NORMAL HUMAN PERIPHERAL-BLOOD MONONUCLEAR-CELLS MOBILIZED WITH GRANULOCYTE-COLONY-STIMULATING FACTOR HAVE INCREASED OSTEOCLASTOGENIC POTENTIAL COMPARED TO NONMOBILIZED BLOOD

Single-cell suspensions of granulocyte colony-stimulating factor (G-CS F)-mobilized peripheral blood mononuclear cells (G-PBMC) cultured in a lpha minimal essential medium (alpha MEM) containing 10% fetal bovine serum formed multicellular aggregates within 24 hours. In six separate experiments, formation of aggregates appeared to be dependent on cell density per surface area, so that 5.8 +/- 1.3 aggregates formed per 1 x 10(5) cells when G-PBMC were cultured at densities greater than or equal to 1 x 10(5) cells/cm(2). The frequency of aggregate formation w as less than 1 per 10(5) cells when G-PBMC were cultured at densities less than 1 x 10(5) cells/cm(2). Once formed, aggregates became adhere nt within 72 hours, and then, over the course of 21 days, released CD3 /CD4/CD25-positive cells into the supernatant. This T-cell production peaked between days 7 and 14, reaching a total of 1,269 +/- 125.9 cell s released per aggregate by day 21. Between days 14 and 21, the aggreg ates also generated macroscopic clusters of adherent mononuclear and g iant multinucleated cells that stained positive for tartrate-resistant acid phosphatase (TRAP). At 4 weeks, the macroscopic foci coalesced i nto monolayers. Multinucleated TRAP-positive cells were distinguished from macrophage polykaryons by the absence of CD14 expression and the presence of osteoclast-specific membrane receptors for calcitonin and alpha(v) beta(3)-vitronectin. The osteoclast nature of these cells was further demonstrated by their ability to form resorption lacunae on d entine slices. Comparable osteoclast formation was not detected in cul tures of normal marrow or normal nonmobilized peripheral blood. (C) 19 96 by The American Society of Hematology.