We wish to thank the study participants for making this work possible and the staff at the NIHR Southampton Biomedical Research Centre and the Wellcome Trust Clinical Research Facility, University Hospital Southampton for assistance with study measurements.
This work was supported by the NIHR Southampton Biomedical Research Centre, Nutrition, the MRC Lifecourse Epidemiology Unit and the University of Southampton. This report is independent research by the NIHR BRC Funding Scheme. The views expressed in this publication are those of the authors and not necessarily those of the NHS, the NIHR or the Department of Health. AMB is supported by a NIHR Clinical Research Fellowship, HPP is supported by the NIHR Southampton Biomedical Research Centre.

伦理学声明:我们按照我们机构内部的肌肉活检过程的说明现在如下。肌肉活检过程已批准的赫特福德郡研究委员会的07 / Q0204 / 68赫特福德郡骨骼肌减少症研究中使用。所有参与者都签署知情同意书20。 注:单个操作员可以执行的程序，由一名助理谁可以以减少焦虑，并确保他们是舒适在任何时候都搞参与者的谈话帮助。处置室应配备一个接收器，可调节高度的沙发，干净的表面和程序手推车。 1.准备参加（图2） <ol><li>请参与者躺在舒适，仰卧在沙发上。从露出腹股沟折痕首选大腿。在露大腿下传播的一次性吸收片。确保腿保持整个过程中直接​​和轻松，但大腿瞬间紧张勾勒出股外侧肌 。 </li><li>标记过程现场大约三分之二向下从髂前上棘的髌骨的假想线。 </li></ol> 2.准备好皮肤（图2） <ol><li>剃皮肤至约4厘米周围拟议活检部位用无菌剃须刀片的直径和清洗用醇棉签皮肤。 注意:这是为了防止在愈合过程中的创伤任何后续毛发滞留。此步骤根据操作者的偏好是可选的。 </li><li>渗入皮肤和筋膜覆以局部麻醉。使用地下25针，以提高2％利多卡因的皮下疱然后渗入更深地进入皮下组织用23政针。旨在穿透到至少一半为一英寸取决于在活检部位皮下组织的内容。该深度应足以确保覆FASCIA被麻醉。允许2 - 麻醉剂才动作5分钟。通过与针或手术刀片末端轻轻探测皮肤确认麻醉。 </li></ol> 3.配制无菌场（图2） <ol><li>用肥皂洗手，保持干净的手，直到无菌礼服穿。在程序小车营造一个无菌区前盖上无菌手套的手。 </li><li>消毒活检部位用2％氯己定/异丙醇溶液或基于碘的皮肤消毒剂。应用的无菌披盖用粘合剂孔以暴露所述活检部位，但也保持无菌区域。 </li></ol> 4.执行程序（图2） <ol><li>随着大小11手术刀，做一个5 - 上表面10毫米的切口，下并通过覆筋膜。插入通过切口的conchotome的封闭咬前端以直角到股骨的长轴线，在深度平均半至一英寸。如果需要使用自由手握住并支持周围活检部位而conchotome的前端被打开和关闭的肌肉内的大腿。 </li><li>通过90旋转conchotome - 180°切割肌肉。撤回conchotome并打开尖到无菌生理盐水浸湿无菌纱布。重复该过程的单个伤口部位内，如果需要的话，无法获得足够的肌肉组织。转移纱布到置于冰上的容器。容器转移至制备实验室根据当地机构协议来处理肌肉。 </li></ol> 5.后步骤（图2） <ol><li>直接压住伤口长达5分钟。通过将它们平行于伤口，而不是直角，以避免潜在的去除瘢痕组织，如果免缝-条被无意中删除，重新打开伤口关闭与免缝，带伤口。广场上的免缝，带无菌剂敷料和领带两层绷带压缩并用胶带固定。 </li><li>解释伤口敷料方法到与会者，他们将必须执行一个换药3 - 手术后4天。确保参与者睡前那天晚上删除加压包扎。 </li></ol> 6.邮政活检咨询<ol><li>通知参与者，这是共同的经历一些大腿刚度可通过温和的运动得到缓解（ 例如 ，行走）。使用简单止痛，如对乙酰氨基酚。警告周围的活检部位，可能会持续长达两周时间以及伤口感染的潜在并发症瞬间麻木的参与者。 </li></ol> 7.说明主题后程序<ol><li>请参与者，以避免72小时剧烈活动（如爬山，跑步，举重）。请参与者，以避免在水中浸泡48 - 72小时。然而，指示participan吨至在淋浴，以保持敷料干燥时包裹保鲜膜或周围的活检部位相似。 </li><li>请参与者后3〜改变敷料 - 第4天，一周后取出免缝-条。 注:参加者应给予活检后护理，备用敷料和负责执行的程序团队的紧急联系方式批示。 </li></ol>

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None declared.

为了保证参与者的安全，高效地进行肌肉活检，至关重要的是要注意协议中的关键步骤。 参加者必须在认真考虑排除标准，以避免并发症，如出血，伤口愈合不良选择。整个手术过程中严格无菌是必不可少的。正确的技术的应用将保证参加者的舒适性，最大限度地减少不必要的创伤参与者的组织，并允许分析足够的样本产量。有时候，可能很难获得肌肉样品，特别是如果参?...

获得骨骼肌的肌病和其他神经肌肉退化性疾病的诊断依赖于安全和有效的方法这是可以接受的，而不是失能向患者1。从历史上看，获得的肌肉组织进行分析的方法包括在全身麻醉或验尸进行开放式活检。这些技术使肌肉的直接可视化，并允许更大的一块肌肉进行活检。给定这些技术必须在患者住院和恢复方面的潜在的缺点，经皮"半开放"肌肉活组织检查方法被引入作为替代。最初是由杜氏（1806年至1875年），谁使用用套针自行建造针，并能获得来自活受试者的肌肉样品没有全身麻醉2中执行了这种技术。自那时以来，各种形式的活组织检查针已经使用3,4,5。然而，活组织检查Ñ通过贝里斯特伦在1962年开发的eedle已在临床实践中最常用的;无论是在儿童和成人，以及在研究6,7,8,9,10。它拥有锐利的套管针，切割套管和推杆驱逐组织后活检。从该过程获得的肌肉的产量已报告从25 -293毫克的范围内重复采样11,9之后。 HENRIKSSON介绍了1979年韦尔- Blakesley conchotome作为替代半开放式的肌肉活检的方法（ 图1）12。该仪器是不像贝里斯特伦结构单一的结构和设计就像一个锋利的尖咬钳子。针尖大小的范围可以从4毫米到6毫米的宽度。当刺骨尖端的两个边缘相对，中空形成确保肌肉的捕获。该conchotome通过5插入 - 10毫米皮肤切口并且避免需要一个套管针穿透肌肉和上覆筋膜作为scalpel可被用来使一个轨道下降到肌肉。 此方法允许受控的组织穿透具有高度的机动性13,14。它是网站，不当的压力可能损坏的神经血管或底层骨结构如特别有用的。在胫骨前15,14的部位。使用conchotome肌肉收益率是可变的，并在我们的实践20 - 200毫克的肌肉已经多次取样后获得。该conchotome过的Bergstrom针的一个潜在的优势，此外，以高度的机动性它提供的，是该咬尖端并不需要被削尖或作为贝里斯特伦的套管针构造16取代经常。 使用韦尔Blakesley conchotome肌肉活检的总体目标是为了获得足够量的肌肉组织，以使能对诊断和resear组织化学和分子分析CH研究。这是一个简单而安全的过程，可以很容易地了解到。利用这种技术，适合于活检肌肉包括二头肌，三头肌，三角肌，腓肠肌，胫骨前肌，比目鱼和sacrospinals 17,15,7,18,14。 股外侧的最外部分是用于活检最常见的部位，因为它避免了大腿内的主要神经血管结构。它被识别大约三分之二向下的假想线接合髂前上棘和髌骨。肌肉形态学数据，例如肌纤维区域，肌纤维的比例，毛细血管密度从股衍生外侧采样是在文献中允许研究11,19之间的比较广泛的应用。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Weil-Blakesley conchotome</td>
			<td>Gebr&#252;der Zepf Medizintechnik, D&#252;rbheim, Germany</td>
			<td>None</td>
			<td>6 mm biting tip</td>
		</tr>
		<tr>
			<td>5 ml 2% lidocaine</td>
			<td>Generic use</td>
			<td>None</td>
			<td>One 5ml vial should suffice for adequate anaesthesia</td>
		</tr>
		<tr>
			<td>Alcohol swab</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>5 mls syringe</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>25G and 23G sterile needles</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile scalpel size 11</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile gauze squares</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Sodium chloride 0.9% 10 ml</td>
			<td>Fannin</td>
			<td>1122/002/001</td>
			<td></td>
		</tr>
		<tr>
			<td>Steri-Drape&#8482; Small Drape with Adhesive Aperture</td>
			<td>3M</td>
			<td>1020</td>
			<td></td>
		</tr>
		<tr>
			<td>ChloraPrep 2% w/v /70% v/v cutaneous solution&#160;</td>
			<td>CareFusion</td>
			<td>270400</td>
			<td>Two</td>
		</tr>
		<tr>
			<td>Steri-Strip&#8482; 1/4 inch x 3 in Reinforced Adhesive Skin Closures</td>
			<td>3M</td>
			<td>R1541</td>
			<td></td>
		</tr>
		<tr>
			<td>PRIMAPORE sterile dressing 15cm x 8cm</td>
			<td>Smith &#38;&#160; 
			Nephew</td>
			<td>66000318</td>
			<td></td>
		</tr>
		<tr>
			<td>Cotton crepe bandage Hospicrepe 233 7,5 cm x 4,5 m</td>
			<td>Hartmann</td>
			<td>915562</td>
			<td></td>
		</tr>
		<tr>
			<td>Flexible cohesive bandage Co-Plus LF</td>
			<td>BSN medical</td>
			<td>2904565</td>
			<td></td>
		</tr>
		<tr>
			<td>Disposable absorbent sheet</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Micropore or transpore tape</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile gown with sterile paper towels</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
		<tr>
			<td>Sterile gloves</td>
			<td>Generic use</td>
			<td>None</td>
			<td></td>
		</tr>
	</tbody>
</table>

human vastus lateralis skeletal muscle biopsy using the weil-blakesley conchotome

使用韦尔 - Blakesley conchotome经皮肌活检是公认的在临床和科研实践。它是一种安全，有效和耐受性良好的技术。在韦尔 - Blakesley conchotome有一个尖锐的针尖刺骨用4 - 第6毫米宽的空洞。它是通过一个5插入 - 10毫米皮肤切口，并且可以被操纵用于受控的组织穿透。尖端被打开，在组织内关闭，然后通过90 -180°旋转切割肌肉。下面重复获得的肌肉量采样可以从20毫克至290毫克，可以为组织学和分子生物学研究进行处理而变化。伤口需要保持干燥和剧烈的身体活动保持最小为大约72小时，虽然活动的正常水平可以立即重新启动之后的过程。当接近注重的是主体，全无菌和后期护理程序选择此过程是安全有效的。右和left-component左侧astus外侧适于活检取决于参与者偏好。

上述过程是安全的，在临床和研究设置接受的。当活检作为诊断过程的一部分来执行，肌肉应根据患者的症状和肌肉无力的迹象来选择。活检部位应该从以前的受伤，挛缩或仪器21.1自由。在研究中，在标准条件如禁食或排斥的糖尿病患者可能需要19。抗凝血剂如华法林需要停止并且过程前凝血时间进行检查。作为我们的经验表明，它不显著增加过量出血<su...

Watch this Scientific Journal Video about Human Vastus Lateralis Skeletal Muscle Biopsy Using the Weil-Blakesley Conchotome at JoVE.com

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 ...

human-vastus-lateralis-skeletal-muscle-biopsy-using-weil-blakesley

Research

JoVE Journal

Medicine

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

16.6K 次观看.  University of Southampton, University Hospital Southampton. This video demonstrates the technique of percutaneous muscle biopsy of the human vastus lateralis using the Weil-Blakesley conchotome.

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

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

In Vivo Canine Muscle Function Assay

We describe a minimally-invasive and reproducible method to measure canine pelvic limb muscle strength and muscle response to repeated eccentric 
contractions. The pelvic limb of an anesthetized dog is immobilized in a stereotactic frame to align the tibia at a right angle to the femur. Adhesive wrap affixes the 
paw to a pedal mounted on the shaft of a servomotor to measure torque. Percutaneous nerve stimulation activates pelvic limb muscles of the paw to either push (extend) or 
pull (flex) against the pedal to generate isometric torque. Percutaneous tibial nerve stimulation activates tibiotarsal extensor muscles. Repeated eccentric (lengthening) 
contractions are induced in the tibiotarsal flexor muscles by percutaneous peroneal nerve stimulation. The eccentric protocol consists of an initial isometric contraction 
followed by a forced stretch imposed by the servomotor. The rotation effectively lengthens the muscle while it contracts, e.g., an eccentric contraction. During 
stimulation flexor muscles are subjected to an 800 msec isometric and 200 msec eccentric contraction. This procedure is repeated every 5 sec. To avoid fatigue, 4 min rest 
follows every 10 contractions with a total of 30 contractions performed.

In Vivo Canine Muscle Function Assay

We describe a minimally-invasive and painless method to measure canine hindlimb muscle strength and muscle response to repeated eccentric contractions.

在犬体内的肌肉功能测定

We describe a minimally-invasive and reproducible method to measure canine pelvic limb muscle strength and muscle response to repeated eccentric 
...

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 ...

Skin Punch Biopsy Explant Culture for Derivation of Primary Human Fibroblasts

Tissues and cell lines derived from an individual with disease are ideal sources to study disease-related cellular phenotypes. Patient-derived fibroblasts in this protocol have been successfully used in the derivation of induced pluripotent stem cells to model disease1. Early passages of these fibroblasts can also be used for cell-based functional assays to study specific disease pathways, mechanisms2 and subsequent drug screening approaches. The advantage of the presented protocol over enzymatic procedures are 1) the reproducibility of the technique from small amounts of tissue derived from older patients, e.g. patients affected with Parkinson's disease, 2) the technically simple approach over more challenging methodologies using enzymatic treatments, and 3) the time consideration: this protocol takes 15-20 min and can be performed immediately after biopsy arrival. Enzymatic treatments can take up to 4 hr and have the problems of overdigestion, reduction of cell viability and subsequent attachment of cells when not handled properly. This protocol describes the dissection and preparation of a 4-mm human skin biopsy for derivation of a fibroblast culture and has a very high success rate which is important when dealing with patient-derived tissue samples. In this culture, keratinocytes migrate out of the biopsy tissue within the first week after preparation. Fibroblasts appear 7-10 days after the first outgrowth of keratinocytes. DMEM high glucose media supplemented with 20% FBS favors the growth of fibroblasts over keratinocytes and fibroblasts will overgrow the keratinocytes. After 2 passages keratinocytes have been diluted out resulting in relatively homogenous fibroblast cultures which expresses the fibroblast marker SERPINH1 (HSP-47). Using this approach, 15-20 million fibroblasts can be derived in 4-8 weeks for cell banking. The skin dissection takes about 15-20 min, cells are then monitored once a day under the microscope, and media is changed every 2-3 days after attachment and outgrowth of cells.

The fibroblast explant culture protocol from human skin punch biopsies is a technically robust and simple way to derive skin cells within 4-8 weeks for banking of about 15-20 million cells at a low passage number.

推导人类纤维原细胞的皮肤穿刺活检植文化

Tissues and cell lines derived from an individual with disease are ideal sources to study disease-related cellular phenotypes. Patient-derived fibroblasts ...

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. ...

High-throughput Flow Cytometry Cell-based Assay to Detect Antibodies to N-Methyl-D-aspartate Receptor or Dopamine-2 Receptor in Human Serum

Over the recent years, antibodies against surface and conformational proteins involved in neurotransmission have been detected in autoimmune CNS diseases in children and adults. These antibodies have been used to guide diagnosis and treatment. Cell-based assays have improved the detection of antibodies in patient serum. They are based on the surface expression of brain antigens on eukaryotic cells, which are then incubated with diluted patient sera followed by fluorochrome-conjugated secondary antibodies. After washing, secondary antibody binding is then analyzed by flow cytometry. Our group has developed a high-throughput flow cytometry live cell-based assay to reliably detect antibodies against specific neurotransmitter receptors. This flow cytometry method is straight forward, quantitative, efficient, and the use of a high-throughput sampler system allows for large patient cohorts to be easily assayed in a short space of time. Additionally, this cell-based assay can be easily adapted to detect antibodies to many different antigenic targets, both from the central nervous system and periphery. Discovering additional novel antibody biomarkers will enable prompt and accurate diagnosis and improve treatment of immune-mediated disorders.

Over the recent years, live cell-based assays have been used successfully to detect antibodies against surface and conformational antigens. Here, we describe a method using high-throughput flow cytometry enabling the analysis of large cohorts of patients. Detection of novel antibodies will improve diagnosis and treatment of immune-mediated disorders.

高通量流式细胞仪检测细胞为基础的试验检测抗体，以N-甲基-D-天冬氨酸受体或多巴胺2受体在人血清

Over the recent years, antibodies against surface and conformational proteins involved in neurotransmission have been detected in autoimmune CNS diseases ...

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 ...