Fig. 2.

The PAI-1 mediated increase in fibronectin matrix assembly does not require uPAR. A - Cell lysates were prepared from control siRNA or uPAR siRNA transfected MG-63 cells. uPAR and β5 integrin levels were analyzed by Western blotting. The membrane was stripped and reprobed with anti β-actin antibody as a loading control. B - Cells layers from control siRNA or uPAR siRNA transfected MG-63 cells were incubated with ¹²⁵I-fibronectin for 6 hours in the absence or presence of PAI-1 (20 nM). Cell layers were extracted with 1% DOC, and ¹²⁵I-fibronectin that was incorporated into the detergent-insoluble matrix was recovered by centrifugation and measured by gamma scintillation. Data are the mean ± S.E. of two experiments performed in duplicate. *Significantly different than non treated cells, t test, p < 0.05 (n=4). C - Cells layers from control siRNA or uPAR siRNA transfected MG-63 cells were incubated with ¹²⁵I-fibronectin for 6 hours in the presence of the uPAR agonist P25 or the control peptide, S25. Cell layers were washed and scraped into 1% DOC and cell layer associated ¹²⁵I-fibronectin was measured by gamma scintillation. Data are the mean ± S.E. of two experiments performed in duplicate. *Significantly different than control cells, t test, p < 0.05 (n=4).