title: Thermo-responsive poly(NiPAAm-co-DEGMA) substrates for gentle harvest of human corneal endothelial cell sheets.
abstract: Gentle harvesting of corneal endothelial cell sheets grown in culture is of interest for the development of cornea replacement strategies. Thin films of a fast responding copolymer of N-isopropylacrylamide (NiPAAm) and diethyleneglycol methacrylate (DEGMA) with a phase transition temperature of 32 degrees C were prepared and evaluated for that purpose. The polymer layers were immobilized onto fluorocarbon substrates using low pressure argon plasma treatment. Cell culture and detachment experiments were performed with L929 mouse fibroblasts and human corneal endothelial cells (HCEC) at standard conditions. The hydrogel-coated supports were found to permit adhesion, spreading, and proliferation of both cell types. Harvesting of cell sheets was achieved upon lowering the temperature to about 30 degrees C. The formation of a closed monolayer as a crucial prerequisite for maintaining ionic pump function in HCEC was proven by ZO-1 immunostainung. Labeling of fibronectin indicated that the vast majority of the extracellular matrix is detached from the hydrogel coatings together with the cell layer. Inspired by this result, the reuse of the hydrogel-coated culture carriers was investigated confirming the suitability of the substrates for repeated cell harvesting. Altogether, the introduced thermoresponsive coating was found advantageous for the efficient generation of HCEC sheets and will be further utilized in transplantation strategies.
authors.affiliation: Leibniz Institute of Polymer Research Dresden, Max Bergmann Center of Biomaterials Dresden, Hohe Strasse 6, 01069 Dresden, Germany. nitschke@ipfdd.de
authors.forename: Mirko
authors.initials: M
authors.lastname: Nitschke
authors.forename: Stefan
authors.initials: S
authors.lastname: Gramm
authors.forename: Thomas
authors.initials: T
authors.lastname: Götze
authors.forename: Monika
authors.initials: M
authors.lastname: Valtink
authors.forename: Juliane
authors.initials: J
authors.lastname: Drichel
authors.forename: Brigitte
authors.initials: B
authors.lastname: Voit
authors.forename: Katrin
authors.initials: K
authors.lastname: Engelmann
authors.forename: Carsten
authors.initials: C
authors.lastname: Werner
mesh_terms: D000818:Animals; D002448:Cell Adhesion; D018929:Cell Culture Techniques; D002461:Cell Line, Transformed; D020099:Coated Materials, Biocompatible; D003315:Cornea / Q000166:cytology*; D003316:Corneal Diseases / Q000628:therapy; D042783:Endothelial Cells / Q000166:cytology*; D005347:Fibroblasts / Q000166:cytology*; D006801:Humans; D008689:Methacrylates / Q000737:chemistry; D051379:Mice; D011109:Polymethacrylic Acids / Q000737:chemistry; D023822:Tissue Engineering
chemical_list: C512139:2-(2-hydroxyethoxy)ethyl methacrylate; D020099:Coated Materials, Biocompatible; D008689:Methacrylates; D011109:Polymethacrylic Acids; C030613:polymethacrylic acid