Preparation And Properties Of Hydrogel For Cartilage Repair And Regeneration

In cartilage regeneration, scaffolds can provide suitable environment for cellgrowth and tissue formation. Hydrogel is a kind of hydrated polymer networks, whichmimics the native extracellular matrix (ECM) of cartilage. The hydrogel can betransplanted into damaged site to realize tissue repair through injection in a minimallyinvasive way. Therefore, the hydrogel is a kind of very important materials incartilage repair and regeneration. In this dissertation, injectable hydrogels wereprepared by photoinitiating polymerization, into which microcarriers,biomacromolecules and growth factors were introduced. The structure and propertiesof the hydrogels were studied. A biomimic hydrogel was designed at last. Theseworks had great significance in designing ideal biomaterials for cartilage repair andregeneration.Based on the previous research work in our laboratory, water-soluble andcrosslinkable chitosan was synthesized by sequentially grafting of methacrylic acid(MA) and lactic acid (LA). Irradiated by 365 nm UV light, Photoinitiator(Irgacure2959) was used to initiate CML polymerization in water solution and therebyobtain chitosan hydrogel. The gelation time was controlled from several minutes totens minutes. When the photoinitiator concentration was lower than 0.05ï¼…, thegelation time was longer and influenced greatly by the photoinitiator concentration.Therefore, in this work, the hydrogel was obtained using 0.05ï¼…photoinitiator. Thedry hydrogels could adsorb water by a factor of tens to hundred. The swelling ratiowas smaller at larger crosslinking density, higher pH value, and in salt solutions,especially in Na₂SO₄ and Na₂HPO₄ solution. Rheological measurement recorded thestorage modulus and loss modulus of 0.8-7 kPa and 10-100 Pa, respectively. With ahigher degree of MA substitution, the resultant hydrogel had a larger modulus anddegraded slowly too. In vitro chondrocyte culture showed that the cytotoxicity of thehydrogel made by Irgacure2959 was much smaller than that of the hydrogel made byammonium persulfate (APS)/ N,N,N',N'-tetramethylethylenediamine (TEMED). In order to promote the chondrocytes growth in hydrogel, gelatin was introducedinto the CML hydrogel. The release behaviors of gelatin as a biomacromoleculartemplate in CML hydrogel were studied. The released gelatin ratio was smaller at alarger crosslinking extent of gelatin, original gelatin content in hydrogel and higherpH value. The released gelatin ratio was smaller at 20℃than that at 37℃. Moreover,the released gelatin ratio was influenced by ions and amino acids in solution. Thestructure and properties of the CML hydrogel were also influenced by the embeddedgelatin. When the embedded gelatin increased, mesh size of the hydrogel was smallerand swelling ratio of the hydrogel decreased. The rheological properties anddegradation properties were also influenced by the embedded gelatin. In vitrochondrocyte culture showed that the gelatin containing hydrogel could promotechondrocyte growth and secrete ECM. Chondrocytes in the CML hydrogel couldmove downward along with the culture time.We modified gelatin with MA and obtained crosslinkable gelatin (GM), whichwas used to form injectable gelatin hydrogel by photoinitiating polymerization. Thepolydispersity of molecular weight broadened and zeta potential increased too aftermodification with MA. There were no double carbon bonds in hydrrogel detected byhydrogen spectrum of high resolution magic angle spinning nuclear magneticresonance (¹H HR-MAS NMR) after irradiation for 20 min. The sol-gel transitionpoint of gelatin before and after modification was characterized by differentialscanning calorimeter (DSC), revealing that it is influenced by MA modification andpolymer concentration. Triple helix structure existed in the gelatin gel and wasconfirmed by circular dichroism (CD), whose content decreased after modification.With polymer concentration increasing, the gelation time decreased, swelling ratiodecreased, storage modulus and loss modulus increased and mesh size decreased.Transform growth factor-β1 (TGF-β1) also combined into the GM hydrogel topromote the chondrocyte growth. In vitro chondrocyte culture showed that the GMhydrogel both with and without TGF-β1 could support the chondrocyte growth andmaintain the chondrocyte phenotype. TGF-β1 could promote chondrocyte growth andmaintaining chondrocytic phenotype. Combination of advantages of the injectable hydrogels and the injectablemicrocarriers, a composite hydrogel was prepared. First, the surface of poly(lactic-co-glycolic acid) (PLGA) microparticles were modified with GM. Themodified particles were then dispersed in CML solution. Under UV irradiation,particles were polymerized with CML to obtain a composite hydrogel, which showedbetter mechanical property and cytoviability than that of the CML hydrogel. Asinjectable materials, the composite hydrogel might have wider potential applications.At last, hyaluronic acid (HA)/gelatin/chondroitin sulfate hydrogel was formed byclick chemistry to mimic the compositions of natural cartilage matrix. HA and CSwere modified with 11-azido-3,6,9-trioxaundecan-1-amine (AA), which contain -N₃group. Gelatin was modified by propargylamine (PA), which contain acetylene group.Catalyzed by Cu (â… ), -N₃ group was reacted with acetylene group to obtain thehydrogel. As Cu (â… ) concentration increasing, the gelation time was shortened until1 mg/mL. The swelling ratio of this hydrogel was 18.8±2.4, larger than that of thegelatin hydrogel. The rheological result showed that this hydrogel was elastomeric.The weight loss of this hydrogel in 4 weeks reached to 50ï¼…. About 20ï¼…gelatin and10ï¼…CS was released from this hydrogel in 2 weeks. The surface of hydrogel filmwas smooth characterized by atomic force microscopy (AFM). In vitro chondrocyteculture showed that the chondrocytes could grow on the surface of the hydrogel andmaintain its phenotype.