8 research outputs found
Coexpression of Kit and the receptors for erythropoietin, interleukin 6 and GM-CSF on hemopoietic cells
The detection of functional growth factor (GF) receptors on subpopulations
of hemopoietic cells may provide a further dissection of immature cell
subsets. Since little information is available about coexpression of
different GF receptors at the level of single hemopoietic cells, we
studied the feasibility of simultaneous cell staining with a combination
of biotin- and digoxigenin-labeled GFs for flow cytometric detection of
functional receptors. Using this methodology, coexpression of Kit and
receptors for erythropoietin (EPO), interleukin 6 (IL-6), and GM-CSF on
hemopoietic cells was studied by triple-staining of rhesus monkey bone
marrow (BM) cells with labeled GFs and antibodies against other cell
surface markers. Most of the immature, CD34+2 cells were Kit+ but did not
display detectable levels of EPO-receptors (EPO-Rs) or GM-CSF-R.
Approximately 60% of these CD34+2/Kit+ cells coexpressed the IL-6-R,
demonstrating that immature cells are heterogeneous with respect to IL-6-R
expression. Maturation of monomyeloid progenitors, as demonstrated by
decreasing CD34 and increasing CD11b expression, is accompanied by a
decline of Kit and an increase in GM-CSF-R expression in such a way that
Kit+/GM-CSF-R+ cells are hardly detectable. IL-6-R expression is
maintained or even increased during monomyeloid differentiation. IL-6-R
and GM-CSF-R were not identified on most CD71+2 cells, which indicated
that these receptors are probably not expressed during erythroid
differentiation. Together with previous results, our data show that both
Kit and CD71 are upregulated with erythroid commitment of immature
progenitors. Upon further differentiation, Kit+/EPO-R-cells lose CD34 and
acquire EPO-R. Maturing erythroid cells eventually lose CD71 and Kit
expression but retain the EPO-R. In conclusion, this approach enables
further characterization of the specificity of GFs for different bone
marrow subpopulations. Apart from insight into the differentiation stages
on which individual GFs may act, information about receptor coexpression
may be used to identify individual cells that can respond to multiple GFs,
and allows for further characterization of the regulation of
lineage-specific differentiation
Facilitated engraftment of human hematopoietic cells in severe combined immunodeficient mice following a single injection of Cl²MDP liposomes
Transplantation of normal and malignant human hematopoietic cells into severe combined immunodeficient (SCID) mice allows for evaluation of long-term growth abilities of these cells and provides a preclinical model for therapeutic interventions. However, large numbers of cells are required for successful engraftment in preirradiated mice due to residual graft resistance, that may be mediated by cells from the mononuclear phagocytic system. Intravenous (i.v.) injection of liposomes containing dichloromethylene diphosphonate (Cl2MDP) may eliminate mouse macrophages in spleen and liver. In this study outgrowth of acute myeloid leukemia (AML) cells and umbilical cord blood (UCB) cells in SCID mice conditioned with a single i.v. injection of Cl2MDP liposomes in addition to sublethal total body irradiation (TBI) was compared to outgrowth of these cells in SCID mice that had received TBI alone. A two- to 10-fold increase in outgrowth of AML cells was observed in four cases of AML. Administration of 107 UCB cells reproducibly engrafted SCID mice that had been conditioned with Cl2MDP liposomes and TBI, whereas human cells were not detected in mice conditioned with TBI alone. As few as 2 x 104 purified CD34+ UCB cells engrafted in all mice treated with Cl2MDP liposomes. In SCID mice treated with macrophage depletion unexpected graft failures were not observed. Histological examination of the spleen showed that TBI and Cl2MDP liposomes i.v. resulted in a transient elimination of all macrophage subsets in the spleen, whereas TBI had a minor effect. Cl2MDP liposomes were easy to use and their application was not associated with appreciable side-effects. Cl2MDP liposome pretreatment in combination with TBI allows for reproducible outgrowth of high numbers of human hematopoietic cells in SCID mice
The efficacy of recombinant thrombopoietin in murine and nonhuman primate models for radiation-induced myelosuppression and stem cell transplantation
Radiation-induced pancytopenia proved to be a suitable model system in
mice and rhesus monkeys for studying thrombopoietin (TPO) target cell
range and efficacy. TPO was highly effective in rhesus monkeys exposed to
the mid-lethal dose of 5 Gy (300 kV x-rays) TBI, a model in which it
alleviated thrombocytopenia, promoted red cell reconstitution, accelerated
reconstitution of immature CD34+ bone marrow cells, and potentiated the
response to growth factors such as GM-CSF and G-CSF. In contrast to the
results in the 5 Gy TBI model, TPO was ineffective following
transplantation of limited numbers of autologous bone marrow or highly
purified stem cells in monkeys conditioned with 8 Gy TBI. In the 5 Gy
model, a single dose of TPO augmented by GM-CSF 24 h after TBI was
effective in preventing thrombocytopenia. The strong erythropoietic
stimulation may result in iron depletion, and TPO treatment should be
accompanied by monitoring of iron status. This preclinical evaluation thus
identified TPO as a potential major therapeutic agent for counteracting
radiation-induced pancytopenia and demonstrated pronounced stimulatory
effects on the reconstitution of immature CD34+ hemopoietic cells with
multilineage potential. The latter observation explains the potentiation
of the hematopoietic responses to G-CSF and GM-CSF when administered
concomitantly. It also predicts the effective use of TPO to accelerate
reconstitution of immature hematopoietic cells as well as possible
synergistic effects in vivo with various other growth factors acting on
immature stem cells and their direct lineage-committed progeny. The
finding that a single dose of TPO might be sufficient for a clinically
significant response emphasizes its potency and is of practical relevance.
The heterogeneity of the TPO response encountered in the various models
used for evaluation points to multiple mechanisms operating on the TPO
response and heterogeneity of its target cells. Mechanistic mouse studies
made apparent that the response of multilineage cells shortly after TBI to
a single administration of TPO is quantitatively more important for
optimal efficacy than the lineage-restricted response obtained at later
intervals after TBI and emphasized the importance of a relatively high
dose of TPO to overcome initial c-mpl-mediated clearance. Further
elucidation of mechanisms determining efficacy might very well result in a
further improvement, e.g., following transplantation of limited numbers of
stem cells. Adverse effects of TPO administration to myelosuppressed or
stem cell transplanted experimental animals were not observed