Rapid and Efficient Generation of Transgene-Free iPSC from a Small Volume of Cryopreserved Blood.
Abstract
Human peripheral blood and umbilical cord blood represent attractive sources of cells
for reprogramming to induced pluripotent stem cells (iPSCs). However, to date, most
of the blood-derived iPSCs were generated using either integrating methods or starting
from T-lymphocytes that have genomic rearrangements thus bearing uncertain consequences
when using iPSC-derived lineages for disease modeling and cell therapies. Recently,
both peripheral blood and cord blood cells have been reprogrammed into transgene-free
iPSC using the Sendai viral vector. Here we demonstrate that peripheral blood can
be utilized for medium-throughput iPSC production without the need to maintain cell
culture prior to reprogramming induction. Cell reprogramming can also be accomplished
with as little as 3000 previously cryopreserved cord blood cells under feeder-free
and chemically defined Xeno-free conditions that are compliant with standard Good
Manufacturing Practice (GMP) regulations. The first iPSC colonies appear 2-3 weeks
faster in comparison to previous reports. Notably, these peripheral blood- and cord
blood-derived iPSCs are free of detectable immunoglobulin heavy chain (IGH) and T
cell receptor (TCR) gene rearrangements, suggesting they did not originate from B-
or T- lymphoid cells. The iPSCs are pluripotent as evaluated by the scorecard assay
and in vitro multi lineage functional cell differentiation. Our data show that small
volumes of cryopreserved peripheral blood or cord blood cells can be reprogrammed
efficiently at a convenient, cost effective and scalable way. In summary, our method
expands the reprogramming potential of limited or archived samples either stored at
blood banks or obtained from pediatric populations that cannot easily provide large
quantities of peripheral blood or a skin biopsy.
Type
Journal articleSubject
Leukocytes, MononuclearPluripotent Stem Cells
Fetal Blood
Humans
Sendai virus
Receptors, Transferrin
Proto-Oncogene Proteins c-myc
Antigens, CD
Cryopreservation
Cell Culture Techniques
Immunohistochemistry
Cell Differentiation
Gene Expression
Cell Lineage
Transgenes
Genetic Vectors
Blood Donors
Octamer Transcription Factor-3
Kruppel-Like Transcription Factors
SOXB1 Transcription Factors
Induced Pluripotent Stem Cells
Cellular Reprogramming
Biomarkers
CD13 Antigens
Kruppel-Like Factor 4
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https://hdl.handle.net/10161/24630Published Version (Please cite this version)
10.1007/s12015-015-9586-8Publication Info
Zhou, Hongyan; Martinez, Hector; Sun, Bruce; Li, Aiqun; Zimmer, Matthew; Katsanis,
Nicholas; ... Chang, Stephen (2015). Rapid and Efficient Generation of Transgene-Free iPSC from a Small Volume of Cryopreserved
Blood. Stem cell reviews and reports, 11(4). pp. 652-665. 10.1007/s12015-015-9586-8. Retrieved from https://hdl.handle.net/10161/24630.This is constructed from limited available data and may be imprecise. To cite this
article, please review & use the official citation provided by the journal.
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Show full item recordScholars@Duke
Erica Ellen Davis
Associate Professor of Pediatrics
Two key questions thematically underscore my research in the Center for Human Disease
Modeling at Duke University: First of all, how can variation at the DNA level be functionally
interpreted beyond the resolution of genetics arguments alone? Secondly, once empowered
with functional information about genetic variants, how can pathogenic alleles be
mapped back to disease phenotypes? Using the ciliary disease module as a model system
of investigation, we are using multidisciplinary tactics to addr
Nicholas Katsanis
Jean and George W. Brumley, Jr., M.D. Professor of Developmental Biology
Joanne Kurtzberg
Jerome S. Harris Distinguished Professor of Pediatrics
Dr. Kurtzberg is an internationally renowned expert in pediatric hematology/oncology,
pediatric blood and marrow transplantation, umbilical cord blood banking and transplantation,
and novel applications of cord blood and birthing tissues in the emerging fields of
cellular therapies and regenerative medicine. Dr. Kurtzberg serves as the Director
of the Marcus Center for Cellular Cures (MC3), Director of the Pediatric Transplant
and Cellular Therapy Program, Director of the Carolina
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