Myocardial repair patch based on multi-material printing and preparation and application thereof

Abstract

This invention relates to a myocardial repair patch based on multi-material printing, its preparation, and its application, belonging to the field of bio-3D printing and regenerative medicine technology. The invention uses PLCL and GelMA hydrogel loaded with active cardiomyocytes as bio-inks, and introduces gelatin gel as a heat buffer material. An orthogonal network structure is constructed using multi-material bio-3D printing technology to prepare a myocardial repair patch with excellent dynamic adaptability to the myocardial interface and good biocompatibility. The myocardial injury repair patch prepared by this invention achieves a structure-function integrated design through the synergistic effect of the three materials, significantly overcoming the limitations of single materials in terms of mechanical properties, biological function, or thermal compatibility. The mechanical properties of the myocardial injury repair patch prepared by this invention can precisely match those of natural myocardium, while possessing high cell activity, effectively inhibiting fibrosis, improving myocardial function, and achieving true active repair. Furthermore, the process conditions are controllable, demonstrating good translational potential.

Description

Technical Field

[0001] This invention relates to the field of bio-3D printing and regenerative medicine technology, specifically to a myocardial injury repair patch based on multi-material printing, its preparation, and its application. This patch integrates active cardiomyocytes, a biodegradable polymer scaffold, and a thermal protection structure, possessing biomimetic mechanical properties, excellent dynamic adaptability to the myocardial interface, and good biocompatibility. It is suitable for the repair and treatment of myocardial defects such as myocardial infarction and cardiac trauma. Background Technology

[0002] Globally, cardiovascular diseases (CVD) are the leading cause of death, with persistently high morbidity and mortality rates. In my country, over 40% of deaths are attributed to CVD. Myocardial infarction is one of the main clinical manifestations of CVD, characterized by the irreversible loss of a large number of functional cardiomyocytes due to severe ischemia. Following infarction, the heart tissue undergoes inflammatory responses, ventricular remodeling, arrhythmias, and progressive dilation, ultimately leading to heart failure. Because adult cardiomyocytes have extremely limited proliferative capacity, the damaged area is replaced by non-contractile fibrotic scar tissue during the repair process, significantly impairing the heart's pumping function.

[0003] Against this backdrop, cardiac repair patches have shown great potential as a postoperative adjuvant therapy strategy. An ideal cardiac patch can not only provide mechanical support but also regulate the local microenvironment, inhibit fibrosis, and promote myocardial regeneration. However, existing patches generally have structural and functional limitations, lacking the anisotropic topological structure of the natural extracellular matrix, making it difficult to guide the directional alignment and synchronous contraction of cardiomyocytes. Although bioprinting technology can precisely construct biomimetic anisotropic scaffolds, effectively promoting the oriented growth and functional maturation of cardiomyocytes, most current printed patches still do not contain active cardiomyocytes and cannot directly participate in the reconstruction of myocardial contractile function.

[0004] In recent years, the rapid development of multi-material bio-3D printing technology has provided a new path to overcome the bottlenecks of traditional cardiac repair materials in terms of structural biomimicry, bioactivity, and mechanical compatibility. This study constructed a novel cardiac repair patch with excellent mechanical properties and high cell viability by fusing two functionally complementary bio-inks: thermoplastic poly(lactide-caprolactone) copolymer (PLCL) and methacrylamide gelatin (GelMA) hydrogel loaded with cardiomyocytes. To address the thermal damage to cells in the adjacent GelMA layer caused by the high-temperature extrusion of PLCL, a novel local cooling technology was introduced to effectively reduce the printing interface temperature and significantly improve cell survival rate. PLCL provides elastic modulus and structural support matching natural myocardium, while the GelMA hydrogel acts as a cell carrier to maintain the survival and function of cardiomyocytes. This patch not only mimics the mechanical and topological properties of myocardial tissue in its microstructure but also adheres stably in vivo and is easy to suture, providing a promising regenerative medicine solution for functional repair after cardiac trauma and myocardial infarction. Summary of the Invention

[0005] The purpose of this invention is to provide a novel method for preparing and using a myocardial injury repair patch. This patch, based on multi-material bio-3D printing technology, innovatively combines two complementary bio-inks: PLCL and GelMA hydrogel loaded with active cardiomyocytes. This results in a composite cardiac repair patch that possesses a biomimetic anisotropic microstructure, excellent dynamic interface adaptability, biodegradability, and high cell activity.

[0006] To achieve the above objectives, the present invention adopts the following technical solution:

[0007] The first aspect of this invention provides a method for preparing a myocardial repair patch based on multi-material printing, comprising the following steps:

[0008] S1. Material preparation: Poly(lactide-caprolactone) copolymer PLCL, methacrylamide gelatin GelMA, and gelatin hydrogel are used as raw materials, and the raw materials are pretreated.

[0009] S1.1. Methacrylamide gelatin (GelMA) loaded with cardiomyocytes was dissolved in phosphate buffer, the pH of the solution was adjusted, and a photoinitiator was added to obtain GelMA hydrogel;

[0010] S1.2. Dissolve the gelatin gel in phosphate buffer to prepare a gelatin hydrogel solution;

[0011] S2. Printing Parameter Setting and Import: Using extrusion-type bio-3D printing technology, a pre-designed three-dimensional model of myocardial repair patch is imported into the bio-3D printer, and printing parameters are set; among them, the printing temperature of PLCL is 140℃~155℃;

[0012] S3. Multi-material collaborative printing:

[0013] S3.1. First layer construction: Print a parallel filament array arranged in a single direction, and then print a GelMA hydrogel containing cardiomyocytes in the gaps between the PLCL filaments to form the first layer;

[0014] S3.2. Second layer construction: On top of the GelMA hydrogel, a layer of gelatin hydrogel is printed along its trajectory as a heat buffer protective layer; then, molten PLCL material is printed in the axial direction perpendicular to the lower PLCL to form a parallel filament array arranged in a single direction, constructing an orthogonal mesh structure to form the scaffold part of the patch; finally, GelMA hydrogel containing cardiomyocytes is printed again in the gaps between the newly printed PLCL filaments to complete the integration of the three materials in the second layer;

[0015] S3.3. Repeat steps S3.1 to S3.2 to obtain the printed patch structure;

[0016] S4. Post-processing: Irradiate the printed structure under ultraviolet light to obtain a myocardial injury repair patch.

[0017] In S1, the molar ratio of lactide to caprolactone in PLCL is 50:50, and the weight-average molecular weight of PLCL is 200 kDa.

[0018] In S1.1, GelMA was dissolved in phosphate buffer, and the concentration of GelMA was 0.05 g / mL; the photoinitiator was lithium phenyl (2,4,6-trimethylbenzoyl) phosphate, and the concentration of the photoinitiator was 0.005 g / mL; the pH of the solution was adjusted to 7.0 before adding the photoinitiator.

[0019] In S1.2, the concentration of the gelatin gel solution is 0.1 g / mL;

[0020] In S2, during the printing process, the printing speed is 1mm / s-10mm / s, the volume ratio of PLCL, GelMA hydrogel, and gelatin hydrogel are set independently, with a setting value of 1~2, and the patch size is set as needed; the printing thickness of each layer is set as needed, and the thickness is adapted to the nozzle size;

[0021] During the printing process, the printing spacing and nozzle diameter are set according to the type of raw material and the requirements of the target patch;

[0022] In S3, the total printing time, starting from the start of printing, shall not exceed 50 minutes.

[0023] In S3.3, repeat steps 3.1 to 3.2 until the patch thickness reaches the target number of layers;

[0024] In S4, the ultraviolet light wavelength is 365nm, and the irradiation time is 25s~55s.

[0025] A second aspect of the present invention provides a myocardial repair patch based on multi-material printing, which is prepared using the above-described method.

[0026] A third aspect of the present invention provides an application of a myocardial repair patch based on multi-material printing.

[0027] The advantages and positive effects of this invention are as follows:

[0028] 1. The novel myocardial injury repair patch of the present invention is designed to achieve integrated structure and function through the synergistic use of three materials (PLCL, GelMA hydrogel, and gelatin hydrogel).

[0029] PLCL provides long-term mechanical support and a scaffold structure that can be controlled to degrade. GelMA hydrogel acts as a bioactive carrier, effectively maintaining cell survival, while gelatin hydrogel serves as a thermal buffer layer, absorbing the heat released by PLCL during the printing process for effective cooling. The three materials work synergistically and complement each other to jointly construct a dual-functional system integrating "structural support + bioactivity," significantly overcoming the limitations of single materials in terms of mechanical properties, biological functions, or thermal compatibility.

[0030] 2. The novel myocardial injury repair patch of the present invention has high cell activity and can achieve true active repair.

[0031] The novel myocardial injury repair patch provided by this invention includes a protective layer structure for thermal protection and a functional design to maintain high cell proliferation. By introducing local cooling technology during the printing process, the temperature rise in the GelMA hydrogel area is effectively controlled, significantly reducing thermal damage and improving the activity of myocardial cells within the patch. This solves the problem of massive cell death caused by thermal stress in traditional hybrid printing, and fully protects the biological function and repair potential of the patch.

[0032] 3. The novel myocardial injury repair patch of the present invention has mechanical properties that can be precisely matched with natural myocardium.

[0033] By leveraging the adjustable mechanical properties of PLCL material and optimizing printing temperature and mesh porosity, a scaffold structure with an elastic modulus highly matched to that of real myocardial tissue was constructed. This structure not only provides sufficient mechanical support to resist ventricular wall stress but also avoids stress shielding caused by excessive stiffness.

[0034] 4. The novel myocardial injury repair patch of the present invention has an anisotropic structure and can guide functional myocardial regeneration.

[0035] By using multi-material 3D printing technology to precisely construct anisotropic structures that mimic the topological features of the extracellular matrix of natural cardiomyocytes, the directional arrangement and synchronous beating of cardiomyocytes can be guided, thereby promoting the maturation and integration of functional myocardial tissue.

[0036] 5. The novel myocardial injury repair patch of the present invention has excellent dynamic interface adaptability and surgical operability.

[0037] The patch combines flexibility, stretchability, and good tissue adhesion, allowing it to closely conform to the complex curves of a beating heart. It can stretch and retract synchronously during heartbeats, facilitating intraoperative suturing and fixation. Its mesh structure design allows the aperture to be slightly larger than that of conventional suture needles, making it easy to suture without damaging the microstructure. It is suitable for minimally invasive or open-chest surgery, significantly improving clinical operability and stability. Animal experiments have shown that it is easy to suture and does not easily shift, making it suitable for minimally invasive or open-chest surgery scenarios and significantly improving clinical applicability.

[0038] 6. The novel myocardial injury repair patch described in this invention can effectively inhibit fibrosis and improve myocardial function.

[0039] Implantation of active cardiomyocytes can effectively inhibit the formation of pathological fibrous scars by regulating the local microenvironment through active cardiomyocytes.

[0040] 7. The novel myocardial injury repair patch of the present invention has controllable process conditions and has good prospects for commercialization.

[0041] The multi-material printing process using PLCL, GelMA hydrogel, and gelatin hydrogel employed in this invention has well-defined parameters and good repeatability. Combined with CT data, it can achieve customized manufacturing for patients. The degradation products are non-toxic and metabolizable, providing an advanced treatment strategy that integrates structural support, bioactivity, mechanical matching, and anti-fibrosis for the repair of myocardial infarction, cardiac trauma, or postoperative myocardial defects. It also has good surgical operability and potential for individualized customization, laying a solid foundation for subsequent clinical translation. Attached Figure Description

[0042] Figure 1 The present invention provides a process flow diagram for the preparation method of a myocardial repair patch based on multi-material printing.

[0043] Figure 2 Images of 66% mesh pore density myocardial injury repair patches printed at different temperatures; (a) at 135℃, (b) at 140℃, (c) at 145℃, (d) at 150℃, and (e) at 155℃.

[0044] Figure 3Stress-strain tests were performed on patches with two mesh porosity densities of 66% and 75%; where (a) is a 66% mesh porosity and (b) is a 75% mesh porosity.

[0045] Figure 4 The stress-strain test results are for the myocardial injury repair patch; where (a) is the 66% mesh porosity and (b) is the 75% mesh porosity.

[0046] Figure 5 Elastic modulus test of ventricular tissue in Bama pigs.

[0047] Figure 6 Comparison of the elastic modulus of cardiac repair patches prepared with different combinations of process parameters and Bama pig ventricular tissue.

[0048] Figure 7 This shows the change in cell survival rate over time within the GelMA hydrogel.

[0049] Figure 8 The diagram shows the printed structures with and without a protective layer; where (a) represents the condition without a protective layer and (b) represents the condition with a protective layer.

[0050] Figure 9 The simulation results of the temperature field of PLCL under the conditions of no protective layer and with protective layer are shown; where (a) is the condition of no protective layer and (b) is the condition of with protective layer.

[0051] Figure 10 The experiment demonstrates the verification of temperature field simulation results during PLCL printing using a thermal imager; (a) represents the condition without a protective layer, and (b) represents the condition with a protective layer.

[0052] Figure 11 The results of staining live / dead cells on cell patches under conditions with and without protective layer are shown. Among them, (a) is under the condition without protective layer, (b) is under the condition with protective layer, and (c) is a comparison of cell viability under the two conditions with and without protective layer.

[0053] Figure 12 The image shows a physical image of a myocardial injury repair patch prepared at a printing temperature of 155°C and a mesh porosity of 75%.

[0054] Figure 13 Microscopic morphology images of PLCL, GelMA hydrogel and gelatin hydrogel in myocardial injury repair patch.

[0055] Figure 14The study shows the comparison of the number of live cells on cell patches printed using local cooling technology after 1, 3 and 7 days of culture; (a) is the result of 1 day of culture, (b) is the result of 3 days of culture, (c) is the result of 7 days of culture, and (d) is the result of survival rate comparison.

[0056] Figure 15 The results are shown in the experimental data one month after the myocardial repair patch was implanted in vivo. Among them, (a) is the tissue section image of the MA group, (b) is the tissue section image of the MB group, (c) is the tissue section image of the MC group, and (d) is the level of apoptosis in the myocardial tissue of each group. Detailed Implementation

[0057] This invention provides a method for preparing a myocardial repair patch based on multi-material printing, the process flow of which is as follows: Figure 1 As shown, it includes the following steps:

[0058] S1. Material preparation: The raw materials are pretreated using poly(lactide-caprolactone) copolymer PLCL (lactide to caprolactone molar ratio of 50:50), methacrylamide gelatin GelMA, and gelatin hydrogel with a weight average molecular weight of 200 kDa.

[0059] S1.1. Methacrylamide gelatin (GelMA) loaded with cardiomyocytes was dissolved in phosphate buffer and the pH of the solution was adjusted to pH=7.0; GelMA hydrogel was obtained by adding 0.005 g / mL of phenyl (2,4,6-trimethylbenzoyl) lithium phosphate as a photoinitiator to the solution.

[0060] S1.2. Dissolve the gelatin hydrogel in phosphate buffer to prepare a gelatin gel solution with a concentration of 0.1 g / mL;

[0061] S2. Printing Parameter Setting and Import: Using extrusion-based bio-3D printing technology, a pre-designed 3D model of a myocardial repair patch is imported into the bio-3D printer, and printing parameters are set. The printing speed is 1mm / s to 10mm / s; the volume ratio of the PLCL is 1 to 2; and the printing temperature is 140℃ to 155℃. The shape and specifications of the patch are set as needed. During PLCL material printing, the distance between the PLCL filament axes and the nozzle diameter are kept constant. The diameter of the printed material is controlled by changing the volume ratio parameter, thus achieving a printing diameter larger than the nozzle diameter. The thickness of each layer is set as needed and is adapted to the nozzle specifications.

[0062] S3. Multi-material collaborative printing:

[0063] S3.1. First layer construction: Print a parallel filament array arranged in a single direction, and then print a GelMA hydrogel containing cardiomyocytes in the gaps between the PLCL filaments to form the first layer;

[0064] S3.2. Second layer construction: On top of the GelMA hydrogel, a layer of gelatin hydrogel is printed along its trajectory as a heat buffer protective layer; then, molten PLCL material is printed in the axial direction perpendicular to the lower PLCL to form a parallel filament array arranged in a single direction, constructing an orthogonal mesh structure to form the scaffold part of the patch; finally, GelMA hydrogel containing cardiomyocytes is printed again in the gaps between the newly printed PLCL filaments to complete the integration of the three materials in the second layer;

[0065] S3.3. Repeat steps S3.1 to S3.2 until the patch thickness reaches the target value to obtain the printed patch structure; the total printing time, calculated from the start of printing, shall not exceed 50 minutes.

[0066] S4. Post-processing: Irradiate the printed structure under ultraviolet light for 25s~55s to obtain a myocardial injury repair patch.

[0067] In the method provided by this invention, the shape and specifications of the patch can be set as needed.

[0068] This invention also provides a myocardial repair patch based on multi-material printing, prepared using the above-described method. The patch provided by this invention is prepared using multi-material bio-3D printing technology, possessing high cell activity, mechanical properties that precisely match natural myocardium, excellent dynamic interface adaptability, and the ability to effectively inhibit fibrosis to improve myocardial function.

[0069] This invention also provides an application of a myocardial repair patch based on multi-material printing.

[0070] The PLCL (poly(lactide-caprolactone) copolymer) used in this embodiment of the invention was purchased from Jinan Dai Biotechnology Co., Ltd., model DG-50LPLCL200, with a lactide to caprolactone molar ratio of 50:50 and a weight-average molecular weight of approximately 200 kDa. This material exhibits good thermoplasticity, adjustable elastic modulus, and controllable biodegradability, making it suitable as a structural support material and applicable to extrusion-based bio-3D printing processes.

[0071] The GelMA (methacrylated gelatin) hydrogel used in this embodiment of the invention was purchased from Suzhou Yongqinquan Intelligent Equipment Co., Ltd., model EFL-GM-60, and was used as a bioactive carrier for loading cardiomyocytes derived from induced pluripotent stem cells (iPSCs).

[0072] The gelatin hydrogel used in this embodiment of the invention was purchased from Sigma-Aldrich, product model V900863-500G, and is type A porcine gelatin.

[0073] The bio-3D printer used in this embodiment of the invention is the commercial bio-3D printer SIA BioPrinter Pro.

[0074] This invention utilizes the BiopDesigner software developed using MATLAB for printing path planning. The software imports an STL model of a myocardial repair patch, processes its complex geometry using a voxelization algorithm, sets the material parameters and motion trajectories for each nozzle, and automatically generates G-code to drive a multi-nozzle bio-3D printer to achieve precise, integrated patch construction. During printing, molten poly(lactide-caprolactone) copolymer (PLCL), methacrylamide gelatin (GelMA) hydrogel loaded with cardiomyocytes, and gelatin hydrogel are used as bio-inks. A gelatin hydrogel is placed on top of the GelMA hydrogel as a localized cooling and protective layer to absorb the heat released during PLCL extrusion, reducing the interface temperature and minimizing thermal damage to the embedded cells, while ensuring effective bonding between the material layers.

[0075] This invention uses PLCL as a structural scaffold to provide stable support that matches the mechanical properties of natural myocardium; GelMA hydrogel loaded with cardiomyocytes forms the functional core layer of the patch, effectively promoting myocardial regeneration and vascularization; and gelatin hydrogel acts as a thermal buffer layer for local cooling, absorbing the heat released by the high-temperature PLCL during the printing process. While ensuring effective adhesion between materials, it significantly reduces the interface temperature, thereby significantly reducing the risk of thermal damage to the cells in the GelMA, increasing the survival rate of cardiomyocytes in the patch to over 91%, and ensuring that it has the ability to repair functional myocardium. This achieves the precise integration and functional synergy of the three materials.

[0076] This invention, through precise control of the mechanical properties of the PLCL scaffold and the synergistic design of the cellular microenvironment within the GelMA hydrogel, effectively overcomes key shortcomings of existing myocardial repair materials, such as a lack of functional cardiomyocytes, insufficient structural biomimicry, poor dynamic surface adhesion, and difficulty in inhibiting fibrosis. The patch can stably integrate into the damaged myocardial region after implantation, providing not only immediate mechanical support but also inhibiting pathological fibrous scar formation through active cell-secreted factors, promoting angiogenesis and functional myocardial tissue regeneration.

[0077] The myocardial repair patch based on multi-material printing provided by this invention has excellent dynamic interface adaptability, can closely conform to the complex curvature of the beating heart, and facilitates intraoperative suturing and fixation. Simultaneously, the patch possesses flexibility, stretchability, and good tissue adhesion, allowing it to stretch and retract synchronously during cardiac pulsation. Its mesh structure design, with pores slightly larger than the diameter of conventional suture needles, facilitates needle insertion and suturing without damaging the microstructure, making it suitable for minimally invasive or open-chest surgery, significantly improving clinical operability and stability. After implantation, the patch can regulate the local microenvironment through active cardiomyocytes, effectively inhibiting pathological fibrosis and improving myocardial function.

[0078] In the method provided by this invention, the overall mechanical properties of the patch are determined by the PLCL. By optimizing the printing temperature range and mesh pore density of the PLCL, a high degree of matching between the overall mechanical properties of the patch and natural myocardial tissue is achieved.

[0079] Printing temperature is a key process parameter for controlling the mechanical properties of PLCL patches. Increasing the printing temperature reduces melt viscosity and enhances molecular chain fluidity, thereby inhibiting the crystallization process and leading to a decrease in the patch's elastic modulus. Conversely, a lower printing temperature restricts chain rearrangement, promotes crystal formation, and thus increases material rigidity. This invention sets the following printing parameters: printing speed 1mm / s-10mm / s, nozzle diameter 0.2mm; filament shaft distance 1mm; mesh porosity 66%; volume ratio: 1.36. PLCL supports in the bottom layer of elliptical patches with a semi-major axis of 10mm and a semi-minor axis of 6mm were printed on the printing platform at printing temperatures of 135℃, 140℃, 145℃, 150℃, and 155℃ respectively. The results are as follows: Figure 2 As shown. From Figure 2 It can be seen that at 135℃, PLCL failed to fully melt to the ideal viscous flow state, the molecular chains were not completely unentangled, the melt flow was poor, and the extruded filament did not fuse well with the cooled lower layer, leading to printing failure. Therefore, in the method provided by this invention, the PLCL printing temperature is set to 140℃~155℃. Within this range, the material can fully melt, the crystallization behavior can be effectively controlled, ensuring good interlayer bonding while taking into account dimensional stability and the required mechanical properties, thereby achieving high-quality printing.

[0080] Cells in GelMA hydrogels, once removed from the culture medium, face problems such as insufficient nutrient and oxygen supply, easily leading to decreased activity or even death. To clarify the tolerance limit of cells in GelMA hydrogels, this invention uses a cell viability rate greater than 90% as the criterion for high-activity printing. Cell-loaded GelMA hydrogels were individually extruded and printed at 10-minute intervals. The GelMA hydrogel printing parameters were set as follows: nozzle diameter: 0.25 mm; filament spacing: 1 mm; volume ratio: 1; printing temperature: 18℃. After printing, the samples were uniformly cultured for 24 hours, followed by live / dead cell staining. The cell viability rate at different printing time points (10 min, 20 min, 30 min, 40 min, 50 min, and 60 min) was observed and quantitatively analyzed using two-photon microscopy. Figure 7 The results showed that cell viability remained high within the first 50 minutes, with an average survival rate of approximately 92%. However, after printing time exceeded 50 minutes, the cell death rate accelerated significantly, and the survival rate plummeted to 59% at 60 minutes. Therefore, to ensure high cell viability during printing, the printing speed parameter of the patch was subsequently set to 6 mm / s to control the single cell-loaded printing operation to within 50 minutes, in order to avoid a significant decrease in cell viability due to prolonged exposure to a non-culture environment.

[0081] The present invention will be further described below with reference to the accompanying drawings and specific embodiments.

[0082] Example 1:

[0083] This embodiment provides a method for preparing a myocardial repair patch based on multi-material printing, including the following steps:

[0084] S1. Material Preparation: Using poly(lactide-caprolactone) copolymer PLCL (lactide to caprolactone molar ratio of 50:50), methacrylamide gelatin (GelMA), and gelatin hydrogel as raw materials, the raw materials are pretreated as follows:

[0085] S1.1. Methacrylamide gelatin (GelMA) loaded with cardiomyocytes was dissolved in phosphate-buffered saline (PBS) to prepare a 0.05 g / mL solution. The pH of the solution was adjusted to pH=7.0; using 0.005 g / mL of phenyl (2,4,6-trimethylbenzoyl) lithium phosphate as a photoinitiator, the GelMA hydrogel was obtained by adding the photoinitiator to the solution.

[0086] S1.2. Dissolve the gelatin hydrogel in phosphate-buffered saline (PBS) to prepare a 0.1 g / mL gelatin hydrogel solution, which serves as a local cooling and protective layer. During the printing process, it can effectively absorb the heat released during the extrusion of high-temperature PLCL material, significantly reducing the impact of heat conduction on the adjacent cell-carrying GelMA hydrogel, and exhibits good thermal buffering capacity, biocompatibility, and biodegradability.

[0087] S2. Printing Parameter Settings and Import:

[0088] The extrusion-based bio-3D printing process was employed, using the commercial bio-3D printer SIA BioPrinter Pro. Specifically, a pre-designed STL format 3D model of the myocardial repair patch was imported, and the following key printing parameters were set:

[0089] Printing speed: 6mm / s; Semi-major axis of elliptical patch: 10mm; Semi-minor axis of elliptical patch: 6mm; Patch thickness: 1mm; Printing thickness per layer: 0.25mm;

[0090] PLCL printing parameter settings: Nozzle diameter: 0.25mm; filament shaft distance: 1mm; mesh porosity: 75%; volume ratio: 1; printing temperature: 155℃; When printing PLCL material, keep the PLCL filament shaft distance and nozzle diameter constant, and control the printing material diameter by changing the volume ratio parameter to achieve the printing diameter requirement of being larger than the nozzle diameter.

[0091] GelMA hydrogel printing parameter settings: Nozzle diameter: 0.25mm; Filament pitch: 1mm; Volume ratio: 1; Printing temperature: 18℃;

[0092] Gelatin hydrogel printing parameter settings: Nozzle diameter: 0.25mm; filament pitch: 1mm; volume ratio: 1; printing temperature: 24℃.

[0093] S3. Multi-material collaborative printing:

[0094] Using bio-3D printing technology, a composite cardiac patch with a biomimetic anisotropic structure is constructed layer by layer according to a preset path: PLCL serves as a structural scaffold, providing stable support that matches the mechanical properties of natural myocardium; GelMA hydrogel loaded with cardiomyocytes constitutes the functional core layer of the patch, effectively promoting myocardial regeneration and vascularization; and gelatin hydrogel acts as a heat buffer protective layer, absorbing the heat released by the high-temperature PLCL during the printing process, significantly reducing the risk of thermal damage to the cells in the GelMA, thereby achieving precise integration and functional synergy of the three materials.

[0095] S3.1. First Layer Construction: The first layer is constructed according to the set printing parameters. First, PLCL material at 155℃ is extruded onto the printing platform to form a parallel filament array arranged in a single direction. The spacing between each PLCL filament is set to 1mm, the filament gap ratio is 75%, and the corresponding PLCL filament volume ratio is set to 1. Subsequently, GelMA hydrogel containing cardiomyocytes is precisely printed in the middle of the PLCL filaments, with a volume ratio of 1. During the printing process of this layer, it is ensured that the GelMA hydrogel does not directly contact the PLCL filaments, thus forming the functional core layer of the patch.

[0096] S3.2. Second Layer Construction: Above the GelMA hydrogel, a circular gelatin hydrogel layer with a diameter of 0.25 mm is printed along its trajectory as a heat buffer protective layer. Then, molten PLCL material is printed along the axial direction perpendicular to the PLCL printed in S3.1, forming a parallel filament array arranged in a single direction, constructing an orthogonal mesh structure to form the scaffold portion of the patch. During this process, the high-temperature PLCL rapidly transfers heat upon contact with the gelatin hydrogel, causing localized liquefaction of the gelatin and effectively absorbing heat. Finally, GelMA hydrogel containing cardiomyocytes is printed again in the gaps between the newly printed PLCL filaments, completing the integration of the three materials in the second layer.

[0097] S3.3. Repeat steps S3.1~S3.2 to achieve subsequent layer stacking: Following the construction process and conditions of the second layer described above, perform the operation on top of the second layer (at this time, the second layer becomes the lower layer). Print the PLCL scaffold, GelMA hydrogel functional filling, and gelatin hydrogel thermal buffer layer in sequence, alternating layers to achieve precise arrangement and functional synergy of PLCL, GelMA, and gelatin hydrogel in three-dimensional space. A total of 4 layers of the patch are printed, requiring 38 minutes of printing time. The preparation time for the cell hydrogel is 10 minutes, and the entire process takes less than 50 minutes.

[0098] S4. Post-processing: The printed structure was irradiated under 365nm ultraviolet light for 30s to allow the GelMA hydrogel to fully photocrosslink, forming a stable three-dimensional network structure and ensuring the structural integrity of the patch and the stability of the cellular microenvironment. By precisely controlling the distribution of the three materials in three-dimensional space, not only was the uniformity of the structure and the interlayer bonding strength guaranteed, but also the organic unity of mechanical adaptation, high cell activity, and biomimetic microstructure was achieved, successfully preparing a novel multifunctional integrated myocardial injury repair patch.

[0099] The microstructure of the novel myocardial injury repair patch was characterized, and the results are as follows: Figure 13As shown, optical microscopic images clearly reveal the spatial distribution and interfacial structure of three materials: PLCL, GelMA hydrogel, and gelatin hydrogel. The gelatin hydrogel incorporates biodegradable green fluorescent nanoparticles, enabling high-contrast imaging under microscopic observation and effectively distinguishing regions of different bio-inks. The gray structure represents PLCL material, forming the orthogonal mesh scaffold of the patch; the cell-carrying GelMA hydrogel is printed within the gaps of the PLCL filaments, exhibiting a uniform and transparent state. The image shows distinct and tightly bonded interfaces among the three materials, indicating that the multi-material co-printing process possesses excellent spatial accuracy and interfacial fusion. The fluorescent nanoparticles used exhibit excellent biocompatibility, having no significant impact on the activity of embedded cells, thus ensuring the functional integrity of the patch while achieving non-destructive visualization.

[0100] Examples 2-4:

[0101] The process conditions for Examples 2 to 4 are the same as those for Example 1, except that the PLCL printing temperatures are set to 140℃, 145℃, and 150℃ respectively.

[0102] Example 5:

[0103] This embodiment provides a method for preparing a myocardial repair patch based on multi-material printing, including the following steps:

[0104] S1. Material Preparation: Using poly(lactide-caprolactone) copolymer PLCL (lactide to caprolactone molar ratio of 50:50), methacrylamide gelatin (GelMA), and gelatin hydrogel as raw materials, the raw materials are pretreated as follows:

[0105] S1.1. Methacrylamide gelatin (GelMA) loaded with cardiomyocytes was dissolved in phosphate-buffered saline (PBS) to prepare a 0.05 g / mL solution. The pH of the solution was adjusted to pH=7.0; using 0.005 g / mL of phenyl (2,4,6-trimethylbenzoyl) lithium phosphate as a photoinitiator, the GelMA hydrogel was obtained by adding the photoinitiator to the solution.

[0106] S1.2. Dissolve the gelatin hydrogel in phosphate-buffered saline (PBS) to prepare a 0.1 g / mL gelatin hydrogel solution, which serves as a local cooling and protective layer. During the printing process, it can effectively absorb the heat released during the extrusion of high-temperature PLCL material, significantly reducing the impact of heat conduction on the adjacent cell-carrying GelMA hydrogel, and exhibits good thermal buffering capacity, biocompatibility, and biodegradability.

[0107] S2. Printing Parameter Settings and Import:

[0108] The process employs extrusion-based bio-3D printing technology and utilizes the commercial bio-3D printer SIA BioPrinter Pro. Specifically, a pre-designed STL format 3D model of a myocardial repair patch is imported (print trajectory parameter programming is shown in the image). Figure 1 ), and set the following key printing parameters:

[0109] Printing speed: 6mm / s; Semi-major axis of elliptical patch: 10mm; Semi-minor axis of elliptical patch: 6mm; Patch thickness: 1mm; Printing thickness per layer: 0.25mm;

[0110] PLCL printing parameter settings: Nozzle diameter: 0.25mm; filament spool distance: 1mm; mesh porosity: 66%; volume ratio: 1.36; printing temperature: 155℃.

[0111] GelMA hydrogel printing parameter settings: Nozzle diameter: 0.25mm; Filament pitch: 1mm; Volume ratio: 1; Printing temperature: 18℃;

[0112] Gelatin hydrogel printing parameter settings: Nozzle diameter: 0.25mm; filament pitch: 1mm; volume ratio: 1; printing temperature: 24℃.

[0113] S3. Multi-material collaborative printing:

[0114] S3.1. First Layer Construction: The first layer is constructed according to the set printing parameters. First, PLCL material at 155℃ is extruded onto the printing platform to form a parallel filament array arranged in a single direction. The spacing between each PLCL filament is set to 1mm, the filament gap ratio is 66%, and the corresponding PLCL filament volume ratio is set to 1.36. Subsequently, GelMA hydrogel containing cardiomyocytes is precisely printed in the middle of the PLCL filaments, with a volume ratio of 1. During the printing process of this layer, it is ensured that the GelMA hydrogel does not directly contact the PLCL filaments, thus forming the functional core layer of the patch.

[0115] S3.2. Second Layer Construction: Above the GelMA hydrogel, a circular gelatin hydrogel layer with a diameter of 0.25 mm is printed along its trajectory as a heat buffer protective layer. Then, molten PLCL material is printed along the axial direction perpendicular to the PLCL printed in S3.1, forming a parallel filament array arranged in a single direction, constructing an orthogonal mesh structure to form the scaffold portion of the patch. During this process, the high-temperature PLCL rapidly transfers heat upon contact with the gelatin hydrogel, causing localized liquefaction of the gelatin and effectively absorbing heat. Finally, GelMA hydrogel containing cardiomyocytes is printed again in the gaps between the newly printed PLCL filaments, completing the integration of the three materials in the second layer.

[0116] S3.3. Repeat steps S3.1~S3.2 to achieve subsequent layer stacking: Following the construction process and conditions of the second layer described above, perform the operation on top of the second layer (at this time, the second layer becomes the lower layer). Print the PLCL scaffold, GelMA hydrogel functional filling, and gelatin hydrogel thermal buffer layer in sequence, alternating layers to achieve precise arrangement and functional synergy of PLCL, GelMA, and gelatin hydrogel in three-dimensional space. A total of 4 layers of the patch are printed, requiring 38 minutes of printing time. The preparation time for the cell hydrogel is 10 minutes, and the entire process takes less than 50 minutes.

[0117] S4. Post-processing: The printed structure was irradiated under 365nm ultraviolet light for 30s to allow the GelMA hydrogel to fully photocrosslink, forming a stable three-dimensional network structure and ensuring the structural integrity of the patch and the stability of the cellular microenvironment. By precisely controlling the distribution of the three materials in three-dimensional space, not only was the uniformity of the structure and the interlayer bonding strength guaranteed, but also the organic unity of mechanical adaptation, high cell activity, and biomimetic microstructure was achieved, successfully preparing a novel multifunctional integrated myocardial injury repair patch.

[0118] Examples 6-8:

[0119] The process conditions for Examples 6-8 are the same as those for Example 5, except that the PLCL printing temperatures are set to 140℃, 145℃, and 150℃ respectively.

[0120] Example 9:

[0121] This comparative example provides a method for preparing a myocardial repair patch based on multi-material printing, including the following steps:

[0122] S1. Material preparation: Poly(lactide-caprolactone) copolymer PLCL with a weight average molecular weight of 200 kDa (lactide to caprolactone molar ratio of 50:50) is used as raw material.

[0123] S2. Printing Parameter Settings and Import:

[0124] The process employs extrusion-based bio-3D printing technology and utilizes the commercial bio-3D printer SIA BioPrinter Pro. Specifically, a pre-designed STL format 3D model of a myocardial repair patch is imported (print trajectory parameter programming is shown in the image). Figure 1 ), and set the following key printing parameters:

[0125] Printing speed: 6mm / s; Semi-major axis of elliptical patch: 10mm; Semi-minor axis of elliptical patch: 6mm; Patch thickness: 1mm; Printing thickness per layer: 0.25mm;

[0126] PLCL printing parameter settings: Nozzle diameter: 0.25mm; filament shaft distance: 1mm; mesh porosity: 75%; volume ratio: 1; printing temperature: 155℃; When printing PLCL material, keep the PLCL filament shaft distance and nozzle diameter constant, and control the printing material diameter by changing the volume ratio parameter to achieve the printing diameter requirement of being larger than the nozzle diameter.

[0127] S3. Bio-3D Printing:

[0128] Using bio-3D printing technology, a composite heart patch with a biomimetic anisotropic structure is constructed layer by layer according to a pre-defined path:

[0129] S3.1. First Layer Construction: The first layer is constructed according to the set printing parameters. First, PLCL material at 155℃ is extruded onto the printing platform, and the PLCL material is printed into two adjacent PLCL filaments according to the set printing parameters. Then, a GelMA hydrogel containing cardiomyocytes is precisely printed in the middle of the PLCL filaments, with a volume ratio of 1. During the printing process, it is ensured that the GelMA hydrogel does not come into direct contact with the PLCL filaments.

[0130] S3.2. Second Layer Construction: Above the GelMA hydrogel, a circular gelatin hydrogel layer with a diameter of 0.25 mm is printed along its trajectory as a heat buffer protective layer. Then, molten PLCL material is printed along the axial direction perpendicular to the PLCL printed in S3.1, forming a parallel filament array arranged in a single direction, constructing an orthogonal mesh structure to form the scaffold portion of the patch. During this process, the high-temperature PLCL rapidly transfers heat upon contact with the gelatin hydrogel, causing localized liquefaction of the gelatin and effectively absorbing heat. Finally, GelMA hydrogel containing cardiomyocytes is printed again in the gaps between the newly printed PLCL filaments, completing the integration of the three materials in the second layer.

[0131] S4. Post-processing: The printed structure was irradiated under 365nm ultraviolet light for 30s to allow the GelMA hydrogel to fully photocrosslink, forming a stable three-dimensional network structure and ensuring the structural integrity of the patch and the stability of the cellular microenvironment. By precisely controlling the distribution of the three materials in three-dimensional space, not only was the uniformity of the structure and the interlayer bonding strength guaranteed, but also the organic unity of mechanical adaptation, high cell activity, and biomimetic microstructure was achieved, successfully preparing a novel multifunctional integrated myocardial injury repair patch.

[0132] Comparative Example 1:

[0133] This comparative example provides a method for preparing a myocardial repair patch based on multi-material printing. The process flow is the same as in Example 9, except that: in the second layer construction process of S3.2, the gelatin hydrogel layer is not printed.

[0134] Comparative Example 2:

[0135] This comparative example provides a method for preparing a myocardial repair patch based on multi-material printing. The process flow is the same as in Example 1, except that the GelMA used is not loaded with cardiomyocytes.

[0136] To further quantify the mechanical properties, this invention uses a texture analyzer (CT3 Texture Analyzer, Brookfield) to perform uniaxial tensile tests on the patches prepared in Examples 1-8 (strain range: 0-20%; tensile rate: 0.5 mm / s; trigger load: 0.008 N), and calculates the elastic modulus based on the stress-strain curves. Figure 3 It can be seen that, under 20% strain conditions, both types of patches exhibited the lowest stress and minimum elastic modulus when printed at 155℃. This is because the molecular chains move more fully at high temperatures, and the crystallization rate slows down and the crystallinity decreases during cooling, thus weakening the material's rigidity. It is worth noting that excessively high elastic modulus may restrict the natural beating of the myocardium and interfere with physiological function; while excessively low stress, although improving flexibility, is detrimental to intraoperative suturing and fixation procedures. Figure 4 ).

[0137] To screen for the printing parameter combination whose mechanical properties most closely resemble those of natural myocardium, further testing was conducted on the ventricular surface tissue of an adult Bama boar (approximately 4 cm² in area). 2 The mechanical properties of ) were determined. Its elastic modulus was measured under the same test conditions (20% strain). Figure 5 The PLCL patches were compared with those prepared in Examples 1-8 at different printing temperatures (140℃~155℃) and pore densities (66%, 75%) to evaluate their mechanical compatibility and physiological fit after implantation. The final results showed that, under the conditions of printing at 155℃ and 75% mesh pore density, the elastic modulus of the PLCL patch was highly compatible with that of Bama pig ventricular tissue. Figure 6 It exhibits excellent biomechanical compatibility, providing key process basis for subsequent in vivo applications.

[0138] To assess the risk of thermal damage to the cell-carrying GelMA hydrogel during high-temperature PLCL printing and to verify the effectiveness of the local cooling technique proposed in this invention, this invention applies the patch (printed structure as shown in Example 9 and Comparative Example 1) prepared in Example 9 to the patch. Figure 8 (As shown) A systematic comparative study was conducted combining numerical simulation and experimental measurement:

[0139] Simulation modeling and thermal imaging verification: Under conditions without a protective layer, finite element simulation shows that the interface temperature at the direct contact between PLCL and GelMA hydrogel reaches as high as 82℃. Figure 9The highest temperature measured using a FLIR A615 thermal imager from the United States reached 86℃. Figure 10 By introducing a 0.25 mm diameter gelatin hydrogel as a thermal buffer layer, the high-temperature PLCL first contacts the upper gelatin layer. Through thermal conduction, some heat is transferred to the gelatin, causing it to locally liquefy and absorb a large amount of heat, thus significantly reducing its own temperature. Subsequently, the cooled PLCL contacts the lower GelMA hydrogel layer. Simulation results show that the interface temperature drops to 39℃, while the measured value from thermal imaging further decreases to 37℃, achieving a temperature drop of approximately 49℃. The thermal imaging measurement and finite element simulation results are in high agreement, fully verifying the reliability of this local cooling mechanism.

[0140] Cell viability assessment: The printed cell-loaded heart patch was sliced, and the locations of PLCL, cell-loaded GelMA, and the protective layer were marked on the slices. After staining for live / dead cells, the cells were observed using a two-photon microscope: red indicates dead cells, and green indicates live cells. Figure 11 Almost all cells in the unprotected patch died; however, with the introduction of the protective layer, the number of viable cells increased significantly, with a cell survival rate exceeding 91%, demonstrating that the gelatin protective layer effectively mitigated heat damage during the printing process and provided significant protection for the cells. According to fluorescence microscopy images, after one day of culture, the cells were in a normal state with a survival rate of 90%. After three days of culture, the cell number increased, with a survival rate of 95%, indicating that the cells not only survived but also continued to proliferate. By day 7, the cell number had further increased, and the survival rate reached 98%. Figure 14 This result indicates that, over time, cells adapted well within the patch, accelerating proliferation and differentiation, achieving normal growth and functional expression. The high temperature at the interface between PLCL and GelMA hydrogel not only caused rapid liquefaction and collapse of the GelMA hydrogel structure but also exposed the embedded cardiomyocytes to a severely thermally damaged environment, resulting in a significant decrease in survival rate. In contrast, the gelatin hydrogel, acting as a thermal buffer layer, significantly inhibited heat transfer to the cell-bearing GelMA layer, effectively ensuring cell survival and functional integrity. Live / dead cell staining combined with two-photon microscopy showed that almost all cells in the unprotected group died (predominantly exhibiting red fluorescence); however, after introducing the gelatin protective layer, the proportion of live cells (green fluorescence) significantly increased, with a cell survival rate exceeding 91%.

[0141] This invention, through triple verification of simulation modeling, thermal imaging measurement, and cell activity assessment, confirms that local cooling technology of gelatin hydrogel can effectively alleviate the thermal damage problem in the process of multi-material bio-3D printing, and provides key process guarantee for constructing myocardial repair patches with high cell activity and integrated structure and function.

[0142] Example 10:

[0143] This embodiment provides an application of a myocardial repair patch based on multi-material printing, wherein the myocardial repair patch used is the product obtained in Example 1, and includes the following steps:

[0144] A left ventricular rupture model was established using healthy Bama pigs (weighing 30-40 kg and approximately 1 year old).

[0145] First, the Bama pig was secured to a large animal operating table and administered general anesthesia via oral injection. Preoperative ultrasound was performed to record basic vital signs, and the procedure was conducted by a professional anesthesia and surgical team, equipped with complete surgical equipment, including a shadowless lamp, anesthesia machine, ventilator, suction device, and related surgical instruments. After routine disinfection of the surgical area, an open-chest surgery was performed to expose the heart, and a cardiac fixator was used to stabilize the left ventricle. Subsequently, a standard 15mm rupture wound was created in the left ventricular wall using a scalpel for subsequent implantation of the repair patch and functional assessment.

[0146] Next, the printed myocardial injury repair patch ( Figure 12 The patch is gently applied to the surface of the heart, ensuring that the functional layer of GelMA containing cardiomyocytes faces the myocardial tissue. Upon contact with blood, the GelMA rapidly absorbs fluid and expands, creating a localized negative pressure on the epicardial surface. This blood infiltration and negative pressure promote a tight fit between the patch and the heart's curvature. Before suturing, the patch is flat; after suturing, it naturally conforms to the curvature of the myocardium, achieving excellent mechanical fit. During cardiac contractions, the patch stretches and retracts synchronously with the myocardium, without displacement, leakage, or peeling.

[0147] The PLCL scaffold layer of the patch has a porosity of 75%, and the mesh pore size is slightly larger than that of conventional suture needles. This facilitates the passage of sutures through the mesh gaps, avoiding direct penetration of the material itself and significantly reducing damage to the patch structure and secondary trauma to the myocardial tissue during the suturing process. At the same time, the cell-loaded GelMA layer effectively supports cell activity and promotes wound repair and tissue regeneration.

[0148] In Example 1 of this invention, the PLCL printing temperature was 155°C and the mesh porosity was 75% when the patch was prepared. The elastic modulus of the patch was closest to that of the surface tissue of the ventricle of Bama pigs, which provided sufficient mechanical support to resist the stress of the ventricle wall and avoided the stress shielding effect caused by excessive stiffness.

[0149] Postoperative observation showed that the patch adhered stably to the surface of the heart with good dynamic fit, and no obvious inflammatory reaction, bleeding or functional impairment was observed.

[0150] Comparative Example 3:

[0151] In this comparative example, the Bama pigs used to establish a left ventricular rupture model underwent only suturing at the injury site in the same surgical environment as in Example 10.

[0152] Comparative Example 4:

[0153] This comparative example provides an application of a myocardial repair patch based on multi-material printing. The process flow is the same as that of Example 10, except that the myocardial repair patch used is the product obtained in Comparative Example 2.

[0154] To further observe the recovery of different damage repair methods, the conditions of Examples 10, 3, and 4 were observed one month after repair. The results are as follows: Figure 15 As shown in the figure. The vertical axis represents the percentage of TUNEL-positive cells (a higher TUNEL positivity rate indicates more severe apoptosis), and the horizontal axis represents the three experimental groups mentioned above. (P < 0.001). The results showed that the MA group (control group with only sutures after injury, Comparative Example 3), the MB group (composite patch without cardiomyocytes implanted in Comparative Example 4), and the MC group (composite patch loaded with active cardiomyocytes implanted in Example 10) all alleviated myocardial fibrosis to some extent, but the MC group showed the most significant repair effect. Quantitative analysis of the apoptosis level in the myocardial tissue of each group was performed. The apoptosis rate in the MA group was approximately 4.5%, and in the MB group it was approximately 4.0%, with no significant difference between the two. However, the apoptosis rate in the MC group was significantly reduced (P < 0.001) to approximately 0.9%. These statistical differences fully demonstrate that the repair patch loaded with active cardiomyocytes has outstanding advantages in inhibiting cardiomyocyte apoptosis, alleviating fibrosis, and promoting tissue regeneration, significantly superior to simple sutures or cell-free scaffolds, fully demonstrating its therapeutic potential in functional myocardial regeneration.

Claims

1. A method for preparing a myocardial repair patch based on multi-material printing, characterized in that, Includes the following steps: S1. Material preparation: Poly(lactide-caprolactone) copolymer PLCL, methacrylamide gelatin GelMA, and gelatin hydrogel are used as raw materials, and the raw materials are pretreated. S1.

1. Methacrylamide gelatin (GelMA) loaded with cardiomyocytes was dissolved in phosphate buffer, the pH of the solution was adjusted, and a photoinitiator was added to obtain GelMA hydrogel; S1.

2. Dissolve the gelatin gel in phosphate buffer to prepare a gelatin hydrogel solution; S2. Printing Parameter Setting and Import: Using extrusion-type bio-3D printing technology, a pre-designed three-dimensional model of myocardial repair patch is imported into the bio-3D printer, and printing parameters are set; among them, the printing temperature of PLCL is 140℃~155℃; S3. Multi-material collaborative printing: S3.

1. First layer construction: Print a parallel filament array arranged in a single direction, and then print a GelMA hydrogel containing cardiomyocytes in the gaps between the PLCL filaments to form the first layer; S3.

2. Second layer construction: On top of the GelMA hydrogel, print a layer of gelatin hydrogel along its trajectory as a heat buffer protective layer; Then, PLCL material is printed along the axial direction perpendicular to the lower PLCL to form a parallel filament array arranged in a single direction, constructing an orthogonal mesh structure to form the scaffold part of the patch; finally, GelMA hydrogel containing cardiomyocytes is printed again in the gaps between the newly printed PLCL filaments to complete the integration of the three materials in the second layer. S3.

3. Repeat steps S3.1 to S3.2 to obtain the printed patch structure; S4. Post-processing: Irradiate the printed structure under ultraviolet light to obtain a myocardial injury repair patch.

2. The method for preparing a myocardial repair patch based on multi-material printing according to claim 1, characterized in that, In S2, during the printing parameter setting process, the printing speed is 1mm / s-10mm / s; The volume ratios of PLCL, GelMA hydrogel, and gelatin hydrogel are set independently, with values ​​ranging from 1 to 2. The printing spacing and nozzle diameter are set according to the type of raw material and the target patch specifications; the printing thickness of each layer is set according to the patch requirements, and the thickness is adapted to the nozzle specifications.

3. A method for preparing a myocardial repair patch based on multi-material printing according to claim 1 or 2, characterized in that, In S1, the weight-average molecular weight of PLCL is 200 kDa, and the molar ratio of lactide to caprolactone in PLCL is 50:

50.

4. A method for preparing a myocardial repair patch based on multi-material printing according to claim 1 or 2, characterized in that, In S1.1, GelMA is dissolved in phosphate buffer to a concentration of 0.05 g / mL; the photoinitiator is lithium phenyl (2,4,6-trimethylbenzoyl) phosphate; the pH of the solution is adjusted to 7.0 before adding the photoinitiator.

5. A method for preparing a myocardial repair patch based on multi-material printing according to claim 1 or 2, characterized in that, In S1.2, the concentration of the gelatin gel solution is 0.1 g / mL.

6. A method for preparing a myocardial repair patch based on multi-material printing according to claim 1 or 2, characterized in that, In S3, the total printing time, starting from the start of printing, shall not exceed 50 minutes.

7. A method for preparing a myocardial repair patch based on multi-material printing according to claim 1 or 2, characterized in that, In S3.3, repeat steps 3.1 to 3.2 until the patch thickness reaches the target number of layers.

8. A method for preparing a myocardial repair patch based on multi-material printing according to claim 1 or 2, characterized in that, In S4, the ultraviolet light wavelength is 365nm, and the irradiation time is 25s~55s.

9. A myocardial repair patch prepared by the method described in claim 1 or 2.

10. An application of a myocardial repair patch prepared by the method of claim 1 or 2.

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