Both direct and collagen-mediated signals are required for active vitamin D3-elicited differentiation of human osteoblastic cells:: Roles of osterix, an osteoblast-related transcription factor
Both direct and collagen-mediated signals are required for active vitamin D3-elicited differentiation of human osteoblastic cells:: Roles of osterix, an osteoblast-related transcription factor
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DOI：
10.1016/j.matbio.2005.09.001
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发表时间：
2006-01-01
期刊：
影响因子：
6.9
通讯作者：
Hata, RI
中科院分区：
文献类型：
--
作者：
Maehata, Y;Takamizawa, S;Hata, RI
为了研究1 α，25（OH）2维生素D-3（VD 3）刺激人成骨细胞分化的机制，我们在存在或不存在VD 3和/或L-抗坏血酸2-磷酸（Asc 2-P）（一种长效维生素C衍生物）的情况下培养MG-63，MG-63是一种人成骨细胞系。细胞生长速率下降的存在下，在培养基中的VD 3。I型胶原合成和碱性磷酸酶（ALP）的活性，这是早期成骨细胞分化的标志物，刺激的存在下，VD 3以及由Asc 2-P。Asc 2-P和VD 3的共同存在的胶原合成和ALP活性的细胞有协同效应。通过向培养基中添加胶原合成抑制剂来抑制胶原合成，减弱了VD 3和Ase 2-P对ALP活性的刺激作用。用COL 1A 1的siRNA表达载体转染细胞，除COL 1A 1的表达水平外，ALP mRNA的表达水平也降低。另一方面，当细胞在I型胶原包被的培养皿上培养时，ALP活性显著增加，并且生长速率降低。当细胞在涂有热变性胶原蛋白的培养皿上培养时，没有观察到这些效果。VD 3还增加了Runx 2和osterix的mRNA水平，它们是成骨细胞分化的关键转录因子，以及分化标志物如骨/肝/肾型ALP、COL 1A 1（a. I型胶原蛋白的I链）和骨钙素。正常人成骨细胞和人骨髓间充质干细胞（hBMSC）对VD 3表现出非常相似的反应。这些结果表明，VD 3刺激的基因表达的I型胶原蛋白和成熟的I型胶原蛋白的存在下产生的Ase 2-P介导的至少一部分的刺激作用的Asc 2-P和VD 3对这些人成骨细胞的分化。在MG-63细胞中，Osterix的表达增加了ALP和COL 1A 1的mRNA水平，但降低了Runx 2的mRNA水平。这些结果还表明，VD 3通过调节成骨细胞相关转录因子的基因表达以及I型胶原蛋白的基因表达来控制人成骨细胞的生长和分化，并且这两种信号的共存对于VD 3表达朝向人成骨细胞分化的全部活性是必需的。(c)2005年Elsevier B. V./国际矩阵生物学学会。All rights reserved.
In order to investigate the mechanisms by which 1 alpha,25(OH)2 vitamin D-3 (VD3) stimulates the differentiation of human osteoblasts, we cultured MG-63, which is a human osteoblastic cell line, in the presence or absence of VD3 and/or L-ascorbic acid 2-phosphate (Asc 2-P), a long-acting vitamin C derivative. The cell growth rate was decreased by the presence of VD3 in the culture medium. Type I collagen synthesis and alkaline phosphatase (ALP) activity, which are markers of early stage osteoblast differentiation, were stimulated by the presence of VD3 as well as by that of Asc 2-P. The co-presence of Asc 2-P and VD3 had a synergistic effect on the collagen synthesis and ALP activity of the cells. Inhibition of collagen synthesis by the addition of inhibitors of collagen synthesis to the medium attenuated the stimulative effect of VD3 and Ase 2-P on the ALP activity. Transfection of the cells with siRNA-expressing vectors for COL I A I decreased the expression level of ALP mRNA in addition to that of COL1A1. On the other hand, ALP activity was significantly increased, and the growth rate was decreased, when the cells were cultured on type I collagen-coated dishes. These effects were not seen when the cells were cultured on dishes coated with heat-denatured collagen. VD3 also increased the mRNA levels for Runx2 and osterix, which are transcription factors critical for osteoblast differentiation, as well as those of differentiation markers such as bone/liver/kidney type ALP, COL1A1, (the gene for the a. I chain of type I collagen), and osteocalcin, in the cells. Normal human osteoblasts and human bone marrow-derived mesenchymal stem cells (hBMSC) showed quite similar responses to VD3. These results indicate that VD3-stimulated gene expression of type I collagen and that mature type I collagen produced in the presence of Ase 2-P mediates at least a part of the stimulative effects of Asc 2-P and VD3 on the differentiation of these human osteoblastic cells. Levels of mRNAs for ALP and COL1A1 were increased, but the level of Runx2 was decreased, by the expression of osterix in MG-63 cells. These results also suggest that VD3 controls the growth and differentiation of human osteoblastic cells by regulating the gene expression of osteoblast-related transcription factors as well as that of type I collagen, and that the co-presence of both signals is essential for VD3 to express full activity toward the differentiation of human osteoblasts. (c) 2005 Elsevier B.V./International Society of Matrix Biology. All rights reserved.