System and method for preparing artificial skin through 3D printing

Abstract

The invention belongs to the technical field of artificial skin preparation, and particularly relates to a system and method for preparing artificial skin through 3D printing, the system integrates key steps of cell extraction, amplification, artificial skin preparation and culture and the like, rapid amplification of skin cells and efficient construction of the artificial skin are achieved, the overall efficiency is remarkably improved, and the cost is reduced. The preparation period is shortened; the integrated square cabin system provided by the invention can move, so that the timeliness of cell extraction is ensured; besides, the prepared artificial skin with the net ridge structure has a certain thickness, the upper surface and the lower surface are flat, and the artificial skin has a wavy staggered three-dimensional structure, so that more epidermal stem cell dryness can be reserved, and the maintenance of epidermal stem cell marker protein is facilitated.

Description

Technical Field

[0001] This invention belongs to the field of artificial skin preparation technology, and more specifically, relates to a system and method for 3D printing artificial skin preparation. Background Technology

[0002] As the body's first line of defense, the skin plays vital roles in immune surveillance, sensory detection, and self-repair, effectively protecting internal organs from external environmental damage. Skin cells, through highly specific arrangement, promote intercellular interactions, triggering paracrine and autocrine signaling to maintain normal physiological functions. However, burns, bacterial infections, trauma, diabetic foot ulcers, and skin cancer can severely damage or even completely slough off skin tissue. Acute skin loss disrupts its crucial semi-permeable barrier function, leading to massive fluid evaporation and potentially life-threatening consequences. Furthermore, patients with extensive skin damage face the risk of both chronic and acute infections, severely impacting their mental health and quality of life. In cases where the skin cannot repair or regenerate on its own, surgical intervention (such as skin grafting or scab formation) is usually required.

[0003] Currently, autologous skin transplantation is considered the best clinical option for skin reconstruction. However, for patients with large-area burns or trauma, healthy skin donors are scarce, transplantation is difficult, and there is a risk of secondary damage and infection to the donor site. While allogeneic skin transplantation can accelerate wound closure, the lack of standardized procedures may trigger inflammation and immune rejection, thus limiting it to a temporary solution. Artificial skin substitutes, as an emerging treatment approach, construct tissue structures similar to natural skin by culturing and expanding seed cells in vitro and then seeding them into biodegradable biological scaffolds. After transplantation, the scaffold gradually degrades, cells proliferate and differentiate, eventually forming tissue similar to normal skin, offering new possibilities for skin defect repair.

[0004] Currently, the research and preparation of artificial skin mainly relies on allogeneic cells, which still carries the risk of immune rejection after transplantation. Furthermore, the preparation of artificial skin remains largely in the laboratory research stage, hindering large-scale clinical application. This is because existing equipment for cell extraction, expansion, artificial skin preparation, and culture is scattered across different laboratories or medical institutions, lacking integrated and automated technological support. This results in long preparation cycles and low efficiency, further limiting its industrialization and clinical application.

[0005] Therefore, developing a modular system capable of rapidly and efficiently completing skin cell extraction, amplification, and artificial skin manufacturing and culture has significant clinical implications and market potential. Summary of the Invention

[0006] This invention provides a system for 3D printing artificial skin, comprising a chamber containing:

[0007] Extraction area, tissue and cell culture area, and artificial skin preparation area;

[0008] The extraction area includes:

[0009] A constant temperature heating device is used for tissue digestion and incubation.

[0010] Refrigeration equipment for reagent storage and tissue digestion;

[0011] Centrifuges are used for centrifugation operations during cell extraction and passage.

[0012] A clean bench is used for aseptic operations in cell extraction, cell and artificial skin culture;

[0013] The tissue and cell culture area includes:

[0014] Cell culture incubator, used for cell and artificial skin culture;

[0015] CO2 gas cylinder, connected to cell culture incubator;

[0016] The artificial skin preparation area includes: a 3D printer for printing artificial skin;

[0017] The cabin also contains storage devices;

[0018] The cabin is also equipped with a power supply device to provide electrical energy to the system.

[0019] The 3D printer is also equipped with an ultraviolet lamp and an ultraviolet curing lamp. The ultraviolet lamp is used to sterilize the 3D printer and ensure a sterile environment in the printing chamber. The ultraviolet curing lamp is used to cure the bio-ink during the printing process.

[0020] The 3D printer has both extrusion and inkjet printing methods.

[0021] The storage device is further divided into an equipment storage layer and a reagent storage layer.

[0022] The refrigeration device stores reagents including: D. Hanks balanced salt solution, culture medium, digestive enzymes, fetal bovine serum, and antibiotic mixture.

[0023] The present invention also provides a method for 3D printing artificial skin using the system described above, comprising:

[0024] Extrusion printing was performed using bio-ink I containing fibroblasts to obtain a fibroblast mesh with a preset filament spacing and height;

[0025] Gelatin ink was deposited into the gaps between fibroblasts using inkjet printing.

[0026] After curing, continue filling the gaps with bio-ink II containing keratinocytes;

[0027] After printing epidermal cells on the surface of a fibroblast mesh using bio-ink II containing keratinocytes, the cells were irradiated with blue light and cultured to obtain artificial skin.

[0028] The artificial skin has a three-dimensional ridge structure with flat upper and lower surfaces and interlaced wavy patterns.

[0029] The fibroblasts and keratinocytes are obtained from the patient's skin tissue through tissue digestion, cell extraction, and culture using the system.

[0030] The height of the fibroblast mesh is 1-1.5 mm, and the deposition height of the gelatin ink is 0.7-1.2 mm.

[0031] The process involves printing epidermal cells to construct an epidermal layer of 0.3-1 mm.

[0032] The beneficial effects of this invention are:

[0033] 1. This invention integrates key steps such as cell extraction, amplification, artificial skin preparation and culture into one unit, realizing rapid amplification of skin cells and efficient construction of artificial skin, significantly improving overall efficiency and shortening the preparation cycle; moreover, the integrated modular system provided by this invention is mobile, ensuring the timeliness of cell extraction.

[0034] 2. This invention uses the patient's own cells for extraction and expansion, thus eliminating the risks of inflammation and immune rejection associated with allogeneic cells.

[0035] 3. The artificial skin with a ridge structure prepared by the present invention has a certain thickness, flat upper and lower surfaces, and a wavy, interlaced three-dimensional structure, which can retain more epidermal cell stemness and is conducive to the maintenance of epidermal stem cell marker proteins. Attached Figure Description

[0036] Figure 1 This is an external schematic diagram of a system for 3D printing artificial skin.

[0037] Figure 2 This is a schematic diagram of the internal structure of a system for 3D printing artificial skin.

[0038] Figure 3 The growth of primary human skin keratinocytes and fibroblasts extracted using this system and their immunofluorescence staining are shown, where (a) represents keratinocytes and (b) represents fibroblasts.

[0039] Figure 4The relative expression of CK15, CK14, and CK5 genes in layered artificial skin and reticular artificial skin.

[0040] Figure 5 Immunofluorescence staining of Ki67 and IVL proteins in flat and reticular artificial skin. Detailed Implementation

[0041] The technical solution of this application will be described in detail below with reference to specific embodiments.

[0042] This invention provides a system for 3D printing artificial skin, such as... Figure 1 , Figure 2 As shown, it includes a cabin 1, which contains:

[0043] Extraction area, tissue and cell culture area, and artificial skin preparation area;

[0044] The extraction area includes:

[0045] Constant temperature heating device 2 is used for tissue digestion incubation;

[0046] Refrigeration unit 3 is used for reagent storage and tissue digestion;

[0047] Centrifuge 4 is used for centrifugation operations during cell extraction and passage.

[0048] The ultra-clean workbench 6 is used for aseptic operations in cell extraction, cell and artificial skin culture;

[0049] The tissue and cell culture area includes:

[0050] Cell incubator 8, used for cell and artificial skin culture;

[0051] CO2 cylinder 9 is connected to the cell culture incubator;

[0052] The artificial skin preparation area includes: 3D printer 7, used for printing artificial skin;

[0053] The cabin 1 is also equipped with a storage device 10;

[0054] The cabin 1 is also equipped with a power supply device 5, which is used to provide electrical energy to the system.

[0055] Furthermore, the 3D printer 7 is also equipped with an ultraviolet lamp and an ultraviolet curing lamp. The ultraviolet lamp is used to sterilize the 3D printer 7 to ensure a sterile environment in the printing chamber; the ultraviolet curing lamp is used to cure the bio-ink during the printing process.

[0056] In addition, the 3D printer 7 has both extrusion and inkjet printing methods.

[0057] In addition, the storage device 10 is further divided into an equipment storage layer and a reagent storage layer.

[0058] The equipment storage layer is used to store surgical instruments, cell sieves, culture dishes or flasks, pipettes, timers, pipette tips, pipettes, etc.

[0059] Surgical instruments: including dermabrasion knives, ophthalmic forceps, and ophthalmic scissors, used for the extraction, cutting, and separation of skin specimens;

[0060] Cell sieve: Used to filter cell suspensions and remove cell clumps, impurities, or undigested tissue fragments;

[0061] Culture dishes or flasks: used for the culture of cells and artificial skin;

[0062] Pipettes: used to transfer liquids;

[0063] Timer: Used for timing processes such as cell extraction and tissue digestion.

[0064] The reagent storage layer is used to store disinfectant reagents and saline solution.

[0065] Disinfectants include: 20g / L iodine tincture and 75% alcohol, used to disinfect the sites from which samples will be taken from the patient;

[0066] Physiological saline: Used to induce swelling of the patient's skin to facilitate subsequent skin harvesting.

[0067] In addition, the refrigeration device 3 stores reagents including: D. Hanks balanced salt solution, culture medium, digestive enzymes, fetal bovine serum, and antibiotic mixture.

[0068] Among them, D. Hanks balanced salt solution is used for short-term preservation and cleaning of skin specimens and cells;

[0069] Culture media: including high-glucose DMEM medium and keratinocyte culture medium, used for cell and artificial skin culture;

[0070] Digestive enzymes: including 0.25%-0.5% neutral protease, 2.5g / L trypsin and 2g / L-3.52g / L collagenase, used for cell extraction and tissue digestion;

[0071] Fetal bovine serum, added to the culture medium, is used to provide the nutrients and environment required for cell growth and artificial skin culture;

[0072] Antibiotic mixtures: including penicillin-streptomycin solution and penicillin-streptomycin-gentamicin solution, added to culture media for the prevention of bacterial contamination in cell and artificial skin cultures.

[0073] The following operations can be performed using this system:

[0074] I. Cell Extraction and Culture

[0075] ① Specimen processing

[0076] Remove fresh full-thickness skin and rinse the specimen repeatedly with D. Hanks balanced salt solution 3-5 times. Cut the specimen into strips 0.5-1 cm wide and place them in a petri dish. Immerse the tissue in 0.25%-0.5% neutral protease solution (neutral protease volume to specimen volume ratio 5:1) and digest at 4°C for 14-16 hours.

[0077] ② Extraction of dermal cells

[0078] Peel off the epidermis and place the dermis in a culture dish. Cut it into a paste using ophthalmic scissors, add 2 g / L-3.52 g / L collagenase to submerge the tissue, and digest it in a 37°C cell culture incubator for 1-4 hours, until the tissue is essentially digested and dispersed, and no tissue fragments are visible. Add D. Hanks' balanced salt solution and centrifuge at 1500 rpm for 5 minutes. Discard the supernatant, repeat 5 times to wash away the collagenase, and the precipitate is fibroblasts (HSF). Discard the supernatant, add DMEM to prepare a cell suspension, and seed it into 50 mL culture flasks. Incubate at 37°C in a 5% CO2 incubator. Change the culture medium after 24 hours, and then change the culture medium every 3 days. After the cells reach confluence, passage them.

[0079] ③ Extraction of epidermal cells

[0080] Aspirate the separating enzyme and rinse the specimen twice with D. Hanks' balanced salt solution. Gently peel off the epidermis with ophthalmic forceps, discarding the dermis. Place the peeled epidermis in a culture dish, add 2-3 mL of 2.5 g / L trypsin (generally, the volume ratio of epidermis to trypsin is 3:1), and digest in a 37°C cell culture incubator for 30 min. Add DMEM containing 10% fetal bovine serum to stop trypsin digestion. Carefully and repeatedly pipette to form a single-cell suspension. Filter through a 200-mesh sieve, centrifuge at 1500 rpm for 5 min, discard the supernatant, add 1 mL of keratinocyte (KC) culture medium, resuspend, and repeat at 1×10⁻⁶. 5 Inoculate one 50mL culture flask and incubate at 37°C in a 5% CO2 incubator. After 4 hours, the cells will begin to adhere to the culture vessel and extend their tentacles. At this point, change the culture medium. Thereafter, change the culture medium every 2 days.

[0081] This invention uses the patient's own cells for extraction and expansion, thus eliminating the risks of inflammation and immune rejection associated with allogeneic cells.

[0082] II. Printing of Artificial Skin

[0083] Use a 3D printer to manufacture biomimetic ridge artificial skin containing HSF cells and KC cells.

[0084] This invention integrates key steps such as cell extraction, amplification, artificial skin preparation and culture into one unit, realizing rapid amplification of skin cells and efficient construction of artificial skin, significantly improving overall efficiency and shortening the preparation cycle; moreover, the integrated modular system provided by this invention is mobile, ensuring the timeliness of cell extraction.

[0085] It should be noted that, Figure 1 , Figure 2 The mobile laboratory shown is merely one specific implementation of the present invention, used to aid in understanding the technical solution, and should not be considered the sole limitation of the invention. Based on the core concept of the present invention, the carrier form of the mobile laboratory can be flexibly adjusted according to actual application needs, such as containerized (using standard or customized container structures for easy transportation and rapid deployment), trailer platform (using towing vehicles for flexible movement, suitable for field or temporary work scenarios), modular structure (achieving free combination of functional modules through standardized interfaces to meet different experimental needs), and other carrier forms with mobility and capable of carrying experimental functions. After understanding the technical concept of the present invention, those skilled in the art can make reasonable adjustments to the specific implementation methods according to actual needs, including but not limited to structural optimization, material replacement, and functional expansion. Equivalent substitutions, adaptive modifications, or reasonable expansions made without departing from the basic concept of the present invention should be considered to fall within the patent protection scope of the present invention.

[0086] The present invention also provides a method for 3D printing artificial skin using the system described above, comprising:

[0087] Bio-inks were formulated using biomaterials such as methacryloyl gelatin (GelMA) / collagen, GelMA / hyaluronic acid (HA), or GelMA / bacterial nanocellulose (BNC). HSF cells and KC cells were resuspended separately to prepare cell concentrations of 1×10⁻⁶. 6 Bio-ink I (containing HSF) and bio-ink II (containing KC) were used at cells / mL. During printing, the nozzle and platform temperatures were controlled at approximately 18°C ​​and 6°C, respectively, and the nozzle inner diameter was 160 μm (30G).

[0088] First, bio-ink I containing fibroblasts is used for extrusion printing to obtain a fibroblast grid with a preset filament spacing and height. The height of the fibroblast grid is 1-1.5 mm, preferably 1.5 mm and the spacing is 1 mm.

[0089] Subsequently, gelatin ink is deposited into the gaps of the fibroblast network using inkjet printing. The deposition height of the gelatin ink is 0.7-1.2 mm, preferably 0.9 mm.

[0090] After curing, the gaps were filled with bio-ink II containing keratinocytes to construct a regular papillary microstructure.

[0091] Subsequently, epidermal cells were printed on the surface of a fibroblast mesh using bio-ink II containing keratinocytes to construct an epidermal layer of 0.3-1 mm. After irradiation with blue light and cultivation, artificial skin was obtained.

[0092] The fibroblasts and keratinocytes mentioned herein are obtained by taking skin tissue from the patient and then using the system for tissue digestion, cell extraction, and culture.

[0093] Preferably, the artificial skin has a three-dimensional ridge structure with flat upper and lower surfaces and wavy interlacing.

[0094] During the skin model printing process, after the monolayer printing of the fibroblast layer and the keratinocyte layer is completed, the cross-linked structure is irradiated with blue light for 10-20 seconds; finally, the blue light irradiation time before culture is 30-60 seconds to ensure complete cross-linking of the structure.

[0095] During the cultivation phase, complete culture medium was added, and the printed artificial skin was placed in a 37°C, 5% CO2 incubator. The gelatin base dissolved automatically, ultimately forming an artificial skin with a biomimetic network microstructure. The culture medium was changed every two days during the cultivation of the artificial skin.

[0096] The artificial skin with a ridge structure prepared by this invention has a certain thickness, flat upper and lower surfaces, and a wavy, interlaced three-dimensional structure, which can retain more epidermal cell stemness and is beneficial to the maintenance of epidermal stem cell marker proteins.

[0097] Example 1

[0098] The preparation method of ridge-structured artificial skin is as follows:

[0099] After cell extraction and culture using the system provided in this invention, fibroblasts (HSF) and keratinocytes (KC) are obtained. A biomimetic reticular artificial skin containing HSF and KC cells is then manufactured using a 3D printer. The printing chamber is sterilized before printing the model. Bio-ink is prepared using selected biomaterials, and the HSF and KC cells are resuspended separately to prepare a cell concentration of 1×10⁻⁶. 6 Bio-ink I (containing HSF) and bio-ink II (containing KC) were used at cells / mL. During printing, the nozzle and platform temperatures were controlled at approximately 18°C ​​and 6°C, respectively, and the nozzle inner diameter was 160 μm (30G).

[0100] First, using bio-ink I containing HSF cells, an HSF mesh with a filament spacing of 1 mm and a height of 1.5 mm was printed at a pressure of 1.5 bar and a printing speed of 12 mm / s. Then, gelatin ink was deposited into the gaps of the HSF mesh using inkjet printing, with a printing fill height of 0.9 mm. The gelatin was then cured at a low temperature for 5–10 min. Next, bio-ink II was used to continue filling the gaps, constructing a regular papillary microstructure. Finally, epidermal cells were printed onto the scaffold surface using bio-ink II to construct an epidermal layer of 0.3–1 mm, ultimately forming a full-thickness skin model with a certain thickness, flat upper and lower surfaces, and a wavy, interwoven three-dimensional structure.

[0101] During the skin model printing process, after the single-layer printing of the HSF and KC layers was completed, the cross-linked structure was irradiated with blue light for 10–20 seconds. After the structure printing was completed, an additional 30–60 seconds of blue light irradiation was applied to ensure complete cross-linking. Finally, complete culture medium was added, and the printed artificial skin was placed in a 37°C, 5% CO2 incubator. The gelatin base dissolved automatically, ultimately forming an artificial skin with a biomimetic network of microstructures. The culture medium was changed every two days during the cultivation of the artificial skin.

[0102] Comparative Example 1

[0103] The preparation method of layered artificial skin is as follows:

[0104] Samples were prepared using a layer-by-layer photocrosslinking technique. Biomaterials were selected to formulate the bio-ink. HSF cells and KC cells were resuspended separately to prepare a cell concentration of 1×10⁻⁶. 6 Bio-ink I (containing HSF) and bio-ink II (containing KC) were prepared at cell / mL. Bio-ink I was injected into a cylindrical mold (2 mm high) to 75% volume (1.5 mm high), and the cross-linked structure was cured by blue light irradiation for 10–20 s to form the dermis layer. Subsequently, bio-ink II was slowly injected into the mold to full volume (0.5 mm high), and the cross-linked structure was again irradiated by blue light for 10–20 s to construct the epidermis layer. After the preparation of the two-layer structure, an additional 30–60 s of blue light irradiation was applied to ensure complete cross-linking of the structure. Finally, the sample was demolded and transferred to a culture dish containing complete culture medium. The prepared layered artificial skin was placed in a 37°C, 5% CO2 incubator, and the culture medium was changed every two days during the culture of the artificial skin.

[0105] Experimental analysis

[0106] 1. Cell characterization

[0107] Extracted keratinocytes (KC cells) and fibroblasts (HSF cells) were observed using an inverted microscope. Figure 3The cells showed good growth, proliferating and confining the culture flask. To further confirm the phenotypes of keratinocytes and fibroblasts, KC cells were stained with CK15 immunofluorescence, and HSF cells were stained with COL I immunofluorescence. The results showed that KC cells expressed CK15, confirming that they were keratinocytes; HSF cells expressed COL I, confirming that they were fibroblasts.

[0108] 2. The influence of microstructure on keratinocyte phenotype

[0109] The artificial skin with a ridged (RR) structure obtained in Example 1 was used as the experimental group, and the artificial skin with a traditional flat structure obtained in Comparative Example 1 was used as the control group. After culturing at the air-liquid interface for 3 days (AL-Day 3) and 7 days (AL-Day 7), total RNA was extracted from cells in each group. The mRNA expression levels of epidermal stem cell marker genes CK15 and basal cell marker genes CK5 and CK14 were detected using RT-PCR. Figure 4 As shown.

[0110] The results showed that the expression of CK15, CK14, and CK5 was significantly higher in cells of the reticular artificial skin structure than in cells of the lamellar artificial skin structure. Furthermore, the expression levels of CK15, CK14, and CK5 were higher after 7 days of air-liquid interface culture than after 3 days. This indicates that cells of the reticular artificial skin structure retained more epidermal stemness, while cells of the lamellar artificial skin structure exhibited a higher degree of differentiation.

[0111] 3. Evaluate the effect of ridge microstructure formation.

[0112] To evaluate the effectiveness of the ridged artificial skin, it was compared with traditional layered artificial skin (flat artificial skin) by performing sectioning and staining analysis. Figure 5 As shown, the upper edge of the reticular microstructure in artificial skin exhibits a horizontal pattern, while the lower edge displays a wavy pattern, similar to the natural reticular ridge morphology of human skin. Cells are tightly integrated with the dermis; compared to lamellar artificial skin, the contact area between cells and the dermis in reticular artificial skin is larger. Cells in both reticular and lamellar artificial skin can form epidermal tissue of a certain thickness and with tight connections.

[0113] To visually observe epidermal development, the expression patterns of epidermal cell proliferation markers (Ki67) and late (IVL) differentiation markers in reticuloendothelial and lamellar artificial skin sections were assessed using immunofluorescence staining.

[0114] like Figure 5 As shown, the staining results indicate that Ki67 is expressed more prominently and abundantly in the ridge structure.

[0115] In both structures, IVL expression is concentrated in the uppermost layer of the artificial skin, while the IVL expression is more abundant in the reticular skin and shows better continuity in the reticular structure. This indicates that the artificial skin with the reticular structure is more likely to differentiate and mature in vitro.

Claims

1. A system for 3D printing artificial skin, characterized in that, Includes a cabin (1), which contains: Extraction area, tissue and cell culture area, and artificial skin preparation area; The extraction area includes: A constant temperature heating device (2) is used for tissue digestion and incubation; Refrigeration unit (3) is used for reagent storage and tissue digestion; Centrifuge (4), used for centrifugation operations during cell extraction and passage; A clean bench (6) is used for aseptic operations in cell extraction, cell and artificial skin culture; The tissue and cell culture area includes: Cell culture incubator (8), used for cell and artificial skin culture; CO2 cylinder (9) is connected to the cell culture incubator; The artificial skin preparation area includes: a 3D printer (7) for printing artificial skin; The cabin (1) is also equipped with a storage device (10); The cabin (1) is also equipped with a power supply device (5) for providing electrical energy to the system.

2. The system for 3D printing artificial skin according to claim 1, characterized in that, The 3D printer (7) is also equipped with an ultraviolet lamp and an ultraviolet curing lamp. The ultraviolet lamp is used to sterilize the 3D printer (7), and the ultraviolet curing lamp is used to cure the bio-ink during the printing process.

3. The system for 3D printing artificial skin according to claim 1, characterized in that, The 3D printer (7) has two printing methods: extrusion and inkjet.

4. The system for 3D printing artificial skin according to claim 1, characterized in that, The storage device (10) is further divided into an equipment storage layer and a reagent storage layer.

5. The system for 3D printing artificial skin according to claim 1, characterized in that, The refrigeration device (3) stores reagents including: D. Hanks balanced salt solution, culture medium, digestive enzymes, fetal bovine serum, and antibiotic mixture.

6. A method for 3D printing artificial skin using the system of claim 1, characterized in that, include: Extrusion printing was performed using bio-ink I containing fibroblasts to obtain a fibroblast mesh with a preset filament spacing and height; Gelatin ink was deposited into the gaps between fibroblasts using inkjet printing. After curing, continue filling the gaps with bio-ink II containing keratinocytes; After printing epidermal cells on the surface of a fibroblast mesh using bio-ink II containing keratinocytes, the cells were irradiated with blue light and cultured to obtain artificial skin.

7. The method for preparing artificial skin by 3D printing according to claim 6, characterized in that, The artificial skin has a three-dimensional ridge structure with flat upper and lower surfaces and interlaced wavy patterns.

8. The method for preparing artificial skin by 3D printing according to claim 6, characterized in that, The fibroblasts and keratinocytes are obtained from the patient's skin tissue through tissue digestion, cell extraction, and culture using the system.

9. The method for preparing artificial skin by 3D printing according to claim 6, characterized in that, The height of the fibroblast mesh is 1-1.5 mm, and the deposition height of the gelatin ink is 0.7-1.2 mm.

10. The method for preparing artificial skin by 3D printing according to claim 6, characterized in that, The process involves printing epidermal cells to construct an epidermal layer of 0.3-1 mm.

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