This work is supported by National Natural Science Foundation of China 81825005 (L.Y.), 82201045 (F.Y.), and 82100982 (F.L.), and by Sichuan Province Science and Technology Program 2021JDRC0144 (F.L.), 2022JDRC0130 (F.Y.).

All 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, obtained from a commercial source (see Table of Materials), were used for the present study.
1. Presurgical preparation
<ol>
	<li>Instrument preparation
	<ol>
		<li>Prepare various needles of disposable syringes (26G, 25G, 23G; see Table of Materials) to be used as elevators. Inflect the needle head approximately 20&#176;-40&#176;, as shown in Figure 1.</li>
		<li>Prepare toothed ophthalmic tweezers as forceps. Ensure that it fits the size of the mice&#39;s molars and can grab the molars firmly. The ideal tweezer size is shown in Figure 1A, B3.</li>
		<li>Obtain a rubber band (which can be torn from a medical latex glove) to be used as a mouth opener, a foam board or corkboard as the surgical platform, a headlamp to light up the surgical area, and a heating pad for postoperative revival. Tear a dry cotton ball into small pieces&#160;and apply it to the surgical site if bleeding occurs&#160;during the procedure.</li>
	</ol>
	</li>
	<li>Anesthesia preparation
	<ol>
		<li>The mouse can be anesthetized by any suitable anesthetic protocol that achieves general anesthesia confirmed through the &#39;toe pinch&#39; method. Following anesthesia, apply veterinary ointment on the eyes of the mouse. 
		&#8203;NOTE: Preferred general anesthetic protocol could be: induction and maintenance with xylazine (10 mg/kg) and ketamine (100 mg/kg) intraperitoneally (IP); 
		For this study,&#160;isoflurane and 1% pentobarbital sodium (50 mg/kg, IP) were used for anesthesia induction and maintenance with additional doses as needed.</li>
	</ol>
	</li>
	<li>Disinfection and Sterilization preparation
	<ol>
		<li>Disinfect the operation platform and the open-air overhead with a 75% ethanol sprayer. Prior to each mouse procedure, the use of a new set of disposable needles is recommended. 
		NOTE: The toothed tweezers are reusable and can be sterilized using any preferred&#160;method such as steam sterilization.</li>
	</ol>
	</li>
</ol>
2. Surgical process
<ol>
	<li>Fixation of the mouse and its mandible
	<ol>
		<li>Tie the mouse to a surgical platform in a supine position using adhesive tape. Pin two 26 G needles in line with the orbital ear plane and another two 26 G needles at ease below the mandible.</li>
		<li>Apply the rubber band around the needles and cross the incisors to hold the mouth open. Slightly pull the tongue out and fix it under the rubber band opposite the surgical side, to prevent it from obstructing the field of view (Figure 1C).</li>
	</ol>
	</li>
	<li>Eliminating the distal resistance
	<ol>
		<li>Hold the molar with tweezers mesially, force a 23 G needle into the buccal alveolar bone of the distal root, and render an interval. 
		NOTE: This step must be taken with great caution, as a needle that is embedded too deep will likely snap the root.</li>
		<li>Next, change to a 25 G needle to continue to expand the interval, and progress delicately toward the periapical area while slowly rotating the needle forward and lingually (with the mouse anatomical position) to press the root out of the alveolar fossa.</li>
	</ol>
	</li>
	<li>Eliminating the mesial resistance
	<ol>
		<li>When there is enough space, use a 23 G needle to insert into the root fork and lift the molar occlusally. After holding the molar tightly, take another 23 G needle and force it into the lingual mesial periodontal membrane to create an interval.</li>
		<li>Then, substitute with a 25 G needle and slowly rotate forward and buccally. If some underlying hindrances impede the luxation of the molar, use a 26 G needle to penetrate the root apex, and repeat the operations.</li>
	</ol>
	</li>
	<li>Final extraction
	<ol>
		<li>Extract the tooth and, while extracting, ensure that the crown rises high above the occlusal plane and that two intact roots can be clearly seen. 
		NOTE: The luxation (dislocation) of the tooth is considered the most painful and distressing moment of the procedure. Prior to dislocating the tooth, the anesthetic depth should be reassessed using the toe pinch test, and accordingly, a moderate dose of an anesthetic agent should be administered if necessary.&#160;</li>
	</ol>
	</li>
	<li>Post-operative care
	<ol>
		<li>After extracting the tooth, apply dry cotton to stop bleeding, reposition the tongue, administer carprofen (5 mg/kg) subcutaneously, and put the mouse on a constant temperature heating pad until recovery from anesthesia.</li>
	</ol>
	</li>
</ol>
3. Imaging of the mouse mandible and the extraction socket
<ol>
	<li>Preparation of the samples
	<ol>
		<li>Euthanize the mouse by cervical dislocation. Use ophthalmic scissors to cut the skeleton muscles attached to the mandible and the zygomatic arch. Cut from the throat along the lower edge of the mandible to the ascending branch and then to the backside of the condyle, and then pull the mandible down and cut along the lower incisor midline. This way, the two separate mandibles are harvested.</li>
		<li>Perform fixation, demineralization, and dehydration following the standard procedure17. When embedding17, ensure that the occlusal plane is parallel with the edge of the cassette (see Table of Materials). Fix the mandible on the bottom of the cassette, with its lower edge cocked (Figure 2). 
		NOTE: This protocol offers the sagittal plane of the mandible area.</li>
	</ol>
	</li>
	<li>Fixing the sample on the microtome
	<ol>
		<li>Regulate the specimen clamp in the microtome to protrude the condyle and the crown side 5&#176;-20&#176; more, to attain an integrated image of crown pulp concomitant with root pulp (Figure 2).</li>
	</ol>
	</li>
	<li>Preparing the sections 
	NOTE: The condyle is always the first anatomical structure to be cut. When it fades away, the molar area can then be cut into several slices.
	<ol>
		<li>Shift the microtome range to 5 &#181;m, collect every eight slices, and scout for dentin in the last slice. If the crown dentin first appears without any root apical dentin, adjust the specimen clamp to make the root area protrude, and vice versa.</li>
	</ol>
	</li>
	<li>Collecting the sections 
	NOTE: The cutting angle is appropriate until the dentin is equally attained in both the crown and apical area.
	<ol>
		<li>Cut along this direction and watch the slices under the microscope until the dental pulp appears in both the crown and the roots. Collect the sections when an obscure contour of molars appears on the surface of the paraffin sample17.</li>
	</ol>
	</li>
</ol>

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E., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intramembranous+bone+healing+process+subsequent+to+tooth+extraction+in+mice:+micro-computed+tomography,+histomorphometric+and+molecular+characterization.">Intramembranous bone healing process subsequent to tooth extraction in mice: micro-computed tomography, histomorphometric and molecular characterization.</a> PLoS One. 10 (5), 0128021 (2015).</li><li>Min, K. -. K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+resveratrol+on+bone-healing+capacity+in+the+mouse+tooth+extraction+socket.">Effects of resveratrol on bone-healing capacity in the mouse tooth extraction socket.</a> Journal of Periodontal Research. 55 (2), 247-257 (2020).</li><li>Yu, F., Li, F., Zheng, L., Ye, L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Epigenetic+controls+of+Sonic+hedgehog+guarantee+fidelity+of+epithelial+adult+stem+cells+trajectory+in+regeneration.">Epigenetic controls of Sonic hedgehog guarantee fidelity of epithelial adult stem cells trajectory in regeneration.</a> Science Advances. 8 (29), (2022).</li><li>Kuroshima, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transplantation+of+noncultured+stromal+vascular+fraction+cells+of+adipose+tissue+ameliorates+osteonecrosis+of+the+jaw-like+lesions+in+mice.">Transplantation of noncultured stromal vascular fraction cells of adipose tissue ameliorates osteonecrosis of the jaw-like lesions in mice.</a> Journal of bone and Mineral Research. 33 (1), 154-166 (2018).</li><li>Ahel, V., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Forces+that+fracture+teeth+during+extraction+with+mandibular+premolar+and+maxillary+incisor+forceps.">Forces that fracture teeth during extraction with mandibular premolar and maxillary incisor forceps.</a> The British Journal of Oral &amp; Maxillofacial Surgery. 53 (10), 982-987 (2015).</li></ol>

All the original data and images are included in this paper. The authors declare no conflict of interest.

The murine socket healing model is an important method for unraveling the underlying mechanisms in bone healing and regeneration, ultimately solving clinical challenges. Existing studies have demonstrated the possibility of the incisor extraction model and the maxillary molar extraction model, whereas studies have not used the mandibular first molar model13,17,18. However, incisors are crucial for rodent living, and their impair...

Socket healing after tooth extraction is a common clinical scenario, which can result in unbecoming complications such as socket hemorrhage, dry socket, or even jaw osteomyelitis under undesirable healing1,2,3. These comorbidities may impair the quality of life of patients and, even worse, vitally challenge prosthetic rehabilitation due to massive bone loss4. Though the socket healing stages have been elucidated, they are inadequate to direct clinical care post-tooth extraction surgery when encountering various prognosis challenges4.
Multiple studies based on animal models have been conducted to gain a better understanding of the underlying mechanisms in the socket healing process and avert the above situations. Sp7 is a master regulator in osteoblast differentiation, playing a vital role in skeleton development, bone hemostasis, and bone regeneration5,6. Rational socket healing models could display the redundancy of Sp7 post-traumatic in bone regeneration. In addition, distinct from long bone fracture healing, only a single osteogenic process, intramembranous ossification, involves the healing process of the extraction socket7. This makes the animal tooth extraction model optimal for studying implant-based therapies, as implant osseointegration obeys the same osteogenic rule8.
For decades, the tooth extraction model has been performed in rats, rabbits, and dogs, since these species have big teeth that are convenient to operate on9,10,11. However, given the flourishing demands for genetic modification and as a more adaptive genetic background to humans, mice are increasingly being used to establish a tooth extraction model. Thenceforward, researchers could unravel a specific cell population&#39;s role in the socket healing process using genome-modified mice instead of observing phenotypes only12. Among murine tooth extraction socket models, preceding studies have demonstrated the establishment and healing process of murine maxillary tooth and incisor extraction sockets13,14,15,16. However, the healing pattern of the prognosis, and the detective and observative time points, may differ across protocols. This appeals to a universal criterion for scholars to establish a murine socket healing model.
This study aimed to constitute a practicable murine socket healing model for the above issues. Mandible molars in mice have distinctive morphological traits compared to maxillary molars and incisors, bringing unique advantages and disadvantages. As the models focusing on the murine mandible are currently vacuum-based, this protocol tried to provide an accomplished method to extract the mandibular first molar in mice. We hope this protocol will enlighten basic researchers with new ideas to uncover underlying mechanisms of socket healing and indicate clinical care.

<table><tbody><tr><td>23/25/26 G needle</td><td>Chengdu Xinjin Shifeng Medical Apparatus &amp;amp; Instruments Co. LTD.</td><td>SB1-074(IV)</td><td></td></tr><tr><td>C57/B6J&amp;nbsp;</td><td>Gempharmatech Experimental Animals Company, Chengdu, China</td><td>C57/B6J</td><td></td></tr><tr><td>DAPI Staining Solution&amp;nbsp;</td><td>Beyotime&amp;nbsp;</td><td>Cat#C1005</td><td></td></tr><tr><td>Embedding Cassettes</td><td>CITOTEST Scientific</td><td>80106-1100-16</td><td></td></tr><tr><td>Hematoxylin and Eosin Stain Kit</td><td>Biosharp</td><td>BL700B</td><td></td></tr><tr><td>Isoflurane</td><td>RWD Life Science Co.,Ltd</td><td>R510-22-10</td><td></td></tr><tr><td>Masson&amp;rsquo;s Trichrome Stain Kit</td><td>Solarbio</td><td>G1340</td><td></td></tr><tr><td>Microtome&amp;nbsp;</td><td>Leica</td><td>RM2235</td><td></td></tr><tr><td>Pentobarbital Sodium</td><td>Huaxia Chemical Reagent Co., Ltd</td><td>2018042001</td><td></td></tr><tr><td>Rabbit polyclonal&amp;nbsp;</td><td>anti-Sp7&amp;nbsp;</td><td>Abcam Cat# ab22552</td><td></td></tr><tr><td>Tweezers</td><td>Chengdu Xinjin Shifeng Medical Apparatus &amp;amp; Instruments Co. LTD.</td><td>SB2-115</td><td></td></tr></tbody></table>

the establishment of a murine mandibular molar extraction socket healing model

This study introduces the development of a molar extraction model in the murine mandible to provide a practicable model for studying alveolar bone regeneration and intramembranous ossification. C57/J6 mice were used to extract the mandibular first molar to establish this model. They were euthanized, and the bilateral mandibles harvested, at 1 week and 4 weeks post-surgery, respectively. Subsequent serial stereoscopic harvest, histological assessment, and immunofluorescence staining were performed to demonstrate successful surgery. Immediately after surgery, the stereoscopic images displayed an empty extraction socket. The hematoxylin and eosin (H&#38;E) at 1 week and Masson staining at 4 weeks post-surgery showed that the area of the original root was partially and fully filled with bone trabeculae, respectively. The immunofluorescence staining showed that, compared with the homeostasis side, the Sp7 expression increased at 1 week post-surgery, suggesting vigorous osteogenesis in the alveolar fossa. All these results demonstrated a practicable murine tooth extraction socket healing model. Upcoming studies revealing the mechanisms of jawbone defect healing or socket healing could adopt this method.

To elucidate the practical use of this method, the right mandibular first molar of two healthy C57BL/6 mice (3 months old, both female) were extracted and followed for 1 week and 4 weeks, respectively. The undamaged left mandibles were used as healthy controls. Figure 1A shows the specific features of the surgical appliance, including 26-23 G needles and a toothed ophthalmic tweezer. The 26 G needle is pinpoint-removed and bent. The 25 G needle is bent at approximately 25&#176; at the pinpoi...

Watch this Scientific Journal Video about The Establishment of a Murine Mandibular Molar Extraction Socket Healing Model at JoVE.com

The Establishment of a Murine Mandibular Molar Extraction Socket Healing Model

This protocol demonstrates step-by-step details of how to extract the mandibular first molar in the mouse. It provides an alternative method for researchers focusing on jawbone healing and regeneration.

This study introduces the development of a molar extraction model in the murine mandible to provide a practicable model for studying alveolar bone ...

the-establishment-murine-mandibular-molar-extraction-socket-healing

Universidad de Cartagena UDEC

Research

JoVE Journal

Biology

3.7K Views.  Sichuan University. This protocol demonstrates step-by-step details of how to extract the mandibular first molar in the mouse. It provides an alternative method for researchers focusing on jawbone healing and regeneration.

Video: The Establishment of a Murine Mandibular Molar Extraction Socket Healing Model

Murine Renal Transplantation Procedure

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell et al over 30 years ago (1) and refined by Zhang et al years later (2), few people in the world have mastered this procedure. In our laboratory we have successfully performed 1200 orthotopic kidney transplantations with &gt; 90% survival rate. The key points for success include stringent control of reperfusion injury, bleeding and thrombosis, both during the procedure and post-transplantation, and use of 10-0 instead of 11-0 suture for anastomoses.
Post-operative care and treatment of the recipient is extremely important to transplant success and evaluation. All renal graft recipients receive antibiotics in the form of an injection of penicillin immediately post-transplant and sulfatrim in the drinking water continually. Overall animal health is evaluated daily and whole blood creatinine analyses are performed routinely with a portable I-STAT machine to assess graft function.

Renal transplantation in mice is a technically challenging procedure that requires careful post-operative care and treatment for success.

Renal orthotopic transplantation in mice is a technically challenging procedure. Although the first kidney transplants in mice were performed by Russell ...

LAD-Ligation: A Murine Model of Myocardial Infarction

Research models of infarction and myocardial ischemia are essential to investigate the acute and chronic pathobiological and pathophysiological processes in myocardial ischemia and to develop and optimize future treatment.
Two different methods of creating myocardial ischemia are performed in laboratory rodents. The first method is to create cryo infarction, a fast but inaccurate technique, where a cryo-pen is applied on the surface of the heart (1-3). Using this method the scientist can not guarantee that the cryo-scar leads to ischemia, also a vast myocardial injury is created that shows pathophysiological side effects that are not related to myocardial infarction. The second method is the permanent ligation of the left anterior descending artery (LAD). Here the LAD is ligated with one single stitch, forming an ischemia that can be seen almost immediately. By closing the LAD, no further blood flow is permitted in that area, while the surrounding myocardial tissue is nearly not affected. This surgical procedure imitates the pathobiological and pathophysiological aspects occurring in infarction-related myocardial ischemia.
The method introduced in this video demonstrates the surgical procedure of a mouse infarction model by ligating the LAD. This model is convenient for pathobiological and pathophysiological as well as immunobiological studies on cardiac infarction. The shown technique provides high accuracy and correlates well with histological sections.

This video demonstrates how to use a fast and reliable model to study pathobiological and pathophysiological processes of myocardial ischemia.

Research models of infarction and myocardial ischemia are essential to investigate the acute and chronic pathobiological and pathophysiological processes ...

Production of Transgenic Xenopus laevis by Restriction Enzyme Mediated Integration and Nuclear Transplantation

Stable integration of cloned gene products into the Xenopus genome is necessary to control the time and place of expression, to express genes at later stages of embryonic development, and to define how enhancers and promoters regulate gene expression within the embryo. The protocol demonstrated here can be used to efficiently produce transgenic Xenopus laevis embryos. This transgenesis approach involves three parts: 1. Sperm nuclei are isolated from adult X. laevis testis by treatment with lysolecithin, which permeabilizes the sperm plasma membrane. 2. Egg extract is prepared by low speed centrifugation, addition of calcium to cause the extract to progress to interphase of the cell cycle, and a high-speed centrifugation to isolate interphase cytosol. 3. Nuclear transplantation: the nuclei and extract are combined with the linearized plasmid DNA to be introduced as the transgene and a small amount of restriction enzyme. During a short reaction, egg extract partially decondenses the sperm chromatin and the restriction enzyme generates chromosomal breaks that promote recombination of the transgene into the genome. The treated sperm nuclei are then transplanted into unfertilized eggs. Integration of the transgene usually occurs prior to the first embryonic cleavage such that the resulting embryos are not chimeric. These embryos can be analyzed without any need to breed to the next generation, allowing for efficient and rapid generation of transgenic embryos for analyses of promoter and gene function. Adult X. laevis resulting from this procedure also propagate the transgene through the germline and can be used to generate lines of transgenic animals for multiple purposes.

Production of Transgenic Xenopus laevis by Restriction Enzyme Mediated Integration and Nuclear Transplantation

This video protocol demonstrates a method for generating transgenic Xenopus laevis by introduction of transgenes into sperm nuclei followed by nuclear transplantation into unfertilized eggs.

Stable integration of cloned gene products into the Xenopus genome is necessary to control the time and place of expression, to express genes at later ...

Calvarial Model of Bone Augmentation in Rabbit for Assessment of Bone Growth and Neovascularization in Bone Substitution Materials

The basic principle of the rabbit calvarial model is to grow new bone tissue vertically on top of the cortical part of the skull. This model allows assessment of bone substitution materials for oral and craniofacial bone regeneration in terms of bone growth and neovascularization support. Once animals are anesthetized and ventilated (endotracheal intubation), four cylinders made of polyether ether ketone (PEEK) are screwed onto the skull, on both sides of the median and coronal sutures. Five intramedullary holes are drilled within the bone area delimited by each cylinder, allowing influx of bone marrow cells. The material samples are placed into the cylinders which are then closed. Finally, the surgical site is sutured, and animals are awaken. Bone growth may be assessed on live animals by using microtomography. Once animals are euthanized, bone growth and neovascularization may be evaluated by using microtomography, immune-histology and immunofluorescence. As the evaluation of a material requires maximum standardization and calibration, the calvarial model appears ideal. Access is very easy, calibration and standardization are facilitated by the use of defined cylinders and four samples may be assessed simultaneously. Furthermore, live tomography may be used and ultimately a large decrease in animals to be euthanized may be anticipated.

Here we present a surgical protocol in rabbits with the aim to assess bone substitution materials in terms of bone regeneration capacities. By using PEEK cylinders fixed onto rabbit skulls, osteoconduction, osteoinduction, osteogenesis and vasculogenesis induced by the materials may be evaluated either on live or euthanized animals.

The basic principle of the rabbit calvarial model is to grow new bone tissue vertically on top of the cortical part of the skull. This model allows ...

Bioengineering

Experimental Model of Ligature-Induced Peri-Implantitis in Mice

Dental implants have a high success and survival rate. However, complications such as peri-implantitis (PI) are highly challenging to treat. PI is characterized by inflammation in the tissues around dental implants with progressive loss of supporting bone. To optimize dental implants' longevity in terms of health and functionality, it is crucial to understand the peri-implantitis pathophysiology. In this regard, using mouse models in research has proven clear benefits in recreating clinical circumstances. This study aimed to describe an experimental model of ligature-induced peri-implantitis in mice and determine whether there is effectiveness in inducing this disease, given the observed bone and tissue changes. The experimental peri-implantitis induction comprehends the following steps: teeth extraction, implant placement, and ligature-inducted PI. A sample of eighteen 3-week-old C57BL/6J male mice was divided into two groups, ligature (N=9) and control non-ligature (N=9). The evaluation of clinical, radiographical, and histological factors was performed. The ligature group showed significantly higher bone loss, increased soft tissue edema, and apical epithelial migration than the non-ligature group. It was concluded that this pre-clinical model can successfully induce peri-implantitis in mice.

This is a report on an experimental model of ligature-induced peri-implantitis in mice. We describe all surgical steps, from pre- and post-operative management of the animals, extractions, implant placement, and ligature-induced peri-implantitis.

Dental implants have a high success and survival rate. However, complications such as peri-implantitis (PI) are highly challenging to treat. PI is ...

Immunology and Infection

Isolation of Murine Coronary Vascular Smooth Muscle Cells

While the isolation and culture of vascular smooth muscle cells (VSMCs) from large vessels is well established, we sought to isolate and culture VSMCs from the coronary circulation. Hearts with intact aortic arches were removed and perfused via retrograde Langendorff with digestion solution containing 300 Units/ml of collagenase type II, 0.1 mg/ml soybean trypsin inhibitor and 1 M CaCl2. The perfusates were collected at 15 min intervals for 90 min, pelleted by centrifugation, resuspended in plating media, and plated on tissue culture dishes. VSMCs were characterized by presence of SM22&#945;, &#945;-SMA, and vimentin. One of the main advantages of using this technique is the ability to isolate VSMCs from the coronary circulation of mice. Although the small number of cells obtained can limit some of the applications for which the cells can be utilized, isolated coronary VSMCs can be used in a variety of well-established cell culture techniques and assays. Studies investigating VSMCs from genetically modified mice can provide further information about structure-function and signaling processes associated with vascular pathologies.

The purpose of this protocol is to demonstrate the isolation and culture techniques of murine primary vascular smooth muscle cells (VSMCs) from the coronary circulation. Once VSMCs have been isolated, they can be used for many standard culture techniques.

While the isolation and culture of vascular smooth muscle cells (VSMCs) from large vessels is well established, we sought to isolate and culture VSMCs ...

Development of an Effective Mice Model for Studying Oronasal Fistulas

Establishment of an Oronasal Fistula Mice Model

This study presents a method utilizing heated ophthalmologic cautery to develop a viable model for investigating oronasal fistulas. C57BL/6 mice were used to establish the oronasal fistula (ONF) model. To create the ONF, the mice were anesthetized, immobilized, and their hard palates were exposed. During the surgical procedure, a 2.0 x 1.5 mm full-thickness mucosal injury was induced in the midline of the hard palate using ophthalmologic cautery. It was crucial to control the size of the ONF and minimize bleeding in order to ensure the success of the experiment. Verification of the ONF model's effectiveness was conducted on the 7th-day post-operation, encompassing both anatomical and functional assessments. The presence of the nasal septum within the oral cavity and the outflow of sterile water from the nostrils upon injection into the oral cavity confirmed the successful establishment of the ONF model. The model demonstrated a practical and successful oronasal fistula, characterized by a low mortality rate, significant weight changes, and minimal variation in ONF size. Future studies may consider adopting this methodology to elucidate the mechanisms of palate wound healing and explore novel treatments for oronasal fistulas.

Author Spotlight: Investigating Wound Healing in Mice Models of Oronasal Fistulas 

This article outlines a step-by-step procedure for establishing a mice model with an oronasal fistula. The oronasal fistula was created by employing heated ophthalmologic cautery to damage the midline portion of the hard palate, resulting in the formation of an opening between the oral and nasal cavities.

This study presents a method utilizing heated ophthalmologic cautery to develop a viable model for investigating oronasal fistulas. C57BL/6 mice were used ...

Medicine

A Murine Maxillary Orthodontic Model Guide to Bone Remodeling Analysis

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.

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.

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

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.

Pulpitis, a common cause of natural tooth loss, leads to necrosis and loss of bioactivity in the inflamed dental pulp. Unraveling the mechanisms ...

Standardized Mandibular Fracture Protocol for Bone Repair in Mice

Less-Invasive Technique for Non-stabilized Mandibular Fracture in Mouse Models

Non-stabilized fractures can be made at mandibular sites in mice, thus making it possible to analyze bone repair using an endochondral ossification mode. To most accurately reflect this process in vivo, it is necessary to have a standardized protocol to avoid excessive bone loss and soft tissue damage, particularly at the mandibular site, an anatomical site characterized by minimal access. To our knowledge, we describe for the first time a less-invasive protocol of non-stabilized mandibular fracture in mice. Adult mice are anesthetized with isoflurane and receive a preoperative dose of buprenorphine by subcutaneous injection. A submandibular approach is performed, with a skin incision made along the inferior border of the mandible. The masseter muscle is elevated in a subperiosteal plane along the mandibular ramus with a periosteal elevator. A vertical and complete fracture of the ramus is performed from the basilar border to the coronoid notch (between condylar and coronoid processes), using a piezoelectric bone surgery device under saline serum irrigation at the basilar edge and scissors to complete the fracture of the mandibular ramus. The skin approach is closed by silk sutures. This detailed procedure for non-stabilized mandibular fractures offers an efficient procedure allowing minimal bone loss and soft tissue damage. This technique ensures successful and consistent results to accurately reflect the process of endochondral bone repair in mouse models.

This protocol presents a comprehensive and efficient method for analyzing mandibular bone repair. We describe a reproducible technique for non-stabilized mandibular fracture in a mouse model, allowing analysis of the process of endochondral bone repair with minimal tissue damage and bone loss.

Non-stabilized fractures can be made at mandibular sites in mice, thus making it possible to analyze bone repair using an endochondral ossification mode. ...

The Establishment of a Murine Mandibular Molar Extraction Socket Healing Model

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Experimental Model of Ligature-Induced Peri-Implantitis in Mice

Isolation of Murine Coronary Vascular Smooth Muscle Cells

Establishment of an Oronasal Fistula Mice Model

The Establishment of a Murine Maxillary Orthodontic Model

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

Less-Invasive Technique for Non-stabilized Mandibular Fracture in Mouse Models