title: 2‑Deoxy‑D‑glucose initiates hepatocyte differentiation in human induced pluripotent stem cells.
abstract: To initiate hepatocyte differentiation in human induced pluripotent stem (iPS) cells, cells are cultured in a medium lacking glucose but supplemented with galactose (hepatocyte selection medium, HSM) or in medium supplemented with oncostatin M and small molecules (hepatocyte differentiation inducer, HDI). In the present study, 2‑Deoxy‑D‑glucose (2DG), an analogue of glucose, was utilized instead of glucose deprivation and the effect of 2DG supplementation on iPS differentiation was examined. First, 201B7 cells, an iPS cell line, were cultured in HSM or HDI media for 2 days and then subjected to reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) in order to analyze expression levels of established hepatocyte markers, including cytosolic aspartate aminotransferase (AST), mitochondrial AST, alanine aminotransferase (ALT), and glycerol kinase. mRNA expression levels of mitochondrial AST, ALT, and glycogen synthase significantly increased following culture in HSM and HDI compared with ReproFF media. Cytosolic AST mRNA expression levels significantly increased following culture in HDI compared with ReproFF media, but not in HSM. To test the effect of 2DG on iPS differentiation, 201B7 cells were cultured in ReproFF, a feeder‑free medium that retains pluripotency, supplemented with 2DG. Following 7 days of culture, the cells were subjected to RT‑qPCR to analyze expression levels of α‑fetoprotein (AFP), a marker of immature hepatocytes. AFP mRNA expression levels significantly increased with the addition of 0.1 µM 2DG in the media, and galactose addition acted synergistically with 2DG to further upregulate AFP expression. In conclusion, the present study demonstrated that hepatocyte differentiation was initiated in iPS cells cultured in HSM and HDI media and that 2DG could be used as a supplement instead of glucose deprivation to initiate hepatocyte differentiation in iPS cells.
authors.affiliation: Department of Gastroenterology, National Hospital Organization, Shimoshizu Hospital, Yotsukaido, Chiba 284‑0003, Japan.
authors.forename: Minoru
authors.initials: M
authors.lastname: Tomizawa
authors.affiliation: Department of Radiology, National Hospital Organization, Shimoshizu Hospital, Yotsukaido, Chiba 284‑0003, Japan.
authors.forename: Fuminobu
authors.initials: F
authors.lastname: Shinozaki
authors.affiliation: Department of Neurology, National Hospital Organization, Shimoshizu Hospital, Yotsukaido, Chiba 284‑0003, Japan.
authors.forename: Yasufumi
authors.initials: Y
authors.lastname: Motoyoshi
authors.affiliation: Department of Rheumatology, National Hospital Organization, Shimoshizu Hospital, Yotsukaido, Chiba 284‑0003, Japan.
authors.forename: Takao
authors.initials: T
authors.lastname: Sugiyama
authors.affiliation: Department of Pediatrics, National Hospital Organization, Shimoshizu Hospital, Yotsukaido, Chiba 284‑0003, Japan.
authors.forename: Shigenori
authors.initials: S
authors.lastname: Yamamoto
authors.affiliation: Department of Neurosurgery, National Hospital Organization, Shimoshizu Hospital, Yotsukaido, Chiba 284‑0003, Japan.
authors.forename: Naoki
authors.initials: N
authors.lastname: Ishige
mesh_terms: D000410:Alanine Transaminase / Q000378:metabolism; D024443:Aspartate Aminotransferase, Cytoplasmic / Q000378:metabolism; D024442:Aspartate Aminotransferase, Mitochondrial / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects*; D002460:Cell Line; D003847:Deoxyglucose / Q000494:pharmacology*; D005690:Galactose / Q000378:metabolism; D005947:Glucose / Q000378:metabolism; D005991:Glycerol Kinase / Q000378:metabolism; D006006:Glycogen Synthase / Q000378:metabolism; D022781:Hepatocytes / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology* / Q000187:drug effects* / Q000378:metabolism; D053683:Oncostatin M / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D060888:Real-Time Polymerase Chain Reaction / Q000379:methods; D000509:alpha-Fetoproteins / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D000509:alpha-Fetoproteins; D053683:Oncostatin M; D003847:Deoxyglucose; D006006:Glycogen Synthase; D024443:Aspartate Aminotransferase, Cytoplasmic; D024442:Aspartate Aminotransferase, Mitochondrial; D000410:Alanine Transaminase; D005991:Glycerol Kinase; D005947:Glucose; D005690:Galactose