Molecular and Cellular Biology of Plasminogen Activation

i014i
Activated Human Neutrophils Rapidly Release the Chemotactically
Active D2D3 Form of the Urokinase-type Plasminogen Activator
Receptor (uPAR/CD87)
Pliyev BK* 1,2 and Tkachuk VA 1,2
1Department of Biological and Medical Chemistry, School of Basic Medicine, Moscow State University,
Moscow, Russia;
2Molecular Endocrinology Laboratory, Institute of Experimental Cardiology, Cardiology Research
Center, Moscow, Russia
Presenting author e-mail: bpliyev@cardio.ru
The urokinase-type plasminogen activator receptor (uPAR/CD87) exists both in cell-bound and
soluble forms. Soluble uPAR (suPAR) is readily detected in blood and is markedly increased
during inflammation and cancer. Neutrophils contain extensive intracellular pools of uPAR that
are translocated to the plasma membrane upon activation. In the present study, we investigated
the ability of human neutrophils to shed uPAR from cell surface following activation and
addressed the possible involvement of the released receptor in inflammatory response. We
first observed that resting neutrophils spontaneously release suPAR. This release was strongly
and rapidly (within minutes) enhanced by calcium ionophore ionomycin and to a lesser extent
when cells were primed with TNF-alpha or LPS and then stimulated with fMLP or IL-8. We
demonstrate that suPAR is produced by resting and activated neutrophils predominantly as a
truncated form devoid of N-terminal D1 domain (D2D3 form) that lacks GPI anchor. Migration
of human embryonic kidney (HEK) 293 cells stably transfected with the human formyl peptide
receptor FPRL1 towards the supernatants harvested from activated neutrophils was significantly
diminished when suPAR was immunodepleted from the supernatants. We conclude that activated
neutrophils release the chemotactically active D2D3 form of suPAR that acts as a ligand of
FPRL1. The release of suPAR by activated cells was significanty but not completely inhibited by
alpha 1-antichymotrypsin, a specific inhibitor of cathepsin G, indicating that cathepsin G acts as
a shedding protease for membrane-bound uPAR. Neutrophils isolated from synovial fluids of
rheumatoid arthritis patients released significantly (p < 0.01) higher amounts of suPAR compared
with cells isolated from pared peripheral blood samples suggesting that the release of suPAR by
neutrophils is increased in vivo in the sites of acute inflammation. We suggest that production of
the chemotactically active D2D3 form of suPAR by activated human neutrophils in vivo could
contribute to the recruitment of monocytes and other formyl peptide receptors-expressing cells to
the sites of acute inflammation where neutrophils accumulation and activation occur.
26 X I t h I n t e r n a t i o n a l W o r k s h o p o n