IADR Abstract Archives
Urokinase Plasminogen Activator (uPA) Activity in OSCC Cell Conditioned Media
Objectives: The protease uPA (urokinase plasminogen activator) and its inhibitor PAI-1 (plasminogen activator inhibitor-1) are biomarker proteins used to manage therapy of cancer patients. High uPA and PAI-1 measured by ELISA in tumor biopsies are associated with an invasive, metastatic phenotype in breast cancer. uPA and PAI-1 are often high in other tumor types including OSCC (oral squamous cell carcinoma). Objectives: (1) Measure uPA and PAI-1 in media from cultured OSCC and normal oral tissue cell lines to determine whether OSCC cells secrete high levels of both uPA and PAI-1. (2) Measure uPA enzyme activity to determine whether PAI-1 blocks uPA activity.
Methods: uPA and PAI-1 were measured by ELISA, and uPA activity was measured with plasminogen as the substrate. A line of normal gingival fibroblasts and Smulow-Glickman gingival epithelial cells were compared with Cal27, SCC-25, and SCC-15 lines derived from squamous cell carcinoma of the tongue (ATCC® CRL-2095, -1628, -1623™, respectively).
Results: (1) uPA was much higher in media from all three OSCC lines than from fibroblasts or epithelial cells. uPA was 1890±27, 834±49, and 401±11 ng/mg protein in Cal27, SCC-25, and SCC-15 media, respectively. In contrast, Cal-27 cells did not secrete PAI-1, and PAI-1 was high in media from only one OSCC cell line (SCC-15). (2) uPA enzyme activity was higher in Cal27 media (241±31 U/mg) than SCC-15 media (39±3 U/mg). This 6-fold difference in activity was consistent with the 5-fold difference in the amount of uPA, indicating that PAI-1 did not block uPA activity. The results suggest that PAI-1 was in the latent form that does not inhibit uPA.
Conclusions: Secretion of high levels of PAI-1 in vivo may not indicate that uPA activity is blocked. Therefore, uPA activity may activate proteases, leading to proteolytic breakdown of the extracellular matrix and increased tumor invasion and metastasis.
Methods: uPA and PAI-1 were measured by ELISA, and uPA activity was measured with plasminogen as the substrate. A line of normal gingival fibroblasts and Smulow-Glickman gingival epithelial cells were compared with Cal27, SCC-25, and SCC-15 lines derived from squamous cell carcinoma of the tongue (ATCC® CRL-2095, -1628, -1623™, respectively).
Results: (1) uPA was much higher in media from all three OSCC lines than from fibroblasts or epithelial cells. uPA was 1890±27, 834±49, and 401±11 ng/mg protein in Cal27, SCC-25, and SCC-15 media, respectively. In contrast, Cal-27 cells did not secrete PAI-1, and PAI-1 was high in media from only one OSCC cell line (SCC-15). (2) uPA enzyme activity was higher in Cal27 media (241±31 U/mg) than SCC-15 media (39±3 U/mg). This 6-fold difference in activity was consistent with the 5-fold difference in the amount of uPA, indicating that PAI-1 did not block uPA activity. The results suggest that PAI-1 was in the latent form that does not inhibit uPA.
Conclusions: Secretion of high levels of PAI-1 in vivo may not indicate that uPA activity is blocked. Therefore, uPA activity may activate proteases, leading to proteolytic breakdown of the extracellular matrix and increased tumor invasion and metastasis.