Two step model for the fusion of macrophages induced by 1alpha, 25-dihydroxyvitamin D_3

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We reported that 1alpha,25-dihydroxyvitamin D_3[1alpha,25(OH)_2D_3] directly induces fusion of mouse alveolar macrophages at a high rate of 70-80% within 3 days. Since then, we have been examining the mechanism of cell fusion as a model system of osteoclast formation. In this study, we examined proteins possibly involved in the fusion. We also examined the role of calcium ions on the progression of macrophage fusion induced by 1alpha,25(OH)_2D_3.1.1alpha,25(OH)_2D_3 enhanced the synthesis of spermidine, which was followed by the synthesis of specific 3 proteins (98, 78 and 50 kD) responsible for the fusion. Interleukin 4 (IL-4) similarly induced macrophage fusion by a mechanism involving spermidine. These 3 proteins were also induced by IL-4.2. We found that the fusion process could be divided into two phases: the 1alpha,25(OH)_2D_3-dependent priming phase (0-18 hr) and the calcium-dependent progression phase (18-72 hr). When macrophages were treated with 1alpha,25(OH)_2D_3 in a low calcium (0.13 mill) medium for 48 hr, no fusion occurred. When the medium was changed to normal calcium (1.85 mill) medium, the macrophages began to fuse within 30 min. The extracellular, but not intracellular calcium ions appeared to be involved in this progression phase.3.1alpha,25(OH)_2D_3 induced the dynthesis of transglutaminase, a calcium-dependent enzyme, by a spermidine-dependent mechanism. The treatment of macrophages with cystamine, an inhibitor of transglutaminase, inhibited the fusion in parallel with the inhibition of transglutaminase activity. IL-4 also increased transglutaminase activity by a spermidine-dependent mechanism.In conclusion, 1alpha,25(OH)_22D_3 first induces spermidine synthesis, which in turn induces transglutaminase in the priming phase. This calcium-dependent enzyme appears to catalyze biological reactions responsible for cell fusion. The extracellular, but not intracellular calcium ions play an important role in the progression phase.