%A Yang,Dong %A Tang,Sai %A Yang,Yan %A Yang,Fan %A Jiang,Wengang %A Liu,Yakun %A Zhang,Fengyun %A Fang,Haoshu %A Wang,Siying %A Zhang,Yuxia %D 2019 %J Frontiers in Oncology %C %F %G English %K MicroRNA-100,CRE/LoxP,Genotype,HCC,SHP-2 %Q %R 10.3389/fonc.2019.00535 %W %L %M %P %7 %8 2019-June-26 %9 Original Research %# %! miR-100 hepatocyte-specific knock-out mice. %* %< %T Generation and Validation of miR-100 Hepatocyte-Specific Knock-Out Mice %U https://www.frontiersin.org/articles/10.3389/fonc.2019.00535 %V 9 %0 JOURNAL ARTICLE %@ 2234-943X %X Background: Inactivation of microRNA-100 (miR-100) is involved in hepatocellular carcinoma (HCC) and miR-100 behaves as a tumor suppressor. To understand miR-100 function in HCC genesis and development in vivo, we developed hepatocyte-specific miR-100 deficient mice.Methods: Mice homozygous for floxed miR-100 allele that carried the Alb-Cre transgene (miR-100flox/floxAlb -Cre+) were developed by mating miR-100flox/flox mice with Alb-Cre+/+mice. The mice tails DNA were genotyped using the primers for LoxP sites and Cre recombinase, respectively. The specific deletion of miR-100 in the livers was verified by quantitative Real-time PCR (qRT-PCR). HE-staining was performed for histology analysis. Liver function was assessed by transaminase activity. The metabolic profiles of the hepatocytes were detected using a Seahorse XFe24 extracellular flux analyzer. The direct targets of miR-100 (such as IGF1R-β, mTOR and CDC25A) and HCC related protein (SHP-2) were detected by qRT-PCR and Western blot in liver tissues.Results: The resultant homozygous knockout mice with genotype of miR-100flox/flox-Alb-Cre+ showed an 80% decrease in hepatic miR-100 expression. In adult mice, miR-100 knockout has no effect on the liver function and morphology. In aged mice, HE staining showed that miR-100 knockout caused infiltration of inflammatory cells and expansion of hepatocellular nuclei. Consistently, liver function was impaired in miR-100 knockout aged mice as indicated by increased serum AST and ALT levels. The metabolic analysis demonstrated that the miR-100 knockout hepatocytes tend to adopt glycolysis. The expressions of the miR-100 target genes, such as IGF1R-β, CDC25A and mTOR, were increased. In addition, the known HCC related protein, SHP-2 also was up-regulated in the knockout livers.Conclusions: We successfully generated a miR-100 hepatocyte-specific knock-out mouse model. The malignant transformation related to HCC were observed in aged mice. Therefore, this model is suitable for investigating the mechanism of miR-100 inactivation contributing to HCC genesis in vivo.