Aim: To investigate the inhibitory effects of plumbagin (5-hydroxy-2 methyl-1,4-naphthoquinone) on the invasion and migration and its correlation with matrix metalloproteinase-2 (MMP-2) and urokinase-plasminogen activator (u-PA) in liver cancer HepG2 cells under non-cytotoxic concentrations.
Methods: The cell viability was determined by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. The adhesion, migration and invasion were measured by cell-matrix adhesion assay and Boyden chamber assay. The MMP-2 and u-PA activities were estimated by gelatin and casein-plasminogen zymography. The mRNA and protein levels of MMP-2, u-PA, urokinase-plasminogen activator receptor (u-PAR), tissue inhibitor of metalloproteinase-2 (TIMP-2), plasminogen activator inhibitor-1 (PAI-1), nuclear factor κ B (NF-κB), c-Fos and c-Jun were evaluated by semi-quantitative reverse transcription polymerase chain reaction and western blotting. Also, the binding abilities of NF-κB and activator protein-1 (AP-1) were analyzed by electrophoretic mobility shift assay (EMSA).
Results: In this study, plumbagin had exhibited an inhibitory effect on the abilities of adhesion, migration and invasion. The results from zymography showed plumbagin treatment may decrease the activities of MMP-2 and u-PA. Further, the mRNA and protein levels of MMP-2, u-PA and u-PAR were significantly reduced, while TIMP-2 and PAI-1 were elevated by plumbagin treatment. Next, plumbagin significantly decreased the nuclear levels of NF-κB, c-Fos and c-Jun. Also, treating HepG2 cells with plumbagin leads to dose-dependent inhibition on the binding abilities of NF-κB and AP-1.
Conclusion: We demonstrated the inhibitory effects of plumbagin on the invasion, migration and adhesion of HepG2 cells, while plumbagin treatment may decrease the expressions of MMP-2 and u-PA and enhance the expressions of TIMP-2 and PAI-1.
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