Immunohistochemical Analysis of the Distribution of RANKL-Expressing Cells and the Expression of Osteoclast-Related Markers in Giant Cell Tumor of Bone

To clarify the mechanism underlying the regulation of osteoclast differentiation and activation in giant cell tumor of bone, we investigated the expression of osteoclast-related markers in osteoclasts, neoplastic cells, and osteoclast precursors comprising giant cell tumor of bone using immunohistochemical analysis, and analyzed the distribution of receptor activator of nuclear factor K-B ligand (RANKL)-expressing cells. We performed serial staining of sections using antibodies against osteoclast-related markers including RANKL. CD68, CD11b, c-Fms, RANK, and cathepsin K and analyzed the presence of the G34W mutation of H3F3A in giant cell tumor of bone. Mononuclear cells were CD11b(+), CD68(+), RANK, and c-Fms(+). In contrast, most multinucleate cells were CD11b(+), CD68(+), RANK(+), and c-Fms(+). RANKL was expressed in mononuclear cells and some multinucleate cells. Conversely, G34W was detected in mononuclear cells. RANKL(+) mononuclear cells were also G34W. RANKL expression was unevenly distributed in giant cell tumor of bone. RANKL(+) cells were frequently localized along blood-containing cavity-like spaces. In areas with a high percentage of RANKL(+) mononuclear cells, large numbers of osteoclasts were observed. In addition, the distribution of multinucleate cells (>100 mu m in diameter) correlated with the distribution of RANKL(+) cells. The distribution of RANKL(+) mononuclear cells is uneven in giant cell tumor of bone and may have some effect on the localization of multinucleate cells. The distribution of mononuclear cells harboring the G34W mutation was identical to that of RANK(+) mononuclear cells.