miRNAs can increase the efficiency of ex vivo platelet generation
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ID: 114524
2012
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Abstract
The process of megakaryopoiesis culminates in the release of platelets, the pivotal cellular component for hemostasis and wound healing. The regulatory architecture including the modulatory role of microRNAs, which underlies megakaryocytic maturation and platelet formation, is incompletely understood, precluding the ex vivo generation of sufficient platelet numbers for transfusion medicine. We derived a highly efficient differentiation protocol to produce mature polyploid megakaryocytes and functional platelets from CD34+-hematopoietic stem and progenitor cells by comparing previously published approaches. Our megakaryocytic culture conditions using the cytokines SCF, TPO, IL-9, and IL-6 include nicotinamide and Rho-associated kinase (ROCK) inhibitor Y27632 as contextual additives. The potency of our novel megakaryocytic differentiation protocol was validated using cord blood and peripheral blood human hematopoietic stem and progenitor cells. Using this novel megakaryocytic differentiation protocol, we characterized the modulatory capacity of several miRNAs highly expressed in normal megakaryocytic cells or malignant blasts from patients with megakaryoblastic leukemia. Overexpression of candidate microRNAs was achieved by lentiviral transduction of CD34+-hematopoietic stem and progenitor cells prior to differentiation. We revealed miR-125b and miR-660 as enhancers of polyploidization, as well as platelet output of megakaryocytes. The oncogene miR-125b markedly expanded the number of megakaryocytes during in vitro culture. Conversely, the miR-23a/27a/24-2 cluster, which is highly expressed in normal megakaryocytes, blocked maturation and platelet formation. Our study on the utilization of microRNAs in conjunction with a highly efficient differentiation protocol constitutes another step towards ex vivo platelet manufacturing on a clinically relevant scale.
| Reference Key |
emmrich2012annalsmirnas
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| Authors | Stephan Emmrich;Kerstin Henke;Jan Hegermann;Matthias Ochs;Dirk Reinhardt;Jan-Henning Klusmann;Stephan Emmrich;Kerstin Henke;Jan Hegermann;Matthias Ochs;Dirk Reinhardt;Jan-Henning Klusmann; |
| Journal | Annals of hematology |
| Year | 2012 |
| DOI |
doi:10.1007/s00277-012-1517-z
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