In vitro replication dynamics of human culture-derived macrophages in a long term serum-free system

Human peripheral blood monocytes maintained in a long term serum-free system were found to undergo extensive replication. Newly replicated culture-derived macrophages initially appeared as colonies of small cells on the adherent monolayer. After the appearance of these colonies, large numbers of nonadherent macrophages were observed. Using PKH26, a fluorescent tracking dye, the increase in cell number was attributed to a replicating pool of cells. Half of the monocytes present in the original monolayer were able to undergo at least one cycle of replication and of these, approximately 16% underwent three or more cycles of replication. Macrophages in the nonadherent state contained a larger proportion of cells that had undergone division compared with adherent cells. However, it appeared that only adherent macrophages were capable of replication, suggesting movement between the adherent and nonadherent states. Culture-derived macrophages were also predisposed to multinucleated giant cell formation; and in the nonadherent state, their capacity to form these cells increased. At the end of the study period, approximately 25% of the cells maintained in a nonadherent state had two or more nuclei, and 3% had 10 or more nuclei. By comparison, the adherent cells, over the same period, had 10% of cells with two or more nuclei and none had 10 or more nuclei. These multinucleated cells were found to arise through cell fusion.