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Identification of Philadelphia-negative granulocyte-macrophage colony-
forming units generated by stroma-adherent cells from chronic myelogenous
leukemia patients
C Carlo-Stella, L Mangoni, G Piovani, D Garau, C Almici and V Rizzoli
Department of Hematology, University of Parma, Italy.
Chronic myelogenous leukemia (CML) is a clonal disorder of the
hematopoietic stem cell characterized by the coexistence of
Philadelphia-negative (Ph-) with Ph+ progenitors. CML progenitor cells have
been shown to be defective in adherence to marrow stroma. The present study
investigated at the cytogenetic level marrow-derived CML clonogenic cells
generated from the stroma-adherent cell fraction. On direct cytogenetic
analysis, the overall mean (+/- SEM) percentage of Ph- metaphases was 3%
+/- 1%. Mononuclear marrow cells from CML patients (n = 18) were incubated
with mafosfamide (100 micrograms/mL) or control medium, seeded onto marrow
stromal layers and allowed to adhere (2 hours, 37 degrees C). After a
short-term (3-day) liquid culture, the cells were harvested, incorporated
in methyl-cellulose, and individual colonies were analyzed by single colony
karyotyping. The mean (+/- SEM) percentage of Ph- colonies generated from
the stroma- adherent fraction was 35% +/- 6%. As compared with marrow
colony- forming unit granulocyte-macrophage plated before any manipulation,
the mean (+/- SEM) percentage of Ph- clones was significantly increased by
stroma adherence (35% +/- 6% v 15% +/- 4%, P < or = .005) and
mafosfamide (100 micrograms/mL) incubation of marrow cells before stroma
adherence (58% +/- 9% v 35% +/- 6%, P < or = .005). An additive effect
was observed by combining mafosfamide treatment and stroma adherence.
Single-colony transfer experiments showed that 50% +/- 4% stroma-adherent
and 70% +/- 4% stroma-adherent mafosfamide-treated progenitors gave rise to
secondary colonies. To further characterize the stroma-adherent fraction,
experiments were performed in which CD34+ marrow cells were used. The mean
(+/- SEM) output of progenitors generated by 10,000 CD34+, stroma-adherent
cells was 888 +/- 188 and 570 +/- 258 for untreated and mafosfamide-treated
cells, respectively. Individual colonies were analyzed by single-colony
karyotyping and fluorescent in situ hybridization using a biotinylated
cosmid DNA probe that hybridize to abl oncogene. The CD34+, stroma-adherent
fraction contained 38% +/- 14% (untreated) and 56% +/- 18%
(mafosfamide-treated) (P < or = .025) Ph- progenitors. In conclusion,
the present data show the possibility to select Ph- clones that (1) have a
maintained capability of stroma adherence, (2) are mafosfamide resistant,
(3) are derived from the CD34+ fraction, and (4) have high-replating
potential.
Volume 83,
Issue 5,
pp. 1373-1380,
03/01/1994
Copyright © 1994 by The American Society of Hematology
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This article has been cited by other articles:
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C. Carlo-Stella, E. Regazzi, G. Sammarelli, S. Colla, D. Garau, A. Gazit, B. Savoldo, D. Cilloni, A. Tabilio, A. Levitzki, et al.
Effects of the Tyrosine Kinase Inhibitor AG957 and an Anti-Fas Receptor Antibody on CD34+ Chronic Myelogenous Leukemia Progenitor Cells
Blood,
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C. Cesana, C. Carlo-Stella, L. Mangoni, E. Regazzi, D. Garau, G. Sammarelli, C. Caramatti, C. Almici, and V. Rizzoli
In Vitro Growth of Mobilized Peripheral Blood Progenitor Cells is Significantly Enhanced by Stem Cell Factor
Stem Cells,
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