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目的研究黑色瘤分化相关基因-7(MDA-7)/白细胞介素-24(IL-24)对食管癌细胞Eca-109与TE-1的增殖抑制作用和杀伤作用。方法应用RT-PCR技术检测MDA-7/IL-24受体复合物在Eca-109与TE-1细胞的表达,构建MDA-7/IL-24基因的重组腺病毒载体,将携带人MDA-7/IL-24基因的腺病毒Ad-mda-7分别感染食管癌细胞Eca-109与TE-1,通过Western-blot检测MDA-7基因的表达,通过CCK-8检测该基因对食管癌细胞的增殖抑制作用,碘化丙啶(PI)染色后流式细胞仪检测MDA-7对食管癌细胞周期的影响,Hoechst/PI双染色后流式细胞仪观察MDA-7对食管癌细胞的诱导凋亡作用。结果 Eca-109与TE-1均有表达MDA-7/IL-24受体复合物IL-22R1/IL-20R2;成功构建包装重组腺病毒,并经Western-blot验证其介导了外源基因MDA-7/IL-24在Eca-109与TE-1细胞中的高效表达;CCK-8实验结果表明,MDA-7/IL-24能显著抑制Eca-109与TE-1细胞的增殖,且随着感染时间的延长及感染剂量的增加而增强抑制作用;细胞周期结果提示,MDA-7/IL-24使大量Eca-109与TE-1细胞被阻滞在G1~G0期;Hoechst/PI双染色结果提示,MDA-7/IL-24能明显促进Eca-109与TE-1细胞的凋亡。结论重组腺病毒能介导MDA-7/IL-24基因通过G1期阻滞和诱导凋亡,从而抑制食管癌细胞的增殖。
Objective To investigate the inhibitory and antitumor effects of melanoma differentiation-related gene-7 (MDA-7)/interleukin-24 (IL-24) on esophageal cancer cells Eca-109 and TE-1. Methods RT-PCR was used to detect the expression of MDA-7/IL-24 receptor complex in Eca-109 and TE-1 cells. The recombinant adenoviral vector containing MDA-7/IL-24 gene was constructed and will carry human MDA- The 7/IL-24 gene adenovirus Ad-mda-7 infects esophageal cancer cells Eca-109 and TE-1 respectively, detects the expression of MDA-7 gene by Western-blot, and detects the gene against esophageal cancer cells by CCK-8. The inhibitory effect of proliferation was detected by propidium iodide (PI) staining. The effect of MDA-7 on the cell cycle of esophageal carcinoma was detected by flow cytometry. Flow cytometry was used to observe the induction of MDA-7 on esophageal carcinoma cells by Hoechst/PI double staining. Apoptosis. Results Both Eca-109 and TE-1 expressed HLA-7/IL-24 receptor complex IL-22R1/IL-20R2. The recombinant adenovirus was successfully constructed and verified by Western-blot. High expression of MDA-7/IL-24 in Eca-109 and TE-1 cells; CCK-8 results showed that MDA-7/IL-24 can significantly inhibit the proliferation of Eca-109 and TE-1 cells, and With the prolongation of infection time and the increase of infection dose, the inhibitory effect was enhanced; cell cycle results suggested that MDA-7/IL-24 caused a large number of Eca-109 and TE-1 cells to be arrested in the G1~G0 phase; Hoechst/PI Double staining results suggested that MDA-7/IL-24 can significantly promote the apoptosis of Eca-109 and TE-1 cells. Conclusion Recombinant adenovirus can mediate MDA-7/IL-24 gene block and induce apoptosis through G1 phase, thus inhibiting the proliferation of esophageal cancer cells.