Retroviral vector-mediated gene transfer into umbilical cord blood-derived megakaryocyte and platelet progenitors.
Optimizing platelet engraftment following hematopoietic stem cell transplantation is essential for minimizing transplant-related morbidity, particularly following umbilical cord blood transplantation (UCBT), where platelet engraftment frequently takes >60 days. One strategy for optimizing platelet engraftment following UCBT is to study or alter the genetic program of megakaryocyte/platelet (Mk/plt) progenitors. Retroviral vector gene transfer has previously proven useful for studying the biology of hematopoietic stem cells; however, procedures for transducing UCB cells of the Mk/plt lineage with retroviral vectors have not been described. We report here that Mk/plt progenitors generated from UCB progenitors can be efficiently transduced with retroviral vectors. Transduced Mk/plt cells were identified and quantitated by expression of a vector transgene encoding a truncated version of the human nerve growth factor receptor (NGFR). Vector-mediated NGFR expression could be readily detected in Mk/plt progenitors defined by immunophenotypic, morphologic, and functional criteria. In addition, NGFR expression persisted in mature anucleate platelets generated from the transduced Mk/plt progenitors. These methods may be useful for introducing genetic elements into Mk/plt progenitors to study various aspects of platelet development and biology and for marking ex vivo expanded Mk/plt progenitors to determine their contribution to engraftment.
Castellino
S M
SM
Department of Pediatrics, Duke University Medical Center, Durham, North Carolina 27710, USA.
Kurtzberg
J
J
Smith
C
C
D018952:Antigens, CD34 / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D001792:Blood Platelets / Q000166:cytology / Q000276:immunology*; D002454:Cell Differentiation / Q000276:immunology / Q000502:physiology; D002455:Cell Division / Q000502:physiology; D002478:Cells, Cultured / Q000502:physiology; D005312:Fetal Blood / Q000187:drug effects / Q000276:immunology* / Q000378:metabolism; D018014:Gene Transfer Techniques; D005822:Genetic Vectors / Q000235:genetics / Q000276:immunology* / Q000494:pharmacology; D006801:Humans; D007231:Infant, Newborn; D008533:Megakaryocytes / Q000166:cytology / Q000276:immunology*; D019039:Platelet Glycoprotein GPIIb-IIIa Complex / Q000187:drug effects / Q000235:genetics / Q000276:immunology; D017475:Receptors, Nerve Growth Factor / Q000378:metabolism; D013234:Stem Cells / Q000166:cytology / Q000276:immunology* / Q000821:virology; D014161:Transduction, Genetic
D018952:Antigens, CD34; D019039:Platelet Glycoprotein GPIIb-IIIa Complex; D017475:Receptors, Nerve Growth Factor