Application of Arctium lappa root polysaccharide in preparing medicine for treating pulmonary hypertension

Abstract

The present invention discloses the application of burdock root polysaccharide in the preparation of a medicament for treating pulmonary hypertension, belonging to the field of pharmaceutical technology. The present invention prepares a medicament for treating pulmonary hypertension with burdock root polysaccharide as the sole active ingredient. The medicament comprises an effective amount of burdock root polysaccharide and pharmaceutically acceptable excipients. Experimental results show that burdock root polysaccharide significantly reduces the increase in pulmonary artery pressure induced by monocrotaline in rats, significantly inhibits the increase in right ventricular hypertrophy index, significantly alleviates pulmonary artery endothelial injury, medial thickening, excessive proliferation of medial cells and infiltration of inflammatory cells in rats with pulmonary hypertension, significantly alleviates the degree of pulmonary artery vascular muscleization in rats with pulmonary hypertension, has the effects of reducing pulmonary artery vascular remodeling and lowering pulmonary artery pressure, and has a significant therapeutic effect on pulmonary hypertension and right ventricular hypertrophy in rats, and can be applied to the preparation of a medicament for treating pulmonary hypertension.

Description

Technical Field

[0001] The present invention belongs to the technical field of medicine, and particularly relates to the application of Arctium lappa L. root polysaccharide in the preparation of drugs for treating pulmonary hypertension. Background Art

[0002] Pulmonary hypertension (PH) is a complex and progressive cardio-pulmonary system disease. The exact pathogenesis is still unclear. The survival period of patients is limited, and there is no effective cure clinically. PH is also regarded as the "cancer" in the cardiovascular field. Approximately 1% of the global population suffers from pulmonary hypertension, and the incidence increases with age, especially significantly increasing to 10% in people over 65 years old, and about 50% of these patients are accompanied by heart failure at the same time. With the gradual aging of our country, PH has become one of the serious problems affecting the health of residents. At present, the treatment of PH mainly focuses on vasodilation, lacking ideal treatment means.

[0003] Arctium lappa L. is a biennial herbaceous plant of the Compositae family and is a common plant with both medicinal and edible uses. Arctium lappa L. root polysaccharide (ALR-PS) is prepared by extracting from its roots. Existing studies have shown that Arctium lappa L. root polysaccharide has various pharmacological activities such as anti-tumor, anti-allergy, and hypoglycemic effects. However, so far, there has been no report on the related research of the therapeutic effect of Arctium lappa L. root polysaccharide on pulmonary hypertension. Summary of the Invention

[0004] In order to solve the problems existing in the prior art, the purpose of the present invention is to provide the application of Arctium lappa L. root polysaccharide in the preparation of drugs for treating pulmonary hypertension. Arctium lappa L. root polysaccharide has the effects of alleviating pulmonary artery vascular remodeling and reducing pulmonary artery pressure, and has a significant therapeutic effect on pulmonary hypertension and right ventricular hypertrophy in rats, and can be applied to the preparation of drugs for treating pulmonary hypertension.

[0005] The purpose of the present invention is achieved in the following way:

[0006] The present invention provides the application of Arctium lappa L. root polysaccharide in the preparation of drugs for treating pulmonary hypertension.

[0007] Based on the above technical solution, further, the Arctium lappa L. root polysaccharide is the only active ingredient.

[0008] Based on the above technical solution, further, the drug includes an effective amount of Arctium lappa L. root polysaccharide and pharmaceutically acceptable excipients.

[0009] Based on the above technical solution, further, the pharmaceutically acceptable excipients include fillers, diluents, binders, disintegrants, emulsifiers, and drug carriers without toxic side effects.

[0010] Based on the above technical solutions, further, the dosage forms of the drug include tablets, granules, oral liquid preparations, drops, injection preparations and capsule preparations.

[0011] Based on the above technical solutions, further, the drug is prepared in the form of a single-dose drug.

[0012] Based on the above technical solutions, further, the single-dose drug contains 100 - 30000 mg of burdock root polysaccharide.

[0013] Based on the above technical solutions, further, the drug can significantly reduce the increase in pulmonary artery pressure, significantly inhibit the increase in right ventricular hypertrophy index, and significantly relieve pulmonary artery vascular remodeling.

[0014] The beneficial effects of the present invention compared with the prior art are as follows:

[0015] The extraction cost of the burdock root polysaccharide of the present invention is low and the safety is high. It has been experimentally confirmed that the burdock root polysaccharide can significantly reduce the increase in pulmonary artery pressure induced by monocrotaline in rats, significantly inhibit the increase in right ventricular hypertrophy index, significantly relieve pulmonary artery endothelial injury, middle layer thickening, excessive proliferation of middle layer cells and inflammatory cell infiltration in rats with pulmonary hypertension, significantly relieve the degree of pulmonary artery vascular muscleization in rats with pulmonary hypertension, and has the effects of reducing pulmonary artery vascular remodeling and pulmonary artery pressure. The burdock root polysaccharide has a significant therapeutic effect on pulmonary hypertension and right ventricular hypertrophy in rats and can be applied to the preparation of drugs for the treatment of pulmonary hypertension. Description of the Drawings

[0016] In order to more clearly illustrate the embodiments of the present invention, the drawings related to the embodiments will be briefly introduced below.

[0017] Figure 1 ALR-PS can significantly reduce the increase in pulmonary artery pressure in rats with pulmonary hypertension induced by monocrotaline (MCT), *p < 0.05, ***p < 0.001.

[0018] Figure 2 ALR-PS can significantly reduce right ventricular hypertrophy in rats with pulmonary hypertension induced by monocrotaline (MCT), *p < 0.05, ***p < 0.001.

[0019] Figure 3The results showed that ALR-PS could significantly alleviate pulmonary artery vascular remodeling in rats induced by monocrotaline (MCT). Among them, A showed HE staining and αSMA immunohistochemistry of the pulmonary artery in rats (20×, scale bar: 50 μm), B showed the percentage of medial thickness WT%, C showed the percentage of medial area WA%, D showed the relative optical density value of αSMA, *p<0.05, **p<0.01, ***p<0.001.

[0020] Figure 4 The results were for the pulmonary artery vascular muscleization of rats in each group. Among them, A showed the immunohistochemical staining map of αSMA (4×, scale bar: 200 μm), B showed the percentage of muscleization, *p<0.05, **p<0.01, ***p<0.001. Specific implementation manners

[0021] The present invention will be described in detail below in conjunction with embodiments, but the implementation manners of the present invention are not limited thereto. Obviously, the embodiments described below are only partial embodiments of the present invention. For those skilled in the art, without creative efforts, obtaining other similar embodiments will fall within the protection scope of the present invention.

[0022] Embodiment 1

[0023] This embodiment provides a preparation method of arctium lappa root polysaccharide (ALR-PS), which includes the following steps:

[0024] (1) Peel, wash and slice the arctium lappa root, place it in an oven and dry at 60°C, and then crush it.

[0025] (2) Add water at a material-liquid ratio of 1:15, boil at 80°C for 3 h, filter successively with gauze and filter paper, collect the filtrate, and continue to add water to cook the filter residue, repeating this step 3-4 times.

[0026] (3) Collect the filtrate and perform suction filtration with ordinary filter paper to preliminarily remove insoluble impurities.

[0027] (4) Concentrate the filtrate by rotary evaporation to an appropriate volume, add 3 volumes of absolute ethanol, add it while stirring, let it stand overnight, and the solution will be layered.

[0028] (5) Collect the lower-layer precipitate, dry it by rotary evaporation, and then add an appropriate amount of water to fully dissolve the arctium lappa root polysaccharide.

[0029] (6) Repeat steps (4)-(5) 3-4 times, collect the aqueous solution of arctium lappa root polysaccharide, and freeze-dry it to obtain arctium lappa root polysaccharide.

[0030] (7) After dissolving the dry product of arctium lappa root polysaccharide in water, filter it with a 0.45-μm microporous filter membrane.

[0031] (8) Collect the filtrate and freeze-dry it to obtain the burdock root polysaccharide powder.

[0032] Example 2

[0033] This example examines the therapeutic effect of the burdock root polysaccharide prepared in Example 1 on rats with monocrotaline-induced pulmonary hypertension.

[0034] (1) Establish a rat model of pulmonary hypertension: Subcutaneously inject 60 mg / kg of monocrotaline (MCT, C101556-1g, Shanghai Aladdin Biochemical Technology Co., Ltd.) into the neck of rats once.

[0035] (2) Grouping of animal experiments

[0036] Adult male SD rats (6-8 weeks old, weighing 180-200 g) were purchased from Liaoning Changsheng Biotechnology Co., Ltd. (certificate number: SCXK(Liao)2020-0001). They were randomly divided into the following groups, with 8 rats in each group.

[0037] a. Normal group: Rats were subcutaneously injected with 10 ml / kg of normal saline into the neck once, weighed daily, and gavaged with ddH2O (10 ml / kg) continuously for 28 days.

[0038] b. Normal drug administration group: Rats were subcutaneously injected with 10 ml / kg of normal saline into the neck once, weighed daily, and gavaged with ALR-PS (600 mg / kg) once continuously for 28 days.

[0039] c. MCT group: Rats were subcutaneously injected with 60 mg / kg of monocrotaline into the neck once, weighed daily, and gavaged with ddH2O once continuously for 28 days.

[0040] d. MCT + ALR-PS (300 mg / kg) group: Rats were subcutaneously injected with 60 mg / kg of monocrotaline into the neck once, weighed daily, and gavaged with ALR-PS (300 mg / kg) once continuously for 28 days.

[0041] e. MCT + ALR-PS (600 mg / kg) group: Rats were subcutaneously injected with 60 mg / kg of monocrotaline into the neck once, weighed daily, and gavaged with ALR-PS (600 mg / kg) once continuously for 28 days.

[0042] (3) Detection of right ventricular systolic pressure (RVSP) in rats

[0043] After 28 days of treatment, isoflurane inhalation anesthesia was applied. The rats were anesthetized and fixed supine. The skin around the neck was prepared. Above the course of the right external jugular vein on the right neck of the rats, a small incision parallel to the long axis of the neck was made using a surgical scalpel. The right external jugular vein was bluntly dissected. The distal end of the right external jugular vein of the rats was ligated using a sterile surgical suture. Under a stereomicroscope, a small "V"-shaped incision was made at the proximal end using a corneal scissors, and a polyethylene catheter filled with heparin solution was inserted. Guided by the change in pressure waveform, it passed through the right external jugular vein, right atrium, and until the right ventricle. The other end of the catheter was connected to a pressure sensor, and the right ventricular systolic pressure (RVSP) of the rats was measured using a BL-420N biological function experimental system to evaluate the pulmonary artery pressure level.

[0044] (4) Detection of right ventricular hypertrophy index (Fulton Index)

[0045] The hearts of the rats were taken out. The large blood vessels, right atrium, and left atrium connected to the heart were removed using ophthalmic scissors. The right ventricle (RV) and left ventricle + interventricular septum (LV+S) were separated. After blotting the excess normal saline with filter paper, each component was weighed, and the Fulton index, which is the ratio of the right ventricular weight to the left ventricular plus interventricular septum weight [RV / (LV+S)], was calculated to represent the degree of right ventricular hypertrophy.

[0046] (5) Morphological examination of pulmonary tissue arterial vascular remodeling

[0047] The anterior chest was lifted by cutting along the two sides of the ribs. Pre-cooled normal saline was injected into the right ventricle at a constant speed using an infusion set. The right auricle was cut open for perfusion. Using a 10 mL syringe, 4% paraformaldehyde was injected into the right lung through the right bronchus. The injection volume was appropriate when the right lung was filled to a slightly inflated state. The right lung tissue was separated and immersed in 4% paraformaldehyde fixative and stored at 4°C for 72 h. Then it was paraffin-embedded, and 4-μm-thick sections were prepared. HE staining (C0105S, Beyotime Biotechnology Co., Ltd.) was performed to evaluate the morphological changes. Using an Image Pro Plus pathological image analysis system, the outer diameter, inner diameter, medial area, and total vascular area of the small pulmonary arteries were measured, and the medial wall thickness percentage WT% = (outer diameter - inner diameter) / (outer diameter) × 100; the area percentage WA% = (medial area) / (total vascular area) × 100 was calculated to represent the degree of pulmonary artery thickening.

[0048] α-Smooth muscle actin (α-SM-actin, αSMA) is a marker protein of smooth muscle cells and also a vascular myogenic factor. The sections were subjected to αSMA immunohistochemistry (ab7817, Abcam), and the expression of αSMA in the pulmonary arteries of rats in each group was evaluated using the relative optical density (relative OD) to determine the proliferation of smooth muscle cells in the middle layer of the pulmonary artery. According to the αSMA positive expression area in the pulmonary artery vessels, the myofibrotic ratio of pulmonary artery vessels with a diameter < 200 μm was statistically analyzed to evaluate the degree of pulmonary artery vessel myofibrosis.

[0049] (6) Results

[0050] Figure 1 The results of the mean right ventricular pressure (RVSP) of rats in each group are shown. The results indicate that the right ventricular systolic pressure of rats in the monocrotaline-induced model group was significantly increased (p < 0.001), and both 300 mg / kg and 600 mg / kg of burdock root polysaccharide could effectively alleviate the increase in RVSP (p < 0.05), and the 600 mg / kg dose group was significantly better than the 300 mg / kg group (p < 0.05), indicating that burdock root polysaccharide could significantly reduce the increase in pulmonary artery pressure induced by monocrotaline in rats.

[0051] Figure 2 The results of the right ventricular hypertrophy index (Fulton Index) of rats in each group are shown. The results indicate that the right ventricular hypertrophy index of rats in the model group was significantly increased (p < 0.001), and the right ventricular hypertrophy index decreased after administration of burdock root polysaccharide. Among them, 600 mg / kg of burdock root polysaccharide could significantly inhibit the increase in the right ventricular hypertrophy index (p < 0.05).

[0052] Figure 3 The results of the lung tissue morphology of rats in each group are as follows Figure 3 The HE staining results of A showed that in the pulmonary artery hypertension model group, the pulmonary artery vascular endothelial cells were damaged, the pulmonary artery lumen was narrowed and occluded, accompanied by a large number of inflammatory cell infiltrations. However, the lung tissue morphology of rats in the group treated with burdock root polysaccharide was significantly improved. Burdock root polysaccharide could significantly protect vascular endothelial cells, inhibit arterial lumen stenosis, and reduce inflammatory cells. Figure 3 B, Figure 3 The results of C showed that the percentage of medial wall thickness (WT%) and the percentage of medial wall area (WA%) in the model group were significantly increased (p < 0.001). After administration of burdock root polysaccharide, the WT% and WA% indexes decreased (p < 0.05), indicating that burdock root polysaccharide could significantly alleviate the thickening of the pulmonary artery middle layer in rats with pulmonary artery hypertension.

[0053] The αSMA immunohistochemical results of the rat pulmonary artery are as Figure 3 A, Figure 3As shown in D, the optical density value of αSMA in the model group of rats increased significantly (p < 0.001), and the optical density value of αSMA decreased after administration of burdock root polysaccharide (p < 0.05). The results indicate that burdock root polysaccharide can significantly alleviate the excessive proliferation of pulmonary artery middle layer cells in rats with pulmonary hypertension.

[0054] Figure 4 The results of pulmonary artery vascular muscleization in rats of each group are as follows. After immunohistochemical staining of αSMA in the pulmonary artery of rats, the ratio of muscularized blood vessels with a blood vessel diameter < 200 μm was statistically analyzed. The results showed that the ratio of muscularized blood vessels in the model group of rats increased significantly (p < 0.001), and the ratio of muscularized blood vessels decreased after administration of burdock root polysaccharide. The results indicate that burdock root polysaccharide can significantly alleviate the degree of pulmonary artery vascular muscleization in rats with pulmonary hypertension.

[0055] Finally, it should be noted that the above embodiments are only used to illustrate the technical solutions of the present invention, rather than to limit them; although the present invention has been described in detail with reference to the foregoing embodiments, those of ordinary skill in the art should understand that they can still modify the technical solutions described in the foregoing embodiments, or perform equivalent replacements for some or all of the technical features; and these modifications or replacements do not make the essence of the corresponding technical solutions deviate from the scope of the technical solutions of the embodiments of the present invention.

Claims

1. Use of burdock root polysaccharide in the preparation of a medicament for treating pulmonary hypertension; The medicament can significantly reduce the increase in pulmonary artery pressure, significantly inhibit the increase in right ventricular hypertrophy index, and significantly relieve pulmonary artery vascular remodeling; The preparation method of the burdock root polysaccharide comprises the following steps: (1) Peel, wash and slice the burdock root, dry it in an oven at 60 °C, and pulverize it; (2) Add water at a material-liquid ratio of 1:15, boil at 80 °C for 3 h, filter successively with gauze and filter paper, collect the filtrate, continue to add water to cook the filter residue, and repeat this step 3-4 times; (3) Collect the filtrate and perform suction filtration with ordinary filter paper to preliminarily remove insoluble impurities; (4) Concentrate the filtrate by rotary evaporation to an appropriate volume, add 3 volumes of anhydrous ethanol, add it while stirring, let it stand overnight, and the solution will be stratified; (5) Collect the lower precipitate, dry it by rotary evaporation, and then add an appropriate amount of water to fully dissolve the burdock root polysaccharide; (6) Repeat steps (4)-(5) 3-4 times, collect the aqueous solution of burdock root polysaccharide, and freeze-dry it to obtain burdock root polysaccharide; (7) Dissolve the dried burdock root polysaccharide in water and filter it through a 0.45 μm microporous filter membrane; (8) Collect the filtrate and freeze-dry it to obtain burdock root polysaccharide powder.

2. The application according to claim 1, wherein The burdock root polysaccharide is the sole active ingredient.

3. The application according to claim 1, wherein The medicament comprises an effective amount of burdock root polysaccharide and a pharmaceutically acceptable excipient.

4. The application according to claim 3, wherein The pharmaceutically acceptable excipients include fillers, diluents, binders, disintegrants and emulsifiers.

5. The application according to claim 1, characterized in that, The dosage form of the medicament includes tablets, granules, oral liquid preparations, drops, injection preparations and capsule preparations.

6. The application according to claim 1, characterized in that The medicament is prepared in the form of a single-dose medicament, and the single-dose medicament contains 100-30000 mg of burdock root polysaccharide.

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