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  • Cell-Based Therapy
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Cell-Based Therapy

The high proliferative potential-quiescent (HPP-Q) cell assay allows an optimized evaluation of gene transfer efficiency into primitive hematopoietic stem/progenitor cells

Gene Therapy volume 7, pages 1790–1794 (2000)Cite this article

Abstract

Various protocols have been described to optimize gene transfer into hematopoietic cells. However, most of these methods do not specify whether they are associated with an improved transduction of the more primitive stem/progenitor cells, the best candidates for long-term engraftment. The majority of these primitive cells remains in quiescence because of the negative control of TGF-β1, effective on these cells at low concentrations (10 pg/ml). In this study, CD34+ cells were activated by a 10 h pretreatment with anti-TGF-β1 followed by four successive retroviral supernatant incubations of 6 h each. After 12 h (two incubations), a significant increase in TGF-β1 mRNA in CD34+ cells was observed. We wondered whether neo-synthesized autocrine TGF-β1 could induce reversion to quiescence of the more primitive CD34+ cells transduced after one cell cycle. This would prevent their subsequent detection in a classic clonal assay. Using the HPP-Q assay comparing a rapid mixed colony assay with or without anti-TGF-β1, we indeed observed, that in clonal growth conditions the more primitive transduced cells were activated and detectable only with anti-TGF-β1. Therefore, this assay represents not only a rapid means to detect quiescent multipotent stem/progenitor cells but also a necessary step for the detection of the more primitive transduced cells which have returned to quiescence after retroviral induction of TGF-β1 secretion.

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Acknowledgements

We thank Dr RC Mulligan for providing the MFG-nlsLacZ A7–21 producer clone. We thank Dr C Paillissé for her assistance in statistical computations. This study was supported by the Centre National de la Recherche Scientifique (CNRS), the Agence Nationale de la Recherche sur le Sida (ANRS), the European contract No. BIO4-CT96–0646. KD and NF have been recipients of fellowships from the Ministère de l’Éducation Nationale, de l'Enseignement Supérieur et de la Recherche (MENESR) and from the Association pour la Recherche sur le Cancer (ARC). SK was supported by the Direction des Recherches Etudes et Techniques (DRET).

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  1. Laboratoire de Biologie des Cellules Souches Somatiques Humaines, Centre National de la Recherche Scientifique (CNRS), UPR 1983, 7 rue Guy Môquet, Villejuif, 94800, France

    K Ducos, A Hatzfeld, A Héron, N Fortunel, S Kisselev, C Héron, M N Monier & J Hatzfeld

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  1. K Ducos
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  2. A Hatzfeld
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  4. N Fortunel
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  7. M N Monier
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Ducos, K., Hatzfeld, A., Héron, A. et al. The high proliferative potential-quiescent (HPP-Q) cell assay allows an optimized evaluation of gene transfer efficiency into primitive hematopoietic stem/progenitor cells. Gene Ther 7, 1790–1794 (2000). https://doi.org/10.1038/sj.gt.3301304

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