A Method for Quantitative Measurement of Mitochondrial Creatine Kinase in Human Skeletal Muscle

1992 ◽  
Vol 29 (2) ◽  
pp. 196-201 ◽  
Author(s):  
J Smeitink ◽  
R Wevers ◽  
J Hulshof ◽  
W Ruitenbeek ◽  
T v Lith ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Creatine Kinase ◽  
Human Skeletal Muscle ◽  
Quantitative Measurement ◽  
Mitochondrial Creatine Kinase ◽  
Biopsy Material ◽  
Skeletal Muscle Biopsy ◽  
Wet Weight ◽  
Enzyme Complexes ◽  
Generating System

Defects in the mitochondrial energy generating system in patients with a mitochondrial myopathy are known to be localized in various enzyme complexes involved in energy production. Such a defect may exist at the level of mitochondrial creatine kinase (Mi-CK). On that account we have developed a method for measurement of the enzyme activity in human skeletal muscle biopsy material (> 10 mg). Interfering creatine kinase isoenzymes are removed by anion exchange and affinity chromatography. The activity of Mi-CK in reference skeletal muscle homogenates amounts to 240±88 mU/mg protein (30±8·0 mU/mg wet weight).

scholarly journals Maturation of Mitochondrial and other Isoenzymes of Creatine Kinase in Skeletal Muscle of Preterm Born Infants

1992 ◽  
Vol 29 (3) ◽  
pp. 302-306 ◽  
Author(s):  
J Smeitink ◽  
W Ruitenbeek ◽  
T v Lith ◽  
R Sengers ◽  
F Trijbels ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Creatine Kinase ◽  
Protein Content ◽  
Human Skeletal Muscle ◽  
Specific Activity ◽  
Prenatal Development ◽  
Quadriceps Muscle ◽  
Mitochondrial Creatine Kinase ◽  
Total Creatine ◽  
Preterm Born

We studied pre- and postnatal changes in total creatine kinase (CK) activity, mitochondrial creatine kinase (Mi-CK) activity and immunochemical reactivity with anti-Mi-CK antibodies in skeletal muscle specimens from 12 infants, 10 of them preterm born, after a pregnancy varying between 28 and 40 weeks. Our results demonstrate that Mi-CK is present in fetal human quadriceps muscle and that the specific activity of Mi-CK increases during prenatal development from week 28 to 40 by a factor about two. Generally, adult levels have not been reached at birth, indicating a further postnatal increase of the activity of the enzyme. The Mi-CK protein content also increases during prenatal development. These results suggest that in human skeletal muscle the expression and accumulation of Mi-CK starts at mid-gestation, later than is known to occur for cytosolic CK.

scholarly journals Mitochondrial creatine kinase activity and phosphate shuttling are acutely regulated by exercise in human skeletal muscle

2012 ◽  
Vol 590 (21) ◽  
pp. 5475-5486 ◽  
Author(s):  
Christopher G. R. Perry ◽  
Daniel A. Kane ◽  
Eric A. F. Herbst ◽  
Kazutaka Mukai ◽  
Daniel S. Lark ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Creatine Kinase ◽  
Human Skeletal Muscle ◽  
Kinase Activity ◽  
Creatine Kinase Activity ◽  
Mitochondrial Creatine Kinase

Fructose transport and GLUT-5 protein in human sarcolemmal vesicles.

1997 ◽  
Vol 273 (3) ◽  
pp. E543 ◽  
Author(s):  
S Kristiansen ◽  
F Darakhshan ◽  
E A Richter ◽  
H S Hundal
Keyword(s):  
Skeletal Muscle ◽  
Protein Concentration ◽  
Human Skeletal Muscle ◽  
Cytochalasin B ◽  
Human Muscle ◽  
Transport Activity ◽  
Biopsy Material ◽  
Skeletal Muscle Biopsy ◽  
Uptake Assay ◽  
Sarcolemmal Vesicles

Sarcolemmal vesicles were produced from human skeletal muscle biopsy material obtained at rest and immediately after maximal dynamic exercise (100% maximal O2 uptake) for analysis of fructose transport and hexose transporter (GLUT-5) protein concentration. Human sarcolemmal vesicles displayed a time-dependent uptake of D-fructose that displayed saturable Michaelis-Menten type kinetics (maximal transport 477 +/- 37 pmol.min-1.mg protein-1; half-maximal concentration constant 8.3 +/- 1.2 mM). At a hexose concentration of 5 mM, vesicle transport rate was eight times faster for glucose than for fructose. Preincubation of human muscle vesicles with 35 microM cytochalasin B before the uptake assay resulted in > 95% inhibition in D-glucose uptake, whereas transport of D-fructose was unaffected. Sarcolemmal vesicles prepared from exercised human muscle showed a significant increase (49%) in vesicle GLUT-4 content (P < 0.03, n = 10), which accounts for the increase in vesicle glucose transport that we have recently reported [S. Kristiansen, M. Hargreaves, and E.A. Richter. Am. J. Physiol. 270 (Endocrinol. Metab. 33): E197-E201, 1996]. In contrast, exercise did not increase the vesicle GLUT-5 protein content or induce changes in vesicle fructose transport activity. In conclusion, we propose that fructose transport into human skeletal muscle occurs via a mechanism distinct from that utilized by glucose on the basis of differences in sensitivity to cytochalasin B and responsiveness to exercise. Furthermore, our findings signify that uptake of fructose in human skeletal muscle is mediated by the GLUT-5 transporter.

Enzymic Analysis of Human Skeletal Muscle Biopsy Samples

1981 ◽  
Vol 60 (3) ◽  
pp. 19P-19P
Author(s):  
F. Martin ◽  
J. Levi ◽  
G. Slavin ◽  
T. J. Peters
Keyword(s):  
Skeletal Muscle ◽  
Muscle Biopsy ◽  
Human Skeletal Muscle ◽  
Skeletal Muscle Biopsy

Adipose triglyceride lipase in human skeletal muscle is upregulated by exercise training

2009 ◽  
Vol 296 (3) ◽  
pp. E445-E453 ◽  
Author(s):  
Thomas J. Alsted ◽  
Lars Nybo ◽  
Martina Schweiger ◽  
Christian Fledelius ◽  
Poul Jacobsen ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Exercise Training ◽  
Endurance Exercise ◽  
Human Skeletal Muscle ◽  
Hormone Sensitive Lipase ◽  
Adipose Triglyceride Lipase ◽  
Skeletal Muscle Biopsy ◽  
Triglyceride Lipase ◽  
Endurance Exercise Training ◽  
Molecule Inhibitor

Mobilization of fatty acids from stored triacylglycerol (TG) in adipose tissue and skeletal muscle [intramyocellular triacylglycerol (IMTG)] requires activity of lipases. Although exercise training increases the lipolytic capacity of skeletal muscle, the expression of hormone-sensitive lipase (HSL) is not changed. Recently, adipose triglyceride lipase (ATGL) was identified as a TG-specific lipase in various rodent tissues. To investigate whether human skeletal muscle ATGL protein is regulated by endurance exercise training, 10 healthy young men completed 8 wk of supervised endurance exercise training. Western blotting analysis on lysates of skeletal muscle biopsy samples revealed that exercise training induced a twofold increase in skeletal muscle ATGL protein content. In contrast to ATGL, expression of comparative gene identification 58 (CGI-58), the activating protein of ATGL, and HSL protein was not significantly changed after the training period. The IMTG concentration was significantly decreased by 28% at termination of the training program compared with before. HSL-phoshorylation at Ser660 was increased, HSL-Ser659 phosporylation was unchanged, and HSL-phoshorylation at Ser565 was decreased altogether, indicating an enhanced basal activity of this lipase. No change was found in the expression of diacylglycerol acyl transferase 1 (DGAT1) after training. Inhibition of HSL with a monospecific, small molecule inhibitor (76-0079) and stimulation of ATGL with CGI-58 revealed that significant ATGL activity is present in human skeletal muscle. These results suggest that ATGL in addition to HSL may be important for human skeletal muscle lipolysis.

Variability of triacylglycerol content in human skeletal muscle biopsy samples

1996 ◽  
Vol 81 (3) ◽  
pp. 1150-1155 ◽  
Author(s):  
P. S. Wendling ◽  
S. J. Peters ◽  
G. J. Heigenhauser ◽  
L. L. Spriet
Keyword(s):  
Skeletal Muscle ◽  
Human Skeletal Muscle ◽  
Vastus Lateralis ◽  
Vastus Lateralis Muscle ◽  
Exercise Time ◽  
Skeletal Muscle Biopsy ◽  
Time Points ◽  
Freeze Dried ◽  
Triacylglycerol Content ◽  
Rest And Exercise

The variability of the triacylglycerol store in human skeletal muscle (TGm) was examined using the needle biopsy technique. In 13 subjects, three biopsies were sampled from the vastus lateralis muscle of one leg at rest and after 90 min of cycling at 65% of maximal O2 uptake on one or two occasions. Visible fat and blood were removed before the samples were frozen, and remaining blood, connective tissue, and fat were removed from freeze-dried fiber bundles. TGm content was measured in two aliquots of powdered muscle from each biopsy. Within-biopsy variability was low at 6%. Despite precautions, many biopsies from inactive subjects were contaminated with adipose tissue. The TGm between-biopsy coefficient of variation (CV) was 23.5 +/- 14.6% (SD, n = 24) for rest and exercise time points where three noncontaminated biopsies existed. The between-biopsy variability at rest (19.8 +/- 7.9%, n = 10) was not significantly different from that at exercise (26.1 +/- 17.4%, n = 14). The muscle glycogen between-biopsy CV for rest and exercise time points was 10.0 +/- 10.3%. The resting TGm content was 26.3 +/- 4.3 mmol/kg dry muscle, and the net utilization during the 90 min of exercise was less than the between-biopsy variability. It is concluded that the TGm store measured in repeated biopsies of human skeletal muscle is variable, with a CV of 20-26%. Therefore, because of this high variability, only changes greater than approximately 24% of resting TGm content may be considered meaningful.

Intramuscular triacylglycerol utilization in human skeletal muscle during exercise: is there a controversy?

2002 ◽  
Vol 93 (4) ◽  
pp. 1185-1195 ◽  
Author(s):  
Matthew J. Watt ◽  
George J. F. Heigenhauser ◽  
Lawrence L. Spriet
Keyword(s):  
Skeletal Muscle ◽  
Muscle Biopsy ◽  
Human Skeletal Muscle ◽  
Moderate Intensity ◽  
Resonance Spectroscopy ◽  
Moderate Exercise ◽  
Moderate Intensity Exercise ◽  
Trained Subjects ◽  
Skeletal Muscle Biopsy ◽  
Isotope Tracer

Intramuscular triacylglyerols (IMTGs) represent a potentially important energy source for contracting human skeletal muscle. Although the majority of evidence from isotope tracer and 1H-magnetic resonance spectroscopy (MRS) studies demonstrate IMTG utilization during exercise, controversy regarding the importance of IMTG as a metabolic substrate persists. The controversy stems from studies that measure IMTG in skeletal muscle biopsy samples and report no significant net IMTG degradation during prolonged moderate-intensity (55–70% maximal O2 consumption) exercise lasting 90–120 min. Although postexercise decrements in IMTG levels are often reported from direct muscle measurements, the marked between-biopsy variability (∼23%) that has been reported with this technique in untrained subjects is larger than the expected decrease in IMTG content, effectively precluding significant findings. In contrast, recent data obtained in endurance-trained subjects demonstrated reduced variability between duplicate biopsies (∼12%), and significant changes in IMTG were detected after 120 min of moderate-intensity exercise. Therefore, it is our contention that the muscle biopsy, isotope tracer, and 1H-MRS techniques report significant and energetically important oxidation of free fatty acids derived from IMTGs during prolonged moderate exercise.

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