Glycidyl Ether-Based Coatings on Polystyrene Culture Substrates for Temperature-Triggered Cell Sheet Fabrication

Abstract

Within this work, thermoresponsive coatings based on poly(glycidyl ether)s (PGEs) were developed for applied polystyrene (PS) tissue culture substrates. Following the “grafting to“ approach, block copolymers comprising a random, high molecular weight, thermoresponsive block and a short, hydrophobic benzophenone (BP) block were synthesized via the sequential, monomer-activated, oxy-anionic ring-opening polymerization (ROP). Ultrathin layers in the sub-nanometer range were immobilized on PS by physical adsorption and UV-induced C, H-insertion of PGE block copolymers via their photo-reactive BP anchor block. The coatings mediated the adhesion of human dermal fibroblasts (HDFs) and allowed the temperature triggered detachment of confluent cell sheets. HDF sheet detachment was found to be induced by the cooperative effects between the partial rehydration of the PGE chains and the cell repellant PS substrate background. In order to improve the performance of PGE monolayers, block copolymers were subsequently self-assembled on PS substrates from dilute aqueous solution under selective solvent conditions. UV immobilization yielded thermoresponsive polymer brushes, which undergo a “pancake-to-brush” transition upon temperature reduction. The improved structure and thermal response of the brush-like PGE coatings as well as the optimization of cell culture parameters facilitated the fabrication of confluent HDF, human aortic smooth muscle cell (HAoSMC) and human umbilical vein endothelial cell (HUVEC) sheets, which constitute the main building blocks of blood vessels. To functionalize PS culture substrates via the “grafting from” approach, a solvent-free, microwave-assisted synthesis of well-defined oligo(glycidyl ether)s (OGEs) was developed. Fast reaction rates could be solely attributed to the high reaction temperatures reached during microwave heating and the obtained oligomers exhibited highly molecular weight- and concentration-dependent CPTs in water. Further, end-functional oligo(glycidyl ether) acrylate (OGEA) macromonomers were synthesized by in situ quenching of the oxy-anionic ROP. Subsequently, a photopolymerization process was developed to graft OGEA macromonomers from PS culture substrates. Surfaceinitiated photografting from bulk macromonomer films yielded porous, rigid, gel-like OGEA coatings with unique bottlebrush properties. Bottlebrushes with optimized structure proved to be functional coatings for the fabrication of HDF sheets. The controlled detachment of cell sheets was found to be triggered by the rehydration of OGEA bottlebrush side chains rather than a macroscopic swelling of the gel-like coatings upon temperature reduction. In summary, this work introduces facile methods for the functionalization of applied PS tissue culture surfaces with thermoresponsive, PGE-based coatings and demonstrates their high potential as functional substrates for cell sheet fabrication.