Figure 5.

NNK Treatment, but Not K-ras Mutation, Suppressed miR-124 Expression in NSCLC

(A) Overexpression of mutant K-ras does not affect the expression of miR-124 in H292 and H1650 cells. Seventy-two hours after of transfection, the indicated cells were subjected to qRT-PCR analysis. (B) Long time treatment of NNK significantly decreased miR-124 expression in NSCLC cells. The indicated cells were treated with or without 100 nM NNK for indicated times, and subsequently the cells were subjected to qRT-PCR analysis. (C) NNK treatment dose dependently inhibited miR-124 expression in NSCLC cells. Indicated cells were treated with indicated concentration of NNK for 7 days then subjected to qRT-PCR analysis. (D) miR-124 expression was measured by qRT-PCR in specimens of NSCLC patients with never smoking (n = 28) or smoking more than 5 years (n = 28). (E) Silencing of DNMT1 abolished NNK-induced inhibition of miR-124 expression in H358 cells. H358 cells were transfected with siRNA of DNMT1. After 12 hr of transfection, cells were treated with 100 nM NNK for 6 days. Then, cells were subjected to western blot and qRT-PCR analysis. (F) A schematic model of regulation of miR-124 expression and miR-124 effect on NSCLC progression. *p < 0.05; **p < 0.01; ***p < 0.001.