微小RNA-335通过靶定人类表皮生长因子3调控乳腺癌细胞的侵袭

MicroRNA-335 inhibits breast cancer cell invasion by targeting human epidermal growth factor receptor 3

目的 观察微小RNA（miR）-335在乳腺癌细胞侵袭中的作用.方法 分别在Basallike类型正永用生化细胞MCF10A和Luminal A乳腺癌细胞MCF7中抑制和过表达miR-335的含量,采用Transwell侵袭实验检测细胞侵袭能力的改变;利用生物信息学网站预测miR-335靶基因,并用荧光报告基因检测实验验证;应用miR-335抑制剂及靶基因siRNA单转染和共转染,验证miR-335是否通过靶基因执行功能.最后利用实时荧光定量聚合酶链反应检测在35例原发性乳腺癌组织中miR-335的表达水平.结果 在MCF10A内敲减miR-335,细胞发生侵袭的数目由对照组51个增加到实验组的67个,促进细胞侵袭的能力（P＜0.05）.在MCF7中过表达miR-335后,由对照组侵袭数目51个降至实验组43个,抑制细胞侵袭的能力（P＜0.05）.miR-335能够靶向结合人类表皮生长因子3（ERBB3）的mRNA的3＇-非翻译区（3＇-UTR）,抑制ERBB3 mRNA及蛋白水平表达（P＜0.05）.敲减ERBB3后能够恢复miR-335低表达造成的细胞侵袭能力的降低（P＜0.05）.miR-335在原发性乳腺癌组织中表达为正常组织的0.75,呈显著低表达（P＜0.01）.结论 在原发性乳腺癌组织样本中,miR-335低表达,并且通过靶向ERBB3影响细胞的侵袭能力.

Objective In this study,we aim to detect microRNA （miR）-335 function in cell invasion.Methods In vitro,we respectively inhibited or over-expressed miR-335 in MCF10A and MCF7 to detect the change of cell invasion by transwell invasion assay.The bioinformatics was used to predict the target of miR-335 and later it was investigated by luciferase reporter assay.Through the single-tranfection or co-transfection of miR-335 inhibitor or human epidermal growth factor receptor 3 （ERBB3） siRNA,we detected whether miR-335 affects invasion by targeting ERBB3.At last,through realtime quantative polymerase chain reaction,we detected the expression level of miR-335 in primary breast cancer specimen.Results Inhibition of miR-335 in MCF10A reduced the ability of cell invasion.Invasion cell numbers enhanced from 51 of control group to 67 of exprimental group.The P value is lower than 0.05.Over-expression of miR-335 enhanced the ability of cell invasion.Invasion cell numbers decreased from 51 of control group to 43 of exprimental group （P 〈 0.05）.miR-335 could target the 3＇ untranslated regions of ERBB3 mRNA and inhibited its mRNA and protein level（P 〈0.05）.Inhibition of ERBB3 could resuce the change of cell invasion that miR-335 inhibitors contributed to （P 〈 0.05）.miR-335 showed lower expression level in primary breast cancer specimen compared to adjacent tissues （P 〈 0.01）.Conclusion miR-335 showed lower expression in primary breast cancer and affected breast cancer cell invasion by targeting ERBB3.