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This Article Full Text Full Text (PDF) All Versions of this Article: 293/2/L328    most recent 00390.2006v1 Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in ISI Web of Science Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Citing Articles Citing Articles via Google Scholar Google Scholar Articles by Stevens, A. P. Articles by Dull, R. O. Search for Related Content PubMed PubMed Citation Articles by Stevens, A. P. Articles by Dull, R. O.

Fluorescence correlation spectroscopy can probe albumin dynamics inside lung endothelial glycocalyx

¹Department of Bioengineering, Proteins and Polymers at Interface Group, University of Utah, Salt Lake City; and ²Department of Anesthesiology, Lung Vascular Biology Laboratory, and ³Department of Pharmaceutics and Pharmaceutical Chemistry, University of Utah School of Medicine, Salt Lake City, Utah

Submitted 3 October 2006 ; accepted in final form 30 April 2007

The endothelial glycocalyx is believed to play a major role in capillary permeability by functioning as a macromolecular barrier overlying the intercellular junction. Little is known about the functional attributes of the glycocalyx (i.e., porosity and permeability) or which constituents contribute to its overall structure-function relationship. In this report, we demonstrate the utility of fluorescence correlation spectroscopy (FCS) to measure albumin diffusion rates and concentration profiles above the cell surface and overlying the intercellular junctions of lung capillary endothelial cells. Albumin diffusion rates and concentration profiles were obtained before and after enzymatic digestion of the glycocalyx with pronase, heparanase, or hyaluronidase. The results suggest a structure interacting with albumin located from 1.0 to 2.0 µm above the cell membrane capable of reducing albumin diffusion by 30% while simultaneously increasing albumin concentration fivefold. Digestion of the glycocalyx with pronase or heparanase resulted in only modest changes in albumin diffusion and concentration profiles. Hyaluronidase digestion completely eliminated albumin-glycocalyx interactions. These data also suggest that hyaluronan is a major determinant for albumin interactions with the lung endothelial glycocalyx. Confocal images of heparan sulfate and hyaluronan confirm a cell-surface layer 2–3 µm in thickness, thus supporting FCS measurements. In summary, we report the first use of FCS to probe extracellular structures and further our understanding of the structure-function relationship of the lung microvascular endothelial glycocalyx.

diffusion; permeability