer progression lack precise regulation of key lipid metabolism nodes. Existing technologies have not reported intervention regimens with APOA4 as a clear target, nor have there been studies on using the "Sophora flavescens-Astragalus membranaceus" herb pair in specific combinations to target and regulate APOA4, thereby achieving precise intervention in lipid metabolism and blocking the "inflammation-cancer" transformation. Therefore, there is an urgent need to develop a novel intervention strategy based on APOA4 regulation to compensate for the shortcomings of existing technologies in targeted lipid metabolism therapy and early intervention for inflammation-related liver diseases. Summary of the Invention

[0006] To address the aforementioned shortcomings of existing technologies, this invention provides the application of a compound extract of Sophora flavescens and Astragalus membranaceus in the preparation of drugs that target and regulate APOA4. The compound extract of Sophora flavescens and Astragalus membranaceus improves abnormal hepatic lipid metabolism, restores lipid homeostasis, and inhibits the progression of chronic inflammation and the inflammation-tumor transformation process by regulating the expression or functional state of APOA4, thereby achieving an intervention effect on the pathological states related to the progression of liver inflammation and cancer.

[0007] This invention provides the application of a compound extract of bitter ginseng and astragalus in the preparation of drugs that target and regulate APOA4.

[0008] Furthermore, the bitter herbs and astragalus were combined according to the mass ratio of raw herbs, mixed, and then extracted by heating and reflux with pure water, filtered, and concentrated to obtain a compound extract solution of bitter herbs and astragalus.

[0009] More preferably, the bitter ginseng and astragalus are combined at a raw drug mass ratio of 3:1, mixed, and then 8 times their total mass of pure water is added and soaked for 40-50 min. After soaking, the mixture is heated to boiling and refluxed for 60-70 min. While still hot, the mixture is filtered through a 200-mesh sieve, and the filtrate and residue are collected. 6 times the mass of pure water is added to the residue, and the mixture is heated and refluxed again for 60-70 min. The mixture is then filtered, and the second filtrate is collected. The two filtrates are combined and concentrated under reduced pressure at 60-70℃ to obtain a bitter ginseng and astragalus compound extract solution.

[0010] In the application described in this invention, the drug is used to upregulate the expression level of APOA4, exerting one or more of the following effects:

[0011] (a) Remodeling lipid molecule metabolism; (b) Inhibits abnormal lipid deposition; (c) Inhibit the expression of inflammatory factors; (d) Improve the lipid metabolism-inflammation coupling pathway; (e) Inhibit the process of chronic inflammation transforming into tumors.

[0012] In the applications described in this invention, the drug can be used for early intervention of the disease, delaying disease progression, preventing disease recurrence, postoperative adjuvant therapy, and combined treatment with other anti-tumor or anti-inflammatory drugs.

[0013] In the application described in this invention, the drug is used to prevent and / or treat diseases associated with decreased APOA4 expression levels.

[0014] Furthermore, the diseases associated with decreased APOA4 expression levels are liver-related diseases.

[0015] Furthermore, the liver-related diseases are one or more of the following: chronic hepatitis, liver fibrosis, cirrhosis, hepatocellular carcinoma, non-alcoholic fatty liver disease (NAFLD), and non-alcoholic steatohepatitis (NASH).

[0016] Furthermore, the drug can upregulate the expression level of APOA4 in liver tissue in a dose-dependent manner at multiple stages of liver disease, including hepatitis, cirrhosis, early-stage hepatocellular carcinoma, and late-stage hepatocellular carcinoma.

[0017] The regulatory effect of the "Sophora flavescens-Astragalus membranaceus" drug pair on APOA4 expression described in this invention has been verified in the diethylnitrosamine (DEN)-induced liver "inflammation-cancer" transformation model in SD rats and the subcutaneous tumor-bearing model in H22 mice.

[0018] The beneficial effects of this invention are: The compound extract of Sophora flavescens and Astragalus membranaceus described in this invention enhances lipid metabolism regulation by dose-dependently upregulating the expression or activity of APOA4, thereby achieving effective intervention in liver inflammation and cancer progression-related diseases. This drug can improve abnormal liver lipid metabolism, reduce the abnormal accumulation of lipid molecules such as triglycerides in tissues, inhibit the pathological process of chronic inflammation progression and its transformation into tumors, and has an inhibitory effect on the occurrence and development of liver tumors. The intervention scheme provided by this invention has clear targeting and dose dependence, which can be used to elucidate the function and mechanism of APOA4 in the process of liver inflammation and tumor development, and provide new experimental evidence and clinical application ideas for targeted intervention strategies for liver inflammation and cancer progression-related diseases based on APOA4. Attached Figure Description

[0019] Figure 1 This refers to the rat animal model establishment and drug administration regimen in Example 1.

[0020] Figure 2 The image shows the H&E staining results of rat liver tissue in Example 1.

[0021] Figure 3 This is a diagram showing the results of lipidomics analysis of rat liver tissue in Example 1.

[0022] Figure 4 The results are from the proteomics analysis of rat liver tissue in Example 1.

[0023] Figure 5 A represents the Western blot analysis results of rat liver tissue in Example 1; Figure 5 B is a bar chart showing the grayscale value analysis of rat liver tissue protein blots.

[0024] Figure 6 This refers to the establishment of the mouse animal model and the drug administration regimen in Example 2.

[0025] Figure 7 The image shows the H&E staining results of mouse tumor tissue in Example 2.

[0026] Figure 8 This is a graph showing the results of lipidomics analysis of mouse tumor tissue in Example 2.

[0027] Figure 9 Figure A shows the results of Western blot analysis of mouse tumor tissue in Example 2; Figure 9 B is a bar chart showing the grayscale value analysis of protein blots from mouse tumor tissue. Detailed Implementation

[0028] The following non-limiting embodiments are intended to enable those skilled in the art to more fully understand the invention, but do not limit the invention in any way.

[0029] Unless otherwise specified, the experimental methods described in the following examples are conventional methods; the reagents and materials described are commercially available unless otherwise specified.

[0030] Example 1 Validation of the regulatory effect of the "Sophora flavescens-Astragalus membranaceus" herb pair on APOA4 expression in a diethylnitrosamine (DEN)-induced liver "inflammation-cancer" transformation model in SD rats. (1) Establishment of animal models Healthy male Wistar rats (200±20 g) were acclimatized for 7 days under controlled conditions (humidity 40%-60%), with free access to drinking water and standard feed. The rats were randomly divided into five groups: a control group, a model group, and low, medium, and high dose "Kushen-Huangqi" treatment groups (2.5, 5.0, and 10.0 g·kg). -1 The model group and the "Sophora flavescens-Astragalus membranaceus" group received weekly intraperitoneal injections of 10% diethylnitrosamine (DEN, 70 mg·kg⁻¹). -1 The control group received an equal volume of physiological saline. From week 6, the "Sophora flavescens-Astragalus membranaceus" group received the medication daily via gavage. At week 13, the model group was further divided into a model group and a fluorouracil-positive control group (20 mg / kg). -1(Once a week). At weeks 8, 12, 16, and 20, liver tissue samples were collected from 6 rats in each group for subsequent analysis. The rat animal model establishment and drug administration regimen are as follows: Figure 1 As shown.

[0031] (2) Preparation of "Sophora flavescens-Astragalus membranaceus" gavage solution (Sophora flavescens and Astragalus membranaceus compound extract solution) Weigh 30 g of Sophora flavescens and 10 g of Astragalus membranaceus, mix them separately, and add 8 times their total weight of water. Soak for 40 min. After soaking, heat to boiling and reflux for 60 min. Filter the hot extract through a 200-mesh sieve and collect the filtrate. Add 6 times the weight of the residue of pure water to the residue, heat again and reflux for 60 min, filter, and collect the second filtrate. Combine the two filtrates and concentrate under reduced pressure at 65℃ to a concentration of 1.0 g·mL⁻¹. -1 The resulting extract was stored at 4°C for later use.

[0032] (3) H&E staining Fresh liver tissue was removed, fixed with 4% paraformaldehyde, embedded in paraffin, and sectioned. Sections were dewaxed in xylene and rehydrated with a series of ethanol solutions. Cell nuclei were stained with hematoxylin, then differentiated in acidic alcohol, and resulcinated under running water. Cytoplasm was counterstained with eosin. Sections were then dehydrated with a series of ethanol solutions, cleared with xylene, and mounted with neutral resin. Histological images were observed and photographed using an optical microscope. The H&E staining results of rat liver tissue are shown below. Figure 2 As shown, liver tissue at all stages of HCC progression showed significant improvement after intervention with "Sophora flavescens-Astragalus membranaceus". From... Figure 2HE staining results showed that the liver tissue of rats in each group exhibited typical pathological evolution at different time points. In the inflammatory phase at week 8, light microscopy revealed hepatocyte swelling, diffuse edema, and extensive inflammatory cell infiltration in the model group rats; treatment with the "Kushen-Huangqi" herbal pair improved the degree of liver damage to varying degrees. In the cirrhosis phase at week 12, the model group showed hepatocyte swelling, lobular destruction, pseudolobule formation, and mild interlobular connective tissue hyperplasia; treatment with the "Kushen-Huangqi" herbal pair significantly improved the degree of hepatocyte damage. In the carcinogenesis phase at week 16, the model group showed disordered liver plate structure, extensive chromatin shrunkenness and necrosis in hepatocytes, with some areas showing vacuolation; carcinogenic hepatocytes varied in size and showed a small number of glandular changes; treatment with the "Kushen-Huangqi" herbal pair and other drugs reduced the number of liver cancer nodules to varying degrees. At week 20, in the late stage of hepatocellular carcinoma, both gross examination and HE staining showed further deterioration of liver tissue, manifested as significant glandular structures and varying degrees of vacuolar degeneration. Treatment with the "Kushen-Huangqi" herbal pair and positive control drugs resulted in slight symptom improvement. In summary, HE staining results visually demonstrated the progressive disease process from the inflammatory stage to the late stage of hepatocellular carcinoma, and confirmed that the "Kushen-Huangqi" herbal pair could improve organic liver damage to varying degrees at different stages of progression.

[0033] (4) Lipidomics analysis Liver tissue samples (80-100 mg) were collected from rats in different experimental groups at the stages of hepatitis, cirrhosis, early HCC, and late HCC. After rinsing with physiological saline and weighing, physiological saline was added at 10 times the weight. The tissue was then cut into small pieces and homogenized using an ultrasonic cell disruptor at 4°C for 1 min (150 W, sonication on for 2 s, off for 2 s). The homogenate was centrifuged at 4°C and 12000 rpm for 10 min, and the supernatant was collected as the tissue homogenate sample. 200 μL of the liver tissue homogenate was placed in a centrifuge tube, 40 μL of methanol and 20 μL of internal standard were added, and the mixture was vortexed for 30 s. A methyl tert-butyl ether-methanol mixture (5:1.5) was then added. v / v 1.3 mL of the mixture was vortexed for 3 min, sonicated for 5 min, and 290 μL of water was added. The mixture was centrifuged at 4°C (12000 r / min) for 5 min. The organic phase was transferred to another centrifuge tube and dried under nitrogen. The residue was reconstituted with 200 μL of methanol, vortexed for 3 min, sonicated for 5 min, and centrifuged at 12000 r / min for 5 min. 5 μL of the supernatant was used for HPLC-MS / MS analysis. After administration of "Kushen-Huangqi" (a traditional Chinese medicine formula), a heatmap was used to show the distribution patterns of lipid types in different groups at each disease stage. The results are shown in [the original text is missing]. Figure 3 It can be seen that at each stage of HCC development, most lipids show a trend of decreasing in the disease group and increasing in the treatment group.

[0034] (5) Protein analysis Liver tissue samples (80-100 mg) from rats in different experimental groups at the stages of hepatitis, cirrhosis, early HCC, and late HCC were collected. After rinsing with physiological saline and weighing, physiological saline was added at 10 times the weight. The tissue was then cut into small pieces and homogenized using an ultrasonic cell disruptor at 4°C for 1 min (150 W, sonication on for 2 s, off for 2 s). The homogenate was centrifuged at 4°C and 12000 rpm for 10 min, and the supernatant was collected as the tissue homogenate sample. The protein concentration of the homogenate was determined using a BCA kit. 200 μg of protein was placed in a 10 kDa centrifuge tube for pretreatment. 100 μL of 8 M urea and 2 μL of 1 M DTT were added to the protein solution, and the mixture was treated at 55°C for 1 h. Subsequently, 5 μL of 1 M IAM was added, and the mixture was reacted at 25°C in the dark for 30 min. After centrifugation, the precipitate was resuspended in 100 μL of 50 mM ammonium bicarbonate solution, and 4 μg of trypsin was added. Digestion was carried out overnight at 37°C, and finally, 2 μL of formic acid was added to terminate the digestion. The solution was then centrifuged at 25°C and 12000 rpm for 5 min, and 5 μL of the supernatant was used for NanoLC-MS / MS analysis. The expression levels of APOA4 protein at different stages of HCC development, as detected by NanoLC-MS, are shown below. Figure 4 As shown in the figure, APOA4 expression levels decreased during the hepatitis, cirrhosis, early HCC, and late HCC stages, and were significantly lower than those in the healthy control group. After intervention with the "Kushen-Huangqi" drug pair, APOA4 levels rebounded in a dose-dependent manner, with a better improvement effect than the positive control group treated with fluorouracil. Therefore, APOA4 was downregulated at all stages of HCC progression, and a dose-dependent rebound occurred after administration of "Kushen-Huangqi".

[0035] (6) Protein immunoassay Liver tissue proteins were extracted using RIPA lysis buffer at 4°C, with PMSF added to inhibit protease activity. Equal volumes of protein were separated by SDS-PAGE and transferred to a polyvinylidene fluoride (PVDF) membrane. The membrane was blocked at room temperature for 15 min and then incubated overnight at 4°C with the corresponding primary antibodies (APOA4 and GAPDH). After washing, the membrane was incubated with secondary antibodies for 1 h in the dark. Protein bands were developed and quantified using ImageJ. The expression levels of APOA4 protein at different stages of HCC development, based on Western blotting, are shown below. Figure 5As shown in the figure, APOA4 expression levels decreased during the hepatitis, cirrhosis, early HCC, and late HCC stages, and were significantly lower than those in the healthy control group. After intervention with the "Sophora flavescens-Astragalus membranaceus" herb pair, APOA4 levels rebounded in a dose-dependent manner, with a better improvement effect than the positive control group treated with fluorouracil. These results are consistent with those obtained using NanoLC-MS.

[0036] (7) Data processing Data analysis was performed using IBM SPSS Statistics 23.0 and SIMCA 14.1 software. Measurement data are expressed as mean ± standard deviation (mean ± SD). The processed data were then analyzed. t Test and orthogonal partial least squares discriminant analysis (OPLS-DA), when p A difference is considered statistically significant when the VIP value is less than 0.05 and VIP > 1.

[0037] Example 2 Validation of the regulatory effect of the "Sophora flavescens-Astragalus membranaceus" herb pair on APOA4 expression in a subcutaneous H22 tumor-bearing model (1) Establishment of animal models Healthy male Balb / c mice (20±2 g) were acclimatized for 7 days under controlled conditions (humidity 40%-60%), with free access to drinking water and standard feed. The subcutaneous H22 tumor-bearing model and administration regimen were as follows: Resuscitated H22 cells were injected intraperitoneally into Balb / c mice and passaged. Ascites fluid was collected and counted after 5-7 days. Approximately 1×10⁻⁶ cells were collected. 6 One surviving cell was inoculated subcutaneously in the axilla of a mouse. The tumor volume reached approximately 100 mm. 3 The administration began at a specific time. Mice were randomly divided into five groups: a model group, and low, medium, and high dose "Kushen-Huangqi" treatment groups (5.0, 10, and 20 g·kg⁻¹). -1 ·d -1 ), and the fluorouracil-positive control group (40 mg·kg) -1 (Administered every other day). After two weeks of continuous administration, plasma and tumor tissue samples were collected for subsequent analysis. The establishment of the mouse animal model and the administration regimen are as follows: Figure 6 As shown.

[0038] (2) Preparation of "Sophora flavescens-Astragalus membranaceus" gavage solution (Sophora flavescens and Astragalus membranaceus compound extract solution) Weigh 30 g of Sophora flavescens and 10 g of Astragalus membranaceus, mix them separately, and add 8 times their total weight of water. Soak for 40 min. After soaking, heat to boiling and reflux for 60 min. Filter the hot extract through a 200-mesh sieve and collect the filtrate. Add 6 times the weight of the residue of the extract to the residue, heat again and reflux for 60 min, filter, and collect the second filtrate. Combine the two filtrates and concentrate under reduced pressure at 65℃ to a concentration of 1.0 g·mL⁻¹. -1 The resulting extract was stored at 4°C for later use.

[0039] (3) H&E staining Fresh tumor tissue was removed, fixed with 4% paraformaldehyde, embedded in paraffin, and sectioned. Sections were dewaxed in xylene and rehydrated with a series of ethanol solutions. Cell nuclei were stained with hematoxylin, then differentiated in acidic alcohol, and restained with sappanhydride under running water. Cytoplasm was counterstained with eosin. Sections were then dehydrated with a series of ethanol solutions, cleared with xylene, and mounted with neutral resin. Histological images were observed and photographed using an optical microscope. The H&E staining results of mouse tumor tissue are shown below. Figure 7 As shown, the tumor cells in the model group were densely packed, with large and regular nuclei and sparse stroma, exhibiting a typical tumor tissue morphology and structure. In contrast, after treatment with SF-AM and 5-FU, the tumor tissue cells were disordered, with decreased cell density, pyknosis or fragmentation of nuclei, destruction of cytoplasmic structure, and obvious necrotic and vacuolated areas.

[0040] (4) Lipidomics analysis Tumor tissue samples (80-100 mg) from mice in different experimental groups were collected. After rinsing with physiological saline and weighing, physiological saline was added at 10 times the weight. The tissue was then cut into small pieces and homogenized using an ultrasonic cell disruptor at 4°C for 1 min (150 W, sonication on for 2 s, off for 2 s). The homogenate was centrifuged at 4°C and 12000 rpm for 10 min, and the supernatant was collected as the tissue homogenate sample. 200 μL of the above tumor tissue homogenate was placed in a centrifuge tube, 40 μL of methanol and 20 μL of internal standard were added, and the mixture was vortexed for 30 s. A methyl tert-butyl ether-methanol mixture (5:1.5) was then added. v / v 1.3 mL of the organic phase was vortexed for 3 min, sonicated for 5 min, and 290 μL of water was added. The mixture was centrifuged at 4°C (12000 r / min) for 5 min. The organic phase was transferred to another centrifuge tube and dried under nitrogen. The residue was reconstituted with 200 μL of methanol, vortexed for 3 min, sonicated for 5 min, and centrifuged at 12000 r / min for 5 min. 5 μL of the supernatant was used for HPLC-MS / MS analysis. The results of lipidomics analysis of mouse tumor tissue are as follows: Figure 8As shown in the figure, in the Balb / c mouse tumor model of H22 cells, the changes in various lipids in tumor tissue after administration of "Kushen-Huangqi" are as follows: most lipids in tumor tissue showed a downregulation trend, with triglycerides (TGs) being the most frequently downregulated, totaling 120 types. The reason may be that tumor cells often abnormally activate lipid synthesis pathways to meet the needs of rapid proliferation. The "Kushen-Huangqi" drug pair may block the de novo synthesis pathway of triglycerides by downregulating the expression or activity of key enzymes such as fatty acid synthase (FASN) and acetyl-CoA carboxylase (ACC); simultaneously, this drug pair may promote lipid catabolism, activating fatty triglyceride lipase (ATGL) or hormone-sensitive lipase (HSL), accelerating the breakdown of triglycerides into free fatty acids for energy. In summary, the broad downregulation effect of this drug pair on triglycerides reflects its unique mechanism in intervening in tumor lipid metabolism remodeling.

[0041] (5) Protein immunoassay Tumor tissue proteins were extracted using RIPA lysis buffer at 4°C, with PMSF added to inhibit protease activity. Equal volumes of proteins were separated by SDS-PAGE and transferred to a polyvinylidene fluoride (PVDF) membrane. The membrane was blocked at room temperature for 15 min and then incubated overnight at 4°C with the corresponding primary antibodies (APOA4 and GAPDH). After washing, the membrane was incubated with secondary antibodies for 1 h in the dark. Protein bands were developed and quantified using ImageJ. The expression levels of APOA4 protein in H22 tumor-bearing mouse tumor tissues, based on Western blotting, are shown below. Figure 9 As shown in the figure. The results showed that after administration of "Kushen-Huangqi", the expression level of APOA4 was upregulated in a dose-dependent manner, consistent with the effect of administration in the rat HCC model.

[0042] (6) Data processing Data analysis was performed using IBM SPSS Statistics 23.0 and SIMCA 14.1 software. Measurement data are expressed as mean ± standard deviation (mean ± SD). The processed data were then analyzed. t Test and orthogonal partial least squares discriminant analysis (OPLS-DA), when p A difference is considered statistically significant when the VIP value is less than 0.05 and VIP > 1.

Claims

1. Application of the compound extract of bitter gourd and astragalus in the preparation of drugs that target and regulate APOA4.

2. The application according to claim 1, characterized in that: The bitter herbs and astragalus were combined according to the mass ratio of raw herbs, and then extracted by heating and reflux with pure water, filtered and concentrated to obtain a compound extract solution of bitter herbs and astragalus.

3. The application according to claim 2, characterized in that: Combine bitter herbs and astragalus at a ratio of 3:1 (crude drug mass). Add 8 times the total mass of pure water and soak for 40-50 minutes. After soaking, heat to boiling and reflux for 60-70 minutes. Filter the mixture through a 200-mesh sieve while hot and collect the filtrate and residue. Add 6 times the mass of pure water to the residue and heat and reflux again for 60-70 minutes. Filter and collect the second filtrate. The filtrates obtained from the two extractions were combined and concentrated under reduced pressure at 60℃~70℃ to obtain a compound extract solution of Sophora flavescens and Astragalus membranaceus.

4. The application according to claim 1, characterized in that: The drug is used to upregulate APOA4 expression levels, exerting one or more of the following effects: (a) Remodeling lipid molecule metabolism; (b) Inhibits abnormal lipid deposition; (c) Inhibit the expression of inflammatory factors; (d) Improve the lipid metabolism-inflammation coupling pathway; (e) Inhibit the process of chronic inflammation transforming into tumors.

5. The application according to claim 1, characterized in that: The drug is used to prevent and / or treat diseases associated with decreased APOA4 expression levels.

6. The application according to claim 5, characterized in that: The diseases associated with decreased APOA4 expression levels are liver-related diseases.

7. The application according to claim 6, characterized in that: The liver-related diseases mentioned are one or more of the following: chronic hepatitis, liver fibrosis, cirrhosis, hepatocellular carcinoma, non-alcoholic fatty liver disease (NAFLD), and non-alcoholic steatohepatitis (NASH).

8. The application according to claim 7, characterized in that: The drug can upregulate the expression level of APOA4 in liver tissue in a dose-dependent manner at multiple stages of liver disease, including hepatitis, cirrhosis, early hepatocellular carcinoma, and late hepatocellular carcinoma.

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