Abstract
Matrix metalloproteinase-20 (MMP20) is expressed by ameloblasts in developing teeth and MMP20 mutations cause enamel malformation. We established a stably transfected Tet-Off Mmp20-inducible ameloblast-lineage cell line and found that MMP20 expression promoted cell invasion. Previously, we engineered transgenic mice (Tg) that drive Mmp20 expression and showed that Mmp20+/+Tg mice had soft enamel. Here we asked if Mmp20 overexpression disrupts ameloblast function. Incisors from Mmp20+/+ mice expressing the Mmp20 Tg had a striking cell infiltrate which nearly replaced the entire enamel layer. A thin layer of enamel-like material remained over the dentin and at the outer tooth surface, but between these regions were invading fibroblasts and epithelial cells that surrounded ectopic bone-like calcifications. Mmp20+/+Tg mice had decreased enamel organ cadherin levels compared to the Mmp20 ablated and WT mice and, instead of predominantly locating adjacent to the ameloblast cell membrane, β-catenin was predominantly present within the nuclei of invading cells. Our data suggest that increased cadherin cleavage by transgenic MMP20 in the WT background releases excess β-catenin, which translocates to ameloblast nuclei to promote cell migration/invasion. Therefore, we conclude that MMP20 plays a role in normal ameloblast migration through tightly controlled Wnt signaling and that MMP20 overexpression disrupts this process.
Highlights
Matrix metalloproteinase-20 (MMP20) is expressed by ameloblasts in developing teeth and MMP20 mutations cause enamel malformation
MMP20 is expressed in teeth[2,3,4,5] and the only non-overlapping function of MMP20 is in enamel formation[6]
When we added a protease inhibitor cocktail to the culture medium, MMP20 was detectable by zymography (Fig. 1b) and by immunoblotting with an antisera specific for the engineered hemagglutinin epitope tag (HA) tag at the C-terminus of the MMP20 protein (Fig. 1c). quantitative real-time PCR (qPCR) analysis of membrane-type-1 Matrix metalloproteinases (MMP) (MT1-MMP, Mmp14) expression served as the negative control for Mmp[20] inducible expression (Fig. 1d)
Summary
Matrix metalloproteinase-20 (MMP20) is expressed by ameloblasts in developing teeth and MMP20 mutations cause enamel malformation. Incisors from Mmp20+/+ mice expressing the Mmp[20] Tg had a striking cell infiltrate which nearly replaced the entire enamel layer. Mmp20+/+Tg mice had decreased enamel organ cadherin levels compared to the Mmp[20] ablated and WT mice and, instead of predominantly locating adjacent to the ameloblast cell membrane, β-catenin was predominantly present within the nuclei of invading cells. Ameloblasts secrete enamel matrix proteins as thin crystallites are induced to grow out from the dentin surface. The crystallites are at their full length and the ameloblasts change into shorter maturation stage cells that typically modulate between ruffle- and smooth-ended morphologies and actively reabsorb the extracellular enamel matrix proteins and their fragments. Part to help neutralize or remove acid stress generated as the enamel crystallites expand in volumetric size during maturation[15]
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