3D printing bio-ink as well as preparation method and application thereof

Abstract

The invention provides 3D printing bio-ink as well as a preparation method and application thereof. The 3D printing bio-ink is prepared from the following components: methacrylic anhydride gelatin, oxidized hyaluronic acid, carboxymethyl chitosan and 2-methacryloyloxyethyl phosphorylcholine. By designing the specific composition of the 3D printing bio-ink, the 3D printing bio-ink which is good in printing performance, mechanical strength and electrical conductivity and high in cell survival rate after printing is prepared, and then a 3D printing stent with high cell-stent interaction is prepared and can be used for preparing a skin injury repair material.

Description

Technical Field

[0001] The present invention belongs to the technical field of medical materials, and specifically relates to a 3D printing biological ink and a preparation method and application thereof. Background Art

[0002] Skin defects caused by external injury or disease can lead to fluid loss, bacterial infection, and secondary complications. According to WHO data, nearly 11 million burn patients are diagnosed annually, and over 6 million suffer from chronic skin ulcers. Chronic skin wounds are unable to progress beyond the inflammatory phase, hindering the proliferation of endothelial cells and fibroblasts and the deposition of collagen matrix, leading to long-term inflammation, angiogenesis deficiency, and cancer. Long-term, unhealed chronic wounds following trauma and burns not only severely impact patient comfort but also impose a significant economic burden.

[0003] Currently, common treatment options for chronic wounds include skin grafting, cytokine therapy, and various wound dressings. However, these methods are limited in the number of transplant donors, are costly, and can lead to contraction and scarring, resulting in very low survival rates after transplantation. Hydrogel dressings are widely used wound dressings that cover wounds, provide a moist environment, promote wound healing, and prevent secondary infection. However, they cannot accurately mimic the complex three-dimensional structure of skin tissue and cannot effectively promote the regeneration of vascular networks in damaged skin. In recent years, the use of 3D bioprinting technology to construct structurally precise and controllable three-dimensional scaffolds has become a research hotspot. 3D bioprinting is a novel additive manufacturing technology that combines biomaterials (such as hydrogels) and biological units (such as cells, DNA, and proteins) to create personalized biofunctional structures using 3D printing, mimicking morphology, biological functions, and cell growth microenvironments. Compared to traditional cell-free 3D printing methods, cell-based 3D bioprinting allows for precise control of cell density and distribution, creating bioactive scaffolds that mimic the structure and function of natural tissue. Research has shown that human skin possesses certain conductive properties. Conductive hydrogels can stimulate cell migration, enhance re-epithelialization, promote dermal angiogenesis, and help restore epithelial potential after skin damage. However, the types of conductive hydrogels currently available for 3D bioprinting are very limited, and they commonly suffer from issues such as poor printability, weak mechanical strength, poor electrical conductivity, low post-printing cell survival, and weak cell-scaffold interactions.

[0004] Therefore, how to provide a new type of 3D printing bio-ink with excellent performance (good printing performance, good mechanical strength, good electrical conductivity, and high cell survival rate after printing), and then use this 3D printing bio-ink to prepare 3D printed scaffolds with strong cell-scaffold interaction, has become a technical problem that needs to be solved urgently. Summary of the Invention

[0005] To address the shortcomings of the existing technology, the present invention aims to provide a 3D printing bio-ink, its preparation method, and its application. By designing the specific composition of the 3D printing bio-ink, the present invention produces a 3D printing bio-ink with excellent printing performance, mechanical strength, electrical conductivity, and high cell survival rate after printing. Furthermore, a 3D printed scaffold with strong cell-scaffold interaction is produced, which can be used to prepare skin damage repair materials.

[0006] To achieve this object, the present invention adopts the following technical solutions:

[0007] In a first aspect, the present invention provides a 3D printing bio-ink, wherein the 3D printing bio-ink comprises the following components: methacrylic anhydride gelatin, oxidized hyaluronic acid, carboxymethyl chitosan, and 2-methacryloyloxyethyl phosphorylcholine.

[0008] The present invention designs the specific composition of 3D printing bio-ink and further uses 2-methacryloyloxyethyl phosphorylcholine to make the 3D printing bio-ink have electrical conductivity, thereby preparing a 3D printing bio-ink with good printing performance, good mechanical strength, good electrical conductivity, and high cell survival rate after printing. Furthermore, a 3D printing scaffold with strong cell-scaffold interaction is prepared, which can be used to prepare skin damage repair materials.

[0009] The 3D printing bio-ink system provided by the present invention can quickly solidify into a gel at 20-37°C. The Schiff base reaction between oxidized hyaluronic acid and carboxymethyl chitosan is used to make the 3D scaffold structure more stable and less prone to collapse. At the same time, the zwitterionic 2-methacryloyloxyethyl phosphorylcholine and methacrylic anhydride gelatin are copolymerized under light, giving the hydrogel the ability to conduct electricity.

[0010] The 3D printing bio-ink provided by the present invention can be used to prepare a hydrogel system with a double network structure, which can improve the mechanical strength of the conductive hydrogel, enhance the conductivity, and optimize the printability. In addition, the use of the zwitterion 2-methacryloyloxyethyl phosphorylcholine is more in line with the human body's electrical signal conduction mode and can achieve strong interactions between cells.

[0011] The 3D printing bio-ink provided by the present invention is suitable for cell-laden biological 3D printing. The mechanical strength of the obtained hydrogel scaffold can match that of skin tissue, promoting the repair of skin damage.

[0012] The following are preferred technical solutions of the present invention, but are not intended to limit the technical solutions provided by the present invention. Through the following preferred technical solutions, the objectives and beneficial effects of the present invention can be better achieved and realized.

[0013] As a preferred technical solution of the present invention, the 3D printing bio-ink comprises the following components in parts by weight:

[0014]

[0015] In the present invention, the comprehensive performance of the 3D printing bio-ink can be further improved by further designing the dosage of each component in the 3D printing bio-ink.

[0016] In the present invention, the performance of 3D printing bio-inks can be further improved by controlling the dosage of 2-methacryloyloxyethyl phosphorylcholine within a specific range. If the dosage is too low, it will fail to stimulate cell proliferation and differentiation, and the conductivity will be too low. If the dosage is too high, it will be cytotoxic and cause cell damage.

[0017] In the present invention, the weight proportion of methacrylic anhydride gelatin in the 3D printing bio-ink can be 5 parts, 6 parts, 7 parts, 8 parts, 9 parts, 10 parts, 11 parts, 12 parts, 13 parts, 14 parts, 15 parts, 16 parts, 17 parts, 18 parts, 19 parts or 20 parts, etc.

[0018] The weight proportion of oxidized hyaluronic acid in the 3D printing bio-ink can be 0.5 parts, 1 parts, 1.2 parts, 1.5 parts, 2 parts, 2.5 parts, 3 parts, 3.5 parts, 4 parts, 4.5 parts or 5 parts, etc.

[0019] The weight proportion of carboxymethyl chitosan in the 3D printing bio-ink can be 0.5 parts, 0.8 parts, 1 part, 2 parts, 3 parts, 4 parts, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts or 10 parts, etc.

[0020] The weight proportions of 2-methacryloyloxyethyl phosphorylcholine in the 3D printing bio-ink can be 1 part, 2 parts, 3 parts, 4 parts, 5 parts, 6 parts, 7 parts, 8 parts, 9 parts or 10 parts, etc.

[0021] As a preferred technical solution of the present invention, the 3D printing bio-ink comprises the following components in parts by weight:

[0022] 7 parts of methacrylic anhydride gelatin, 1.2 parts of oxidized hyaluronic acid, 0.8 parts of carboxymethyl chitosan and 2 parts of 2-methacryloyloxyethyl phosphorylcholine.

[0023] In the present invention, the 3D printing bio-ink prepared by further selecting specific amounts of each component has good printing performance, mechanical strength and electrical conductivity, and the cell survival rate after printing is high.

[0024] As a preferred technical solution of the present invention, the raw materials for preparing the methacrylic anhydride gelatin include gelatin and methacrylic anhydride.

[0025] Preferably, the mass ratio of gelatin to methacrylic anhydride is 1:(0.2-1), for example, it can be 1:0.2, 1:0.3, 1:0.4, 1:0.5, 1:0.6, 1:0.7, 1:0.8, 1:0.9 or 1:1, etc.

[0026] It should be noted that the present invention does not have any special requirements for the weight-average molecular weight of gelatin, and all gelatins commonly used in the art are applicable, including but not limited to: 10,000 to 70,000, for example, it can be 10,000, 15,000, 20,000, 25,000, 30,000, 35,000, 40,000, 45,000, 50,000, 55,000, 65,000 or 70,000, etc.

[0027] Preferably, the weight average molecular weight of the methacrylic anhydride gelatin is ≥14,000, for example, it can be 14,000, 15,000, 16,000, 17,000, 14,000, 18,000, 19,000, 20,000, 21,000, 22,000, 23,000 or 24,000.

[0028] In the present invention, the methacrylic anhydride gelatin can be a commercially available product or can be homemade. The present invention does not impose any particular restrictions on the preparation method for methacrylic anhydride gelatin, and any commonly used preparation method in the art is applicable. The present invention provides a preparation method for methacrylic anhydride gelatin, which is as follows:

[0029]

[0030] 10 g of gelatin was dissolved in 100 mL of PBS buffer solution at 50°C to obtain a 10% (w / v) gelatin solution. 2-10 g of methacrylic anhydride was added with stirring, and the mixture was reacted at 50°C for 4 hours. The reaction solution was collected and dialyzed for 96 hours using a dialysis bag with a molecular weight cutoff of 14,000 to remove unreacted reactants. The dialyzate was adjusted to a neutral pH with 0.1 mol / L sodium hydroxide solution and freeze-dried at -60°C to obtain a white spongy solid methacrylic anhydride gelatin.

[0031] As a preferred technical solution of the present invention, the weight average molecular weight of the oxidized hyaluronic acid is ≥14,000, for example, it can be 14,000, 15,000, 16,000, 17,000, 14,000, 18,000, 19,000, 20,000, 21,000, 22,000, 23,000 or 24,000.

[0032] It should also be noted that, in the present invention, the oxidized hyaluronic acid can be a commercially available product or can be homemade. There are no special restrictions on the preparation method of the oxidized hyaluronic acid in the present invention, and the preparation methods commonly used in the art are applicable. The present invention provides a preparation method of oxidized hyaluronic acid, and the preparation method is as follows:

[0033]

[0034] 5 g of hyaluronic acid powder was dissolved in 100 mL of deionized water, and 0.321 g of sodium periodate powder was dissolved in 5 mL of deionized water to obtain a sodium periodate solution. 5 mL of the sodium periodate solution was added to 100 mL of the hyaluronic acid solution, and the mixture was stirred in the dark at 20°C for 1 h. After that, 100 μL of glycerol was added dropwise, and the reaction was terminated by stirring in the dark for 3 h. The mixture was dialyzed for 96 h using a dialysis bag with a molecular weight cutoff of 14,000. The pH value of the dialysate was then adjusted to neutral with 0.1 mol / L sodium hydroxide solution, and the mixture was freeze-dried at -60°C for 48 h to obtain oxidized hyaluronic acid.

[0035] As a preferred technical solution of the present invention, the 3D printing bio-ink also includes a solvent.

[0036] Preferably, the solvent comprises a phosphate buffer solution.

[0037] In the present invention, there is no special restriction on the amount of solvent used. For example, the weight parts of the solvent are 10 to 30 parts, for example, 10 parts, 12 parts, 14 parts, 16 parts, 18 parts, 20 parts, 22 parts, 24 parts, 26 parts, 28 parts or 30 parts, etc.

[0038] As a preferred technical solution of the present invention, the 3D printing bio-ink also includes a photoinitiator.

[0039] Preferably, the weight portion of the photoinitiator is 0.3 to 0.5 parts, for example, it can be 0.3 parts, 0.32 parts, 0.34 parts, 0.36 parts, 0.38 parts, 0.4 parts, 0.42 parts, 0.44 parts, 0.46 parts, 0.48 parts or 0.5 parts.

[0040] It should be noted that the present invention does not have any special restrictions on the specific type of photoinitiator, and all commonly used photoinitiators in the field are applicable, including but not limited to LAP (phenyl (2,4,6-trimethylbenzoyl) phosphate lithium salt) and MBF (methyl benzoylformate).

[0041] In a second aspect, the present invention provides a method for preparing the 3D printing bio-ink as described in the first aspect, the preparation method comprising the following steps:

[0042] Dissolve methacrylic anhydride gelatin, carboxymethyl chitosan, 2-methacryloyloxyethyl phosphorylcholine, part of the solvent and an optional photoinitiator to obtain a solution A;

[0043] mixing the oxidized hyaluronic acid and the remaining solvent to obtain solution B;

[0044] The solution A and the solution B are mixed evenly to obtain the 3D printing bio-ink.

[0045] In the present invention, the mixing temperature when preparing solution A is 37-50°C, for example, 37°C, 38°C, 39°C, 40°C, 42°C, 44°C, 46°C, 48°C or 50°C.

[0046] In a third aspect, the present invention provides a 3D printed scaffold, wherein the raw materials for preparing the 3D printed scaffold include the 3D printed biological ink and cells as described in the first aspect;

[0047] Preferably, the biological unit includes any one of cells, DNA or proteins, or a combination of at least two of them.

[0048] Preferably, the cells include any one or a combination of at least two of skin tissue cells, human umbilical vein endothelial cells and keratinocytes.

[0049] Preferably, the cell density in the mixed solution of the 3D printing bio-ink and the bio-unit is (1.5-2.5)×10 6 cells / mL, for example, 1.5×10 6 cells / mL、1.6×10 6 cells / mL、1.7×10 6 cells / mL、1.8×10 6 cells / mL、1.9×10 6 cells / mL, 2.0×10 6 cells / mL、2.1×10 6 cells / mL、2.2×10 6 cells / mL、2.3×10 6 cells / mL、2.4×10 6 cells / mL or 2.5×10 6 cells / mL, etc.

[0050] In a fourth aspect, the present invention provides an application of the 3D printing bio-ink as described in the first aspect or the 3D printing scaffold as described in the third aspect, wherein the application includes use in preparing skin damage repair materials.

[0051] Compared with the prior art, the present invention has the following beneficial effects:

[0052] The present invention designs the specific composition of 3D printing bio-ink and further uses 2-methacryloyloxyethyl phosphorylcholine to make the 3D printing bio-ink have electrical conductivity, thereby preparing a 3D printing bio-ink with good printing performance, good mechanical strength, good electrical conductivity, and high cell survival rate after printing. Furthermore, a 3D printing scaffold with strong cell-scaffold interaction is prepared, which can be used to prepare skin damage repair materials. The storage modulus is 918.2-1024.0Pa, the loss modulus is 77.18-111.26Pa, and the electrical conductivity is 0.026-0.280S·m -1 , cell survival rate ≥ 90%, specifically 90-98%. BRIEF DESCRIPTION OF THE DRAWINGS

[0053] Figure 1 A graph showing the test results of the storage modulus and loss modulus of the printed stent provided in Application Example 1 of the present invention;

[0054] Figure 2 Schematic diagram of live and dead staining of fibroblasts in a 3D printed scaffold for staining the 3D printed scaffold provided in Application Example 1 of the present invention;

[0055] Figure 3 A schematic diagram of vascularization of a 3D printed stent provided in Application Example 1 of the present invention;

[0056] Among them, blue is cell nucleus staining, green is vascular endothelial cell staining, and red is CD31 staining. DETAILED DESCRIPTION

[0057] The technical solution of the present invention will be further described below with reference to the accompanying drawings and through specific embodiments. It should be understood by those skilled in the art that the embodiments are merely to help understand the present invention and should not be regarded as specific limitations of the present invention.

[0058] The sources of some components in the following examples and comparative examples are as follows:

[0059] Methacrylic anhydride gelatin 1: 10 g of gelatin (purchased from Sigma V900863, the same below) was dissolved in 100 mL of PBS buffer at 50°C to obtain a 10% (w / v) gelatin solution. 3.1 g of methacrylic anhydride was added with stirring, and the mixture was reacted at 50°C for 4 h. The reaction solution was collected and dialyzed using a dialysis bag with a molecular weight cutoff of 14,000 for 96 h to remove unreacted reactants. The dialyzate was adjusted to a neutral pH with 0.1 mol / L sodium hydroxide solution and freeze-dried at -60°C to obtain a white spongy solid methacrylic anhydride gelatin.

[0060] Methacrylic anhydride gelatin 2: 10 g of gelatin was dissolved in 100 mL of PBS buffer solution at 50°C to obtain a 10% (w / v) gelatin solution. 6.2 g of methacrylic anhydride was added with stirring, and the mixture was reacted at 50°C for 4 h. The reaction solution was collected and dialyzed using a dialysis bag with a molecular weight cutoff of 14,000 for 96 h to remove unreacted reactants. The dialyzate was adjusted to a neutral pH with 0.1 mol / L sodium hydroxide solution and freeze-dried at -60°C to obtain a white spongy solid methacrylic anhydride gelatin.

[0061] Methacrylic anhydride gelatin 3: 10 g of gelatin was dissolved in 100 mL of PBS buffer solution at 50°C to obtain a 10% (w / v) gelatin solution. 9.3 g of methacrylic anhydride was added with stirring, and the mixture was reacted at 50°C for 4 h. The reaction solution was collected and dialyzed for 96 h using a dialysis bag with a molecular weight cutoff of 14,000 to remove unreacted reactants. The pH of the dialysate was adjusted to neutral with 0.1 mol / L sodium hydroxide solution and freeze-dried at -60°C to obtain a white spongy solid methacrylic anhydride gelatin.

[0062] Oxidized hyaluronic acid: dissolve 5 g of hyaluronic acid powder in 100 mL of deionized water, and dissolve 0.321 g of sodium periodate powder in 5 mL of deionized water to obtain a sodium periodate solution; add 5 mL of the sodium periodate solution to 100 mL of the hyaluronic acid solution, stir in the dark at 20°C for 1 h, then add 100 μL of glycerol dropwise, stir in the dark for 3 h to terminate the reaction, and dialyze using a dialysis bag with a molecular weight cutoff of 14,000 for 96 h. Then, adjust the pH of the dialysate to neutral with 0.1 mol / L sodium hydroxide solution, and freeze-dry at -60°C for 48 h to obtain oxidized hyaluronic acid.

[0063] Phosphate buffer solution: purchased from Sevier;

[0064] Photoinitiator: LAP, purchased from Anaiji Chemical, model number E062362;

[0065] Fibroblasts: purchased from Fenghui Biotechnology, model HFF-1.

[0066] Examples 1-8 and Comparative Example 1

[0067] Examples 1-8 and Comparative Example 1 respectively provide a 3D printing bio-ink, the specific composition of which is shown in Table 1 below. The amount of each component in Table 1 is in parts by weight.

[0068] The preparation method of the 3D printing bio-ink is as follows:

[0069] Methacrylic anhydride gelatin, carboxymethyl chitosan, 2-methacryloyloxyethyl phosphorylcholine and a photoinitiator were added to a portion of the phosphate buffer solution and stirred in a 37°C water bath until completely dissolved to obtain solution A.

[0070] Add oxidized hyaluronic acid to a portion of the phosphate buffer solution, stir and dissolve, to obtain solution B;

[0071] The solution A and the solution B are mixed evenly to obtain the 3D printing bio-ink.

[0072] Table 1

[0073]

[0074] Application Example 1

[0075] Application Example 1 provides a 3D printed scaffold and the same, wherein the raw materials for preparing the 3D printed scaffold include the 3D printed bio-ink provided in Example 1, fibroblasts, human umbilical vein endothelial cells and keratinocytes.

[0076] The preparation method of the 3D printed bracket is as follows:

[0077] The 3D printing bio-ink provided in Example 1 was divided into 10 equal parts, 3 of which were mixed evenly with fibroblasts (cell density was 2.0×10 6 cells / mL) to obtain precursor 1; 2 of them were mixed evenly with human umbilical vein endothelial cells (HUVECs) (cell density was 2.0×10 6 cells / mL) to obtain precursor 2; the remaining 5 parts were mixed evenly with keratinocytes (HaCaTs) (cell density was 2.0×10 6 cells / mL) to obtain precursor 3;

[0078] 3D printing was performed using a GeSim Bioscaffold 3.2 printer (manufacturer: GeSim). Precursor 1 and Precursor 2 were used to alternately print four layers (each layer was 0.2 mm thick) to simulate the dermis. Precursor 3 was used to 3D print two layers (each layer was 0.2 mm thick) to simulate the dermis, resulting in a 3D-printed circular double-layer scaffold with a diameter of 1.5 mm.

[0079] The results of testing the storage modulus and loss modulus of the 3D printed bracket provided in Application Example 1 of the present invention are as follows: Figure 1 As shown. Figure 1 It can be seen that the storage modulus (G') of the hydrogels is greater than the loss modulus (G"), showing a viscoelastic behavior dominated by elasticity, indicating the formation of a stable gel.

[0080] Calcein-AM (0.5uL / 1mL culture medium) and propidium iodide (PI) (0.5uL / 1mL culture medium) were used to stain fibroblasts for live and dead cells. Live cells are green. Figure 2 It can be seen that biological 3D printing with bio-ink mixed with cells will not have a negative impact on cells, the cell survival rate is high, and the cells can proliferate normally.

[0081] Immunofluorescence staining was used to characterize the vascularization of the printed skin scaffolds. Figure 3 In the figure, blue is the staining of cell nuclei, green is the staining of vascular endothelial cells, and red is the staining of CD31, which is a marker protein for new blood vessels. Figure 3 It can be seen that bio-ink mixed cell printing can complete vascularization, which is helpful for subsequent damage repair research.

[0082] Application Examples 2-8 and Comparative Application Example 1

[0083] Application Examples 2-8 and Comparative Application Example 1 each provide a 3D printed stent. The only difference from Application Example 1 is that the 3D printed bio-ink provided in Example 1 used in Application Example 1 is replaced with the 3D printed bio-ink provided in Examples 2-8 and Comparative Example 1, respectively. Other conditions are the same as those in Application Example 1.

[0084] The performance of the 3D printing bio-ink provided in the above embodiments and comparative examples, and the 3D printing scaffold provided in the above application examples and comparative application examples were characterized. The specific testing methods are as follows:

[0085] Printing performance: Structural analysis of the printed mesh support;

[0086] Storage modulus and loss modulus: A rotational rheometer was used to test the storage modulus and loss modulus of the 3D printed scaffold.

[0087] Electrical conductivity: The electrical conductivity of the 3D printed bracket was tested using a Keithley 2400 field source instrument.

[0088] Cell survival rate in 3D printed scaffolds: After live-dead staining of cells, the number of live and dead cells was counted using ImageJ, and the survival rate was calculated.

[0089] The above performance test results are detailed in Table 2 below.

[0090] Table 2

[0091]

[0092] From the above content, it can be seen that the present invention designs the specific composition of 3D printing bio-ink and further uses 2-methacryloyloxyethyl phosphorylcholine to make the 3D printing bio-ink have electrical conductivity, thereby preparing a 3D printing bio-ink with good printing performance, good mechanical strength, good electrical conductivity, and high cell survival rate after printing. Furthermore, a 3D printing scaffold with strong cell-scaffold interaction is prepared, which can be used to prepare skin damage repair materials. Its storage modulus is 918.2~1024.0Pa, the loss modulus is 77.18~111.26Pa, and the electrical conductivity is 0.026~0.280S·m -1 , cell survival rate ≥ 90%, specifically 90-98%.

[0093] From the data of Examples 1 and 5-8, it can be seen that the present invention can further improve the comprehensive performance of 3D printing bio-ink by controlling the amount of 2-methacryloyloxyethyl phosphorylcholine in the 3D printing bio-ink within a specific range.

[0094] If 2-methacryloyloxyethyl phosphorylcholine is not used in the 3D printing bio-ink (Comparative Example 1), the performance of the prepared 3D printing bio-ink is poor.

[0095] In summary, the present invention designs the specific composition of 3D printed bio-ink to make the 3D printed bio-ink have excellent comprehensive performance, and then prepares a 3D printed scaffold with strong cell-scaffold interaction, which can be used to prepare skin damage repair materials.

[0096] The applicant declares that the present invention uses the above-described embodiments to illustrate the detailed process flow of the present invention, but the present invention is not limited to the above-described detailed process flow, that is, it does not mean that the present invention must rely on the above-described detailed process flow to be implemented. Those skilled in the art should understand that any improvements to the present invention, equivalent replacements for various raw materials in the products of the present invention, addition of auxiliary ingredients, and selection of specific methods, etc., are all within the scope of protection and disclosure of the present invention.

Claims

1. A 3D printing bio-ink, characterized in that: The 3D printing bio-ink comprises the following components: methacrylic anhydride gelatin, oxidized hyaluronic acid, carboxymethyl chitosan and 2-methacryloyloxyethyl phosphorylcholine.

2. The 3D printing bio-ink according to claim 1, characterized in that The 3D printing bio-ink comprises the following components in parts by weight:

3. The 3D printing bio-ink according to claim 2, characterized in that The 3D printing bio-ink comprises the following components in parts by weight: 7 parts of methacrylic anhydride gelatin, 1.2 parts of oxidized hyaluronic acid, 0.8 parts of carboxymethyl chitosan and 2 parts of 2-methacryloyloxyethyl phosphorylcholine.

4. The 3D printing bio-ink according to any one of claims 1 to 3, characterized in that The raw materials for preparing the methacrylic anhydride gelatin include gelatin and methacrylic anhydride; Preferably, the mass ratio of gelatin to methacrylic anhydride is 1:(0.2-1); Preferably, the weight average molecular weight of the methacrylic anhydride gelatin is ≥14,000.

5. The 3D printing bio-ink according to any one of claims 1 to 4, characterized in that The weight average molecular weight of the oxidized hyaluronic acid is ≥14,000.

6. The 3D printing bio-ink according to any one of claims 1 to 5, characterized in that The 3D printing bio-ink also includes a solvent; Preferably, the solvent comprises a phosphate buffer solution.

7. The 3D printing bio-ink according to any one of claims 1 to 6, characterized in that The 3D printing bio-ink also includes a photoinitiator.

8. A method for preparing a 3D printing bio-ink according to any one of claims 1 to 7, characterized in that: The preparation method comprises the following steps: Dissolve methacrylic anhydride gelatin, carboxymethyl chitosan, 2-methacryloyloxyethyl phosphorylcholine, part of the solvent and an optional photoinitiator to obtain a solution A; mixing the oxidized hyaluronic acid and the remaining solvent to obtain solution B; The solution A and the solution B are mixed evenly to obtain the 3D printing bio-ink.

9. A 3D printed bracket, characterized in that: The raw materials for preparing the 3D printed scaffold include the 3D printed bio-ink and bio-unit according to any one of claims 1 to 7; Preferably, the biological unit comprises any one of cells, DNA or proteins, or a combination of at least two; Preferably, the cells include any one or a combination of at least two of skin tissue cells, human umbilical vein endothelial cells, and keratinocytes.

10. A use of the 3D printing bio-ink according to any one of claims 1 to 7 or the 3D printing scaffold according to claim 9, characterized in that: The application includes use in preparing skin damage repair materials.

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