Proteomic Analysis of the Initial Oral Pellicle in Caries‐Active and Caries‐Free Individuals

Abstract

Purpose: To 1) elucidate individual proteomic profiles of the 3-min biofilm of caries-active and caries-free individuals and 2) compare these proteomic profiles against the background of caries. Experimental design: The initial oral pellicle of 12 caries-active and 12 caries-free individuals is generated in situ on ceramics specimens. The individual, host-specific proteomic profiles of this basic pellicle layer are analyzed by a chemical elution protocol combined with an elaborate mass spectrometry and evaluated bioinformatically. Results: A total of 1188 different proteins are identified. Additionally, 68 proteins are present in the profiles of all individuals, suggesting them as ubiquitously occurring base-proteins of the initial human pellicle. Thereof, the single profiles exhibit high inter-individual differences independent of their group affiliation, stating the initial pellicle to represent a rather "individual fingerprint". Quantitative analyses imply slight indication for 23 proteins potentially capable of counting for caries-specific biomarkers. Conclusions and clinical relevance: The introduced protocol enables the individual analysis of minimal protein amounts and allows for highly precise characterizations and comparisons of individual proteomic profiles. The results contain a considerable higher extent of protein identifications and might serve as a base for future large scale analyzes to identify discrimination factors for the development of caries susceptibility tests.

Deep Proteomic Insights into the Individual Short‐Term Pellicle Formation on Enamel—An In Situ Pilot Study

“…Next, the list of identified proteins was compared to the one recently published in another study from our group, where individual proteomic profiles of the 3‐min pellicle of 12 caries‐free and 12 caries‐active subjects on ceramics were analyzed. [ 33 ] In doing so, 65 of the 68 proteins proposed in the study as base proteins of the 3‐min pellicle on ceramics were recovered. For two of the missing proteins, namely Ig kappa chain C region and Immunoglobulin lambda‐like polypeptide 5, a multitude of proteins representing family members with similar properties and functions were identified in the current data sets.…”

Targeted metabolomics of pellicle and saliva in children with different caries activity

“…Aside from analysis of children's saliva samples, similar analytical approaches were performed for their pellicle samples, which were expected to display a more stable picture. The analysis of pellicle for diagnostic purposes has increased over the last years, as former limitations, such as small sample amounts, appear manageable by the development of efficient pellicle collection methods and increased sensitivity of analytical instruments 23 . The pellicle layer is a valuable analytical target because marker metabolites might be enriched on the tooth surface where the caries infection starts.…”

“…Eventually, pellicle was successfully sampled using custom-made "sampling lollipops" by fixing ceramic slabs (total surface of 8 cm²) onto "lollipop"-shaped silicone forms and placing three per participant buccally into the lower jaw for 10 min. The application of the ceramic material as substrate for pellicle formation is a standardised procedure that has successfully been used previously in a proteomic study on the initial pellicle 23 . The adsorption properties of this material are mainly based on electrostatic interactions, being very similar to tooth enamel, because hydrophilic feldspar ceramic is also negatively charged at physiological pH conditions and has comparable contact angles and the same isoelectric point (3.6 ± 0.2) 24 .…”

“…The preparation steps for the saliva samples were limited to a minimum with the removal of mucins and bacterial debris by centrifugation and proteins by solvent-mediated precipitation. Samples of oral biofilms (pellicle) were thoroughly rinsed with water to remove adherent saliva and subsequently desorbed from the orally exposed ceramic surfaces using Triton-X and radioimmunoprecipitation assay (RIPA) solutions 23 . Clear saliva and pellicle desorbates were spiked with the stable isotope-labelled internal standards (IS) for quantitative purposes and directly analysed by UPLC-MS/MS in the case of amino acids, and after derivatisation in the case of organic acids 25,26 and carbohydrates 27,28 , respectively, to improve ionisation and discrete fragmentation in MS/MS analysis.…”

Bioadhesion in the oral cavity and approaches for biofilm management by surface modifications

“…The pellicle is mainly composed of selectively absorbed salivary proteins and peptides but also contains proteins and other macromolecules from gingival crevicular fluid, blood, bacteria, mucosa, and diet [ 62 , 64 – 66 ]. Despite a large variability in individuals’ profiles, a core set of 68 proteins present in the pellicle proteome of 24 individuals was identified including among others antibacterial proteins (e.g., lysozyme, lactotransferrin, lactoperoxidase, and cystatins), lubricants (e.g., mucin 7), proteins promoting protein–substrate interactions (e.g., S100 family members, annexin A1, or elongation factor 2) or protein–protein interactions (e.g., 14-3-3 protein family or gamma-glutamyltransferase E), and proteins promoting pellicle integrity (e.g., carbonic anhydrase 6) [ 67 , 68 ]. The pellicle provides many beneficial functions [ 61 ].…”