Dataset Information

Proteomic analysis of human dental cementum and alveolar bone.

ABSTRACT: UNLABELLED:Dental cementum (DC) is a bone-like tissue covering the tooth root and responsible for attaching the tooth to the alveolar bone (AB) via the periodontal ligament (PDL). Studies have unsuccessfully tried to identify factors specific to DC versus AB, in an effort to better understand DC development and regeneration. The present study aimed to use matched human DC and AB samples (n=7) to generate their proteomes for comparative analysis. Bone samples were harvested from tooth extraction sites, whereas DC samples were obtained from the apical root portion of extracted third molars. Samples were denatured, followed by protein extraction reduction, alkylation and digestion for analysis by nanoAcquity HPLC system and LTQ-FT Ultra. Data analysis demonstrated that a total of 318 proteins were identified in AB and DC. In addition to shared proteins between these tissues, 105 and 83 proteins exclusive to AB or DC were identified, respectively. This is the first report analyzing the proteomic composition of human DC matrix and identifying putative unique and enriched proteins in comparison to alveolar bone. These findings may provide novel insights into developmental differences between DC and AB, and identify candidate biomarkers that may lead to more efficient and predictable therapies for periodontal regeneration. BIOLOGICAL SIGNIFICANCE:Periodontal disease is a highly prevalent disease affecting the world population, which involves breakdown of the tooth supporting tissues, the periodontal ligament, alveolar bone, and dental cementum. The lack of knowledge on specific factors that differentiate alveolar bone and dental cementum limits the development of more efficient and predictable reconstructive therapies. In order to better understand cementum development and potentially identify factors to improve therapeutic outcomes, we took the unique approach of using matched patient samples of dental cementum and alveolar bone to generate and compare a proteome list for each tissue. A potential biomarker for dental cementum was identified, superoxide dismutase 3 (SOD3), which is found in cementum and cementum-associated cells in mouse, pig, and human tissues. These findings may provide novel insights into developmental differences between alveolar bone and dental cementum, and represent the basis for improved and more predictable therapies.

SUBMITTER: Salmon CR

PROVIDER: S-EPMC3873800 | biostudies-literature | 2013 Oct

REPOSITORIES: biostudies-literature

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Proteomic analysis of human dental cementum and alveolar bone.

Salmon Cristiane R CR Tomazela Daniela M DM Ruiz Karina Gonzales Silvério KG Foster Brian L BL Paes Leme Adriana Franco AF Sallum Enilson Antonio EA Somerman Martha J MJ Nociti Francisco H FH

Journal of proteomics 20130902

Dental cementum (DC) is a bone-like tissue covering the tooth root and responsible for attaching the tooth to the alveolar bone (AB) via the periodontal ligament (PDL). Studies have unsuccessfully tried to identify factors specific to DC versus AB, in an effort to better understand DC development and regeneration. The present study aimed to use matched human DC and AB samples (n=7) to generate their proteomes for comparative analysis. Bone samples were harvested from tooth extraction sites, wher ...[more]

PMID: 24007660

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Project description:Background and objectivesDental cementum (DC) is a mineralized tissue covering tooth roots that plays a critical role in dental attachment. Differences in deciduous vs. permanent tooth DC have not been explored. We hypothesized that proteomic analysis of DC matrix would identify compositional differences in deciduous (DecDC) vs. permanent (PermDC) cementum that might reflect physiological or pathological differences, such as root resorption that is physiological in deciduous teeth but can be pathological in the permanent dentition.MethodsProtein extracts from deciduous (n = 25) and permanent (n = 12) teeth were pooled (five pools of DecDC, five teeth each; four pools of PermDC, three teeth each). Samples were denatured, and proteins were extracted, reduced, alkylated, digested, and analyzed by liquid chromatography-mass spectrometry (LC-MS/MS). The beta-binomial statistical test was applied to normalized spectrum counts with 5% significance level to determine differentially expressed proteins. Immunohistochemistry was used to validate selected proteins.ResultsA total of 510 proteins were identified: 123 (24.1%) exclusive to DecDC; 128 (25.1%) exclusive to PermDC; 259 (50.8%) commonly expressed in both DecDC and PermDC. Out of 60 differentially expressed proteins, 17 (28.3%) were detected in DecDC, including myeloperoxidase (MPO), whereas 43 (71.7%) were detected in PermDC, including decorin (DCN) and osteocalcin (BGLAP). Overall, Gene Ontology (GO) analysis indicated that all expressed proteins were related to GO biological processes that included localization and response to stress, and the GO molecular function of differentially expressed proteins was enriched in cell adhesion, molecular binding, cytoskeletal protein binding, structural molecular activity, and macromolecular complex binding. Immunohistochemistry confirmed the trends for selected differentially expressed proteins in human teeth.ConclusionsClear differences were found between the proteomes of DecDC and PermDC. These findings may lead to new insights into developmental differences between DecDC and PermDC, as well as to a better understanding of physiological/pathological events such as root resorption.

Project description:The WNT/β-catenin signaling pathway plays a central role in the biology of the periodontium, yet the function of specific extracellular WNT ligands remains poorly understood. By using a Wnt1-inducible transgenic mouse model targeting Col1a1-expressing alveolar osteoblasts, odontoblasts, and cementoblasts, we demonstrate that the WNT ligand WNT1 is a strong promoter of cementum and alveolar bone formation in vivo. We induced Wnt1 expression for 1, 3, or 9 wk in Wnt1Tg mice and analyzed them at the age of 6 wk and 12 wk. Micro-computed tomography (CT) analyses of the mandibles revealed a 1.8-fold increased bone volume after 1 and 3 wk of Wnt1 expression and a 3-fold increased bone volume after 9 wk of Wnt1 expression compared to controls. In addition, the alveolar ridges were higher in Wnt1Tg mice as compared to controls. Nondecalcified histology demonstrated increased acellular cementum thickness and cellular cementum volume after 3 and 9 wk of Wnt1 expression. However, 9 wk of Wnt1 expression was also associated with periodontal breakdown and ectopic mineralization of the pulp. The composition of this ectopic matrix was comparable to those of cellular cementum as demonstrated by quantitative backscattered electron imaging and immunohistochemistry for noncollagenous proteins. Our analyses of 52-wk-old mice after 9 wk of Wnt1 expression revealed that Wnt1 expression affects mandibular bone and growing incisors but not molar teeth, indicating that Wnt1 influences only growing tissues. To further investigate the effect of Wnt1 on cementoblasts, we stably transfected the cementoblast cell line (OCCM-30) with a vector expressing Wnt1-HA and performed proliferation as well as differentiation experiments. These experiments demonstrated that Wnt1 promotes proliferation but not differentiation of cementoblasts. Taken together, our findings identify, for the first time, Wnt1 as a critical regulator of alveolar bone and cementum formation, as well as provide important insights for harnessing the WNT signal pathway in regenerative dentistry.

Project description:A naturally graded interface due to functional demands can deviate toward a discontinuous interface, eventually decreasing the functional efficiency of a dynamic joint. It is this characteristic feature in a human bone-tooth fibrous joint bone-PDL-tooth complex that will be discussed through histochemistry, and site-specific high resolution microscopy, micro tomography(Micro XCT™), X-ray fluorescence imaging and wet nanoindentation techniques. Results demonstrated two causes for the occurrence of 5-50 μm narrowed PDL-space: 1) microscopic scalloped regions at the PDL-insertion sites and macro-scale stratified layers of bone with rich basophilic lines, and 2) macroscopic bony protrusions. Narrowed PDL-complexes illustrated patchy appearance of asporin, and when imaged under wet conditions using an atomic force microscope (AFM), demonstrated structural reorganization of the PDL, collagen periodicity, organic-dominant areas at the PDL-cementum and PDL-bone entheses and within cementum and bone. Scanning electron microscopy (SEM) results confirmed AFM results. Despite the narrowed PDL, continuity between PDL and vasculature in endosteal spaces of bone was demonstrated using a Micro XCT™. The higher levels of Ca and P X-ray fluorescence using a microprobe were correlated with higher elastic modulus values of 0.1-1.4 and 0.1-1.2 GPa for PDL-bone and PDL-cementum using wet nanoindentation. The ranges in elastic modulus values for PDL-bone and PDL-cementum entheses in 150-380 μm wide PDL-complex were 0.1-1.0 and 0.1-0.6 GPa. Based on these results we propose that strain amplification at the entheses could be minimized with a gradual change in modulus profile, a characteristic of 150-380 μm wide functional PDL-space. However, a discontinuity in modulus profile, a characteristic of 5-50 μm wide narrowed PDL-space would cause compromised mechanotransduction. The constrictions or narrowed sites within the bone-tooth fibrous joint will become the new "load bearing sites" that eventually could cause direct local fusion of bone with cementum.

Project description:UnlabelledDental cementum (DC) covers the tooth root and has important functions in tooth attachment and position. DC can be lost to disease, and regeneration is currently unpredictable due to limited understanding of DC formation. This study used a model of experimentally-induced apposition (EIA) in mice to identify proteins associated with new DC formation. Mandibular first molars were induced to super-erupt for 6 and 21days after extracting opposing maxillary molars. Decalcified and formalin-fixed paraffin-embedded mandible sections were prepared for laser capture microdissection. Microdissected protein extracts were analyzed by liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS), and the data submitted to repeated measure ANOVA test (RM-ANOVA, alpha=5%). A total of 519 proteins were identified, with 97 (18.6%) proteins found exclusively in EIA sites and 50 (9.6%) proteins exclusively expressed in control sites. Fifty six (10.7%) proteins were differentially regulated by RM-ANOVA (p<0.05), with 24 regulated by the exclusive effect of EIA (12 proteins) or the interaction between EIA and time (12 proteins), including serpin 1a, procollagen C-endopeptidase enhancer, tenascin X (TNX), and asporin (ASPN). In conclusion, proteomic analysis demonstrated significantly altered protein profile in DC under EIA, providing new insights on DC biology and potential candidates for tissue engineering applications.SignificanceDental cementum (DC) is a mineralized tissue that covers the tooth root surface and has important functions in tooth attachment and position. DC and other periodontal tissues can be lost to disease, and regeneration is currently unpredictable due to lack of understanding of DC formation. This study used a model of experimentally-induced apposition (EIA) in mice to promote new cementum formation, followed by laser capture microdissection (LCM) and liquid chromatography coupled to tandem mass spectrometry (LC-MS/MS) proteomic analysis. This approach identified proteins associated with new cementum formation that may be targets for promoting cementum regeneration.

Project description:The objective of this study was to evaluate age related changes in physical (structure/mechanical properties) and chemical (elemental/inorganic mineral content) properties of cementum layers interfacing dentin.Human mandibular molars (N=43) were collected and sorted by age (younger=19-39, middle=40-60, older=61-81 years). The structures of primary and secondary cementum (PC, SC) types were evaluated using light and atomic force microscopy (AFM) techniques. Chemical composition of cementum layers were characterized through gravimetric analysis by estimating ash weight and concentrations of Ca, Mn, and Zn trace elements in the analytes through inductively coupled plasma mass spectroscopy. The hardness of PC and SC was determined using microindentation and site-specific reduced elastic modulus properties were determined using nanoindentation techniques.PC contained fibrous 1-3 µm wide hygroscopic radial PDL-inserts. SC illustrated PC-like structure adjacent to a multilayered architecture composing of regions that contained mineral dominant lamellae. The width of the cementum dentin junction (CDJ) decreased as measured from the cementum enamel junction (CEJ) to the tooth apex (49-21 µm), and significantly decreased with age (44-23 µm; p<0.05). The inorganic ratio defined as the ratio of post-burn to pre-burn weight increased with age within primary cementum (PC) and secondary cementum (SC). Cementum showed an increase in hardness with age (PC (0.40-0.46 GPa), SC (0.37-0.43 GPa)), while dentin showed a decreasing trend (coronal dentin (0.70-0.72 GPa); apical dentin (0.63-0.73 GPa)).The observed physicochemical changes are indicative of increased mineralization of cementum and CDJ over time. Changes in tissue properties of teeth can alter overall tooth biomechanics and in turn the entire bone-tooth complex including the periodontal ligament. This study provides baseline information about the changes in physicochemical properties of cementum with age, which can be identified as adaptive in nature.

Project description:BackgroundDentin is a permeable tubular composite and complex structure, and in weight, it is composed of 20% organic matrix, 10% water, and 70% hydroxyapatite crystalline matrix. Demineralization of dentin with gradient concentrations of ethylene diamine tetraacetic acid, 0.6 N hydrochloric acid, or 2% nitric acid removes a major part of the crystalline apatite and maintains a majority of collagen type I and non-collagenous proteins, which creates an osteoinductive scaffold containing numerous matrix elements and growth factors. Therefore, demineralized dentin should be considered as an excellent naturally-derived bioactive material to enhance dental and alveolar bone tissues regeneration.MethodThe PubMed and Midline databases were searched in October 2021 for the relevant articles on treated dentin matrix (TDM)/demineralized dentin matrix (DDM) and their potential roles in tissue regeneration.ResultsSeveral studies with different study designs evaluating the effect of TDM/DDM on dental and bone tissues regeneration were found. TDM/DDM was obtained from human or animal sources and processed in different forms (particles, liquid extract, hydrogel, and paste) and different shapes (sheets, slices, disc-shaped, root-shaped, and barrier membranes), with variable sizes measured in micrometers or millimeters, demineralized with different protocols regarding the concentration of demineralizing agents and exposure time, and then sterilized and preserved with different techniques. In the act of biomimetic acellular material, TDM/DDM was used for the regeneration of the dentin-pulp complex through direct pulp capping technique, and it was found to possess the ability to activate the odontogenic differentiation of stem cells resident in the pulp tissues and induce reparative dentin formation. TDM/DDM was also considered for alveolar ridge and maxillary sinus floor augmentations, socket preservation, furcation perforation repair, guided bone, and bioroot regenerations as well as bone and cartilage healing.ConclusionTo our knowledge, there are no standard procedures to adopt a specific form for a specific purpose; therefore, future studies are required to come up with a well-characterized TDM/DDM for each specific application. Likely as decellularized dermal matrix and prospectively, if the TDM/DDM is supplied in proper consistency, forms, and in different sizes with good biological properties, it can be used efficiently instead of some widely-used regenerative biomaterials.

Dataset Information

Proteomic analysis of human dental cementum and alveolar bone.

Publications

Proteomic analysis of human dental cementum and alveolar bone.

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