Analysis of Minerals Produced by hFOB 1.19 and Saos-2 Cells Using Transmission Electron Microscopy with Energy Dispersive X-ray Microanalysis
Analysis of Minerals Produced by hFOB 1.19 and Saos-2 Cells Using Transmission Electron Microscopy with Energy Dispersive X-ray Microanalysis
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DOI：
10.3791/57423
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发表时间：
2018-06-01
影响因子：
1.2
通讯作者：
Strzelecka-Kiliszek, Agnieszka
中科院分区：
文献类型：
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作者：
Bozycki, Lukasz;Komiazyk, Magdalena;Strzelecka-Kiliszek, Agnieszka
本视频介绍了使用透射电子显微镜和能量色散X射线微量分析（TEM-EDX）来比较两种人骨细胞系hFOB 1.19和Saos-2释放的囊泡中矿物质的状态。这些细胞系在用抗坏血酸（AA）和β-甘油磷酸盐（β-GP）处理后，通过茜素红-S（AR-S）染色和紫外线（UV）分析细胞裂解液中的矿物质组成，光下或囊泡中使用TEM成像，然后进行EDX定量和离子绘图，我们可以推断骨肉瘤Saos-2和成骨细胞hFOB 1.19细胞显示出不同的矿化特征。Saos-2细胞比hFOB 1.19细胞更有效地矿化，并产生更大的矿物沉积物，这些矿物沉积物在紫外光下不可见，但与羟基磷灰石（HA）相似，因为它们具有更多的Ca和F取代。使用这些技术获得的结果使我们能够得出结论，矿化过程取决于细胞类型。我们认为，在细胞水平上，囊泡的起源和性质预先决定了矿物质的类型。
This video presents the use of transmission electron microscopy with energy dispersive X-ray microanalysis (TEM-EDX) to compare the state of minerals in vesicles released by two human bone cell lines: hFOB 1.19 and Saos-2. These cell lines, after treatment with ascorbic acid (AA) and beta-glycerophosphate (beta-GP), undergo complete osteogenic transdifferentiation from proliferation to mineralization and produce matrix vesicles (MVs) that trigger apatite nucleation in the extracellular matrix (ECM).Based on Alizarin Red-S (AR-S) staining and analysis of the composition of minerals in cell lysates using ultraviolet (UV) light or in vesicles using TEM imaging followed by EDX quantitation and ion mapping, we can infer that osteosarcoma Saos-2 and osteoblastic hFOB 1.19 cells reveal distinct mineralization profiles. Saos-2 cells mineralize more efficiently than hFOB 1.19 cells and produce larger mineral deposits that are not visible under UV light but are similar to hydroxyapatite (HA) in that they have more Ca and F substitutions.The results obtained using these techniques allow us to conclude that the process of mineralization differs depending on the cell type. We propose that, at the cellular level, the origin and properties of vesicles predetermine the type of minerals.