The Study Of Zwitterionic Molecules For Cell Cryopreservation

Living cells are critical in some important technologies,such as cell therapy and tissue engineering.Cell preservation that can maintain the activity and functions of cells for a considerable period,has become the support of these important technologies.Cryopreservation is a most reliable technology for cell preservation,but it suffers from the intrinsic toxicity of current state-of-the-art cryoprotectant(CPA),dimethyl sulfoxide(DMSO).In this thesis,aiming to the scientific issue of cell cryoinjuries induced by ice crystals,a variety of novel efficiently and biocompatible CPA compositions based on zwitterionic molecules were designed and developed,and their abilities of cryoprotection were systematically investigated,achieving DMSO-free cryopreservation of human cells.The effective preservation of chondrocytes is a support for cartilage tissue engineering technology.In this thesis,we firstly used zwitterionic osmoprotectant betaine that can enter cells as intracellular CPA,combined with zwitterionic polymers,poly(sulfobetainemethacrylate)(PSBMA)or poly(carboxybetaine methacrylate)(PCBMA)as extracellular CPA.The ability of osmotic regulation,the prevention of ice formation/growth,and cytotoxicity of zwitterionic compositions were systematically evaluated.The optimum propotion of zwitterionic molecule and polymer can achieve?90% post-thaw survival efficiency of chondrocytes.Meanwhile,zwitterionic combination presented no cytotoxicity,superior to DMSO.To solve the problems of current complex CPA removal process inducing the risk of sample contamination,the dual thermosensitive zwitterionic polymer poly(SBMAco-NIPAAm)was designed and synthesized to achieve a phase transition into a gel state below upper critical solution temperature(UCST)(freezing temperature of 0 ?)and above lower critical solution temperature(LCST)(physical temperature of 37 ?).The results showed below the freezing point temperature,the phase transition of thermosensitive polymer could effectively inhibit the formation of ice crystals,resisting the extracellular cryoinjuries,and the polymer was combined with the zwitterionic betaine as an intracellular CPA for effective GLC-82 cryopreservation.The post-thaw survival efficiency of GLC-82 cells was as high as 96%.After recovery to physiological temperature,the polymer underwent a phase change to form gel-embedded cells without the need of CPA removal,expected to be directly applied for tissue engineering.This thesis was based on zwitterionic molecules to develop the novel biocompatible and highly effcient CPAs,providing strong support for cell-based applications including tissue engineering and cell therapy.