A hydrogel with skin damage repairing effect

Abstract

The application belongs to the field of 3D biological printing and particularly relates to a hydrogel with skin damage repair function. 3+ The application proposes a preparation method of Eu 2+ mediated collagen polypeptide / sodium alginate composite hydrogel, successfully applies the hydrogel to 3D printing skin wound dressing, uses the hydrogel as biological ink, prepares skin full-layer damage dressing through 3D printing technology, and finally uses Ca 2+ The coordination between the Eu 2+ and sodium alginate enhances the stability of the hydrogel. The 3D printing skin wound dressing constructed by the application has good function of promoting skin full-layer damage repair and has application potential in the fields of tissue engineering and regenerative medicine.

Description

TECHNICAL FIELD

[0001] The present application belongs to the field of biological functional materials, and particularly relates to a hydrogel with skin damage repair function. BACKGROUND

[0002] Skin damage can destroy the protective function of the skin to the internal environment of the human body, and serious skin wounds are difficult to heal in a natural state. Skin is the largest organ of the human body and plays an important role in protecting the human body from the external environment and maintaining the balance of the body. After the skin is damaged, it can cause serious health problems such as viral infection, blood and tissue fluid loss, and impaired function of the tissue structure. Skin wound repair is a complex physiological process, mainly including four stages of hemostasis, inflammation, proliferation and remodeling. Large-area full-thickness skin defects can damage the epidermis, dermis and subcutaneous tissue, and such skin wounds are difficult to heal by themselves, so clinical treatment is needed to promote the repair. Skin wound dressings can prevent the skin wound from being further damaged and provide a good environment for the healing of the wound, thereby effectively promoting the healing of full-thickness skin wounds.

[0003] 3D printing technology can prepare materials that can conform to the shape and depth of the wound, and this technology has attracted more and more attention in the preparation of skin wound dressings. Sodium alginate (SA) is a natural polymer extracted from brown algae and can form a gel by coordinating with metal ions at room temperature, and is a widely used 3D printing bio-ink. Patent CN107139447A discloses a sol-like sodium alginate as a printing raw material, which is extruded and then cross-linked with Ca 2+ to prepare a hydrogel scaffold with a three-dimensional structure. Patent CN114470336A discloses a mixed hydrogel composed of sodium alginate and gelatin or methacrylamide-based gelatin, which can be printed into a scaffold structure as a 3D printing bio-ink, and then cross-linked with Ca 2+ to prepare a hydrogel scaffold. However, SA itself does not have biological activity, and it is urgent to construct functionalized SA for 3D printing of biological functional materials. SUMMARY

[0004] In view of the above technical problems, the present application proposes a preparation method of Eu 3+ mediated collagen polypeptide / sodium alginate composite hydrogel, which is used as a bio-ink to prepare a skin full-thickness damage dressing through 3D printing technology, and finally the stability of the dressing is enhanced through coordination between Ca 2+ and sodium alginate. The 3D printed skin wound dressing constructed by the present application has good function of promoting the repair of skin full-thickness damage, and has application potential in the fields of tissue engineering and regenerative medicine. Specifically, the present application includes the following contents:

[0005] In a first aspect, the present application provides a hydrogel having skin damage repair effect, the hydrogel comprising the following components: sodium alginate, europium ion (Eu 3+ ), collagen polypeptide; the sequence of the collagen polypeptide is:

[0006] (G-X-Y) m1 -collagen functional sequence-(G-X-Y) m2 -(D) n ;

[0007] wherein G is glycine, and the D is aspartic acid;

[0008] X and Y are selected from the same or different amino acids;

[0009] m1 and m2 are any integer greater than or equal to 1, and the sum of m1 and m2 is greater than or equal to 6;

[0010] n is any integer greater than or equal to 2;

[0011] The collagen functional sequence is selected from any one of integrin binding site GFOGER, integrin binding site GROGER, integrin binding site GLOGER, integrin binding site GMOGER, integrin binding site GAOGER, integrin binding site GLKGEN, integrin binding site GLOGEN, DDR binding site GVMGFO, MMP enzyme cutting site GPLGIAGITGAR, MMP enzyme cutting site GPQGIAGQRGVV, MMP enzyme cutting site GPQGLLGAOGIL, MMP enzyme cutting site GPQGLAGQRGIV, heparin binding site GRPGKRGKQGQK, and fibronectin binding site GLPGQRGER.

[0012] Preferably, X is proline P, and Y is hydroxyproline O; or both X and Y are hydroxyproline O.

[0013] Preferably, the sequence of the collagen polypeptide is: (G-P-O)3-GFOGER-(G-P-O)3-(D)2.

[0014] Preferably, the concentration ratio of sodium alginate, europium ion (Eu 3+ ), collagen polypeptide is: 40 mg / mL: 2 mM: 6 mM.

[0015] Preferably, the preparation method of the hydrogel is: dissolving sodium alginate in a collagen polypeptide solution to obtain a mixed solution of collagen polypeptide and sodium alginate; and mixing the mixed solution with an Eu 3+ solution in equal volume to prepare the hydrogel.

[0016] In a second aspect, the present application provides a skin wound dressing prepared by 3D printing technology using the hydrogel of the first aspect.

[0017] In a third aspect, the present application provides a preparation method of the skin wound dressing of the second aspect, which comprises the following steps:

[0018] (1) preparing the hydrogel of the first aspect;

[0019] (2) 3D printing: using the hydrogel prepared in step (1) as bio-ink, printing a gel material by 3D printing technology, and atomizing CaCl2 solution on the surface of the 3D-printed gel material to obtain a skin wound dressing.

[0020] Preferably, the 3D printing technology in step (2) uses an extrusion type 3D printer.

[0021] Preferably, the concentration of the CaCl2 solution used in step (2) is 170 mg / mL.

[0022] Preferably, the skin wound dressing is a disc-shaped material with a diameter of 15 mm and a height of 2.5 mm.

[0023] In a fourth aspect, the present application provides the use of the hydrogel of the first aspect or the skin wound dressing of the second aspect in the preparation of a skin damage repair product.

[0024] Preferably, the skin damage is full-thickness skin damage.

[0025] The present application has the following beneficial effects: ① Sodium alginate is a widely used 3D printing ink, but it does not have biological function. The present application proposes a preparation method of Eu 3+ mediated collagen polypeptide / sodium alginate composite hydrogel, which modifies sodium alginate and forms a composite hydrogel with biological function; ② Sodium alginate cannot print a structure with fine details, but the composite hydrogel provided by the present application improves its 3D printability and successfully applies it to 3D printed skin wound dressings; ③ The skin wound dressing prepared by 3D printing technology using the composite hydrogel as bio-ink effectively promotes the healing of full-thickness skin defects in SD rats; ④ The prepared skin full-thickness wound dressing has no risk of virus transmission and immunogenicity, and has good biocompatibility; ⑤ The preparation process of the composite hydrogel is simple and the reaction conditions are mild. BRIEF DESCRIPTION OF DRAWINGS

[0026] The present application will be further described below in conjunction with the drawings and examples.

[0027] Figure 1 For Eu 3+ The 3D printable results of the collagen polypeptide / sodium alginate hydrogel formed by mediation; Wherein A is the grid material photo and SEM picture printed based on the gel SA-C, SA-E-C and SA-E-D-C; B is the hollow cylinder printed based on the gel SA-E-D-C; C is the star printed based on the gel SA-E-D-C; D is the meniscus printed based on the gel SA-E-D-C; E is the model picture, top view and side view of the ear printed based on the gel SA-E-D-C;

[0028] Figure 2 For Eu 3+ And Ca 2+ The physicochemical property characterization results of the collagen polypeptide / sodium alginate hydrogel formed by mediation; Wherein A-C are SEM images of gels SA-C, SA-E-C and SA-E-D-C respectively; D is the porosity of the gels SA-C, SA-E-C and SA-E-D-C; E is the compression modulus of the gel; F is the curve of the swelling rate of the gel changing with time; G is the frequency scanning curve of the gel; H is the strain scanning curve; I is the change of G' and G" with time under the alternating scanning of 1% and 80% strain;

[0029] Figure 3 The healing conditions of the full-thickness skin wounds of SD rats at different time points after applying different dressings; Wherein A is the photo of the skin wound of the SD rat; B is the change of the wound contour; C is the wound healing rate;

[0030] Figure 4 The H&E and Masson staining results of the skin tissues of the SD rats in each group at different time points;

[0031] Figure 5 The collagen fiber expression generation rate of the skin wound site of the SD rats in each group at different time points. DETAILED DESCRIPTION

[0032] The technical solutions of the present application are further described below in combination with the examples, but the protection scope of the present application is not limited to the following.

[0033] In the following examples, the GPO is Gly( glycine)-Pro( proline)-Hyp( hydroxyproline); the GFOGER is Gly( glycine)-Phe( phenylalanine)-Hyp( hydroxyproline)-Gly( glycine)-Glu( glutamic acid)-Arg( arginine); and the SA is sodium alginate.

[0034] The sequence of the collagen polypeptide in the following examples is (GPO)3GFOGER(GPO)3DD; the collagen polypeptide comprises a (GPO)3repeat sequence for stabilizing the triple helix structure of the polypeptide; the collagen polypeptide comprises the cell integrin binding sequence GFOGER in the native collagen protein; the N-terminus of the collagen polypeptide comprises two amino acids D, so that the amino acids at this position are coordinated with Eu 3+ coordinated.

[0035] The gel preparation method described in the following examples is: mixing SA and Eu(NO3)3solution, so as to prepare a polypeptide-modified composite gel formed by the mediation of Eu 3+ The collagen polypeptide-modified gel is prepared by dissolving SA with a collagen polypeptide solution, and then mixing with an Eu(NO3)3solution to form a polypeptide-modified composite gel; the CaCl2solution is atomized into droplets using an atomizer, and is covered on the surface of the SA solution and the above two hydrogels, respectively, to prepare Ca 2+ solidified gels SA-C, SA-E-C and SA-E-D-C.

[0036] The method for characterizing the 3D printability of the gel described in the following examples is: using the gel that has not been Ca 2+ solidified as a bio-ink, printing it into a grid shape with fine structure and other various shapes.

[0037] The method for characterizing the physical and chemical properties of the composite hydrogel described in the following examples includes: rheological test, field emission scanning electron microscope (SEM) test, compression resistance test and swelling performance test.

[0038] The preparation method of the skin wound dressing described in the following examples is: using the gel that has not been Ca 2+ solidified as a bio-ink, using an extrusion type 3D printer to print a cylindrical shape with a length and width of 15 mm and a height of 2.5 mm, and then Ca 2+ solidifying it.

[0039] The experimental model for verifying the efficacy of the skin wound dressing described in the following examples is: a full-thickness skin injury model of SD rats.

[0040] skin injury model.

[0041] Example 1 Eu 3+ mediated collagen polypeptide / sodium alginate composite hydrogel

[0042] SA and Eu(NO3)3·6H2O were respectively dissolved in 100mM pH 7.0 Hepes buffer to prepare SA and Eu 3+The initial solutions were prepared to achieve initial concentrations of 80 mg / mL and 4 mM, respectively. The initial SA solutions were diluted to 100 mg / mL using 100 mM Hepes buffer (pH 7.0) to prepare the SA working solution. SA and Eu... 3+ The initial solutions were mixed at a 1:1 volume ratio to form gel SA-E, with an SA concentration of 40 mg / mL and Eu... 3+ The concentration was 2 mM. An initial CMP-DD solution was prepared by dissolving the peptide CMP-DD [(GPO)3GFOGER(GPO)3DD] in 100 mM pH 7.0 Hepes buffer, with an initial concentration of 12 mM. SA was then dissolved in the initial peptide solution to prepare an initial SA / CMP-DD mixture, resulting in an initial SA concentration of 80 mg / mL. This initial SA / CMP-DD mixture was then mixed with Eu... 3+ The initial solutions were mixed in a 1:1 ratio to form a composite gel, SA-ED. At this point, SA and Eu... 3+ The final concentrations of the peptide CMP-DD were 40 mg / mL, 2 mM, and 6 mM, respectively. SA working solution, gel SA-E, and SA-ED were used as bio-inks, transferred into printing syringes, and 3D printed using an extrusion 3D printer through a tapered plastic needle, thus demonstrating the printability of SA solution, gel SA-E, and SA-ED. The printing speed was set to 2 mm / s, and the extrusion speed to 1.8 mm / s. 3 / s. When printing the mesh material, a block model was selected, with the model's length and width both set to 20mm, height to 3mm, layer height to 0.6mm, and line width to 3mm. After the material printing was completed, droplets of a 170mg / mL CaCl2 solution were used to cover the material surface using an atomizer, and the solution was allowed to soak for 10 minutes to prepare 3D printing materials of different shapes: SA-C, SA-EC, and SA-EDC.

[0043] The results are as follows Figure 1 As shown in the photographs and SEM characterization of the printed models, the mesh structure printed with SA solution is prone to collapse and cannot form a regular shape; using gel SA / Eu 3+ It can print fine and regular meshes, and when using peptide CMP-DD modified gel SA-ED as a bio-ink, its printing accuracy is consistent with that of gel SA-E. Figure 1 As shown in Figure A). We further printed hollow cylinders using gel SA-ED (as shown in Figure A). Figure 1 As shown in B), star-shaped ( Figure 1 As shown in C), meniscus ( Figure 1 As shown in D) and the ear model ( Figure 1The results show that the gel SA-E + The printing precision is good, and the introduction of the polypeptide CMP-DD does not affect the printing effect.

[0044] Example 2 Eu 3+ and Ca 2+ Characterization of the physicochemical properties of the collagen polypeptide / sodium alginate hydrogel formed by mediation

[0045] 1. Field emission scanning electron microscope (SEM) and porosity characterization of the composite gel

[0046] Using the method in Example 1, SA working solution, gel SA-E and SA-E-D were prepared, and using an atomizer, the CaCl2 solution was atomized into droplets and covered on the surface of the SA working solution, gel SA-E and SA-E-D, respectively, to prepare the gels SA-C, SA-E-C and SA-E-D-C. The composite gels SA-C, SA-E-C and SA-E-D-C were freeze-dried using a freeze dryer, the freeze-dried samples were cut into thin slices with a thickness of 2-3 mm, and were pasted on the conductive glue on the surface of the metal disc, and were sputtered with gold for 2 minutes under vacuum conditions, and the morphology was characterized using a field emission scanning electron microscope.

[0047] The mass of the freeze-dried gels SA-C, SA-E-C and SA-E-D-C was marked as m1, and the volume was marked as V1. The materials were soaked in ultrapure water for 10 minutes, and the wet weight of the materials was recorded as m2, the weight of water in the materials was recorded as m3 (m3 = m2-m1), the volume of water was recorded as V2 (V2 = m3 / ρ), and the porosity P = V2 / V1 x 100%. Each sample was determined in triplicate.

[0048] The results are shown in Table 2. Figure 2 As shown in Table 2, the three gels all formed a porous structure, among which the gel SA-C formed irregular pores with the largest pore size; the gel SA-E-C had regular pores with a reduced pore size; and the composite gel SA-E-D-C formed a regular porous structure with a smaller pore size. The results of the porosity showed that the gel SA-C had the largest porosity (207%), the porosity of the gel SA-E-C was reduced to 187%, and the porosity of the SA-E-D-C was further reduced to 131%. The above results show that the Eu 3+ After the polypeptide CMP and SA are combined, they will form a more regular porous structure.

[0049] 2. Characterization of the compression resistance of the composite gel

[0050] SA working solution, gel SA-E, and SA-ED were prepared using the method described in Example 1. The SA working solution, gel SA-E, and SA-ED were used as bio-inks, transferred into printing syringes, and cylindrical models were printed using an extrusion 3D printer. The printing speed was set to 2 mm / s, and the extrusion speed was 1.8 mm / s. 3 The model was set to 10mm in length and width, 6mm in height, 0.6mm in layer height, and 1mm in line width. After the material was printed, droplets of a 170mg / mL CaCl2 solution were sprayed onto the material surface using an atomizer and allowed to soak for approximately 10 minutes. A compression test was performed using a universal testing machine at a compression rate of 1mm / min and a strain of 70%. The compressive modulus was calculated using the average slope of the stress-strain curve at 15% strain.

[0051] The results are as follows Figure 2 As shown in Figure E, the compressive modulus of gel SA-C is only 10.5 kPa. Introducing Eu into the gel system... 3+ After that, Eu 3+ and Ca 2+ The combined effect of these factors improved the gelling properties of SA, thus increasing the compressive modulus of gels SA-EC and SA-EDC to 16.5 kPa and 13.8 kPa, respectively. These results indicate that Eu... 3 and Ca 2+ The SA composite hydrogel formed under the mediation has good compressibility.

[0052] 3. Characterization of the swelling properties of the composite gel

[0053] Gels SA-C, SA-EC, and SA-EDC were prepared using the method described in section 1 above. The mass M0 (g) of each gel was weighed, and they were then immersed in ultrapure water, ensuring complete submersion, and placed in a 37°C constant temperature water bath. The gels were weighed at specific time points, and their mass was recorded as M. t Before weighing, wipe the surface of the gel dry and calculate the swelling ratio SR of the composite gel [SR = (M t -M0) / M0].

[0054] The results are as follows Figure 2 As shown in Figure F, the swelling ratios of all three gels reached equilibrium around 2 hours, with gel SA-C reaching a swelling ratio of 116.8 ± 4.3%. The introduction of Eu into the gel system... 3+ After that, Eu 3+ and Ca 2+The mechanical properties of SA were improved by the synergistic effect of Eu 3+ and Ca 2+ The SA composite hydrogel formed under the mediation of Eu

[0055] 4. Rheological test of the composite gel and characterization of self-healing performance

[0056] Gel SA-C, SA-E-C and SA-E-D-C were prepared using the method in 1 above; the composite gels SA-C, SA-E-C and SA-E-D-C were respectively placed on the sample table of the rheometer, the shear strain was set to 1%, the storage modulus G'(Pa) and the loss modulus G"(Pa) of the gel in the angular frequency range of 1-100 rad / s were tested by frequency scanning; then the angular frequency was fixed at 1 rad / s, and the changes of G'(Pa) and G"(Pa) of the gel in the shear strain range of 1-100% were detected. By fixing the angular frequency at 1 rad / s, the shear strain was alternately scanned at 1% and 80% to detect the self-healing performance of the gel.

[0057] The frequency scanning results are shown in G of Figure 2 The results show that the storage modulus (G') of gels SA-C, SA-E-C and SA-E-D-C is greater than the loss modulus (G"), and their G' increases with the increase of frequency, showing the typical rheological properties of hydrogel materials. When the shear frequency reaches 80 rad / s and higher, the G' of gel SA-C reaches about 6300 Pa, while the G' of gels SA-E-C and SA-E-D-C increases to about 9800 Pa and 7200 Pa respectively, proving that gels SA-E-C and SA-E-D-C have stronger gelation performance and mechanical strength than SA-C. Figure 2 The results show that with the increase of strain, the G' of gels SA-C, SA-E-C and SA-E-D-C gradually decreases, while the G" continuously increases. When the strain is greater than 30%, the value of G" exceeds G'. It proves that the three gels have typical shear thinning behavior. The results of the alternate scanning of gels SA-C, SA-E-C and SA-E-D-C at low strain of 1% and high strain of 80% are shown in H of Figure 2As shown in Figure I, when the strain is 1%, the G' and G” values ​​of gels SA-C, SA-EC, and SA-EDC are all around 5000 Pa and 600 Pa, respectively, at which point they all maintain gel characteristics. However, under 80% high-amplitude oscillatory shear, G' drops to around 480 Pa, and at this point, G” is greater than G', thus the gels all transform into a quasi-liquid state. After the shear strain is restored to 1%, the G' and G” values ​​of the gels return to their initial state, and this process can be repeated for several cycles, demonstrating that all three gels have good self-healing properties.

[0058] Example 3: Full-thickness skin damage repair function of 3D printed skin dressings

[0059] SA working solution, gel SA-E, and SA-ED were prepared using the method in Example 1. These were then used as bio-inks to print cylindrical models using an extrusion 3D printer, with the printing speed set to 2 mm / s and the extrusion speed to 1.8 mm / s. 3 The model was set to 15mm in length and width, 2.5mm in height, 0.6mm in layer height, and 1mm in line width. After the material was printed, droplets of a 170mg / mL CaCl2 solution were applied to the material surface using an atomizer and allowed to soak for approximately 10 minutes to prepare cylindrical gel materials SA-C, SA-EC, and SA-EDC. These materials were then stored at 4℃ for later use.

[0060] SD female rats with a body weight of 150-200 g were anesthetized by intraperitoneal injection of 10% chloral hydrate at a dose of 0.3 mL / 100 g. The hair on the back of the rats was shaved using a hair clipper, and the involved hair was removed using depilatory cream, and then the back of the rats was cleaned with 0.9% sodium chloride injection. The exposed skin on the back of the rats was disinfected with iodophor, and then a 1.5 cm diameter circular area was marked on the exposed skin using the opening of a 15 mL centrifuge tube, the upper skin tissue was completely cut off using surgical scissors to form a circular wound area, the wound reached the fascia, and hemostasis was performed. Then the rats were randomly divided into 5 groups, and the skin wound of the blank group was not dressed, the skin wound of the control group was dressed with gel material SA-C, and the skin wound of the experimental group was treated with gel dressings SA-E-C and SA-E-D-C respectively. The dressings were fixed using adhesive tape and elastic bandage, and each group of rats was individually raised, and the wound condition was observed every day. The wound was photographed on the 4th, 7th, 11th and 21st day after the operation, and a scale accurate ruler was placed at the wound site to calibrate the multiple of the picture. Then the rats were sacrificed by cervical dislocation, and the wound edge tissue was removed, fixed with 10% formaldehyde, and treated with conventional paraffin embedding, sectioning and H&E staining and Masson staining. The skin full-thickness wound map of each group of rats at each time point was obtained by splicing using Adobe Photoshop software. The area of the rat skin wound was calculated using Image J software, and the collagen fiber expression generation rate in the Masson staining result picture was analyzed. The wound healing rate and collagen fiber expression generation rate of the rat skin wound were calculated by the following formula respectively:

[0061] Wound healing rate = wound area at each time point / initial wound area;

[0062] CF% = A C / A0;

[0063] wherein A C is the collagen fiber area, and A0is the total area of the skin tissue.

[0064] The healing results of the full-thickness skin wound of the SD rats after dressing with different dressings at each time point are shown in Table 1, and the photographs of the skin wound of the SD rats are shown in Figure 1. Figure 3 Figure 3 ​As shown in Figure A), on day 4, the skin wounds of rats in the control group, SA-C group, and SA-EC group were bright red and showed signs of bleeding. The control group and SA-C group had larger wound areas with slight infection and swelling at the edges. The SA-EC group had smaller wound areas and no infection. The wound area in the SA-EC group further decreased, and scabs formed, making the wound smoother. On day 7, the skin wound area of ​​all groups decreased and scabs formed. The scabs in the control group and SA-C group were larger, with uneven surfaces and darker colors. The scabs in the SA-EC group were smaller and flatter. The scabs in the SA-EC group were darker and irregularly shaped, while the scabs in the SA-EDC group were lighter in color and more regular in shape. On day 11, the scabs on the wounds of rats in the SA-EDC group fell off, and the wounds became flat and lighter in color. Darker scabs remained on the wounds of the control and SA-C groups, with the scab area in the control and SA-C groups being larger than that in the SA-EC group. On day 21, the skin of rats in the control and SA-C groups still had obvious skin depressions, and these areas were a darker red. The skin of rats in the SA-EC group also had depressions, but these areas were light pink. The skin wounds of rats in the SA-EDC group were basically healed, and the skin was relatively smooth; the original wound site was white with a small amount of light pink. Changes in the edge and area of ​​the rat skin wounds at different time points (e.g., Figure 3 As shown in Figure B, compared with the control group and the SA-C group, the reduction rate of skin wound area in the SA-EC and SA-EDC groups was significantly faster, and the wound healing rate in the SA-EDC group was also faster than that in the SA-EC group. At 21 days, the wound area in the SA-EDC group decreased from 268.13 mm². 2 Dropped to 2.40mm 2 The value was significantly smaller than that of the SA-EC group (245.33 mm). 2 Reduced by 14.07mm 2 ), SA-C group (248.13mm) 2 Reduced by 19.40mm 2 ) and control group (by 239.55mm) 2 It dropped to 15.27mm 2 The results of the wound healing rate are as follows: Figure 3 As shown in Figure C, the wound healing rates of rats in the SA-EC and SA-EDC groups were significantly higher than those in the control and SA-C groups, with the SA-EDC group exhibiting the highest wound healing rate. These results demonstrate that the scaffold material SA-EDC has the most significant effect in promoting wound closure.

[0065] The H&E staining results of the skin tissues of the SD rats are shown in Figure 4 At day 4, the skin wound tissues of each group all had inflammatory cell infiltration, the dermis of the skin of the control group and the SA-C group was thin, the granulation tissue was sparse, and the wound was severely sunken, the skin of the SA-E-C and SA-E-D-C groups had granulation tissue to fill the wound, making the surface flat, among which the wound of the SA-E-D-C group had scab tissue, the density of the granulation tissue was obviously higher than that of the SA-E-C group, and the dermis of the skin of this group began to form. At day 11, the skin of the control group and the SA-C group had scab tissue, the SA-E-C group began to form a relatively thick epidermis, and the density of the granulation tissue of the skin of the three groups all increased, but there was still inflammatory cell infiltration, and the arrangement of the tissue fibers was relatively disordered, the skin of the SA-E-D-C group formed a relatively thin epidermis, the tissue fibers arranged regularly and densely along the growth direction of the epidermis, and the inflammatory cells obviously decreased. At day 21, the skin of the control and SA-C groups had a relatively thick epidermis layer, and the arrangement of the tissue fibers began to become regular, the thickness of the epidermis layer of the skin of the SA-E-C and SA-E-D-C groups further decreased, the density of the fibrous tissue further increased, and was higher than that of the control and SA-C groups, and a relatively mature dermis layer was formed, among which the epidermis layer of the skin of the SA-E-D-C group was the most regular, blood vessels appeared in the dermis layer, the density of the tissue fibers was significantly higher than that of the SA-E-C group, and was in bundles and arranged regularly along the growth direction of the epidermis. The above results show that the wound dressings SA-E-C and SA-E-D-C have the functions of promoting the regeneration of granulation and epidermis, and can inhibit the proliferation of the epidermis, and the effect of SA-E-D-C is the most obvious.

[0066] The Masson staining results of the skin tissues of the SD rats are shown in Figure 4As shown, at day 4, the collagen fibers in the skin of the control group and the SA-C group were arranged in a scattered and sparse manner, and even the collagen fibers were broken. The collagen fibers in the skin of the SA-E-C group and the SA-E-D-C group began to arrange in a relatively dense manner, and the number of the collagen fibers in the skin of the SA-E-D-C group was obviously more than that in the other groups. At day 11, the number of the collagen fibers in the skin of the control group and the SA-C group further increased, but was still obviously less than that in the SA-E-C group and the SA-E-D-C group. The collagen fibers in the skin of the SA-E-C group and the SA-E-D-C group formed relatively thick fiber bundles and arranged in a dense and regular manner. The collagen fibers in the skin of the SA-E-D-C group interwove to form a network and arranged in parallel along the epidermis. At day 21, the collagen fibers in the skin of the control group and the SA-C group arranged in a scattered manner. The density of the collagen fibers in the skin of the SA-E-C group and the SA-E-D-C group was obviously greater than that in the other two groups and arranged regularly under the epidermis. The collagen fibers in the skin of the SA-E-D-C group formed thicker fiber bundles and presented a wave-like fluctuation. The above results prove that the skin dressings SA-E-C and SA-E-D-C can promote the formation of collagen fibers at the skin wound site, and the effect of the SA-E-D-C is more significant.

[0067] The generation rate of the collagen fibers in the Masson staining results is shown in the following table: Figure 5 As shown, the generation rate of the collagen fibers in the skin of the SA-E-C group and the SA-E-D-C group at each time point was obviously greater than that in the blank group and the SA-C group, and the generation rate of the collagen in the SA-E-D-C group was the highest. At day 4, the generation rate of the collagen in the blank group and the SA-C group was lower than 10%, while the generation rate of the collagen in the skin of the SA-E-C group and the SA-E-D-C group reached about 20% and 40%, respectively. At day 11, the generation rate of the collagen in the skin of each group showed an upward trend, but the generation rate of the collagen fibers in the blank group and the SA-C group was obviously lower than that in the SA-E-C group and the SA-E-D-C group, and the generation rate of the collagen in the SA-E-D-C group was obviously higher than that in the other three groups. At day 21, the generation rate of the collagen in the blank group and the SA-C group was about 65%, while the generation rate of the collagen in the skin of the SA-E-C group and the SA-E-D-C group was close to 75% and 80%, respectively. The above results prove that the skin dressings SA-E-C and SA-E-D-C can obviously improve the generation rate of the collagen fibers at the skin wound site, and the promoting effect of the SA-E-D-C is more significant.

[0068] The above examples are only used to illustrate the technical solutions of the present application, and in the specific implementation and application process of the present application, the technical solutions can be changed by the person skilled in the art according to the specific experimental conditions and requirements, and are not used to limit the present application. Any within the spirit and principles of the present application shall be included in the protection scope of the present application.

Claims

1. A hydrogel with skin damage repair function, characterized in that, The hydrogel comprises the following components: sodium alginate, europium ions (Eu). 3+ Collagen polypeptide; the sequence of the collagen polypeptide is: (GXY) m1 -Collagen functional sequence- (GXY) m2 -(D) n ; Wherein, G is glycine, and D is aspartic acid; X and Y are selected from the same or different amino acids; Both m1 and m2 are integers greater than or equal to 1, and the sum of m1 and m2 is greater than or equal to 6; n is any integer greater than or equal to 2; The collagen functional sequence is selected from any one of the following: integrin binding site GFOGER, integrin binding site GROGER, integrin binding site GLOGER, integrin binding site GMOGER, integrin binding site GAOGER, integrin binding site GLKGEN, integrin binding site GLOGEN, DDR binding site GVMGFO, MMP restriction site GPLGIAGITGAR, MMP restriction site GPQGIAGQRGVV, MMP restriction site GPQGLLGAOGIL, MMP restriction site GPQGLAGQRGIV, heparin binding site GRPGKRGKQGQK, and fibronectin binding site GLPGQRGER.

2. The hydrogel as described in claim 1, characterized in that, X is proline (P), and Y is hydroxyproline (O); or both X and Y are hydroxyproline (O).

3. The hydrogel as described in claim 2, characterized in that, The sequence of the collagen polypeptide is: (GPO)3-GFOGER-(GPO)3-(D)2.

4. The hydrogel according to claim 3, characterized in that, The sodium alginate and europium ions (Eu) 3+ The concentration ratio of collagen peptides was 40 mg / mL: 2 mM: 6 mM.

5. The hydrogel as described in claim 4, characterized in that, The hydrogel is prepared by dissolving sodium alginate in a collagen polypeptide solution to obtain a mixed solution of collagen polypeptide and sodium alginate; this mixed solution is then reacted with Eu... 3+ Hydrogels were prepared by mixing solutions of equal volume.

6. A skin wound dressing, characterized in that, The skin wound dressing is prepared by 3D printing technology from the hydrogel described in any one of claims 1-5.

7. The method for preparing the skin wound dressing as described in claim 6, characterized in that, The method includes the following steps: (1) Prepare the hydrogel according to any one of claims 1-5; (2) 3D printing: The hydrogel prepared in step (1) is used as bio-ink to print the gel material using 3D printing technology. CaCl2 solution is atomized and dropped onto the surface of the 3D printed gel material to obtain a skin wound dressing.

8. The method for preparing the skin wound dressing as described in claim 7, characterized in that, In step (2), the 3D printing technology uses an extrusion 3D printer.

9. The method for preparing the skin wound dressing as described in claim 7, characterized in that, The concentration of the CaCl2 solution used in step (2) is 170 mg / mL.

10. The use of the hydrogel of claim 1 or the skin wound dressing of claim 6 in the preparation of skin damage repair products.

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