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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.
OBJECTIVE: To use RNA interference (siRNA) to block the expression of 5-LOX gene and to observe its effect on inhibiting proliferation and inducing apoptosis of human pancreatic cancer cells.Methods: siRNA plasmid targeting 5-LOX was constructed and transfected by Lipofectamine -2000 were transfected into human pancreatic cancer cell line SW1990.The expression of 5-LOX mRNA was detected by RT-PCR, the proliferation inhibition rate was detected by MTT assay and the apoptosis rate was detected by flow cytometry.Results: LOX sequence-specific three si RNAs (group 1, group 2 and group 3) could effectively inhibit the expression of 5-LOX gene in SW1990 cells with the inhibitory rates of 19.6% ± 1.9%, 55.4% ± 2.6% and 55.2% ± 2.7% .The transfection of three plasmids targeting 5-LOX siRNA plasmid could significantly inhibit the proliferation of SW1990 cells.The inhibitory rates of proliferation of SW1990 cells were 5.37% ± 1.19%, 11.63% ± 1.25% and 13.67% ± 1.04% respectively. After 48h, the proliferation inhibition rates were 16.13% ± 1.5%, 26.63% ± 1.22% and 25.47% ± 1.67%, respectively. The proliferation inhibition rate of each siRNA group was higher than that of blank group and negative control group 0.05). The apoptotic rates of the three groups at 24 h and 48 h after transfection were 5.56% ± 1.05%, 11.45% ± 1.44%, 12.13% ± 1.36% and 7.37% ± 1.23%, 18.7 5% ± 1.5% and 22.02% ± 1.45%, respectively, which were higher than that of the blank control group and the negative control group (P <0.05) .Conclusion: The constructed siRNA targeting 5-LOX plasmid expression vector can effectively block SW1990 cells 5 -LOX gene expression, significantly inhibited SW1990 cell proliferation, and to some extent induced its apoptosis.