Inhibiting the mu opioid receptor (MOR), Src, Gab-1, PI3 kinase or STAT3 attenuates opioid and growth factor-induced epithelial mesenchymal transition (EMT) in human lung cancer cells.

<p><b>Panel A</b>: Graphical representation of the % control vimentin expression. H358 human NSCLC cells were either untreated, treated with 100 ng/ml epidermal growth factor (EGF), 100 nM DAMGO, morphine or fentanyl or 100 ng/ml insulin growth factor (IGF) for 96 hours with or without pretreatment of cells with the peripheral MOR antagonist, methylnaltrexone (MNTX, 100 nM), MOR siRNA, Gab-1 siRNA, Src siRNA, the PI3 kinase inhibitor LY294002 (10 uM), Akt Inhibitor X (5 uM), the Src family kinase inhibitor PP2 (100 nM) or the STAT3 inhibitor Stattic (10 uM). Cell lysates were then obtained and immunoblotted with the EMT marker anti-vimentin antibody. Experiments were repeated in triplicate and immunoreactive bands were analyzed using computer-assisted densitometry. There is a statistically significant difference (p<0.05 indicated by asterisks (*)) between control and treatment groups with error bars = standard deviation. An increase in vimentin expression is suggestive of an epithelial mesenchymal transition. <b>Panel B</b>: Graphical representation of the % control claudin-1 expression. H358 human NSCLC cells were either untreated, treated with 100 ng/ml epidermal growth factor (EGF), 100 nM DAMGO, morphine or fentanyl or 100 ng/ml insulin growth factor (IGF) for 96 hours with or without pretreatment of cells with the peripheral MOR antagonist, methylnaltrexone (MNTX, 100 nM), MOR siRNA, Gab-1 siRNA, Src siRNA, the PI3 kinase inhibitor LY294002 (10 uM), Akt Inhibitor X (5 uM), the Src family kinase inhibitor PP2 (100 nM) or the STAT3 inhibitor Stattic (10 uM). Cell lysates were then obtained and immunoblotted with the EMT marker anti-claudin-1 antibody. Three independent experiments per condition were performed and immunoreactive bands were analyzed using computer-assisted densitometry. There is a statistically significant difference (p<0.05 indicated by asterisks (*)) between control and treatment groups with error bars = standard deviation. A decrease in claudin-1 expression is suggestive of an epithelial mesenchymal transition.