Name: 作者聚焦：在肌腱病研究中整合机械和生物分析
Uploaded: 2024-03-01T13:00:00
Description: Watch this Scientific Journal Video about Integrating Mechanical and Biological Analysis in Tendinopathy Research at JoVE.com

我们要感谢我们的资金支持：Joe Fallon 研究基金、Louis Meeks 博士 BIDMC 运动医学实习生研究基金和校内赠款 （AN），均来自 BIDMC 骨科，以及美国国立卫生研究院的支持 （2T32AR055885 （PMW））。

这项研究是根据机构动物护理和使用委员会 （IACUC） 在贝丝以色列女执事医疗中心的批准进行的。使用5%异氟醚麻醉动物进行诱导，2.5%异氟醚维持，并注意避免体温过低。
1. 设置测试系统
<ol><li>通过步进电机控制被动踝关节旋转，以施加一致的旋转和扭矩。使用微控制器控制步进电机。使用 3D 位置和方向系统的输入来标记旋转度数。使用扭矩传感器的输出提?...

循环机械负荷诱导大鼠跟腱肌腱病

<ol>
	<li>Kaux, J. F., Forthomme, B., Goff, C. L., Crielaard, J. M., Croisier, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+opinions+on+tendinopathy.">Current opinions on tendinopathy.</a> J Sports Sci Med. 10 (2), 238-253 (2011).</li><li>Maffulli, N., Longo, U. G., Kadakia, A., Spiezia, F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Achilles+tendinopathy.">Achilles tendinopathy.</a> Foot Ankle Surg. 26 (3), 240-249 (2020).</li><li>Teunis, T., Lubberts, B., Reilly, B. T., Ring, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+systematic+review+and+pooled+analysis+of+the+prevalence+of+rotator+cuff+disease+with+increasing+age.">A systematic review and pooled analysis of the prevalence of rotator cuff disease with increasing age.</a> J Shoulder Elbow Surg. 23 (12), 1913-1921 (2014).</li><li>von Rickenbach, K. J., Borgstrom, H., Tenforde, A., Borg-Stein, J., McInnis, K. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Achilles+tendinopathy:+evaluation,+rehabilitation,+and+prevention.">Achilles tendinopathy: evaluation, rehabilitation, and prevention.</a> Curr Sports Med Rep. 20 (6), 327-334 (2021).</li><li>Aicale, R., Oliviero, A., Maffulli, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Management+of+Achilles+and+patellar+tendinopathy:+what+we+know,+what+we+can+do.">Management of Achilles and patellar tendinopathy: what we know, what we can do.</a> J Foot Ankle Res. 13 (1), 59 (2020).</li><li>Jarvinen, T. A., 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=Achilles+tendon+injuries.">Achilles tendon injuries.</a> Curr Opin Rheumatol. 13 (2), 150-155 (2001).</li><li>Silbernagel, K. G., Hanlon, S., Sprague, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Current+clinical+concepts:+conservative+management+of+Achilles+tendinopathy.">Current clinical concepts: conservative management of Achilles tendinopathy.</a> J Athl Train. 55 (5), 438-447 (2020).</li><li>Tayfur, A., 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=Are+landing+patterns+in+jumping+athletes+associated+with+patellar+tendinopathy?+A+systematic+review+with+evidence+gap+map+and+meta-analysis.">Are landing patterns in jumping athletes associated with patellar tendinopathy? A systematic review with evidence gap map and meta-analysis.</a> Sports Med. 52 (1), 123-137 (2022).</li><li>Malliaras, P., Cook, J. L., Kent, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reduced+ankle+dorsiflexion+range+may+increase+the+risk+of+patellar+tendon+injury+among+volleyball+players.">Reduced ankle dorsiflexion range may increase the risk of patellar tendon injury among volleyball players.</a> J Sci Med Sport. 9 (4), 304-309 (2006).</li><li>Backman, L. J., Danielson, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Low+range+of+ankle+dorsiflexion+predisposes+for+patellar+tendinopathy+in+junior+elite+basketball+players:+a+1-year+prospective+study.">Low range of ankle dorsiflexion predisposes for patellar tendinopathy in junior elite basketball players: a 1-year prospective study.</a> Am J Sports Med. 39 (12), 2626-2633 (2011).</li><li>Glazebrook, M. A., Wright, J. R., Langman, M., Stanish, W. D., Lee, J. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Histological+analysis+of+achilles+tendons+in+an+overuse+rat+model.">Histological analysis of achilles tendons in an overuse rat model.</a> J Orthop Res. 26 (6), 840-846 (2008).</li><li>Carpenter, J. E., Flanagan, C. L., Thomopoulos, S., Yian, E. H., Soslowsky, L. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effects+of+overuse+combined+with+intrinsic+or+extrinsic+alterations+in+an+animal+model+of+rotator+cuff+tendinosis.">The effects of overuse combined with intrinsic or extrinsic alterations in an animal model of rotator cuff tendinosis.</a> Am J Sports Med. 26 (6), 801-807 (1998).</li><li>Gao, H. G., 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=Increased+serum+and+musculotendinous+fibrogenic+proteins+following+persistent+low-grade+inflammation+in+a+rat+model+of+long-term+upper+extremity+overuse.">Increased serum and musculotendinous fibrogenic proteins following persistent low-grade inflammation in a rat model of long-term upper extremity overuse.</a> PLoS One. 8 (8), e71875 (2013).</li><li>Fung, D., 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+to+tendon+fatigue+damage+accumulation+in+a+novel+in+vivo+model.">Early response to tendon fatigue damage accumulation in a novel in vivo model.</a> J Biomech. 43 (2), 274-279 (2010).</li><li>Ueda, Y., 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=Molecular+changes+to+tendons+after+collagenase-induced+acute+tendon+injury+in+a+senescence-accelerated+mouse+model.">Molecular changes to tendons after collagenase-induced acute tendon injury in a senescence-accelerated mouse model.</a> BMC Musculoskelet Disord. 20 (1), 120 (2019).</li><li>Bloom, E., 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=Overload+in+a+rat+in+vivo+model+of+synergist+ablation+induces+tendon+multi-scale+structural+and+functional+degeneration.">Overload in a rat in vivo model of synergist ablation induces tendon multi-scale structural and functional degeneration.</a> J Biomech Eng. 145 (8), 081003 (2023).</li><li>Williamson, P. M., 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=A+passive+ankle+dorsiflexion+testing+system+to+assess+mechanobiological+and+structural+response+to+cyclic+loading+in+rat+Achilles+tendon.">A passive ankle dorsiflexion testing system to assess mechanobiological and structural response to cyclic loading in rat Achilles tendon.</a> J Biomech. 156, 111664 (2023).</li><li>Oliveira, L. F., Peixinho, C. C., Silva, G. A., Menegaldo, L. L. <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+passive+mechanical+properties+estimation+of+Achilles+tendon+using+ultrasound.">In vivo passive mechanical properties estimation of Achilles tendon using ultrasound.</a> J Biomech. 49 (4), 507-513 (2016).</li></ol>

本研究提出了一种使用被动踝关节背屈系统循环加载大鼠跟腱的方法，用于 体内 过度使用诱导的肌腱病模型。该系统的重要性在于它能够隔离跟腱，在不通过手术进入肌腱的情况下施加可量化的载荷，并测量 体内 肌腱特性。 
2010 年，Fung 等人提出了大鼠髌腱疲劳模型，并建立了定制测试系统14。他们的研究提出了一种通过暴露肌腱直接加载髌...

由于肌腱将肌肉与骨骼连接起来，并在其一生中经历日常重复运动，因此它们极易出现过度使用性损伤，这些损伤是疼痛和限制性的，并导致机械功能受损，影响 30-50% 的人口1。肌腱病是一种慢性疾病，由于重复的疲劳运动和愈合不足到受伤前的水平而被认为是过度使用性损伤。上肢和下肢均常受累，包括肩袖、肘部、跟腱和髌腱2、3、4、5。跟腱病常见于涉及跑步和跳跃的活动中，尤其是参与田径、中长跑、网球和其他球类运动的运动员，影响 7-9% 的跑步者 6,7。跑步和跳跃造成的损伤也可能导致踝关节背屈受限，这是跟腱和髌腱病的危险因素 8,9,10。因此，需要更好地评估和表征肌腱病，本研究可以将其作为过度使用性跟腱损伤的被动踝背屈的大鼠模型。
以前使用小动物模型的工作旨在研究肌腱病的发展和标志物。这些包括跑步机运动、重复伸展、直接肌腱负荷、胶原酶注射、手术和体外研究 11,12,13,14,15,16。尽管文献受益于使用这些肌腱病模型来识别损伤标志物，但局限性包括在不生理相关的关节运动中加载肌腱，例如肌腱的直接加载，不直接测量施加的负载，例如跑步机研究，以及不使用生理过度使用，例如胶原酶注射， 除其他外。为此，本研究旨在开发一种系统，该系统可无创地将量化负荷施加到跟腱上，并应用于过度使用诱导的肌腱病研究，以填补先前开发的肌腱病小动物模型的空白。我们进行了一项试点研究，以证明该系统在一系列加载周期内会引起机械性能的可重复变化。该系统使生理相关的运动和负荷能够诱发过度使用，同时量化和测量在负荷方案中施加在肌腱上和承受的力。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
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			<td>1/32&#39;&#39; Aluminum beads</td>
			<td></td>
			<td></td>
			<td></td>
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		<tr>
			<td>2.5% isoflurane</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>3D digitizing pen</td>
			<td>Polhemus, Vermont, NH, USA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>3D electromagnetic positioning and orientation sensor</td>
			<td>Polhemus, Vermont, NH, USA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>5% isoflurane</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Customized device: 1) Assembly, sensors, 3D printed animal bed and ankle mount actuator</td>
			<td></td>
			<td></td>
			<td>Assembled as described in manuscript</td>
		</tr>
		<tr>
			<td>MATLAB code</td>
			<td>MATLAB, Natick, MA, USA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Microcontroller</td>
			<td>Ivrea, Italy</td>
			<td>Arduino UNO, Rev3&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Nose cone</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Scalpel and scalpel holder</td>
			<td></td>
			<td></td>
			<td>No. 11 scalpel</td>
		</tr>
		<tr>
			<td>Sprague-Dawley rats</td>
			<td>Charles River Laboratories, Wilmington, MA, USA</td>
			<td></td>
			<td>11-13 weeks old</td>
		</tr>
		<tr>
			<td>Stepper driver</td>
			<td>SparkFun Electronics, Niwot, CO 80503</td>
			<td>DM542T</td>
			<td></td>
		</tr>
		<tr>
			<td>Stepper motor</td>
			<td>SparkFun Electronics, Niwot, CO 80503</td>
			<td>23HE30-2804S</td>
			<td></td>
		</tr>
		<tr>
			<td>Straight forceps</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Torque sensor assembly</td>
			<td>Futek Inc., Irvine, CA, USA&#160;</td>
			<td>FSH03985, FSH04473, FSH03927</td>
			<td></td>
		</tr>
		<tr>
			<td>Water heating pad</td>
			<td></td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
</table>

a passive ankle dorsiflexion testing system for an in vivo model of overuse-induced tendinopathy

肌腱病是一种慢性肌腱疾病，会导致疼痛和功能丧失，由肌腱反复超负荷和恢复时间有限引起。该协议描述了一种测试系统，该系统通过被动背屈对大鼠跟腱周期性地施加机械载荷。定制编写的代码包括循环加载前和循环加载后测量，以评估加载协议以及基于反馈控制的循环疲劳加载方案的影响。 
我们使用了 25 只 Sprague-Dawley 大鼠进行这项研究，每组 5 只大鼠接受 500、1,000、2,000、3,600 或 7,200 个循环的疲劳负荷。计算了滞后、峰值应力和加载和卸载模量的循环加载前和循环加载后测量值之间的百分比差异。结果表明，该系统可以根据施加的载荷数量对跟腱造成不同程度的损伤。该系统提供了一种创新方法，可将量化和生理上不同程度的循环载荷应用于跟腱，用于疲劳引起的过度使用肌腱损伤 的体内 模型。

As tendons connect muscle to bone and experience daily repetitive motions throughout their lifetime, they are highly prone to overuse injuries that are painful and limiting and result in impaired mechanical function, affecting 30-50% of the population1. Tendinopathies are chronic conditions considered overuse injuries due to repetitive fatigue motions and inadequate healing to pre-injury levels. Both upper and lower extremities are commonly affected, including the rotator cuff, elbow, Achilles tendon, and patellar tendon2,3,4,5. Achilles tendinopathy is common in activities involving running and jumping, especially athletes involved in track and field, middle- and long-distance running, tennis, and other ball sports, affecting 7-9% of runners6,7. Injuries from running and jumping may also cause limited ankle dorsiflexion, which is a risk factor for Achilles and patellar tendinopathies8,9,10. Thus, there is a need for a better assessment and characterization of tendinopathy, which this study can provide as a rat model of passive ankle dorsiflexion for overuse Achilles tendon injuries.
Previous work using small animal models has been aimed at studying the development and markers of tendinopathy. These include treadmill exercise, repetitive reaching, direct tendon loading, collagenase injections, surgery, and in vitro studies11,12,13,14,15,16. Although the literature has benefited from the identification of damage markers from employing these tendinopathy models, limitations include loading the tendon in non-physiologically relevant joint motions, as in the case of direct loading of the tendon, not directly measuring applied loads, such as for treadmill studies, and not using physiological overuse, as in the case for collagenase injections, among others. To that end, this study aimed to develop a system that noninvasively applies quantified loads to the Achilles tendon with the application for overuse-induced tendinopathy studies to fill the gaps in previously developed small animal models for tendinopathy. We performed a pilot study to demonstrate that the system induces reproducible changes in mechanical properties over a range of loading cycles. This system enables physiologically relevant motion and loading to induce overuse while simultaneously quantifying and measuring the forces applied to and experienced by the tendon during the loading regimen.

作者聚焦：在肌腱病研究中整合机械和生物分析 

用于过度使用诱导的肌腱病 体内 模型的被动踝关节背屈测试系统

Our work addresses the mechano biological factors involved in the initiation and progression of overuse induced tendinopathy. The questions we are trying to answer include what factors of exercise, such as frequency, magnitude, and duration of load affect the tendon's response to injury. Current tools such as mechanical measures, histology, and biological assays to evaluate outcome measures are centered and limited to end stage tissue harvesting.With this model, we can explore in vivo continuous measurements with tools such as ultrasound and MRI In the field, in vivo models load the tendon and physiologically relevant manners, while ex vivo models allow for the measurement of biological responses to injuries. Our protocol combines these two measures for direct measurement of tendon stresses and strains during injuries and the associated biological changes. We've established an adaptive system to induce tendon injuries, which allows for simultaneous exploration of mechanical and biological factors.Having established this model, future directions include assessing different parameters, therapeutic avenues, and rehabilitation measures.

随着应用循环次数的增加， 体内 肌腱机械性能的降低幅度更大。与 3,600 和 7,200 周期组相比，500 周期组的滞后和加载和卸载模量的降低显着降低 （p < 0.05）（图 2）。虽然从500循环组到3,600循环组，每个循环的峰值应力显着降低，但500和7,200循环组之间没有显着减少。3,600 和 7,200 循环组的滞后、峰值应力以及加载和卸载模量的百分比一致下降。苏木精、曙红和 Masson ?...

Watch this Scientific Journal Video about Integrating Mechanical and Biological Analysis in Tendinopathy Research at JoVE.com

该协议提出了一种测试系统，用于在大鼠跟腱中诱导可量化和受控的疲劳损伤，用于过度使用诱导的肌腱病 的体内 模型。该过程包括将大鼠的脚踝固定到关节致动器上，该致动器使用定制编写的 MATLAB 脚本执行被动踝关节背屈。

Tendinopathy is a chronic tendon condition that results in pain and loss of function and is caused by repeated overload of the tendon and limited recovery ...

我们的工作涉及过度使用诱导的肌腱病的发生和发展所涉及的机械生物学因素。我们试图回答的问题包括哪些运动因素，如负荷的频率、大小和持续时间会影响肌腱对损伤的反应。目前用于评估结果测量的机械测量、组织学和生物测定等工具以终末期组织采集为中心，仅限于终末期组织采集。有了这个模型，我们可以用超声和MRI等工具探索体内连续测量 在现场，体内模型加载肌腱和生理相关方式，而离体模型允许测量对损伤的生物反应。我们的方案结合了这两种措施，用于直接测量损伤期间的肌腱应力和应变以及相关的生物学变化。我们已经建立了一个诱发肌腱损伤的适应性系统，可以同时探索机械和生物因素。建立该模型后，未来的方向包括评估不同的参数、治疗途径和康复措施。

a-passive-ankle-dorsiflexion-testing-system-for-an-vivo-model-overuse

Research

JoVE Journal

Bioengineering

A Passive Ankle Dorsiflexion Testing System for an In Vivo Model of Overuse-induced Tendinopathy

Tendinopathy is a chronic tendon condition that results in pain and loss of function and is caused by repeated overload of the tendon and limited recovery time. This protocol describes a testing system that cyclically applies mechanical loads via passive dorsiflexion to the rat Achilles tendon. The custom-written code consists of pre- and post-cyclic loading measurements to assess the effects of the loading protocol along with the feedback control-based cyclic fatigue loading regimen. 
We used 25 Sprague-Dawley rats for this study, with 5 rats per group receiving either 500, 1,000, 2,000, 3,600, or 7,200 cycles of fatigue loads. The percentage differences between the pre- and post-cyclic loading measurements of the hysteresis, peak stress, and loading and unloading moduli were calculated. The results demonstrate that the system can induce varying degrees of damage to the Achilles tendon based on the number of loads applied. This system offers an innovative approach to apply quantified and physiological varying degrees of cyclic loads to the Achilles tendon for an in vivo model of fatigue-induced overuse tendon injury.

This protocol presents a testing system used to induce quantifiable and controlled fatigue injuries in a rat Achilles tendon for an in-vivo model of overuse-induced tendinopathy. The procedure consists of securing the rat's ankle to a joint actuator that performs passive ankle dorsiflexion with a custom-written MATLAB script.