Interactions between Myeloma Cells and Osteoclasts Are Controlled by Unique Histological Structures Involving Vascular Compartments.

Although myeloma development and enhanced bone resorption are intimately related, the mechanism responsible of this relation is not known. In vitro studies have stressed the critical role of direct cell contacts between myeloma cells and osteoclast. In vivo, however, little is known about the organization of the cells present at osteolytic lesions, because of the complexity of cell-cell interactions in the bone marrow of myeloma patients. Therefore, we conducted an immunohistochemical study with multiple stainings allowing the simultaneous identification of different cell types at resorption sites of bone marrow biopsies of myeloma patients.

The biopsies showed that in average 1% of the bone surface was lined by mature multinucleated TRAP+ osteoclasts, but that only 6% of these osteoclasts showed direct contacts with myeloma cells. The biopsies showed also TRAP+ mononucleated pre-osteoclasts in the bone marrow compartment, and 40% of these pre-osteoclasts showed direct contacts with myeloma cells. Bone marrow pre-osteoclasts show thus much more frequent contacts with myeloma cells, compared with mature osteoclasts lining bone surfaces. These respective values remained unchanged, whether the myeloma cells were identified through CD138 or through light chain expression (counts in biopsies from 13 patients). Importantly, we found that 80% of the osteoclasts lining the bone surfaces, were separated from the bone marrow compartment by a specialized cell wall (seen in all biopsies of the 15 patients, that were analyzed). This wall consists of a single layer of (sometimes very) flattened cells lining the bone marrow, and expressing NCAM, propeptide of type III collagen, and osteocalcin, but not CD34. When performing 3D reconstructions by using serial sections, this wall appeared as a continuous roof covering the bone surfaces undergoing remodeling, and connected to the bone lining cells at its periphery. Furthermore, CD34 staining revealed that capillaries are abundant at the bone marrow side of this cell wall, and that some show an opening into the wall. These capillaries may thus allow communication between the bone marrow and the bone surfaces undergoing remodeling, and render the bone remodeling compartments vascular-like. In addition, the TRAP+ preosteoclast detected in the bone marrow space, were positioned along the capillaries leading to the vascular/remodeling compartments.

In conclusion, this study shows that in vivo, interactions between myeloma cells and osteoclasts are mediated only rarely through direct cell contacts, and identifies for the first time unique cell arrangements that are likely to play a role in these interactions:

• a specialized cell wall separates the bone marrow from the vascular/remodeling compartments in most resorption sites (80% of the osteoclasts), and the cells of this wall are thus in a privileged situation to control myeloma-osteoclast interactions;

• capillaries connect the marrow cavity with the vascular/remodeling compartment, thereby allowing guidance of pre-osteoclasts from the bone marrow to the resorption sites;

• the generation of these pre-osteoclasts may be stimulated by the high incidence of their direct contacts with myeloma cells in the bone marrow.