小鼠结扎诱导种植体周围炎的实验模型
Summary
这是一份关于小鼠结扎诱导的种植体周围炎实验模型的报告。我们描述了所有手术步骤，从动物的术前和术后管理、拔牙、植入物放置和结扎诱导的种植体周围炎。
Abstract
种植牙的成功率和存活率都很高。然而，种植体周围炎 （PI） 等并发症的治疗非常具有挑战性。PI 的特征是种植牙周围组织发炎，支撑骨逐渐丧失。为了优化种植牙在健康和功能方面的寿命，了解种植体周围炎的病理生理学至关重要。在这方面，在研究中使用小鼠模型已被证明在重建临床环境方面有明显的好处。本研究旨在描述小鼠结扎诱导的种植体周围炎的实验模型，并确定在观察到的骨骼和组织变化的情况下是否有效诱导这种疾病。实验性种植体周围炎诱导包括以下步骤：拔牙、种植体放置和结扎诱导 PI。将18只3周龄C57BL/6J雄性小鼠的样本分为结扎组（N=9）和对照非结扎组（N=9）两组。对临床、影像学和组织学因素进行评估。与非结扎组相比，结扎组的骨质流失、软组织水肿增加和顶端上皮迁移明显更高。得出的结论是，这种临床前模型可以成功诱导小鼠种植体周围炎。
Introduction
种植牙作为替代缺失牙齿的理想选择越来越普遍1.到 2026 年，美国成年人口中种植牙的普及率预计将增加到 23%2。根据 Grand View Research 的市场分析报告（2022 年），预计 2022 年全球种植牙市场规模将达到约 46 亿美元。此外，预计到 2030 年，它将呈现出 10% 左右的稳定年增长率3.不幸的是，使用种植牙会导致并发症，例如种植体周围炎。种植体周围炎被定义为一种生物膜诱发的疾病，其特征是种植体周围粘膜发炎，随后支撑骨逐渐丧失4。
一项系统评价发现，患者水平种植体周围炎的平均患病率为 19.53%（95% 置信区间 [CI]，12.87-26.19%），种植体水平为 12.53%（95% CI 11.67-13.39%）5。由于种植体失败的增加，以及因此产生的大量治疗费用，种植体周围炎代表了日益增长的公共卫生6.
了解种植体周围炎的发病机制对于开发一种系统的方法以防止其发生和发展以及最大限度地延长种植体的美学和功能至关重要7,8。从这个意义上说，在牙科研究中使用小鼠模型已被证明是有利的，因为小鼠与人类共享超过95%的基因9,10，可用的在线遗传数据库的数量，以及重现临床场景的能力11。所有描述的优点都允许剖析不同疾病中的遗传机制12，可获得的调节和管理，以及作为人体panel广泛使用的抗体，超出了用于炎症组织评估和疾病定位的遗传修饰可用性（例如，敲除和过表达）13。虽然有利，但很少有关于小鼠种植体周围炎的出版物。这是由于方法学上的挑战，其中包括获得或安装微型植入物的困难。
为了在小鼠中发生种植体周围炎，已经描述了许多方案，例如结扎诱导的种植体周围炎、细菌诱导的种植体周围炎14、脂多糖 （LPS） 诱导的种植体周围炎15 或联合 LPS + 结扎诱导的种植体周围炎16。在这里，我们将重点介绍结扎模型，因为它是诱发牙周炎 17,18,19 和最近的种植体周围炎20,21 的最广泛接受的方法。在植入物周围放置在粘膜下位置的结扎会刺激斑块积聚，从而刺激组织发炎。因此，这种方法的发展是基于一种可行的成本效益技术，用于种植体周围疾病的临床前研究。本研究旨在描述小鼠结扎诱导的种植体周围炎的实验模型，并确定根据观察到的骨骼和组织变化是否有效诱导这种疾病。
本文的总体目标是报告应用于通过结扎诱导小鼠种植体周围炎的方案，并通过组织评估和植入物周围的骨质流失来观察其有效性。
Protocol
涉及动物受试者的程序已获得加州大学洛杉矶分校校长动物研究委员会（ARC 协议编号 2002-125）和动物研究：报告体内实验 （ARRIVE）22 的批准。对于这种方法，使用18只3周龄的C57BL / 6J雄性小鼠并进行了拔牙，种植体放置和种植体周围炎诱导。所有牙科手术均在10×显微放大倍率下进行，并由经过培训和校准的操作员进行（图1A）。 1….
Representative Results
对于这种方法，使用18只3周龄的C57BL / 6J雄性小鼠并进行了拔牙，种植体放置和种植体周围炎诱导。每组有9只动物，具有统计学意义，考虑到线性骨质流失达到80%的功效，15%的标准差（σ）和95%的置信区间（α = 0.05）。在实验期间，小鼠被 随意 喂食软饮食。9只小鼠接受结扎（结扎诱导的种植体周围炎实验组），9只小鼠未接受结扎（对照组）。 种植体骨整合的成?…
Discussion
该协议提供了利用小鼠结扎模型诱导种植体周围炎的外科手术的描述性报告。与小鼠一起工作具有优势，例如具有成本效益，考虑到许多背景23等方面，广泛的遗传阵列的可用性24,25。多年来，一些研究成功地将小鼠用于医学和牙科领域，包括种植体周围炎26,27,28。<…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作得到了NIH/NIDCR DE031431的支持。我们要感谢加州大学洛杉矶分校转化病理学核心实验室在准备脱钙组织学切片方面的帮助。
Materials
| #5 dental explorer | Hu-Friedy, Chicago, IL | 392-0911 | Dental luxation |
| 15c blade and surgical scalpel | Henry Schein Inc., Melville, NY | 1126186 | Tissue incision |
| 6-0 silk ligatures | Fisher Scientific, Hampton, NH | NC9201232 | Ligature |
| Amoxicillin 50μg/mL | Zoetis, San Diego, CA | TS/DRUGS/57/2003 | Oral suspension |
| Bacon Soft Diet | Bio Serve®, Frenchtown, NJ | 14-726-701 | – |
| C57BL/6J male mice | The Jackson Laboratories, Bar Harbor, ME, USA | 000664 | Age: 3-week-old |
| CTAn software | V.1.16 Bruker, Billerica, MA | – | Volumetric analysis |
| Dolphin software | Navantis, Toronto, CA | – | Linear bone analysis |
| Implant carrier & Tip | D. P. Machining Inc., La Verne, CA | Unique product | Implant holder |
| Implant support | D. P. Machining Inc., La Verne, CA | Unique product | Implant capture |
| Isoflurane | Vet One, Boise, ID | NDC13985-528-60 | Inhalational anesthetic |
| Micro-CT scan 1172 | SkyScan, Kontich, Belgium | – | μCT scans |
| Nrecon Software | Bruker Corporation, Billerica, MA | – | Images reconstruction |
| Ø 0.3mm – L 2.5mm Micro Drills | Sphinx, Hoffman Estates, IL | ART. 50699 | Osteotomy |
| Ø 0.5mm – L 1.0mm Titanium implants | D. P. Machining Inc., La Verne, CA | Unique product | – |
| Ophthalmic lubricant | Apexa, Ontario, CA | NDC13985-600-03 | Artificial tears |
| Pin Vise | General Tools, Secaucus, NJ | 90 | Osteotomy |
| Rimadyl 50mg/ml | Zoetis, San Diego, CA | 4019449 | Anti-inflammatory |
| Sterile cotton tipped | Dynarex, Glendale, AZ | 4304-1 | Hemostasis |
| Tip forceps | Fine Science Tools, Foster City, CA | 11071-10 | Dental Extraction |
| Tying forceps | Fine Science Tools, Foster City, CA | 18025-10 | Ligature placement |
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Cite This Article
de Araújo Silva, D. N., Casarin, M., Monajemzadeh, S., Menezes da Silveira, T., Lubben, J., Bezerra, B., Pirih, F. Q. Experimental Model of Ligature-Induced Peri-Implantitis in Mice. J. Vis. Exp. (207), e66316, doi:10.3791/66316 (2024).