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Normal human peripheral blood mononuclear cells mobilized with granulocyte
colony-stimulating factor have increased osteoclastogenic potential
compared to nonmobilized blood
LE Purton, MY Lee and B Torok-Storb
Program in Transplantation Biology, Clinical Research Division, Fred
Hutchinson Cancer Research Center, Seattle, WA, USA.
Single-cell suspensions of granulocyte colony-stimulating factor (G-
CSF)-mobilized peripheral blood mononuclear cells (G-PBMC) cultured in
alpha minimal essential medium (alphaMEM) containing 10% fetal bovine serum
formed multicellular aggregates within 24 hours. In six separate
experiments, formation of aggregates appeared to be dependent on cell
density per surface area, so that 5.8 +/- 1.3 aggregates formed per 1 x
10(5) cells when G-PBMC were cultured at densities greater than or equal to
1 x 10(5) cells/cm2. The frequency of aggregate formation was less than 1
per 10(5) cells when G-PBMC were cultured at densities less than 1 x 10(5)
cells/cm2. Once formed, aggregates became adherent within 72 hours, and
then, over the course of 21 days, released CD3/CD4/CD25-positive cells into
the supernatant. This T-cell production peaked between days 7 and 14,
reaching a total of 1,269 +/- 125.9 cells released per aggregate by day 21.
Between days 14 and 21, the aggregates also generated macroscopic clusters
of adherent mononuclear and giant multinucleated cells that stained
positive for tartrate-resistant acid phosphatase (TRAP). At 4 weeks, the
macroscopic foci coalesced into monolayers. Multinucleated TRAP-positive
cells were distinguished from macrophage polykaryons by the absence of CD14
expression and the presence of osteoclast-specific membrane receptors for
calcitonin and alphavbeta3-vitronectin. The osteoclast nature of these
cells was further demonstrated by their ability to form resorption lacunae
on dentine slices. Comparable osteoclast formation was not detected in
cultures of normal marrow or normal nonmobilized peripheral blood.
Volume 87,
Issue 5,
pp. 1802-1808,
03/01/1996
Copyright © 1996 by The American Society of Hematology
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