Application of quercetin or derivative thereof in anti-vocal cord fibrosis medicine and preparation

Abstract

The invention relates to an application of Quercetin (QUE) or a derivative thereof in a medicine for resisting vocal cord fibrosis and a preparation of the Quercetin or the derivative thereof. The quercetin can significantly inhibit abnormal deposition of extracellular matrix (ECM) after vocal cord injury, improve reduction of hyaluronic acid (HA) and elastic fibers (EF), and reduce overexpression of type I collagen (COL1A1) and fibronectin (FN1). Besides, by regulating PI3K / AKT and MAPK signal channels, the quercetin inhibits proliferation and migration of fibroblasts and promotes apoptosis of the fibroblasts, so that vocal cord fibrosis is effectively relieved. The invention also provides an application of a preparation of the quercetin and the derivative thereof combined or not combined with other anti-fibrosis drugs (such as 5-fluorouracil and dexamethasone) in treatment of vocal cord fibrosis, wherein the dosage forms of the preparation comprise an oral preparation, an injection, a spray or a local gel preparation.

Description

TECHNICAL FIELD

[0001] The application belongs to the field of medicine, and particularly relates to application and preparation of quercetin or its derivative in anti-vocal fold fibrosis drugs. BACKGROUND

[0002] The vocal fold (VF) is vital to human beings, which is the key organ of phonation, enabling us to communicate emotions and express ideas through language. For special professions such as singing, broadcasting and teaching, the vocal fold is the tool for survival, directly determining the development of their careers. The vocal fold can be divided into five layers from the outside to the inside: stratified squamous epithelium layer (surface protection), Reinke's layer (loose connective tissue), elastic fiber layer (provides elasticity), collagen fiber layer (forms the vocal ligament together with the elastic fiber), and vocal fold muscle (controls tension). The vocal fold body-cover layer vibration theory divides the vocal fold structure into two independent vibration systems, the cover layer and the body layer. The cover layer includes mucosal epithelium and superficial layer of lamina propria (containing elastic fiber network), which realizes the wave-like movement of the edge of the vocal fold through mucosal wave; the body layer is composed of deep layer of lamina propria (mainly collagen fibers) and vocal fold muscle, which accounts for more than 80% of the volume of the vocal fold, and adjusts the tension and length of the vocal fold through muscle contraction. This layered structure is rare in other organs of the human body. When speaking, the vocal fold needs to coordinate vibration and closure, and the mechanical properties of each layer are significantly different. If the vocal fold is damaged, it is difficult to manage in treatment.

[0003] The unique microstructure of the vocal fold is the basis for normal phonation. Vocal fold injury (VFI) or continuous exposure to adverse stimuli can cause excessive proliferation and disordered arrangement of extracellular matrix in the vocal fold lamina propria, and the microstructure is damaged to varying degrees, such as reduced hyaluronic acid content, shortened and disordered arrangement of elastic fibers, and further destruction of the vocal fold lamina propria structure. These histological changes lead to weakening of the vocal fold mucosal wave, affecting the vocal fold vibration function, and thus affecting phonation, and even may worsen into life-threatening airway obstruction. The consequences of vocal fold injury have a negative impact on the physical and mental health of patients, so vocal fold injury, scar formation and subsequent voice quality are the problems that clinicians and patients are most concerned about.

[0004] At present, the treatment methods for vocal fold scar include voice training, surgical treatment, physical therapy and drug therapy. The clinical treatment methods for vocal fold fibrosis are still limited. Traditional drug therapy, such as glucocorticoid therapy, has the risk of systemic adverse reactions, and surgical intervention also faces the challenge of postoperative scar recurrence, so it is urgent to explore safe and effective new treatment strategies.

[0005] Natural medicine has unique advantages in the prevention and treatment of fibrosis due to its multi-component and multi-target characteristics. Quercetin, a flavonoid compound widely found in fruits and vegetables, has been shown to have anti-inflammatory, antioxidant, and cell proliferation and apoptosis regulation activities. It has also been shown to have some intervention effects in liver, kidney, and lung fibrosis models. However, its mechanism in vocal cord fibrosis has not been clearly defined.

[0006] Network pharmacology, as a new research method integrating multiple disciplines, can systematically analyze the action targets and pathways of natural medicine by constructing a "drug-target-disease" network, and provide new ideas for the mechanism research of complex diseases. Based on this, the present application first combines network pharmacology with in vitro and in vivo experiments to disclose the molecular mechanism of quercetin in resisting vocal cord fibrosis, and provides a theoretical basis for clinical diagnosis and treatment.

[0007] Therefore, the present application provides an application of quercetin or its derivative in an anti-vocal cord fibrosis drug and a preparation thereof, which has great significance in applying quercetin to resisting vocal cord fibrosis.

[0008] Based on the above technical background, the present application provides an application of quercetin or its derivative in an anti-vocal cord fibrosis drug and a preparation thereof.

[0009] The first aspect of the present application discloses that quercetin or its derivative can inhibit the reduction of hyaluronic acid and elastic fibers after vocal cord injury, and reduce the abnormal overexpression of collagen and fibronectin.

[0010] The second aspect of the present application discloses that quercetin or its derivative has a multi-target and multi-pathway anti-fibrosis mechanism.

[0011] The third aspect of the present application discloses that quercetin or its derivative can regulate the behavior of fibroblasts and inflammatory response by inhibiting the PI3K / AKT and MAPK pathways.

[0012] The fourth aspect of the present application discloses that quercetin or its derivative can inhibit the proliferation and migration of fibroblasts and promote their apoptosis.

[0013] The technical solution of the present application is that quercetin or its derivative is applied in the preparation of a vocal cord preparation, especially in the preparation of a composition for preventing or treating vocal cord fibrosis.

[0014] Quercetin is a naturally occurring flavonoid compound, specifically belonging to the flavonol subclass. The IUPAC name is 2-(3,4-dihydroxyphenyl)-3,5,7-trihydroxy-4H-chromen-4-one.

[0015] Molecular formula: C 15 H 10 O7

[0016]

[0017] Core structure: Its basic structure consists of three rings (A, C, B rings).

[0018] A ring: There are two hydroxyl groups (-OH) at positions 5 and 7.

[0019] C ring: There is a carbonyl group (C=O) at position 4, and a double bond between positions 2-3 (this is a characteristic of flavonols).

[0020] B ring: There are two adjacent hydroxyl groups (-OH) at positions 3' and 4', known as catechol or catechin structure, which is the key to many biological activities of quercetin.

[0021] Quercetin is a yellow pigment widely found in plants, and is a common component in many fruits, vegetables, leaves, grains, and traditional Chinese medicines. It is hailed as one of the most effective and widely studied flavonoid antioxidants.

[0022] Main characteristics: Strong antioxidant properties: able to neutralize free radicals and reduce oxidative stress. Anti-inflammatory properties: exerts anti-inflammatory effects by regulating various inflammatory pathways (such as NF-κB). Widely found in plants such as apples, onions, berries, broccoli, tea leaves, ginkgo leaves, and St. John's wort.

[0023] 2. Quercetin derivatives: Quercetin derivatives are compounds formed by substituting or modifying one or more hydroxyl groups (-OH) in the molecular structure of quercetin with other chemical groups. These modifications usually occur at the hydroxyl groups of quercetin, especially at positions 3' and 4' of the B ring, and at positions 3, 5, and 7 of the A ring. These modifications greatly affect the solubility, stability, bioavailability, and biological activity of quercetin.

[0024] Common quercetin derivatives are mainly divided into the following categories: a) Glycoside derivatives This is the most common form in nature. One of the hydroxyl groups of quercetin (usually the 3-OH group) is connected to one or more sugar molecules (such as glucose, rhamnose, rutinose, etc.) through a glycosidic bond. Representative examples: Rutin: 3-O-rutinose glycoside of quercetin (rutinose = glucose + rhamnose). Widely present in buckwheat, asparagus, and citrus fruits. Isoquercitrin: 3-O-glucoside of quercetin. Present in many plants. Quercitrin: 3-O-rhamnoside of quercetin. One of the main components of ginkgo biloba leaf extract. Characteristics: Glycosylation increases the water solubility of quercetin, but it needs to be hydrolyzed by intestinal flora into quercetin aglycone (i.e. quercetin itself) in the human body to be fully absorbed. b) Methylated derivatives One or more hydroxyl groups of quercetin are replaced by methyl groups (-CH3). Representative examples: Isoquercitrin: 3'-O-methylated derivative of quercetin. Myricetin: 3',5'-O-dimethylated derivative of quercetin. Characteristics: After methylation, the polarity and hydrogen bonding ability of the molecule decrease, and the liposolubility is usually better, the metabolism in the body is more stable, and the half-life is longer. This may improve its oral bioavailability. c) Sulfated or glucuronidated metabolites This is the main form of quercetin after phase II metabolism in the human body (mainly in the liver and intestine). Characteristics: Quercetin or its partial metabolites are combined with sulfate or glucuronate. d) Semi-synthetic or synthetic derivatives In order to improve the low water solubility and low bioavailability of quercetin itself, scientists have synthesized many new derivatives through chemical methods. Examples: Quercetin complexes: such as complexes with phospholipids (quercetin phospholipid complexes), or inclusion complexes with cyclodextrins, which can significantly improve its absorption. Esterified or etherified derivatives: modification of hydroxyl groups into other lipophilic groups.

[0025] Preferably, the composition comprises a pharmaceutical composition.

[0026] Preferably, the composition comprises a pharmaceutically acceptable carrier.

[0027] Preferably, the dosage form of the composition is an oral preparation, an injection, a spray, or a topical gel preparation.

[0028] Preferably, the composition comprises a quercetin nano-preparation.

[0029] The technical solution of the present application has the following beneficial effects:

[0030] The quercetin in the product of this invention has the effect of significantly inhibiting the reduction of hyaluronic acid and elastic fibers after vocal cord injury, reducing the abnormal overexpression of collagen and fibronectin, and alleviating vocal cord fibrosis; (2) Quercetin exerts its effect by regulating hormone response, cell migration and extracellular matrix components, and has a multi-target and multi-pathway anti-fibrotic mechanism; (3) Quercetin exerts its anti-fibrotic effect by inhibiting PI3K / AKT and MAPK pathways, regulating fibroblast behavior and inflammatory response, and does not cause damage to the liver and kidneys; (4) The inhibitory effect of quercetin on fibroblast proliferation is significantly better than or equivalent to that of 5-fluorouracil and dexamethasone. Attached Figure Description

[0031] Figure 1 A comparison image of hyaluronic acid and elastic fibers after staining.

[0032] Figure 2 The percentage of hyaluronic acid and elastic fiber dyed area.

[0033] Figure 3 This is a bar chart showing the expression levels of FN and COL1A1 in the vocal cords.

[0034] Figure 4 This is a pathological section of vocal cord tissue.

[0035] Figure 5 Box plot comparing texture parameters of four groups of vocal cord pathology images.

[0036] Figure 6 The effect of QUE on the expression levels of PI3K, p-PI3K, AKT, p-AKT, p38 and p-p38 proteins in vocal cord tissue.

[0037] Figure 7 To investigate the effects of QUE on the expression of PI3K, AKT, and p38MAPK mRNA in vocal cord tissue.

[0038] Figure 8 The results show the effect of QUE on the proliferation of vocal cord fibroblasts.

[0039] Figure 9 The effect of QUE on vocal cord fibroblast migration.

[0040] Figure 10 Effects of QUE on apoptosis of vocal cord fibroblasts

[0041] Figure 11 The effects of QUE on the liver and kidneys of SD rats.

[0042] Figure 12 The effects of different concentrations of three drugs on the expression levels of COL1A1 and FN1 proteins in RVFF.

[0043] Figure 13 Results of CCK-8 method for measuring the effects of the three drugs on the proliferation of RVFF. DETAILED DESCRIPTION

[0044] The following illustrates some embodiments of the present application, which further details the above content of the present application but should not be understood as limiting the above subject matter of the present application to only the following embodiments.

[0045] Example 1 Therapeutic effect of quercetin on vocal cord fibrosis

[0046] In this example, 36 SD rats were randomly divided into a normal control group (NC), an injury group (VFI), a low-dose quercetin treatment group (QUEL, 100 mg / kg), and a high-dose quercetin treatment group (QUEH, 200 mg / kg). QUEL and QUEH were treated on the basis of injury, and the administration time was 7 days before injury to 3 days after injury, a total of 10 days. The content and arrangement changes of ECM components (hyaluronic acid (HA), elastic fiber (EF), type I collagen fiber (COL1A1), and fibronectin (FN1)) were detected by Alcian Blue staining, modified EVG staining, Western Blot, and image texture analysis.

[0047] The results of this example show that QUE has a significant inhibitory effect on the reduction of hyaluronic acid in the vocal cord tissue: in the VFI group and the NC group, the VFI group had a HA staining area ratio of 13.09% ± 5.09%, and the NC group had a HA staining area ratio of 28.16% ± 4.54%. Compared with the NC group, the VFI group had a significantly decreased HA staining area ratio in the lamina propria (P < 0.0001). In the two QUE treatment groups, the QUEL group had a HA staining area ratio of 19.88% ± 2.14%, and the QUEH group had a HA staining area ratio of 19.88% ± 2.14%. The HA staining area ratio content of both groups was higher than that of VFI (QUEL: P = 0.0135, QUEH: P = 0.0003).

[0048] In the VFI group and the NC group, the VFI group had an EF staining area ratio of 0.47% ± 0.03%, and the NC group had an EF staining area ratio of 1.00% ± 0.12%. Compared with the NC group, the VFI group had a significantly decreased EF staining area ratio in the lamina propria (P < 0.0001). This indicates that after vocal cord injury, the content of extracellular matrix EF in the vocal cord tissue decreases.

[0049] The results of this example show that QUE has a significant inhibitory effect on the reduction of elastic fibers in the vocal cord tissue: in the two QUE treatment groups, the EF staining area ratio of QUEL group was 0.71%±0.06%, and the EF staining area ratio of QUEH group was 0.86%±0.05%. The EF staining area ratio of the two groups was higher than that of VFI (QUEL: P=0.0001, QUEH: P<0.0001). This shows that QUE has an inhibitory effect on the decrease of extracellular matrix EF content after vocal cord injury, and the higher the concentration, the more obvious the inhibitory effect.

[0050] The results of this example show that QUE significantly reduces the abnormal overexpression of collagen and fibronectin: compared with the NC group, the expression of FN1 and COL1A1 in the vocal cord of the VFI group increased (PFN1=0.0059, PCOL1A1=0.0247). This shows that after vocal cord injury, the expression of COL1A1 and FN1 proteins in the vocal cord tissue increases. In the two QUE treatment groups, the expression of FN1 and COL1A1 in the vocal cord was higher than that in the VFI group (QUEL: PFN1=0.0217, PCOL1A1=0.0067; QUEH: PFN1=0.0074, PCOL1A1=0.0121). This shows that QUE has an inhibitory effect on the increase of COL1A1 and FN1 protein expression after vocal cord injury, and the higher the concentration, the more obvious the inhibitory effect.

[0051] The results of this example show that QUE has an inhibitory effect on the proliferation of blood vessels in the lamina propria of the vocal cord: 56 days after injury, the HE staining results show that the normal vocal cord pathological tissue is characterized by uniform distribution of fibrous connective tissue in the lamina propria without abnormal angiogenesis; while the injured vocal cord has thick collagen fibers, significantly disordered arrangement, and abnormal angiogenesis. No abnormal angiogenesis was observed in the treatment group. According to the image texture analysis results, compared with the NC group, the energy (Energy) of the vocal cord pathological image of VFI was significantly lower than that of the NC group (P<0.0001), the entropy (Entropy) was significantly higher than that of the NC group (P<0.0001), the contrast (Contrast) was higher than that of the NC side (P=0.0385), the correlation (Correlation) was significantly lower than that of the NC group (P=0.008), and the homogeneity (Homogeneity) was significantly lower than that of the NC group (P<0.0001).

[0052] The Energy of the vocal cord pathological picture of the QUEL group was higher than that of the VFI group (P=0.0019), the Entropy was lower than that of the VFI group (P=0.0172), and the differences in the Contrast, Correlation and Homogeneity parameters were not statistically significant (Contrast: P=0.5090, Correlation: P=0.2940, Homogeneity: P=0.1641).

[0053] Compared with the VFI group, the Energy of the vocal cord pathological picture of the QUEH group was higher (P=0.0345), the Entropy was significantly lower (P<0.0001), the Contrast was lower (P=0.0178), the Correlation was significantly higher (P=0.0008), and the Homogeneity was higher (P=0.0046).

[0054] The results of the example are shown in Figures 1-5 .

[0055] Example 2 Mechanism of quercetin in treating vocal cord fibrosis

[0056] Based on the TCMSP and Gene Cards databases, 2968 potential intersection target points of QUE (quercetin) against VFF (vocal cord fibrosis) were screened, a protein interaction network (PPI) was constructed, and 14 core target points were identified, namely AKT1, IL6, TNF, TP53, IL1B, EGFR, MMP9, CASP3, PTGS2, JUN, BCL2, MYC, EGF and PPARG. A "disease + drug-target-signal pathway" relationship network was constructed by combining GO function annotation and KEGG pathway enrichment analysis. The core target points have significant significance in the PI3K / AKT and MAPK signal pathways. The molecular docking verified that the binding energy of QUE with PI3K, AKT1 and p38 proteins was all less than -5 kcal / mol. According to the KEGG enrichment analysis and the molecular docking results, the Western Blot and RT-PCR were further used to explore the inhibitory effect of QUE on the PI3K / AKT and MAPK signal pathways.

[0057] The in vivo experiment of the example shows that, by the Western Blot experiment, the protein expression amounts of PI3K, p-PI3K, AKT, p-AKT, p38 and p-p38 in the vocal cord tissue of the SD rats were detected, and the results are shown in Figure 6The results show that the expression of PI3K, p-PI3K, AKT, p-AKT, p38 and p-p38 proteins in the vocal cords of the VFI group is significantly higher than that of the NC group (P<0.05). In the QUEL group, the expression of PI3K, p-PI3K, AKT, p38 and p-p38 proteins is significantly lower than that of the VFI group (P<0.05), while the expression of p-AKT has no statistically significant difference with that of the VFI group (P=0.2251). In the QUEH group, the expression of PI3K, p-PI3K, AKT, p-AKT, p38 and p-p38 proteins is significantly lower than that of the VFI group (P<0.05).

[0058] Meanwhile, the expression levels of PI3K, AKT and p38MAPK mRNA in the vocal cord tissues of SD rats were detected by RT-PCR in this embodiment, and the results are shown in Figure 7 The results show that the trend is consistent with the Western Blot results.

[0059] This embodiment further verifies that quercetin reduces the proliferation of fibroblasts by inhibiting the PI3K / AKT and MAPK signaling pathways: we treated RVFF with 10 ng / mL TGF-β1 and added 0 uM, 20 uM and 100 uM of QUE in the culture medium respectively, and treated for 1 day, and then detected the proliferation in the next 4 days (from the end of QUE treatment on the 0th day) by CCK-8. The results in CCK8 show that on the 1st day, 20 uM and 100 uM of QUE significantly inhibited the proliferation of RVFF treated with 10 ng / mL TGF-β1 (P<0.05). It was observed in the following days, and the effect of inhibiting the proliferation of RVFF was more obvious with the increase of the dose of QUE treatment and the number of days (P<0.05). The results are shown in Figure 8

[0060] This embodiment further verifies that quercetin reduces the migration of fibroblasts by inhibiting the PI3K / AKT and MAPK signaling pathways: RVFF was induced by 10 ng / mL TGF-β1, and then inoculated into the upper chamber of the Transwells chamber, and 0 uM, 20 uM and 100 uM of QUE were added in the culture medium of the upper chamber respectively, and then the lower chamber of the Transwells chamber was stained with crystal violet after two days of treatment to evaluate the migration ability. The results are shown in Figure 9 Compared with the normal group, the migration ability of RVFF induced by 10 ng / mL TGF-β1 is significantly enhanced (P<0.05). But after QUE treatment, the amount of RVFF cells migrated to the lower chamber is significantly reduced (P<0.05), and there is a dose-dependent effect.

[0061] ​This example further verifies that quercetin promotes apoptosis of in vitro vocal cord fibroblasts: we treated RVFF with 10 ng / mL TGF-β1 and added 0 uM, 20 uM and 100 uM of QUE in the culture medium, respectively, for 1 day, and evaluated the effect of QUE on TGF-β1-induced apoptosis of RVFF using Annexin V / PI staining cell fluorescence analysis, and the results are shown in Figure 10 The flow cytometry results show that the level of apoptosis of RVFF induced by 10 ng / mL TGF-β1 did not change significantly (P < 0.05) compared with the normal group. However, after treatment with QUE, the level of apoptosis of RVFF increased (P < 0.05) in a dose-dependent manner. This indicates that QUE can reduce vocal cord fibrosis by inducing apoptosis of RVFF cells.

[0062] Example 3 Safety of quercetin for treating vocal cord fibrosis

[0063] This example further verifies the safety of QUE at the animal level: we collected blood samples from rats after 10 days of continuous gavage administration of 0.5% CMC-Na solution (injury group, 0 mg / kg), 100 mg / kg quercetin solution and 200 mg / kg quercetin solution (6 rats in each group), and detected biochemical indicators reflecting liver and kidney function. Six biochemical indicators were selected as evaluation criteria, three liver function indicators, ALP, ALT and AST; three kidney function indicators, UA, UREA and CREA. Statistical analysis of the biochemical test results is shown in Figure 11 Statistical analysis shows that there is no statistically significant difference (P > 0.05) in liver and kidney function indicators between the two treatment groups and the control group. This further indicates that, at the dosages used in this study, QUE has no toxic effect on the liver and kidneys of SD rats.

[0064] Example 4 Preparation of quercetin preparation

[0065] 4.1 Preparation of quercetin laryngeal spray

[0066] Formulation:

[0067]

[0068] Preparation process:

[0069] 1) High-pressure homogenization method to prepare quercetin nanocrystals;

[0070] 2) Ultrasonic mixing with chitosan solution;

[0071] 3) After sterilization filtration, dispense into a spray device.

[0072] The product of this embodiment is subjected to accelerated test at 40°C / 75% condition, no pathogenic bacteria is detected within 6 months, the solution has no precipitation or stratification; the retention rate of active ingredient is >92% within 6 months, the degradation product is <2%, the particle size increase is controllable (PDI <0.25), and the permeability of the throat mucosa is ensured.

[0073] 4.2 Preparation of quercetin oral dissolving film

[0074]

[0075] Preparation process:

[0076] 1) Quercetin, sucralose, glycerol, and sodium dodecyl sulfate were added to an aqueous ethanol solution and stirred to dissolve;

[0077] 2) Hydroxypropyl methylcellulose was added to the solution of 1) and stirred to dissolve;

[0078] 3) The film solution was subjected to standing / vacuum extraction / centrifugal defoaming;

[0079] 4) The defoamed film solution was coated and dried, and the film was cut into appropriate size to obtain quercetin oral dissolving film.

[0080] The product of this embodiment is placed in the oral cavity without water, and is completely wetted within 5s, swells and disintegrates into flocculent particles within 30s, can be rapidly disintegrated and absorbed in the oral cavity, and plays a role in resisting vocal cord fibrosis.

[0081] 4.3 Preparation of quercetin oral solution

[0082]

[0083] Preparation process:

[0084] 1) Quercetin raw material (particle size ≤50 μm) was mixed with Tween 80, and ball milling was performed for 30 min to form a uniform paste;

[0085] 2) CMC-Na was added to purified water, and stirred in a water bath at 60°C until completely dissolved;

[0086] 3) The quercetin-Tween 80 paste was added to the CMC-Na solution and homogenized at high speed;

[0087] 4) Ascorbic acid was added, and purified water was added to constant volume, and the pH was adjusted to 6.5-7.0;

[0088] 5) 0.22 μm microporous filter was used for sterilization, and the solution was divided into brown glass bottles.

[0089] The product of this example was subjected to accelerated test at 40°C / 75% condition, no bacteria / mold was detected within 6 months, after 6 months, the active ingredient retention was 93.7%; degradation product <1%, in line with the liver and kidney non-toxic threshold; sedimentation volume ratio: ≥0.90, shake well for reconstitution within 30s. The product can be valid for 24 months.

[0090] Example 5 Comparison of application of quercetin with other drugs

[0091] In this example, it was verified that quercetin had the same or better effect on inhibiting the expression of COL1A1 and FN1 of vocal cord fibroblasts than other drugs: three drugs, QUE, 5-Fu (5-fluorouracil), and DXMS (dexamethasone), were set at 25 μM, 50 μM, 100 μM, and 200 μM for a total of four drug treatment concentrations to evaluate the regulatory effect of the three drugs on the markers of vocal cord fibrosis, COL1A1 and FN1, induced by 10 ng / mL TGF-β1, and the results are shown in Figure 12 The results showed that at 25 μM, 50 μM, 100 μM, and 200 μM, the expression levels of COL1A1 and FN1 of RVFF (rat vocal cord fibroblasts) induced by 10 ng / mL TGF-β1 were up-regulated compared with the normal group (P<0.05). Compared with the TGFB1 treatment group, the expression levels of FN1 and COL1A1 were significantly reduced after treatment with the four concentrations of QUE (P<0.05), indicating that QUE can effectively inhibit the expression of fibrosis-related proteins of RVFF cells induced by 10 ng / mL TGF-β1.

[0092] This example further verified that the inhibitory effect of QUE on the proliferation of RVFF cells was more significant than that of other drugs: 10 ng / mL TGF-β1 was used to induce RVFF, and 25 μM of QUE, 5-Fu, and DXMS were added to the culture medium, respectively, for 1 day of treatment, and the relative proliferation of cells within 0-4 days was observed (day 0 was after the drug treatment of cells). The results are shown in Figure 13The relative proliferation level of the control group RVFF cells gradually increased over time. The relative proliferation rate of the TGF-β1 group RVFF cells was significantly higher than that of the control group (P<0.05), and the relative proliferation level was close to 15 times that of the 0th day on the 4th day, indicating that TGF-β1 had a strong promoting effect on the proliferation of RVFF cells. The relative proliferation level of the QUE group RVFF cells was significantly lower than that of the TGF-β1 group (P<0.05), and the growth was very slow, and was also significantly lower than that of the control group, which fully indicated that QUE could effectively inhibit the proliferation of cells in the RVFF cells stimulated by 10 ng / mL TGF-β1. The 5-Fu group also showed an inhibitory effect on cell proliferation (P<0.05), but the inhibitory effect of the QUE group was similar to that of the 5-Fu group or even better at some time points (P<0.05). The relative proliferation level of the DXMS group cells was lower than that of the TGF-β1 group (P<0.05), but higher than that of the QUE group (P<0.05), indicating that its ability to inhibit cell proliferation was weak.

[0093] The above only describes the preferred embodiments of the present application. It should be noted that those skilled in the art can make several improvements and refinements without departing from the principles of the present application, and these improvements and refinements should also be considered within the scope of the present application.

Claims

1. Use of quercetin or a pharmaceutically acceptable derivative thereof in a medicament for combating fibrosis of the vocal cords, characterized in that: Quercetin or its derivative preparation can inhibit PI3K / AKT and MAPK signaling pathways, reduce the proliferation and migration of fibroblasts, and promote the apoptosis of fibroblasts, thereby improving vocal cord fibrosis.

2. Use according to claim 1, characterized in that: The drug can inhibit the reduction of hyaluronic acid (HA) and elastic fibers (EF) after vocal cord injury.

3. Use according to claim 1, characterized in that: The drug can reduce the overexpression of type I collagen (COL1A1) and fibronectin (FN1).

4. Use according to claim 1, characterized in that: The drug has an inhibitory effect on the proliferation of blood vessels in the vocal cord lamina propria.

5. The use according to claim 1, characterized in that: Quercetin preparation can be used for the prevention and treatment of anti-vocal cord fibrosis.

6. Quercetin or a pharmaceutically acceptable derivative thereof as a medicament for anti-vocal cord fibrosis, characterized by: Quercetin or its derivative preparation can inhibit PI3K / AKT and MAPK signaling pathways, reduce the proliferation and migration of fibroblasts, and promote the apoptosis of fibroblasts, thereby improving vocal cord fibrosis.

7. The medicament according to claim 6, characterized in that: The dosage form of the drug is oral preparation, injection, spray or topical gel preparation.

8. The medicament according to claim 6, characterized in that: The drug has high safety and no damage to the liver and kidney.

9. The medicament according to claim 6, characterized in that: The drug can further contain or not contain other anti-fibrosis drugs, preferably 5-fluorouracil or dexamethasone.

10. The medicament according to claim 6, characterized in that: The administration dose of quercetin is 5-25 mg / day.

Images