Changes in the Proteomic Profile of Acquired Enamel Pellicles as a Function of Their Time of Formation and Hydrochloric Acid Exposure.

OBJECTIVE: Changes in the protein profile of acquired enamel pellicles (AEP) formed in vivo over different time periods were evaluated after the application of hydrochloric acid (HCl). METHODS: Nine subjects were submitted to dental prophylaxis with pumice. After 3 or 120 min, the teeth were isolated with cotton rolls and 50 µL of 0.1 M HCl (pH 1.0), 0.01 M HCl (pH 2.0), or deionized water were applied on the buccal surface of the teeth for 10 s. The AEP was then collected using an electrode filter paper presoaked in 3% citric acid. After protein extraction, the samples were submitted to reverse-phase liquid chromatography coupled to mass spectrometry (nano LC-ESI-MS/MS). Label-free quantification was performed (Protein Lynx Global Service software). RESULTS: A total of 180 proteins were successfully identified in the AEP samples. The number of identified proteins increased with the time of pellicle formation. Only 4 proteins were present in all the groups (isoforms of IgA, serum albumin, and statherin). The greatest number of proteins identified uniquely in one of the groups was obtained for the groups treated with HCl after 2 h of pellicle formation (approx. 50 proteins). CONCLUSION: Proteins resistant to removal by HCl, such as serum albumin and statherin, were identified even in the short-term AEP. In addition, 120-min pellicles present many proteins that are resistant to removal by HCl. This suggests an increase in protection against intrinsic acids with the time of pellicle formation, which should be evaluated in future studies.

Combined Effect of Enamel Deproteinization and Intermediate Bonding in the Retention of Pit and Fissure Sealants in Children: A Randomized Clinical Trial.

OBJECTIVE: This manuscript shows if enamel deproteinization along with an intermediate layer of bonding enhances the retention of pit and fissure sealants.. STUDY DESIGN: Two hundred six mandibular first permanent molars were allocated to Group I (n=103) and Group II (n=103). Group I underwent deproteinization, acid etching, bonding agent application and pit & fissure sealant placement while Group II treated with acid etching followed by pit & fissure sealant application only. Clinical analysis of all the teeth in the two groups was performed at 1, 3, 6, 9 and 12 months respectively. Pearson's chi - square test was utilized to evaluate the success of both treatment procedures (p<0.05). RESULTS: At 12 months follow up the differences between the groups pertaining to Marginal integrity, Marginal discoloration and Anatomical form were statistically significant suggesting enhanced retention in Group I. CONCLUSIONS: Enamel deproteinization along with the use of intermediate bonding layer significantly enhances the retention of pit and fissure sealants in terms of enhanced marginal integrity, decreased marginal discoloration and preserving the anatomical form.

Deproteinization Effectiveness on Occlusal Enamel Surfaces and Resultant Acid Etching Patterns: An in vitro Study.

PURPOSE: The goal of this in vitro study was to identify whether occlusal enamel deproteinization is effective in the removal of organic material in order to obtain quality etching patterns using phosphoric acid (H3PO4) and sodium hypochlorite (NaOCl) compared to phosphoric acid alone. STUDY DESIGN: Nine extracted third permanent molars were polished with pumice and water. Every pit and fissure was evaluated as a unit, resulting in 40 individual units and then these were divided into five treatment groups. The occlusal enamel surface of each group was subjected to the following treatments: Group 1 (C) Control: No treatment; Group 2 (P): Polish and rinse; Group 3 (PD): Polish, rinse, and sodium hypochlorite (NaOCl) 5.25% for 60 seconds; Group 4 (PA): Polish, rinse, and acid etching with H3PO4 37% for 15 seconds; and Group 5 (PDA): Polish, rinse, sodium hypochlorite (NaOCl) 5.25% for 60 seconds, and acid etching with H3PO4 37% for 15 seconds. Results showed no significant statistical difference in the organic material present between groups 1 (C) (30.18%) and 2 (P) (36.61%), but there was a statistical difference (p <0.002) between Groups 1 and 2, and Group 3 (PD) (16.50%). In the acid etching group, the undesirable Type-III pattern (discussed later) was found in Group 4 (PA) (33.54%), while this was only 7.70% in Group 5, nearly five times more than Group 4, with a significant statistical difference (0.05). When differences were sought for Types I and II etch patterns (discussed later) for Groups 4 and 5, Group 4 (PA) obtained 26.29% (Type I) and 1.75% (Type II) etch patterns, compared to Group 5 (PDA) with 33.4% (Type I) and 38.97% (Type II) etch patterns. CONCLUSIONS: The enamel deproteinization technique is an effective way to remove organic material on the occlusal surfaces of teeth, obtaining after phosphoric acid application, up to 72.38% of Types I and II etch patterns. Etching Types I or II can also be determined by the removal of organic material in between enamel crystals.

Effect of deproteinization before and after acid etching on the surface roughness of immature permanent enamel.

OBJECTIVE: The purpose of this in vitro investigation was to assess the effect of deproteinization before and after acid etching on the surface roughness of immature human enamel of permanent teeth compared to acid etching alone using noncontact three-dimensional (3D) optical profilometer. MATERIALS AND METHODS: Forty-eight enamel blocks were randomly distributed into 4 groups (12 each) according to the surface treatment in the form of deproteinized with 2.5% sodium hypochlorite (NaOCl) before and after acid etching with 32% phosphoric acid (H3PO4) compared to application of H3PO4 alone. The surface roughness (Sa) was measured using a 3D optical noncontact surface profiler. Two specimens from each group were selected and prepared for scanning electron microscopic (SEM) analysis. Shapiro-Wilk test, one-way analysis of variance, and Tukey's honest significance difference test were used. All statistical analyses were established with a significance level of P < 0.05. RESULTS: The highest surface roughness (Sa) was recorded for Group 3/NaOCl ± H3PO4 and the lowest Sa was recorded for Group 1 (control). All surface treatments applied showed significantly greater values of surface roughness (Sa) than the enamel surfaces with no surface treatment (control). There was significant difference between control group and Group 2/H3PO4 (P = 0.002), Group 3/NaOCl ± H3PO4 (P = 0.0001), and Group 4/H3PO4 ± NaOCl (P = 0.017). There was no significant difference between Group 2/H3PO4 and Group 4/H3PO4 ± NaOCl. SEM evaluation showed different topographical features of deproteinized enamel surface. CONCLUSIONS: Deproteinizing the enamel of immature permanent teeth with 2.5% NaOCl before and after acid etching with 32% H3PO4 increased surface roughness compared to the application of H3PO4 alone.

A Clinical Evaluation of Deproteinization and Different Cavity Designs on Resin Restoration Performance in MIH-Affected Molars: Two-Year Results.

BACKGROUND: The aim of this study was to evaluate the clinical effects of deproteinization of the hypomineralized enamel and different cavity designs on the performance of the composite resin restorations(CRRs) placed into the cavities of MIH (molar incisor hypomineralization)-affected molars. STUDY DESIGN: 95 MIH-affected permanent first molars (PFMs) and 31 caries but not MIH-affected PFMs (126 teeth in total) were included in the study. The MIH-affected molars were divided into three groups. In Group I, all hypomineralized tissue was removed until healthy enamel was reached. In Group II, carious and cheesy hypomineralized tissue was removed until a reasonable resistance was detected in the hypomineralized tissue. In Group III, cavities designed as Group II, differently from this group deproteinization of the left hypomineralized tissue was performed prior to the placement of CRRs. Group IV served as the control group consisting of unaffected carious PFMs. Restorations were evaluated according to modified USPHS criteria for 24 months. RESULTS: The retention rates were 93.7% for Group I, 80.7% for Group II, 93.5% for Group III and 100% for Group IV. The success rate for the restorations in Group II proved significantly lower (p<0.05) than that of the other three groups. No significant difference in success rates was observed between Group I, Group III and Group IV (p>0.05) at the end of 24 months. CONCLUSIONS: Failure of the restorations was predominant in the group that the hypomineralized tissue was left surrounding the cavities. Deproteinization of the hypomineralized enamel was found to enhance the retention rates of CRRs.

TLR signalling can modify the mineralization of tooth germ.

OBJECTIVE: The aim of this work is to investigate the possible role of Toll-like receptor 4 (TLR4) during the development of mouse tooth germ. TLR4 is well known to inhibit mineralization and cause inflammation in mature odontoblasts and dental pulp cells. However, unlike these pathological functions of TLR4, little is known about the developmental function(s) of TLR4 during tooth development. MATERIALS AND METHODS: TLR4 expression was studied via Western blot in developing lower mouse incisors from E13.5 to E18.5. To generate functional data about the effects of TLR4, a specific agonist (LPS) was applied to the medium of in vitro tooth germ cultures, followed by Western blot, histochemical staining, ELISA assay, in situ hybridization and RT-qPCR. RESULTS: Increased accumulation of biotin-labelled LPS was detected in the enamel organ and in preodontoblasts. LPS treatment induced degradation of the inhibitor molecule (IκB) of the NF-κB signalling pathway. However, no morphological alterations were detected in cultured tissue after LPS addition at the applied dosage. Activation of TLR4 inhibited the mineralization of enamel and dentin, as demonstrated by alizarin red staining and as decreased levels of collagen type X. mRNA expression of ameloblastin was elevated after LPS administration. CONCLUSION: These results demonstrate that TLR4 may decrease the mineralization of hard tissues of the tooth germ and may trigger the maturation of ameloblasts; it can give valuable information to understand better congenital tooth abnormalities.

Amoxicillin diminishes the thickness of the enamel matrix that is deposited during the secretory stage in rats.

BACKGROUND: The use of amoxicillin during early childhood has been associated with molar incisor hypomineralization. AIM: The objective of this study was to determine whether the use of amoxicillin interferes with enamel development, during secretion and early mineralization stages. DESIGN: Fifteen pregnant rats were randomly assigned to three groups that received physiological solution (sham group), 100 mg/kg/day amoxicillin (A100G), and 500 mg/kg/day amoxicillin (A500G). After birth, the pups in each group received the same treatment until post-natal day 7 or 12. The upper first molars were analyzed histomorphometrical and immunostaining with amelogenin on day 7, and MMP-20 on day 12 was performed using a semiquantitative method (H-score). RESULTS: At 7 days, several vacuolar structures were observed in the ameloblasts in the A100G and A500G groups. A significant reduction of the enamel thickness (P < 0.001) was found in amoxicillin-treated rats compared with the sham group. Significant differences were not observed in enamel thickness (P > 0.05) between the groups of 12-day-old rats. Moreover, significant differences were not observed in the number of amelogenin- and MMP-20-immunolabeled ameloblasts (P > 0.05) between groups. CONCLUSION: The present results suggest that amoxicillin interferes with the initial stages of amelogenesis by causing structural changes in the ameloblasts and a reduction of the enamel matrix.

Effect of bromelain and papain gel on enamel deproteinisation before orthodontic bracket bonding.

AIM: To test the hypothesis that enamel surface deproteinisation with different concentrations of bromelain in association with 10% papain increases the shear bond strength (SBS) of brackets bonded with orthodontic composite and resin modified glass ionomer cement (RMGIC). MATERIALS AND METHODS: Orthodontic brackets were attached according to the following protocols to 195 bovine incisors, which were acquired and divided into 13 groups: 1) Transbond XT (TXT) according to the manufacturer's recommendations; 2) Deproteinisation with 3% bromelain (BD) plus 10% papain and TXT; 3) 6% BD plus 10% Papain and TXT; 4) RMGIC, without enamel deproteinisation and without acid etching; 5) RMGIC, with 3% BD plus 10% papain and without acid etching; 6) RMGIC, with 6% BD plus 10% papain and without acid etching; 7) attachment using RMGIC following etching with polyacrylic acid; 8) 3% BD plus 10% papain, attachment using RMGIC and etching with polyacrylic acid; 9) 6% BD plus 10% papain, and attachment using RMGIC following etching with polyacrylic acid; 10) etching with 37% phosphoric acid and attachment using RMGIC; 11) 3% BD plus 10% papain, etching with 37% phosphoric acid and attachment using RMGIC; 12) 6% BD plus 10% papain, etching with 37% phosphoric acid and attachment using RMGIC; 13) deproteinisation with 2.5% sodium hypochlorite (NaOCl), etching with polyacrylic acid and RMGIC. After bonding, the brackets were removed by a universal mechanical testing machine, which recorded shear bond strength at failure. The material remaining on the tooth was assessed using the adhesive remnant index (ARI). RESULTS: Deproteinisation with 3% and 6% bromelain gel plus papain significantly increased the shear bond strength (p < 0.05), when acid etching was performed with phosphoric acid, followed by primer application and attachment using Transbond XT (Group 3) and when attached with RMGIC without etching. Deproteinisation with 6% bromelain gel plus papain significantly increased (p < 0.05) the ARI score only when attachment was performed using RMGIC, without etching (Group 6). CONCLUSIONS: Deproteinisation with bromelain associated with papain in a gel increased the shear bond strength and is recommended before orthodontic bracket attachment.

Enamel Deproteinization using Papacarie and 10% Papain Gel on Shear Bond Strength of Orthodontic Brackets Before and After Acid Etching.

OBJECTIVE: To suggest Papacarie(®) as a new deproteinizing agent in comparison with indigenously prepared 10% papain gel before and after acid etching that may enhance the quality of the bond between enamel surface and composite resin complex. STUDY DESIGN: One hundred and twenty five extracted human premolars were utilized and divided into five groups: In the group 1, enamel surface was etched and primer was applied. In group 2, treatment with papacarie(®) for 60 seconds followed by etching and primer application. In group 3, etching followed by treatment with papacarie(®) for 60 seconds and primer application. In group 4, treatment with 10% papain gel for 60 seconds followed by etching and primer application. In group 5, etching followed by treatment with 10% papain gel for 60 seconds and primer application . After bonding the brackets, the mechanical testing was performed using a Universal testing machine. The failure mode was analyzed using an adhesive remnant index. The etching patterns before and after application of papacarie(®) and 10% papain gel was also evaluated using SEM. The values obtained for shear bond strength were submitted to analysis of variance and Tukey test (p < 0.05). RESULTS: It was observed that group 2 and group 4 had the highest shear bond strength and was statistically significant from other groups (p=0.001). Regarding Adhesive remnant index no statistical difference was seen between the groups (p=0.538). CONCLUSION: Papacarie(®) or 10% papain gel can be used to deproteinize the enamel surface before acid etching to enhance the bond strength of orthodontic brackets.

Excessive fluoride reduces Foxo1 expression in dental epithelial cells of the rat incisor.

Enamel fluorosis is characterized by hypomineralization, and forkhead box O1 (Foxo1) is essential for mouse enamel biomineralization. This study investigated the effect of fluoride on Foxo1 expression and its implications for enamel fluorosis. Mandibular incisors were extracted from Sprague Dawley rats treated for 3 months with water containing 0, 50, or 100 p.p.m. F⁻. Immunohistochemistry was used to localize and quantify FOXO1 expression in dental epithelial layer cells of the incisors. The effect of fluoride on expression of Foxo1, kallikrein-4 (Klk4), and amelotin (Amtn) mRNAs was analyzed by real-time RT-PCR, and western blotting was used to measure total and nuclear FOXO1 protein levels in mature dental epithelial cells. The results revealed that nuclear FOXO1 was mainly localized in the transition and the mature ameloblasts and exhibited weaker expression in the rats exposed to fluoride. In addition to the reduced levels of Foxo1, Klk4, and AmtnmRNAs, the protein levels of total and nuclearFOXO1 were decreased in the mature dental epithelial cells exposed to fluoride. Thus, excessive fluoride may have an effect on the expression levels of Foxo1 in dental epithelial cells and thereby affect hypomineralization of the enamel during fluorosis.

Acidulated phosphate fluoride application changes the protein composition of human acquired enamel pellicle.

We evaluated, by proteomic analysis, whether the chemical changes provoked on enamel by acidulated phosphate fluoride (APF) application alter the protein composition of acquired enamel pellicle. Enamel slabs, pretreated with distilled water (negative control), phosphoric acid (active control) or APF solution, were immersed in human saliva for pellicle formation. The adsorbed proteins were extracted and analyzed by liquid chromatography-mass spectrometry/mass spectrometry. Fifty-six proteins were identified, 12 exclusive to APF and 11 to phosphoric acid. APF decreased the concentration of histatin-1, but increased the concentration of S100-A9, which is confirmed by immunoblotting. The findings suggest that APF application changes the acquired enamel pellicle composition.

Deproteinization treatment on bond strengths of primary, mature and immature permanent tooth enamel.

OBJECTIVE: The aim of this study was to examine the effect of pre-post deproteinization treatment with 5% sodium hypochloride on shear bond strength (sbs) of adhesive resin to primary, immature and mature permanent teeth enamel. METHOD: 30 teeth were used for each of primary, immature and mature permanent teeth groups. (totally 90). In control groups, enamel was etched for 60s with 37% phosphoric acid (3M) and rinsed for 10s (Procedure A). In experimental groups, deproteinization was applied with 5% NaOCI solution for 120s before (Procedure D+A) and after acid-etching (Procedure A+D). Gluma Comfort Bond (Heraeus-Kulzer) and Charisma (Heraeus-Kulzer) composite resin were applied to etched enamel surfaces. Data were determined with Two-Way ANOVA and LSD Multiple Comparison Test (p < 0.05). RESULTS: SBS was significantly lower in primary and immature permanent teeth than mature permanent teeth (p < 0.05). "Procedure A+D" statistically increased sbs values in primary and immature permanent teeth (p < 0.05). CONCLUSION: Deproteinization after acid etching significantly enhanced the shear bond strength values in primary and immature permanent teeth.

Serotonin and fluoxetine receptors are expressed in enamel organs and LS8 cells and modulate gene expression in LS8 cells.

The selective serotonin re-uptake inhibitor (SSRI) fluoxetine is widely used in the treatment of depression in children and fertile women, but its effect on developing tissues has been sparsely investigated. The aim of this study was to investigate if enamel organs and ameloblast-derived cells express serotonin receptors that are affected by peripherally circulating serotonin or fluoxetine. Using RT-PCR and western blot analysis we found that enamel organs from 3-d-old mice and ameloblast-like cells (LS8 cells) express functional serotonin receptors, the rate-limiting enzyme in serotonin synthesis (Thp1), as well as the serotonin transporter (5HTT), indicating that enamel organs and ameloblasts are able to respond to serotonin and regulate serotonin availability. Fluoxetine and serotonin enhanced the alkaline phosphatase activity in the cell culture medium from cultured LS8 cells, whereas the expression of enamelin (Enam), amelogenin (Amel), and matrix metalloproteinase-20 (MMP-20) were all significantly down-regulated. The secretion of vascular endothelial growth factor (VEGF), monocyte chemotactic protein 1 (MCP-1), and interferon-inducible protein 10 (IP-10) was also reduced compared with controls. In conclusion, enamel organs and ameloblast-like cells express functional serotonin receptors. Reduced transcription of enamel proteins and secretion of vascular factors may indicate possible adverse effects of fluoxetine on amelogenesis.

Enamel deproteinization before acid etching--a scanning electron microscopic observation.

OBJECTIVES: This study was undertaken to evaluate the topographical features of enamel surface deproteinized with sodium hypochlorite (NaOCl) and etched with phosphoric acid (H3PO4) compared to phosphoric acid alone using Scanning Electron Microscopic (SEM) Analysis. STUDY DESIGN: 30 enamel blocks of 1 mm2 from ten human sound extracted permanent molars were obtained and treated as under: Group 1 (10 blocks): Enamel surface was etched with 37% H3PO4 gel for 15 seconds. Group 2 (10 blocks): Enamel surface was treated with 5.25% NaOCl for 60 seconds and then etched with 37% H3PO4 gel for 15 seconds. 10 enamel blocks were included in the control group where no treatment was carried out. The samples were subjected to SEM analysis and 5 microphotographs of each sample were obtained at 500X magnification and evaluated for the quality of etching pattern of the enamel in percentage (%) using Auto-CAD 2007 software. RESULTS: Mean values of etching pattern in Group 1 being 55.76% and Group 2 being 53.58%. No significant difference was observed between the two groups. CONCLUSION: The use of 37% phosphoric acid for 15 seconds still remains the best method for pretreatment of enamel.

Deproteinization of primary enamel with sodium hypochlorite before phosphoric acid etching.

The aim of this study was to evaluate the deproteinization of primary enamel by analyzing etching pattern types, with and without the application of 5% NaOCl before acid etching with 37% H3PO4. Fifteen extracted human primary molars were randomly selected for the present in vitro study; 1mm x 1mm blocks were prepared and divided into two groups (n = 21). These groups were treated as follows: Group A- Acid Etching with 37% H3PO4 gel for 15 s; Group B- 5% NaOCl for 60 s + Acid Etching with 37% H3POfor 15 s. The specimens were prepared for scanning electron microscopy analysis. The images were evaluated for quality types I and II etching of the enamel surface using ImageJ software. Datasets were checked for normality by Kolgomorv-Smirnov test and the nonparametric unpaired Mann-Whitney test was applied. The mean surface area of type I and II etching pattern values was 1922.314 µm2for Group A and 3840.473 µm2Group B. We conclude that deproteinization with 5% NaOCl prior to acid etching can be used to increase the area of adhesion and the quality of the etching pattern.

El objetivo del estudio fue evaluar la desproteinización del esmalte primario a través de los tipos de patrones de grabado, con y sin NaOCl 5% utilizado antes del grabado ácido con H3PO4 37%. Quince dientes primarios humanos extraídos se seleccionaron al azar para el presente estudio in vitro, se prepararon bloques de 1mm x 1 mm y se dividieron en dos grupos (n = 21). Estos grupos se trataron de la siguiente manera: Grupo A: Grabado ácido con H3PO4 37% en gel durante 15 segundos; Grupo B: NaOCl 5% durante 60 segundos + Grabado ácido con H3PO4 37% durante 15 segundos. Las muestras se prepararon para el análisis de microscopía electrónica de barrido. Las imágenes obtenidas se evaluaron principalmente por la calidad de los grabados tipo I y II de la superficie del esmalte primario, utilizando el software Image J. Los datos se analizaron en cuanto a su normalidad mediante la prueba de Kolgomorv-Smirnov, se utilizó pruebas no paramétricas: Prueba de Mann-Whitney no pareada. Como resultado, se encontró que el área de superficie media de los valores de patrón de grabado de tipo I y II para el Grupo A era 1922,314 µm2 y el Grupo B era 3840,473 µm2. Finalmente, llegamos a la conclusión de que se puede usar la desproteinización con NaOCl 5% antes del grabado ácido para aumentar el área de adhesión y la calidad del patrón de grabado.

Enamel deproteinization and its effect on acid etching: an in vitro study.

PURPOSE: The goal of this in vitro study was to identify the topographical features of the enamel surface deproteinized and etched with phosphoric acid (H3PO4) compared to phosphoric acid alone. MATERIALS AND METHOD: Ten extracted lower first and second permanent molars were polished with pumice and water, and then divided into 4 equal buccal sections having similar physical and chemical properties. The enamel surfaces of each group were subjected to the following treatments: Group A: Acid Etching with H3PO4 37% for 15 seconds. Group AH1: Sodium Hypochlorite (NaOCl) 5.25% for 30 seconds followed by Acid Etching with H3PO4 37% for 15 seconds. Group AH2; Sodium Hypochlorite (NaOCl) 5.25% for 60 seconds followed by Acid Etching with H3PO4 37% for 15 seconds. Results showed that group AH2 etching technique reached an area of 76.6 mm2 of the total surface, with a 71.8 mm2 (94.47%), type 1 and 2 etching pattern, followed by group AH1 with 55.9 mm2 out of 75.12 mm2 (74.1%), and finally group A with only 36.8 mm2 (48.83%) out of an area of 72.7 mm2. A significant statistical difference (P < 0.05) existed between all groups, leading to the conclusion that enamel deproteinization with 5.25% NaOCl for 1 minute before H3PO4, etching increases the enamel conditioning surface as well as the quality of the etching pattern.

Comparison of deproteinization agents on bonding to developmentally hypomineralized enamel.

OBJECTIVE: To compare bonding of dental adhesive to hypomineralized enamel (HE) after pre-treatment with either 5% sodium hypochlorite (NaOCl) solution or papain-based papacarie gel. METHODS: Normal enamel (NE) and HE obtained from hypomineralized first permanent molars were acid-etched with 32% phosphoric acid and randomly allocated into no deproteinization, deproteinization using 5% NaOCl, or deproteinization usping papacarie gel groups. Subsequently, the specimens were bonded, packed with composite resins and subjected to micro-shear bond strength (MSBS) testing and the data analysed using 2-way ANOVA and Tukey tests. Furthermore, specimens from all groups were subjected for qualitative analysis using scanning electron microscope. RESULTS: Two way-ANOVA showed that the factor "enamel substrate" was significant (p<0.001), "enamel pre-treatment" was not significant and interaction of the two factors was significant (p=0.005). HE produced inferior bonding with dental adhesive compared to NE. Enamel pre-treatment with deproteinization agents enhanced bonding to HE. No significant difference in MSBS was evident between the two deproteinization agents (p>0.05). Qualitative analysis of acid-etched moderate HE showed barely visible enamel rods with irregular etching pattern. Following acid etching and deproteinization, Type I and II etching patterns were observed in moderate HE; while a porous enamel surface with more profound etching patterns in severe HE. CONCLUSIONS: Papain-based papacarie could be an alternative deproteinization agent for bonding dental adhesive to HE. CLINICAL SIGNIFICANCE: Papain-based papacarie, a natural deproteinization agent and a proven chemo-mechanical caries removal agent could be an alternative to NaOCl for enhancement of bond durability of adhesive restorations to HE.

A western-blotting study of enamel glycoproteins in rat experimental fluorosis.

Experimental fluorosis was induced in order to get information on enamel protein glycosylation, using Western-blotting methodology with peroxidase-labelled concanavalin A. Fluoride inhibited amelogenin degradation, especially the production of intermediate forms. Within the non-amelogenin family of proteins there were changes in both the conventionally stainable components and the glycoconjugates revealed by lectin only. Fluoride influenced the whole extracellular processing of enamel proteins including movement between the mineral and non-mineral compartments. A different degradation scheme of enamel proteins, which also affects the glycoconjugates, might be of importance in the properties of the fluorosed enamel surface and its interactions with the oral environment.

X-linked amelogenesis imperfecta may result from decreased formation of tyrosine rich amelogenin peptide (TRAP).

Amelogenesis imperfecta (AI) is a group of inherited disorders with defective tooth enamel formation caused by various gene mutations. One of the mutations substitutes a cytidine for an adenine in exon 6 of the X-chromosomal amelogenin gene, which results in a proline to threonine change in the expressed amelogenin. This transformation is four amino acids N-terminal to the cleavage site for enamel matrix metalloproteinase-20 (MMP-20) in amelogenin. MMP-20 releases the tyrosine rich amelogenin peptide (TRAP) from amelogenin. This study evaluated the rate at which MMP-20 hydrolyses mutated amelogenin relative to unmutated amelogenin. A full-length recombinant human amelogenin and a mutated amelogenin with a substitution of proline by threonine were expressed and purified by ammonium sulphate precipitation and reverse phase HPLC. Recombinant metalloproteinase-20 (rMMP-20) was used to digest the recombinant proteins, which resulted in fragments with a mass predicted for TRAP. The proteolytic site was also modelled as substrates by two synthetic peptides, SYGYEPMGGWLHHQ and SYGYETMGGWLHHQ, selected from residues 36 to 49 of the amino acid sequence for amelogenin and the respective X-linked amelogenin mutant. These two peptides were labelled at their N- and C-termini respectively by using rhodamine and biotin. After digestion with MMP-20, the truncated peptides were separated by avidin-labelled magnetic Dynal beads and were identified by mass spectrometry. These results demonstrated that both oligopeptides were cleaved between tryptophan and leucine, matching the TRAP cutting site found in tooth enamel. Enzyme kinetics showed that the k(cat)/K(m) of rMMP-20 against the unmutated amelogenin peptide was 21 times greater than that against the mutated peptide. This study suggests that the reduced rate of TRAP formation by a single amino acid substitution alters enamel matrix hydrolysis by MMP-20, which may result in amelogenesis imperfecta.

An immunohistochemical study of the effects of fluoride on enamel development in the rat incisor.

A monoclonal antiamelogenin antibody was used to investigate the effects of fluoride on enamel development in the rat incisor. The results suggested that during secretion the enamel matrix molecules are arranged in such a way as to mask the epitope recognized by the monoclonal antibody. However, during the transition stage of development as the matrix begins to be degraded the epitope becomes exposed and labelling intensity increases to reach a maximum at the end of transition/start of maturation. The effect of fluoride is to delay the appearance of labelling within the enamel matrix until the end of transition. This suggests that the fluoride may inhibit enzymatic degradation or disaggregation of the matrix, the resulting residual matrix then inhibiting crystal growth.