Study Of Enzymatic Responsive Short Peptide Self-assemly Hydrogels And Its Application In Cell Culture

Hydrogel was a kind of gel in which water played the role of dispersing medium.Since the structure of hydrogel was a three dimensional entangled network from microscale perspect,it could provide proper mechanical support.Meanwhile,proper porous properties of the net-work guaranteed the permeability of this kind of material.So hydrogel was considered holding good potential in tissue engineering,regenerative medicine and controlled drug release.Accor-dign to current works,self-assembly of small molecules was a common method in preparing hydrogels,in which short peptides were a competitive choice for these small molecules owing to the convenience of synthesis,diversity in functionalization,tunable morphology etc.Peptides could self-assembled into nanostructures with large length-diameter ratio like nano-ribbon and nano-tapes,and further led to hydrogelation under the control of non-covalent interactions in-cluding hydrophobic interaction,electrostatic interaction,H-bond and so on.By rational insert specific recognition sequence of enzyme,peptides could be designed into enzymatic responsive molecules with proper self-assembly characters.The enzymatic responsive peptide self-assem-bly system could perform several functions like gelation,degradation and functionalization un-der the endogenous stimulations of enzymes,meanwhile these systems could participated in the biochemical reactions,which provided a new choice for their application.In this thesis,we introduce a series of enzymatic responsive short peptides based on the self-assemly model of Ac-I₃K-NH₂.After characterization of their self-assembly behaviours,the enzymatic response of these molecules were testified together with influence of enzymes on morphology,secondary structure and rheological characters.Then these systems were used as support in cell culture.The details are as follows:Firstly,we designed and synthesized a series of transglutaminase responsive peptides,then characterized their nanotape conjugates from morphological aspect and beta-sheet from sec-ondary structure aspect in HEPES buffer.We illustrated that their response towards transglu-taminase was dimerization.Furthermore,we exhibited that the gelation mechanism was the self-assembly of emzymatically catalyzed product.Then we evaluated the cytotoxicity of short peptide and their hydrogel and further applied these hydrogels in liver hemostatic model of SD rats.Based on the hemostasic data in virto and in vivo,we proposed a possible mechanism of hemostatic process of designed peptides.Secondly,we designed a matrix metalloproteinase II responsive peptide Ac-I₃SLKG-NH₂and its control molecule Ac-I₃SLGK-NH₂ and further characterized their nanofiber morphology and beta-sheet secondary structure.Then we proposed and testified the mechanism of biodeg-radation?including irregulation of nanofibers,deself-assembly of secondary structures and drop of storage modulus?and studied the biodegradation process.Finally we used normal cell lines to evaluate the cytotoxicity of these hydrogels and migration of cells.Furthermore,we success-fully built a hydrogel hydrophilic drug controlled and selectively release system and testified its selectivity and function.Thirdly,we design and synthesize two dual-enzymatic responsive peptides Ac-I₃SLKGK-NH₂?response to plasma amino oxidase and matrix metalloproteinase II?and Ac-I₃SLKGQ-NH₂?response to transglutaminase and matrix metalloproteinase II?.Then we add plasma amino oxidase,matrix metalloproteinase II and transglutaminase,matrix metalloproteinase to each system sequentially and characterize hydrogelation and degradation of two systems.To illustrate two processes in detail,we characterize changes in two systems from perspective of morphology,secondary structure and rheological properties.Owing to the dual-enzymatic re-sponse of two systems,their hydrogelation can be triggered by enzymes and support the spon-taneously migration of cells.In conclusion,two mono-enzymatic responsive short peptide self-assembly systems and two dual-enzymatic responsive short peptide self-assembly systems are designed and synthe-sized,and their cytotoxicity and applications are briefly investigated.Further,enzymatically catalyzed degradation and dual-enzymatic responsiveness in short peptide self-assembly model are archieved,which provide new solution for the tissue engineering materials.