Data on t-PA release into serum fro

Data on t-PA release into serum from hydrogels of different DCL are shown in Fig. 3. When the hydrogels were incubated in serum without thrombin, only small amounts of t-PA were released,possiblybydiffusion.Muchgreatert-PAreleaseoccurred on exposure to thrombin-containing serum, suggesting that t-PA was released mainly by thrombin-triggered hydrogel degradation rather than by simple diffusion. The release of t-PA from all three hydrogels increased with increasing thrombin concentration and the release rates were essentially linear. Moreover, there was no initial burst release or lag phase, presumably because the hydrogels were degraded at a constant rate. Such enzyme-triggered release is thus superior to release driven by diffusion, where an initial burst release phase is normally experienced.25 The thrombin-responsive release of t-PA was also investigated using PBS instead of serum as the medium (Fig. S6, ESI†). Release kinetics in the two media were essentially the same, indicating that the ‘‘non-specific’’ proteins in the serum did not influence hydrogel degradation and t-PA release. t-PA release rates at different thrombin concentrations in serum were estimated from the data in Fig. 3a–c. The release rate decreased with increasing DCL, presumably due to the lower degradation rates at higher DCL (Fig. 3d). Moreover, the release rate forthe hydrogel with thelowest DCL increased more rapidly as the thrombin concentration increased than for the mediumand high-DCL gels. It appears, therefore, that not only the release rate but also the responsiveness to the thrombin concentration is dependent on the degree of crosslinking. To investigate whether the thrombin-responsive release of t-PA can be switched on and off in response, respectively, to the presenceandabsenceofthrombin,experimentswerecarriedout inwhich, at a designated time, the releasemedium was changed from thrombin-containing serum to normal serum. As shown in Fig. 3e, t-PA was released from the hydrogels at different rates, depending on DCL, in the presence of serum containing 10 U mL1 thrombin. When after 2 h the hydrogels were placed in normal serum without thrombin, t-PA release stopped for all three gels.