靶向5-LOX的siRNA诱导胰腺癌细胞的凋亡

Small interfering RNA targeted on 5-lipoxygenase induces apoptosis of pancreatic cancer cells

目的:采用RNA干扰技术(siRNA)阻断5-LOX基因的表达,观察其抑制人胰腺癌细胞增殖及诱导细胞凋亡的作用.方法:构建靶向5-LOX的siRNA质粒表达载体,采用Lipofectamine-2000转染人胰腺癌细胞株SW1990,采用RT-PCR检测RNA干扰后5-LOX mRNA表达,MTT法检测细胞的增殖抑制率,流式细胞仪检测细胞凋亡率.结果:靶向5-L O X序列特异性的三条s i R N A(组1、组2、组3)可以有效地抑制SW1990细胞5-LOX基因表达,其表达抑制率分别为19.6%±1.9%、55.4%±2.6%和55.2%±2.7%.转染靶向5-LOX siRNA的三个质粒表达载体可以显著抑制SW1990细胞的增殖,细胞接种24h后,三组增殖抑制率分别为5.37%±1.19%、11.63%±1.25%和13.67%±1.04%;48h后其增殖抑制率分别为16.13%±1.5%、26.63%±1.22%和25.47%±1.67%,各siRNA组增殖抑制率高于空白组和阴性对照组(均P<0.05),转染后24h和48h三组细胞凋亡率分别为5.56%±1.05%、11.45%±1.44%、12.13%±1.36%和7.37%±1.23%、18.75%±1.5%和22.02%±1.45%,均高于空白对照组和阴性对照组(P<0.05).结论:所构建的靶向5-LOX的siRNA质粒表达载体可以有效阻断SW1990细胞5-LOX基因表达,显著地抑制SW1990细胞增殖,并在一定程度上诱导其凋亡.

AIM： To investigate the effects of siRNA （small interfering RNA） on cell proliferation, apoptosis in human pancreatic cancer cell line SW1990 through knockout of 5-1ipoxygenase （5-LOX）. METHODS： The siRNA expression vectors targeted at 5-LOX gene were designed and DNA template was synthesized, then siRNA was obtained by in vitro transcription. After siRNA was transfected into cell line SW1990 with Lipofectamine-2000, the expression level of 5-LOX mRNA was detected using RT-PCR, SW1990 cell proliferation inhibition was evaluated by MTT assay and apoptosis rate was measured by flow cytometry. RESULTS： The sequence-specific siRNA effectively suppressed 5-LOX mRNA expression in cell line SW1990. The expression inhibition rates （19.6% ± 1.9%, 55.4% ± 2.6% and 55.2% ± 2.7%） of 5-LOX mRNA in group 1-3 were all higher than those in negative control groups （2.9%）. At 24 h and 48 h after cell seeding, the proliferation inhibition rates （5.37% ± 1.19%, 11.63% ± 1.25% and 13.67% ± 1.04%; 16.13% ± 1.5%, 26.63% ± 1.22% and 25.47% ± 1.67%） of cell line SW1990 in group 1-3 were significantly higher than those in blank control group and negative control group, respectively （P 〈 0.05）. Apoptosis inhibition rates （5.56% ± 1.05%, 11.45% ±1.44% and 12.13%± 1.36%; 7.37% ± 1.23%, 18.75% ± 1.5% and 22.02% ± 1.45%） of cell line SW1990 in group 1-3 were also higher than those in blank control group and negative control group （P 〈 0.05）. CONCLUSION： siRNA plasmid expression vectors targeted at 5-LOX in vitro effectively block 5-LOX gene expression, significantly inhibit cell proliferation, and induce cell apoptosis to some extent of cell line SW1990.