282315 The Kinetics of Fibrin Clot Formation On Surfaces Immobilized with Cell and Fibronectin Binding Domains

Thursday, November 1, 2012: 8:30 AM
Pennsylvania East (Westin )
Anand Ramanathan and Nancy Wangechi Karuri, Chemical and Biological Engineering, Illinois Institute of Technology, Chicago, IL

Fibronectin (FN) is essential for the formation of a fibrin clot, an extracellular matrix that arrests bleeding. In the clot, FN binds fibrin and provides a scaffold for wound healing to occur. Our laboratory had previously shown that the ratio between the amount of a FN and cell binding domains, III1-2 and III9-10 respectively, immobilized on a surface influences the amount of FN in the ECM of cells. We investigated the effect of surfaces with different amounts of III1-2 and III9-10 on the kinetics of fibrin clot formation in a cell free system. Affinity purified glutathione tagged III1-2 and III9-10 were coated on polystyrene dishes. ELISA assays were used to identify the limits of protein loading on the surface and to quantify the levels of the individual proteins in mixtures. Fibrinogen was added to surfaces immobilized with III1-2 and III9-10, fibrin clot formation was proteolytically activated by the addition of thrombin and followed through dynamic light scattering. The kinetics of clot formation were studied in the presence and absence of full-length FN and with or without the covalent cross-linker, Factor XIIIA. We found that the rate of fibrin clot assembly was dependent on the ratio of III1-2 to III9-10 and higher rates were obtained when the ratio of III1-2 to III9-10 was high.  Moreover, the effect of the III1-2 and III9-10 coated surfaces on the rate of clot formation was significantly enhanced by FN and factor XIIIA. These results in a cell free system coupled together with our previous findings provide new avenues for the design of materials that can be used to accelerate wound healing.

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