Immobilization of isolated functionalities of thrombomodulin on surfaces for improved hemocompatibility and evaluation of their physiological relevance
Biomaterials
Final Report Abstract
Methods for conjugation and immobilization of the proteases Activated Protein C (APC) and thrombin with low loss of activity were developed. A number of inhibitor molecules with affinity for thrombin and for APC have been tested concerning inhibitor constants and have been surface immobilized. A thrombomodulin‐like thrombin‐triggered APC release system could be set up using these inhibitor im‐ mobilized surfaces. The successful construction was proven in buffer systems with the respective proteins. The system however did not provide sufficient anticoagulant capacity in whole blood. A non‐intended alternative system was set up, where the inhibitor (heparin) is structural component of a hydrogel. Thrombin responsitivity was achieved by linker peptides with a thrombin‐cleavable sequence. This system also copies the thrombomodulin function, in the sense that an anticoagulant (here heparin instead of APC) is released only in response to the trigger thrombin, i.e. to activated coagulation. Similar to APC, the inhibitor released by the thrombin/thrombomodulin complex, heparin also can inhibit an overstoichiometric number of coagulation factors. Hydrogels with different gel characteristics were pre‐ pared and evaluated concerning thrombin affinity, accessibility of the gel structure for thrombin and for antithrombin, stability and specifity for the degradation by thrombin. In whole blood it exhibits excellent anticoagulant properties, it can quench coagulation also initiated by other sources and is the only material in the test, which could maintain non‐anticoagulated blood liquid for more than two hours. This concept does not only copy the thrombomodulin function, but it also presents one of the few so far developed bioresponsive feedback‐control systems, where the response has direct influence on its trigger. The comparison of various linker peptides in the hydrogel with different turn‐over speed and sensitivity also to the more upstream coagulation factor FXa indicates that response to early coagulation factors is more efficient than a fast release of the anticoagulant by thrombin. The physiological model thrombomodulin induces anticoagulant APC formation in response to thrombin; systems which mimic the thrombomodulin function may even surpass their original by the response to other coagulation factors.
Publications
- Application of reversible inhibitors for hemocompatible release systems (9th World Biomaterials Congress 2012, Chengdu)
Manfred F. Maitz, Anne Kleppisius, Carsten Werner
- Application of reversible inhibitors for hemocompatible release systems (Jahrestagung der Gesellschaft für Thrombose und Hämostaseforschung, 2012)
Manfred F. Maitz, Anne Kleppisius, Carsten Werner
- Bio-responsive anticoagulant hydrogels for anticoagulant surfaces, 9th World Biomaterials Congress 2012, Chengdu
Manfred F. Maitz, Uwe Freudenberg, Mikhail V. Tsurkan, Theresa Beyrich, Carsten Werner
- Thrombin responsive biofeedback system as thromboresistant material (Jahrestagung der Gesellschaft für Thrombose und Hämostaseforschung, 2012)
M.F. Maitz, T. Beyrich, M. Tsurkan, U. Freudenberg, C. Werner
- Bio‐responsive polymer hydrogels homeostatically regulate blood coagulation. Nature Commun 4:2168 (2013)
Manfred F. Maitz, Freudenberg U., Tsurkan M.V., Fischer M., Beyrich T., Werner C.
(See online at https://doi.org/10.1038/ncomms3168)
