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J Biol Chem, Vol. 275, Issue 20, 14939-14948, May 19, 2000
From the Laboratory of Molecular Pathology, Department of
Pathology, the University of Texas Southwestern Medical Center,
Dallas, Texas 75235-9072
CD44 on lymphocytes binding to its
carbohydrate ligand hyaluronan can mediate primary adhesion (rolling
interactions) of lymphocytes on vascular endothelial cells. This
adhesion pathway is utilized in the extravasation of activated T cells
from the blood into sites of inflammation and therefore influences
patterns of lymphocyte homing and inflammation. Hyaluronan is a
glycosaminoglycan found in the extracellular matrix and is involved in
a number of biological processes. We have shown that the expression of
hyaluronan on the surface of endothelial cells is inducible by
proinflammatory cytokines. However, the manner through which hyaluronan
is anchored to the endothelial cell surface so that it can resist shear
forces and the mechanism of the regulation of the level of hyaluronan on the cell surface has not been investigated. In order to characterize potential hyaluronan receptors on endothelial cells, we performed analyses of cell surface staining by flow cytometry on intact endothelial cells and ligand blotting assays using membrane fractions. Hyaluronan binding activity was detected as a major species
corresponding to the size of CD44, and this was confirmed to be the
same by Western blotting and immunoprecipitation. Moreover, alterations in the surface level of hyaluronan after tumor necrosis factor-
Hyaluronan Anchoring and Regulation on the Surface of
Vascular Endothelial Cells Is Mediated through the Functionally
Active Form of CD44*
stimulation is regulated primarily by changes in the cell surface levels of the hyaluronan-binding form of CD44. In laminar flow assays,
lymphoid cells specifically roll on hyaluronan anchored by purified
CD44 coated on glass tubes, indicating that the avidity of the
endothelial CD44/hyaluronan interaction is sufficient to support
rolling adhesions under conditions mimicking physiologic shear forces.
Together these studies show that CD44 serves to anchor hyaluronan on
endothelial cell surfaces, that activation of CD44 is a major regulator
of endothelial surface hyaluronan expression, and that the non-covalent
interaction between CD44 and hyaluronan is sufficient to provide
resistance to shear under physiologic conditions and thereby support
the initial steps of lymphocyte extravasation.
*
This work was supported by National Institutes of Health
Grants R01 CA57571 and HL56746 and the Arthritis Foundation.The costs of publication of this
article were defrayed in part by the
payment of page charges. The article
must therefore be hereby marked
"advertisement" in
accordance with 18 U.S.C. Section
1734 solely to indicate this fact.
Established Investigator of the American Heart Association and a
recipient of a Clinical Scientist award from the Burroughs Wellcome
Fund. To whom reprint requests should be addressed: Dept. of Pathology,
the University of Texas Southwestern Medical Center, 5323 Harry Hines
Blvd., Dallas, TX 75390-9072. Tel.: 214-648-4121; Fax: 214-648-4070;
E-mail: siegelman@utsw.swmed.edu.
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