Deciphering Black Extrinsic Tooth Stain Composition in Children Using Metaproteomics.

The present study focuses on the use of a metaproteomic approach to analyze Black Extrinsic Tooth Stains, a specific type of pigmented extrinsic substance. Metaproteomics is a powerful emerging technology that successfully enabled human protein and bacterial identification of this specific dental biofilm using high-resolution tandem mass spectrometry. A total of 1600 bacterial proteins were identified in black stain (BS) samples and 2058 proteins in dental plaque (DP) samples, whereas 607 and 582 human proteins were identified in BS and DP samples, respectively. A large diversity of bacteria genera (142) in BS and DP was identified, showing a high prevalence of Rothia, Kingella, Neisseria, and Pseudopropionibacterium in black stain samples. In this work, the high diversity of the dental microbiota and its proteome is highlighted, including significant differences between black stain and dental plaque samples.

The potential impact of salivary peptides in periodontitis.

Periodontitis is a complex immune-inflammatory condition characterized by the disruption of the periodontal ligament and subsequent formation of periodontal pockets, and by alveolar bone loss, often resulting in tooth loss. A myriad of factors, namely, genetic, metabolic, immunological, and inflammatory, is associated with progression of periodontitis. Periodontitis is also associated with systemic conditions such as neoplastic disorders, obesity, and diabetes. The current diagnosis of this disease relies on clinical measurements such as clinical attachment loss and probing depth, which have poor precision due to patient, operator and probe-related factors. Thus, there is a need to develop reliable, objective, and reproducible biomarkers for early diagnosis of periodontitis. In this regard, saliva, with contributions from the gingival crevicular fluid, holds great potential. However, most of the information on biomarkers of periodontium-related salivary proteins has come from studies on the molecular pathogenesis of periodontitis. In periodontitis, a more holistic approach, such as the use of -omics technologies, for biomarker discovery, is needed. Herein, we review the biomarkers proposed to date for the assessment of periodontitis, with emphasis on the role of salivary peptides in periodontitis and their assessment by high-throughput saliva proteomics. We also discuss the challenges pertaining to the identification of new periodontitis biomarkers in saliva.

[Dental stem cells: myth or hope in neuroregenerative medicine?] / Cellules souches dentaires : mythe ou espoir en médecine neurorégénératrice ?

The use of dental stem cells has raised many hopes in the development of new treatments for neurodegenerative diseases. According to current statistics, about 1 in 6 people in the world would be affected by a neurological disease. This number continues to increase as the world's population ages, making neurodegenerative diseases probably the one of the major challenges of public health in the 21st century. Neurodegenerative diseases are characterized mainly by a progressive loss of cognitive abilities and patient autonomy related to loss and degeneration of neurons in brain structures. Unfortunately, today, the only treatments available for this type of disease do only relieve the symptoms, they do not treat them, and few clinical trials have been truly convincing to date. Hence, hope lies for these diseases in the development of other therapeutic approaches. As such, dental stem cells could be a promising area of research because of their rapid growth, their great capacity for differentiation into different types of cells (among neuronal ones for some of them) and how easy they can be obtained, without raising ethical issues as for example for embryonic stem cells.

Variation of human salivary alpha-amylase proteoforms in three stimulation models.

OBJECTIVES: To evaluate the sAA proteoforms' expression during different stimulation situations. MATERIALS AND METHODS: This study evaluated the salivary alpha-amylase (sAA) proteoforms' behavior by western blot (WB) analysis and high-resolution mass spectrometry (LC-MS/MS) in different situations that produce increases in sAA activity. For this purpose, six healthy women with a similar body mass index, age, and fit, underwent different sAA stimulation tests, such as acetic acid stimulation, psychological stress using the standardized Trier social stress test, and physical effort using the Cooper treadmill test. RESULTS: The three models showed an increase in sAA activity. The WB demonstrated seven common bands observed in the six women (band one at 59 kDa, two at 56 kDa, three at 48 kDa, four at 45 kDa, five at 41 kDa, six at 36 kDa, and seven at 14 kDa), in which sAA protein was identified. The individual WB analysis showed that band two, which corresponded to the native non-glycosylated sAA proteoform, had a higher increase after the three sAA stimulation inducers, and this band was also the only proteoform correlated with sAA activity (r = 0.56, P = 0.001). In addition, when the label-free quantification analysis was performed, the different proteoforms showed different responses depending on the type of stimulation. CONCLUSIONS: This preliminary study showed that the diverse sAA proteoforms' expression depends on the different stimulation models. CLINICAL RELEVANCE: This study opens new perspectives and challenges for the use of the different alpha-amylase proteoforms as possible biomarkers in addition to the sAA activity.

Dental stem cells as a promising source for cell therapies in neurological diseases.

One of the main hallmarks of neurodegenerative diseases is the loss of neurons in the brain and/or spinal cord. The clinical characteristics of neurodegenerative diseases depend on the specific regions of the central nervous system that have undergone cell loss. These diseases place an enormous burden on society due to the degree of human suffering and their substantial economic cost. Indeed, approximately 1 in 6 individuals worldwide suffer from neurological disorders. As the world's population ages, the impact of neurological disorders is expected to increase and likely become one of the main public health and medical challenges of the coming century. There is still no cure for neurodegenerative diseases and currently available therapies only provide symptomatic relief. Hence, there is a pressing need to identify alternative therapeutic approaches to treat neurodegenerative diseases. Considering the global impact of these diseases and the need for new therapies, stem cell therapies have emerged as a promising treatment for neurodegenerative diseases. Notably, dental stem cells are an optimal candidate for cell-based therapeutic approaches, due primarily to the ease with which they can be obtained and their high regenerative potential. In this review, we summarize the potential of dental stem cells for use as a neurodegenerative disease therapy.

Assessing a multiplex-targeted proteomics approach for the clinical diagnosis of periodontitis using saliva samples.

AIM: The present study focused on the research of new biomarkers based on the liquid chromatography-multiple-reaction monitoring (MRM) proteomic profile in whole saliva of patients with periodontitis compared with periodontal healthy patients. METHODS: A 30-min multiplexed liquid chromatography-MRM method was used for absolute quantification of 35 plasma biomarkers in saliva from control patients and patients with periodontitis. RESULTS: Three proteins namely hemopexin, plasminogen and α-fibrinogen were shown to be clearly related to the presence of periodontitis compared with healthy patients. Apolipoprotein H was found to discriminate for the first time chronic and aggressive periodontitis. CONCLUSION: Our results indicate that this innovative MRM method could be used to screen for periodontitis in clinical environment. Furthermore, apolipoprotein H was found to be a discriminant biomarker of aggressive periodontitis.

Absolute quantification of 35 plasma biomarkers in human saliva using targeted MS.

BACKGROUND: Although the use of human saliva for diagnosing disease has been known to be of great clinical potential, few attempts have been made so far to develop its use. In this work, we developed an MRM-MS approach for 35 plasma biomarkers using human saliva in a clinical environment. METHODS & RESULTS: A 30-min micro LC-MS/MS run in MRM mode was conducted in order to quantify the 35 plasma proteins in human saliva. Sample preparation procedures were performed in quadruplicate and analyzed in duplicate. Results show that 32 of the 35 plasma proteins were quantified in human saliva using calibration curves in the 2- log10 dynamic ranges with excellent linearity. DISCUSSION/CONCLUSION: Our MRM method is compatible with routine measurements in daily clinical practice.

Adhesion and proliferation of human mesenchymal stem cells from dental pulp on porous silicon scaffolds.

In regenerative medicine, stem-cell-based therapy often requires a scaffold to deliver cells and/or growth factors to the injured site. Porous silicon (pSi) is a promising biomaterial for tissue engineering as it is both nontoxic and bioresorbable. Moreover, surface modification can offer control over the degradation rate of pSi and can also promote cell adhesion. Dental pulp stem cells (DPSC) are pluripotent mesenchymal stem cells found within the teeth and constitute a readily source of stem cells. Thus, coupling the good proliferation and differentiation capacities of DPSC with the textural and chemical properties of the pSi substrates provides an interesting approach for therapeutic use. In this study, the behavior of human DPSC is analyzed on pSi substrates presenting pores of various sizes, 10 ± 2 nm, 36 ± 4 nm, and 1.0 ± 0.1 µm, and undergoing different chemical treatments, thermal oxidation, silanization with aminopropyltriethoxysilane (APTES), and hydrosilylation with undecenoic acid or semicarbazide. DPSC adhesion and proliferation were followed for up to 72 h by fluorescence microscopy, scanning electron microscopy (SEM), enzymatic activity assay, and BrdU assay for mitotic activity. Porous silicon with 36 nm pore size was found to offer the best adhesion and the fastest growth rate for DPSC compared to pSi comporting smaller pore size (10 nm) or larger pore size (1 µm), especially after silanization with APTES. Hydrosilylation with semicarbazide favored cell adhesion and proliferation, especially mitosis after cell adhesion, but such chemical modification has been found to led to a scaffold that is stable for only 24-48 h in culture medium. Thus, semicarbazide-treated pSi appeared to be an appropriate scaffold for stem cell adhesion and immediate in vivo transplantation, whereas APTES-treated pSi was found to be more suitable for long-term in vitro culture, for stem cell proliferation and differentiation.

Influence of temperature and relative humidity on dentin and enamel bonding: a critical review of the literature. Part 1. Laboratory studies.

PURPOSE: The aim of this literature review was to investigate the results from in vitro laboratory studies on the influence of temperature and relative humidity present before polymerization on enamel and dentin bonding systems. MATERIALS AND METHODS: A systematic search was carried out including articles published in English, in peer reviewed journals, and indexed in MEDLINE/PubMed database. The search was carried out using the terms: relative AND humidity AND dental. In vitro studies were retrieved and divided into laboratory simulation studies and studies on physical properties. Laboratory simulation studies were addressed by subtopic: resin-enamel bond strength, resin-dentin bond strength, and dentin-enamel microleakage. Studies on physical properties tested the influence of humidity and temperature through polymerization contraction, flexural strength, and dentin wettability. RESULTS: Laboratory simulation studies demonstrated a strong influence of humidity and temperature on dentin and enamel bond strength and microleakage with dental adhesives systems. The studies on physical properties failed to demonstrate any influence of humidity on the adhesion performance, except for wettability measurement. CONCLUSION: The clinical relevance of these in vitro results remains to be demonstrated. A review of in vivo clinical studies will complete the literature data presented here.

MS characterization of multiple forms of alpha-amylase in human saliva.

Alpha-amylase is a major and well-characterized component of human saliva. Recent proteomic studies suggested that this protein could be observed in more than twenty spots on 2-D gels of salivary proteins. The aim of this work was to investigate this unexpected redundancy. 2-D gel electrophoresis was combined with systematic MALDI-TOF MS analysis. More than 140 protein spots identifying the alpha-amylase were shown to constitute a stable but very complex pattern. Careful analysis of mass spectra and simultaneous hierarchical clustering of the observed peptides and of the electrophoretic features of spots allowed one to define three major groups. A main class grouping 90 spots was shown to correspond to full length alpha-amylases that can be assumed to include isoforms and post-translationally modified forms, a subset of this class being demonstrated to be N-glycosylated. A second group included short alpha-amylases that are differently truncated in a non-random manner, very likely in the oral cavity. The last class grouped alpha-amylase forms showing both the N- and C-terminal sequences of the enzyme but displaying a molecular weight that was up to 50% lower than that of the native protein. It is speculated that the last group of alpha-amylase spots could correspond to proteins submitted to internal deletions prior to the secretion.