1 The Forsyth Institute, Cambridge, MA, USA.
2 Department of Developmental Biology, Harvard School of Dental Medicine, Boston, MA, USA.
J Dent Res. 2019 Apr;98(4):468-475. doi: 10.1177/0022034518823537. Epub 2019 Feb 11.
The present study was conducted to investigate the role of proteolysis by matrix metalloproteinase 20 (MMP20) in regulating the initial formation of the enamel mineral structure during the secretory stage of amelogenesis, utilizing Mmp20-null mice that lack this essential protease. Ultrathin sagittal sections of maxillary incisors from 8-wk-old wild-type (WT), Mmp20-null (KO), and heterozygous (HET) littermates were prepared. Secretory-stage enamel ultrastructures from each genotype as a function of development were compared using transmission electron microscopy, selected area electron diffraction, and Raman microspectroscopy. Characteristic rod structures observed in WT enamel exhibited amorphous features in newly deposited enamel, which subsequently transformed into apatite-like crystals in older enamel. Surprisingly, initial mineral formation in KO enamel was found to proceed in the same manner as in the WT. However, soon after a rod structure began to form, large plate-like crystals appeared randomly within the developing KO enamel layer. As development continued, observed plate-like crystals became dominant and obscured the appearance of the enamel rod structure. Upon formation of these plate-like crystals, the KO enamel layer stopped growing in thickness, unlike WT and HET enamel layers that continued to grow at the same rate. Raman results indicated that Mmp20-KO enamel contains a significant portion of octacalcium phosphate, unlike WT enamel. Although normal in all other respects, large, randomly dispersed mineral crystals were observed in secretory HET enamel, although to a lesser extent than that seen in KO enamel, indicating that the level of MMP20 expression has a proportional effect on suppressing aberrant mineral formation. In conclusion, we found that proteolysis of extracellular enamel matrix proteins by MMP20 is not required for the initial development of the enamel rod structure during the early secretory stage of amelogenesis. Proteolysis by MMP20, however, is essential for the prevention of abnormal crystal formation during amelogenesis.
本研究旨在探讨基质金属蛋白酶 20（MMP20）的蛋白水解作用在调节成釉细胞分泌期牙釉质初始矿化结构形成中的作用，利用缺乏这种必需蛋白酶的 Mmp20 基因敲除（KO）小鼠。从 8 周龄野生型（WT）、Mmp20 基因敲除（KO）和杂合（HET）同窝仔鼠的上颌切牙中制备超薄矢状切片。利用透射电子显微镜、选区电子衍射和拉曼微光谱比较了每种基因型随发育而变化的分泌期牙釉质超微结构。在 WT 牙釉质中观察到的特征棒状结构在新沉积的牙釉质中表现出无定形特征，随后在较老的牙釉质中转化为磷灰石样晶体。令人惊讶的是，在 KO 牙釉质中发现初始矿化形成的方式与 WT 相同。然而，在棒状结构开始形成后不久，在发育中的 KO 牙釉质层中随机出现大的板状晶体。随着发育的继续，观察到的板状晶体变得占主导地位，并掩盖了牙釉质棒状结构的外观。当这些板状晶体形成后，KO 牙釉质层停止增厚，而 WT 和 HET 牙釉质层仍以相同的速度继续生长。拉曼结果表明，与 WT 牙釉质不同，Mmp20-KO 牙釉质含有相当一部分八钙磷酸盐。尽管在所有其他方面都是正常的，但在分泌型 HET 牙釉质中观察到大的、随机分散的矿物质晶体，尽管程度低于 KO 牙釉质，但这表明 MMP20 表达水平对抑制异常矿化形成有比例效应。总之，我们发现 MMP20 对细胞外牙釉质基质蛋白的蛋白水解作用对于成釉细胞分泌期牙釉质早期棒状结构的初始发育不是必需的。然而，MMP20 的蛋白水解作用对于防止成釉过程中异常晶体形成是必不可少的。
