preparation of the mouth gag for a mouse pulpitis model

새로운 구강 재갈을 사용하여 Pulp Inflammation Mouse Model 만들기 

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 therapies2,3. 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 the neural feedback offered by healthy pulp4. This situation has led researchers to explore alternative solutions for managing inflamed pulp beyond mere removal.
In 2007, Murray et al. initiated the application of tissue engineering in regenerative endodontics, thereby sparking increased interest in pulp preservation and regeneration5. However, inflamed pulp tissue poses a challenge as cells release inflammatory factors such as IL-6, which recruit inflammatory cells and results in cell necrosis, loss of pulp vitality, and complications in functional recovery6,7. Understanding inflammation and the associated cell death is, therefore, crucial for advancements in the preservation of vital pulp. There are a number of experiments that have been conducted to explore the molecular biology of the inflame pulp in vivo or in vitro8,9. Though in vitro&#160;experiments like 2D or 3D cell cultures have been developed for years and are becoming mature and widely used to test reactions of pulp cells to inflammatory factors, these experiments cannot reflect the interaction between pulp tissue and the systemic immune system10. If the phenomenon being studied is derived from cells of other tissue origin like immune, vascular and nervous system, then pure pulp cell culture will lead to a dead end. Therefore, in vivo experiments are very necessary and referential.
Mice have increasingly become a common choice in inflammation research in vivo&#160;due to their cost-effectiveness, high fertility, and vitality. However, a comprehensive protocol for mice pulpitis model is currently absent, which can serve as a reference. The small size of mice and their sensitivity to stimulation pose significant challenges during experimental procedures. Observing the minuscule teeth concealed deep within the mouse mouth often necessitates the use of a cantilever microscope, notwithstanding the more common presence of desktop microscopes in laboratories. The absence of a mouth opener requires assistance from others. To address this, the group has devised a mouth-gag using readily available materials which aims to provide a standardized and reproducible protocol for constructing the mice pulpitis model. This article details the procedure, covering preoperative preparation, immobilization, pulp exposure surgery, and sample collection on C57 mice. This protocol recommends the use of the mouth-gag, providing information on its structure, production, and application to facilitate other researchers in replicating the procedure.

저자 스포트라이트: 개선된 마우스 모델을 통한 치과용 펄프 연구 강화

Pulpitis 연구를 위한 새로운 구강 개그를 이용한 Murine Pulp Exposure Model 구축

Individual experiments like 2D or 3D cell cultures have been widely used to test the reactions of dental pulp cells. These experiments cannot reflect the interaction between pulp tissue and the systemic immune system. Therefore, in vivo experiments are becoming increasingly popular in endodontic research.C57 mice have increasingly become a common choice in inflammation research on dental pulp due to their cost effectiveness, high fertility and vitality. However, the small size of mice and lack of coordination posed significant challenges during experimental procedures. The fixation process is an important factor that higher the technical threshold.We devised a novel mouth gag using readily available materials and simple assemblies to assist researchers in pulp exposure surgery. According to our research, this mouth gag can significantly shorten the time required for beginners to immobilize the mouth and expose the maxillary first molar of mice.

Mouse Pulpitis 모델에 대한 구강 재갈 준비

시작하려면 왼손으로 8미크론 두께의 교정 아치 와이어를 곧게 펴고 오른손은 와이어 아크에 약간 구부린 상태로 유지합니다. Young loop bending 플라이어를 사용하여 활 와이어의 중간 지점에서 사다리꼴의 상단 가장자리를 약 8mm 정도 구부립니다. 점 A가 플라이어 부리의 가장자리에 있는지 확인하십시오.이제 왼손으로 펜치를 잡습니다. Clamp 오른쪽 엄지와 집게 손가락으로 활 와이어의 다른 쪽 끝을 만든 다음 A 지점에서 활 와이어를 구부려 120도 각도를 만듭니다. 아치 와이어를 수평 테이블에 놓고 단일 평면에 있는지 확인합니다.와이어의 양쪽에 약 9mm를 남겨두고 펜치를 사용하여 자유 끝을 75도 각도로 구부립니다. 이제 점 B에서 약 5mm 떨어진 점 C를 찾습니다. 점 C에서 와이어를 구부려 105도 각도를 만듭니다. 마찬가지로, 점 D와 E에서 와이어를 구부려 IK 및 KJ 부품을 동시에 클램프합니다.점 L, J 및 K에서 구부리는 A, B, C를 복제한 다음 점 J에서 자유 끝을 구부려 IKJ 평면에 수직으로 만듭니다. 하악 부분을 위해 여분의 혀 압압기를 구부립니다. 점 J에서 5mm 떨어진 지점 I을 클램프하고 자유 끝을 구부려 IKJ 평면과 CD 부품 모두에 평행하게 만듭니다.다음으로, 점 I 및 점 H에서 5mm 간격을 두고 아치를 IH 및 JIH 평면에 수직으로 구부립니다. H에서 5mm 떨어진 지점 G를 찾아 JIHG 레인을 고정합니다. 그런 다음 점 G에서 아치를 구부리고 이제 KJ 평면에 대칭인 자유 끝을 구부립니다.점 F 뒤의 자유단은 점 E의 자유단과 대칭이어야 하며, 마지막으로 자유단에 고무 캡을 놓습니다.

preparation-of-the-mouth-gag-for-a-mouse-pulpitis-model

Research

JoVE Journal

Medicine

Preparation of the Mouth Gag for a Mouse Pulpitis Model

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.

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 ...

연구

의학

To begin, straighten an 8 micron thick orthodontic arch wire with your left hand while keeping the right hand slightly bent against the wire arc. Use a Young loop bending plier to bend the top edge of the trapezoid about eight millimeters at the midpoint of the bow wire. Ensure that point A is on the edge of the plier beak.Now hold the plier with the left hand. Clamp the free end of the bow wire with the right thumb and forefinger, then bend the bow wire from point A to make a 120-degree angle. Place the arch wire on a horizontal table to check if it is on a single plane.Leave about nine millimeters on each side of the wire and use pliers to bend the free end to a 75-degree angle. Now locate point C which is about five millimeters from point B.Bend the wire at point C to make a 105-degree angle. Similarly, bend the wire at points D and E.Clamp the IK and KJ parts simultaneously.Duplicate the A, B, C bending at points L, J, and K, then bend the free end at point J to make it vertical to the IKJ plane. Bend an extra tongue depressor for the mandibular part. Clamp point I five millimeters from point J and bend the free end to make it parallel to both the IKJ plane and the CD part.Next, leave a five millimeter gap from point I and at point H, bend the arch vertically to the IH and JIH planes. Locate point G five millimeters from H and clamp the JIHG lane. Then bend the arch at point G.Now bend the free end symmetrical to the KJ plane.The free end after point F should be symmetrical to the free end at point E.Finally, place rubber caps on the free ends.

Using A Mouth Gag for Pulpitis Assessment in Mice

To begin, fix an anesthetized mouse in a supine position on a fixation plate. Secure the limbs with skin tape. Compress the free ends of a prepared mouth gag with the thumb and the index finger.Fix the mouse's front incisors in the trapezoidal groove between the two arms of the mouth gag. Ensure the arm with the tongue depressor is placed on the mandibula. Check the maxillary first molar to ensure it is free of dental caries, trauma, and odontogenesis.Assess the surrounding gingiva to ensure absence of redness, swelling, or fistula. Then, check the health of the opposite teeth. Next, with a dental burr, drill the occlusal side of the maxillary first molar at 20, 000 RPM.Remove the enamel. With a syringe, drop normal saline every three minutes over the tooth to prevent overheating. Put a number 8 C+file on the lowest position of the drilled pit and pierce the last dentine to expose the pulp chamber.Clean the fragments around the tooth, then remove the mouth gag. The time taken to fix the mouth with the mouth gag and to find the maxillary first molar was significantly shortened relative to the traditional method. The pulp exposure was confirmed through micro CT and reconstruction modeling.At 24 hours post-surgery, most of the pulp tissue retained its morphology.