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CN118750659B - Preparation method of hydrogel with tissue repair and antioxidation functions and hydrogel thereof - Google Patents

Preparation method of hydrogel with tissue repair and antioxidation functions and hydrogel thereof Download PDF

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CN118750659B
CN118750659B CN202411241750.8A CN202411241750A CN118750659B CN 118750659 B CN118750659 B CN 118750659B CN 202411241750 A CN202411241750 A CN 202411241750A CN 118750659 B CN118750659 B CN 118750659B
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recombinant human
collagen
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hydrogel
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CN118750659A (en
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徐祥波
张昕
王树芳
陈西华
贺斌
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Institute Of Science And Technology National Health Commission
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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/04Macromolecular materials
    • A61L31/043Proteins; Polypeptides; Degradation products thereof
    • A61L31/044Collagen
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
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    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/14Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L31/145Hydrogels or hydrocolloids
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L31/00Materials for other surgical articles, e.g. stents, stent-grafts, shunts, surgical drapes, guide wires, materials for adhesion prevention, occluding devices, surgical gloves, tissue fixation devices
    • A61L31/14Materials characterised by their function or physical properties, e.g. injectable or lubricating compositions, shape-memory materials, surface modified materials
    • A61L31/16Biologically active materials, e.g. therapeutic substances
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
    • A61L2300/00Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices
    • A61L2300/40Biologically active materials used in bandages, wound dressings, absorbent pads or medical devices characterised by a specific therapeutic activity or mode of action
    • AHUMAN NECESSITIES
    • A61MEDICAL OR VETERINARY SCIENCE; HYGIENE
    • A61LMETHODS OR APPARATUS FOR STERILISING MATERIALS OR OBJECTS IN GENERAL; DISINFECTION, STERILISATION OR DEODORISATION OF AIR; CHEMICAL ASPECTS OF BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES; MATERIALS FOR BANDAGES, DRESSINGS, ABSORBENT PADS OR SURGICAL ARTICLES
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    • A61L2400/06Flowable or injectable implant compositions

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Abstract

The invention provides a preparation method of hydrogel with tissue repair and antioxidation functions and the hydrogel thereof, wherein the hydrogel is a solution comprising the following components: recombinant human type I collagen, recombinant human type III collagen, or a mixture of recombinant human type I collagen and recombinant human type III collagen, a solution of methacryloylated chitosan and total arasaponin containing a photoinitiator; the mass and volume concentration of the total saponins of panax notoginseng is not more than 0.6 percent. The hydrogel with tissue repair and antioxidation functions is suitable for various tissue repair, antibiosis and fibrosis prevention, and can simultaneously exhibit the functions of antioxidation and free radical removal.

Description

Preparation method of hydrogel with tissue repair and antioxidation functions and hydrogel thereof
Technical Field
The invention relates to the field of materials, in particular to a preparation method of hydrogel with tissue repair and antioxidation functions and the hydrogel.
Background
The decrease of endometrial receptivity caused by the damage of endometrium due to congenital or acquired diseases, operation and the like is one of the main reasons for infertility of women of childbearing age. Uterine cavity adhesions (intrauterine adhesion, IUA), also known AS Asherman's Syndrome (AS), are usually caused by mechanical injuries such AS induced abortion surgery, uterine curettage or infections, and are manifested AS endometrial basal layer lesions, which in turn cause tissue adhesions and fibrosis, ultimately leading to reduced fertility.
The main clinical treatment method is hysteroscopic surgery treatment, namely, the adhesion part is separated through surgery, and the method has high recurrence rate and is easy to cause secondary damage. Drug infusion or oral drug therapy also suffers from significant drawbacks such as: the active ingredients are easy to lose, and the method is only suitable for immediate injury treatment and can not solve the subsequent healing and repairing problems; and also easily causes post-wound adhesion, wound infection, and the like.
Active oxygen aggregation and angiogenesis disorder can be caused after injury of a wound site, so that healing is difficult, oxygen free radical removal, inflammatory reaction inhibition and angiogenesis induction are key to treatment of the wound site, and therefore a novel material capable of resisting oxidation and scavenging free radicals is needed to better promote wound healing.
Disclosure of Invention
In order to solve the problems, the invention provides a preparation method of hydrogel with tissue repair and antioxidation functions, wherein the hydrogel is a solution comprising the following components: a solution of recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen, comprising a photoinitiator of methacryloylated chitosan and total arasaponin, said solution having a viscosity of 3000±3000 x 15% mpa.s to 15000±15000 x 15% mpa.s, preferably, said viscosity of 3000±3000 x 10% mpa.s to 15000±15000 x 10% mpa.s, more preferably, said viscosity of 3000±3000 x 5% mpa.s to 15000±15000 x 5% mpa.s when tested at a shear rate of 1/s; when the solution is applied to corresponding tissues, ultraviolet light or visible blue light is used for external irradiation, and the hydrogel is prepared after the solution becomes gel; the mass and volume concentration of the total saponins of panax notoginseng is not more than 0.6 percent, namely the mass of the total saponins of panax notoginseng in the 100ml solution is not more than 0.6: 0.6 g.
The human collagen is divided into more than 28 kinds of collagen according to different tissue parts, physiological functions and molecular structures, wherein the collagen is mainly type I collagen and type III collagen which are closely related to the skin injury repair process and repair quality, and the type I collagen is mainly present in adult skin, tendons and bone tissues from the distribution; type III collagen is mainly present in infant skin or in the intima and intestinal tract. The collagen type I and III in human body maintains the normal tissue structure of skin, forms an extracellular matrix network structure, plays roles in supporting organs and protecting organisms, and is also related to cell attachment and cell migration. The balance of the two also determines the hardness and tenderness of the skin, as well as the degree of wound healing and scarring. Therefore, when the recombinant human collagen is used in tissue repair, the type I collagen and the type III collagen can effectively promote repair and tissue regeneration.
In one embodiment, the total saponins of panax notoginseng mass volume concentration is 0.4% -0.6%.
In one embodiment, the photoinitiator is phenyl-2, 4, 6-trimethylbenzoyl lithium phosphite or (2-hydroxy-2-methyl-1- [4- (2-hydroxyethoxy) phenyl ] -1-propanone.
In one embodiment, the hydrogel has a viscosity of 10025.85 mPa-3163.85 mPa s when tested at 37 ℃ and a shear rate of 1/s.
In one embodiment, when the mass volume concentration of the methacryloylated chitosan is 8%, i.e., when the 100 ml solution contains 8 g methacryloylated chitosan, and when the mass volume concentration of the recombinant human type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen is 30%, i.e., when the 100 ml solution contains 30 g recombinant type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen, the volume ratio of the methacryloylated chitosan to the recombinant type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen is 3:1 to 1:1.
In one embodiment, the volume ratio of the methacryloylated chitosan to the recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen is 1:1, and the total saponins of panax notoginseng mass volume concentration is 0.6%.
In one embodiment, a hydrogel for controlling endometrial damage is provided, the hydrogel prepared by the method described above.
In one embodiment, a hydrogel for preventing and treating intrauterine adhesion and thin endometrium is provided, which is prepared by the above method.
In one embodiment, there is provided a hydrogel for preventing and treating intrauterine adhesion and thin endometrium, the hydrogel is a solution comprising recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen, methacryloylated chitosan containing a photoinitiator, and total saponins of panax notoginseng, and when the solution is applied to corresponding tissues, the hydrogel is prepared by externally irradiating with ultraviolet light or visible blue light, and stopping after the solution becomes gel-like; when the viscosity of the solution is tested under the conditions of 37 ℃ and a shear rate of 1/s, the viscosity is 3000 mPa & s-15000 mPa & s; the mass and volume concentration of the total saponins of panax notoginseng is not more than 0.6 percent, namely the mass of the total saponins of panax notoginseng in the 100 ml solution is not more than 0.6: 0.6 g.
In one embodiment, when the mass volume concentration of the methacryloylated chitosan is 8%, when the mass volume concentration of the recombinant human type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen is 30%, the volume ratio of the methacryloylated chitosan to the recombinant human type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen is 1:1 to 1:1, and the total mass volume concentration of the pseudo-ginseng saponin is 0.6%.
Tissue damage can occur in various organs or tissues of the human body, and generally includes external wounds, bacterial infections, surgical side effects, and the like. If not treated effectively in time, the wound may be infected, healed slowly, scar hyperplasia and even necrosis. The invention provides a preparation method of hydrogel with tissue repair and antioxidation functions and hydrogel thereof based on methacryloyl chitosan, collagen and panax notoginseng saponins, the material can be injected to corresponding tissues in a liquid state, can be photocrosslinked and solidified to form gel under the irradiation of ultraviolet light or visible light, can be firmly attached to wounds, promotes the wound healing, and can simultaneously exhibit the functions of resisting oxidation and scavenging free radicals. The invention can be applied to various tissue repair, antibiosis and prevention of fibrosis, including viscera injury repair, endometrium repair, cartilage repair, skin superficial repair and the like.
Drawings
In order to more clearly illustrate the technical solutions of the embodiments of the present application, the drawings that are needed in the embodiments will be briefly described below, and it is obvious that the drawings in the following description are only some embodiments described in the present application, and other drawings may be obtained according to these drawings without inventive effort for a person skilled in the art.
FIG. 1 is a graph showing the effect of a hydrogel of the present invention on repairing rat endometrial lesions.
Detailed Description
In order that those skilled in the art will better understand the technical solutions of the present application, the present application will be further described with reference to the following examples, and it is apparent that the described examples are only some, but not all, examples of the present application. All other embodiments, which can be made by those skilled in the art based on the embodiments of the application without making any inventive effort, shall fall within the scope of the application. The application is further described below with reference to the drawings and examples.
Example one preparation of the raw materials of the invention
Preparation of methacryloylated Chitosan (CSMA)
Adding chitosan powder into 0.1 mol/L acetic acid solution, wherein the mass volume fraction is as follows: 0.5 And (5%) to 5%, and stirring at normal temperature until the mixture is completely dissolved. The chitosan solution is placed in a water bath at 60 ℃, methacrylic anhydride is added dropwise, wherein the gram weight ratio of chitosan to methacrylic anhydride is 1:2-1:3. After the dripping is finished, stirring and reacting at 60 ℃ for 6-12 h. After the reaction was completed, 0.5 mol/L NaOH solution was slowly added dropwise to the reaction solution, and the pH of the reaction solution was adjusted to be neutral. The neutral reactant solution was dialyzed 72 h against deionized water, changing water once every 6: 6 h. The dialyzed solution is frozen at the temperature of minus 80 ℃ and then is freeze-dried in vacuum for 72 h, and the obtained freeze-dried powder product is the methacryloyl chitosan.
Preparation of recombinant human collagen type I or III (RC) solution
Dissolving recombinant human collagen type I or III powder in PBS buffer solution, and stirring at 37deg.C until completely dissolving.
Preparation of Panax Notoginseng Saponins (PNS) solution
Dissolving Notoginseng radix total saponin powder in PBS buffer solution, and stirring at room temperature until it is completely dissolved.
Preparation of a precursor solution of Tetramethacryloylated chitosan-collagen+Panax notoginsenosides hydrogel (Hydrogel)
The methacryloylated chitosan powder was taken and dissolved in PBS buffer, with the addition of 0.25% photoinitiator, and stirred at ambient temperature for 12h until complete dissolution. Then adding the collagen solution and the total arasaponin solution into the mixture, and stirring the mixture at the temperature of 4 ℃ in a dark place. Wherein, the mass volume concentration of the methacryloyl chitosan is 8% (w/v), the mass volume concentration of the collagen is 30% (w/v), and the mass volume concentration of the total sanchi saponin solution is 10% (w/v).
Fifth, hydrogel preparation method
When the methacryloylated chitosan-collagen+panax notoginseng saponins hydrogel precursor solution is used for repairing the injury, the solution is injected and filled into corresponding tissues, then ultraviolet light (365 nm) or visible blue light (405 nm) is used for irradiating the corresponding tissues externally, and the filled solution is stopped after the filled solution becomes gel. At this time, the hydrogel is highly adhered in gel form, preventing tissue adhesion; meanwhile, the repairing effect of various components can be better exerted, and the repairing and regenerating of damaged tissues can be promoted. The material and the application method are suitable for repairing various wounds, especially wounds with irregular shapes, such as epidermis injury, endometrium injury, cartilage regeneration, vaginal mucosa damage and the like, and can remove excessive free radicals at the oxidative stress part of the wound. The mixed solution can be changed from liquid to gel rapidly under irradiation of ultraviolet light (365 nm) or visible blue light (405 nm).
EXAMPLE two viscosity test of methacryloylated chitosan-collagen solution at different volume ratios
The methacryloyl Chitosan (CSMA) -collagen (RC) solutions with different volume ratios are dripped on a rheometer sample stage, and the viscosity test is carried out under the conditions of 37 ℃ and the shearing rate of 1/s.
TABLE 1 CSMA to RC solution viscosity at different ratios
According to viscosity detection data, the change of the volume ratio of the methacryloyl chitosan and the collagen can directly influence the viscosity of the solution, when the ratio of CSMA to RC is more than 3:1, the viscosity is lower than 3000 mPa & s, and the viscosity is lower, the fluidity is strong, and the solution is unfavorable for residence in the uterine cavity; when the ratio of CSMA to RC is smaller than 1:1, the viscosity is higher than 15000 mPa.s, the viscosity is larger, the injectable type is difficult to achieve, the damaged tissue is difficult to be completely covered, and the treatment effect cannot be achieved. Therefore, the solution viscosity is considered to be suitable for being used as a drug loading platform for subsequent experiments when CSMA is rc=1-3:1.
Example three experiments of loading Panax notoginsenosides with different ratios on cell compatibility of composite hydrogels
In the volume proportion CSMA, RC=1:1 of methacryloylated chitosan-collagen hydrogel precursor solution, 10% of total sanchi saponin solution with mass volume concentration is loaded in different proportions to form 10% of total sanchi saponin hydrogel precursor solution with mass volume concentration of 10% of total sanchi saponin solution with mass volume concentration of 2%, 4%, 6%, 8% and 10% of total 100ml of solution, namely the total 100ml of solution contains 2 ml, 4 ml, 6 ml, 8 ml and 10 ml of total sanchi saponin solution with mass volume concentration of 10%.
Each group of precursor solutions was injected into a six-well plate, and after sterilization by uv irradiation for 30min, methacryloylated chitosan-collagen + total arasaponin hydrogels (denoted as 2% hydrogel, 4% hydrogel, 6% hydrogel, 8% hydrogel, 10% hydrogel, respectively) were formed. Each set of hydrogels and DMEM complete broth (DMEM basal medium, 10% fetal bovine serum, and 1% diabody, which is streptomycin and penicillin) were placed in 50mL sterile centrifuge tubes to give samples with a hydrogel concentration of 0.1 g/mL, and the centrifuge tubes with the samples were placed in a 37 ℃ 5% CO 2 incubator for 72 h to give a hydrogel extract. Human endometrial stromal cells HESCs were seeded at a density of 1X 10 3/100. Mu.L of cell suspension per well in 96-well plates and cultured in a 5% CO 2 incubator at 37℃for 24h. The medium was aspirated and 200. Mu.L of the extract per well was added and set as the experimental group. While the control group was further incubated with 200. Mu.L of DMEM complete medium per well. Three time points of 24h, 48 h and 72 h were simultaneously cultured for each group and 5 duplicate wells were set for each time point. For the measurement, the old medium was discarded, 100. Mu.L of DMEM complete medium and 10. Mu.L of CCK8 solution were added to each well, the mixture was blown up uniformly, and the absorbance was measured at 450 nm using an microplate reader after incubation at 37℃for 1.5 h.
The calculation formula of the cell viability (Cell Viability) is as follows:
The calculated average cell viability is shown in the following table, and it is found from the data in the table that the cell viability of the 8% hydrogel group after 72h and the cell viability of the 10% hydrogel group after 48 h are less than 95%, so that the total saponins loading ratio of pseudo-ginseng is considered to be unsuitable for normal growth of cells and subsequent in vivo experiments.
Therefore, when the loading ratio of the total saponins of panax notoginseng is not more than 8%, the methacryloylated chitosan-collagen + total saponins of panax notoginseng hydrogel has good cell compatibility, is suitable for being used on wounds and is beneficial to wound healing.
TABLE 2 influence of hydrogels with different loading ratios of Panax notoginsenosides on cell viability
Example IV Effect of loading Panax notoginsenosides at different concentrations on the antioxidant capacity of hydrogels
Oxidative stress refers to a state in which highly active molecules such as active oxygen and active nitrogen radicals are produced excessively in the body when the body is subjected to various harmful stimuli, the oxidation degree exceeds the oxide scavenging ability, and the oxidation system and the antioxidant system are dynamically unbalanced, thereby causing damage to cells and tissues. Research shows that the oxidative stress caused by unbalanced level of oxidation and reduction in the organism is the pathophysiological basis of a plurality of diseases, and the oxidation stress is defended, and the oxidation resistance system in the organism is mainly relied on. DPPH, diphenyl picrylphenylhydrazine, is a stable nitrogen-centered chromogenic radical that produces a characteristic absorption peak at 515 nm wavelengths. The antioxidant substance in the sample can reduce the absorbance of DPPH until the absorbance disappears, and the change amount of the absorbance is in linear relation with the content of the antioxidant substance in a certain interval.
In the volume proportion CSMA, RC=1:1, the methacryloylated chitosan-collagen hydrogel precursor solution is loaded with several kinds of total arasaponin solutions in different proportions to form the methacryloylated chitosan-collagen+total arasaponin hydrogel precursor solution with different proportions, wherein the loading proportion of the total arasaponin solution is 2%, 4% and 6%.
Each set of precursor solutions was injected into a six-well plate, cured by uv irradiation and sterilized for 30 min a, forming methacryloylated chitosan-collagen + total saponins of panax notoginseng hydrogels (2% hydrogel, 4% hydrogel, 6% hydrogel, respectively). And taking out each group of hydrogel, quick-freezing for 1h in a refrigerator at the temperature of minus 80 ℃, and then placing the hydrogel into a freeze vacuum dryer for vacuum drying for 48-72 h to prepare freeze-dried samples. The dried sample of 0.1 g is placed at the bottom of a centrifuge tube, added with 1mL PBS buffer, placed in a 37 ℃ incubator for incubation of 0.5-1 h, and then the extract is taken out for centrifugation (6000 rmp, 10 min), and the supernatant is taken. DPPH reagent with concentration of 0.04 mg/mL is prepared. 950. Mu.L DPPH reagent and 50. Mu.L sample leach supernatant were added to the EP tube. A blank of 950. Mu.L DPPH reagent and 50. Mu.L PBS buffer was used, and a control of 950. Mu.L DPPH reagent and 50. Mu.L absolute ethanol was used. After thoroughly mixing, 100 μl was added to a 96-well plate, and detection was performed with an enzyme-labeled instrument at wavelength 515 nm, and OD values were recorded, with 5 replicate wells per group.
DPPH radical clearance = (OD blank-OD sample+od control)/OD blank 100%
The DPPH radical scavenging rate of each group is shown in the following table, and it is found that the average DPPH radical scavenging rate of the 2% hydrogel group is only 1.1904%, and the oxidation stress state of the wound part is hardly improved. The 4% hydrogel and the 6% hydrogel have remarkable free radical scavenging effect, and can largely defend against oxidative stress. Therefore, when the loading ratio of the total saponins of panax notoginseng is more than 4%, the composite hydrogel can have better antioxidation effect.
TABLE 3 hydrogel DPPH radical scavenging Rate for different Panax notoginsenosides loading ratios
Example four effects of hydrogels of methacryloylated chitosan-collagen on the repair Capacity of rat endometrial lesions
An endometrial injury repair experiment was established for rats, and female SD rats, 8 weeks old, weighing about 250 g, and sexually mature, were selected and adapted for one week. After the rats are anesthetized by isoflurane, the abdomen is prepared for skin, and the abdomen is cut off to find out the Y-shaped uterus. The upper and lower ends of the uterus are clamped by hemostatic clips respectively, a 1mL syringe is inserted into the uterus of a rat, 95% alcohol is injected into the uterus to be full, and the uterus is washed with physiological saline for several times after 60 s. At this time, the methacryloylated chitosan-collagen + panax notoginseng saponins hydrogel precursor solutions (CRP 1, CRP2, CRP 3) at different concentrations were injected into the injured uterus, and then externally irradiated with ultraviolet light or blue light at about 60 s, which was seen to be gel-like. And (5) sterilizing after suturing, and continuing feeding for observation. An alcohol damage Control group (designated as Control) was also set. All rats were sacrificed 7 days post-surgery, uterine tissue was photographed and fixed in 4% paraformaldehyde at 48 h, and then observed after ethanol gradient dehydration, paraffin embedding, and slice staining.
As shown in figure 1, the uterus of the alcohol damage control group can be obviously blood stasis, and each group of uterus of CRP1, CRP2 and CRP3 has no obvious wound, and has little material residue in the uterus and is gel. From the H & E staining results, the rat uterine epithelium after alcohol injury is lost, the number of matrix cells is small, the matrix cells are loosely arranged, the rat uterine epithelium presents thin intima symptoms, and the repairing effect is poor. The CRP1, CRP2 and CRP3 have the advantages of repairing each group to a certain extent, increasing the endometrium thickness, increasing the number of matrix cells, and completely coating obvious glands and uterine cavity epithelium. Among them, CRP1 group repair effect was more remarkable. The results show that the CRP1 group, namely CSMA: RC=1:1, and the methacryloylated chitosan-collagen+Panax notoginsenosides hydrogel loaded with 6% of Panax notoginsenosides has good antioxidant capacity and also has the capacity of promoting endometrial injury repair and wound healing.
It is to be understood that this invention is not limited to the particular methodology, protocols, and materials described, as these may vary. It is also to be understood that the terminology used herein is for the purpose of describing particular embodiments only, and is not intended to limit the scope of the present invention which will be limited only by the appended claims.
Those skilled in the art will also recognize, or be able to ascertain using no more than routine experimentation, many equivalents to the specific embodiments of the invention described herein. Such equivalents are also encompassed by the appended claims.

Claims (9)

1. A solution for preparing a hydrogel with tissue repair and antioxidant functions, wherein the solution for preparing the hydrogel is a solution comprising: recombinant human type I collagen, recombinant human type III collagen or a mixture of recombinant human type I collagen and recombinant human type III collagen, a solution of methacryloylated chitosan and total saponins of panax notoginseng containing a photoinitiator, wherein the viscosity is 3000 mPa-15000 mPa-s when the solution is subjected to viscosity test under the conditions of 37 ℃ and a shear rate of 1/s; when the solution is applied to corresponding tissues, ultraviolet light or visible blue light is used for external irradiation, and the hydrogel is prepared after the solution becomes gel; the mass and volume concentration of the total saponins of panax notoginseng is 0.4% -0.6%.
2. The solution according to claim 1, wherein the photoinitiator is lithium phenyl-2, 4, 6-trimethylbenzoyl phosphite or 2-hydroxy-2-methyl-1- [4- (2-hydroxyethoxy) phenyl ] -1-propanone.
3. The solution according to claim 1, wherein the viscosity is 10025.85 mPa-3163.85 mPa s when the solution is subjected to a viscosity test at 37 ℃ at a shear rate of 1/s.
4. The solution of claim 1, wherein the 100ml solution comprises 8g methacryloylated chitosan and the 100ml solution comprises 30 g recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen when the mass volume concentration of the recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen is 30%, and the volume ratio of the methacryloylated chitosan to the recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen is 3:1 to 1:1.
5. The solution according to claim 4, wherein the volume ratio of the methacryloylated chitosan to the recombinant human type i collagen, recombinant human type iii collagen, or a mixture of recombinant human type i collagen and recombinant human type iii collagen is 1:1, and the total saponins of panax notoginseng mass volume concentration is 0.6%.
6. A hydrogel for preventing and treating endometrium injury, wherein the hydrogel is prepared by externally irradiating the solution according to any one of claims 1 to 5 with ultraviolet light or visible blue light, and stopping the application after the solution becomes gel-like.
7. A hydrogel for preventing and treating intrauterine adhesion and thin endometrium, characterized in that when the solution according to any one of claims 1-5 is applied to the corresponding tissue, the solution is irradiated with ultraviolet light or visible blue light from outside and stopped after the solution becomes gel-like, thus preparing the hydrogel.
8. The hydrogel for preventing and treating intrauterine adhesion and thin endometrium is characterized in that the solution for preparing the hydrogel comprises recombinant human type I collagen, recombinant human type III collagen or a mixture of the recombinant human type I collagen and the recombinant human type III collagen, a solution containing a photoinitiator, methacryloylated chitosan and total saponins of pseudo-ginseng, when the solution is applied to corresponding tissues, ultraviolet light or visible blue light is used for irradiation outside, and the hydrogel is prepared after the solution becomes gel; when the solution is subjected to viscosity test under the conditions of 37 ℃ and a shearing rate of 1/s, the viscosity is 3000 mPa.s-15000 mPa.s, and the mass and volume concentration of the total saponins of panax notoginseng is 0.4-0.6%.
9. The hydrogel of claim 8, wherein when the methacryloylated chitosan has a mass to volume concentration of 8%, the methacryloylated chitosan and the recombinant human type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen has a mass to volume concentration of 30%, the ratio of the methacryloylated chitosan to the recombinant human type i collagen, the recombinant human type iii collagen, or the mixture of the recombinant human type i collagen and the recombinant human type iii collagen is 1:1, and the total saponins of panax notoginseng has a mass to volume concentration of 0.6%.
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