The antifibrotic effects of plasminogen activation occur via prostaglandin E2 synthesis in humans and mice
Kristy A. Bauman, Scott H. Wettlaufer, Katsuhide Okunishi, Kevin M. Vannella, Joshua S. Stoolman, Steven K. Huang, Anthony J. Courey, Eric S. White, Cory M. Hogaboam, Richard H. Simon, Galen B. Toews, Thomas H. Sisson, Bethany B. Moore, Marc Peters-Golden
Kristy A. Bauman, Scott H. Wettlaufer, Katsuhide Okunishi, Kevin M. Vannella, Joshua S. Stoolman, Steven K. Huang, Anthony J. Courey, Eric S. White, Cory M. Hogaboam, Richard H. Simon, Galen B. Toews, Thomas H. Sisson, Bethany B. Moore, Marc Peters-Golden
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Research Article Pulmonology

The antifibrotic effects of plasminogen activation occur via prostaglandin E2 synthesis in humans and mice

Abstract

Plasminogen activation to plasmin protects from lung fibrosis, but the mechanism underlying this antifibrotic effect remains unclear. We found that mice lacking plasminogen activation inhibitor–1 (PAI-1), which are protected from bleomycin-induced pulmonary fibrosis, exhibit lung overproduction of the antifibrotic lipid mediator prostaglandin E2 (PGE2). Plasminogen activation upregulated PGE2 synthesis in alveolar epithelial cells, lung fibroblasts, and lung fibrocytes from saline- and bleomycin-treated mice, as well as in normal fetal and adult primary human lung fibroblasts. This response was exaggerated in cells from Pai1–/– mice. Although enhanced PGE2 formation required the generation of plasmin, it was independent of proteinase-activated receptor 1 (PAR-1) and instead reflected proteolytic activation and release of HGF with subsequent induction of COX-2. That the HGF/COX-2/PGE2 axis mediates in vivo protection from fibrosis in Pai1–/– mice was demonstrated by experiments showing that a selective inhibitor of the HGF receptor c-Met increased lung collagen to WT levels while reducing COX-2 protein and PGE2 levels. Of clinical interest, fibroblasts from patients with idiopathic pulmonary fibrosis were found to be defective in their ability to induce COX-2 and, therefore, unable to upregulate PGE2 synthesis in response to plasmin or HGF. These studies demonstrate crosstalk between plasminogen activation and PGE2 generation in the lung and provide a mechanism for the well-known antifibrotic actions of the fibrinolytic pathway.

Authors

Kristy A. Bauman, Scott H. Wettlaufer, Katsuhide Okunishi, Kevin M. Vannella, Joshua S. Stoolman, Steven K. Huang, Anthony J. Courey, Eric S. White, Cory M. Hogaboam, Richard H. Simon, Galen B. Toews, Thomas H. Sisson, Bethany B. Moore, Marc Peters-Golden

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Figure 6

Plasminogen activation stimulates PGE2 release in AECs.

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Plasminogen activation stimulates PGE2 release in AECs. (A) AECs fro...
(A) AECs from saline-treated mice were cultured at 1.5 × 106/ml on fibronectin-coated plates and serum starved for 24 hours. Cells were then treated with SFM, 10 U/ml uPA, 45 mU/ml PLG, or uPA + PLG for 24 hours. PGE2 was then measured by ELISA in cell supernatants; n = 4, **P < 0.01. (B) Mice were given i.t. saline or i.t. bleomycin on day 0. On day 14, AECs were purified. AECs from saline-treated, normal and bleomycin-treated mice (N-AEC and B-AEC, B) were cultured in SFM or with uPA plus PLG for 24 hours, and PGE2 was measured; n = 3 or more in all groups, **P < 0.01, ***P < 0.001. (C) AECs were purified from WT or Pai1–/– mice and were treated with SFM or uPA plus PLG for 24 hours before culture supernatants were analyzed for PGE2 production via ELISA; n = 4 or more in each group, *P < 0.05, **P < 0.01, ***P < 0.001. (D) AECs were purified from saline-treated mice and cultured at 1.5 × 106/ml on fibronectin-coated plates. Cells were serum starved overnight and then cultured in SFM, 10 U/ml uPA plus 45 mU/ml PLG, or with 50 mU/ml plasmin. Total RNA was prepared and analyzed for Cox2 via real-time RT-PCR. Values for each sample were first normalized to β-actin, then the mean value for the SFM group was normalized to 1. n = 2 per group, representative of 2 experiments.