Hyaluronic Acid Hydrogels Covalently Crosslinked By Diglycidyl Ethers Via Lyophilization

Hyaluronic acid(HA)is a naturally occurring polysaccharide,widely distributed in various parts of the body,such as eyes,synovial fluid of joint cavity,connective tissue and skin.Hyaluronic acid is readily degraded by hyaluronidase in the body,shortening its residence time in the body,thus limiting its biomedical applications.The chemical modification and crosslinking of hyaluronic acid are the primary approaches to solve the above problem,thereby extending its residence time in the human body.It is generally believed that the diglycidyl ether-type crosslinkers crosslink HA by the formation of a stable ether bond between the primary alcohol groups of hyaluronic acid and the epoxides of the diglycidyl ethers at elevated temperature,however,we reported here the covalently crosslinked HA hydrogels,prepared for the first time via lyophilization of a series of diglycidyl ethers with different concentrations and HA with different MW and concentrations in aqueous solutions,which enabled the efficient crosslinking of HA molecules to obtain HA hydrogels 1-72 after addition of aqueous solutions into the resulted aerogels while those in the solutions at room temperature were not chemically crosslinked during the same time period as lyophilization.Among the 72 hydrogel samples,24 of hyaluronic acid hydrogels that were more stable were selected for furthur characterization after screening of their stability in the PBS buffer solution.For the selected 24 hydrogels,their properties suach as water absorption in the form of aerogels,rheology,degradation time in plasma,degradation time by hyaluronidase in vitro,cytotoxicity and cell proliferation were studied.The results indicated that water absorption of the hydrogels in the form of aerogels weres 22-44 times of their selfies;lyophilization enabled the formation of hyaluronic acid hydrogels covalently crosslinked by diglycidyl ethers while the identical solutions kept at room temperature remained as liquid,or became physical gels,not chemically crosslinked gels.The stability time of the hydrogels in plasma and in the solution containing hyaluronidase reached 152 hr and 264 hr,respectively;Cytotoxicity of those hydrogels were not significantly increased;Cell proliferation experiments showed that the HA hydrogels retained their biocompatability.In conclusion,the hydrogels prepared in this study have good biocompatibility,mechanical properties and anti-enzymatic degradation properties.The hydrogel obtained in this study can not only serve as a cell scaffold,transport cells into the body;can also be used as medical dressings,wound healing to provide a humid environment,promote wound healing.It is expected that the covalently crosslinked hydrogels reported in this study could be used in drug delivery,cell transplantation,tissue filling,tissue repair.