Enamel Matrix Derivative Protein Stimulated Wound Healing via Phosphoinositide 3-Kinase
Enamel Matrix Derivative Protein Stimulated Wound Healing via Phosphoinositide 3-Kinase
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DOI：
10.1902/jop.2009.090127
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发表时间：
2009-10-01
影响因子：
4.3
通讯作者：
Domae, Naochika
中科院分区：
文献类型：
--
作者：
Goda, Seiji;Kaneshita, Yuki;Domae, Naochika
背景：牙釉质基质衍生物（Enamel matrix derivative，EMD）蛋白已被临床用于牙周组织再生，但其分子机制尚不清楚。以往的研究表明，磷脂酰肌醇3-激酶（PI 3-kinase）的激活在促进细胞迁移中起着关键作用。鉴于成骨细胞的迁移是创伤愈合过程中的关键步骤之一，我们假设EMD蛋白可以通过激活PI 3-激酶来刺激成骨细胞迁移。在这项研究中，我们测试了这一假设，使用MG-63细胞作为模型系统，以评估机制的迁移刺激EMD protein.Methods：融合MG-63细胞被机械划伤使用无菌I-mm移液器尖端，删除一个圆形区域内的细胞。在有或没有PI 3-激酶抑制剂渥曼青霉素（1、10和100 nM）或LY 294002（1、10和100 μ M）的各种刺激条件（25、50或100 μ g/ml EMD蛋白）下，将威尔斯孔温育24小时。通过从一个孔中随机选择三个区域来计数伤口切片中的迁移细胞。结果：EMD蛋白对MG-63细胞增殖无影响，但能促进MG-63细胞向伤口的迁移。我们发现EMD蛋白以剂量依赖的方式增强Akt的磷酸化。结论：PI 3激酶的激活在EMD蛋白刺激成骨细胞迁移过程中起重要作用。牙周病学杂志2009;80：1631-1637。
Background: Enamel matrix derivative (EMD) protein has been clinically used for periodontal regeneration, but the molecular mechanisms are not clear. Previous studies suggested that the activation of phosphoinositide 3-kinase (PI 3-kinase) plays a key role in facilitating cell migration. Given that the migration of osteoblasts is one of the key steps in the wound-healing processes, we hypothesized that EMD protein would stimulate osteoblast migration by activating PI 3-kinase. In this study, we tested this hypothesis using MG-63 cells as model systems to evaluate mechanisms of migration by stimulation with EMD protein.Methods: Confluent MG-63 cells were mechanically scratched using a sterilized I-mm pipette tip that removed the cells within a circular area. The wells were incubated for 24 hours in various stimulation conditions (25, 50, or 100 mu g/ml EMD protein) with or without the PI 3-kinase inhibitor wortmannin (1, 10, and 100 nM) or LY294002 (1, 10, and 100 mu M). Migrated cells in the wound section were counted by randomly selecting three areas from one well. The activation of PI 3-kinase by EMD protein was evaluated by the phosphorylation of Akt using Western blot analysis.Results: Although EMD protein did not affect proliferation, it enhanced migration into wounds on MG-63 cells. We showed that EMD protein enhanced the phosphorylation of Akt in a dose-dependent manner. We demonstrated that the PI 3-kinase inhibitors wortmannin and LY294002 blocked migration into wounds and the phosphorylation of Akt enhanced by EMD protein in MG-63 cells.Conclusion: These results demonstrated that the activation of PI 3-kinase plays a key role in the EMD protein-stimulated migration of osteoblasts. J Periodontol 2009;80:1631-1637.