Aluminum-induced mitogenesis in MC3T3-E1 osteoblasts: potential mechanism underlying neoosteogenesis

We recently reported that aluminum administration to beagles stimulates uncoupled bone formation in the marrow cavity which increases trabecular bone volume and generates new osseous networks within the axial skeleton. To investigate whether this osteogenic process results from direct stimulation of bone cell replication, we examined the mitogenic effects of aluminum on undifferentiated osteoblasts derived from the MC3T3-E1 clonal cell line. Addition of AlCl3 (1-50 microM) to serum-free culture medium of quiescent osteoblasts resulted in a dose-dependent increase in [3H]thymidine incorporation into DNA and a concordant increase in cell number to 48% of the density achieved at the maximum replicative rate induced by fetal bovine serum (FBS). The time course of aluminum-induced mitogenesis was similar to that of FBS, with onset of DNA synthesis detectable by 12 h and progressive increases in replicative rates observed over a 48-h study period. Moreover, maximal stimulation of DNA synthesis by AlCl3 and that by FBS were not additive, whereas aluminum exerted additional effects on cell replication when combined with low FBS concentrations. Analysis of cell cycle kinetics indicated that aluminum, analogous to FBS, influences the osteoblast replicative activity by inducing transition from the G0 to the S phase of the cell cycle. In addition, exposure of cells to aluminum resulted in rapid accumulation of c-fos mRNA by 30 min, indicating that aluminum, like fetal bovine serum, induced expression of growth-regulating genes. Deferoxamine mesylate, a chelator of aluminum, blocked the replicative actions of aluminum in a dose-dependent fashion. In contrast, pertussis toxin, a specific inhibitor of certain G-proteins, had no effect on the mitogenic effects of aluminum, indicating that aluminum-induced mitogenesis occurs by a pertussis toxin-insensitive pathway. Though the particular cellular pathway remains to be defined, these data provide initial evidence that aluminum-induced neoosteogenesis may depend upon direct stimulation of osteoblast replication.