Ascl2 knockdown in colon cancer LS174T cells led to expression change of EMT-associated microRNA / 重庆医学

ABSTRACT

Objective To investigate the effects of transcription factor achaete scute-like 2(Ascl2)on epithelial-mesenchymal transition (EM T ) associated microRNAs .Methods Colon cancer LS174T cells were transfected with shRNA-Ascl2 vector and shRNA-control vector respectively ,then the transfected cells were selected with G 418 and stably transfected cell lines were estab-lished .Real-time PCR and Western-blot analysis were used to determine the interference effect .Transwell invasion experiment were used to observe the effects of Ascl2 RNA interference on cell invasion capability in vitro .MicroRNA chip analysis was used to de-tect the change of EMT-associated microRNA expression ,and real-time PCR experiment was used to validate the microarray re-sults .Results The mRNA and protein expressions were significantly reduced after Ascl2 interference (P<0 .01) .The numbers of invading cells were significantly decreased after Ascl2 interference (P<0 .01) .MicroRNA chip analysis found microRNA-200 fami-ly (including microRNA-200b ,microRNA-200a ,microRNA-429 ,microRNA-200c ,microRNA-141) was more than 2-fold upregula-tion after Ascl2 interference (P< 0 .01) .Conclusion Ascl2 regulates the invasion and metastasis of colon cancer cell ,possibly through transcription regulation of microRNA-200 family ,and then regulation of EM T .