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Ex vivo expansion and long-term hematopoietic reconstitution ability of sorted CD34+CD59+ cells from patients with paroxysmal nocturnal hemoglobinuria

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Abstract

Autologous bone marrow transplantation (ABMT) for paroxysmal nocturnal hemoglobinuria (PNH) remains difficult so far. To expand residual normal CD34+CD59+ cells isolated from patients with PNH and observe the long-term hematopoietic reconstruction ability of the expanded cells both ex vivo and in vivo, CD34+CD59+ cells from 13 PNH patients and CD34+ cells from 11 normal controls were separated from bone marrow mononuclear cells first by immunomagnetic microbeads and then by flow cytometry autoclone sorting. The cells were then cultivated under different conditions. The long-term hematopoietic supporting ability of expanded CD34+CD59+ cells was evaluated by long-term culture in semi-solid medium in vitro and long-term engraftment in irradiated severe combined immunodeficiency (SCID) mice in vivo. The best combination of hematopoietic growth factors for ex vivo expansion was SCF + IL-3 + IL-6 + FL + Tpo + Epo. The most suitable time for harvest was on day 7. CD34+CD59+ PNH cells retained strong colony-forming capacity even after expansion. The survival rate, complete blood cell count recovery on day 90, and human CD45 expression in different organs were similar between the irradiated SCID mice transplanted with expanded CD34+CD59+ PNH cells and those with normal CD34+ cells (P > 0.05) both in primary and secondary transplantation. These data provided a new potential way of managing PNH with ABMT.

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Acknowledgments

We thank National Important Laboratory of Chinese Basic Medicine Institute, Chinese Academy of Medical Science and Central Laboratory of Peking Union Medical College Hospital where most of our study was done, and we also thank Profs Zhi-nan Zhang and Ti Shen (Peking Union Medical College, Chinese Academy of Medical Science) for valuable comments. This work was supported by grant of Clinic and Basic Research program from Chinese Health Department and by research funds of Peking Union Medical College Hospital, Chinese Academy of Medical Science.

Conflict of interest statement

The authors declare no competing financial interests.

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Authors and Affiliations

  1. Department of Pediatrics, Peking Union Medical College Hospital, Chinese Academe of Medical Science, Beijing, 100730, China

    Juan Xiao

  2. Department of Hematology, Peking Union Medical College Hospital, Chinese Academe of Medical Science, Beijing, China

    Bing Han, Yong-ji Wu, Yu-ping Zhong & Wan-ling Sun

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  1. Juan Xiao
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  2. Bing Han
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Correspondence to Bing Han.

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Xiao, J., Han, B., Wu, Yj. et al. Ex vivo expansion and long-term hematopoietic reconstitution ability of sorted CD34+CD59+ cells from patients with paroxysmal nocturnal hemoglobinuria. Int J Hematol 92, 58–67 (2010). https://doi.org/10.1007/s12185-010-0628-3

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  • DOI: https://doi.org/10.1007/s12185-010-0628-3

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