This work was supported by the National Natural Science Foundation of China grant 82100982 to F.L.

The animal procedures in this study were reviewed and approved by the Ethical Committee of the West China School of Stomatology, Sichuan University (WCHSIRB-D-2017-041). Adult C57BL/6 mice were used in this study (see the Table of Materials). This protocol adds mechanical load to the right maxillary first molar (M1) for mesial movement where a highly coordinated bone remodeling process is induced by coupling of bone resorption and bone formation (Figure 1).
1. Preoperative preparation
<ol>
	<li>Surgical items
	<ol>
		<li>Prepare the following surgical items for the operation: surgical platform (Figure 2A), fastener (Figure 2B), surgical instruments (Figure 2C and Supplementary Figure S1), orthodontic supplies (Figure 2C), and dental restoration supplies (Figure 2D). 
		NOTE: The customized coil spring is custom-made and provides a force of 10cN when stretched to 10 mm.</li>
	</ol>
	</li>
	<li>Sterilization
	<ol>
		<li>Sterilize the surgical instruments by autoclaving and all the surgical items with ultraviolet irradiation for at least 30 min.</li>
	</ol>
	</li>
	<li>Anesthesia
	<ol>
		<li>Anesthetize the mouse by administering ketamine (100 mg/kg) and diazepam (5 mg/kg) by intraperitoneal injection.</li>
		<li>Apply vet ointment to the eyes of the murine with a cotton stick to avoid eye dryness.</li>
		<li>Proceed with the surgery only when the mouse does not respond when its toes are pinched with forceps.</li>
	</ol>
	</li>
</ol>
2. Surgical process
<ol>
	<li>Spread and tape the limbs of the anesthetized mouse in a supine position to the surgical platform using adhesive tape.</li>
	<li>Pin a 27 G needle on either side above the head and another 27 G needle on either side below the axilla.</li>
	<li>Wind a rubber band around the above two needles and the upper incisors and another one around another two&#160;needles and the lower incisors. Change the needle positions to control the degree of opening and the orientation of the mouth (Figure 3A). 
	NOTE: For the orthodontic tooth movement operation, keep the mouth open to the maximum extent before the buccinator becomes completely tight. The tongue should be pulled towards the non-operative side to expose the surgical field and prevent ischemia.</li>
	<li>Bend the 1.5 mm end of a 3 cm 304 stainless steel wire and push the bent end through the interproximate space between the M1 and the maxillary second molar (M2) from the buccal side with curved ophthalmic tweezers (Figure 3B). When the palatal end of the ligature wire is seen from the palatal side, pull it out up to about half of its length and pass it through one end of the customized coil spring.</li>
	<li>Tie a square knot with the two ends of the ligature wire in the mesial direction of the maxillary M1 until the spring is firmly fixed to the tooth (Figure 3C). Subtract the excess wire.</li>
	<li>Similarly, pierce a second 3 cm 304 stainless steel wire through the other end of the coil spring.</li>
	<li>Clean and dry the incisors&#39; surfaces with cotton balls. Apply adhesives on all those surfaces with cotton sticks and light-cure them.</li>
	<li>Push the second stainless steel wire through the interproximate space between the maxillary incisors and tie a slip knot in the labial direction (Figure 3D). Subtract the excess wire and make the rest of the wire lie close to the tooth surface.</li>
	<li>Inject light-cured resin to cover the knot and incisors; light-cure the resin (Figure 3E).</li>
</ol>
3. Postoperative management
<ol>
	<li>After surgery, inject the mice with 0.05 mg/kg buprenorphine intraperitoneally&#160;for postoperative analgesia.</li>
	<li>Place the anesthetized mouse on a 37 &#176;C thermostatic electric blanket. When the murine regains consciousness with ambulation, return it to a separate housing cage.</li>
	<li>Due to the limited functioning of the incisors after the surgery, replace regular hard fodder with only a soft diet.</li>
	<li>Check the orthodontic appliances every day. If any condition are observed during the inspection that affects the conduction of orthodontic force, such as spring deformation, spring loosening, and device falling off, the mouse should be excluded from the experiment.</li>
	<li>In order to maintain the comparability of experiments, assess the weight of the mice daily post-surgery. Any mice exhibiting a weight loss exceeding 30% of their preoperative weight must be excluded from the experiment.</li>
</ol>

A Murine Maxillary Orthodontic Model Guide to Bone Remodeling Analysis

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The authors declare no conflicts of interest.

In this paper, we tried to describe the simplest orthodontic tooth movement protocol on murine maxillary model step by step to study the latent mechanisms of mechanical load-induced bone remodeling. Apart from research on bone remodeling, there are some other mainstream applications of this method: 1) methodological research on the acceleration of orthodontic tooth movement; 2) research on orthodontic root resorption; 3) biological mechanisms of orthodontic tooth movement and pain; 4) research on the transgenic model.</p...

Bone is a highly active reconstructed tissue that adapts its size, shape, and properties through the lifetime of the individual1,2. In addition to hormones, aging, nutrition, and other biological or biochemical factors3, the idea that mechanical load is the most determining factor has garnered general acceptance4,5. Under some circumstances with abnormal mechanical load, the imbalance between bone resorption and bone formation may lead to abnormal bone remodeling and bone disorders. Bone diseases such as disuse osteoporosis and bone loss during long-term bed rest or in the presence of microgravity at spaceflight have a close relationship with abnormal mechanical load6,7,8.
Mechanical load has also been used to treat bone-related diseases such as distraction treatment and orthodontic treatment. Distraction treatment has been used in developmental diseases such as craniosynostosis and mandibular hypoplasia9,10, while orthodontic treatment has been widely used to rectify abnormal teeth position and any malocclusion11. The core of orthodontic treatment is also the management of mechanical load. When the bone tissue is subjected to mechanical load, a highly coordinated bone remodeling process is induced by coupling of bone resorption followed by bone formation, which can move teeth to achieve the orthodontic purpose12,13.
Although orthodontic treatment has been widely applied for clinical practice, as our knowledge of the biological effects of mechanical load is limited, the results of orthodontic treatment are uncontrollable. To overcome these limitations, several animal models such as mouse, rat, rabbit, cat, dog, monkey, and pig have been established to investigate the underlying mechanism of mechanical load-induced bone remodeling (Table 1)14,15,16,17,18,19,20,21,22,23,24,25,26,27,28,29,30,31,32. Large animals such as dogs, monkeys, and pigs have some advantages over small animals in orthodontic operation-they have more human-like teeth and dentition so that the surgical procedure is easy to replicate in humans. Additionally, a wide view can reduce the operation difficulty and make it possible to apply a variety of orthodontic schemes33,34. However, large animals are difficult to obtain, leading to challenges related to sample size, and they are subject to ethical restrictions35. Furthermore, routine extraction procedures and complex instruments make the experiments difficult to perform due to which large animals are rarely used.
Under such circumstances, rodents are mainly used to establish orthodontic models. Among these models, rats and rabbits have lower operating difficulty and more tooth movement schemes compared with mice. However, the murine model has the unique advantage that there are a large number of genetically modified mice available, which is especially crucial for investigating the underlying mechanisms36. However, the murine model is the most difficult model to manipulate because of its small size. Reviewing the current methods, moving the first molar in the mesial direction is the only practical method for an orthodontic model. Two devices are mainly used to move the tooth-coil spring and elastic band. Using an elastic band is easier, but the orthodontic force varies greatly, which makes it difficult to obtain stable results.
Xu et al.15 have established a murine model with a coil spring on the mandible. However, due to the mobility of the mandible and the obstructive nature of the tongue, operation on the maxilla is always the first choice for both intraoperative and postoperative considerations. Taddei et al.16 described a more detailed protocol on the murine maxilla 10 years ago and more visual and pellucid details should be added. In summary, this protocol has systematically described a detailed orthodontic tooth movement protocol in a murine maxillary model to help researchers master the modeling method in a standardized way and enable the comparative evaluation between different studies.

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Experimental Models: Mouse Lines</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>C57/B6J&#160;</td>
			<td>Gempharmatech Experimental Animals Company&#160;</td>
			<td>C57/B6J</td>
			<td></td>
		</tr>
		<tr>
			<td>Critical Commercial Assays</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Hematoxylin and Eosin Stain Kit</td>
			<td>Biosharp</td>
			<td>BL700B</td>
			<td></td>
		</tr>
		<tr>
			<td>Masson&#8217;s Trichrome Stain Kit</td>
			<td>Solarbio</td>
			<td>G1340</td>
			<td></td>
		</tr>
		<tr>
			<td>Instruments</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>27 G needle</td>
			<td>Chengdu Xinjin Shifeng Medical Apparatus &#38; Instruments Co. LTD.</td>
			<td>SB1-074(IV)</td>
			<td></td>
		</tr>
		<tr>
			<td>Adhesives</td>
			<td>Minnesota Mining and Manufacturing Co., Ltd.</td>
			<td>41282</td>
			<td></td>
		</tr>
		<tr>
			<td>Corkboard</td>
			<td>DELI Group Co., Ltd.</td>
			<td>8705</td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton balls</td>
			<td>Haishi Hainuo Group Co.,&#160; Ltd.</td>
			<td>20120047</td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton sticks</td>
			<td>Lakong Medical Devices Co., Ltd.</td>
			<td>M6500R</td>
			<td></td>
		</tr>
		<tr>
			<td>Customized coil spring</td>
			<td>Chengdu Mingxing Spring Co., Ltd.</td>
			<td>1109-02</td>
			<td></td>
		</tr>
		<tr>
			<td>Forceps</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>none</td>
			<td></td>
		</tr>
		<tr>
			<td>Light-cured fluid resin</td>
			<td>Shofu Dental Trading (SHANGHAI) Co., Ltd.</td>
			<td>518785</td>
			<td></td>
		</tr>
		<tr>
			<td>Light curer</td>
			<td>Liang Ya Dental Equipment Co., Ltd.</td>
			<td>LY-A180</td>
			<td></td>
		</tr>
		<tr>
			<td>Medical adhesive tapes&#160;</td>
			<td>Haishi Hainuo Group Co.,&#160; Ltd.</td>
			<td>0008-2014</td>
			<td></td>
		</tr>
		<tr>
			<td>Medical non-woven fabric</td>
			<td>Henan Yadu Industrial Co., Ltd.</td>
			<td>01011500018</td>
			<td></td>
		</tr>
		<tr>
			<td>Needle holders</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>none</td>
			<td></td>
		</tr>
		<tr>
			<td>Rubber bands</td>
			<td>Haishi Hainuo Group Co.,&#160; Ltd.</td>
			<td>32X1</td>
			<td></td>
		</tr>
		<tr>
			<td>Surgical scissors</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>none</td>
			<td></td>
		</tr>
		<tr>
			<td>Tweezers</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>none</td>
			<td></td>
		</tr>
	</tbody>
</table>

the establishment of a murine maxillary orthodontic model

Due to the lack of reproducible protocols for establishing a murine maxillary orthodontic model, we present a reliable and reproducible protocol to provide researchers with a feasible tool to analyze mechanical loading-associated bone remodeling. This study presents a detailed flowchart in addition to different types of schematic diagrams, operation photos, and videos. We performed this protocol on 11 adult wide-type C57/B6J mice and harvested samples on postoperative days 3, 8, and 14. The micro-CT and histopathological data have proven the success of tooth movements coupled with bone remodeling using this protocol. Furthermore, according to the micro-CT results on days 3, 8, and 14, we have divided bone modeling into three stages: preparation stage, bone resorption stage, and bone formation stage. These stages are expected to help researchers concerned with different stages to set sample collection time reasonably. This protocol can equip researchers with a tool to carry out regenerative analysis of bone remodeling.

We have performed the OTM surgery on 11 adult male mice (C57/BL6, 3 months old). They were euthanized for results on days 3, 8, and 14 post surgery. In these experiments, the right maxillary side is the operation side, while the left maxillary side is the control side. The micro-CT showed that there was a temporal consecutive increase in the distance between M1 and M2: 30 &#181;m, 70 &#181;m, and 110 &#181;m at days 3, 8, and 14 post surgery, respectively (Figure 4). The low-density periodon...

Watch this Scientific Journal Video about Author Spotlight: Insights into an Efficient Murine Maxillary Orthodontic Model Protocol at JoVE.com

Author Spotlight: Insights into an Efficient Murine Maxillary Orthodontic Model Protocol

Here we demonstrate step-by-step a manageable, orthodontic tooth movement protocol operated on a murine maxillary model. With the explicit explanation of each step and visual demonstration, researchers can master this model and apply it to their experimental needs with a few modifications.

The Establishment of a Murine Maxillary Orthodontic Model

Due to the lack of reproducible protocols for establishing a murine maxillary orthodontic model, we present a reliable and reproducible protocol to ...

As we have reviewed, murine maxillary orthodontic model is widely used, but simply described. It is difficult to repeat and to compare experimental data among different studies based on current methods. Therefore, this protocol will help researchers master the modeling method in a standardized way and enable comparative evaluations between different studies.This protocol is actually the simplest modeling method in murine to date, and our video has shown a lot of practical modeling details. This method is based on murine and the narrowing operation space will definitely bring great challenges to beginners. A certain amount of practice can be helpful.This protocol provides orthodontic load in the simplest way, so experiments on this protocol will be time-saving and easy to repeat. There are also lots of advantages for novices, such as wide surgical field of view and visual surgical procedures. Zero-based learner can master this model easier with our protocol.

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JoVE Journal

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590 Views.  Sichuan University. As we have reviewed, murine maxillary orthodontic model is widely used, but simply described. It is difficult to repeat and to compare experimental data among different studies based on current methods. Therefore, this protocol will help researchers master the modeling method in a standardized way and enable comparative evaluations between different studies.This protocol is actually the simplest modeling method in murine to date, and our video has shown a lot of practical modeling deta...

Video: Author Spotlight: Insights into an Efficient Murine Maxillary Orthodontic Model Protocol

Murine Model of Wound Healing

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. Impaired wound healing, which occurs in diabetic patients for example, can lead to severe unfavorable outcomes such as amputation. There is, therefore, an increasing impetus to develop novel agents that promote wound repair. The testing of these has been limited to large animal models such as swine, which are often impractical. Mice represent the ideal preclinical model, as they are economical and amenable to genetic manipulation, which allows for mechanistic investigation. However, wound healing in a mouse is fundamentally different to that of humans as it primarily occurs via contraction. Our murine model overcomes this by incorporating a splint around the wound. By splinting the wound, the repair process is then dependent on epithelialization, cellular proliferation and angiogenesis, which closely mirror the biological processes of human wound healing. Whilst requiring consistency and care, this murine model does not involve complicated surgical techniques and allows for the robust testing of promising agents that may, for example, promote angiogenesis or inhibit inflammation. Furthermore, each mouse acts as its own control as two wounds are prepared, enabling the application of both the test compound and the vehicle control on the same animal. In conclusion, we demonstrate a practical, easy-to-learn, and robust model of wound healing, which is comparable to that of humans.

A murine model of cutaneous wound healing that can be used to assess therapeutic compounds in physiological and pathophysiological settings.

Wound healing and repair are the most complex biological processes that occur in human life. After injury, multiple biological pathways become activated. ...

A Preclinical Murine Model of Hepatic Metastases

Numerous murine models have been developed to study human cancers and advance the understanding of cancer treatment and development. Here, a preclinical, murine pancreatic tumor model of hepatic metastases via a hemispleen injection of syngeneic murine pancreatic tumor cells is described. This model mimics many of the clinical conditions in patients with metastatic disease to the liver. Mice consistently develop metastases in the liver allowing for investigation of the metastatic process, experimental therapy testing, and tumor immunology research.

A preclinical, murine model of hepatic metastases performed via a hemispleen injection technique.

Numerous murine models have been developed to study human cancers and advance the understanding of cancer treatment and development. Here, a preclinical, ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

3D Imaging of PDL Collagen Fibers during Orthodontic Tooth Movement in Mandibular Murine Model

Orthodontic tooth movement is a complex biological process of altered soft and hard tissue remodeling as a result of external forces. In order to understand these complex remodeling processes, it is critical to study the tooth and periodontal tissues within their 3D context and therefore minimize any sectioning and tissue artefacts. Mouse models are often utilized in developmental and structural biology, as well as in biomechanics due to their small size, high metabolic rate, genetics and ease of handling. In principle this also makes them excellent models for dental related studies. However, a major impediment is their small tooth size, the molars in particular. This paper is aimed at providing a step by step protocol for generating orthodontic tooth movement and two methods for 3D imaging of the periodontal ligament fibrous component of a mouse mandibular molar. The first method presented is based on a micro-CT setup enabling phase enhancement&#160;imaging of fresh collagen tissues. The second method is a bone clearing method using ethyl cinnamate that enables imaging through the bone without sectioning and preserves endogenous fluorescence. Combining this clearing method with reporter mice like Flk1-Cre;TdTomato provided a first of its kind opportunity to image the 3D vasculature in the PDL and alveolar bone.

We present a protocol for generating orthodontic tooth movement in mice and methods for 3D visualization of the collagen fibers and blood vessels of periodontal ligament without sectioning.

Orthodontic tooth movement is a complex biological process of altered soft and hard tissue remodeling as a result of external forces. In order to ...

Establishment and Evaluation of a Porcine Vein Graft Disease Model

Venous graft disease (VGD) is the leading cause of coronary artery bypass graft (CABG) failure. Large animal models of CABG-VGD are needed for the investigation of disease mechanisms and the development of therapeutic strategies. 
To perform the surgery, we enter the cardiac chamber through the third intercostal space and carefully dissect the internal mammary vein and immerse it in normal saline. The right main coronary artery is then treated for ischemia. The target vessel is incised, a shunt plug is placed, and the distal end of the graft vein is anastomosed. The ascending aorta is partially blocked, and the proximal end of the graft vein is anastomosed after perforation. The graft vein is checked for patency, and the proximal right coronary artery is ligated. 
CABG surgery is performed in minipigs to harvest the left internal mammary vein for its use as a vascular graft. Serum biochemical tests are used to evaluate the physiological status of the animals after surgery. Ultrasound examination shows that the proximal, middle, and distal end of the graft vessel are unobstructed. In the surgical model, turbulent blood flow in the graft is observed upon histological examination after the CABG surgery, and venous graft stenosis associated with intimal hyperplasia is observed in the graft. The study here provides detailed surgical procedures for the establishment of a repeatable CABG-induced VGD model.

In this protocol, novel pig vein bypass grafting was performed through a small incision in the left chest wall without cardiopulmonary bypass. A postoperative pathology study was done, which showed intimal thickening.

Venous graft disease (VGD) is the leading cause of coronary artery bypass graft (CABG) failure. Large animal models of CABG-VGD are needed for the ...

Establishment of a Simple and Effective Rat Model for Intraoperative Parathyroid Gland Imaging

Parathyroid gland (PG) identification is a critical unmet need in thyroidectomy. The identification of the PG is challenging in thyroid surgery as it is similar in color to the thyroid gland. The lack of effective animal models in preclinical research is a severe limitation for the development of PG identification techniques. This protocol allows for the establishment of a simple and effective rat model for PG identification. In this model, black iron oxide nanoparticles (IONPs) are injected locally in the thyroid gland and rapidly diffuse within the thyroid gland but not the PG. A negatively stained PG and a positively stained thyroid gland can be easily identified by the naked eye without requiring external microscopes. The position of the PG can be identified by increasing the color contrast between the thyroid gland and the PG, based on the color of the black IONPs. This rat model is low-cost and convenient for PG identification, and the IONPs are a novel PG contrast agent.

To date, the development of parathyroid gland (PG) identification methods is limited by the lack of animal models in preclinical research. Here, we establish a simple and effective rat model for intraoperative PG imaging and evaluate its effectiveness by using iron oxide nanoparticles as a novel PG contrast agent.

Parathyroid gland (PG) identification is a critical unmet need in thyroidectomy. The identification of the PG is challenging in thyroid surgery as it is ...

Analyzing Alveolar Bone Remodeling to Explore Skeletal Mechanical Biology

Using Inducible Osteoblastic Lineage-Specific Stat3 Knockout Mice to Study Alveolar Bone Remodeling During Orthodontic Tooth Movement

The alveolar bone, with a high turnover rate, is the most actively-remodeling bone in the body. Orthodontic tooth movement (OTM) is a common artificial process of alveolar bone remodeling in response to mechanical force, but the underlying mechanism remains elusive. Previous studies have been unable to reveal the precise mechanism of bone remodeling in any time and space due to animal model-related restrictions. The signal transducer and activator of transcription 3 (STAT3) is important in bone metabolism, but its role in osteoblasts during OTM is unclear. To provide in vivo evidence that STAT3 participates in OTM at specific time points and in particular cells during OTM, we generated a tamoxifen-inducible osteoblast lineage-specific Stat3 knockout mouse model, applied orthodontic force, and analyzed the alveolar bone phenotype. 
Micro-computed tomography (Micro-CT) and stereo microscopy were used to access OTM distance. Histological analysis selected the area located within three roots of the first molar (M1) in the cross-section of the maxillary bone as the region of interest (ROI) to evaluate the metabolic activity of osteoblasts and osteoclasts, indicating the effect of orthodontic force on alveolar bone. In short, we provide a protocol for using inducible osteoblast lineage-specific Stat3 knockout mice to study bone remodeling under orthodontic force and describe methods for analyzing alveolar bone remodeling during OTM, thus shedding new light on skeletal mechanical biology.

Author Spotlight: Comparing Alveolar and Long Bone Remodeling to Explore OTM Model Potential 

This study provides a protocol for using inducible osteoblast lineage-specific Stat3 knockout mice to study bone remodeling under orthodontic force and describes methods for analyzing alveolar bone remodeling during orthodontic tooth movement, thus shedding light on skeletal mechanical biology.

The alveolar bone, with a high turnover rate, is the most actively-remodeling bone in the body. Orthodontic tooth movement (OTM) is a common artificial ...

Creating a Pulp Inflammation Mouse Model Using a Novel Mouth-Gag 

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

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.

Author Spotlight: Enhancing Dental Pulp Research with Improved Mouse Models

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