High resolution respirometry to assess function of mitochondria in native homogenates of human heart muscle

Impaired myocardial bioenergetics is a hallmark of many cardiac diseases. There is a need of a simple and reproducible method of assessment of mitochondrial function from small human myocardial tissue samples. In this study we adopted high-resolution respirometry to homogenates of fresh human cardia...

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Bibliographic Details
Published in:PloS one 2020-01, Vol.15 (1), p.e0226142-e0226142
Main Authors: Krajčová, Adéla, Urban, Tomáš, Megvinet, David, Waldauf, Petr, Balík, Martin, Hlavička, Jan, Budera, Petr, Janoušek, Libor, Pokorná, Eva, Duška, František
Format: Article
Language:English
Subjects:
ADP
Adult
Aged
Atria
Bioenergetics
Biology and Life Sciences
Brain death
Brain research
Cardiac muscle
Citrate (si)-Synthase - metabolism
Coefficient of variation
Consent
Coronary artery disease
Cryopreservation
Cytochrome
Cytochrome c
Cytochromes
Energy Metabolism
Enzymes
Fatty Acids - metabolism
Female
Funding
Heart diseases
Heart surgery
High resolution
Hospitals
Humans
Intensive care
Laboratories
Malate
Male
Medicine
Medicine and Health Sciences
Metabolism
Microscopy
Middle Aged
Mitochondria
Mitochondria, Heart - metabolism
Mitochondrial Membranes - metabolism
Muscles
Myocardium
Oligomycin
Organ donors
Oxidation-Reduction
Oxidative phosphorylation
Oxygen
Oxygen - metabolism
Oxygen consumption
Phosphorylation
Physiology
Research and Analysis Methods
Respirometry
Rotenone
Sodium azide
Sodium azides
Surgery
Ventricle
Ventricles (cerebral)
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Summary:Impaired myocardial bioenergetics is a hallmark of many cardiac diseases. There is a need of a simple and reproducible method of assessment of mitochondrial function from small human myocardial tissue samples. In this study we adopted high-resolution respirometry to homogenates of fresh human cardiac muscle and compare it with isolated mitochondria. We used atria resected during cardiac surgery (n = 18) and atria and left ventricles from brain-dead organ donors (n = 12). The protocol we developed consisting of two-step homogenization and exposure of 2.5% homogenate in a respirometer to sequential addition of 2.5 mM malate, 15 mM glutamate, 2.5 mM ADP, 10 μM cytochrome c, 10 mM succinate, 2.5 μM oligomycin, 1.5 μM FCCP, 3.5 μM rotenone, 4 μM antimycin and 1 mM KCN or 100 mM Sodium Azide. We found a linear dependency of oxygen consumption on oxygen concentration. This technique requires < 20 mg of myocardium and the preparation of the sample takes
ISSN:1932-6203
1932-6203
DOI:10.1371/journal.pone.0226142