|Year : 2015 | Volume
| Issue : 3 | Page : 82-85
Mussel adhesive protein coating: A potential therapeutic method for self-healing of cracked teeth
Li Bo-Lin1, Cao Ying2, Li Quan-Li2
1 Hefei No 7 High School, Anhui Medical University, Hefei, China
2 College and Hospital of Stomatology, Anhui Medical University, Hefei, China
|Date of Web Publication||28-Aug-2015|
69#, Meishan Road, Hefei - 230 032
Source of Support: None, Conflict of Interest: None
Introduction: Nowadays, cracked tooth syndrome is the third main cause of tooth extraction, following caries and periodontal diseases, done in almost all the dental clinics. Nevertheless, the diagnosis and treatment of this condition remain controversial. All candidate therapeutics, such as occlusal adjustment, preventive filling, root canal therapy (RCT), and crown restoration, provide unpredictable outcomes. As such, methods to prevent further crack development and to induce crack self-healing must be developed. The Hypothesis: Mussels secreting adhesive foot protein (Mafp) can attach to various surfaces under aqueous conditions. In nature, mussels adhere to stones and deposit layer by layer through mineralization, thereby forming mussel-stone composites with excellent mechanical property. Given the natural process of mussel-stone complex formation, we hypothesize that application of Mafp coating at the crack interface may mineralize the cracks by capturing calcium and phosphate ions from the saliva. This process consequently leads to crack self-healing and complete restoration of the tooth structure. Evaluation of the Hypothesis: To test our hypothesis, we need to develop a model in vivo. Cracked teeth disks are adhered together using Mafp solution. Then, the tooth disks are sutured on the interior side of the cheeks. After regular intervals, the disks are removed and characterized. Scanning electron microscopy is performed to evaluate the morphology of the crack interface. Microhardness and shear bond strength are used to evaluate the mechanical property of the healing cracked zone. Transmission electron microscopy is also conducted to evaluate the crystallinity of the crack interface.
Keywords: Biomimetic mineralization, biomineralization, cracked tooth syndrome, mussel adhesive protein, self-healing
|How to cite this article:|
Bo-Lin L, Ying C, Quan-Li L. Mussel adhesive protein coating: A potential therapeutic method for self-healing of cracked teeth. Dent Hypotheses 2015;6:82-5
| Introduction|| |
A cracked tooth is an incomplete fracture of a vital tooth. At the early stage of this condition, the crack is often difficult to distinguish by the naked eyes. The diagnosis is also complicated and primarily based on symptoms. Superficial enamel cracks generally present no obvious symptoms [Figure 1]a], whereas cracks [Figure 1]b] invading the deep dentin typically present localized pain during chewing or biting and unexplained sensitivity to temperature stimuli, particularly cold. When a cracked tooth becomes symptomatic, the condition can lead to pulpitis and the tooth may need to be subjected to root canal therapy (RCT) [Figure 1]c] and crown restoration. Even after RCT, the status of the cracked tooth remains unpredictable and some cracked teeth may finally need extraction. The crack often results in the reinfection of root canal system of unsealed teeth. To date, few evidence-based guidelines are available regarding the prevention, diagnosis, and treatment of cracked teeth. , Therefore, in clinical dentistry, superficial cracks must be repaired by self-healing to prevent the crack to invade the deep dentin and the pulp.
|Figure 1: (a) craking in the incisor enamel, (b) craking in the molor, (c) cracking after root canal therapy|
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Enamel is the exterior layer of teeth exposed to the mouth environment. The enamel in the crown and cementum in the root cover dentin form the main body of a tooth. Enamel is composed of 96% hydroxyapatite (HA) crystals, which are inorganic minerals. The HA crystals assemble into a prism-like structure, which are tightly packed in an organized pattern to form a special enamel microstructure. Structurally, the basic microstructure of dentin comprises a calcified collagen matrix containing HA (approximately 70% by weight), organic matrix (mainly collagen, 20%), and water (10%). The organic substance of dentin is type-I collagen, which self-assembles into fibrils to form collagen matrix scaffolds.  As the enamel and the external part of dentin do not contain cells, they cannot self-repair once damaged. Therefore, a remineralizing strategy must be established to allow self-healing of cracks and prevention of superficial crack development.
Mussels can attach to various surfaces, ranging from natural inorganic and organic materials to synthetic materials, under aqueous conditions. This bioadhesive property is dependent on the mussels secreting adhesive foot protein (Mafp) that exhibits good biocompatibility and can be commercially produced. , The Mafp products are used in medicine as a bioadhesive for adhering skin cut and fractured bones. Mafp should be mineralize prior to applications in repairing cracked teeth. An Mafp derivative has been reported to induce biomineralization. ,, In this study, we hypothesize that Mafp coating applied to adhere to a cracked tooth causes biomimetic mineralization, thereby inducing the cracked tooth to self-repair.
| The Hypothesis|| |
In nature, mussels commonly adhere to stones and deposit layer by layer through mineralization [Figure 2]. Mussel-stone composites exhibit excellent mechanical property, which hardly deprives mussels of the stone body. Since tooth is a mineral composite, we used Mafp to treat the cracked tooth interface. The protein should adhere to the crack tooth and repair the crack through mineralization by capturing calcium and phosphate ions from the saliva.
L-3-(3,4-dihydroxyphenylalanine) (L-DOPA) is a major component of Mafp, and studies have suggested that DOPA is primarily responsible for the adhesive characteristics of the protein. Direct interactions between the catechol side chains of DOPA and the contacting surfaces have been considered as the mechanism of DOPA adhesion in the early stages. DOPA oxidation catalyzed by catechol oxidase with oxidizing agents produces DOPA-quinone or DOPA-semiquinone, which participates in Mafp cross-linking reactions via aryl-aryl coupling (formation of di-DOPA) or Michael additions with amine-containing protein residues. These reactions lead to the solidification of proteins. 
A universal biomimetic mineralization route, known as polydopamine-assisted hydroxylapatite (HA) formation (pHAF), has been established based on the natural mussel-adhesion mechanism. In this process, the material surface is activated by polydopamine coating (catecholamines located at the interfaces are involved in the binding). Subsequently, surface-anchored catecholamine moieties (catecholamines that do not participate in substrate adhesion) bind to calcium ions and enrich the interface with these ions. This process facilitates the formation of HA crystals that are aligned to the c-axis, parallel to the polydopamine layer. pHAF is an effective approach to create novel HA-based organic-inorganic hybrid biomaterials, regardless of the type, size, and shape of the hybridized counterpart materials. , Thus, Mafp may induce mineralization to form Mafp-HA composite to repair cracked teeth. The protocol for inducing self-repairing of cracked tooth by the application of Mafp is shown as follows:
- The tooth surface around the crack line is polished to remove contaminants such as biofilms. The surface is then etched with 20% H 3 PO 4 for approximately 20 s to freshen the interface of the crack line. Etching the tooth surface with H 3 PO 4 is a common method for dental adhesion in clinical dentistry.
- The pH of 1% concentration of the Mafp stock solution is adjusted to approximately 8.5 and FeCl 3 solution is added into the fresh Mafp solution with a final concentration of 3% FeCl 3 . A drop of the mixed fresh Mafp solution is applied on the tooth surface around the crack line. As the Mafp remains stable under acidic pH, the stock solution is acidic with 1% acetic acid. However, bioadhesion requires Mafp to polymerize, which usually occurs under alkaline pH. Given that the pH of ocean is approximately 8.5, we adjusted the Mafp solution to this pH to promote Mafp polymerization and adhesion to the tooth surface. , Ferric ions can promote Mafp polymerization  and dentin adhesion because they condition the dentin surface. 
- The cracked line self-heals. The Mafp permeates into the crack line interface and polymerizes to adhere to the cracked tooth. The polymerized Mafp captures calcium ions from the saliva and the mineralization of Mafp is gradually induced by pHAF. , Finally, the interface between the cracked teeth completely mineralizes to self-repair the cracked tooth, which is similar to bone fracture healing [Figure 3].
| Evaluation of the Hypothesis|| |
Preparation of the cracked tooth model
Extracted sound human molars are collected and 2-mm-thick dental slices, including enamel and dentin, are obtained perpendicular to the longitudinal axis of each tooth by using a low-speed diamond saw. The dental slices are polished with a series of silicon carbide papers and then ultrasonically cleaned with acetone, ethanol, and deionized water. The slices are stored in a polyethylene tube at 4°C prior to use.
The dental slices are fractured along the middle line and the fragments are treated with manual diaplasis and splint immobilization with a 0.2-mm stainless wire.
The as-prepared cracked tooth model can be divided into two groups. For the experimental group, half of the teeth are etched with 20% H 3 PO 4 for approximately 20 s and then coated with fresh Mafp solution. For the control group, the remaining teeth are etched with 20% H 3 PO 4 for approximately 20 s.
In vivo experiments are performed on male Beagle dogs. The tooth disks are sutured on the interior side of the cheeks, with the experimental disk on the left side and the control disk on the right side. The disks are removed after 2 weeks, 4 weeks, and 8 weeks. The disks are then thoroughly rinsed with deionized water, air-dried and then evaluated.
Evaluation of crack healing
Scanning electron microscopy is performed to evaluate the morphology of the crack interface. Microhardness and shear bond strength are used to evaluate the mechanical property of the healing cracked zone. Transmission electron microscopy is also conducted to evaluate the crystallinity of the crack interface.
Financial support and sponsorship
This work was supported by NSFC grant no. 81400559.
Conflicts of interest
There are no conflicts of interest.
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[Figure 1], [Figure 2], [Figure 3]