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Establishment of an immortalized human erythroid cell line sustaining differentiation potential without inducible gene expression system


Ex vivo manufactured red blood cells (RBC) generated from immortalized erythroid cell lines which can continuously grow are expected to become a significant alternative in future transfusion therapies. The ectopic expression of human papilloma virus (HPV) E6/E7 gene has successfully been employed to establish these cell lines. To induce differentiation and maturation of the immortalized cell lines, terminating the HPV-E6/E7 expression through a gene induction system has been believed to be essential. Here, we report that erythroid cell lines established from human bone marrow using simple expression of HPV-E6/E7 are capable of normal erythroid differentiation, without turning gene expression off. Through simply changing cell culture conditions, a newly established cell line, Erythroid Line from Lund University (ELLU), is able to differentiate toward mature cells, including enucleated reticulocytes. ELLU is heterogeneous and, unexpectedly, clones expressing adult hemoglobin rapidly differentiate and produce fragile cells. Upon differentiation, other ELLU clones shift from fetal to adult hemoglobin expression, giving rise to more mature cells. Our findings propose that it is not necessary to employ gene induction systems to establish immortalized erythroid cell lines sustaining differentiation potential and describe novel cellular characteristics for desired functionally competent clones.

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We thank Martin L Olsson, Johan Flygare, and Jenny Hansson for scientific discussion, and Yukio Nakamura and Alexander Doyle for critical reading. This work was supported by the Swedish Research Council (K.M.), the Swedish Cancer Society (K.M.), and Royal Physiographic Society of Lund (S.S.). K.M. was funded by StemTherapy program at Lund University. The Lund Stem Cell Center was supported by a Center of Excellence grant in life sciences from the Swedish Foundation for Strategic Research.


Vetenskapsrådet (The Swedish Research Council), Cancerfonden (The Swedish Cancer Society), Kungl. Fysiografiska Sällskapet i Lund (Royal Physiographic Society of Lund).

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KM designed the project. SS and KM planned experiments. SS performed experiments. RK provided materials. HÅ supported experiments. NK gave specialists opinions. SS and KM wrote the manuscript.

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Correspondence to Kenichi Miharada.

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Human bone marrow was donated from a healthy donor with a written informed consent.

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Supplementary file1 Supplementary Figure 1 (A) RT-PCR analysis of expression of HPV16-E6, HPV16-E7 and GAPDH genes in ELLU clones at steady state and on day 14 of differentiation. (B) Representative plots of flow cytometry analysis for the frequencies of Band3+CD49d+ and Band3+CD49d- cells in the differentiating ELLU clones (PDF 2734 kb)

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Soboleva, S., Kurita, R., Kajitani, N. et al. Establishment of an immortalized human erythroid cell line sustaining differentiation potential without inducible gene expression system. Human Cell 35, 408–417 (2022).

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  • Red blood cells
  • Immortalized erythroid cell lines
  • HPV-E6/E7
  • Enucleation
  • Hemoglobin