Osteoclast Markers Accumulate on Cells Developing from Human Peripheral Blood Mononuclear Precursors

Recent studies show that human osteoclasts develop in vitro from hematopoietic cells; however, special cultures conditions and/or cytokine mobilized peripheral blood are apparently required. Here, we report that cells expressing osteoclast markers differentiate from precursors present in nonmobilized peripheral blood mononuclear cells (PBMC), without the addition of stromal cells, growth factors, cytokines or steroids; and characterize their phenotype. Three days after establishing high-density PBMC cultures (1.5 x 10(6) cells/cm2), in serum-containing medium, small adherent colonies of tartrate resistant acid phosphatase positive (TRAP+) cells emerge, amidst massive monocyte cell death. These adherent cells have an eccentrically placed, round nucleus, and express low levels of TRAP and sodium fluoride-resistant- alpha-naphthyl-acetate-esterase (NaF-R-NSE). Over the next week, this cell population accumulates phenotypic markers of osteoclasts (vitronectin receptor [VR], calcitonin receptor, TRAP, cathepsin K protein, and mRNA) with increased nuclearity, covering the entire surface by 15 days. When cultured on bone, VR+, TRAP+ cells of low multinuclearity appear and cover up to 50% of the surface. Resorption lacunae can be observed by day 22. Although these pits are not nearly as numerous as the cells of preosteoclast phenotype, they do represent the activity of a subset of osteoclast-like cells that has achieved osteoclastic maturity under these culture conditions. Transcripts for osteoprotegerin ligand (OPGL), an osteoclast differentiation factor (also known as RANKL and TRANCE) are expressed, likely by adherent cells. Thus, an adherent population of cells, with preosteoclast/osteoclast phenotypic properties, arises selectively under simple culture conditions from normal PBMC. Further characterization of these cells should identify factors involved in the growth, terminal differentiation and activation of osteoclasts.