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Biophys. J. BioFAST: First Published April 7, 2006. doi:10.1529/biophysj.105.075283
© 2006 by the Biophysical Society.

A more recent version of this article appeared on July 1, 2006.
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SPECTROSCOPY, IMAGING, OTHER TECHNIQUES

DIFFUSIONAL ANISOTROPY IN COLLAGENOUS TISSUES: FLUORESCENCE IMAGING OF CONTINUOUS POINT PHOTOBLEACHING

Holly A Leddy 1, Mansoor A. Haider 2 and Farshid Guilak 1*

1 Duke University Medical Center
2 North Carolina State University

* To whom correspondence should be addressed. E-mail: guilak{at}duke.edu.

Submitted on September 30, 2005
Revised on November 11, 2005
Accepted on 22 February 2006


  Abstract
Molecular transport in avascular collagenous tissues such as articular cartilage occurs primarily via diffusion. The presence of ordered structures in the extracellular matrix may influence the local transport of macromolecules, leading to anisotropic diffusion depending on the relative size of the molecule and that of extracellular matrix structures. Here we present a novel photobleaching technique for measuring the anisotropic diffusivity of macromolecules in collagenous tissues. We hypothesized that macromolecular diffusion is anisotropic in collagenous tissues, depending on molecular size and the local organization of the collagen structure. A theoretical model and experimental protocol for fluorescence imaging of continuous point photobleaching (FICOPP) was developed to measure diffusional anisotropy. Significant anisotropy was observed in highly ordered collagenous tissues such as ligament, with diffusivity ratios greater than 2 along the fiber direction compared to the perpendicular direction. In less ordered tissues such as articular cartilage, diffusional anisotropy was dependent on site in the tissue and size of the diffusing molecule. Anisotropic diffusion was also dependent on the size of the diffusing molecule, with greatest anisotropy observed for larger molecules. These findings suggest that diffusional transport of macromolecules is anisotropic in collagenous tissues, with higher rates of diffusion along primary orientation of collagen fibers.

Key Words: FRAP, collagen, diffusion, microphotolysis, proteoglycan




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Copyright © 2006 by the Biophysical Society.