Altered Mineralization of Human Osteoarthritic Osteoblasts Is Attributable to Abnormal Type I Collagen Production
Altered Mineralization of Human Osteoarthritic Osteoblasts Is Attributable to Abnormal Type I Collagen Production
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DOI：
10.1002/art.24489
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发表时间：
2009-05-01
影响因子：
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通讯作者：
Lajeunesse, Daniel
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文献类型：
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作者：
Couchourel, Denis;Aubry, Isabelle;Lajeunesse, Daniel
目标。骨关节炎（OA）的骨组织由大量矿化程度较低的类骨基质组成。本研究的目的是利用体外OA软骨下成骨细胞来探讨这种异常基质的形成机制。从胫骨平台制备正常和OA成骨细胞。采用碱性磷酸酶活性测定表型，采用酶联免疫吸附法测定骨钙素、骨桥蛋白、前列腺素E-2 （PGE(2)）和转化生长因子β 1 （TGF β 1）。实时聚合酶链反应检测COL1A1和COL1A2的表达。通过其c端前肽的释放和Western blot分析来确定I型胶原蛋白的产生。用茜素红染色评价体外矿化。利用小干扰RNA技术抑制TGF β 1的表达。与正常成骨细胞相比，OA成骨细胞的矿化程度降低，即使存在骨形态发生蛋白2 （BMP-2）。与正常成骨细胞相比，OA成骨细胞的碱性磷酸酶和骨钙素水平升高，而骨桥蛋白水平相似。OA成骨细胞的col1a1 - col1a2信使RNA比正常成骨细胞高3倍，OA成骨细胞的胶原生成增加。因为TGF β 1抑制bmp -2依赖性矿化，并且因为OA成骨细胞中TGF β 1水平比正常成骨细胞高4倍，抑制TGF β 1。OA成骨细胞中COL1A1-to-COL1A2比值异常，茜素红染色升高。OA成骨细胞中TGF1 β 1水平升高是导致COL1A1 / COL1A2比例异常以及成熟I型胶原生成异常的部分原因。这种异常的col1a1 -to - col1a2比值会产生一种基质，使OA成骨细胞的矿化变得迟钝。
Objective. Bone tissue in osteoarthritis (OA) is composed of abundant undermineralized osteoid matrix. The aim of this study was to investigate the mechanisms responsible for this abnormal matrix, using in vitro OA subchondral osteoblasts.Methods. Primary normal and OA osteoblasts were prepared from tibial plateaus. Phenotype was determined by alkaline phosphatase activity, and osteocalcin, osteopontin, prostaglandin E-2 (PGE(2)), and transforming growth factor beta 1 (TGF beta 1) were assessed by enzyme-linked immunosorbent assay. Expression of COL1A1 and COL1A2 was determined by real-time polymerase chain reaction. The production of type I collagen was determined by the release of its C-terminal propeptide and Western blot analysis. In vitro mineralization was evaluated by alizarin red staining. Inhibition of TGF beta 1 expression was performed using a small interfering RNA technique.Results. Mineralization of OA osteoblasts was reduced compared with mineralization of normal osteoblasts, even in the presence of bone morphogenetic protein 2 (BMP-2). Alkaline phosphatase and osteocalcin levels were elevated in OA osteoblasts compared with normal osteoblasts, whereas osteopontin levels were similar. The COL1A1-to-COL1A2 messenger RNA ratio was 3-fold higher in OA osteoblasts compared with normal osteoblasts, and the production of collagen by OA osteoblasts was increased. Because TGF beta 1 inhibits BMP-2-dependent mineralization, and because TGF beta 1 levels are similar to 4-fold higher in OA osteoblasts than in normal osteoblasts, inhibiting TGF beta 1. levels in OA osteoblasts corrected the abnormal COL1A1-to-COL1A2 ratio and increased alizarin red staining.Conclusion. Elevated TGF1 beta 1 levels in OA osteoblasts are responsible, in part, for the abnormal ratio of COL1A1 to COL1A2 and for the abnormal production of mature type I collagen. This abnormal COL1A1-toCOL1A2 ratio generates a matrix that blunts mineralization in OA osteoblasts.