Assessing mitochondrial respiration in permeabilized fibres and biomarkers for mitochondrial content in human skeletal muscle

2022 ◽  
Author(s):  
Jujiao Kuang ◽  
Nicholas J Saner ◽  
Javier Botella ◽  
Matthew J‐C Lee ◽  
Cesare Granata ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Respiration ◽  
Human Skeletal Muscle ◽  
Mitochondrial Content

scholarly journals Methodological considerations when assessing mitochondrial respiration and biomarkers for mitochondrial content in human skeletal muscle

2021 ◽  
Author(s):  
Jujiao Kuang ◽  
Nicholas J Saner ◽  
Javier Botella ◽  
Matthew J-C Lee ◽  
Cesare Granata ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Respiration ◽  
Coefficient Of Variation ◽  
Human Skeletal Muscle ◽  
Citrate Synthase ◽  
Membrane Integrity ◽  
Mitochondrial Outer Membrane ◽  
Muscle Fibres ◽  
Biological Variability ◽  
Mitochondrial Content

Background: The assessment of mitochondrial respiration and mitochondrial content are two common measurements in the fields of skeletal muscle research and exercise science. However, to verify the validity of the observed changes in both mitochondrial respiration and mitochondrial content following an intervention such as exercise training, it is important to determine the reliability and reproducibility of the experimental design and/or techniques employed. We examined the repeatability of widely used methodologies for assessing mitochondrial respiration and mitochondrial content, respectively; the measurement of maximal mitochondrial oxidative phosphorylation in permeabilized muscle fibres using high-resolution respirometry, and the measurement of citrate synthase activity as a biomarker for mitochondrial content in a microplate with spectrophotometer. Result: For mitochondrial respiration, the coefficient of variation for repeated measurements using muscle sampled from same biopsy decreased from 12.7% to 11% when measured in triplicate with outliers excluded, rather than in duplicate. The coefficient of variation was 9.7% for repeated muscle biopsies sampled across two separated days. For measurements of citrate synthase activity, the coefficient of variation was 3.5% of three technical repeats on the same plate, 10.2% for duplicate analyses using the same muscle lysate when conducted in the same day, and 30.5% when conducted four weeks apart. Conclusion: We have provided evidence for important technical considerations when measuring mitochondrial respiration with human skeletal muscle: 1) the relatively large technical variability can be reduced by increasing technical repeats and excluding outliers; 2) the biological variability and absolute mitochondrial respiration value of the participants should be considered when estimating the required sample size; 3) a new threshold of 15% for the increase in respiration rate after the addition of cytochrome c test for testing mitochondrial outer membrane integrity. When analysing citrate synthase activity, our evidence suggests it is important to consider the following: 1) all samples from the same study should be homogenized and measured at the same time using the same batch of freshly made chemical reagents; 2) biological variability should be considered when detecting small change in mitochondrial content; 3) the relative change should be used to compare the outcomes from different studies.

Does impaired mitochondrial function affect insulin signaling and action in cultured human skeletal muscle cells?

2008 ◽  
Vol 294 (1) ◽  
pp. E97-E102 ◽  
Author(s):  
Audrey E. Brown ◽  
Matthias Elstner ◽  
Stephen J. Yeaman ◽  
Douglass M. Turnbull ◽  
Mark Walker
Keyword(s):  
Skeletal Muscle ◽  
Glucose Uptake ◽  
Insulin Signaling ◽  
Mitochondrial Respiration ◽  
Respiratory Function ◽  
Insulin Action ◽  
Human Skeletal Muscle ◽  
Human Skeletal Muscle Cells ◽  
Basal Glucose Uptake

Insulin-resistant type 2 diabetic patients have been reported to have impaired skeletal muscle mitochondrial respiratory function. A key question is whether decreased mitochondrial respiration contributes directly to the decreased insulin action. To address this, a model of impaired cellular respiratory function was established by incubating human skeletal muscle cell cultures with the mitochondrial inhibitor sodium azide and examining the effects on insulin action. Incubation of human skeletal muscle cells with 50 and 75 μM azide resulted in 48 ± 3% and 56 ± 1% decreases, respectively, in respiration compared with untreated cells mimicking the level of impairment seen in type 2 diabetes. Under conditions of decreased respiratory chain function, insulin-independent (basal) glucose uptake was significantly increased. Basal glucose uptake was 325 ± 39 pmol/min/mg (mean ± SE) in untreated cells. This increased to 669 ± 69 and 823 ± 83 pmol/min/mg in cells treated with 50 and 75 μM azide, respectively (vs. untreated, both P < 0.0001). Azide treatment was also accompanied by an increase in basal glycogen synthesis and phosphorylation of AMP-activated protein kinase. However, there was no decrease in glucose uptake following insulin exposure, and insulin-stimulated phosphorylation of Akt was normal under these conditions. GLUT1 mRNA expression remained unchanged, whereas GLUT4 mRNA expression increased following azide treatment. In conclusion, under conditions of impaired mitochondrial respiration there was no evidence of impaired insulin signaling or glucose uptake following insulin exposure in this model system.

scholarly journals Mitochondrial respiration variability and simulations in human skeletal muscle: The Gene SMART study

2020 ◽  
Vol 34 (2) ◽  
pp. 2978-2986 ◽  
Author(s):  
Macsue Jacques ◽  
Jujiao Kuang ◽  
David J. Bishop ◽  
Xu Yan ◽  
Javier Alvarez‐Romero ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Respiration ◽  
Human Skeletal Muscle ◽  
Smart Study

scholarly journals Training-Induced Changes in Mitochondrial Content and Respiratory Function in Human Skeletal Muscle

2018 ◽  
Vol 48 (8) ◽  
pp. 1809-1828 ◽  
Author(s):  
Cesare Granata ◽  
Nicholas A. Jamnick ◽  
David J. Bishop
Keyword(s):  
Skeletal Muscle ◽  
Respiratory Function ◽  
Human Skeletal Muscle ◽  
Mitochondrial Content ◽  
Induced Changes

scholarly journals Physical activity changes the regulation of mitochondrial respiration in human skeletal muscle

2002 ◽  
Vol 543 (1) ◽  
pp. 191-200 ◽  
Author(s):  
J. Zoll ◽  
H. Sanchez ◽  
B. N'Guessan ◽  
F. Ribera ◽  
E. Lampert ◽  
...  
Keyword(s):  
Physical Activity ◽  
Skeletal Muscle ◽  
Mitochondrial Respiration ◽  
Human Skeletal Muscle

The role of phosphorylcreatine and creatine in the regulation of mitochondrial respiration in human skeletal muscle

2001 ◽  
Vol 537 (3) ◽  
pp. 971-978 ◽  
Author(s):  
B Walsh ◽  
M Tonkonogi ◽  
K Soderlund ◽  
E Hultman ◽  
V Saks ◽  
...  
Keyword(s):  
Skeletal Muscle ◽  
Mitochondrial Respiration ◽  
Human Skeletal Muscle

scholarly journals Dietary restriction increases skeletal muscle mitochondrial respiration but not mitochondrial content in C57BL/6 mice

2012 ◽  
Vol 133 (1) ◽  
pp. 37-45 ◽  
Author(s):  
Sarah Hempenstall ◽  
Melissa M. Page ◽  
Katrina R. Wallen ◽  
Colin Selman
Keyword(s):  
Skeletal Muscle ◽  
Dietary Restriction ◽  
Mitochondrial Respiration ◽  
Mitochondrial Content
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