Sjoevall 2013 Mitochondrion

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SjΓΆvall F, Ehinger JK, Marelsson SE, Morota S, Asander Frostner E, Uchino H, Lundgren J, ArnbjΓΆrnsson E, Hansson Magnus J, Fellman V, ElmΓ©r E (2013) Mitochondrial respiration in human viable platelets - methodology and influence of gender, age and storage. Mitochondrion 13:7-14.

Β» PMID: 23164798 Open Access

Sjoevall F, Ehinger JK, Marelsson SE, Morota S, Asander Frostner E, Uchino H, Lundgren J, Arnbjoernsson E, Hansson Magnus J, Fellman V, Elmer E (2013) Mitochondrion

Abstract: Studying whole cell preparations with intact mitochondria and respiratory complexes has a clear benefit compared to isolated or disrupted mitochondria due to the dynamic interplay between mitochondria and other cellular compartments. Platelet mitochondria have a potential to serve as a source of human viable mitochondria when studying mitochondrial physiology and pathogenic mechanisms, as well as for the diagnostics of mitochondrial diseases. The objective of the present study was to perform a detailed evaluation of platelet mitochondrial respiration using high-resolution respirometry. Further, we aimed to explore the limits of sample size and the impact of storage as well as to establish a wide range of reference data from different pediatric and adult cohorts. Our results indicate that platelet mitochondria are well suited for ex-vivo analysis with the need for minute sample amounts and excellent reproducibility and stability. β€’ Keywords: Diagnostics; Human; Platelets; Mitochondria; OXPHOS; High-resolution respirometry; Mitochondrial disorders

β€’ O2k-Network Lab: SE Lund Elmer E, JP Tokyo Uchino H

Coupling control and the Q-junction

Mitochondrial coupling control states are measured without simultaneous change of a selected pathway control state, i.e. coupling control is separated from pathway control. Biochemical coupling efficiencies (E-L coupling efficiencies) and P-L coupling efficiencies are, therefore, studied at a defined pathway control state that must not change between measurement of LEAK respiration L, OXPHOS capacity P, and electron transfer capacity E.
A physiologically relevant pathway control state for partial reconstitution of TCA cycle function is obtained by supply of NADH-linked substrates (e.g. pyruvate&malate PM; N-pathway) in combination with succinate (S; S-pathway), supporting convergent electron transfer through Complexes I and II into the Q-junction (NS-pathway). OXPHOS- and ET-capacities are higher in the combined NS-pathway than in the separate N- or S-pathway (Gnaiger 2020). Is the NS-pathway control state appropriate for the analysis of coupling control?
Partial additivity in OXPHOS capacity NSP or ET capacity NSE implies that there is competition between the N- and S-pathway, when the NS-pathway capacity is less than the arithmetic sum of the constituent pathway capacities. In mitochondria with lower OXPHOS than ET capacity (P<E; when the phosphorylation system is limiting), the competition in NSE is increasingly pronounced in NSP, and when respiration is further reduced by complete inhibition of the phosphorylation system (e.g. by oligomycin), competition between the N- and S-pathways is maximal in LEAK respiration. Different levels of competition imply that the ratio of the effective N- and S-pathway in the NS-pathway state may shift to the extent that the dominant pathway may fully outcompete the other in the LEAK state. Convergent electron input into the Q-junction in NSE, therefore, may shift to single electron input through either the dominant N- or S-pathway in NSL, which then would effectively correspond to either NL or SL. This has deep implications on LEAK respiration, since the N-pathway has three coupling sites (H+ pumps: CI, CIII, CIV) with a correspondingly higher H+/O2 ratio compared to the S-pathway with two coupling sites (H+ pumps: CIII, CIV). A higher rate of the proton leak is implied when measuring the same rate of LEAK respiration in NL than when observing an identical oxygen consumption rate in SL.
When inhibiting O2 consumption by oligomycin in the NS-pathway state, the relative contribution of the N- and S-pathways to LEAK respiration is not known. By subsequent uncoupler titrations, the relative contribution of these pathways is likely to change, thus obtaining an undefined combination of pathway control and coupling control. In conclusion, the NS-pathway state is not appropriate for studying coupling control. Coupling control is best studied in the separate N- or S-pathway (Gnaiger et al 2000; 2015).
  1. Gnaiger E (2020) Mitochondrial pathways and respiratory control. An introduction to OXPHOS analysis. 5th ed. Bioenerg Commun 2020.2.
  2. Gnaiger E, Boushel R, SΓΈndergaard H, Munch-Andersen T, Damsgaard R, Hagen C, DΓ­ez-SΓ‘nchez C, Ara I, Wright-Paradis C, Schrauwen P, Hesselink M, Calbet JAL, Christiansen M, Helge JW, Saltin B (2015) Mitochondrial coupling and capacity of oxidative phosphorylation in skeletal muscle of Inuit and caucasians in the arctic winter.
  3. Gnaiger E, MΓ©ndez G, Hand SC (2000) High phosphorylation efficiency and depression of uncoupled respiration in mitochondria under hypoxia. Proc Natl Acad Sci U S A 97:11080-5.


Β» Bioblast_alert_2014(02)

Labels: MiParea: Respiration, Instruments;methods, mt-Medicine, Patients  Pathology: Aging;senescence, Neurodegenerative  Stress:Ischemia-reperfusion  Organism: Human  Tissue;cell: Blood cells, Platelet  Preparation: Permeabilized cells, Intact cells 

Coupling state: LEAK, ROUTINE, OXPHOS, ET  Pathway: N, ROX  HRR: Oxygraph-2k 

SE, JP, MitoEAGLE blood cells data 

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