Insulin-like Growth Factor-I signaling mechanisms, type I collagen and alpha smooth muscle actin in human fetal lung fibroblasts.

UNLABELLED

Bronchial wall remodeling is a major morbidity component in oxidant injury in bronchopulmonary dysplasia (BPD) and asthma.

HYPOTHESIS

IGF-1 enhances alpha smooth muscle expression and collagen synthesis in developing lung fibroblasts leading to fibrosis through nuclear NF-(k)B -dependent transcription. We studied NF-(k)B dependent transcription by transfecting HFLF with a NF-(k)B responsive promoter driving the luciferase gene and treating with IGF-1 (100 ng/mL) and measuring luciferase activity. We exposed cells to the PI-3 kinase inhibitor or the Erk1/2 inhibitor one hr before stimulating with IGF-1. We also used IGF-1 receptor antibody to inhibit the action of IGF-1 and studied its effect on alpha-sma and type I collagen. IGF-1 treatment significantly increased luciferase activity. This was attenuated by PI-3 kinase and MAP-Kinase inhibitors. Western blot analysis showed PI-3 kinase mediates IGF-1 activation of NF-(k)B independent of I(K)B phosphorylation. We found an up-regulation of phospho NF-kB in the nuclear extract compared with total NFKB showing that IGF-1 regulates NF-(k)B transcriptional activity downstream of NF-(k)B nuclear translocation. IGF-1-induced increase in alpha-sma expression and type-I collagen was significantly inhibited by pretreatment with LY294002 and IGF-1 receptor antibody. IGF-1 cell signaling leading to collagen synthesis in fetal lung fibroblasts is mediated by PI3 Kinase acting through NF-(k)B in HFLF.