Inhibition of Lnk in mouse induced pluripotent stem cells promotes hematopoietic cell generation.
Embryonic stem (ES) cell- and induced pluripotent stem (iPS) cell-derived hematopoietic stem/progenitor cells (HSPCs) are considered as an unlimited source for HSPC transplantation. However, production of immature hematopoietic cells, especially HSPCs, from ES and iPS cells has been challenging. The adaptor protein Lnk has been shown to negatively regulate HSPC function via the inhibition of thrombopoietin (TPO) and stem cell factor signaling, and Lnk-deficient HSPCs show an enhanced self-renewal and repopulation capacity. In this study, we examined the role of Lnk on the hematopoietic differentiation from mouse ES and iPS cells by the inhibition of Lnk using a dominant-negative mutant of the Lnk (DN-Lnk) gene. We generated mouse ES and iPS cells stably expressing a DN-Lnk, and found that enforced expression of a DN-Lnk in ES and iPS cells led to an enhanced generation of Flk-1-positive mesodermal cells, thereby could increase in the expression of hematopoietic transcription factors, including Scl and Runx1. We also showed that the number of both total hematopoietic cells and immature hematopoietic cells with colony-forming potential in DN-Lnk-expressing cells was significantly increased in comparison with that in control cells. Furthermore, Lnk inhibition by the overexpression of the DN-Lnk gene augmented the TPO-induced phosphorylation of Erk1/2 and Akt, indicating the enhanced sensitivity to TPO. Adenovirus vector-mediated transient DN-Lnk gene expression in ES and iPS cells could also increase the hematopoietic cell production. Our data clearly showed that the inhibition of Lnk in ES and iPS cells could result in the efficient generation and expansion of hematopoietic cells.
Laboratory of Stem Cell Regulation, National Institute of Biomedical Innovation, Osaka, Japan.
Katsuhisa
K
Tashiro
Miyuki
M
Omori
Kenji
K
Kawabata
Nobue
N
Hirata
Tomoko
T
Yamaguchi
Fuminori
F
Sakurai
Satoshi
S
Takaki
Hiroyuki
H
Mizuguchi
D048868:Adaptor Proteins, Signal Transducing; D000818:Animals; D051792:Basic Helix-Loop-Helix Proteins / Q000096:biosynthesis; D002454:Cell Differentiation / Q000235:genetics*; D002460:Cell Line; D050676:Core Binding Factor Alpha 2 Subunit / Q000096:biosynthesis; D048049:Extracellular Signal-Regulated MAP Kinases / Q000378:metabolism; D006412:Hematopoietic Stem Cells; D057026:Induced Pluripotent Stem Cells / Q000378:metabolism*; D047908:Intracellular Signaling Peptides and Proteins / Q000235:genetics*; D008565:Membrane Proteins; D051379:Mice; D010766:Phosphorylation; D011518:Proto-Oncogene Proteins / Q000096:biosynthesis; D051057:Proto-Oncogene Proteins c-akt / Q000378:metabolism; D015398:Signal Transduction / Q000235:genetics; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; D013926:Thrombopoietin / Q000378:metabolism; D040301:Vascular Endothelial Growth Factor Receptor-2 / Q000096:biosynthesis
D048868:Adaptor Proteins, Signal Transducing; D051792:Basic Helix-Loop-Helix Proteins; D050676:Core Binding Factor Alpha 2 Subunit; D048049:Extracellular Signal-Regulated MAP Kinases; D047908:Intracellular Signaling Peptides and Proteins; D008565:Membrane Proteins; D011518:Proto-Oncogene Proteins; D051057:Proto-Oncogene Proteins c-akt; D000073942:T-Cell Acute Lymphocytic Leukemia Protein 1; D013926:Thrombopoietin; D040301:Vascular Endothelial Growth Factor Receptor-2; C490777:Lnk protein, mouse; C493726:Runx1 protein, mouse; C489366:Tal1 protein, mouse