Department of Oral Anatomy, School of Dentistry, Kyung Hee University, Seoul, Korea.
Clin Oral Implants Res. 2010 Jul;21(7):741-6. doi: 10.1111/j.1600-0501.2009.01901.x.
This study was designed to investigate the effect of enamel derivative matrix (EMD) on the proliferation, mineralization, and differentiation of human mesenchymal stem cells (hMSCs).
For the proliferation assay, water-soluble tetrazolium salt-8 tests were carried out after culturing for 24 and 48 h. For the evaluation of mineralization, Alizarin red S (ARS) tests were performed after 21 days of culturing in an osteogenic medium. In order to investigate some of the bone-related proteins, namely type I collagen (Col I A2), bone sialoprotein (BSP), and bone gamma-carboxyglutamate (Gla) protein (BGLAP, osteocalcin), real-time polymerase chain reaction (RT-PCR) tests were carried out after 2, 3, and 4 weeks of culturing, respectively.
The activity of proliferation and mineralization increased significantly depending on the concentration of EMD (P<0.05). In the control group, the expression of Col I A2 decreased, but EMD enhanced its expression over time and was correlated to the concentration. The amount of expression of BSP in this group increased over time, but EMD strikingly suppressed its expression in the fourth week. As well, the amount of expression of BGLAP increased as the culture duration lengthened in the control group. However, the expression of BGLAP was suppressed in the experimental group with EMD.
Within the limits of this study, EMD enhanced the proliferation of hMSCs. After evaluation with ARS staining, EMD seemed to enhance mineralization, and the RT-PCR test revealed that EMD promoted early-stage osteoblast differentiation by enhancing Col I A2 expression, but exerted an inhibitory effect on the mineralization by lowering the gene expression of BSP and BGLAP. Mineralized nodules formed with EMD may be composed of substances other than normal bone. Because most of the organic matrix of bone is type I collagen, which acts as the mineralization site, bone or bone-like mineralized mass might have been formed in spite of the different components of the non-collagenous proteins.
本研究旨在探讨釉原蛋白基质（EMD）对人间充质干细胞（hMSCs）增殖、矿化和分化的影响。
采用水溶性四唑盐（WST-8）法在 24 小时和 48 小时后进行增殖检测。在成骨培养基中培养 21 天后，采用茜素红 S（ARS）法进行矿化评价。为了研究一些与骨相关的蛋白，即 I 型胶原（Col I A2）、骨涎蛋白（BSP）和骨γ-羧基谷氨酸（Gla）蛋白（BGLAP，骨钙素），在培养 2、3 和 4 周后分别进行实时聚合酶链反应（RT-PCR）检测。
增殖和矿化活性随 EMD 浓度的增加而显著增加（P<0.05）。在对照组中，Col I A2 的表达减少，但 EMD 随着时间的推移增强了其表达，并与浓度相关。该组中 BSP 的表达量随着时间的推移而增加，但 EMD 在第四周显著抑制了其表达。同样，在对照组中，随着培养时间的延长，BGLAP 的表达量增加。然而，用 EMD 处理的实验组中，BGLAP 的表达受到抑制。
在本研究范围内，EMD 增强了 hMSCs 的增殖。通过 ARS 染色评估，EMD 似乎增强了矿化，RT-PCR 检测显示，EMD 通过增强 Col I A2 的表达促进早期成骨细胞分化，但通过降低 BSP 和 BGLAP 的基因表达对矿化产生抑制作用。用 EMD 形成的矿化结节可能由不同于正常骨的物质组成。因为骨的大多数有机基质是 I 型胶原，它作为矿化部位，尽管非胶原蛋白的成分不同，但可能形成了骨或类骨质矿化物质。
