【摘要】 目的探讨RNAi沉默Rac1基因对结肠癌细胞增殖及裸鼠成瘤的影响。方法 1细胞实验:体外培养结肠癌细胞SW480,随机分为空白对照组、阴性对照组、shRNA-Rac1组,阴性对照组、shRNA-Rac1组分别转染空载质粒慢病毒、shRNA-Rac1慢病毒。采用RT-PCR技术、Western blotting法检测各组Rac1 mRNA及其蛋白表达;采用MTT法检测各组转染24、48、72、96 h的细胞增殖情况。2动物实验:取shRNA-Rac1组、阴性对照组转染后细胞分别接种于裸鼠皮下,建立结肠癌裸鼠移植瘤模型(分别记为观察组、对照组),观察裸鼠成瘤及移植瘤生长情况,喂养35天脱臼处死,完整剥离肿瘤组织,称取瘤体质量并测量肿瘤体积。同时检测瘤体Rac1蛋白表达情况。结果 1细胞实验:与阴性对照组比较,shRNA-Rac1组Rac1 mRNA及其蛋白表达明显降低(P均<0.05);与阴性对照组比较,shRNA-Rac1组各时间点细胞增殖速度明显降低(P均<0.05)。2动物实验:对照组癌细胞接种4~8天均可见瘤体形成,观察组接种35天仅2只成瘤,且瘤体形成时间较对照组晚10天。接种35天,观察组、对照组瘤体体积分别为(32.54±43.13)、(948.13±523.50)mm3,瘤体质量分别为(0.023±0.031)、(0.873±0.372)g,两组比较P均<0.01。与对照组比较,观察组瘤体组织Rac1蛋白阳性表达率明显降低(P<0.05)。结论抑制Rac1表达能降低结肠癌细胞增殖速度,抑制裸鼠移植瘤生长。更多还原

【Abstract】 Objective To investigate the effects of Rac1 gene silenced by RNA interference( RNAi) on cell proliferation and growth of subcutaneous tumor of colon cancer SW480 cells in nude mice. Methods 1 Cell experiment: colon cancer SW480 cells cultured in vitro were randomly divided into the blank control group,negative control group,and shRNARac1 group. Recombinant lentivirus virus vector( NC-shRNA and Rac1-shRNA) were constructed and were respectively transfected into negative control group and shRNA-Rac1 group. The expression levels of Rac1 mRNA and protein were measured by quantitative real-time polymerase chain reaction( qRT-PCR),and Western blotting,respectively. Cell proliferation at 24,48,72,and 96 h after transfection was determined by MTT. 2 Animal experiment: cells from shRNA-Rac1 group and negative control group were planted into nude mice to establish mice models of colon cancer( observation group and control group). Formulation and growth of the tumor was observed. After the mice were sacrificed by cervical dislocation,tumor tissues were removed completely and we measured its weight and size. Meanwhile,Rac1 expression was also detected by immunohistocytochemistry. Results 1 cell experiment: Compared with the blank control group and negative control group,the expression of Rac1 mRNA and protein in the shRNA-Rac1 group decreased significantly( all P < 0. 05). The proliferation ability of colon cancer cells was significantly inhibited in the shRNA-Rac1 group at all detecting time points as compared with that of the control group( all P < 0. 05). 2 Animal experiment: tumor was observed in all cases of the control group 4-8 days after the transplantation. While in the observation group,tumor was only observed in 2 cases 35 days after the transplantation,and it took 10 days longer to form a tumor than that in the control group. On the 35 th day of transplantation,average volume of the tumor in the observation group and control group was( 948. 13 ± 523. 50) and( 32. 54 ± 43. 13) mm3,and the tumor average weight was respectively( 0. 873 ± 0. 372) and( 0. 023 ± 0. 031) g,and significant difference was found between the two groups( all P < 0. 01). Besides,compared with the control group,the expression rate of Rac1 in the observation group was significantly lower than that in the control group( P < 0. 01). Conclusion Silencing the expression of Rac1 decreases the proliferation of colon cancer cells SW480 and inhibits the tumor formation in nude mice.更多还原