A traditional Chinese medicine composition for treating warfarin-induced hemorrhagic syndrome, a preparation method and application thereof

Abstract

The application discloses a traditional Chinese medicine composition for treating warfarin-induced hemorrhagic syndrome, a preparation method and application thereof, and belongs to the field of veterinary medicine preparation. The traditional Chinese medicine composition comprises the following components in parts by mass: 20-25 parts of Astragalus membranaceus, 10-15 parts of Atractylodes macrocephala, 10-15 parts of Codonopsis pilosula, 10-13 parts of Zingiber officinale, 10-13 parts of Syzygium aromaticum, 8-12 parts of Panax notoginseng, 5-7 parts of Citrus reticulata, 5-7 parts of Angelica sinensis and 5-7 parts of raw licorice. The traditional Chinese medicine composition has the effects of warming kidney and assisting yang, tonifying qi and absorbing blood, promoting blood circulation and stopping bleeding, has a good treatment effect on the warfarin-induced hemorrhagic syndrome, and has no toxic side effects.

Description

Technical Field

[0001] This invention belongs to the field of veterinary drug preparation technology, and relates to a traditional Chinese medicine composition for treating warfarin-induced hemorrhage syndrome, its preparation method and application. Background Technology

[0002] Warfarin is a commonly used vitamin K antagonist, primarily used to prevent and treat thrombotic diseases such as atrial fibrillation, deep vein thrombosis, and pulmonary embolism. It exerts its anticoagulant effect by inhibiting vitamin K epoxide reductase complex subunit 1 (VKORC1), thus blocking the activation of coagulation factors II, VII, IX, and X. Warfarin has a narrow therapeutic window, and individual dosage variations are significant. Its anticoagulant effect is easily affected by diet, concomitant medications, genetic factors, and liver and kidney function. Overuse or excessively strong anticoagulant effects can cause coagulation disorders and bleeding, manifesting as skin ecchymosis, nosebleeds, gingival bleeding, hematuria, etc. In severe cases, gastrointestinal bleeding, intracranial hemorrhage, or retroperitoneal hemorrhage can occur, even endangering life. Furthermore, animals that ingest rodenticides containing warfarin may also experience severe bleeding and death.

[0003] Currently, warfarin-related bleeding is mainly treated by discontinuing warfarin, supplementing with vitamin K, and transfusing fresh frozen plasma or prothrombin complex concentrate. Vitamin K (VK) has a relatively slow onset of action and is difficult to meet the emergency treatment needs of severe bleeding on its own. Fresh frozen plasma transfusion requires a large volume and carries risks such as allergies and circulatory overload. Although prothrombin complex concentrate has a faster onset of action, it is expensive and carries a certain risk of thrombosis, thus limiting its clinical application.

[0004] Therefore, existing treatments still have shortcomings in terms of onset speed, cost, safety, and accessibility. Developing a traditional Chinese medicine composition that is easy to prepare, cost-effective, safe, and efficient, and capable of improving warfarin-induced coagulation dysfunction and bleeding symptoms, is of great significance for the prevention and treatment of warfarin-induced bleeding syndrome. Summary of the Invention

[0005] Purpose of the invention: The purpose of this invention is to provide a traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome, which aims to improve coagulation dysfunction and bleeding symptoms caused by warfarin, and to provide a safe, effective, and easy-to-prepare intervention for warfarin-related bleeding, thereby overcoming the shortcomings of existing treatment methods in terms of onset time, cost, accessibility, and safety.

[0006] Another object of the present invention is to provide a method for preparing the traditional Chinese medicine composition.

[0007] A third objective of this invention is to provide specific applications of this traditional Chinese medicine composition.

[0008] Technical solution: The present invention provides a traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome, wherein the composition comprises the following components by weight: Astragalus membranaceus 20-25 parts, Atractylodes macrocephala 10-15 parts, Codonopsis pilosula 10-15 parts, Zingiber officinale (processed) 10-15 parts, Syzygium aromaticum 10-15 parts, Panax notoginseng 8-12 parts, Citrus reticulata 5-7 parts, Angelica sinensis 5-7 parts, and Glycyrrhiza uralensis (raw) 5-7 parts.

[0009] Furthermore, the composition of the traditional Chinese medicine is as follows: 20 parts Astragalus membranaceus, 10 parts Atractylodes macrocephala, 15 parts Codonopsis pilosula, 15 parts Zingiber officinale (processed), 15 parts Clove, 10 parts Panax notoginseng, 7 parts Citrus reticulata peel, 7 parts Angelica sinensis, and 7 parts Glycyrrhiza uralensis.

[0010] This invention also provides a method for preparing the aforementioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome, the preparation steps of which are as follows: Step (1) Soak the ingredients to be decocted in 1.5 times their volume of distilled water for 1 hour to allow them to fully absorb the water; After soaking in step (2), bring the raw materials to a boil over high heat. After boiling, immediately reduce to a simmer and continue simmering for 2 hours. After simmering, filter the dregs through an 80-mesh sieve and collect the liquid. The filtered dregs will be simmered again. Step (3) Add 1.5 times the volume of distilled water to the dregs, bring the dregs to a boil over high heat, and immediately reduce to a simmer and continue simmering for 2 hours. After simmering, filter the dregs through an 80-mesh sieve and discard them, then collect the liquid. Combine the two decoctions and concentrate the liquid over a simmer until it is equal in volume to the raw materials to make the final decoction.

[0011] The present invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome in the preparation of drugs that improve clinical indicators such as mouse body weight, average daily food intake, and average daily water intake.

[0012] The present invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced hemorrhage syndrome in the preparation of a drug to reduce mouse mortality.

[0013] This invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome in the preparation of drugs that improve tongue color, fecal signs and scoring in mice; The tongue color and fecal signs scores refer to the following: daily observation and scoring of the tongue color and feces of experimental mice. A pale red or pink tongue with a glossy appearance is recorded as 1 point; a slightly reddish-brown tongue with slightly reduced gloss is recorded as 2 points; a distinctly reddish-brown or dark red tongue without gloss is recorded as 3 points; a deep purple or dark purple tongue with visible punctate or patchy ecchymosis is recorded as 4 points; a tongue color score >2 points indicates bleeding, while ≤2 points indicates normal. A formed, pale yellow stool is recorded as 1 point; a soft or unformed stool with a small amount of bright red blood is recorded as 2 points; a soft, unformed, black (tarry) stool is recorded as 3 points; a hard, formed stool with a large amount of dark black tarry stool is recorded as 4 points; a fecal score >2 points indicates bleeding, while ≤2 points indicates normal.

[0014] The present invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome in the preparation of drugs that improve the visual and microscopic bleeding phenomena of the liver and intestines in mice.

[0015] The present invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome in the preparation of drugs that improve organ index indicators in mice.

[0016] This invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome in the preparation of drugs that improve the four coagulation parameters and international normalized ratio (INR) of peripheral blood in mice; The coagulation parameters include activated partial thromboplastin time (APTT), prothrombin time (PT), thrombin time (TT), and fibrinogen (FIB).

[0017] The present invention also provides the application of the above-mentioned traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome in the preparation of drugs that improve the expression of peripheral blood vitamin K (VK) content, liver tissue vitamin K epoxide reductase complex 1 (VKORC1), and γ-glutamyl carboxylase (GGCX).

[0018] The traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome consists of Astragalus membranaceus, Atractylodes macrocephala, Codonopsis pilosula, prepared ginger, clove, Panax notoginseng, tangerine peel, Angelica sinensis, and raw licorice. Among them, Astragalus membranaceus has the effects of invigorating qi and nourishing blood, and invigorating qi and strengthening the spleen; Atractylodes macrocephala has the effects of strengthening the spleen and replenishing qi, and strengthening the spleen and stomach. Codonopsis pilosula has the effects of tonifying the middle energizer and replenishing qi, strengthening the spleen and nourishing the stomach; Prepared ginger has the effects of warming the middle jiao and dispelling cold, stopping bleeding and improving bleeding. Cloves have the effect of warming the kidneys and invigorating yang. Panax notoginseng has the effects of promoting blood circulation, removing blood stasis, and stopping bleeding without leaving blood stasis. Angelica sinensis has the effects of nourishing blood, promoting blood circulation, and improving blood stasis in the tongue. Dried tangerine peel has the effects of regulating qi, strengthening the spleen, and enhancing immunity; Raw licorice has the effects of tonifying the spleen and replenishing qi, and harmonizing various medicines.

[0019] Beneficial Effects: Compared with the prior art, the present invention has the following significant advantages: 1. The traditional Chinese medicine composition of the present invention has the effects of warming the kidney and assisting yang, replenishing qi and stopping bleeding, and promoting blood circulation and stopping bleeding. It can improve warfarin-induced coagulation dysfunction and bleeding symptoms, reduce liver damage, and improve survival rate and overall body condition. Its effects may be related to regulating the activity of coagulation factors, increasing plasma vitamin K levels, and regulating the expression of VKORC1 and GGCX genes. Experimental results show that its therapeutic effect is better than that of the vitamin K treatment group. 2. The traditional Chinese medicine composition of the present invention provides a new intervention method for the prevention and treatment of warfarin-induced bleeding syndrome. It has the characteristics of simple preparation, low cost and convenient application. It can reduce the dependence on peripheral blood products such as prothrombin complex and fresh frozen plasma, reduce treatment costs and transfusion-related risks, and improve the prognosis of patients or diseased animals. Attached Figure Description

[0020] Figure 1 This is a comparison chart of the changes in clinical indicators such as body weight, average daily food intake, and average daily water intake of mice in each group according to embodiments of the present invention; in the chart, A is a comparison chart of the changes in body weight of mice in each group; B is a comparison chart of the changes in average daily food intake of mice in each group; C is a comparison chart of the changes in average daily water intake of mice in each group; (Note: The Model group is compared with each other, * P <0.05); Figure 2 This is a comparison chart of the mortality rates of mice in each group in the embodiments of the present invention; Figure 3 These are comparative charts showing the changes in tongue color and feces of mice in each group on days 1, 3, 6, and 9, and the tongue color and feces physical examination score chart on day 9 in this embodiment of the invention. In the charts, A is a comparative chart showing the changes in tongue color of mice in each group on days 1, 3, 6, and 9; B is a chart showing the tongue color physical examination score chart on day 9; C is a comparative chart showing the changes in feces of mice in each group on days 1, 3, 6, and 9; and D is a chart showing the feces physical examination score chart on day 9. (Note: When comparing groups, the same letter indicates no significant difference.) P >0.05, different letters indicate a significant difference. P <0.05;); Figure 4 These are visual comparison images of liver and intestinal bleeding in mice in different groups in this invention embodiment; Figure 5 These are comparative microscopic images of hemorrhage in the liver and intestines of mice stained with hematoxylin and eosin (HE) in various groups according to embodiments of the present invention; red arrows indicate hemorrhage. Figure 6These are comparative charts showing the changes in the indices of the heart, liver, spleen, lungs, and kidneys in mice across different groups in this invention embodiment. In the charts, A is a comparison chart of the heart organ indices in different groups of mice; B is a comparison chart of the liver organ indices in different groups of mice; C is a comparison chart of the spleen organ indices in different groups of mice; D is a comparison chart of the lung organ indices in different groups of mice; and E is a comparison chart of the kidney organ indices in different groups of mice. (Note: When comparing groups, the same letter indicates no significant difference.) P >0.05, different letters indicate a significant difference. P <0.05;); Figure 7 This is a comparison chart of the four coagulation indicators and international normalized ratio (INR) of mouse plasma in each group of embodiments of the present invention; in the chart, A is a comparison chart of activated partial thromboplastin time (APTT) of mouse plasma in each group of mice; B is a comparison chart of thrombin time (TT) of mouse plasma in each group of mice; C is a comparison chart of prothrombin time (PT) of mouse plasma in each group of mice; D is a comparison chart of fibrinogen (FIB) of mouse plasma in each group of mice; E is a comparison chart of international normalized ratio (INR) of mice in each group of mice; (Note: When comparing groups, the same letter indicates no significant difference.) P >0.05, different letters indicate a significant difference. P <0.05;); Figure 8 These are comparative graphs of peripheral blood vitamin K content, liver tissue vitamin VKORC1, and GGCX expression in mice in various groups according to embodiments of the present invention; Figure A is a comparative graph of peripheral blood vitamin K content in mice in various groups; Figure B is a comparative graph of liver tissue vitamin VKORC1 expression; Figure C is a comparative graph of GGCX expression; (Note: When comparing groups, the same letter indicates no significant difference) P >0.05, different letters indicate a significant difference. P <0.05). Detailed Implementation

[0021] The specific technical solution of the present invention will be further described in detail below with reference to specific examples.

[0022] This invention discloses a traditional Chinese medicine composition for treating warfarin-induced hemorrhage syndrome in large-scale farming. The composition comprises the following herbs in proportion: 20g Astragalus membranaceus, 10g Atractylodes macrocephala, 15g Codonopsis pilosula, 15g Zingiber officinale (processed), 15g Syzygium aromaticum, 10g Panax notoginseng, 7g Citrus reticulata peel, 7g Angelica sinensis, and 7g Glycyrrhiza uralensis. These herbs are soaked in 1.5 times their volume of distilled water for 1.5 hours. After boiling, the mixture is simmered for 2 hours, and the liquid is filtered through an 80-mesh sieve. Another 1.5 times their volume of distilled water is added to the dregs, and the mixture is simmered for another 2 hours, then filtered through an 80-mesh sieve. The two decoctions are combined and concentrated to 107mL over a low flame. After filtration, a 1g / mL aqueous decoction is prepared and administered via gavage.

[0023] The traditional Chinese medicine preparation of the present invention has the effects of warming kidney and reinforcing yang, supplementing qi to control bleeding, and activating blood circulation to stop bleeding. It has a good therapeutic effect on warfarin-induced bleeding syndrome with small toxic and side effects.

[0024] Example 1 Therapeutic effect of different traditional Chinese medicine compositions on warfarin-induced bleeding syndrome 1. Methods 1.1. Experimental animals Seventy-five healthy SPF grade male ICR mice aged 4-6 weeks, with a body weight of (20 ± 2) g, were selected, purchased from and housed in the experimental animal center of a certain university. The certificate number of animal experimental facilities is SCXK (Su) 2022-0009, and the use permit is SYXK (Su) 2022-0044. Housing conditions: (20 ± 1)°C, 30% - 40% humidity, free access to food and water, 12 h light cycle, and the experiment was carried out after 7 days of adaptive feeding. All operations in the feeding process of experimental mice and other experimental operations complied with the requirements of laboratory animal ethics (approval number: 202505038) and the Regulations on the Administration of Laboratory Animals of the People's Republic of China. Three parallel experiments were carried out under the same conditions in this study. 1.2. Composition of traditional Chinese medicine composition and preparation of water decoction The traditional Chinese medicine composition comprises 20 g of Astragali Radix, 10 g of Atractylodis Macrocephalae Rhizoma, 15 g of Codonopsis Radix, 15 g of Zingiberis Rhizoma Preparatum, 15 g of Caryophylli Flos, 10 g of Notoginseng Radix et Rhizoma, 7 g of Citri Reticulatae Pericarpium, 7 g of Angelicae Sinensis Radix and 7 g of Glycyrrhizae Radix et Rhizoma Cruda, and has the effects of warming kidney and reinforcing yang, supplementing qi to control bleeding, and activating blood circulation to stop bleeding. A total of 107 g of the traditional Chinese medicine composition proportionally weighed out was taken, added with 1.5 times the volume of distilled water and soaked for 1.5 h. After boiling, the mixture was decocted with slow fire for 2 h, and the medicinal solution was filtered through an 80-mesh filter screen. 1.5 times the volume of distilled water was added to the medicinal residue, the mixture was continuously decocted with slow fire for 2 h, and the medicinal solution was filtered through an 80-mesh filter screen. The two decoctions were combined and concentrated to 107 mL with slow fire, and filtered to prepare a 1 g / mL water decoction. 1.3. Experimental grouping and treatment methods Seventy-five healthy SPF-grade male ICR mice were randomly divided into four groups: a control group, a model group, a traditional Chinese medicine (TCM) group, a vitamin K group, and a TCM combined with vitamin K group (TCM+VK). Except for the control group, mice in all other groups were administered 0.1 mL of 0.025% warfarin solution by gavage daily for 1-3 days. From 4-10 days, mice in the model group were administered 0.2 mL of physiological saline by gavage, mice in the TCM group were administered 0.2 mL of TCM decoction by gavage, mice in the VK group were injected with 0.2 mL of VK injection intramuscularly, and mice in the TCM+VK group were administered 0.2 mL of TCM decoction by gavage and 0.2 mL of VK injection intramuscularly. mL; Mice in each group were sampled on day 9, and the survival rate, body weight, food intake, water intake, and bleeding status of each group were statistically observed; Prothrombin time (PT), activated partial thromboplastin time (APTT), fibrinogen (FIB), and thrombin time (TT) in the plasma of each group of mice were detected by kits; The expression levels of GGCX and VKORC1 genes in mouse liver tissue were detected by real-time quantitative PCR (RT-qPCR); The content of VK in plasma was detected by enzyme-linked immunosorbent assay (ELISA); All medications were administered according to the requirements in Table 1. The concentration of the decoction of traditional Chinese medicine was 1 g / mL, administered by gavage; the concentration of vitamin K was 0.75 ng / mL, administered by intramuscular injection; and the concentration of warfarin aqueous solution was 0.025 g / L, administered by oral gavage. Table 1 Grouping and treatment of experimental animals ; 1.4. The traditional Chinese medicine composition improved the body weight, average daily food intake, and average daily water intake of hemorrhagic mice. Daily records were kept of the body weight, average daily food intake, and average daily water intake of mice in each group. 1.5. Traditional Chinese medicine composition reduces mortality in hemorrhagic mice. The number of mice that died in each group was recorded daily, and the mortality rate of each group was calculated. 1.6. Traditional Chinese medicine composition improves tongue color and feces in hemorrhagic mice and reduces physical examination scores. The tongue color and feces of the experimental mice were observed and scored daily. A pale red or pink tongue with a glossy appearance was recorded as 1 point; a slightly reddish-brown tongue with slightly reduced gloss was recorded as 2 points; a distinctly reddish-brown or dark red tongue without gloss was recorded as 3 points; a deep purple or dark purple tongue with visible punctate or patchy ecchymosis was recorded as 4 points. A tongue color score >2 points indicated bleeding, while a score ≤2 points was considered normal. Formed, pale yellow feces were recorded as 1 point; soft or unformed feces with a small amount of bright red blood were recorded as 2 points; soft or unformed feces with a black (tarry) appearance was recorded as 3 points; hard, formed feces with a large amount of dark black tarry stool was recorded as 4 points. A fecal score >2 points indicated bleeding, while a score ≤2 points was considered normal. Changes in tongue color and feces were compared among the groups. 1.7. Traditional Chinese medicine composition improves abnormal hemorrhage in the liver and intestines of hemorrhagic mice as observed by eye and microscopic examination. After the experiment, mice in each group were sacrificed, and the abdominal cavity was immediately opened to observe the color, shape and bleeding of the liver and intestines. The focus was on recording changes such as liver enlargement, darkening of color, petechiae or ecchymosis on the surface, as well as intestinal congestion, edema, bleeding and abnormal color of intestinal contents. Images were collected for comparison and analysis. Liver and intestinal tissues from the same location were collected from mice in each group. The tissues were gently rinsed with physiological saline to remove residual peripheral blood and intestinal contents. After drying, the tissues were fixed in 4% paraformaldehyde fixative. Following fixation, the tissues were dehydrated, cleared, embedded in paraffin, paraffin-embedded, and sectioned. Section thickness was 4–5 μm. Hematoxylin-eosin (HE) staining was used to observe pathological changes in liver tissue structure, hepatocyte degeneration and necrosis, inflammatory cell infiltration, vasodilation, and tissue hemorrhage under a microscope. Simultaneously, the integrity of the intestinal mucosa, villous structure, mucosal edema, inflammatory cell infiltration, and hemorrhage were observed. The pathological evaluation was carried out based on the degree of tissue congestion, hemorrhage, inflammatory cell infiltration and tissue structural damage. The visual and histopathological changes of the liver and intestines in each group were compared to evaluate the ameliorative effect of the traditional Chinese medicine composition on warfarin-induced tissue hemorrhage and damage. 1.8. Traditional Chinese medicine composition improves abnormal organ indexes of heart, liver, spleen, lungs, and kidneys in hemorrhagic mice. After the experiment, the final weight of mice in each group was measured. The mice were then sacrificed and dissected quickly. The heart, liver, spleen, lungs, and kidneys were completely separated, and surrounding fat, fascia, and connective tissue were removed. The residual peripheral blood on the surface of the organs was washed with physiological saline, and the surface moisture was blotted dry with filter paper before the wet weight of each organ was measured. Calculate the indices of each organ using the following formula: Organ index (mg / g) = organ wet weight (mg) ÷ mouse final body weight (g).

[0025] The cardiac index, liver index, spleen index, lung index, and kidney index of mice in each group were calculated and compared between groups. By analyzing the changes in organ indices, the effects of warfarin-induced hemorrhage on major organs and the ameliorative effect of the traditional Chinese medicine composition on abnormal changes such as organ enlargement, atrophy, congestion, edema, or damage were evaluated. 1.9. Traditional Chinese medicine composition improves abnormal plasma coagulation parameters and international normalized ratio in hemorrhagic mice. Collect plasma samples, follow the instructions in the kit, and use a semi-automatic coagulation analyzer to measure and analyze the data. 1.10. Traditional Chinese medicine composition improves abnormal levels of vitamin K in peripheral blood and VKORC1 and GGCX expression in liver tissue of hemorrhaging mice. Mouse plasma samples were collected, and vitamin K levels were measured and analyzed according to the manufacturer's instructions. 20 mg of frozen liver tissue was weighed, total RNA was extracted, and the total RNA concentration was determined and reverse transcribed into cDNA. The obtained cDNA was immediately used for RT-qPCR detection. A 20 μL reaction mixture was premixed according to the PCR kit instructions. The reaction conditions were: 95℃ pre-denaturation for 30 s, 60–95℃ annealing for 40 s, 72℃ extension for 30 s, and a final extension at 72℃ for 5 min after all PCR cycles. Each sample was tested three times, and the cDNA copy number was calculated using the 2-ΔΔCt method. The primer sequences are as follows: GGCX gene primer sequence: F: 5'TACACCATCATGTTTCTGGTCCC 3'---R: 3'GGTGAGGAGAGGGTTCAGGT 5'; VKORC1 gene primer sequences: F: 5' ACAGCTGTTGTTAGGTTGCTT 3'---R: 3'GGTGGTAATGCACACAATGCAG 5'; 1.11 Data Analysis Statistical analysis was performed using IBM SPSS Statistics 17.0 software, and graphs were created using Graphpad Prism 10.5.1. All data are expressed as mean ± standard error (Mean ± SEM). One-way ANOVA was used for comparisons between groups. The criterion for statistical significance was... P <0.05, indicating a significant difference between different letters.

[0026] 2. Experimental Results 2.1. Body weight, average daily food intake, and average daily water intake of mice in each group. like Figure 1 As shown, the Model group mice exhibited weight loss, decreased appetite, and reduced water intake; after treatment, all treatment groups showed varying degrees of remission, with the TCM group mice showing significantly higher average daily food intake and water intake than the Model group. P <0.05), and weight also recovered significantly ( P <0.05); 2.2 Mortality rate of mice in each group like Figure 2 As shown, no mice died in the Control group, while the mortality rate in the Model group was 40.0%. The mortality rates in the VK group, TCM group, and TCM+VK group were 33.3%, 13.3%, and 20.0%, respectively, with the TCM group having the lowest mortality rate. 2.3 Changes in tongue color, feces, and physical signs in mice of each group. like Figure 3 As shown, the mice in the Model group exhibited symptoms such as dark red tongue, congestion, and black stool, with significantly elevated tongue color and fecal scores. P <0.05); symptoms were relieved in all treatment groups, with the TCM+VK group showing the most significant improvement; 2.4. Visual and microscopic examination of hemorrhage in the liver and intestines of mice in each group. like Figure 4-5 As shown, the liver and intestines of the Model group mice showed obvious congestion, hemorrhage and tissue damage; after treatment, the tissue lesions in all groups were reduced, with the TCM group and TCM+VK group showing more significant improvement. 2.5 Results of organ indices of heart, liver, spleen, lungs, and kidneys in each group of mice like Figure 6 As shown, compared with the Control group, the indices of organs such as the heart, liver, and spleen were significantly increased in the Model group; after treatment, the organ indices of all treatment groups decreased to varying degrees, with the TCM group and the TCM+VK group showing more significant improvement. P <0.05); 2.6 Results of plasma coagulation parameters and international normalized ratio in mice of each group like Figure 7 As shown, the APTT, PT, and INR ratios were significantly increased in the Model group mice, while the FIB content and TT were significantly decreased; compared with the Model group, the APTT, PT, and INR were significantly decreased in all treatment groups; the FIB content was significantly increased in all treatment groups, and the TT was significantly increased in the TCM and TCM + VK groups. P <0.05); 2.7 Results of vitamin K content in peripheral blood and VKORC1 and GGCX expression in liver tissue of mice in each group like Figure 8 As shown, the vitamin K content in peripheral blood and the expression of VKORC1 and GGCX genes in liver tissue were significantly decreased in the Model group mice; while the vitamin K content in peripheral blood and the expression of VKORC1 and GGCX genes in liver tissue were significantly increased in the TCM and TCM+VK groups. P <0.05); 3. Conclusion: This traditional Chinese medicine composition showed good therapeutic effects in treating warfarin-induced hemorrhage syndrome in mice, significantly improving bleeding symptoms and reducing hemorrhage-related mortality. The composition also had a positive effect on the recovery of coagulation function and liver damage in hemorrhaging mice, possibly by increasing plasma vitamin K levels, upregulating VKORC1 and GGCX gene expression in liver tissue, promoting vitamin K metabolism and restoring coagulation factor activity, thereby promoting coagulation and reducing tissue damage, and thus promoting the recovery of hemorrhaging mice. Simultaneously, the composition improved liver and intestinal pathological damage and promoted the recovery of body weight, food intake, and water consumption. Compared with vitamin K alone, the composition showed better comprehensive therapeutic effects, indicating that it may be an effective intervention method for warfarin-induced hemorrhage syndrome.

Claims

1. A traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome, characterized in that, The composition of the traditional Chinese medicine is as follows: Astragalus membranaceus 20-25 parts, Atractylodes macrocephala 10-15 parts, Codonopsis pilosula 10-15 parts, Zingiber officinale (processed) 10-15 parts, Syzygium aromaticum 10-15 parts, Panax notoginseng 8-12 parts, Citrus reticulata 5-7 parts, Angelica sinensis 5-7 parts, Glycyrrhiza uralensis (raw) 5-7 parts.

2. The method for preparing the traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome as described in claim 1, characterized in that, Includes the following steps: Step (1) Soak the raw materials to be decocted in 1.5 times their volume of distilled water; Step (2) After soaking, bring the soaked raw materials to a boil over high heat. After boiling, reduce to a simmer and continue simmering. After simmering, filter the dregs through an 80-mesh filter and collect the liquid. The filtered dregs will be simmered again. Step (3) Add 1.5 times the volume of distilled water to the filtered residue, bring to a boil over high heat, then reduce to a simmer and continue simmering. After simmering, filter the residue through an 80-mesh sieve and discard it, then collect the liquid. Combine the two decoctions and simmer over low heat to concentrate the liquid to the same volume as the raw materials to make the final decoction.

3. The method for preparing the traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome according to claim 2, characterized in that, The soaking time described in step (1) is 1 hour. The simmering time mentioned in step (2) is 2 hours; The simmering time in step (3) is 2 hours.

4. The application of a traditional Chinese medicine composition for treating warfarin-induced hemorrhage syndrome prepared according to the preparation method described in claim 2 in the preparation of drugs that improve clinical indicators such as mouse body weight, average daily food intake, and average daily water intake.

5. The application of a traditional Chinese medicine composition for treating warfarin-induced hemorrhage syndrome prepared according to the preparation method described in claim 2 in the preparation of a drug to reduce mouse mortality.

6. The application of a traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome prepared according to the preparation method described in claim 3 in the preparation of drugs that improve tongue color, changes in fecal signs, and scoring.

7. The application of a traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome prepared according to the preparation method described in claim 2 in the preparation of drugs that improve bleeding indicators in mouse liver and intestines under microscopic examination.

8. The application of a traditional Chinese medicine composition for treating warfarin-induced hemorrhage syndrome prepared according to the preparation method described in claim 2 in the preparation of drugs that improve organ index indicators in mice.

9. The application of a traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome prepared according to the preparation method described in claim 2 in the preparation of drugs that improve the four coagulation parameters and international normalized ratio in mouse peripheral blood.

10. The application of a traditional Chinese medicine composition for treating warfarin-induced bleeding syndrome prepared according to the preparation method described in claim 2 in the preparation of drugs that improve the expression indicators of peripheral blood vitamin K content, liver tissue vitamin K epoxide reductase complex 1, and γ-glutamyl carboxylase.

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