McKee M D, Glimcher M J, Nanci A
Department of Stomatology, Université de Montréal, Québec, Canada.
Anat Rec. 1992 Dec;234(4):479-92. doi: 10.1002/ar.1092340404.
The ultrastructural distribution of two noncollagenous proteins, osteopontin (OPN) and osteocalcin (OC), originally extracted from bone matrix and proposed to play an important role in bone formation, was examined in the matrices of bone and cartilage from embryonic and postnatal chicken tibial growth plates by high-resolution immunocytochemistry using the colloidal gold technique. In bone, immunolabeling patterns using polyclonal antibodies against chicken OPN and OC were generally similar in that both showed an intense, but regionally variable, labeling of mineralized bone matrix and small mineralization loci dispersed throughout the osteoid and containing prominent condensed organic material. Unmineralized osteoid showed weak-to-moderate labeling. In the mineralized bone matrix proper, labeling was predominantly associated with amorphous, electron-dense patches of organic material among the collagen fibrils. In growth plate cartilage, both proteins first appeared related to calcified cartilage in the hypertrophic zone, although the labeling patterns were somewhat different. For OPN, gold particles were mostly associated with an organic lamina limitans-like density containing condensed, filamentous organic matrix at the periphery of small nodules and large masses of calcified cartilage, with additional moderate labeling throughout the interior of the calcified cartilage. For OC, labeling was observed over filamentous structures throughout the calcified cartilage matrix, with some, but less, labeling at the periphery. In the lowermost zones of the growth plate, the major reaction using both antibodies was found over a layer of dense, amorphous organic material at the periphery of the calcified cartilage at the future bone/calcified cartilage interface, a labeling pattern that persisted following bone deposition at these sites. OPN and to a lesser extent OC were also concentrated in cement (resting, reversal) lines. Throughout the bone and cartilage of the tibia, cells of both the osteoblastic and the osteoclastic lineages were found directly apposed to labeled surfaces and lamina limitans of organic matrix containing OPN and OC. In summary, it is concluded from the immunocytochemical data presented here that the association of OPN and OC with mineralized regions of the extracellular matrices of bone and cartilage and the accumulation of these proteins at tissue surfaces and interfaces are consistent with the hypotheses that they play a role in the extracellular mineralization process per se and/or that they may mediate cell adhesion and dynamics.
利用胶体金技术通过高分辨率免疫细胞化学方法，检测了最初从骨基质中提取并被认为在骨形成中起重要作用的两种非胶原蛋白——骨桥蛋白（OPN）和骨钙素（OC）在胚胎期和出生后鸡胫骨生长板的骨和软骨基质中的超微结构分布。在骨中，使用抗鸡OPN和OC的多克隆抗体的免疫标记模式总体相似，即两者均显示矿化骨基质有强烈但区域可变的标记，以及分散在类骨质中且含有突出浓缩有机物质的小矿化位点。未矿化的类骨质显示弱阳性至中度标记。在矿化的骨基质中，标记主要与胶原纤维之间无定形、电子致密的有机物质斑块相关。在生长板软骨中，两种蛋白质最初都出现在肥大区与钙化软骨相关的部位，尽管标记模式有所不同。对于OPN，金颗粒大多与类似有机界膜的密度相关，在小结节和大量钙化软骨的周边含有浓缩的丝状有机基质，在钙化软骨内部也有中度标记。对于OC，在整个钙化软骨基质的丝状结构上观察到标记，在周边有一些但较少的标记。在生长板的最下部区域，使用两种抗体的主要反应出现在未来骨/钙化软骨界面处钙化软骨周边一层致密的无定形有机物质上，这种标记模式在这些部位骨沉积后持续存在。OPN以及程度较轻的OC也集中在黏合（静止、反转）线中。在胫骨的整个骨和软骨中，成骨细胞系和破骨细胞系的细胞都直接贴附在含有OPN和OC的有机基质的标记表面和界膜上。总之，从这里呈现的免疫细胞化学数据可以得出结论，OPN和OC与骨和软骨细胞外基质的矿化区域的关联以及这些蛋白质在组织表面和界面的积累，与它们在细胞外矿化过程本身中起作用和/或它们可能介导细胞黏附和动态变化的假设是一致的。
