No abstract

Tooth enamel is a unique mineralized tissue in that it is acellular, is more highly mineralized, and is comprised of individual crystallites that are larger and more oriented than other mineralized tissues. Dental enamel forms by matrix-mediated biomineralization. Enamel crystallites precipitate from a supersaturated solution within a well-delineated biological compartment. Mature enamel crystallites are comprised of non-stoichiometric carbonated calcium hydroxyapatite. The earliest crystallites appear suddenly at the dentino-enamel junction (DEJ) as rapidly growing thin ribbons. The shape and growth patterns of these crystallites can be interpreted as evidence for a precursor phase of octacalcium phosphate (OCP). An OCP crystal displays on its (100) face a surface that may act as a template for hydroxyapatite (OHAp) precipitation. Octacalcium phosphate is less stable than hydroxyapatite and can hydrolyze to OHAp. During this process, one unit cell of octacalcium phosphate is converted into two unit cells of hydroxyapatite. During the precipitation of the mineral phase, the degree of saturation of the enamel fluid is regulated. Proteins in the enamel matrix may buffer calcium and hydrogen ion concentrations as a strategy to preclude the precipitation of competing calcium phosphate solid phases. Tuftelin is an acidic enamel protein that concentrates at the DEJ and may participate in the nucleation of enamel crystals. Other enamel proteins may regulate crystal habit by binding to specific faces of the mineral and inhibiting growth. Structural analyses of recombinant amelogenin are consistent with a functional role in establishing and maintaining the spacing between enamel crystallites.

Enamelysin is a tooth-specific matrix metalloproteinase that is expressed during the early through middle stages of enamel development. The enamel matrix proteins amelogenin, ameloblastin, and enamelin are also expressed during this same approximate developmental time period, suggesting that enamelysin may play a role in their hydrolysis. In support of this interpretation, recombinant enamelysin was previously demonstrated to cleave recombinant amelogenin at virtually all of the precise sites known to occur in vivo. Thus, enamelysin is likely an important amelogenin-processing enzyme. To characterize the in vivo biological role of enamelysin during tooth development, we generated an enamelysindeficient mouse by gene targeting. Although mice heterozygous for the mutation have no apparent phenotype, the enamelysin null mouse has a severe and profound tooth phenotype. Specifically, the null mouse does not process amelogenin properly, possesses an altered enamel matrix and rod pattern, has hypoplastic enamel that delaminates from the dentin, and has a deteriorating enamel organ morphology as development progresses. Our findings demonstrate that enamelysin activity is essential for proper enamel development.Dental enamel covers the crown of the tooth and is unique among mineralized tissues because of its high mineral content, large crystals, and organized prism pattern. Other mineralized tissues such as bone, dentin, and cementum are composed of ϳ20% organic material. In contrast, mature enamel has less than 1% organic matter by weight (1, 2). Moreover, enamel crystallites possess a volume that is 100 times greater than the volume of crystallites found in other mineralizing tissues. These enamel crystallites form enamel rods that, in turn, form a unique interlacing (decussating) prism pattern. As a result, dental enamel is the hardest substance in the body. Its hardness is intermediate between that of iron and carbon steel, yet it also has a high elasticity (3).Although mature enamel is a very hard protein-free tissue, it does not start this way. Enamel development (amelogenesis) consists of several stages that include the secretory, transition, and maturation stages. During the secretory stage, enamel crystallites elongate into long thin ribbons that are only a few apatitic unit cells in thickness (about 10 nm) with a width of ϳ30 nm (4, 5). The ribbons are evenly spaced, are oriented parallel to each other, and grow in length but very little in width and thickness. Ultimately, enamel crystal length determines the final thickness of the enamel layer as a whole (for review, see Ref. 6). It is during the secretory stage that the columnar-shaped ameloblast cells, located adjacent to the forming enamel, secrete specialized enamel proteins into the enamel matrix. These proteins include amelogenin (7), ameloblastin (8), and enamelin (9). Amelogenin is the predominant component and comprises ϳ90% of total enamel matrix protein (10). Interestingly, the full-length enamel proteins are found only at the mineralizing front, su...

For almost three decades, proteinases have been known to reside within developing dental enamel. However, identification and characterization of these proteinases have been slow and difficult, because they are present in very small quantities and they are difficult to purify directly from the mineralizing enamel. Enamel matrix proteins such as amelogenin, ameloblastin, and enamelin are cleaved by proteinases soon after they are secreted, and their cleavage products accumulate in the deeper, more mature enamel layers, while the full-length proteins are observed only at the surface. These results suggest that proteinases are necessary for "activating" enamel proteins so the parent proteins and their cleavage products may perform different functions. A novel matrix metalloproteinase named enamelysin (MMP-20) was recently cloned from tooth tissues and was later shown to localize primarily within the most recently formed enamel. Furthermore, recombinant porcine enamelysin was demonstrated to cleave recombinant porcine amelogenin at virtually all of the sites that have previously been described in vivo. Therefore, enamelysin is at least one enzyme that may be important during early enamel development. As enamel development progresses to the later stages, a profound decrease in the enamel protein content is observed. Proteinases have traditionally been assumed to degrade the organic matrix prior to its removal from the enamel. Recently, a novel serine proteinase named enamel matrix serine proteinase-l (EMSPI) was cloned from enamel organ epithelia. EMSP1 localizes primarily to the early maturation stage enamel and may, therefore, be involved in the degradation of proteins prior to their removal from the maturing enamel. Other, as yet unidentified, proteinases and proteinase inhibitors are almost certainly present within the forming enamel and await discovery.

Dental enamel is the epithelial-derived hard tissue covering the crowns of teeth. It is the most highly mineralized and hardest tissue in the body. Dental enamel is acellular and has no physiological means of repair outside of the protective and remineralization potential provided by saliva. Enamel is comprised of highly organized hydroxyapatite crystals that form in a defined extracellular space, the contents of which are supplied and regulated by ameloblasts. The entire process is under genetic instruction. The genetic control of amelogenesis is poorly understood, but requires the activities of multiple components that are uniquely important for dental enamel formation. Amelogenesis imperfecta (AI) is a collective designation for the variety of inherited conditions displaying isolated enamel malformations, but the designation is also used to indicate the presence of an enamel phenotype in syndromes. Recently, genetic studies have demonstrated the importance of genes encoding enamel matrix proteins in the etiology of isolated AI. Here we review the essential elements of dental enamel formation and the results of genetic analyses that have identified disease-causing mutations in genes encoding enamel matrix proteins. In addition, we provide a fresh perspective on the roles matrix proteins play in catalyzing the biomineralization of dental enamel.

Epithelial-mesenchymal interactions guide tooth development through its early stages and establish the morphology of the dentin surface upon which enamel will be deposited. Starting with the onset of amelogenesis beneath the future cusp tip, the shape of the enamel layer covering the crown is determined by five growth parameters. Appositional growth occurs at a mineralization front along the ameloblast distal membrane in which amorphous calcium phosphate (ACP) ribbons form and lengthen. The ACP ribbons convert to calcium hydroxyapatite as the ribbons elongate. Appositional growth involves a secretory cycle that leaves an imprint of incremental lines. A potentially important function of enamel proteins is to ensure alignment of successive mineral increments on the tips of enamel ribbons deposited in the previous cycle so the crystallites lengthen with each cycle. Enamel crystallites harden in a maturation process that involves mineral deposition on the sides of existing crystallites until they interlock with adjacent crystallites. Neutralization of acidity generated by hydroxyapatite formation is a key part of the mechanism. Here we review the growth parameters that determine the shape of the enamel crown as well as the mechanisms of enamel appositional growth and maturation.

By the Shields classification, articulated over 30 years ago, inherited dentin defects are divided into 5 types: 3 types of dentinogenesis imperfecta (DGI), and 2 types of dentin dysplasia (DD). DGI type I is osteogenesis imperfecta (OI) with DGI. OI with DGI is caused, in most cases, by mutations in the 2 genes encoding type I collagen. Many genes are required to generate the enzymes that catalyze collagen's diverse post-translational modifications and its assembly into fibers, fibrils, bundles, and networks. Rare inherited diseases of bone are caused by defects in these genes, and some are occasionally found to include DGI as a feature. Appreciation of the complicated genetic etiology of DGI associated with bony defects splintered the DGI type I description into a multitude of more precisely defined entities, all with their own designations. In contrast, DD-II, DGI-II, and DGI-III, each with its own pattern of inherited defects limited to the dentition, have been found to be caused by various defects in DSPP (dentin sialophosphoprotein), a gene encoding the major non-collagenous proteins of dentin. Only DD-I, an exceedingly rare condition featuring short, blunt roots with obliterated pulp chambers, remains untouched by the revolution in genetics, and its etiology is still a mystery. A major surprise in the characterization of genes underlying inherited dentin defects is the apparent lack of roles played by the genes encoding the less-abundant non-collagenous proteins in dentin, such as dentin matrix protein 1 (DMP1), integrin-binding sialoprotein (IBSP), matrix extracellular phosphoglycoprotein (MEPE), and secreted phosphoprotein-1, or osteopontin (SPP1, OPN). This review discusses the development of the dentin extracellular matrix in the context of its evolution, and discusses the phenotypes and clinical classifications of isolated hereditary defects of tooth dentin in the context of recent genetic data respecting their genetic etiologies.

Enamel is a highly organized hierarchical nanocomposite, which consists of parallel arrays of elongated apatitic crystallites forming an intricate three-dimensional microstructure. Amelogenin, the major extracellular matrix protein of dental enamel, regulates the formation of these crystalline arrays via cooperative interactions with forming mineral phase. Using cryoelectron microscopy, we demonstrate that amelogenin undergoes stepwise hierarchical self-assembly. Furthermore, our results indicate that interactions between amelogenin hydrophilic C-terminal telopeptides are essential for oligomer formation and for subsequent steps of hierarchical self-assembly. We further show that amelogenin assemblies stabilize mineral prenucleation clusters and guide their arrangement into linear chains that organize as parallel arrays. The prenucleation clusters subsequently fuse together to form needle-shaped mineral particles, leading to the formation of bundles of crystallites, the hallmark structural organization of the forming enamel at the nanoscale. These findings provide unique insight into the regulation of biological mineralization by specialized macromolecules and an inspiration for bottom-up strategies for the materials design.