Designed Fabrication Of Sericin Based Materials And Its Application In Skin Wound Repair

Wound healing is a complex physiological process,including four overlapping stages of hemostasis,inflammation,proliferation and remodeling.Chronic wounds caused by severe bacterial infections often cannot successfully complete these four stages,and need long-term treatment,which brings great physical and psychological pain to patients.For severe wound infection caused by drug-resistant bacteria,if the drug-resistant bacteria in the wound site are not killed,it may lead to amputation or death of the patient.Therefore,it is necessary to kill the resistant bacteria in the wound to avoid serious inflammation and edema.At the same time,if it can accelerate the proliferation and remodeling stage of wound healing,the time for wound healing could be reduced.So,it is necessary to develop wound repair dressings with antibacterial properties and accelerating wound healing.With the development of nanotechnology,organic/inorganic nanocomposites have provided new ideas for the creation of multifunctional wound repair materials.Sericin is a natural protein with good biocompatibility and biodegradability.In particular,sericin has cytoprotective and mitogenic effects on fibroblasts and keratinocytes,making it attractive for the development of skin and tissue repair materials.It could be processed into films,gels,sponges and other forms.In addition,sericin containing various amino acids and special protein sequences,could be used as a protein template for biomimetic synthesis of inorganic nanomaterials.In this study,we used sericin to prepare sericin-inorganic nanocomposites with antibacterial properties through biomimetic synthesis strategy.Further,a wound repair material based on sericin with antibacterial effects was constructed.This paper includes the following three parts:1.Sericin has biological functions to promote wound repairCell proliferation,cell migration,collagen deposition and angiogenesis are important events in the proliferation and remodeling stages of wound healing.A varietyof cells,cytokines,growth factors and complex cell signals play an important role in this process.If the signal pathways related to cell proliferation,cell migration,collagen production,and angiogenesis can be activated,the process of wound healing could be accelerated.We cultured mouse fibroblasts(NIH/3T3)cells with sericin extracted by high temperature alkaline water(Sericin~A)and sericin extracted by high temperature and high pressure(Sericin~P).The results showed that sericin could promote cell proliferation in serum-free and low serum(0.5%v/v)medium,and Sericin~Apromotes cell proliferation more effectively than Sericin~P.We further found that sericin can promote the PI3K/AKT,SAPK/JNK,ERK,p38 MAPK,and NF-κB signaling pathways in NIH/3T3 cells.These signaling pathways are related to cell proliferation,cell migration,angiogenesis,and collagen deposition.Further,we found that Sericin~A and Sericin~P can promote the expression of Cyclin D1,Vimentin and CollagenⅢ,and Sericin~A is dose-dependent in the concentration range of 0.02%to 0.10%(w/v).At the transcription level,Sericin~A and Sericin~P can promote the expression of genes related to cell migration(Fibronectin1,Lamininβ1,Vimentin),collagen production(TGF-β1,TGF-β1,Collagen I,Collagen III),and angiogenesis(EGF and VEGF).We have also found that sericin can promote the expression of integrinβ1,as well as the expression of FAK and its phosphorylation.In summary,our data indicate that sericin can activate the PI3K/AKT,MAPK,NF-κB pathways,and promote the production of Cyclin D1,Vimentin,and Collagen,which has positive effect for the proliferation and remodeling stages of wound healing.2.Bioinspired design of Ag NPs embedded silk sericin-based sponges for efficiently combating bacteria and promoting wound healingSericin has excellent biological function and could be used as a potential wound repair material.It has special protein sequences that can be used as protein templates to biomimetic synthesis inorganic nanomaterials.The amino acids in sericin can be used as reducing agents and stabilizers for the synthesis of nanomaterials.Silver nanoparticles(Ag NPs),as a broad-spectrum antibacterial agent,has excellent antibacterial properties.In this chapter,we developed a novel,simple,one-step biosynthesis method to prepare sericin-silver nanoparticle composite in situ in solution.The synthesized silver nanoparticle had good dispersibility and long-term stability at room temperature.Light irradiation was essential for the preparation of sericin-silver nanoparticle composite.Further,in order to prepare antibacterial sponge dressing based on sericin-silver nanoparticle,we developed a green strategy for in situ biomimetic synthesis of Ag NPs within sericin/poly(vinyl alcohol)hydrogel.Ag NPs were in situ synthesized by the redox property of tyrosine residues in sericin without additional chemicals,similar to a biomineralization process.Ag NPs-sericin/poly(vinyl alcohol)dressing was demonstrated to bear desired high porosity,good wettability,hygroscopicity,and mechanical properties.The evaluation of cytocompatibility and immunotoxicity of the composite with NIH/3T3,HEK-293,and RAW264.7 cells further demonstrated its excellent biocompatibility.The composite also showed an effective antibacterial activity against S.aureus,E.coli,P.aeruginosa and could inhibit biofilm formation caused by S.aureus and P.aeruginosa.Finally,the animal experiment suggested that the composite dressing could facilitate re-epithelialization and collagen deposition to promote wound healing.Our work shows that the composite dressing has great potential as a competitive alternative for traditional wound dressings.3.Preparation of sericin-gold nanoclusters-DAPT nanofibers and its promotion of wound healing of drug-resistant bacterial infectionsChronic infection caused by drug-resistant bacteria poses a great challenge to wound care,but it is often neglected in clinical treatment.Inorganic nanomaterials have attracted great attention for their unique physical and chemical properties in resisting drug-resistant bacterial infection.Gold nanomaterials have broad application prospects in biomedical field due to their excellent biocompatibility,good stability and easy modification.Our research found that sericin could prepare gold nanomaterials with different particle sizes and shapes in situ in different p H environments.Among them,Sericin-Au NC nanoclusters have broad-spectrum antibacterial activity.In order to further enhance the antibacterial performance of Sericin-Au NC,we use Au NC as drug carrier to load drug small molecules.And then,Sericin-Au NC-DAPT composite nanomaterials were prepared in situ in sericin solution.We have tested the antibacterial properties of the composite nanomaterials,and found that the MIC of the composite nanomaterials to four common bacteria was 8μg/m L.SEM and TEM were used to observe the morphology of bacteria treated with composite nanomaterials and found that the composite nanomaterials could effectively destroy the cell membrane of bacteria.After wounds are infected by drug-resistant bacteria,it needs wound repair materials with excellent antibacterial properties to kill bacteria.It also requires wound repair materials have appropriate surface microstructures and biochemical properties to promote cell adhesion,proliferation and differentiation.Organic/inorganic antibacterial nanofibers with biological activity could effectively kill drug-resistant bacteria and promote wound healing.We prepared a nanofiber dressing(PVA/Se-Au-D)based on sericin-Au NC-DAPT by electrospinning.In vitro study showed that nanofiber dressing can effectively kill drug-resistant bacteria and has excellent biocompatibility.MDR E.coli and MRSA wound infection rat models show that PVA/Se-Au-D can significantly promote wound healing.H&E staining and massion staining showed that PVA/Se-Au-D nanofibers could enhance collagen deposition and skin regeneration.Immunohistochemistry showed that sericin nanofiber dressing could reduce inflammation caused by bacterial infection.Immunofluorescence showed that nanofiber dressing can promote angiogenesis.Therefore,PVA/Se-Au-D nanofiber matrix could be used as a multifunctional dressing for treating wounds with drug-resistant bacteria infection and promoting wound healing.In conclusion,we have proved that sericin promotes cell proliferation,cell migration,collagen production and angiogenesis through PI3K/AKT,MAPK and NF-κB pathways.Further,sericin is used as a protein template for biomimetic synthesis of inorganic nanomaterials,and sericin-Ag NPs and sericin-Au NCs organic/inorganic nano composite materials are constructed through an in-situ green strategy.Antibacterial sponge and antibacterial nanofibers based on sericin were prepared by electrospinning and freeze crosslinking methods.According to the experimental results of drug-resistant bacterial infection model,it can be concluded that the sericin-based organic/inorganic composite dressing can significantly promote wound healing.