CN115197442B - Injectable self-healing hydrogel dressing for treating gastric perforation, preparation method and application - Google Patents

Abstract

The present invention relates to an injectable self-healing hydrogel dressing for treating gastric perforation, wherein the hydrogel is a triblock polymer PTOPT thermosensitive block dissolved in a pH 2.5 solution, which is self-assembled by temperature induction to form micelles containing ortho-amino groups and aromatic ring components, and a PEG long chain is used as a bridge to connect the micelles to form a three-dimensional network structure. The hydrogel has the characteristics of injectable, thermosensitive, self-healing, adhesion, anti-bacterial adhesion and biocompatibility. It can play an active and effective therapeutic effect in the gastric perforation model and is a promising hydrogel dressing.

Description

Injectable self-healing hydrogel dressing for treating gastric perforation, preparation method and use thereof

Technical Field

The invention relates to the field of biomedical materials, and in particular to an injectable self-healing hydrogel dressing for treating gastric perforation, a preparation method and application thereof.

Background technique

Hydrogels are promising natural extracellular matrix substitutes with exciting prospects in medical biomaterials. Injectable hydrogels, in particular, can minimize biological invasion and pain during treatment when treating wounds where hydrogels cannot be placed directly (such as irregular wounds in the body). At present, researchers have designed and developed injectable hydrogels for wound dressings through physical or chemical methods. However, most hydrogels lack adaptability to some complex biological environments, especially to the acidic gastric environment. Because most hydrogels will be hydrolyzed under such strong acidic conditions, resulting in damage to the hydrogel and loss of its original therapeutic function, it may even cause various inflammatory reactions, delay wound healing, and in severe cases may lead to death. Therefore, in the clinical treatment of gastric perforation, it is necessary to develop and design injectable hydrogels that can adapt to the complex environment of the stomach.

Among the many injectable hydrogel wound dressings, self-healing hydrogels with self-healing ability can self-repair after being subjected to external forces, which can greatly extend their service life in practical applications. The application of injectable hydrogels with self-healing properties in biomedical materials has become a research hotspot. Therefore, it is necessary to design and develop a bioadaptive injectable self-healing hydrogel to promote the healing of gastric perforation.

Summary of the invention

The purpose of the present invention is to solve the current lack of bioadaptive injectable self-healing hydrogel and to be used for the treatment of gastric perforation.

The objective of the present invention is achieved through the following technical solutions:

A method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation comprises the following steps:

S1. Synthesis of Pentafluorophenol Acrylate (PFPA)

Pentafluorophenol and dichloromethane are mixed, triethylamine and acryloyl chloride are added dropwise under ice bath conditions, and then reacted at room temperature, filtered, concentrated, washed, dried, and purified to obtain transparent liquid pentafluorophenol acrylate (PFPA);

S2. Synthesis of 2-((3-oxo-3-(phenylethylamino)propyl)amino)ethyl)carbamate (TBC)

Phenylethylamine and tert-butyl N-(2-aminoethyl)carbamate were dissolved in anhydrous ethanol, stirred vigorously, reacted, vacuum dried and purified to obtain an orange oily liquid 2-((3-oxo-3-(phenylethylamino)propyl)amino)ethyl)carbamate (TBC);

S3. Synthesis of macromolecular chain transfer agent PEG-CTA

Trithioester, 2-(dodecyl trithiocarbonate)-2-isobutyric acid (RAFT ₃₆₅ ) and oxalyl chloride were added into a three-necked flask and dissolved in anhydrous dichloromethane for reaction. After the reaction, the remaining oxalyl chloride and dichloromethane were dried by spin drying, and then PEG and anhydrous dichloromethane were added for reaction. After the reaction, the mixture was precipitated and dried to obtain a light yellow product PEG-CTA.

S4. Synthesis of triblock polymer Poly(EG-co-PFPA)-b-PEO-b-Poly(EG-co-PFPA) (GPOGP)

PEG-CTA prepared by S3, oligoethylene glycol methacrylate (PEG-MA), PFPA prepared by S1 and azobisisobutyronitrile (AIBN) were dissolved in 1,4-dioxane, and nitrogen was bubbled into the entire reaction system for reaction; after the reaction was completed, a white solid triblock polymer Poly(EG-co-PFPA)-b-PEO-b-Poly(EG-co-PFPA)(GPOGP) was obtained after precipitation, filtration, purification and drying;

S5. Synthesis of the precursor triblock polymer Poly(EG-co-TBC)-b-PEO-b-Poly(EG-co-TBC)(PTOPT-Boc)

The TBC prepared in S2 and the triblock polymer GPOGP prepared in S4 were dissolved in tetrahydrofuran, and nitrogen was bubbled into the reaction system for reaction; after the reaction, a slightly yellow solid PTOPT-Boc was obtained by precipitation, purification, filtration and drying;

S6. Preparation of PTOPT Polymer

The precursor triblock polymer PTOPT-Boc is dissolved in dichloromethane, and then trifluoroacetic acid is added to the solution to react at room temperature. After the reaction is completed, the PTOPT polymer is obtained by precipitation, purification, and drying.

S7. Preparation of hydrogel dressing

Dissolving PTOPT polymer in acidic ice water yields a biocompatible, injectable, self-healing hydrogel dressing.

Specifically, in S1, pentafluorophenol and dichloromethane are added to a three-necked flask, and then triethylamine and acryloyl chloride are added dropwise at -20 to 10°C for 20 to 60 minutes; then the ice bath is removed, and the reaction is carried out at 20 to 40°C for 3 to 8 hours, and triethylamine hydrochloride is removed by filtration using a Buchner funnel, and the filtrate is concentrated, and then washed with HCl solution, saturated NaHCO ₃ and brine; the organic phase is dried and dehydrated with NaSO ₄ or MgSO ₄ , concentrated, and petroleum ether, dichloromethane or n-hexane is used as an eluent, and the residual liquid is purified by column chromatography to obtain a transparent liquid; wherein the mass ratio of pentafluorophenol to acryloyl chloride is (5 to 10): (3 to 5). The amount of dichloromethane added is 50 to 100 mL; the concentration of HCl is 0.1 mol/L to 1 mol/L.

Specifically, in S2, phenethylamine and tert-butyl N-(2-aminoethyl)carbamate are dissolved in 10-50 mL of anhydrous ethanol and stirred vigorously; the mixture is stirred under reflux at 60-80° C. and reacted for 24-48 hours; the solvent ethanol is dried in a vacuum; and the residual liquid is purified by column chromatography using petroleum ether, dichloromethane, ethyl acetate or methanol as an eluent to obtain a TBC product; wherein the mass ratio of phenethylamine to tert-butyl N-(2-aminoethyl)carbamate is (2-10):(3-9).

Specifically, in S3, the mass ratio of RAFT ₃₆₅ , oxalyl chloride and PEG is (1-3): (3-7): (5-20); wherein the molecular weight of PEG is between 2000 and 40000; and the amounts of anhydrous dichloromethane added for the first and second times are 5-20 mL and 50-200 mL, respectively.

Specifically, in S4, after the reaction is completed, the solution is added dropwise into ether or n-hexane for precipitation, and the purification process is repeated twice; the product is filtered and dried overnight under vacuum to obtain a white solid GPOGP; wherein the mass ratio of PEG-CTA, oligoethylene glycol methacrylate (PEG-MA), PFPA, azobisisobutyronitrile (AIBN) and 1,4-dioxane is (1-3): (2-8): (0.5-2): (8-30).

Specifically, in S5, after the reaction is completed, the reaction solution is added dropwise to ether or n-hexane precipitant for precipitation, and the purification process is repeated twice; the product is filtered and dried under vacuum overnight to obtain a slightly yellow solid PTOPT-Boc; wherein the mass ratio of TBC, GPOGP and tetrahydrofuran is (1-3): (2-10): (20-50).

Specifically, in S6, the mass ratio of PTOPT-Boc, dichloromethane and trifluoroacetic acid is (1-3): (10-20): (1-2).

Specifically, in S7, the prepared triblock polymer PTOPT with a concentration of 50 mg/mL to 200 mg/mL is dissolved in a solution of acetic acid/potassium nitrate or hydrochloric acid with a pH of 2.5, and placed in a refrigerator for 10 to 30 hours to prepare a hydrogel dressing.

The injectable self-healing hydrogel dressing for treating gastric perforation prepared by the above method has good biocompatibility and can effectively reduce the risk during use.

The prepared hydrogel dressing was used in the gastric perforation wound model and other complex wound healing.

The present invention develops a hydrogel (PTOPT) adapted to the gastric environment as an in vivo wound dressing. By adapting to physiological conditions, the hydrogel dressing can be simply injected at the target site through the supramolecular self-assembly of ABA triblock copolymers. The formed supramolecular hydrogel can adapt to the gastric environment, and through the synergistic interaction of hydrophobic and cationic-π, it obtains rapid self-healing repair properties, reduces the risk of damage during use, and effectively extends the service life of the material. At the same time, the PTOPT hydrogel dressing can also effectively prevent the accumulation of microorganisms on its surface and has excellent anti-bacterial adhesion properties. And we also measured the cytotoxicity of the hydrogel by CCK-8, and the hydrogel had minimal toxicity to gastric mucosal cells and L929 cells. And the hydrogel was used to perform 2D/3D cell culture on gastric mucosal cells and L929 cells, and the data showed that the hydrogel had good biocompatibility. In addition, compared with traditional gastric perforation treatment, PTOPT hydrogel dressing has state-of-the-art advantages, including simplified surgical procedures, reduced postoperative adhesion and promoted angiogenesis. The design of our PTOPT hydrogel provides useful insights into the development of bio-environment-adaptive functional materials for various biomedical applications.

The structural formula of PFPA prepared by S1 of the present invention is shown below:

The structural formula of TBC prepared by S2 of the present invention is shown below:

The structural formula of the macromolecular chain transfer agent PEG-CTA prepared by S3 of the present invention is shown below:

The structural formula of the triblock polymer GPOGP with active ester prepared by S4 of the present invention is as follows:

The structural formula of the precursor triblock polymer PTOPT-Boc prepared by S5 of the present invention is as follows:

The structural formula of the target triblock polymer PTOPT prepared by S6 of the present invention is as follows:

The beneficial effects of the present invention are:

(1) The PTOPT hydrogel in the present invention has good temperature sensitivity, and the sol-gel transition temperature of the hydrogel is about 22°C, which is lower than the human body temperature (~37°C). It can form a stable gel under the induction of human body temperature without the need for other complex external stimulation.

(2) The PTOPT hydrogel in the present invention has a fascinating injectable property. When the temperature is below 22°C, the hydrogel is in a sol state. The liquid-like polymer solution can be well injected into the target site through a syringe and induced to become a stable hydrogel by body temperature. In addition, the relatively shear thinning behavior of the hydrogel also shows that even a stable hydrogel has a fascinating injectability, which can minimize biological invasion and pain during treatment.

(3) The PTOPT hydrogel in the present invention has excellent gastric environment-adaptive self-healing properties. In an environment with a pH of 2.5 that matches the gastric environment, a cation-π interaction occurs between the protonated amino groups in the hydrogel and the aromatic benzene rings, enabling the hydrogel to quickly self-heal after being damaged, thereby extending the service life of the material and reducing the risks during use.

(4) The PTOPT hydrogel in the present invention has good antibacterial adhesion properties against Escherichia coli (E. coli) and Staphylococcus aureus (S. aureus). It is crucial to prevent microorganisms from forming microbial films on the surface of implanted biomaterials. The presence of PEG segments in the hydrogel will form a strong hydration layer at the wound site, acting as a barrier to bacteria and effectively preventing bacterial adhesion. This can effectively prevent the immune response or inflammation that may be caused by the microbial film, which inhibits the healing process.

(5) The PTOPT hydrogel of the present invention exhibits good adhesion effects due to cation-π electrostatic interactions, hydrogen bonds, π-π stacking and hydrophobic interactions with the tissue surface. When the hydrogel forms a gel in situ on wounds of different shapes, especially in a wet and dynamic human body environment, it will be a promising wound dressing.

(6) The PTOPT hydrogel of the present invention has good biocompatibility. In vitro cytotoxicity shows that the PTOPT hydrogel is non-toxic and can promote cell growth and proliferation. In addition, the PTOPT hydrogel was used for 2D/3D cell culture of mouse fibroblasts (L 929) and rat gastric epithelial cells (GES), indicating that the PTOPT hydrogel did not reduce cell proliferation and had good biocompatibility.

BRIEF DESCRIPTION OF THE DRAWINGS

Figure 1 (a) PTOPT hydrogel modulus changes with temperature (4°C to 45°C); (b) PTOPT hydrogel modulus changes under three thermal cycles of heating (37°C) and cooling (12°C);

FIG2 is a sample of the hydrogel dressing PTOPT liquefied sample injected into 37° C. deionized water; (hydrogel stained with Rhodamine B);

Figure 3 (a) Modulus changes with increasing strain (0.1%-1000%) (left) and instantaneous recovery after 1000% strain deformation (right); (b) Dynamic strain cycle test of PTOPT hydrogel at 37°C (γ = 1% or 300%) showing self-healing behavior; (c) Demonstration of the self-healing properties of PTOPT hydrogel;

Figure 4 (a) Colony survival rate images of glass slides and PTOPT hydrogel-modified glass slides after 3 hours of co-culture of E. coli and Staphylococcus aureus; (B) Quantitative data of E. coli and Staphylococcus aureus adhesion on different glass slides;

FIG5 (a) Stress-strain curves of the PTOPT hydrogel of the present invention on different tissues; (b) Adhesion strength of the PTOPT hydrogel on different tissues;

Figure 6 (a) Cell viability of L929 cells after 1 and 7 days of culture with PTOPT hydrogels; (b) Cell viability of GES cells after 1 and 7 days of culture with PTOPT hydrogels, results are shown as mean ± SD (n = 6); (c), (d) 2D and 3D culture of L929 cells using 15 w/v% hydrogels. Representative live and dead staining of L929 cells on day 1 after culture with hydrogels. (e), (f) 2D and 3D culture of GES cells using 15 w/v% hydrogels. Representative live and dead staining of GES cells on day 1 after culture with hydrogels.

Figure 7 (a) Schematic diagram of gastric perforation treatment; (b) Optical photograph of gastric perforation treatment; (c) Optical photograph of the stomach 7 days after gastric perforation treatment; (d) Tissue adhesion after surgery; (e) H&E staining of the stomach 7 days after gastric perforation treatment. Scale bar: 1000 μm.

Figure 8 Representative immunofluorescence images of (a) CD31 and (b) CD34 staining at the wound edge. Brown staining represents blood vessels. Scale bar represents 50 μm. (c) Quantification of CD31 and CD34 staining.

Detailed ways

The technical solution of the present invention is further described in detail below in conjunction with the accompanying drawings, but the protection scope of the present invention is not limited to the following.

Example 1

S1: Synthesis of pentafluorophenol acrylate (PFPA): Add pentafluorophenol (5.52 g, 30 mmol) and 50 mL of dichloromethane to a 100 mL three-necked flask. Then, triethylamine (3.24 g, 32 mmol) and acryloyl chloride (2.9 g, 32 mmol) were added dropwise under ice bath conditions for more than 30 minutes. Then, the ice bath was removed and the reaction was carried out at 25°C for 4 hours. The triethylamine hydrochloride was removed by filtration using a Buchner funnel, and the filtrate was concentrated. Then, it was washed with 0.1 M HCl solution (1×75 mL), saturated NaHCO ₃ (1×75 mL) and brine (1×75 mL). The organic phase was dried with MgSO ₄ to remove water and concentrated. Petroleum ether was used as the eluent, and the residue was purified by column chromatography to obtain a transparent liquid.

S2: Synthesis of 2-((3-oxo-3-(phenylethylamino)propyl)amino)ethyl)carbamate (TBC) containing both amine (cation) and aromatic (π): Phenethylamine (5.25 g, 30 mmol) and tert-butyl N-(2-aminoethyl)carbamate (5.92 g, 37 mmol) were dissolved in 25 mL of anhydrous ethanol and stirred vigorously. The mixture was stirred under reflux at 80°C and reacted for 48 hours. The solvent ethanol was dried in a vacuum. The residue was purified by column chromatography using EAC and methanol (V:V=10:1, 1 vol.% TEA) as eluents to obtain the TBC product as an orange oily liquid.

S3: Synthesis of macromolecular chain transfer agent PEG-CTA: Add 1g trithioester 2-(dodecyl trithiocarbonate)-2-isobutyric acid (RAFT ₃₆₅ ) and 2ml oxalyl chloride to a three-necked flask and dissolve in 5mL anhydrous dichloromethane for reaction. After the reaction is completed, the remaining oxalyl chloride and dichloromethane in the flask are dried by spin drying. Then add 5g of PEG (Mn 20000) and 50mL anhydrous dichloromethane for reaction. After the reaction is completed, n-hexane is used as a precipitant for precipitation and dried to obtain a light yellow product PEG-CTA.

S4: Synthesis of triblock polymer Poly(EG-co-PFPA) -b-PEO-b-Poly(EG-co-PFPA) (GPOGP) with active ester by RAFT polymerization: PEG-CTA (0.518 g, 0.025 mmol), PEG-MA (1.018 g, 9 mmol), PFPA (0.238 g, 1 mmol) and AIBN (0.002 g, 0.0125 mmol) were dissolved in 8 mL of 1,4-dioxane. Nitrogen was bubbled through the entire reaction system for 15 minutes and reacted at 70°C for 12 hours. The solution was then added drop by drop to a large amount of ether for precipitation, and the purification process was repeated twice. The product was filtered and dried overnight under vacuum to obtain a white solid.

S5: Synthesis of the precursor triblock polymer Poly(EG-co-TBC)-b-PEO-b-Poly(EG-co-TBC) (PTOPT-Boc) by transesterification of TBC with a triblock polymer GPOGP having an active ester: TBC (0.67 g, 2 mmol) and GPOGP (1.54 g) were dissolved in 20 mL of tetrahydrofuran, and nitrogen was bubbled into the reaction system for 20 minutes. The reaction was carried out in an oil bath at 50°C overnight. After the reaction was completed, the reaction solution was added drop by drop to a large amount of ether for precipitation, and the purification process was repeated twice. The product was filtered and dried overnight under vacuum to obtain a slightly yellow solid.

S6: The precursor triblock polymer PTOPT-Boc was hydrolyzed by trifluoroacetic acid to obtain a PTOPT polymer: the precursor triblock polymer PTOPT-Boc was dissolved in 10 ml of dichloromethane, and then 2 ml of trifluoroacetic acid was added to the solution, and the reaction was carried out at 25°C for 4 hours. After the reaction was completed, it was purified twice by ether precipitation and dried in a vacuum oven at room temperature overnight to obtain the target product triblock polymer PTOPT.

S7: Dissolve the PTOPT polymer in ice water at pH = 2.5 to obtain a biocompatible injectable self-healing hydrogel: Dissolve the prepared triblock polymer PTOPT at a mass fraction of 15% in a buffer solution of 100 mmol acetic acid and 250 mmol KNO ₃ at pH 2.5 and place in a refrigerator for 12 hours.

The hydrogel wound dressing PTOPT prepared by the method of the present invention, which is injectable, thermosensitive, self-healing, adhesive, anti-bacterial and biocompatible, can play an active and effective therapeutic effect in the gastric perforation model. The following is a detailed analysis based on the experimental data.

As shown in Figure 1(a), the prepared PTOPT hydrogel exhibits excellent thermal responsiveness. The storage modulus (G′) and loss modulus (G″) of the hydrogel intersect at 22°C. When the temperature is below 22°C, G′ is greater than G″, and the PTOPT mixture exhibits the properties of a sol. When the temperature of the solution exceeds 22°C, G′ will intersect with G″ and exceed it, and the solution is in a gel state. Therefore, when heated to 37°C, the PLOPL solution can become an independent gel within 1 minute. This shows that the hydrogel has a clear sol-gel transition temperature (TSG). As shown in Figure 1(b), this gel-sol-gel transition is completely reversible. When the temperature is below the TSG temperature (12°C), G′ is less than G″ and is in a sol state. After the temperature is immediately raised to above the TSG temperature (37°C), G′ is less than G″ and is in a gel state. And it can be repeated for several cycles. These data show that the hydrogel has good temperature-sensitive properties.

Figure 2 shows the injectability of the PTOPT hydrogel in Example 1. Using a 23G x 3/4″ syringe, the polymer solution stored at a temperature below TSG (4°C) was easily injected into a 37°C water bath and immediately formed a stable hydrogel.

FIG3 shows the self-healing properties of the PTOPT hydrogel in Example 1. As shown in FIG3(a), within the strain range of 0.1% to 10%, G′ and G″ remain essentially unchanged, and G′ is greater than G″, showing gel properties. When the strain increases to 25%, G′ and G″ intersect, and this strain is the critical strain of the hydrogel. Further increasing the strain, G″ is greater than G′, showing sol-like properties. This is because when the strain exceeds the critical strain, the polymer chains are destroyed and the three-dimensional network of the hydrogel is destroyed. However, after the three-dimensional network of the hydrogel was destroyed under large strain (γ = 1000%), a small strain (γ = 1%) was immediately applied, and G′ and G″ quickly recovered to the initial values (the self-recovery efficiency reached 100% within 3 minutes). This shows that after the large strain is removed, the mechanical properties of the hydrogel sample will recover quickly. In addition, continuous step strain changes between 300% and 1% were performed to evaluate the healing ability of the hydrogel. As shown in Figure 3(b), when a large destructive strain of 300% was applied for 50 seconds, the G″ of the hydrogel was greater than G′, indicating that the hydrogel network was destroyed and a gel-sol transition occurred. However, when the hydrogel was under a lower strain (γ = 1%) for several seconds, the G′ of the hydrogel quickly recovered to the initial value, indicating that the broken hydrogel network can be rebuilt through reversible cation-π interactions. In addition, the self-healing properties of the hydrogel can also be observed macroscopically. As shown in Figure 3(c), a circular hydrogel is cut into two semicircles, and then the two semicircles are placed together. After 3 minutes of contact without external stimulation, the two semicircles heal themselves into a whole and can support their own weight.

FIG4 shows the antibacterial adhesion performance of the PTOPT hydrogel in Example 1. As shown in FIG4(a), the colony counts of Escherichia coli and Staphylococcus aureus on the blank glass slide are much higher than those on the glass slide modified with hydrogel, which indicates that bacteria can easily adhere to the surface of the glass slide and form a microbial film, while the glass slide modified with PTOPT hydrogel has good antibacterial adhesion performance. This shows that bacteria can easily adhere to the surface of the glass slide and form a microbial film, while the glass slide modified with PTOPT hydrogel has excellent anti-adhesion properties and can effectively prevent the formation of microbial films. As shown in FIG4(b), the quantitative data of the colonies show that there are far fewer bacteria on the glass slide modified with hydrogel, indicating that it has excellent antibacterial adhesion properties. This is due to the presence of the PEG segment in the hydrogel, which can form a strong hydration layer on the glass slide, acting as a barrier to bacteria and effectively preventing bacterial adhesion.

FIG5 shows the tissue adhesion performance of the PTOPT hydrogel in Example 1. As shown in FIG5(a) and (b), it can be seen that the adhesion strength of the hydrogel to pig intestine, pig stomach, pig skin and pig heart is 15.3, 2.8, 20.5 and 3.3 kPa respectively, which shows that the hydrogel has good adhesion strength to various tissues. The hydrogel has good adhesion to various tissues due to the ordered cation-π sequence in the hydrogel molecular structure. The hydrogel adhesion mechanism diagram is shown in FIG3-8. The hydrogel and the tissue surface undergo cation-π mediated electrostatic interaction, hydrogen bonding, π-π stacking and hydrophobic interaction, and exhibit good adhesion. These results show that when the hydrogel forms a gel in situ on wounds of different shapes, especially in a wet and dynamic human body environment, it will be a promising wound dressing.

FIG6 shows the biocompatibility of the PTOPT hydrogel in Example 1. As can be seen from FIG6(a) and (b), within a certain concentration range, the viability of L 929 cells and GES cells is almost stable, and all activities are maintained at around 90% or above. In vitro cytotoxicity shows that the PTOPT hydrogel is non-toxic and can promote cell growth and proliferation. In addition, we examined the cell compatibility of the PTOPT hydrogel by using it for 2D/3D cell culture of L 929 cells and GES cells. Cells without any treatment were used as normal controls. As shown in FIG6(c), (d), (e) and (f), after 24 hours of culture, very few dead cells were observed in both the 2D culture group and the 3D group. There was no significant difference in the evaluation of live cell density between the normal group and the hydrogel group, indicating that the PTOPT hydrogel did not reduce cell proliferation and had good biocompatibility.

FIG7 shows the use of PTOPT hydrogel in Example 1 to treat gastric perforation. As shown in FIG7(a), an in vivo rat gastric perforation model was constructed to demonstrate that PTOPT hydrogel can be used as a sealant for treating gastrointestinal perforation. According to the above method, a 5 mm perforation was made in the gastric antrum, as shown in FIG7(b). On the 7th day after surgery, macroscopic images of the harvested gastric tissue incision showed that the wound area treated with PTOPT hydrogel was significantly reduced, indicating that the wound healing effect was good, as shown in FIG7(c). We also took a representative macroscopic image of abdominal adhesions. There was almost no abdominal adhesion treated with PTOPT hydrogel, as shown in FIG7(d). H&E staining showed that the PTOPT hydrogel treatment was almost healed, as shown in FIG7(e).

FIG8 shows the angiogenesis around the wound of gastric perforation treated with PTOPT hydrogel in Example 1. We evaluated the fluorescence intensity of the wound edge by IF staining of CD31 and CD34 (markers of endothelial cells and angiogenesis). As shown in FIG8 , in the results of CD31 and CD34, the intensity of positively stained capillaries in the PTOPT hydrogel group was significantly higher than that in the suture group (P<0.01) and the incision group (P<0.05).

The experimental results show that the hydrogel wound dressing PTOPT prepared by the method of the present invention, which is injectable, thermosensitive, self-healing, adhesive, anti-bacterial adhesion and biocompatible, can play an active and effective therapeutic effect in the gastric perforation model. The results of postoperative tissue adhesion, granulation tissue growth, and angiogenesis around the wound were further evaluated, and the hydrogel showed good therapeutic effects. In addition, these hydrogels have stable rheological properties, moduli close to human soft tissues, good adhesion properties, self-healing properties, anti-bacterial adhesion, biocompatibility, etc. Therefore, the hydrogel dressing PTOPT has a good application prospect in the treatment of gastric perforation healing.

The above is only a preferred embodiment of the present invention. It should be understood that the present invention is not limited to the form disclosed herein, and should not be regarded as excluding other embodiments, but can be used in various other combinations, modifications and replacements, and can be within the scope of the concept described herein. The changes and modifications made by those skilled in the art do not deviate from the spirit and scope of the present invention and should be within the scope of protection of the claims attached to the present invention.

Claims (10)

1. A method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation, characterized in that it comprises the following steps: S1. Synthesis of Pentafluorophenol Acrylate (PFPA) Pentafluorophenol and dichloromethane are mixed, triethylamine and acryloyl chloride are added dropwise under ice bath conditions, and then reacted at room temperature, filtered, concentrated, washed, dried, and purified to obtain transparent liquid pentafluorophenol acrylate (PFPA); S2. Synthesis of 2-((3-oxo-3-(phenylethylamino)propyl)amino)ethyl)carbamate (TBC) Phenylethylamine and tert-butyl N-(2-aminoethyl)carbamate were dissolved in anhydrous ethanol, stirred vigorously, reacted, vacuum dried and purified to obtain an orange oily liquid 2-((3-oxo-3-(phenylethylamino)propyl)amino)ethyl)carbamate (TBC); S3. Synthesis of macromolecular chain transfer agent PEG-CTA Trithioester, 2-(dodecyl trithiocarbonate)-2-isobutyric acid and oxalyl chloride were added into a three-necked flask and dissolved in anhydrous dichloromethane for reaction. After the reaction, the remaining oxalyl chloride and dichloromethane were dried by spin drying, and polyethylene glycol (PEG) and anhydrous dichloromethane were added for reaction. After the reaction, the mixture was precipitated and dried to obtain a light yellow product PEG-CTA. S4. Synthesis of triblock polymer Poly(EG-co-PFPA)-b-PEO-b-Poly(EG-co-PFPA)(GPOGP) PEG-CTA prepared by S3, oligoethylene glycol methacrylate (PEG-MA), PFPA prepared by S1 and azobisisobutyronitrile (AIBN) were dissolved in 1,4-dioxane, and nitrogen was bubbled into the entire reaction system for reaction; after the reaction was completed, a white solid triblock polymer was obtained after precipitation, filtration, purification and drying; S5. Synthesis of the precursor triblock polymer Poly(EO-co-TBC)-b-PEO-b-Poly(EO-co-TBC)(PTOPT-Boc) The TBC prepared in S2 and the triblock polymer GPOGP prepared in S4 were dissolved in tetrahydrofuran, and nitrogen was bubbled into the reaction system for reaction; after the reaction, a slightly yellow solid PTOPT-Boc was obtained by precipitation, purification, filtration and drying; S6. Preparation of PTOPT Polymer The precursor triblock polymer PTOPT-Boc is dissolved in dichloromethane, and then trifluoroacetic acid is added to the solution to react at room temperature. After the reaction is completed, the PTOPT polymer is obtained by precipitation, purification, and drying. S7. Preparation of hydrogel dressing Dissolving PTOPT polymer in acidic ice water yields a biocompatible, injectable, self-healing hydrogel dressing. 2. A method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S1, pentafluorophenol and dichloromethane are added to a three-necked flask, and then triethylamine and acryloyl chloride are added dropwise at -20 to 10°C for 20 to 60 minutes; then the ice bath is removed, and the mixture is reacted at 20 to 40°C for 3 to 8 hours, and the triethylamine hydrochloride is removed by filtration using a Buchner funnel, the filtrate is concentrated, and then washed with HCl solution, saturated _NaHCO3 and brine; the organic phase is dried and dehydrated with _NaSO4 or _MgSO4 , concentrated, and petroleum ether, dichloromethane or n-hexane is used as an eluent, and the residual liquid is purified by column chromatography to obtain a transparent liquid; wherein the mass ratio of pentafluorophenol to acryloyl chloride is (5 to 10): (3 to 5). 3. A method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S2, phenylethylamine and N-(2-aminoethyl) tert-butyl carbamate are dissolved in 10-50 mL of anhydrous ethanol and stirred vigorously; the mixture is stirred under reflux at 60-80° C. and reacted for 24-48 hours; the solvent ethanol is dried in a vacuum; and the residual liquid is purified by column chromatography using petroleum ether, dichloromethane, ethyl acetate or methanol as an eluent to obtain a TBC product; wherein the mass ratio of phenylethylamine to N-(2-aminoethyl) tert-butyl carbamate is (2-10):(3-9). 4. The method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S3, the mass ratio of 2-(dodecyl trithiocarbonate)-2-isobutyric acid, oxalyl chloride and PEG is (1-3): (3-7): (5-20). 5. The method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S4, after the reaction is completed, the solution is added dropwise into ether or n-hexane for precipitation, and the purification process is repeated twice; the product is filtered and dried under vacuum overnight to obtain a white solid GPOGP. 6. The method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S5, after the reaction is completed, the reaction solution is added drop by drop into ether or n-hexane precipitant for precipitation, and the purification process is repeated twice; the product is filtered and dried under vacuum overnight to obtain a slightly yellow solid PTOPT-Boc; wherein the mass ratio of TBC, GPOGP and tetrahydrofuran is (1-3): (2-10): (20-50). 7. The method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S6, the mass ratio of PTOPT-Boc, dichloromethane and trifluoroacetic acid is (1-3): (10-20): (1-2). 8. The method for preparing an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 1, characterized in that: in S7, the prepared triblock polymer PTOPT with a concentration of 50 mg/mL to 200 mg/mL is dissolved in a solution prepared with acetic acid/potassium nitrate or hydrochloric acid and the pH is 2.5, and the solution is placed in a refrigerator for 10 to 30 hours to prepare the hydrogel dressing. 9. An injectable self-healing hydrogel dressing for treating gastric perforation, characterized in that: the injectable self-healing hydrogel dressing for treating gastric perforation is prepared by the method according to any one of claims 1 to 8. 10. Use of an injectable self-healing hydrogel dressing for treating gastric perforation according to claim 9, characterized in that the hydrogel dressing is used in gastric perforation wound models and other complex wound healing dressings.