Plasminogen activator inhibitor-1 mRNA expression in cultured pigmented ciliary epithelial cells of the porcine eye

Abstract

Background. Plasminogen activator inhibitor-1 (PAI-1) is a main regulator of the fibrinolytic system. PAI-1 inhibits tissue plasminogen activator and urokinase-type plasminogen activator, resulting in reduced plasminogen activity and attenuated fibrinolysis and proteolysis. The present study was performed to determine the gene expression encoding for PAI-1 in cultured pigmented ciliary epithelial cells of the porcine eye and to detect PAI-1 activity in cell culture supernatants.

Methods. Total mRNA of respective confluent primary cultures of porcine ciliary epithelial cells, porcine liver cells and porcine kidney cells was isolated. Reverse transcribed PAI-1 mRNA was measured by real-time polymerase chain reaction (TaqMan PCR) with PAI-1 primers and probes deduced from the human PAI-1 gene. PAI-1 activity in supernatants of the cell cultures was determined by a specific chromogenic test (Coatest PAI).

Results. PAI-1 mRNA was localized in all samples of primary cultures of porcine pigmented ciliary epithelial cells. As a negative control we analyzed total mRNA of porcine kidney cells. PAI-1 mRNA was not detectable in these cells. On the other hand, we established PAI-1 mRNA in porcine liver cells as a positive control. High levels of PAI-1 activity were found in all samples of cell culture supernatants.

Conclusions. Our results indicate that PAI-1 is produced and secreted by the porcine ciliary epithelium. We suggest that PAI-1 together with components of the fibrin/fibrinolytic system may be involved in aqueous humor outflow. Overproduction of PAI-1 may induce less fibrinolysis and extracellular proteolysis in aqueous humor and trabecular meshwork, which could result in an elevated intraocular pressure by increasing the outflow obstruction. Therefore stimulation of PAI-1 production may perhaps contribute to the pathogenesis of primary open-angle glaucoma.