Test of a new technique for the diagnosis of carpal tunnel syndrome.

Several new techniques for carpal tunnel syndrome diagnosis have been developed in the last few years. This work tests a technique that compares the distal motor latency of the median nerve to the second lumbrical muscle (2L) with the distal motor latency of the ulnar nerve to the interossei muscle (INT). Results from 40 normal hands give the superior limit of the normal difference (2L-INT) as 0. 26 ms (&xmacr;+3 SD). In 55 hands with different levels of carpal tunnel syndrome, this new technique was more sensitive and accurate than the conventional test which uses the distal motor latency of the median nerve to the abductor pollicis brevis muscle (APB), especially in the less severe cases. With the absence of the compound muscle action potentials of the APB muscle caused by severe thenar atrophy, it is much easier to obtain the potential from the 2L muscle. We concluded that this is a sensitive, simple, rapid, and non-invasive new technique, and therefore, it should be incorporated as part of the routine ENMG procedures for carpal tunnel syndrome diagnosis.     

Isolation of a slowly adhering cell fraction containing stem cells from murine skeletal muscle by the preplate technique

This protocol details a procedure, known as the modified preplate technique, which is currently used in our laboratory to isolate muscle cells on the basis of selective adhesion to collagen-coated tissue culture plates. By employing this technique to murine skeletal muscle, we have been able to isolate a rapidly adhering cell (RAC) fraction within the earlier stages of the process, whereas a slowly adhering cell (SAC) fraction containing muscle-derived stem cells is obtained from the later stages of the process. This protocol outlines the methods and materials needed to isolate RAC and SAC populations from murine skeletal muscle. The procedure involves mechanical and enzymatic digestion of skeletal muscle tissue with collagenase XI, dispase and trypsin followed by plating the resultant muscle slurry on collagen type I-coated flasks where the cells adhere at different rates. The entire preplate technique requires 5 d to obtain the final preplate SAC population. Two to three additional days are usually required before this population is properly established. We also detail additional methodologies designed to further enrich the resultant cell population by continuing the modified preplating process on the SAC population. This process is known as replating and requires further time.     

The influence of limb alignment and transfemoral amputation technique on muscle capacity during gait

Many factors influence successful outcomes following transfemoral amputation. One factor is surgical technique. In this study, the influence of limb alignment and surgical technique on a muscle’s capacity to generate force was examined using musculoskeletal modeling. Non-amputee and transfemoral amputee models were analyzed while hip adduction, femur length, and reattached muscle wrap position, tension and stabilization technique were systematically varied. With muscle tension preserved, wrap position and femur length had little influence on muscle capacity. However, limb alignment, muscle tension and stabilization technique notably influenced muscle capacity. Overall, myodesis stabilization provided greater muscle balance and function than myoplasty stabilization.     

A Novel Application of Musculoskeletal Ultrasound Imaging

Ultrasound is an attractive modality for imaging muscle and tendon motion during dynamic tasks and can provide a complementary methodological approach for biomechanical studies in a clinical or laboratory setting. Towards this goal, methods for quantification of muscle kinematics from ultrasound imagery are being developed based on image processing. The temporal resolution of these methods is typically not sufficient for highly dynamic tasks, such as drop-landing. We propose a new approach that utilizes a Doppler method for quantifying muscle kinematics. We have developed a novel vector tissue Doppler imaging (vTDI) technique that can be used to measure musculoskeletal contraction velocity, strain and strain rate with sub-millisecond temporal resolution during dynamic activities using ultrasound. The goal of this preliminary study was to investigate the repeatability and potential applicability of the vTDI technique in measuring musculoskeletal velocities during a drop-landing task, in healthy subjects. The vTDI measurements can be performed concurrently with other biomechanical techniques, such as 3D motion capture for joint kinematics and kinetics, electromyography for timing of muscle activation and force plates for ground reaction force. Integration of these complementary techniques could lead to a better understanding of dynamic muscle function and dysfunction underlying the pathogenesis and pathophysiology of musculoskeletal disorders.     

Reconstruction of the maxilla with a double musculoperiosteal flap in connection with a composite calvarial bone graft

A new technique is shown for a one-stage reconstruction of the mucosa of the floors of the nose and maxillary sinus, the bone structures of the maxilla and the hard palate, as well as the mucosal layers of the hard and soft palates and vestibulum. To accomplish this coverage, a vascularized calvarial bone graft with temporal muscle from one side is combined with a vascularized temporal muscle flap from the other side to achieve a three-layer "sandwich" plasty. The advantage of this procedure is reconstruction of the complete maxillary defect with the possibility of denture rehabilitation and the avoidance of oronasal fenestration. Besides the possible complication of insufficient vascularization of the bone and muscle grafts, the donor defect in the calvarial bone and the missing muscle for mastication are to be considered.     

Accumulation of deletions in human mitochondrial DNA during normal aging: analysis by quantitative PCR

We have developed a quantitative PCR technique to measure the amount of a specific mitochondrial DNA deletion (ΔmtDNA), the so-called ‘common deletion’, in human tissues. Using this method, we estimate that there is a 10 000-fold increase in this ΔmtDNA species in muscle during the course of the normal human lifespan. The maximum amount of common deletion observed in aged muscle was approx 0.1%. Tissues that turn-over slowly, such as skeletal muscle and heart, contained more ΔmtDNA than more rapidly dividing tissues, such as a liver, in agreement with studies performed by others.     

The effect on butyryl-CoA dehydrogenase of reagents specific for nucleophilic sulphur.

The technique of erythrocyte-mediated microinjection has been successfully adapted for use with cultured muscle cells. Erythrocytes were fused with primary chick myotube cultures with poly(ethylene glycol), and fluorescent antibodies to haemoglobin demonstrated that this protein was injected into the sarcoplasm of myotubes. The microinjection treatment did not significantly alter protein metabolism in the muscle cells as monitored by rates of synthesis and degradation of muscle proteins. 125I-labelled ribonuclease A and bovine serum albumin were degraded with the expected exponential decay kinetics after microinjection into muscle cells, and the half-life of ribonuclease A (40 h) was approximately twice that of bovine serum albumin (17 h). The degradation of ribonuclease A in the muscle cells was enhanced 1.6-fold in the absence of horse serum and chick-embryo extract, whereas the degradation of bovine serum albumin was not altered during deprivation. These results are characteristic of the breakdown of microinjected ribonuclease A and bovine serum albumin in other cell types. Therefore, our experiments indicate the erythrocyte-mediated microinjection is a valid technique to study protein degradation in primary chick muscle cultures.     

A new histochemical method for magnesium actomyosin adenosine triphosphatase at physiological pH

A new lead-precipitation technique for demonstrating magnesium-activated actomyosin adenosine triphosphatase (ATPase) at physiological pH and electrolyte levels in fixed skeletal muscle sections is reported. This method is compared with standard acid- and alkali-denatured muscle stained for calcium myosin ATPase as well as calcium-formalin denatured and pyrophosphate-formalin denatured muscle also stained for calcium myosin ATPase. The technique was developed using hamster skeletal muscle; however, it has also been applied to human, rat, and cat muscle. The fiber-type staining intensities of the formalin-denatured magnesium actomyosin ATPase closely resemble those of the formalin-denatured calcium myosin ATPase in rodents, but intensities in Type 1 fibers are reversed relative to calcium myosin ATPase in human muscle. Cat muscle shows intermediate characteristics.     

Picrosirius red staining of cardiac muscle following phosphomolybdic acid treatment

When the picrosirius red technique was applied to cardiac muscle sections, intense yellow myocyte staining sometimes obscured thin collagenous septa. The picrosirius red technique was modified to include treatment of the sections in 0.2% (w/v) aqueous phosphomolybdic acid prior to staining. With 1-5 min treatment, cytoplasmic staining was eradicated; diminution of collagen staining occurred only with long treatments at much higher concentrations of phosphomolybdic acid. Using this phosphomolybdic acid-picrosirius red technique, collagenous septa as thin as 0.2-0.5 micron and fine collagen fibers making up the septa were clearly discernible. The technique also worked well on sections stained by other techniques and then destained. The phosphomolybdic acid-picrosirius red technique should be useful in experiments designed to investigate the effects of collagen distribution on the electrical and mechanical behavior of cardiac muscle.     

Picrosirius Red Staining of Cardiac Muscle Following Phosphomolybdic Acid Treatment

When the picrosirius red technique was applied to cardiac muscle sections, intense yellow myocyte staining sometimes obscured thin collagenous septa. The picrosirius red technique was modified to include treatment of the sections in 0.2% (w/v) aqueous phosphomolybdic acid prior to staining. With 1-5 min treatment, cytoplasmic staining was eradicated; diminution of collagen staining occurred only with long treatments at much higher concentrations of phosphomolybdic acid. Using this phosphomolybdic acid-picrosirius red technique, collagenous septa as thin as 0.2-0.5 /im and fine collagen fibers making up the septa were clearly discernible. The technique also worked well on sections stained by other techniques and then destained. The phosphomolybdic acid-picrosirius red technique should be useful in experiments designed to investigate the effects of collagen distribution on the electrical and mechanical behavior of cardiac muscle.     

Quantitative histochemical investigations of semipermeable membrane techniques for the assay of acid phosphatase in skeletal muscle

Summary The reproducibility, specificity and validity of Meijer's semipermeable membrane simultaneous coupling technique for the assay of acid phosphatase activity in sections of skeletal muscle have been investigated quantitatively, using naphthol AS-BI phosphate as the substrate and hexazotised Pararosanaline (HPRA) as the coupler. With this technique, unlike conventional techniques, presumed specific final reaction product (FRP) is evident in three different histological sites in normal skeletal muscle; first, as intensely coloured red granules within muscle fibres; second, as a diffuse reddish colouration throughout the sarcoplasm of all muscle fibres (the intrafibre areas); and third, in certain connective tissue elements between the muscle fibres (the interfibre areas). The mean absorbance of the FRP (at its absorption maximum, 530 nm) formed in each of these sites after a constant incubation time does not differ significantly in serial sections. 6 mM sodium molybdate, an acid phosphatase inhibitor, reduces the mean absorbance by 50% in the intrafibre areas, but in the interfibre connective tissue areas, 1 mM is sufficient. In contrast, 10 mM EDTA, an alkaline phosphatase inhibitor, has a negligible effect on the formation of specific FRP. Thus, Meijer's technique appears to be reproducible and specific. The mean absorbance of the FRP formed in each of the three reactive histological areas increases linearly with incubation time and section thickness. The maximum amount of FRP is formed at pH 5 and when the substrate concentration is above about 4 mM. However, some of the FRP in the intrafibre areas is unspecific, and arises from the transformation of adsorbed HPRA to a purple-coloured product having an absorption maximum at 570 nm. Much of the non-specific FRP appears after the incubation has been terminated with formalin, and reaches a maximum several hours after the sections have been subsequently mounted. As a consequence, Meijer's technique is not entirely valid.     

Muscle-sparing latissimus dorsi myocutaneous flap with maintenance of muscle innervation,function, and aesthetic appearance of the donor site

In this report, the authors describe the application of a muscle-sparing technique to harvest a myocutaneous latissimus dorsi muscle flap, including only a tiny lateral muscle segment but carrying a large skin paddle, with the advantage of leaving intact innervation and function of the remaining latissimus dorsi muscle. According to the experiences and complications associated with the pure thoracodorsal artery perforator harvest at the authors' institution, the necessity of increasing the reliability of the vascular pedicle demands that a small muscle strip be left embedding the perforator vessels attached to the skin paddle. This procedure was applied in eight cases with only one minor complication, which was a distal flap tip necrosis in the largest flap used. The muscle function and aesthetic contour of the posterior axillary fold were preserved in every case. Harvesting a large skin paddle flap that is carried by a diminutive longitudinal segment of latissimus dorsi muscle circumvents thoracodorsal nerve damage and maintains muscle function. In contrast to a thoracodorsal artery perforator flap without muscle, the harvesting of which is a delicate procedure, this procedure is regarded as easier and safer.     

Free gracilis muscle transplantation, with microneurovascular anastomoses for the treatment of facial paralysis. A preliminary report.

A clinical operative technique for free muscle transplantation by microneurovascular anastomoses is presented. Two cases of free transfer of the gracilis muscle for dynamic reconstruction of facial paralysis are described, including a follow-up study with electromyography, light microscopy, and electron microscopy. We feel this new technique will have a wide range of application in reconstructive surgery.     

Voltage clamp methods for the study of membrane currents and SR Ca(2+) release in adult skeletal muscle fibres

Skeletal muscle excitation-contraction (E-C)(1) coupling is a process composed of multiple sequential stages, by which an action potential triggers sarcoplasmic reticulum (SR)(2) Ca(2+) release and subsequent contractile activation. The various steps in the E-C coupling process in skeletal muscle can be studied using different techniques. The simultaneous recordings of sarcolemmal electrical signals and the accompanying elevation in myoplasmic Ca(2+), due to depolarization-initiated SR Ca(2+) release in skeletal muscle fibres, have been useful to obtain a better understanding of muscle function. In studying the origin and mechanism of voltage dependency of E-C coupling a variety of different techniques have been used to control the voltage in adult skeletal fibres. Pioneering work in muscles isolated from amphibians or crustaceans used microelectrodes or 'high resistance gap' techniques to manipulate the voltage in the muscle fibres. The development of the patch clamp technique and its variant, the whole-cell clamp configuration that facilitates the manipulation of the intracellular environment, allowed the use of the voltage clamp techniques in different cell types, including skeletal muscle fibres. The aim of this article is to present an historical perspective of the voltage clamp methods used to study skeletal muscle E-C coupling as well as to describe the current status of using the whole-cell patch clamp technique in studies in which the electrical and Ca(2+) signalling properties of mouse skeletal muscle membranes are being investigated.     

Glut 4 content in the plasma membrane of rat skeletal muscle: comparative studies of the subcellular fractionation method and the exofacial photolabelling technique using ATB-BMPA

Employing subcellular membrane fractionation methods it has been shown that insulin induces a 2-fold increase in the Glut 4 protein content in the plasma membrane of skeletal muscle from rats. Data based upon this technique are, however, impeded by poor plasma membrane recovery and cross-contamination with intracellular membrane vesicles. The present study was undertaken to compare the subcellular fractionation technique with the technique using [³H]ATB-BMPA exofacial photolabelling and immunoprecipitation of Glut 4 on soleus muscles from 3-week-old Wistar rats. Maximal insulin stimulation resulted in a 6-fold increase in 3-O-methylglucose uptake, and studies based on the subcellular fractionation method showed a 2-fold increase in Glut 4 content in the plasma membrane, whereas the exofacial photolabelling demonstrated a 6- to 7-fold rise in cell surface associated Glut 4 protein. Glucose transport activity was positively correlated with cell surface Glut 4 content as estimated by exofacial labelling. In conclusion: (1) the increase in glucose uptake in muscle after insulin exposure is caused by an augmented concentration of Glut 4 protein on the cell surface membrane, (2) at maximal insulin stimulation (20 mU/ml) approximately 40% of the muscle cell content of Glut 4 is at the cell surface, and (3) the exofacial labelling technique is more sensitive than the subcellular fractionation technique in measuring the amount of glucose transporters on muscle cell surface.     

Preparation and properties of vertebrate smooth-muscle myofibrils and actomyosin.

A new technique for obtaining a myofibril-like preparation from vertebrate smooth muscle has been developed. An actomyosin can be readily extracted from these myofibrils at low ionic strength and in yields 20 times as high as previously reported. The protein composition of all preparations has been monitored using dodecylsulfate-gel electrophoresis. By this method smooth muscle actomyosin showed primarily only the major proteins, myosin, actin and tropomyosin, while the myofibrils contained, additionally, three new proteins not previously described with polypeptide chain weights of 60000, 110000 and 130000. The ATPase activities of both the myofibrils and actomyosin preparations are considerably higher than previously described for vertebrate smooth muscle. They are sensitive to micromolar Ca2+ ion concentrations to the same degree as comparable skeletal and cardiac muscle preparations, even though troponin-like proteins could not be identified in these smooth muscle preparations. From the latter observation and the presence of Ca2+-sensitivity in tropomyosin-free actomyosin it is suggested that this calcium sensitivity is, as in some invertebrate muscles, a property of the myosin molecule.     

大鼠输精管平滑肌细胞的体外培养及形态学特征(英文）

建立大鼠输精管平滑肌细胞的培养方法。取大鼠输精管,剥离外膜和内膜,用组织块法进行体外培养。用抗α-SMA(anti α-smooth muscle actin)免疫组化染色的方法鉴定培养的细胞。结果显示,在倒置显微镜下观察细胞形态多样,表现为长梭形或星形,细胞伸出突起互相接触,彼此融合,部分区域细胞多层重叠,部分区域细胞单层高低起伏,呈"峰-谷"状生长。免疫组化染色鉴定呈阳性反应,用该方法所分离、培养的输精管平滑肌细胞纯度达99%以上。应用组织块法培养大鼠输精管平滑肌细胞,操作简单,结果稳定。     

Technique for interpretation of electromyography for concentric and eccentric contractions in gait

We present a technique to combine muscle shortening and lengthening velocity information with electromyographic (EMG) profiles during gait. A biomechanical model was developed so that each muscle's length could be readily calculated over time as a function of angles of the joints it crossed. The velocity of shortening and lengthening of the muscle fiber was then calculated, and with computer graphics this information was overlaid on the EMG profiles. Thus, researchers and clinicians were not only able to interpret the processed EMG signal as level of activity (tension) but also to gain insight as to the muscles' role as generators (muscle shortening) or absorbers (muscle lengthening) of energy. Six common muscles are documented, using database profiles; soleus (SOL), medial gastrocnemius (MG), tibialis anterior (TA), vastus lateralis (VL), rectus femoris (RF), and semitendinosus (ST). The protocol thus demonstrates a relatively simple technique for calculating muscle fiber velocity and for combining that velocity information with EMG activity profiles.     

Use of uncalibrated biplanar radiography for the measurement of skeletal coordinates around the shoulder girdle

Mechanical modelling of the musculoskeletal system is dependent upon information regarding the bony attachments of the relevant muscles; in order to study the biomechanics of the shoulder girdle the authors have identified the muscle attachments in three embalmed cadavers. A simple biplanar radiographic technique was then used to determine the attachment coordinates using frames of reference defined for each bone. This technique, using hand positioning without special fixtures was believed to be sufficiently accurate, bearing in mind the likely degree of biological variation. In order to test this assumption, the accuracy of the technique has been studied by measuring the agreement between the two measurements of the common coordinate in the pairs of radiographs. it was found that for the trunk, the errors in the common coordinate were always less than the natural variation; for the scapula they were of a similar magnitude but, for the humerus, the measurement errors frequently exceeded the variation in the coordinates of muscle attachments. It was concluded that, in general, uncalibrated biplanar radiography was sufficiently accurate for the determination of the spatial coordinates of muscle attachments.