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HEMATOPOIESIS
From the Sidney Kimmel Cancer Center and Department of
Mathematics, University of California at San Diego, CA; and the
Department of Microbiology and Immunology, University of Miami Medical
School, FL.
Most current theories assume that self-renewal and differentiation
of hematolymphoid stem cells (HSCs) is randomly regulated by intrinsic
and environmental influences. A direct corollary of these tenets is
that self-renewal will continuously generate functionally heterogeneous
daughter HSCs. Decisions about self-renewal versus commitment are made
by individual, single HSCs and, thus, require examination on the clonal
level. We followed the behavior of individual, clonally derived HSCs
through long-term, serial repopulation experiments. These studies
showed that daughter HSCs derived from individual clones were
remarkably similar to each other in the extent and kinetics of
repopulation. Moreover, daughter HSCs within a clone showed equivalent
contributions to the myeloid or lymphoid lineages. Lineage contribution
could be followed because of the discovery of a new subset of HSCs that
gave rise stably to skewed ratios of myeloid and lymphoid cells.
Overall, the data argue that self-renewal does not contribute to the
heterogeneity of the adult HSC compartment. Rather, all HSCs in a clone
follow a predetermined fate, consistent with the generation-age
hypothesis. By extension, this suggests that the self-renewal and
differentiation behavior of HSCs in adult bone marrow is more
predetermined than previously thought.
(Blood. 2002;100:1302-1309) Hematopoietic stem cells (HSCs) replenish all
lineages of mature blood cells through a cascade of differentiation
steps. Differentiation is associated with a loss of self-renewal
capacity, requiring HSCs as a population to self-renew to maintain the
HSC pool. Thus, the ability to commit and to self-renew are defining
properties of HSCs. How the balance of self-renewal and commitment to
differentiation is regulated is incompletely understood.
Most of our knowledge of HSCs derives from population-based studies.
However, commitment and self-renewal decisions are made by single HSCs
and thus require examination on the clonal level. Clonal approaches,
such as injecting limited numbers of purified or retrovirally marked
HSCs, revealed extensive heterogeneity in the HSC
compartment.1-11 HSC heterogeneity can be visualized by
the kinetics of repopulation in transplantation assays: primitive HSCs
need several proliferation and differentiation steps until they give
rise to mature progeny. This causes a delay in the appearance of mature
progeny in the periphery. However, the large clone size generated
through the multiple steps leads to long-term, sustained repopulation.
In contrast, less primitive HSCs require fewer steps, generating small
clones. These less primitive HSCs contribute early to peripheral
hematopoiesis, but their contribution declines over time. Primitive and
less primitive HSCs can be separated to some extent,12,13
demonstrating that these different functions are derived from distinct
subsets of HSCs. When hosts receive mixtures of several types of HSCs,
the kinetics of repopulation will be a combination of the behaviors of
the individual HSCs. This leads to kinetics of repopulation that change
little over time.
Although the functional diversity of the HSC compartment has been
demonstrated unequivocally, it is less clear how such diversity is
generated. The prevailing view is that HSC heterogeneity is regulated
by extrinsic and intrinsic events.14,15 Extrinsic (environmental) signals are derived predominantly from stromal cells
and their products. Stromal cells are organized into niches that differ
in their ability to maintain HSCs.16,17 Thus, homing of
HSCs to different types of stromal cell niches should contribute to HSC
heterogeneity. In addition to extrinsic signals, intrinsic mechanisms
control HSC decisions. Intrinsic mechanisms could induce HSC
heterogeneity if HSCs make random decisions at the time of each HSC
division. For example, each HSC has a choice of many fates, including
self-renewal, differentiation apoptosis, and migration. Evidence for
random processes, termed stochastic, comes from the analysis of
the differentiation of precursors of the myeloid and lymphoid
lineages.18,19 For example, multipotent myeloid precursors
generate more restricted precursors in an apparently random
fashion.18
There is also increasing evidence that intrinsic mechanisms can be
nonrandom but can preprogram the behavior of HSCs. For example, the
size of the HSC compartment and the aging of HSCs are regulated by
mechanisms that are HSC intrinsic and encoded in the germ
line.20-23 Such genetic mechanisms predetermine the behavior of HSCs and thus should limit the generation of HSC heterogeneity.
The hypothesis that HSCs are regulated by random intrinsic or
environmental mechanisms makes a testable prediction, namely that the
generation of HSC heterogeneity is a continual process, and
self-renewal should result in daughter HSCs that are functionally different. In contrast, the idea of predetermined HSC behavior predicts
that daughter HSCs derived from a clone will be similar to each other.
As outlined above, functional diversity is resolvable through the
behavior of clonal HSCs in repopulation experiments.
To test this prediction, an efficient method to examine HSC behavior on
the clonal level was needed. We took advantage of a simple method to
isolate clonally derived HSCs.24 In this system, HSCs,
cultured on the supportive stromal cell line S17, give rise to colonies
of small granulocytic cells. Statistical considerations, verified
experimentally, indicated that 90% of the colonies were derived from a
single initiating HSC. Approximately half of the cultures contained
repopulating HSCs,24 making this a highly efficient method
for enriching repopulating HSCs. We used this system to isolate HSC
clones from bone marrow (BM) and have analyzed the self-renewal and
differentiation of individual HSC clones in long-term transplantation
experiments. The experimental design and the hypothesis tested are
outlined in Figure 1. Unexpectedly, self-renewal of the clonal HSCs generated daughter HSCs that behaved similarly to each other in terms of repopulation kinetics and lineage
contribution. This suggests that HSC behaviors in adult BM are more
predetermined than previously thought. Our data are consistent with the
generation-age hypothesis, an inherently deterministic view of HSC
function.
Clonal HSC cultures
Repopulation assays
All mice were bled in regular intervals, and white blood cells were
prepared as described.16,24,26 Immunofluorescence was used
to measure the percentage of donor-type cells (Ly5.1+) in
the lymphocyte (B220, Thy-1) and myeloid (Gr-1, Mac-1) lineages as
described.16,24,26 Each data point is the mean of 3 measurements of the percentage of Ly5.1+ cells in a blood
sample. For secondary transfers, 5 × 106 BMC from the
primary hosts were injected into lethally irradiated (1062 cGy in 2 doses) B6 mice or B6 mice in which the CD45 locus was
ablated.27 The B6-CD45 Symbolic analysis Symbolic analysis quantifies qualitative aspects of dynamic time series and was used here for a numerical comparison of the kinetics of the repopulation curves of daughter HSCs derived from clonal or multiclonal grafts. First, the slopes for each segment of each curve were calculated. Next, we assigned the plus sign (+) for a segment whose slope was positive and > , the minus sign ( ) for a segment
whose slope was found negative and <, and the approximate sign
(~) for a segment with an absolute slope |s|  .
Here, denotes a cutoff parameter (expressed as percentage). The
size of was set to equal the larger of the 2 standard deviations of
each of the 2 data points that defined a segment of a curve. The result
is that each repopulation curve is described as a string over the
symbols A = {+,,~}. Given
strings
S1 = { 1,...,n}
and
S2 = { 1,...,n} of length n over this alphabet, we defined their matrix of
pairings as M = ((i,
j))1 i,jn. Using the mapping
Ham1(S1,S2) 1
and
Ham1(S1,S2) = 0  S1 = S2.
These properties allow canonical clustering of the elements of any
given set of strings according to their similarity or dissimilarity.
Suppose that
S1 = {+,,~,~,*}, S2 = {+,,+,~,~}.
Then
Statistical analysis Mann-Whitney U nonparametric analysis was performed with Instat (GraphPad Software, San Diego, CA).
Serial transfers show concerted repopulation behavior of HSC clones A hallmark of clonal HSC analysis is the ability to resolve the functional heterogeneity of the HSC compartment. Clonal HSCs, isolated from limiting-dilution cultures, were injected individually into ablated mice. The contribution of each clone to peripheral hematopoiesis was followed by staining white blood cells for the donor-type allele of the CD45/Ly5 antigen. The data in Figure 2 demonstrate extensive heterogeneity among the clonally derived HSCs with respect to the onset, duration, and extent of repopulation.
The heterogeneity of HSC behaviors seen in the primary hosts is
consistent with the view that random and environmental signals constantly generated functionally diverse HSCs. If this was a general
principle, then the daughter HSCs derived from each of the HSC clones
should exhibit additional heterogeneous repopulation behavior, and HSCs
of different primitiveness would be expected in the progeny of a single
HSC clone. To test this, we performed serial BM transplantation.
Overall, 13 individual HSC clones were tested in secondary transplants
(Table 1).
Of the 13 clones tested, 8 clones generated daughter HSCs in the
primary host that repopulated secondary host(s). Of these 8 clones,
daughter HSCs from 7 clones were injected into at least 2 secondary
hosts, and these 7 clones are depicted in Figure
3. For comparison, we also show clone
r27.B3, a clone that failed to generate daughter HSCs in the primary
host. Clonal heterogeneity in the repopulation patterns seen in the
primary hosts (Figure 2) was evident also in the secondary hosts
(Figure 3). For example, clone B2 showed high levels, though clone B4
had only moderate levels, of secondary repopulation (Figure 3D). In
contrast to this inter-clonal heterogeneity, daughter HSCs derived from
any given clone behaved remarkably similarly to each other. For
example, the daughter HSCs derived from clone B2 showed repopulation
kinetics so similar to each other that 3 of 4 of the data points from
the pairs of secondary hosts overlap (Figure 3D). Similarly, the
kinetics of repopulation in the 4 secondary recipients of clone r38.2
daughter HSCs were almost indistinguishable (Figure 3C). Other clones, such as C2 or r16.4 (Figure 3C,D), varied in the extent of
repopulation. However, even for these clones the kinetics with which
donor-type cells were generated were similar. For example, HSCs derived
from clone r16.3 were injected into 3 secondary recipients. Donor-type contribution in all 3 hosts first increased, then sharply decreased, at
the same time. HSCs derived originally from clones B2 and C2 were
tested in tertiary transfers, and the similarity in repopulation patterns became even more pronounced in these recipients (Figure 4). All recipients of daughter HSCs,
derived from clone B2, showed high levels of donor-type cells, and the
kinetics of repopulation was similar in the 3 mice we followed for 7 months. HSCs from clone C2 failed to repopulate any of the 4 tertiary
recipients (Figure 4), indicating that all HSCs in clone C2 had ceased
to self-renew in both secondary hosts.
As a control for the primary repopulation experiments, we had originally injected some W41W41 hosts with 105 freshly explanted BMCs (Figure 2). This dose of BM contains on average 5 HSCs defined as W41W41 repopulating units.29 BMCs from these control mice were transplanted into secondary hosts in parallel to the clonally repopulated mice. Multiclonal HSC grafts showed comparable levels of repopulation in the secondary hosts. However, HSCs derived from the control mice varied noticeably more in their kinetics of repopulation than the clonally derived HSCs (Figure 3A). For example, in one secondary host, HSCs derived from the BM2 mouse showed steadily declining levels of donor-type cells. However, the other host showed an initial decrease, followed by an increase of donor-type cells. HSCs from the BM1 mouse appeared more similar in the secondary hosts. However, in the tertiary hosts, variation in the repopulation kinetics became pronounced (Figure 4). We also tested HSCs that had been cultured as mixed populations (bulk) on the stromal cell line S17 as described.16,26 Primary recipients were injected with cultured cells (equivalent of 2 × 105 cells originally seeded). As reported previously, HSCs cultured on the stromal cell line S17 are qualitatively and quantitatively similar to freshly explanted BM HSCs.16,26 After 8 months, BMCs from 2 of these primary mice (derived from independent experiments and designated bulk1 and bulk2) were injected into 3 and 4 secondary recipients, respectively. The repopulation kinetics of HSCs derived from the bulk grafts were diverse (Figure 3B), indicating that the concerted repopulation behaviors of the clonal HSCs were not caused by culture on line S17. Rather, the data argue that the distinct kinetics of the control HSCs reflect the original heterogeneity of the multiclonal graft. Overall, the data show that clonally, but not multiclonally, derived daughter HSCs give rise to daughter HSCs with strikingly similar repopulation behaviors. Although the similarities in the repopulation curves for daughter HSCs
from some clones (eg, r38.2, C2) are intuitively obvious, the patterns
of other curves are less conspicuous. To quantify the similarities and
differences in the repopulation kinetics of daughter HSCs from each
clone, we performed symbolic analysis. Secondary repopulation data were
transformed into symbol strings according to the slope of each segment
of each of the curves. Next, we determined similarity and dissimilarity
between pairs of symbolized kinetics using normalized Hamming
distances.28 This determines the number of distinct symbol
pairs between 2 strings and, when divided by the length of the strings,
will yield a probability measure of how different 2 strings are. A
normalized Hamming distance of 0 means that 2 strings are completely
similar, whereas a Hamming distance of 1 means complete dissimilarity. When more than 2 secondary hosts were analyzed per clone, pairs of each
permutation were compared (Figure 5).
Symbolic analysis of the secondary recipients shows (Figure 5A) that
the kinetics of daughter HSCs derived from multiclonal grafts differ in
shape, with a mean relative Hamming distance of 0.7. Parenthetically, a
Hamming distance of 0.66 is found when randomly simulated data are
examined (H.B.S., manuscript in preparation), indicating that the kinetics of repopulation by HSCs derived from multiclonal grafts
are not related.
One secondary host of daughter HSCs derived from the clone 7of9 also showed divergent kinetics of repopulation when compared with the other 3 recipients in that group. However, the other curves in the 7of9 group showed low Hamming scores, indicating that their shapes were similar by the criteria used. Low Hamming distances were also derived from the comparison of daughter HSCs derived from all other clones tested (Figure 5). The Hamming distances derived from clonal and multiclonal daughter HSCs, respectively, were statistically highly significant (P = .001). In the tertiary recipients, the kinetics of daughter HSCs from clone B2 generated Hamming distances of 0.33 and 0.0, respectively. A mean Hamming distance of 0.7 was found when the kinetics of daughter HSCs from the multiclonal BM1 graft were compared. The comparison is not quite statistically significant (P = .052), probably because of the low number of points in this analysis. Overall, symbolic analysis confirms the interpretation that the kinetics of repopulation of clonal daughter HSCs are more similar to each other than are daughter HSCs from multiclonal grafts. Clonal analysis reveals lineage-dominant HSCs All clones examined here gave rise to myeloid and lymphoid cells. Clonal analysis revealed an additional unique feature a subpopulation
of HSCs that gave rise to unusual ratios of myeloid and lymphoid cells.
To define the typical or balanced ratios of myeloid to lymphoid cells,
data from 50 unmanipulated B6 mice were analyzed. A mean value of
14.1% ± 3.9% myeloid cells in blood was obtained. B and T
lymphocytes make up the remaining portion of white blood cells. The
mean ratio of lymphoid-to-myeloid cells in these mice is 6.0 ± 2.0
(95% confidence level, 0.57). This pattern of lymphoid-to-myeloid cells we call balanced. A comparable ratio of myeloid-to-lymphoid cells
is obtained in stably repopulated mice that received multiclonal BM
grafts (Figure 6). HSC clones that
contributed to balanced ratios of myeloid and lymphoid cells were
found. Clones A3 and r38.2 (Figure 6) are examples of such balanced
lineage repopulation. However, a subset of the clones differed in their
contribution to the myeloid and lymphoid lineages. For example, clones
r16.4, B2, B4, and 7of9 generated low ratios (
Among 38 repopulating HSC clones in primary recipients, we found 4 myeloid-dominant HSCs in 3 independent experiments. These clones had
self-renewal capacity, as judged by their ability to generate daughter
HSCs that repopulated secondary hosts (Figure 3). At first glance, the
incidence of lymphoid-dominant HSCs appears to be higher at almost 60%
of the clones in primary recipients. However, the enumeration of
lymphoid-dominant HSCs is equivocal because long-lived lymphocytes can
dominate the periphery when an HSC ceases to contribute to
repopulation. Indeed, most of the primary HSC clones with low levels of
donor-type repopulation appear as lymphoid dominant, and 3 of 3 such
clones failed to repopulate secondary hosts, indicting stem cell
failure (Table 1 and data not shown). We also tested 3 lymphoid-dominant clones with high levels of donor-type levels (more
than 40%). Of these, one clone (r27.B3) failed to repopulate secondary
recipients (Figure 3). However, 2 clones (C2 and r16.3) repopulated
secondary recipients and retained lymphoid dominance (Figures 3,
7). This suggests that lymphoid-dominant
HSC clones are also infrequent.
Lineage dominance is stable, HSC-intrinsic behavior In HSC clones that self-renewed, lineage dominance persisted through serial transfers (Figures 6-8). For example, clones C2 and r16.3 were lymphoid dominant in the primary and the secondary hosts. Similarly, clones B2, B4, and r16.4 gave rise to myeloid-dominant progeny in the primary, secondary, and tertiary hosts (Figures 6-8). Unfortunately, lineage data for the secondary transplant of clone 7of9 were made uninterpretable by an ear mite infection (or its treatment) that caused myelopenia. Overall, the data indicate that lineage dominance is a stable property of HSCs. Similar to balanced HSCs, lineage-dominant HSC clones generated daughter HSCs with concerted repopulation kinetics (Figure 3). The rate of appearance of myeloid and lymphoid cells in blood following transplantation appears to be more similar for the daughter HSCs within each clone in secondary, and particularly in tertiary, recipients than for the multiclonal grafts (Figures 7, 8). In contrast, the secondary and tertiary recipients of multiclonally repopulated control mice showed diverse repopulation kinetics and diverse lineage contributions. For example, in tertiary mice that received HSCs from the BM1 control mouse, one secondary host appeared as balanced and the other 2 as myeloid dominant (Figure 8). To quantify this observation, we performed symbolic analysis of the kinetics with which myeloid cells were generated (Figure 9). The Hamming distances of clonal and multiclonal grafts were statistically significantly different, with P = .0276 in secondary hosts. In tertiary animals the differences in the Hamming distance approaches (but does not attain) significance with P = .0581. This is probably because of the low number of points examined here. Overall, the data support the conclusion that clonally derived daughter HSCs behaved more alike in repopulation kinetics and lineage contribution than HSCs derived from multiclonal grafts.
One possible interpretation for lineage dominance could be that these
HSCs are leukemic. However, this seems to be unlikely for the following
reasons: the hosts had normal-sized, and -formed organs (spleen, BM,
thymus, heart, liver, kidney), revealing no overt signs of leukemia or
hyper-proliferative diseases even after 3 rounds of transplantation.
The ability to respond to low concentrations of cytokines is a hallmark
of some types of leukemia. We found that the myeloid progeny of the
myeloid-dominant HSC clone B2 required the same concentration of GM-CSF
for proliferation, as do cells from unmanipulated control BM (data not
shown). Composition of the T-cell-receptor repertoire can be
indicative of abnormalities in the hematopoietic system.30
Judged by the representation of T-cell-receptor v
A clonal transplantation system was used to examine the generation of HSC heterogeneity. Our data demonstrate an unexpected similarity among the daughter HSCs of clonally derived HSCs. The similarity encompassed all HSC behaviors measured: self-renewal, primitiveness, and lineage contribution. Stability of the lineage contribution was especially evident in the lineage-dominant HSCs. These data suggest that HSC behavior in adult bone marrow is largely predetermined. Daughter HSCs from multiclonal grafts showed more heterogeneous behavior in the kinetics of repopulation and in the lineage contribution. The most likely explanation for these observations is that HSC behavior is preprogrammed and that self-renewal does not induce HSC heterogeneity. The heterogeneity of the HSCs derived from multiclonal grafts is likely a reflection of the heterogeneous clonal composition of the mixture of HSCs, initially transplanted into the control primary host. The clonal transplantation system that was used here readily distinguished different types of HSCs. Those HSCs that had self-renewed in the primary host generated an unknown number of daughter HSCs. If an HSC had self-renewed extensively, the secondary grafts could have contained many HSCs. This would lead to a composite, and thus level, repopulation kinetics, comparable to that generated by the multiclonal BM grafts in the primary recipients. Repopulation kinetics of daughter HSCs derived from clone 7of9 are consistent (though not conclusively so) with such a scenario. However, daughter HSCs derived from other HSC clones repopulated poorly or transiently, consistent with a limited number of HSCs in the secondary grafts. In this situation, HSC heterogeneity should have been discernible in the secondary recipients. Indeed, heterogeneity was seen in the daughter HSCs derived from multiclonal grafts. This supports the interpretation that the secondary grafts (of most clonal and multiclonal HSC transplants) contained sufficiently low numbers of HSCs to readily resolve different types of HSCs in the grafts. This is supported by the observation that lineage dominance was observable through serial transplantation. Because the frequency of lineage-dominant HSCs is low, a multiclonal HSC graft should have obscured the lineage dominance. The interpretation that HSCs were limiting in the serial transplants is consistent with published studies5,31,32 showing that the HSC compartment is not fully regenerated after transplantation. Overall, the data indicate that daughter HSCs from the clonally derived HSCs behaved similarly to each other, supporting the view that the HSC behaviors measured here are preprogrammed. The similarity in the behaviors of the daughter HSCs is further emphasized by the synchronous extinction of several clones in secondary or tertiary recipients. Concerted cessation of clone r16.3 in the secondary mice suggests that clonal extinction is unlikely caused solely by the transplantation procedure. Collectively, the data suggest that the lifespan of each HSC clone is predetermined and that clonal extinction results from a concurrent cessation of the self-renewal of all HSCs within a clone. Consistent with a predetermined lifespan is the observation that most clones showed less primitive repopulation kinetics in the secondary than in the primary hosts. This suggests that within a clone all daughter HSCs progress synchronously from a primitive to a less primitive state. This agrees with the central tenet of the generation-age hypothesis33 that HSCs age and lose primitiveness with each proliferation step. That the history of a cell determines its behavior is an inherently deterministic view. The HSCs examined here self-renewed and differentiated in vivo, in a complete and physiological environment. We did not examine whether HSC programs can be changed by extrinsic signals. However, on transplantation, HSCs should have had access to functionally different niches. Our data suggest that this does not contribute to the HSC heterogeneity. However, it is possible that each HSC clone is preprogrammed to express a distinct set of homing molecules, directing all HSCs within a clone to the same type of niche. Subsets of HSCs with distinct patterns of homing receptors have been isolated,34 supporting the plausibility of this interpretation. Alternatively, each niche could be composed of several types of stromal cells. This is consistent with the structure of the hematon, an aggregate isolated from BM, thought to represent in situ niches.17 Such mixed niches would be functionally fairly homogeneous and would be permissive for the deterministic programs of different types of HSC. In either case, HSCs would be dependent on stromal niches for the execution of their programs. Although the daughter HSCs within each clone behaved similarly, different HSC clones showed clearly distinguishable repopulation patterns. Thus, the HSC compartment in adult BM is heterogeneous. Our data indicate that this heterogeneity is not generated continuously. This suggests that stem cell heterogeneity is generated earlier in development, perhaps when stem cells seed to the BM during development. Alternatively, as predicted by the generation-age hypothesis, HSC heterogeneity may be generated by the preprogrammed aging of HSCs. At the time of analysis, a snapshot of the current state of the HSC compartment would then reveal HSCs at different stages in the aging process. Another unique feature of clonal analysis was the identification of infrequent lineage-dominant HSCs. In the regenerating hematopoietic system, the appearance of mature cells in blood progresses through a reproducible sequence.5,6 Early after transplantation, myeloid cells are overrepresented in the periphery, followed by bursts of B and then of T lymphocytes. The sequence with which these cells appear in the periphery reflects the time it takes for these lineages to mature. Lineage-dominant repopulation patterns, similar to those described here, were noted in the analysis of retrovirally marked HSCs.4 Although a population of HSCs that generated stably and long-term skewed ratios of myeloid and lymphoid cells was found,4 the data were interpreted to represent the dynamic changes of lineage contribution after transplantation. We followed the repopulation behavior of lineage-dominant HSCs through serial transplantations, and the data demonstrate that lineage dominance is a stable behavior of a subset of HSCs. Many of the lineage-dominant HSCs had self-renewal capacity, indicating that lineage dominance is not associated with a loss of primitiveness. Because loss of self-renewal capacity is a hallmark of lineage commitment, lineage dominance is unlikely to be a reflection of lineage commitment. Rather, lineage-dominant HSCs are a normal subset of stem cells with distinct developmental potential. Myeloid-dominant HSCs have been attributed to the aging of the HSC compartment.35 We and others4,5 demonstrated lineage-dominant HSCs through clonal analysis in young mice. Thus, myeloid dominance is not associated with the pathology of aging. Perhaps a selective loss of lymphoid-dominant stem cells could account for the imbalance seen in the aged animals. The stability of the behavior of the adult HSC points toward epigenetic effects that may determine the developmental potential of each HSC. Epigenetic events, such as stable changes in chromatin and DNA methylation, cause long-lasting changes in gene expression. The generation of all tissue-specific stem cells from embryonic stem cells likely involves epigenetic changes that ensure a stable supply of functionally restricted stem cells.36,37 Such changes also could cause modifications in the expression of genes required for the optimal establishment of precursors or mature cells in the underrepresented lineage and could thereby account for stable lineage dominance. Our results open the possibility that distinct subsets of HSCs will be optimal for different applications. For example, lineage-dominant stem cells could be used to selectively remedy lineage-specific cytopenias. Moreover, HSC heterogeneity, fixed through epigenetic mechanisms, may well account for the heterogeneity seen in stem-cell leukemias.38 Characterizing the mechanisms of lineage dominance may not only be biologically important, it may have therapeutic usefulness.
We thank Drs P. Linton, C. Bonifer, and J. Phillips for valuable discussions and suggestions.
Submitted September 17, 2001; accepted April 8, 2002.
Supported by National Institutes of Health grants DK48015, AG09948, and AG17564.
The publication costs of this article were defrayed in part by page charge payment. Therefore, and solely to indicate this fact, this article is hereby marked "advertisement" in accordance with 18 U.S.C. section 1734.
Reprints: Christa E. Müller-Sieburg, Sidney Kimmel Cancer Center, 10835 Altman Row, San Diego, CA 92121; e-mail: cmuller{at}skcc.org.
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  I. Roeder, K. Horn, H.-B. Sieburg, R. Cho, C. Muller-Sieburg, and M. Loeffler Characterization and quantification of clonal heterogeneity among hematopoietic stem cells: a model-based approach Blood, December 15, 2008; 112(13): 4874 - 4883. [Abstract] [Full Text] [PDF]
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D. G. Kent, B. J. Dykstra, J. Cheyne, E. Ma, and C. J. Eaves Steel factor coordinately regulates the molecular signature and biologic function of hematopoietic stem cells Blood, August 1, 2008; 112(3): 560 - 567. [Abstract] [Full Text] [PDF]
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R. H. Cho, H. B. Sieburg, and C. E. Muller-Sieburg A new mechanism for the aging of hematopoietic stem cells: aging changes the clonal composition of the stem cell compartment but not individual stem cells Blood, June 15, 2008; 111(12): 5553 - 5561. [Abstract] [Full Text] [PDF]
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D. C. Weksberg, S. M. Chambers, N. C. Boles, and M. A. Goodell CD150- side population cells represent a functionally distinct population of long-term hematopoietic stem cells Blood, February 15, 2008; 111(4): 2444 - 2451. [Abstract] [Full Text] [PDF]
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M. A. Kirkland, P. J. Quesenberry, I. Roeder, H. Sieburg, R. Cho, B. Dykstra, N. Uchida, C. Eaves, and C. Muller-Sieburg Discrete stem cells: subsets or a continuum? Blood, December 15, 2006; 108(12): 3949 - 3950. [Full Text] [PDF]
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H. B. Sieburg, R. H. Cho, B. Dykstra, N. Uchida, C. J. Eaves, and C. E. Muller-Sieburg The hematopoietic stem compartment consists of a limited number of discrete stem cell subsets Blood, March 15, 2006; 107(6): 2311 - 2316. [Abstract] [Full Text] [PDF]
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R. Kumar, J. C. Langer, and H.-W. Snoeck Transforming growth factor-beta2 is involved in quantitative genetic variation in thymic involution Blood, March 1, 2006; 107(5): 1974 - 1979. [Abstract] [Full Text] [PDF]
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E. F. Srour, X. Tong, K. W. Sung, P. A. Plett, S. Rice, J. Daggy, C. T. Yiannoutsos, R. Abonour, and C. M. Orschell Modulation of in vitro proliferation kinetics and primitive hematopoietic potential of individual human CD34+CD38-/lo cells in G0 Blood, April 15, 2005; 105(8): 3109 - 3116. [Abstract] [Full Text] [PDF]
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I. Roeder, L. M. Kamminga, K. Braesel, B. Dontje, G. de Haan, and M. Loeffler Competitive clonal hematopoiesis in mouse chimeras explained by a stochastic model of stem cell organization Blood, January 15, 2005; 105(2): 609 - 616. [Abstract] [Full Text] [PDF]
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 E. Henckaerts, J. C. Langer, J. Orenstein, and H.-W. Snoeck The Positive Regulatory Effect of TGF-{beta}2 on Primitive Murine Hemopoietic Stem and Progenitor Cells Is Dependent on Age, Genetic Background, and Serum Factors J. Immunol., August 15, 2004; 173(4): 2486 - 2493. [Abstract] [Full Text] [PDF]
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C. E. Muller-Sieburg, R. H. Cho, L. Karlsson, J.-F. Huang, and H. B. Sieburg Myeloid-biased hematopoietic stem cells have extensive self-renewal capacity but generate diminished lymphoid progeny with impaired IL-7 responsiveness Blood, June 1, 2004; 103(11): 4111 - 4118. [Abstract] [Full Text] [PDF]
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C. D. Helgason, J. Antonchuk, C. Bodner, and R. K. Humphries Homeostasis and regeneration of the hematopoietic stem cell pool are altered in SHIP-deficient mice Blood, November 15, 2003; 102(10): 3541 - 3547. [Abstract] [Full Text] [PDF]
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Abstract
Introduction
/
10) of lymphoid-to-myeloid cells, appearing as
lymphoid dominant (Figure 
