Department of Periodontology, Division of Oral Rehabilitation, Faculty of Dental Science, Kyushu University, Fukuoka, Japan.
Department of Periodontology, Division of Oral Rehabilitation, Faculty of Dental Science, Kyushu University, Fukuoka, Japan.
Arch Oral Biol. 2017 Nov;83:241-251. doi: 10.1016/j.archoralbio.2017.08.005. Epub 2017 Aug 12.
Amelogenin, the major component of the enamel matrix derivative (EMD), has been suggested as a bioactive candidate for periodontal regeneration. Apart from producing a regenerative effect on periodontal tissues, amelogenin has also been reported to have an anti-inflammatory effect. However, the precise molecular mechanisms underlying these effects remain unclear. In the present study, we examined the immunomodulatory effects of amelogenin on macrophages.
Human phorbol 12-myristate 13-acetate (PMA)-differentiated U937 macrophages and CD14+ peripheral blood-derived monocytes (PBMC)-derived macrophages were stimulated with recombinant amelogenin (rM180). After performing a detailed microarray analysis, the effects of rM180 on macrophage phenotype and signal transduction pathways were evaluated by real-time polymerase chain reaction, enzyme-linked immunosorbent assay, confocal microscopy and flow cytometry.
The microarray analysis demonstrated that rM180 increased the expression of anti-inflammatory genes in lipopolysaccharide (LPS)-challenged macrophages after 24h, while it temporarily up-regulated inflammatory responses at 4h. rM180 significantly enhanced the expression of M2 macrophage markers (CD163 and CD206). rM180-induced M2 macrophage polarisation was associated with morphological changes as well as vascular endothelial growth factor (VEGF) production. rM180 enhanced prostaglandin E2 (PGE2) expression, and the activation of the cAMP/cAMP-responsive element binding (CREB) signaling pathway was involved in amelogenin-induced M2 macrophage polarisation. Blocking of PGE2 signaling by indomethacin specifically abrogated rM180 with or without LPS-induced M2 shift in PBMC-derived macrophages.
Amelogenin could reprogram macrophages into the anti-inflammatory M2 phenotype. It could therefore contribute to the early resolution of inflammation in periodontal lesions and provide a suitable environment for remodeling-periodontal tissues.
釉原蛋白是牙釉质基质衍生物（EMD）的主要成分，被认为是牙周再生的生物活性候选物。釉原蛋白除了对牙周组织具有再生作用外，还具有抗炎作用。然而，这些作用的确切分子机制尚不清楚。在本研究中，我们研究了釉原蛋白对巨噬细胞的免疫调节作用。
用重组釉原蛋白（rM180）刺激佛波醇 12-肉豆蔻酸 13-乙酸酯（PMA）分化的 U937 巨噬细胞和 CD14+外周血衍生单核细胞（PBMC）衍生的巨噬细胞。在进行详细的微阵列分析后，通过实时聚合酶链反应、酶联免疫吸附试验、共聚焦显微镜和流式细胞术评估 rM180 对巨噬细胞表型和信号转导途径的影响。
微阵列分析表明，rM180 在脂多糖（LPS）刺激后 24 小时增加了 LPS 刺激的巨噬细胞中抗炎基因的表达，而在 4 小时暂时上调了炎症反应。rM180 显著增强了 M2 巨噬细胞标志物（CD163 和 CD206）的表达。rM180 诱导的 M2 巨噬细胞极化与形态变化以及血管内皮生长因子（VEGF）的产生有关。rM180 增强前列腺素 E2（PGE2）的表达，cAMP/cAMP 反应元件结合（CREB）信号通路的激活参与了釉原蛋白诱导的 M2 巨噬细胞极化。吲哚美辛特异性阻断 PGE2 信号转导可特异性阻断 rM180 诱导的 PBMC 衍生巨噬细胞中的 M2 转移，无论是否有 LPS 诱导。
釉原蛋白可以将巨噬细胞重新编程为抗炎的 M2 表型。因此，它可能有助于牙周病变中炎症的早期消退，并为重塑牙周组织提供合适的环境。
