NAC inhibits NSCLC cell proliferation through reduction of PDK1 protein expression. A-B, A549 NSCLC cells exposed to increased concentrations of NAC for up to 48 h (A), or NSCLC cell lines indicated were treated with NAC (5 mM) for up to 48 h (B). Afterwards, cell proliferation was determined by Luminescent Cell Viability Assay. C, NSCLC cell lines indicated were treated with NAC (5 mM) for up to 48 h. Afterwards, cell proliferation was determined by MTT assays. Data are means ± SD from 3 separate experiments. * p < 0.01, compared with untreated cells (CTR). D-E, Cellular protein was isolated from A549 cells that were cultured with increased concentrations of NAC as indicated for 24 h (D) or cultured with NAC (5 mM) for the indicated time period (E) followed by Western blot analysis with antibodies against PDK1 protein. The bar graphs represent the mean ± SD of PDK1/GAPDH of at least three independent experiments. *indicates significant difference from untreated control (0). F-G, Several NSCLC cells as indicated were treated with NAC (5 mM) for 24 h followed by Western blot for detecting PDK1 protein. (F) or A549 cells were transfected with control or overexpression of PDK1 vectors for 24 h, followed by exposure of the cells to NAC for an additional 24 h. Afterwards, the luminescence of viable cells was detected using Cell Titer-Glo Luminescent Cell Viability Assay Kit. The upper panels represent protein levels of PDK1 by Western blot (G). All data were depicted as mean ± SD. *indicates significant difference as compared to the untreated control cells (CTR).