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Open Access The Assessment of Human Erythroid Output in NOD/SCID Mice Reconstituted With Human Hematopoietic Stem Cells

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Abstract:

The third-generation NOD/LtSz-scid/IL2Rnull (NOD/SCID IL2Rnull) mouse represents a significantly improved xenograft model allowing high levels of human leukocyte engraftment over extended follow up. One remaining limitation of this mouse model, however, is the low level of circulating human erythrocytes. We established a practical ex vivo erythroid culture system of xenograft marrow progenitors to enrich for human erythroid progeny. At various time points after transplant, erythroid cells were easily assayed after 17 days of ex vivo culture of xenograft marrow, with nearly all nucleated cells of human origin and approximately 60% human GPA or CD71 positive. We then transplanted cord blood CD34+ cells marked with a lentiviral vector encoding green fluorescent protein (GFP). Three months later, ex vivo culture of xenograft marrow progenitors showed 41.3% of the cultured erythroid cells were positive for GFP and human CD71, and 56.2% were positive for GFP and human GPA, similar to that of circulating leukocytes at the same time point. Next, G-CSF mobilized peripheral blood CD34+ cells from a sickle cell trait subject were infused in this mouse model to determine if the hemoglobin pattern could be modeled. CD34+ cells from the sickle cell trait subject engrafted equally compared to CD34+ cells from normal subjects, establishing the sickle cell trait phenotype. Lastly, a comparison of adult-derived peripheral blood CD34+ cells and cord blood-derived CD34+ cells xenografted mice was made, and long term follow-up demonstrated a recapitulation of the fetal to adult hemoglobin switch. This approach should prove a useful tool for testing strategies for genetic manipulation of erythroid progeny and the study of hemoglobin switching.

Keywords: Erythropoiesis; GFP; HbF; HbS; Hemoglobin switching; Lentiviral vector; NOD/LtSz-scid/IL2Rnull mice; Sickle cell; Xenograft mouse model

Document Type: Research Article

DOI: http://dx.doi.org/10.3727/096368910X314161

Affiliations: Molecular and Clinical Hematology Branch, National Institutes of Diabetes and Digestive and Kidney Disorders (NIDDK) and National Heart, Lung, and Blood Institute (NHLBI), National Institutes of Health, Bethesda, MD, USA

Publication date: November 1, 2010

More about this publication?
  • Cell Transplantation publishes original, peer-reviewed research and review articles on the subject of cell transplantation and its application to human diseases. To ensure high-quality contributions from all areas of transplantation, separate section editors and editorial boards have been established. Articles deal with a wide range of topics including physiological, medical, preclinical, tissue engineering, and device-oriented aspects of transplantation of nervous system, endocrine, growth factor-secreting, bone marrow, epithelial, endothelial, and genetically engineered cells, among others. Basic clinical studies and immunological research papers are also featured. To provide complete coverage of this revolutionary field, Cell Transplantation will report on relevant technological advances, and ethical and regulatory considerations of cell transplants. Cell Transplantation is now an Open Access journal starting with volume 18 in 2009, and therefore there will be an inexpensive publication charge, which is dependent on the number of pages, in addition to the charge for color figures. This will allow work to be disseminated to a wider audience and also entitle the corresponding author to a free PDF, as well as prepublication of an unedited version of the manuscript.
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