| Collagen mimetic peptide (CMP) which was designed according to the special amino acid sequence of collagen could simulate part of characteristic structure or biological function of nature collagen. It could be used to study the special structure and function of collagen as molecular models, and also to explore well biocompatibility biomaterials. This would bring new thoughts to the further research of collagen and biomaterials based on collagen.In this study, several CMPs based on the characteristic POG triplets of collagen were designed. We studied triple-helix and thermostability regulation with lengths and sequences. The CMP with cysteine could couple multiarm star PEG, and then assembling to hybrid hydrogels by triple-helix of CMP. With studying the structure, biodegradability and cytocompatibility of this system, we provided some reference for building and researching novel peptide bionic hydrogels used for3D cell clutrue.The main contents and results of this work were as follows:1. Triple-helix, thermostability and cytocompatibility of CMPsFive CMPs were designed and synthetized varying the number of POG triplets and incorporating an integrin α2β1binding motif GFOGER. CD spectroscopy from260to190nm was recorded to confirm the existence of triple-helix conformation at room temperature, while thermal melting and thermal annealing of triple-helix (thermal unfolding and refolding of triple-helix, respectively) was characterized by monitoring ellipticity at225nm as a function of temperature. The results demonstrated that all the CMPs adopted triple-helix conformation, and the thermal stability of the CMPs enhanced with increasing number of POG triplets. In contrast to natural collagen, the thermal denaturation processes of CMPs were reversible. Meanwhile, the phenomenon of "hysteresis" was observed by comparing melting and thermal curves. The cytotoxicity to L929detected by CCK-8, results showed that CMPs couldn’t obviously inhibit the growth of cells.2. Preparation and characterization of hybrid hydrogels based on CMP and multiarm star PEGThe CMP both with cysteine and RGD was synthesized, it could couple multiarm (4-arm or8-arm) star PEG with terminal maleimide functionality (MAL-PEG), then assembling to hydrogels by triple-helix. Different precursor concentrations and mole ratios of hydrogels forming were determined to seek suitable conditions. Reaction product was characterized by nuclear magnetic resonance (NMR) to confirm the yield. Viscoelastic properties and stiffness were tested by rheometer. Structural feature was measured by atomic force microscopy (AFM), transmission electron microscopy (TEM) and scanning electron microscope (SEM). The results showed that the reaction yield was over80%, and the hydrogel with6%precursor concentration and1:1mole ratio was best, both of4-arm MAL-PEG and8-arm MAL-PEG could couple CMP, then forming hydrogel, and having the similar rheological performance. It had honeycomb-like network structure with round compartments or pores.3. Cytobiology study of hybrid hydrogels based on CMP and multiarm star PEGWe studied cytotoxicity and adhesion of CMPs by L929and degradation of MAL-PEG-CMP hydrogels by collagenase, packaged cells during hydrogel forming to see its3D culture cytocompatibility. The results showed that CMPs had no obvious cytotoxicity and MAL-PEG-CMP hydrogels could be degraded by collagenase. There was no obvious cytotoxicity in3D culture.In conclusion, MAL-PEG-CMP hydrogels based on the tripe-helix of CMP had good cytocompatibility and could be degraded by collagenase. These findings added new insights to search for cell responsive bionic hydrogels and ideas of bioactive materials based on peptides. |
PDF Full Text Request