Anti-hepatic fibrosis small molecule compound and application thereof

Abstract

The invention discloses an anti-hepatic fibrosis small molecule compound and application, the anti-hepatic fibrosis small molecule compound is composed of 3 beta-ethylphthalein and 6-hydroxykaempferol, and the anti-hepatic fibrosis small molecule compound is used as an anti-hepatic fibrosis drug after splenectomy. The traditional Chinese medicine composition can improve the liver fibrosis degree (relieving liver inflammation, improving liver zymogram, reducing bilirubin and reducing collagen deposition) after splenectomy, and the liver fibrosis is caused by at least one of alcoholic hepatitis, fatty liver, viral hepatitis, drug-induced hepatitis, autoimmune hepatitis and the like.

Description

Technical Field

[0001] The present invention relates to the field of biomedicine technology, and in particular to a small molecule compound for resisting liver fibrosis and its application. Background Art

[0002] Chronic liver disease is an important global health problem, and cirrhosis is the leading cause of death from chronic liver disease worldwide, accounting for 2.2% of the global death toll in 2016, and is also the 11th leading cause of death and the 15th leading cause of morbidity in the world; cirrhosis is the terminal stage of liver fibrosis progression, which seriously threatens human health, and there is no satisfactory treatment so far. Cirrhosis and its secondary hypersplenism promote and influence each other; the degree of liver fibrosis also increases with the increase in the degree of hypersplenism. Splenectomy is the main treatment method; however, patients often experience symptoms of spleen deficiency and blood stasis such as jaundice, fatigue, and varicose veins after surgery, leading to liver damage and worsening of liver fibrosis. Cirrhosis is the terminal stage of liver fibrosis progression, and there is no ideal treatment so far. Splenectomy is the main treatment method for hypersplenism in cirrhosis, and is widely used in the clinical treatment of patients with hypersplenism in cirrhosis; however, patients often experience symptoms of spleen deficiency and blood stasis such as jaundice, fatigue, and varicose veins after surgery, leading to liver damage and worsening of liver fibrosis. At present, modern medicine is still in the exploratory stage for chemical drugs to combat liver fibrosis after splenectomy, and there are no anti-liver fibrosis drugs for patients after splenectomy in clinical practice.

[0003] As an important measure for the clinical treatment of liver cirrhosis, traditional Chinese medicine against liver fibrosis has shown unique advantages and potential. The Golden Chamber states: "Those who treat diseases before they occur should see liver diseases, know that liver transmits to spleen, and should strengthen spleen first", which laid the foundation for the concept of "treating diseases before they occur" in later generations. The concept of "treating diseases before they occur" is an important principle of traditional Chinese medicine treatment. The Golden Chamber starts with "strengthening spleen" to prevent and treat "liver diseases". Patients with splenectomy for liver cirrhosis often present with symptoms of spleen deficiency, such as fatigue, abdominal distension, poor appetite, fatigue, plump and tender tongue, red tongue, thin white tongue coating, and thin or weak pulse; at the same time, patients have dark tongue, petechiae or ecchymosis, varicose sublingual collaterals, weak and large pulse, late or astringent pulse, which are symptoms of "blood stasis", indicating that patients with splenectomy for liver cirrhosis often present with the symptoms of spleen deficiency and blood stasis, so the clinical treatment method of invigorating qi and strengthening spleen, and promoting blood circulation and removing blood stasis is adopted, and Atractylodes macrocephala and safflower are representative prescriptions for invigorating qi and strengthening spleen, and promoting blood circulation and removing blood stasis.

[0004] 3β-Acetoxyatractylone is derived from Atractylodes macrocephala, a plant of the Asteraceae family. English: 3β-Acetoxyatractylone, CAS 61206-10-8, molecular weight 274.355, C 17 H 22 O 3 , the structural formula is Figure 1 shown.

[0005] 6-Hydroxykaempferol, derived from safflower of Asteraceae, safflower genus, English: 6-Hydroxykaempferol, CAS4324-55-4, molecular weight 302.25, C 15 H 10 O 7 The structural formula is Figure 2 shown.

[0006] The above two substances have not been used to treat liver fibrosis. Using them to treat liver fibrosis after splenectomy can provide new ideas and directions for the development and application of drugs for liver fibrosis. Summary of the invention

[0007] The purpose of the present invention is to overcome the defects and deficiencies of the prior art, provide a small molecule compound for resisting liver fibrosis and its application, and solve various problems existing in the prior art.

[0008] To achieve the above object, the present invention provides the following technical solutions:

[0009] The purpose of the present invention is to provide a significant targeted therapeutic drug for the treatment of liver fibrosis after splenectomy, to provide new ideas and directions for the development and application of drugs for liver fibrosis, and to use small molecule compounds to target the AKT / NF-κB pathway, inhibit liver function, and reduce the degree of liver fibrosis.

[0010] The present invention provides a small molecule compound for resisting liver fibrosis, and its purpose can be achieved by the following technical scheme:

[0011] A small molecule compound for resisting liver fibrosis is composed of 3β-acetyloxy atractylodesone and 6-hydroxykaempferol.

[0012] The small molecule compound for resisting liver fibrosis is composed of 3β-acetyloxy atractylodesone and 6-hydroxykaempferol in a weight ratio of 1:0.8-1.2.

[0013] The application of the small molecule compound for resisting liver fibrosis can be used as an anti-liver fibrosis drug for patients after splenectomy.

[0014] The application of the small molecule compound for resisting liver fibrosis in resisting liver fibrosis, and the dosage form of the anti-liver fibrosis drug is an oral preparation.

[0015] The application of the small molecule compound for resisting liver fibrosis in resisting liver fibrosis requires the addition of a pharmaceutically acceptable carrier to the small molecule compound.

[0016] The application of the small molecule compound for preventing liver fibrosis in preventing liver fibrosis requires the addition of a pharmaceutically acceptable carrier including saline, buffer, glucose, water and a combination thereof to the small molecule compound.

[0017] The application of the small molecule compound for anti-liver fibrosis in anti-liver fibrosis generally requires that the pharmaceutical preparation should match the administration method, and the pharmaceutical composition should be manufactured under sterile conditions; the oral dosage form of the small molecule compound includes one of tablets, capsules, granules, syrups, solutions, and suspensions.

[0018] The application of the small molecule compound for resisting liver fibrosis in resisting liver fibrosis comprises the following medicine dosages: 10-40 mg / kg of 3β-acetyloxy atractylodesone and 10-40 mg / kg of 6-hydroxykaempferol.

[0019] The beneficial effects of the present invention are:

[0020] (1) The present invention can improve the degree of liver fibrosis after splenectomy (reducing liver inflammation, improving liver enzyme spectrum, reducing bilirubin, and reducing collagen deposition). The causes of liver fibrosis include at least one of alcoholic hepatitis, fatty liver, viral hepatitis, drug-induced hepatitis, autoimmune hepatitis, etc.

[0021] (2) The drug is easy to use, has a low preparation process cost, and is easy to promote. BRIEF DESCRIPTION OF THE DRAWINGS

[0022] Figure 1 It is the chemical formula of 3β-acetyloxy atractylodesinone;

[0023] Figure 2 is the chemical formula of 6-hydroxykaempferol;

[0024] Figure 3 Drug active ingredient-disease-target network diagram;

[0025] Figure 4 Gene ontology GO enrichment histogram of intersection targets;

[0026] Figure 5 It is the Kyoto Encyclopedia of Genes and Genomes KEGG enrichment bubble map;

[0027] Figure 6 Schematic diagram of molecular docking of 3β-Acetoxyatractylone and CHRM1 in the simulated molecular docking of drug active ingredients and key proteins of AKT / NF-κB pathway;

[0028] Figure 7 Schematic diagram of molecular docking of 3β-Acetoxyatractylone and CHRM2 in the simulated molecular docking of active pharmaceutical ingredients and key proteins of AKT / NF-κB pathway;

[0029] Figure 8Molecular docking of 3β-Acetoxyatractylone and PTGS2 in the simulated molecular docking of drug active ingredients and key proteins in the AKT / NF-κB pathway;

[0030] Fig. 9 Molecular docking of 3β-Acetoxyatractylone and RXRA in the simulated molecular docking of drug active ingredients and key proteins of AKT / NF-κB pathway;

[0031] Fig.10 Molecular docking of 6-Hydroxykaempferol and HSP90AB1 in the simulated molecular docking of drug active ingredients and key proteins of AKT / NF-κB pathway;

[0032] Fig.11 Molecular docking of 6-Hydroxykaempferol and PTGS2 in the simulated molecular docking of drug active ingredients and key proteins of AKT / NF-κB pathway;

[0033] Fig.12 This is a graph showing the improvement of liver function indicators in cirrhotic rabbits by compounds;

[0034] Fig.13 This is a picture of the pathological changes of liver tissue in the control group observed under an optical microscope;

[0035] Fig.14 This is a picture of the pathological changes of liver cirrhosis model tissue observed under an optical microscope;

[0036] Fig.15 This is a picture of the pathological changes of liver tissue in the splenectomy group of liver cirrhosis observed under an optical microscope;

[0037] Fig.16 This is a picture of the pathological changes of liver tissue in the low-dose drug group observed under an optical microscope;

[0038] Fig.17 This is a picture of the pathological changes in liver tissue of the medium-dose group observed under an optical microscope;

[0039] Fig.18 This is a picture of the pathological changes in liver tissue in the high-dose drug group observed under an optical microscope;

[0040] Fig.19 Western Blot method was used to detect the expression levels of Col-I, Col-III, and α-SMA in the three groups;

[0041] Fig. 20 Histogram of the relative expression levels of Col-I, Col-III, and α-SMA proteins in the three groups of LX-2 cells detected by Western Blot. DETAILED DESCRIPTION

[0042] The following will be combined with the drawings in the embodiments of the present invention to clearly and completely describe the technical solutions in the embodiments of the present invention. Obviously, the described embodiments are only part of the embodiments of the present invention, not all of the embodiments. Based on the embodiments of the present invention, all other embodiments obtained by ordinary technicians in this field without creative work are within the scope of protection of the present invention.

[0043] Embodiment 1,

[0044] A small molecule compound for resisting liver fibrosis, comprising 3β-acetyloxy atractylone and 6-hydroxykaempferol in a weight ratio of 1:1; wherein the chemical formula of 3β-acetyloxy atractylone is as follows Figure 1 As shown, the chemical formula of 6-hydroxykaempferol is as follows Figure 2 shown.

[0045] Its preparation method is:

[0046] Weigh 30 mg of 3β-acetyloxyatractyl ketone and 30 mg of 6-hydroxykaempferol in a 1:1 weight ratio, and dissolve them in 10 ml of normal saline to prepare a small molecule drug combination solution.

[0047] Embodiment 2,

[0048] A small molecule compound for resisting liver fibrosis is composed of 3β-acetyloxy atractylodesone and 6-hydroxykaempferol in a weight ratio of 1:1.2.

[0049] Weigh 30 mg of 3β-acetyloxyatractyl ketone, weigh 30 mg of 6-hydroxykaempferol according to a weight ratio of 1:1.2, and weigh 36 mg of 6-hydroxykaempferol according to a weight ratio of 1:1.2, and add them into 10 ml of normal saline to prepare a small molecule drug combination solution.

[0050] Embodiment 3,

[0051] A small molecule compound for resisting liver fibrosis is composed of 3β-acetyloxy atractylodesone and 6-hydroxykaempferol in a weight ratio of 1:0.8.

[0052] Weigh 30 mg of 3β-acetyloxyatractyl ketone, weigh 24 mg of 6-hydroxykaempferol in a weight ratio of 1:0.8, and dissolve them in 10 ml of normal saline to prepare a small molecule drug combination solution.

[0053] The following experiment proves the effect of the drug in Example 1 of the present application

[0054] Experiment 1:

[0055] The physicochemical structures and targets of 3β-acetyloxyatractyl ketone and 6-hydroxykaempferol were obtained by using the Traditional Chinese Medicine Systems Pharmacology Database and Analysis Platform (http: / / tcmspw.com / tcmsp.php, TCMSP).

[0056] The disease name was corrected on the NCBI website with "Liver Fibrosis", and the obtained name was used as a keyword to search for disease targets in the Drug Target Database (TTD) (http: / / db.idrblab.net / ttd / ), the Online Mendelian Inheritance in Man (OMIM) database (https: / / omim.org / ), the GeneCards database (https: / / www.genecards.org / ), the PharmGKB database (https: / / www.pharmgkb.org / ), and the DrugBank database (https: / / go.drugbank.com / ), and the relevant literature was consulted to supplement the targets. After all targets were deduplicated, the gene ID was corrected through the Uniprot database (https: / / www.uniprot.org / ) to obtain the therapeutic targets corresponding to liver fibrosis diseases (Table 1).

[0057] The common targets of 3β-acetyloxyatractyl ketone and 6-hydroxykaempferol and liver fibrosis were observed, and the drug active ingredient-disease-target network diagram was constructed using Cytoscape3.8.0 software (e.g. Figure 3 ).

[0058] The main targets of AKT / NF-κB pathway acted by 3β-acetyloxyatractylone and 6-hydroxykaempferol were observed, and the gene ontology (GO) and Kyoto Encyclopedia of Genes and Genomes (KEGG) enrichment of the common targets were observed (e.g. Figure 4 , Figure 5 ).

[0059] The AutoDockVina software was used to perform molecular docking of 3β-acetyloxy-atractyl-1-one and 6-hydroxy-kaempferol on CHRM1, HSP90AB1PTGS2, and RXRA targets to verify their mutual binding activity. First, the 2D structure file of the small molecule ligand was downloaded from the PubChem website and converted into a 3D structure. The pdb format file of the core target protein was downloaded from the PDB website (http: / / www.rcsb.org / ). The corresponding ligands and receptors were processed and operated routinely using AutoDockTools. AutoDockVina was used for molecular docking, and the receptor and ligand were docked with the minimum binding energy (at least less than -5kcal / mol) to obtain a molecular docking diagram (as shown in Figure 1). Figure 6-Figure 11) and the corresponding ligand-receptor binding energy (Table 1).

[0060]

[0061] Figure 6-Figure 11 This is a simulated molecular docking diagram of the active ingredients of the drug and the key proteins of the AKT / NF-κB pathway.

[0062] Experiment 2

[0063] 80 New Zealand white rabbits aged 3 months, weighing 2.5-3.0 kg, aged 90-120 days, 40 males and 40 females were selected. The animals were raised in stacked stainless steel wire cages, fed with standard granular feed, and had free access to tap water. The rabbit room was well ventilated and kept clean, with an air humidity of 50%-70% and a temperature of 20℃-29℃. After acclimatization for 1 week, blood was drawn from the central ear artery after fasting for 12 hours to check liver function. Color Doppler examination showed no lesions in the liver and spleen. Those with normal indicators in each group were included in the study.

[0064] The cirrhosis model was prepared by the thioacetamide (TAA) method. TAA was added to normal saline to prepare a 5% solution, which was injected intraperitoneally at a dose of 30 mg / kg, twice a week, for 16 weeks. Dose adjustment plan: ① For those whose body weight gain or loss is within 5% within 1 week, maintain the original dose. ② For those whose body weight drops by more than 5% within 1 week, reduce the TAA dose by 20 mg / kg. ③ For those whose body weight drops by more than 10% within 1 week, suspend the injection 1-2 times, and adjust the dose after the weight stops dropping to avoid death. ④ For those whose body weight increases by more than 5% continuously within 1 week, increase the TAA dose by 10 mg / kg.

[0065] 1. Splenectomy model establishment plan

[0066] Penicillin was injected one day before splenectomy, and the rabbit was fasted for 24 hours before the operation (drinking water was allowed). 20% urethane solution was injected intraperitoneally at a dose of 5 ml / kg for anesthesia. The rabbit was fixed on a fixed frame with four limbs tied to the four corners of the fixed frame with thin ropes. After the vital signs were stable, the rabbit hair on the abdomen was shaved with pet electric clippers, and the skin was disinfected with complex iodine and alcohol. A midline incision of about 6.0 cm was made under the xiphoid process. The skin was cut layer by layer to separate the subcutaneous tissue to the abdominal cavity. Sterile gauze was used to protect the gastrointestinal tract and pulled open to fully expose the field of vision. After finding the spleen, the splenic artery and vein were ligated, the spleen was removed, and the abdomen was closed layer by layer after careful hemostasis. The operation was completed. The dressing was changed once a day after the operation, and penicillin was injected intramuscularly at a dose of 40,000 units / kg / time, twice a day, for 5 consecutive days.

[0067] 2 Grouping scheme

[0068] Eighty New Zealand white rabbits were randomly divided into four groups using random codes generated by software, with 20 rabbits in each group: (1) blank control group; (2) liver cirrhosis group; (3) liver cirrhosis splenectomy group; (4) compound group (three subgroups): ① low-dose compound group; ② medium-dose compound group; ③ high-dose compound group.

[0069] 4 Intervention plans for each group

[0070] 4.1 Preparation of Chinese medicine and determination of its infusion dosage

[0071] 4.2 Intervention plan

[0072] (1) Blank control group: The mice were not given any treatment, and were allowed to drink water and eat freely. They were not given any saline or Chinese medicine.

[0073] (2) Cirrhosis group: Normal New Zealand white rabbits were given TAA intraperitoneal injection to establish a cirrhosis with hypersplenism model. The model was established after 16 weeks. The rabbits were free to move around in the cage and had free access to water and food. They were given normal saline by gavage at 10 g / (kg.d) twice a day, at 9:00 and 15:00, for 21 consecutive days.

[0074] (3) Cirrhosis and splenectomy group: Normal New Zealand white rabbits were given TAA intraperitoneal injection to establish a cirrhosis and hypersplenism model. Splenectomy was performed 16 weeks after model establishment. The rabbits were free to move around in the cage after awakening and fasted on the day of surgery. From the first day after surgery, the rabbits were allowed to drink water and eat freely. On the second day after surgery, they were given normal saline by gavage at a rate of 10 g / (kg.d) twice a day, at 9:00 and 15:00, for 21 consecutive days.

[0075] (4) Compound group

[0076] ① Low-dose group: normal New Zealand white rabbits, TAA intraperitoneal injection to prepare liver cirrhosis with hypersplenism model, splenectomy after 16 weeks of modeling, free movement in the cage after awakening, fasting on the day of surgery, free drinking water and food from the first day after surgery, the second day after surgery to give the agent prepared in Example 1, 3β-acetyloxy atractylone 10 mg / kg and 6-hydroxykaempferol 10 mg / kg gavage, twice a day, respectively at 9:00 and 15:00, for 21 consecutive days.

[0077] ②Medium-dose group: Same as low-dose group, except that the dose was changed to 20 mg / (kg.d).

[0078] ③ High-dose group: Same as the low-dose group, except that the dose was changed to 40 mg / (kg.d).

[0079] 5. Specimen Collection

[0080] After the treatment, the rabbits were anesthetized and killed by air embolization of the ear vein. After specimen collection, the bodies were sent to the animal center for unified incineration. Specimen collection: (1) The cleaned complete liver specimens were removed and collected and stored at -80℃ for later use; (2) 5ml of blood was drawn from the portal vein and inferior vena cava and stored at -80℃ for later use. The time point for killing and specimen collection was 21 days after treatment.

[0081] 6.1.3 Blood index examination

[0082] ① Liver function indicators (alanine aminotransferase, aspartate aminotransferase, total bilirubin, direct bilirubin, indirect bilirubin, albumin indicators): All groups adopted fasting venous blood collection, added anticoagulant, and stored at low temperature. The blood biochemical indicators were detected by fully automatic biochemical analyzer (HITACHI7600 made in Japan).

[0083] ② Detection of liver fibrosis indicators (hyaluronidase, laminin, type III procollagen, type IV collagen, glycocholic acid): All groups adopted fasting venous blood collection, added anticoagulant, and stored at low temperature. All indicators were detected by fully automatic chemiluminescence immunoassay.

[0084]

[0085]

[0086] The low-dose compound group showed improved alanine aminotransferase, indirect bilirubin, and hyaluronidase compared with the cirrhosis splenectomy group; the medium-dose compound group showed improved alanine aminotransferase, indirect bilirubin, and hyaluronidase compared with the cirrhosis splenectomy group; the high-dose compound group showed improved alanine aminotransferase, alkaline phosphatase, total bilirubin, indirect bilirubin, hyaluronidase, and type III collagen compared with the cirrhosis splenectomy group, and the results were statistically significant (P<0.05). Fig.12 )

[0087] 6.1.4 Observation of liver tissue microstructure using light and electron microscopes

[0088] Liver tissue specimens from the same part were fixed with 40 g / L formaldehyde solution for 24 h, dehydrated, transparent, embedded in paraffin and sliced. 5 μm continuous sections were transparent with xylene, dehydrated with gradient ethanol, stained with hematoxylin and eosin, sealed with neutral gum, and the pathological changes of liver tissue were observed under an optical microscope.

[0089] like Figure 13-18 As shown in the figure, the liver lobule structure of the rabbits in the control group was intact, the hepatic cords were neatly arranged, the portal area was clear, the hepatic plates were distributed in a spoke-like manner, the hepatocytes were slightly edematous, and the hepatic sinusoids were slightly dilated (red arrows) (as shown in the figure). Fig.13); The liver lobule structure of the rabbit liver tissue in the cirrhosis model group disappeared, and pseudolobules of varying sizes were formed. There was inflammatory cell infiltration and bile duct cell proliferation, showing the characteristics of cirrhosis (such as Fig.14 ); In the cirrhosis splenectomy group, the normal liver lobule structure disappeared, the hepatocytes became edematous and balloon-like, the hepatic sinusoids were dilated and congested, and the pathological damage was alleviated (such as Fig.15 ); In the low-dose group, the normal liver lobule structure disappeared, the hepatic sinusoids expanded and congested, and inflammatory cell infiltration was observed (such as Fig.16 ); The liver tissue of rabbits in the medium-dose group was edematous to balloon-like degeneration, the cytoplasm was loose and vacuolated, the hepatic sinusoids were dilated and congested, and no other obvious abnormalities were observed (such as Fig.17 ); In the high-dose group, the rabbits had mild edema of perivenous hepatocytes, slight dilatation and congestion of hepatic sinusoids, and significantly reduced pathological damage (such as Fig.18 ).

[0090] Experiment 3

[0091] Cell culture and passaging

[0092] Human hepatic stellate cell line LX-2 cells were cultured in a sterile environment of 37°C and 5% CO2 with high-glucose DMEM (complete culture medium) containing 10% FBS. When the cell density reached about 90%, the original culture medium was aspirated, and the cells were washed twice with phosphate buffer. 1 mL of trypsin was added to each 90 mm culture dish. After incubation in an incubator for 1-2 min, 2 mL of complete culture medium was added to terminate digestion. After pipetting, the cells were transferred into a 15 mL centrifuge tube and centrifuged at 5000 r / min for 5 min. The upper culture medium was aspirated, 2 mL of complete culture medium was added, the cells were gently pipetted, and 1 mL was transferred into new culture dishes after mixing. The complete culture medium in each dish was supplemented to 6 mL, and the cells were transferred into an incubator after mixing.

[0093] Preparation of small molecule drug combination solution

[0094] Weigh 3 mg of 3β-acetyloxy atractylone and 3 mg of 6-hydroxykaempferol in a 1:1 weight ratio, and dissolve in 1 ml of saline to prepare a 3 mg / mL small molecule drug combination solution. Place the prepared combination solution in a cryovial and freeze it in a -80°C refrigerator for later use.

[0095] Western Blot was used to detect the effect of Chinese herbal medicine combination on the expression levels of Col-I, Col-III and α-SMA

[0096] The cells were cultured in a 6-well plate. When the cell density reached 80%, the original culture medium was aspirated and the cells were washed twice with PBS. Only complete culture medium (high-glucose DMEM containing 10% FBS) was added to the control group, complete culture medium containing 5ng / mL TGF-β1 was added to the model group, and complete culture medium containing 5ng / mL TGF-β1 and 3mg / mL small molecule drug combination liquid powder was added to the compound group. After 48h of culture, the cells were digested, centrifuged, and the culture medium was removed. 100uLSDS lysis buffer (containing 1% PMSF) was added to each well of cells, lysed on ice for 30min, centrifuged, and the supernatant was taken for BCA protein quantitative detection. 5× loading buffer was added at a ratio of 4:1, heated at 100°C for 8 min, and 20 ug of the denatured protein sample was subjected to gel electrophoresis, transferred to the membrane, and blocked. Primary antibodies (α-smooth muscle actin, α-SMA; collagen I, Col-I; collagen III, Col-III) were added and incubated overnight at 4°C, and goat anti-rabbit IgG secondary antibodies were added and incubated at room temperature for 1.5 h. After each step, the cells were washed three times with TBST for 5 min each time, and then the bands were detected by chemiluminescence, and the grayscale values ​​of Col-I, Col-III, and α-SMA bands were analyzed using Image J. Fig.19 Western Blot was used to detect the expression levels of Col-I, Col-III, and α-SMA in the three groups; Fig. 20 Western Blot was used to detect the relative expression levels of Col-I, Col-III and α-SMA proteins in the three groups of LX-2 cells.

[0097] The above description is only a preferred embodiment of the present invention and is not intended to limit the present invention. Any modifications, equivalent substitutions, improvements, etc. made within the spirit and principles of the present invention should be included in the protection scope of the present invention.

Claims

1. A small molecule compound for resisting liver fibrosis, characterized in that: It is composed of 3β-acetyloxy atractylodesone and 6-hydroxykaempferol.

2. The small molecule compound for resisting liver fibrosis according to claim 1, characterized in that: The invention is composed of 3β-acetyloxy atractylodesone and 6-hydroxykaempferol in a weight ratio of 1:0.8-1.

2.

3. Use of the small molecule anti-liver fibrosis compound according to claim 1 in anti-liver fibrosis, characterized in that: It can be used as an anti-liver fibrosis drug after splenectomy.

4. The use of the small molecule anti-liver fibrosis compound according to claim 3 in anti-liver fibrosis, characterized in that: The dosage form of the anti-liver fibrosis drug is an oral preparation.

5. Use of the small molecule anti-liver fibrosis compound according to claim 3 in anti-liver fibrosis, characterized in that: When used, a pharmaceutically acceptable carrier needs to be added to the small molecule compound.

6. Use of the small molecule anti-liver fibrosis compound according to claim 5 in anti-liver fibrosis, characterized in that: Pharmaceutically acceptable carriers that need to be added to small molecule compounds include saline, buffer, glucose, water and combinations thereof.

7. Use of the small molecule anti-liver fibrosis compound according to claim 6 in anti-liver fibrosis, characterized in that: Generally, the pharmaceutical preparation should match the mode of administration, and the pharmaceutical composition is preferably manufactured under sterile conditions; the oral dosage form of the small molecule compound includes one of tablets, capsules, granules, syrups, solutions, and suspensions.

8. Use of the small molecule anti-liver fibrosis compound according to claim 7 in anti-liver fibrosis, characterized in that: The drug dosage is 10-40 mg / kg of 3β-acetyloxy atractylodesone and 10-40 mg / kg of 6-hydroxykaempferol.

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