The Mechanism Of Self-assembling Peptide In Skin Wound Rapid Repairing

This thesis focuses on the effects of self-assembling peptide Sciobio 3D cell culture matrix II and Sciobio 3D cell culture matrix III on skin wound rapid of repairing,and its mechanism.The self-assembling peptides were composed of 16 amino acids.For peptides' physical-chemical properties and self-assembling performance part: Mass spectrometry and high performance liquid chromatography determine whether the peptides meet experimental requirements;Circular dichroism spectroscopic investigate of secondary structures,consider the secondary structures influenced by the assembled times and growth factor(TGF-?1).Congo red staining macroscopic study of dynamic self-assembling process and gelation ability;Atomic force microscope microscopic obtains the microstructure of peptides.In the part of cell experiments: Detection the cell viability of HFF-1 cells in 3D culture microenvironment;q-PCR comparing the mRNA expression amount of related growth factors of cells constructed in 3D and 2D culture.In the part of animal experiments: Established a rapid skin wound repairing SD rat model,identified the impact of self-assembling peptides on wound healing from wound area,HE staining,immunohistochemical staining,q-PCR,etc.Then make a hypothesis to check what happen to wound healing process.Pfu DNA polymerase as a model to study the controlled release and protection of self-assembling peptides on proteins associated with the healing process.The results show: Both peptides are stable ?-sheet structure.The purity of the two short peptides is > 95% and meets the experimental requirement.24 hours after self-assembled,both peptides can form a stable,uniform membrane-film structure.Microscopically,the peptides with nanofiber network structure,can be used a nano-matrix for 3D cell culture.And,the added growth factor has not significantly influenced to the secondary structure of short peptides.HFF-1 cells grow well in 3D culture microenvironment.The growth rate of 1-5 days is slightly lower than that of 2D cultured cells.After the 7th day,cells in the 3D environment grow faster.q-PCR tests have the similar result: Culture 1-5 days,cell growth factors of 3D group are low expression level.However,on the 7th day,3D group cells content exceeded that of 2D control.Erythrocyte membrane lysis test results didn't find the destructive effect on biofilms at various concentrations and can be used as animal experiment material.In SD rat skin wound repairing model,self-assembling peptides group has been obtained much more positive result.The peptide group had basically completed the wound repair process on the 11 th day.On the 7th day,the non-healing area of the control group was 2 times than that of the peptide group,3 times on the 11 th day,and 2.5 times on the 14 th day.The results of HE staining and immunohistochemistry confirmed that self-assembling peptides can accelerate the dissipating of inflammation during skin wound repair.The expression of EGF,VEGF,TGF-?2 and other related growth factors was increasing in cells and tissues.The results of protection and controlled release experiments using Pfu enzyme as a representative protein show: When 0-4 ?l of peptide was added to 25 ?l PCR system,the amplification efficiency of Pfu enzyme could be improved.The controlled release rate of Pfu DNA polymerase was 94% for the 5 mg/ml short peptide,81% for Taq DNA polymerase,and 97% for TGF-?1.This result suggested the self-assembling peptides can not only control the release of proteins,but also increase the amplification efficiency of Pfu enzyme in PCR.In summary,the self-assembling short peptides utilized here are suitable for 3D cell culture,accelerate skin wound repairing with less inflammation,and can promote the expression of repair-related growth factors,ensuring the quality of wound repairing while promote the rapid development of skin wounds.The study cut the frontier-cross of the health field of in nano-repair and regenerative medicine,has a widely application prospects for the clinic and beyond,provide new options,new ideas and new strategies for the treatment of wound repairing,has important theoretical and social significance value.