<i>Background:</i> The –675 4G/5G plasminogen activator inhibitor-1 (PAI-1) polymorphism is linked with asthma and bronchial hyperreactivity. The aim of this study was to evaluate the effect of allergen challenge on plasma PAI-1 concentration in relation to the –675 4G/5G PAI-1 gene polymorphism in house dust mite-allergic asthma patients (HDM-AAs). <i>Materials and Methods:</i> The study was performed in 54 HDM-AAs and 54 healthy nonatopic controls (HCs). Plasma samples were collected in HDM-AAs before, as well as 30 min, 6 h and 24 h after allergen challenge and at corresponding time points in sham-challenged HCs. <i>Results:</i> In subjects carrying the individual PAI-1 genotype, the mean baseline plasma PAI-1 concentration was greater in HDM-AAs than in HCs. At 30 min the mean increase in plasma PAI-1 concentration was significantly greater in HDM-AAs (14.4 ± 12.9 ng/ml) than in HCs (3.4 ± 3.2 ng/ml; p < 0.001). At 6 h, plasma PAI-1 concentration greater than before challenge was found in only 4 HCs (7.4%) but in 48 HDM-AAs (88.9%; p < 0.0001). An increase in plasma PAI-1 concentration at 6 h was found in all HDM-AAs carrying the 4G allele but only in 33.3% of the 5G homozygotes (p < 0.0001). The strongest correlation was found between log PC₂₀ and PAI-1 plasma concentration over the period of 24 h (r = –0.507; p = 0.0001). <i>Conclusion:</i> Changes in plasma PAI-1 concentration associated with allergen-induced bronchoconstriction are modulated by the –675 4G/5G polymorphism of the PAI-1 gene. Allergen-induced upregulation of PAI-1 synthesis may participate in the development of bronchial hyperreactivity in HDM-AAs.