这项工作得到了国防部 (DoD) 美国陆军医学研究和装备司令部 (USAMRMC) 国会定向医学研究项目 (CDMRP) (PR121604) 和国家关节炎和肌肉骨骼研究所赠款的支持。和皮肤病 (结缔组织国立研究院), NIH R01 AR068332 桑卡尔。

以下程序是由印第安纳大学医学院的动物保育和使用委员会 (IACUC) 批准的。所有的生存手术都是在无菌条件下进行的, 如 NIH 指南所述。疼痛和感染的风险管理与适当的止痛药和抗生素, 以确保成功的结果。
1. 麻醉和准备
<ol><li>用腹腔 (ip) 路线对老鼠进行称量, 并麻醉氯胺酮 (100 毫克/千克) 和甲苯噻嗪 (10 毫克/千克) 的混合物。将鼠标放在一个空笼子里并监视它, 直到它完全镇静下来。</li>
	<li>使用脚趾捏反射确保鼠标镇静。将眼膏涂抹在眼睛上, 防止其干燥。</li>
	<li>从右肢上取出皮毛。用碘基擦洗和70% 乙醇擦拭手术部位。从膝盖中心开始擦洗手术部位, 并向外进行环形清扫。重复这3x 与新鲜的磨砂, 结束70% 乙醇。</li>
	<li>术前剂量盐酸丁丙诺啡镇痛 (0.03 毫克/千克) 皮下即刻疼痛管理。</li>
	<li>将鼠标放在无菌手术垫覆盖的加热垫上。</li>
</ol>2. 手术方法
注: 骨折前, 应在手术前对小鼠的特定应变、年龄和性别进行经验性测定。这种手术方法是为 C57BJ6 雄性小鼠在10周的年龄进行优化。
<ol><li>将鼠标放在背部, 弯曲手术腿的膝盖。使用手术刀刀片, 使一个1.5 厘米的切口集中在膝关节。</li>
	<li>用镊子将髌骨侧向移位以暴露股骨远端。在滑车槽的中心插入一个1.5 长25口径不锈钢皮下注射针, 沿着髓质管的长度逆行, 并通过股骨的近端。采取 X 射线, 以确保正确放置 pin。 注: 针应从鼠标的背侧离开以创建导丝的路径。</li>
	<li>通过针的轴, 通过4长36口径的钨导丝, 进入中心的股骨远端和退出的斜角在鼠标的背侧。</li>
	<li>在成功放置导丝后, 小心地取出25口径的针, 轻轻地拉在轮毂上, 同时保持四肢和导丝到位。通过 x 射线确认导丝的位置。</li>
	<li>从撞击盘上方34.6 厘米的高度保持391克重量 (图 1A)。将股骨横放在两个支撑点上, 使股骨的股骨粗隆间和髁上区域在支撑铁砧 (图 1B) 和肢体的侧边上, 面对加载点 (图1C).放下重量, 并小心地从设备上取出鼠标后立即断开。</li>
	<li>用 X 射线确认骨折位置。</li>
	<li>在导丝上插入24口径不锈钢皮下导管, 以稳定股骨骨折。 注: 此应用程序可能需要一些力, 因为进入点是使用一个较小的直径针产生。这种直径的差异有效地防止了24尺杆通过股骨近端的潜在迁移。插入深度可以手动感觉, 因为钝管满足较大的转子皮质骨。</li>
	<li>在去除导丝前, 用 X 射线确定钢棒的位置和股骨骨折的稳定性。</li>
	<li>用钢丝钳切开股骨远端的多余油管。用镊子将暴露的油管埋在髁表面下, 用钳子轻轻向下用力, 小心不要使膝关节脱臼。</li>
	<li>用镊子重新定位髌骨。用5-0 可吸收缝线关闭切口部位。</li>
</ol>3. 术后管理
<ol><li>手术后, 小鼠可通过ip 路线注射多达500µL 的无菌生理盐水, 以帮助他们在术后恢复。</li>
	<li>在加热的恢复床上监视动物, 直到他们从手术中醒来。一旦走动, 把他们送回笼子里去。</li>
	<li>在手术后的几天里, 继续密切监视老鼠, 以确保他们恢复正常, 恢复机动性。管理盐酸丁丙诺啡镇痛 (0.03 毫克/千克) 皮下每6小时皮下手术后3天, 并根据需要此后。避免使用非甾体抗炎药物, 因为它们已经被证明会损害手术后的骨愈合。</li>
</ol>

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这篇手稿的作者没有什么可透露的。作者进一步指出, 对本手稿所报告的研究中使用的所有材料的完全访问没有任何限制。

本手术的目的是在小鼠中产生标准的闭合性股骨骨折。该模型的一个主要优点是, 内固定发生在骨折产生后, 从而避免了髓内棒的角度。也许这个协议最关键的方面是在股骨中段形成一个标准化的横向骨折, 因为骨折几何取决于施加的弯曲力和后肢的位置。在弯矩时, 股骨的定位不当会导致偏斜或粉碎性骨折。重量和下落高度必须是经验主义地预定的, 因为他们取决于老鼠的年龄、性和劳损。采用装...

骨折是肌肉骨骼系统中最常见的损伤之一, 与巨大的社会经济负担有关, 包括预计在美国每年超过250亿美元的治疗费用1, 2。虽然大多数骨折愈合无事故, 愈合与大量停机和生产力损失。大约 5-10% 的骨折导致延迟愈合或不愈合, 由于年龄或其他基本的慢性健康状况, 如骨质疏松症和糖尿病3,4,5。目前尚无 FDA 批准的药理治疗, 可促进有效的骨愈合, 缩短恢复时间。
骨折愈合是一个复杂和高度动态的过程, 涉及多种细胞类型的协调。因此, 全面了解与骨再生相关的细胞和分子事件, 对于确定加速这一过程的治疗目标至关重要。与其他人类疾病一样, 建立一个高度顺从和可再生的动物模型是研究骨愈合的关键。较大的动物, 如绵羊和猪, 有骨重塑的性质和生物力学类似于人类, 但昂贵, 需要大量的愈合时间, 并不容易服从基因操纵6。另一方面, 小动物模型, 如大鼠和小鼠, 提供了许多好处, 包括易于处理, 维护成本低, 繁殖周期短, 愈合时间缩短7。此外, 鼠标基因组是完全有序的, 允许快速操作和生成的遗传变异。因此, 鼠标是一个强大的模型系统, 研究人类疾病, 伤害和修复8。在人类中, 像骨质疏松症和糖尿病这样的并发症增加了延迟愈合的可能性。现有的一些小鼠模型可以用来研究骨质疏松症和糖尿病并发症对骨损伤和愈合的影响。患有骨质疏松症的患者在骨折愈合后的晚期骨形成明显减少9。去卵巢 (OVX) 小鼠表现出快速骨丢失和延迟骨愈合类似于绝经后骨质疏松症10,11。此外, 许多类型的 I 型和 II 型糖尿病的小鼠模型模仿的低骨质量表表和受损骨折愈合在人类看到11。此外, 小鼠骨折模型是研究愈伤组织复杂生物过程的多功能平台, 探索加速骨组织再生的新的治疗策略。
尽管骨结构和新陈代谢有差异, 但在小鼠和人类中, 骨折愈合的整个过程仍然非常相似, 包括软骨和膜内骨化的结合, 然后是骨重塑。软骨骨化包括招募祖细胞到较不机械稳定的区域周围的骨折间隙, 他们分化成软骨细胞, 肥大和矿化软骨产生柔软的愈伤组织。第二波祖细胞浸润愈伤组织, 分化为成熟的成骨细胞, 分泌新的骨基质12,13,14,15。在膜内骨化过程中, 骨膜和骨内表面的祖细胞直接分化为基质, 从而促进骨折间隙9、11、12 的衔接. ,13。在一起, 软骨和膜内血肿骨化相接导致了坚硬愈伤组织的发展, 随着时间的推移进一步改建, 形成一个强大的辅助骨, 能够支持机械负荷13,14 ,15。在健康的人, 愈合过程需要大约3月, 与仅35天在老鼠16。
骨折愈合通常被研究使用开放或闭合的外科模型17。开放性手术方法, 如产生严重的缺陷或完整的截骨, 规范损伤位置和几何, 以减少粉碎性骨折造成的偏差。截骨术作为一个优秀的模型来研究不愈合背后的基础机制, 因为愈合往往延迟比较闭合性骨折。此外, 需要硬外固定, 以稳定 osteotomized 骨, 这意味着再生将主要依赖于膜内骨化。开放式手术方法使用的设备, 如锁定钉, 别针夹子, 锁定板, 以提供轴向和旋转稳定性的骨折肢体;然而, 这些设备是昂贵的, 需要大量的时间在手术18,19,20,21。另一方面, 闭合模型是稳定的一个简单的髓内固定装置, 允许足够的不稳定, 以刺激软骨愈合。因此, 闭合断裂模型不容易模仿非联合的条件。内固定技术, 如髓内针, 指甲, 和加压螺钉, 是有利的, 因为他们便宜, 易于使用, 并尽量减少手术时间21,22,23。在某些情况下, 在骨折前插入髓内针, 但是髓内针的弯曲会导致股骨骨折的角或移位, 从而造成变的愈伤组织大小和愈合。在闭合模型中, 断裂位置和几何更难标准化, 因为它们是使用三点弯曲装置产生的, 其中骨干的重量被丢弃。然而, 用适当的技术, 这种手术方法提供了快速和一致的结果。此外, 闭合性骨折模型作为临床相关工具, 用于研究由高强度撞击或机械应力引起的骨折22。
这项手术协议是根据以前描述的方法, 使用髓内针稳定骨折股骨大鼠和小鼠22,24,25。首先, 通过髁间凹槽插入一个小直径的髓内针, 以建立一个入口点, 并在使用重力依赖性三点的股骨中段上产生横向骨折之前引入导丝。弯曲装置。在成功地产生闭合性股骨骨折后, 将一条直径较大的髓内棒结合在导丝上, 以稳定股骨骨折。这种方法避免了骨折时髓内针角引起的延迟愈合的风险, 因为在骨折后放置杆后, 可以重新定位和优化股骨损伤的稳定性。

<table>
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			<td>Thermo Fisher Scientific</td>
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			<td>Ultra-High-Temperature Tungsten Wire,</td>
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			<td>0.005&#34; Diameter, 1/16 lb. Spool, 380&#39; Long</td>
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			<td>304 stainless steel, 24G thin walled tubing</td>
			<td>Microgroup Inc</td>
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			<td>Fine Science Tools</td>
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			<td>#10 Scalpel Blades</td>
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			<td>10010-00</td>
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			<td>Narrow Pattern Forceps</td>
			<td>Fine Science Tools</td>
			<td>11002-12</td>
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			<td>Fine Science Tools</td>
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			<td>Fine Science Tools</td>
			<td>14058-11</td>
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			<td>ACE Surgical Supply Co., Inc.</td>
			<td>08-051-90</td>
			<td>ACE #150 Wire Cutter, tungsten carbide tips</td>
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			<td>Ethicon Suture</td>
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			<td>N/A</td>
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			<td>Custom Built</td>
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	</tbody>
</table>

the generation of closed femoral fractures in mice: a model to study bone healing

骨骨折给病人带来了巨大的社会经济负担, 除了严重影响他们的生活质量。促进有效骨愈合的治疗策略是不存在和高需求的。为了了解与骨再生相关的复杂生物过程, 需要有效和可再生的骨折愈合动物模型。许多动物模型的骨折愈合已经产生了多年;然而, 小鼠骨折模型最近成为研究骨愈合的有力工具。各种开放和封闭的模型已经开发, 但闭合性股骨骨折模型突出作为一个简单的方法, 以产生快速和重现性的结果, 在生理上相关的方式。该手术方案的目的是在小鼠中产生单边闭合性股骨骨折, 并通过插入髓内钢棒促进股骨骨折后稳定。虽然诸如钉子或螺钉等设备提供了更大的轴向和旋转稳定性, 但使用髓内棒可以为一致愈合结果提供足够的稳定, 而不会产生新的骨组织缺陷或损害附近的软组织。影像学是用来监测愈伤组织的形成, 骨愈合, 并随后重塑骨痂的进展。骨愈合的结果通常与愈合的骨强度和测量的扭转试验。然而, 了解骨折修复相关的早期细胞和分子事件是研究骨组织再生的关键。髓内固定小鼠闭合性股骨骨折模型是研究骨折愈合的一个有吸引力的平台, 并对加速愈合的治疗策略进行评价。

通过影像学检查, 对手术过程的成功实施进行了监测。关键步骤包括插入髓内针, 放置导丝, 在股骨中段诱导横断, 并与髓内杆适当稳定 (图 2Ai - 2Aiv)。在手术后28天内用每周影像影像监测骨折愈伤组织的愈合进展 (图 2B)。10-16 天骨折后, 软骨细胞发生肥厚, 产生矿化软骨, 形成突出的柔软愈伤组织。
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Watch this Scientific Journal Video about The Generation of Closed Femoral Fractures in Mice: A Model to Study Bone Healing at JoVE.com

The Generation of Closed Femoral Fractures in Mice: A Model to Study Bone Healing

小鼠闭合性股骨骨折模型是研究骨折愈合的有力平台, 是加速骨再生的新的治疗策略。这项手术的目的是在小鼠中产生单侧闭合性股骨骨折, 用髓内钢棒稳定股骨。

小鼠闭合性股骨骨折的形成: 骨愈合模型的研究

Bone fractures impose a tremendous socio-economic burden on patients, in addition to significantly affecting their quality of life. Therapeutic strategies ...

the-generation-closed-femoral-fractures-mice-model-to-study-bone

Research

Education

JoVE Core

JoVE Journal

Anatomy and Physiology

Medicine

Unit 1

Bone Tissue and the Skeletal System

SCIENTISTS IN ACTION

15.1K 次观看.  Indiana University School of Medicine. 小鼠闭合性股骨骨折模型是研究骨折愈合的有力平台, 是加速骨再生的新的治疗策略。这项手术的目的是在小鼠中产生单侧闭合性股骨骨折, 用髓内钢棒稳定股骨。

视频: The Generation of Closed Femoral Fractures in Mice: A Model to Study Bone Healing

Biomarkers in an Animal Model for Revealing Neural, Hematologic, and Behavioral Correlates of PTSD

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for objective diagnosis but also for the evaluation of therapeutic efficacy and resilience to trauma. Ongoing research is directed at identifying molecular biomarkers for PTSD, including traumatic stress induced proteins, transcriptomes, genomic variances and genetic modulators, using biologic samples from subjects' blood, saliva, urine, and postmortem brain tissues. However, the correlation of these biomarker molecules in peripheral or postmortem samples to altered brain functions associated with psychiatric symptoms in PTSD remains unresolved. Here, we present an animal model of PTSD in which both peripheral blood and central brain biomarkers, as well as behavioral phenotype, can be collected and measured, thus providing the needed correlation of the central biomarkers of PTSD, which are mechanistic and pathognomonic but cannot be collected from people, with the peripheral biomarkers and behavioral phenotypes, which can.
Our animal model of PTSD employs restraint and tail shocks repeated for three continuous days - the inescapable tail-shock model (ITS) in rats. This ITS model mimics the pathophysiology of PTSD 17, 7, 4, 10. We and others have verified that the ITS model induces behavioral and neurobiological alterations similar to those found in PTSD subjects 17, 7, 10, 9. Specifically, these stressed rats exhibit (1) a delayed and exaggerated startle response appearing several days after stressor cessation, which given the compressed time scale of the rat's life compared to a humans, corresponds to the one to three months delay of symptoms in PTSD patients (DSM-IV-TR PTSD Criterian D/E 13), (2) enhanced plasma corticosterone (CORT) for several days, indicating compromise of the hypothalamopituitary axis (HPA), and (3) retarded body weight gain after stressor cessation, indicating dysfunction of metabolic regulation.
The experimental paradigms employed for this model are: (1) a learned helplessness paradigm in the rat assayed by measurement of acoustic startle response (ASR) and a charting of body mass; (2) microdissection of the rat brain into regions and nuclei; (3) enzyme-linked immunosorbent assay (ELISA) for blood levels of CORT; (4) a gene expression microarray plus related bioinformatics tools 18. This microarray, dubbed rMNChip, focuses on mitochondrial and mitochondria-related nuclear genes in the rat so as to specifically address the neuronal bioenergetics hypothesized to be involved in PTSD.

We describe a rat model of post traumatic stress disorder (PTSD) that reveals the persistent alterations in neuroendocrine function and the delayed long-term, exaggerated fear response, characteristic of PTSD patients. The animal model and methods described here are useful for correlating biomarkers in brain nuclei, which are mechanistic but cannot be measured in patients, with biomarkers in peripheral white blood cells, which can.

在动物模型揭示神经，血液，创伤后应激障碍和行为相关因素的生物标志物

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for ...

科研

医学

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.

伤口愈合的小鼠模型

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

Femoral Bone Marrow Aspiration in Live Mice

Serial sampling of the cellular composition of bone marrow (BM) is a routine procedure critical to clinical hematology. This protocol describes a detailed step-by-step technical procedure for an analogous procedure in live mice which allows for serial characterization of cells present in the BM. This procedure facilitates studies aimed to detect the presence of exogenously administered cells within the BM of mice as would be done in xenograft studies for instance. Moreover, this procedure allows for the retrieval and characterization of cells enriched in the BM such as hematopoietic stem and progenitor cells (HSPCs) without sacrifice of mice. Given that the cellular composition of peripheral blood is not necessarily reflective of proportions and types of stem and progenitor cells present in the marrow, procedures which provide access to this compartment without requiring termination of the mice are very helpful. The use of femoral bone marrow aspiration is illustrated here for cytological analysis of marrow cells, flow cytometric characterization of the hematopoietic stem/progenitor compartment, and culture of sorted HSPCs obtained by femoral BM aspiration compared with conventional marrow harvest.

This protocol describes a procedure for serial sampling of femoral bone marrow (BM) without requiring the sacrifice of mice. This procedure facilitates longitudinal studies of the BM composition of mice over time and provides serial access to cells within the BM for ex vivo and transplantation studies.

股骨骨髓的愿望在活体小鼠

Serial sampling of the cellular composition of bone marrow (BM) is a routine procedure critical to clinical hematology. This protocol describes a detailed ...

A Simple Critical-sized Femoral Defect Model in Mice

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all limb bone fractures fail to heal completely due to the extent of the trauma, disease, or age of the patient. Our ability to improve bone regenerative strategies is critically dependent on the ability to mimic serious bone trauma in test animals, but the generation and stabilization of large bone lesions is technically challenging. In most cases, serious long bone trauma is mimicked experimentally by establishing a defect that will not naturally heal. This is achieved by complete removal of a bone segment that is larger than 1.5 times the diameter of the bone cross-section. The bone is then stabilized with a metal implant to maintain proper orientation of the fracture edges and allow for mobility. 
Due to their small size and the fragility of their long bones, establishment of such lesions in mice are beyond the capabilities of most research groups. As such, long bone defect models are confined to rats and larger animals. Nevertheless, mice afford significant research advantages in that they can be genetically modified and bred as immune-compromised strains that do not reject human cells and tissue.
Herein, we demonstrate a technique that facilitates the generation of a segmental defect in mouse femora using standard laboratory and veterinary equipment. With practice, fabrication of the fixation device and surgical implantation is feasible for the majority of trained veterinarians and animal research personnel. Using example data, we also provide methodologies for the quantitative analysis of bone healing for the model.

Animal models are frequently employed to mimic serious bone injury in biomedical research. Due to their small size, establishment of stabilized bone lesions in mice are beyond the capabilities of most research groups. Herein, we describe a simple method for establishing and analyzing experimental femoral defects in mice.

一个简单的临界尺寸股骨缺损模型小鼠

While bone has a remarkable capacity for regeneration, serious bone trauma often results in damage that does not properly heal. In fact, one tenth of all ...

Facial Nerve Axotomy in Mice: A Model to Study Motoneuron Response to Injury

The goal of this surgical protocol is to expose the facial nerve, which innervates the facial musculature, at its exit from the stylomastoid foramen and either cut or crush it to induce peripheral nerve injury. Advantages of this surgery are its simplicity, high reproducibility, and the lack of effect on vital functions or mobility from the subsequent facial paralysis, thus resulting in a relatively mild surgical outcome compared to other nerve injury models. A major advantage of using a cranial nerve injury model is that the motoneurons reside in a relatively homogenous population in the facial motor nucleus in the pons, simplifying the study of the motoneuron cell bodies. Because of the symmetrical nature of facial nerve innervation and the lack of crosstalk between the facial motor nuclei, the operation can be performed unilaterally with the unaxotomized side serving as a paired internal control. A variety of analyses can be performed postoperatively to assess the physiologic response, details of which are beyond the scope of this article. For example, recovery of muscle function can serve as a behavioral marker for reinnervation, or the motoneurons can be quantified to measure cell survival. Additionally, the motoneurons can be accurately captured using laser microdissection for molecular analysis. Because the facial nerve axotomy is minimally invasive and well tolerated, it can be utilized on a wide variety of genetically modified mice. Also, this surgery model can be used to analyze the effectiveness of peripheral nerve injury treatments. Facial nerve injury provides a means for investigating not only motoneurons, but also the responses of the central and peripheral glial microenvironment, immune system, and target musculature. The facial nerve injury model is a widely accepted peripheral nerve injury model that serves as a powerful tool for studying nerve injury and regeneration.

We present a surgical protocol detailing how to perform a cut or crush axotomy on the facial nerve in the mouse. The facial nerve axotomy can be employed to study the physiological response to nerve injury and test therapeutic techniques.

面神经干切断小鼠:模型，研究运动神经元损伤反应

The goal of this surgical protocol is to expose the facial nerve, which innervates the facial musculature, at its exit from the stylomastoid foramen and ...

Establishment of a Segmental Femoral Critical-size Defect Model in Mice Stabilized by Plate Osteosynthesis

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a model organism, the mouse has already been widely used in bone-related research. Large diaphyseal bone defect models in mice, however, are sparse and often use bone fixation which fills the bone marrow cavity and does not provide optimal mechanical stability. The objectives of the current study were to develop a critical-size, segmental, femoral defect in nude mice. A 3.5-mm mid-diaphyseal femoral ostectomy (approximately 25% of the femur length) was performed using a dedicated jig, and was stabilized with an anterior located locking plate and 4 locking screws. The bone defect was subsequently either left empty or filled with a bone substitute (syngenic bone graft or coralline scaffold). Bone healing was monitored noninvasively using radiography and in vivo micro-computed-tomography and was subsequently assessed by ex vivo micro-computed-tomography and undecalcified histology after animal sacrifice, 10 weeks postoperatively. The recovery of all mice was excellent, a full-weight-bearing was observed within one day following the surgical procedure. Furthermore, stable bone fixation and consistent fixation of the implanted materials were achieved in all animals tested throughout the study. When the bone defects were left empty, non-union was consistently obtained. In contrast, when the bone defects were filled with syngenic bone grafts, bone union was always observed. When the bone defects were filled with coralline scaffolds, newly-formed bone was observed in the interface between bone resection edges and the scaffold, as well as within a short distance within the scaffold. 
The present model describes a reproducible critical-size femoral defect stabilized by plate osteosynthesis with low morbidity in mice. The new load-bearing segmental bone defect model could be useful for studying the underlying mechanisms in bone regeneration pertinent to orthopaedic applications.

Although mouse models are invaluable tools for bone tissue engineering, models of long bone defects are sparse. This need motivated development of the present protocol which uses a locking plate with four screws and a dedicated jig to perform and stabilize a reproducible, femoral, critical-size defect with low morbidity.

在小鼠节段性股临界尺寸缺陷模型的建立接骨板稳定

The use of tissue-engineered bone constructs is an appealing strategy to overcome drawbacks of autografts for the treatment of massive bone defects. As a ...

An Intramedullary Locking Nail for Standardized Fixation of Femur Osteotomies to Analyze Normal and Defective Bone Healing in Mice

Bone healing models are essential to the development of new therapeutic strategies for clinical fracture treatment. Furthermore, mouse models are becoming more commonly used in trauma research. They offer a large number of mutant strains and antibodies for the analysis of the molecular mechanisms behind the highly differentiated process of bone healing. To control the biomechanical environment, standardized and well-characterized osteosynthesis techniques are mandatory in mice. Here, we report on the design and use of an intramedullary nail to stabilize open femur osteotomies in mice. The nail, made of medical-grade stainless steel, provides high axial and rotational stiffness. The implant further allows the creation of defined, constant osteotomy gap sizes from 0.00 mm to 2.00 mm. Intramedullary locking nail stabilization of femur osteotomies with gap sizes of 0.00 mm and 0.25 mm result in adequate bone healing through endochondral and intramembranous ossification. Stabilization of femur osteotomies with a gap size of 2.00 mm results in atrophic non-union. Thus, the intramedullary locking nail can be used in healing and non-healing models. A further advantage of the use of the nail compared to other open bone healing models is the possibility to adequately fix bone substitutes and scaffolds in order to study the process of osseous integration. A disadvantage of the use of the intramedullary nail is the more invasive surgical procedure, inherent to all open procedures compared to closed models. A further disadvantage may be the induction of some damage to the intramedullary cavity, inherent to all intramedullary stabilization techniques compared to extramedullary stabilization procedures.

This protocol describes an osteosynthesis technique using an intramedullary locking nail for standardized fixation of femur osteotomies, which can be used to analyze normal and defective bone healing in mice.

髓内锁钉股骨截骨标准化固定用于分析正常和有缺陷的骨愈合小鼠

Bone healing models are essential to the development of new therapeutic strategies for clinical fracture treatment. Furthermore, mouse models are becoming ...

Development of a Direct Pulp-capping Model for the Evaluation of Pulpal Wound Healing and Reparative Dentin Formation in Mice

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is often capped with biocompatible materials in order to expedite pulpal wound healing. The ultimate goal is to regenerate reparative dentin, a physical barrier that functions as a "biological seal" and protects the underlying pulp tissue. Although this direct pulp-capping procedure has long been used in dentistry, the underlying molecular mechanism of pulpal wound healing and reparative dentin formation is still poorly understood. To induce reparative dentin, pulp capping has been performed experimentally in large animals, but less so in mice, presumably due to their small sizes and the ensuing technical difficulties. Here, we present a detailed, step-by-step method of performing a pulp-capping procedure in mice, including the preparation of a Class-I-like cavity, the placement of pulp-capping materials, and the restoration procedure using dental composite. Our pulp-capping mouse model will be instrumental in investigating the fundamental molecular mechanisms of pulpal wound healing in the context of reparative dentin in vivo by enabling the use of transgenic or knockout mice that are widely available in the research community.

We describe a step-by-step method of performing direct pulp capping on mice teeth for the evaluation of pulpal wound healing and reparative dentin formation in vivo.

伤口的牙髓评价一个直接盖髓模型的开发和治疗小鼠中修复性牙本质形成

Dental pulp is a vital organ of a tooth fully protected by enamel and dentin. When the pulp is exposed due to cariogenic or iatrogenic injuries, it is ...

A Minimally Invasive Model to Analyze Endochondral Fracture Healing in Mice Under Standardized Biomechanical Conditions

Bone healing models are necessary to analyze the complex mechanisms of fracture healing to improve clinical fracture treatment. During the last decade, an increased use of mouse models in orthopedic research was noted, most probably because mouse models offer a large number of genetically-modified strains and special antibodies for the analysis of molecular mechanisms of fracture healing. To control the biomechanical conditions, well-characterized osteosynthesis techniques are mandatory, also in mice. Here, we report on the design and use of a closed bone healing model to stabilize femur fractures in mice. The intramedullary screw, made of medical-grade stainless steel, provides through fracture compression an axial and rotational stability compared to the mostly used simple intramedullary pins, which show a complete lack of axial and rotational stability. The stability achieved by the intramedullary screw allows the analysis of endochondral healing. A large amount of callus tissue, received after stabilization with the screw, offers ideal conditions to harvest tissue for biochemical and molecular analyses. A further advantage of the use of the screw is the fact that the screw can be inserted into the femur with a minimally invasive technique without inducing damage to the soft tissue. In conclusion, the screw is a unique implant that can ideally be used in closed fracture healing models offering standardized biomechanical conditions.

This protocol describes a minimally invasive osteosynthesis technique using an intramedullary screw for standardized stabilization of femur fractures, which can be used to analyze endochondral bone healing in mice.

标准化生物力学条件下小鼠软骨骨折愈合的微创模型分析

Bone healing models are necessary to analyze the complex mechanisms of fracture healing to improve clinical fracture treatment. During the last decade, an ...

A Closed-chest Model to Induce Transverse Aortic Constriction in Mice

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to perform a partial thoracotomy to visualize the transverse aortic arch. However, the surgical trauma caused by the thoracotomy in open-chest models changes the respiratory physiology as the ribs are dissected and left unattached after chest closure. To prevent this, we established a minimally invasive, closed chest approach via lateral thoracotomy. Herein we approach the aortic arch via the 2nd intercostal space without entering the chest cavities, leaving the mouse with a less traumatic injury to recover from. We perform this operation using standard laboratory settings for open chest TAC procedures with equal survival rates. Apart from maintaining physiological breathing patterns due to the closed chest approach, the mice seem to benefit by showing rapid recovery, as the less invasive technique appears to facilitate a fast healing process and to reduce immune response after trauma.

Here, we present a protocol of Transverse Aortic constriction (TAC) via a lateral thoracotomy. This technique is a minimally invasive, closed chest surgical procedure aiming to simulate pressure overload and heart failure in mice utilizing standard TAC laboratory settings.

诱导小鼠横向主动脉狭窄的闭合胸模型

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to ...