Search the repository

A+ | A- | | SLO | ENG

Abstract: Background: We recently demonstrated that following a 10-day exposure to inactivity/simulated microgravity impairments ofoxidative metabolism were located ‘upstream’ of mitochondrial function, as evaluated by maximal ADP-stimulated mitochon-drial respiration (JO 2max) determined ex vivo. The aim of this study was to evaluate mitochondrial sensitivity to submaximal[ADP] by an alternative approach aimed at identifying responses associated with fibre type composition.Methods: Isolated permeabilized vastus lateralis fibres were analysed by high-resolution respirometry in 9 young males beforeand after a 10-day horizontal bed rest. Eleven submaximal titrations of ADP (from 12.5 to 10 000 μM) were utilized to assess com-plex I + II-linked ADP sensitivity. We applied to JO 2 versus [ADP] data a traditional Michaelis–Menten kinetics equation, withthe calculation of the apparent K m and maximal respiration (Vmax), and two ‘sequential’ hyperbolic equations, yielding two K mand Vmax values. The two-hyperbolic equations were solved and the [ADP] value corresponding to 50% of JO 2max was calculated.Isoform expression of myosin heavy chains (MyHC) 1, 2A and 2X was also determined. Control experiments were also carriedout on rat skeletal muscle samples with different percentages of MyHC isoforms.Results: The two hyperbolic equations provided an alternative fitting of data and identified two distinct phases of the JO 2 versus[ADP] response: a first phase characterized by low Vmax (Vmax1 , 28 ± 10 pmol s−1 mg−1) and apparent K m (K m1 , 62 ± 54 μM) anda second phase characterized by higher Vmax (Vmax2 , 61 ± 16 pmol s−1 mg−1) and K m (K m2 , 1784 ± 833 μM). Data were confirmedin control experiments carried out in rat muscle samples with different percentages of MyHC isoforms. Correlation and receiveroperating characteristics analyses suggest that the two phases of the response were related to the % of MyHC isoforms.Conclusions: A novel mathematical approach (two sequential hyperbolic functions) for the fitting of JO 2 versus [ADP] data ob-tained by high-resolution respirometry on permeabilized skeletal muscle fibres, obtained in humans and rats, provided an alter-native fitting of the experimental data compared to the traditional Michaelis–Menten kinetics equation. This alternative modelallowed the identification of two distinct phases in the responses, which were related to fibre type composition. A first phase, characterized by low apparent K m and Vmax values, was correlated with the percentage of less oxidative (Type 2A + 2X) MyHCisoforms. A second phase, characterized by high apparent K m and Vmax , was related to more oxidative (Type 1) MyHC isoforms.
Keywords: ADP, bed rest, mitochondrial sensitivity, myosin heavy chains, skeletal muscle mitochondria
Published in DiRROS: 05.05.2025; Views: 530; Downloads: 298
Full text (3,59 MB)
This document has many files! More...