The Effect Of CDK8-siRNA On The Growth And Apoptosis Of HCT116 Cells

Objiective CDK8(cyclin-dependent kinase 8) is a member of CDK(cyclin-dependent kinase) gene family,which plays an important role in gene transcription.The result show that overexpression of CDK8 could promote the expression ofβ-catenin. Through regulating WNT/β-catenin signaling pathway ,CDK8 plays an important role in the formation and development of tumor. RNA interference (RNAi) could specifically reduce gene expression efficiency, has been widely applied in cancer therapy. The aim of present study was to employ the sequence-sepcific siRNA silencing the CDK8 gene and investigate its effects on human colorectal cancer line HCT116.Methods 1,Structure of CDK8 mRNA was analysed by software to select targeted interference sites. The CDK8-siRNA was transfected into colorectal cancer line HCT116 by positive ion liposome Lipofectamine 2000.Transfect efficiency was assessed by flow cytometry. 2, After human colon cancer cell line HCT116 was transfected by CDK8 small interfering RNA (siRNA),the mRNA and protein of CDK8 andβ-catenin were determined by RT-PCR and Western blot assay respectively. 3, The cell growth inhibition was assesses by MTT. Besides,apoptosis and cell cycle were measured by flow cytometry.Results 1 Negative control FAM-siRNA was successfully transfected into human colorectal cancer cell line HCT116.The transfection efficiency was about 97.2% detected by FCM with cell density at 1.5*105/well and siRNA concentration with 80 pmol in 6 well play. 2, After CDK8-siRNA were successfully transfected into HCT116 cell 48h, detected by RT-PCR,CDK8 andβ-catenin mRNA levels in cells was significantly inhibited. Contrasted with negative control and blank control, the expression of CDK8 andβ-catenin protein were obviously decreased 72h after CDK8-siRNA transfection. 3, CDK8-siRNA could effectively inhibit the growth of HCTl16 cells and accelerate apoptosis in vitro, it could also remain cell cycle in GO/G1 phase.Conclution CDK8-siRNA is successfully constructed and efficiently transfected into HCT116 cells. CDK8 - siRNA can effectively silence the overexpression of CDK8 in colon cancer cell lines HCT116. CDK8-siRNA can affect the transcription of WNT/β-catenin pathway in colon cancer cell lines HCT116.Using CDK8-siRNA transfection could inhibit the growth of HCT116 cells and accelerate apoptosis, it could also change cell cycle.