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目的:探讨联合应用凋亡素(apoptin)基因、新城疫病毒(newcastle disease virus,NDV)血凝素-神经氨酸酶(he-magglutinin-neuramidinase,HN)基因及人白介素18(hIL-18)基因体外对人结肠癌细胞HCT-116的抑制效应。方法:重组质粒pIRApoptinHNIL18通过脂质体介导法体外转染人结肠癌细胞HCT-116,通过Western blotting和RT-PCR检测外源基因表达;采用AO/EB染色法检测pIRApoptinHNIL18对人结肠癌细胞HCT-116抑制作用;利用流式细胞术分析pIRApoptinHNIL18对人结肠癌细胞HCT-116线粒体膜电位(mitochondrial transmembrane potential,△Ψ)和活性氧(reactive oxygen species,ROS)水平;采用3,5-二羟基甲苯法测定唾液酸含量。结果:pIRApoptinHNIL18转染人结肠癌细胞HCT-116后,外源基因能够有效表达;肿瘤细胞生长受到抑制;线粒体△Ψ由(94.41±8.17)%下降到(30.70±8.01)%(P<0.05);唾液酸含量由(0.33±0.06)mmol/L下降到(0.09±0.03)mmol/L(P<0.01);ROS水平由(52.48±6.09)%升高到(68.98±7.26)%(P<0.05)。结论:pIRApoptinHNIL18可通过线粒体途径诱导细胞凋亡,从而抑制人结肠癌细胞HCT-116的生长。
Objective: To investigate the combination of apoptin gene, hecastle-neuramidinase (HN) gene and human interleukin 18 (hIL-18) in newcastle disease virus (NDV) Inhibitory Effect of Genes on Human Colon Cancer Cells HCT-116 in Vitro. Methods: The recombinant plasmid pIRApoptinHNIL18 was transfected into human colon cancer cell line HCT-116 by liposome-mediated method. The expression of foreign gene was detected by Western blotting and RT-PCR. The expression of pIRApoptinHNIL18 in human colon cancer cell HCT -116. The mitochondrial transmembrane potential (△ Ψ) and reactive oxygen species (ROS) levels of HCT-116 human colon cancer cells were analyzed by flow cytometry. The cytotoxicity of pIRApoptinHNIL18 against 3,5-dihydroxy Toluene determination of sialic acid content. Results: The transfected human colon cancer cell line HCT-116 could express the foreign gene efficiently and the growth of tumor cells was inhibited. The △ Ψ of mitochondria decreased from (94.41 ± 8.17)% to (30.70 ± 8.01)% (P <0.05) ; Sialic acid content decreased from (0.33 ± 0.06) mmol / L to (0.09 ± 0.03) mmol / L (P <0.01); ROS level increased from (52.48 ± 6.09)% to (68.98 ± 7.26)% 0.05). Conclusion: pIRApoptinHNIL18 can induce apoptosis in the mitochondria and inhibit the growth of human colon cancer cell line HCT-116.