Lilly Research Laboratory , Indianapolis , IN .
Blood 84 : 3700-8 ( 1994)
Abstract
Bovine vascular smooth muscle cells ( SMC ) express the urokinase-type plasminogen activator receptor ( u-PAR ) claimed to be important in cell invasion .
Receptor numbers and affinity are regulated by thrombin and several other mitogens involved in SMC proliferation .
We investigated the effects of these mitogens on u-PAR mRNA levels .
On continuous thrombin stimulation the u-PAR message in SMC was 10 +/- 2.3-fold elevated reaching a maximum between 6 and 9 hours and declining to control values within 48 hours .
Thrombin present for 30 minutes on the cell surface produced similar effects .
Stimulation with the thrombin receptor activation peptide S-F-L-L-R-N representing the NH2-terminus of the tethered ligand also increased u-PAR mRNA levels with an identical time course .
D-Phe-Pro-Arg-chloromethyl ketone ( PPACK ) active site blocked thrombin and the catalytically inactive thrombin mutant S205A did not affect u-PAR mRNA levels .
Thrombin stimulation also resulted in a 2 +/- 0.2-fold transient increase in thrombin receptor mRNA preceding the rise in u-PAR message .
Transforming growth factor beta 1 ( TGF beta ) and platelet-derived growth factor ( PDGF ) showed similar time courses for the elevation of u-PAR mRNA levels with a maximal 5.5 +/- 0.9 and +/- 2.5-fold increase , respectively .
Basic fibroblast growth factor ( bFGF ) and phorbol myristate acetate ( PMA ) showed a more prolonged effect increasing u-PAR mRNA levels 8 +/- 2.0-fold and 12.3 +/- . 5-fold , respectively , within 6 hours but remaining 5 to 10-fold elevated at 48 hours .
In order to decide if the u-PAR mRNA increase was due to message stabilization or a consequence of transcriptional activation we used the RNA polymerase II inhibitor , 6-dichloro-1-beta-D-ribofuranosyl benzimidazole ( DRB ) during the stimulation experiments .
u-PAR mRNA levels on TGF beta 1 stimulation of SMC decayed after the addition of DRB indicating that enhancement of transcriptional activity was involved in the induction .
In contrast , the time course of u-PAR mRNA elevation on thrombin , bFGF , and PMA stimulation was not significantly altered in the presence of DRB suggesting that in these latter cases u-PAR mRNA message accumulation was at least in part due to mRNA stabilization .
Increased transcriptional activity , mRNA stabilization and expression of u-PAR protein on the SMC surface in response to growth factors may facilitate enhanced cell surface protease activity , cell migration , and development of atheromatous lesions .