The most significantly upregulated gene by both cmvIL-10 and hIL-10 was PAI-1, or plasminogen activator inhibitor 1. PAI-1 is a 43 kDa glycoprotein that inhibits the function of urokinase plasminogen activator (uPA), a serine protease that catalyzes the conversion of inactive plasminogen to plasmin and has been implicated in many aspects of tumor progression . The activity of uPA system is regulated by the receptor uPAR and two endogenous inhibitors, PAI-1 and PAI-2 . PAI-1 is constitutively secreted by many cell types and high levels have been found to inhibit cell adhesion and promote migration [63, 64]. In order to confirm that changes in gene expression identified by the qPCR array correlated with protein expression, MDA cells were treated with cmvIL-10 or hIL-10 and PAI-1 levels measured by ELISA. As expected, PAI-1 was produced by untreated cells, however, the amount of protein secreted was significantly increased by cmvIL-10 after 12 h of exposure (Fig. 3a). After 24 h, both cmvIL-10 and hIL-10 stimulated a significant increase in PAI-1 production, and this was maintained over 72 h. In addition, we examined uPAR protein levels and found that they were also elevated upon exposure to cmvIL-10 or hIL-10 (Fig. 3b). These results demonstrate that expression of two elements of the uPA serine protease system, its receptor uPAR and its serpin inhibitor PAI-1, are significantly increased by cmvIL-10 and hIL-10 in human breast cancer cells.Fig. 3PAI-1 and uPAR levels are elevated upon exposure to cmvIL-10 or hIL-10. a MDA-MB-231 cells were cultivated in the presence of 10 ng/ml purified recombinant cmvIL-10 or hIL-10. At the indicated time points, culture supernatants were collected and levels of PAI-1 measured by ELISA. b MDA cells were cultured as above and levels of uPAR in the supernatant were determined by ELISA. Error bars represent standard error among three replicates for each data point. *p < 0.05. These results are representative of three independent experiments