RNA干扰RSK4基因表达对裸鼠移植瘤生长与转移影响观察

Effects of RSK4 gene down-regulation by RNA interference on xenograft and metastasis of MCF-7 cell in nude mice

目的:应用慢病毒干扰载体(RSK4-RNAi-LV)干扰RSK4基因在乳腺癌细胞株MCF-7中的表达,研究RSK4基因被干扰后对裸鼠移植瘤生长及转移的影响。方法:将转染了siRNA(RSK4-RNAi-LV)的MCF-7细胞组(实验组)、转染了siRNA(NC-GFP-LV)的MCF-7细胞组(阴性对照组)和未转染的MCF-7细胞组(空白对照组)分别接种至裸鼠乳腺脂肪垫下,建立裸鼠移植瘤模型,观察每组裸鼠移植瘤生长情况;应用实时荧光定量PCR和蛋白质印迹法检测3组移植瘤组织中RSK4mRNA及其蛋白的表达;HE染色观察3组裸鼠内脏转移情况。结果:实验组的移植瘤平均体积为(2 264.08±367.47)mm3,明显大于阴性对照组(843.67±318.13)mm3及空白对照组的(720.45±241.35)mm3差异有统计学意义,F=112.425,P=0.02;实验组瘤体平均体质量为(1.44±0.25)g,明显重于阴性对照组(0.73±0.20)g及空白对照组的(0.70±0.21)g,差异有统计学意义,F=89.54,P=0.01。实验组裸鼠移植瘤肺转移6只,空白对照及阴性对照组各1只。实验组肿瘤组织RSK4mRNA的相对表达量为0.140±0.843,明显低于阴性对照组的1.000±0.000(P=0.008)和空白对照组的0.878±0.689(P=0.005)。实验组、阴性对照组和空白对照组RSK4蛋白的表达量分别为0.138±0.023、0.532±0.032和0.465±0.057,3组间差异有统计学意义,F=73.1,P=0.003;实验组RSK4蛋白表达量明显低于阴性对照组(P=0.006)和空白对照组(P=0.012)。结论:慢病毒干扰MCF-7细胞中RSK4基因表达能促进MCF-7细胞裸鼠移植瘤的生长及转移。

OBJECTIVE：To interfere the expression of RSK4 gene in the breast cancer cell line MCF-7 with lentiviral RNAi vector （RSK4-RNAi-LV）, and investigate the effect of proliferation of transplantation tumor and metastasis. METHODS：The experimental group of nude mice was subcutaneously injected with MCF-7 cells transfected with siRNA （RSK4-RNAi-LV） whlile the negative control o{ nude mice was subcutaneously injected with MCF-7 cells transfected with siRNA （NC-GFP-LV）,the blank contol group of nude mice was subcutaneously injected with MCF-7 cells alone to establish vivo tumor model. Investigate the proliferation of transplantation tumor of the three groups of nude mice. The ex- pression of RSK4 of transplantation tumor were detected by real-time PCR,western blot analysis. The visceral metastasis in nude mice was observed by HE pathology. RESULTS： The proliferation of transplantation tumor of mice in experimental group was significantly faster compared with the negative control group and blank control group. The mice in RSK4- RNAi LV group （2 264.08±367.47） mm2 had substantially greater diameter of tumors compared to that in the NC-GFP- LV group （（720.45±241.35） mm2 ,P=0. 018] and that in the control group [（843.67±318.13） mma ,P=0. 003]. The mean tumor weight was significantly large in the RSK4-RNAi-LV group [（1.44 ± 0.25） g,P=0. 027] compared to that in the negative control [（0.70±0.21） g,P=0. 015] and that the blank control group （0. 73±0. 20） g,P=0.01]. In six athymic nude mice suffer lung metastases in the RSK4-RNAi LV group, the mean tumor weight was significantly large compared with that of NC-GFP LV group and control group. The RSK4 mRNA expression was significantly lower in the RSK4-RNAi-LV group （0. 140 ± 0. 084 3） compared to that in the negative control （ 1. 000 ±0. 000, P = 0. 008） andthat in the blank control group （0. 878 ± 0. 068 9, P = 0. 005）. The RSK4 protein expression of tumor tissue in RSK4- RNAi-LV group,NC-GFP-LV group and blank control group were 0. 138±0. 023,0. 465±0. 057,0. 532±0. 032 ,respec tively. Significant difference was observed in the three groups （F= 73.1, P= 0. 003）. The RSK4 protein expression was significantly lower in the RSK4-RNAi-LV group （0. 138 ±0. 023） compared to that in the negative control （0. 465± 0. 057,P〈0. 006） or the blank control group （0. 532±0. 032,P=0. 012）. CONCLUSION.. Our date indicated that RNAi against RSK4 gene promoted the prolieration of transplantation tumor and metastasis of breast carcinoma efectively.