Difference between revisions of "Pham 2014 Am J Physiol"
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|journal=Am J Physiol | |journal=Am J Physiol | ||
|abstract=As approximately 80% of diabetics die from heart failure, understanding diabetic cardiomyopathy is crucial. Mitochondria occupy 35-40% of mammalian cardiomyocyte volume, supply 95% of the hearts' ATP, and diabetic heart mitochondria show impaired structure, arrangement and function. We predict that bioenergetic efficiencies are present in diabetic heart mitochondria; therefore we explored mitochondrial proton and electron handling by linking oxygen flux within streptozotocin (STZ)-induced-diabetic Sprague-Dawley rat heart tissues, to steady-state ATP synthesis, Reactive Oxygen Species (ROS) production, and mitochondrial membrane potential (Δ''Ψ''). By coupling high-resolution respirometers with purpose-built fluorometers, we followed Magnesium Green (ATP synthesis), Amplex Ultra Red (ROS production), and safranin-O (Δ''Ψ''). Relative to control rats, the mass-specific respiration of STZ-diabetic hearts was depressed in oxidative phosphorylating (OXPHOS) states. Steady-state ATP synthesis capacity was almost a third lower in STZ-diabetic heart and relative to O2 flux, this equates to an estimated 12% depression in OXPHOS efficiency. However, with anoxic transition, STZ-diabetic and control heart tissues showed similar ATP hydrolysis capacities through reversal of the F1/F0 ATP-synthase. STZ-diabetic cardiac mitochondria also produced more net ROS relative to oxygen flux (ROS/O) in OXP. While Δ''Ψ'' did not differ between groups, the time to develop Δ''Ψ'' with the onset of OXPHOS was protracted in STZ-diabetic mitochondria. ROS/O is higher in life-like OXPHOS states and potential delays in the time to develop Δ''Ψ'' may delay ATP synthesis with inter-beat fluctuations in ADP concentrations. Whereas diabetic cardiac mitochondria produce less ATP in normoxia, they consume as much ATP in anoxic infarct-like states. | |abstract=As approximately 80% of diabetics die from heart failure, understanding diabetic cardiomyopathy is crucial. Mitochondria occupy 35-40% of mammalian cardiomyocyte volume, supply 95% of the hearts' ATP, and diabetic heart mitochondria show impaired structure, arrangement and function. We predict that bioenergetic efficiencies are present in diabetic heart mitochondria; therefore we explored mitochondrial proton and electron handling by linking oxygen flux within streptozotocin (STZ)-induced-diabetic Sprague-Dawley rat heart tissues, to steady-state ATP synthesis, Reactive Oxygen Species (ROS) production, and mitochondrial membrane potential (Δ''Ψ''). By coupling high-resolution respirometers with purpose-built fluorometers, we followed Magnesium Green (ATP synthesis), Amplex Ultra Red (ROS production), and safranin-O (Δ''Ψ''). Relative to control rats, the mass-specific respiration of STZ-diabetic hearts was depressed in oxidative phosphorylating (OXPHOS) states. Steady-state ATP synthesis capacity was almost a third lower in STZ-diabetic heart and relative to O2 flux, this equates to an estimated 12% depression in OXPHOS efficiency. However, with anoxic transition, STZ-diabetic and control heart tissues showed similar ATP hydrolysis capacities through reversal of the F1/F0 ATP-synthase. STZ-diabetic cardiac mitochondria also produced more net ROS relative to oxygen flux (ROS/O) in OXP. While Δ''Ψ'' did not differ between groups, the time to develop Δ''Ψ'' with the onset of OXPHOS was protracted in STZ-diabetic mitochondria. ROS/O is higher in life-like OXPHOS states and potential delays in the time to develop Δ''Ψ'' may delay ATP synthesis with inter-beat fluctuations in ADP concentrations. Whereas diabetic cardiac mitochondria produce less ATP in normoxia, they consume as much ATP in anoxic infarct-like states. | ||
|keywords=Diabetic cardiomypathy, Anoxia, Efficiency, Mitochondria, Oxidative phosphorylation, Safranin, Amplex Red, Magnesium | |keywords=Diabetic cardiomypathy, Anoxia, Efficiency, Mitochondria, Oxidative phosphorylation, Safranin, Amplex Red, Magnesium Green | ||
|mipnetlab=NZ Auckland Hickey AJ | |mipnetlab=NZ Auckland Hickey AJ | ||
}} | }} | ||
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|substratestates=CI, CI&II, ROX | |substratestates=CI, CI&II, ROX | ||
|instruments=Oxygraph-2k, O2k-Fluorometer | |instruments=Oxygraph-2k, O2k-Fluorometer | ||
|additional=Safranin, Amplex Red, Magnesium Green | |||
}} | }} | ||
Demonstration of using Amplex Red, Safranin and Magnesium green methods with the O2k-Fluoresececen Module. | Demonstration of using Amplex Red, Safranin and Magnesium green methods with the O2k-Fluoresececen Module. |
Revision as of 11:55, 1 April 2016
Pham T, Loiselle D, Power A, Hickey AJ (2014) Mitochondrial inefficiencies and anoxic ATP hydrolysis capacities in diabetic rat heart. Am J Physiol 307:C499–507. |
Pham T, Loiselle D, Power A, Hickey AJ (2014) Am J Physiol
Abstract: As approximately 80% of diabetics die from heart failure, understanding diabetic cardiomyopathy is crucial. Mitochondria occupy 35-40% of mammalian cardiomyocyte volume, supply 95% of the hearts' ATP, and diabetic heart mitochondria show impaired structure, arrangement and function. We predict that bioenergetic efficiencies are present in diabetic heart mitochondria; therefore we explored mitochondrial proton and electron handling by linking oxygen flux within streptozotocin (STZ)-induced-diabetic Sprague-Dawley rat heart tissues, to steady-state ATP synthesis, Reactive Oxygen Species (ROS) production, and mitochondrial membrane potential (ΔΨ). By coupling high-resolution respirometers with purpose-built fluorometers, we followed Magnesium Green (ATP synthesis), Amplex Ultra Red (ROS production), and safranin-O (ΔΨ). Relative to control rats, the mass-specific respiration of STZ-diabetic hearts was depressed in oxidative phosphorylating (OXPHOS) states. Steady-state ATP synthesis capacity was almost a third lower in STZ-diabetic heart and relative to O2 flux, this equates to an estimated 12% depression in OXPHOS efficiency. However, with anoxic transition, STZ-diabetic and control heart tissues showed similar ATP hydrolysis capacities through reversal of the F1/F0 ATP-synthase. STZ-diabetic cardiac mitochondria also produced more net ROS relative to oxygen flux (ROS/O) in OXP. While ΔΨ did not differ between groups, the time to develop ΔΨ with the onset of OXPHOS was protracted in STZ-diabetic mitochondria. ROS/O is higher in life-like OXPHOS states and potential delays in the time to develop ΔΨ may delay ATP synthesis with inter-beat fluctuations in ADP concentrations. Whereas diabetic cardiac mitochondria produce less ATP in normoxia, they consume as much ATP in anoxic infarct-like states. • Keywords: Diabetic cardiomypathy, Anoxia, Efficiency, Mitochondria, Oxidative phosphorylation, Safranin, Amplex Red, Magnesium Green
• O2k-Network Lab: NZ Auckland Hickey AJ
Labels: MiParea: Respiration, mt-Medicine
Pathology: Diabetes, Myopathy
Stress:Ischemia-reperfusion, Oxidative stress;RONS, Mitochondrial disease
Organism: Rat
Tissue;cell: Heart
Preparation: Homogenate
Regulation: Coupling efficiency;uncoupling, mt-Membrane potential Coupling state: LEAK, OXPHOS, ETS"ETS" is not in the list (LEAK, ROUTINE, OXPHOS, ET) of allowed values for the "Coupling states" property.
HRR: Oxygraph-2k, O2k-Fluorometer
Safranin, Amplex Red, Magnesium Green
Demonstration of using Amplex Red, Safranin and Magnesium green methods with the O2k-Fluoresececen Module.
Correction
An OROBOROS Oxygraph-2k was used in this publication, whereas the Anton Paar/OROBOROS Oxygraph was the first-generation instrument for high-resolution respirometry, which was replaced by the Oxygraph-2k in 2002.
- Further details: Gnaiger 2012 Abstract Bioblast-Gentle Science