Bone sialoprotein expression enhances osteoblast differentiation and matrix mineralization in vitro
Bone sialoprotein expression enhances osteoblast differentiation and matrix mineralization in vitro
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DOI：
10.1016/j.bone.2007.04.191
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发表时间：
2007-09-01
期刊：
影响因子：
4.1
通讯作者：
Goldberg, Harvey A.
中科院分区：
文献类型：
--
作者：
Gordon, Jonathan A. R.;Tye, Coralee E.;Goldberg, Harvey A.
骨唾液蛋白（BSP）是一种在矿化组织中大量表达的空气酸性非胶原糖蛋白。虽然BSP经常被用作成骨细胞分化的标记物，但该蛋白在成骨细胞功能中的作用尚不清楚。BSP属于RGD-containing matrix proteins的SIBLING （Small integrin binding LIgand N-linked Glycoprotein，小整合素结合配体n-链接糖蛋白）家族，其中一些成员已被证明影响细胞分化。在MC3T3E1细胞系中，cmv介导的腺病毒在原代成骨细胞中过表达或RNA干扰抑制可特异性改变成骨细胞中BSP的正常表达水平。或者，在成骨细胞培养物中添加重组BSP蛋白。采用实时荧光定量PCR检测成骨相关转录因子Osterix和Runx2以及成骨特异性基因骨钙素的mRNA水平。作为成骨细胞分化的标志，碱性磷酸酶活性、runx2 -荧光素酶报告活性和钙黄蛋白掺入矿化培养物也被测量。BSP的过表达增加了成骨细胞相关基因的表达以及钙的掺入和成骨细胞结节的形成。同样，在成骨细胞培养物中添加重组BSP也增加了成骨细胞分化的几种标志物。相反，通过小发夹rna编码质粒抑制BSP的表达可抑制成骨细胞标志物的表达和结节的形成。BSP的几个功能域突变体的过表达表明，在BSP过表达模型中观察到的成骨相关基因表达和基质矿化的增加是由蛋白质c端附近的整合素结合RGD基序介导的。这些结果表明BSP可能作为基质相关信号直接促进成骨细胞分化，导致矿化基质的产生增加。(C) 2007爱思唯尔公司版权所有。
Bone sialoprotein (BSP) is air acidic, noncollagenous glycoprotein abundantly expressed in mineralized tissues. Although BSP is frequently used as a marker of osteoblast differentiation, the role of the protein in osteoblast function is unclear. BSP belongs to the SIBLING (Small Integrin-binding LIgand N-linked Glycoprotein) family of RGD-containing matrix proteins, several members of which have been shown to affect cell differentiation. The normal levels of BSP expression in osteoblasts were specifically altered by CMV-mediated adenoviral overexpression in primary osteoblasts or inhibition by an RNA interference-based strategy in the MC3T3E1 cell line. Alternatively, osteoblast cultures were supplemented with recombinant BSP protein. Quantitative real-time PCR was used to monitor the mRNA levels of the osteoblast-related transcription factors Osterix and Runx2 as well as the osteoblast-specific gene osteocalcin. As markers of osteoblast differentiation, alkaline phosphatase enzyme activity, Runx2-luciferase reporter activity and calcein incorporation into mineralized cultures were also measured. The overexpression of BSP increased osteoblast-related gene expression as well as calcium incorporation and nodule formation by osteoblast cultures. Similarly, supplementation of osteoblast cultures with recombinant BSP increased several markers of osteoblast differentiation. Conversely, suppression of BSP expression by small-hairpin RNA-encoding plasmids inhibited expression of osteoblast markers and nodule formation. Overexpression of several functional-domain mutants of BSP demonstrated that increases in osteoblast-related gene expression and matrix mineralization observed in BSP overexpression models are mediated by the integrin-binding RGD motif found near the C-terminus of the protein. These results demonstrate that BSP may serve as a matrix-associated signal directly promoting osteoblast differentiation resulting in the increased production of a mineralized matrix. (C) 2007 Elsevier Inc. All rights reserved.