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Blood, 15 April 2001, Vol. 97, No. 8, pp. 2257-2268
HEMATOPOIESIS
Resolution of pluripotential intermediates in murine
hematopoietic differentiation by global complementary DNA amplification
from single cells: confirmation of assignments by expression profiling
of cytokine receptor transcripts
Filio Billia,
Mary Barbara,
Jon McEwen,
Maryanne Trevisan, and
Norman
N. Iscove
From the Ontario Cancer Institute and the Department of
Medical Biophysics, University of Toronto, Ontario, Canada.
Although hematopoiesis is known to proceed from stem cells through
a graded series of multipotent, oligopotent, and unipotent precursor
cells, it has been difficult to resolve these cells physically
one from another. There is, therefore, corresponding uncertainty about
the exact distribution and timing of the expression of genes known to
be important in hematopoietic differentiation. In earlier work, the
generation of a set of amplified complementary DNAs (cDNAs) from
single precursor cells was described, whose biologic potential
was determined by the outcome of cultured sibling cells. In this study,
the new acquisition of cDNA from multipotent myeloid precursor cells is
described, as is the mapping of RNA-level expression of 17 distinct
cytokine receptors (c-kit, Flk-1, Flk-2/Flt-3, c-fms,
gp130, erythropoietin receptor, GM-CSFR , G-CSFR, TNFR1, IL-1RI,
IL-1RII, IL-2R , IL-3-specific receptor, IL-4R, IL-6R, IL-7R, and IL-11R) to the enlarged sample set, spanning stages from pentapotent precursors through oligopotent intermediates to
committed and maturing cells in the myeloid and lymphoid lineages. Although the enhanced scope and resolving power of the analysis yielded
previously unreported observations, there was overall agreement with
known biologic responsiveness at individual stages, and major
contradictions did not arise. Moreover, each precursor category
displayed a unique overall pattern of hybridization to the matrix of 17 receptor probes, supporting the notion that each sample pool indeed
reflected a unique precursor stage. Collectively, the results provide
supportive evidence for the validity of the cDNA assignments to
particular stages, the depth of the information captured,
and the unique capacity of the sample matrix to resolve individual stages in the hematopoietic hierarchy.
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