Evaluation of mitochondrial respiratory function in small biopsies of liver

AV Kuznetsov, D Strobl, E Ruttmann… - Analytical …, 2002 - Elsevier
AV Kuznetsov, D Strobl, E Ruttmann, A Königsrainer, R Margreiter, E Gnaiger
Analytical biochemistry, 2002Elsevier
Mitochondrial respiratory function was studied in permeabilized pig liver biopsies. The cell
membrane was permeabilized mechanically in tissue samples of 2–7 mg, for application of a
standardized substrate/inhibitor titration protocol in high-resolution respirometry. Specific
respirometric tests demonstrated complete plasma membrane permeabilization and
accessibility of substrates to intact mitochondria. High respiratory adenylate control ratios
and cytochrome c conservation in the tissue preparation were comparable or even better …
Mitochondrial respiratory function was studied in permeabilized pig liver biopsies. The cell membrane was permeabilized mechanically in tissue samples of 2–7 mg, for application of a standardized substrate/inhibitor titration protocol in high-resolution respirometry. Specific respirometric tests demonstrated complete plasma membrane permeabilization and accessibility of substrates to intact mitochondria. High respiratory adenylate control ratios and cytochrome c conservation in the tissue preparation were comparable or even better than in isolated mitochondria. Citrate synthase and cytochrome c oxidase activities remained at 85% of controls after up to 98 h storage of liver tissue at 0°C in histidine–tryptophan–ketoglutarate solution. Multiple mitochondrial defects, however, were indicated after 48 h cold storage by the decline in respiratory capacity, which was lowered to a larger extent with complex I substrates compared to respiration with substrates for complex II or IV, measured in the absence of cytochrome c. After prolonged ischemia, the adenylate control ratio was significantly reduced, and cytochrome c depletion was detected by the stimulatory effect of cytochrome c. High-resolution respirometry allows the assessment of mitochondrial function in a few milligrams of permeabilized liver tissue, without isolation of mitochondria. This provides a basis for the analysis of mitochondrial function in human liver biopsies.
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