本研究由国家自然科学基金 82101000 （H. W.）、U21A20368 （L. Y.） 和 82100982 （F. L.） 资助，并由四川省科技计划 2023NSFSC1499（H. W.） 资助。

本研究所有动物手术均由四川大学华西口腔医学院伦理委员会（WCHSIRB-D-2021-597）审查批准。本研究使用Sprague-Dawley大鼠（雄性，体重300g）。
1. 术前准备
<ol><li>仪器准备<ol><li>本研究使用的手术器械见 图1A ：电动剃须刀、组织剪刀、眼科剪刀、眼科镊子、一次性使用手术刀、骨膜分离器、口腔低速手机、口腔探头、一次性使用冲洗吸尘器、针架、3.0缝合线。</li>
		<li>准备一个口腔探针，并根据缺陷的直径用记号笔标记探针的大弯曲端（图1B）。使用它来确定手术过程中缺陷的大小。</li>
		<li>使用前对用于执行该程序的所有手术材料和器械进行消毒。将所需材料包装在折叠的布或包装纸中，并用高压灭菌胶带密封以进行蒸汽灭菌（125-135°C 20-25 分钟）。</li>
		<li>手术区域准备：喷洒75%酒精，对手术台和手术台周围环境进行消毒。在手术台上创建一个大约 60 cm x 90 cm 的无菌区域，用高压灭菌的窗帘。</li>
	</ol></li>
	<li>麻醉准备<ol><li>用10%盐酸氯胺酮（50-100mg / kg）和2%甲苯噻嗪2mg / kg腹膜内麻醉大鼠。使用皮下注射卡洛芬（5 mg/kg）进行术前和术中镇痛。通过脚趾捏试验检查麻醉深度。麻醉后，将无菌眼膏涂抹在眼睛上，以防止眼睛干涩和角膜损伤。</li>
	</ol></li>
</ol>2. 外科手术
<ol><li>将大鼠置于无菌手术台上的侧卧位置，并用电动剃须刀去除下肢毛发。</li>
	<li>使用5%的碘伏溶液和75%的酒精对手术区域的皮肤组织进行消毒。</li>
	<li>找到股骨近端和远端，沿股骨的长轴做一个2.5厘米的切口，切开大鼠的皮肤组织。<ol><li>用眼科镊子和组织剪刀将皮肤层与筋膜分开，并通过股二头肌和股外侧肌肉暴露到股骨的外侧通路。</li>
		<li>找到两块肌肉间隔的交叉点（一条白色组织线），并用一次性手术刀片沿着肌肉边界小心地分开，直到到达股骨表面。 注意：使用一次性刀片分离肌肉时，重要的是沿着肌肉隔膜分离，并注意在软组织内引起血管损伤。初学者和不熟悉解剖结构的人必须使用眼科钳子和骨膜分离器在两个肌肉块之间进行钝性解剖。</li>
	</ol></li>
	<li>应用骨膜分离器将股骨表面肌肉钝性分离，并暴露股骨干中部。</li>
	<li>使用无菌记号笔标记位于股骨头外侧 1/3 斜嵴顶部的股骨干中表面的缺损部位区域。</li>
	<li>使用口腔低速手机搭配直径为 1.2 毫米的慢动作球钻，在标记部位垂直于骨表面钻一个小孔，破坏骨膜和骨皮质，深度足以到达骨髓腔。在此钻孔深度水平上，向所有方向扩展平行于该深度的孔的尺寸，修整以实现箱形腔形状。</li>
	<li>使用平行于缺陷边缘的标记口腔探针来确定制备过程中和之后的缺陷直径和形态。</li>
	<li>分别用 3-0 条单丝可吸收缝合线闭合肌肉层和皮肤层，并从内到外用 5% 的碘伏对手术区域进行消毒。</li>
</ol>3. 术后护理
<ol><li>手术后，皮下给予卡洛芬（5mg / kg），并将大鼠置于恒温加热垫上，直至从麻醉中恢复。当老鼠恢复意识时，轻轻地将其移至装有干燥高压灭菌垫料的笼子中。</li>
	<li>术后继续镇痛 24 小时，术后监测 1 周。</li>
</ol>4. 样品采集和分析
<ol><li>手术后通过腹膜内注射戊巴比妥钠100-200mg / kg对大鼠进行人性安乐死。小心地分离股骨表面的肌肉和筋膜组织，并完全切除手术侧的股骨。术后0天（图2A，B），2周，4周和6周收集标本。</li>
	<li>将股骨标本固定在4%多聚甲醛中24小时。使用显微计算机断层扫描分析股骨的结构。设置扫描参数如下：X射线管电位，70 kVp;过滤器，AL 0.5 mm;X 射线强度，0.2 mA;体素尺寸，17 μm;积分时间为 1 × 300 ms。使用位图数据重建 3D 模型图像。</li>
	<li>在10%EDTA中将标本脱钙8周，然后在一系列分级乙醇稀释液中对股骨标本进行脱水。然后，将样品嵌入石蜡15 中。<ol><li>将嵌入的样品从矢状面切成5μm石蜡切片。</li>
		<li>使用苏木精和伊红 （H&E） 染色试剂盒和 Masson 染色试剂盒对切片进行染色。通过组织病理学观察缺损区域的愈合情况。</li>
	</ol></li>
</ol>

大鼠股骨盒的建立和评估：空腔缺陷模型

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所有原始数据和图像均包含在本文中。作者声明没有利益冲突

临床前动物模型对于检查骨愈合和生物材料对骨再生的影响至关重要。该协议说明了股骨箱腔缺损模型，该模型复制了与临床骨再生相关的膜内骨形成过程。使用预先标记的口腔探针对缺陷区域进行术中标准化。显微 CT 和组织病理学染色结果显示，6 周内逐渐愈合，骨膜增厚，新小梁骨形成，随后皮质骨形成致密。在愈合过程中未观察到软骨组织，表明膜内成骨。这一发现为研究生物材料或药物?...

骨折和有缺陷的骨头通常由外伤、肿瘤、炎症和先天性畸形引起 1,2。尽管年轻健康个体的骨组织通常具有强大的再生能力3，但超过临界尺寸的缺陷或由于全身性疾病（例如糖尿病、骨质疏松症和感染）导致的愈合障碍仍可能导致并发症，例如骨骼不连续性或愈合受损4。为了应对这一临床挑战，骨移植或生物材料通常用于替换严重有缺陷的骨骼或重建大骨段。然而，这些治疗有局限性。例如，尽管被认为是金标准，但自体骨移植受到供体供应限制和潜在供体部位并发症的影响 5,6。同种异体移植物也存在一定的风险，例如免疫介导的排斥反应、疾病的潜在传播以及对移植物的生物力学和生物学特性的负面影响7.
近年来，专注于骨缺损愈合机制的研究激增。替代生物材料的使用和组织工程的进步已成为骨再生领域的突出话题8.在这些生物材料可以应用于人体治疗之前，它们必须在体外和体内进行测试，以确保其有效性和安全性。然而，体外环境复杂性的降低以及免疫和炎症反应的缺乏限制了各种生物材料的体外评估。因此，需要为各种类型的骨组织缺损建立动物模型9.动物模型允许在不同负载条件下评估生物材料，促进对物种特异性骨骼特征的理解，并深入了解动物模型与人类临床情况之间的相似性。这些优势对于研究骨-支架相互作用并将研究结果转化为临床实践至关重要 9,10。
目前，机械骨缺损动物模型被广泛用于验证生物材料的性能，其中颅骨缺损模型和节段性骨缺损模型是最常用的方法11。节段性骨缺损模型通常用于模拟以骨不连结束的严重长骨或胫骨创伤，由于其均匀的尺寸和明确的解剖位置而具有优势，简化了新骨形成和血运重建的放射学或组织学评估。然而，这些模型需要金属植入物来稳定双侧骨折段，并且需要涉及软骨内和膜内骨化的复杂愈合过程12。另一方面，颅骨缺损模型因其标准化的缺损直径、方便的手术通路以及硬脑膜和软组织的支持功能而成为评估生物材料的主要筛选工具13。尽管它们被广泛用于在临床相关场景中模拟膜内骨形成，但由于它们在愈合过程中的非承重性质，它们不适合评估生物力学负荷条件下的骨愈合14。
为了解决这些局限性，我们在大鼠股骨干组织中建立了箱腔皮质骨缺损模型。利用显微计算机断层扫描（CT）、三维（3D）重建和组织病理学染色（苏木精和伊红[HE]和Masson），分析了该模型在止血条件下的愈合过程。我们旨在为评估生物材料在生物力学负载条件下的性能以及研究骨再生相对于膜内骨化的生物工程和机制提供新的见解。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>1.2 mm slow speed ball drill</td>
			<td>Dreybird Medical Equipment Co., Ltd.</td>
			<td>RA3-012</td>
			<td>For preparation of box cavity defects</td>
		</tr>
		<tr>
			<td>3.0 suture</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>None</td>
			<td>For suturing wounds</td>
		</tr>
		<tr>
			<td>4% paraformaldehyde</td>
			<td>Biosharp</td>
			<td>BL539A</td>
			<td>For fix the femoral specimens</td>
		</tr>
		<tr>
			<td>Cotton balls</td>
			<td>Haishi Hainuo Group Co.,&#160; Ltd.</td>
			<td>20120047</td>
			<td>For skin sterilization and cleaning of surgical field</td>
		</tr>
		<tr>
			<td>Cotton sticks</td>
			<td>Lakong Medical Devices Co., Ltd.</td>
			<td>M6500R</td>
			<td>For skin disinfection</td>
		</tr>
		<tr>
			<td>Dental technician grinding machine</td>
			<td>Marathon</td>
			<td>N3-140232</td>
			<td>For preparation of box cavity defects</td>
		</tr>
		<tr>
			<td>Disposable scalpel</td>
			<td>Hangzhou Huawei Medical Supplies Co., Ltd.</td>
			<td>20100227</td>
			<td>For creating skin incisions as well as to sharply separate muscle tissue</td>
		</tr>
		<tr>
			<td>Electric shaver</td>
			<td>JASE</td>
			<td>BM320210</td>
			<td>Removal of hair tissue from the surgical area</td>
		</tr>
		<tr>
			<td>Hematoxylin and Eosin Stain kit</td>
			<td>Biosharp</td>
			<td>C1005</td>
			<td>For the histological analysis of the specimens</td>
		</tr>
		<tr>
			<td>Masson&#8217;s Trichrome Stain Kit</td>
			<td>Solarbio</td>
			<td>G1340</td>
			<td>For the histological analysis of the specimens</td>
		</tr>
		<tr>
			<td>Micro CT</td>
			<td>Scanco medical ag</td>
			<td>&#181;CT 45</td>
			<td>For analyzing the healing of defects in femoral samples</td>
		</tr>
		<tr>
			<td>Needle holder</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>None</td>
			<td>For suture-holding needles</td>
		</tr>
		<tr>
			<td>Olympus Research Grade Whole Slide Scanning System VS200</td>
			<td>Chengdu Knowledge Technology Co.</td>
			<td>VS200</td>
			<td>For analyzing the results of HE staining and Masson staining</td>
		</tr>
		<tr>
			<td>Ophthalmic forceps</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>None</td>
			<td>For clamping skin, muscle tissue</td>
		</tr>
		<tr>
			<td>Ophthalmic scissors</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>None</td>
			<td>For forming a skin incision approach</td>
		</tr>
		<tr>
			<td>Oral low-speed handpiece</td>
			<td>Marathon</td>
			<td>Y221101003</td>
			<td>For preparation of box cavity defects</td>
		</tr>
		<tr>
			<td>Oral probe</td>
			<td>Shanghai Sangda Medical Insurance Co., Ltd.</td>
			<td>20000143</td>
			<td>For measuring the diameter of defects</td>
		</tr>
		<tr>
			<td>Periosteal separator</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>None</td>
			<td>For blunt separation of muscle tissue</td>
		</tr>
		<tr>
			<td>Sprague&#8211;Dawley rats</td>
			<td>Byrness Weil Biotech Ltd</td>
			<td>None</td>
			<td>For the establishment of femoral bone boxy cavitary defect</td>
		</tr>
		<tr>
			<td>Tissue scissors</td>
			<td>Chengdu Shifeng Co., Ltd.</td>
			<td>None</td>
			<td>For forming a skin incision approach</td>
		</tr>
	</tbody>
</table>

creating a box-cavity defect model in the cortical bone of rat femora

严重的骨缺损或复杂骨折会导致严重的并发症，例如骨不连或骨愈合不足。组织工程涉及细胞、支架和细胞因子的应用，被认为是骨再生的有前途的解决方案。因此，模拟骨骼缺陷的各种动物模型在探索组织工程治疗骨愈合的潜力方面发挥着至关重要的作用。本研究建立了大鼠股骨中部的箱形皮质骨缺损模型，可作为评估生物材料促进骨愈合功能的理想模型。这种箱形皮质骨缺损是使用口腔低速手机钻孔的，并通过车床针进行成型。术后立即进行显微CT分析，确认箱腔皮质骨缺损成功建立。然后在手术后的多个时间点（0 天、2 周、4 周和 6 周）收获大鼠手术侧的股骨。使用micro-CT，苏木精和伊红（H&E）染色以及Masson三色染色评估了每个样品缺陷区域的愈合过程。这些结果表明，愈合模式与膜内骨化一致，6 周内基本愈合完成。这种动物模型的愈合过程的分类提供了一种有效的 体内 方法，用于研究在骨组织缺损愈合过程中靶向膜内骨化的新型生物材料和药物。

在该协议中，我们通过钻孔成功建立了尺寸为4.5 mm x 1.5 mm的大鼠股骨箱腔缺陷模型。为了分析手术侧的愈合过程，我们在术后0 d、2周、4周和6周采集了手术侧股组织，这是大鼠股骨创伤愈合过程中软骨内骨化、膜内骨化和骨重塑的关键时间点2。在术后第 0 天，从显微 CT 位图数据重建 3D 模型显示，我们成功模拟了股骨中的箱腔缺损，其尺寸为 4.5 mm x 1.5 mm...

Watch this Scientific Journal Video about Creating a Box-Cavity Defect Model in the Cortical Bone of Rat Femora at JoVE.com

作者聚焦：用于增强生物材料和骨再生评估的箱腔皮质方法

在这里，我们提出了一种在大鼠股骨干组织中产生箱腔缺损的方案。该模型可以评估生物材料在生物力学应力下的性能，并探索与膜内成骨相关的骨再生机制。

在大鼠股骨皮质骨中创建箱腔缺损模型

Severe bone defects or complex fractures can result in serious complications such as nonunion or insufficient bone healing. Tissue engineering, which ...

各种具有骨缺损的动物模型被应用于探索组织工程对骨愈合的治疗潜力。然而，它们中的大多数在生物力学负载条件下无法产生标准缺陷，这对于评估生物材料的骨愈合效果具有重要意义。与传统的骨缺损模型相比，最近的箱腔皮质骨缺损模型提供了显着的进步。它不仅可以提供标准化的骨缺损尺寸，以便更准确地量化骨愈合，而且还允许在生物力学负载条件下评估生物材料，而无需额外固定。我们的方案提供了一种全面且可推广的方法，用于在骨再生模型的体内研究中研究膜内骨形成。这对骨组织工程领域特别有益。

creating-a-box-cavity-defect-model-in-the-cortical-bone-of-rat-femora

Research

JoVE Journal

Medicine

Creating a Box-Cavity Defect Model in the Cortical Bone of Rat Femora

576 次观看.  West China Hospital of Stomatology, Sichuan University. 各种具有骨缺损的动物模型被应用于探索组织工程对骨愈合的治疗潜力。然而，它们中的大多数在生物力学负载条件下无法产生标准缺陷，这对于评估生物材料的骨愈合效果具有重要意义。与传统的骨缺损模型相比，最近的箱腔皮质骨缺损模型提供了显着的进步。它不仅可以提供标准化的骨缺损尺寸，以便更准确地量化骨愈合，而且还允许在生物力学负载条件下评估生?...

视频: 作者聚焦：用于增强生物材料和骨再生评估的箱腔皮质方法

Multi-modal Imaging of Angiogenesis in a Nude Rat Model of Breast Cancer Bone Metastasis Using Magnetic Resonance Imaging, Volumetric Computed Tomography and Ultrasound

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell proliferation in the bone marrow cavity as well as for interaction of tumor and bone cells resulting in local bone destruction. Our aim was to develop a model of experimental bone metastasis that allows in vivo assessment of angiogenesis in skeletal lesions using non-invasive imaging techniques.
For this purpose, we injected 105 MDA-MB-231 human breast cancer cells into the superficial epigastric artery, which precludes the growth of metastases in body areas other than the respective hind leg1. Following 25-30 days after tumor cell inoculation, site-specific bone metastases develop, restricted to the distal femur, proximal tibia and proximal fibula1. Morphological and functional aspects of angiogenesis can be investigated longitudinally in bone metastases using magnetic resonance imaging (MRI), volumetric computed tomography (VCT) and ultrasound (US).
MRI displays morphologic information on the soft tissue part of bone metastases that is initially confined to the bone marrow cavity and subsequently exceeds cortical bone while progressing. Using dynamic contrast-enhanced MRI (DCE-MRI) functional data including regional blood volume, perfusion and vessel permeability can be obtained and quantified2-4. Bone destruction is captured in high resolution using morphological VCT imaging. Complementary to MRI findings, osteolytic lesions can be located adjacent to sites of intramedullary tumor growth. After contrast agent application, VCT angiography reveals the macrovessel architecture in bone metastases in high resolution, and DCE-VCT enables insight in the microcirculation of these lesions5,6. US is applicable to assess morphological and functional features from skeletal lesions due to local osteolysis of cortical bone. Using B-mode and Doppler techniques, structure and perfusion of the soft tissue metastases can be evaluated, respectively. DCE-US allows for real-time imaging of vascularization in bone metastases after injection of microbubbles7.
In conclusion, in a model of site-specific breast cancer bone metastases multi-modal imaging techniques including MRI, VCT and US offer complementary information on morphology and functional parameters of angiogenesis in these skeletal lesions.

In the pathogenesis of bone metastasis, angiogenesis is a crucial process and therefore represents a target for imaging and therapy. Here, we present a rat model of site-specific breast cancer bone metastasis and describe strategies to non-invasively image angiogenesis in vivo using magnetic resonance imaging, volumetric computed tomography and ultrasound.

利用磁共振成像，容积电脑断层扫描及超音波的裸鼠乳腺癌骨转移大鼠模型血管生成的多模态成像

Angiogenesis is an essential feature of cancer growth and metastasis formation. In bone metastasis, angiogenic factors are pivotal for tumor cell ...

科研

医学

A Model of Chronic Nutrient Infusion in the Rat

Chronic exposure to excessive levels of nutrients is postulated to affect the function of several organs and tissues and to contribute to the development of the many complications associated with obesity and the metabolic syndrome, including type 2 diabetes. To study the mechanisms by which excessive levels of glucose and fatty acids affect the pancreatic beta-cell and the secretion of insulin, we have established a chronic nutrient infusion model in the rat. The procedure consists of catheterizing the right jugular vein and left carotid artery under general anesthesia; allowing a 7-day recuperation period; connecting the catheters to the pumps using a swivel and counterweight system that enables the animal to move freely in the cage; and infusing glucose and/or Intralipid (a soybean oil emulsion which generates a mixture of approximately 80% unsaturated/20% saturated fatty acids when infused with heparin) for 72 hr. This model offers several advantages, including the possibility to finely modulate the target levels of circulating glucose and fatty acids; the option to co-infuse pharmacological compounds; and the relatively short time frame as opposed to dietary models. It can be used to examine the mechanisms of nutrient-induced dysfunction in a variety of organs and to test the effectiveness of drugs in this context.

A protocol for chronic infusions of glucose and Intralipid in rats is described. This model can be used to study the impact of nutrient excess on organ function and physiological parameters.

在大鼠模型慢性营养输液

Chronic exposure to excessive levels of nutrients is postulated to  affect the function of several organs and tissues and to contribute to the  ...

Adjustable Stiffness, External Fixator for the Rat Femur Osteotomy and Segmental Bone Defect Models

The mechanical environment around the healing of broken bone is very important as it determines the way the fracture will heal. Over the past decade there has been great clinical interest in improving bone healing by altering the mechanical environment through the fixation stability around the lesion. One constraint of preclinical animal research in this area is the lack of experimental control over the local mechanical environment within a large segmental defect as well as osteotomies as they heal. In this paper we report on the design and use of an external fixator to study the healing of large segmental bone defects or osteotomies. This device not only allows for controlled axial stiffness on the bone lesion as it heals, but it also enables the change of stiffness during the healing process in vivo. The conducted experiments have shown that the fixators were able to maintain a 5 mm femoral defect gap in rats in vivo during unrestricted cage activity for at least 8 weeks. Likewise, we observed no distortion or infections, including pin infections during the entire healing period. These results demonstrate that our newly developed external fixator was able to achieve reproducible and standardized stabilization, and the alteration of the mechanical environment of in vivo rat large bone defects and various size osteotomies. This confirms that the external fixation device is well suited for preclinical research investigations using a rat model in the field of bone regeneration and repair.

One constraint of preclinical research in the field of bone repair is the lack of experimental control over the local mechanical environment within a healing bone lesion. We report the design and use of an external fixator for bone repair with the ability to change fixator stiffness in vivo.

可调节刚度，外固定支架的大鼠股骨截骨和节段性骨缺损模型

The mechanical environment around the healing of broken bone is very important as it determines the way the fracture will heal. Over the past decade there ...

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 ...

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 ...

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.

用线 Myography 测定小鼠肠系膜动脉的等轴收缩力

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

Using a Knee Arthrometer to Evaluate Tissue-specific Contributions to Knee Flexion Contracture in the Rat

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ROM is called a joint contracture and results in increased morbidity. Due to the difficulty of reversing established knee contractures, early detection is important, and hence, knowing risk factors for their development is essential. The rat represents a good model with which the effect of an intervention can be studied due to the similarity of rat knee anatomy to that of humans, the rat's ability to tolerate long durations of knee immobilization in flexion, and because mechanical data can be correlated with histologic and biochemical analysis of knee tissue.
Using an automated arthrometer, we demonstrate a validated, precise, reproducible, user-independent method of measuring the extension ROM of the rat knee joint at specific torques. This arthrometer can be used to determine the effects of interventions on knee joint ROM in the rat.

The goal of the protocol is to measure the extension range of motion of the rat knee. The effects of various diseases that increase the stiffness of the knee joint and the effectiveness of treatments can be quantified.

用膝关节关节仪评价大鼠膝关节屈曲收缩的特定组织贡献

Normal knee range of motion (ROM) is critical to well-being and allows one to perform basic activities such as walking, climbing stairs and sitting. Lost ...

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

Bone fractures impose a tremendous socio-economic burden on patients, in addition to significantly affecting their quality of life. Therapeutic strategies that promote efficient bone healing are non-existent and in high demand. Effective and reproducible animal models of fractures healing are needed to understand the complex biological processes associated with bone regeneration. Many animal models of fracture healing have been generated over the years; however, murine fracture models have recently emerged as powerful tools to study bone healing. A variety of open and closed models have been developed, but the closed femoral fracture model stands out as a simple method for generating rapid and reproducible results in a physiologically relevant manner. The goal of this surgical protocol is to generate unilateral closed femoral fractures in mice and facilitate a post-fracture stabilization of the femur by inserting an intramedullary steel rod. Although devices such as a nail or a screw offer greater axial and rotational stability, the use of an intramedullary rod provides a sufficient stabilization for consistent healing outcomes without producing new defects in the bone tissue or damaging nearby soft tissue. Radiographic imaging is used to monitor the progression of callus formation, bony union, and subsequent remodeling of the bony callus. Bone healing outcomes are typically associated with the strength of the healed bone and measured with torsional testing. Still, understanding the early cellular and molecular events associated with fracture repair is critical in the study of bone tissue regeneration. The closed femoral fracture model in mice with intramedullary fixation serves as an attractive platform to study bone fracture healing and evaluate therapeutic strategies to accelerate healing.

The murine closed femoral fracture model is a powerful platform to study fracture healing and novel therapeutic strategies to accelerate bone regeneration. The goal of this surgical protocol is to generate unilateral closed femoral fractures in mice using an intramedullary steel rod to stabilize the femur.

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 ...

Acupuncture in a Rat Model of Asthma

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes methods for the fabrication of the high platform, establishes a rat model of asthma and measures changes in respiratory function using a noninvasive and real-time whole-body plethysmography (WBP) system.

哮喘鼠模型中的针灸

A high platform can fix rats without restriction and completely expose the acupoints on the back during acupuncture manipulation. This article describes ...

Retinal Pathophysiological Evaluation in a Rat Model

A posterior segment eye disease like diabetic retinopathy alters the physiology of the retina. Diabetic retinopathy is characterized by a retinal detachment, breakdown of the blood-retinal barrier (BRB), and retinal angiogenesis. An in vivo rat model is a valuable experimental tool to examine the changes in the structure and function of the retina. We propose three different experimental techniques in the rat model to identify morphological changes of retinal cells, retinal vasculature, and compromised BRB. Retinal histology is used to study the morphology of various retinal cells. Also, quantitative measurement is performed by retinal cell count and thickness measurement of different retinal layers. A BRB breakdown assay is used to determine the leakage of extraocular proteins from the plasma to vitreous tissue due to the breakdown of BRB. Fluorescence angiography is used to study angiogenesis and leakage of blood vessels by visualizing retinal vasculature using FITC-dextran dye.

Diabetic retinopathy is one of the leading causes of blindness. Histology, blood-retinal barrier breakdown assay, and fluorescence angiography are valuable techniques to understand the pathophysiology of the retina, which could further enhance the efficient drug screening against diabetic retinopathy.

Methods Article

在大鼠股骨皮质骨中创建箱腔缺损模型