lithium-chloride and Carcinoma--Non-Small-Cell-Lung

Disregulation of dickkopf-related protein 1 (DKK1) has been reported in a variety of human cancers. However, how DKK1 functions in Non-small cell lung cancer has not been revealed. In the current study, DKK1 was knocked out by the lentivirus-mediated short hairpin RNA interference approach in H1299 and 95C non-small cell lung cancer cell lines. Subsequently, the migration and invasion ability were assessed by wound-healing and transwell assays. In addition, epithelial-mesenchymal transition markers and β-catenin were examined by Western blot analysis. The signaling pathway downstream of DKK1 was characterized using the Wnt signaling pathway inhibitor, IWP2, and glycogen synthase kinase 3 beta inhibitor, LiCl. Immunofluorescence analysis investigated the subcellular localization of β-catenin. The results suggested that knockdown of DKK1 caused reduced migration and invasion ability of H1299 and 95C cells. DKK1 silencing resulted in the downregulation of epithelial-mesenchymal transition-related proteins, such as Snail and zinc finger E-box binding homeobox 1. Besides, DKK1 silencing inhibited β-catenin and promoted the phosphorylation of β-catenin. Mechanism results indicated that the expression of β-catenin was reduced in H1299 or 95C cells after being treated with Wnt signaling inhibitor, IWP2. In addition, the inhibition of β-catenin phosphorylation by glycogen synthase kinase 3 beta inhibitor, LiCl, significantly enhanced the migration and invasion capacities in DKK1-knockdown cell lines. Furthermore, cell immunofluorescence revealed that nuclear β-catenin was reduced when DKK1 was knocked down. Taken together, these findings suggest that DKK1 induces the occurrence of epithelial-mesenchymal transition and promotes migration and invasion in non-small cell lung cancer cells. Mechanically, β-catenin plays a vital role in DKK1-induced non-small cell lung cancer cell migration and invasion, and DKK1 inhibits the phosphorylation of β-catenin, resulting in the increased nuclear localization of β-catenin.

Topics: Benzothiazoles; beta Catenin; Carcinoma, Non-Small-Cell Lung; Cell Line, Tumor; Cell Movement; Epithelial-Mesenchymal Transition; Gene Expression Regulation, Neoplastic; Gene Knockdown Techniques; Glycogen Synthase Kinase 3 beta; Humans; Intercellular Signaling Peptides and Proteins; Lentivirus; Lithium Chloride; Neoplasm Invasiveness; Phosphorylation; RNA, Small Interfering; Wnt Signaling Pathway

Non-small cell lung cancer (NSCLC) A549 cells are resistant to tumor necrosis factor-related apoptosis-inducing ligand (TRAIL)-induced apoptosis. Therefore, combination therapy using sensitizing agents to overcome TRAIL resistance may provide new strategies for treatment of NSCLC. Here, we investigated whether lithium chloride (LiCl), a drug for mental illness, could sensitize A549 cells to TRAIL-induced apoptosis. We observed that LiCl significantly enhanced A549 cells apoptosis through up-regulation of death receptors DR4 and DR5 and activation of caspase cascades. In addition, G2/M arrest induced by LiCl also contributed to TRAIL-induced apoptosis. Concomitantly, LiCl strongly inhibited the activity of c-Jun N-terminal kinases (JNKs), and the inhibition of JNKs by SP600125 also induced G2/M arrest and augmented cell death caused by TRAIL or TRAIL plus LiCl. However, glycogen synthase kinase-3β (GSK3β) inhibition was not involved in TRAIL sensitization induced by LiCl. Collectively, these findings indicated that LiCl sensitized A549 cells to TRAIL-induced apoptosis through caspases-dependent apoptotic pathway via death receptors signaling and G2/M arrest induced by inhibition of JNK activation, but independent of GSK3β.

Topics: Apoptosis; Carcinoma, Non-Small-Cell Lung; Caspases; Cell Line, Tumor; Gene Expression Regulation, Neoplastic; Humans; JNK Mitogen-Activated Protein Kinases; Lithium Chloride; Lung Neoplasms; MAP Kinase Kinase 4; TNF-Related Apoptosis-Inducing Ligand; Tumor Necrosis Factor-alpha; Up-Regulation

Aberrant activations of Wnt and Notch signaling pathways are individually reported to be associated with the pathogenesis of non-small-cell lung cancer (NSCLC). However, the data about the cross talk between the two signaling pathways are still limited. To elucidate potential Wnt/Notch cross talk within NSCLC, we examined the impact of Notch3 activity on LiCl-induced cell cycle changes.. The lung cancer cell lines were treated with LiCl, a Wnt activator, in the absence or presence of Notch3-siRNA. Cell cycles and the expression of the regulators of cell cycle, c-MYC, p21 and Skp2 (S phase kinase-associated protein 2) were measured after treatment.. The treatment with LiCl increased the percent of cells at S phase and G phase and the expression of c-MYC and Skp2 and decreased the expression of p21. Moreover, the expression of Notch3 and its down-stream genes, HES-1 and HEYL, was up-regulated by LiCl. Notch3-siRNA weakened the effect of LiCl on the cell cycle and resulted in attenuation of the LiCl-induced increment of c-MYC and Skp2 and the LiCl-induced decrement of p21.. These data suggest that Notch3 activation cooperatively takes part in the LiCl-induced cell cycle changes, at least partially, associated with c-MYC, Skp2 and p21.

Topics: Carcinoma, Non-Small-Cell Lung; Cell Cycle; Cell Line, Tumor; Cyclin-Dependent Kinase Inhibitor p21; Glycogen Synthase Kinase 3; Glycogen Synthase Kinase 3 beta; Humans; Lithium Chloride; Proto-Oncogene Proteins c-myc; Receptor, Notch3; Receptors, Notch; RNA, Small Interfering; S-Phase Kinase-Associated Proteins; Signal Transduction; Wnt Proteins