阅读说明：本技术 一种促进骨修复复合水凝胶及其制备方法与应用 (Composite hydrogel for promoting bone repair and preparation method and application thereof ) 是由 林海燕 周艳 王维倩 涂业颖 于 2020-04-14 设计创作，主要内容包括：本发明公开了一种促进骨修复复合水凝胶,其包括如下组分：甲基丙烯酸改性明胶和多巴胺改性海藻酸钠；所述复合水凝胶中,所述甲基丙烯酸改性明胶的含量为8～12％w/v,所述多巴胺改性海藻酸钠的含量为0.5％w/v～2％w/v。多巴胺改性海藻酸钠的分子结构中含有邻苯二酚结构,具有优异的粘附性能。本发明以甲基丙烯酸改性明胶作为水凝胶的基材,并通过掺入多巴胺改性海藻酸钠来改善水凝胶的力学性能和粘附性能,使其具有优异的粘性及适宜的柔软度和弹性,适用于骨修复治疗。体外实验结果表明,本发明的复合水凝胶能负载并缓释促进骨修复的生物活性因子和/或药物,并在骨缺损模型中表现出良好的修复效果,能够显著促进骨缺损伤口的愈合。(The invention discloses a composite hydrogel for promoting bone repair, which comprises the following components: methacrylic acid modified gelatin and dopamine modified sodium alginate; in the composite hydrogel, the content of the methacrylic acid modified gelatin is 8-12% w/v, and the content of the dopamine modified sodium alginate is 0.5-2% w/v. The molecular structure of the dopamine modified sodium alginate contains a catechol structure, and the dopamine modified sodium alginate has excellent adhesion performance. According to the invention, methacrylic acid modified gelatin is used as a base material of the hydrogel, and dopamine modified sodium alginate is doped to improve the mechanical property and the adhesion property of the hydrogel, so that the hydrogel has excellent viscosity, appropriate softness and elasticity, and is suitable for bone repair treatment. In-vitro experiment results show that the composite hydrogel can load and slowly release bioactive factors and/or medicines for promoting bone repair, shows a good repair effect in a bone defect model, and can remarkably promote the healing of bone defect wounds.)

1. The composite hydrogel for promoting bone repair is characterized by comprising the following components: methacrylic acid modified gelatin and dopamine modified sodium alginate; in the composite hydrogel, the content of the methacrylic acid modified gelatin is 8-12% w/v, and the content of the dopamine modified sodium alginate is 0.5-2% w/v.

2. The composite hydrogel for promoting bone repair of claim 1, wherein the composite hydrogel comprises 10% w/v of methacrylic acid modified gelatin and 1% w/v of dopamine modified sodium alginate.

3. A method for preparing the composite hydrogel for promoting bone repair according to claim 1 or 2, comprising the steps of: respectively dissolving methacrylic acid modified gelatin and dopamine modified sodium alginate in a solvent to prepare a methacrylic acid modified gelatin solution and a dopamine modified sodium alginate solution, and then preparing the bone repair promoting composite hydrogel through photo-crosslinking.

4. The method for preparing a composite hydrogel for promoting bone repair according to claim 3, wherein the solvent is a PBS solution or a MES solution.

5. The method for preparing a composite hydrogel for promoting bone repair according to claim 3, wherein the photoinitiator for photocrosslinking is lithium phenyl-2, 4, 6-trimethylbenzoylphosphite.

6. The use of the composite hydrogel for promoting bone repair of claim 1 in a biological scaffold material.

7. The use of the composite hydrogel for promoting bone repair of claim 1 in a bone repair scaffold material.

8. A bone repair scaffold material formed from the bone repair promoting composite hydrogel of claim 1 or 2.

9. The bone repair scaffold material of claim 8, further loaded with bioactive factors and/or drugs that promote bone repair.

10. A method of preparing a bone repair scaffold material according to claim 8 or 9, comprising the steps of: respectively dissolving methacrylic acid modified gelatin and dopamine modified sodium alginate in a solvent to prepare a methacrylic acid modified gelatin solution and a dopamine modified sodium alginate solution, mixing the methacrylic acid modified gelatin solution and the dopamine modified sodium alginate solution, adding bioactive factors and/or medicines for promoting bone repair, and then preparing the bone repair scaffold material through photo-crosslinking.

Technical Field

The invention relates to the technical field of medical biomaterials, in particular to a composite hydrogel for promoting bone repair and a preparation method and application thereof.

Background

Bone defects are common diseases in clinic, and are often caused by that fracture blocks penetrate out of limbs in trauma or that inactivated bones are removed in debridement of open fractures. The need for bone repair is prevalent in orthopedic and neurological surgery, as well as oral maxillofacial surgery.

The existing methods for treating bone defects include autologous bone and allogeneic bone transplantation, tissue engineering technology, gene therapy rejection methods, growth factors, auxiliary therapy of physical therapy and the like. Biomedical implants are an important component of modern medicine, are used to replace missing structures or functions of the human body, and play an important role in a variety of clinical applications such as tissue repair and reconstruction, fracture fixation, hip and knee replacement, electronic sensing or monitoring of organ transplantation. However, implant-related infections and delayed tissue regeneration are two major clinical complications. Especially for orthopedic implants, can lead to dysfunction, implant failure, additional surgery, and even death. An ideal implant should have good colonization and induce an appropriate cellular response to promote osteointegration without sacrificing the desired structural and physicochemical properties and functions.

The gelatin is derived from collagen in connective tissues such as bones, bonds, cartilages, skins, sarcolemmas and the like of animals, is an irreversible degradation product of the collagen, still keeps good biocompatibility, and has wide application in the fields of medicines and biological materials. Gelatin is a typical amphiphilic polyelectrolyte, and an aqueous solution of gelatin forms a thermo-reversible gel when the temperature drops below 35 ℃. Hydrogels made from gelatin alone are not stable and can be modified by grafting other groups to enhance their stability. Since the gelatin molecule contains a large amount of amino, methacrylic acid anhydride can be used for grafting modification to prepare methacrylic acid modified gelatin (GelMA). However, the hydrogel formed by GelMA has hard texture, lacks elasticity and viscosity, has poor colonization effect, and limits the application of the hydrogel.

Disclosure of Invention

In order to overcome the defects and shortcomings in the prior art, the invention aims to provide a composite hydrogel for promoting bone repair and a preparation method and application thereof. The composite hydrogel has excellent adhesion performance, proper mechanical property, good biocompatibility and degradation performance, is an ideal bone repair scaffold material, can be used for loading and slowly releasing medicines and/or growth factors and the like beneficial to bone repair, and is used for bone repair treatment.

In order to realize the purpose, the technical scheme adopted by the invention is as follows:

the invention provides a composite hydrogel for promoting bone repair, which comprises the following components: methacrylic acid modified gelatin and dopamine modified sodium alginate; in the composite hydrogel, the content of the methacrylic acid modified gelatin is 8-12% w/v, and the content of the dopamine modified sodium alginate is 0.5-2% w/v.

Most preferably, the content of the methacrylic acid modified gelatin in the composite hydrogel is 10% w/v, and the content of the dopamine modified sodium alginate in the composite hydrogel is 1% w/v.

The molecular structure of the dopamine modified sodium alginate contains a catechol structure, and the dopamine modified sodium alginate has excellent adhesion performance. According to the invention, methacrylic acid modified gelatin is used as a base material of the hydrogel, and dopamine modified sodium alginate is doped to improve the mechanical property and the adhesion property of the hydrogel, so that the hydrogel has excellent viscosity, appropriate softness and elasticity, and is suitable for bone repair treatment.

Preferably, the preparation method of the composite hydrogel for promoting bone repair comprises the following steps: respectively dissolving methacrylic acid modified gelatin and dopamine modified sodium alginate in a solvent to prepare a methacrylic acid modified gelatin solution and a dopamine modified sodium alginate solution, and then preparing the bone repair promoting composite hydrogel through photo-crosslinking.

Preferably, the solvent is a PBS solution or a MES solution.

Preferably, the photoinitiator used for photocrosslinking is lithium phenyl-2, 4, 6-trimethylbenzoylphosphite (photoinitiator LAP). The photoinitiator LAP is an aqueous ultraviolet photoinitiator, the photopolymerization reaction condition is mild, and the photocrosslinking time can be greatly shortened. Preferably, the photoinitiator LAP is added in an amount of 0.08% w/v to 1.5% w/v.

Preferably, the method for preparing the methacrylic acid modified gelatin (GelMA) comprises the following steps: dissolving gelatin in water to prepare a gelatin solution, dissolving methacrylic anhydride in water to prepare a methacrylic anhydride solution, slowly dripping the methacrylic anhydride solution into the gelatin solution, stirring in a water bath to react, diluting the mixed solution after the reaction is finished, dialyzing, and finally freeze-drying to obtain GelMA.

Preferably, the concentration of the gelatin solution is 4-6% w/v, the dosage ratio of the methacrylic anhydride to the gelatin is 0.54-0.64 mL/g, the temperature of the water bath stirring reaction is 55-65 ℃, the time is 3-6 h, and the concentration of the gelatin diluted by the mixed solution is 1.5-3% w/v.

Preferably, the preparation method of the dopamine modified sodium alginate (SA-DA) comprises the following steps: dissolving sodium alginate in water to prepare a sodium alginate solution, adding 1- (3-dimethylaminopropyl) -3-ethylcarbodiimide hydrochloride (English name: EDC), adjusting the pH value of the solution to subacidity, activating, adding N-hydroxysuccinimide (English name: NHS), uniformly stirring, adding dopamine hydrochloride, keeping the pH value of a reaction system at 5-6, reacting under the protection of inert gas, dialyzing with a subacidity hydrochloric acid solution after the reaction is finished, dialyzing with distilled water, and finally freeze-drying to obtain SA-DA. In the present application, the purpose of the activation is to activate the carboxyl group of sodium alginate to cause amidation with dopamine.

Sodium Alginate (SA) has good biodegradability and biocompatibility, is easy to dissolve in water, has good gelatinization performance, strong hygroscopicity and good water retention performance. As the SA molecular structure contains carboxylate radical molecules, the SA is subjected to reactive grafting by using dopamine hydrochloride (DA) under an acidic condition, and the prepared SA-DA molecular structure contains a catechol structure. Researches show that the catechol structure plays a key role in strong adhesion and can be firmly and stably adhered to the surfaces of various base materials.

Preferably, the concentration of the sodium alginate solution is 0.8-1.2% w/v, and the molecular weight cut off of the distilled water dialysis is 8000-15000 Da. In the preparation method of SA-DA, EDC and NHS are used as catalysts, and the addition amount of EDC and NHS is not strictly required, so that effective catalytic action can be exerted, and EDC and NHS can be removed by dialysis in subsequent steps. And the dopamine hydrochloride can be added in excess, and the excess unreacted dopamine hydrochloride can be removed in subsequent dialysis. For reference, the addition amount of EDC is 1% g/mL, the addition amount of NHS is 1.5% g/mL, and the addition amount of dopamine hydrochloride is 0.5% g/mL. When the three substances are added, in the preparation method, the catalyst can exert a good catalytic effect, so that the reaction grafting rate is high, the addition amount of dopamine hydrochloride is sufficient, the yield of the modified product SA-DA is high, excessive dopamine hydrochloride waste cannot be caused, and the preparation cost is low.

The invention also provides application of the composite hydrogel for promoting bone repair, which comprises application of the composite hydrogel for promoting bone repair in a biological scaffold material, preferably application in a bone repair scaffold material.

The invention also provides a bone repair scaffold material, which is formed by the bone repair promoting composite hydrogel.

Preferably, the bone repair scaffold material is also loaded with bioactive factors and/or drugs for promoting bone repair.

Preferably, the bioactive factor comprises Platelet Derived Growth Factor (PDGF). PDGF has important functions of promoting migration and recruitment, division and proliferation, differentiation of bone forming cells, local blood circulation reconstruction and the like, is combined with the composite hydrogel to form a slow release system, and can effectively accelerate fracture healing and promote connection of bone defects.

Preferably, the preparation method of the bone repair scaffold material comprises the following steps: respectively dissolving methacrylic acid modified gelatin and dopamine modified sodium alginate in a solvent to prepare a methacrylic acid modified gelatin solution and a dopamine modified sodium alginate solution, mixing the methacrylic acid modified gelatin solution and the dopamine modified sodium alginate solution, adding bioactive factors and/or medicines for promoting bone repair, and then preparing the bone repair scaffold material through photo-crosslinking.

Preferably, the addition amount of the bioactive factor and/or the medicament is 0.15% w/v-0.3% w/v. When the addition amount of the bioactive factor and the medicament is controlled in the range, the hydrogel has a good bone repair effect and does not generate side effects. If the addition amount of the bioactive factor and the drug is too high, side effects may be caused by too high a drug concentration, and if the addition amount is too low, the bone repair effect of the hydrogel may be affected.

Compared with the prior art, the invention has the beneficial effects that: the composite hydrogel is an ideal bone repair scaffold material, has suitable mechanical property and excellent viscosity, can ensure that the hydrogel does not fall off from a wound, can also ensure that the hydrogel is not broken due to too small mechanical property, can not be too hard due to too large mechanical property, can cause discomfort of the wound and the like, has good biocompatibility and degradation property, and is safe to use. In-vitro experiment results show that the composite hydrogel can load and slowly release bioactive factors and/or medicines for promoting bone repair, shows a good repair effect in a bone defect model, and can remarkably promote the healing of bone defect wounds.

Drawings

FIG. 1 is a diagram of the ultraviolet absorption spectra of SA, DA and SA-DA;

FIG. 2 is a nuclear magnetic hydrogen spectrum of Gel and GelMA;

FIG. 3 is a graph of swelling performance of GelMA hydrogel prepared in example 3 and GelMA/0.5% SA-DA composite hydrogel prepared in example 4;

FIG. 4 is a graph of the degradation profiles of GelMA hydrogel prepared in example 3 and GelMA/0.5% SA-DA composite hydrogel prepared in example 4;

FIG. 5 is a graph showing the compressive strength test of GelMA hydrogel prepared in example 3 and composite hydrogels prepared in examples 4 to 7;

FIG. 6 is a graph showing the adhesion strength test of GelMA hydrogel prepared in example 3 and composite hydrogels prepared in examples 4 to 7;

FIG. 7 shows the results of the biocompatibility tests of GelMA hydrogel prepared in example 3, GelMA/1% SA-DA composite hydrogel prepared in example 5, and GelMA// 1% SA-DA @ PDGF bone repair scaffold material prepared in example 8;

FIG. 8 is an in vitro drug release profile of GelMA// 1% SA-DA @ PDGF bone repair scaffold material.

Detailed Description

To better illustrate the objects, technical solutions and advantages of the present invention, the present invention is further described in detail below with reference to the following embodiments and the accompanying drawings. It is apparent that the following examples are only a part of the embodiments of the present invention, and not all of them. It should be understood that the embodiments of the present invention are only for illustrating the technical effects of the present invention, and are not intended to limit the scope of the present invention.