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目的观察通过RNA干扰抑制裸鼠移植瘤细胞中Vav1基因表达对肿瘤生长的影响,并探讨其机制。方法培养人结肠癌细胞株HT29,通过皮下注射制备裸鼠移植瘤模型。造模成功后将裸鼠随机分为实验组和对照组,分别给予Vav1-siRNA或生理盐水进行瘤内注射每3日一次进行干预,测量肿瘤体积;干预5次后处死动物取出肿瘤,比较肿瘤体积及重量;MTT法检测细胞活性;荧光定量PCR(QPCR)和免疫组化法(IHC)检测肿瘤组织中周期蛋白D1(Cyclin D1)、基质金属蛋白酶-2(MMP-2)、基质金属蛋白酶组织抑制剂-1(TIMP-1)的表达。结果与对照组比较,实验组裸鼠肿瘤的体积和重量都低于对照组(t=-2.429、-4.407,P<0.05)。与对照组比较,实验组肿瘤细胞活性明显降低(t=-10.706,P<0.05)。QPCR和IHC结果均显示,实验组Cyclin D1、MMP-2的mRNA和蛋白表达低于对照组,而TIMP-1表达高于对照组(P<0.05),差异均有统计学意义。结论抑制Vav1基因表达后裸鼠移植瘤的生长明显受到抑制,这可能与Vav1基因能调节Cyclin D1、MMP-2、TIMP-1表达有关。
Objective To observe the effect of Vav1 gene expression on the growth of tumor cells in nude mice by RNA interference and to explore its mechanism. Methods Human colon cancer cell line HT29 was cultured and transplanted into nude mice by subcutaneous injection. After successful modeling, the nude mice were randomly divided into experimental group and control group. The mice were injected with Vav1-siRNA or saline intramuscularly once every three days to measure the tumor volume. After 5 times intervention, the animals were sacrificed and the tumors were removed. Volume and weight of cells were detected by MTT assay. Cell viability was detected by MTT assay. Cyclin D1, MMP-2 and MMP-2 were detected by quantitative polymerase chain reaction (QPCR) and immunohistochemistry (IHC) Tissue inhibitor-1 (TIMP-1) expression. Results Compared with the control group, the tumor volume and weight of the experimental group were lower than those of the control group (t = -2.429, -4.407, P <0.05). Compared with the control group, the activity of tumor cells in experimental group was significantly lower (t = -10.706, P <0.05). The results of QPCR and IHC showed that the mRNA and protein expressions of Cyclin D1 and MMP-2 in the experimental group were lower than those in the control group, while the expression of TIMP-1 in the experimental group was higher than that in the control group (P <0.05). Conclusion The growth inhibition of Vav1 gene in nude mice was significantly inhibited, which may be related to the regulation of the expression of Cyclin D1, MMP-2 and TIMP-1 by Vav1 gene.