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Generation of metabolically functional hepatocyte-like cells from dedifferentiated fat cells by Foxa2, Hnf4a and Sall1 transduction.

ABSTRACT: Mature adipocyte-derived dedifferentiated fat (DFAT) cells have been identified to possess similar multipotency to mesenchymal stem cells, but a method for converting DFAT cells into hepatocytes was previously unknown. Here, using comprehensive analysis of gene expression profiles, we have extracted three transcription factors, namely Foxa2, Hnf4a and Sall1 (FHS), that can convert DFAT cells into hepatocytes. Hepatogenic induction has converted FHS-infected DFAT cells into an epithelial-like morphological state and promoted the expression of hepatocyte-specific features. Furthermore, the DFAT-derived hepatocyte-like (D-Hep) cells catalyzed the detoxification of several compounds. These results indicate that the transduction of DFAT cells with three genes, which were extracted by comprehensive gene expression analysis, efficiently generated D-Hep cells with detoxification abilities similar to those of primary hepatocytes. Thus, D-Hep cells may be useful as a new cell source for surrogate hepatocytes and may be applied to drug discovery studies, such as hepatotoxicity screening and drug metabolism tests.

SUBMITTER: Hagiwara R

PROVIDER: S-EPMC7894465 | biostudies-literature | 2020 Dec

REPOSITORIES: biostudies-literature

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Generation of metabolically functional hepatocyte-like cells from dedifferentiated fat cells by Foxa2, Hnf4a and Sall1 transduction.

Hagiwara Reiko R Oki Yoshinao Y Matsumaru Takashi T Ibayashi Shiho S Kano Koichiro K

Genes to cells : devoted to molecular & cellular mechanisms 20201110 12

Mature adipocyte-derived dedifferentiated fat (DFAT) cells have been identified to possess similar multipotency to mesenchymal stem cells, but a method for converting DFAT cells into hepatocytes was previously unknown. Here, using comprehensive analysis of gene expression profiles, we have extracted three transcription factors, namely Foxa2, Hnf4a and Sall1 (FHS), that can convert DFAT cells into hepatocytes. Hepatogenic induction has converted FHS-infected DFAT cells into an epithelial-like mor ...[more]

PMID: 33064855

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Project description:Somatic cell reprogramming has generated enormous interest after the first report by Yamanaka and his coworkers in 2006 on the generation of induced pluripotent stem cells (iPSCs) from mouse fibroblasts. Here we report the generation of stable iPSCs from mouse fibroblasts by recombinant protein transduction (Klf4, Oct4, Sox2, and c-Myc), a procedure designed to circumvent the risks caused by integration of exogenous sequences in the target cell genome associated with gene delivery systems. The recombinant proteins were fused in the frame to the glutathione-S-transferase tag for affinity purification and to the transactivator transcription-nuclear localization signal polypeptide to facilitate membrane penetration and nuclear localization. We performed the reprogramming procedure on embryonic fibroblasts from inbred (C57BL6) and outbred (ICR) mouse strains. The cells were treated with purified proteins four times, at 48-h intervals, and cultured on mitomycin C treated mouse embryonic fibroblast (MEF) cells in complete embryonic stem cell (ESC) medium until colonies formed. The iPSCs generated from the outbred fibroblasts exhibited similar morphology and growth properties to ESCs and were sustained in an undifferentiated state for more than 20 passages. The cells were checked for pluripotency-related markers (Oct4, Sox2, Klf4, cMyc, Nanog) by immunocytochemistry and by reverse transcription-polymerase chain reaction. The protein iPSCs (piPSCs) formed embryoid bodies and subsequently differentiated towards all three germ layer lineages. Importantly, the piPSCs could incorporate into the blastocyst and led to variable degrees of chimerism in newborn mice. These data show that recombinant purified cell-penetrating proteins are capable of reprogramming MEFs to iPSCs. We also demonstrated that the cells of the generated cell line satisfied all the requirements of bona fide mouse ESCs: form round colonies with defined boundaries; have a tendency to attach together with high nuclear/cytoplasmic ratio; express key pluripotency markers; and are capable of in vitro differentiation into ecto-, endo-, and mesoderm, and in vivo chimera formation.

Dataset Information

Generation of metabolically functional hepatocyte-like cells from dedifferentiated fat cells by Foxa2, Hnf4a and Sall1 transduction.

Publications

Generation of metabolically functional hepatocyte-like cells from dedifferentiated fat cells by Foxa2, Hnf4a and Sall1 transduction.

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