Cell line-dependent differentiation of induced pluripotent stem cells into cardiomyocytes in mice.
OBJECTIVE
Mouse and human fibroblasts can be directly reprogrammed to pluripotency by the ectopic expression of four transcription factors (Oct3/4, Sox2, Klf4, and c-Myc) to yield induced pluripotent stem (iPS) cells. iPS cells can be generated even without the expression of c-Myc. The present study examined patterns of differentiation of mouse iPS cells into cardiomyocytes in three different cell lines reprogrammed by three or four factors.


RESULTS
During the induction of differentiation on feeder-free gelatinized dishes, genes involved in cardiogenesis were expressed as in embryonic stem cells and myogenic contraction occurred in two iPS cell lines. However, in one iPS cell line (20D17) generated by four factors, the expression of cardiac-specific genes and the beating activity were extremely low. Treating iPS cells with trichostatin A (TSA), a histone deacetylase (HDAC) inhibitor, increased Nkx2.5 expression in all iPS cell lines. While the basal Nkx2.5 expression was very low in 20D17, the TSA-induced increase was the greatest. TSA also induced the expression of contractile proteins in 20D17. Furthermore, we demonstrated the increased mRNA level of Oct3/4 and nuclear protein level of HDAC4 in 20D17 compared with the other two iPS cell lines. DNA microarray analysis identified genes whose expression is up- or down-regulated in 20D17.


CONCLUSIONS
Mouse iPS cells differentiate into cardiomyocytes in a cell line-dependent manner. TSA induces myocardial differentiation in mouse iPS cells and might be useful to overcome cell line variation in the differentiation efficiency.
Kaichi
Shinji
S
Department of Pediatrics, Graduate School of Medicine, Kyoto University, Kyoto, Japan. koj@kuhp.kyoto-u.ac.jp
Hasegawa
Koji
K
Takaya
Tomohide
T
Yokoo
Noritaka
N
Mima
Takahiro
T
Kawamura
Teruhisa
T
Morimoto
Tatsuya
T
Ono
Koh
K
Baba
Shiro
S
Doi
Hiraku
H
Yamanaka
Shinya
S
Nakahata
Tatsutoshi
T
Heike
Toshio
T
D000107:Acetylation; D000818:Animals; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation / Q000187:drug effects; D002460:Cell Line; D019070:Cell Lineage; D002469:Cell Separation / Q000379:methods; D018920:Coculture Techniques; D053595:Embryonic Stem Cells / Q000187:drug effects / Q000502:physiology*; D005434:Flow Cytometry; D020869:Gene Expression Profiling / Q000379:methods; D018507:Gene Expression Regulation, Developmental; D056572:Histone Deacetylase Inhibitors / Q000494:pharmacology; D006655:Histone Deacetylases / Q000378:metabolism; D006657:Histones / Q000378:metabolism; D006877:Hydroxamic Acids / Q000494:pharmacology; D007150:Immunohistochemistry; D057026:Induced Pluripotent Stem Cells / Q000187:drug effects / Q000502:physiology*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000378:metabolism; D051379:Mice; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000187:drug effects / Q000502:physiology*; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000378:metabolism; D020411:Oligonucleotide Array Sequence Analysis; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000378:metabolism; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000378:metabolism; D013997:Time Factors; D014162:Transfection
D015415:Biomarkers; D056572:Histone Deacetylase Inhibitors; D006657:Histones; D006877:Hydroxamic Acids; C000715467:KLF4 protein, human; C000715468:Klf4 protein, mouse; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; D050814:Octamer Transcription Factor-3; D016271:Proto-Oncogene Proteins c-myc; D055748:SOXB1 Transcription Factors; C012589:trichostatin A; C498618:Hdac5 protein, mouse; D006655:Histone Deacetylases