Production of human osteoclasts in a three-dimensional bone marrow culture system

Osteoclasts, which are derived from hemopoietic stem cells, play important roles in bone remodeling and resorption. Osteoclast development is critically dependent on the bone microenvironment. We have developed a novel human ex vivo bone marrow model that mimics bone marrow both structurally and functionally by providing artificial scaffolding, thus obtaining a three-dimensional growth configuration with high cell density and intimate physical contact between hemopoietic and stromal cells. In this study, we utilized the 3-D culture system to produce multinucleated cells (MNCs) from human bone marrow cells cultured in the presence of Vitamin D-3 and the absence of hydrocortisone and any exogenous growth factors. These multinucleated cells had the phenotypic and functional characteristics of osteoclasts as determined by their morphology, expression of tartarate resistant acid phosphatase (TRAP), and the ability to resorb bone. Scoring of the resorption pits revealed a dose response to Vitamin D3 stimulation with 10(-7) M being the optimal concentration. Furthermore, addition of parathyroid hormone (10(-8) M) resulted in an up-to three-fold enhancement of bone resorption. These findings suggest that the 3-D culture system represent a physiologically relevant model to study osteoclastogenesis and elucidate the molecular and cellular signals associated with this process. (C) 2003 Elsevier B.V. All rights reserved.