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Role of microRNA‐155 at early stages of hepatocarcinogenesis induced by choline‐deficient and amino acid–defined diet in C57BL/6 mice

MicroRNAs (miRs) are conserved, small (20‐25 nucleotide) noncoding RNAs that negatively regulate expression of messenger RNAs (mRNAs) at the posttranscriptional level. Aberrant expression of certain microRNAs plays a causal role in tumorigenesis. Here, we report identification of hepatic microRNAs that are dysregulated at early stages of feeding C57BL/6 mice choline‐deficient and amino acid–defined (CDAA) diet that is known to promote nonalcoholic steatohepatitis (NASH)‐induced hepatocarcinogenesis after 84 weeks. Microarray analysis identified 30 hepatic microRNAs that are significantly (P ≤ 0.01) altered in mice fed CDAA diet for 6, 18, 32, and 65 weeks compared with those fed choline‐sufficient and amino acid–defined (CSAA) diet. Real‐time reverse transcription polymerase chain reaction (RT‐PCR) analysis demonstrated up‐regulation of oncogenic miR‐155, miR‐221/222, and miR‐21 and down‐regulation of the most abundant liver‐specific miR‐122 at early stages of hepatocarcinogenesis. Western blot analysis showed reduced expression of hepatic phosphatase and tensin homolog (PTEN) and CCAAT/enhancer binding protein beta (C/EBPβ), respective targets of miR‐21 and miR‐155, in these mice at early stages. DNA binding activity of nuclear factor kappa B (NF‐κB) that transactivates miR‐155 gene was significantly (P = 0.002) elevated in the liver nuclear extract of mice fed CDAA diet. Furthermore, the expression of miR‐155, as measured by in situ hybridization and real‐time RT‐PCR, correlated with diet‐induced histopathological changes in the liver. Ectopic expression of miR‐155 promoted growth of hepatocellular carcinoma (HCC) cells, whereas its depletion inhibited cell growth. Notably, miR‐155 was significantly (P = 0.0004) up‐regulated in primary human HCCs with a concomitant decrease (P = 0.02) in C/EBPβ level compared with matching liver tissues. Conclusion: Temporal changes in microRNA profile occur at early stages of CDAA diet‐induced hepatocarcinogenesis. Reciprocal regulation of specific oncomirs and their tumor suppressor targets implicate their role in NASH‐induced hepatocarcinogenesis and suggest their use in the diagnosis, prognosis, and therapy of liver cancer. (HEPATOLOGY 2009.)