Jacques Jaime, Hotton Dominique, De la Dure-Molla Muriel, Petit Stephane, Asselin Audrey, Kulkarni Ashok B, Gibson Carolyn Winters, Brookes Steven Joseph, Berdal Ariane, Isaac Juliane
Laboratory of Molecular Oral Pathophysiology, INSERM UMRS 1138, Team Berdal, Cordeliers Research Center, Pierre and Marie Curie University - Paris 6, Paris Descartes University - Paris 5, Paris, France; UFR d'Odontologie, Paris Diderot University - Paris 7, Paris, France; Unit of Periodontology, Department of Stomatology, University of Talca, Talca, Chile.
Laboratory of Molecular Oral Pathophysiology, INSERM UMRS 1138, Team Berdal, Cordeliers Research Center, Pierre and Marie Curie University - Paris 6, Paris Descartes University - Paris 5, Paris, France.
PLoS One. 2014 Jun 16;9(6):e99626. doi: 10.1371/journal.pone.0099626. eCollection 2014.
Research on enamel matrix proteins (EMPs) is centered on understanding their role in enamel biomineralization and their bioactivity for tissue engineering. While therapeutic application of EMPs has been widely documented, their expression and biological function in non-enamel tissues is unclear. Our first aim was to screen for amelogenin (AMELX) and ameloblastin (AMBN) gene expression in mandibular bones and soft tissues isolated from adult mice (15 weeks old). Using RT-PCR, we showed mRNA expression of AMELX and AMBN in mandibular alveolar and basal bones and, at low levels, in several soft tissues; eyes and ovaries were RNA-positive for AMELX and eyes, tongues and testicles for AMBN. Moreover, in mandibular tissues AMELX and AMBN mRNA levels varied according to two parameters: 1) ontogenic stage (decreasing with age), and 2) tissue-type (e.g. higher level in dental epithelial cells and alveolar bone when compared to basal bone and dental mesenchymal cells in 1 week old mice). In situ hybridization and immunohistodetection were performed in mandibular tissues using AMELX KO mice as controls. We identified AMELX-producing (RNA-positive) cells lining the adjacent alveolar bone and AMBN and AMELX proteins in the microenvironment surrounding EMPs-producing cells. Western blotting of proteins extracted by non-dissociative means revealed that AMELX and AMBN are not exclusive to mineralized matrix; they are present to some degree in a solubilized state in mandibular bone and presumably have some capacity to diffuse. Our data support the notion that AMELX and AMBN may function as growth factor-like molecules solubilized in the aqueous microenvironment. In jaws, they might play some role in bone physiology through autocrine/paracrine pathways, particularly during development and stress-induced remodeling.
对釉基质蛋白（EMPs）的研究主要集中在了解它们在釉质生物矿化中的作用及其在组织工程中的生物活性。虽然EMPs的治疗应用已有广泛记载，但其在非釉质组织中的表达和生物学功能尚不清楚。我们的首要目标是筛选从成年小鼠（15周龄）分离的下颌骨和软组织中釉原蛋白（AMELX）和成釉蛋白（AMBN）基因的表达。使用逆转录聚合酶链反应（RT-PCR），我们发现AMELX和AMBN在下颌牙槽骨和基骨中有mRNA表达，在一些软组织中表达水平较低；眼睛和卵巢的AMELX RNA呈阳性，眼睛、舌头和睾丸的AMBN RNA呈阳性。此外，在下颌组织中，AMELX和AMBN的mRNA水平根据两个参数而变化：1）个体发育阶段（随年龄增长而降低），以及2）组织类型（例如，与1周龄小鼠的基骨和牙间充质细胞相比，牙上皮细胞和牙槽骨中的水平更高）。使用AMELX基因敲除小鼠作为对照，在下颌组织中进行原位杂交和免疫检测。我们在相邻牙槽骨内衬中鉴定出产生AMELX的（RNA阳性）细胞，以及在产生EMPs的细胞周围微环境中的AMBN和AMELX蛋白。通过非解离方法提取的蛋白质的蛋白质印迹分析表明，AMELX和AMBN并非矿化基质所特有；它们在一定程度上以溶解状态存在于下颌骨中，并且可能具有一定的扩散能力。我们的数据支持这样一种观点，即AMELX和AMBN可能作为溶解在水性微环境中的生长因子样分子发挥作用。在颌骨中，它们可能通过自分泌/旁分泌途径在骨生理学中发挥某些作用，特别是在发育和应激诱导的重塑过程中。
