The Mechanism of the Formation of Nonadhesive Fibrinogen Matrices Examined by Atomic Force Microscopy

Abstract

The plasma protein fibrinogen prevents adhesion of blood cells such as platelets and leukocytes to various surfaces, including fibrin gels. We have previously reported that the decrease in cell adhesion results from fibrinogen aggregation and the formation of an extensible multilayered matrix incapable of transducing strong mechanical forces through cellular integrins1,2. Here, using the various capabilities of Atomic Force Microscopy (AFM), we have examined the physical properties of matrices produced from human plasma fibrinogen (hFg) and two recombinant fibrinogens: recombinant normal fibrinogen (rFg) and fibrinogen with truncated αC domains (FgAα251). We have determined the thickness of matrices using AFM lithography, and also performed the high-resolution topographical visualization of single molecules within the matrices. With force spectroscopy, we have measured the extensibility, adhesion forces and the energy of AFM tip-fibrinogen matrix interactions. Our results indicate that hFg and rFg adsorbed at high concentrations form an extensible matrix containing 8-9 molecular layers. In contrast, FgAα251 forms only 2-3 molecular layers. These data indicate that the αC domains are critically involved in the formation of a fibrinogen multilayer. Furthermore, the inability of FgAα251 to form a thick multilayered matrix results in sustained cell adhesion, unlike hFg which completely prevents cell adhesion. To evaluate the contribution of the αC domains to the formation of multilayered matrices, we developed a model based on intermolecular domain interactions to simulate the adsorption process for hFg and FgAα251. These findings have implications for processes where deposition of fibrinogen occurs, such as thrombus formation and adsorption of fibrinogen on implanted biomaterials.1. Podolnikova, N. P. et al., Biochemistry, 49, 68-77 (2010).2. Yermolenko, I. S. et al., Langmuir, 26, 17269-17277 (2010).