SASAKI Takahisa Showa Univ., School of Dentistry Associate Professor, 歯学部, 助教授 (50129839)
TANAKA Hirofumi Showa Univ., School of Dentistry Assistant, 歯学部, 助手 (30146899)
SUDA Tatsuo Showa Univ., School of Dentistry Professor, 歯学部, 教授 (90014034)
|Budget Amount *help
¥5,800,000 (Direct Cost: ¥5,800,000)
Fiscal Year 1990: ¥1,900,000 (Direct Cost: ¥1,900,000)
Fiscal Year 1989: ¥3,900,000 (Direct Cost: ¥3,900,000)
(1) Studies on stromal cell lines which support osteoclast formation in vitro
We previously reported that osteoclast-like multinucleated cells (MNCs) were formed in response to 1alpha, 25 (OH) _2D_3 in co-cultures of mouse spleen cells and osteoblastic cells freshly isolated from mouse calvariae (Endocrinology, 123 : 2600, 1988). In this study, we examined whether such primary osteoblastic cells can be replaced by established cell lines in inducing osteoclast-like MNC formation. Two bone marrow-derived stromal cell line, MC3T3-G2/PA6 and ST2, were found to suppoort osteoclast-like MNC formation in the co-culture with spleen cells. In addition, a new stromal cell line (KS4), which supports osteoclast-like MNC formation in the co-cultures, was established from mouse calvariae. KS4 cells exhibited typical phenotypes of osteoblasts such as intense alkaline phophatase activity, PTH-dependent cAMP production, and nodule formation in a long-term culture.
(2) Origin of osteoclasts
We examined the
origin of osteoclasts using the co-culture system with ST2 cells. Not only immature hematopoietic cells in bone marrow but also monocytoes in peripheral blood and macrophages in alveolar tissues differentiated into osteoclast-like MNCs in the presence of ST2 cells. This indicates that osteoclasts are the cells of the monocyte-macrophage family.
(3) Effects of colony stimulating factors (CSFs) on osteoclast development
We examined the effects of CSFs (IL-3, GM-CSF, M-CSF, G-CSF) on the proliferation and the differentiation of osteoclast progenitors. None of the CSFs stimulated differentiation of the progenitors into osteoclast-like MNCs. However, the CSFs, especially M-CSF, markedly stimulated proliferation of the progenitors.
The mutation of osteopetrotic (op/op) mice was recently found in the coding region of the M-CSF gene. This indicates that osteoclast deficiency in op/op mice is due to failure of the production of functional M-CSF. We examined osteoclast formation in op/op mice using the co-colture system. We found that osteoblastic cells from op/op mice did not support osteoclast formation in the co-cultures with normal spleen cells. Adding M-CSF to the co-cultures induced the appearance of osteoclast-like cells. These results indicate that M-CSF produced by osteoblastic cells is essential for osteoclast development. Less