Matsumoto T, Igarashi C, Takeuchi Y, Harada S, Kikuchi T, Yamato H, Ogata E
Fourth Department of Internal Medicine, University of Tokyo School of Medicine, Japan.
Bone. 1991;12(1):27-32. doi: 10.1016/8756-3282(91)90051-j.
Although vitamin D is essential for mineralization of bone, it is as yet unclear whether vitamin D has a direct stimulatory effect on the bone mineralization process. In the present study, the effect of 1,25-dihydroxyvitamin D3 [1,25(OH)2D3] on in vitro mineralization mediated by osteoblast-like MC3T3-E1 cells was examined. MC3T3-E1 cells continued to grow after they reached confluency, and DNA content and alkaline phosphatase activity increased linearly until about 16 days of culture, whereas 45Ca accumulation into cell and matrix layer remained low. After this period, DNA content plateaued, and 45Ca accumulation increased sharply. Histological examination by von Kossa staining revealed that calcium was accumulated into extracellular matrix. In addition, needle-shaped mineral crystals similar to hydroxyapatite crystals could be demonstrated in between collagen fibrils by electron microscopy. Thus, MC3T3-E1 cells differentiate in vitro into cells with osteoblastic phenotype and exhibit mineralization. When MC3T3-E1 cells were treated with 1,25(OH)2D3 at this stage of culture, there was a dose-dependent stimulation of 45Ca accumulation by 1,25(OH)2D3, and a significant stimulation of 45Ca accumulation was observed with 3 x 10(-10) M 1,25(OH)2D3. Although 1,25(OH)2D3 enhanced alkaline phosphatase activity and collagen synthesis at the early phase of culture, it did not affect any of these parameters at the late phase when 1,25(OH)2D3 stimulated mineralization. Neither 24,25-dihydroxyvitamin D3 nor human PTH(1-34) affected mineralization in the presence or absence of 1,25(OH)2D3. These results demonstrate that 1,25(OH)2D3 stimulates matrix mineralization induced by osteoblastic MC3T3-E1 cells, and are consistent with the possibility that 1,25(OH)2D3 has a direct stimulatory effect on bone mineralization process.(ABSTRACT TRUNCATED AT 250 WORDS)
尽管维生素D对骨矿化至关重要，但维生素D是否对骨矿化过程具有直接刺激作用仍不清楚。在本研究中，检测了1,25-二羟基维生素D3[1,25(OH)2D3]对成骨样MC3T3-E1细胞介导的体外矿化的影响。MC3T3-E1细胞汇合后继续生长，DNA含量和碱性磷酸酶活性在培养约16天前呈线性增加，而45Ca在细胞和基质层中的积累仍较低。在此之后，DNA含量趋于平稳，45Ca积累急剧增加。通过冯·科萨染色进行的组织学检查显示钙积累到细胞外基质中。此外，通过电子显微镜可在胶原纤维之间证实存在类似于羟基磷灰石晶体的针状矿物晶体。因此，MC3T3-E1细胞在体外分化为具有成骨细胞表型的细胞并表现出矿化。当在培养的这个阶段用1,25(OH)2D3处理MC3T3-E1细胞时，1,25(OH)2D3对45Ca积累有剂量依赖性刺激作用，在3×10(-10)M 1,25(OH)2D3时观察到对45Ca积累有显著刺激作用。尽管1,25(OH)2D3在培养早期增强了碱性磷酸酶活性和胶原蛋白合成，但在1,25(OH)2D3刺激矿化的后期，它对这些参数均无影响。在有或没有1,25(OH)2D3的情况下，24,25-二羟基维生素D3和人甲状旁腺激素(1-34)均不影响矿化。这些结果表明1,25(OH)2D3刺激成骨细胞MC3T3-E1细胞诱导的基质矿化，并且与1,25(OH)2D3对骨矿化过程具有直接刺激作用的可能性一致。(摘要截短为250字)
