卵巢上皮性癌组织中Jagged1 mRNA的表达及沉默Jagged1基因的表达对裸鼠移植瘤生长的影响

Expression of Jagged1 mRNA in human epithelial ovarian carcinoma tissues and effect of RNA interference of Jagged1 on growth of xenograft in nude mice

目的：探讨Notch基因的配体——Jagged1 mRNA在卵巢上皮性癌（卵巢癌）组织中的表达及沉默Jagged1基因的表达对卵巢癌裸鼠移植瘤生长的影响。方法（1）收集2011年2月至2014年3月在河南省新乡市中心医院行手术治疗的48例卵巢癌及30例良性卵巢上皮性肿瘤患者的存档组织标本，采用实时荧光定量逆转录（RT）-PCR技术检测肿瘤组织中Jagged1、Notch受体——Notch1及其下游靶基因Hes1、Hey1 mRNA的表达。（2）培养卵巢浆液性囊腺癌细胞系SKOV3细胞接种于裸鼠腋窝皮下，构建卵巢癌裸鼠移植瘤模型，随机将裸鼠分为3组，Jagged1干扰组、空载体组和对照组，每组10只裸鼠，分别转染重组载体pcDNA3.1（＋）-siRNA-Jagged1质粒（特异性沉默Jagged1基因的表达）、空载体pDC3.1质粒和磷酸盐缓冲液。转染结束后14 d，测量3组裸鼠移植瘤体积和瘤体质量，计算抑瘤率；采用实时荧光定量RT-PCR技术和蛋白印迹（western blot）法分别检测裸鼠移植瘤组织中Jagged1、Notch1、Hes1和Hey1 mRNA和蛋白的表达。结果（1）实时荧光定量RT-PCR技术检测显示，卵巢癌组织中Jagged1、Notch1、Hes1和Hey1 mRNA的表达水平均高于良性卵巢上皮性肿瘤组织，分别比较，差异均有统计学意义（P〈0.01）。（2）转染结束后14 d，Jagged1干扰组裸鼠的移植瘤体积和瘤体质量分别为（491±68）mm3和（2.6±0.4）g，均明显小于空载体组[分别为（842±88）mm3和（4.4±0.8）g]和对照组[分别为（851±90）mm3和（4.5±0.9）g；P〈0.05]；Jagged1干扰组裸鼠的抑瘤率为42.2%，显著高于空载体组和对照组（分别为2.2%、0；P〈0.05）；Jagged1干扰组裸鼠移植瘤组织中Jagged1、Hes1和Hey1 mRNA和蛋白的表达水平均明显低于空载体组和对照组（P〈0.05），而3组间Notch1 mRNA和蛋白的表达水平无明显差异（P〉0.05）。结论 Jagged1 mRNA在卵巢癌组织中呈高表达；特异性沉默Jagged1基因的表达后，可抑制裸鼠移植瘤的生长，这可能是通过抑制Notch1信号通路而发挥抑癌作用。

Objective To investigate the expression of Jagged1 in human epithelial ovarian carcinoma tissues and the effect of Jagged1 on growth of xenograft in nude mice. Methods （1） Forty-eight cases of ovarian cancer and 30 cases of patients with benign epithelial ovarian tumor in the Henan Province Xinxiang Central Hospital during Feb. 2011 to Mar. 2014 were enrolled in this study. The mRNA expression of Jagged1, Notch1 and the downstream target genes Hes1, Hey1 were analyzed by using realtime PCR method. （2） The ovarian cancer xenograft models in nude mice were constructed by injecting SKOV3 cells in axillary subcutaneouswere. The nude mice were randomly divided into Jagged1 interference group, blank plasmid group and control group. Each group had 10 mice. They were transfected with pcDNA3.1（＋）-siRNA-Jagged1, blank plasmid pDC3.1 and phosphate buffer, respectively. The tumor volumes and tumor masses were measured 14 days after transfection and the inhibition rate was calculated. The relative mRNA expression of Jagged1, Notch1, Hes1 and Hey1 in xenograft tissues after transfection in each group was detected by using realtime PCR technique and the relative protein expression of Jagged1, Notch1, Hes1 and Hey1 in xenograft tissues was detected by utilizing western blot method. Results （1） The relative mRNA expression of Jagged1, Notch1, Hes1 and Hey1 in ovarian cancer tissues were higher than benign ovarian tumor tissues, the differences were statistically significant （P〈0.01）. （2） The tumor volume was （491 ± 68） mm3 and tumor mass was （2.6±0.4） g in Jagged1 interference group, which were significantly lower than that in the blank plasmid group [（842 ± 88） mm3 and （4.4 ± 0.8） g, respectively] and that in the control group [（851 ± 90） mm3 and （4.5 ± 0.9） g, respectively;P〈0.05], the tumor inhibition rate was 42.2% in Jagged1 interference group, which was significantly higher than that in the blank plasmid group and that in the control group （2.2%and 0, respectively）, the differences were statistically significant （P〈0.05）. The relative mRNA and protein expression of Jagged1, Hes1 and Hey1 in xenograft tissues of nude micein Jagged1 interference group were lower than that in the other two groups, the differences were statistically significant （P〈0.05）. There were no differences of relative mRNA and protein expression of Notch1 in xenograft tissues of nude mice among the three groups （P〉0.05）. Conclusions Jagged1 is highly expressed in epithelial ovarian carcinoma. Jagged1 gene interference in xenograft tumor can inhibit ovarian cancer cell growth and improve tumor suppressor rate, which probably play roles by inhibiting Notch1 signaling pathway.