Overexpression of miR-122 promotes the hepatic differentiation and maturation of mouse ESCs through a miR-122/FoxA1/HNF4a-positive feedback loop

• Published in: Liver international 2014-02, Vol.34 (2), p.281-295
• Main Authors: Deng, Xiao-Geng, Qiu, Rong-Lin, Wu, Yao-Hao, Li, Zhi-Xi, Xie, Ping, Zhang, Jie, Zhou, Jia-Jia, Zeng, Le-Xiang, Tang, Jing, Maharjan, Anu, Deng, Jie-Min
• Format: Article
• Language: English
• Subjects: Animals; Cell Differentiation - physiology; differentiation; embryonic stem cells; Embryonic Stem Cells - cytology; Embryonic Stem Cells - metabolism; Feedback, Physiological - physiology; Flow Cytometry; Hepatocyte Nuclear Factor 3-alpha - metabolism; Hepatocyte Nuclear Factor 4 - metabolism; hepatocytes; Hepatocytes - cytology; Hepatocytes - metabolism; mesenchymal-to-epithelial transition; Mice; Microarray Analysis; MicroRNAs - metabolism; Microscopy, Fluorescence; miR-122; Real-Time Polymerase Chain Reaction
• ISSN: 1478-3223; 1478-3231
• DOI: 10.1111/liv.12239

Background & Aims microRNA‐122 is the only identified liver‐specific miRNA and plays a crucial role in liver development, maintenance of hepatic homeostasis as well as tumourigenesis. In our previous differentiation of ESCs into hepatocytes, microRNA‐122 (miR‐122) was expressed at a relatively low level. Here, we aim to elucidate the effect and underlying mechanisms of miR‐122 during differentiation of ESCs into hepatocytes. Methods Mouse ESCs were initially induced towards HPCs by activin A, FGF‐4 and sodium butyrate and were subsequently transfected with a recombinant adenovirus expressing vector pAV.Ex1d‐CMV>miR‐122/IRES/eGFP 9 days after induction. Cells were analysed by real‐time PCR, immunofluorescence, flow cytometry, microscopy and functional assays. Furthermore, microarray analysis was performed. Results We demonstrated that overexpression of miR‐122 could effectively promote hepatic differentiation and maturation, as assessed by morphological and functional tests. The microarray analysis revealed that 323 genes were down‐regulated, whereas 59 were up‐regulated. Particularly, two liver‐specific transcription factors, FoxA1 and HNF4a, were significantly up‐regulated. Moreover, the expression of E‐cadherin was dramatically increased and the proliferation of HPCs was suppressed, whereas knockdown of FoxA1 reduced E‐cadherin expression and increased the proliferation of HPCs. In addition, the expression levels of FoxA1, HNF4a and E‐cadherin in time‐course transfection experiments with miR‐122 were not significantly increased except in cells in which transfection with miR‐122 occurred 9 days after induction. Conclusion Overexpression of miR‐122 at an appropriate stage could promote hepatic differentiation and maturation by regulating the balance between proliferation and differentiation, as well as the balance between EMT and MET, partially through a miR‐122/FoxA1/HNF4a‐positive feedback loop.