Post-Eruptive Changes in Protein Content Concur With Fast Enamel Hardening

Objectives: Enamel formation in pigs is far shorter than in humans. Consequently, pig enamel is less mineralized with more protein at eruption, similar to hypomineralized human enamel. Yet, it hardens quickly after eruption without enzyme secretion by ameloblasts. The change in enamel proteome that concurs with post-eruptive maturation is not well understood. This study aimed to characterize the changes in porcine enamel proteome to elucidate post-eruptive enamel hardening.
Methods: Animals at three different ages (n=3 animals/age) were used to study the same tooth type at eruption (PE-0), two weeks and six weeks post-eruption (PE-2 and PE-6). Whole saliva was collected, and animals were sacrificed and exsanguinated to avoid blood contamination. Erupted enamel was collected using slow-speed burs. Following in-gel tryptic digestion, samples were analyzed by LC-MS/MS.
Results: The number of proteins in enamel decreases from 1035 at PE-0 to 456 at PE-2, and 250 in PE-6 enamel. The difference in protein composition was biggest between PE-0 and PE-2, followed by the difference between PE-0 and PE-6. 473 proteins were common between the enamel and saliva of animals at PE-0. This number decreases with post-eruptive time to 243 in PE-2 and 138 in PE-6. Unique peptides from enamel proteins decrease after eruption, with a mean number of 17 in PE-0, 5 in PE-2, and 11 in PE-6. The number of unique peptides remains equal from PE-0 to PE-6 for KLK4 (3 peptides), decreases fast for MMP20 (from 22 to 12) and less for serum albumin in enamel (from 27 to 20).
Conclusions: Pig enamel rapidly loses protein abundance within two weeks after eruption, including serum albumin, which has been reported in hypomineralized human enamel. This model shows that acellular protein removal is effective for fast enamel hardening in porcine teeth. This study highlights the translational potential for treating hypomineralized human enamel.