A fast method for high resolution oxymetry study of skeletal muscle mitochondrial respiratory chain complexes

作者：Vienne Jean Claude; Cimetta Catherine; Dubois Marie; Duburcq Thibault; Favory Raphael; Dessein Anne Frederique; Fontaine Monique; Curt Marie Joncquel Chevalier; Cuisset Jean Marie; Douillard Claire; Mention Mulliez Karine; Dobbelaere Dries; Vamecq Joseph^*

来源：Analytical Biochemistry, 2017, 528: 57-62.

DOI：10.1016/j.ab.2017.04.015

• Ultradispersion
• Saponin-permeabilized fibers
• Respiratory chain complex
• High resolution oxymetry
• FCCP
• Electron transfer system (ETS)
• Muscle needle biopsy
• MIR05 buffer

High resolution oxymetry study (HROS) of skeletal muscle usually requires 90-120 min preparative phase (dissection, permeabilization and washing). This work reports on the suitability of a rapid muscle preparation which by-passes this long preparation. For a few seconds only, muscle biopsy from pigs is submitted to gentle homogenization at 8000 rotations per minute using an ultra-dispersor apparatus. Subsequent HROS is performed using FCCP instead of ADP, compounds crossing and not plasma membrane, respectively. This simplified procedure compares favorably with classical (permeabilized fibers) HROS in terms of respiratory chain complex activities. Mitochondria from cells undergoing ultra dispersion were functionally preserved as attested by relative inefficacy of added cytochrome C (not crossing intact mitochondrial outer membrane) to stimulate mitochondrial respiration. Responsiveness of respiration to ADP (in the absence of FCCP) suggested that these intact mitochondria were outside cells disrupted by ultradispersion or within cells permeated by this procedure.

• 出版日期2017-7-1