MicroRNAs are the regulatory molecules in post-transcriptional regulation of gene expression, which affect diverse biological processes and have been found to play important roles in regulating stem cell character in plants and animals. The aim of this study was to identify the role of miR-122 during hepatic differentiation of human adipose tissue-derived stem cells (hADSCs), and also to investigate whether overexpression of miR-122 could enhance differentiation of hADSCs toward functional hepatocyte-like cells without any extrinsic factor. To investigate this, the level of miR-122 was monitored by quantitative real-time PCR (qRT-PCR) at specific time intervals following hepatic differentiation of hADSCs using growth factors. For the next step, lentiviral transduction was applied to overexpress miR-122 in hADSCs for up to 21 days. Hepatic functionality was evaluated by analyzing specific hepatocyte genes and biochemical markers at different time points of differentiation induction. The qRT-PCR results revealed that miR-122 was upregulated during hepatic differentiation of hADSCs. Additionally, the stable miR-122 overexpression in hADSCs resulted in increased expression of specific hepatocyte markers such as ALB, AFP, CK18, CK19, and HNF4a compared with the negative control cells. Moreover, urea and albumin production as well as glycogen deposits were observed in the treated cells. Therefore, our findings demonstrate that the hepatic differentiation process could be improved by the overexpression of miR-122 in hADSCs, making it a potential therapeutic resource for liver disorders.

MicroRNAs (miRNAs) are noncoding RNAs involved in the regulation of the diverse biological processes such as metabolism, proliferation, and cell cycle, in addition to regulation of differentiation. So far, some miRNAs have been recognized to have important role in regulating hepatic functions. Statistically, let-7f has been revealed as a negative regulator of hepatic differentiation. In the present study, we investigated the effect of let-7f on hepatic differentiation of human adipose tissue-derived stem cells (hADSCs). hADSCs were transduced with recombinant lentivirus containing human inhibitor let-7 f. The expression of hepatocyte nuclear factors alpha (HNF4a), albumin (ALB), alpha fetoprotein (AFP), cytokeratin 18 (CK18), and cytokeratin 19 (CK19) was evaluated using quantitative real-time PCR (qRT-PCR). Immunocytochemistry was used to investigate the expression levels of the hepatocyte markers including ALB, AFP, and HNF4a, and biochemical analysis was implemented for hepatic function, glycogen deposition, and urea secretion. qRT-PCR showed significant upregulation in HNF4a, ALB, AFP, CK18, and CK19 expression in cells transduced with let-7f inhibitor lentiviruses. Moreover, positive staining was detected for ALB, AFP, and HNF4a using immunocytochemistry. Urea production and glycogen deposits were also found in the treated cells, the two specific features of the hepatic cells. Therefore, let-7f silencing led to the increased expression of the hepatocyte-specific factors and the accelerated hADSCs hepatic differentiation. Summing all these finding together, our present report has provided evidences that inhibition of let-7f would facilitate induction of hADSCs into hepatocyte-like cells and possibly in regenerative therapy of the liver disease in a wider spectrum.

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Human adipose tissue-derived stem cells (hADSCs) have been considered as a promising source for cell therapy of liver diseases due to their accessibility, abundance, and expression of hepatocyte markers. Currently, the important role of certain microRNAs (miRNAs) has been reported during hepatic differentiation of stem cells. However, the combination effect of miRNAs on hepatic differentiation of these cells needs to be more investigated. The present study seeks to determine whether the combination of miRNAs can enhance hepatic differentiation of hADSCs in the absence of any other stimulation. First, lentiviral transduction was used to overexpress miR-122 and silence let-7f in hADSCs for up to 21 days. Then, hepatic functionality was evaluated by analyzing specific hepatocyte genes and biochemical markers at different time points of differentiation induction. Stable miR-122 overexpression and let-7f silencing together in hADSCs resulted in increased expression of hepatocyte markers including ALB, AFP, CK18, CK19, and HNF4a. In addition, urea and albumin production, immunocytochemistry, and glycogen staining confirmed that the treated cells differentiated toward hepatocyte-like cells. Therefore, our findings demonstrate the possibility of using microRNAs to induce hADSCs into functional hepatocyte-like cells.

There are numerous studies which provide support for the use of human adipose tissue-derived stem cells (hASCs) to generate hepatocyte-like cells. However, the produced cells exhibit only a certain level of differentiation, mainly due to inefficient induction conditions. Therefore, based on the important role of insulin-like growth factor (IGF-I) in hepatic function and development, in the current study we evaluated the differentiation efficacy of the mentioned factor to induce hASCs into functional hepatocyte-like cells. To investigate this, using a two-step protocol, hASCs were treated with a combination of HGF, Dex, and OSM in the presence or absence of IGF-I up to 21 days. Hepatic differentiation was evaluated by analyzing specific hepatocyte markers at different time points of differentiation induction. Increased expression of hepatocyte-specific genes including ALB, AFP, CK18, and HNF4a, downregulation of bile duct cells marker (CK19), the higher number of ALB positive cells, increased urea production together with higher glycogen deposit was observed upon the treatment of hASCs with the induction medium containing IGF-I compared to the other treatment. In conclusion, our findings suggest IGF-I as a potent inducer of hepatic differentiation of hASCs and its potential to generate more functional hepatocyte-like cells.