Role of p53 up-regulated modulator of apoptosis in gemcitabine-induced apoptosis of human pancreatic cancer cell lines

Objective:To investigate the effect of gemcitabine on the proliferation of human pancreatic cancer AsPC-1 cells in vitro and elucidate the mechanism.Methods:The vector of antisense oligoneucleotides of p53 up-regulated modulator of apoptosis (PUMA) pcDNA3.1-PUMMAS and empty vector pcDNA3.1 was transfected into AsPC-1 cells by lipofectAMINE 2000 mediation,respectively. The stable clone was selected by G418. pcDNA3.1-PUMAAS-transfected,pcDNA3.1-transfected,and control AsPC-1 cells were treated with serial concentrations(1,5,10,and 15 μmol/L)of gemcitabine for 72 h.The expressions of PUMA mRNA and protein were detected by RT-PCR and Western blotting.MTT assay was used to measure the inhibitory actions of gemcitabine on AsPC-1 cells.The apoptosis was detected by flow cytometry,Hoechst 33258 staining,and TUNEL staining.Results:Gemcitabine induced apoptosis of AsPC-1 cells and inhibited the proliferation in a dose-dependent manner. The apoptosis induced by gemcitabine was accompanied with the up-regulation of PUMA. In pcDNA3.1-PUMAAS-transfected cells,the up-regulation of PUMA induced by gemcitabine was significantly decreased compared with AsPC-1 control cells and pcDNA3.1 transfected cells.The apoptosis was inhibited and the proliferation was increased after transfection of pcDNA3.1-PUMAAS.Conclusion:Gemcitabine induced the apoptosis of AsPC-1 cells and inhibited the proliferation in vitro. The apoptosis-inducing effect was related to up-regulation of PUMA expression.