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Stem cell biology

Reprogramming human B cells into induced pluripotent stem cells and its enhancement by C/EBPα

Leukemia volume 30, pages 674–682 (2016)Cite this article

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Abstract

B cells have been shown to be refractory to reprogramming and B-cell-derived induced pluripotent stem cells (iPSC) have only been generated from murine B cells engineered to carry doxycycline-inducible Oct4, Sox2, Klf4 and Myc (OSKM) cassette in every tissue and from EBV/SV40LT-immortalized lymphoblastoid cell lines. Here, we show for the first time that freshly isolated non-cultured human cord blood (CB)- and peripheral blood (PB)-derived CD19+CD20+ B cells can be reprogrammed to iPSCs carrying complete VDJH immunoglobulin (Ig) gene monoclonal rearrangements using non-integrative tetracistronic, but not monocistronic, OSKM-expressing Sendai Virus. Co-expression of C/EBPα with OSKM facilitates iPSC generation from both CB- and PB-derived B cells. We also demonstrate that myeloid cells are much easier to reprogram than B and T lymphocytes. Differentiation potential back into the cell type of their origin of B-cell-, T-cell-, myeloid- and fibroblast-iPSCs is not skewed, suggesting that their differentiation does not seem influenced by ‘epigenetic memory’. Our data reflect the actual cell-autonomous reprogramming capacity of human primary B cells because biased reprogramming was avoided by using freshly isolated primary cells, not exposed to cytokine cocktails favoring proliferation, differentiation or survival. The ability to reprogram CB/PB-derived primary human B cells offers an unprecedented opportunity for studying developmental B lymphopoiesis and modeling B-cell malignancies.

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Acknowledgements

This work was supported by the European Research Council to P.M (ERC-2014-CoG-646903), the ISCIII/FEDER (E-Rare-2 Call PI12/03112 to PM, CP11/00131 to AFF and PI14/01191 to CB), MINECO (SAF2013-43065 to PM and SAF2012-37167 to TG), The Foundation TV3 Marato (120410 to TG), The Spanish Association Against Cancer to PM and MCC, and The Generalitat de Catalunya (SGR330 to PM and SGR1136 to TG). CB is supported by a Miguel Servet II contract (CPII13/00011). DRM and AM-L are supported by PFIS (FI11/0511) and FPI (BES-2014-067844) scholarships, respectively. PM also acknowledges the financial support from The Obra Social La Caixa-Fundaciò Josep Carreras. TG acknowledges the support by the Technology and Business Development Office of CRG within the framework of its proof-of-concept programs. CB, PM, JCS and OQ-B are investigators of the Spanish Cell Therapy cooperative network (TERCEL). Special thanks to Dr Lee Carpenter (University of Oxford, UK) and Dr Igor Slukvin (University of Wisconsin, MA) for providing MS5 and OP9 cells, and Dra. Dolors Comoler (IDIBAPS, Barcelona) for valuable help with the VDJH assays.

Author contributions

CB, JLS: conceived the study, designed and performed experiments, analyzed data and wrote the manuscript. KN, MO, MN: contributed key reagents and interpreted the data. BDS, DR-M, AM-L, MCC, AB, LA, JC, AH, OQ-B, JCS, MFF, AF, IG: performed experiments. TG: conceived the study. PM: conceived the study, designed experiments, analyzed data and wrote the manuscript.

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Author notes

  1. C Bueno and J L Sardina: These authors contributed equally to this work.

Authors and Affiliations

  1. Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain

    C Bueno, D Romero-Moya, A Muñoz-López, L Ariza, J Castaño, I Granada & P Menendez

  2. Gene Regulation, Stem Cells and Cancer Program, Centre for Genomic Regulation (CRG) and University Pompeu Fabra (UPF),, Barcelona, Spain

    J L Sardina, B Di Stefano & T Graf

  3. Department of Hematology, University Hospital of Salamanca, Instituto Biosanitario de Salamanca (IBSAL), Salamanca, Spain

    M C Chillón & A Balanzategui

  4. Servei d’Oncologia Radioteràpica, Hospital Clinic, Barcelona, Spain

    A Herreros

  5. Cancer Epigenetics Laboratory, Instituto Universitario de Oncología del Principado de Asturias, Universidad de Oviedo, Asturias, Spain

    M F Fraga & A Fernández

  6. Hematology Department, Hospital Germans Trias i Pujol, Institut Català d'Oncologia, Badalona, Spain

    I Granada

  7. Hematopoietic Innovative Therapy Division, Differentiation and Cytometry Unit, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Madrid, Spain

    O Quintana-Bustamante & J C Segovia

  8. Faculty of Medicine, Laboratory of Gene Regulation, University of Tsukuba, Tsukuba, Ibaraki, Japan

    K Nishimura

  9. Research Center for Stem Cell Engineering and National Institute of Advanced Industrial Science and Technology (AIST),, Tsukuba, Ibaraki, Japan

    M Ohtaka & M Nakanishi

  10. Instituciò Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain

    P Menendez

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  1. C Bueno
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  2. J L Sardina
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Correspondence to C Bueno or P Menendez.

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Bueno, C., Sardina, J., Di Stefano, B. et al. Reprogramming human B cells into induced pluripotent stem cells and its enhancement by C/EBPα. Leukemia 30, 674–682 (2016). https://doi.org/10.1038/leu.2015.294

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