Bile Duct Carcinoma Associated Micrornas The Expression And Function Of The Preliminary Study

Background:Biliary tract cancer is highly fatal cancers of the biliary epithelium, arising within the liver (intrahepatic cholangiocarcinoma; ICC), in the extrahepatic bile ducts (extrahepatic cholangiocarcinoma; ECC). Recent data show that the incidence and mortality rates of biliary tract cancer have been increasing in several areas worldwide. MicroRNAs (miRNAs) are small,20-to24-nucleotide and non-coding RNAs found in diverse organisms and have a broad impact on gene expression through translational repression or post-transcriptional suppression. Recent studies showed that dsyregulation of miRNAs was commonly linked to tumorigenesis and indicated that miRNAs could function as tumor suppressors and oncogenes. Therefore, it is of great importance to focus on the expression and function of miRNAs, associated with tumor formation and progression. Leading us to propose that suppression of miRNAs may be a novel approach for the treatment of Cholangiocarcinoma.Objective:We intend to investigate the expression profile of miRNAs down-regualted in human extrahepatic and intrahepatic cholangiocarcinoma tissues and probe the effect on cell growth of four of these miRNAs; To research the expressionsignature and function of miRNA-21in Cholangiocarcinoma and indentify its functional target mRNA.Methods:Up-regulated、 Down-regulated miRNAs in extrahepatic or intrahepatic cholangiocarcinoma tissues were analyzed by using miRNA-microarray, which was confirmed by using miRNA Real-Time PCR analysis. Based on these findings, differences and representative of these up-regulated、 Down-regulated miNRAs were chosen to perform function investigation. The miRNAs mimics were transfected into QBC939cells and cell proliferation assay was performed by using MTT, which would demonstrate the relationship between down-regulated miRNAs and cholangiocarcinoma cell growth. MiR-21expression in human cholangiocarcinoma tissues and QBC939cell line is measured by using Real-Time PCR and Northern Blot, respectively. Cell growth and apoptosis was analyzed in QBC939after transfected with Anti-miR-21. Specific target analysis is performed by using dual-reporter gene assay, western blot and FACS. In Vitro invasion assay is performed to probe the effect of miR-21on QBC939invasiveness. Results:28miRNAs and21miRNAs were up-regulated in extrahepatic cholangiocarcinoma and intrahepatic cholangiocarcinoma, respectively. There were12miRNAs down-regulated both in two types of cholangiocarcinoma. MiR-125b, miR-19a, miR-21, and miR-378*were inhibited in QBC939cells, which indicated a significant inhibitory effect on cell growth.25miRNAs and15miRNAs were down-regulated in extrahepatic cholangiocarcinoma and intrahepatic cholangiocarcinoma, respectively. There were6miRNAs down-regulated both in two types of cholangiocarcinoma. miR-181a, miR-596, miR-492, and miR-602were overexpressed in QBC939cells, which indicated a significant inhibitory effect on cell growth. Expression analysis reveals that miR-21levels depicted a significant up-regulation as compared to the matched normal bile duct. Silencing of miR-21in QBC939by using anti-miR-21decreases cell growth and induces cell apoptosis. PTEN、 PDCD4、 RECK is identified as a direct effector of miR-21and miR-21promoted QBC939cell invasion by inhibiting the expression level of RECK.Conclusions:The miRNAs expression patterns in human extrahepatic and intrahepatic cholangiocarcinoma tissues are different and up-regulated miRNAs act as oncomirs on cholangiocarcinoma cell growth. The miRNAs expression patterns in human extrahepatic and intrahepatic cholangiocarcinoma tissues are different and Down-regulated miRNAs act as suppressors on cholangiocarcinoma cell growth. Targeting of miR-21is sufficient to limit cholangiocarcinoma progress. Therefore, leading us to propose that suppression of miRNAs may be a novel approach for the treatment of Cholangiocarcinoma.