Preservation of heparin-binding EGF-like growth factor activity on heparin-modified poly(-isopropylacrylamide)-grafted surfaces.

A heparin-modified poly(-isopropylacrylamide) (PIPAAm)-grafted surface bound with heparin-binding epidermal growth factor-like growth factor (HB-EGF) was able to culture hepatocytes maintaining high albumin secretion and high expression of hepatocyte-specific genes. However, the activity of HB-EGF on the surface and its binding effects on hepatocytes remain unclear. In this study, we investigated the temperature-dependent interactions of HB-EGF and EGF receptor (EGFR) with heparin-modified PIPAAm to evaluate the activity of HB-EGF on the surface. Quartz crystal microbalance (QCM) measurements revealed that the amounts of adsorbed HB-EGF on either the heparin-modified PIPAAm-grafted surface (heparin-IC1) or PIPAAm-grafted surfaces were almost the same regardless of swelling/deswelling of grafted PIPAAm chains. The heparin-IC1 surface bound to HB-EGF at 37 °C had the ability to bind to hepatocytes through specific affinity interaction with EGFR, whose activation was confirmed by western blotting. However, the physisorbed HB-EGF on the PIPAAm surface greatly diminished its activity. Taken together, the introduction of heparin into grafted PIPAAm chains on the surface plays a pivotal role in holding HB-EGF while preserving its activity. Hydration and swelling of surface-grafted PIPAAm chains at 20 °C greatly diminished the attachment of hepatocytes with HB-EGF bound to heparin-IC1, whereas hepatocytes were able to bind to HB-EGF bound to heparin-IC1 at 37 °C. Thus, the equilibrated affinity interaction between EGFRs and surface-bound HB-EGF was considered to be attenuated by steric hindrance due to hydration and/or swelling of grafted PIPAAm chains.