Development of a Direct Pulp-capping Model for the Evaluation of Pulpal Wound Healing and Reparative Dentin Formation in Mice

Summary

We describe a step-by-step method of performing direct pulp capping on mice teeth for the evaluation of pulpal wound healing and reparative dentin formation 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 individuals^1,2. 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 i....

Protocol

Mice were purchased from Jackson Laboratory and kept in a pathogen-free vivarium in the UCLA Division of Laboratory Animal Medicine (DLAM). The experiments were performed according to the approved institutional guidelines from the Chancellor's Animal Research Committee (ARC#2016-037).

1. Mouse Anesthetization

1. Use eight-week-old female C57/BL6 mice (n = 3).
2. Anesthetize the mice using ketamine (80-120 mg/kg of mouse weight)/xylazine (5 mg/kg of mouse weight) solutions and administer intraperitoneally (ip) at a dose of 10 mL/kg.
3. Prepare ketamine (80 - 120 mg/kg)/xylazine (5 mg/kg) solutions and administer them intraperitone....

Results

Here, we showed the step-by-step procedures to perform pulp capping on mice teeth. One of the key aspects of pulp capping in mice is to have the appropriate apparatus. In this regard, having the microscope with a 10X power magnification is essential (Figure 1A). To create a Class-I-like preparation in the tooth, we used a ¼-round burr in an electric high speed handpiece at 200,000 rpm (Figure 1B). Alternatively, any other engines, including those that use.......

Discussion

Currently, there are several different experimental models available to validate the in vivo effects of dental materials, scaffolds, or growth factors on odontogenic differentiation of dental pulp stem cells (DPSCs)¹³. These models include ectopic autologous transplantation of DPSCs into an organ, such as the renal capsule, or subcutaneous transplantation of DPSCs into immunocompromised mice with scaffolds^14,15. However, these methods are limited in that their odontogenic effect on DPSCs is.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
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