Comparison of protein profiles of the pellicle, gingival crevicular fluid, and saliva: possible origin of pellicle proteins

BACKGROUND: The pellicle, the acellular organic material deposited on the surface of tooth enamel, has been thought to be derived from saliva. In this study, protein compositions of the pellicle, gingival crevicular fluid, and saliva collected from healthy adults were compared to elucidate the origin of pellicle proteins. RESULTS: The pellicle, gingival crevicular fluid, and saliva from the parotid gland or mixed gland were collected; subsequently, protein expression in samples from the respective individual was compared by SDS-PAGE and mass spectrometry. Following SDS-PAGE, proteins in the major bands were identified by mass spectrometry. The band pattern of pellicle proteins appeared different from those of gingival crevicular fluid, or saliva samples. Using mass spectrometry, 13 proteins in these samples were identified. The relative abundance of the proteins was quantitatively analyzed using mass spectrometry coupled with stable isotope labeling and by western blot. Cystatin S and α-amylase detected in pellicle were enriched in saliva samples, but not in gingival crevicular fluid, by western blot, and their abundance ratios were high in saliva and low in gingival crevicular fluid when analyzed by stable isotope labeling. Serotransferrin, however, was found only in the pellicle and gingival crevicular fluid by western blot and its abundance ratio was low in saliva. CONCLUSIONS: Our study revealed that the gingival crevicular fluid appears to contribute to pellicle formation in addition to saliva.

Proteomic analysis of the acquired enamel pellicle formed on human and bovine tooth: a study using the Bauru in situ pellicle model (BISPM)

Abstract The acquired enamel pellicle (AEP) is an organic film, bacteria-free, formed in vivo as a result of the selective adsorption of salivary proteins and glycoproteins to the solid surfaces exposed to the oral environment. Objective: This study aimed to compare the proteomic profile of AEP formed in situ on human and bovine enamel using a new intraoral device (Bauru in situ pellicle model - BISPM). Material and Methods: One hundred and eight samples of human and bovine enamel were prepared (4×4 mm). Nine subjects with good oral conditions wore a removable jaw appliance (BISPM) with 6 slabs of each substrate randomly allocated. The AEP was formed during the morning, for 120 minutes, and collected with an electrode filter paper soaked in 3% citric acid. This procedure was conducted in triplicate and the pellicle collected was processed for analysis by LC-ESI-MS/MS. The obtained mass spectrometry MS/MS spectra were searched against human protein database (SWISS-PROT). Results: The use of BISPM allowed the collection of enough proteins amount for proper analysis. A total of 51 proteins were found in the AEP collected from the substrates. Among them, 15 were common to both groups, 14 were exclusive of the bovine enamel, and 22 were exclusive of the human enamel. Proteins typically found in the AEP were identified, such as Histatin-1, Ig alpha-1, Ig alpha 2, Lysozyme C, Statherin and Submaxillary gland androgen-regulated protein 3B. Proteins not previously described in the AEP, such as metabolism, cell signaling, cell adhesion, cell division, transport, protein synthesis and degradation were also identified. Conclusion: These results demonstrate that the proteins typically found in the AEP appeared in both groups, regardless the substrate. The BISPM revealed to be a good device to be used in studies involving proteomic analysis of the AEP.

Saliva and dental erosion

Dental erosion is a multifactorial condition. The consideration of chemical, biological and behavioral factors is fundamental for its prevention and therapy. Among the biological factors, saliva is one of the most important parameters in the protection against erosive wear. Objective: This review discusses the role of salivary factors on the development of dental erosion. Material and Methods: A search was undertaken on MeDLINe website for papers from 1969 to 2010. The keywords used in the research were "saliva", "acquired pellicle", "salivary flow", "salivary buffering capacity" and "dental erosion". Inclusion of studies, data extraction and quality assessment were undertaken independently and in duplicate by two members of the review team. Disagreements were solved by discussion and consensus or by a third party. Results: Several characteristics and properties of saliva play an important role in dental erosion. Salivary clearance gradually eliminates the acids through swallowing and saliva presents buffering capacity causing neutralization and buffering of dietary acids. Salivary flow allows dilution of the acids. In addition, saliva is supersaturated with respect to tooth mineral, providing calcium, phosphate and fluoride necessary for remineralization after an erosive challenge. Furthermore, many proteins present in saliva and acquired pellicle play an important role in dental erosion. Conclusions: Saliva is the most important biological factor affecting the progression of dental erosion. Knowledge of its components and properties involved in this protective role can drive the development of preventive measures targeting to enhance its known beneficial effects.

Película adquirida do esmalte - a importância de seus componentes para a proteção contra a desmineralização do esmalte dentário / Enamel pellicle (AEP) formed by different components against the in vitro dental enamel

O presente estudo teve como objetivo avaliar o efeito protetor da película adquirida do esmalte (PAE), formada por diferentes componentes, contra a desmineralização in vitro do esmalte dentário. Na primeira etapa, uma revisão sistemática da literatura buscou evidências sobre a relação entre proteínas salivares e a doença cárie. Foram selecionados estudos observacionais controlados comparando a presença e/ou concentração de proteínas salivares entre indivíduos cárie-resistentes (CPO=0) e cárie-susceptíveis (CPO>0). A busca eletrônica pelos estudos foi feita nas bases de dados PubMed, Ovid Medline, ISI Web of Science, Medline, Cochrane, Lilacs, Scielo, BBO, Paho e Wholis, usando os descritores “dental caries” OU “tooth demineralization” OU “dental caries susceptibility” OU “dental enamel solubility” E“salivary proteins and peptides” OU “saliva” E “proteins”. De 188 estudos identificados, somente nove foram incluídos na revisão sistemática e cinco apresentaram evidência de relação entre proteínas salivares e cárie dental. Contudo, apesar de alta evidência encontrada, mais estudos com metodologia bem definida são necessários para que se possa identificar proteínas salivares como biomarcadores para a doença cárie. Na segunda etapa, um estudo in vitro avaliou o efeito protetor da PAE, formada por diferentes amostras salivares, contra a desmineralização do esmalte. Espécimes dentários foram expostos, por duas horas, à saliva total (ST), saliva da parótida (SP), ST dializada e SP dializada, permitindo a adsorção de proteínas sobre o esmalte dentário e, conseqüentemente, a formação de PAE. A diálise das amostras salivares foi realizada com membrana de peso molecular de corte igual a 1 kDa, o que permitiu a remoção de íons, principalmente, o cálcio e fosfato, da amostras salivares. Para o grupo controle foi empregada água deionizada (sem a presença de íons ou proteínas), não havendo assim a formação de PAE sobre os espécimes dentários deste grupo...

Película adquirida salival: revisión de la literatura / Acquired salivary pellicle: a review of the literature

En este artículo de revisión se presenta y analiza la información actualizada disponible sobre la composición química, mecanismo de formación y factores que afectan la producción de la película adquirida salival. Asimismo se discuten aspectos vinculados con la función que cumple dicho integumento, en especial la relacionada con su desempeño como antecesor de la placa bacteriana de la cual dependen las afecciones de mayor prevalencia e incidencia en odontología, como son la caries dental y la enfermedad periodontal.

In this article is presented and analyzes the information brought up to date on the chemical composition, mechanism of formation and factors that affect the production of salivary acquired pellicle. Likewise aspects related to the function are discussed that complies said integument, especially it related to its performance as the ancestor of the bacterial plaque of which the affections of grater prevalence and incidence in dentistry they depend, like are the dental decay and the periodontal illness.

Adhesion of salivary components to Streptococcus mutans peptides

Streptococcus mutans es el principal microorganismo asociadoa la caries dental, esta bacteria se une al esmalte a través de su interacción con las proteínas de la película adquirida yla proteína de superficie celular comúnmente denominada PAc. Por lo menos dos sitios de PAc interactúan in vitro con los receptores salivales, uno está dentro de la región más conservadade esta proteína que comprende los residuos de 816-1213 y el otro dentro de la secuencia rica en Alanina, residuos186-469. El objetivo del presente trabajo fue establecer similitudes o diferencias en la interacción de péptidos de PAc con los componentes salivales de individuos con y sin experienciade caries, para lo cual se tomaron muestras de saliva por salivación espontánea de 20 individuos con caries y 20 sin caries. A partir de las muestras de saliva se extrajeron las proteínas de la película adquirida (PA) utilizando hidroxilapatita sintética y fueron sometidas a la interacción con trespéptidos sintéticos de los segmentos de unión de PAc con los componentes salivales: PAc (301-319), PAc (365-377) y PAc (1024-1044). Los resultados muestran una baja interacciónentre los componentes de la PA y los péptidos en todos los individuos, sugiriendo que con base en las similitudes entre los individuos sanos y los individuos con la enfermedad lospéptidos de PAc estudiados no son relevantes en la adhesión

Streptococcus mutans is the main microorganism associated to dental caries; it adheres to the dental enamel by interacting with the acquired film’s proteins and the cell surface adhesin, called variously antigen PAc. At least two distinct sites in PAc interact with salivary receptors in vitro, these are within residues 816-1213, the most conserved portion of PAc, and within residues 186-469, the alanine-rich sequence. Our purpose was to establish differences or similarities in PAc’s peptides interactions with the salivary components of individuals with and without previous caries experience. 40 saliva samples were obtained from patients with (n=20) and without (n=20) caries. The acquired film’s proteins were extracted using hydroxyapatite, and subjected to interaction with three synthetic PAc peptides (PAc (301-319), PAc (365-377), and PAc (1025-1044)) synthesized from PAc’s bonding sites to the salivary components. The results show low interaction between the acquired pellicle components and the peptides in all patients. This suggests that the examined PAc’s are not relevant as far as the initial adhesion of Streptococcus mutans to the tooth’s surface is concerned, as defined by the similarities in the results for healthy and affected individuals.