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目的应用能同时表达3种不同基因(血管内皮生长因子VEGF、人端粒酶逆转录酶hTERT、抗凋亡基因bcl-x1)的短发夹RNA(short hairpin RNA,shRNA)的重组腺病毒对胃癌细胞系(BGC-823)进行RNA干扰,观察重组腺病毒对胃癌细胞增殖活性及细胞周期的影响。方法构建能同时表达3种不同的的重组腺病毒Ad-shVEGF-shTERT-sh bcl-xl,同时构建单独表达上述3个基因的重组腺病毒,将上述4种重组腺病毒同时感染BGC-823,MTT法检测BGC-823细胞增殖活性,流式细胞仪检测细胞凋亡情况,蛋白印迹检测目的蛋白的表达。结果重组腺病毒感染胃癌细胞后,3个目的基因的蛋白表达显著下调,细胞增殖明显受抑制,大量细胞凋亡。表达3个基因的重组腺病毒生长增殖抑制作用强于表达单个基因的重组腺病毒。结论相对于单独沉默1个基因,同时沉默3种不同基因,对胃癌细胞的生长抑制作用更强。同时沉默多个基因的表达是胃癌基因治疗研究的一个新的有效途径。
OBJECTIVE: To construct a recombinant adenovirus vector expressing short hairpin RNA (shRNA) capable of simultaneously expressing three different genes (vascular endothelial growth factor VEGF, human telomerase reverse transcriptase hTERT, anti-apoptotic gene bcl-x1) Gastric cancer cell line (BGC-823) RNA interference, observe the effect of recombinant adenovirus on the proliferation of gastric cancer cells and cell cycle. METHODS: Three recombinant adenoviruses, Ad-shVEGF-shTERT-sh bcl-xl, were constructed and constructed. The recombinant adenovirus expressing these three genes was constructed simultaneously. The four recombinant adenoviruses were simultaneously infected with BGC-823, The proliferation of BGC-823 cells was detected by MTT assay. The apoptosis of BGC-823 cells was detected by flow cytometry. The expression of target protein was detected by Western blot. Results After the recombinant adenovirus was infected into gastric cancer cells, the protein expression of the three genes was significantly down-regulated, the cell proliferation was significantly inhibited, and a large number of apoptosis was observed. The recombinant adenovirus expressing the three genes showed stronger growth inhibition than the recombinant adenovirus expressing the single gene. Conclusion Compared with silencing a single gene alone and silencing three different genes simultaneously, the growth inhibition of gastric cancer cells is stronger. Simultaneous silencing of the expression of multiple genes is a new and effective approach for gene therapy of gastric cancer.