Porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis and preparation method thereof

Abstract

The invention provides a porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis and a preparation method, and relates to the technical field of biological materials. The porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis comprises a needle body and a matrix layer, the needle body is a porous drug-loading autolysis hyaluronic acid microneedle body, and the matrix layer is a magnetic hyaluronic acid matrix. The invention has the following beneficial effects: 1, a soluble biocompatible microneedle structure is adopted, so that the injury caused by the breakage of a metal microneedle is avoided, and the medicine can directly reach the subcutaneous part; 2, a porous structure is constructed through ice crystal recrystallization, and the drug loading content is increased through a secondary drying technology; 3, the drug carrier is added through the nano magnetic particles, so that diffusion and absorption of drug components are promoted; 4, the operation is simple, tedious steps such as injection in the articular cavity are not needed, and the infection risk in the articular cavity is reduced; 5, batch mold production can be realized, and the production cost is reduced; 6, the administration amount is increased through microneedle perforation administration and matrix layer permeation micropore administration.

Description

Technical Field

[0001] The invention relates to the technical field of biomaterials, and in particular to a porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis and a preparation method thereof. Background Art

[0002] Knee osteoarthritis (KOA) is a common joint disease that develops from degenerated articular cartilage and has become an increasingly serious problem in the elderly population. Among the clinical manifestations of KOA, pain is the most common symptom, which directly affects the patient's lower limb physical function.

[0003] The main goals of clinical management of KOA are pain relief, cartilage regeneration, and functional restoration. Intra-articular injections (IAI), involving medications such as corticosteroids (CSs), can provide modest pain relief and slight functional improvements, although treatment outcomes are poor in patients with moderate to severe KOA. Physical therapy (PT) with medications and exercise training have been identified as the most promising interventions for reducing pain and improving mobility in the early stages of KOA. In addition, a number of biologics have been developed and used for cartilage repair in patients with KOA; among these biologics, hyaluronic acid (HA), platelet-rich plasma (PRP), and mesenchymal stem cells (MSCs) have been approved in clinical trials. In a multidisciplinary approach to treating KOA, a combination therapy regimen of IAI medications plus PT may be considered the optimal strategy, which can significantly improve pain and function in all disease stages until surgical treatment is required.

[0004] However, the prior art has the following technical problems: 1. Due to the closed negative pressure environment in the knee joint cavity, injection into the joint capsule has high requirements for aseptic properties. 2. The joint capsule lacks vascular structures, and the articular cartilage and joint capsule tissue have little blood supply. Oral administration and other routes of administration cannot achieve effective blood drug concentrations, and once infected, it is difficult to cure. 3. The treatment of knee osteoarthritis requires long-term and stable administration, which most patients cannot adhere to. Therefore, how to treat knee osteoarthritis more effectively has become a technical problem that needs to be solved urgently. Summary of the invention

[0005] In order to solve the technical problems existing in the prior art, the embodiment of the present invention provides a porous magnetic hyaluronic acid microneedle and a preparation method for treating knee osteoarthritis. The technical solution is as follows:

[0006] A method for preparing porous magnetic hyaluronic acid microneedles for treating knee osteoarthritis comprises the following steps:

[0007] 1) Take a certain amount of hyaluronic acid, add purified water, swell at 25-35°C, remove the bubbles in the solution, and obtain a uniform hyaluronic acid clear solution;

[0008] 2) Take the clarified hyaluronic acid solution and naturally cast it onto the PDMS microneedle mold;

[0009] 3) The PDMS microneedle mold coated with the clear hyaluronic acid solution is placed in a vacuum drying oven, and after repeated vacuuming, pressure holding and degassing operations, it is taken out and quickly frozen in liquid nitrogen to -80°C to -100°C to form polycrystalline ice crystals, and then the ice crystals are remelted at -5°C to -10°C;

[0010] 4) placing the material of step 3) in a vacuum drying oven and drying at room temperature until the microneedle is demolded, adding a high concentration solution containing the drug into the mold and drying to obtain a porous drug-loaded self-soluble hyaluronic acid microneedle body;

[0011] 5) Take a certain amount of hyaluronic acid, add purified water, swell at 25-35°C, add a solution of ferroferric oxide nanopowder, remove the bubbles in the solution to obtain a uniform hyaluronic acid solution; and

[0012] 6) The microneedle carrier is poured again to form a magnetic matrix, which is placed in a vacuum drying oven and dried at room temperature until the microneedles are demoulded, thereby obtaining the porous magnetic hyaluronic acid microneedles for treating knee osteoarthritis.

[0013] Optionally, in step 1), the molecular weight of the hyaluronic acid is 1-10 KDa.

[0014] Optionally, in step 1), the mass volume ratio (g / mL) of the hyaluronic acid to the purified water is 1:50.

[0015] Optionally, the vacuum drying conditions in step 3) are: 30° C., evacuate to 0.06 MPa, maintain pressure for 2 to 3 minutes, and slowly release air, and repeat the vacuuming, pressure maintaining and releasing operations under the above conditions for 3 times;

[0016] And / or, the PDMS microneedle mold is placed in liquid nitrogen and quickly frozen to -90°C to form polycrystalline ice crystals, and then the ice crystals are re-thawed at -6°C.

[0017] Optionally, in step 4), the drug is hyaluronic acid and a pre-designed drug carrier, and the drug carrier is selected from one or more of the group consisting of diclofenac diethylamide, celecoxib and indolecoxib.

[0018] Optionally, in step 4), the concentration of the high-concentration drug-containing solution is 0.01-1 g / ml.

[0019] Optionally, in step 4), the concentration of the high-concentration drug-containing solution is 0.1 g / ml.

[0020] Optionally, in step 5), the concentration of the solution of the ferrosoferric oxide nanopowder is 0.1 g / mL, and / or the diameter of the ferrosoferric oxide nanopowder is less than 50 µm.

[0021] The porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis prepared according to the method comprises a needle body and a matrix layer, wherein the needle body is a porous drug-loaded self-soluble hyaluronic acid microneedle needle body, and the matrix layer is a magnetic hyaluronic acid matrix.

[0022] Application of the porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis in preparing a medicine for treating knee osteoarthritis.

[0023] The beneficial effects brought about by the technical solution provided by the embodiment of the present invention include at least:

[0024] 1. The use of a soluble biocompatible microneedle structure avoids damage caused by metal microneedle breakage and allows the drug to reach the subcutaneous tissue directly; 2. A porous structure is constructed through ice crystal recrystallization, and the secondary drying technology is used to increase the drug content; 3. Nanomagnetic particles are added to the drug carrier to promote the diffusion and absorption of drug ingredients; 4. The operation is simple, without the need for tedious steps such as intra-articular injection, and the risk of intra-articular infection is reduced; 5. Mass mold production can be achieved, reducing production costs; 6. The drug administration is increased through microneedle perforation and matrix layer penetration micropore administration. BRIEF DESCRIPTION OF THE DRAWINGS

[0025] In order to more clearly illustrate the technical solutions in the embodiments of the present invention, the following briefly introduces the drawings required for use in the description of the embodiments. Obviously, the drawings described below are only some embodiments of the present invention. For ordinary technicians in this field, other drawings can be obtained based on these drawings without creative work.

[0026] Figure 1 Schematic diagram of a microneedle and a mold provided by an embodiment of the present invention;

[0027] Figure 2 It is a virtual electron microscope schematic diagram of the microneedle ice crystal after recrystallization provided by the present invention;

[0028] Figure 3 This is a microscopic photograph of the microneedle puncturing the sealing film. DETAILED DESCRIPTION

[0029] The technical solution of the present invention is described below in conjunction with the accompanying drawings.

[0030] Microneedles (MNs) are a new type of physical penetration technology that can directly penetrate the skin to deliver drugs. They can allow drug molecules to directly cross the stratum corneum barrier in a minimally painful and non-invasive manner. They have outstanding properties such as good therapeutic effects, relative safety, and low cost. They have attracted widespread attention in the field of transdermal drug delivery in the past few decades. Soluble microneedles in microneedles are widely used because they can encapsulate drugs and dissolve after being inserted into the skin, and have good biocompatibility. Different soluble microneedles can be made according to treatment needs.

[0031] The embodiments of the present application provide a method and application for preparing magnetically thermal drug-loaded hyaluronic acid microneedles based on ice crystal recrystallization.

[0032] The four characteristics of hyaluronic acid: originating from the human body, strong biocompatibility, strong water retention, and viscoelastic and lubricating properties, make it an ideal material for preparing soluble microneedles.

[0033] The purpose of the present invention is to provide a preparation method and application of magnetic thermal drug-carrying hyaluronic acid microneedles based on ice crystal recrystallization, and to test its performance.

[0034] To achieve the above object, the technical solution adopted by the present invention is as follows:

[0035] Example 1

[0036] A method for preparing self-soluble hyaluronic acid microneedles comprises the following steps:

[0037] Take a certain amount of hyaluronic acid, add appropriate amount of purified water, swell in a water bath at 25-35℃ for 25-35min, let stand at room temperature for 10min, and then remove the bubbles in the solution under vacuum (30℃, evacuate to 0.06MPa, maintain pressure for 4min, slowly release air, repeat the above conditions of evacuation, pressure maintenance and release for 3 times). Obtain a uniform clear hyaluronic acid solution for use.

[0038] Clean the PDMS microneedle mold, air-dry it, and set aside.

[0039] Take the clear hyaluronic acid solution and naturally cast it onto the PDMS microneedle mold, shaking it back and forth to ensure uniform coating;

[0040] The PDMS microneedle mold coated with a clear hyaluronic acid solution is placed in a vacuum drying oven, and after repeated vacuuming, pressure maintenance and degassing operations, it is taken out and quickly frozen in liquid nitrogen to -90°C to form polycrystalline ice crystals, and then the ice crystals are remelted at -6°C (the purpose of ice crystal recrystallization is to increase the drug loading capacity).

[0041] The material is then placed in a vacuum drying oven, desiccant is added, and dried at room temperature for 24 hours until the microneedles are demolded. The pre-applied high-concentration drug-containing solution is then added to the mold, and then dried immediately to obtain porous drug-loaded self-soluble hyaluronic acid microneedles (the principle of this step is that ice crystals first crystallize to form cavities, which are then dried to remove water, and then the drug solution is filled in. After drying, the drug crystallizes in the cavities).

[0042] Take a certain amount of hyaluronic acid, add an appropriate amount of purified water, swell in a water bath at 25-35°C for 25-35 minutes, let stand at room temperature for 10 minutes, add 1g of ferrosoferric oxide nanopowder with a diameter less than 50µm and mix with 10ml of the solution, then vacuum remove the bubbles in the solution to obtain a uniform hyaluronic acid solution for use.

[0043] The above-mentioned microneedle carrier is poured again to form a magnetic matrix, and then the material is placed in a vacuum drying oven, a desiccant is added, and dried at room temperature for 24 hours until the microneedles are demolded to obtain porous magnetic hyaluronic acid microneedles.

[0044] The molecular weight of the hyaluronic acid is 1-10 KDa, the mass volume ratio (g / mL) of hyaluronic acid to the purified water is 1:50, and the vacuum drying conditions of all the steps are: 30°C, vacuuming to 0.06 MPa, maintaining pressure for 2-3 minutes, and slowly releasing air, and repeating the vacuuming, maintaining pressure and releasing air under the above conditions for 3 times.

[0045] The PDMS microneedle mold is easy to demould, easy to use, and can be reused repeatedly.

[0046] Example 2 Simulated human skin test

[0047] Taking into account the elasticity of the skin, a single-layer Parafilm sealing film was used to simulate the skin. The microneedle prepared by the present invention was pressed vertically on the sealing film with a force of 5N by the thumb for 2 minutes and then pulled out. The holes left on the sealing film were observed under a microscope, and the number of pinholes left after the microneedle passed through the sealing film was counted to evaluate the mechanical properties of the microneedle.

[0048] After three experiments, the number of pinholes / microneedles was >90%, and the surface microneedles could penetrate a single layer of Parafilm sealing film, that is, they could penetrate the human stratum corneum.

[0049] The results of simulated human skin tests have shown that the microneedles prepared by the method of the present invention have sufficient mechanical strength to pierce the epidermis of the skin to form micron-scale channels, thereby increasing the transdermal permeability of hyaluronic acid.

[0050] Example 3 Testing the Autodissolution Performance of Microneedles Using Ethanol of Different Concentrations

[0051] Since the water content of the skin is different at different depths, in order to study the solubility of the self-dissolving microneedles prepared by the method of the present invention at different positions of the skin, the inventors prepared ethanol aqueous solutions with mass fractions of 80wt%, 60wt%, 40wt%, and 20wt% to simulate skin tissue fluid, that is, the water content in the solution was 20wt%, 40wt%, 60wt%, and 80wt%, respectively. The prepared hyaluronic acid self-dissolving microneedles were placed in a 96-well plate, and 200 μL of ethanol solutions with water contents of 20wt%, 40wt%, 60wt%, and 80wt% were added respectively; optical microscope (1 40 times) to observe and record the dissolution process of the microneedles, and record the time when the microneedles are completely dissolved. Figure 3 shown.

[0052] It can be concluded from the above experiment that the hyaluronic acid microneedle is insoluble in 100% ethanol, dissolves quickly in pure water, and can dissolve in a solution of 40% ethanol (i.e., 60% water content) in 90s, because the water content of the skin at different depths varies from 20% to 70%, for example, the water content of the stratum corneum is 20%-30wt%, and increases in a gradient from the outside to the inside, and the water content of the deeper skin granular layer is as high as 70%. This verifies that the hyaluronic acid microneedle prepared by the method of the present invention can dissolve by itself through the epidermis of the skin.

[0053] Application method of the microneedle of the present invention

[0054] Place the microneedle on the skin on both sides of the patella, and press the microneedle vertically with a force of 5N for 2 minutes with the thumb. Then place the microneedle in a changing magnetic field to make the ferroferric oxide nanopowder heat up to promote blood circulation in the surrounding skin and drug absorption, so as to achieve the purpose of treatment.

[0055] The above is only a specific embodiment of the present invention, but the protection scope of the present invention is not limited thereto. Any person skilled in the art can easily think of changes or substitutions within the technical scope disclosed by the present invention, which should be included in the protection scope of the present invention. Therefore, the protection scope of the present invention should be based on the protection scope of the claims.

Claims

1. A method for preparing porous magnetic hyaluronic acid microneedles for treating knee osteoarthritis, characterized in that: The steps include: 1) Take a certain amount of hyaluronic acid, add purified water, swell at 25-35°C, remove the bubbles in the solution, and obtain a uniform hyaluronic acid clear solution; 2) Take the clarified hyaluronic acid solution and naturally cast it onto the PDMS microneedle mold; 3) The PDMS microneedle mold coated with the clear hyaluronic acid solution is placed in a vacuum drying oven, and after repeated vacuuming, pressure holding and degassing operations, it is taken out and quickly frozen in liquid nitrogen to -80°C to -100°C to form polycrystalline ice crystals, and then the ice crystals are remelted at -5°C to -10°C; 4) placing the material of step 3) in a vacuum drying oven and drying at room temperature until the microneedle is demolded, adding a high concentration solution containing the drug into the mold and drying to obtain a porous drug-loaded self-soluble hyaluronic acid microneedle body; 5) Take a certain amount of hyaluronic acid, add purified water, swell at 25-35°C, add the solution of ferroferric oxide nanopowder, remove the bubbles in the solution to obtain a uniform hyaluronic acid solution; as well as 6) The microneedle carrier is poured again to form a magnetic matrix, which is placed in a vacuum drying oven and dried at room temperature until the microneedles are demoulded, thereby obtaining the porous magnetic hyaluronic acid microneedles for treating knee osteoarthritis.

2. The method according to claim 1, characterized in that In step 1), the molecular weight of the hyaluronic acid is 1-10 KDa.

3. The method according to claim 1, characterized in that In step 1), the mass volume ratio (g / mL) of the hyaluronic acid to the purified water is 1:

50.

4. The method according to claim 1, characterized in that: Step 3) Medium vacuum drying conditions: 30°C, evacuate to 0.06MPa, maintain pressure for 2-3 minutes, and slowly release air. Repeat the above conditions of evacuating, maintaining pressure and releasing air 3 times; And / or, the PDMS microneedle mold is placed in liquid nitrogen and quickly frozen to -90°C to form polycrystalline ice crystals, and then the ice crystals are re-thawed at -6°C.

5. The method according to claim 1, characterized in that In step 4), the drug is hyaluronic acid and a pre-designed drug carrier, and the drug carrier is selected from one or more of the group consisting of diclofenac diethylamide, celecoxib and indolecoxib.

6. The method according to claim 1, characterized in that In step 4), the concentration of the high concentration solution containing the drug is 0.01-1 g / ml.

7. The method according to claim 6, characterized in that In step 4), the concentration of the high concentration solution containing the drug is 0.1 g / ml.

8. The method according to claim 1, characterized in that In step 5), the concentration of the solution of the ferrosoferric oxide nanopowder is 0.1 g / mL, and / or the diameter of the ferrosoferric oxide nanopowder is less than 50 µm.

9. The porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis prepared according to the method of any one of claims 1 to 8, characterized in that: The porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis comprises a needle body and a matrix layer. The needle body is a porous drug-loaded self-soluble hyaluronic acid microneedle needle body, and the matrix layer is a magnetic hyaluronic acid matrix.

10. Use of the porous magnetic hyaluronic acid microneedle for treating knee osteoarthritis according to claim 9 in preparing a drug for treating knee osteoarthritis.

Images