Traditional Chinese medicine composition for relieving or treating vascular endothelial injury of Kawasaki disease as well as preparation method and application of traditional Chinese medicine composition

Abstract

The invention discloses a traditional Chinese medicine composition for relieving or treating vascular endothelial injury of Kawasaki disease. The traditional Chinese medicine composition comprises the following components in parts by weight: 50-80 parts of gypsum, 10-30 parts of rhizoma anemarrhenae, 5-10 parts of honey-fried licorice root, 5-10 parts of polished round-grained rice, 10-20 parts of codonopsis pilosula and 10-20 parts of cortex moutan. The traditional Chinese medicine compound for treating or relieving the vascular endothelial injury of the Kawasaki disease adopts a scientific ratio, all the components can be mutually coordinated and promoted, and the traditional Chinese medicine compound is more suitable for children. A new way is provided for clinical treatment of the Kawasaki disease, a new preparation method of the traditional Chinese medicine composition is provided, if the traditional Chinese medicine composition is prepared into a standard Chinese patent medicine preparation, application of the traditional Chinese medicine in Kawasaki disease can be promoted, and the advantages of the traditional Chinese medicine in Kawasaki disease treatment are exerted.

Description

Technical Field

[0001] The present invention relates to a Chinese medicine composition, in particular to a Chinese medicine composition for alleviating or treating vascular endothelial damage caused by Kawasaki disease. Background Art

[0002] Kawasaki disease (KD) is an acute febrile rash characterized by systemic vasculitis. Its most serious complication is coronary artery damage, making it a leading cause of acquired heart disease in children. KD presents with characteristic clinical manifestations: persistent fever for more than five days, bilateral non-purulent conjunctival congestion, tangled tongue, a polymorphic rash, unilateral lymphadenopathy, edema and annular excoriation of the fingertips, and systemic vascular damage. Severe cases can lead to coronary artery stenosis and thrombosis, myocardial infarction, and even sudden death. The pro-inflammatory-anti-inflammatory imbalance and the resulting endothelial damage in the acute phase of KD are key pathological manifestations of KD.

[0003] Early treatment for KD focuses on anti-inflammatory drugs, with IVIG and aspirin as the primary treatments. These are merely symptomatic treatments targeting clinical manifestations. Even after aggressive Western medication, some children still develop coronary artery damage or even coronary aneurysms, requiring long-term or even lifelong medication, severely impacting their quality of life. Circulating endothelial microparticle levels are significantly elevated during the acute and subacute phases of KD. Regardless of the presence of coronary artery damage, brachial artery flow-mediated vasodilation in KD children at all stages of the disease course is significantly decreased compared with healthy controls, suggesting persistent endothelial damage in KD. Therefore, seeking new anti-inflammatory treatments targeting endothelial damage is an urgent clinical challenge for KD.

[0004] Kawasaki disease falls under the category of febrile diseases in Traditional Chinese Medicine (TCM). Baihu Tang (Baihu Tang) is a classic formula for treating these diseases. Traditionally, Baihu Tang consists of gypsum, rhizome of Anemarrhena asphodeloides, roasted licorice root, and glutinous rice. Young children have delicate spleens and stomachs, and the pungent and cooling properties of Baihu Tang can easily damage them. In clinical treatment, Baihu Tang is often modified to prevent and treat severe cold damage, making it more suitable for pediatric use. Summary of the Invention

[0005] Purpose of the invention: The technical problem to be solved by the present invention is to address the deficiencies of the existing technology. In order to be more suitable for the special population of pediatrics, the present invention provides a modified Baihu Decoction based on the traditional Baihu Decoction. This Chinese medicine combination can significantly alleviate or treat vascular endothelial damage in Kawasaki disease.

[0006] In order to solve the above technical problems, the present invention discloses a traditional Chinese medicine composition for alleviating or treating vascular endothelial damage in Kawasaki disease, comprising the following components in parts by weight: 50-80 parts of gypsum, 10-30 parts of rhizoma anemarrhenae, 5-10 parts of roasted licorice, 5-10 parts of polished rice, 10-20 parts of codonopsis pilosula and 10-20 parts of cortex moutan.

[0007] Preferably, the traditional Chinese medicine composition comprises the following components in parts by weight: 50-60 parts of gypsum, 15-20 parts of rhizoma anemarrhenae, 5-8 parts of licorice root, 6-10 parts of polished rice, 10-15 parts of codonopsis pilosula, and 10-15 parts of cortex moutan.

[0008] In a preferred embodiment, the traditional Chinese medicine composition comprises the following components in parts by weight: 50 parts of gypsum, 18 parts of rhizoma anemarrhenae, 6 parts of licorice root, 9 parts of polished rice, 12 parts of codonopsis pilosula, and 12 parts of cortex moutan.

[0009] The present invention also provides a preparation method of the above-mentioned traditional Chinese medicine composition, comprising the following steps: decocting a formula amount of gypsum, soaking rhizome of anemarrhena, roasted licorice, japonica rice, codonopsis pilosula, and cortex moutan in water, then decocting them several times, combining the water extracts, and concentrating the water extracts under reduced pressure to obtain an extract.

[0010] Preferably, the soaking time is 30-60 minutes, and the medicinal materials are decocted with water in an amount of 8 to 10 times the weight of the total medicinal materials.

[0011] Preferably, decoction is performed 2-3 times, each time for 30-60 minutes.

[0012] The present invention also discloses a traditional Chinese medicine preparation, which comprises the traditional Chinese medicine composition and pharmaceutically acceptable excipients and / or carriers.

[0013] The dosage form of the preparation is any one of water decoction, tablet, capsule, granule, pill, powder and paste.

[0014] When the traditional Chinese medicine compound of the present invention is made into tablets, the extract of the traditional Chinese medicine compound raw materials and carrier lactose or corn starch are granulated and dried, and lubricant magnesium stearate is added when necessary, mixed evenly, and then compressed into tablets.

[0015] When the Chinese medicinal compound of the present invention is prepared into capsules, the extract of the Chinese medicinal compound raw materials is uniformly mixed with carrier lactose or corn starch, granulated, and then filled into capsules to prepare the capsules.

[0016] When the traditional Chinese medicine compound of the present invention is prepared into granules, the extract of the traditional Chinese medicine compound raw materials is uniformly mixed with carrier lactose or corn starch, granulated, and dried to prepare the granules.

[0017] The present invention also discloses the use of the above-mentioned traditional Chinese medicine composition or the above-mentioned traditional Chinese medicine preparation in preparing a medicine for alleviating vascular endothelial damage caused by Kawasaki disease.

[0018] Traditional Chinese Medicine scholars classify Kawasaki disease as a "febrile disease," and clinical diagnosis and treatment of KD are often based on the "Wei Qi Ying Xue" dialectical framework. The pathogenesis of KD is characterized by exposure to a warm-heat pathogenic toxin that enters the child through the mouth and nose, invading the lung's Wei, then invading the Qi, and gradually entering the Ying and Blood, leading to illness. This aligns with the opening statement in Ye Tianshi's Treatise on Warm-Heat Diseases: "Warm pathogens, when received from above, first invade the lungs and then travel back to the pericardium. The lung governs Qi and belongs to the Wei, while the heart governs Blood and belongs to the Ying..." The typical clinical manifestations of KD, such as high fever, rash, chapped lips, conjunctival congestion, and swollen hands and feet, suggest that its progression generally conforms to the Wei Qi Ying Xue pattern of febrile diseases. If left untreated during the period of intense struggle between the pathogenic and righteous forces, KD can retrogradely spread to the pericardium, a key approach to Traditional Chinese Medicine treatment. The pathogenesis is characterized by a dual Qi and Ying burning syndrome (heat entering the Qi and gradually entering the Ying), and treatment focuses on clearing Qi and cooling the Ying. Baihu Decoction is a classic prescription for treating febrile diseases. It is a pungent and cooling formula. It is used for symptoms of both Qi and Ying burning to clear Qi and cool Ying, allowing pathogenic factors to escape. Gypsum is pungent, sweet, and extremely cold, clearing heat and relieving restlessness; Anemarrhena asphodeloides clears stomach heat, nourishes Yin and moistens dryness; Roasted Licorice root nourishes the stomach and promotes fluid production, harmonizing the other herbs and preventing severe cold from damaging the middle. Young children have delicate spleens and stomachs, which can be easily damaged by pungent and cooling formulas. Adding Codonopsis pilosula to nourish the spleen and lungs, nourishes blood and promotes fluid production, protects the spleen and stomach, and prevents damage to vital Qi. Adding Cortex Moutan Bark to the formula clears heat and cools blood, activates blood circulation, and dissipates stasis, preventing pathogenic factors from penetrating deeply into the bloodstream. Gypsum and Anemarrhena asphodeloides complement each other, particularly potent in clearing heat, relieving restlessness, promoting fluid production, and quenching thirst, making them an optimal combination for treating severe Qi heat. Japonica rice, roasted licorice root, Codonopsis pilosula, and Cortex Moutan bark nourish the stomach and promote fluid production, mitigating the bitter, cold, and descending properties of Gypsum and Anemarrhena asphodeloides, preventing severe cold from damaging the middle, and retaining the medicinal properties. These ingredients serve as adjuvants. Roasted Licorice root also serves as a harmonizer. Therefore, the formula has the effect of strengthening the body, eliminating evil, and preventing the disease from worsening.

[0019] Beneficial effects: The Chinese herbal compound for alleviating or treating vascular endothelial damage in Kawasaki disease of the present invention adopts a scientific ratio, and the various components can coordinate and promote each other, and are more suitable for children. Traditional Chinese medicine believes that the whole course of Kawasaki disease basically conforms to the transmission of Wei Qi, Ying Qi and Blood, and the clinical manifestations of Kawasaki disease in the acute stage are consistent with the characteristics of febrile diseases. Animal experiments have shown that the Chinese herbal combination of modified Baihu Decoction provided by the present invention can show an anti-inflammatory effect comparable to that of honeysuckle and forsythia, and effectively alleviates cardiac inflammatory infiltration and vascular elastic fiber destruction in Kawasaki disease model mice. In addition, it was observed that the Chinese herbal combination of modified Baihu Decoction reversed the changes in cardiac vascular fibrosis in Kawasaki disease mice. The present invention provides a new approach for the clinical treatment of Kawasaki disease, and provides a new method for preparing a Chinese herbal composition. If it is made into a standardized Chinese patent medicine preparation, it can promote the application of Chinese medicine in Kawasaki disease and give play to the advantages of Chinese medicine in the treatment of Kawasaki disease. BRIEF DESCRIPTION OF THE DRAWINGS

[0020] Figure 1 The total ion chromatograms of the modified Baihu decoction extract in Example 1 in positive and negative ion modes are shown;

[0021] Figure 2 The effect of the modified Baihu decoction in Example 2 on inflammatory infiltration and elastic fibers in the endothelial cells of the heart vessels of mice;

[0022] Figure 3 The effect of the modified Baihu decoction in Example 2 on the expression levels of inflammatory factors TNFα (A) and IL-1β (B) in the heart of mice;

[0023] Figure 4 The effect of the modified Baihu decoction in Example 2 on myocardial and cardiovascular fibrosis in mice;

[0024] Figure 5 This is the effect of the modified Baihu Decoction in Example 2 on the fibrosis markers αSMA and COL1 in the heart vessels of mice. DETAILED DESCRIPTION

[0025] The present invention will be further described below in conjunction with the accompanying drawings and specific embodiments, and the above and / or other advantages of the present invention will become more apparent.

[0026] Example 1: A traditional Chinese medicine compound for alleviating or treating vascular endothelial damage in Kawasaki disease, comprising the following components in parts by weight: 50g of gypsum, 18g of rhizoma anemarrhenae, 6g of roasted licorice root, 9g of polished rice, 12g of codonopsis pilosula, and 12g of moutan bark. The composition ratio of Baihu decoction (50g of gypsum, 18g of rhizoma anemarrhenae, 6g of roasted licorice root, and 9g of polished rice) is derived from the Treatise on Febrile Diseases. The dosages of codonopsis pilosula and moutan bark were determined by syndrome differentiation and addition by multiple pediatric experts from Jiangsu Provincial Hospital of Traditional Chinese Medicine, Suzhou Hospital of Traditional Chinese Medicine, and Children's Hospital Affiliated to Soochow University. All medicinal materials were purchased from Suzhou Tianling Chinese Medicine Pieces Co., Ltd. The comparative example of the honeysuckle and forsythia pair: 40g of honeysuckle and 40g of forsythia were purchased from Suzhou Chunhuitang Pharmaceutical Co., Ltd.

[0027] The preparation method is as follows:

[0028] Modified Baihu Decoction Chinese herbal composition: First, decoct the formulated amount of gypsum (wrapped in cloth for decoction), soak Rhizoma Anemarrhenae, Radix Glycyrrhizae, glutinous rice (wrapped in cloth for decoction), Codonopsis pilosula, and Cortex Moutan for 30 minutes, decoct twice with water 10 times the weight of the total medicinal materials, each time for 30 minutes, combine the aqueous extracts, and concentrate the aqueous extracts under reduced pressure to obtain an extract, i.e., the Chinese herbal compound extract, with a final concentration of 11.89 g crude drug / mL.

[0029] Honeysuckle and Forsythia: Take 5 times the amount of the prescribed medicinal materials (200g honeysuckle and 200g forsythia), add 3 times the amount of water, and soak for 30 minutes; then add 5 times the amount of water, reflux extract for 60 minutes, and filter while hot; add 3 times the amount of water to the residue, reflux extract for 30 minutes, and filter while hot; combine the filtrate, rotary evaporation and concentrate to an appropriate amount; freeze-dry to obtain the honeysuckle-forsythia extract.

[0030] The components of the modified Baihu Tang Chinese medicine compound were determined and analyzed using UHPLC-MS / MS. Compounddiscover 3.2 software was used to extract the characteristic peaks of the raw mass spectrometry data, and the mzcloud online database and mzVault Chinese medicine natural product database were used for characteristic peak identification. Figure 1 The main compounds are shown in Table 1. For example, the peak graph shows that the retention time is 24.04 min for timosaponin B II, 20.59 min for paeoniflorin, 28.73 min for paeonol, and 21.69 min for liquiritin.

[0031] Table 1 Identification results of main compounds (partial)

[0032]

[0033]

[0034] Example 2: Protective effect of the Chinese medicine composition prepared in Example 1 on the heart and blood vessels of mice.

[0035] Four-week-old, SPF-grade male C57BL / 6 mice were used for the experiments. These mice were provided by Suzhou Zhaoyan New Drug Research Center Co., Ltd., with Laboratory Animal Production License No. SCXK(Su)2018-0006 and Animal Quality Certificate No. 202005247. This animal experiment was approved by the Ethics Committee of Soochow University, with approval number SUDA20220906A01. Candida albicans cell wall extracts (CAWS) for modeling were purchased from the China Industrial Microbiology Culture Collection.

[0036] Method for constructing a Kawasaki disease mouse model: 0.2 mL CAWS (20 mg / mL) was injected intraperitoneally once a day for 5 consecutive days to construct a Kawasaki disease mouse model.

[0037] Yinqiao Powder is another classic prescription for Kawasaki disease. It is a pungent, cool, and neutral herb. The honeysuckle and forsythia combination is the core herb pair of Yinqiao Powder and is a commonly used Chinese herbal combination in the clinical treatment of Kawasaki disease. In this study, honeysuckle and forsythia were used as a comparative example. The pungent, cool, and neutral Baihu Decoction, after being modified, demonstrated comparable efficacy in improving Kawasaki disease vasculitis as the pungent, cool, and neutral honeysuckle and forsythia combination.

[0038] Grouping and Dosing: Mice in the model (CAWS) group, honeysuckle and forsythia (JYH-LQ) group, and modified white tiger decoction (JWBHT) group received intraperitoneal injections of 0.2 mL of CAWS (20 mg / mL) once daily for 5 consecutive days to establish a Kawasaki disease mouse model. Mice in the control (control) group received intraperitoneal injections of 0.2 mL of PBS. Simultaneously with model establishment, mice in the JYH-LQ group received oral gavage of 0.2 mL of an aqueous solution of honeysuckle and forsythia extract (raw drug dosage 2.6 g / kg), mice in the JWBHT group received oral gavage of 0.2 mL of a diluted solution of modified white tiger decoction concentrate (raw drug dosage 13.91 g / kg), and mice in the control and model groups received oral gavage of an equal volume of PBS. Following the final injection, mice were fed for 14 days, at which time their hearts were harvested and fixed with 4% paraformaldehyde solution.

[0039] Observation indicators:

[0040] Histological staining: Mouse heart tissue was fixed with 4% paraformaldehyde solution, dehydrated, embedded in paraffin, and sectioned. Hematoxylin and eosin, EVG, Masson's, and picrosirius red staining were performed. After sealing, the sections were observed and images were collected under an optical microscope.

[0041] Immunohistochemical staining: After dewaxing and antigen retrieval, add 20 μL of inactivation blocking buffer (2-in-1) to cover the tissue. Incubate at room temperature for 15 minutes. Rinse and drain for 30 seconds. Circle the tissue with an immunohistochemical pen and add the primary antibody CD31 to completely cover the tissue. Incubate in a 4°C humidified chamber overnight, then rewarm at 37°C for 30 minutes. Rinse and drain for 30 seconds. Add 20 μL of HRP polyvalent mouse / rabbit secondary antibody working solution to cover the tissue and incubate at room temperature for 30 minutes. Add 20 μL of 594-labeled Tyramide working solution to cover the tissue and incubate at room temperature for 10 minutes. Rinse, place the section in a retrieval chamber, add antibody stripping solution, microwave on high for 3 minutes until boiling, then reduce to low for 15 minutes. Allow to cool naturally. Repeat the primary and secondary antibody incubation steps, switching the fluorescent dye to 488-labeled Tyramide and incubating with a second primary antibody, αSMA or COL1. Add 40ul of anti-fade DAPI sealing two-in-one solution on the tissue, cover with a coverslip after 5 minutes, complete the sealing, and scan to collect images.

[0042] (1) Effects of Modified Baihu Decoction on Inflammatory Infiltration and Elastic Fibers in the Endothelial Cells of Mouse Cardiac Vascular Endothelial Cells

[0043] HE staining of pathological sections of mouse hearts showed that the vascular endothelial cells of mice in the normal group were intact and smooth, without inflammatory cell infiltration, while the vascular endothelial cells of mice in the model group were rough, with incomplete structure and disordered cell arrangement. Extensive infiltration of inflammatory cells was observed in the blood vessels and surrounding myocardial tissues. The vascular endothelial structures of the JWBHT-treated group and the JYH-LQ-treated group were relatively intact, with less inflammatory cell infiltration. EVG staining of pathological sections of mouse hearts showed that the elastic fibers of the vascular walls of mice in the control group were regularly arranged and curled in an orderly manner, while the elastic fibers of the vascular walls of mice in the model group were disorderly arranged, loose and broken, while the elastic fibers of the JWBHT-treated group and the JYH-LQ-treated group were relatively regular, with no obvious breakage. This proves that the modified Baihu Decoction Chinese medicine composition provided by the present invention can exhibit anti-inflammatory efficacy comparable to that of honeysuckle and forsythia, and effectively reduce inflammatory infiltration of the heart vessels and destruction of vascular elastic fibers in Kawasaki disease model mice. The results are as follows. Figure 2 shown.

[0044] (2) Effect of Modified Baihu Decoction on the Expression Levels of Inflammatory Factors TNFα and IL-1β in Mouse Hearts

[0045] like Figure 3 As shown, normal mice showed no cytokine expression in the cardiovascular region. However, IL-1β and TNF-α expression was elevated in the cardiovascular tissues of model mice, but decreased after JWBHT administration. This demonstrates that the modified Baihu Tang Chinese medicine composition of the present invention can effectively reduce the expression of inflammatory factors in the cardiovascular region of Kawasaki disease model mice.

[0046] like Figure 4 As shown, Masson and picrosirius red staining of mouse cardiac pathological sections revealed essentially no collagen deposition or collagen fibrosis in normal mouse myocardial tissue. However, model mouse heart tissue exhibited varying degrees of collagen deposition and severe myocardial and perivascular fibrosis, with JWBHT administration significantly improving fibrosis. This demonstrates that the modified Baihu Tang Chinese herbal composition provided by the present invention can effectively alleviate cardiac fibrosis in Kawasaki disease model mice.

[0047] (3) Effects of Modified Baihu Decoction on αSMA and COL1 fibrosis markers in mouse heart vessels

[0048] like Figure 5 As shown, compared with normal mice, CD31 was downregulated, αSMA and COL1 were upregulated in the cardiovascular region of the model mice, and the expression trend of these markers was reversed after JWBHT administration. This shows that the treatment of the modified Baihu Decoction Chinese medicine composition of the present invention can effectively reduce the expression of cardiovascular fibrosis markers in Kawasaki disease model mice.

[0049] The present invention provides a concept and method for a traditional Chinese medicine composition for alleviating or treating vascular endothelial damage in Kawasaki disease. Numerous methods and approaches exist for implementing this technical solution. The foregoing merely represents a preferred embodiment of the present invention. It should be noted that those skilled in the art may make various improvements and modifications without departing from the principles of the present invention, and such improvements and modifications are also within the scope of protection of the present invention. Any components not specified in this embodiment may be implemented using existing technologies.

Claims

1. A Chinese medicine composition for alleviating or treating vascular endothelial damage in Kawasaki disease, characterized in that: The invention comprises the following components in parts by weight: 50-80 parts of gypsum, 10-30 parts of rhizoma anemarrhenae, 5-10 parts of licorice root, 5-10 parts of polished rice, 10-20 parts of codonopsis pilosula and 10-20 parts of cortex moutan.

2. The Chinese medicine composition according to claim 1, characterized in that The invention comprises the following components in parts by weight: 50-60 parts of gypsum, 15-20 parts of rhizoma anemarrhenae, 5-8 parts of licorice root, 6-10 parts of polished rice, 10-15 parts of codonopsis pilosula, and 10-15 parts of cortex moutan.

3. The Chinese medicine composition according to claim 1, characterized in that The invention comprises the following components in parts by weight: 50 parts of gypsum, 18 parts of rhizoma anemarrhenae, 6 parts of licorice root, 9 parts of polished rice, 12 parts of codonopsis pilosula and 12 parts of cortex moutan.

4. The method for preparing the Chinese medicine composition according to claim 1, characterized in that: The method comprises the following steps: decocting a prescribed amount of gypsum, soaking rhizome of anemarrhena, roasted liquorice, polished rice, codonopsis pilosula and cortex moutan in water, decocting the mixture for several times, combining water extracts and concentrating the water extracts under reduced pressure to obtain an extract.

5. The preparation method according to claim 4, characterized in that The soaking time is 30-60 minutes, and the herbs are decocted with water 8 to 10 times the weight of the total herbs.

6. The preparation method according to claim 4, characterized in that Decoction 2-3 times, 30-60 minutes each time.

7. A Chinese medicine preparation, characterized in that The traditional Chinese medicine preparation comprises the traditional Chinese medicine composition according to claim 1 and pharmaceutically acceptable excipients and / or carriers.

8. The Chinese medicine preparation according to claim 7, characterized in that The dosage form of the preparation is any one of water decoction, tablet, capsule, granule, pill, powder and paste.

9. Use of the traditional Chinese medicine composition according to claim 1 or the traditional Chinese medicine preparation according to claim 7 in the preparation of a medicament for alleviating or treating vascular endothelial damage in Kawasaki disease.

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