Hydrogels are good forbiologyCompatibility and structural similarity to extracellular matrix components are favored delivery vehicles in the field of drug delivery. However, conventional single-network (SN) hydrogels suffer from poor mechanical properties and easy burst release of drugs.Recently, Zhao Chenjia, a doctoral student from the biomanufacturing team of Tsinghua University, is the first author. Together with the team of Professor Jing Yang from the University of Nottingham, UK, they published a paper entitled “Affinity-Controlled Double-Network Hydrogel Facilitates Long-Term Release of Anti- “Human Papillomavirus Protein”, reports a novel dual-network (DN) hydrogel, a material system that not only possesses tunable mechanical strength and long-term controllable delivery of antiviral proteins, but can also be personalized on demand3D printinghas great application potential in the field of tissue engineering.
Background introduction
The bioengineered protein drug market has been one of the fastest growing segments of the pharmaceutical and biotech market in recent years.However, the basic dosage form of therapeutic proteins is lyophilisate, which has low absorption efficiency and short biological half-life.clinicalThe efficacy has been greatly limited. Traditional protein formulations often require frequent injections to achieve the desired therapeutic effect, but this approach increases the risk of adverse side effects associated with overdose and reduces patient compliance. Therefore, the development of in situ delivery systems capable of long-term controlled release of protein drugs is of great significance in reducing adverse reactions and improving patients’ quality of life.
The team of Professor Sun Wei of Tsinghua University and the team of Professor Jing Yang of the University of Nottingham, UK have jointly developed a new dual network (DN) hydrogel system, which can achieve anti-virus by using the principle of affinity controlled release and the dense network structure in dual network materials. Long-term smooth release of protein. The system uses an alginic acid (ALG)/polyethylene glycol diacryloyl ester (PEGDA) network to improve the mechanical properties and network density of the hydrogel, adding a positively charged monomer 2-(acryloyloxy)ethyl ]Trimethylammonium chloride (AETAC) electrostatically interacts with a model protein electronegative anti-human papillomavirus (HPV) protein (3-hydroxyphthalic anhydride-modified bovine β-lactoglobulin) to achieve Delayed release effect. Compared with PEGDA SN hydrogel and ALG SN hydrogel, ALG/PEGDA-AETAC DN hydrogel has the characteristics of high stability, good mechanical strength and good flexibility. Meanwhile, the DN hydrogel significantly alleviated the burst release of antiviral proteins, extending the release time to more than 14 days.Combined with the biomimetic cervical reconstruction model designed by the researchers in the previous study, digital light processing (DLP) technology was used to print DN hydrogels, and a personalized drug-loaded cervical biomimetic could be constructed.Implants, which provides the possibility for in situ tissue repair and antiviral therapy in patients with cervical cancer after conization. The ALG/PEGDA-AETAC DN hydrogel developed in this study can be used as a carrier for the long-term release of negatively charged proteins. The good mechanical properties and biocompatibility show the potential for future application in soft tissue in situ delivery.
Experimental process and results
1. Preparation of double network gels
The researchers used a two-step cross-linking method to prepare double-network hydrogels. First, sodium alginate and calcium chloride were ionically cross-linked to form a brittle network. After UV light irradiation, PEGDA and AETAC monomers undergo chemical crosslinking under the action of photoinitiators to form a flexible network.
Figure 1. Preparation principle and process of dual network hydrogels
2. Characterization of double network hydrogels
The mechanical test results of the hydrogel showed that the compressive strength of the ALG/PEGDA-AETAC DN hydrogel was higher than that of the PEGDA-AETAC SN hydrogel. When the concentration of Igracure 2959 was not lower than 0.2% w/v, the compressive modulus of ALG/PEGDA-AETAC DN hydrogel was more than twice that of PEGDA-AETAC SN. In addition variable Igracure 2959 concentrations provided tunable compressibility of the hydrogel. In addition, compared with SN hydrogels, DN hydrogels have a lower swelling coefficient and a denser pore structure, the formation of which further delays the drug release and plays a key role in regulating drug release. effect.
Figure 2. Mechanical properties, swelling coefficients and micromorphological characterizations of dual-network hydrogels
3 Cytotoxicity testing of dual network hydrogels
Biocompatibility is a prerequisite to determine whether the hydrogel can be used further. In this study, we evaluated the growth of NIH 3T3 cells and HUVECs in SN and DN hydrogel extracts to verify their cytotoxicity. As shown in Figure 3, both CCK8 detection and cell live/dead staining demonstrated that ALG/PEGDA-AETAC and PEGDA-AETAC hydrogels had little cytotoxicity.Cytotoxicity classification according to ISO 10993-5:2009standardthe cytotoxicity level is one, it is non-toxic, and it provides the possibility for future clinical application.
Figure 3. Cytotoxicity characterization of SN and DN hydrogels
4 Protein release properties of double network hydrogels
Subsequently, the researchers compared the protein release kinetic curves of hydrogels with different compositions, and found that the controlled release effect of DN hydrogels against HPV proteins was stronger than that of SN hydrogels. In addition, the addition of the positively charged group AETAC enhanced the controlled release effect of the hydrogel, which was mainly due to the electrostatic interaction of the positively charged group of AETAC with the negatively charged anti-HPV protein. Therefore, ALG/PEGDA-AETAC has the best controlled release effect of anti-HPV protein, and can achieve stable release of anti-HPV protein for up to 14 days.
Figure 4. Release kinetics of antiviral proteins in different hydrogels
5 Printability characterization of dual network hydrogels
The study also explored the printability and potential applications of DN hydrogels after DLP
3D printingThe predetermined structure can then be formed.In order to verify the feasibility, the researchers printed letter structures and cones with hollow structures, and it was observed that ALG/PEGDA-AETAC DN hydrogels could form complex structures according to preset models, and the above-mentioned cone structures could be used as cervical cancer Cone cutOperationPost tissue patch. In addition, the researchers also found that the as-printed structure could recover its original shape after being compressed by more than 50%, which indicated that the DN hydrogel has good flexibility and great application potential in the field of soft tissue engineering.
3D printingThe predetermined structure can then be formed.In order to verify the feasibility, the researchers printed letter structures and cones with hollow structures, and it was observed that ALG/PEGDA-AETAC DN hydrogels could form complex structures according to preset models, and the above-mentioned cone structures could be used as cervical cancer Cone cutOperationPost tissue patch. In addition, the researchers also found that the as-printed structure could recover its original shape after being compressed by more than 50%, which indicated that the DN hydrogel has good flexibility and great application potential in the field of soft tissue engineering.
Figure 5. Printability test of DN hydrogels
6 in conclusion
This paper reports a novel DN hydrogel whose main components are PEGDA, sodium alginate, and positively charged monomer AETAC. By adjusting the concentration of photoinitiator, DN hydrogels matching the modulus of human soft tissue can be prepared. Furthermore, cytotoxicity tests showed that the prepared DN hydrogels supported cell growth. Based on an affinity-controlled delivery mechanism inspired by controlled release in the ECM, the positively charged group AETAC in the DN hydrogel can inhibit the burst release of negatively charged proteins through electrostatic interactions and achieve a prolonged release (over 14 days). Compared with SN hydrogels, the denser network structure in DN hydrogels also contributed to the sustained release of proteins. In addition, the material can pass DLP
3D printing technologyThe construction of complex structures with high elasticity has great potential for application in tissue engineering.
3D printing technologyThe construction of complex structures with high elasticity has great potential for application in tissue engineering.
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