Plumbagin from Plumbago Zeylanica L Induces Apoptosis in Human Non-small Cell Lung Cancer Cell Lines through NFkB Inactivation

Abstract

<br /><strong>Objective</strong>: To detect effects of plumbagin on proliferation and apoptosis in non-small cell lung cancer celllines, and investigate the underlying mechanisms. Materials and <br /><strong>Methods</strong>: Human non-small cell lung cancercell lines A549, H292 and H460 were treated with various concentrations of plumbagin. Cell proliferationrates was determined using both cell counting kit-8 (CCK-8) and clonogenic assays. Apoptosis was detected byannexin V/propidium iodide double-labeled flow cytometry and TUNEL assay. The levels of reactive oxygenspecies (ROS) were detected by flow cytometry. Activity of NF-kB was examined by electrophoretic mobilityshift assay (EMSA) and luciferase reporter assay. Western blotting was used to assess the expression of bothNF-kB regulated apoptotic-related gene and activation of p65 and IkBk. <br /><strong>Results</strong>: Plumbagin dose-dependentlyinhibited proliferation of the lung cancer cells. The IC50 values of plumbagin in A549, H292, and H460 cells were10.3 μmol/L, 7.3 μmol/L, and 6.1 μmol/L for 12 hours, respectively. The compound concentration-dependentlyinduced apoptosis of the three cell lines. Treatment with plumbagin increased the intracellular level of ROS, andinhibited the activation of NK-kB. In addition to inhibition of NF-kB/p65 nuclear translocation, the compoundalso suppressed the degradation of IkBk. ROS scavenger NAC highly reversed the effect of plumbagin onapoptosis and inactivation of NK-kB in H460 cell line. Treatment with plumbagin also increased the activity ofcaspase-9 and caspase-3, downregulated the expression of Bcl-2, upregulated the expression of Bax, Bak, andCytC. <br /><strong>Conclusions</strong>: Plumbagin inhibits cell growth and induces apoptosis in human lung cancer cells throughan NF-kB-regulated mitochondrial-mediated pathway, involving activation of ROS.