Rationale Genistein is a dietary isoflavone and a broad-spectrum tyrosine kinase inhibitor contained in soy products. An epidemiologic study of asthma revealed that subjects with high consumption of dietary soy isoflavones had better lung function than those with lower intake. In a guinea pig model of allergic asthma, genistein reduced methacholine-induced bronchoconstriction. To explore the mechanisms underlying these observations, we tested genistein's ability to block lung myofibroblast differentiation, a key phenotypic change in asthmatic airway remodeling.
Methods Human fetal lung fibroblasts (IMR-90) were grown to subconfluence in DMEM containing 10% FBS, serum-deprived for 24 hours, and treated with genistein (10 μM) for an additional 24 hurs. Cells were then stimulated with TGF-b1 (2 ng/mL) for 24 hours, and a-smooth muscle actin (aSMA) expression, a marker of the myofibroblast phenotype, was assessed by both immunoblot and immunofluorescence microscopy. To determine the intracellular mechanism of inhibitory actions by genistein, we also assessed phosphorylation of Smad2 by immunoblot in the presence and absence of genistein after 30-minute stimulation with TGF-b1.
Results Treatment with genistein resulted in 54.9 ± 17.1% reduction in TGF-b1 induced expression of aSMA. Immunofluorescence microscopy revealed a decrease in aSMA staining intensity and stress fiber formation in genistein-treated cells. In addition, Smad2 phosphorylation was inhibited by 71.1 ± 16.7% in genistein-treated cells.
Conclusions These results demonstrate that genistein interferes with TGF-b1-stimulated myofibroblast differentiation and suggest that tyrosine kinase inhibition may have a role in modulation of asthmatic airway remodeling.
Funded by NIH RO1HL072891, T32HL076139, The CHEST Foundation-GSK Clinical Research Trainee Award.
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