Hydrogel for resisting collagen proliferative diseases as well as preparation method and application thereof

Abstract

The invention belongs to the technical field of medicines, and particularly relates to hydrogel for resisting collagen proliferative diseases as well as a preparation method and application of the hydrogel. According to the hydrogel for resisting the collagen proliferative diseases, by optimizing the composition and proportion of the hydrogel, the salvianic acid hydrogel can effectively prevent and treat hypertrophic scars. The salvianic acid hydrogel dosage form provided by the invention is used for local administration, is convenient to operate, and can improve the compliance of patients compared with oral medicines (such as compound salvia miltiorrhiza dropping pills). The salvianic acid hydrogel dosage form can be uniformly smeared on the surface of a wound and quickly forms a film for covering, so that the local concentration of salvianic acid is increased, and a relatively good treatment effect is achieved. And the tanshinol hydrogel dosage form is simple and convenient in production process and low in cost, is an economical and effective treatment scheme, and has a good prospect in prevention or treatment of collagen hyperplasia diseases.

Description

Technical Field

[0001] The present invention belongs to the field of pharmaceutical technology, and particularly relates to a hydrogel for treating collagen hyperplasia diseases, and a preparation method and use thereof. Background Art

[0002] Collagen hyperplasia diseases are a group of diseases caused by abnormal hyperplasia or metabolic disorders of collagen fibers in connective tissues due to genetic or acquired factors. These diseases are mainly caused by gene mutations leading to disorders in the synthesis or function of specific types of collagen, resulting in excessive hyperplasia or deposition of collagen fibers, thereby affecting the normal structure and function of tissues. Among them, pathological scars (such as hypertrophic scars and keloids) are typical manifestations of abnormal collagen fiber hyperplasia. Their formation is affected by various factors such as genetics, inflammation, and infection.

[0003] A scar is a general term for the appearance and histopathological changes of normal skin tissue caused by various skin injuries, and is an inevitable product in the process of human wound repair, manifested as changes in appearance, pain, itching, etc. During the wound healing process, the balance between the synthesis and degradation metabolism of collagen caused by various reasons can form pathological scars. Hypertrophic scars are the most common type of pathological scars clinically, accompanied by abnormal tissue hyperplasia during the wound healing process. The overall incidence of hypertrophic scars after skin trauma is as high as 40%-70%, and the incidence of hypertrophic scars after burns even exceeds 80%. Hypertrophic scars usually present as thickening, redness, and hardness, bringing obvious appearance, function, and psychological burdens to patients. Its formation mechanism has not been fully elucidated, and the process involves complex processes such as inflammatory reactions, fibroblast activation, and excessive collagen deposition. Especially in the maxillofacial region, hypertrophic scars have a particularly significant impact on aesthetics and function. With the increase in the number of surgical, traumatic, and burn cases, the incidence of hypertrophic scars is gradually rising. How to effectively control the formation of scars and inhibit hyperplasia has become a difficult point in clinical treatment.

[0004] Currently, clinical treatment methods are relatively limited. For example, local silicone application, compression or massage therapy, intralesional injection of corticosteroids or 5-fluorouracil, laser ablation, and surgery are the most commonly used methods for preventing and treating hypertrophic scars. However, the above-mentioned therapies lack evidence of effectiveness and safety, have a high recurrence rate, and the mechanism of action is still unclear. Moreover, these methods all have certain limitations. For example, surgery may lead to recurrence of scar hyperplasia, the effect of laser treatment is unstable, drug injection is prone to cause adverse reactions or requires multiple operations, and patient compliance is poor. Therefore, there is an urgent need for a safe, effective, and long-lasting treatment method to better inhibit the occurrence and development of hypertrophic scars.

[0005] Searching for a more minimally invasive and convenient drug therapy with sustained and stable curative effects and fewer adverse reactions remains one of the urgent goals pursued by clinicians. Currently, it is believed that myofibroblasts play a key role in the process of scar formation. A large number of myofibroblasts proliferate, escape apoptosis, and secrete a large amount of collagen extracellular matrix, leading to local tissue hyperplasia. Inhibiting the synthesis and secretion of collagen and reducing the formation of extracellular matrix in scar tissue are one of the strategies for treating scars, but there is currently no reported exact and effective treatment plan.

[0006] Previous studies have shown that the research on Salvia miltiorrhiza and its preparations has mainly focused on their applications in visceral fibrosis diseases, and their clinical applications in hypertrophic scars still need to be verified by a large number of studies. Moreover, due to the complex effective components of traditional Chinese medicine, the effective components of Salvia miltiorrhiza include tanshinones, such as tanshinone IIA and tanshinone I, and salvianolic acids, such as salvianolic acid A and danshensu. In addition, the types and concentrations of various effective components and excipients in pharmaceutical preparations have a great impact on their efficacy.

[0007] Therefore, the administration methods of the effective components of Salvia miltiorrhiza for treating collagen hyperplasia diseases, such as dosage forms and concentrations, are problems that still need to be explored and solved. Summary of the Invention

[0008] Aiming at the defects of the existing technology, the present invention provides a hydrogel for treating collagen hyperplasia diseases, aiming to effectively prevent and treat collagen hyperplasia diseases.

[0009] The present invention provides a hydrogel for treating collagen hyperplasia diseases, which is composed of the following raw materials in mass concentration:

[0010] Active ingredient 0.019 - 0.08 mg / mL, moisturizing and repairing agent 15 - 25 mg / mL, gelling agent 5 - 15 mg / mL, thickening agent 5 - 10 mg / mL, antioxidant 0.1 - 1 mg / mL;

[0011] The active ingredient is danshensu.

[0012] Preferably, the moisturizing and repairing agent is selected from at least one of sodium hyaluronate, glycerol, trehalose, chitosan, and collagen; the gelling agent is at least one of carbomer and poloxamer; the thickening agent is selected from at least one of carboxymethyl cellulose and polyvinyl alcohol.

[0013] Preferably, the sodium hyaluronate is a mixture of sodium hyaluronates with different molecular weights, and the mixture is a combination composed of sodium hyaluronate with a molecular weight of 100 - 200 kDa, sodium hyaluronate with a molecular weight of 200 - 400 kDa, and sodium hyaluronate with a molecular weight of 1000 - 1500 kDa in a weight ratio of 1 - 10:1 - 10:1 - 10;

[0014] And / or, the carbomer is at least one of carbomer 940, carbomer 934, and carbomer 980, and the antioxidant is at least one of vitamin C, lipoic acid, and glutathione.

[0015] The present invention provides a method for preparing the hydrogel for treating collagen hyperplasia diseases as described in any one of the above, which is characterized in that it includes:

[0016] Mix and stir the active ingredient, moisturizing and repairing agent, gelling agent, and thickening agent to obtain the product.

[0017] The present invention provides the use of the hydrogel for treating collagen hyperplasia diseases as described in any one of the above in the preparation of a drug for preventing or treating collagen hyperplasia diseases.

[0018] Preferably, the collagen hyperplasia disease is a pathological scar disease.

[0019] Preferably, the pathological scar disease is a hypertrophic scar disease.

[0020] The term "moisturizing and repairing agent" refers to a reagent having moisturizing properties and tissue repair properties, such as sodium hyaluronate.

[0021] The term "gelling agent" refers to the main matrix component in the hydrogel material, which provides structure and stability, such as carbomer.

[0022] The term "thickening agent" refers to a reagent that provides viscosity to the hydrogel, such as carboxymethyl cellulose.

[0023] The types and formulations of the excipients of the present invention are obtained through screening and optimization. In the screening experiment of the moisturizing and repairing agent, glycerol, propylene glycol, butylene glycol, diglycerol, dipropylene glycol, polyethylene glycol, trehalose, betaine, erythritol, chitosan, collagen, glycerol polyether, hyaluronic acid with different molecular weights and their ratios, etc. were compared; in the screening experiment of the gelling agent, xanthan gum, carbomer, poloxamer with different molecular weights or models, etc. were compared; in the screening experiment of the thickening agent, carboxymethyl cellulose, hydroxymethyl cellulose, guar gum, carrageenan, hydroxypropyl methyl cellulose, arabic gum, polyvinyl alcohol, etc. were compared. Only under the preferred types and formulations of the excipients can the efficient treatment of collagen hyperplasia diseases be achieved.

[0024] In the present invention, by screening the formulation components and their ratios of the hydrogel, danshensu hydrogels with different concentrations were prepared respectively. Experiments showed that the danshensu hydrogel could effectively prevent and treat hypertrophic scars, and the prevention and treatment effect was the best when danshensu was at an appropriate concentration. The danshensu hydrogel dosage form was used for topical administration, which was convenient to operate. Compared with treatment methods such as oral medications (such as compound danshen dripping pills) and radiotherapy, it could improve the compliance of patients. The danshensu hydrogel dosage form could be evenly applied on the surface of the wound and quickly form a film to cover it, prolonging the local action time of the drug, increasing the local concentration of the danshensu drug, and achieving a better treatment effect. Moreover, the production process of the danshensu hydrogel dosage form was simple and the cost was low, which could provide an economical and effective treatment plan for patients and had good prospects in the prevention or treatment of collagen hyperplasia diseases.

[0025] Obviously, based on the above content of the present invention, according to the common general knowledge and customary means in the art, without departing from the above basic technical idea of the present invention, various other forms of modifications, substitutions or changes can also be made.

[0026] The following is a further detailed description of the above content of the present invention through specific embodiments in the form of examples. However, this should not be construed as limiting the scope of the above subject matter of the present invention to the following examples. All technologies implemented based on the above content of the present invention fall within the scope of the present invention. BRIEF DESCRIPTION OF THE DRAWINGS

[0027] Figure 1 It is the result diagram of rabbit ear scars; among them, A is the gross observation result diagram; among them, B is the statistical result diagram of the scar thickness, where * indicates P≤0.05, *** indicates P≤0.001, **** indicates P≤0.0001, and ns indicates P>0.05.

[0028] Figure 2 It is the statistical result diagram of the scar elevation index (SEI), where * indicates P≤0.05, ** indicates P≤0.01, **** indicates P≤0.0001, and ns indicates P>0.05.

[0029] Figure 3 It is the staining result diagram of the scar tissue; among them, A is the hematoxylin-eosin (HE) staining result diagram; among them, B is the Masson staining result diagram. DETAILED DESCRIPTION OF THE EMBODIMENTS

[0030] In the following examples and experimental examples, the reagents and materials not specifically described are all commercially available products.

[0031] Carboxymethyl cellulose: CAS number is 9004-32-4, brand is Hushi, product number is 30036328, CP pure. The relative molecular mass of carbomer 940 is 280.7.

[0032] Example 1 Danshensu Hydrogel and Its Preparation

[0033] The danshensu hydrogel (100 μM) of this example was prepared by the following method:

[0034] An appropriate amount of danshensu (CAS No.: 76822-21-4) was added to 1 ml of deionized water and fully dissolved to prepare a 100 μM danshensu solution (i.e., 0.0198 mg / ml). Then, a total of 20 mg of sodium hyaluronate powder with different molecular weights, 5 mg of carbomer, 5 mg of carboxymethyl cellulose, and 1 mg of vitamin C powder were weighed. The powders were added to the above danshensu solution and continuously stirred and mixed to form a transparent viscous danshensu hydrogel. The main active ingredient of this hydrogel is danshensu. The excipients are sodium hyaluronate with different molecular weights, namely: low molecular weight sodium hyaluronate (molecular weight 100-200 kDa), medium molecular weight sodium hyaluronate (molecular weight 200-400 kDa), and high molecular weight sodium hyaluronate (molecular weight 1000-1500 kDa), and their mass ratio is 1:1:6, and the total concentration of sodium hyaluronate is 20 mg / ml; the carbomer is carbomer 940; the antioxidant is vitamin C.

[0035] Example 2 Danshensu Hydrogel and Its Preparation

[0036] The danshensu hydrogel (400 μM) of this example was prepared by the following method:

[0037] An appropriate amount of danshensu (CAS No.: 76822-21-4) was added to 1 ml of deionized water and fully dissolved to prepare a 400 μM danshensu solution (i.e., 0.0792 mg / ml). Then, a total of 20 mg of sodium hyaluronate powder with different molecular weights, 5 mg of carbomer, 5 mg of carboxymethyl cellulose, and 1 mg of vitamin C powder were weighed. The powders were added to the above danshensu solution and continuously stirred and mixed to form a transparent viscous danshensu hydrogel. The main active ingredient of this hydrogel is danshensu. The excipients are sodium hyaluronate with different molecular weights, namely: low molecular weight sodium hyaluronate (molecular weight 100-200 kDa), medium molecular weight sodium hyaluronate (molecular weight 200-400 kDa), and high molecular weight sodium hyaluronate (molecular weight 1000-1500 kDa), and their mass ratio is 1:1:6, and the total concentration of sodium hyaluronate is 20 mg / ml; the carbomer is carbomer 940; the antioxidant is vitamin C.

[0038] Example 3 Danshensu Hydrogel and Its Preparation

[0039] The danshensu hydrogel (800 μM) of this example was prepared by the following method:

[0040] An appropriate amount of danshensu (CAS No.: 76822-21-4) was added to 1 ml of deionized water and fully dissolved to prepare a 800 μM danshensu solution (i.e., 0.1585 mg / ml). Then, 20 mg of sodium hyaluronate powder with different molecular weights, 5 mg of carbomer, 5 mg of carboxymethyl cellulose, and 1 mg of vitamin C powder were weighed. The powders were added to the above danshensu solution and continuously stirred and mixed to form a transparent viscous danshensu hydrogel. The main active ingredient of this hydrogel is danshensu. The excipients are sodium hyaluronate with different molecular weights, namely: low molecular weight sodium hyaluronate (molecular weight 100 - 200 kDa), medium molecular weight sodium hyaluronate (molecular weight 200 - 400 kDa), and high molecular weight sodium hyaluronate (molecular weight 1000 - 1500 kDa), and their mass ratio is 1:1:6. The total concentration of sodium hyaluronate is 20 mg / ml; the carbomer is carbomer 940; the antioxidant is vitamin C.

[0041] The technical solution of the present invention will be further described through experiments below. The sample danshensu hydrogel used in the following experimental examples was prepared by the methods of Examples 1 - 3 above.

[0042] Experimental Example 1 Construction of a scar animal model and study on the intervention effect of danshensu hydrogel

[0043] I. Experimental methods

[0044] 1. Construction of a hypertrophic scar animal model and experimental grouping

[0045] Six New Zealand rabbits weighing 2 - 3 kg, regardless of gender, were selected as experimental animals. The rabbits were fed in an environment with a temperature of 23 ± 3°C and a humidity of 50 ± 10%, and could freely access water and food. The rabbits were anesthetized by injecting 3% pentobarbital sodium at a dose of 1 ml / kg through the marginal ear vein. After the rabbits entered a deep anesthesia state, the epidermis, dermis, and perichondrium were removed on the ventral surface of the rabbit ear using a square punch with a diameter of 1 cm and a No. 11 disposable sterile plastic handle scalpel, forming 2 square full-thickness incisions with a diameter of 1 cm on each ear.

[0046] The wounds on the rabbit ears were immediately disinfected with povidone-iodine solution. The wound on the left ear of the rabbit was used as the control group, which was only disinfected with povidone-iodine solution and bandaged with gauze. The wounds on the right ear of the rabbit were used as the experimental group and randomly divided into 3 groups. After povidone-iodine disinfection, they were treated by applying 100 μM, 400 μM, and 800 μM danshensu hydrogel respectively, and the dressing was changed once every two days. If scabs appeared during the re-epithelialization process of the rabbit ear wounds, they were removed in a timely manner.

[0047] On the 21st day after surgery, the scar in the surgical area was photographed, its thickness was measured, and tissue samples were taken.

[0048] 2. Measurement of Scar Thickness

[0049] Measure the thickness of the scar and the thickness of the normal skin adjacent to the scar with a vernier caliper, and subtract the two values to obtain the scar thickness.

[0050] 3. Calculation of Scar Elevation Index (SEI)

[0051] Compare the thickness of the scar area with the thickness of the normal skin area adjacent to the scar and calculate the ratio. The formula is:

[0052] SEI = Thickness of Scar Area / Thickness of Normal Skin

[0053] The higher the SEI value, the more severe the scar hyperplasia.

[0054] 4. Histological Analysis

[0055] Samples need to collect both the scar and the normal skin tissue adjacent to the scar. The tissue is fixed in 4% paraformaldehyde solution for 24 hours, and then the tissue is soaked in decalcifying solution. The decalcifying solution is changed every 2 - 3 days. After 3 weeks, the tissue decalcification is completed. Then the tissue is rinsed overnight with running water, dehydrated, embedded, sectioned, and then stained with hematoxylin - eosin (HE) and Masson.

[0056] 5. HE Staining

[0057] The specific steps of HE staining are as follows: The paraffin sections are first baked in an oven at 65°C for 1 hour, and then the sections are successively placed in xylene I (15 minutes) - xylene II (15 minutes) - absolute ethanol I (5 minutes) - absolute ethanol II (5 minutes) - 95% ethanol (5 minutes) - 85% ethanol (5 minutes) - 70% ethanol (5 minutes) for dewaxing. Stain with hematoxylin solution for 5 seconds, differentiate with 1% hydrochloric acid alcohol for several seconds, and blue with 1% ammonia water solution for several seconds. Then the sections are successively dehydrated in gradient alcohols of 70%, 85%, and 95%, and stained with eosin solution for 1 minute. The sections are successively placed in absolute ethanol I (5 minutes) - absolute ethanol II (5 minutes) - absolute ethanol III (5 minutes) - xylene I (5 minutes) - xylene II (5 minutes) for clearing. Mount the sections with neutral gum, scan the sections, and read the sections.

[0058] 6. Masson Staining

[0059] The specific steps of Masson staining are as follows: After dewaxing the paraffin sections to water, they are placed in a 2.5% potassium dichromate mordant in an oven at 65 °C for 30 minutes. Then the sections are immediately washed in running water for 30 s until the yellow color on the tissue fades. Subsequently, the sections are immersed in Weigert iron hematoxylin staining solution for 5 s and differentiated in 1% hydrochloric acid alcohol for 1 min until the cell nuclei are grayish black and the background is light gray or colorless. Then, the sections are immersed in preheated ponceau acidic fuchsin solution for 5 min and rinsed with running water for about 20 s until the water flowing from the sections is colorless. The sections are slightly drained of water and then immersed in 1% phosphomolybdic acid solution for differentiation. Observation under the microscope shows that the differentiation is completed when the collagen fibers turn light red. Next, the sections are stained in 2.5% aniline blue solution for 20 s and rinsed and differentiated with 1% aqueous acetic acid solution. Finally, after dehydration with absolute ethanol, the sections are cleared in xylene for 5 min, sealed with neutral gum, scanned, and read.

[0060] II. Experimental Results

[0061] 1. Scar appearance and thickness results

[0062] The scar appearance results are as shown in A of Figure 1 : By macroscopic observation of the wound area, it can be found that the scars in the control group are firm, red, and protrude from the skin surface, while the scars in the experimental group are significantly softer, flatter, and the color is closer to that of the surrounding skin in the general view. This indicates that the scars after treatment with danshensu hydrogel are significantly removed, and the best effects are achieved when the concentration of danshensu is 100 μM and 400 μM.

[0063] The scar thickness results are as shown in B of Figure 1 : The results of measuring the scar thickness with a vernier caliper show that compared with the control group, the scar thickness in the experimental group is significantly reduced. Among them, when the concentration of danshensu is 100 μM and 400 μM, the scar thickness is significantly lower than that at 800 μM.

[0064] The scar elevation index results are as shown in Figure 2 : Compared with the control group, when the concentration of danshensu is 100 μM, 400 μM, and 800 μM, the scar elevation index (SEI) of rabbit ear scars is significantly reduced. In particular, the higher the concentration of danshensu, the greater the increase in the SEI index. This indicates that when treating hypertrophic scars, it is preferred to use a concentration of 100 μM of danshensu, which can effectively reduce the scar elevation index.

[0065] 2. Histological analysis results

[0066] HE and Masson staining are as shown in Figure 3Shown as follows: The scar collagen in the control group was dense, and large collagen fiber bundles could be seen. After treating the rabbit ear scars with danshensu hydrogels at 100 μM, 400 μM, and 800 μM, the collagen was sparser and no obvious large collagen fiber bundles were seen. It is indicated that danshensu hydrogel can effectively inhibit the proliferation and aggregation of collagen.

[0067] The above results show that danshensu hydrogel significantly reduces the thickness of scar hyperplasia and effectively inhibits the proliferation of collagen, and can be used for the prevention and treatment of hypertrophic scars. In particular, when the concentration of danshensu in the hydrogel is 100 μM and 400 μM, the treatment effect is optimal.

[0068] The danshensu hydrogel dosage form is used for local administration, which is convenient for operation. Compared with treatment methods such as oral drugs (such as compound danshen dripping pills) and radiotherapy, it can improve the compliance of patients. The danshensu hydrogel dosage form can be evenly applied to the surface of the wound and quickly form a film to cover it, so as to extend the local action time of the drug, increase the local concentration of danshensu drug, and achieve a better treatment effect. And the production process of danshensu and the danshensu hydrogel dosage form is simple, with low cost, and can provide an economical and effective treatment plan for patients.

[0069] As can be seen from the above examples and experimental examples, in the present invention, by screening the formulation composition and its ratio of the hydrogel, danshensu hydrogels with different concentrations were respectively prepared. Experiments show that danshensu hydrogel can effectively prevent and treat hypertrophic scars. The danshensu hydrogel dosage form is used for local administration, which is convenient for operation. Compared with treatment methods such as oral drugs (such as compound danshen dripping pills) and radiotherapy, it can improve the compliance of patients. The danshensu hydrogel dosage form can be evenly applied to the surface of the wound and quickly form a film to cover it, extending the local action time of the drug, increasing the local concentration of danshensu drug, and achieving a better treatment effect. And the production process of the danshensu hydrogel dosage form is simple, with low cost, and can provide an economical and effective treatment plan for patients, having good prospects in the prevention or treatment of collagen hyperplastic diseases.

Claims

1. A hydrogel for treating collagen hyperplasia diseases, characterized in that, It consists of raw materials with the following mass concentrations: Active ingredient 0.019 - 0.08 mg / mL, moisturizing and repairing agent 15 - 25 mg / mL, gelling agent 5 - 15 mg / mL, thickening agent 5 - 10 mg / mL, antioxidant 0.1 - 1 mg / mL; The active ingredient is danshensu.

2. The hydrogel for treating collagen hyperplastic diseases according to claim 1, characterized in that: The moisturizing and repairing agent is selected from at least one of sodium hyaluronate, glycerol, trehalose, chitosan, and collagen; the gelling agent is at least one of carbomer and poloxamer; the thickening agent is selected from at least one of carboxymethyl cellulose and polyvinyl alcohol.

3. The hydrogel for treating collagen hyperplastic diseases according to claim 2, wherein: The sodium hyaluronate is a mixture of sodium hyaluronates with different molecular weights, and the mixture is a combination of sodium hyaluronate with a molecular weight of 100 - 200 kDa, sodium hyaluronate with a molecular weight of 200 - 400 kDa, and sodium hyaluronate with a molecular weight of 1000 - 1500 kDa in a weight ratio of 1 - 10:1 - 10:1 - 10; And / or, the carbomer is at least one of carbomer 940, carbomer 934, and carbomer 980; the antioxidant is at least one of vitamin C, lipoic acid, and glutathione.

4. The preparation method of the hydrogel for treating collagen hyperplasia diseases according to any one of claims 1-3, characterized in that, It includes: Mix and stir the active ingredient, moisturizing and repairing agent, gelling agent, and thickening agent to obtain it.

5. Use of the hydrogel for preventing or treating collagen hyperplasia diseases according to any one of claims 1 - 3 in the preparation of a drug for preventing or treating collagen hyperplasia diseases.

6. Use of the hydrogel for preventing or treating collagen hyperplasia diseases according to claim 5 in the preparation of a medicament for preventing or treating collagen hyperplasia diseases, characterized in that: The collagen hyperplasia disease is a pathological scar disease.

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