Effects of simulated bleeding in an in vitro nasal fibroblast wound healing model

Abstract

Background: We investigated the effect of simulated bleeding on plasminogen activity, matrix metalloproteinase (MMP) expression, and wound healing using a human fibroblast model. Methods: Nasal fibroblasts from three chronic rhinosinusitis (CRS) patients with nasal polyps and three controls were grown in culture and a standardized injury was created using a punch. To mimic bleeding, fibroblasts were stimulated with plasminogen (100 μg/mL), plasminogen + tranexamic acid (TA; 100 μg/mL) or media only. At 24, 48, and 72 hours after injury, we measured urokinase plasminogen activator (uPA) and tissue-type plasminogen activator (tPA) activities and inactive and active MMP-2 and -9 expression. Results: Injury stimulated the nasal fibroblasts to express uPA and tPA and active and inactive MMP-2 and -9. In CRS patients, plasminogen significantly decreased MMP-9 expression after 48 hours (p < 0.04). In untreated fibroblasts, we observed a decrease in active MMP-9 expression, whereas plasminogen increased active MMP-9 expression after 48 hours (p < 0.04). At 24 hours, active MMP-9 expression was reduced by plasminogen ± TA (p < 0.02). Plasminogen also stimulated uPA expression in CRS patient fibroblasts after 48 hours (p < 0.04). Fibroblast proliferation occurred when exposed to plasminogen and was strongly modulated by uPA and inactive and active MMP-2. The quality of wound healing was affected by inactive MMP-2, uPA and tPA, simulation, and inhibition of bleeding. Conclusion: Activation of the plasminogen pathway and inactive MMP-2 expression tended to increase both proliferation of nasal fibroblasts and MMP-9 expression as a marker for deterioration of the quality of wound healing.