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Blood, 15 December 2000, Vol. 96, No. 13, pp. 4335-4343
PHAGOCYTES
An adherent condition is required for formation of multinuclear
osteoclasts in the presence of macrophage colony-stimulating factor
and receptor activator of nuclear factor  B ligand
Takeshi Miyamoto,
Fumio Arai,
Osamu Ohneda,
Katsumasa Takagi,
Dirk M. Anderson, and
Toshio Suda
From the Department of Cell Differentiation, Institute
of Molecular Embryology and Genetics, and the Department of Orthopedic
Surgery, Kumamoto University School of Medicine, Kumamoto, Japan; and
the Department of Molecular Biology, Immunex Corporation, Seattle, WA.
Identification of receptor activator of nuclear factor- B
(RANK) and RANK-ligand (RANKL) has provided new insights into the osteoclast differentiation pathway. Osteoclast precursor cells were isolated using monoclonal antibodies against c-Fms and RANK, and
the effect of adherence on the in vitro differentiation and proliferation of these cells was examined in 2 different types of
stromal-cell-free culture systems: a semisolid culture medium (a
nonadherent system) and a liquid culture medium (an adherent system).
Osteoclast precursor cells were not able to differentiate into mature
osteoclasts efficiently in the semisolid culture system. Trimerized
RANKL enhanced osteoclast differentiation in semisolid cultures, but
not to the extent seen when cells were allowed to adhere to plastic.
Initial precursor cells were capable of differentiating into
macrophages or osteoclasts. Once these cells were transferred to
adherent conditions, striking differentiation was induced. Multinuclear
cells were observed even after they had displayed phagocytic activity,
which suggests that cell adhesion plays an important role in the
differentiation of osteoclast precursor cells. Integrins, especially
the arginine-glycine-aspartic acid (RGD)-recognizing integrins
 v and  3, were needed for
osteoclast-committed precursor cells to proliferate in order to form
multinuclear osteoclasts, and the increase in cell density affected the
formation of multinuclear cells. A model of osteoclast differentiation
with 2 stages of precursor development is proposed: (1) a
first stage, in which precursor cells are bipotential and
capable of anchorage-independent growth, and (2) a second stage, in
which the further proliferation and differentiation of
osteoclast-committed precursor cells is anchorage-dependent.
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