The covalent grafting of fibronectin at the surface of poly(ethylene terephthalate) track-etched membranes improves adhesion of rat hepatocytes

Abstract

[H-3]methylated fibronectin (FN) has been immobilized on the surface of poly(ethylene terephthalate) track-etched membranes (PET), carboxylated and then activated or not with water-soluble carbodiimide (WSC). Upon washing in 10% SDS or in the nutritive medium used for hepatocytes cultivation (supplemented with 15% newborn calf serum), respectively, 76% and 43% of [H-3]FN are released from the unactivated PET membranes and those WSC activated. This difference is almost totally abolished when the -NH2 functions of FN have been fully acetylated, to impair their reaction with activated -COOH groups. These results strongly suggest that part of FN is covalently grafted to the activated -COOH of PET but that, in addition, FN is also adsorbed on this surface. Rat hepatocytes were inoculated on PET membranes on which FN had been adsorbed and/or grafted. Image analysis clearly indicates that, during the first hours of culture, the FN immobilization on WSC activated, carboxylated PET membranes, significantly favours the adhesion of hepatocytes. After 24 h, the difference between these substrates decreases, probably due to the reconditioning of the PET surface by extracellular matrix constituents secreted by the hepatocytes. Our results confirm that the nature of the protein-polymer interaction strongly affects cell behaviour.