Drug-loaded acupuncture needle as well as preparation method and application thereof

Abstract

The invention relates to a medicine-carrying acupuncture needle and a preparation method and application thereof in the technical field of combined treatment of traditional Chinese medicine and western medicine, the medicine-carrying acupuncture needle comprises a needle body, and the surface of the needle body is subjected to glow discharge treatment; the drug-loaded hydrogel is at least uniformly coated on the surface near the tip of the needle body and is cured and attached to the surface of the part of the needle body; the drug-loaded hydrogel comprises a dual-network hydrogel, a conductive polymer and drug molecules which are uniformly mixed. According to the medicine-carrying acupuncture needle, glow discharge treatment is carried out on the surface of the needle body, the multifunctional hydrogel medicine-carrying coating is constructed, active ingredients of icariin and the acupuncture needle are organically combined, and an accurate treatment mode of needle and medicine cooperation is achieved; the medicine carrying acupuncture needle system has the advantages of being high in medicine carrying rate, controllable in release and the like, intelligent medicine administration can be achieved under the electrical stimulation condition, achilles tendon tissue repair can be remarkably promoted, inflammatory response can be inhibited, the curative effect is better than that of a single treatment mode, and a safe and effective innovative treatment scheme is provided for motor system injuries.

Description

Technical Field

[0001] This application relates to the field of integrated traditional Chinese and Western medicine treatment technology, and in particular to a drug-loaded acupuncture needle, its preparation method and application. Background Technology

[0002] Tendonopathy is a degenerative condition of the tendon caused by multiple factors. Its main pathological features are degeneration of tendon tissue and metabolic imbalance of the extracellular matrix. Achilles tendinitis is a common musculoskeletal disorder. When the Achilles tendon is subjected to excessive pressure in a short period, it may suffer strain, contusion, or tear, leading to aseptic Achilles tendinitis. Achilles tendinitis has a high incidence rate among people who exercise frequently, especially track and field athletes, with an incidence of approximately 5.6%. Current clinical treatments primarily focus on conventional rehabilitation therapies such as pain relief and reducing local inflammation, as well as surgery. However, these treatments are slow to take effect, difficult to cure, and have a high recurrence rate.

[0003] As the core tool of acupuncture treatment, acupuncture needles are carefully designed in shape and material to precisely stimulate acupoints, regulate the flow of qi and blood, and achieve the purpose of treating diseases. However, traditional acupuncture needles only have the function of physical stimulation. Although they can be functionally modified by coating the surface of acupuncture needles with drug-loaded gel, there are still problems such as low drug loading capacity of the drug-loaded gel, easy detachment of hydrogel from the acupuncture needle, and lack of controllable release function, which makes their therapeutic effect on tendon degenerative diseases such as Achilles tendinitis very limited. Summary of the Invention

[0004] The main objective of this invention is to provide a drug-loaded acupuncture needle, its preparation method, and its application, aiming to solve at least one of the above-mentioned technical problems.

[0005] To achieve the above objectives, the present invention proposes a drug-loaded acupuncture needle, comprising: a needle body, the surface of which is treated with glow discharge; a drug-loaded hydrogel, wherein the drug-loaded hydrogel is uniformly coated at least on the surface near the tip of the needle body and is solidified and attached to that portion of the needle body surface; the drug-loaded hydrogel comprises a uniformly mixed dual-network hydrogel, a conductive polymer, and drug molecules.

[0006] In some embodiments of the present invention, the dual-network hydrogel is constructed using carboxymethyl chitosan-gluconolactone and methacrylamide silk fibroin.

[0007] In some embodiments of the present invention, the conductive polymer component is PEDOT:PSS.

[0008] In some embodiments of the present invention, the drug molecule is icariin.

[0009] To achieve the above objectives, the present invention also proposes a method for preparing the above-mentioned drug-loaded acupuncture needle, comprising the following steps: preparing a needle body and subjecting the needle body to glow discharge treatment; uniformly mixing a dual-network hydrogel, a conductive polymer, and drug molecules to obtain a drug-loaded hydrogel; uniformly coating the drug-loaded hydrogel onto the surface of the glow discharge-treated needle body and fixing it in place to obtain the drug-loaded acupuncture needle.

[0010] In some embodiments of the present invention, the needle body is alkali washed before undergoing glow discharge treatment.

[0011] In some embodiments of the present invention, the parameters of the glow discharge treatment are: gas pressure of 0.36 mbar to 0.42 mbar, discharge current of 10 mA to 20 mA, and time of 20 s to 90 s.

[0012] In some embodiments of the present invention, the preparation of the drug-loaded hydrogel includes the following steps: dissolving carboxymethyl chitosan in ammonia and calcium hydroxide solution, stirring to obtain a carboxymethyl chitosan solution; mixing gluconate-δ-lactone with the carboxymethyl chitosan solution, stirring until the system becomes a transparent gel to obtain a first gel solution; adding icariin to a dispersion of poly(3,4-ethylenedioxythiophene):polystyrene sulfonate, ultrasonically dispersing to obtain a mixed solution; adding methacrylamide silk fibroin to the mixed solution, stirring until completely dissolved to obtain a second gel solution; mixing the first gel solution and the second gel solution, adding a photoinitiator, stirring evenly to obtain the drug-loaded hydrogel.

[0013] In some embodiments of the present invention, the fixation and bonding of the needle body after coating with drug-loaded hydrogel includes the following steps: irradiating the needle body with an ultraviolet lamp, and drying the needle body after the surface of the needle body has solidified.

[0014] To achieve the above objectives, the present invention also proposes the application of the above-mentioned drug-loaded acupuncture needle or the drug-loaded acupuncture needle prepared by the above-mentioned method in the preparation of medical devices for treating Achilles tendonopathy.

[0015] Compared with the prior art, the present invention has the following beneficial effects: This invention addresses the insufficient synergy between acupuncture and medication in the treatment of Achilles tendinitis by proposing a novel electroresponsive drug-loaded acupuncture system. Compared to traditional acupuncture treatment that relies solely on mechanical stimulation and conventional drug delivery suffers from poor targeting, this invention constructs a multifunctional hydrogel drug-loaded coating to organically combine the active ingredient of icariin with acupuncture needles, achieving a precise treatment mode of "acupuncture-drug synergy." This system features high drug loading rate and controllable release, enabling intelligent drug delivery under electrostimulation conditions. Animal experiments have demonstrated that this drug-loaded acupuncture needle can significantly promote Achilles tendon tissue repair and inhibit inflammatory responses, with efficacy superior to single treatment methods, providing a safe and effective innovative treatment option for musculoskeletal injuries. Attached Figure Description

[0016] Various other advantages and benefits will become apparent to those skilled in the art upon reading the following detailed description of preferred embodiments. The accompanying drawings are for illustrative purposes only and are not intended to limit the scope of this application. Furthermore, the same reference numerals denote the same parts throughout the drawings. In the drawings: Figure 1 This is a schematic diagram of the structure of the drug-loaded acupuncture needle of the present invention; Figure 2 This is a flowchart illustrating the preparation process of the drug-loaded acupuncture needle of the present invention; Figure 3 Scanning electron microscope images of various hydrogel synthesis methods; Figure 4 Infrared spectra of various hydrogels synthesized; Figure 5 Scanning electron microscope and elemental distribution map of each step in the synthesis of icariin-loaded acupuncture needles; Figure 6 Figure showing the swelling results of the hydrogel in Test Example 4; Figure 7 Natural and electro-responsive drug release curves of icariin-loaded hydrogels; Figure 8 The drug release curves of the icariin-loaded hydrogel at different constant potentials; Figure 9 Comparison of the surface adhesion strength of various hydrogels with glow discharge treated or untreated acupuncture needle body materials; Figure 10 A diagram illustrating the biocompatibility test of icariin-loaded hydrogels; Figure 11 This is a diagram illustrating the biosafety testing of drug-loaded acupuncture. Figure 12 A graph showing the hemolytic effect of drug-loaded acupuncture on rabbit erythrocytes; Figure 13 This is a behavioral assessment of mice after drug-loaded acupuncture combined with treatment. Figure 14Pathological staining images of Achilles tendon tissues after different treatments. Detailed Implementation

[0017] The present invention will be further described below with reference to specific embodiments. It should be understood that the specific embodiments described herein are for illustrative purposes only and are not intended to limit the scope of the invention. Those skilled in the art will readily identify several non-critical parameters that can be varied or modified without yielding substantially the same results.

[0018] Unless otherwise specified, the techniques or conditions described in the examples shall be performed in accordance with the techniques or conditions described in the literature in this field, or in accordance with the product instructions. Reagents or instruments whose manufacturers are not specified are all conventional products that can be purchased through legitimate channels. Unless otherwise specified, the experimental methods in the following examples are conventional methods. Unless otherwise specified, the experimental materials used in the following examples are all commercially available products.

[0019] Currently, materials science and traditional Chinese medicine are mostly combined in terms of drug structure or dosage form. There are few existing integrated drug delivery systems for acupuncture needles, which face problems such as easy drug detachment, low drug loading capacity, complex manufacturing process, and weak bonding with the needle body, which affects subsequent development and application. Therefore, this invention proposes an integrated drug delivery system using a surface-hydroxylated acupuncture needle and a multi-hydroxyl dual-network conductive hydrogel. This system can achieve the dual purpose of precise drug delivery and exerting the effects of acupuncture. This process is easy to industrialize and can also be applied to carry other drugs, which is of great significance for broadening the types of clinical diseases and improving the efficacy.

[0020] This application discloses a drug-loaded acupuncture needle. For example... Figure 1 As shown, the drug-loaded acupuncture needle includes a needle body 1 and a drug-loaded hydrogel 2; wherein, the surface of the needle body 1 is treated with glow discharge; the drug-loaded hydrogel 2 is uniformly coated on the surface near the tip of the needle body 1 and is solidified and attached to this part of the needle body 1 surface; the drug-loaded hydrogel 2 includes a uniformly mixed double network hydrogel, a conductive polymer and drug molecules.

[0021] In this invention, by performing glow discharge treatment on the needle surface, the number of functional groups available for chemical modification on the needle surface can be significantly increased. Based on the interaction between the dual-network hydrogel and the conductive polymer with the functional groups on the needle surface (inter-functional forces), the adhesion performance between the drug-loaded hydrogel and the needle surface can be significantly enhanced, making the drug-loaded hydrogel firmly bonded to the needle and effectively reducing the possibility of hydrogel detachment. Moreover, the dual-network hydrogel and the conductive polymer have a rich three-dimensional network structure, which effectively increases the drug loading capacity. At the same time, the introduction of the conductive polymer into the drug-loaded hydrogel as a carrier for electro-responsive drug release can achieve controlled drug release by utilizing the structure and charge changes induced by the electric field, effectively avoiding non-responsive release. Thus, by utilizing the firm bond between the drug-loaded hydrogel and the needle and the electro-responsive controlled drug release, the precise delivery and controlled release of the drug can be achieved, combining the therapeutic effects of acupoint stimulation and drug synergy, and exhibiting good biocompatibility and safety, thereby significantly improving the efficacy of acupuncture in the treatment of Achilles tendonopathy.

[0022] In some embodiments of the present invention, the dual-network hydrogel is constructed using carboxymethyl chitosan-gluconolactone (CMCS-GDL) and methacrylamide silk fibroin (SilMA).

[0023] In this embodiment, firstly, the multi-hydroxyl molecular structure of CMCS-GDL has an optimizable gel viscosity, which can significantly enhance the adhesion performance between the drug-loaded hydrogel and the needle surface. Secondly, by utilizing the photocuring properties of SilMA, the drug-loaded hydrogel can be rapidly molded and its mechanical strength can be improved. Thus, while ensuring interfacial adhesion, the gel is endowed with excellent mechanical strength, effectively solving the problems of drug burst release and insufficient mechanical properties of traditional drug-loaded acupuncture needles. At the same time, the inherent tissue regeneration and repair effect and stem cell regulation function of silk fibroin can synergistically enhance the therapeutic effect. In addition, the three main components of this dual-network hydrogel system (CMCS, GDL and SilMA) are all derived from natural materials and have excellent biocompatibility, which can effectively avoid immune reactions during the treatment process.

[0024] In some embodiments of the present invention, the conductive polymer component is poly(3,4-ethylenedioxythiophene):polystyrene sulfonate (PEDOT:PSS).

[0025] In this embodiment, PEDOT:PSS serves as a carrier for the electroresponsive release of drugs. This conductive polymer material can achieve effective drug loading through multiple mechanisms such as electrostatic interaction and physical encapsulation, and can utilize electric field-induced structural and charge changes to achieve effective and controllable drug release.

[0026] In some embodiments of the present invention, the drug molecule is icariin, and the molecular formula of icariin is shown below:

[0027] Icariin is the main active ingredient extracted from Epimedium and has a variety of pharmacological activities. In this invention, icariin can inhibit Achilles tendon sensory nerves by downregulating glutamate and its receptors, thereby inhibiting central sensitization and relieving and treating Achilles tendon end pain. Combined with conductive double-network hydrogel, the directional and controllable release of icariin is achieved under the stimulation of electrical signals, improving efficacy and reducing toxic side effects.

[0028] In some embodiments of the present invention, the needle body can be made of austenitic stainless steel, preferably 06Cr19Ni10 (SUS304), which has good conductivity and can be well used with electronic diagnostic instruments.

[0029] The drug-loaded acupuncture needle of this invention uses austenitic stainless steel as the needle body and combines it with a drug-loaded conductive double-network hydrogel. It has a rich and high-strength three-dimensional network structure, excellent biocompatibility, and abundant active groups. Icariin is stably loaded onto the surface of the acupuncture needle to construct a hydrogel-acupuncture needle composite system with minimally invasive and precise positioning characteristics. The intelligent hydrogel with electroresponsive characteristics realizes the controllable release of drugs under the stimulation of an external electric field, which significantly improves the application effect of acupuncture needles in the targeted treatment of deep tissues such as Achilles tendinitis.

[0030] This embodiment also proposes a preparation method for preparing the above-mentioned drug-loaded acupuncture needles, such as... Figure 2 As shown, the preparation method includes the following steps: S1. Prepare the needle body and perform glow discharge treatment on the needle body.

[0031] S2. The dual-network hydrogel, conductive hydrogel and drug molecules are uniformly mixed to obtain a drug-loaded hydrogel.

[0032] S3. The drug-loaded hydrogel is uniformly coated onto the surface of the needle body treated with glow discharge and fixed together to obtain the drug-loaded acupuncture needle.

[0033] The preparation method of this invention first involves treating the needle body with glow discharge, then uniformly mixing a double-network hydrogel, a conductive polymer, and drug molecules to obtain a drug-loaded hydrogel. The drug-loaded hydrogel is then uniformly coated onto the glow discharge-treated needle surface and fixed in place to obtain a drug-loaded acupuncture needle. The overall manufacturing process is simple and effectively combines the needle and drug, achieving multiple optimizations including high drug loading, strong hydrogel bonding, high hydrogel strength, and controllable drug release. On one hand, electrical stimulation of acupoints allows the body to exert its internal regulatory function; on the other hand, the drug exerts its therapeutic effect. Acupoint stimulation and drug treatment have synergistic and complementary effects, thereby significantly improving the treatment effect on Achilles tendonitis.

[0034] In some embodiments of the present invention, the needle body is alkali washed before undergoing glow discharge treatment.

[0035] The alkaline washing process of the present invention can not only effectively remove stains from the needle surface, but also provide alkaline conditions for the needle surface to attach -OH groups, thereby providing favorable conditions for the subsequent stable bonding and uniform distribution of the drug-loaded hydrogel with the needle, and further improving the bonding effect between the drug-loaded hydrogel and the needle.

[0036] In some embodiments of the present invention, the needle body alkaline washing process includes: firstly, ultrasonically cleaning the needle body in anhydrous ethanol for 5 min, then immersing it in sodium hydroxide solution, and then drying it with nitrogen gas.

[0037] Furthermore, the pH value of the sodium hydroxide solution is preferably 9-11, and the soaking time of the needle in the sodium hydroxide solution is preferably 30 s.

[0038] In some embodiments of the present invention, the parameters for glow discharge treatment are: gas pressure of 0.36 mbar to 0.42 mbar, discharge current of 10 mA to 20 mA, and time of 20 s to 90 s; for example, the gas pressure can be 0.36 mbar, 0.37 mbar, 0.38 mbar, 0.39 mbar, 0.40 mbar, 0.41 mbar, 0.42 mbar, etc.; the discharge current can be 10 mA, 11 mA, 12 mA, 13 mA, 14 mA, 15 mA, 16 mA, 17 mA, 18 mA, 19 mA, 20 mA, etc.; and the time can be 20 s, 30 s, 40 s, 50 s, 60 s, 70 s, 80 s, 90 s, etc.

[0039] In this invention, by performing glow discharge treatment on the needle surface using the above parameters, the needle can be hydroxylated, thereby forming a stable hydroxyl (-OH) functional group coating layer on the needle surface. At this time, the drug-loaded hydrogel rich in hydroxyl groups, especially the multi-hydroxyl CMCS-GDL, can achieve strong bonding with the hydroxyl-rich needle through intermolecular interactions such as hydrogen bonds, hydrophilic matching, and physical intercalation, ensuring high strength and high stability of the interface bonding, and further ensuring the stable bonding and uniform distribution of the drug-loaded hydrogel and the needle.

[0040] In some embodiments of the present invention, the needle hydroxylation process includes: placing the alkali-washed needle into a plasma glow discharge instrument and discharging it for 20 s to 90 s under the parameters of a gas pressure of 0.36 mbar to 0.42 mbar and a discharge current of 10 mA to 20 mA.

[0041] Furthermore, the glow discharge treatment can be repeated multiple times, preferably 2 to 4 times.

[0042] Furthermore, the needle treated with glow discharge should be used for subsequent operations as soon as possible, that is, the drug-loaded hydrogel should be applied to the surface of the needle as soon as possible.

[0043] In some embodiments of the present invention, the preparation of the drug-loaded hydrogel includes the following steps: S21. First, dissolve carboxymethyl chitosan in ammonia water and calcium hydroxide solution, stir, and obtain carboxymethyl chitosan solution.

[0044] S22. Then, gluconate-δ-lactone and carboxymethyl chitosan solution are mixed and stirred until the system becomes a transparent gel, thus obtaining the first gel solution.

[0045] S23. Add icariin (ICA) to the dispersion of poly(3,4-ethylenedioxythiophene):polystyrene sulfonate, and ultrasonically disperse to obtain a mixed solution.

[0046] S24. Add methacrylamide silk fibroin to the mixed solution and stir until completely dissolved to obtain the second gel solution.

[0047] S25. Mix the first gel solution and the second gel solution, add the photoinitiator, and stir evenly to obtain the drug-loaded hydrogel.

[0048] In some embodiments of the present invention, ammonia water and calcium hydroxide solution refer to the mixed system formed by adding calcium hydroxide (Ca(OH)2) to ammonia water.

[0049] Furthermore, the pH value of the ammonia water is preferably 7.4 to 11. Ca(OH)2 is mixed in the ammonia water by ultrasonic treatment, preferably for 10 minutes, so that Ca(OH)2 cannot be completely dissolved.

[0050] In some embodiments of the present invention, after carboxymethyl chitosan (CMCS) is dissolved in ammonia and calcium hydroxide solution in step S21, preferably, the mass fraction of CMCS is 5% to 15%, for example, the mass fraction of CMCS is 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, etc.

[0051] After adding carboxymethyl chitosan to ammonia and calcium hydroxide solution, stir for 30 to 60 minutes to fully dissolve it.

[0052] Furthermore, the degree of substitution of the selected CMCS is preferably greater than 80%.

[0053] In some embodiments of the present invention, after glucono-δ-lactone (GDL) is dissolved in carboxymethyl chitosan solution in step S22, preferably, the mass fraction of GDL is 5% to 15%, for example, the mass fraction of GDL is 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, etc.

[0054] When glucono-δ-lactone is mixed with a carboxymethyl chitosan solution, and the mixture is rapidly stirred, GDL reacts with Ca(OH)₂, causing the solution to become transparent and gradually releasing Ca. 2+ It cross-links with CMCS molecules to form a network structure.

[0055] It should be noted that in the aforementioned steps, an appropriate amount of Ca(OH)2 should be added to the ammonia solution. This ensures that calcium hydroxide effectively neutralizes gluconic acid-δ-lactone, releasing calcium ions for crosslinking of carboxymethyl chitosan, while avoiding excessive addition of Ca(OH)2. In other words, excessive Ca(OH)2 should be avoided as it cannot react with gluconic acid-δ-lactone, which would lead to problems such as reduced hydrogel transparency and decreased mechanical strength.

[0056] In some embodiments of the present invention, in step S23, ultrasonic dispersion treatment for 5 min to 20 min is preferred. Since the solubility of ICA in water is very small, and the equilibrium solubility of icariin in water at 25 °C is 15.04 mg / L, ultrasonic dispersion treatment for 5 to 20 min can make icariin uniformly dispersed in the mixture.

[0057] In some embodiments of the present invention, after the methacrylamide silk fibroin (SilMA) is dissolved in the mixed solution of step S23 in step S24, preferably, the mass fraction of SilMA is 5% to 15%, for example, the mass fraction of SilMA is 5%, 6%, 7%, 8%, 9%, 10%, 11%, 12%, 13%, 14%, 15%, etc.

[0058] When methacrylamide silk fibroin is fully dissolved in a mixed solution of icariin and poly(3,4-ethylenedioxythiophene):polystyrene sulfonate, the system is dark blue and slightly viscous.

[0059] Furthermore, when dissolving SilMA, it is important to cut it into small pieces beforehand, and the dissolution process can be accelerated by appropriately increasing the temperature.

[0060] In some embodiments of the present invention, in step S25, the ratio of the first gel solution and the second gel solution can be 1:1, and the photoinitiator can be lithium phenyl (2,4,6-trimethylbenzoyl) phosphate (LAP). The mass fraction of LAP after dissolution is preferably 0.25% to 1%, for example, the mass fraction of LAP is 0.25%, 0.30%, 0.35%, 0.40%, 0.45%, 0.50%, 0.55%, 0.60%, 0.65%, 0.70%, 0.75%, 0.80%, 0.85%, 0.90%, 0.95%, 1%, etc.

[0061] In some embodiments of the present invention, before using the drug-loaded hydrogel, the glass bottle containing the drug-loaded hydrogel can be wrapped with aluminum foil to block light and placed at 4 ℃ for short-term storage to prevent premature light curing.

[0062] In some embodiments of the present invention, the fixation and bonding of the needle body after coating with drug-loaded hydrogel includes the following steps: The needle is irradiated with ultraviolet light, and after the surface of the needle has solidified, the needle is dried.

[0063] In some embodiments of the present invention, the ultraviolet lamp includes short-wave ultraviolet light (e.g., 254nm), medium-wave ultraviolet light, or long-wave ultraviolet light (e.g., 365nm).

[0064] In some embodiments of the present invention, the needle body can be irradiated with a 254 nm ultraviolet lamp for 5 to 60 minutes to solidify the drug-loaded hydrogel on the needle body surface. Then, it is placed in an oven and dried at 37 °C for 30 to 120 minutes. During the heating and drying process, the hydroxyl groups of the drug-loaded hydrogel and the hydroxyl groups on the needle body surface interact with each other through strong intermolecular interactions such as hydrogen bonds, thereby achieving efficient adhesion. The dried drug-loaded hydrogel has sufficient adhesion to the surface of the acupuncture needle to ensure that the gel layer will not fall off the surface of the needle body during acupuncture, so that the drug can be accurately delivered to specific acupoints to exert its effect.

[0065] In some embodiments of the present invention, in step S3, the needle body can be inverted (needle tip facing up) and inserted into the foam plate for fixation. 5 ~ 10 μL of the prepared drug-loaded hydrogel is transferred to the needle tip using a micropipette, and spread out with a scraper to evenly coat the needle body surface at the front third of the distance from the needle tip.

[0066] At this point, the drug-loaded hydrogel still has a certain degree of fluidity. Under the action of surface tension, it will gather into droplets on the surface of the needle tip. In order to ensure that the drug-loaded hydrogel can penetrate the skin during acupuncture, the foam board can be placed on its side during the application process to prevent the droplets from sliding down to the middle of the needle due to gravity. Repeat the application of the drug-loaded hydrogel 2 to 4 times. The next application can be done after the drug-loaded hydrogel applied in the previous application has dried slightly.

[0067] This embodiment also proposes the application of a drug-loaded acupuncture needle in the preparation of a medical device for treating Achilles tendonopathy, which is prepared using the drug-loaded acupuncture needle described above or the preparation method of the drug-loaded acupuncture needle described above.

[0068] The following detailed description, in conjunction with specific embodiments, further illustrates the drug-loaded acupuncture needles and their preparation method as described in this application.

[0069] Example 1: Pre-acupuncture treatment Acupuncture needles (made of 304 austenitic stainless steel, 0.25 mm in diameter, 13 mm in length) were ultrasonically cleaned in ethanol for 5 min, dried with nitrogen, then immersed in sodium hydroxide solution at pH 10 for 30 s, rinsed with deionized water, and dried with nitrogen. The needles were then inserted tip-up into a fixing plate and discharged for 60 s under the parameters of 0.40 mbar pressure and 10 mA discharge current. After removal, the needles were quickly placed in a sealed plastic bag and vacuum-sealed for storage.

[0070] Example 2: Pre-treatment for acupuncture Acupuncture needles (made of 304 austenitic stainless steel, 0.25 mm in diameter, 13 mm in length) were ultrasonically cleaned in ethanol for 5 min, dried with nitrogen, then immersed in sodium hydroxide solution with a pH of 10 for 30 s, rinsed with deionized water, and dried with nitrogen. The needles were then inserted into a fixing plate with the needle tip facing upwards and discharged for 60 s under the parameters of 0.36 mbar air pressure and 10 mA discharge current. After removal, the acupuncture needles were quickly placed in a plastic sealed bag and vacuum-sealed for storage.

[0071] Example 3: Pre-treatment for acupuncture Acupuncture needles (made of 304 austenitic stainless steel, 0.25 mm in diameter, 13 mm in length) were ultrasonically cleaned in ethanol for 5 min, dried with nitrogen, then immersed in sodium hydroxide solution with a pH of 10 for 30 s, rinsed with deionized water, and dried with nitrogen. The needles were then inserted into a fixing plate with the needle tip facing upwards and discharged for 60 s under the parameters of 0.40 mbar pressure and 20 mA discharge current. After removal, the needles were quickly placed in a plastic-sealed bag and vacuum-sealed for storage.

[0072] Example 4: Pre-acupuncture treatment Acupuncture needles (made of 304 austenitic stainless steel, 0.25 mm in diameter, 13 mm in length) were ultrasonically cleaned in ethanol for 5 min, dried with nitrogen, then immersed in sodium hydroxide solution with a pH of 10 for 30 s, rinsed with deionized water, and dried with nitrogen. The needles were then inserted into a fixing plate with the needle tip facing upwards and discharged for 20 s under the parameters of 0.40 mbar air pressure and 10 mA discharge current. After removal, the acupuncture needles were quickly placed in a plastic sealed bag and vacuum-sealed for storage.

[0073] Comparative Example 1: Pre-treatment for acupuncture The acupuncture needles (made of 304 austenitic stainless steel, 0.25 mm in diameter and 13 mm in length) were ultrasonically cleaned in ethanol for 5 minutes, dried with nitrogen, then immersed in a sodium hydroxide solution with a pH of 10 for 30 seconds, rinsed with deionized water, and dried with nitrogen. The acupuncture needles were then placed directly into a sealed plastic bag and vacuum-sealed for storage.

[0074] Example 5: Preparation of the precursor portion of a multi-hydroxyl hydrogel First, add 2.0 μL of concentrated ammonia solution (28% by mass) to 1 mL of deionized water, and then add 10 mg of Ca(OH)₂. Sonicate the mixture for 10 min. Next, weigh 60 mg of carboxymethyl chitosan (CMCS) and add it to the system. The selected CMCS has a substitution degree greater than 80%. Stir the mixture further for 30 min to dissolve and uniformly disperse the CMCS; at this point, the mixture is a milky white viscous consistency. Then, weigh 60 mg of glucono-δ-lactone (GDL) and add it to the system. Stir the mixture rapidly until it becomes a clear, transparent gel.

[0075] Example 6: Preparation of the precursor portion of a multi-hydroxyl hydrogel Add 2.0 μL of concentrated ammonia solution (28% by mass) to 1 mL of deionized water, then add 10 mg of Ca(OH)₂ and sonicate the mixture for 10 min. Next, weigh 80 mg of carboxymethyl chitosan (CMCS) and add it to the system (CMCS degree of substitution greater than 80%). Stir the mixture further for 30 min to dissolve and uniformly disperse the CMCS; at this point, the mixture is a milky white viscous consistency. Then, weigh 80 mg of glucono-δ-lactone (GDL) and add it to the system. Stir the mixture rapidly until it becomes a clear, transparent gel.

[0076] Example 7: Preparation of CMCS-GDL / SilMA hydrogel precursor First, add 2.0 μL of concentrated ammonia solution (28% by mass) to 1 mL of deionized water, and then add 10 mg of Ca(OH)₂. Sonicate the mixture for 10 min. Next, weigh 60 mg of carboxymethyl chitosan (CMCS) with a substitution degree greater than 80% and add it to the system. Stir the mixture further for 30 min to dissolve and uniformly disperse the CMCS, resulting in a milky white, viscous mixture. Then, weigh 60 mg of glucono-δ-lactone (GDL) and add it to the system. Stir the mixture rapidly until it becomes a clear, transparent gel. Finally, add 50 mg of SilMA and 5 mg of photosensitizer to the system to synthesize a hydrogel.

[0077] Example 8: Preparation of the precursor portion of a drug-loaded conductive hydrogel Take 350 μL of 1.7wt% PEDOT:PSS dispersion and dilute with deionized water to a total volume of 1 mL. Weigh 2 mg of icariin (ICA) and add it to the solution, then sonicate for 10 min to ensure uniform dispersion of the drug in the mixture. Weigh 100 mg of methacrylamide silk fibroin (SilMA), cut it into small particles, and add it to the system, stirring until completely dissolved. At this point, the system is dark blue and slightly viscous.

[0078] Example 9: Preparation of the precursor portion of a drug-loaded conductive hydrogel Take 500 μL of 1.7 wt% PEDOT:PSS dispersion and dilute with deionized water to a total volume of 1 mL. Weigh 2 mg of icariin (ICA) and add it to the solution, then sonicate for 10 min to ensure uniform dispersion of the drug in the mixture. Weigh 120 mg of methacrylamide silk fibroin (SilMA), cut it into small particles, and add it to the system, stirring until completely dissolved. At this point, the system is dark blue and slightly viscous.

[0079] Example 10: Preparation of the conductive hydrogel precursor Take 350 μL of 1.7 wt% PEDOT:PSS dispersion and dilute with deionized water to a total volume of 1 mL. Weigh 100 mg of methacrylamide silk fibroin (SilMA), cut it into small particles, and add it to the system. Stir until completely dissolved. At this point, the system is dark blue and slightly viscous.

[0080] Example 11: Preparation of drug-loaded conductive dual-network hydrogel 500 μL each of the polyhydroxy hydrogel precursor solution from Example 5 and the drug-loaded conductive hydrogel precursor solution from Example 8 were mixed uniformly. 5 mg of the photoinitiator lithium phenyl (2,4,6-trimethylbenzoyl) phosphate (LAP) was weighed and added to the mixture, which was then stirred until homogeneous. The final hydrogel composition was calculated as follows: CMCS / GDL (6 wt%), SilMA (5 wt%), PEDOT:PSS (0.3 wt%), ICA (1 mg / mL), and LAP (0.5 wt%).

[0081] Example 12: Preparation of conductive dual-network hydrogel 500 μL each of the polyhydroxy hydrogel precursor solution from Example 5 and the conductive hydrogel precursor solution from Example 10 were mixed uniformly. 5 mg of the photoinitiator lithium phenyl (2,4,6-trimethylbenzoyl) phosphate (LAP) was weighed and added to the mixture, and then stirred until homogeneous.

[0082] Example 13: Preparation of drug-loaded acupuncture needles (drug-loaded needles) The acupuncture needles of Example 1 were inverted (tip facing upwards) and fixed in a foam board. 5 μL of the hydrogel prepared in Example 11 was transferred to the needle tip using a micropipette. The hydrogel was then spread evenly on the needle surface, covering the area 4 mm from the needle tip (the depth to which it will penetrate the skin). The hydrogel was applied three times, each time 10 minutes after the previous application when the gel had dried at room temperature. After hydrogel coating, the needles were irradiated with a 254 nm UV lamp for 10 minutes to solidify the coating, and then dried in a 37 °C oven for 60 minutes. The resulting drug-loaded acupuncture needles were then sterilized under UV light on a cell manipulation table for 30 minutes, placed in a vacuum-sealed plastic bag, and stored at 4 °C.

[0083] Example 14: Preparation of hydrogel-loaded needles (drug-free) The acupuncture needles from Example 1 were inverted (tip facing upwards) and fixed in a foam board. 5 μL of the hydrogel prepared in Example 12 was transferred to the needle tip using a micropipette. The hydrogel was then spread evenly on the needle surface, covering the area 4 mm from the needle tip (the depth to which it will penetrate the skin). This process was repeated three times, each time after the previous application had dried at room temperature for 10 minutes. After hydrogel coating, the needles were irradiated with a 254 nm UV lamp for 10 minutes to solidify the gel, and then dried in a 37 °C oven for 60 minutes. The resulting drug-loaded acupuncture needles were then sterilized under UV light on a cell manipulation table for 30 minutes, placed in a vacuum-sealed plastic bag, and stored at 4 °C.

[0084] Comparative Example 2: Preparation of Drug-Loaded Acupuncture Needles The acupuncture needles were inverted (tip facing upwards) and fixed in a foam board. The hydrogel precursor solution from Example 8 was diluted by half, and 5 μL of this solution was transferred using a micropipette and applied to the needle tip. The solution was then spread evenly on the needle surface, covering the area 4 mm from the needle tip (the depth to which it will penetrate the skin). The hydrogel was applied three times, each time after the previous application had dried at room temperature for 10 min. After hydrogel coating, the needles were irradiated with a 254 nm UV lamp for 10 min to solidify the hydrogel on the needle surface, and then placed in an oven at 37 °C for 60 min. The resulting drug-loaded acupuncture needles were then sterilized under UV light on a cell manipulation table for 30 min, placed in a vacuum-sealed plastic bag, and stored at 4 °C.

[0085] Test Example 1: Characterization of Surface Changes After Plasma Treatment Preparation 1: Stainless steel sheets (304 austenitic stainless steel, square with sides of 1 cm) of the same material as acupuncture needles were ultrasonically cleaned in ethanol for 5 min, dried with nitrogen, then immersed in sodium hydroxide solution at pH 10 for 30 s, rinsed with deionized water, and dried with nitrogen. The stainless steel sheets were placed on clean filter paper and subjected to glow discharge treatment for 10 s, 20 s, 30 s, 60 s, 90 s, and 120 s at pressures of 0.26 mbar, 0.30 mbar, 0.32 mbar, 0.36 mbar, 0.40 mbar, and 0.42 mbar, and discharge currents of 10 mA, 15 mA, and 20 mA, respectively. After removal, the stainless steel sheets were quickly placed in a plastic-sealed bag, vacuum-sealed, and the contact angle changes with water and drug-loaded hydrogels were tested within 30 minutes.

[0086] Preparation 2: Stainless steel sheets (304 austenitic stainless steel, square with sides of 1 cm) of the same material as acupuncture needles were ultrasonically cleaned in ethanol for 5 min, dried with nitrogen, then immersed in sodium hydroxide solution with a pH of 10 for 30 s, rinsed with deionized water, and dried with nitrogen. The stainless steel sheets were then placed directly into a plastic sealed bag and vacuum-sealed for storage.

[0087] Test: A 10 μL water droplet or a 10 μL drug-loaded conductive double-network hydrogel droplet prepared in Example 11 was placed on the surface of a stainless steel sheet treated under each of the preparation schemes (corresponding to Examples 1 to 4 and other treatment parameters, respectively) or the surface of a stainless steel sheet treated under the preparation scheme 2 (corresponding to Example 1). The contact angle was measured using a contact angle meter and recorded using ImageJ software. Some results are shown in Table 1 below.

[0088] Table 1 Contact Angle Test Results

[0089] As shown in Table 1, the stainless steel sheet (acupuncture needle) of the present invention, after glow discharge treatment, has a significantly improved contact angle compared to stainless steel sheet that has only undergone alkaline washing. This makes the needle body more hydrophilic, which is beneficial for the spread of gel at the needle tip during the preparation process. It also greatly optimizes the adhesion of the drug-loaded hydrogel to the surface of the acupuncture needle.

[0090] Test Example 2: Characterization of Hydrogel Preparation The hydrogels from Examples 5 (A: CMCS+GDL), 7 (B: CMCS+GDL+SilMA), 12 (C: CMCS+GDL+SilMA+PEDOT:PSS), and 11 (D: CMCS+GDL+SilMA+PEDOT:PSS+ICA) were placed in 24-well plates with a diameter of 16 mm and irradiated with a 254 nm UV lamp for 10 min. The hydrogel discs were then removed and freeze-dried in liquid nitrogen. The morphological changes of the synthesized hydrogels at each step were observed using a scanning electron microscope. The results are as follows: Figure 3 As shown. The results, obtained using changes in the infrared absorption spectrum, are as follows. Figure 4 As shown, this demonstrates the crosslinking changes of the hydrogel and the drug loading during each step.

[0091] like Figure 3 As shown, comparison Figure 3 D and Figure 3 A, Figure 3 B and Figure 3 C. The changes in the micromorphology of the hydrogel demonstrate that the drug-loaded conductive dual-network hydrogel of this invention achieves comprehensive improvements in mechanical properties, conductivity, and drug delivery capabilities. When silk fibroin and PEDOT:PSS are added, the pore structure of the hydrogel becomes more complex, the pore walls thicken, and the surface roughness increases, which is beneficial for improving drug loading and the mechanical strength of the hydrogel. Figure 4 As shown, the changes in infrared spectral peaks demonstrate the successful synthesis of the composite hydrogel and the loading of the drug according to this invention.

[0092] Test Example 3: Characterization of Drug-Loaded Acupuncture Needles The morphological changes of drug-loaded acupuncture needles at different preparation stages, including empty needles (Example 1), glue-loaded needles (Example 14), and drug-loaded needles (Example 13), were characterized using SEM. Elemental analysis was performed on the surfaces of acupuncture needles under different modification conditions, including empty needles, glue-loaded needles, and drug-loaded needles. Specific results are as follows: Figure 5 As shown. By Figure 5 It can be seen that a clear hydrogel coating structure appeared on the surface of the acupuncture needle. Combined with the change process of the elements on the surface of the acupuncture needle, it proves the uniform coating of hydrogel and the successful loading of drugs.

[0093] Test Example 4: Swelling Performance Test of Drug-Loaded Conductive Dual-Network Hydrogel The hydrogel precursor prepared in Example 11 was placed in a 24-well plate with a diameter of 16 mm and irradiated with a 254 nm UV lamp for 10 min. The hydrogel discs were then removed and dried in a 37 °C oven until constant weight. The hydrogels were then immersed in phosphate buffer and subjected to swelling performance testing at 37 °C. After 5 min, 10 min, 15 min, 20 min, 30 min, 45 min, 60 min, 120 min, and 480 min, the hydrogels were removed, free liquid was removed with filter paper, and the weight was calculated to determine the swelling ratio.

[0094] like Figure 6 As shown, the drug-loaded hydrogel material of the present invention exhibits excellent swelling properties, can quickly reach swelling equilibrium, and has a high swelling rate and good stability, which is beneficial to the full dissolution and release of the drug.

[0095] Test Example 5: Drug Release Performance Test of Drug-Loaded Conductive Dual-Network Hydrogel Spontaneous Release Characteristics: The hydrogel precursor prepared in Example 11 was placed in a 48-well plate with a diameter of 10.2 mm and irradiated with a 254 nm UV lamp for 10 min. The hydrogel discs were then removed and dried in a 37 ℃ oven until constant weight. The hydrogel discs were placed in centrifuge tubes containing 3 mL of pH 7.4 PBS and placed in a shaker at 37 ℃. 0.5 mL of the solution was collected and centrifuged at 10 min, 20 min, 30 min, 60 min, 24 h, and 48 h, respectively, while simultaneously adding the same volume of PBS to the original centrifuge tube. Five gel discs were used as parallel samples in each test. 200 μL of the test sample was added to each well of a 96-well plate, and the absorbance was measured at 378 nm. The concentration of the drug in the sample was calculated by referring to the standard curve, and the spontaneously released drug amount and release rate were calculated.

[0096] Electrodischarge characteristics: The hydrogel precursor prepared in Example 11 was placed in a 48-well plate with a diameter of 10.2 mm and irradiated with a 254 nm UV lamp for 10 min. The hydrogel discs were then removed and dried in an oven at 37 ℃ until constant weight. The hydrogel discs were fixed onto a copper conductive adhesive as the working electrode, a silver-silver chloride electrode as the reference electrode, and a platinum electrode as the counter electrode. 3 mL of pH 7.4 PBS was added to the electrolytic cell as the electrolyte. Constant voltages of 0.6 V, 0.75 V, 0.85 V, 1 V, and 1.2 V were applied. At 15 min, 30 min, 45 min, and 60 min, 1 mL of electrolyte was aspirated as a sample, and the same volume of PBS was added to maintain a constant electrolyte volume. Five gel discs were used as parallel samples in each test. Add 200 μL of the sample to each well of a 96-well plate, measure the absorbance at 378 nm, calculate the drug concentration in the sample by referring to the standard curve, and calculate the amount and rate of drug released by electrical stimulation.

[0097] In one test example of this invention, in the same medium, such as Figure 7 As shown, the electrostatic release of the drug is significantly stronger than its spontaneous release. In one test example of this invention, in the same medium, such as... Figure 8 As shown, the intensity of drug release upon electrophoresis is positively correlated with the applied voltage. In the various test examples of this invention, the voltage range used was 0.6 ~ 1.2 V, and the current applied within this voltage range was 0.3 ~ 0.8 mA. At 60 minutes, the release rates of each group ranged from 42% to 94%.

[0098] Test Example 6: Bonding Strength Test between Drug-Loaded Conductive Dual-Network Hydrogel and Needle Surface The adhesion between hydrogels with different proportions and treated stainless steel materials was evaluated using a 180° peel test. 1 mL of each of the hydrogel precursors from Examples 12 (CGC / S / P), 5 (CGC), and 10 (SilMA / P) were evenly spread into 6-well plates with a diameter of 34.8 mm. After irradiation with a 254 nm UV lamp for 10 min, the hydrogel discs were gently removed and cut into 2 × 0.5 cm rectangles. These rectangular slices were divided into two groups, and the two groups were adhered to glow discharge-treated and conventionally treated stainless steel plates, respectively. Transparent tape was adhered to the back of the hydrogel strips as an additional protective layer to prevent excessive stretching during the test. Shear adhesion tests were performed using a universal testing machine at a tensile rate of 50 mm / min. When the peeling process reached a steady state, no further increase in the measured peel force was observed, indicating a plateau. Interfacial toughness (J / m) was measured. 2The force Fpeel at the platform is calculated by dividing the width of the hydrogel sample being tested.

[0099] like Figure 9 As shown, S and SilMA represent methacrylamide silk fibroin, P represents PEDOT:PSS, CGC represents CMCS-GDL hydrogel, and Glow represents a stainless steel sheet treated with glow discharge. The results of this invention show that by adjusting the hydrophilicity / hydrophobicity of the needle material surface and the enhanced hydrogen bonding of the conductive dual-network hydrogel through glow discharge, the adhesion between the hydrogel and the metal surface is significantly improved, enabling it to adhere firmly to the needle tip and effectively preventing the coating from falling off during acupuncture. Furthermore, the adhesion of the conductive dual-network hydrogel is stronger than that of a single-component hydrogel.

[0100] Test Example 7: Biosafety ① Toxicity of drug-loaded needle extract on primary mouse tendon cells Prepare cell culture medium (containing 89% DMEM high glucose medium, 10% fetal bovine serum, and 1% penicillin-streptomycin solution). Incubate 10 drug-loaded needles and 10 hydrogel-loaded needles (without drug loading) in 5 mL of cell culture medium at 37 °C for 24 h. The resulting extract is the 100% extract. Dilute the 100% extract with cell culture medium to prepare 70%, 50%, 25%, and 5% extracts. Weigh phenol crystals.

[0101] Primary mouse tendon cells were cultured to 80% confluence, digested, and passaged into three 96-well plates (0.1 mL per well). Cells were cultured in a cell culture incubator until adherence. The three 96-well plates were divided into three time groups: 12 h, 24 h, and 48 h. Each time group was further divided into a blank control group and five experimental groups (100% extract, 70% extract, 50% extract, 25% extract, and 5% extract), with six wells in each group.

[0102] The original culture medium of primary mouse tendon cells was discarded. 100 μL / well of each of the experimental groups (100%, 70%, 50%, 25%, and 5%) extracts was added, while 100 μL / well of cell culture medium was added to the blank control group. Cells were cultured in a cell culture incubator for 12 h, 24 h, and 48 h, respectively. After culture, 10 μL of CCK-8 reagent was added to each well, and the cells were incubated at 37 ℃ for 1 h. The absorbance was measured at 450 nm using a microplate reader to compare cell growth in each group. Relative cell viability was calculated using the formula: Relative cell viability = (OD value of experimental group - OD value of blank well) / (OD value of blank control group - OD value of blank well) × 100%.

[0103] like Figure 10As shown, the drug-loaded needle of the present invention exhibits significantly higher relative survival rate of mouse primary tendon cells at various concentrations of extract and at different time points, which can effectively reduce the cytotoxicity of the material and significantly improve its biosafety.

[0104] ② Effects of drug-loaded needle extract on the survival and death of primary mouse tendon cells Primary mouse tendon cells were cultured to 80% confluence, digested, and passaged into three 6-well plates (2 mL per well). Cells were cultured in a cell culture incubator until adherence. Three 12-well plates were divided into drug-loaded needle and hydrogel-loaded needle groups (no drug loading). The original culture medium of the primary mouse tendon cells was discarded, and 2 mL / well of 100% extract was added to each experimental group. Cells were cultured in a cell culture incubator for 24 h. The culture medium was aspirated, the cells were washed once with PBS, and 500 μL of live / dead cell (staining) working solution was added. The cells were incubated at 37 ℃ in the dark for 10 min. The staining solution was removed, the cells were washed once with PBS, and the culture medium was replaced with complete medium. High-content imaging was performed to compare the cell viability and mortality of each group.

[0105] like Figure 11 As shown, the mouse primary tendon cells treated with the drug-loaded needle extract of the present invention have higher survival rate and more intact morphology, which significantly improves the biocompatibility and better biosafety of the material.

[0106] Test Example 8: Hemolytic Effect of Drug-Loaded Target on Rabbit Erythrocytes Six 1.5 mL centrifuge tubes were added to seven groups of 6 rabbit erythrocyte suspensions. Five groups were experimental groups, and the other two were positive control and blank control groups, respectively. One, two, or ten drug-loaded needles and one hydrogel-loaded needle (without drug loading) were added to each of the five experimental groups. The positive control group received 0.3 mL of 10% Triton solution, and the blank control group received 0.3 mL of PBS solution. All centrifuge tubes were heated in a 37 ℃ water bath for 1 h, centrifuged at 3000 r / min for 5 min, and 100 μL of the supernatant from each group was added to a 96-well plate. The absorbance was measured at 540 nm. The hemolysis rate was calculated using the formula: Hemolysis rate = [(OD value of experimental group - OD value of blank control group) / (OD value of positive control group - OD value of blank control group)] × 100%.

[0107] like Figure 12 As shown, the drug-loaded needle of the present invention exhibits significantly lower hemolysis rates at all test concentrations, effectively improving the blood compatibility of the material and significantly reducing the risk of hemolysis when in contact with blood.

[0108] Test Example 9: Behavioral Test ① Construction of Achilles tendinitis model: Wild-type male C57 / BL mice aged 6-8 weeks were randomly divided into 5 groups: blank control group (NC), Achilles tendinitis group (TEN), acupuncture treatment group (EA), ICA treatment group (ICA: direct injection of ICA drug solution), and drug-loaded acupuncture combined with treatment group (EA+ICA), with 10 mice in each group. After 7 days of acclimatization feeding, 20 μL (concentration of 15 mg / mL) of type I collagenase was injected into each of the two Achilles tendons of the Achilles tendinitis group mice to prepare a collagenase-induced Achilles tendon micro-injury model.

[0109] ②von Frey: A von Frey filament was applied to the sole of the animal's hind paw with increasing force. Rapid withdrawal of the hind paw was recorded as a positive response. Ten trials were conducted at a given force, with a 30-second interval between trials, and the number of positive responses for each von Frey filament stimulus was recorded.

[0110] like Figure 13 As shown in the (mechanical pain threshold) diagram, the drug-loaded needle combined treatment group of the present invention significantly relieved the pain in mice, and the effect was better than that of single acupuncture or ICA treatment group, which fully demonstrates the synergistic effect of the combined treatment strategy adopted in the present invention in relieving tendinopathy pain.

[0111] ③ Hot Plate Test: The thermal pain threshold in mice was determined using an IITC (IITC Life Science Inc.) hot and cold plate test. Mice were placed in a transparent acrylic test chamber and allowed approximately 15 minutes to acclimatize. The initial temperature of the hot plate was set to 30 °C, with a heating rate of 10 °C per minute. The maximum temperature was set to 55 °C to prevent irreversible tissue damage to the mice. A positive response was indicated by the mouse lifting or licking its paw. The temperature from the start of the test to the moment the mouse lifted or licked its paw was recorded. The test was repeated three times, with a 5-minute interval between each stimulation. The average of the three measurements was calculated as the thermal pain threshold.

[0112] like Figure 13 As shown in the (thermal pain threshold) diagram, the drug-loaded needle combined treatment group of the present invention can effectively relieve pain hypersensitivity induced by thermal stimulation, highlighting the synergistic therapeutic advantage of the present invention in comprehensively improving multiple pain modalities of tendinopathy.

[0113] ④ Rotador test: Before the test, allow the animals to acclimatize to the rotador for 5-10 minutes. Set the acceleration of the rotador to 20 rpm / min, and the test time to 5-10 minutes. Record the time the mice spend on the rotador. By comparing the time spent on the rotador in different groups, assess their motor coordination and balance.

[0114] like Figure 13As shown in (rotary rod fatigue), the drug-loaded needle combined treatment group of the present invention can effectively improve its motor coordination and balance function, further verifying the synergistic effect of the combined treatment strategy of the present invention in promoting the functional recovery of tendinopathy.

[0115] ⑤ Lower limb grip strength test: Calibrate and zero-point adjust the mouse grip strength meter, remove the mouse from the cage, lower the mouse above the grid, keep its torso parallel to the grid, and allow the mouse's forepaws and hind paws to grip the grid. Gently pull the mouse with its tail to make it grip the probe forcefully, and record the maximum grip strength value displayed on the screen. To obtain accurate data, this procedure needs to be repeated three times to obtain three measurements of hind limb grip strength.

[0116] like Figure 13 As shown in (lower limb grip strength), the drug-loaded needle combined treatment group of the present invention can effectively improve muscle function and motor ability, proving the synergistic therapeutic advantage of the combined treatment of the present invention in improving tendonopathy-related motor dysfunction.

[0117] Test Example 10: Pathological Staining The Achilles tendon tissue samples taken at the final time point were fixed, dehydrated, paraffin-impregnated, embedded, and sectioned in paraffin. H&E staining was used to assess the degree of Achilles tendon tissue damage and degeneration. Masson staining and Sirius red staining were used to assess the degree of tissue damage and fibrosis by analyzing the distribution and proportion of type I and type III collagen fibers in the Achilles tendon tissue.

[0118] like Figure 14 As shown, the drug-loaded needle combined treatment group of the present invention can effectively promote tendon repair and inhibit pathological fibrosis, highlighting the synergistic therapeutic advantages of the present invention in improving the pathological remodeling of tendon tissue.

[0119] The above description is merely a preferred embodiment of the present invention, but the scope of protection of the present invention is not limited thereto. Any variations or substitutions that can be easily conceived by those skilled in the art within the technical scope disclosed in the present invention should be included within the scope of protection of the present invention. Therefore, the scope of protection of the present invention should be determined by the scope of the claims.

Claims

1. A drug-loaded acupuncture needle, characterized in that, include: The needle body, the surface of which is treated with glow discharge; A drug-loaded hydrogel is uniformly coated on the surface near the tip of the needle and is cured and adhered to that portion of the needle surface. The drug-loaded hydrogel comprises a uniformly mixed dual-network hydrogel, a conductive polymer, and drug molecules.

2. The drug-loaded acupuncture needle according to claim 1, characterized in that, The dual-network hydrogel is constructed using carboxymethyl chitosan-gluconolactone and methacrylamide silk fibroin.

3. The drug-loaded acupuncture needle according to claim 1, characterized in that, The conductive polymer is composed of PEDOT:PSS.

4. The drug-loaded acupuncture needle according to claim 1, characterized in that, The drug molecule is icariin.

5. A method for preparing a drug-loaded acupuncture needle according to any one of claims 1 to 4, characterized in that, Includes the following steps: Prepare the needle body and perform glow discharge treatment on the needle body; A drug-loaded hydrogel is obtained by uniformly mixing a dual-network hydrogel, a conductive polymer, and drug molecules. The drug-loaded hydrogel is uniformly coated onto the surface of the needle body treated with glow discharge and then fixed in place to obtain the drug-loaded acupuncture needle.

6. The preparation method according to claim 5, characterized in that, Before performing glow discharge treatment on the needle body, the needle body is subjected to alkaline washing.

7. The preparation method according to claim 5, characterized in that, The parameters for the glow discharge treatment are: gas pressure of 0.36 mbar to 0.42 mbar, discharge current of 10 mA to 20 mA, and time of 20 s to 90 s.

8. The preparation method according to claim 5, characterized in that, The preparation of the drug-loaded hydrogel includes the following steps: Carboxymethyl chitosan was dissolved in ammonia and calcium hydroxide solution and stirred to obtain a carboxymethyl chitosan solution. Glucono-δ-lactone is mixed with the carboxymethyl chitosan solution and stirred until the system becomes a transparent gel to obtain the first gel solution. Icariin was added to a dispersion of poly(3,4-ethylenedioxythiophene):polystyrene sulfonate and ultrasonically dispersed to obtain a mixed solution. Add methacrylamide silk fibroin to the mixed solution and stir until completely dissolved to obtain a second gel solution; The first gel solution and the second gel solution are mixed, a photoinitiator is added, and the mixture is stirred until homogeneous to obtain the drug-loaded hydrogel.

9. The preparation method according to claim 8, characterized in that, The fixation and bonding of the needle body after coating it with drug-loaded hydrogel includes the following steps: The needle is irradiated with ultraviolet light, and after the surface of the needle has solidified, the needle is dried.

10. The use of the drug-loaded acupuncture needle prepared by the method of any one of claims 1 to 4 or any one of claims 5 to 9 in the preparation of a medical device for treating Achilles tendonopathy.

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