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

本文报道1 α，25-二羟维生素D_3[1 α，25（OH）_2D_3]直接诱导小鼠肺泡巨噬细胞融合，融合率在3天内高达70-80%。从那时起，我们一直在研究细胞融合的机制作为破骨细胞形成的模型系统。在这项研究中，我们研究了可能参与融合的蛋白质。1.1alpha，25（OH）_2D_3能促进精脒的合成，并促进精脒合成的同时，还能促进精脒合成98、78和50 kD的特异性蛋白质。白细胞介素4（IL-4）同样诱导巨噬细胞融合的机制，涉及亚精胺。这3种蛋白也可被IL-4.2诱导。我们发现，融合过程可分为两个阶段：1 α，25（OH）_2D_3依赖的启动阶段（0-18小时）和钙依赖的进展阶段（18-72小时）。在低钙（0.13mil）培养液中，用1 α，25（OH）_2D_3处理巨噬细胞48小时，无融合发生。当培养基改为正常钙（1.85 mill）培养基时，巨噬细胞在30 min内开始融合。细胞外的钙离子参与了这一过程，而细胞内的钙离子则不参与。3.1 α，25（OH）_2D_3通过亚精胺依赖的机制诱导钙依赖性酶--转氨酶的合成。用胱胺（一种转氨酶抑制剂）处理巨噬细胞，与抑制转氨酶活性平行地抑制融合。IL-4还通过亚精胺依赖性机制增加转氨酶活性，因此，1 α，25（OH）_22D_3首先诱导亚精胺的合成，亚精胺在启动期又诱导转氨酶的合成。这种钙依赖性酶似乎催化负责细胞融合的生物反应。细胞外而非细胞内钙离子在进展阶段起重要作用。

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.