Mammalian enamel maturation: Crystallographic changes prior to tooth eruption

Abstract

Using the distal molar of a minipig as a model, we studied changes in the microstructural characteristics of apatite crystallites during enamel maturation (16-23 months of postnatal age), and their effects upon the mechanical properties of the enamel coat. The slow rate of tooth development in a pig model enabled us to reveal essential heterochronies in particular components of the maturation process. The maturation changes began along the enamel-dentine junction (EDJ) of the trigonid, spreading subsequently to the outer layers of the enamel coat to appear at the surface zone with a 2-month delay. Correspondingly, at the distal part of the tooth the timing of maturation processes is delayed by 3-5 month compared to the mesial part of the tooth. The early stage of enamel maturation (16-20 months), when the enamel coat is composed almost exclusively of radial prismatic enamel, is characterized by a gradual increase in crystallite thickness (by a mean monthly increment of 3.8 nm); and an increase in the prism width and thickness of crystals composed of elementary crystallites. The late stage of maturation (the last two months prior to tooth eruption), marked with the rapid appearance of the interprismatic matrix (IPM) during which the crystals densely infill spaces between prisms, is characterized by an abrupt decrease in microstrain and abrupt changes in the micromechanical properties of the enamel: a rapid increase in its ability to resist long-term load and its considerable hardening. The results suggest that in terms of crystallization dynamics the processes characterizing the early and late stage of mammalian enamel maturation represent distinct entities. In regards to common features with enamel formation in the tribosphenic molar we argue that the separation of these processes could be a common apomorphy of mammalian amelogenetic dynamics in general.

Fig 1. Mandibular dentition and molar eruption in subadult (SA) and adult (A) minipig representing the age span covered in this study.

Note the embryonic stage of m3 development (entirely covered by a vascularized dental sac) in subadult individuals.

Fig 2. Cross-section of the mesial part (protoconid-metaconid) of m3 (SEM) in (a) an immature (17-month-old) and (b) mature (30-month-old) minipig.

The immature tooth illustrates the heterotopy of enamel maturation: the simultaneous appearance of mature compact enamel close to the EDJ (1), a partly mature enamel segment (2), and outer immature enamel (3); EDJ = enamel-dentine junction. (c) longitudinal section of semimature m3 (20-month-old) individual with a macroscopic view of Hunter-Schreger bands (HSB) and outer aprismatic enamel.

Fig 3. Techniques of measuring prism width (a) and thickness of crystallite aggregates (b).

Fig 4. Crystallite thickness (a,c) and crystallite length (b,d) revealed by X-ray diffraction of the inner and outer enamel (a,b), and the mesial and distal enamel (c,d) plotted against the postnatal age of particular individuals (abscissa, lower) and day of m3 calcification (abscissa, upper)—mean values of particular individuals with SD error bars (mostly covered by the marks).

The blue band indicates the range of mature enamel crystallite size estimated both from our previous observations and from data by Daculsi and Kerebel [31], Daculsi et al. [58], and Simmer and Fincham [8]. The beginning of calcification (the zero point of the upper abscissa scale) is taken from Tonge and McCance [47] and Wang et al. [48]. The ovals indicate sampling areas for either inner and outer enamel and the mesial or distal tooth part, respectively.

Fig 5. Variation of microstrain during the maturation stage of calcification in (a) the inner and outer enamel parts, (b) the mesial and distal tooth parts.

Mean values of particular individuals with SD error bars (mostly covered by the marks).

Fig 6. Mechanical parameters (a) H_IT, (b) E_IT, (c) η_IT, and (d) C_IT and their relationships to enamel formation.

Each point represents an average value calculated from 10 microindentation points. For d) the error function parameter a from the inner enamel part was set to the value of the same parameter for the outer enamel to prevent instability in the fit. Mean values of particular individuals with SD error bars (mostly covered by the signs).

Fig 7. A cross-section of the mesial part of m3 in a 17-month-old individual (a) and the structure of the enamel at different parts of the enamel coat (b-h), all at the same magnification, SEM, acid etched, scale bar 10 μm.

Note the apparent differences in enamel texture and prism width between the mature inner enamel (b-e) and the immature outer enamel (f-h). HSB—Hunter-Schregers bands.

Fig 8. Mean width of radial prisms (a) and mean thickness of core crystals (b) at particular distances from the EDJ measured from SEM images (2000x magnification).

Each point represents an average value calculated from at least 30 measurements, in some cases up to 240 measurements, with SD error bars.

Fig 9. Optical macrophotography of the post-talonid part of the m3 crown in a 20-month-old individual.

Note the dense arrangement of prisms, the absence of IPM and APE, and the structural fissures among compact blocks of prisms, partly infilled by a non/mineralized compound (red arrowheads). Natural surface, no polishing, no acid etching, gold coating. Blue arrowhead: isolated IPM aggregates at inner part of enamel prisms.

Fig 10. Enamel microstructure at identical loci of the mesial part of m3 in 16- and 17-month-old individuals (SEM, no acid etching).

Note the complete development of dense IPM at the inner part of the crown while only the first signs of IPM appear at the outer part.

Fig 11. SEM images of selected tooth parts of a 17-month-old individual showing the transition zone between dentine and enamel (EDJ) and similarities between the shape and size of the dentine and enamel tubules (acid etched).