אנו מודים Yutian גן לשימוש באיור 3. עבודה זו נתמכה על ידי הקרן הלאומית למדע 0420971; המכללה סמית Blakeslee, Wilens וקרנות הולמס; לבין הווארד יוז רפואי במכון.

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	<li>Fehrenbach, E., Mooren, F. C., V&#246;lker, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Molecular and Cellular Exercise Physiology. , 199-217 (2005).</li><li>Mooren, F. C., Mooren, F. C., V&#246;lker, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Cell.">The Cell.</a> Molecular and Cellular Exercise Physiology. , 3-18 (2005).</li><li>Thompson, H. S., Clarkson, P. M., Scordilis, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+repeated+bout+effect+and+heat+shock+proteins:+intra-muscular+HSP27+and+HSP70+expression+following+two+bouts+of+eccentric+exercise+in+humans.">The repeated bout effect and heat shock proteins: intra-muscular HSP27 and HSP70 expression following two bouts of eccentric exercise in humans.</a> Acta. Physiol. Scand. 174, 47-56 (2002).</li><li>McHugh, M. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recent+advances+in+the+understanding+of+the+repeated+bout+effect:+the+protective+effect+against+muscle+damage+from+a+single+bout+of+eccentric+exercise.">Recent advances in the understanding of the repeated bout effect: the protective effect against muscle damage from a single bout of eccentric exercise.</a> Scand. J. Med. Sci. Sports. 13, 88-97 (2003).</li><li>Metskas, L. A., Kulp, M., Scordilis, S. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Gender+Dimorphism+in+the+Exercise-Naïve+Murine+Skeletal+Muscle+Proteome.">Gender Dimorphism in the Exercise-Naïve Murine Skeletal Muscle Proteome.</a> Cell Molec. Biol. Lett. 15, 507-516 (2010).</li><li>Lopez, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two-dimensional+electrophoresis+in+proteome+expression+analysis.">Two-dimensional electrophoresis in proteome expression analysis.</a> J. Chromatogr. B. 849, 190-202 (2007).</li><li>Lowry, O. H., Rosenburg, N. J., Farr, A. L., Randall, R. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Protein+measurement+with+the+Folin+phenol+reagent.">Protein measurement with the Folin phenol reagent.</a> J. Biol. Chem. 193, 265-275 (1951).</li><li>Smith, P. K., Krohn, R. I., Hermanson, G. T., Mallia, A. K., Gartner, F. H., Provenzano, M. D., Fujimoto, E. K., Goeke, N. M., Olson, B. J., Klenk, D. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Measurement+of+Protein+Using+Bicinchoninic+Acid.">Measurement of Protein Using Bicinchoninic Acid.</a> Anal. Biochem. 150, 76-85 (1985).</li><li>Norton, J. P., Clarkson, P. M., Graves, J. E., Litchfield, P. L., Kirwan, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Serum+creatine+kinase+activity+and+body+composition+in+males+and+females.">Serum creatine kinase activity and body composition in males and females.</a> Hum. Biol. 57, 591-598 (1985).</li><li>Amelink, G. J., Kamp, H. H., B&#228;r, P. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Creatine+kinase+isoenzyme+profiles+after+exercise+in+the+rat:+sex-linked+differences+in+leakage+of+CK-MM.">Creatine kinase isoenzyme profiles after exercise in the rat: sex-linked differences in leakage of CK-MM.</a> Pfl&#252;gers. Arch. 412, 417-421 (1988).</li><li>Kendall, B., Eston, R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Exercise-induced+muscle+damage+and+the+potential+protective+role+of+estrogen.">Exercise-induced muscle damage and the potential protective role of estrogen.</a> Sports Med. 32 (2), 103-123 (2002).</li><li>Tiidus, P. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Can+oestrogen+influence+skeletal+muscle+damage,+inflammation,+and+repair?.">Can oestrogen influence skeletal muscle damage, inflammation, and repair?.</a> Br. J. Sports Med. 39, 251-253 (2005).</li></ol>

אין ניגודי אינטרסים הכריז.

LC / MS proteomics השיטה המוצגת כאן הוא פרוטוקול אמין ביותר לשחזור לניתוח מהיר של הרמה הראשונה של proteome שרירי השלד. זה מאפשר השוואה אמצעי סביר הספציפיות המינים. שפע חלבונים נמוכה ידרוש חלוקה של המדגם שרירים כדי להסיר כמה שיותר את החלבונים כויץ ככל האפשר, ובכך להגדיל את שפע חלבונים נמוך. חייטות פרופיל proteome יכול להיות מושלם עם רצועות שונות בטווח pH IPG ו חלופי צבע ג&#39;ל אחוז אם תרצה בכך. הכתמים ניאון רגיש עוד קיים, אך אנו ממליצים כי כל מעבדה לקבוע את הליניאריות של כתמים אלה במערכת שלך. לצורך ניתוח קפדני יותר של ביטוי החלבון מערכת DIGE (דו מימדי ב-ג&#39;ל אלקטרופורזה המערכת, GE Healthcare מדעי החיים) יכול לשמש אך זה אינו מוסיף לרמת המורכבות למתודולוגיה. יתר על כן, פרוטוקול המתואר במסמך ניתן להשתמש עם ani ביותרmal הרקמות אשר כבר רצף הגנום, כמו גם רצף דה נובו של חלבון מכל מקור שהוא, כל עוד זה ניתן לפתור על ג&#39;ל דו מימדי, שכן שברי האנזימטית חופפים ניתן להפיק באמצעות אנזימים טוהר גבוהה בנוסף כדי טריפסין. דוגמאות ממקורות רקמות אחרות עשויות לדרוש שינויים המתודולוגיה מיצוי, במיוחד אם מחפשים חלבונים בממברנה ו הידרופובי, לעומת זאת, היו לנו תוצאות טובות באמצעות פרוטוקול זה עם לב, כבד, כליות ורקמות במוח. אחרים שלאחר translational שינויים עשויים להיות מזוהים על ידי שינוי בבסיס הנתונים מסה ספקטרום קריטריון החיפוש. לסיכום, הצגנו פרוטוקול ראשוני מפורט סביר לניתוח פרופיל proteome.

<table border="1" style=";text-align:right;direction:rtl"><tbody><tr><th> שם מגיב </th><th> סוג </th><th> חברה </th><th> מספר קטלוגי </th><th> תגובות </th></tr><tr><td> PepClean C-18 Spin עמודות </td><td> מתכלים </td><td> ThermoFisher מדעי </td><td> PI-89870 </td><td></td></tr><tr><td> מונוליטי Pepswift עמודה (100um x 5 ס&quot;מ) </td><td> מתכלים </td><td> Dionex </td><td> 162348 </td><td></td></tr><tr><td> קריטריון מראש יצוק 10.5% -14% טריס HCl-SDS polyacrylamide ג&#39;לים </td><td> מתכלים </td><td> BioRad </td><td> 345-0106 </td><td></td></tr><tr><td> Readystrip רצועות IPG </td><td> מתכלים </td><td> BioRad </td><td> משתנה ה-pH טווחי </td><td></td></tr><tr><td> חומצה אצטית </td><td> מגיב </td><td> פישר סיינטיפיק </td><td> A465-1 </td><td></td></tr><tr><td> אצטון </td><td> מגיב </td><td> Pharmco </td><td> 329000 </td><td></td></tr><tr><td> אצטוניטריל </td><td> מגיב </td><td> פישר סיינטיפיק </td><td> A955 </td><td></td></tr><tr><td>garose </td><td> מגיב </td><td> BioRad </td><td> 163-2111 </td><td> כיסוי פתרון </td></tr><tr><td> אמוניום ביקרבונט </td><td> מגיב </td><td> Fluka </td><td> 40867 </td><td></td></tr><tr><td> Bromophenol כחול </td><td> מגיב </td><td> פישר סיינטיפיק </td><td> BP114 </td><td></td></tr><tr><td> Bio-Lyte Ampholytes </td><td> מגיב </td><td> BioRad </td><td></td><td> משתנה ה-pH טווחי </td></tr><tr><td> בחורים </td><td> מגיב </td><td> USB </td><td> 13361 </td><td></td></tr><tr><td> Commassie כחול R-250 </td><td> מגיב </td><td> ThermoFisher מדעי </td><td> 20278 </td><td></td></tr><tr><td> Dithiothreitol (DTT) </td><td> מגיב </td><td> USB </td><td> 15395 </td><td></td></tr><tr><td> חומצה פורמית </td><td> מגיב </td><td> ThermoFisher מדעי </td><td> 28905 </td><td></td></tr><tr><td> גליצרול </td><td> מגיב </td><td> Sigma-Aldrich </td><td> G6279 </td><td></td></tr><tr><td> גליצין </td><td> מגיב </td><td> USB </td><td> 16407 </td><td></td&gt; </tr><tr><td> Iodoacetamide </td><td> מגיב </td><td> Sigma-Aldrich </td><td> I6125 </td><td></td></tr><tr><td> מתנול </td><td> מגיב </td><td> פישר סיינטיפיק </td><td> A452 </td><td></td></tr><tr><td> שמן מינרלי </td><td> מגיב </td><td> BioRad </td><td> 163-2129 </td><td></td></tr><tr><td> Dodecyl נתרן גופרתי </td><td> מגיב </td><td> Sigma-Aldrich </td><td> L6026 </td><td></td></tr><tr><td> טריס בסיס </td><td> מגיב </td><td> Sigma-Aldrich </td><td> 93349 </td><td></td></tr><tr><td> טריס [2-carboexyethyl] phosphine </td><td> מגיב </td><td> ThermoFisher מדעי </td><td> 77720 </td><td></td></tr><tr><td> טריפסין Endoproteinase, TPCK מטופלים, MS כיתה </td><td> מגיב </td><td> לנקב </td><td> 90055 </td><td> שונה </td></tr><tr><td> דשן </td><td> מגיב </td><td> USB </td><td> 75826 </td><td></td></tr><tr><td> מים </td><td> מגיב </td><td> Burdick &amp; ג&#39;קסון </td><td> 365 </td><td></td></tr><tr><td> Centrivap מרוכז </td><td> Tool </td><td> Labconco </td><td></td><td></td></tr><tr><td> Exquest חותך נקודה </td><td> כלי </td><td> BioRad </td><td></td><td></td></tr><tr><td> LCQ Deca XP מקס יון ספקטרומטר מסה מלכודת </td><td> כלי </td><td> ThermoFisher מדעי </td><td></td><td></td></tr><tr><td> ממונע העלי </td><td> כלי </td><td> Kontes קורפ </td><td></td><td></td></tr><tr><td> פוליפרופילן בחישה </td><td> כלי </td><td> BioSpec Prod </td><td></td><td></td></tr><tr><td> מוטות </td><td></td><td></td><td></td><td></td></tr><tr><td> פושט צורה ולובש צורה תא IEF </td><td> כלי </td><td> BioRad </td><td></td><td></td></tr><tr><td> Sonicator </td><td> כלי </td><td> ברנסון </td><td></td><td></td></tr><tr><td> המודד פלוס HPLC מערכת </td><td> כלי </td><td> ThermoFisher מדעי </td><td></td><td></td></tr><tr><td> VersaDoc הדמיה מערכת </td><td> כלי </td><td> BioRad </td><td></td><td></td></tr></tbody></table>

skeletal muscle gender dimorphism from proteomics

התכווצות שרירי השלד הגולמי נקבעת בעיקר על ידי מספר קטן יחסית של חלבונים התכווצות, אך רקמה זו גם וישימה להפליא גורמים סביבתיים כגון 1 היפרטרופיה על ידי תרגילי התנגדות וניוון של חוסר שימוש. זה ובכך מפגין שיפוץ והתאמות ללחצים (חום, איסכמיה, מתכות כבדות וכו &#39;.) 2,3. נזק עלול להיגרם לשרירים על ידי כוח השריר מתארך תוך כדי הפעלת, 4 שנקרא כיווץ אקסצנטרי. חלבונים התכווצות ניתן נפגע מאמצים כאלה צריכים לתקן, מושפל ו / או resynthesized; פונקציות אלה אינם חלק של חלבונים התכווצות, אך שפע של חלבונים הרבה פחות אחרים בתא. כדי לקבוע מה משנה של חלבונים מעורב שיפור של סוג זה של נזק, proteome גלובלי חייב להיות שהוקמו לפני תרגיל 5 ואחר כך המשכתי לאחר התרגיל כדי לקבוע את ההפרשביטוי החלבון ובכך לסמן חלבונים מועמד ההתאמות נזק ותיקון שלה. יתר על כן, רוב המחקרים של שרירי השלד נערכו על הזכר של המין ולכן לא יכול להיות נציג של שריר נשית. במאמר זה אנו מציגים שיטה להפקת חלבונים מהשרירים reproducibly זכר ונקבה, ומפריד אותם על ידי אלקטרופורזה דו מימדי ג&#39;ל ואחריו ברזולוציה גבוהה הדמיה דיגיטלית 6. פרוטוקול זה מספק עבור כתמים (חלבונים המזוהים לאחר מכן), כי להראות מובהקות סטטיסטית (p &lt;0.05) כפולה להגדיל או להקטין, להופיע או להיעלם מן המדינה שליטה. אלה נכרת אז מתעכל עם טריפסין ו מופרדים על ידי כרומטוגרפיה נוזלית בלחץ גבוה מצמידים את ספקטרומטר מסה (LC / MS) לזיהוי חלבון (LC / MS / MS) 5. מתודולוגיה זו (איור 1) ניתן להשתמש ברקמות רבות עם קצת שינוי לא (כבד, מוח, לשמועלא וכו &#39;.).

Watch this Scientific Journal Video about Skeletal Muscle Gender Dimorphism from Proteomics at JoVE.com

Skeletal Muscle Gender Dimorphism from Proteomics

קבוצת ישר קדימה שיטות לבודד לקבוע את זהותו של החלבונים הנפוץ ביותר בא לידי ביטוי בשרירי השלד. אודות 800 נקודות הם להבחין על ג&#39;ל דו מימדי של השריר 10 מ&quot;ג; זה מאפשר להגדרה של ביטוי המגדר חלבון ספציפי. שיטות אלה ייתן תוצאות המקבילה רוב הרקמות.

שרירי השלד מין Dimorphism מ Proteomics

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

skeletal-muscle-gender-dimorphism-from-proteomics

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JoVE Journal

Medicine

12.4K Views.  Smith College. קבוצת ישר קדימה שיטות לבודד לקבוע את זהותו של החלבונים הנפוץ ביותר בא לידי ביטוי בשרירי השלד. אודות 800 נקודות הם להבחין על ג'ל דו מימדי של השריר 10 מ"ג; זה מאפשר להגדרה של ביטוי המגדר חלבון ספציפי. שיטות אלה ייתן תוצאות המקבילה רוב הרקמות.

Video: Skeletal Muscle Gender Dimorphism from Proteomics

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.

מיו מכני ניתוח בשרירי השלד מבודדת

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 ...

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;

חיתוך רוחב עצב הטיביאלי - מודל סטנדרטי לניוון שרירים שלד מושרה denervation בעכברים

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.

נהלי ביופסיה של שריר שלד אנושי באמצעות טכניקת Bergstrom השתנה

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.

ההכנה וRespirometric הערכה של המיטוכונדריה מבודדת מרקמת שריר שלד מתקבלת על ידי Percutaneous מחט ביופסיה

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.

דימות תהודה מגנטית כמותי של מחלת שריר שלד

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.

בן אנוש<em&gt; Vastus lateralis</em&gt; ביופסיה בשרירי השלד שימוש וייל-בלייסלי 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.

אינדוקציה הערכת נזק שרירי השלד Exertional בבני אדם

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

Phosphorus-31 Magnetic Resonance Spectroscopy: A Tool for Measuring In Vivo Mitochondrial Oxidative Phosphorylation Capacity in Human Skeletal Muscle

Skeletal muscle mitochondrial oxidative phosphorylation (OXPHOS) capacity, which is critically important in health and disease, can be measured in vivo and noninvasively in humans via phosphorus-31 magnetic resonance spectroscopy (31PMRS). However, the approach has not been widely adopted in translational and clinical research, with variations in methodology and limited guidance from the literature. Increased optimization, standardization, and dissemination of methods for in vivo 31PMRS would facilitate the development of targeted therapies to improve OXPHOS capacity and could ultimately favorably impact cardiovascular health. 31PMRS produces a noninvasive, in vivo measure of OXPHOS capacity in human skeletal muscle, as opposed to alternative measures obtained from explanted and potentially altered mitochondria via muscle biopsy. It relies upon only modest additional instrumentation beyond what is already in place on magnetic resonance scanners available for clinical and translational research at most institutions. In this work, we outline a method to measure in vivo skeletal muscle OXPHOS. The technique is demonstrated using a 1.5 Tesla whole-body MR scanner equipped with the suitable hardware and software for 31PMRS, and we explain a simple and robust protocol for in-magnet resistive exercise to rapidly fatigue the quadriceps muscle. Reproducibility and feasibility are demonstrated in volunteers as well as subjects over a wide range of functional capacities.

Phosphorus-31 Magnetic Resonance Spectroscopy: A Tool for Measuring In Vivo Mitochondrial Oxidative Phosphorylation Capacity in Human Skeletal Muscle

This work demonstrates the feasibility of an in vivo phosphorus-31 magnetic resonance spectroscopy (31PMRS) technique to quantify mitochondrial oxidative phosphorylation (OXPHOS) capacity in human skeletal muscle.

זרחן-31 ספקטרוסקופיה תהודה מגנטית: כלי למדידת<em&gt; In vivo</em&gt; קיבולת זרחון חמצוני מיטוכונדריאלי של שריר שלד אדם

Skeletal muscle mitochondrial oxidative phosphorylation (OXPHOS) capacity, which is critically important in health and disease, can be measured in vivo ...

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.

ניתוח חצי אוטומטי של מורפולוגיה שרירי השלד העכבר והרכב סיבים מסוג

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.

צימוד נוירו-וסקולריים שרירי השלד, קיבולת חמצוני, ופועלים Microvascular עם 'אחד עצור חנות' ספקטרוסקופיית אינפרא אדום

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