Traditional Chinese medicine composition for preventing or treating pulmonary nodules and application thereof

Abstract

The invention provides a traditional Chinese medicine composition for preventing or treating pulmonary nodules and application thereof. The traditional Chinese medicine composition comprises or consists of the following traditional Chinese medicine raw materials in parts by weight: 1-20 parts of ginseng, 6-30 parts of gynostemma pentaphyllum, 3-30 parts of rhizoma pinelliae preparata, 2-20 parts of thunberg fritillary bulb, 3-30 parts of moutan bark, 3-30 parts of selfheal, 3-30 parts of vinegar-processed curcuma zedoary, 5-50 parts of coix seed, 3-20 parts of bran-fried immature bitter orange, 3-30 parts of pericarpium citri reticulatae, 1-30 parts of roasted herba epimedii and 3-20 parts of roasted liquorice. The traditional Chinese medicine composition is rich in raw materials and scientific and advanced in formula, is effectively used for preventing or treating pulmonary nodules, can be developed into a new medicine for treating pulmonary nodules, and has relatively high economic and social benefits.

Description

Technical Field

[0001] The present invention belongs to the field of medicine, and in particular, relates to a Chinese medicine composition and its application in preventing or treating pulmonary nodules. Background Art

[0002] Pulmonary nodules refer to focal, round, and high-density lung shadows with a diameter of ≤3 cm on imaging. They may be isolated or multiple, without atelectasis, hilar lymphadenopathy, or pleural effusion. Their detection rate is increasing year by year, and some develop into lung cancer, which can easily cause anxiety and depression in patients, seriously endangering public health. The etiology of pulmonary nodules is complex and may be related to environmental pollution, occupational exposure, infection, tumors, etc. There are generally no obvious clinical symptoms, and a few manifest as cough, sputum, or even hemoptysis.

[0003] Malignant pulmonary nodules are very likely to progress to lung cancer if they are not intervened early. Lung cancer ranks second in the global cancer spectrum, with an incidence and mortality rate of 11.4% and 18% respectively, which has caused a heavy socioeconomic burden. Pulmonary nodules generally have no clinical symptoms, and are mainly manifested by lung CT nodule shadows. The pathogenesis is complex, mainly involving inflammatory response, abnormal cell proliferation, oxidative stress, DNA damage, cell apoptosis, increased diversity of respiratory flora, gene mutation, etc. The diagnosis and treatment guidelines for pulmonary nodules mainly distinguish between high- and medium-risk nodules, and use a diameter of 8mm as the limit of the treatment strategy, that is, those with a nodule diameter of less than 8mm are mainly followed up regularly, and those who find that the nodules are enlarged (diameter ≥8mm) or have malignant signs after reexamination of lung computed tomography (CT) are surgically removed. The repeated overdiagnosis and treatment will continue to increase the psychological and economic burden on patients. Therefore, it is of great significance to carry out clinical and basic research on pulmonary nodules and explore effective treatment measures.

[0004] In recent years, a large number of clinical studies have confirmed that traditional Chinese medicine has obvious advantages in treating lung nodules. It can shrink nodules to a certain extent, reduce nodule density, promote the absorption of inflammatory lesions, reduce the probability of malignant transformation, relieve patients' anxiety, and reduce psychological pressure and economic burden. At present, there is an urgent need to develop a traditional Chinese medicine composition for the prevention or treatment of lung nodules. Summary of the invention

[0005] In view of the above-mentioned defects of the prior art, the present invention provides a traditional Chinese medicine composition for preventing or treating lung nodules, which is composed of ginseng, gynostemma pentaphyllum, pinellia tuber, thunbergia thunbergii, peony bark, selfheal, vinegar-curd zedoaria, coix seed, stir-fried immature bitter orange, tangerine peel, roasted epimedium, and roasted licorice, which is beneficial for shrinking nodules, improving lung inflammation, and alleviating nodule progression.

[0006] The compatibility principles in the prescription of the present invention are as follows:

[0007] (1) Immature Bitter Orange stir-fried with bran and Thunberg Fritillary Bulb are the sovereign drugs. Immature Bitter Orange promotes qi movement to resolve phlegm-dampness, reduces accumulation and dredges stuffiness. After stir-frying with bran, its nature becomes milder, and the power of promoting qi movement and relieving stuffiness remains undiminished. Thunberg Fritillary Bulb is good at clearing and resolving heat-phlegm, descending and dispersing the lung qi, resolving phlegm and dissipating nodules. They are the sovereign drugs together.

[0008] (2) Coix Seed, Ginseng, Pinellia Tuber, Rhizoma Curcumae Aromaticae, and Tangerine Peel are the ministerial drugs together. Coix Seed promotes diuresis, percolates dampness, and strengthens the spleen, helping to eliminate internal dampness, and can also detoxify and dissipate nodules. Ginseng gently supplements the qi of various zang-organs, tonifies the lung and spleen, helps Immature Bitter Orange and Thunberg Fritillary Bulb to regulate qi movement, transform and transport phlegm-dampness, and also alleviates the bitter-cold nature of the sovereign drugs; Tangerine Peel promotes qi movement with pungency and warmth, dredges qi mechanism, disperses the lung qi and relieves cough, dries dampness and resolves phlegm, helping the sovereign drugs to regulate qi movement and resolve phlegm-dampness; Pinellia Tuber dries dampness and resolves phlegm, dissipates fullness and nodules. When combined with Tangerine Peel, qi movement is smooth and phlegm will naturally be eliminated; Rhizoma Curcumae Aromaticae breaks blood, promotes qi movement, reduces accumulation, activates blood circulation and dredges collaterals, dissipates masses and nodules.

[0009] (3) Gynostemma Pentaphyllum, Cortex Moutan, Epimedium Sagittatum, and Spica Prunellae are the adjuvant drugs together. Gynostemma Pentaphyllum benefits qi and strengthens the spleen, reduces phlegm and relieves cough, assisting Ginseng to supplement the qi of the lung and spleen, and assisting Thunberg Fritillary Bulb, Tangerine Peel, and Pinellia Tuber to relieve the concern of phlegm-dampness; Cortex Moutan dissipates stagnant qi, when combined with Rhizoma Curcumae Aromaticae, they jointly break the binding of phlegm and stasis, to achieve the state of blood stasis, and reach the effect of dissipating nodules and dredging collaterals; Epimedium Sagittatum warms the kidney and boosts yang, taking the effect of generating qi with gentle fire, to assist in regulating the qi mechanism of the triple energizer; Spica Prunellae clears heat and purges fire, softens hardness and dissipates nodules. When combined with Thunberg Fritillary Bulb and Pinellia Tuber, phlegm and stasis are cleared and nodules are dissipated.

[0010] (4) Prepared Licorice Root is the guiding drug. Prepared Licorice Root enters the lung meridian, benefits the lung and relieves cough, and harmonizes all the drugs as the guiding drug.

[0011] The whole formula together exerts the effects of promoting qi movement, removing dampness, resolving phlegm and dissipating nodules, activating blood circulation and reducing accumulation.

[0012] On the one hand, the present invention provides a traditional Chinese medicine composition, comprising the following traditional Chinese medicine raw materials by weight parts or consisting of them: Ginseng 1 - 20 parts, Gynostemma Pentaphyllum 6 - 30 parts, Pinellia Tuber 3 - 30 parts, Thunberg Fritillary Bulb 2 - 20 parts, Cortex Moutan 3 - 30 parts, Spica Prunellae 3 - 30 parts, Rhizoma Curcumae Aromaticae 3 - 30 parts, Coix Seed 5 - 50 parts, Immature Bitter Orange stir-fried with bran 3 - 20 parts, Tangerine Peel 3 - 30 parts, Epimedium Sagittatum 1 - 30 parts, Prepared Licorice Root 3 - 20 parts.

[0013] In some embodiments, the traditional Chinese medicine composition comprises the following traditional Chinese medicine raw materials by weight parts or consists of them: Ginseng 1 - 11 parts, Gynostemma Pentaphyllum 10 - 20 parts, Pinellia Tuber 7 - 17 parts, Thunberg Fritillary Bulb 4 - 13 parts, Cortex Moutan 7 - 17 parts, Spica Prunellae 7 - 17 parts, Rhizoma Curcumae Aromaticae 7 - 17 parts, Coix Seed 15 - 25 parts, Immature Bitter Orange stir-fried with bran 7 - 17 parts, Tangerine Peel 7 - 17 parts, Epimedium Sagittatum 1 - 11 parts, Prepared Licorice Root 4 - 13 parts.

[0014] In some embodiments, the traditional Chinese medicine composition comprises the following traditional Chinese medicine raw materials by weight parts or consists of them: ginseng 4.8 - 7.2 parts, gynostemma pentaphyllum 12 - 18 parts, pinellia ternata 9.6 - 14.4 parts, fritillaria thunbergii 7.2 - 10.8 parts, cortex moutan 9.6 - 14.4 parts, prunella vulgaris 9.6 - 14.4 parts, zedoary turmeric 7.2 - 10.8 parts, coix seed 16 - 24 parts, stir-fried fructus aurantii immaturus 7.2 - 10.8 parts, dried tangerine peel 9.6 - 14.4 parts, epimedium 4.8 - 7.2 parts, roasted licorice root 7.2 - 10.8 parts.

[0015] In some embodiments, the traditional Chinese medicine composition comprises the following traditional Chinese medicine raw materials by weight parts or consists of them: ginseng 5.5 - 6.5 parts, gynostemma pentaphyllum 14 - 16 parts, pinellia ternata 11 - 13 parts, fritillaria thunbergii 8 - 10 parts, cortex moutan 11 - 13 parts, prunella vulgaris 11 - 13 parts, zedoary turmeric 8 - 10 parts, coix seed 18 - 22 parts, stir-fried fructus aurantii immaturus 8 - 10 parts, dried tangerine peel 11 - 13 parts, epimedium 5.5 - 6.5 parts, roasted licorice root 8 - 10 parts.

[0016] In some embodiments, the traditional Chinese medicine composition comprises the following traditional Chinese medicine raw materials by weight parts or consists of them: ginseng 6 parts, gynostemma pentaphyllum 15 parts, pinellia ternata 12 parts, fritillaria thunbergii 9 parts, cortex moutan 12 parts, prunella vulgaris 12 parts, zedoary turmeric 9 parts, coix seed 20 parts, stir-fried fructus aurantii immaturus 9 parts, dried tangerine peel 12 parts, epimedium 6 parts, roasted licorice root 9 parts.

[0017] On the other hand, the present invention provides a traditional Chinese medicine preparation obtained from the above traditional Chinese medicine composition.

[0018] In some embodiments, the traditional Chinese medicine preparation may include granule, tablet, capsule, pill, powder, syrup, oral solution, mixture, decoction paste, fluid extract, extract, etc.

[0019] Still on the other hand, the present invention provides a method for preparing an extract from the above traditional Chinese medicine composition, comprising the following steps:

[0020] The above raw materials are decocted with 10 - 12 times the amount of water for 2 times, 1 hour each time, filtered, concentrated, and refrigerated for later use.

[0021] In some embodiments, the traditional Chinese medicine preparation can be prepared by methods well-known in the art.

[0022] In particular, in some embodiments, the granule can be prepared by a method comprising the following steps: the traditional Chinese medicine raw materials of the traditional Chinese medicine composition are decocted with water for 2 times, 1 hour each time, the decoction liquids are combined, filtered, the filtrate is concentrated to an extract, auxiliary materials are added, granules are made, and dried to obtain.

[0023] In some embodiments, the tablets can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water for 1 hour each time, combining the decoctions, filtering, concentrating the filtrate to an extract, adding excipients, making into granules, drying, tabletting, and coating to obtain the tablets.

[0024] In some embodiments, the capsules can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water for 1 hour each time, combining the decoctions, filtering, concentrating the filtrate to an extract, drying, mixing the dried extract powder with excipients, sieving, and filling into capsules to obtain the capsules.

[0025] In some embodiments, the pills can be prepared by a method comprising the following steps: pulverizing the traditional Chinese medicine raw materials of the traditional Chinese medicine composition into fine powder, sieving, mixing to obtain powder; making the powder into water pills by using water and drying; or making the powder into honeyed water pills by using refined honey and water and drying; or making the powder into big honey pills or small honey pills by adding refined honey to obtain the pills.

[0026] In some embodiments, the powders can be prepared by a method comprising the following steps: pulverizing the traditional Chinese medicine raw materials of the traditional Chinese medicine composition into fine powder, sieving, and mixing to obtain the powders.

[0027] In some embodiments, the syrups can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water for 1 hour each time, combining the decoctions, filtering, concentrating the filtrate, adding appropriate amount of sucrose and preservatives, boiling to dissolve, filtering, adding water to an appropriate amount, and stirring evenly to obtain the syrups.

[0028] In some embodiments, the oral solutions can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water for 1 hour each time, combining the decoctions, standing, filtering, concentrating the filtrate, adding a preservative (sodium benzoate), adding water to an appropriate amount, stirring evenly, and filling into containers to obtain the oral solutions.

[0029] In some embodiments, the mixtures can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water for 1 hour each time, combining the decoctions, filtering, concentrating the filtrate to an extract, adding excipients such as a preservative (sodium benzoate) and sucrose, adding water to an appropriate amount, filtering, and sealing, or sealing and sterilizing to obtain the mixtures.

[0030] In some embodiments, the extract pastes can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water for 1 hour each time, combining the decoctions, filtering, concentrating the filtrate to a clear extract, adding sucrose (or honey) to the clear extract, heating and melting, mixing evenly, and concentrating to obtain the extract pastes.

[0031] In some embodiments, the fluid extract can be prepared by a method comprising the following steps: using the traditional Chinese medicine raw materials of the traditional Chinese medicine composition, using 70% ethanol as a solvent, impregnating for 24 hours and then performing percolation, collecting the initial percolate, storing it separately, continuing percolation until all soluble components are completely percolated out, concentrating the subsequent percolate to a thick paste below 60°C, adding the initial percolate, mixing evenly, diluting with 70% ethanol to 1000 ml, standing, and filtering to obtain the product.

[0032] In some embodiments, the extract can be prepared by a method comprising the following steps: decocting the traditional Chinese medicine raw materials of the traditional Chinese medicine composition twice with water, each time for 1 hour, combining the decoction liquids, filtering, and concentrating the filtrate under reduced pressure to a thick paste; or drying the extract, pulverizing, and sieving to obtain extract powder.

[0033] On the other hand, the present invention provides the use of the above traditional Chinese medicine composition or traditional Chinese medicine preparation in the preparation of a drug for preventing or treating pulmonary nodules.

[0034] In the present invention, the pulmonary nodules refer to inflammatory nodules, ground-glass nodules, and carcinoma in situ.

[0035] In some embodiments, the pulmonary nodules are inflammatory nodules.

[0036] In some embodiments, the pulmonary nodules are pulmonary ground-glass nodules, especially pure pulmonary ground-glass nodules.

[0037] In some embodiments, the pulmonary nodules are carcinoma in situ.

[0038] On another hand, the present invention provides a method for preventing or treating pulmonary nodules, comprising the step of administering to a subject in need thereof a prophylactically or therapeutically effective amount of the traditional Chinese medicine composition or traditional Chinese medicine preparation of the present invention.

[0039] As used herein, the term "prophylactically or therapeutically effective amount" means an amount that has a prophylactic or therapeutic effect and can be used to prevent or treat the specific diseases, disorders, or conditions described herein. For example, a "prophylactically or therapeutically effective amount" may refer to the amount required to provide a prophylactic or therapeutic or desired effect in an individual receiving treatment. As is known to those skilled in the art, the prophylactically or therapeutically effective amount may vary depending on the route of administration, the use of excipients, and the possibility of co-therapy with other treatments.

[0040] The raw materials of the present invention are rich, the formula is scientific and advanced, and it is effectively used for preventing or treating pulmonary nodules. It can be developed into a new drug for treating pulmonary nodules, and has high economic and social benefits. BRIEF DESCRIPTION OF THE DRAWINGS

[0041] Figure 1 It is a flowchart showing the experiment of the effect of Yifei Sanjie Prescription in intervening in pulmonary nodule mice induced by B(a)p combined with LPS in Example 2.

[0042] Figure 2 Photographs showing the pulmonary nodule appearance in each group of mice (benzo[a]pyrene combined with LPS-induced nodule model).

[0043] Figure 3 Imaging diagrams showing the CT conditions of the lungs of each group of mice (benzo[a]pyrene combined with LPS-induced nodule model).

[0044] Figure 4 HE staining section diagrams showing the pulmonary tissue pathology (HE, ×200) of each group of mice (benzo[a]pyrene combined with LPS-induced nodule model).

[0045] Figure 5 Immunohistochemical diagrams showing the CD68 expression in the pulmonary tissue of each group of mice (IHC, ×200) (benzo[a]pyrene combined with LPS-induced nodule model).

[0046] Figure 6 and 7 Imaging diagrams showing the comparison of CT conditions before and after 2-year follow-up of two patients with pulmonary nodules in Example 3. Figure 6 Among them, the shooting time of the upper left picture is August 20, 2022, the shooting time of the upper middle picture is April 17, 2023, and the shooting time of the upper right picture is March 26, 2024. Figure 7 Among them, the shooting time of the upper left picture is August 30, 2021, the shooting time of the upper middle picture is February 6, 2022, and the shooting time of the upper right picture is January 4, 2023.

[0047] Figure 8 Imaging diagrams showing the CT conditions of the lungs of each group of mice in Example 4 (the upper row is the CT imaging diagram, and the lower row is the CT imaging 3D diagram) (Kras gene mouse induction model).

[0048] Figure 9 Photograph diagrams showing the pulmonary appearance of each group of mice (Kras gene mouse induction model).

[0049] Figure 10 HE staining section diagrams showing the pulmonary tissue pathology (HE, ×200) of each group of mice (Kras gene mouse induction model).

[0050] Figure 11 Immunohistochemical diagrams showing the Ki-67 expression in the pulmonary tissue of each group of mice (IHC, ×200) (Kras gene mouse induction model). Detailed implementation manners

[0051] In the following, the present invention will be described in detail through examples. However, the examples provided herein are for illustrative purposes only and do not limit the present invention.

[0052] Unless otherwise specified, the experimental methods used in the following examples are all conventional methods.

[0053] Unless otherwise specified, the materials, reagents, etc. used in the following examples are all commercially available.

[0054] Example

[0055] Example 1: Preparation of Yifei Sanjie Fang preparation

[0056] Traditional Chinese medicine raw materials: Ginseng, Pinellia ternata, Fritillaria thunbergii, Paeonia suffruticosa, Prunella vulgaris, Rhizoma Curcumae, Coix lacryma-jobi, Fructus Aurantii Immaturus, Pericarpium Citri Reticulatae, and Radix Glycyrrhizae Preparata were provided by Zhengzhou Ruilong Pharmaceutical Co., Ltd., Epimedium brevicornu Preparata was provided by Yuzhou Kaixuan Pharmaceutical Co., Ltd., and Gynostemma pentaphyllum was provided by Anguo Runde Pharmaceutical Co., Ltd.

[0057] Preparation method: 6 parts of ginseng, 15 parts of Gynostemma pentaphyllum, 12 parts of Pinellia ternata, 12 parts of Prunella vulgaris, 20 parts of Coix lacryma-jobi, 9 parts of Radix Glycyrrhizae Preparata, 9 parts of Fritillaria thunbergii, 12 parts of Paeonia suffruticosa, 9 parts of Rhizoma Curcumae, 9 parts of Fructus Aurantii Immaturus, 12 parts of Pericarpium Citri Reticulatae, and 6 parts of Epimedium brevicornu Preparata were added with 10 - 12 times the amount of water and decocted twice, each time for 1 hour. The decoctions were combined, filtered, and the filtrate was concentrated under reduced pressure to a thick paste to obtain an extract; or the extract was dried, pulverized, and sieved to obtain an extract powder.

[0058] Example 2: Effect of Yifei Sanjie Fang on lung nodule mice induced by B(a)p combined with LPS

[0059] 1. Experimental materials

[0060] 1.1 Animals

[0061] 90 SPF-grade C57BL / 6J mice, half male and half female (age: 6 - 8 weeks; animal quality certificate number: 110324221105785761), were purchased from Henan Chengxin Youkang (Beijing Sibefu) Company (SCXK(Beijing)2019 - 0010).

[0062] 1.2 Drugs

[0063] ① Benzo[a]pyrene (B(a)p, CAS No.50 - 32 - 8) was provided by Sigma - Aldrich Company.

[0064] ② Lipopolysaccharide (LPS, CAS No.93572 - 42 - 0) was provided by Sigma - Aldrich Company.

[0065] ③ Glycerol trioctanoate (CAS No.538 - 23 - 8) was provided by Sigma - Aldrich Company.

[0066] ④ Yifei Sanjie Fang: Prepared in Example 1.

[0067] 1.3 Reagents:

[0068] The IL-6 (M0044c) ELISA kit was provided by Elabscience Biotechnology Co., Ltd.; the TNF-α (M3063) ELISA kit was provided by Elabscience Biotechnology Co., Ltd.; CD68 (DF-7518) was provided by Affinity Bioscience.

[0069] 1.4 Instruments:

[0070] Small animal Micro CT in vivo imaging system (Pingsheng Medical Technology Co., Ltd., Kunshan); slide scanner (BUXCO, USA); 3D Histech digital slide scanning and analysis system (3D Histech, Hungary); Multiskan GO full wavelength microplate reader (Thermo Scientific, USA), etc.

[0071] 2. Experimental methods

[0072] Figure 1 It is the experimental flow chart showing the effect of Yifei Sanjie Prescription on intervening in B(a)p combined with LPS-induced lung nodule mice.

[0073] 2.1 Model preparation:

[0074] After the mice were purchased, they were adaptively fed for 7 days. The feeding conditions were: room temperature (25±1) °C, relative humidity (50±10)%, air change rate 10 - 15 times / h, ammonia concentration ≤ 14 mg / m 3 , noise ≤ 60 db. They were fed with sterilized feed and freely drank sterilized water. The purification operating system was regularly inspected to keep the environment quiet. The mice in the model group were instilled with 50 μl of B(a)p (40 mg / kg, dissolved in glyceryl trioctanoate) through the trachea once a week for 4 consecutive weeks; starting from the 6th week, 50 μl of LPS (2.5 μg / kg, dissolved in sterile water) was instilled through the trachea once every two weeks for 5 consecutive times; the blank control group was instilled with an equal volume of glyceryl trioctanoate and sterile water. The operation method was that after the mice were anesthetized by inhaling isoflurane (1 - 3%), the upper incisors of the mice were hung with a silk thread to make them hang naturally on a self-made plastic rack. The left hand pulled out the tongue of the mice, and the right hand held a pipette gun to inject the prepared compound into the mouth. At this time, the tongue was continuously pulled, and the nasal cavity of the mice was gently pinched for about 5 - 10 seconds to make them actively inhale the instilled liquid into the lungs through breathing.

[0075] 2.2 Grouping and administration:

[0076] Ninety mice were randomly divided into a normal group (10 mice), a model group (20 mice), a high-dose Yifei Sanjie formula group (20 mice), a medium-dose Yifei Sanjie formula group (20 mice), and a low-dose Yifei Sanjie formula group (20 mice). Corresponding drug interventions were carried out from 21 to 36 weeks. The blank group and the model group were given pure water (25 mL / kg / d) by gavage, and the high-dose group, medium-dose group, and low-dose group were given suspensions of different doses of the Yifei Sanjie formula dissolved in pure water (34.06 mL / kg / d; 17.03 mL / kg / d; 8.52 mL / kg / d) by gavage. The body weight was measured weekly to adjust the drug dosage, and the drug dosage was calculated using the equivalent dose coefficient conversion formula. The formula is Drat = Dhuman × (Hirat / Hihuman) × (Wrat / Whuman). 2 / 3 . D: dosage, HI: body size coefficient, W: body weight. At the 36th week, the mice were sacrificed after analyzing the formation of lung nodules by Micro-CT scanning, and lung tissue, blood, etc. were taken.

[0077] 2.3 Specimen collection and processing

[0078] 2.3.1 General conditions:

[0079] The mental state, activity, fur, food and water intake, and body weight of the mice were observed.

[0080] 2.3.2 Micro-CT:

[0081] Micro-CT scanning was used to analyze the formation of lung nodules in mice of each group at 36 weeks. Before the formal scan, the mice were anesthetized by respiratory anesthesia with 2% isoflurane and 98% oxygen, and the thoracic cavity field of view was selected for CT scanning. The scanning parameters were a voltage of 60 kV, a current of 400 μA, a 360° rotational scan, an angular gain of 1°, a continuous rotational scan of 284 s, an effective voxel of 10.34 μm, a field of view of 10.58 mm × 10.58 mm, and the scanning time for each mouse was about 5 min. After the scan was completed, the overall structure reconstruction was completed using the software attached to the system. The 3D Slicer software was used for image analysis. The maximum long-axis diameter (A) of each nodule was measured in the transverse plane, and the maximum short-axis diameter (B) measured perpendicular to the long axis in the same plane was measured. The average diameter of each nodule (d = (A + B) / 2) and the nodule area (S = A × B) were calculated.

[0082] 2.3.3 Lung coefficient:

[0083] After the lung was completely removed from the mouse, the wet weight of the lung was weighed. The calculation formula is: lung coefficient = wet weight of lung (mg) / body weight (g) × 100%.

[0084] 2.3.4 Lung appearance:

[0085] Place the intact mouse lung tissue on a white bottom plate, observe the lung nodules under a magnifying glass, take pictures and count them.

[0086] 2.3.5 Lung tissue pathology:

[0087] The left lung of the mouse was perfused with 4% paraformaldehyde and fixed for 72 h, and the formaldehyde fixative was changed every 24 h. After the tissue was completely fixed, tissue blocks with a thickness of about 3 mm were cut for paraffin embedding. The embedded wax blocks were fixed on a microtome and thin sections with a thickness of 4 μm were cut for hematoxylin and eosin staining (HE). Pathological scoring was performed on the lung tissue, and the scoring criteria for alveolitis are shown in Table 1. Classification basis for the pathological types of lung tissue: The microscopic lesions were divided into inflammation and AAH. Inflammation showed thickening of the alveolar wall with obvious rupture and fusion, mainly with an increase in macrophages and lymphocytes. AAH was focal and diffuse lesions involving alveoli and terminal bronchioles, consisting of relatively uniform atypical cubic to columnar cells, with dense chromatin, cellular and nuclear atypia.

[0088] Table 1 Grading and scoring criteria for alveolitis

[0089]

[0090] 2.3.6 CD68, a surface marker of macrophages in lung tissue:

[0091] The expression of cluster of differentiation 68 (CD68) in lung tissue was detected by immunohistochemistry (IHC), and the concentration of the primary antibody was 1:500; the integral optical density (IOD) of the image was calculated using the Image-Pro Plus 6.0 professional image acquisition and analysis system.

[0092] 2.3.7 IL-6 and TNF-α in BALF:

[0093] The expressions of IL-6 and TNF-α in BALF were detected by enzyme-linked immunosorbent assay (ELISA).

[0094] 2.4 Statistical analysis:

[0095] Statistical analysis was performed using SPSS 22.0. One-way analysis of variance (One-Way ANOVA) was used for between-group comparisons. For those with homogeneous variances, the least significant difference method was used, and for those with heterogeneous variances, Dunnett's T3 method was used. The results are presented as the mean value. The significance level was set at α = 0.05.

[0096] 3. Experimental results

[0097] 3.1 General conditions

[0098] The mice in the blank group were in good spirits, had normal food and water intake, were active, responsive, and had smooth and shiny hair. After induction with B(a)p combined with LPS, the mice in the model group had poor spirits, reduced food and water intake, less activity, were unresponsive, had dull hair, and were lethargic and preferred to lie down. After drug intervention, the above symptoms of the mice in the different-dose traditional Chinese medicine groups were improved to varying degrees, and the mental state of the low- and medium-dose groups was significantly better than that of the high-dose group.

[0099] 3.2 Lung appearance

[0100] Compared with the blank group, the nodule rate and the number of nodules in the lungs of the mice in the model group were significantly increased (P < 0.01); compared with the model group, the nodule rates of the different-dose traditional Chinese medicine groups were all decreased, and the low- and medium-dose groups were more obvious; the number of nodules in the low- and medium-dose groups was also decreased ( Figure 2 , Table 2).

[0101] Table 2 Lung nodule conditions of mice in each group

[0102]

[0103] Note: n = 10 - 19, n is the number of mice; compared with the blank group, P a < 0.05, P aa < 0.01.

[0104] 3.3 Lung coefficient

[0105] Compared with the blank group, the lung coefficient of the mice in the model group was significantly increased (P < 0.01); compared with the model group, the lung coefficients of the low- and medium-dose groups were significantly decreased (P < 0.05) (Table 3).

[0106] Table 3 Lung coefficient conditions of mice in each group

[0107]

[0108]

[0109] Note: n = 10 - 19, n is the number of mice. Compared with the blank group, P a< 0.05, P aa < 0.01; Compared with the model group, P b < 0.05, P bb < 0.01.

[0110] 3.4 Lung CT

[0111] No abnormalities were seen in the lung CT of the mice in the blank group, the texture was clear, and no nodular shadows were seen; nodular shadows were partially visible in the mice in the model group, and the lung texture was thickened ( Figure 3 ), and the nodule rate, the number of nodules, the nodule diameter and the area were all significantly increased (P < 0.01); compared with the model group, the nodule diameter in the high-dose group was significantly decreased (P < 0.05), the number of nodules, the nodule diameter and the area in the medium-dose group were all significantly decreased (P < 0.05), and the number of nodules, the nodule diameter and the area in the low-dose group were all significantly lower (P < 0.01, P < 0.05); the number of nodules in the low-dose group was significantly lower than that in the high-dose group (P < 0.05) (Table 4).

[0112] Table 4 Lung CT conditions of mice in each group

[0113]

[0114] Note: n = 10 - 19, n is the number of mice. Compared with the blank group, P a < 0.05, P aa < 0.01; compared with the model group, P b < 0.05, P bb < 0.01; compared with the high-dose group, P c < 0.05, P cc < 0.01.

[0115] 3.5 Lung tissue pathology

[0116] The alveolar structure of the mice in the blank group was intact, no alveolar inflammation was seen, and the alveolar wall was not thickened. The alveolar structure of the mice in the model group was damaged, with obvious fracture and fusion phenomena, alveolar inflammatory infiltration was visible, and the alveolar interstitium was thickened. After drug intervention, the inflammatory infiltration in the Chinese medicine groups with different doses was reduced ( Figure 4 ).

[0117] Through pathological scoring, it was found that the alveolitis score in the model group was significantly higher than that in the blank group (P < 0.01); the alveolitis scores in the Chinese medicine groups with different doses were all significantly reduced (P < 0.01, P < 0.05) (Table 5).

[0118] From the perspective of pathological types, compared with the blank group, the proportion of inflammation in the model group increased, and atypical adenomatous hyperplasia appeared; compared with the model group, the proportions of inflammation and atypical adenomatous hyperplasia in each administration group decreased after 16 weeks of Chinese medicine intervention (Table 6).

[0119] Alveolar inflammation scores of mice in each group in Table 5

[0120]

[0121]

[0122] Note: n = 10 - 19, where n is the number of mice. Compared with the blank group, P a < 0.05, P aa < 0.01; compared with the model group, P b < 0.05, P bb < 0.01.

[0123] Proportion of pathological types of mice in each group (%) in Table 6

[0124]

[0125] Note: n = 10 - 19, where n is the number of mice.

[0126] 3.6 CD68 expression

[0127] Compared with the blank group, the CD68 expression in the model group of mice was significantly increased (P < 0.01). Compared with the model group, the CD68 expression in the low-dose group was significantly decreased (P < 0.01), and was lower than that in the high-dose group and the medium-dose group (P < 0.01, P < 0.05)( Figure 5 , Table 7).

[0128] CD68 expression in lung tissues of mice in each group in Table 7

[0129]

[0130] Note: n = 6, where n is the number of mice. Compared with the blank group, P a < 0.05, P aa < 0.01; compared with the model group, P b < 0.05, P bb < 0.01; compared with the high-dose group, P c < 0.05, P cc < 0.01; compared with the medium-dose group, P d < 0.05, P dd < 0.01.

[0131] 3.7 Contents of IL-6 and TNF-α in bronchoalveolar lavage fluid (BALF)

[0132] Compared with the blank group, the levels of IL-6 and TNF-α in the BALF of mice in the model group were significantly increased (P<0.01); compared with the model group, the level of IL-6 in the medium and low dose groups was significantly decreased (P<0.05), and the levels of TNF-α in each dose group were significantly decreased (P<0.01) (Table 8).

[0133] Table 8 Expression of IL-6 and TNF-α in BALF of mice in each group

[0134]

[0135] Note: Compared with the blank group, P a <0.05, P aa <0.01; compared with the model group, P b <0.05, P bb <0.01.

[0136] 4. Conclusion

[0137] In summary, the traditional Chinese medicine composition of the present invention has a certain curative effect in the treatment of pulmonary nodules, can significantly reduce the nodule rate, nodule number and size of mice with pulmonary nodules, can improve alveolitis and lung coefficient of lung tissue, reduce the expression of CD68 in lung tissue and the contents of IL-6 and TNF-α in BALF, effectively relieve the progression of pulmonary nodules, and is an innovation in the long-term treatment of pulmonary nodules, with high practical value.

[0138] Example 3: Study on the intervention of Yifei Sanjie Prescription in pure ground-glass pulmonary nodules

[0139] 1. Research objects

[0140] 1.1 Source of cases: 780 patients with pure ground-glass nodules in the lungs of the Third Affiliated Hospital of Henan University of Traditional Chinese Medicine, the First Affiliated Hospital of Henan University of Traditional Chinese Medicine, Henan Provincial People's Hospital, Xinzheng People's Hospital, Shangcheng County Traditional Chinese Medicine Hospital, etc. from January 2021 to November 2024 were included.

[0141] 1.2 Diagnostic criteria

[0142] 1.2.1 Western medicine diagnostic criteria: Refer to the "Chinese Expert Consensus on the Diagnosis and Treatment of Pulmonary Nodules (2018 Edition)", the "Diagnosis and Treatment Consensus on Ground-Glass Nodules in Early Lung Adenocarcinoma of Shanghai Pulmonary Hospital (First Edition)", and the "Thoracic Surgery Expert Consensus on the Surgical Methods for Lung Cancer with Ground-Glass Nodules ≤2 cm Based on High-Resolution CT Imaging Guidance (2019 Edition)".

[0143] 1.2.2 Inclusion criteria: ① Patients diagnosed with pure ground-glass pulmonary nodules by thin-layer (slice thickness 1 mm) chest spiral CT scan; ② 18 < age < 80 years old; ③ Good communication and understanding ability; ④ Voluntarily sign the informed consent form.

[0144] 1.2.3 Exclusion criteria: ① patients with pure ground-glass nodules in the lungs who meet surgical indications; ② patients with metastatic lung nodules; ③ patients with diffuse interstitial lung lesions, such as interstitial pneumonia and pulmonary fibrosis; ④ patients with organ failure caused by diseases of various systems; ⑤ patients who are pregnant or lactating.

[0145] 1.2.4 Termination criteria: ① Patients whose nodules change and require biopsy or surgical resection; ② Patients with severe adverse drug reactions; ③ Patients who request surgical resection on their own initiative.

[0146] Post-termination measures: For those who meet termination criterion ①, a multidisciplinary consultation is required to decide on the next steps; for those who meet termination criterion ②, records should be kept truthfully and active measures should be taken immediately until the adverse reactions disappear; for those who meet termination criterion ③, the postoperative pathological results of the patients should be tracked.

[0147] 2. Study Design

[0148] 2.1 Study type: Prospective cohort study.

[0149] 2.2 Sample size estimation

[0150] This study was a cohort study, and the primary outcome measure was nodule growth rate. Previous literature reported that the two-year natural growth rate of ground-glass nodules in the lung was 13.5%, which was the growth rate of the non-exposed group; based on previous clinical observations, the two-year growth rate of ground-glass nodules in the lung after Chinese medicine intervention was expected to be 6.75%. Assuming the test level α = 0.05 (bilateral), the power 1-β = 0.80, and the exposure group: non-exposed group = 1:1, the PASS software was input for calculation, and the sample size was corrected according to the 20% loss rate. The estimated results showed that there were 390 cases in each of the exposed group and the non-exposed group, for a total of 780 cases.

[0151] 2.3 Queue Formation

[0152] 2.3.1 Exposure factors: Treatment with Yifei Sanjie prescription was taken as the exposure factor.

[0153] 2.3.2 Formation of cohorts: Two cohorts were naturally formed according to the patients' wishes. Follow-up plus Yifei Sanjie Fang was the Yifei Sanjie Fang cohort (exposed group); only follow-up was the follow-up cohort (non-exposed group).

[0154] 2.3.3 Cohort migration: If patients in the exposed group change their will and no longer continue the treatment with Yifei Sanjie Fang, this is not considered cohort migration and they should be grouped according to the patient's exposure intensity; if patients in the non-exposed group change their will and reselect Yifei Sanjie Fang for treatment, this is considered cohort migration, but the total follow-up time remains unchanged and they should be grouped according to the patient's exposure intensity. The division of exposure intensity should be combined with clinical practice.

[0155] The researchers should control the cohort migration, such as building the patients' confidence in treatment, strengthening the trust between doctors and patients, and truthfully recording the process and reasons for treatment changes. After the study is completed, it will be analyzed jointly by clinical experts and statistical experts.

[0156] 2.3.4 Cohort contamination: Patients in the exposure group may take other traditional Chinese medicines or patent medicines for treating pulmonary nodules without permission during the treatment process, which is cohort contamination. If it occurs, the researchers should truthfully record the drug composition, efficacy, administration time, etc., and continue the follow-up. After the study is completed, it will be analyzed jointly by clinical experts and statistical experts.

[0157] 3. Research protocol

[0158] Exposure group: Follow-up + Yifei Sanjie Granules, dosage: one bag each time, three times a day;

[0159] Yifei Sanjie Granules are uniformly prepared by the Preparation Room of the Third Affiliated Hospital of Henan University of Traditional Chinese Medicine, with independent small packages. Preparation method: Add 6 parts of ginseng, 15 parts of gynostemma pentaphyllum, 12 parts of Pinellia ternata, 12 parts of Prunella vulgaris, 20 parts of coix seed, 9 parts of roasted licorice root, 9 parts of fritillaria thunbergii, 12 parts of moutan cortex, 9 parts of zedoary turmeric processed with vinegar, 9 parts of immature bitter orange stir-fried with bran, 12 parts of tangerine peel, and 6 parts of epimedium processed with honey, add 10 - 12 times the amount of water and decoct twice, 1.5 hours each time. Filter, and concentrate the filtrate under reduced pressure to a clear paste with a relative density of about 1.25 - 1.35 (60 - 70 °C). Add appropriate amounts of dextrin, xylitol and the above-mentioned fine powder, make granules with 80% ethanol, dry, and package (8 g / bag) to obtain the product.

[0160] Non-exposure group: Follow-up.

[0161] 4. Follow-up

[0162] 4.1 Follow-up content: General items, traditional Chinese medicine syndromes, medication conditions, drug adverse reactions, adverse events, thin-layer chest spiral CT (slice thickness 1 mm) scan, Self-Rating Anxiety Scale, Self-Rating Depression Scale.

[0163] 4.2 Follow-up time points:

[0164] The follow-up time points are formulated according to the inclusion time and follow-up content. General items are followed up once at the time of inclusion; thin-layer chest spiral CT (slice thickness 1 mm) scan is followed up at the time of inclusion, 6 months, 12 months, and 24 months after inclusion; traditional Chinese medicine syndromes, medication conditions, drug adverse reactions, adverse events, Self-Rating Anxiety Scale, and Self-Rating Depression Scale are followed up at the time of inclusion and 3 months, 6 months, 9 months, 12 months, 18 months, and 24 months after inclusion. The time window is ±15 days.

[0165] To ensure the scientificity and rigor of this study and at the same time reduce the radiation dose received by patients, the project team stipulates that if a patient has undergone a thin-slice (slice thickness 1 mm) chest spiral CT scan within 1 month before being included in the study, there is no need for a re-scan at the time of inclusion and the patient can directly enter this study; otherwise, a re-scan is required to enter this study.

[0166] 4.3 Follow-up termination:

[0167] There are two cases: ① If the termination criteria occur during the study, the follow-up is terminated; ② If the termination criteria do not occur during the study, the follow-up should be completed according to the follow-up time points.

[0168] 4.4 Follow-up method:

[0169] The researchers inform and make an appointment for the follow-up time. After completing the follow-up, the researchers record and analyze the results and truthfully fill in the case observation form. If all the results are normal, the time for the next follow-up is determined; if the results are abnormal, the researchers inform in a timely manner and take further treatment measures.

[0170] 5. Outcome measures

[0171] 5.1 Primary outcome measure: Nodule growth rate.

[0172] 5.2 Secondary outcome measures ① The maximum transverse diameter of the nodule, ② The volume of the nodule.

[0173] 5.3 Drug adverse reactions / adverse events

[0174] 5.3.1 Adverse drug reactions:

[0175] Adverse drug reactions refer to harmful rather than expected reactions that occur during the normal use of a drug at the prescribed dose and are causally related to the use of the drug, and should be recorded at any time. The expected adverse drug reactions in this study include: dry throat, thirst, stomach discomfort, decreased appetite, etc.

[0176] 5.3.2 Adverse events:

[0177] An adverse event refers to any adverse medical event that occurs after a subject receives a drug, but it is not necessarily causally related to the drug used. Adverse events may be: a new disease; or deterioration of a concomitant disease; the effect of a control drug; unrelated to participation in the trial; a combination of one or more factors. A serious adverse event is an adverse event that occurs at any dose of the investigational drug or at any time during the observation period, including: resulting in death; immediately life-threatening, requiring hospitalization or prolonging hospitalization; resulting in permanent or severe disability; causing cancer; having important medical significance (referring to those events that do not immediately endanger life or cause death or require hospitalization, but may harm the patient or require measures to prevent one of the consequences defined above); requiring medical treatment to prevent permanent injury or damage. It should be recorded truthfully.

[0178] 6. Data Statistics and Analysis

[0179] 6.1 Establishment of the database Based on the above work, a data collection platform and a management platform are established. Data entry is carried out by two independent persons. Problems found during the data entry process need to be registered, reported, and processed in a timely manner. After the data entry is completed, some CRF forms are randomly checked to ensure the data quality, and then the data administrator performs data conversion, cleaning, and locking, and hands it over to the statisticians for statistics.

[0180] 6.2 Statistics and Analysis The statistical software selected is SPSS 21.0 software. Qualitative data is analyzed using χ 2 test or CMH test, etc.; quantitative data is analyzed using t-test, Wilcoxon rank sum test, or repeated measures analysis of variance (ANOVA), etc. All statistical tests use two-sided tests, and P < 0.05 indicates that the difference is statistically significant.

[0181] 7. Research Results

[0182] 7.1 Nodule Growth Rate

[0183] Growth definition: ① The volume increases by ≥25% compared with the previous one; ② The appearance of a solid component during follow-up or the CT value increases by ≥30% compared with the previous one; ③ The appearance of malignant signs.

[0184] After 2 years of follow-up, among 398 patients in the traditional Chinese medicine group, 27 cases (6.8%) had nodule growth, and among 382 patients in the non-traditional Chinese medicine group, 48 cases (12.6%) had nodule growth. Comparison between the two groups showed that the nodule growth rate in the traditional Chinese medicine group was significantly lower than that in the non-traditional Chinese medicine group (P < 0.05), as shown in Table 9.

[0185] Table 9 Comparison of nodule growth rates between the two groups (%)

[0186]

[0187] 7.2 Nodule Volume

[0188] Before follow-up, there was no significant difference in the nodule volume between the two groups (P>0.05). After 2 years of follow-up, the nodule volume in the traditional Chinese medicine group decreased slightly compared with that before follow-up, but the difference was not statistically significant (P>0.05); the nodule volume in the non-traditional Chinese medicine group increased significantly compared with that before follow-up, and the difference was statistically significant (P<0.001); there was no statistically significant difference in the nodule volume between the two groups (P>0.05). Comparing the differences in nodule volume before and after follow-up between the two groups, the difference was statistically significant (P<0.001). See Table 10.

[0189] Table 10 Comparison of nodule volume between two groups (mm 3 )

[0190]

[0191] Note: a: Comparison between two groups; b: Comparison before and after follow-up of two groups.

[0192] 7.3 Comparison of the maximum diameter of nodules

[0193] Before follow-up, there was no significant difference in the maximum diameter of nodules between the two groups (P>0.05). After 2 years of follow-up, the maximum diameter of nodules in the traditional Chinese medicine group decreased compared with that before follow-up, and the difference was statistically significant (P<0.05); the maximum diameter of nodules in the non-traditional Chinese medicine group increased compared with that before follow-up, and the difference was statistically significant (P<0.05); there was no statistically significant difference in the maximum diameter of nodules between the two groups (P>0.05). Comparing the differences in the maximum diameter of nodules before and after follow-up between the two groups, the difference was not statistically significant (P>0.05). See Table 11.

[0194] Table 11 Comparison of the maximum diameter of nodules between two groups (mm)

[0195]

[0196] Note: a: Comparison between two groups; b: Comparison before and after follow-up of two groups.

[0197] 8. Summary

[0198] Figure 6 and 7 are imaging diagrams showing the contrast CT images of two patients with pulmonary nodules before and after 2 years of follow-up. Figure 6 is the comparison of CT images of the patient at 7 months and 1.5 years of treatment. The nodule volume gradually decreased, and the nodule shrank by 34% at 1.5 years. Figure 7 is the comparison of the images of the patient at 6 months and 1.5 years of treatment. The nodule shrank by 53% at 1.5 years of treatment. Yifei Sanjie Recipe can reduce the growth rate of ground-glass nodules, shrink the nodule volume and size, and promote the regression of nodules, showing good curative effects.

[0199] Example 4: Effect of Yifei Sanjie Recipe on Kras mutant mouse models of in situ lung cancer

[0200] 1. Experimental materials

[0201] 1.1 Animals:

[0202] Six SPF-grade LSL-K-ras G12D mice (age: 34 - 41 days; animal quality certificate number: 320818220100231857) and six Sftpc-MerCreMer mice (age: 41 days; animal quality certificate number: 320981230100072185) were purchased from Cyagen Biosciences (Suzhou) Inc.

[0203] 1.2 Drugs:

[0204] ① Tamoxifen (CAS: 10540-29-1) was provided by Sigma-Aldrich.

[0205] ② Corn oil (Cat: IC9000) was provided by Solarbio.

[0206] ③ Yifei Sanjie Formula: The preparation method was the same as that in Example 1.

[0207] 1.3 Reagents:

[0208] One Step Mouse Genotyping Kit (PD101) was provided by Novoprotein; Ki-67 (GB111141) was provided by Wuhan Sevier.

[0209] 1.4 Instruments:

[0210] Small animal Micro CT in vivo imaging system (Pingsheng Medical Technology Co., Ltd., Kunshan);

[0211] Slide scanner (BUXCO, USA); 3D Histech digital slide scanning and analysis system (3D Histech, Hungary).

[0212] 2. Experimental methods

[0213] 2.1 Establishment and identification of SPC-CRE-Kras double-positive mice:

[0214] The heterozygous LSL-K-ras G12D mice were crossed with homozygous Sftpc-MerCreMer mice to obtain SPC-CRE-Kras double-positive transgenic mice and their littermate negative mice for experiments. When the mice were 21 - 28 days old, their tails were cut off, and DNA was extracted using the OneStep Mouse Genotyping Kit and subjected to PCR amplification. Specific primers were used to identify the genotypes of the offspring transgenic mice. The upstream primer for Kras identification was 5’-GCAGGTCGAGGGACCTAATA-3’ (SEQ ID NO:1), and the downstream primer was 5’-CTGCATAGTACGCTATACCCTGT-3’ (SEQ ID NO:2). The upstream primer for Sftpc identification was 5’-AGGTGAGCAACAGTACCTTTCAG-3’ (SEQ ID NO:3), and the downstream primer was 5’-CCAAAAGACGGCAATATGGTGGAA-3’ (SEQ ID NO:4). The PCR reaction conditions were: pre-denaturation at 94°C for 3 min, denaturation at 94°C for 30 s, annealing at 62°C for 35 s, extension at 72°C for 35 s, with 33 amplification cycles, and the reaction system was 25 μL.

[0215] 2.2 Model establishment and drug administration:

[0216] Nineteen mice were divided into a normal group (7 mice), a model group (6 mice), and a Yifei Sanjie Recipe group (6 mice). Tamoxifen (75 mg / kg) was intraperitoneally injected to induce Kras gene mutation once a day for 5 consecutive days. After the last injection of tamoxifen, drug intervention was carried out. The blank group and the model group were given pure water (0.1 ml / 10 g) by gavage, and the traditional Chinese medicine group was given Yifei Sanjie Recipe (17.03 mL / kg / d) by gavage. At the 6th week, the mice were sacrificed after Micro-CT scanning of their lungs.

[0217] 2.3 Sample collection and processing

[0218] 2.3.1 General conditions:

[0219] The mental state, activities, fur, food and water intake of the mice were observed.

[0220] 2.3.2 Lung CT:

[0221] Micro-CT was used to scan the lungs of mice in each group. Before the formal scan, the mice were anesthetized with 2% isoflurane and 98% oxygen, and the chest field was selected for CT scanning. The scanning parameters were voltage 60kV, current 400μA, 360° rotation scanning, angle gain 1°, continuous rotation scanning 284s, effective voxel 10.34μm, field of view 10.58mm×10.58mm, and the scanning time for each mouse was about 5min. After the scan was completed, the overall structure was reconstructed using the software provided with the system, and the image analysis was performed using 3DSlicer software.

[0222] 2.3.3 Lung appearance:

[0223] Place the intact lung tissue on a white base and observe the lung appearance under a magnifying glass.

[0224] 2.3.4 Lung coefficient:

[0225] The lungs were completely removed from the mice and the wet weight of the lungs was measured using the following calculation formula: lung coefficient = wet weight of lungs (g) / body weight (kg) × 100%.

[0226] 2.3.5 Lung tissue pathology:

[0227] The left lung of the mouse was perfused with 4% paraformaldehyde and fixed for 72 hours, and the formaldehyde fixative was changed every 24 hours. After the tissue was completely fixed, it was embedded in paraffin. The embedded wax block was fixed on a microtome and sliced ​​into 4 μm thick slices for hematoxylin and eosin staining (HE).

[0228] 2.3.6 Ki-67 positive cell rate:

[0229] Immunohistochemistry (IHC) was used to detect the expression of Ki-67 in lung tissue, with the primary antibody concentration of 1:500. The Image-Pro Plus 6.0 professional image acquisition and analysis system was used to calculate the proportion of positive cells.

[0230] 2.4 Statistical analysis

[0231] SPSS 22.0 was used for statistical analysis. One-way ANOVA was used for comparison among groups. The least significant difference method was used for homogeneous variances, and Dunnett's T3 method was used for unequal variances. The results were expressed as mean ± SD. The significance level is α = 0.05.

[0232] 3. Experimental results

[0233] 3.1 General situation

[0234] The mice in the blank control group were active, responsive, with shiny fur and no fighting or biting; the mice in the model group began to show slow movement, dull response, listlessness and a preference for lying down 4 weeks after induction, and 1 mouse died 6 weeks after induction; compared with the model group, the above symptoms of the mice in the traditional Chinese medicine group were alleviated, and there was no abnormal death.

[0235] 3.2 Lung CT

[0236] The lung CT texture of the mice in the blank control group was clear, and the transparency of the lung field was normal; the lung volume of the mice in the model group was significantly increased, the transparency of the lung field was decreased, air bronchogram could be seen, and pulmonary consolidation occurred ( Figure 8 ); compared with the model group, the lung volume of the mice in the traditional Chinese medicine group was decreased, the transparency of the lung field was increased, and the proportion of the consolidated lung volume was decreased (P < 0.05) (Table 12).

[0237] Table 12 Lung volume and proportion of consolidation in each group of mice (%)

[0238]

[0239] Note: Compared with the blank control group, P a <0.05, P aa <0.01; compared with the model group, P b <0.05, P bb <0.01.

[0240] 3.3 Lung appearance and lung coefficient

[0241] The lungs of the mice in the blank control group were soft, light red in color, and normal in size; the lungs of the mice in the model group were slightly hard, whitish in color, dense and enlarged, and the lung coefficient was significantly increased (P < 0.01); after traditional Chinese medicine intervention, the lung coefficient was significantly decreased ( Figure 9 , Table 13).

[0242] Table 13 Lung coefficients of each group of mice

[0243]

[0244] Note: Compared with the blank control group, P a <0.05, P aa <0.01; compared with the model group, P b <0.05, P bb <0.01.

[0245] 3.4 Lung tissue pathology

[0246] In the blank control group of mice, the alveolar structure was intact, the alveolar wall was not thickened, and no abnormal cells were seen. In the model group, the alveolar structure was damaged, with obvious fractures and fusions, and cancer nests appeared. After traditional Chinese medicine intervention, it was significantly alleviated ( Figure 10 ).

[0247] 3.5 Ki-67 expression

[0248] Compared with the blank control group, the positive cell rate of Ki-67 in the model group of mice was significantly increased (P < 0.01). After traditional Chinese medicine intervention, the positive cell rate was significantly decreased (P < 0.01)( Figure 11 , Table 14).

[0249] Table 14 Positive cell rate of Ki-67 in mice of each group (%)

[0250]

[0251] Note: Compared with the blank control group, P a < 0.05, P aa < 0.01; compared with the model group, P b < 0.05, P bb < 0.01.

[0252] In summary, the traditional Chinese medicine Yifei Sanjie Formula has a certain effect on improving inflammatory nodules, inhibiting the growth of pure ground-glass nodules in the lung, promoting the regression of pure ground-glass nodules in the lung, reducing the size of in-situ cancer, and inhibiting cell proliferation. This study provides more treatment options for patients with lung nodules who have not reached the surgical indication, can delay or even avoid surgical operation, breaks through the previous binary diagnosis and treatment mode of "observation - operation" for lung nodules, and provides a clinical basis for further exploring the intervention of traditional Chinese medicine in lung nodules, screening the advantageous population for traditional Chinese medicine intervention, and determining the best timing for traditional Chinese medicine intervention.

[0253] The above-described embodiments are only preferred embodiments of the present invention and do not impose any form of limitation on the present invention. Although the present invention has been disclosed above with preferred embodiments, it is not limited to the present invention. Any person skilled in the art can make some modifications or decorations within the scope of the technical solution of the present invention to make equivalent changes to equivalent embodiments. However, as long as it does not depart from the technical solution of the present invention, any simple modification, equivalent change and decoration made to the above embodiments according to the technical essence of the present invention shall fall within the scope of the technical solution of the present invention.

Claims

1. A Chinese medicine composition comprising or consisting of the following Chinese medicine raw materials in parts by weight: 1-20 parts of ginseng, 6-30 parts of gynostemma pentaphyllum, 3-30 parts of pinellia tuber, 2-20 parts of thunbergii bulb, 3-30 parts of peony bark, 3-30 parts of selfheal, 3-30 parts of vinegar-curd zedoaria, 5-50 parts of coix seed, 3-20 parts of stir-fried aurantium immaturum with bran, 3-30 parts of tangerine peel, 1-30 parts of roasted epimedium, and 3-20 parts of roasted liquorice.

2. The Chinese medicine composition according to claim 1, wherein The Chinese medicine composition comprises or is composed of the following Chinese medicine raw materials in parts by weight: 1-11 parts of ginseng, 10-20 parts of gynostemma pentaphyllum, 7-17 parts of pinellia tuber, 4-13 parts of thunbergii bulb, 7-17 parts of peony bark, 7-17 parts of selfheal, 7-17 parts of vinegared zedoaria, 15-25 parts of coix seed, 7-17 parts of stir-fried aurantium immaturum with bran, 7-17 parts of tangerine peel, 1-11 parts of roasted epimedium, and 4-13 parts of roasted licorice.

3. The Chinese medicine composition according to claim 1, wherein The Chinese medicine composition comprises or is composed of the following Chinese medicine raw materials in parts by weight: 4.8-7.2 parts of ginseng, 12-18 parts of gynostemma pentaphyllum, 9.6-14.4 parts of pinellia tuber, 7.2-10.8 parts of thunbergii, 9.6-14.4 parts of peony bark, 9.6-14.4 parts of selfheal, 7.2-10.8 parts of vinegared zedoaria, 16-24 parts of coix seed, 7.2-10.8 parts of stir-fried immature bitter orange, 9.6-14.4 parts of dried tangerine peel, 4.8-7.2 parts of roasted epimedium, and 7.2-10.8 parts of roasted liquorice. In particular, the Chinese medicine composition includes or is composed of the following Chinese medicine raw materials in parts by weight: 5.5-6.5 parts of ginseng, 14-16 parts of gynostemma pentaphyllum, 11-13 parts of pinellia tuber, 8-10 parts of thunbergii bulb, 11-13 parts of peony bark, 11-13 parts of selfheal, 8-10 parts of vinegar-curd zedoaria, 18-22 parts of coix seed, 8-10 parts of stir-fried aurantium immaturum with bran, 11-13 parts of tangerine peel, 5.5-6.5 parts of roasted epimedium, and 8-10 parts of roasted licorice.

4. The Chinese medicine composition according to claim 1, wherein The Chinese medicine composition comprises or is composed of the following Chinese medicine raw materials in parts by weight: 6 parts of ginseng, 15 parts of gynostemma pentaphyllum, 12 parts of pinellia tuber, 9 parts of thunbergii bulb, 12 parts of peony bark, 12 parts of selfheal, 9 parts of vinegared zedoaria, 20 parts of coix seed, 9 parts of stir-fried immature bitter orange, 12 parts of tangerine peel, 6 parts of roasted epimedium, and 9 parts of roasted liquorice.

5. A Chinese medicine preparation obtained from the Chinese medicine composition according to any one of claims 1 to 4.

6. The Chinese medicine preparation according to claim 5, wherein The Chinese medicine preparations include granules, tablets, capsules, pills, powders, syrups, oral solutions, mixtures, decoctions, fluid extracts, and extracts.

7. Use of the Chinese medicine composition according to any one of claims 1 to 4 or the Chinese medicine preparation according to claim 5 or 6 in the preparation of a medicament for preventing or treating pulmonary nodules.

8. The use according to claim 7, wherein: The pulmonary nodules are inflammatory nodules.

9. The use according to claim 7, wherein: The pulmonary nodules are pulmonary ground glass nodules, particularly pure pulmonary ground glass nodules.

10. The use according to claim 7, wherein: The pulmonary nodule is carcinoma in situ.

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