Body temperature response type double-sided patch as well as preparation method and application thereof

Abstract

The invention belongs to the field of biomedical materials, and particularly relates to a body temperature response type double-sided patch and a preparation method and application thereof, and the body temperature response type double-sided patch is composed of a shape memory polymer and a zinc ion slow-release bonding layer; the shape memory polymer is prepared by crosslinking diisopropenyl benzene, lipoic acid, cellulose acetate and iron ions. The zinc ion slow-release adhesive layer is obtained by dissolving zinc ions in a lipoic acid solution, wound repair can be adaptively promoted at the body temperature, and compared with the prior art, the healing time is shortened, complications are reduced, the wound healing quality is improved, and regenerative repair is achieved.

Description

Technical Field

[0001] This invention belongs to the field of biomedical materials, specifically relating to a body temperature responsive double-sided patch, its preparation method, and its application. Background Technology

[0002] Diabetic chronic wound healing faces three major challenges: lack of mechanical tension at the wound margin, persistent high oxidative stress, and susceptibility to secondary infection, ultimately leading to delayed healing and difficulty in tissue regeneration. Mechanical tension at the wound margin is the initiating factor for wound closure, but it is significantly weakened in diabetic wounds, hindering cell migration and proliferation, and is a major factor in the prolonged healing of diabetic wounds. Simultaneously, the persistent high oxidative stress and inflammatory response in the diabetic wound microenvironment exacerbate cellular dysfunction, reduce collagen synthesis and angiogenesis, promote infection, and further delay wound healing. Managing diabetic chronic wounds through convenient methods such as patches and dressings is crucial. However, existing dressings only provide passive barrier protection or perform only a single function such as anti-infection. They neglect the mechanical microenvironment required for wound contraction and cannot provide multiple functions such as regulating oxidative stress, inhibiting inflammation and infection, making it difficult to meet the complex needs of dynamic repair in diabetic wounds and unable to achieve functional regeneration with skin appendages. Therefore, there is an urgent need to develop a novel wound patch that can simultaneously provide mechanical tension, regulate oxidative stress, and inhibit infection, in order to effectively promote rapid healing and functional tissue regeneration of diabetic wounds.

[0003] Existing technologies such as traditional gauze and hydrogel dressings primarily provide a passive protective barrier for chronic diabetic wounds. In recent years, technologies have been developed to improve hydrogel dressings based on the characteristics of the wound's biochemical environment. These dressings inhibit wound infection and promote wound healing by carrying antibacterial drugs and exosomes. However, this improved method still has the following drawbacks: 1) it lacks effective stimulation against mechanical tension and cell proliferation; 2) it is not suitable for the moist wound environment.

[0004] In summary, existing technologies cannot fully repair wounds due to a lack of mechanical tension and incompatibility with moist wound environments. Summary of the Invention

[0005] To address the aforementioned technical problems, this invention provides a body temperature-responsive double-sided patch, its preparation method, and its application.

[0006] A body temperature responsive double-sided patch is composed of a shape memory polymer and a zinc ion slow-release adhesive layer; The shape memory polymer is prepared by crosslinking diisopropylbenzene, lipoic acid, cellulose acetate and iron ions; the mass ratio of diisopropylbenzene, lipoic acid, cellulose acetate and iron-containing compounds is 0.5~0.8:2~4:0.15~0.75:0.05~0.15; The zinc ion slow-release adhesive layer is obtained by dissolving zinc ions in a thioctic acid solution.

[0007] In the body temperature responsive double-sided patch of the present invention, the shape memory polymer triggers shape recovery at body temperature, providing active mechanical contraction force to promote the tightening and healing of wound edges. The zinc ion slow-release adhesive layer provides strong wetting adhesion in the wound environment and continuously releases zinc ions to promote antioxidant stress and angiogenesis during the wound healing process, improve the quality of wound healing, and achieve regenerative repair.

[0008] Preferably, the preparation steps of the shape memory polymer are as follows: diisopropylbenzene and lipoic acid are used as fixed components, and dissolved together with cellulose acetate and iron-containing compounds in N,N-dimethylformamide to obtain a mixed solution. The mixed solution is then heated and cured to obtain the shape memory polymer.

[0009] Preferably, the heating and curing step is as follows: the mixed solution is heated sequentially at 70℃~90℃ for 3h~6h, at 90℃~110℃ for 1h~2h, and at 130℃~150℃ for 3h~5h.

[0010] Preferably, the preparation steps of the zinc ion slow-release adhesive layer are as follows: add zinc nitrate hexahydrate to thioctic acid solution, dissolve and pour into a mold, dry to obtain the zinc ion slow-release adhesive layer; The mass ratio of lipoic acid to zinc nitrate hexahydrate is 1:0.01~0.5.

[0011] The method for preparing the body temperature responsive double-sided patch involves pouring a solution of the zinc ion slow-release adhesive layer onto the surface of the shape memory polymer and drying it to obtain the body temperature responsive double-sided patch.

[0012] The application of the aforementioned temperature-responsive double-sided patch in the preparation of drugs for chronic wound healing in diabetes.

[0013] Preferably, the drug is used to reduce the wound area.

[0014] Preferably, the drug is used to improve the speed of wound healing.

[0015] Preferably, the drug is used to promote tissue and functional regeneration in the wound.

[0016] Preferably, the tissue and functional regeneration includes hair follicle regeneration, blood vessel regeneration, and sebaceous gland regeneration.

[0017] Compared with the prior art, the beneficial effects of the present invention are as follows: Promotes wound contraction: Through the thermal response of shape memory polymers, mechanical contraction force is automatically generated, significantly accelerating the initial closure of wounds.

[0018] Antibacterial effect: The release of zinc ions has a strong antibacterial effect and effectively inhibits wound infection.

[0019] Promotes regeneration: Through the synergistic effect of mechanical contraction and Zn²⁺ release, it promotes tissue repair and functional regeneration of wounds, including the regeneration of hair follicles and sebaceous glands.

[0020] Advantages: By integrating mechanical and biochemical treatment strategies, this invention can adaptively promote wound repair at body temperature, shorten healing time, reduce complications, improve wound healing quality, and achieve regenerative repair compared to existing technologies. Attached Figure Description

[0021] Figure 1 The image shows the preparation and characterization of the shape memory polymer LCAFe. In the image, A is a schematic diagram of LCAFe preparation, B is an infrared spectrum, C is a partial infrared spectrum, D is a stress-strain curve, E is the shape fixation rate and shape recovery, F shows self-healing, G is the stress-strain curve of the healed LCAFe network, and H is...

[0022] Figure 2 is the glass transition temperature of LCAFe.

[0023] Figure 3 The structure and characterization of the LA-Zn adhesive membrane are shown in Figure 1. A is a schematic diagram of the reaction of the LA-Zn adhesive membrane, B is a scanning electron microscope and energy dispersive X-ray diffraction analysis, (i) in B is a cross-sectional view, (ii) is a longitudinal cross-sectional scanning electron microscope view of the SMART patch, (iii) is a cross-sectional scanning electron microscope view of the LA-Zn adhesive membrane, C is an infrared spectrum, D is a release curve, E is the adhesion performance, and F is the quantitative evaluation results of the bioadhesion performance, (i) in F is a schematic diagram, (ii) is the shear force of the adhesive membrane, and (iii) is the wound closure strength.

[0024] Figure 4 Characterization of SMART patches, where A represents shape memory performance, B represents antibacterial effect, and C represents biocompatibility. In C, (i) represents live / dead cell staining results, and (ii) represents CCK-8 assay results.

[0025] Figure 5The SMART patch was used to promote rapid wound healing, hair follicle and sebaceous gland regeneration in diabetic mice. In the figure, A is a schematic diagram of the construction of the diabetic mouse wound model, B is the wound area, C is the wound area reduction rate, D is the wound closure time, E is the staining result, and F is the tissue remodeling. In F, (i) is collagen regeneration, (ii) is hair follicle regeneration, and (iii) is sebaceous gland regeneration.

[0026] Figure 6 To analyze the molecular mechanism of SMART patch regulation of diabetic wounds through transcriptome sequencing, A is a volcano plot, B is the LA-CA@Fe SMP GO enrichment analysis results, C is a heatmap, D is the SMART patch GO enrichment analysis, and E is the heatmap analysis verification of related differentially expressed genes.

[0027] Figure 7 For KEGG pathway analysis, A is LA-CA@Fe SMP and B is SMART patch.

[0028] Figure 8 To verify the effects of SMART patches on hair follicle regeneration, angiogenesis, macrophage immune regulation, and reactive oxygen species scavenging in vivo, A represents hair follicle regeneration, B represents pro-angiogenic characteristics, C represents immune regulation effect, D represents reactive oxygen species level, E represents [missing information], F represents statistical results of B, G represents statistical results of CD86, H represents statistical results of CD206, I represents statistical results of D, and J represents a schematic diagram of the effect. Detailed Implementation

[0029] The specific embodiments of the present invention are described in detail below, but it should be understood that the scope of protection of the present invention is not limited to the specific embodiments. All other embodiments obtained by those skilled in the art based on the embodiments of the present invention without inventive effort are within the scope of protection of the present invention. Unless otherwise specified, the experimental methods described in the embodiments of the present invention are conventional methods.

[0030] This project innovatively proposes a smart SMART patch, which solves three key problems of wound mechanical tension supply, reactive oxygen species clearance, and infection control by integrating a shape memory polymer layer with thermo-responsive mechanical contraction function and a zinc ion sustained-release adhesive layer that provides biochemical regulation. This smart patch provides a novel treatment strategy for diabetic wounds with active temporal regulation by simulating the mechanical and biochemical timing process of natural wound healing. (1) Can a smart Janus structure integrating a thermo-responsive shape memory polymer upper layer and a zinc ion sustained-release adhesive lower layer be successfully constructed? (2) Can the smart SMART patch material provide the force required for tissue closure, reverse the high oxidative stress environment, resist infection, accelerate the closure of chronic wounds, and achieve regenerative repair? (3) Does the regenerative repair of this diabetic chronic wound rely on the mechanical-biochemical synergistic mechanism of the smart SMART patch?

[0031] This invention provides a SMART patch comprising two layers of material: Shape memory polymer: This layer of material (LA-CA@Fe SMP) triggers shape recovery at body temperature, providing active mechanical contraction force to promote the tightening and healing of wound edges.

[0032] Zinc ion sustained-release adhesive layer: This layer (LA-Zn adhesive membrane) provides strong wetting adhesion in the wound environment and continuously releases zinc ions (Zn²⁺) to promote antioxidant stress and angiogenesis during the wound healing process.

[0033] Example 1 Preparation of shape memory polymers: Shape memory polymers are prepared by crosslinking lipoic acid (LA) and cellulose acetate (CA) with iron ions (Fe³⁺), and are denoted as LCAFe.

[0034] First, diisopropylbenzene (DIB, 0.6 g, D806978, Macklin, Shanghai, China) and lipoic acid (LA, 3.0 g, A835604, Macklin, Shanghai, China) were used as fixed components and dissolved in 15 mL of N,N-dimethylformamide (DMF, D757658, Macklin, Shanghai, China) along with varying amounts of cellulose acetate (CA, C804765, Macklin, Shanghai, China) and ferric chloride hexahydrate (FeCl3·6H2O, F2877, Sigma-Aldrich, MA, USA). The mixture was stirred at 80 °C for 40–60 minutes until completely dissolved. Subsequently, the solution was poured into a polytetrafluoroethylene mold and subjected to a programmed curing process: 80 °C (4 h), 100 °C (2 h), and 140 °C (4 h). After heat treatment, the system is allowed to cool naturally to room temperature to obtain the final LCAFe film, which is a flexible cross-linked network.

[0035] Physicochemical property characterization of shape memory polymers: This invention systematically prepared multiple groups of samples by varying the amounts of added CA and FeCl3·6H2O, aiming to investigate the independent and synergistic effects of CA content and Fe³⁺ crosslinking degree on material properties. The naming convention "LCAx Fey" was adopted, where x represents 20 / 3 times (g) of the added CA mass and y represents 20 / 3 times (g) of the added FeCl3·6H2O mass.

[0036] CA variable series (FeCl3·6H2O kept constant at 0.45g): LCA0Fe3 (CA 0g), LCA1Fe3 (CA 0.15g), LCA2Fe3 (CA 0.3g), LCA3Fe3 (CA 0.45g), LCA4Fe3 (CA 0.6g), LCA5Fe3 (CA 0.75g).

[0037] Fe variable series (CA remains constant at 0.6g): LCA4Fe0 (FeCl3·6H2O 0g), LCA4Fe1 (FeCl3·6H2O 0.05g), LCA4Fe2 (FeCl3·6H2O 0.10g), LCA4Fe3 (FeCl3·6H2O 0.15g).

[0038] Control group: LCA1Fe0 (CA 0.15g, FeCl3·6H2O 0g).

[0039] See the schematic diagram for the synthesis. Figure 1 A 1. Fourier transform infrared spectroscopy Different groups of LCAFe, LA-Zn adhesives, LA polymers, and LA powders were measured using an attenuated total reflectance-Fourier transform infrared spectrometer (ATR-FTIR; FTIR-8400S, Shimadzu, Tokyo, Japan). Infrared spectra were acquired in the range of 4000–800 cm⁻¹, scanned 20 times, and had a resolution of 4 cm⁻¹.

[0040] The results are as follows Figure 1 B and 1C show that at 1589 cm⁻¹, Fe³⁺ in the polymer network forms a coordination bond with the carboxylate group. As the cellulose acetate content increases, the -C=O peak slightly redshifts from 1703 cm⁻¹ to 1700 cm⁻¹, indicating the formation of hydrogen bonds.

[0041] 2. Mechanical properties Stress-strain curves were obtained using an electronic universal testing machine (EZ-SX, Shimadzu, Japan). The test sample size was 20 mm × 5 mm × 0.6 mm, the gauge length was 10 mm, and the crosshead speed was 5 mm / min.

[0042] The results are as follows Figure 1 D shows that the mechanical properties of the LCAFe copolymer gradually increase with the increase of cellulose acetate content and Fe³⁺ content, indicating the high bonding strength of the coordination bonds formed between Fe³⁺ and carboxylate groups.

[0043] 3. Shape memory performance To investigate heat-induced shape memory behavior, a rectangular polymer film with dimensions of 20 mm × 5 mm × 0.6 mm was heated to 60 °C and held, then fully stretched as a temporary shape. Subsequently, the temporary shape of the film was fixed at 0 °C for 10 min, and then exposed to room temperature to obtain the shape retention rate (Rf). Shape recovery (Rr) of the deformed sample was triggered by heating. Rf and Rr were calculated using the following formula:

[0044] R f = R r = The results are as follows Figure 1 As shown in E, the LCAFe copolymer exhibits a high shape retention rate, effectively locking in temporary deformed shapes after cooling and unloading. Subsequently, upon reheating to 120°C, the LCAFe copolymer demonstrates excellent shape recovery properties, with a recovery rate approaching its original permanent shape.

[0045] 4. Self-healing performance By connecting the newly cut interfaces, the two severed rectangular LCAFe copolymers undergo self-healing at 120°C, forming a stretchable healing network. Figure 1 F). Furthermore, this self-healing network, after being stretched and fixed at room temperature, was able to recover its pre-stretched shape at 60°C, indicating that the self-healing process effectively restored structural integrity and shape memory function. The stress-strain curve of the healed LCAFe network recovered approximately 80% of its original elastic modulus within 0.5 hours and overlapped with the original curve after 1 hour of self-healing. Figure 1 G).

[0046] 5. Glass transition temperature The glass transition temperature (Tg) of the LCAFe samples was characterized using differential scanning calorimetry (DSC 204, NETZSCH, Bavaria, Germany). Under a nitrogen atmosphere, the samples were first heated from room temperature to 120 °C at a constant rate of 10 °C / min and held for 5 min, then cooled to -20 °C at a constant rate of 10 °C / min and held for 2 min, and finally heated to 200 °C at a constant rate of 10 °C / min.

[0047] The results are as follows Figure 2 LCA1Fe3 has a glass transition temperature (Tg) of 37°C, close to human body temperature (37°C). This key property enables body temperature-triggered shape recovery, making it ideal for triggering wound contraction. Therefore, LCA1Fe3 was chosen as the shape memory layer for subsequent use and named LA-CA@Fe SMP.

[0048] Example 2 Synthesis of Zinc Ion Slow-Release Adhesive Layer (LA-Zn Adhesive Film) The zinc ion slow-release adhesive layer was prepared by dissolving 1 g of LA in 3 mL of anhydrous ethanol under gentle stirring. Subsequently, three different mass-volume percentages (1%wt, 5%wt, and 10%wt) of zinc nitrate hexahydrate (Zn(NO3)2·6H2O, Z118841, Aladdin, Shanghai, China) were added to the solution to obtain a mixture. The mixture was sonicated for 10 minutes to ensure complete dissolution, resulting in solution A. Solution A was poured into a polytetrafluoroethylene mold and dried at room temperature to obtain the LA-Zn adhesive film.

[0049] Characterization of zinc ion slow-release adhesive layer 1. Scanning electron microscopy and energy-dispersive X-ray analysis The longitudinal section and upper and lower surfaces of the SMART patch were characterized using a scanning electron microscope (SEM, S-4800, Hitachi, Tokyo, Japan) at 5 kV. The samples underwent gold / palladium sputtering coating prior to inspection. Elemental analysis was performed by energy-dispersive X-ray spectroscopy.

[0050] The longitudinal section shows a clear layered structure. Figure 3 B(ii)). Examination of the two surfaces showed that the LA-CA@Fe SMP layer had a uniform porous morphology, while the LA-Zn adhesive film layer appeared dense and non-porous. Figure 3 B(ii)). Furthermore, EDAX results indicate that the LA-Zn adhesive film has a higher zinc content in its elemental composition compared to the LA-CA@Fe SMP layer ( Figure 3 B(iii)).

[0051] 2. Fourier transform infrared spectroscopy The detection method is the same as before, and the results are as follows: Figure 3 C. In the LA-Zn adhesive film, the -OH peak at 1251 cm⁻¹ associated with the carboxyl group disappears, and the -C=O peak shifts from 1696 cm⁻¹ to 1703 cm⁻¹, indicating that a coordinate bond is formed between Zn²⁺ and the carboxyl group.

[0052] 3. Inductively Coupled Plasma Mass Spectrometry To investigate the Zn²⁺ release behavior, each sample group was immersed in PBS (phosphate-buffered saline) and incubated at 37°C for 4 weeks. Subsequently, the PBS solution was collected weekly and replaced with fresh PBS for each group. The Zn²⁺ release rate was measured using inductively coupled plasma mass spectrometry (ICP-MS NexION 350D, PerkinElmer, USA). Release curves of the adhesion layers with different Zn²⁺ concentrations (1%, 5%, and 10%) were evaluated over four weeks.

[0053] Inductively coupled plasma mass spectrometry (ICP-MS) analysis showed that the adhesion layers containing 5% and 10% Zn²⁺ promoted sustained release during the first week, with the 5% group exhibiting the highest cumulative release rate in the fourth week. Figure 3 D).

[0054] 4. Adhesion performance evaluation The adhesive ability of the LA-Zn adhesive film (2cm × 1cm) was qualitatively demonstrated by bonding it to fingers and pigskin substrates (in air and underwater). Adhesive performance was recorded by photographic documentation. Quantitative shear strength was measured according to the modified ASTM F2255 standard. In short, the LA-Zn adhesive film (2cm × 1cm) was bonded to two pieces of pigskin, and slight compression (~1kPa) was applied for 30 seconds to form an adhesive joint (n=3). Shear strength was measured using a universal testing machine at a constant deformation rate of 5mm / min. The maximum shear force before failure was recorded, and the shear strength (kPa) was calculated by dividing this force by the initial adhesive area.

[0055] To simulate the ability of the SMART patch to hold wound edges together, a wound closure strength test was conducted. A piece of pigskin was cut in half and different Zn... 2+ LA-Zn adhesive films (2cm × 1cm) at concentrations (1%, 5%, and 10%) and commercial adhesive Tegaderm TM Two pieces of pigskin were bridged and bonded together. The overlapping area was fixed for 10 minutes (n=3). The maximum force required to separate the bonded skin pieces was measured at a constant deformation rate of 5 mm / min and defined as the wound closure strength.

[0056] The results are as follows Figure 3As shown in Figure E, the LA-Zn adhesive membrane achieved good skin adhesion under both dry and humid conditions. In the quantitative evaluation of bioadhesion performance results, such as... Figure 3 As shown in Figure F, the LA-Zn adhesive film in the 5% group had the highest shear force, reaching 22.30 ± 0.85 kPa. Figure 3 F(ii)). Furthermore, the 5% group showed a wound closure strength of up to 0.48 N on wet pigskin, superior to commercially available wound dressings (Tegaderm). TM () Figure 3 F(iii)). The strong adhesion between the LA-Zn adhesive film and the skin tissue is attributed to the fact that the carboxyl groups in the adhesive and Zn²⁺ promote the formation of hydrogen bonds and Zn²⁺-carboxylate coordination bonds at the interface. Figure 3 F(i)). In summary, this invention selects a zinc ion content of 5%.

[0057] Example 3 Preparation of SMART patches In this invention, the SMART patch has a Janus structure. SMART stands for shape memory adhesive regenerative therapeutic.

[0058] A LA-Zn adhesion film solution with a 5% zinc ion content was directly poured onto the surface of a pre-formed LA-CA@Fe SMP film. The system was then dried at room temperature to obtain the final SMART patch, as shown in the figure. Figure 3 A.

[0059] In this embodiment, the preparation steps of the LA-Zn adhesion membrane solution with a zinc ion content of 5% are as follows: 1g of LA is dissolved in 3mL of anhydrous ethanol under gentle stirring. Then, 5%wt% zinc nitrate hexahydrate (Zn(NO3)2·6H2O, Z118841, Aladdin, Shanghai, China) is added to the solution to obtain a mixture. The mixture is ultrasonically treated for 10 minutes to ensure complete dissolution, thereby obtaining the LA-Zn adhesion membrane solution with a zinc ion content of 5%.

[0060] Characterization of SMART patches 1. Shape memory performance The shape memory performance of the SMART patch was tested according to the method described in Example 1.

[0061] The results showed that the SMART patch retained excellent shape memory properties and could recover its original shape when exposed to body temperature after stretching and fixing. Figure 4 A).

[0062] 2. Antibacterial properties Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus) were cultured and amplified in liquid lysogenic broth. LA-Zn adhesion membranes, LA-CA@Fe SMP, and SMART patches were sterilized by UV irradiation in 24-well plates for 24 hours. Subsequently, 10 μL of bacterial suspension (10 8 The initial dilution (CFU / mL) was pipetted onto the surface of each material and incubated at 37°C for 2 hours. After incubation, 2 mL of PBS was added to each well for dilution; then 20 μL of this initial dilution was transferred to 5 mL of fresh PBS, and 20 μL of the second dilution was spread onto lysogenic broth agar plates. The plates were incubated at 37°C for 12 hours, after which the colony-forming units (CFU) on each plate were counted and recorded as Ce (CFU of the material exposure group) (n=3). Cn represents the CFU count of the corresponding control group of unexposed material. The bacterial inhibition rate was calculated using the following formula:

[0063] Bacterial inhibition rate = (Cn - Ce) / Cn Figure 4 The results showed that both the adhesion layer and the shape memory layer had excellent antibacterial effects, with the adhesion layer having a significantly higher antibacterial rate than the shape memory layer. The synergistic effect of the two layers enabled the SMART patch to exhibit a 100% antibacterial rate.

[0064] 3. Biocompatibility L929 cells were co-cultured with LA-Zn adhesion membranes, LA-CA@Fe SMP, and SMART patches for 48 hours. Live / dead cell viability / toxicity assays were performed using a live / dead cell viability / toxicity assay kit (PF00007, Proteintech, Wuhan, China). Green and red fluorescence were observed using a confocal laser scanning microscope (Nikon, Tokyo, Japan). Cell proliferation (n=3) was assessed using a cell counting kit-8 (CCK-8, C0037, Beyotime, Shanghai, China). The optical density (OD) of the samples was recorded at 450 nm. Cell proliferation activity was calculated using the following formula: Proliferation activity (%) = [A(treatment) - A(blank)] / [A(control) - A(blank)] × 100%. Live / dead cell staining results are shown below. Figure 4 C(i) showed minimal cell death, indicating good biocompatibility. Furthermore, the CCK-8 assay results were as follows: Figure 4 C(ii) shows that both the single component (LA-Zn adhesive membrane, LA-CA@Fe SMP) and the integrated SMART patch promoted cell proliferation at 24, 48 and 72 hours.

[0065] The effect of SMART patches on diabetic wound healing 1. Construction of a diabetic mouse wound model Six-week-old C57BL / 6J mice were purchased from the Laboratory Animal Research Center of the School of Stomatology, Fourth Military Medical University, Xi'an, China, and housed in a specific pathogen-free environment. Animal experiments were conducted at the Laboratory Animal Center of the Fourth Military Medical University. All procedures followed the university's guidelines for laboratory animal care and use. Mice were fed a high-fat diet containing 60% fat for four weeks, followed by intraperitoneal injection of streptozotocin (STZ, 50 mg / kg body weight) to establish a diabetic mouse model. Mice injected with STZ continued to be fed a high-fat diet. To create skin wounds, diabetic mice were anesthetized by intraperitoneal injection of 1% sodium pentobarbital (50 mg / kg). In vivo, a full-thickness wound defect model of circular skin lesions (7 mm in diameter) was constructed on the backs of mice (n=3). A schematic diagram is shown below. Figure 5 A.

[0066] 2. Evaluate the efficacy of SMART patches on diabetic chronic wounds. (1) Assessment of wound size and healing speed Commercial adhesive Tegaderm was applied to the wound sites of diabetic mice. TM LA-Zn adhesive membrane, LA-CA@FeSMP, and SMART patch were used. Wounds were photographed with a digital camera on days 0, 3, 6, 10, and 13. ImageJ software was used to analyze the images to assess wound area. Results are as follows: Figure 5 B. On day 3, a significant reduction in wound size was observed in the LA-CA@Fe SMP and SMART patch groups, while this was not observed in the LA-Zn adhesive membrane and Tegaderm groups. This early wound closure (days 0–3) can be attributed to the shape memory properties of the LA-CA@Fe SMP layer, which generates a mechanical contractile force upon exposure to body temperature. This rapid mechanical contraction is crucial because it immediately reduces the wound area, thereby reducing the risk of infection and fluid loss. This aligns with a core principle of mechanobiology, recognizing that mechanical forces are the fundamental driving force for cell proliferation and migration during the initial healing phase. On day 6, the SMART patch group showed a further significant reduction in wound area. Interestingly, during days 3–6, the wound area reduction rate in the LA-Zn adhesive membrane group was significantly higher than that in the LA-CA@Fe SMP group (…). Figure 5 C). The second phase of accelerated healing (days 3-6) is primarily mediated by the sustained release of Zn²⁺, which regulates cellular biological behavior. This sequential treatment, from initial mechanical contraction to subsequent biochemical regulation, aligns with the natural chronological order of wound healing, achieving a complex level of regulation that cannot be replicated by a single mechanism strategy. Therefore, the SMART patch group, synergistically combining both mechanisms, consistently exhibited the smallest wound area ( Figure 5B, 5C). On day 13, the SMART patch group showed complete wound closure, with the wound surface covered by hair. In contrast, the Tegaderm™ group required an average of 26 days for complete closure. Figure 5 B, 5D).

[0067] (2) Histological staining of the wound after healing To assess the histological structure of the healing wound, tissue samples were taken on day 21 post-injury for histological analysis. Figure 5 E-staining Masson and hematoxylin-eosin (H&E) results showed complete wound closure at 21 days in the SMART patch, LA-CA@Fe SMP, and LA-Zn adhesive membrane groups. However, tissue treated with Tegaderm™ showed epidermal crusting and incomplete dermal closure. Figure 5 E). Masson staining showed enhanced collagen deposition in all treatment groups compared to the Tegaderm™ group. The SMART patch and LA-CA@Fe SMP groups showed the most significant collagen regeneration, reflecting the positive effect of mechanical force on tissue remodeling. Figure 5 F(i)). Notably, SMART patch and LA-CA@Fe SMP treatment promoted the regeneration of skin appendages, with numerous immature hair follicles (black arrows) and newly formed sebaceous glands (green asterisks) observed in the HE staining results. Figure 5 E, 5F (ii, iii)). Tissue treated with LA-Zn adhesive membranes showed subepithelial cysts (yellow asterisks), which, although not forming hair follicle structures, at least represented initial sites for the development of some new hair follicle morphology. In contrast, almost no new hair follicles and sebaceous glands were observed in the Tegaderm™ group. Figure 5 E).

[0068] Molecular mechanisms by which SMART patches promote rapid closure and regenerative healing of diabetic wounds 1. Transcriptome sequencing analysis of the effects of SMART patches on diabetic wounds at the RNA level Skin wounds were constructed in diabetic mice using the same method described above. LA-CA@FeSMP, LA-Zn adhesive membranes, and SMART patches were used at the wound sites. Wounds in the control group were left untreated. On day 13, skin wound tissue from the backs of diabetic mice was collected for RNA sequencing (n=3). Total RNA extraction, RNA-seq library construction, Illumina sequencing, and bioinformatics analysis were performed by Shanghai Sangon Biotech Co., Ltd. (Shbio, Shanghai, China). In short, gene expression profiles were obtained using the Illumina HiSeq sequencing platform. Quality control checks, principal component analysis, GO enrichment analysis, and KEGG enrichment analysis were performed. Differential gene expression analysis was performed using DESeq2, with a significant change defined as P < 0.05 and log2 (fold change) > |1|.

[0069] To explore the specific mechanism by which SMART patches promote the regeneration of diabetic wounds, this invention compares LA-CA@FeSMP with a control group to analyze the effect of mechanical force in wound healing. In addition, this invention also compares SMART patches and LA-CA@Fe SMP to explain the effect of Zn²⁺ release in wound healing. Figure 6 Volcano plot A shows differentially expressed genes among the control group, SMART patch group, and LA-CA@Fe SMP group. Figure 6 GO enrichment analysis of B showed that, compared with the control group, the genes upregulated in the LA-CA@Fe SMP group were associated with mechanotransduction, extracellular matrix deposition, hair follicle regeneration, angiogenesis, and macrophage immune regulation. Figure 6 The heatmap of C focuses on genes related to mechanoconstriction, extracellular matrix deposition, and hair follicle regeneration, all showing a significant upregulation pattern. For example, key genes involved in mechanotransduction and contraction (such as Myl6, Tnnc1, and Krtap), ECM components (such as Col5al), and hair follicle morphogenesis (such as keratin-related protein genes) are significantly upregulated, providing a molecular basis for the observed physical wound contraction, collagen regeneration, and appendage recovery. To elucidate the unique biochemical mechanism mediated by sustained Zn²⁺ release, this invention compares the transcriptomes of the SMART patch group and the LA-CA@Fe SMP group. Figure 6 GO enrichment analysis of D showed that the SMART patch enhanced pathways involved in reactive oxygen species (ROS) responses, inflammation regulation, positive regulation of wound healing, and angiogenesis. Figure 6E validated this finding through heatmap analysis of differentially expressed genes related to these pathways, showing downregulation of oxidative stress-related genes (such as Cxcl1 and Itgam) and pro-inflammatory genes (such as Il1b and Tnfrsf1b) in the SMART patch group, indicating successful mitigation of oxidative stress and inflammation. Simultaneously, the upregulation of key growth factors and their receptors involved in proliferation and migration provided a direct molecular explanation for accelerated wound healing. Furthermore, the upregulation of vascularization-related genes indicated that the SMART patch could promote angiogenesis in vivo.

[0070] KEGG pathway analysis revealed more details about the SMART patch's healing mechanism via a dual mechano-biochemical pathway. Figure 7 A KEGG results showed that LA-CA@Fe SMP significantly enriched pathways related to cell adhesion and extracellular matrix-receptor interactions compared to the control group, confirming that the shape memory polymer layer activated mechanotransduction in these cells. The PI3K-Akt and cAMP-PKG signaling pathways were significantly upregulated. The PI3K-Akt pathway is a well-established mechanoresponsive pathway known to be activated by integrin binding and cytoskeletal tension, thereby promoting cell survival, proliferation, and migration—processes crucial for wound closure. The cAMP-PKG pathway, stimulated by mechanical stress, leads to changes in second messenger dynamics, a process involved in enhancing endothelial barrier function and angiogenesis. Therefore, the KEGG results indicate that the mechanoconstriction of the SMART patch is sensed by cells through the adhesion complex, triggering downstream signaling via the PI3K-Akt and cAMP-PKG pathways to regulate cellular responses and tissue regeneration. Figure 7 KEGG pathway analysis of B further elucidated the unique biochemical regulatory role of the adhesion layer. Comparison between the SMART patch group and the LA-CA@Fe SMP group revealed the downregulation of pathways related to inflammation and oxidative stress. Among the downregulated pathways, the NF-κB signaling pathway, as a major regulator of inflammation, controls the expression of pro-inflammatory cytokines, chemokines, and enzymes involved in ROS production. Its downregulation indicates that Zn²⁺ continuously released from the adhesion layer exerts its therapeutic effect through the NF-κB signaling pathway. In summary, transcriptomic analysis shows that the SMART patch regulates the regeneration and repair of diabetic wounds through a synergistic yet distinct mechano-biochemical dual pathway.

[0071] 2. In vivo validation of the mechano-biochemical synergistic mechanism of SMART patch (1) In vivo verification of hair follicle regeneration The promoting effect of SMART patches on hair follicle regeneration was verified by immunofluorescence staining targeting cytokeratin 5 (KRT5, a marker of hair follicle stem cells and progenitor cells). Figure 8 As shown in Figure A, the abundance of KRT5-positive cells was significantly higher in wounds treated with SMART patches. Figure 8 The semi-quantitative statistical plot of E shows that, compared with the LA-Zn adhesive membrane group and the control group, the KRT5 fluorescence intensity of the SMART patch group and the LA-CA@Fe SMP group was significantly increased, and the SMART patch had the most significant promoting effect. This indicates that mechanical contraction is the main driving factor for activating hair follicle stem cells.

[0072] (2) Verification of pro-angiogenic characteristics The pro-angiogenic properties of the SMART patch were verified by immunofluorescence staining of CD31. Figure 8 As shown in Figure B, a denser network of CD31-positive microvessels was observed in the SMART patch group, such as... Figure 8 The semi-quantitative results shown in F further confirm the immunofluorescence effect.

[0073] (3) Verification of immune regulatory effects The effect of SMART patches on macrophage immune regulation was detected by immunofluorescence staining of macrophage markers CD86 and CD206. Results are as follows: Figure 8 C showed that the SMART patch group exhibited the most significant differentiation of macrophages into the M2 repair phenotype, while inhibiting their differentiation into the pro-inflammatory M1 phenotype. Furthermore, the LA-CA@Fe SMP group also showed a trend towards M2 macrophage differentiation, while the LA-Zn adhesive membrane group and the control group showed no significant difference in M2 macrophage differentiation. Figure 8 (G, H). This finding provides direct evidence that mechanical force itself can directly drive macrophages to polarize towards a pro-repair surface.

[0074] (4) Verification of antioxidant stress capacity The levels of reactive oxygen species (ROS) at the wound site after different treatment groups were detected using a reactive oxygen species assay kit. Figure 8 As shown in D, 8I, a significant reduction in ROS signaling was observed in the LA-Zn adhesion membrane group, demonstrating the most effective extracellular matrix ROS clearance. In the LA-CA@Fe SMP group, ROS remained in the extracellular matrix, but it was more concentrated around newly formed hair follicles. The SMART patch group exhibited a combined effect: reduced extracellular ROS, while ROS accumulated around newly formed hair follicles. This indicates that the LA-Zn adhesion membrane is primarily responsible for clearing extracellular ROS, while mechanical force alone does not contribute significantly to this process. Furthermore, ROS plays a dual role in diabetic wound healing; its accumulation in the extracellular matrix impairs the migration ability of keratinocytes and the collagen secretion ability of fibroblasts, thus slowing the wound closure process. However, ROS enriched around hair follicle stem cells can promote stem cell proliferation and activation, which is beneficial for hair follicle development.

[0075] It should be noted that when numerical ranges are mentioned in the claims of this invention, it should be understood that the two endpoints of each numerical range and any value between the two endpoints can be selected. To avoid redundancy, the present invention describes preferred embodiments.

[0076] Although preferred embodiments of the invention have been described, those skilled in the art, upon learning the basic inventive concept, can make other changes and modifications to these embodiments. Therefore, the appended claims are intended to be interpreted as including both the preferred embodiments and all changes and modifications falling within the scope of the invention.

[0077] Obviously, those skilled in the art can make various modifications and variations to this invention without departing from its spirit and scope. Therefore, if these modifications and variations fall within the scope of the claims of this invention and their equivalents, this invention also intends to include these modifications and variations.

Claims

1. A body temperature responsive double-sided patch, characterized by, The body temperature responsive double-sided patch is composed of a shape memory polymer and a zinc ion sustained-release adhesive layer. The shape memory polymer is prepared by cross-linking diisopropenyl benzene, thioctic acid, cellulose acetate and iron ions; the mass ratio of diisopropenyl benzene, thioctic acid, cellulose acetate and the compound containing iron ions is 0.5-0.8:2-4:0.15-0.75:0.05-0.

15. The zinc ion sustained-release adhesive layer is obtained by dissolving zinc ions in a thioctic acid solution.

2. The body temperature-responsive double-sided patch according to claim 1, characterized by The preparation steps of the shape memory polymer are as follows: diisopropenyl benzene and thioctic acid are used as fixed components, and cellulose acetate and the compound containing iron ions are dissolved in N,N-dimethylformamide to obtain a mixed solution; the mixed solution is heated and solidified to obtain the shape memory polymer.

3. The body temperature-responsive double-sided patch according to claim 1, characterized by The heating and solidification step is as follows: the mixed solution is sequentially heated at 70-90℃ for 3-6h, 90-110℃ for 1-2h and 130-150℃ for 3-5h.

4. The body temperature-responsive double-sided patch according to claim 1, characterized by The preparation steps of the zinc ion sustained-release adhesive layer are as follows: zinc nitrate hexahydrate is added to the thioctic acid solution, and the solution is poured into a mold after dissolution, dried to obtain the zinc ion sustained-release adhesive layer. The mass ratio of the thioctic acid to the zinc nitrate hexahydrate is 1:0.01-0.

5.

5. The method of claim 1, wherein the body temperature responsive double-sided patch is prepared by the steps of: The solution of the zinc ion sustained-release adhesive layer is poured onto the surface of the shape memory polymer, and dried to obtain the body temperature responsive double-sided patch.

6. The use of the body temperature responsive double-sided patch of claim 1 in the preparation of a drug for the healing of chronic wounds of diabetes.

7. Use according to claim 6, characterized in that, The drug is used to reduce the wound area.

8. Use according to claim 6, characterized in that, The drug is used to improve the wound healing speed.

9. Use according to claim 6, characterized in that, The drug is used to promote the tissue and functional regeneration of the wound.

10. Use according to claim 9, characterized in that, The tissue and functional regeneration includes hair follicle regeneration, blood vessel regeneration and sebaceous gland regeneration.

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