Smart biomaterials for cell sheet engineering
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The aim of this instigation was to develop thermoresponsive polymer films for cell culture use. Conventional cell detachment methods involve the use of proteolytic enzymes, such as trypsin, or mechanical scrapping. These detachment methods yield either individual cells or cell clusters, neither of which is attractive from a tissue engineering perspective. Temperature controlled cell detachment via thermoresponsive polymer films is an attractive alternative, as upon temperature reduction below the chosen polymers lower critical solution temperature (LCST) the polymer film will either swell or dissolve facilitating the detachment of a cell sheet with intact cell-cell junctions. This work describes the use of physical adsorption as a method of thermoresponsive film preparation. Physical adsorption is routinely used to coat surfaces with cell adhesion promoters and it is one of the simplest methods available for altering surface properties. Four polymers were used in this study; (1) poly N-(isopropylacrylamide) (pNIPAm), (2) NIPAm-co-N-tertbutylacrylamide (NtBAm), (3) NIPAm-co-acrylamidobenzophenone (AcBzPh) and (4) NIPAm-co-NtBAm-co-AcBzPh. These were chosen due to their unique advantages: (1) polymerization circumvention, (2 and 4) hydrophobicity and (3 and 4) crosslinking capability. Polymer films were formed both above and below the LCST in order to determine whether or not the temperature induced changes in the polymers’ conformation affected the film’s physical and chemical properties and their ability to successfully host cells. The physically adsorbed film’s roughness, thickness, percentage composition and wettability were determined. Human pulmonary microvascular endothelial cells were seeded on the prepared films and cell viability was assessed qualitatively using light microscopy and quantitatively using cell viability assays. Electron microscopy was used to investigate the detached cell sheets. The cell culture potential of a new class of thermoresponsive copolymers was investigated. These copolymers swell rather than dissolving upon temperature reduction without the need for a covalent bond between the polymer layer and the underlying substrate.