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Efficient retroviral transduction of human B-lymphoid and myeloid progenitors: marked inhibition of their growth by the Pax5 transgene

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Abstract

We applied a coculture system for the genetic manipulation of human B-lymphoid and myeloid progenitor cells using murine bone marrow stromal cell support, and investigated the effects of forced Pax5 expression in both cell types. Cytokine-stimulated cord blood CD34+ cells could be transduced at 85% efficiency and 95% cell viability by a single 24-h infection with RD114-pseudotyped retroviral vectors, produced by the packaging cell line Plat-F and bicistronic vector plasmids pMXs-Ig, pMYs-Ig, or pMCs-Ig, encoding EGFP. Infected CD34+ cells were seeded onto HESS-5 cells in the presence of stem cell factor and granulocyte colony-stimulating factor, allowing the extensive production of B progenitors and granulocytic cells. We examined the cell number and CD34, CD33, CD19, and CD20 lambda and kappa expressions by flow cytometry. Ectopic expression of Pax5 in CD34+ cells resulted in small myeloid progenitors coexpressing CD33 and CD19 and inhibited myeloid differentiation. After 6 weeks, the number of Pax5-transduced CD19+ cells was 40-fold lower than that of control cells. However, the expression of CD20 and the κ/λ chain on Pax5-transduced CD19+ cells suggests that the Pax5 transgene may not interfere with their differentiation. This report is the first to describe the effects of forced Pax5 expression in human hematopoietic progenitors.

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Acknowledgments

This study was supported in part by a Grant-in-aid from the Ministry of Education, Science, Sports, and Culture of Japan. We thank Asuka Tajima for technical assistance and the Tokyo Cord Blood Bank for cord blood samples.

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Authors and Affiliations

  1. Division of Molecular Therapy, Advanced Clinical Research Center, Institute of Medical Science, University of Tokyo, 4-6-1 Shirokanedai, Minato-ku, Tokyo, 108-8639, Japan

    Rieko Sekine & Arinobu Tojo

  2. Division of Cellular Therapy, Institute of Medical Science, University of Tokyo, Tokyo, Japan

    Toshio Kitamura

  3. Department of Industrial Science and Technology, Science University of Tokyo, Noda, Japan

    Takashi Tsuji

Authors
  1. Rieko Sekine
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  2. Toshio Kitamura
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  3. Takashi Tsuji
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  4. Arinobu Tojo
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Correspondence to Arinobu Tojo.

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Sekine, R., Kitamura, T., Tsuji, T. et al. Efficient retroviral transduction of human B-lymphoid and myeloid progenitors: marked inhibition of their growth by the Pax5 transgene. Int J Hematol 87, 351–362 (2008). https://doi.org/10.1007/s12185-008-0082-7

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  • DOI: https://doi.org/10.1007/s12185-008-0082-7

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