-The alveolar fibrinolytic system is altered in acute lung injury (ALI). Levels of the fibrinolytic protease inhibitor, plasminogen activator inhibitor-1 (PAI-1), are too low in bronchoalveolar lavage to address its prognostic significance. This study was performed to assess whether PAI-1 antigen in undiluted pulmonary edema fluid levels can identify patients with ALI and predict their outcome. PAI-1 antigen levels in both plasma and edema fluid were higher in ALI compared with hydrostatic edema, and edema fluid PAI-1 values identified those with ALI with high sensitivity and specificity. Both the high plasma and edema fluid PAI-1 antigen values were associated with a higher mortality rate and fewer days of unassisted ventilation in patients with ALI. Differences in PAI-1 activity were concordant with levels of PAI-1 antigen. Although the fibrinderived alveolar D-dimer levels were strikingly similar in both groups, ALI patients had a higher relative proportion of D-monomer. In conclusion, PAI-1 levels in edema fluid and plasma identify those with ALI that have a poor prognosis. The data indicate that fibrin turnover in early ALI is a consequence of a rapid fibrinogen influx and fractional fibrinolytic inhibition.fibrinolysis; prognosis CURRENT ESTIMATES INDICATE that the incidence of acute lung injury (ALI) is 20-75 per 100,000 persons, with a mortality rate of 20-60% (48). Prior studies that have attempted to identify prognostic markers in bronchoalveolar lavage (BAL) fluid obtained from patients with ALI in the first several days after intubation have met with only modest success (7,8,15,37,38). Several obstacles exist in identifying valuable and reliable prognostic markers in ALI. For instance, it is clear that the lung inflammation evolves rapidly in ALI, making the timing of the analysis crucial. Second, there are both soluble activators and inhibitors for numerous cytokines/chemokines in the inflammatory milieu. Third, the BAL procedure itself dilutes the alveolar contents 100-fold, making it difficult to quantify potentially useful markers of ALI. Finally, the heterogeneity of underlying diseases and choice of control groups add to the difficulty in extrapolating the findings to disease pathobiology. We have overcome some of these obstacles by utilizing undiluted pulmonary edema fluid, not BAL fluid, that was obtained from patients with clinical ALI and well-defined hydrostatic pulmonary edema (HYDRO) controls within hours of intubation and initiation of mechanical ventilation.ALI is characterized by the deposition of fibrin in the alveolar space. Deposition of fibrin in the alveolar space is the net result of an alteration in the balance of coagulation and fibrinolytic proteases [plasmin and urokinase type plasminogen activators (u-PA)] and antiproteases [plasminogen activator inhibitor-1 (PAI-1) and ␣ 2 -antiplasmin] and the availability of plasmaderived fibrinogen (31,35). Upon fibrinogen influx into the alveolar compartment, coagulation/fibrinolysis proteases/antiproteases act to generate insoluble cr...

Although the fibroproliferative response to lung injury occurs with a high frequency in patients with clinical acute lung injury, the mechanisms that initiate this response are largely unknown. This study was undertaken first to identify fibroblast mitogenic factors in pulmonary edema fluid, and second to examine the human lung fibroblast’s gene expression profile in response to pulmonary edema fluid. The edema fluid obtained from patients with early lung injury has an eightfold higher concentration of IL-1β and a twofold greater IL-1β-dependent mitogenic effect than does fluid obtained from control patients with hydrostatic pulmonary edema. Furthermore, fibroblasts responded to acute lung injury patient-derived edema fluid through production of soluble mediators that possess an autocrine mitogenic effect. Gene array analysis reveals that acute lung injury edema fluid induces several inflammation-modulating and proliferation-related genes in fibroblasts, whose inductions are similarly dependent on bioactive IL-1β. Most notably, the 20-fold induction of IL-6 mRNA and protein was completely blocked by IL-1 receptor antagonist. The combined addition of IL-1β and IL-6 was mitogenic, and the proliferative response to conditioned medium from IL-1β-exposed cells was blocked by antagonistically acting Abs to IL-6 or to gp130. These novel findings indicate that soluble IL-1β bioactivity and autocrine IL-1β-dependent IL-6 up-regulation are critical initiators of fibroblast activation and proliferation and that they likely play a role in the fibroproliferative response seen in human acute lung injury.

Pulmonary inflammation and fibrosis are characterized by increased turnover and production of the extracellular matrix as well as an impairment of lung fibrinolytic activity. Although fragments of the extracellular matrix component hyaluronan induce macrophage production of inflammatory mediators, the effect of hyaluronan on the fibrinolytic mediators plasminogen activator inhibitor (PAI)-1 and urokinase-type plasminogen activator (uPA) is unknown. This study demonstrates that hyaluronan fragments augment steady-state mRNA, protein, and inhibitory activity of PAI-1 as well as diminish the baseline levels of uPA mRNA and inhibit uPA activity in an alveolar macrophage cell line. Hyaluronan fragments alter macrophage expression of PAI-1 and uPA at the level of gene transcription. Similarly, hyaluronan fragments augment PAI-1 and diminish uPA mRNA levels in freshly isolated inflammatory alveolar macrophages from bleomycin-treated rats. These data suggest that hyaluronan fragments influence alveolar macrophage expression of PAI-1 and uPA and may be a mechanism for regulating fibrinolytic activity during lung inflammation.

The fibroproliferative response to acute lung injury (ALI) results in severe, persistent respiratory dysfunction. We have reported that IL-1β is elevated in pulmonary edema fluid in those with ALI and mediates an autocrine-acting, fibroblast mitogenic pathway. In this study, we examine the role of IL-1β-mediated induction of cyclooxygenase-2 and PGE2, and evaluate the significance of individual E prostanoid (EP) receptors in mediating the fibroproliferative effects of IL-1β in ALI. Blocking studies on human lung fibroblasts indicate that IL-1β is the major cyclooxygenase-2 mRNA and PGE2-inducing factor in pulmonary edema fluid and accounts for the differential PGE2 induction noted in samples from ALI patients. Surprisingly, we found that PGE2 produced by IL-1β-stimulated fibroblasts enhances fibroblast proliferation. Further studies revealed that the effect of fibroblast proliferation is biphasic, with the promitogenic effect of PGE2 noted at concentrations close to that detected in pulmonary edema fluid from ALI patients. The suppressive effects of PGE2 were mimicked by the EP2-selective receptor agonist, butaprost, by cAMP activation, and were lost in murine lung fibroblasts that lack EP2. Conversely, the promitogenic effects of mid-range concentrations of PGE2 were mimicked by the EP3-selective agent, sulprostone, by cAMP reduction, and lost upon inhibition of Gi-mediated signaling with pertussis toxin. Taken together, these data demonstrate that PGE2 can stimulate or inhibit fibroblast proliferation at clinically relevant concentrations, via preferential signaling through EP3 or EP2 receptors, respectively. Such mechanisms may drive the fibroproliferative response to ALI.

Background:Fibroblasts from patients with idiopathic pulmonary fibrosis (IPF) overexpress the urokinase-type plasminogen activator receptor (uPAR) and are hypermotile. Results: uPAR ligation increases fibroblast motility by localizing ␣5␤1 integrin-Fyn signaling complexes to lipid rafts. Conclusion:The hypermotile phenotype of IPF fibroblasts is due to lipid raft-localized uPAR-integrin-Fyn signaling complexes. Significance: These unique lipid raft signals may be therapeutic targets for IPF.