小干扰RNA抑制ΔNp63α表达对人食管鳞癌细胞生物学特性的影响

Effect of small interfering RNA targetingΔNp63αgene on the biological characteristics of human esophageal squamous carcinoma cells

目的通过腺相关病毒介导的小干扰RNA(siRNA)抑制人食管鳞癌细胞Eca109中ΔNp63α的表达,观察其对食管鳞癌细胞生物学特性的影响。方法构建H1启动子驱动的表达针对ΔNp63α的siRNA重组腺相关病毒AAV-ΔNp63αshRNA并感染Eca109细胞。同时以感染空腺相关病毒AAV-Null的细胞及不加腺相关病毒的细胞分别作为对照组和空白对照组,观察siRNA干扰ΔNp63α表达在体外及体内对Eca109细胞生长、增殖、致瘤及凋亡的影响。结果与对照组和空白对照组比较,Eca109细胞感染AAV-ΔNp63αshRNA后ΔNp63α蛋白表达下降(P均<0.05),细胞生长速度减慢(P均<0.05),细胞增殖指数(proliferation index,PI)降低(P均<0.01),裸鼠瘤体质量减轻(P均<0.05),细胞凋亡增加(P均<0.05)。结论腺相关病毒介导的siRNA干扰能显著抑制Eca109细胞ΔNp63α的表达,减慢肿瘤细胞的增殖,促进肿瘤细胞凋亡,抑制肿瘤生长。

Objective To construct a small interfering RNA （siRNA） vector targeting ANp63a and investigate △Np63a gene interference on the proliferation and apoptosis of human esophageal squamous carcinoma Ecal09 cell line. Methods Adeno-associated virus （AAV）-△Np63ahRNA driven by H1 promoter was constructed and was used to infect Ecal09 cells. AAV Null and normal ceil lines were utilized in the control group and blank control group, respectively. The influence of siRNA interference of△Np63a expression on the growth, proliferation, tumorigenic efficiency and apoptosis of Ecal09 cells were analyzed in vitro and in vivo. Results Compared with the two control groups, the specific siRNA targeting ANp63a gene significantly down-regulated the expression of △Np63a protein levels in Eeal09 cells （all P〈0.05）. The growth of Ecal09 cells infected with AV-△Np63ahRNA was significantly lower than those in the two control groups （all P^0.05）. Cell cycle analysis showed the proliferation index （PI） of AAV-△Np63ahRNA infected cell line was significantly lower compared with the two control groups （all P〈0.01）. In vivo experiment exhibited that AAV-△Np63ashRmn infected cells resulted in a lower tumor weight in nude mice compared with the cells in the two control groups （all P〈0.05）. In addition, the apoptosis index （AI） of AAV-ANp63ahRNa infected cells were significantly higher than those of the other cell lines （P〈0.05）. Conclusion AAV- mediated expression of siRNA can significantly reduce △Np63a expression in Ecal09 cells, slowing down the proliferation, promoting the apoptosis, and subsequently inhibiting the growth of tumor.