这项工作是支持公共卫生服务津贴（HL086513）从NHLBI私立教育机构，以及来自加州再生医学（RC1 - 00104），公共卫生服务津贴（HL085377）从NHLBI研究所的综合研究资助和礼品波林基金会HSB 资深大律师是一个加州再生医学的桥梁研究所的支持干细胞研究奖（TB1 - 01194）旧金山州立大学。

加州大学旧金山分校的机构动物护理和使用委员会（IACUC）批准该协议。 1。鼠标趾长伸肌 （EDL）肌肉的解剖<ol><li>按照机构准则执行的所有动物的程序。 </li><li>安乐死与200 mg / kg腹腔巴比妥/颈椎脱位，肌肉收获10前的动物。剥离应实行，使肌肉可收获和“安乐死”的15分钟内，安装在张力换能器。 </li><li>夹层纸盒和引脚腿到托盘上排列胴体仰卧。 </li><li>在解剖镜下，开放的皮肤，小心地打开筋膜（图1a），和剥离胫骨从脚踝向上暴露的EDL（图1b） 。使用滴乳酸林格氏液，以保持肌肉湿润和收获期间的缓冲。 </li><li>删除的EDL，保留尽可能多的肌腱上，每年年底，并把到一个Petri菜含有乳酸林格氏液。铁缝合肌腱（ 图1c）。重要的是，肌纤维不能触及，或在清扫不安。 </li></ol> 2。鼠标EDL的安装在肌肉地带肌动描记<ol><li>对于这些研究中，一个组织洗澡是需要的，固定的肌肉，同时允许它在恒温沐浴在生理溶液中连续充氧。洗澡加上一个力传感器的测量肌肉紧张。我们采用从丹麦MYO技术（DMT型号820MS）的综合肌肉地带肌动描记洗澡为此。此外，一个方形脉冲电刺激（草型号S48）和数据采集平台（ADInstruments PowerLab系统的数据采集系统和LabChart软件）引出，记录和分析MYO机械的反应，分别需要。大唐820MS集成到盒盖定位肌肉两侧肌肉地带中旬部分，铂电极。其他myographs可能需要特别注意电极放置。 </li><li>肌动描记浴填写5毫升克雷布斯Henseleit解决方案11。加热至25 ° C。泡泡O 2 / CO 2（95％/ 5％），通过洗澡前15分钟使用。 </li><li>使用缝线的EDL肌动描记夹具之间的延长和夹具之间的EDL肌筋（Fig.1D五 ）安全。要小心不要夹肌肉本身。 </li><li>肌动描记浴保持在25 ° C。 </li></ol> 3。肌的力学分析 A.抽搐紧张<ol><li>在洗澡时设置的初始长度，以便有没有肌肉松弛。 </li><li>确定通过调整电压，以获得最大的抽搐紧张的最大刺激（持续时间为0.5毫秒），然后设定在20％以上最大的刺激（实现超最大刺激）。在我们的研究，超最大的刺激，通常在40伏的输出实现。 </li><li>验证从使用示波器的刺激输出。 </li><li>确定最佳长度逐渐伸展的肌肉，直到没有抽搐的紧张局势进一步增加。 </li><li>允许3分钟的肌肉平衡。 </li><li>交付平方米超最大的刺激在最佳使用电子刺激S44草的长度（0.5毫秒），并记录输出。 </li><li>记录：抽搐张力曲线（P T随时间; 图2A）。 </li></ol> B.破伤风紧张<ol><li>让肌肉休息3分钟。 </li><li>应用最佳使用电子刺激S44草的长度在150赫兹的超最大刺激的列车为300毫秒，并记录输出。 </li><li>记录：破伤风张力曲线（P O随时间; 图2B）。 </li></ol> C.部队频率<ol><li>让肌肉休息3分钟。 </li><li>部队频率：适用于30，60，100，140和160赫兹的超最大刺激的列车，彼此之间的刺激（图3）休息3分钟。此外，火车可应用于15，25，35，45，55，65，75，100，140和160赫兹在较低的频率更高的分辨率，其中力变化显着。 </li><li>剧情：力频率的关系（％最大力量与刺激频率）。 </li></ol> D.疲劳<ol><li>应用短破伤风的火车：300毫秒（或调整，以产生50％的峰值力的频率），每10分钟3秒60赫兹。 10分钟，强直力下降的一个〜15％的初始值（ 图4）高原水平。 </li><li>剧情：低频疲劳（％最大力量与时间）。 </li></ol> E.在协议结束额外的数据收集<ol><li>在卸载肌肉的肌动描记之前，设置MUSC乐在最佳长度确定的步骤III.A.4，使用显微镜或一个眼，用游标卡尺测量其直径。计算截面积（2微米）。 </li><li>通过去除缝线，体重肌肉测量肌肉质量（毫克）。 </li><li>称重鼠标评估体重（GM）。 </li></ol> 4。计算<ol><li>肌肉：体重比= 肌肉质量/体质量</li><li>抽搐紧张，P T（MN）= 在抽搐产生的最大张力</li><li>具体抽搐张力（N /厘米2）= 抽搐张力（MN）/截面积（2微米）× 10 5 / MN•微米2 /厘米2 </li><li>到峰值张力时间（毫秒）= 从收缩发病时间最大张力</li><li>半弛豫时间（毫秒）= 从峰值紧张到峰值张力的50％ </li><li>强直紧张，P O（MN）= 在破伤风产生的最大张力</li><li>具体强直张力（N /厘米2）= 强直张力（MN）/截面积（2微米）× 10 5 / MN•微米2 /厘米2 </li><li>破伤风上升最大速率（N / S）= 期间，破伤风，即紧张，强直张力曲线的最大斜率（或DP的O / DT）上升的紧张局势加剧的最高税率</li><li>半松弛强直张力（MS）= 时间从停止刺激，在紧张刺激终止的50％ </li><li>抽搐紧张强直张力比，P - T / P O = 最大抽搐张力/最大强直张力</li><li>疲劳指数= 经过两分钟的低频疲劳等长张力最大的张力比</li></ol> 5。代表性的成果<img src="/files/ftp_upload/2582/2582fig1.jpg" alt="图1" /> 图1。剖析的EDL肌肉。 EDL的一个 muscles.TA，后肢胫前，B，EDL（趾长伸肌）肌曝光，曝光C，EDL的肌腱缝合的附件，D，张力传感器浴（从侧面看），E，安装在洗澡（从上面）。在肌肉未完全沉浸在缓冲区，仅供;在实践中，应完全沉浸肌肉，防止干燥。 <img src="/files/ftp_upload/2582/2582fig2.jpg" alt="图2" /> 图2。张力曲线的例子 。 的抽搐紧张说明抽搐最大张力（P T），收缩时间（CT）和半弛豫时间（HRT）的曲线，例如酒吧，1秒，B，强直张力曲线显示最大强直张力（P O）的例子，半松弛强直张力（HRTT）。酒吧，1S。 <img src="/files/ftp_upload/2582/2582fig3.jpg" alt="图3" /> 图3。力频率的关系分析的例子 。 ，增量刺激频率产生的紧张局势，B，例如，在30MHz的脉冲序列。酒吧，80毫秒，C，例如，在140MHz的脉冲序列。酒吧，80毫秒，D，从A 中显示的数据派生的力-频率曲线的例子。力 - 频率曲线的形状是肌肉力量的特点，可以比较来自不同动物的肌肉之间。 <img src="/files/ftp_upload/2582/2582fig4.jpg" alt="图4" /> 图4。低频疲劳分析的例子 。超过stimulation.Examples在指定的时间点（B，C，D）的低频脉冲串期间产生了递减的紧张局势图所示，E，从 A中显示的数据产生的低频疲劳曲线的范例。低频疲劳曲线的形状是肌肉力量的特点，可以比较来自不同动物的肌肉之间。

<ol>
	<li>Harcourt, L. J., Schertzer, J. D., Ryall, J. G., Lynch, G. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Low+dose+formoterol+administration+improves+muscle+function+in+dystrophic+mdx+mice+without+increasing+fatigue.">Low dose formoterol administration improves muscle function in dystrophic mdx mice without increasing fatigue.</a> Neuromuscul Disord. 17, 47-55 (2007).</li><li>Messina, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=VEGF+overexpression+via+adeno-associated+virus+gene+transfer+promotes+skeletal+muscle+regeneration+and+enhances+muscle+function+in+mdx+mice.">VEGF overexpression via adeno-associated virus gene transfer promotes skeletal muscle regeneration and enhances muscle function in mdx mice.</a> FASEB J. 21, 3737-3746 (2007).</li><li>Danieli-Betto, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Sphingosine+1-phosphate+protects+mouse+extensor+digitorum+longus+skeletal+muscle+during+fatigue.">Sphingosine 1-phosphate protects mouse extensor digitorum longus skeletal muscle during fatigue.</a> Am J Physiol Cell Physiol. 288, C1367-C1373 (2005).</li><li>MacIntosh, B. R., Willis, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Force-frequency+relationship+and+potentiation+in+mammalian+skeletal+muscle.">Force-frequency relationship and potentiation in mammalian skeletal muscle.</a> J Appl Physiol. 88, 2088-2096 (2000).</li><li>Hayes, A., Williams, D. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contractile+properties+of+clenbuterol-treated+mdx+muscle+are+enhanced+by+low-intensity+swimming.">Contractile properties of clenbuterol-treated mdx muscle are enhanced by low-intensity swimming.</a> J Appl Physiol. 82, 435-439 (1997).</li><li>Coulton, G. R., Curtin, N. A., Morgan, J. E., Partridge, T. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+mdx+mouse+skeletal+muscle+myopathy:+II.+Contractile+properties.">The mdx mouse skeletal muscle myopathy: II. Contractile properties.</a> Neuropathol Appl Neurobiol. 14, 299-314 (1988).</li><li>Danieli-Betto, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deficiency+of+alpha-sarcoglycan+differently+affects+fast-+and+slow-twitch+skeletal+muscles.">Deficiency of alpha-sarcoglycan differently affects fast- and slow-twitch skeletal muscles.</a> Am J Physiol Regul Integr Comp Physiol. 289, R1328-R1337 (2005).</li><li>Chan, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+gene+for+speed:+contractile+properties+of+isolated+whole+EDL+muscle+from+an+alpha-actinin-3+knockout+mouse.">A gene for speed: contractile properties of isolated whole EDL muscle from an alpha-actinin-3 knockout mouse.</a> Am J Physiol Cell Physiol. 295, C897-C904 (2008).</li><li>Personius, K. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Grip+force,+EDL+contractile+properties,+and+voluntary+wheel+running+after+postdevelopmental+myostatin+depletion+in+mice.">Grip force, EDL contractile properties, and voluntary wheel running after postdevelopmental myostatin depletion in mice.</a> J Appl. , (2010).</li><li>Donovan, J., Brown, P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Euthanasia.">Euthanasia.</a> Curr Protoc Immunol. Chap 1, (2006).</li><li>Beekley, M. D., Wetzel, P., Kubis, P., Gros, G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Contractile+properties+of+skeletal+muscle+fibre+bundles+from+mice+deficient+in+carbonic+anhydrase+II.">Contractile properties of skeletal muscle fibre bundles from mice deficient in carbonic anhydrase II.</a> Pflugers Arch. 452, 453-463 (2006).</li></ol>

我们描述了一个详细的评估，在explanted从小鼠后肢肌肉的肌机械性能的方法。 EDL的，而更难以解剖，因为其背后的胫前肌后的位置，比胫前，因为其突出的腱附件的脚踝和膝盖关节更容易评估。这些筋便于安装在肌肉地带肌动描记。相反，更容易访问的胫前在膝关节广泛，几乎atendinous附件，异常艰难的两个剖析不影响肌肉的前提下，并安装在肌动描记安全。我们还指出，迅速安装在生理缓冲液和温度中的含氧浴肌肉，保持肌肉的力学性能是必不可少的。我们发现，我们可以长达30分钟重复这种分析没有显着变化，在这种条件下肌肉反应。最后，它是必不可少的，肌纤维不被感动在解剖和安装程序，可以有肌肉功能上的不利影响，并导致低估肌机械力。遵循这些程序，这种分析提供了可靠的定量方法来评估的肌肉功能的基因改造6,7,8,9影响，以及在肌肉疾病的小鼠模型1,2,3,5治疗性干预之间的比较。

解决方案和媒体 乳酸林格氏液 <ul><li> 100毫米氯化钠</li><li> 30毫米CH3CH（OH）COONa（乳酸钠） </li><li> 4 mM的氯化钾</li><li> 1毫米氯化钙2 2H 2 O（二水氯化钙） <ul><li>调节pH值至6.75 </li></ul></li></ul> 克雷布斯Henseleit解决方案<ul><li> 118毫米氯化钠</li><li> 4.7 mM的氯化钾</li><li> 1.25毫米氯化钙2 </li><li> 1.2毫米氯化镁2 </li><li> 1.2毫米的KH 2 PO 4 </li><li> 25毫米碳酸氢钠3 </li><li> 11毫米葡萄糖<ul><li>与O 2 / CO 2（95％/ 5％）气体平衡，调整pH值到7.2-7.4 </li></ul></li></ul> 戊巴比妥 <ul><li> 5 mg / mL的工作液在无菌水</li></ul>

myo-mechanical analysis of isolated skeletal muscle

在体内治疗肌肉疾病1,2,3的治疗干预的影响评估，定量分析方法，测量力的产生和治疗肌肉疲劳性。我们描述了一个新鲜explanted从小鼠后肢肌肉中的肌机械性能评估的详细方法。我们描述了小鼠趾长伸肌肌肉的非创伤性的收获，安装的肌肉，在肌肉地带肌动描记器（型号820MS;丹麦MYO技术）和最大抽搐，强直张力，收缩时间，半弛豫时间的测量，使用方波脉冲刺激器（型号S48;草技术）。使用这些测量，我们展示了具体的抽搐和规范化肌肉的横截面积，抽搐，强直紧张的比例，力-频率关系曲线和低频疲劳曲线 4强直张力的计算。这种分析方法提供了一个为1,2,3,5肌肉疾病的小鼠模型，以及治疗干预的肌肉功能的基因改造6,7,8,9影响比较之间的定量比较。

Watch this Scientific Journal Video about Myo-mechanical Analysis of Isolated Skeletal Muscle at JoVE.com

Myo-mechanical Analysis of Isolated Skeletal Muscle

为了评估<em&gt;在体内</em&gt;肌肉疾病的治疗性干预的影响，方法是需要定量力量的产生和治疗肌肉疲劳性。我们详细评估explanted小鼠后肢肌肉的肌力学性能的方法。这种分析提供了一个强大的方法，以定量的影响肌肉功能的基因改造，以及在治疗肌肉疾病的小鼠模型的比较。

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 ...

myo-mechanical-analysis-of-isolated-skeletal-muscle

Research

JoVE Journal

Medicine

26.6K 次观看.  University of California San Francisco. 为了评估在体内肌肉疾病的治疗性干预的影响，方法是需要定量力量的产生和治疗肌肉疲劳性。我们详细评估explanted小鼠后肢肌肉的肌力学性能的方法。这种分析提供了一个强大的方法，以定量的影响肌肉功能的基因改造，以及在治疗肌肉疾病的小鼠模型的比较。

视频: Myo-mechanical Analysis of Isolated Skeletal Muscle

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 ...

科研

医学

Tibial Nerve Transection - A Standardized Model for Denervation-induced Skeletal Muscle Atrophy in Mice

The tibial nerve transection model is a well-tolerated, validated, and reproducible model of denervation-induced skeletal muscle atrophy in rodents. Although originally developed and used extensively in the rat due to its larger size, the tibial nerve in mice is big enough that it can be easily manipulated with either crush or transection, leaving the peroneal and sural nerve branches of the sciatic nerve intact and thereby preserving their target muscles. Thus, this model offers the advantages of inducing less morbidity and impediment of ambulation than the sciatic nerve transection model and also allows investigators to study the physiologic, cellular and molecular biologic mechanisms regulating the process of muscle atrophy in genetically engineered mice. The tibial nerve supplies the gastrocnemius, soleus and plantaris muscles, so its transection permits the study of denervated skeletal muscle composed of fast twitch type II fibers and/or slow twitch type I fibers. Here we demonstrate the tibial nerve transection model in the C57Black6 mouse. We assess the atrophy of the gastrocnemius muscle, as a representative muscle, at 1, 2, and 4 weeks post-denervation by measuring muscle weights and fiber type specific cross-sectional area on paraffin-embedded histologic sections immunostained for fast twitch myosin.

The tibial nerve transection model is a well-tolerated, validated, and reproducible model of skeletal muscle atrophy. The model surgical protocol is described and demonstrated in C57Black6 mice. &#160;

胫骨神经切断术 - 一个标准化的模型失神经诱导小鼠骨骼肌

The tibial nerve transection model is a well-tolerated, validated, and reproducible model of denervation-induced skeletal muscle atrophy in rodents. ...

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.

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

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 ...

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 ...

Induction and Assessment of Exertional Skeletal Muscle Damage in Humans

Contraction-induced muscle damage via voluntary eccentric (lengthening) contractions offers an excellent model for studying muscle adaptation and recovery in humans. Herein we discuss the design of an eccentric exercise protocol to induce damage in the quadriceps muscles, marked by changes in strength, soreness, and plasma creatine kinase levels. This method is simple, ethical, and widely applicable since it is performed in human participants and eliminates the interspecies translation of the results. Subjects perform 300 maximal eccentric contractions of the knee extensor muscles at a speed of 120&#176;/sec on an isokinetic dynamometer. The extent of the damage is measurable using relatively non-invasive isokinetic and isometric measures of strength loss, soreness, and plasma creatine kinase levels over several days following the exercise. Therefore, its application can be directed to specific populations in an attempt to identify mechanisms for muscle adaptation and regeneration.

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 ...

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 ...

Skeletal Muscle Neurovascular Coupling, Oxidative Capacity, and Microvascular Function with 'One Stop Shop' Near-infrared Spectroscopy

Exercise represents a major hemodynamic stress that demands a highly coordinated neurovascular response in order to match oxygen delivery to metabolic demand. Reactive hyperemia (in response to a brief period of tissue ischemia) is an independent predictor of cardiovascular events and provides important insight into vascular health and vasodilatory capacity. Skeletal muscle oxidative capacity is equally important in health and disease, as it determines the energy supply for myocellular processes. Here, we describe a simple, non-invasive approach using near-infrared spectroscopy to assess each of these major clinical endpoints (reactive hyperemia, neurovascular coupling, and muscle oxidative capacity) during a single clinic or laboratory visit. Unlike Doppler ultrasound, magnetic resonance images/spectroscopy, or invasive catheter-based flow measurements or muscle biopsies, our approach is less operator-dependent, low-cost, and completely non-invasive. Representative data from our lab taken together with summary data from previously published literature illustrate the utility of each of these end-points. Once this technique is mastered, application to clinical populations will provide important mechanistic insight into exercise intolerance and cardiovascular dysfunction.

Skeletal Muscle Neurovascular Coupling, Oxidative Capacity, and Microvascular Function with &#039;One Stop Shop&#039; Near-infrared Spectroscopy

Here, we describe a simple, non-invasive approach using near-infrared spectroscopy to assess reactive hyperemia, neurovascular coupling and skeletal muscle oxidative capacity in a single clinic or laboratory visit.

"一站式" 近红外光谱对骨骼肌神经血管的耦合、氧化能力及微血管功能的作用

Exercise represents a major hemodynamic stress that demands a highly coordinated neurovascular response in order to match oxygen delivery to metabolic ...

Obtaining Quality Extended Field-of-View Ultrasound Images of Skeletal Muscle to Measure Muscle Fascicle Length

Muscle fascicle length, which is commonly measured in vivo using traditional ultrasound, is an important parameter defining a muscle&#8217;s force generating capacity. However, over 90% of all upper limb muscles and 85% of all lower limb muscles have optimal fascicle lengths longer than the field-of-view of common traditional ultrasound (T-US) probes. A newer, less frequently adopted method called extended field-of-view ultrasound (EFOV-US) can enable direct measurement of fascicles longer than the field-of-view of a single T-US image. This method, which automatically fits together a sequence of T-US images from a dynamic scan, has been demonstrated to be valid and reliable for obtaining muscle fascicle lengths in vivo. Despite the numerous skeletal muscles with long fascicles and the validity of the EFOV-US method for making measurements of such fascicles, few published studies have utilized this method. In this study, we demonstrate both how to implement the EFOV-US method to obtain high quality musculoskeletal images and how to quantify fascicle lengths from those images. We expect that this demonstration will encourage the use of the EFOV-US method to increase the pool of muscles, both in healthy and impaired populations, for which we have in vivo muscle fascicle length data.

This study describes how to obtain high quality musculoskeletal images using the extended field-of-view ultrasound (EFOV-US) method for the purpose of making muscle fascicle length measures. We apply this method to muscles with fascicles that extend past the field-of-view of common traditional ultrasound (T-US) probes.