Anisotropic properties of the enamel organic extracellular matrix

Enamel biosynthesis is initiated by the secretion, processing, and self-assembly of a complex mixture of proteins. This supramolecular ensemble controls the nucleation of the crystalline mineral phase. The detection of anisotropic properties by polarizing microscopy has been extensively used to detect macromolecular organizations in ordinary histological sections. The aim of this work was to study the birefringence of enamel organic matrix during the development of rat molar and incisor teeth. Incisor and molar teeth of rats were fixed in 2% paraformaldehyde/0.5% glutaraldehyde in 0.2 M phosphate-buffered saline (PBS), pH 7.2, and decalcified in 5% nitric acid/4% formaldehyde. After paraffin embedding, 5- µ m-thick sections were obtained, treated with xylene, and hydrated. Form birefringence curves were obtained after measuring optical retardations in imbibing media, with different refractive indices. Our observations showed that enamel organic matrix of rat incisor and molar teeth is strongly birefringent, presenting an ordered supramolecular structure. The birefringence starts during the early secretion phase and disappears at the maturation phase. The analysis of enamel organic matrix birefringence may be used to detect the effects of genetic and environmental factors on the supramolecular orientation of enamel matrix and their effects on the structure of mature enamel.     

Fluoride-binding by the organic matrix of developing bovine enamel.

Newly deposited enamel was obtained from foetal bovine incisors and demineralized by flow dialysis against 0.1 M EDTA. Samples of the demineralized enamel matrix were subjected to equilibrium dialysis for 24 h against a 0.2 M Nacl solution containing 100 or 200 parts/10⁶ fluoride. The results showed that 40.7 ± 1.2 μg F was bound per mg of demineralized matrix. Samples of freeze-dried, demineralized matrix were dissolved in saline containing carrier-free ¹⁸F^? and chromatographed on a BioGel P-150 column. Most of the bound ¹⁸F activity was identified with the second, and largest, protein peak. The results suggest that a partial explanation for the relatively high concentrations of fluoride in newly deposited enamel matrix is a capacity of early enamel matrix to selectively bind significant amounts of F^?.     

Calcium binding by organic matrix of developing bovine enamel

Flow-rate dialysis was used to define the Ca-binding parameters of demineralized fetal bovine enamel matrix in the presence and absence of physiologic concentrations of F. There were 5.45 × 10^?5 mol/g protein of high-affinity Ca-binding sites with a dissociation constant of 1.95 × 10^?6 mol/l. The presence of F had no effect on these parameters. The binding of Ca by the organic matrix may be important in heterogeneous nucleation of the mineral phase in developing enamel.     

Immunoblotting demonstration of antibodies to the organic matrix of developing bovine enamel

Antiserum was raised against an acetic acid extract of partly mineralized bovine enamel. By use of immunoblotting it was shown that rabbit antibodies react with both high and low molecular weight proteins. Most but not all of the enamel proteins isolated by different extraction solutions were antigenic identical. The blotting time was found to be important when immunoblotting of enamel proteins is carried out.     

Absence of in-vitro fluoride-binding by the organic matrix of developing bovine enamel

The in-vitro binding of fluoride to isolated organic matrix of secretory bovine enamel was studied by direct fluoride measurement and equilibrium dialysis. Over a wide range of protein and fluoride concentrations there was no indication of fluoride binding by the matrix in contrast to earlier reports.     

Immunoblotting demonstration of antibodies to the organic matrix of developing bovine enamel

Antiserum was raised against an acetic acid extract of partly mineralized bovine enamel. By use of immunoblotting it was shown that rabbit antibodies react with both high and low molecular weight proteins. Most but not all of the enamel proteins isolated by different extraction solutions were antigenic identical. The blotting time was found to be important when immunoblotting of enamel proteins is carried out.     

Comparison of demineralization of different organic acid to enamel]

The rates of demineralization of 5 organic acids (mathanoic acid, formic acid, propionic acid, Lactic acid, acetic acid, mixed acid) to the bovine enamel were tested and analysed with the self-made calcium ionselective microelectrodes(Ca(2+)-ISME) basing on a neutral carriers of ETH1001. The results showed; 1. The difference between the rates of demineralization of formic acid and lactic acid, formic acid and propionic acid, formic acid and acetic acid, acetic acid and mixed acid, acetic acid and lactic acid, propionic acid and mixed acid, propionic acid and lactic acid, lactic acid and mixed acid were of great significance (P < 0.01); 2. The rates of demineralization of acetic and mixed acid decreased with time, due to saturation of the solution during demineralization; 3. Ca(2+)-ISME was of the advantages of simplicity, rapidity, sensitivity and accuracy. The results suggest that the cariogenic potential is related to different acid products of different cariogenic bacteria, and the degree of mineral saturation within solution affects the rate of demineralization.     

Effects of Galla chinensis on inhibition of demineralization of regular bovine enamel or enamel disposed of organic matrix

Objective

This in vitro study was undertaken to assess the effects of Galla chinensis extract on inhibition of enamel caries-like demineralization and to elucidate the role of the organic matrix of enamel in this process.

Design

Either regular or enamel disposed of its organic matrix both of bovine origin were exposed to a demineralizing solution for 3 days (pH 4.5). Specimes were additionally treated with either 4 g/L of G. chinensis extraction (GCE) or double distilled water (DDW) four times daily for 5 min each time. Regular enamel exposed accordingly to sodium fluoride (1 g/L) during the demineralizing period served as positive control. After exposure mineral loss and lesion depth of all samples were analysed by transversal microradiography. One-way ANOVA and Student-Newman-Keuls test were used to compare the differences amongst groups. A factorial ANOVA was chosen to test the interaction between GCE and enamel organic matrix.

Results

Mineral loss and lesion depth of specimens in the positive control group were significantly lower compared to all other groups. Regular enamel treated with GCE showed significantly lower values compared to regular enamel treated with DDW or to enamel disposed of its organic matrix (p < 0.05). These three groups revealed similar values (p > 0.05). Significant interaction between GCE and enamel organic matrix with respect to both outcomes could be observed (p < 0.05).

Conclusions

G. chinensis inhibits enamel caries-like demineralization in vitro. However, its potential seems to be weaker compared to sodium fluoride. The organic matrix of enamel was shown to play a substantial role in the observed mechanism.     

The physiological and pathological role of some organic dentine and enamel structures

The enamel is the toughest human tissue. The major component of the inorganic part is hydroxyl apatite (90-92%). The organic part of enamel is formed by proteins, proteoglycans and lipoids, and represents only 1-2% of the entire weight. The organic components are organized, forming histological structures like enamel lamellae, enamel rods sheaths, enamel spindles and tufts. The authors, with the aid of the scanning electron microscope and histochemical staining, have demonstrated that enamel lamellae presented a true histological structure, contrary to some opinions that consider this entities developmental failures or simple cracks. In the opinion of the authors, these lamellae confer elasticity to the enamel when exposed to lateral or tangential forces, or even torque. The lamellae are also considered pathways for the progression of dental caries. The dentine-enamel junction is another non-mineralized tooth-structure which functions like an elastic support for the tough enamel, opposing unfortunately a very low resistance in the face of dental caries progression. In such cases we talk about secondary enamel caries. The dentinal tubules with the organic structures inside are essential in maintaining the vitality of the dentine and enamel; but in pathological circumstances they represent pathways for pathological stimuli heading toward the pulp, and they are the weakest points in front of caries progression.