In-vitro Remineralization of Human Enamel with Amelogenin-derived Peptide ADP5

Objectives: Incorporating biomimetic mineral layer to the molecular structure of enamel has been a long-standing challenge. Aim of this in-vitro study was to evaluate ADP5-guided remineralization of enamel and ability to control the biomimetic mineral layer properties.
Methods: Extracted healthy human molars were used to create white spot lesions on enamel with acetic acid. Samples were incubated in a 15-AA long ADP5 amelogenin-derived peptide solution for 10 mins at 37°C. Samples were treated with Ca²⁺/PO4^-3 for 1 h at 37°C (Group 1). Samples treated with Ca²⁺/PO4^-3 without ADP5 served as positive controls (Group 2). Enamel without any treatment were used as negative control. Scanning electron microscope (SEM) was used to evaluate the morphology and thickness of new mineral layer. Energy dispersive X-ray spectroscopy used to measure chemical composition of new layer. Structural characterization was done by transmission electron microscopy (TEM). Nanoindentation measurements and microhardness testing were performed to assess mechanical properties. The average values and standard deviations were calculated and expressed as mean ± standard error.

Results: SEM images showed continuous layer of plate-like crystals growing from the surface of the underlying enamel lesion in Group 1 with a 10μm-thick continuous remineralized layer. Ca²⁺/PO4^-3clusters without substantial remineralization were observed in Group 2. TEM showed hydroxyapatite (HAp) crystals of 30− 50 nm in healthy enamel tissue which correspond to plate-shape mineral found in Group 1. Elemental analysis of the samples from Group 1 revealed Ca/P ratio of 1.54±0.12. The microhardness and nanomechanical revealed the values between that of untreated enamel and dentin in Group 1.
Conclusions: This study demonstrated crystalline mineral layer is formed on an artificially created lesion on human enamel in the presence of Ca²⁺/PO₄³⁻ions under physiologically viable conditions by using ADP5.