Item Details

title

IMPORTANCE OF THE FIBRINOGEN αC-CONNECTOR IN FIBRIN FIBER MECHANICAL PROPERTIES AND AUTOMATED FIBER DIAMETER MEASUREMENTS IN SEM IMAGES OF FIBRIN CLOTS

author

Daraei, Ali

abstract

Fibrinogen is the key mechanical protein in blood coagulation since it is the building block of fibrin fibers, and these 100 nm thick fibers provide mechanical and structural stability to blood clots as they stem the flow of blood. Fibrinogen has two large, intrinsically unfolded regions, termed αC regions, which comprise 27% of its molecular weight. The role of these unfolded regions has long puzzled scientists. We hypothesize that they contribute significantly to the mechanical properties of fibrin fibers, which, in turn, determine the stability of blood clots.We used a nano-mechanical manipulation method, based on a combined atomic force and optical microscope, to determine the mechanical properties of native fibrin fibers, and of fibers formed from a variant in which amino acids 264-391 in the αC-connector were deleted. Compared to native fibrin fibers, fibers formed from this variant showed dramatically different mechanical properties. The extensibility (fracture strain) was reduced by a factor of 1.7 (from 2.1 to 1.26) and the modulus (stiffness) decreased by a factor of 3 (from 3 MPa to 1 MPa). We also evaluated the accuracy of a plug-in for the open-source, NIH-supported image analysis program ImageJ, called DiameterJ, in determining the diameters of fibers in Scanning Electron Microscopy (SEM) images. We found that for SEM images with an optimal pixel size of 8-10 nm (13-16 pixels/fiber) several DiameterJ algorithms strongly correlated with the manual measurements. We also evaluated the emerging best subsets of the 24 algorithms on two patient data sets to see if the trends in samples from healthy individuals and patients that were detected by manual measurements can be reliably detected by these automated algorithms. Finally, by combining experimental AFM-based single-molecule dynamic force assays with a theoretical reconstruction of the force-extension curves, we were able to resolve the structural underpinnings of the mechanical deformation and rupture of fibrin fibers. We performed AFM-based mechanical testing of single uncrosslinked fibrin fibers in vitro and in silico to explore their nanomechanical and material properties.

subject

Alpha C connector

Extensibility

Fibrin Fibers

Fibrinogen

Modulus

Strain Stiffening

contributor

Guthold, Martin M (committee chair)

Kim-Shapiro, Daniel D (committee member)

Bonin, Keith K (committee member)

Baker, Stephen S (committee member)

Hudson, Nathan N (committee member)

date

2021-01-13T09:35:30Z (accessioned)

2021-01-13T09:35:30Z (available)

2020 (issued)

degree

Physics (discipline)

identifier

http://hdl.handle.net/10339/97961 (uri)

language

en (iso)

publisher

Wake Forest University

type

Dissertation