摘要
目的探讨HOXC-AS3通过微小RNA(miR)-1269对大肠癌细胞转移特性的影响及其机制。方法采用定量荧光聚合酶链式反应(QF-PCR)分析人大肠癌细胞株HT-29、LoVo和SW620中HOXC-AS3表达水平;采用脂质体将NC-短发卡RNA(shRNA)和HOXC-AS3 shRNA转染至SW620细胞后,分为NC-shRNA组和HOXC-AS3 shRNA组,采用划痕实验、Transwell实验和蛋白质印迹法(Western blot)检测细胞迁移、侵袭和上皮-间充质转化能力;采用生物信息学和双荧光素酶报告基因分析HOXC-AS3的靶基因;并采用QF-PCR分析细胞中HOXC-AS3的靶基因miR-1269的表达水平。组间计量数据比较采用t检验。结果大肠癌细胞株SW620细胞HOXC-AS3表达水平(1.67±0.11)明显高于大肠癌细胞株HT-29和LoVo细胞(1.20±0.13;1.08±0.12),差异有统计学意义(t=6.703、8.478,P<0.05)。NC-shRNA组细胞HOXC-AS3表达水平(0.98±0.08)明显高于HOXC-AS3 shRNA组水平(0.28±0.06),差异有统计学意义(t=17.560,P<0.05)。NC-shRNA组细胞划痕愈合率[(84.75±4.36)%]明显高于HOXC-AS3 shRNA组[(40.28±7.67)%],差异有统计学意义(t=12.340,P<0.05)。NC-shRNA组细胞迁移数量[(129.50±13.07)个]明显高于HOXC-AS3 shRNA组[(70.83±9.32)个],差异有统计学意义(t=8.956,P<0.05)。NC-shRNA组上皮细胞标志物E-钙黏蛋白(E-cadherin)表达水平(0.69±0.10)明显低于HOXC-AS3 shRNA组(1.27±0.07),差异有统计学意义(t=11.580,P<0.05)。NC-shRNA组细胞波形蛋白(Vimentin)和N-钙黏蛋白(N-cadherin)表达水平(1.30±0.10、1.35±0.07)明显高于HOXC-AS3 shRNA组(0.72±0.09、0.99±0.13),差异有统计学意义(t=10.440、5.822,P<0.05)。miR-1269是HOXC-AS3的靶基因。NC-shRNA组细胞miR-1269表达水平(1.07±0.14)明显低于HOXC-AS3 shRNA组(1.79±0.13),差异有统计学意义(t=9.267,P<0.05)。结论敲降HOXC-AS3可显著抑制大肠癌细胞迁移、侵袭和上皮-间充质转化能力,其机制可能与靶向调控miR-1269的表达有关。
Objective To investigate the effect of HOXC-AS3 on the metastatic characteristics of colorectal cancer cells through microRNA(miR)-1269 and the underlying mechanism.Methods The expression level of HOXC-AS3 in colorectal cancer cell lines HT-29,LoVo and SW620 was analyzed by fluorescence quantitative polymerase chain reaction(PCR).NC short hairpin RNA(shRNA)and HOXC-AS3 shRNA were transfected into SW620 cells with liposomes,and then divided into NC shRNA group and HOXC-AS3 shRNA group.The ability of cell migration,invasion and epithelial mesenchymal transformation was analyzed by wound healing assay,Transwell test and Western blotting respectively.The target gene of HOXC-AS3 was analyzed by bioinformatics and double luciferase reporter gene.The expression level of the target gene miR-1269 of HOXC-AS3 in cells was analyzed by fluorescence quantitative PCR.The comparison of measurement data between groups was conducted using t-test.Results The expression level of HOXC-AS3 in colorectal cancer cell line SW620(1.67±0.11)was significantly higher than that in colorectal cancer cell lines HT-29 and LoVo(1.20±0.13;1.08±0.12,t=6.703,8.478,P<0.05).The expression level of HOXC-AS3 in NC shRNA group(0.98±0.08)was significantly higher than that in HOXC-AS3 shRNA group(0.28±0.06,t=17.560,P<0.05).The wound healing rate in NC shRNA group[(84.75±4.36)%]was significantly higher than that in HOXC-AS3 shRNA group[(40.28±7.67)%,t=12.340,P<0.05].The number of migrating cells in NC shRNA group[(129.50±13.07)cells]was significantly greater than that in HOXC-AS3 shRNA group[(70.83±9.32)cells,t=8.956,P<0.05].The expression level of E-cadherin in the NC shRNA group(0.69±0.10)was significantly lower than that in the HOXC-AS3 shRNA group(1.27±0.07,t=11.580,P<0.05).The expression levels of Vimentin and N-cadherin in NC shRNA group(1.30±0.10,1.35±0.07)were significantly higher than those in HOXC-AS3 shRNA group(0.72±0.09,0.99±0.13,t=10.440,5.822,P<0.05).The expression level of miR-1269 in NC shRNA group(1.07±0.14)was significantly lower than that in HOXC-AS3 shRNA group(1.79±0.13,t=9.267,P<0.05).Conclusion Knockdown of HOXC-AS3 can significantly inhibit the migration,invasion and epithelial mesenchymal transformation of colorectal cancer cells,which may be related to the targeted regulation of miR-1269 expression.
作者
毛争强
杜波涛
孙航
宰守峰
Mao Zhengqiang;Du Botao;Sun Hang;Zai shoufeng(Department of Surgical Oncology,the Fourth Clinical College of Xinxiang Medical University,Xinxiang Central Hospital,Xinxiang 453000,China)
出处
《中华实验外科杂志》
CAS
北大核心
2023年第9期1765-1768,共4页
Chinese Journal of Experimental Surgery
