Reprogramming human B cells into induced pluripotent stem cells and its enhancement by C/EBPα.
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.
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.
C
C
Bueno
Gene Regulation, Stem Cells and Cancer Program, Centre for Genomic Regulation (CRG) and University Pompeu Fabra (UPF), Barcelona, Spain.
J L
JL
Sardina
Gene Regulation, Stem Cells and Cancer Program, Centre for Genomic Regulation (CRG) and University Pompeu Fabra (UPF), Barcelona, Spain.
B
B
Di Stefano
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.
D
D
Romero-Moya
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.
A
A
Muñoz-López
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.
L
L
Ariza
Department of Hematology, University Hospital of Salamanca, Instituto Biosanitario de Salamanca (IBSAL), Salamanca, Spain.
M C
MC
Chillón
Department of Hematology, University Hospital of Salamanca, Instituto Biosanitario de Salamanca (IBSAL), Salamanca, Spain.
A
A
Balanzategui
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.
J
J
Castaño
Servei d'Oncologia Radioteràpica, Hospital Clinic, Barcelona, Spain.
A
A
Herreros
Cancer Epigenetics Laboratory, Instituto Universitario de Oncología del Principado de Asturias, Universidad de Oviedo, Asturias, Spain.
M F
MF
Fraga
Cancer Epigenetics Laboratory, Instituto Universitario de Oncología del Principado de Asturias, Universidad de Oviedo, Asturias, Spain.
A
A
Fernández
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.|Hematology Department, Hospital Germans Trias i Pujol, Institut Català d'Oncologia, Badalona, Spain.
I
I
Granada
Differentiation and Cytometry Unit, Hematopoietic Innovative Therapy Division, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Madrid, Spain.
O
O
Quintana-Bustamante
Differentiation and Cytometry Unit, Hematopoietic Innovative Therapy Division, Centro de Investigaciones Energéticas, Medioambientales y Tecnológicas (CIEMAT), Madrid, Spain.
J C
JC
Segovia
Faculty of Medicine, Laboratory of Gene Regulation, University of Tsukuba, Tsukuba, Ibaraki, Japan.
K
K
Nishimura
Research Center for Stem Cell Engineering and National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, Japan.
M
M
Ohtaka
Research Center for Stem Cell Engineering and National Institute of Advanced Industrial Science and Technology (AIST), Tsukuba, Ibaraki, Japan.
M
M
Nakanishi
Gene Regulation, Stem Cells and Cancer Program, Centre for Genomic Regulation (CRG) and University Pompeu Fabra (UPF), Barcelona, Spain.
T
T
Graf
Josep Carreras Leukemia Research Institute and School of Medicine, University of Barcelona, Barcelona, Spain.|Instituciò Catalana de Recerca i Estudis Avançats (ICREA), Barcelona, Spain.
P
P
Menendez
D001402:B-Lymphocytes / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D001483:Base Sequence; D022762:CCAAT-Enhancer-Binding Proteins / Q000235:genetics* / Q000276:immunology; D002454:Cell Differentiation; D002469:Cell Separation; D065150:Cellular Reprogramming / Q000235:genetics* / Q000276:immunology; D005312:Fetal Blood / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D015870:Gene Expression; D005822:Genetic Vectors; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors / Q000235:genetics / Q000276:immunology; D007963:Leukocytes, Mononuclear / Q000166:cytology / Q000276:immunology / Q000378:metabolism*; D008969:Molecular Sequence Data; D022423:Myeloid Cells / Q000166:cytology / Q000276:immunology / Q000378:metabolism; D050814:Octamer Transcription Factor-3 / Q000235:genetics / Q000276:immunology; D061251:Primary Cell Culture; D016271:Proto-Oncogene Proteins c-myc / Q000235:genetics / Q000276:immunology; D055748:SOXB1 Transcription Factors / Q000235:genetics / Q000276:immunology; D029082:Sendai virus / Q000235:genetics; D060152:V(D)J Recombination / Q000276:immunology
D022762:CCAAT-Enhancer-Binding Proteins; C515848:CEBPA protein, human; C000715467:KLF4 protein, human; D000090062:Kruppel-Like Factor 4; D051741:Kruppel-Like Transcription Factors; C489427:MYC protein, human; D050814:Octamer Transcription Factor-3; C494173:POU5F1 protein, human; D016271:Proto-Oncogene Proteins c-myc; C527782:SOX2 protein, human; D055748:SOXB1 Transcription Factors