Ct. Jordan et al., LONG-TERM REPOPULATING ABILITIES OF ENRICHED FETAL LIVER STEM-CELLS MEASURED BY COMPETITIVE REPOPULATION, Experimental hematology, 23(9), 1995, pp. 1011-1015
To characterize hematopoietic cell biology, many investigators have us
ed protocols that enrich for primitive hematopoietic stem cells (PHSC)
. In this study, we quantified the long-term repopulating ability (LTR
A) of enriched and discarded fractions of PHSC from day-14 murine feta
l liver using the competitive repopulation assay. We fractionated popu
lations of fetal cells using the antigenic markers AA4.1(+), AA4.1(+)/
Sca(+), and AA4.1(+)/Lin(low)/Sca(+). Differentiating and repopulating
abilities of each of these populations were directly compared using c
ompetitive repopulation. Adult bone marrow was mixed with fetal cell f
ractions from congenic donors having genetically distinguishable marke
rs, and mixtures were given to irradiated recipients. Differentiating
and repopulating abilities of the enriched donor cells were measured b
y the proportions of myeloid and lymphoid cells having donor markers t
hat repopulated the recipients. LTRA was found primarily in the AA4.1(
+) and AA4.1(+)/Sca(+) subpopulations. Further fractionation of the AA
4.1(+) cells to derive an AA4.1(+)/Lin(low)/Sca(+) fraction showed tha
t virtually all of the long-term stem cell activity was found in this
subpopulation. These cells were 1400- to 1600-fold enriched in long-te
rm functional ability compared to fresh marrow. This very high multili
neage repopulating ability per cell was directly measured using a long
-term functional assay in vivo. Importantly, the measured repopulating
ability for AA4.1(+)/Lin(low)/Sca(+) cells was about five-fold less t
han expected from the fraction of cells enriched and remained two- to
three-fold less even after compensating for repopulating ability in di
scarded fractions. This illustrates that long-term functional abilitie
s of enriched PHSC cannot be estimated from fractions enriched but sho
uld be quantitatively assayed.