Establishment of a Murine Pulp Exposure Model with a Novel Mouth-Gag for Pulpitis Research

Summary

This article presents a streamlined protocol for establishing a pulpitis model in mice using an innovative mouth-gag, followed by subsequent histological analysis.

Abstract

Pulpitis, a common cause of natural tooth loss, leads to necrosis and loss of bioactivity in the inflamed dental pulp. Unraveling the mechanisms underlying pulpitis and its efficient treatment is an ongoing focus of endodontic research. Therefore, understanding the inflammatory process within the dental pulp is vital for improving pulp preservation. Compared to other in vitro experiments, a murine pulpitis model offers a more authentic and genetically diverse context to observe the pathological progression of pulpitis. However, using mice, despite their cost-effectiveness and accessibility, poses difficulties due to their small size, poor coordination, and low tolerance, complicating intraoral and dental procedures. This protocol introduces a novel design and application of a mouth-gag to expose mouse pulp, facilitating more efficient intraoral procedures. The mouth-gag, comprised of a dental arch readily available to most dentists and can significantly expedite surgical preparation, even for first-time procedures. Micro-CT, hematoxylin-eosin (HE) staining, and immunofluorescence staining were used to identify changes in morphology and cell expression. The aim of this article is to help researchers establish a more reproducible and less demanding procedure for creating a pulp inflammation model using this novel mouth-gag.

Introduction

The dental pulp, an integral part of the tooth, is responsible for multiple essential functions such as nutrient supply, dentin formation, sensory function, and defense reactions¹. Nevertheless, the dental pulp, surrounded by hard tissue, is susceptible to injuries and damages from deep caries, pulpitis, trauma, or subsequent therapies^2,3. The absence of functional dental pulp increases the risk of tooth fragility⁴. Moreover, the loss of pulp vitality in young permanent teeth can adversely affect tooth maturation, and current denture techniques fail to restore th....

Protocol

The experimental procedures in this study were approved by the ethical committee of the West China School of Stomatology, Sichuan University (WCHSIRB-D-2021-125). Adult C57BL/6 mice were obtained from Gempharmatech Experimental Animals Company, Chengdu, China. The whole crown of the maxillary first molar erupts 21 days after birth. Mice for surgery should be older than 21 days with normal vitality¹¹. Here, 6 to 8-week-old mice were used for modeling. Figure 1 is a flo.......

Discussion

As the solitary soft tissue within teeth, dental pulp fulfills a crucial role in maintaining bioactivity of tooth but remains highly sensitive. The preservation of this vital pulp has become the preferred initial approach in recent endodontic treatments, necessitating a comprehensive understanding of the inflammatory mechanisms of dental pulp¹⁶. The spatiotemporal fluctuation of the inflammatory microenvironment and interactions between resident cell types in pulpitis complicates its investigation.......

Materials

Name	Company	Catalog Number	Comments
Animal
C57/B6J mice	Gempharmatech Experimental Animals Company	C57/B6J	For the establishment of pulp exposure
Equipment
1 mL syringe	Chengdu Xinjin Shifeng Medical Apparatus & Instruments Co. LTD.	SB1-074(IV)	Apply in drug injection.
8# C+ file	Readysteel	0010047	Apply in exposing the roof of pulp chamber.
Anesthesia Mix solution			10% ketamine hydrochloride+ 5% xylazine + 85% sterile isotonic saline.
DAPI Staining Solution	Beyotime	C1005	Apply in immunofluorescence staining for counter-staining of nucleus.
Dental high-speed dental handpiece	Jing yuan electronic commerce technology	WJ-422	Apply in pulp exposure.
Heavy wire cutter	Jirui Medical Instrument Co., Ltd.	none	Apply inarc cutting.
Hematoxylin and Eosin Stain kit	Biosharp	BL700B	For the histological analysis of the slides.
IL-6 antibody	Novus	NBP2-89149	Apply in immunofluorescence staining to detect the inflammation of the dental pulp.
Ketamine(Ketamine hydrochloride)	Vet One, Boise, Idaho, USA	C3N VT1	100mg/kg, IP. Apply in nesthetization.
Medical tap	3M	1530	Apply in mice immobilization.
Orthodontic arch wire	Shanghai Wei Rong Medical Apparatus Co. LTD.	K417	Diameter of 8µm
Round dental burr (0.6 mm)	Shofu global	072208	Apply in removing enamel and shallow layer of dentin.
Young loop bending plier	Jirui Medical Instrument Co., Ltd.	none	Apply in arc bending.