Calzia 2015 Abstract MiP2015
|Mitochondrial respiration in homogenized small tissue biopsies from the M. Vastus lateralis of patients with Huntington's Disease before and after cycling exercise.
Mitochondrial respiration is assumed to be severely affected by the presence of mutant huntingtin in HD patients. However, mitochondrial function remains difficult to be quantified in-vivo. Therefore, we used minimal volume tissue biopsies (4-8 mg) obtained from the m. vastus lateralis of HD subjects (mutation carriers) who volunteered to participate to our study, for quantifying mitochondrial respiration by means of the high-resolution respirometry before and after a standardized cycling exercise.
All patients gave written consent to participate to our study; the study protocol has been approved by the ethical committee of our institution. The tissue samples homogenates were put into the chambers of the O2K®-Oxygraph (Oroboros Instruments, Austria) and continuously stirred at 37°C. Mitochondrial respiration was quantified by adding complex I (10 mM Pyruvate, 5 mM Malate, and 10 mM Glutamate) and complex II (10 mM Succinate) substrates in the medium containing the homogenate and 5 mM ADP. Then 5 µM oligomycine was added to inhibit the ATP-synthase in order to obtain the LEAK-respiration state as an indicator of mitochondrial coupling. This step was followed by the addition of 1 µM of the uncoupler FCCP in order to achive the maximum respiration in the uncoupled state and the coupling (LEAK/ET-pathway)-ratio. After blocking mitochondrial respiration by 0.5 µM rotenone and 5 µM Antimycine A, the complex IV activity was selectively quantified by adding 2 mM Ascorbate and 0,5 mM TMPD. Here we present preliminary data from the first 3 patients included into the study.
A typical experiment as well as the preliminary data obtained yet in healthy controls and HD mutation carriers are presented in the figures below.
Our preliminary data do not allow definitive conclusions yet but they suggest that mitochondrial respiration can be reliably quantified in minimal volume needle biopsies from the m. vastus lateralis of human subjects. This test may therefore be used to quantify mitochondrial dysfunction as well as the effects of physical training during the course of the disease.
• O2k-Network Lab: DE Ulm Radermacher P
Labels: MiParea: Respiration, Exercise physiology;nutrition;life style, Patients Pathology: Other
Organism: Human Tissue;cell: Skeletal muscle
Coupling state: LEAK, OXPHOS, ET Pathway: N, S, ROX HRR: Oxygraph-2k Event: E1, Oral MiP2015
1-Ulm Univ Hospital, Dept Neurology; 2-Division Sports Rehabilitation Med, Dept Internal Medicine II; 3-Dept Anesthesiological Pathophysiology Process Development, Ulm, Germany. - [email protected]