Role of curing temperature of poly(glycerol sebacate) substrates on protein-cell interaction and early cell adhesion

Abstract

A novel procedure to obtain smooth, continuous polymeric surfaces from poly(glycerol sebacate) (PGS) has been developed with the spin-coating technique. This method proves useful for separating the effect of the chemistry and morphology of the networks (that can be obtained by varying the synthesis parameters) on cell-protein-substrate interactions from that of structural variables. Solutions of the PGS pre-polymer can be spin-coated, to then be cured. Curing under variable temperatures has been shown to lead to PGS networks with different chemical properties and topographies, conditioning their use as a biomaterial. Particularly, higher synthesis temperatures yield denser networks with fewer polar terminal groups available on the surface. Material-protein interactions were characterised by using extracellular matrix proteins such as fibronectin (Fn) and collagen type I (Col I), to unveil the biological interface profile of PGS substrates. To that end, atomic force microscopy (AFM) images and quantification of protein adsorbed in single, sequential and competitive protein incubations were used. Results reveal that Fn is adsorbed in the form of clusters, while Col I forms a characteristic fibrillar network. Fn has an inhibitory effect when incubated prior to Col I. Human umbilical endothelial cells (HUVECs) were also cultured on PGS surfaces to reveal the effect of synthesis temperature on cell behaviour. To this effect, early focal adhesions (FAs) were analysed using immunofluorescence techniques. In light of the results, 130 °C seems to be the optimal curing temperature since a preliminary treatment with Col I or a Fn:Col I solution facilitates the formation of early focal adhesions and growth of HUVECs.