阅读说明：本技术 仿生皮肤支架及其制备方法 (Bionic skin stent and preparation method thereof ) 是由 杨熙 牛长梅 赵伟新 周晨 于 2021-08-27 设计创作，主要内容包括：本申请提供了一种仿生皮肤支架及其制备方法,涉及仿生皮肤支架技术领域。该仿生皮肤支架包括从上到下依次设置的致密层、表皮层和真皮层。真皮层上具有贯穿真皮层上下两个表面的多个第一通孔和多个第二通孔,表皮层上具有贯穿表皮层上下两个表面的多个第三通孔,一个第一通孔和一个第三通孔对应设置并连通,第二通孔与第三通孔错位设置且不连通,多个第一通孔的截面孔的孔面积总和与多个第二通孔的截面孔的孔面积总和之和大于多个第三通孔的截面孔的孔面积总和。该仿生皮肤支架模拟全层皮肤结构,具有不同梯度的孔隙,有利于血管的生成,加速创面修复。(The application provides a bionic skin stent and a preparation method thereof, and relates to the technical field of bionic skin stents. The bionic skin bracket comprises a compact layer, an epidermal layer and a dermal layer which are arranged from top to bottom in sequence. The dermis layer is provided with a plurality of first through holes and a plurality of second through holes which penetrate through the upper surface and the lower surface of the dermis layer, the epidermis layer is provided with a plurality of third through holes which penetrate through the upper surface and the lower surface of the epidermis layer, one first through hole and one third through hole are correspondingly arranged and communicated, the second through holes and the third through holes are arranged in a staggered mode and are not communicated, and the sum of the hole areas of the section holes of the first through holes and the sum of the hole areas of the section holes of the second through holes is larger than the sum of the hole areas of the section holes of the third through holes. The bionic skin bracket simulates a full-layer skin structure, has pores with different gradients, is beneficial to the generation of blood vessels and accelerates the repair of wound surfaces.)

1. A bionic skin stent is characterized by comprising a compact layer, an epidermal layer and a dermal layer which are sequentially arranged from top to bottom;

the dermis layer is provided with a plurality of first through holes and a plurality of second through holes which penetrate through the upper surface and the lower surface of the dermis layer, the epidermis layer is provided with a plurality of third through holes which penetrate through the upper surface and the lower surface of the epidermis layer, one first through hole and one third through hole are correspondingly arranged and communicated, the second through holes and the third through holes are arranged in a staggered mode and are not communicated, and the sum of the hole areas of the section holes of the first through holes and the sum of the hole areas of the section holes of the second through holes is larger than the sum of the hole areas of the section holes of the third through holes.

2. The biomimetic skin scaffold according to claim 1, wherein the first through hole and the third through hole which are mutually through are coaxially arranged and have the same aperture.

3. The bionic skin scaffold as claimed in claim 2, wherein the aperture of the first through hole is 200-600 μm, and the distance between two adjacent first through holes is 1-20 mm;

the aperture of the second through hole is 200-1000 μm, and the distance between two adjacent second through holes is 1-10 mm.

4. The biomimetic skin scaffold according to claim 3, wherein the thickness of the dermal layer is 1-4mm, the sum of the thicknesses of the epidermal layer and the dermal layer is 1.5-5mm, and the thickness of the dense layer is 10-300 μm; the thickness of the epidermis layer is smaller than that of the dermis layer, and the thickness of the compact layer is smaller than that of the epidermis layer.

5. The biomimetic skin scaffold according to any one of claims 1-4, wherein the dense layer is a dense gel layer, a PU membrane, or a silicone membrane.

6. The biomimetic skin scaffold according to any one of claims 1-4, wherein the epidermis layer and the dermis layer are gel layers having different porosity structures.

7. A method of making a biomimetic skin scaffold according to any of claims 1-6, comprising:

sequentially injecting the corium layer gel solution, the epidermis layer gel solution and the compact layer gel solution into a mould, crosslinking for 1-48h by a composite crosslinking agent after the corium layer gel solution, the epidermis layer gel solution and the compact layer gel solution are shaped, and then demoulding;

the mould is internally provided with a plurality of first cylinders and a plurality of second cylinders, the height of the first cylinders is larger than that of the second cylinders so as to form corresponding layer structures of the bionic skin scaffold, and corresponding through holes are formed in the dermis layer and the epidermis layer.

8. The method of claim 7, wherein the setting is performed by freeze-drying for 12-48 hours.

9. The method of claim 7, wherein the dermal gel solution comprises 1-40mg/ml sodium alginate, 5-40mg/ml collagen, 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 1-40mg/ml gelatin, and 1-10mg/ml sodium hyaluronate;

the epidermal layer gel solution comprises 5-40mg/ml sodium alginate, 1-30mg/ml collagen, 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 5-40mg/ml gelatin and 1-10mg/ml sodium hyaluronate;

the compact layer gel solution comprises 10-40mg/ml sodium alginate, 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 5-20mg/ml gelatin, 1-10mg/ml sodium hyaluronate and 1-10% of glycerol by volume percentage.

10. The method of claim 9, wherein the composite crosslinking agent comprises a crosslinking agent a component and a crosslinking agent B component;

the crosslinking agent A component is one or more of formaldehyde, glutaraldehyde, carbodiimide, hydroxysuccinimide and genipin;

the crosslinking agent B component is calcium chloride.

Technical Field

The application relates to the technical field of bionic skin stents, in particular to a bionic skin stent and a preparation method thereof.

Background

The skin is the largest organ in human body area and has important physiological functions. The treatment of large-area skin defects caused by wounds, burns, scalds and the like is a great clinical problem. The traditional autologous skin transplantation operation is still an important clinical treatment method for treating large-area severe full-thickness skin injury at present, but has the problems of source shortage, secondary injury and the like.

In order to alleviate the problems, an artificial skin stent is used for clinical treatment, the existing artificial skin stent simulates the dermal structure of the skin to a certain extent to achieve the clinical treatment effect, but the problems of insufficient bionic degree of the tissue structure and the like still exist, and a plurality of challenges exist in the quality of wound healing.

The existing artificial skin stent is generally prepared according to the structure of skin, and the artificial skin stent is provided with an epidermal layer and a dermal layer, wherein the dermal layer is provided with through holes so as to promote the proliferation of blood vessels. However, the porosity of the dermal layer is uniform and does not match the tissue generation rate when repairing skin.

Disclosure of Invention

The application aims to provide a bionic skin stent and a preparation method thereof so as to improve the repair effect of a wound surface.

In a first aspect, an embodiment of the present application provides a biomimetic skin scaffold, which includes a compact layer, an epidermal layer, and a dermal layer, which are sequentially disposed from top to bottom. The dermis layer is provided with a plurality of first through holes and a plurality of second through holes which penetrate through the upper surface and the lower surface of the dermis layer, the epidermis layer is provided with a plurality of third through holes which penetrate through the upper surface and the lower surface of the epidermis layer, one first through hole and one third through hole are correspondingly arranged and communicated, the second through holes and the third through holes are arranged in a staggered mode and are not communicated, and the sum of the hole areas of the section holes of the first through holes and the sum of the hole areas of the section holes of the second through holes is larger than the sum of the hole areas of the section holes of the third through holes.

In the bionic skin bracket, the dermis layer is attached to the wound surface, the dermis layer and the epidermis layer are both provided with through holes, and the through holes of the dermis layer are denser, so that the degradation speed of the dermis layer is higher, the degradation speed of the epidermis layer is slower, different gradient degradation rates can be formed, the degradation rates are matched with the tissue generation rate, and the repair of skin wounds is promoted; and the through holes of the epidermis layer are communicated with partial through holes of the dermis layer, which is beneficial to the growth of blood vessels and accelerates the repair of skin wound surfaces. And the compact layer is not provided with the through holes, can provide a physical barrier, has the effects of locking water, protecting the wound surface, preventing infection and the like, provides a moist microenvironment for wound surface repair, and is beneficial to the regeneration of skin tissues.

In a possible implementation manner, the first through hole and the third through hole which are communicated with each other are coaxially arranged and have the same aperture.

On one hand, the blood vessel is favorable for growing in; on the other hand, the degradation rate of the dermis layer and the epidermis layer can be better regulated and matched with the skin repair process.

In a possible implementation manner, the aperture of the first through hole is 200-600 μm, and the distance between two adjacent first through holes is 1-20 mm. The aperture of the second through hole is 200-1000 μm, and the distance between two adjacent second through holes is 1-10 mm.

In one possible implementation, the thickness of the dermis layer is 1-4mm, the sum of the thicknesses of the epidermis layer and the dermis layer is 1.5-5mm, and the thickness of the dense layer is 10-300 μm. The thickness of the epidermis layer is smaller than that of the dermis layer, and the thickness of the dense layer is smaller than that of the epidermis layer. The thickness of each layer and the thickness relation among the layers are more adaptive to the skin of a human body, and the effects of locking water, protecting the wound surface, preventing infection and the like are better.

In one possible implementation, the dense layer is a dense gel layer, a PU film, or a silicone film. The compact layer formed by the materials has better physical barrier function, is beneficial to protecting the wound surface and preventing infection.

In one possible implementation, the epidermis layer and the dermis layer are gel layers with different porosities.

In a second aspect, the present application provides a method for preparing a biomimetic skin scaffold, comprising: sequentially injecting the corium layer gel solution, the epidermis layer gel solution and the compact layer gel solution into a mould, crosslinking for 1-48h by a composite crosslinking agent after the corium layer gel solution, the epidermis layer gel solution and the compact layer gel solution are shaped, and then demoulding. The mould is internally provided with a plurality of first cylinders and a plurality of second cylinders, the heights of the first cylinders are greater than those of the second cylinders so as to form corresponding layer structures of the bionic skin bracket, and corresponding through holes are formed in the dermis layer and the epidermis layer.

According to the application, the bionic skin scaffold is prepared by using a gel solution, and on one hand, the structure of the bionic skin scaffold can better repair a skin wound; on the other hand, after the gel solution is injected, the cross-linking reaction is carried out to form hydrogel, so that the three-layer structure of the bionic skin bracket has no obvious boundary, has better mechanical property and is closer to the skin structure of a human body.

In one possible implementation, the setting mode is freeze drying for 12-48 h. The skin-care product is shaped by a freeze drying mode, so that a micropore structure is formed in the skin-care product, the transportation of nutrient substances and metabolites is facilitated, the adhesion and proliferation of cells are facilitated, and the regeneration of skin tissues is accelerated.

In one possible implementation, the dermal gel solution includes 1-40mg/ml sodium alginate, 5-40mg/ml collagen, 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 1-40mg/ml gelatin, and 1-10mg/ml sodium hyaluronate. The epidermal layer gel solution comprises 5-40mg/ml sodium alginate, 1-30mg/ml collagen, 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 5-40mg/ml gelatin and 1-10mg/ml sodium hyaluronate. The dense layer gel solution comprises 10-40mg/ml sodium alginate, 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 5-20mg/ml gelatin, 1-10mg/ml sodium hyaluronate and 1-10% of glycerol by volume percentage.

The biological materials such as sodium alginate, collagen, chitosan and the like have good biocompatibility, can promote the proliferation, adhesion and migration of cells, provide a microenvironment for the growth of the cells, and can effectively promote wound healing and skin regeneration.

In one possible implementation, the composite crosslinking agent includes a crosslinking agent a component and a crosslinking agent B component. The component A of the cross-linking agent is one or more of formaldehyde, glutaraldehyde, carbodiimide, hydroxysuccinimide and genipin. The component B of the cross-linking agent is calcium chloride.

Drawings

In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings that are required to be used in the embodiments will be briefly described below, it should be understood that the following drawings only illustrate some embodiments of the present application and therefore should not be considered as limiting the scope, and for those skilled in the art, other related drawings can be obtained from the drawings without inventive effort.

Fig. 1 is a schematic layer structure diagram of a biomimetic skin scaffold provided in an embodiment of the present application;

FIG. 2 is a cross-sectional view of a skin mold provided in an embodiment of the present application;

fig. 3 is a first scanning electron microscope image of the biomimetic skin scaffold provided in embodiment 1 of the present application;

fig. 4 is a second scanning electron microscope image of the biomimetic skin scaffold provided in embodiment 1 of the present application.

Icon: 110-a dense layer; 120-a skin layer; 130-dermis layer; 131-a first via; 132-a second via; 121-a third via; 210-a peripheral frame; 220-inner mold core; 230-a groove structure; 221-a backplane; 222-a first cylinder; 223-a second column; 211-first glue injection hole; 212-second glue injection hole; 213-third glue injection hole.

Detailed Description

In order to make the objects, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions in the embodiments of the present application will be clearly and completely described below with reference to the drawings in the embodiments of the present application.

Fig. 1 is a schematic layer structure diagram of a biomimetic skin scaffold provided in an embodiment of the present application. Referring to fig. 1, the biomimetic skin scaffold provided by the present application includes a compact layer 110, an epidermal layer 120, and a dermal layer 130, which are sequentially disposed from top to bottom. The dermis layer 130 is provided with a plurality of first through holes 131 and a plurality of second through holes 132 which penetrate through the upper surface and the lower surface of the dermis layer 130, the epidermis layer 120 is provided with a plurality of third through holes 121 which penetrate through the upper surface and the lower surface of the epidermis layer 120, one first through hole 131 and one third through hole 121 are correspondingly arranged and communicated, the second through holes and the third through holes are arranged in a staggered mode and are not communicated, and the sum of the hole areas of the cross-section holes of the first through holes 131 and the sum of the hole areas of the cross-section holes of the second through holes 132 is larger than the sum of the hole areas of the cross-section holes of the third through holes 121.

It should be noted that: the aperture of each first through hole 131 may be the same or different; the aperture of each second through hole 132 may be the same or different; the aperture of each third through hole 121 may be the same or different, and the application is not limited thereto. A hole communicating with the third through hole 121 is taken as the first through hole 131, and a hole not communicating with the third through hole 121 is taken as the second through hole 132.

One first through hole 131 has a cross section having a hole area, and the sum of the hole areas is the sum of the hole areas of the sectional holes of the plurality of first through holes 131. A second through hole 132 has a cross section with a hole area, and the sum of the hole areas is the sum of the hole areas of the sectional holes of the second through holes 132. One third through hole 121 has a cross section having a hole area, and the sum of the hole areas is the sum of the hole areas of the sectional holes of the third through holes 121.

In the bionic skin stent, through holes are formed in the dermis layer 130 and the epidermis layer 120, and the through holes in the dermis layer 130 are denser, so that the degradation speed of the dermis layer 130 is higher, the degradation speed of the epidermis layer 120 is slightly slower, different gradient degradation rates can be formed, the degradation rates are matched with tissue generation rates, and the repair of skin wounds is promoted; and the through holes of the epidermis layer 120 are communicated with part of the through holes of the dermis layer 130, which is beneficial for the blood vessels to grow from the dermis layer 130 to the epidermis layer 120 and is beneficial for the repair of the skin. And compact layer 110 does not set up the through-hole, can provide physical barrier, has effects such as lock water, protection surface of a wound, prevention infection, provides moist microenvironment for the surface of a wound restoration, does benefit to the regeneration of skin tissue.

Alternatively, the first through hole 131 and the third through hole 121 are coaxially disposed and have the same aperture. On one hand, the blood vessel is favorable for growing in; on the other hand, the degradation rates of the dermal layer 130 and epidermal layer 120 may be better controlled to match the skin repair process.

Further, the aperture of the first through hole 131 is 200-600 μm, and the distance between two adjacent first through holes 131 is 1-20 mm. The aperture of the second through hole 132 is 200-. So as to precisely control the degradation rate of the cortex layer 120 and the dermis layer 130.

It should be noted that the plurality of first through holes 131 may be arranged in an array, and the distance between two adjacent first through holes 131 may be the distance between two adjacent first through holes 131 arranged in an array, or the distance between two diagonal first through holes 131; the plurality of second through holes 132 may be arranged in an array, and the distance between two adjacent second through holes 132 may be the distance between two adjacent second through holes 132 arranged in an array, or the distance between two diagonal second through holes 132.

Illustratively, the distance between two adjacent first through holes 131 is 1mm, 2mm, 3mm, 4mm, 5mm, 10mm or 20 mm; the distance between two adjacent second through holes 132 is 1mm, 2mm, 3mm, 5mm, 8mm or 10 mm.

The first through-hole 131 and the third through-hole 121 have an aperture of 200 μm, 300 μm, 500 μm, 800 μm, or 1000 μm; the aperture of the second via 132 is 200 μm, 300 μm, 400 μm, 500 μm, or 600 μm. The first through hole 131 and the third through hole 121 may have the same or different apertures; the first through hole 131 and the second through hole 132 may have the same or different hole diameters, and the application is not limited thereto.

Alternatively, the thickness of the dermal layer 130 is 1-4mm, the sum of the thicknesses of the epidermal layer 120 and the dermal layer 130 is 1.5-5mm, and the thickness of the dense layer 110 is 10-300 μm. The thickness of the epidermis layer 120 is less than that of the dermis layer 130, and the thickness of the dense layer 110 is less than that of the epidermis layer 120. The thickness is more adaptive to the skin of a human body, and the effects of locking water, protecting the wound surface, preventing infection and the like are better.

Illustratively, the thickness of the dermal layer 130 is 1mm, the thickness of the epidermal layer 120 is 0.5mm, the thickness of the dense layer 110 is 10 μm, and the sum of the thicknesses of the dermal layer 130 and the epidermal layer 120 is 1.5 mm; or the thickness of the dermis layer 130 is 2mm, the thickness of the epidermis layer 120 is 0.6mm, the thickness of the compact layer 110 is 50 μm, and the sum of the thicknesses of the dermis layer 130 and the epidermis layer 120 is 2.6 mm; or the thickness of the dermis layer 130 is 3mm, the thickness of the epidermis layer 120 is 1mm, the thickness of the compact layer 110 is 100 μm, and the sum of the thicknesses of the dermis layer 130 and the epidermis layer 120 is 5 mm; or the thickness of the dermal layer 130 is 4mm, the thickness of the epidermal layer 120 is 0.8mm, the thickness of the dense layer 110 is 300 μm, and the sum of the thicknesses of the dermal layer 130 and the epidermal layer 120 is 4.8 mm.

In the present application, the dense layer 110 is a dense gel layer, a PU film, or a silica gel film. The dense layer 110 formed by the materials has better physical barrier function, is beneficial to protecting wound surface wounds and preventing infection. Optionally, the compact layer 110, the epidermal layer 120, and the dermal layer 130 are all freeze-dried gel layers.

The preparation method of the bionic skin stent comprises the following steps: sequentially injecting the corium layer gel solution, the epidermis layer gel solution and the compact layer gel solution into a mould, crosslinking for 1-48h by a composite crosslinking agent after the corium layer gel solution, the epidermis layer gel solution and the compact layer gel solution are shaped, and then demoulding.

The bionic skin scaffold is prepared by using the gel solution, and on one hand, the structure of the bionic skin scaffold can better repair the skin wound; on the other hand, after the gel solution is injected, the cross-linking reaction is carried out to form hydrogel, so that the three-layer structure of the bionic skin bracket has no obvious boundary, has better mechanical property and is closer to the skin structure of a human body.

In order to more conveniently prepare the bionic skin stent with the structure, the structure of the die can be improved. Fig. 2 is a schematic structural diagram of a skin mold according to an embodiment of the present application, please refer to fig. 1 and fig. 2, the skin mold includes a peripheral frame 210 and an inner core 220, the inner core 220 is disposed in the peripheral frame 210, so that a groove structure 230 is formed between the peripheral frame 210 and the inner core 220, so as to contain a gel solution and perform a cross-linking reaction to obtain a hydrogel, thereby obtaining a biomimetic skin scaffold.

The inner mold core 220 comprises a bottom plate 221, a plurality of first columns 222 and a plurality of second columns 223, the shape of the bottom plate 221 can be matched with the shape of a bionic skin support to be formed, the shape of the bottom plate 221 is square, and the square bionic skin support can be obtained; the bottom plate 221 is circular in shape, so that a circular bionic skin stent can be obtained; the shape of the bottom plate 221 is elliptical, and an elliptical bionic skin stent can be obtained.

The shape of the peripheral frame 210 is matched with the shape of the bottom plate 221, the peripheral frame 210 is mounted on the bottom plate 221, and a groove structure 230 is formed between the peripheral frame 210 and the bottom plate 221. Each first column 222 is connected to the bottom plate 221 and extends away from the bottom plate 221; each second post 223 is also connected to the base plate 221 and extends away from the base plate 221.

Alternatively, the axis of the first cylinder 222 is substantially perpendicular to the bottom plate 221, the axis of the second cylinder 223 is substantially perpendicular to the bottom plate 221, and the first cylinders 222 and the second cylinders 223 are located inside the groove structure 230. The plurality of first columns 222 are used for forming the plurality of first through holes 131, the heights of the plurality of first columns 222 are basically consistent, and the height of each first column 222 is the same as the sum of the height of each first through hole 131 and the height of the through third through hole 121; the second columns 223 are used for forming a plurality of second through holes 132, the heights of the second columns 223 are basically consistent, and the height of each second column 223 is the same as that of each second through hole 132; the aperture of the first through hole 131 and the aperture of the third through hole 121 are the same as the outer diameter of the first cylinder 222, and the aperture of the second through hole 132 is the same as the outer diameter of the second cylinder 223.

The height of the first column 222 is greater than that of the second column 223, and the height of the first column 222 is less than that of the peripheral frame 210. When the bionic skin stent is prepared, when the dermis gel solution is injected into the groove in the skin mold, the height of the dermis gel solution is flush with the upper end of the second column 223, then the epidermis gel solution is continuously injected, the height of the epidermis gel solution is flush with the upper end of the first column 222, and then the dense gel solution is injected, so that the bionic skin stent with the hole structure is formed.

Three glue injection holes can be respectively arranged on the peripheral frame 210, wherein the three glue injection holes are sequentially arranged from top to bottom and respectively include a first glue injection hole 211, a second glue injection hole 212 and a third glue injection hole 213, the height of the third glue injection hole 213 is consistent with that of the second column 223, and after the gel solution in the dermis is injected into the groove through the third glue injection hole 213, the height of the gel solution in the dermis is flush with the upper end of the second column 223; the height of the second glue injection holes 212 is consistent with that of the first cylinders 222, and after the skin layer gel solution is injected into the grooves through the second glue injection holes 212, the height of the skin layer gel solution is flush with the upper ends of the first cylinders 222; after the dense layer gel solution is injected into the groove through the first glue injection hole 211, the height of the dense layer gel solution is consistent with the thickness of the dense layer 110, so as to form the bionic skin stent.

In the application, the dermal layer gel solution comprises 1-40mg/ml sodium alginate (or oxidized sodium alginate), 5-40mg/ml collagen (which can be recombinant human collagen), 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 1-40mg/ml gelatin and 1-10mg/ml sodium hyaluronate. The epidermal layer gel solution comprises 5-40mg/ml sodium alginate (or oxidized sodium alginate), 1-30mg/ml collagen (which can be recombinant human collagen), 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethylcellulose, 5-40mg/ml gelatin and 1-10mg/ml sodium hyaluronate. The dense layer gel solution comprises 10-40mg/ml sodium alginate (or oxidized sodium alginate), 1-10mg/ml chitosan, 1-30mg/ml sodium carboxymethyl cellulose, 5-20mg/ml gelatin, 1-10mg/ml sodium hyaluronate and 1-10% glycerol by volume percentage.

Through the matching of the materials, the bionic skin stent has good biocompatibility, the layer structure interfaces with different gradients are tightly combined, and the bionic skin stent has a complete integrated structure and meets the requirements of a skin multilayer complex structure. In this application, the setting mode may be: and (3) placing the skin mould together with the gel solution inside in an environment with the temperature of-80 ℃ for precooling, and then carrying out freeze drying for 12-48 h. The skin-care product is shaped by a freeze drying mode, so that a micropore structure is formed in the skin-care product, the transportation of nutrient substances and metabolites is facilitated, the adhesion and proliferation of cells are facilitated, and the regeneration of skin tissues is accelerated.

In other embodiments, the setting mode may also be: and (3) placing the skin mould together with the gel solution inside in an environment at 4 ℃ for 1-4h for shaping.

In the present application, the crosslinking may be performed by: and injecting the composite cross-linking agent into the groove of the skin mold through the glue injection hole, and then carrying out cross-linking reaction. In other embodiments, the manner of crosslinking may also be: and immersing the shaped skin mould together with the gel inside into the composite cross-linking agent for cross-linking reaction.

In order to perform a crosslinking reaction on the gel solution to obtain the hydrogel, optionally, the composite crosslinking agent comprises a crosslinking agent A component and a crosslinking agent B component. The component A of the cross-linking agent is one or more of formaldehyde, glutaraldehyde, carbodiimide, hydroxysuccinimide and genipin; the component B of the cross-linking agent is calcium chloride.

And demolding after crosslinking is finished, cleaning the bionic skin stent for multiple times after demolding is finished, removing redundant crosslinking agents, then placing the bionic skin stent in an environment of 80 ℃ below zero for precooling, and then freeze-drying for 12-48 hours to obtain the freeze-dried bionic skin stent. After being packaged, the product can be preserved for a long time after being irradiated and sterilized.

The bionic skin stent obtained by matching the method with the mold has through holes on the dermis layer 130 and the epidermis layer 120, and the through holes of the dermis layer 130 are denser, so that the degradation speed of the dermis layer 130 is higher, the degradation speed of the epidermis layer 120 is slightly slower, different gradient degradation rates can be formed, the degradation rates are matched with the tissue generation rate, and the repair of skin wounds is promoted; and the through holes of the epidermis layer 120 are communicated with part of the through holes of the dermis layer 130, which is beneficial for the blood vessels to grow from the dermis layer 130 to the epidermis layer 120 and is beneficial for the repair of the skin. And compact layer 110 does not set up the through-hole, can provide physical barrier, has effects such as lock water, protection surface of a wound, prevention infection, provides moist microenvironment for the surface of a wound restoration, does benefit to the regeneration of skin tissue.

In order to make the objects, technical solutions and advantages of the embodiments of the present application clearer, the technical solutions of the embodiments of the present application will be clearly and completely described below. The examples, in which specific conditions are not specified, were conducted under conventional conditions or conditions recommended by the manufacturer. The reagents or instruments used are not indicated by the manufacturer, and are all conventional products available commercially.

Example 1

The preparation method of the bionic skin stent comprises the following steps:

(1) preparing a compact layer gel solution: 25mg/ml sodium alginate, 5mg/ml chitosan, 5mg/ml sodium carboxymethylcellulose, 5mg/ml gelatin, 5mg/ml sodium hyaluronate and 5% glycerol.

Preparing a skin layer gel solution: 20mg/ml sodium alginate, 10mg/ml collagen, 5mg/ml chitosan, 5mg/ml sodium carboxymethylcellulose, 10mg/ml gelatin and 3mg/ml sodium hyaluronate.

Preparing a gel solution of a dermis layer: 15mg/ml sodium alginate, 20mg/ml collagen, 2mg/ml chitosan, 2mg/ml sodium carboxymethylcellulose, 20mg/ml gelatin and 2mg/ml sodium hyaluronate.

Preparing a composite cross-linking agent: the crosslinking agent A comprises 1 percent of glutaraldehyde; the component B of the cross-linking agent is 8 percent of calcium chloride.

(2) Firstly, injecting a certain amount of dermal layer gel solution into a skin mould and leveling; then injecting a certain amount of epidermal layer gel solution into a skin mould and leveling; finally, injecting a certain amount of compact layer gel solution into a skin mould and leveling. And (3) placing the skin mould together with the gel solution inside in an environment with the temperature of-80 ℃ for precooling, and then carrying out freeze drying for 24h for shaping. And injecting the composite cross-linking agent into the groove of the skin mold through the glue injection hole for a cross-linking reaction for 12 hours, and demolding after the cross-linking is finished. And (3) washing with deionized water for multiple times, precooling in an environment at-80 ℃, and freeze-drying for 12 hours to obtain the bionic skin scaffold. After being packaged, the product can be preserved for a long time after being irradiated and sterilized.

In the bionic skin stent obtained by shape adaptation of the die, a plurality of second through holes are arranged in an array, the row spacing and the column spacing of two adjacent second through holes of the array are both 1mm, and the aperture of each second through hole is 500 micrometers; the plurality of first through holes and the corresponding third through holes are arranged in an array, the row spacing and the column spacing of two adjacent first through holes of the array are both 1.1mm, and the aperture of each first through hole and the aperture of each third through hole are 400 micrometers; the thickness of the dermis layer is 3mm, the thickness of the epidermis layer is 1mm, and the thickness of the dense layer is 50 μm.

Example 2

Example 2 is a modification of example 1, and example 2 is different in that: pre-cooling the skin mould and the gel solution inside the skin mould in the embodiment 1 in an environment of-80 ℃, and then freeze-drying for 24h for shaping; the modification is as follows: the skin mould together with the gel solution inside was kept in an environment at 4 ℃ for 3h for styling.

Example 3

Example 3 is a modification of example 1, and example 3 is different in that: the dense layer gel solution was without added glycerol.

Example 4

Example 4 is a modification of example 1, and example 4 is different in that: the aperture of each of the first through hole and the third through hole is 200 μm.

Example 5

Example 5 is a modification of example 1, and example 5 is different in that: the aperture of the first through hole is 400 μm, and the aperture of the third through hole is 200 μm.

Comparative example 1

Comparative example 1 is an improvement over example 1, with comparative example 1 differing in that: the second through hole is not arranged on the dermis layer.

Experimental example 1

Fig. 3 is a first scanning electron microscope image of the biomimetic skin scaffold provided in embodiment 1 of the present application; fig. 4 is a second scanning electron microscope image of the biomimetic skin scaffold provided in embodiment 1 of the present application. As can be seen from FIG. 3, the biomimetic skin scaffold provided in example 1 has a certain micropore structure in addition to the larger through holes, and as can be seen from FIG. 4, the pore size of the micropore structure is substantially 40-80 μm, which is beneficial to the transportation of nutrients and metabolites, and is also beneficial to the adhesion and proliferation of cells, and the regeneration of skin tissues is accelerated.

The above description is only a preferred embodiment of the present application and is not intended to limit the present application, and various modifications and changes may be made by those skilled in the art. Any modification, equivalent replacement, improvement and the like made within the spirit and principle of the present application shall be included in the protection scope of the present application.