Chang-zheng Liu 1 , Wei Liu 2 , Yi Zheng 2 , Jin-mei Su 3 , Jing-jing Li 2 , Lan Yu 2 , Xiao-dong He 2 , and Song-sen Chen 1 1 Department of Biochemistry, National Laboratory of Medical Molecular Biology, Institute of Basic Medical Sciences, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing 100005, China 2 Department of General Surgery, 3 Department of Rheumatology, Peking Union Medical College Hospital, Chinese Academy of Medical Sciences & Peking Union Medical College, Beijing, 100730, China
Objective To investigate the expression profile of microRNA-21 in human cholangiocarcinoma tissues and to validate its bona fide targets in human cholangiocarcinoma cells. Methods The expression profile of microRNA-21 in human cholangiocarcinoma tissues and cholangiocarcinoma cell line, QBC939, was evaluated by using real-time PCR analysis. The bona fide targets of microRNA-21 were analyzed and confirmed by dual luciferase reporter gene assay and western blot, respectively. The expressional correlation of microRNA-21 and its targets was probed in human cholangiocarcinoma tissues by using real-time PCR, locked nucleic acid in situ hybridization (LNA-ISH), and immunohistochemistry analysis. Results Real-time PCR analysis revealed that microRNA-21 expression depicted a significant up-regulation in human cholangiocarcinoma tissues about 5.6-fold as compared to the matched normal bileduct tissues (P<0.05). The dual luciferase reporter gene assay revealed endogenous microRNA-21 in cholangiocarcinoma cell line, QBC939, inhibited the luciferase reporter activities of wild-type PTEN (P<0.01) and PDCD4 (P<0.05) and had no this effect on mutated PTEN and PDCD4. Moreover, loss of microRNA-21 function led to a significant increase of PTEN and PDCD4 protein levels in QBC939 cells. Elevated microRNA-21 levels were accompanied by marked reductions of PTEN and PDCD4 expression in the same cholangiocarcinoma tissue. Conclusion microRNA-21 expression is up-regulated in human cholangiocarcinoma and PTEN, PDCD4 are direct effectors of microRNA-21.
cholangiocarcinoma; microRNA-21; phosphatase and tensin homolog; programmed cell death 4
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