Name: a non-invasive method for generating the cyclic loading-induced intra-articular cartilage lesion model of the rat knee
Uploaded: 2021-07-05T14:30:00
Description: Watch this Scientific Journal Video about A Non-Invasive Method for Generating the Cyclic Loading-Induced Intra-Articular Cartilage Lesion Model of the Rat Knee at JoVE.com

这项研究得到了JSPS KAKENHI资助（编号JP18H03129和JP18K19739）的部分支持。 这项研究还获得了再生康复研究与培训联盟（AR3T）的资助，该联盟得到了Eunice Kennedy Shriver国家儿童健康与人类发展研究所（NICHD），国家神经疾病和中风研究所（NINDS）和美国国立卫生研究院国家生物医学成像和生物工程研究所（NIBIB）的支持，奖励号为P2CHD086843。内容完全由作者负责，并不一定代表美国国立卫生研究院的官方观点。

该方案已获得京都大学动物研究委员会的批准（批准号：Med kyo 17616）。
1.对大鼠膝盖进行 体内 循环加压
<ol><li>诱导实验动物麻醉<ol><li>通过在麻醉盒中吸入5%异氟醚溶液，在12周龄的Wistar大鼠（256.8±8.7g）中诱导麻醉。</li>
		<li>腹膜内注射三种麻醉剂9的混合物，包括美托咪定，咪达唑仑和布托酚，以2mg / kg的大鼠体重，并剃除右膝关节周围的区域。?...

<ol>
	<li>De Souza, R. L., 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=Non-invasive+axial+loading+of+mouse+tibiae+increases+cortical+bone+formation+and+modifies+trabecular+organization:+a+new+model+to+study+cortical+and+cancellous+compartments+in+a+single+loaded+element.">Non-invasive axial loading of mouse tibiae increases cortical bone formation and modifies trabecular organization: a new model to study cortical and cancellous compartments in a single loaded element.</a> Bone. 37 (6), 810-818 (2005).</li><li>Poulet, B., Hamilton, R. W., Shefelbine, S., Pitsillides, A. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterizing+a+novel+and+adjustable+noninvasive+murine+joint+loading+model.">Characterizing a novel and adjustable noninvasive murine joint loading model.</a> Arthritis and Rheumatism. 63 (1), 137-147 (2011).</li><li>Wu, P., 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=Early+response+of+mouse+joint+tissue+to+noninvasive+knee+injury+suggests+treatment+targets.">Early response of mouse joint tissue to noninvasive knee injury suggests treatment targets.</a> Arthritis and Rheumatism. 66 (5), 1256-1265 (2014).</li><li>Poulet, B., 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=Intermittent+applied+mechanical+loading+induces+subchondral+bone+thickening+that+may+be+intensified+locally+by+contiguous+articular+cartilage+lesions.">Intermittent applied mechanical loading induces subchondral bone thickening that may be intensified locally by contiguous articular cartilage lesions.</a> Osteoarthritis Cartilage. 23 (6), 940-948 (2015).</li><li>Ko, F. C., 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=Progressive+cell-mediated+changes+in+articular+cartilage+and+bone+in+mice+are+initiated+by+a+single+session+of+controlled+cyclic+compressive+loading.">Progressive cell-mediated changes in articular cartilage and bone in mice are initiated by a single session of controlled cyclic compressive loading.</a> Journal of Orthopaedic Research. 34 (11), 1941-1949 (2016).</li><li>Adebayo, O. O., 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=Role+of+subchondral+bone+properties+and+changes+in+development+of+load-induced+osteoarthritis+in+mice.">Role of subchondral bone properties and changes in development of load-induced osteoarthritis in mice.</a> Osteoarthritis Cartilage. 25 (12), 2108-2118 (2017).</li><li>Ko, F. C., 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=In+vivo+cyclic+compression+causes+cartilage+degeneration+and+subchondral+bone+changes+in+mouse+tibiae.">In vivo cyclic compression causes cartilage degeneration and subchondral bone changes in mouse tibiae.</a> Arthritis and Rheumatism. 65 (6), 1569-1578 (2013).</li><li>Ji, X., 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+in+vivo+cyclic+compressive+loading+on+the+distribution+of+local+Col2+and+superficial+lubricin+in+rat+knee+cartilage.">Effects of in vivo cyclic compressive loading on the distribution of local Col2 and superficial lubricin in rat knee cartilage.</a> Journal of Orthopaedic Research. 39 (3), 543-552 (2021).</li><li>Kawai, S., Takagi, Y., Kaneko, S., Kurosawa, T. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+three+types+of+mixed+anesthetic+agents+alternate+to+ketamine+in+mice.">Effect of three types of mixed anesthetic agents alternate to ketamine in mice.</a> Experimental Animals. 60 (5), 481-487 (2011).</li><li>Iijima, H., 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=Destabilization+of+the+medial+meniscus+leads+to+subchondral+bone+defects+and+site-specific+cartilage+degeneration+in+an+experimental+rat+model.">Destabilization of the medial meniscus leads to subchondral bone defects and site-specific cartilage degeneration in an experimental rat model.</a> Osteoarthritis Cartilage. 22 (7), 1036-1043 (2014).</li></ol>

目前的协议首次展示了如何在大鼠股骨外侧髁上建立负荷诱导的软骨病变模型，类似于小鼠等较小啮齿动物的关节内损伤模型2。然而，小鼠的负荷方案导致严重的骨赘形成和交叉韧带病变，这对于评估循环压缩的效果并不理想。目前的方案在大鼠中产生了局灶性软骨病变，负荷力要低得多。正确的加载方法设置对于协议至关重要，因为只有适当的应力大小、速度和持续时间才能...

传统上，前交叉韧带 （ACL） 横断或内侧半月板不稳定被认为是研究小动物创伤后骨关节炎 （PTOA） 的最佳方法。近年来，非侵入性循环压缩模型已被用于研究PTOA。该模型最初设计用于研究松质骨对机械负荷的反应1，然后被修改为PTOA研究的非手术动物模型2，3，4，5，6。其基本原理是通过施加周期性的外力来碰撞关节软骨，从而引发一系列炎症反应。然而，该模型仅应用于小鼠，尚未讨论较大动物的适当负荷量。
先前模型的另一个问题是，高容量方案包含太多周期，这导致软骨下骨过度增厚，这是几个样品中不必要的副作用7。因此，开发了一种对大型动物具有适当幅度和较低负荷副作用的循环压缩新方法8。本文的总体目标是描述大鼠无创循环压缩模型的方案，并观察软骨变性的代表性结果。目前的协议将帮助对非侵入性循环压缩模型在大鼠上的应用感兴趣的读者。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Anesthetic Apparatus for Small Animals</td>
			<td>SHINANO MFG CO.,LTD.</td>
			<td>SN-487-0T</td>
			<td></td>
		</tr>
		<tr>
			<td>Autograph AG-X</td>
			<td>Shimadzu Corp</td>
			<td>N.A.</td>
			<td>Precision Universal / Tensile Tester</td>
		</tr>
		<tr>
			<td>Fluoview FV10i microscope</td>
			<td>Olympus Corp</td>
			<td>N.A.</td>
			<td>A fully automated confocal laser-scanning microscope</td>
		</tr>
		<tr>
			<td>ISOFLURANE Inhalation Solution</td>
			<td>Pfizer Japan Inc.</td>
			<td>(01)14987114133400</td>
			<td></td>
		</tr>
		<tr>
			<td>LIVE/DEA Viability/Cytotoxicity Kit</td>
			<td>Thermo Fisher Scientific Japan Inc</td>
			<td>L3224</td>
			<td>A quick and easy two-color assay to determine viability of cells</td>
		</tr>
		<tr>
			<td>TRAPEZIUM X Software</td>
			<td>Shimadzu Corp</td>
			<td>N.A.</td>
			<td>Data processing software for Autograph AG-X</td>
		</tr>
	</tbody>
</table>

a non-invasive method for generating the cyclic loading-induced intra-articular cartilage lesion model of the rat knee

原发性骨关节炎 （OA） 的病理生理学仍不清楚。然而，相对年轻年龄组的 OA 的特定亚分类可能与关节软骨损伤和韧带撕脱史相关。膝关节OA的外科动物模型在了解创伤后OA的发生和进展方面起着重要作用，并有助于开发这种疾病的新疗法。然而，最近已考虑使用非手术模型来避免可能影响干预评估的创伤性炎症。 
本研究建立了 体内 循环压缩负荷诱导的关节内软骨病变大鼠模型，使研究人员能够（1）确定可能导致局灶性软骨损伤的最佳负荷量、速度和持续时间;（2）评估软骨细胞活力的创伤后时空病理变化;（3）评估参与关节压缩负荷的适应和修复机制的破坏性或保护性分子的组织学表达。本报告描述了大鼠模型中这种新型软骨病变的实验方案。

获得了承受20 N循环负荷的样品中软骨细胞活力短期变化（1 h和12 h）的代表性结果。如图 3所示，创伤后12小时死亡软骨细胞（红色荧光）的数量增加。相反，活软骨细胞的数量（绿色荧光）继续减少，一些样本在受影响区域不含活软骨细胞。
组织学显示，经历20 N动态负荷的大鼠膝关节软骨受损，所有样本中股骨外侧髁均确认一个局灶性病变区（<strong cl...

Watch this Scientific Journal Video about A Non-Invasive Method for Generating the Cyclic Loading-Induced Intra-Articular Cartilage Lesion Model of the Rat Knee at JoVE.com

A Non-Invasive Method for Generating the Cyclic Loading-Induced Intra-Articular Cartilage Lesion Model of the Rat Knee

在这里，我们提出了大鼠膝关节的循环负荷诱导的关节内软骨病变模型，该模型由60次超过20 N的循环按压产生，导致大鼠股骨髁软骨受损。

一种生成周期负荷诱导的大鼠膝关节内软骨病变模型的非侵入性方法

The pathophysiology of primary osteoarthritis (OA) remains unclear. However, a specific subclassification of OA in relatively younger age groups is likely ...

该协议表明，有可能在动物中引起软骨损伤，较大的动物无需任何手术干预或对动物膝盖进行结构化破坏。最大的优点是，即使没有经验，操作人员也证明是重复的，这是任何手术方法都难以实现的。首先将麻醉的大鼠安装在固定装置上。将麻醉的大鼠放在底板上，将其右膝盖连接到一小块带有凹槽的树脂上。将右后肢置于髋关节伸展、膝关节屈曲和踝关节伸展位置，膝关节弯曲约 140 度。将老鼠的脚跟放在可移动夹具上的楔形凹槽上。将固定装置移至应力和拉伸测试仪器上，并确保不与称重传感器接触。打开应力和拉伸测试仪器控制软件，然后单击校准按钮。校准后，小心地将框架顶部连接到称重传感器上。缓慢打开可移动主操作面板上的旋钮，直到预紧力达到五牛顿，以保持膝关节与框架紧密连接。在主菜单上，单击创建新方法和系统标签。将测试模式设置为循环，将测试类型设置为压缩。单击传感器标签并选择测试选项卡以检查极限是否在 60 牛顿以内。然后选择描边选项卡并检查限制是否在 500 毫米以内。在测试控制标签下，选择增长原点以 0.3% 或满量程启动主程序。将第一和第三部分控制行程速度设置为每秒1毫米，然后将第二部分的最大测试力设置为20牛顿，将第四部分的最小测试力设置为5牛顿。将整个持续时间设置为峰值负载的 0.5 秒和最小负载的 10 秒。在页面底部的预紧选项卡中，确保选中ON，挠度去除速度设置为每分钟100毫米，最大力为5牛顿。在试样标签中将材料设置为金属。在主菜单中的“选择方法和测试”部分下，选择刚刚构建的方法，然后单击“开始”开始测试。将周期数设置为 60。装载后将大鼠放回笼子里，并在具有足够空间和随意食物的笼子中保持 12-12 小时的浅色黑暗时间表，直到需要分析为止。研究了承受20牛顿循环负荷的样品中软骨细胞活力的短期变化。创伤后12小时死亡软骨细胞的数量增加。相反，活软骨细胞的数量继续减少。经历20牛顿动态负荷的大鼠膝关节软骨受损，所有样本股骨外侧髁均确认1个局灶性病变区。在八周的观察期内，病变大小没有逐渐增加。在受影响的区域可以观察到对应于病变界面和未受影响的软骨的边界。最重要的是测试力和循环次数的正确设置。通过这种技术，人们可以控制血液关节覆盖的机械负荷，为探索机械负荷如何影响骨关节炎的发展或预防铺平了道路。

Research

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的隔离方法<em&gt;离体</em&gt;肺灌注的大鼠模型：教训制定鼠EVLP课程中学

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科研

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Major hepatic surgery with inflow occlusion, and liver transplantation, necessitate a period of warm ischemia, and a period of reperfusion leading to ...

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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

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Here, based on a clinician&rsquo;s point-of-view, we propose a two-model lower body positive pressure (LBPP) protocol (walking and squatting models) in ...

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Acute Compartment Syndrome is a devastating consequence of musculoskeletal trauma. Currently the diagnosis is based on clinical signs and symptoms, and ...

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The current animal models of osteoarthritis (OA) can be divided into spontaneous models and induced models, both of which aim to simulate the ...

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Parathyroid gland (PG) identification is a critical unmet need in thyroidectomy. The identification of the PG is challenging in thyroid surgery as it is ...

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Cartilage defects of the knee joint caused by trauma are a common sports joint injury in the clinic, and these defects result in joint pain, impaired ...