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慢病毒介导的CXCR4 RNA干扰对结直肠癌肝转移的影响

The influence of lentivirus-mediated CXCR4 RNA interference on hepatic metastasis of colorectal cancer.

作者信息

Wang Tian-Bao, Hu Bao-Guang, Liu Da-Wei, Shi Han-Ping, Dong Wen-Guang

机构信息

Department of Surgery, The First Affiliated Hospital, Sun Yat-sen University, Guangzhou, Guangdong 510080, P.R. China.

Department of Surgery, Prince of Wales Hospital, The Chinese University of Hong Kong, Shatin, Hong Kong, P.R. China.

出版信息

Int J Oncol. 2014 Jun;44(6):1861-9. doi: 10.3892/ijo.2014.2348. Epub 2014 Mar 19.

Abstract

The aim of this study was to construct a lentiviral vector of CXCR4-siRNA (Lenti-CXCR4-siRNA) and investigate whether the vector can inhibit the growth, migration, invasion and hepatic metastasis of colorectal cancer (CRC). RT-PCR and western blotting were employed to identify the ideal RNA interference sequence. Lenti-CXCR4-siRNA was constructed and transfected into the SW480 cell line. We used RT-PCR and western blotting to measure the expression of CXCR4 RNA and protein, respectively; the MTS assay to assess the proliferation of SW480 cells; transwell chambers to estimate the inhibitory effect on migration and invasion; and the Balb/c nude mouse model of CRC to examine the inhibition of hepatic metastasis. The relative expression of the CXCR4 gene and protein was 5.4 and 18.95%, respectively, in the siCXCR4 group. The genes in the expression plasmid pLenti-CXCR4-siRNA were in the correct order. In the SW480, nonsense control (NC) and the Lenti-CXCR4-siRNA groups CXCR4 RNA levels were, respectively, 0.54±0.06, 1.00±0.03 and 0.11±0.04 (P=0.0001); CXCR4 protein levels were 0.60±0.03, 0.72±0.03 and 0.18±0.02 (P=0.0001); the OD value was 1.38±0.04 (P=0.0050), 1.28±0.05 (P=0.0256) and 0.92±0.06; SW480 cell number in migration test was 32±6.85, 32.63±1.69 and 0.75±0.71 (P=0.0000); SW480 cell number in the invasion test was 29.13±10.3, 30.38±6.09 and 0.63±0.74 (P=0.0000); hepatic metastasis number was 7.10±3.98 (P=0.034), 7.50±4.09 (P=0.019) and (3.50±2.51); hepatic metastasis mean weight (in g) was 2.25±2.51 (P=0.000), 2.11±2.38 (P=0.000) and 1.45±2.07. Lenti-CXCR4-siRNA constructs were correctly constructed and effectively inhibit the expression of CXCR4 RNA and protein, reducing the proliferation, migration, invasion capacity of SW480 cells and hepatic metastasis of CRC.

摘要

本研究旨在构建CXCR4-siRNA慢病毒载体(Lenti-CXCR4-siRNA),并研究该载体是否能抑制结直肠癌(CRC)的生长、迁移、侵袭及肝转移。采用RT-PCR和蛋白质印迹法鉴定理想的RNA干扰序列。构建Lenti-CXCR4-siRNA并转染至SW480细胞系。我们分别用RT-PCR和蛋白质印迹法检测CXCR4 RNA和蛋白质的表达;用MTS法评估SW480细胞的增殖;用Transwell小室评估对迁移和侵袭的抑制作用;用CRC的Balb/c裸鼠模型检测对肝转移的抑制作用。在siCXCR4组中,CXCR4基因和蛋白质的相对表达分别为5.4%和18.95%。表达质粒pLenti-CXCR4-siRNA中的基因顺序正确。在SW480、无义对照(NC)和Lenti-CXCR4-siRNA组中,CXCR4 RNA水平分别为0.54±0.06、1.00±0.03和0.11±0.04(P = 0.0001);CXCR4蛋白质水平分别为0.60±0.03、0.72±0.03和0.18±0.02(P = 0.0001);OD值分别为1.38±0.04(P = 0.0050)、1.28±0.05(P = 0.0256)和0.92±0.06;迁移试验中SW480细胞数分别为32±6.85、32.63±1.69和0.75±0.71(P = 0.0000);侵袭试验中SW480细胞数分别为29.13±10.3、30.38±6.09和0.63±0.74(P = 0.0000);肝转移数分别为7.10±3.98(P = 0.034)、7.50±4.09(P = 0.019)和(3.50±2.51);肝转移平均重量(g)分别为2.25±2.51(P = 0.000)、2.11±2.38(P = 0.000)和1.45±2.07。Lenti-CXCR4-siRNA构建体构建正确,能有效抑制CXCR4 RNA和蛋白质的表达,降低SW480细胞的增殖、迁移、侵袭能力及CRC的肝转移。

https://cdn.ncbi.nlm.nih.gov/pmc/blobs/2c3b/4063541/87ad5209bb54/IJO-44-06-1861-g00.jpg

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