Tissue engineering of the human corneal endothelium: From primary cell culture to transplantation of highly functional cell sheets

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The proposed research project aims at in vitro cultivation and subsequent transfer of human corneal endothelial cell (HCEC) sheets for tissue engineering purposes. To allow a straightforward translation into a pre-clinical setting, primary, non-immortalized HCEC cultures are investigated. Isolation protocols and culture conditions are adapted to ensure an appropriate cell proliferation and maintenance. Highly functional and viable HCEC sheets are generated by methods which comply with essential requirements for later clinical application (e.g. animal compound free culture conditions). A tunable thermo-responsive cell culture carrier that was previously developed by the applicants and that allows to meet particular requirements of HCEC is employed. Upon stimulated detachment the cell sheets need to preserve intact intercellular contacts as well as extracellular matrix to stably survive transfer, storage and transplantation. Towards this goal a comprehensive range of characterization techniques for cell sheet morphology, viability, and physiological function is applied. Gentle manipulation techniques as well as dedicated transfer tools are developed using an in vitro transplantation model. More significant criteria for the transplant quality beyond the sole endothelial cell density are identified and evaluated.