Currently, volunteer donors serve as the primary source of platelets for transfusion. Donor-derived platelets have an extremely short shelf life, are not always available, and have potential to transmit infectious diseases to the recipient. Due to these limitations, induced pluripotent stem (iPS) cells and embryonic stem (ES) cells have been explored as renewable platelet sources for clinical applications. Platelets arise from megakaryocyte-erythroid progenitors (MEPs), and mutations in the gene encoding the transcription factor GATA1 lead to the MEP accumulation in both mice and humans. While GATA1-deficient murine ES cells can serve as a renewable source of MEPs in vitro, these progenitors do not fully mature into platelet-producing megakaryocytes following induced overexpression of GATA1. Ji-Yoon Noh and colleagues at the Children’s Hospital of Philadelphia developed a protocol that restores endogenous GATA1 levels in MEPs derived from murine ES cells in which Gata1 mRNA is inducibly targeted for degradation, resulting in the generation of megakaryocytes capable of platelet generation. Specifically, murine ES cells were engineered to conditionally suppress Gata1 in the presence of doxycycline (dox), as the result of a Gata1-targeting shRNA under the control of a tetracycline response element. In the presence of dox, these engineered murine ES cells expressed hematopoiesis-supporting factors, developed an MEP-like profile, and expanded robustly. Dox removal restored endogenous Gata1 expression, with a concomitant differentiation of MEP-like cells to mature megakaryocytes that were able to produce clinically relevant numbers of functional platelets. In a companion Commentary, Matthew Canver, Daniel Bauer, and Stuart Orkin at Harvard Medical School discuss potential next steps towards the clinical realization of stem cell-derived platelet therapies. The accompanying movie shows platelets from murine ES cell-derived MEPs (green) incorporating into developing thrombi at the site of arteriole injury. Note that these cells express the activation marker P-selectin (blue) following clot lodging, similar to endogenous platelets.
Transfusion of donor-derived platelets is commonly used for thrombocytopenia, which results from a variety of clinical conditions and relies on a constant donor supply due to the limited shelf life of these cells. Embryonic stem (ES) and induced pluripotent stem (iPS) cells represent a potential source of megakaryocytes and platelets for transfusion therapies; however, the majority of current ES/iPS cell differentiation protocols are limited by low yields of hematopoietic progeny. In both mice and humans, mutations in the gene-encoding transcription factor GATA1 cause an accumulation of proliferating, developmentally arrested megakaryocytes, suggesting that GATA1 suppression in ES and iPS cell–derived hematopoietic progenitors may enhance megakaryocyte production. Here, we engineered ES cells from WT mice to express a doxycycline-regulated (dox-regulated) shRNA that targets
Ji-Yoon Noh, Shilpa Gandre-Babbe, Yuhuan Wang, Vincent Hayes, Yu Yao, Paul Gadue, Spencer K. Sullivan, Stella T. Chou, Kellie R. Machlus, Joseph E. Italiano Jr., Michael Kyba, David Finkelstein, Jacob C. Ulirsch, Vijay G. Sankaran, Deborah L. French, Mortimer Poncz, Mitchell J. Weiss
The creation of a donor-independent source of platelets has been challenging; however, recent advances show growing promise for alternative platelet sources. Pluripotent stem cells have the capacity to differentiate into mature megakaryocytes with the ability to produce functional platelets. In this issue of
Matthew C. Canver, Daniel E. Bauer, Stuart H. Orkin