SNAI2在结直肠癌中的表达及其对细胞增殖和侵袭能力的影响

Expression of SNAI2 in colorectal cancer and its effect on cell proliferation and invasion

目的明确锌指转录因子2(SNAI2)在结直肠癌组织中的表达及对结直肠癌细胞增殖和侵袭能力的影响。方法收集海南省人民医院胃肠外科2015年9月至2016年4月间收治的50例临床结直肠癌组织样本及癌旁组织,使用反转录-聚合酶链反应(RT-PCR)技术检测其中SNAI2的表达水平,t检验比较两组SNAI2水平的差异。针对SNAI2基因,设计四组干扰RNA序列和一组无任何靶向位点的随机插入序列。将各组序列剪切到慢病毒载体上,用二代包装系统包装成慢病毒悬液。将HT29接种至6孔板,分别标记为对照组、sh SNAI2 1#组、sh SNAI2 2#组、sh SNAI2 3#组、sh SNAI2 4#组。对照组注入插入随机序列载体的慢病毒悬液,sh SNAI2 1#组、sh SNAI2 2#组、sh SNAI2 3#组、sh SNAI2 4#组分别注入插入四组干扰RNA序列的慢病毒悬液。挑选SNAI2表达最高的SW480细胞系进行慢病毒感染,获得稳定knock-down SNAI2细胞系。在高表达SNAI2的SW480细胞系中,采用慢病毒感染的方法构建稳定干扰SNAI2细胞株。通过噻唑蓝(MTS)比色法检测细胞增殖能力,划痕实验检测细胞侵袭迁移能力。同时Western blot检测E-钙黏蛋白(E-cadherin)、波形蛋白(Vimentin)的表达。结果癌旁组织SNAI2的表达量低于结直肠癌组织,差异有统计学意义(P<0.05);与对照组比较,细胞黏附分子E-cadherin表达量升高,细胞骨架蛋白、Vimentin表达量降低,差异均有统计学意义(P<0.05);si SNAI2的SW480细胞系中,与对照组比较,细胞增殖能力、侵袭能力显著降低,差异有统计学意义(P<0.05)。结论 SNAI2在人结直肠癌中表达量升高,作为一种癌基因,其可促进结直肠癌细胞的体外增殖、侵袭和体内转移能力。

Objective To define the expression of SNAI2 in colorectal cancer and its effect on the proliferationand invasion of colorectal cancer cells. Methods The expression level of SNAI2 was detected by reverse transcriptionpolymerase chain reaction（RT-PCR） in 50 cases of colorectal cancer tissues and adjacent tissues from Department of Gas-trointestinal Surgery in our hospital during Sep. 2015 and Apr. 2016, and the difference between the two groups was detect-ed by t test. For the SNAI2 gene, four groups of interfering RNA sequences and a group of random insertion sequences with-out any targeting sites were designed. The sequence of each group was sheared onto lentiviral vector and packaged into len-tivirus suspension by two generation packaging system. HT29 was inoculated into 6 orifice plates and labeled as controlgroup, sh SNAI2 1#group, sh SNAI2 2#group, sh SNAI2 3#group, and sh SNAI2 4#group. The control group was injected withlentivirus suspension inserted into the random sequence vector. Sh SNAI2 1#group, sh SNAI2 2#group, sh SNAI2 3#groupand sh SNAI2 4#group were injected with four groups of lentivirus suspension which interfered with RNA sequence. SNAI2 cell line with the highest expression of SW480 was selected for lentivirus infection, and stable knock-down SNAI2 cell linewas obtained. In SW480 cells with the high expression of SNAI2, the SNAI2 interference of stable cell lines was estab-lished through slow infection. The proliferation ability of cell was detected by thiazole blue（MTS） colorimetry method. Thecell invasion ability was detected by scarification test. Western blot was used to detect the expression of E-cadherin and Vi-mentin. Results The expression of SNAI2 in normal adjacent tissues was less than that in colorectal cancer tissues com-pared with normal adjacent tissues, and the difference was statistically significant（P〈0.05）. The expression of cell adhesionmolecule E-cadherin increased compared with the control group, while the expression of cytoskeletal protein and Vimentindecreased, both with statistically significant difference（P〈0.05）. In the SW480 si SNAI2 cell line, compared with the control group, the cell invasion and proliferation ability decreased significantly（P〈0.05）. Conclusion SNAI2 is highly ex-pressed in human colorectal cancer tissue, and it could promote the proliferation, invasion and migration of colorectal can-cer cells in vivo.