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Int. J. Mol. Sci. 2016, 17(5), 728; https://doi.org/10.3390/ijms17050728

Review
Advances of Proteomic Sciences in Dentistry
1
Department of Prosthodontics and Dental Materials, School of Dentistry, King Faisal University, Al-Hofuf 31982, Saudi Arabia
2
Department of Biomedical Engineering, College of Engineering, King Faisal University, Al-Hofuf 31982, Saudi Arabia
3
Department of Restorative Dental Sciences, Al Farabi Colleges, Riyadh 11313, Saudi Arabia
4
Department of Restorative Dentistry, College of Dentistry, Taibah University, Madina Munawwarrah 41311, Saudi Arabia
5
Department of Materials Science and Engineering, The Kroto Research Institute, The University of Sheffield, North Campus, Broad Lane, Sheffield S3 7HQ, UK
*
Author to whom correspondence should be addressed.
Academic Editor: Mohamed N. Rahaman
Received: 18 April 2016 / Accepted: 9 May 2016 / Published: 13 May 2016

Abstract

:
Applications of proteomics tools revolutionized various biomedical disciplines such as genetics, molecular biology, medicine, and dentistry. The aim of this review is to highlight the major milestones in proteomics in dentistry during the last fifteen years. Human oral cavity contains hard and soft tissues and various biofluids including saliva and crevicular fluid. Proteomics has brought revolution in dentistry by helping in the early diagnosis of various diseases identified by the detection of numerous biomarkers present in the oral fluids. This paper covers the role of proteomics tools for the analysis of oral tissues. In addition, dental materials proteomics and their future directions are discussed.
Keywords:
proteomics; dentistry; enamel; dentin; saliva; gingival crevicular fluids and dental materials

1. Introduction

Every living thing contains fascinating molecules called proteins [1]. Proteins are building blocks for the living matrix and perform various functions [2]. The human body contains a number of different proteins which are structural, catalytic, regulatory, transport and storage, and transducer proteins. Each of these proteins plays a specific functional role [3]. Proteins belong to biological macromolecules that exist as three-dimensional structures because of the sequences involving the twenty different amino acids [4,5]. These amino acids are linked with each other by peptide bonds. The word “proteome”, first coined by Mark Wilkins in 1961, is used to describe a mixture of proteins [6]. All proteomes arise from mRNA and can be used to describe a cell’s protein content [7]. In simple terms, proteomics is the study of the distribution and interaction of proteins in time and space in a cell, organisms, or an ecosystem. In recent years, a number of proteomical studies on human body fluid and tissues (diseased and non-diseased) have been carried out by several researchers to analyze the chemistry in order to understand the life processes at the molecular as well as the cellular levels [8,9].
Proteomic tools have the ability to analyze human body samples such as blood, saliva, serums, urine, cervico-vaginal fluid (CVF), sperm cells, gingival crevicular fluids (GCF), microorganisms, and different tissues (enamel, dentine, cementum, pulp, gingiva, bone ligaments, stem cells, and mucosa) in both pathological and normal physiological states [10,11,12]. Quite a few studies have reported the analysis of dental tissues by means of proteomic tools. Approximately, 64% of human oral tissues samples have been studied for proteomics analysis as compared to 11% of animal dental tissues which signifies the clinical importance of proteomics [13]. A number of studies have explored human saliva due to its ease of accessibility and non-invasive method of collection. A significant number of studies (30%) have explored the salivary proteome during oral pathological conditions e.g., caries, periodontitis, gingivitis, dental abscess, endodontic lesions, and oral carcinomas [14]. Two methods exist to conduct proteomics of body fluids. In top down proteomics, intact proteins are analyzed by Electrospray Ionization (ESI) or Matrix-Assisted Laser Desorption/Ionization (MALDI) and the peptides are generated by a gas phase fragmentation method [15]. In contrast, bottom-up approach of proteomics is commonly used to analyze peptides produced through chemical or enzymatic cleavage of proteins, and with post-translational modification as well as through liquid chromatography (LC) in conjunction with mass spectrometry (MS). The bottom-up analysis, the more conventional method, has been sometimes also called “shotgun proteomics” [16]. Both approaches are commonly used in proteomics procedures utilizing mass spectrometry.
Table 1 presents a description of proteomics studies carried out on whole mouth saliva (WS), unstimulated whole mouth saliva (USWS) parotid gland secretions, submandibular and sublingual gland secretions, salivary gland ductal secretions, enamel, dentine, pulp tissues, gingival crevicular fluid (GCF), cementum, alveolar bone, periodontal fibers (PDL), and dental materials through top-down and bottom up approaches.
Recent developments in dental proteomic have helped uncover previously unknown details regarding the unique protein structures and their function for the diagnosis, defense mechanisms, and regeneration of dental tissues, tissue calcification, and repairing of dental tissues [40]. The aim of this paper is to elaborate on the currently available techniques, their reported applications for dental tissues. Furthermore, the current status of dental proteomical analysis and the discovered biomarkers is discussed in detail.

2. Dental Hard Tissue Proteomics

The tooth is the strongest calcified tissue of the human body due to its special architecture and compositions. It is composed of three distinct mineralized hard tissues: enamel, dentine, and cementum. Enamel is the hardest tissue of the human body and contains 96% minerals, 1% proteins and the remainder being water. The adequate mechanical properties of enamel suit its primary function: mastication of food. Enamel, the only dental hard tissue formed before eruption of teeth, is formed by cells called ameloblasts. Histologically, the inorganic component of enamel is composed of micro-rods and inter-rods of hydroxyapatite (HA) crystals embedded in protein matrix, the organic phase [41]. To date, the major enamel proteins that have been recognized are amelogenin, ameloblastin, enamelin, and tuftelin [42]. Additionally, a total of 42 proteins has been identified during enamel formation (secretory phase and maturation phase) by two dimensional electrophoresis (2-DE) and MS. These proteins include ERp29 which is involved in secretory protein synthesis and calcium binding protein (calbindin) and play a role in tooth maturation [43,44,45,46]. It has been concluded that amelogenin takes part in enamel formation and cementum development by guiding cells. It also regulates initiation and growth of HA crystals during the mineralization front across the carboxyl terminals [47,48]. Very recently, a novel organic protein containing enamel matrix was reported in an adult human tooth with thickness of 100–400 µm which could provide important protein transportation or biochemical linkage between enamel and dentin [49]. Ameloblasts secrete enamel specific extracellular matrix protein called ameloblastin and its expression is also detected during the initial development of craniofacial bones and dental hard tissues of mesenchymal origin [50]. The precise role of ameloblastin is not known but it has been hypothesized that it may control the enamel mineralization process during tooth development alongside growth of enamel mineral crystals [51].
The bulk structure of a tooth is made from dentin which possesses neurogenic and regenerative capabilities. By weight, dentin contains 70% minerals (mainly hydroxyapatite), 20% organic component, and 10% water. In proteomics, dentin has been particularly found useful for the identification of collagenous and non-collagenous proteins [52]. Its formation and biomineralization (dentinogenesis) is dynamically complex. Odontoblasts develop and secrete extracellular matrix followed by mineralization in an organized fashion [53]. Most abundant collagenous proteins present in dentin matrix are collagen (type I, III, V, VI, and XII) providing a three dimensional (3D) template for the mineralization of apatite crystals. Fibronectin and matrix metalloproteinase (MMP) 2, 9, and 20 are associated with predentin collagen fibrils [36]. Park et al. [36] performed Sodium-Dodecyl-Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE) followed by an LC-MS/MS method for identifying dentin proteins. The outcome of these experiments revealed the presence of 233 proteins and was confirmed using Western blot technique and immunohistochemical staining. This study was the first to provide dentin protein classification such as: metabolic enzymes, signal transduction, cellular organization, transport, immune response, transcription factor activity, cell growth/maintenance, chaperone/stress response, nucleic acid binding, and unknowns function. Another study reported by Jagr et al. [54], 2-DE and nano-LC-MS/MS was used to identify 289 proteins overall of which 90 had been previously unknown. In this study nine novel proteins were identified and were classified as immunoglobulins which help in the formation of extracellular matrix, formation of the cytoskeleton, cell adhesion molecule activity, cytoskeleton protein binding, immune responses, and peptidase activity. These findings may provide deep insight for the regenerative and rehabilitation of dental tissues. Moreover, only a few studies reported the proteomics analysis of cementum and alveolar bone. A total of 235 and 213 proteins have been recognized in the alveolar bone and cementum respectively using LC-MS/MS with LTQ-FT (Ultra) due to their high resolution and high accuracy [33]. Previously, proteins including osteocalcin (BGLAP), TNN, FN, VIM, CHAD, vitronectin VTN, and LUM were identified as non-collagenous extracellular proteins in cementum and alveolar bone [55,56,57].

3. Oral Fluid Proteomics

Compared to dental hard tissues, whole mouth saliva (WMS) and GCF have been studied more for proteomical analysis due to their non-invasive collection technique, minimal patient discomfort and anxiety as compare to blood collection for serum or plasma [14]. WMS is not only composed of major and minor salivary glands secretions but also contains mucosal transudates from all surfaces of the mouth, lymphoid tissues, oropharynx, and GCFs [58]. Proteomics studies on human saliva revealed 1000 plus proteins and peptides (Figure 1).
Numerous studies have been conducted on WMS to evaluate various body physiological and pathological conditions and have proven it as a diagnostic as well as a maintenance test fluid. The WMS was isolated from different diseases such as dental caries, Sjögren’s syndrome, diabetic patients, breast cancer patients, squamous cell carcinoma patients, and graft-versus-host disease patients. The WMS has been analyzed successfully by proteomical tools (electrophorically and chromatographically) [59,60,61,62].
Human gingival crevicular fluid (GCF) has been analyzed extensively. GCF has a variable protein composition based on periodontal health and diseases. GCF contains serum transudate (found in gingival sulcus), broken products of host epithelial or connective tissues, subgingival microbial plaque, extracellular proteins, host inflammatory mediators and cells [63]. GCF provides medium for the transportation of bacterial byproducts into the periodontal microenvironment and also helps to drive off host derived products [64]. It has been reported that GCF volume for biochemical and proteomics analysis is limited due to severity of tissue inflammation [65]. Many methods are available for the collection of GCF such as paper strips, capillary tubes, gingival wash, and paper cones [63]. In the last decade researchers have favored using paper strip in their research work due to easy insertion into the gingival crevice up to 1 mm of depth without bleeding from periodontal pockets [35]. After collection of the GCF sample it goes through different steps for proteomics analysis, as illustrated in Figure 2.
Variety of proteolytic enzymes are identified in GCF, such as collagenase, elastase, and cathepsin B, D, H, and L [66]. These proteolytic enzymes have been reported as destructors of periodontal tissues and have the capability to degrade type-I collagen and glycoproteins [67]. Table 2 describes detailed profiling of GCF proteins, proteomic tools used, and the number of proteins identified. Most commonly reported identified proteins from GCF are actin, keratins, histones, annexins, proteins S100-A9, apolipoprotein A-1, albumin, salivary gland antimicrobial peptides (histatins, HNP-1, -2 & -3, LL-37, statherin), and cystatin B [68,69]. Some immune related proteins present in GCF such as; Ig γ-1 chain C region, Ig γ-3 chain C region, lactoferroxin-C, leukocyte elastase inhibitor, α 1 antitrypsin, heat shock protein β-1, and coronin-1A [70].
A protein based oral biofilm, the acquired enamel pellicle (AEP), is formed on tooth surfaces within seconds after mechanical cleaning of the tooth surfaces [75]. It consists predominantly of proteins secreted from major and minor salivary glands, carbohydrates, ions, exogenous proteins, and lipids [76]. Lee and co-workers investigated AEP layer on enamel and quantified 50 proteins through Liquid Chromatography- Electrospray Ionization-Mass Spectrometry(LC-ESI-MS/MS) [77]. This layer amount is approximately 0.5–1 µg per tooth surface and its formation is crucial and the dynamic process is influenced by many factors such as; circadian cycle, biochemical properties of tooth surfaces, proteolytic capacity of the oral micro environment, and the oral microbiota [78]. The fatty acids (FAs) identified in AEP play an essential role in the pellicle formation, bacterial adhesion and protection against pellicle [79]. In-situ study reveals qualitatively and quantitatively a wide range of FAs (C12–C24) through gas-chromatography- electrospray ionization/ mass spectrometry (GC-EI/MS), in this study pellicle were formed in-situ on bovine enamel slabs mounted on upper jaw splints and inserted in the mouth of 10 subjects for 3–240 min. Several methods have been used for the collection of AEP over the last four decades e.g., palatal appliances, chemical solubilization techniques, mechanical techniques and soaked membranes method [80]. All these methods reported different compositions due to different routes of collection. Mayhall et al. [81] remounted freshly extracted discs of teeth crowns in a palatal appliance worn by the subject for 1 h. After AEP formation on the specimens, they dipped the appliances in 2% HCl and detected glutamic acid, serine, and glycine but a low amount of proline [81]. In another in vivo study on AEP composition it revealed a high level of glutamic acid and alanine but a significant amount of hexosamines. This study also determined that a different approach of AEP collection varies the composition of AEP [82]. AEP has many function in the oral cavity such as lubrication, regulation of mineral homeostasis, providing defense against microbes and microbial colonization through specific receptors. Siqueira et al. [75] identified 100 plus proteins and peptides from in vivo AEP, and suggested that all play an active role in maintaining oral health. Similarly, histatin peptide has shown protective mechanism against demineralization of the tooth [83]. A total of 130 proteins were identified from AEP using LC-ESI-MS/MS with high confidence which allowed the classification of AEP proteins according to nature of origin, chemical properties, and biological function as shown in Figure 3 [39]. Very recently, another group of researchers has identified 76 proteins from in vivo AEP present on deciduous teeth through mass spectrometry which opens up a diagnostic frontier in pediatric dentistry [84].

4. Dental Soft Tissue Proteomics

This dental pulp is a soft connective tissue that is composed of cells (mesenchymal, odontoblasts, fibroblasts) neural fibers, blood vessels, and lymphatics [85]. Tooth development, nourishment, sensitivity, defense reactions, repair, and regeneration are the main functions of dental pulp [86]. Its unique composition helps in nutrition as well as sensation for external stimuli [87]. Robertson et al. [88] investigated the calcification response from dental pulp against various external stimuli including dental trauma, caries, abrasion or attrition, and tooth retransplantation. Similarly, Yamazoe et al. [89] harvested dental pulp cells in subcutaneous tissues and analyzed its calcified tissues through proteomics. The reason behind this is that stem cells have the potential to form deciduous or permanent pulp cells. In the last decade, dental pulp stem cells have proven value in repairing dentin-pulp complex [90]. Dental pulp contains unique tissue specific proteins and small leucine-rich proteoglycans (biglycan, lumican, and mimecan) [91]. Pääkkönen et al. [92] analyzed gene and protein expression in healthy and carious dental pulp organs for the first time using cDNA microarray and 2D-gel electrophoresis. In their study, slight expression changes were reported due to the high amount of healthy pulp in both conditions. In addition, a total of 96 proteins were identified through 2-DE gel followed by MS/MS techniques. In the same experiment, cDNA microarrays explored the difference of gene expression in carious tissues and no gene differences were detected in 96 detected proteins. In another in vitro study on proteome mapping of odontoblasts-like dental pulp revealed 23 total proteins by 2-DE gel followed by MS [93]. These proteins are comprised of various types of peptides such as cell membrane bound molecules, cytoskeleton, and nuclear proteins and are involved in matrix synthesis and enzyme metabolism. The expression of various recognized proteins (annexin VI, heteronuclear ribonuclear proteins C, collagen type VI, matrilin-2) were confirmed using western blotting (WB) technique and real time- polymerase chain reaction (RT-PCR) analysis. The RNA amplification technique was successfully used to analyze gene expression and protein encoding linked to physiology of dental pulp. Microarray analysis disclosed a total of 362 genes related to pulp expression specifically hence, further classified as protoncogenes, tooth morphogenesis, genes of collagen, DNAse, metallopeptidases, and growth factors [94]. McLachlan et al. [95] studied dental pulp tissues for detailed characterization and molecular changes due to dental caries. A total of 445 genes were identified with two fold or greater difference in the expression level. At least 85 genes were reported abundant in health and 360 more abundant in disease suggesting that this approach may contribute to improved future diagnosis and treatment. Another comprehensive study on human tooth pulp was done by 2D-gel electrophoresis followed by nano-liquid chromatography tandem mass spectrometry (LC/MS). This approach detected 342 proteins in total with a high confidence, and two proteins were distinguished in human samples [37]. Very recently, Eckhard et al. [96] attempted in depth dental pulp proteome with N-Terminome by the help of the terminal amine isotropic labelling of substrates (TAILS) approach and identified 17 missing protein candidates for the Chromosome-centric Human Proteome Project (C-HPP; www.c-hpp.og). Missing proteins can be defined as proteins that show only transcriptomic evidences and an expected sequence (or suggested by homology) or partly detected proteins. Furthermore, there are transcript evidences for the survival of the corresponding proteins available without conclusive mass spectrometry data [97].
Periodontal ligament (PDL) is another fibrous connective tissue containing heterogeneous cell population and type 1collagen fibers abundantly. They play a key role in maintaining PDL space, homeostasis, and anchorage, as well as maintaining and providing regeneration or repair of periodontium in response to disease and mechanical trauma. Only a few studies reported on PDL cellular components at genomics and proteomics level but it is very essential to understand the unique features and functions. Reichenberg et al. [38] reported a first study on periodontal ligament (PDL) fibroblast proteome for understanding physiology and regulation of PDL and identifying disease related protein markers. In this study 900 spots were detected and 117 proteins spot identified with 74 different genes. In another study on exploring the early osteogenic differential protein-profile in human PDL cells [98], 29 differentially expressed proteins during osteogenic differentiations were reported [98] which have been primarily linked to the cell membrane-binding, cytoskeleton, nuclear regulations, matrix synthesis signal conduction and metabolic enzymes [99]. Proteomics may shed light on these complex functional details of these intra- and inter-cellular processes.

5. Dental Materials Proteomics

Concurrent application of genomics and proteomics have revolutionized dentistry by allowing the identification and characterization of oral tissues (soft, hard, and liquid), and also help in understanding them on the molecular level [10]. By definition, dental materials are those materials or devices which interact with the oral environment in physio-chemical, mechanical, and biological aspects [100,101,102,103,104,105,106]. Hence, dental materials should be biocompatible and interact without causing any toxicity. Many approaches have been used previously to analyze the success of dental materials and failure at a cellular level. Very recently, Ryta et al. [107] studied elution of unreacted triethylene glycol dimethacrylate (TEGDMA) from Smart Dentine Replacement (SDR™), Dentsply International, UK bulk-fill dental composite by using HPLC. In this study, polymerized specimens were treated with four solutions (100% ethanol, 75% ethanol, distilled water, and 100% methanol) with different concentration to evaluate direct dental pulp toxicity of unreacted TEGDMA monomer. It was confirmed through HPLC that the toxicity of unreacted TEGDMA towards dental pulp established during the first hour after the placement of resin. Dental adhesive systems were analyzed by a research group for the quantification of monomer elution and carbon–carbon double bonds in dental adhesive system using reverse-phase HPLC, and observed that no correlation exists between the resin dentin bonding of adhesives and the elution of unreacted monomers [108]. However, further proteomic analysis of materials on the molecular level is needed to understand the changes in proteomes of failed or successful implants. Some of the studies reported in the last decade on proteomics of dental materials are listed in Table 3.

6. Conclusions

With the help of “omics” (genomics, transcriptomics, proteomics, metabolomics, and metagenomics) many hidden compositions, behavior and metabolisms of dental tissues and oral fluids have been analyzed in the last fifteen years. These scientific disciplines helped the gathering of valuable information of the human proteome and will complete the Human Proteome Project (HPP) [117]. Proteomics tools have provided remarkable information regarding dental tissues and oral fluids [118]. The overall analysis on proteomics in dentistry shows that more studies directed toward structural formation, diagnosis, and pathogenesis but very limited studies on evaluation of treatment, prevention of diseases, and prognosis of interventions. To sum up, all proteomic tools can help to fill the gaps of the unexplored aspects of oral health and dental sciences.

Author Contributions

Zohaib Khurshid performed the literature search, wrote the manuscript. The information to create Table 1 and Table 2 was done by Sana Zohaib. Shariq Najeeb helped in the development of all figures and also in manuscript write up. Muhammad Sohail Zafar and Rabia Rehman performed literature search, and helped in the compilation of Table 3. Ihtesham Ur Rehman performed literature search, provide the guidelines in order to prepare the manuscript and finalized the manuscript.

Conflicts of Interest

The authors declare no conflict of interest.

Abbreviations

BGLAPBone Gamma Carboxyglutamate Protein
cDNAComplementary Deoxyribonucleic Acid
CHADChondroadherin
C-HPPChromosome-Centric Human Proteome Project
CsACyclosporin A
DNAseDeoxyribonuclease
EMDEnamel Matrix Derivative
ESIElectrospray Ionization
FNFibronectin
FNAFine Needle Aspiration
HGFHepatocyte Growth Factor
GCFGingival Crevicular Fluid
GVHDGraft Versus Host Disease
HAHydroxyapatite
HPPHuman Proteome Project
LC/MSLiquid chromatography/mass spectrometry
LTQ-FTLinear Ion Trap Mass Spectrometer
LUMLumican
mRNAmessenger Ribonucleic Acid
MSMass-Spectrometry
MALDIMatrix-Assisted Laser Desorption/Ionization
PDLPeriodontal Ligament
PEEKPolyetheretherketone
R-PCRRealtime-Polymerase Chain Reaction
SDS-PAGESodium dodecyl sulfate polyacrylamide gel electrophoresis
TAILSTerminal Amine Isotropic Labelling of Substrates
TNNTenascin
USWSUnstimulated Whole-Mouth Saliva
VIMVimentin
VTNVitronectin
WSWhole-Mouth saliva
WBWestern Blotting
2-DE2-Dimensional Electrophoresis

References

  1. Bhattacharyya, M. Protein structure and function: Looking through the network of side-chain interactions. Curr. Protein Pept. Sci. 2015, 17, 4–25. [Google Scholar] [CrossRef]
  2. Alberts, B. The cell as a collection of protein machines: Preparing the next generation of molecular biologists. Cell 1998, 92, 291–294. [Google Scholar] [CrossRef]
  3. Murzin, A.G.; Brenner, S.E.; Hubbard, T.; Chothia, C. SCOP: A structural classification of proteins database for the investigation of sequences and structures. J. Mol. Biol. 1995, 247, 536–540. [Google Scholar] [CrossRef]
  4. Luscombe, N.M. Amino acid-base interactions: A three-dimensional analysis of protein-DNA interactions at an atomic level. Nucleic Acids Res. 2001, 29, 2860–2874. [Google Scholar] [CrossRef] [PubMed]
  5. Nussinov, R.; Wolfson, H.J. Efficient detection of three-dimensional structural motifs in biological macromolecules by computer vision techniques. Proc. Natl. Acad. Sci. USA 1991, 88, 10495–10499. [Google Scholar] [CrossRef] [PubMed]
  6. Wilkins, M. Proteomics data mining. Exp. Rev. Proteom. 2009, 6, 599–603. [Google Scholar] [CrossRef] [PubMed]
  7. Altelaar, A.F.M.; Munoz, J.; Heck, A.J.R. Next-generation proteomics: Towards an integrative view of proteome dynamics. Nat. Rev. Genet. 2013, 14, 35–48. [Google Scholar] [CrossRef] [PubMed]
  8. Latterich, M.; Abramovitz, M.; Leyland-Jones, B. Proteomics: New technologies and clinical applications. Eur. J. Cancer 2008, 44, 2737–2741. [Google Scholar] [CrossRef] [PubMed]
  9. De Vries, S.; Bonvin, A. How proteins get in touch: Interface prediction in the study of biomolecular complexes. Curr. Protein Pept. Sci. 2008, 9, 394–406. [Google Scholar] [CrossRef] [PubMed]
  10. Chiappelli, F.; Covani, U.; Giacomelli, L. Proteomics as it pertains to oral pathologies and dental research. Bioinformation 2010, 5, 277. [Google Scholar] [CrossRef]
  11. Hubbard, M.J.; Faught, M.J.; Carlisle, B.H.; Stockwell, P.A. ToothPrint, a proteomic database for dental tissues. Proteomics 2001, 1, 132–135. [Google Scholar] [CrossRef]
  12. Jágr, M.; Eckhardt, A.; Pataridis, S.; Broukal, Z.; Dušková, J.; Mikšík, I. Proteomics of human teeth and saliva. Physiol. Res. 2014, 63, S141–S154. [Google Scholar] [PubMed]
  13. Rezende, T.M.B.; Lima, S.M.F.; Petriz, B.A.; Silva, O.N.; Freire, M.S.; Franco, O.L. Dentistry proteomics: From laboratory development to clinical practice. J. Cell. Physiol. 2013, 228, 2271–2284. [Google Scholar] [CrossRef] [PubMed]
  14. Amado, F.M.L.; Ferreira, R.P.; Vitorino, R. One decade of salivary proteomics: Current approaches and outstanding challenges. Clin. Biochem. 2013, 46, 506–517. [Google Scholar] [CrossRef] [PubMed]
  15. Cabras, T.; Iavarone, F.; Manconi, B.; Olianas, A.; Sanna, M.T.; Castagnola, M.; Messana, I. Top-down analytical platforms for the characterization of the human salivary proteome. Bioanalysis 2014, 6, 563–581. [Google Scholar] [CrossRef] [PubMed]
  16. Seema, S.; Krishnan, M.; Harith, A.K.; Sahai, K.; Iyer, S.R.; Arora, V.; Tripathi, R.P. Laser ionization mass spectrometry in oral squamous cell carcinoma. J. Oral Pathol. Med. 2014, 43, 471–483. [Google Scholar] [CrossRef] [PubMed]
  17. Hu, S.; Arellano, M.; Boontheung, P.; Wang, J.; Zhou, H.; Jiang, J.; Elashoff, D.; Wei, R.; Loo, J.A.; Wong, D.T. Salivary proteomics for oral cancer biomarker discovery. Clin. Cancer Res. 2008, 14, 6246–6252. [Google Scholar] [CrossRef] [PubMed]
  18. Bencharit, S.; Altarawneh, S.K.; Baxter, S.S.; Carlson, J.; Ross, G.F.; Border, M.B.; Mack, C.R.; Byrd, W.C.; Dibble, C.F.; Barros, S.; et al. Elucidating role of salivary proteins in denture stomatitis using a proteomic approach. Mol. BioSyst. 2012, 8, 3216–3223. [Google Scholar] [CrossRef] [PubMed]
  19. Baldini, C.; Giusti, L.; Ciregia, F.; da Valle, Y.; Giacomelli, C.; Donadio, E.; Sernissi, F.; Bazzichi, L.; Giannaccini, G.; Bombardieri, S.; et al. Proteomic analysis of saliva: A unique tool to distinguish primary Sjögren’s syndrome from secondary Sjögren’s syndrome and other sicca syndromes. Arthritis Res. Ther. 2011, 13, R194. [Google Scholar] [CrossRef] [PubMed]
  20. Baldini, C.; Laura, G.; Laura, B.; Antonio, L.; Stefano, B. Proteomic analysis of the saliva: A clue for understanding primary from secondary Sjögren’s syndrome? Autoimmun. Rev. 2008, 7, 185–191. [Google Scholar] [CrossRef] [PubMed]
  21. Devic, I.; Shi, M.; Schubert, M.M.; Lloid, M.; Izutsu, K.T.; Pan, C.; Missaghi, M.; Morton, T.H.; Mancl, L.A.; Zhang, J.; et al. Proteomic analysis of saliva from patients with oral chronic graft-versus-host disease. Biol. Blood Marrow Transplant. 2014, 20, 1048–1055. [Google Scholar] [CrossRef] [PubMed]
  22. Fonteles, C.S.R.; dos Santos, C.F.; da Silva Alves, K.S.; de Miranda Mota, A.C.; Damasceno, J.X.; Fonteles, M.C. Comparative proteomic analysis of human whole saliva of children with protein-energy undernutrition. Nutrition 2012, 28, 744–748. [Google Scholar] [CrossRef] [PubMed]
  23. Dowling, P.; Robert, W.; Paula, M.; Michael, H.; Aongus, C.; Martin, C. Analysis of the saliva proteome from patients with head and neck squamous cell carcinoma reveals differences in abundance levels of proteins associated with tumour progression and metastasis. J. Proteom. 2008, 71, 168–175. [Google Scholar] [CrossRef] [PubMed]
  24. Rao, P.V.; Reddy, A.P.; Lu, X.; Dasari, S.; Krishnaprasad, A.; Biggs, E.; Roberts, C.T.; Nagalla, S.R. Proteomic identification of salivary biomarkers of type-2 diabetes. J. Proteome Res. 2009, 8, 239–245. [Google Scholar] [CrossRef] [PubMed]
  25. Border, M.B.; Schwartz, S.; Carlson, J.; Dibble, C.F.; Kohltfarber, H.; Offenbacher, S.; Buse, J.B.; Bencharit, S. Exploring salivary proteomes in edentulous patients with type 2 diabetes. Mol. BioSyst. 2012, 8, 1304–1310. [Google Scholar] [CrossRef] [PubMed]
  26. Wang, Q.; Gao, P.; Wang, X.; Duan, Y. Investigation and identification of potential biomarkers in human saliva for the early diagnosis of oral squamous cell carcinoma. Clin. Chim. Acta 2014, 427, 79–85. [Google Scholar] [CrossRef] [PubMed]
  27. Yan, G.; Huang, W.; Xue, H.; Jia, Y.; Yang, D. [Relationship between dental caries and salivary proteome by electrospray ionization ion-trap tandem mass spectrometry in children aged 6 to 8 years]. Hua Xi Kou Qiang Yi Xue Za Zhi 2014, 32, 297–302. [Google Scholar] [PubMed]
  28. Ellias, M.F.; Zainal Ariffin, S.H.; Karsani, S.A.; Abdul Rahman, M.; Senafi, S.; Megat Abdul Wahab, R. Proteomic analysis of saliva identifies potential biomarkers for orthodontic tooth movement. Sci. World J. 2012, 2012, 1–6. [Google Scholar] [CrossRef] [PubMed]
  29. Wu, Y.; Shu, R.; Luo, L.-J.; Ge, L.-H.; Xie, Y.-F. Initial comparison of proteomic profiles of whole unstimulated saliva obtained from generalized aggressive periodontitis patients and healthy control subjects. J. Periodontal Res. 2009, 44, 636–644. [Google Scholar] [CrossRef] [PubMed]
  30. Gonçalves, L.D.R.; Soares, M.R.; Nogueira, F.C.S.; Garcia, C.; Camisasca, D.R.; Domont, G.; Feitosa, A.C.R.; Pereira, D.D.A.; Zingali, R.B.; Alves, G. Comparative proteomic analysis of whole saliva from chronic periodontitis patients. J. Proteom. 2010, 73, 1334–1341. [Google Scholar] [CrossRef] [PubMed]
  31. Rangé, H.; Léger, T.; Huchon, C.; Ciangura, C.; Diallo, D.; Poitou, C.; Meilhac, O.; Bouchard, P.; Chaussain, C. Salivary proteome modifications associated with periodontitis in obese patients. J. Clin. Periodontol. 2012, 39, 799–806. [Google Scholar] [CrossRef] [PubMed]
  32. Gonçalves, L.D.R.; Soares, M.R.; Nogueira, F.C.S.; Garcia, C.H.S.; Camisasca, D.R.; Domont, G.; Feitosa, A.C.R.; Pereira, D.A.; Zingali, R.B.; Alves, G. Analysis of the salivary proteome in gingivitis patients. J. Periodontal Res. 2011, 46, 599–606. [Google Scholar] [CrossRef] [PubMed]
  33. Salmon, C.R.; Tomazela, D.M.; Ruiz, K.G.S.; Foster, B.L.; Paes Leme, A.F.; Sallum, E.A.; Somerman, M.J.; Nociti, F.H. Proteomic analysis of human dental cementum and alveolar bone. J. Proteom. 2013, 91, 544–555. [Google Scholar] [CrossRef] [PubMed]
  34. Donadio, E.; Giusti, L.; Seccia, V.; Ciregia, F.; da Valle, Y.; Dallan, I.; Ventroni, T.; Giannaccini, G.; Sellari-Franceschini, S.; Lucacchini, A. New insight into benign tumours of major salivary glands by proteomic approach. PLoS ONE 2013, 8, e71874. [Google Scholar] [CrossRef] [PubMed]
  35. Huynh, A H.S.; Veith, P.D.; McGregor, N.R.; Adams, G.G.; Chen, D.; Reynolds, E.C.; Ngo, L.H.; Darby, I.B. Gingival crevicular fluid proteomes in health, gingivitis and chronic periodontitis. J. Periodontal Res. 2015, 50, 637–649. [Google Scholar]
  36. Park, E.-S.; Cho, H.-S.; Kwon, T.-G.; Jang, S.-N.; Lee, S.-H.; An, C.-H.; Shin, H.-I.; Kim, J.-Y.; Cho, J.-Y. Proteomics analysis of human dentin reveals distinct protein expression profiles. J. Proteome Res. 2009, 8, 1338–1346. [Google Scholar] [CrossRef] [PubMed]
  37. Eckhardt, A.; Jágr, M.; Pataridis, S.; Mikšík, I. Proteomic analysis of human tooth pulp: Proteomics of human tooth. J. Endod. 2014, 40, 1961–1966. [Google Scholar] [CrossRef] [PubMed]
  38. Reichenberg, E.; Redlich, M.; Cancemi, P.; Zaks, B.; Pitaru, S.; Fontana, S.; Pucci-Minafra, I.; Palmon, A. Proteomic analysis of protein components in periodontal ligament fibroblasts. J. Periodontol. 2005, 76, 1645–1653. [Google Scholar] [CrossRef] [PubMed]
  39. Siqueira, W.; Zhang, W.; Helmerhorst, E.J.; Gygi, S.; Oppenheim, F.G. Identification of protein components in in vivo human acquired enamel pellicle using LC-ESI-MS/MS. J. Proteome Res. 2007, 6, 2152–2160. [Google Scholar] [CrossRef] [PubMed]
  40. Khurshid, Z.; Naseem, M.; Sheikh, Z.; Najeeb, S.; Shahab, S.; Zafar, M.S. Oral antimicrobial peptides: Types and role in the oral cavity. Saudi Pharm. J. 2015. [Google Scholar] [CrossRef]
  41. He, L.H.; Swain, M.V. Understanding the mechanical behaviour of human enamel from its structural and compositional characteristics. J. Mech. Behav. Biomed. Mater. 2008, 1, 18–29. [Google Scholar] [CrossRef] [PubMed]
  42. Hubbard, M.J.; Kon, J.C. Proteomic analysis of dental tissues. J. Chromatogr. B 2002, 771, 211–220. [Google Scholar] [CrossRef]
  43. Hubbard, M.J.; McHugh, N.J.; Mangum, J.E. Exclusion of all three calbindins from a calcium-ferry role in rat enamel cells. Eur. J. Oral Sci. 2011, 119, 112–119. [Google Scholar] [CrossRef] [PubMed]
  44. Hubbard, M.J.; McHugh, N.J.; Carne, D.L. Isolation of ERp29, a novel endoplasmic reticulum protein, from rat enamel cells. Eur. J. Biochem. 2000, 267, 1945–1957. [Google Scholar] [CrossRef] [PubMed]
  45. Hubbard, M.J. Abundant calcium homeostasis machinery in rat dental enamel cells. Up-regulation of calcium store proteins during enamel mineralization implicates the endoplasmic reticulum in calcium transcytosis. Eur. J. Biochem. 1996, 239, 611–623. [Google Scholar] [CrossRef] [PubMed]
  46. Hubbard, M.J. Calbindin28kDa and calmodulin are hyperabundant in rat dental enamel cells. Identification of the protein phosphatase calcineurin as a principal calmodulin target and of a secretion-related role for calbindin28kDa. Eur. J. Biochem. 1995, 230, 68–79. [Google Scholar] [CrossRef] [PubMed]
  47. Moradian-Oldak, J.; Goldberg, M. Amelogenin supra-molecular assembly in vitro compared with the architecture of the forming enamel matrix. Cells Tissues Organs 2005, 181, 202–218. [Google Scholar] [CrossRef] [PubMed]
  48. Bartlett, J.D.; Bernhard, G.; Michel, G.; Janet, M.-O.; Michael, L.P.; Malcolm, L.S.; Xin, W.; Shane, N.W.; Yan, L.Z. Protein–protein interactions of the developing enamel matrix. Curr. Top. Dev. Biol. 2006, 74, 57–115. [Google Scholar] [PubMed]
  49. Dusevich, V.; Xu, C.; Wang, Y.; Walker, M.P.; Gorski, J.P. Identification of a protein-containing enamel matrix layer which bridges with the dentine-enamel junction of adult human teeth. Arch. Oral Biol. 2012, 57, 1585–1594. [Google Scholar] [CrossRef] [PubMed]
  50. Vymetal, J.; Slabý, I.; Spahr, A.; Vondrásek, J.; Lyngstadaas, S.P. Bioinformatic analysis and molecular modelling of human ameloblastin suggest a two-domain intrinsically unstructured calcium-binding protein. Eur. J. Oral Sci. 2008, 116, 124–134. [Google Scholar] [CrossRef] [PubMed]
  51. Wald, T.; Bednárová, L.; Osička, R.; Pachl, P.; Sulc, M.; Lyngstadaas, S.P.; Slaby, I.; Vondrášek, J. Biophysical characterization of recombinant human ameloblastin. Eur. J. Oral Sci. 2011, 119, 261–269. [Google Scholar] [CrossRef] [PubMed]
  52. Smith, A.J.; Scheven, B.A.; Takahashi, Y.; Ferracane, J.L.; Shelton, R.M.; Cooper, P.R. Dentine as a bioactive extracellular matrix. Arch. Oral Biol. 2012, 57, 109–121. [Google Scholar] [CrossRef] [PubMed]
  53. Petersson, U.; Hultenby, K.; Wendel, M. Identification, distribution and expression of osteoadherin during tooth formation. Eur. J. Oral Sci. 2003, 111, 128–136. [Google Scholar] [CrossRef] [PubMed]
  54. Jágr, M.; Eckhardt, A.; Pataridis, S.; Mikšík, I. Comprehensive proteomic analysis of human dentin. Eur. J. Oral Sci. 2012, 120, 259–268. [Google Scholar] [CrossRef] [PubMed]
  55. Robey, P.G. Vertebrate mineralized matrix proteins: Structure and function. Connect. Tissue Res. 1996, 35, 131–136. [Google Scholar] [CrossRef] [PubMed]
  56. Bosshardt, D.D.; Selvig, K.A. Dental cementum: The dynamic tissue covering of the root. Periodontol. 2000 1997, 13, 41–75. [Google Scholar] [CrossRef] [PubMed]
  57. Hammarström, L.; Alatli, I.; Fong, C.D. Origins of cementum. Oral Dis. 1996, 2, 63–69. [Google Scholar] [CrossRef] [PubMed]
  58. Campisi, G.; Fede, O.D.; Roccia, P.; Nicola, F.D.; Falaschini, S.; Muzio, L. Lo Saliva: Its value as a biological matrix and current methods of sampling. Eur. J. Inflamm. 2006, 4, 11–19. [Google Scholar]
  59. Bassim, C.W.; Ambatipudi, K.S.; Mays, J.W.; Edwards, D.A.; Swatkoski, S.; Fassil, H.; Baird, K.; Gucek, M.; Melvin, J.E.; Pavletic, S.Z. Quantitative salivary proteomic differences in oral chronic graft-versus-host disease. J. Clin. Immunol. 2012, 32, 1390–1399. [Google Scholar] [CrossRef] [PubMed]
  60. Chianeh, Y.R.; Prabhu, K. Biochemical markers in saliva of patients with oral squamous cell carcinoma. Asian Pac. J. Trop. Dis. 2014, 4, S33–S40. [Google Scholar] [CrossRef]
  61. Levine, M. Susceptibility to dental caries and the salivary proline-rich proteins. Int. J. Dent. 2011, 2011, 1–13. [Google Scholar] [CrossRef] [PubMed]
  62. Zhang, L.; Xiao, H.; Karlan, S.; Zhou, H.; Gross, J.; Elashoff, D.; Akin, D.; Yan, X.; Chia, D.; Karlan, B.; et al. Discovery and preclinical validation of salivary transcriptomic and proteomic biomarkers for the non-invasive detection of breast cancer. PLoS ONE 2010, 5, e15573. [Google Scholar] [CrossRef] [PubMed]
  63. Lamster, I.B. Evaluation of components of gingival crevicular fluid as diagnostic tests. Ann. Periodontol. 1997, 2, 123–137. [Google Scholar] [CrossRef] [PubMed]
  64. Tsuchida, S.; Satoh, M.; Umemura, H.; Sogawa, K.; Kawashima, Y.; Kado, S.; Sawai, S.; Nishimura, M.; Kodera, Y.; Matsushita, K.; et al. Proteomic analysis of gingival crevicular fluid for discovery of novel periodontal disease markers. Proteomics 2012, 12, 2190–2202. [Google Scholar] [CrossRef] [PubMed]
  65. Tsuchida, S.; Satoh, M.; Sogawa, K.; Kawashima, Y.; Kado, S.; Ishige, T.; Beppu, M.; Sawai, S.; Nishimura, M.; Kodera, Y.; et al. Application of proteomic technologies to discover and identify biomarkers for periodontal diseases in gingival crevicular fluid: A review. PROTEOM.-Clin. Appl. 2014, 8, 232–240. [Google Scholar] [CrossRef]
  66. Kunimatsu, K. Cathepsins B, H and L activities in gingival crevicular fluid from chronic adult periodontitis patients and exprimental gingivitis subjects. J. Periodontal Res. 1990, 25, 69–73. [Google Scholar] [CrossRef] [PubMed]
  67. Dannies, P.S. Protein degradation in health and disease. Ciba foundation symposium 75 (new series). Yale J. Biol. Med. 1982, 55, 75. [Google Scholar]
  68. Baliban, R.C.; Sakellari, D.; Li, Z.; DiMaggio, P.A.; Garcia, B.A.; Floudas, C.A. Novel protein identification methods for biomarker discovery via a proteomic analysis of periodontally healthy and diseased gingival crevicular fluid samples. J. Clin. Periodontol. 2012, 39, 203–212. [Google Scholar] [CrossRef] [PubMed]
  69. Ngo, L.H.; Darby, I.B.; Veith, P.D.; Locke, A.G.; Reynolds, E.C. Mass spectrometric analysis of gingival crevicular fluid biomarkers can predict periodontal disease progression. J. Periodontal Res. 2013, 48, 331–341. [Google Scholar] [CrossRef] [PubMed]
  70. Silva-Boghossian, C.M.; Colombo, A.P.V.; Tanaka, M.; Rayo, C.; Xiao, Y.; Siqueira, W.L. Quantitative proteomic analysis of gingival crevicular fluid in different periodontal conditions. PLoS ONE 2013, 8, e75898. [Google Scholar] [CrossRef] [PubMed]
  71. Carneiro, L.G.; Venuleo, C.; Oppenheim, F.G.; Salih, E. Proteome data set of human gingival crevicular fluid from healthy periodontium sites by multidimensional protein separation and mass spectrometry. J. Periodontal Res. 2012, 47, 248–262. [Google Scholar] [CrossRef] [PubMed]
  72. Carneiro, L.G.; Nouh, H.; Salih, E. Quantitative gingival crevicular fluid proteome in health and periodontal disease using stable isotope chemistries and mass spectrometry. J. Clin. Periodontol. 2014, 41, 733–747. [Google Scholar] [CrossRef] [PubMed]
  73. Rody, W.J., Jr.; Holliday, L.S.; McHugh, K.P.; Wallet, S.M.; Spicer, V.; Krokhin, O. Mass spectrometry analysis of gingival crevicular fluid in the presence of external root resorption. Am. J. Orthod. Dentofac. Orthop. 2014, 145, 787–798. [Google Scholar] [CrossRef] [PubMed]
  74. Kinney, J.S.; Morelli, T.; Oh, M.; Braun, T.M.; Ramseier, C.A.; Sugai, J.V.; Giannobile, W.V. Crevicular fluid biomarkers and periodontal disease progression. J. Clin. Periodontol. 2014, 41, 113–120. [Google Scholar] [CrossRef] [PubMed]
  75. Siqueira, W.L.; Custodio, W.; McDonald, E.E. New insights into the composition and functions of the acquired enamel pellicle. J. Dent. Res. 2012, 91, 1110–1118. [Google Scholar] [CrossRef] [PubMed]
  76. Vukosavljevic, D.; Custodio, W.; Buzalaf, M.A.R.; Hara, A.T.; Siqueira, W.L. Acquired pellicle as a modulator for dental erosion. Arch. Oral Biol. 2014, 59, 631–638. [Google Scholar] [CrossRef] [PubMed]
  77. Lee, Y.H.; Zimmerman, J.N.; Custodio, W.; Xiao, Y.; Basiri, T.; Hatibovic-Kofman, S.; Siqueira, W.L. Proteomic evaluation of acquired enamel pellicle during in vivo formation. PLoS ONE 2013, 8, e67919. [Google Scholar] [CrossRef] [PubMed]
  78. Lendenmann, U.; Grogan, J.; Oppenheim, F.G. Saliva and dental pellicle—A review. Adv. Dent. Res. 2000, 14, 22–28. [Google Scholar] [CrossRef] [PubMed]
  79. Reich, M.; Hannig, C.; Al-Ahmad, A.; Bolek, R.; Kummerer, K. A comprehensive method for determination of fatty acids in the initial oral biofilm (pellicle). J. Lipid Res. 2012, 53, 2226–2230. [Google Scholar] [CrossRef] [PubMed]
  80. Siqueira, W.L.; Helmerhorst, E.J.; Zhang, W.; Salih, E.; Oppenheim, F.G. Acquired enamel pellicle and its potential role in oral diagnostics. Ann. N. Y. Acad. Sci. 2007, 1098, 504–509. [Google Scholar] [CrossRef] [PubMed]
  81. Mayhall, C.W. Concerning the composition and source of the acquired enamel pellicle of human teeth. Arch. Oral Biol. 1970, 15, 1327–1341. [Google Scholar] [CrossRef]
  82. Armstrong, W.G. Amino-acid composition of the acquired pellicle of human tooth enamel. Nature 1966, 210, 197–198. [Google Scholar] [CrossRef] [PubMed]
  83. Siqueira, W.L.; Margolis, H.C.; Helmerhorst, E.J.; Mendes, F.M.; Oppenheim, F.G. Evidence of intact histatins in the in vivo acquired enamel pellicle. J. Dent. Res. 2010, 89, 626–630. [Google Scholar] [CrossRef] [PubMed]
  84. Zimmerman, J.N.; Custodio, W.; Hatibovic-Kofman, S.; Lee, Y.H.; Xiao, Y.; Siqueira, W.L. Proteome and peptidome of human acquired enamel pellicle on deciduous teeth. Int. J. Mol. Sci. 2013, 14, 920–934. [Google Scholar] [CrossRef] [PubMed]
  85. Yu, T.; Volponi, A.A.; Babb, R.; An, Z.; Sharpe, P.T. Stem cells in tooth development, growth, repair, and regeneration. Curr. Top. Dev. Biol. 2015, 115, 187–212. [Google Scholar] [PubMed]
  86. Mjör, I.A.; Sveen, O.B.; Heyeraas, K.J. Pulp-dentin biology in restorative dentistry. Part 1: Normal structure and physiology. Quintessence Int. 2001, 32, 427–446. [Google Scholar] [PubMed]
  87. Farges, J.-C.; Alliot-Licht, B.; Renard, E.; Ducret, M.; Gaudin, A.; Smith, A.J.; Cooper, P.R. Dental pulp defence and repair mechanisms in dental caries. Mediat. Inflamm. 2015, 2015, 230251. [Google Scholar] [CrossRef] [PubMed]
  88. Robertson, A.; Lundgren, T.; Andreasen, J.O.; Dietz, W.; Hoyer, I.; Norén, J.G. Pulp calcifications in traumatized primary incisors. A morphological and inductive analysis study. Eur. J. Oral Sci. 1997, 105, 196–206. [Google Scholar] [CrossRef] [PubMed]
  89. Yamazoe, T.; Aoki, K.; Simokawa, H.; Ohya, K.; Takagi, Y. Gene expression of bone matrix proteins in a calcified tissue appeared in subcutaneously transplanted rat dental pulp. J. Med. Dent. Sci. 2002, 49, 57–66. [Google Scholar] [PubMed]
  90. Zafar, M.S.; Khurshid, Z.; Almas, K. Oral tissue engineering progress and challenges. Tissue Eng. Regen. Med. 2015, 12, 387–397. [Google Scholar] [CrossRef]
  91. Sandanayake, N.S.; Sinclair, J.; Andreola, F.; Chapman, M.H.; Xue, A.; Webster, G.J.; Clarkson, A.; Gill, A.; Norton, I.D.; Smith, R.C.; et al. A combination of serum leucine-rich α-2-glycoprotein 1, CA19–9 and interleukin-6 differentiate biliary tract cancer from benign biliary strictures. Br. J. Cancer 2011, 105, 1370–1378. [Google Scholar] [CrossRef] [PubMed]
  92. Pääkkönen, V.; Ohlmeier, S.; Bergmann, U.; Larmas, M.; Salo, T.; Tjäderhane, L. Analysis of gene and protein expression in healthy and carious tooth pulp with cDNA microarray and two-dimensional gel electrophoresis. Eur. J. Oral Sci. 2005, 113, 369–379. [Google Scholar] [CrossRef] [PubMed]
  93. Wei, X.; Wu, L.; Ling, J.; Liu, L.; Liu, S.; Liu, W.; Li, M.; Xiao, Y. Differentially expressed protein profile of human dental pulp cells in the early process of odontoblast-like differentiation in vitro. J. Endod. 2008, 34, 1077–1084. [Google Scholar] [CrossRef] [PubMed]
  94. Tete, S.; Mastrangelo, F.; Scioletti, A.P.; Tranasi, M.; Raicu, F.; Paolantonio, M.; Stuppia, L.; Vinci, R.; Gherlone, E.; Ciampoli, C.; et al. Microarray expression profiling of human dental pulp from single subject. Clin. Investig. Med. 2008, 31, 55–61. [Google Scholar]
  95. McLachlan, J.L.; Smith, A.J.; Bujalska, I.J.; Cooper, P.R. Gene expression profiling of pulpal tissue reveals the molecular complexity of dental caries. Biochim. Biophys. Acta Mol. Basis Dis. 2005, 1741, 271–281. [Google Scholar] [CrossRef] [PubMed]
  96. Eckhard, U.; Marino, G.; Abbey, S.R.; Tharmarajah, G.; Matthew, I.; Overall, C.M. The human dental pulp proteome and N-terminome: Levering the unexplored potential of semitryptic peptides enriched by TAILS to identify missing proteins in the human proteome project in underexplored tissues. J. Proteome Res. 2015, 14, 3568–3582. [Google Scholar] [CrossRef] [PubMed]
  97. Paik, Y.-K.; Hancock, W.S. Uniting ENCODE with genome-wide proteomics. Nat. Biotechnol. 2012, 30, 1065–1067. [Google Scholar] [CrossRef] [PubMed]
  98. Wu, L.; Wei, X.; Ling, J.; Liu, L.; Liu, S.; Li, M.; Xiao, Y. Early osteogenic differential protein profile detected by proteomic analysis in human periodontal ligament cells. J. Periodontal Res. 2009, 44, 645–656. [Google Scholar] [CrossRef] [PubMed][Green Version]
  99. Wu, L.; Wei, X.; Ling, J.; Liu, L. A differential expression proteomic study of human periodontal ligament cell during osteogenic differentiation. Zhong Hua Kou Qiang Yi Xue Za Zhi 2009, 44, 212–216. [Google Scholar]
  100. Khurshid, Z.; Zafar, M.; Qasim, S.; Shahab, S.; Naseem, M.; AbuReqaiba, A. Advances in nanotechnology for restorative dentistry. Materials 2015, 8, 717–731. [Google Scholar] [CrossRef]
  101. Sheikh, Z.; Najeeb, S.; Khurshid, Z.; Verma, V.; Rashid, H.; Glogauer, M. Biodegradable materials for bone repair and tissue engineering applications. Materials 2015, 8, 5744–5794. [Google Scholar] [CrossRef]
  102. Najeeb, S.; Zafar, M.S.; Khurshid, Z.; Siddiqui, F. Applications of polyetheretherketone (PEEK) in oral implantology and prosthodontics. J. Prosthodont. Res. 2016, 60, 12–19. [Google Scholar] [CrossRef] [PubMed]
  103. Matinlinna, J.P.; Zeeshan, S.; Mohamed-Nur, A.; Nader, H.; Mohammad, A.J.; Zohaib, K. Barrier membranes for periodontal guided tissue regeneration applications. In Handbook of Oral Biomaterials; Pan Stanford Publishing: Boca Raton, FL, USA, 2014; pp. 605–636. [Google Scholar]
  104. Najeeb, S.; Khurshid, Z.; Matinlinna, J.P.; Siddiqui, F.; Nassani, M.Z.; Baroudi, K. Nanomodified peek dental implants: Bioactive composites and surface modification—A review. Int. J. Dent. 2015, 2015. [Google Scholar] [CrossRef] [PubMed]
  105. Najeeb, S.; Khurshid, Z.; Zafar, M.S.; Ajlal, S. Applications of light amplification by stimulated emission of radiation (lasers) for restorative dentistry. Med. Princ. Pract. 2016, 25, 201–211. [Google Scholar] [CrossRef] [PubMed]
  106. Naseem, M.; Khurshid, Z.; Khan, H.A.; Niazi, F.; Shahab, S.; Zafar, M.S. Oral health challenges in pregnant women: Recommendations for dental care professionals. Saudi J. Dent. Res. 2015. [Google Scholar] [CrossRef]
  107. Łagocka, R.; Jakubowska, K.; Chlubek, D.; Buczkowska-Radlińska, J. Elution study of unreacted TEGDMA from bulk-fill composite (SDR™ Dentsply) using HPLC. Adv. Med. Sci. 2015, 60, 191–198. [Google Scholar] [CrossRef] [PubMed]
  108. Miletic, V.; Santini, A.; Trkulja, I. Quantification of monomer elution and carbon–carbon double bonds in dental adhesive systems using HPLC and micro-Raman spectroscopy. J. Dent. 2009, 37, 177–184. [Google Scholar] [CrossRef] [PubMed]
  109. Boyan, B.D.; Weesner, T.C.; Lohmann, C.H.; Andreacchio, D.; Carnes, D.L.; Dean, D.D.; Cochran, D.L.; Schwartz, Z. Porcine fetal enamel matrix derivative enhances bone formation induced by demineralized freeze dried bone allograft in vivo. J. Periodontol. 2000, 71, 1278–1286. [Google Scholar] [CrossRef] [PubMed]
  110. Derhami, K.; Zheng, J.; Li, L.; Wolfaardt, J.F.; Scott, P.G. Proteomic analysis of human skin fibroblasts grown on titanium: Novel approach to study molecular biocompatibility. J. Biomed. Mater. Res. 2001, 56, 234–244. [Google Scholar] [CrossRef]
  111. Koin, P.J.; Kilislioglu, A.; Zhou, M.; Drummond, J.L.; Hanley, L. Analysis of the degradation of a model dental composite. J. Dent. Res. 2008, 87, 661–665. [Google Scholar] [CrossRef] [PubMed]
  112. Jung, J.Y.; Kang, G.C.; Jeong, Y.J.; Kim, S.H.; Kwak, Y.G.; Kim, W.J. Proteomic analysis in cyclosporin a-induced overgrowth of human gingival fibroblasts. Biol. Pharm. Bull. 2009, 32, 1480–1485. [Google Scholar] [CrossRef] [PubMed]
  113. Taiyoji, M.; Shitomi, Y.; Taniguchi, M.; Saitoh, E.; Ohtsubo, S. Identification of proteinaceous inhibitors of a cysteine proteinase (an Arg-specific gingipain) from Porphyromonas gingivalis in rice grain, using targeted-proteomics approaches. J. Proteome Res. 2009, 8, 5165–5174. [Google Scholar] [CrossRef] [PubMed]
  114. Zilm, P.S.; Bartold, P.M. Proteomic identification of proteinase inhibitors in the porcine enamel matrix derivative, EMD®. J. Periodontal Res. 2011, 46, 111–117. [Google Scholar] [CrossRef] [PubMed]
  115. Dorkhan, M.; Chávez de Paz, L.E.; Skepö, M.; Svensäter, G.; Davies, J.R. Effects of saliva or serum coating on adherence of Streptococcus oralis strains to titanium. Microbiology 2012, 158, 390–397. [Google Scholar] [CrossRef] [PubMed]
  116. Zhao, M.; An, M.; Wang, Q.; Liu, X.; Lai, W.; Zhao, X.; Wei, S.; Ji, J. Quantitative proteomic analysis of human osteoblast-like MG-63 cells in response to bioinert implant material titanium and polyetheretherketone. J. Proteom. 2012, 75, 3560–3573. [Google Scholar] [CrossRef] [PubMed]
  117. Oakley, M.; Barthel, D.; Bykov, Y.; Garibaldi, J.; Burke, E.; Krasnogor, N.; Hirst, J. Search strategies in structural bioinformatics. Curr. Protein Pept. Sci. 2008, 9, 260–274. [Google Scholar] [CrossRef] [PubMed]
  118. Khurshid, Z.; Shariq, N.; Maria, M.; Syed, F.M.; Syed, Q.R.; Sana, Z.; Farshid, S.; Muhammad, S.Z. Histatin peptides: Pharmacological functions and its applications in dentistry. Saudi Pharm. J. 2016. [Google Scholar] [CrossRef]
Figure 1. Illustration representing human salivary drop proteins and peptides.
Figure 1. Illustration representing human salivary drop proteins and peptides.
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Figure 2. Illustration representing the steps of gingival crevicular fluids (GCF) proteomics analysis.
Figure 2. Illustration representing the steps of gingival crevicular fluids (GCF) proteomics analysis.
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Figure 3. Classification of acquired enamel pellicle (AEP) proteins according to their origin, chemical function, and by their biological functions proposed by Siqueira et al. [84].
Figure 3. Classification of acquired enamel pellicle (AEP) proteins according to their origin, chemical function, and by their biological functions proposed by Siqueira et al. [84].
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Table 1. Detailed discussion of oral diseases protein analysis using proteomic tools.
Table 1. Detailed discussion of oral diseases protein analysis using proteomic tools.
SampleDisease ConditionProteomic ToolsIdentified MarkersReferences
Whole mouth saliva (WS)Oral squamous cell carcinomaShotgun proteome analysis, Western blotting (WB) and Enzyme Linked Immuno-Sorbent Assay (ELISA)MRP14, M2BP, CD59, catalase, profilin, M2BP, involucrin, histone H1, S100A12, and S100P[17]
WSDenture stomatitisSurface-Enhanced Laser Desorption/Ionization (SELDI) time-of-flight-(TOF)/ mass spectrometry (MS), liquid chromatography (LC)- Matrix-Assisted Laser Desorption/Ionization (MALDI)-TOF-MSStatherin, desmocollin-2, kininogen-1, carbonic anhydrase-6, cystatin SN, cystatin c, peptidyl-prolyl cis-trans isomerase and immunoglobulin fragments[18]
WSPrimary Sjögren’s syndrometwo dimensional electrophoresis (2-DE), MALDI-TOF/MS, WB and ELISACarbonic anhydrase VI, α-amylases precursor, epidermal fatty acid binding protein (E-FABP), macroglobulin (b-2), immunoglobulin k light chain (IGK-light chain) and glyceraldehydes-3-phosphate dehydrogenase (G3PDH)[19]
WSSecondary Sjögren’s syndrome2-DE, MALDI-TOF-MS, WB and ELISADecrease ↓ Proline rich proteins (PRPs), ↓ Cystatin C, ↓ Lysozyme C and histatin, Increase ↑ Kallikrein and defensins[20]
WSGVHDTandem MS & ELISAIL-1 receptor antagonist and cystatin B[21]
WSProtein-energy undernutrition2-DE Gel and Image Master two dimensional 2DCyclic-dependent protein kinase[22]
WSSquamous cell carcinoma (head and neck region)C4 Reverse Phase-High Pressure Liquid Chromatography (RP-HPLC), and LC-MS/MSMRP14, Profilin, CD59, catalase and M2BP[23]
WSDiabetes (type-2)2D-LC-MS/MS, WB and ELISAα-1-antitrypsin (A1AT), α-2 macroglobulin (A2MG), transthyretin (TTR), salivary α -amylase (AMYS), cystatin C (Cys-C)[24]
WSEdentulous patient with type-2 diabetes2D-LC-MS/MSSerum amyloid-A and glyceraldehyde-3-phosphate dehydrogenase are increased, serotransferrin and amylase, palate, lung and nasal epithelium associated proteins (PLUNC) are reduced[25]
Unstimulated whole mouth saliva (USWS)Squamous cell carcinoma (oral mucosa)Ultra-Pressure Liquid Chromatography (UPLC-MS), Hydrophilic Interaction Liquid Chromatography (HILIC)l-carnitine, choline, betaine and pipecolinic acid[26]
Saliva (parotid glands)CariesHPLC-MS/MSmatrix metalloproteinase-9 (MMP9), mucin-7 (MUC7), lactotransferrin (LTF), carbonic anhydrase 6 (CA6), azurocidin (AZU), and cold agglutinin[27]
WSOrthodontic tooth movement2-DE, MALDI-TOF/tandem mass spectrometry (TEM)Protein S100-A9, CRISP-3, Immunoglobulin J chain and Ig α-1 chain C region[28]
USWSAggressive periodontitis2-DE/HPLC–Electrospray Ionization (ESI)-MSIncrease in serum albumin, immunoglobulin Ig γ2, α2 chain C region, zinc-α2 glycoprotein, salivary α-amylase and vitamin D-binding proteins.
Decrease in lactotransferrin, carbonic anhydrase 6, elongation factor 2, 14-3-3 sigma, short palate, lung and nasal epithelium carcinoma-associated protein-2
[29]
USWSPeriodontitis chronic2-DE/MALDI-TOF/TOF MS and nLC-Q-TOFRise in serum albumin, hemoglobin, immunoglobulin and α-amylase[30]
WSPeriodontitis in obese patientSELDI-TOF-MSAlbumin, haemoglobin (α and β chains) and α-defensins (1, 2 & 3)[31]
USWSGingivitis2-DE/MALDI-TOF/TOF MS and nLC-Q-TOFZymogen granule protein-16 homolog B mucin, S100-A9, histatin, proline-rich-protein, 3, lipocalin-1 precursor, carbonic anhydrase 6, prolactin-induced protein, cystatin, keratins[32]
Dental cementum-Nano-Acuity HPLC and LTQ-FT ultraOsteomodulin (OMD), biglycan (BGN), insulin-like growth factor II (IGF2), pigment epithelium-derived factor (SERPINF1) and POSTN[33]
Fine Needle Aspiration (FNA) fluidParotid gland tumor (Benign origin)Nano LC-ESI-MS/MS and LTQ-Qrbitrap velos analysis and Western blot analysisIg γ-1 and kappa chain and Ig α-1 chain C regions, S100A9, macrophage capping proteins, apolipoprotein E and α crystalline B chain, annexin (A1 and A4)[34]
Gingival crevicular fluids (GCF)Gingivitis and chronic periodontitis2-DE-LC-ESI-MS and Nano-LC-ESI-MSFibronectin, keratin, neutrophil, defensin3, Immunoglobulins, lactotransferrin precursor, 14-3-3 protein ζ/δ and α-actinin[35]
Dentin-LC-MS/MSBiglycan, osteoglycin, osteopontin, osteocalcin, asporin, lumican, mimecan, DSPP and SOD3[36]
Dental pulp-2-DE, Nano-LCMS/MS342 proteins identified[37]
Periodontal fibers (PDL)-2-DE, MALDI-TOF, Western blot,117 proteins identified[38]
Acquired enamel pellicle (AEP)-LC-ESI-MS/MS130 proteins identified[39]
Table 2. Profiling and proteomic tools used for the detection and characterization of gingival crevicular fluid (GCF) proteins.
Table 2. Profiling and proteomic tools used for the detection and characterization of gingival crevicular fluid (GCF) proteins.
Author Sample Collection SitesCollection MethodProteomic ToolNumber of Identified ProteinsOutcome of StudyReference
Baliban et al.Collected from pre-selected sites with probing depth >6 mm and <8 mm in periodontitis patients and for periodontaly health from mesio-buccal sites of first molarFilter strips (Periopapers®, Interstate Drug. Exchange, Amityville, NY, USA)Protein digest with trypsin, HPLC, fragmented analysis with tandem mass spectrometry (MS/MS)432 human proteins identified (120 new)Study identified novel biomarkers from GCF of periodontaly healthy and chronic periodontitis patients[68]
Tsuchida et al.Labial side of maxillary incisors without crown and restorationAbsorbent paper points (ZIPPERER®, Munich, Germany)2-DE, Sodium-Dodecyl-Sulfate-Polyacrylamide Gel Electrophoresis (SDS-PAGE), WB analysis, HPLC with LTQ-XL, HPLC with LTQ-Orbitrap XL, LC-MS/MS327 proteins identifiedSOD1 and DCD were significantly increase ↑ in GCF of periodontal patients[64]
Carneiro et al.Healthy gingival sulcus of the second and third molar teethPeriopapers®, USATrypsin digested followed by nano-flow liquid chromatography electrospray ionization tandem mass spectrometry (LC-ESI-MS/MS) analysis and ELISA for human albumin analysis199 proteins identified Provide proteins analysis of healthy periodontium and explore GCF composition with new groups of proteins specific to GCF microenvironment[71]
Ngo et al.Five deepest sites and molar sites except mesial surface were excludedMicrocaps (glass micocapillary tubes); Drummed Scientific, Brookmall, PA, USAMatrix-assisted laser desposition/ionization time-of-flight (MALDI-TOF) mass spectrometry (MS) GCF mass spectra could be best for analyzing attachment loss and diagnosis of periodontal diseases[69]
Carina, et al.Chronic Periodontitis patients sample were taken from different sites (5 deep sites, 5 shallow sites with gingivitis, and 4 without bleeding on probing sites)Periopaper strip (ProFlow Inc., Amityville, NY, USA)Reversed- phase (RP) LC-ESi-MS/MS and ELISA230 proteins identifiedConcluded marked differences in GCF proteomics in different disease profiles[70]
Carneiro et al.The pre-selected specific sites with moderate and severe chronic periodontal disease were defined by pocket depth of 5–7 mm (24 patients) and >7 mm (16 patients)Periopaper strips (Oraflow, Plainview, NY, USA)SDS-PAGE, Isotope-Coded-Affinity-Tag (ICAT) labelling, mTRAQ labelling, Nano-LC-ESI-MS/MS, Human Albumin ELISA Kit, and S100-A9 protein quantification by ELISA238 proteins IdentifiedInnovative approach concluded the novel changes in host and microbial derived GCF proteome of periodontal patients[72]
Rody Jr et al.Collected from a deciduous second molar with radiographic evidence of root resorption on 1 quadrant (experimental site) and from the permanent first molar on the contralateral quadrant (control site) in the same jawPeriopaper strips (Oraflow, Plainview, NY, USA)One dimensional LC-MS and Two dimensional (2D) LC-MS2789 proteins in control group and 2421 proteins in root resorption groupMass spectrometry is useful tool for analyzing external root resorption[73]
Kinney et al.Collection from the mesio-buccal aspect of each site (tooth) for up-to 28 teeth per patientMethylcellulose strip (Pro Flow, Inc., Amityville, NY, USA)ELISA and Quantibody Human Cytokine Array (HCA) This method offer improved patient monitoring and disease control[74]
Huynh et al.Collection were chosen based on how well they represented the healthy, gingivitis and chronic periodontitis inclusion criteriaGlass-microcapillary tube (Drummond Scientific, Brookmall, PA, USA)One dimensional Gel-Electrophoresis and Nano-LC-ESI-MS121 proteins identifiedConcluded various biomarkers which differentiate between healthy periodontium, gingivitis and chronic periodontitis[35]
Table 3. Use of proteomics techniques for dental materials analysis.
Table 3. Use of proteomics techniques for dental materials analysis.
Author NameTitle of StudyOutcomesReferences
Boyan et al.Porcine fetal enamel matrix derivative enhances bone formation induced by demineralized freeze dried bone allograft in vivoEmdogain contains a number of low-molecular-weight proteins (mainly amelogenins), associated with cementogenesis and osteogenesis during tooth development[109]
Derhami et al.Proteomic analysis of human skin fibroblasts grown on titanium: Novel approach to study molecular biocompatibilityGain a better understanding of the molecular basis of biocompatibility of human skin fibroblast on titanium[110]
Koin et al.Analysis of the degradation of a model dental compositeLiquid chromatography mass spectrometry (LC-MS) found leaching of intact BisGMA and several degradation products that contained the bisphenol A moiety from the overlayer into distilled water after 2 weeks of aging[111]
Jung et al.Proteomic analysis in cyclosporin A (CsA)-induced overgrowth of human gingival fibroblast (HGF)The CsA-treated HGF demonstrated that Prx 1 may play a crucial role in the HGF proliferation induced by CsA and proteomic analysis data provide an efficient approach in understanding the mechanisms of HGF proliferation by CsA[112]
Taiyoji et al.Identification of proteinaceous inhibitors of a cysteine proteinase (an Arg-specific gingipain) from Porphyromonas gingivalis in rice grain, using targeted-proteomics approachesThese results suggest that these rice proteins may be useful as nutraceutical ingredients for the prevention and management of periodontal diseases[113]
Haigh et al.Alterations in the salivary proteome associated with periodontitisResults highlight the predominant involvement of S100 proteins in the host response during periodontitis[113]
Zilm and Bartold et al.Proteomic identification of proteinase inhibitors in the porcine enamel matrix derivative, EMD®Enamel matrix derivatives (EMD) contains a number of high-molecular-weight compounds which include the proteinase inhibitors, fetuin A and α1-antichymotrypsin[114]
Dorkhan et al.Effects of saliva or serum coating on adherence of Streptococcus oralis strains to titaniumThe adherence of LA11 and 89C strain to the moderately rough surfaces coated with saliva was more than twice that seen on the smooth saliva coated surfaces. This clearly demonstrates that surface topography is, at least to some degree, maintained in the presence of a saliva coating[115]
Zhao et al.Quantitative proteomic analysis of human osteoblast-like MG-63 cells in response to bio-inert implant material titanium and polyetheretherketone (PEEK)Titanium and polyetheretherketone (PEEK) induces similar response in osteoblast proteome and PEEK causing worse proliferation was related to mRNA processing[116]
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