Congenital amegakaryocytic thrombocytopenia iPS cells exhibit defective MPL-mediated signaling
Shinji Hirata, … , Shinji Kunishima, Koji Eto
Shinji Hirata, … , Shinji Kunishima, Koji Eto
Published August 1, 2013
Citation Information: J Clin Invest. 2013;123(9):3802-3814. https://doi.org/10.1172/JCI64721.
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Congenital amegakaryocytic thrombocytopenia iPS cells exhibit defective MPL-mediated signaling

Abstract

Congenital amegakaryocytic thrombocytopenia (CAMT) is caused by the loss of thrombopoietin receptor–mediated (MPL-mediated) signaling, which causes severe pancytopenia leading to bone marrow failure with onset of thrombocytopenia and anemia prior to leukopenia. Because Mpl–/– mice do not exhibit the human disease phenotype, we used an in vitro disease tracing system with induced pluripotent stem cells (iPSCs) derived from a CAMT patient (CAMT iPSCs) and normal iPSCs to investigate the role of MPL signaling in hematopoiesis. We found that MPL signaling is essential for maintenance of the CD34+ multipotent hematopoietic progenitor (MPP) population and development of the CD41+GPA+ megakaryocyte-erythrocyte progenitor (MEP) population, and its role in the fate decision leading differentiation toward megakaryopoiesis or erythropoiesis differs considerably between normal and CAMT cells. Surprisingly, complimentary transduction of MPL into normal or CAMT iPSCs using a retroviral vector showed that MPL overexpression promoted erythropoiesis in normal CD34+ hematopoietic progenitor cells (HPCs), but impaired erythropoiesis and increased aberrant megakaryocyte production in CAMT iPSC–derived CD34+ HPCs, reflecting a difference in the expression of the transcription factor FLI1. These results demonstrate that impaired transcriptional regulation of the MPL signaling that normally governs megakaryopoiesis and erythropoiesis underlies CAMT.

Authors

Shinji Hirata, Naoya Takayama, Ryoko Jono-Ohnishi, Hiroshi Endo, Sou Nakamura, Takeaki Dohda, Masanori Nishi, Yuhei Hamazaki, Ei-ichi Ishii, Shin Kaneko, Makoto Otsu, Hiromitsu Nakauchi, Shinji Kunishima, Koji Eto

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Figure 1

Disease-specific iPSCs recapitulate the disease phenotype manifested in a patient with CAMT.

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Disease-specific iPSCs recapitulate the disease phenotype manifested in ...
(A) Maternal mutation Q186X (C-to-T transition at the cDNA nucleotide position 556) in exon 4 and paternal mutation 1,499delT (single nucleotide deletion of thymine at position 1,499) in exon 10 in a patient with CAMT. SP, signal peptide; CY, cytoplasmic domain. (B) Generation of MKs and platelets from normal or CAMT iPSC–derived HPCs cultured on C3H10T1/2 feeder cells for 10 days in the presence of SCF (50 ng/ml), TPO (100 ng/ml), and heparin (25 U/ml). All CAMT iPSC clones generated few MKs or platelets. (C) Flow cytometric analysis of MPL-mediated downstream signaling in normal iPSC– (red lines) or CAMT iPSC–derived (blue lines) CD34+ HPCs stimulated with 100 ng/ml TPO for 10 minutes (solid lines) or with vehicle control (dotted lines). No response was observed with CAMT iPSCs. *P < 0.05.