Ontwikkeling van een Direct-Pulp aftopping model voor de evaluatie van pulpal wondgenezing en Reparative Dentin Formation in Muizen
Summary
We beschrijven een stap voor stap methode om direct pulp aftopping op muizen tanden voor de evaluatie van pulpa wondgenezing en vorming herstellende dentine in vivo.
Abstract
Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is often capped with biocompatible materials in order to expedite pulpal wound healing. The ultimate goal is to regenerate reparative dentin, a physical barrier that functions as a “biological seal” and protects the underlying pulp tissue. Although this direct pulp-capping procedure has long been used in dentistry, the underlying molecular mechanism of pulpal wound healing and reparative dentin formation is still poorly understood. To induce reparative dentin, pulp capping has been performed experimentally in large animals, but less so in mice, presumably due to their small sizes and the ensuing technical difficulties. Here, we present a detailed, step-by-step method of performing a pulp-capping procedure in mice, including the preparation of a Class-I-like cavity, the placement of pulp-capping materials, and the restoration procedure using dental composite. Our pulp-capping mouse model will be instrumental in investigating the fundamental molecular mechanisms of pulpal wound healing in the context of reparative dentin in vivo by enabling the use of transgenic or knockout mice that are widely available in the research community.
Introduction
Dental caries are one of the most prevalent oral diseases and the leading cause of surgical interventions to dentitions in almost all individuals1,2. The prognosis of surgical interventions and restorations of a tooth largely depends upon proper pulpal response and successful wound healing. Indeed, dental caries that penetrate deeply through the enamel and dentin frequently lead to the exposure of the underlying pulp tissue that is often “capped” with dental materials, such as calcium hydroxide (Ca(OH)2) or hydraulic calcium-silicate cements (HCSCs), including mineral trioxide aggregates (MTA). The ultimate goal of such a pulp-capping procedure is to expedite pulpal wound healing by regenerating reparative dentin, a physical barrier that functions as a “biological seal” to protect the underlying pulp tissue and to increase the life expectancy of the tooth and the overall oral health. However, the underlying mechanism of pulpal wound healing and reparative dentin formation is not fully understood.
To better understand the mechanisms of pulpal wound healing and reparative dentin formation in vivo, several animals were previously used, including monkeys, dogs, and pigs3-5. Among them, rats are frequently used because they are relatively smaller in sizes compared to the other animals, but their teeth are large enough to perform direct pulp capping without any technical difficulties6-10. These animal models are ideal alternatives to human studies for examining pulpal responses and reparative dentin formation. However, their utilization is limited to observational studies at the cellular level, and they scarcely provide mechanistic insights during reparative dentin formation at the molecular level.
Recent technical advances in genetic engineering provided invaluable and indispensable research tools-mice that harbor a gene that is either overexpressed or deleted-that are instrumental to studying molecular mechanisms of human diseases in vivo. The numbers of different strains of transgenic or knockout mice that are strategically inducible in a cell-specific manner are continually growing in the scientific community. Therefore, examining pulpal wound healing and reparative dentin regeneration in these mice would greatly help to expedite our understanding of these processes at the molecular level. However, the use of mice is significantly dampened, as performing a pulp-capping procedure on a mouse tooth is technically challenging due to its miniature size. Here, we present our reproducible method of performing direct pulp capping in mice for the evaluation of pulpal wound healing and reparative dentin formation in vivo.
Protocol
Representative Results
Discussion
Momenteel zijn er verschillende experimentele modellen voor de in vivo effecten van tandheelkundige materialen, scaffolds of groeifactoren op odontogene differentiatie van dentale pulp stamcellen (DPSC's) 13 te valideren. Deze modellen omvatten ectopische autologe transplantatie van DPSC's in een orgaan, zoals de renale capsule of subcutane transplantatie van DPSC in immuungecompromitteerde muizen met steigers 14,15. Echter, deze methoden beperkt hun odontogene effect op DPSC's…
Disclosures
The authors have nothing to disclose.
Acknowledgements
Deze studie werd ondersteund door R01DE023348 (RHK) van NIDCR / NIH en de Faculteit Research Grant (RHK) van de Raad voor onderzoek van de Academische Senaat van de afdeling Los Angeles van de Universiteit van Californië.
Materials
| BM-LED stereo microscope | MEIJI Techno | Microscope | |
| Optima MCX-LED | Bien Air Dental | 1700588-001 | Electic motor engine |
| isoflurane | Henry schein animal health | NDC 11695-0500-2 | |
| 1/4 round bur | Brasseler | 001092T0 | |
| Endodontic K-file | Roydent | 98947 | |
| ProRoot MTA | Dentsply | PROROOT5W | MTA |
| Paper point | Henry schein | 100-3941 | |
| Ultra-Etch | Ultradent product Inc. | Phosphoric acid etchant | |
| OptiBond SoloPlus | Kerr | 29669 | Adhesives |
| Coltolux LED | Coltene/whaledent Inc. | C7970100115 | Curing light unit |
| Characterization tint | Bisco | T-14012 | Flowable composite |
| Skyscan | Breuker | 1275 | uCT scanner |
| Microm | Thermo | HM355S | Microtome |
| Hematoxyline-1 | Thermo Scientific | 7221 | |
| Eosin-Y | Thermo Scientific | 7111 | |
| Cytoseal 60 | Thermo Scientific | 8310-16 | Mounting solution |
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