The authors have no acknowledgements.

以下程序均符合杨百翰大学机构审查委员会（IRB）的标准。 1.准备收缩协议注意:下面的协议说明基于所述Biodex Advantage软件。导航软件和如果使用不同的系统操作所述测力计将是不同的。 <ol><li>等速强度测试协议<ol><li>为了使等速协议，开测功机控制软件的计算机上，并选择"协议"选项卡，然后在屏幕顶部的工具栏上的"记录"选项卡。从下拉列表中选择"新建协议"。 </li><li>在"学习型"框中，选择"测试"选项。 </li><li>从下拉菜单中选择以下参数:在"模式"菜单中选择等速。在"联合"菜单中选择膝盖。在下面 ＆＃34;模式"菜单中选择扩展/屈选择两个伸屈运动心收缩选择。"销售代表"下的"同归"下拉菜单。 </li><li>在60°/秒，为离开那里，向收缩既有速度"单方面"标签计划1套3次重复。要做到这一点，输入"1"变成了"＃组"字段和"结束通过销售代表"行中输入"3"。然后在这两个"速度与千寻"和"速度走向"行键入60。 </li><li>使用90°为解剖参考点，并设置繁多的运动限制，以约10°，距离110°朝向（0°=全面推广，135° =完全屈曲）。为了做到这一点，选择在屏幕左侧的"设置光盘"按钮。校准杠杆臂，使得90°是平行于地面（如使用所提供的计算机屏幕上的杠杆臂角度指示器指示）。然后用90°作为所述解剖重ference点，向（0°=全面推广，135°=完全屈曲）的运动限制的范围内设定为约10°，距离110°。 注:此处给出的首选关节角度设置，但这些设置应根据具体情况逐案与固定屈曲畸形或其他限制对象的工作特别是当修改。 </li><li>在名称在屏幕左下方的"描述"字段的协议，并选择保存顶部菜单栏上的按钮。 </li></ol></li><li>等距强度测试协议<ol><li>重复步骤1.1.1开始制作等距协议。 </li><li>从"学习型"中的"测试"选项。 </li><li>从下拉菜单中选择以下参数:在"模式"菜单中选择"等距"。在"联合"菜单中选择"拐点"。在"模式"菜单中选择"扩展/屈"。下的"ContractioN的方向"菜单中选择"离开"，在"同归"菜单中选择"代表"。 </li><li>程序与1组3个等距收缩持续每5秒，在70°的关节角度进行测试。要做到这一点，输入"1"变成了"位置"字段中的"单方面"选项卡。在"结束由众议员"行中的位置＃1列中，键入"3"。在标有"角度"行选择"70"。 </li><li>选择90°为解剖参考点和远限制到70°。 </li><li>在名称在屏幕左下方的"描述"字段的协议，并选择保存顶部菜单栏上的按钮。 </li></ol></li><li>肌肉损坏了离心运动协议<ol><li>为了使离心运动的协议，开测功机控制软件的计算机上，并选择"协议"选项卡。然后选择屏幕顶部的工具栏上的"记录"选项卡。现在，点击"新建协议＆＃34;从下拉列表中。 </li><li>从"学习型"中的"运动"选项。 </li><li>从下拉菜单中选择以下参数:在"模式"菜单中选择"等速"。在"联合"菜单中选择"拐点"。在"模式"菜单中选择"扩展/屈"。在"收缩"菜单中选择"CON / ECC"。 </li><li>在"单方面"选项卡中，节目10套重复10次180°/秒，距离120°/秒速度向与集之间的60秒休息。 <ol><li>输入"10"变成了"＃设置"字段。 </li><li>在第一列（标示为＃1），标记该行中输入"10"，"通过重复次数结束"。然后，标有"速度与千寻"下一行中输入"180"。在标有"速度走向"下一行，键入"120"。在标有"扭矩"的行，键入200到600取决于S的预期强度ubject。 </li><li>通过＃10＃套2重复步骤1.3.4.2。 </li><li> 60类中的"休息时间秒"字段。 </li></ol></li><li>的90°到"解剖参考"字段。 </li><li>选择在屏幕的左侧向（0°=完全伸展，135°=完全屈曲）"设置的ROM"按钮的运动限制到40°，距离110°设置范围。这些设置提供运动，通过它的主题必须提供抵抗测功机的臂的范围内。为安全起见，测功机将停止，如果这个问题停止这种指定的运动范围内提供移动性。 <ol><li>设定的运动范围中，选择在屏幕左侧的"设定ROM"按钮。校准杠杆臂，使得90°是平行于地面（如使用所提供的计算机屏幕上的杠杆臂角度指示器指示）。然后用90°作为解剖参考点，S等运动限制到40°，距离110°朝向范围（0°=全面推广，135°=完全屈曲）。 注:这些关节的角度设置是一个典型的健康主题的建议。这些应该进行修改，以适应能力，从一个人到另一个。 </li></ol></li><li>名称中的"描述"字段的协议，并在顶部工具栏中选择"保存"按钮。 </li></ol></li></ol> 2.测量基线<ol><li>测量酸痛<ol><li>在页面上绘制水平100毫米线做一个视觉模拟评分。 </li><li>上线的左侧表示"否疼痛"，并在该行的右手侧表示"极端疼痛"（ 图1）。此方法通常用于评估延迟性肌肉酸痛1,10,11。 </li><li>指示主题做两体重蹲。执行武器举行straigh都蹲在肩膀和双脚与肩同宽，前面T出。当上腿与地板平行返回到开始位置之前达到适当的深度。 </li><li>让拍摄对象表示他或她在股四头肌觉得通过对视觉模拟评分适当的位置画一条垂直线的股四头肌在深蹲肌肉酸痛的力度。 </li><li>通过测量受试者的印记从视觉模拟评分的无酸痛结束毫米的距离量化酸痛的主题的水平。 </li></ol></li><li>血清肌酸激酶 注意:处理血液之前戴防护手套，并在锐器容器按医院或大学政策使用后立即针的处理。 <ol><li>遵循标准化放血过程从禁食条件下受验者的手臂的肘前区域撤出血液以获得5至6毫升血液12。 </li><li>离心机在15 XG血液样本在3 15分钟 °C到从血细胞中分离血清。 </li><li>用移液管，转移血清样品放入适当标记的主题和时间点离心管。 </li><li>储存在-80℃以备后用的样品。 </li><li>测定血清样品用于使用根据制造商的协议市售的试剂盒的肌酸激酶活性。可替代地，血清样品可以运输到当地的医疗诊断实验室进行分析。 </li></ol></li><li>主题准备<ol><li>前主体的访问四十八小时，指示他们要避免剧烈的或不熟悉的运动，非甾体类抗炎药和酒精。 </li><li>当拍摄对象到达时，收集体重，身高，性别和之前安装的腿主导地位的信息。腿霸主地位通过询问用哪条腿，他们会选择踢球的主题被确定。 </li><li>有subjec在50进行的强度试验上前t升温到150瓦用于在自行车测功计10分钟。 </li><li>热身后，问这个问题的测力计的椅子上坐下。调整倾斜，高度，深度，斜倚，并且座椅和轴臂的旋转，以适应受试者。使用股骨外侧髁作为参考并对齐膝的旋转轴与该测力计轴臂的。调整轴臂长度，使得它仅仅是接近内踝，并用带固定。主题应能舒适地跖屈全运动范围。 </li><li>离开座位的前边缘和腿部的后面之间有足够的空间（25毫米），以允许膝关节的最大屈曲，而不会挤压或不适。 </li><li>受稳定与肩，髋，踝关节和大腿绑带。以分离膝伸肌和避免辅助肌受累，至关重要的是，该主题被牢固地稳定。 </li><li>有主题演出3-5实践分机之前开始收缩协议可能存在的，并根据需要调整膝关节/屈曲机动，运动的监控范围和不适的迹象。 </li><li>现在，测功机的可调部件正确安装到这个问题，录制在以后的访问重复设置的座椅位置设置和轴手臂的位置。 注:重要的是，受试者是安全和舒适。测功机配备有用于在紧急的情况下，主体和操作都紧急停止按钮。 </li></ol></li><li>力测量<ol><li>这个问题是在座位上正确定向和等距强度试验加载协议之后，指导如何执行测试的主题。 <ol><li>告诉他或她将被收缩对杠杆臂，将留固定的主题。请告诉主体按照计算机屏幕做3宫缩持续5秒EAC的提示H。 </li><li>告诉主体维护每个收缩的最大努力。告诉主体在宫缩时呼吸。让拍摄对象在测试过程中交叉于胸前交叉双臂。 </li><li>启动等距协议，允许受实践与次最大努力的考验。这种做法审判将在熟悉受制于测试，以及提供特定的测试热身。允许受试者练试验2至3倍。 </li></ol></li><li>一旦受试者是准备好了，开始测试和口头鼓励受试者以提供每个收缩的最大努力。 </li><li>记录扭矩，功率，和工作。该软件还计算变异系数（基于性能重复性试验有效性的统计表示，较低的值表示较高的再现性）。如果变异系数超过15％，重复测试。重复测试高达4倍。如果主体是无法生成系数Ø˚F方差后4尝试小于15％使用从产生方差值的最低系数的集合的数据。 </li><li>允许受到两到三分钟休息，然后加载等速协议来获取等速强度数据。 </li><li>这个问题被固定在座位上后，指导如何进行等速测试。 <ol><li>告诉他们会做对杠杆臂，将在一个固定的速度动议三项屈伸收缩的主题。给该主题的目标是在整个每个收缩运动的整个范围内为有力地收缩。 </li><li>告诉他们，当他们到达运动范围的结束杠杆臂将停止。让拍摄对象在测试过程中交叉于胸前交叉双臂。提醒受不宫缩时屏住呼吸 注意:当从收缩的屈曲部件收集的数据可能并不感兴趣的研究者，主体谁为屈伸演习提供最大的努力学分倾向于延长演习中，以便获得更好的力量的一致性。 </li></ol></li><li>开始测试和口头鼓励受提供最大的努力。 </li><li>记录扭矩，功率，和工作。该软件还计算变异系数（基于性能重复性试验有效性的统计表示，较低的值表示较高的再现性）。如果变异系数超过15％，重复测试。 <ol><li>重复测试高达4倍。如果主体是无法产生偏差小于15％的系数后4次尝试，从产生偏差值系数最低的一组使用的数据。 </li></ol></li></ol></li></ol> 3.感应损坏<ol><li>一到第2部分的基线测量三天后，让主体回归到实验室进行的损伤诱导部协议</li><li>对一个周期测功计在50受试者热身10分钟，以150瓦</li><li>经过热身，东方和使用记录在2.3.7步调整设置固定在测功机的主题，加载肌肉损伤离心运动的协议和指导如何正确地执行演习的主题。 <ol><li>告诉它们将收缩对杠杆臂，以启动轴臂的移动对每个收缩的主题。告诉主体，由于轴臂以固定利率向他们移动，他们的目标是尽可能有力地收缩反对。 </li><li>允许受试者把握位于座椅的任一侧的手柄。 </li><li>告诉主体在宫缩时呼吸;在偏心阶段呼气。 </li></ol></li><li>当拍摄对象准备就绪后，按下屏幕上的启动按钮启动第一套收缩。 </li><li>口头鼓励的主题，以保持最大的EF堡垒每个收缩。 </li><li>按照指导的主题，通过10套10重复与集之间有1分钟的休息时间在屏幕上的提示进行操作。 </li><li>一旦10套完成允许受试者1至5分钟，以其余重复协议之前。 注:套偏心协议下可用的最大数量是10然而这可以根据所使用的测力计和软件变化。其余部分提供时间如有必要，运营商重新加载协议。 </li><li>重复该过程两次，总共300主要离心收缩。 </li></ol> 4.肌肉损伤评估<ol><li>重复两等速和作为有害运动后立即在后24，48，72和96小时在2.4中描述等角强度试验。 </li><li>重复测量疼痛于24，48，72部分2.1所述，破坏运动后96小时。一定要使用新的视觉模拟评分对每个评估使主体不能看到他们以前的响应。 </li><li>重复抽血破坏运动后48和120小时为肌酸激酶检测部分2.2所描述的，或由实验方案规定的时间点。 </li><li>绘制关于对时间的曲线图上的x轴的y轴的肌酸激酶，强度和酸痛数据。 </li><li>观察这些变量是如何偏离，并在运动后的日子里向基线值回归，说明损伤修复，分别为。 </li></ol>

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J., Bendahan, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Comparative+MRI+analysis+of+T2+changes+associated+with+single+and+repeated+bouts+of+downhill+running+leading+to+eccentric-induced+muscle+damage.">Comparative MRI analysis of T2 changes associated with single and repeated bouts of downhill running leading to eccentric-induced muscle damage.</a> J. Appl. Physiol. 105 (1), 299-307 (2008).</li><li>Crameri, R. 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=Myofibre+damage+in+human+skeletal+muscle:+effects+of+electrical+stimulation+versus+voluntary+contraction.">Myofibre damage in human skeletal muscle: effects of electrical stimulation versus voluntary contraction.</a> J. Physiol. 583 (Pt 1), 365-380 (2007).</li><li>Yu, J. G., Malm, C., Thornell, L. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Eccentric+contractions+leading+to+DOMS+do+not+cause+loss+of+desmin+nor+fibre+necrosis+in+human+muscle.">Eccentric contractions leading to DOMS do not cause loss of desmin nor fibre necrosis in human muscle.</a> Histochem. Cell Biol. 118 (1), 29-34 (2002).</li><li>Jamurtas, A. Z., 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=Comparison+between+leg+and+arm+eccentric+exercises+of+the+same+relative+intensity+on+indices+of+muscle+damage.">Comparison between leg and arm eccentric exercises of the same relative intensity on indices of muscle damage.</a> Eur. J. Appl. Physiol. 95 (2-3), 179-185 (2005).</li></ol>

The authors have nothing to disclose.

有几个步骤是获得本协议的预期效果的关键。首先，受试者必须充分熟悉到收缩的协议，特别是测力。要确保拍摄理解他们应该做的，给他们练习之前，数据采集强度测试的机会到底是什么。谁不熟悉充分使用这些程序的主体会显示损伤诱导下过天的学习曲线。这可以是一个混杂变量呈现强度测量无效。 图5从谁可能没有被正确熟悉的个人资料显示。本课题显示，由于越来越多的实力...

The overall goal of this procedure is to induce exertional damage to the quadriceps femoris muscles using voluntary lengthening (eccentric) contractions in human subjects.
Contraction-induced skeletal muscle damage is a common consequence of exercise that is marked by delayed onset muscle soreness1, transient strength loss, and elevated muscle-specific enzymes in the blood2. Exertional muscle damage is most pronounced following exercise to which the subject is unaccustomed, particularly when eccentric contractions are involved3. Exertional muscle damage is typically benign. Soreness subsides, and both serum proteins and strength typically return to pre-damage levels within a few days to weeks after the damaging insult. In extreme cases, exertional muscle damage can lead to a life-threatening syndrome know as rhabdomyolysis. However, exertional muscle damage is usually insufficient to cause clinical rhabdomyolysis in healthy individuals4 in the absence of compounding factors including heat stress, dehydration5, infection6 or rare genetic predispositions7. 
Contraction-induced muscle damage is typically less severe than toxin-induced or freezing-induced injury, methods often used in rodent studies8,9. Yet, contraction-induced injury provides a useful method to study the muscle damage response with notable advantages. First, it is a safe and ethical method for use with human subjects1-3. Thus, interspecies translation of the results is not needed as data can be obtained directly from human subjects. Moreover, translating data obtained from rodent studies is very difficult given that the severity of injury seen in the rodent injury models exceeds the level of damage that would be ethical to induce in human subjects. Second, contraction-induced damage is commonly experienced and a natural process of exercise. Therefore, this mode of damage induction is useful for studying muscle damage in the context of exercise, adaptation to exercise as well as overt muscle injury. Here we describe a safe and reliable method to induce and evaluate skeletal muscle damage using lengthening contractions in humans.

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			<td style="width:167px;">Biodex Medical Systems</td>
			<td style="width:119px;">850-000</td>
			<td style="width:423px;">Other models are available and should produce similar results</td>
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		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Creatine Kinase kit</td>
			<td style="width:167px;">Sigma-Aldrich&#160;</td>
			<td>MAK116</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Serum Vacutainers</td>
			<td style="width:167px;">BD Bioscience</td>
			<td style="width:119px;">367812</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">Winged safety push button blood collection set</td>
			<td style="width:167px;">BD Bioscience</td>
			<td style="width:119px;">367338</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Cryogenic vials</td>
			<td style="width:167px;">Sigma-Aldrich&#160;</td>
			<td style="width:119px;">V5007</td>
			<td>We use the 2mL vials to store serum aliquots</td>
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induction and assessment of exertional skeletal muscle damage in humans

通过自愿偏心（延长）收缩收缩引起的肌肉损伤提供了研究人类肌肉的适应和恢复的优秀典范。在这里我们讨论的离心运动协议的设计诱导四头肌肌肉，在力量，疼痛，和血浆肌酸激酶水平的变化标志着损害。因为它是在人参加演出，并消除了结果的种间翻译此法简单，伦理和适用广泛。科目的速度进行伸膝肌群的最大300离心收缩 120°/上等速测力秒。损害的程度是衡量使用强度的损失，疼痛，和血浆肌酸激酶水平的相对非侵入等速和等距的措施在几天后的锻炼。因此，它的应用，可向特定人群，以试图确定肌肉机制适应和再生。

使用这里介绍的方法，基线酸胀，血清肌酸激酶活性和强度（等距和等速）测量是在7未受过训练的年轻男子。第二天，受试者接受了肌肉损伤上述离心收缩协议。为了提供肌肉损伤的指标，跟进的实力，疼痛和血清肌酸激酶活性的评估进行的。强度运动后以及24，48，72，和96小时后，立即进行测定。疼痛测定24，48，72，和运动后96小时。血清肌酸激酶在48和120小时的运动后...

Watch this Scientific Journal Video about Induction and Assessment of Exertional Skeletal Muscle Damage in Humans at JoVE.com

Induction and Assessment of Exertional Skeletal Muscle Damage in Humans

This article describes a safe and reliable method to induce and quantify exertional skeletal muscle damage in human subjects.

诱导和人类劳力骨骼肌损伤评估

Contraction-induced muscle damage via voluntary eccentric (lengthening) contractions offers an excellent model for studying muscle adaptation and recovery ...

induction-assessment-exertional-skeletal-muscle-damage

Research

JoVE Journal

Medicine

7.4K 次观看.  Brigham Young University. This article describes a safe and reliable method to induce and quantify exertional skeletal muscle damage in human subjects.

视频: Induction and Assessment of Exertional Skeletal Muscle Damage in Humans

Myo-mechanical Analysis of Isolated Skeletal Muscle

To assess the in vivo effects of therapeutic interventions for the treatment of muscle disease 1,2,3, quantitative methods are needed that measure force generation and fatigability in treated muscle. We describe a detailed approach to evaluating myo-mechanical properties in freshly explanted hindlimb muscle from the mouse. We describe the atraumatic harvest of mouse extensor digitorum longus muscle, mounting the muscle in a muscle strip myograph (Model 820MS; Danish Myo Technology), and the measurement of maximal twitch and tetanic tension, contraction time, and half-relaxation time, using a square pulse stimulator (Model S48; Grass Technologies). Using these measurements, we demonstrate the calculation of specific twitch and tetanic tension normalized to muscle cross-sectional area, the twitch-to-tetanic tension ratio, the force-frequency relationship curve and the low frequency fatigue curve 4. This analysis provides a method for quantitative comparison between therapeutic interventions in mouse models of muscle disease 1,2,3,5, as well as comparison of the effects of genetic modification on muscle function 6,7,8,9.

To assess the in vivo effects of therapeutic interventions for muscle disease, methods are needed to quantitate force generation and fatigability in treated muscle. We detail an approach to evaluating myo-mechanical properties in explanted mouse hindlimb muscle. This analysis provides a robust approach to quantitating the effects of genetic modification on muscle function, as well as comparison of therapies in mouse models of muscle disease.

离体骨骼肌肌的力学分析

To assess the in vivo effects of therapeutic interventions for the treatment of muscle disease 1,2,3, quantitative methods are needed that measure force ...

科研

医学

Skeletal Muscle Gender Dimorphism from Proteomics

Gross contraction in skeletal muscle is primarily determined by a relatively small number of contractile proteins, however this tissue is also remarkably adaptable to environmental factors1 such as hypertrophy by resistance exercise and atrophy by disuse. It thereby exhibits remodeling and adaptations to stressors (heat, ischemia, heavy metals, etc.)2,3. Damage can occur to muscle by a muscle exerting force while lengthening, the so-called eccentric contraction4. The contractile proteins can be damaged in such exertions and need to be repaired, degraded and/or resynthesized; these functions are not part of the contractile proteins, but of other much less abundant proteins in the cell. To determine what subset of proteins is involved in the amelioration of this type of damage, a global proteome must be established prior to exercise5 and then followed subsequent to the exercise to determine the differential protein expression and thereby highlight candidate proteins in the adaptations to damage and its repair. Furthermore, most studies of skeletal muscle have been conducted on the male of the species and hence may not be representative of female muscle. 
In this article we present a method for extracting proteins reproducibly from male and female muscles, and separating them by two-dimensional gel electrophoresis followed by high resolution digital imaging6. This provides a protocol for spots (and subsequently identified proteins) that show a statistically significant (p &lt; 0.05) two-fold increase or decrease, appear or disappear from the control state. These are then excised, digested with trypsin and separated by high-pressure liquid chromatography coupled to a mass spectrometer (LC/MS) for protein identification (LC/MS/MS)5. This methodology (Figure 1) can be used on many tissues with little to no modification (liver, brain, heart etc.).

A straight-forward set of methods to isolate and determine the identity of the most abundant proteins expressed in skeletal muscle. About 800 spots are discerned on a two-dimensional gel from 10 mg muscle; this allows for the determination of gender-specific protein expression. These methods will give equivalent results in most tissues.

骨骼肌蛋白质组学性别异形

Gross contraction in skeletal muscle is primarily determined by a relatively small number of contractile proteins, however this tissue is also remarkably ...

Assessment of Right Ventricular Structure and Function in Mouse Model of Pulmonary Artery Constriction by Transthoracic Echocardiography

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, the RV is significantly affected in pulmonary diseases such as pulmonary artery hypertension (PAH). In addition, the RV is remarkably sensitive to cardiac pathologies, including left ventricular (LV) dysfunction, valvular disease or RV infarction4. To understand the role of RV in the pathogenesis of cardiac diseases, a reliable and noninvasive method to access the RV structurally and functionally is essential.
A noninvasive trans-thoracic echocardiography (TTE) based methodology was established and validated for monitoring dynamic changes in RV structure and function in adult mice. To impose RV stress, we employed a surgical model of pulmonary artery constriction (PAC) and measured the RV response over a 7-day period using a high-frequency ultrasound microimaging system. Sham operated mice were used as controls. Images were acquired in lightly anesthetized mice at baseline (before surgery), day 0 (immediately post-surgery), day 3, and day 7 (post-surgery). Data was analyzed offline using software.
Several acoustic windows (B, M, and Color Doppler modes), which can be consistently obtained in mice, allowed for reliable and reproducible measurement of RV structure (including RV wall thickness, end-diastolic&#160;and end-systolic dimensions), and function (fractional area change, fractional shortening, PA peak velocity, and peak pressure gradient) in normal mice and following PAC.
Using this method, the pressure-gradient resulting from PAC was accurately measured in real-time using Color Doppler mode and was comparable to direct pressure measurements performed with a Millar high-fidelity microtip catheter. Taken together, these data demonstrate that RV measurements obtained from various complimentary views using echocardiography are reliable, reproducible and can provide insights regarding RV structure and function. This method will enable a better understanding of the role of RV cardiac dysfunction.

Right ventricle (RV) dysfunction is critical to the pathogenesis of cardiovascular disease, yet limited methodologies are available for its evaluation. Recent advances in ultrasound imaging provide a noninvasive and accurate option for longitudinal RV study. Herein, we detail a step-by-step echocardiographic method using a murine model of RV pressure overload.

右心室结构和功能的肺动脉缩窄经胸超声小鼠模型的评估

Emerging clinical data support the notion that RV dysfunction is critical to the pathogenesis of cardiovascular disease and heart failure1-3. Moreover, ...

Human Skeletal Muscle Biopsy Procedures Using the Modified Bergstr&#246;m Technique

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly effective. This video describes the percutaneous biopsy technique using a modified Bergstr&#246;m needle to obtain skeletal muscle tissue samples from the vastus lateralis of human subjects. The Bergstr&#246;m needle consists of an outer cannula with a small opening (&#8216;window&#8217;) at the side of the tip and an inner trocar with a cutting blade at the distal end. Under local anesthesia and aseptic conditions, the needle is advanced into the skeletal muscle through an incision in the skin, subcutaneous tissue, and fascia. Next, suction is applied to the inner trocar, the outer trocar is pulled back, skeletal muscle tissue is drawn into the window of the outer cannula by the suction, and the inner trocar is rapidly closed, thus cutting or clipping the skeletal muscle tissue sample. The needle is rotated 90&#176;&#160;and another cut is made. This process may be repeated three more times. This multiple cutting technique typically produces a sample of 100-200 mg or more in healthy subjects and can be done immediately before, during, and after a bout of exercise or other intervention. Following post-biopsy dressing of the incision site, subjects typically resume their activities of daily living right away and can fully participate in vigorous physical activity within 48-72 hr. Subjects should avoid heavy resistance exercise for 48 hr to reduce the risk of herniation of the muscle through the incision in the fascia.

The purpose of this video is to present the modified Bergstr&#246;m skeletal muscle biopsy technique on human subjects.

人骨骼肌活检手术使用修改伯格斯坦技术

The percutaneous biopsy technique enables researchers and clinicians to collect skeletal muscle tissue samples. The technique is safe and highly ...

Preparation and Respirometric Assessment of Mitochondria Isolated from Skeletal Muscle Tissue Obtained by Percutaneous Needle Biopsy

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining samples of human Vastus lateralis, isolating mitochondria from minimal amounts of skeletal muscle tissue, and plate based respirometric profiling using an extracellular flux (XF) analyzer. Comparison of respirometric profiles obtained using 1.0, 2.5 and 5.0 &#956;g of mitochondria indicate that 1.0 &#956;g is sufficient to measure respiration and that 5.0 &#956;g provides most consistent results based on comparison of standard errors. Western blot analysis of isolated mitochondria for mitochondrial marker COX IV and non-mitochondrial tissue marker GAPDH indicate that there is limited non-mitochondrial contamination using this protocol. The ability to study mitochondrial respirometry in as little as 20 mg of muscle tissue allows users to utilize individual biopsies for multiple study endpoints in clinical research projects.

Methods for biopsy of Vastus lateralis, preparation of purified mitochondria, and respirometric profiling are described. The use of small muscle volume makes this technique suitable for clinical research applications.

线粒体的制备及透气性评估，从骨骼肌组织通过穿刺活检获得的分离

Respirometric profiling of isolated mitochondria is commonly used to investigate electron transport chain function. We describe a method for obtaining ...

Quantitative Magnetic Resonance Imaging of Skeletal Muscle Disease

Quantitative magnetic resonance imaging (qMRI) describes the development and use of MRI to quantify physical, chemical, and/or biological properties of living systems. Neuromuscular diseases often exhibit a temporally varying, spatially heterogeneous, and multi-faceted pathology. The goal of this protocol is to characterize this pathology using qMRI methods. The MRI acquisition protocol begins with localizer images (used to locate the position of the body and tissue of interest within the MRI system), quality control measurements of relevant magnetic field distributions, and structural imaging for general anatomical characterization. The qMRI portion of the protocol includes measurements of the longitudinal and transverse relaxation time constants (T1 and T2, respectively). Also acquired are diffusion-tensor MRI data, in which water diffusivity is measured and used to infer pathological processes such as edema. Quantitative magnetization transfer imaging is used to characterize the relative tissue content of macromolecular and free water protons. Lastly, fat-water MRI methods are used to characterize fibro-adipose tissue replacement of muscle. In addition to describing the data acquisition and analysis procedures, this paper also discusses the potential problems associated with these methods, the analysis and interpretation of the data, MRI safety, and strategies for artifact reduction and protocol optimization.

Neuromuscular diseases often exhibit a temporally varying, spatially heterogeneous, and multi-faceted pathology. The goal of this protocol is to characterize this pathology using non-invasive magnetic resonance imaging methods.

骨骼肌疾病的定量磁共振成像

Quantitative magnetic resonance imaging (qMRI) describes the development and use of MRI to quantify physical, chemical, and/or biological properties of ...

Ultrasound Based Assessment of Coronary Artery Flow and Coronary Flow Reserve Using the Pressure Overload Model in Mice

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow reserve (CFR) in humans. Reduced CFR is accompanied by marked intramyocardial and pericoronary fibrosis and is used as an indication of the severity of dysfunction. This study explores, step-by-step, the real-time changes measured in the coronary flow velocity, CFR and systolic to diastolic peak velocity (S/D) ratio in the setting of an aortic banding model in mice. By using a Doppler transthoracic imaging technique that yields reproducible and reliable data, the method assesses changes in flow in the septal coronary artery (SCA), for a period of over two weeks in mice, that previously either underwent aortic banding or thoracotomy. 
During imaging, hyperemia in all mice was induced by isoflurane, an anesthetic that increased coronary flow velocity when compared with resting flow. All images were acquired by a single imager. Two ratios, (1) CFR, the ratio between hyperemic and baseline flow velocities, and (2) systolic (S) to diastolic (D) flow were determined, using a proprietary software and by two independent observers. Importantly, the observed changes in coronary flow preceded LV dysfunction as evidenced by normal LV mass and fractional shortening (FS). 
The method was benchmarked against the current gold standard of coronary assessment, histopathology. The latter technique showed clear pathologic changes in the coronary artery in the form of peri-coronary fibrosis that correlated to the flow changes as assessed by echocardiography. 
The study underscores the value of using a non-invasive technique to monitor coronary circulation in mouse hearts. The method minimizes redundant use of research animals and demonstrates that advanced ultrasound-based indices, such as CFR and S/D ratios, can serve as viable diagnostic tools in a variety of investigational protocols including drug studies and the study of genetically modified strains.

Coronary flow reserve (CFR) is useful for assessment of myocardial oxygen demand and evaluation of cardiovascular risk. This study establishes a step-by-step transthoracic Doppler echocardiographic (TTDE) method for longitudinal monitoring of the changes in CFR, as measured from coronary artery in mice, under the experimental pressure overload of aortic banding.

冠脉流量和冠状动脉血流储备用小鼠压力超负荷基于模型的超声评估

Transthoracic Doppler echocardiography (TTDE) is a clinically useful, noninvasive tool for studying coronary artery flow velocity and coronary flow ...

Induction and Assessment of Ischemia-reperfusion Injury in Langendorff-perfused Rat Hearts

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for myocardial infarction and cardiac complications of thoracic surgery. The ability of certain drugs to precondition the myocardium against ischemia reperfusion injury has led to multiple clinical trials, with little success. The isolated heart model allows acute observation of the functional effects of ischemia reperfusion injury in real time, including the effects of various pharmacological interventions administered at any time-point before or within the ischemia-reperfusion injury window. Since brief periods of ischemia can precondition the heart against ischemic injury, in situ aortic cannulation is performed to allow for functional assessment of non-preconditioned myocardium. A saline filled balloon is placed into the left ventricle to allow for real-time measurement of pressure generation. Ischemic injury is simulated by the cessation of perfusion buffer flow, followed by reperfusion. The duration of both ischemia and reperfusion can be modulated to examine biochemical events at any given time-point. Although the Langendorff isolated heart model does not allow for the consideration of systemic events affecting ischemia and reperfusion, it is an excellent model for the examination of acute functional and biochemical events within the window of ischemia reperfusion injury as well as the effect of pharmacological intervention on cardiac pre- and postconditioning. The goal of this protocol is to demonstrate how to perform in situ aortic cannulation and heart excision followed by ischemia/reperfusion injury in the Langendorff model.

The isolated rat heart is an enduring model for ischemia reperfusion injury. Here, we describe the process of harvesting the beating heart from a rat via in situ aortic cannulation, Langendorff perfusion of the heart, simulated ischemia-reperfusion injury, and infarct staining to confirm the extent of ischemic insult.

诱导和缺血再灌注损伤评估中的Langendorff灌注鼠心

The biochemical events surrounding ischemia reperfusion injury in the acute setting are of great importance to furthering novel treatment options for ...

Human Vastus Lateralis Skeletal Muscle Biopsy Using the Weil-Blakesley Conchotome

Percutaneous muscle biopsy using the Weil-Blakesley conchotome is well established in both clinical and research practice. It is a safe, effective and well tolerated technique. The Weil-Blakesley conchotome has a sharp biting tip with a 4 - 6 mm wide hollow. It is inserted through a 5 - 10 mm skin incision and can be maneuvered for controlled tissue penetration. The tip is opened and closed within the tissue and then rotated through 90 -180&#176; to cut the muscle. The amount of muscle obtained following repeated sampling can vary from 20 mg to 290 mg which can be processed for both histology and molecular studies. The wound needs to be kept dry and vigorous physical activity kept to a minimum for approximately 72 hr although normal levels of activity can restart immediately following the procedure. This procedure is safe and effective when close attention is paid to the selection of subjects, full asepsis and post procedure care. &#160;Both right and left vastus lateralis are suitable for biopsy dependent on participant preference.

Human Vastus Lateralis Skeletal Muscle Biopsy Using the Weil-Blakesley Conchotome

This video demonstrates the technique of percutaneous muscle biopsy of the human vastus lateralis using the Weil-Blakesley conchotome.

人的<em&gt;股外侧肌</em&gt;骨骼肌活检使用韦尔 - Blakesley Conchotome

Percutaneous muscle biopsy using the Weil-Blakesley conchotome is well established in both clinical and research practice. It is a safe, effective and ...

Semi-automated Analysis of Mouse Skeletal Muscle Morphology and Fiber-type Composition

For years, distinctions between skeletal muscle fiber types were best visualized by myosin-ATPase staining. More recently, immunohistochemical staining of myosin heavy chain (MyHC) isoforms has emerged as a finer discriminator of fiber-type. Type I, type IIA, type IIX and type IIB fibers can now be identified with precision based on their MyHC profile; however, manual analysis of these data can be slow and down-right tedious. In this regard, rapid, accurate assessment of fiber-type composition and morphology is a very desirable tool. Here, we present a protocol for state-of-the-art immunohistochemical staining of MyHCs in frozen sections obtained from mouse hindlimb muscle in concert with a novel semi-automated algorithm that accelerates analysis of fiber-type and fiber morphology. As expected, the soleus muscle displayed staining for type I and type IIA fibers, but not for type IIX or type IIB fibers. On the other hand, the tibialis anterior muscle was composed predominantly of type IIX and type IIB fibers, a small fraction of type IIA fibers and little or no type I fibers. Several image transformations were used to generate probability maps for the purpose of measuring different aspects of fiber morphology (i.e., cross-sectional area (CSA), maximal and minimal Feret diameter). The values obtained for these parameters were then compared with manually-obtained values. No significant differences were observed between either mode of analysis with regards to CSA, maximal or minimal Feret diameter (all p &#62; 0.05), indicating the accuracy of our method. Thus, our immunostaining analysis protocol may be applied to the investigation of effects on muscle composition in many models of aging and myopathy.

Immunohistochemical staining of myosin heavy chain isoforms has emerged as the state-of-the-art discriminator of skeletal muscle fiber-type (i.e., type I, type IIA, type IIX, type IIB). Here, we present a staining protocol along with a novel semi-automated algorithm that facilitates rapid assessment of fiber-type and fiber morphology.

小鼠骨骼肌形态及纤维型成分的半自动化分析

For years, distinctions between skeletal muscle fiber types were best visualized by myosin-ATPase staining. More recently, immunohistochemical staining of ...