TNF- has been shown to induce matrix metalloproteinase-9 expression, which in turn degrades extracellular matrix in the inflammatory responses. However, the inductive mechanisms of the MMP-9 by TNF- remain unclear. In human tracheal smooth muscle cells, TNF- induced MMP-9 expression and Akt phosphorylation in a time-dependent manner, which was attenuated by inhibitors including Src (PP1), EGFR (AG1478), PDGFR (AG1296), and PI3K (LY294002), respectively, revealed by reporter gene assay, RT-PCR, zymographic and Western blotting analyses. Transfection with dominant negative mutants of c-Src (KM), p85 and Akt (KA) also reduced MMP-9 expression. These findings indicated that MMP-9 expression was regulated by PI3K/Akt via the transactivation of growth factor receptors. Furthermore, LY294002 or wortmannin inhibited Akt phosphorylation, but had no effect on NF-B translocation which was blocked by helenalin. Mutated NF-B DNA binding element in the MMP-9 promoter and helenalin also attenuated MMP-9 expression, suggesting that PI3K/Akt and NF-B regulated MMP-9 expression. To support this notion, immunofluorescence staining and immunoprecipitation were applied to characterize the transcription factors involved in these responses. The results showed that LY294002 and curcumin blocked Akt translocation into nucleus. In contrast, the p300, acetyl-histone (H3), and NF-B p65 were found to be co-immunoprecipitated with the phosphorylated Akt, indicating that these components associated with the mmp-9 promoter revealed by chromatin immunoprecipitation assays. Thus, our study provides a new insight into the molecular mechanisms that phosphorylation of Akt by growth factor receptors transactivation may eventually stimulate p300 activity and assemble transcription factors (p65) and lead to MMP-9 expression induce by TNF-.