Difference between revisions of "SUIT-015"
From Bioblast
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::: '''[[Categories of SUIT protocols |SUIT-category]]:''' FNS(Oct,PGM) | ::: '''[[Categories of SUIT protocols |SUIT-category]]:''' FNS(Oct,PGM) | ||
::: '''[[SUIT protocol pattern]]:''' diametral | ::: '''[[SUIT protocol pattern]]:''' diametral 1OctM;2D;3G;4P;5S;6U;7Rot | ||
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|sort=Was published in year | |sort=Was published in year | ||
|order=descending | |order=descending | ||
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{{Template:SUIT-0XX}} | |||
== Strengths and limitations == | |||
:::+ The protocol provides information on FAO capacity in the absence of other, potentially interfering pathways, both in the LEAK state and in OXPHOS. | |||
:::+ FNS OXPHOS capacity comprises the most important pathways in many cell types and thus provides a physiologically relevant estimate of maximum mitochondrial respiratory capacity. | |||
:::+ FNS ET capacity is a good estimate of overal ET capacity in may cell types. | |||
:::+ Application of the cytochrome ''c'' test early in the protocol ensures comparability of all states in case of any effect of ''c''. | |||
:::+ Reasonable duration of the experiment. | |||
:::- SRot(E) may be underestimated if S is not saturating. | |||
:::- CIV activity is not measured, to save experimental time. | |||
:::- GM and PM yield typically identical fluxes in human skeletal muscle fibres. However, PM is the superior alternative to GM: the fraction of the N-pathway is lower and of the S-pathway is higher with GM compared to PM (GM<sub>''P''</sub> is inhibited by the CII inhibitor malonic acid to a larger extent than PM<sub>''P''</sub>). PM, therefore, yields a more sensitive assay for the diagnosis of injuries in the N-pathway, since an impairment of N-pathway capacity can be compensated partially by activation of the S-pathway. This is a disadvantage compared to SUIT-004 and SUIT-008 for diagnosis of N-capacity. | |||
:::- To detect an additive effect of P after GM<sub>''P''</sub>, pyruvate would have to be added as step 3 (before S). However, inhibition of respiration was observed after titration of P (5 mM) in horse skeletal muscle fibres (Votion et al 2012), which was not the case when P was titrated in steps of 1 mM. | |||
:::- When evaluating the additive effect of the N- and S-pathway, it has to be considered that NS<sub>''P''</sub>- and NS<sub>''E''</sub>-capacities can only be compared with N<sub>''P''</sub>- and S<sub>''E''</sub>-capacities. This is not a problem when NS<sub>''P''</sub> = NS<sub>''E''</sub> (Gnaiger 2009). Otherwise, it may be assumed that S<sub>''P''</sub> = S<sub>''E''</sub> (Votion et al 2012), such that NS<sub>''P''</sub> can be compared with N<sub>''P''</sub> + S<sub>''P''</sub>. SUIT-004 should be chosen for the additive effect in the ET-state. | |||
:::- ''Rox'' may be lower in substrate states earlier in the SUIT protocol. Therefore, this ''Rox'' measurement is frequently taken as a methodological control rather than as the final basis of ''Rox'' correction of mitochondrial respiration (mt). | |||
:::- Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler. | |||
== Compare SUIT protocols == | |||
::::* GM and PM yield typically identical fluxes in human skeletal muscle fibres. | |||
::::* [[SUIT-004]] 1PM;2D;3U;4S;5Rot- | |||
::::* [[SUIT-008]] 1PM;2D;3G;4S;5U;6Rot- | |||
::::* [[1PM;2D;3U;4G;5S;6Oct;7Rot;8Gp-]] | |||
::::* [[1PGM;2D;3S;4U;5Rot-]] | |||
[[File:1GM;2D;3S;3c;4U;5Rot-.jpg|300px]] 1GM;2D;3S;3c;4U;5Rot;6Ama | |||
{{MitoPedia concepts | |||
|mitopedia concept=SUIT protocol, SUIT A | |||
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Revision as of 12:28, 14 January 2019
Description
Abbreviation: FNS(Oct,PGM)
Reference: A Schoepf 2016 FEBS J
MitoPedia concepts:
SUIT protocol,
SUIT B
- SUIT-category: FNS(Oct,PGM)
- SUIT protocol pattern: diametral 1OctM;2D;3G;4P;5S;6U;7Rot
References
Year | Reference | Organism | Tissue;cell | |
---|---|---|---|---|
Schoepf 2016 FEBS J | 2016 | Schöpf B, Schäfer G, Weber A, Talasz H, Eder IE, Klocker H, Gnaiger E (2016) Oxidative phosphorylation and mitochondrial function differ between human prostate tissue and cultured cells. https://doi.org/10.1111/febs.13733 | Human | Endothelial;epithelial;mesothelial cell Genital Other cell lines Fibroblast |
Strengths and limitations
- + The protocol provides information on FAO capacity in the absence of other, potentially interfering pathways, both in the LEAK state and in OXPHOS.
- + FNS OXPHOS capacity comprises the most important pathways in many cell types and thus provides a physiologically relevant estimate of maximum mitochondrial respiratory capacity.
- + FNS ET capacity is a good estimate of overal ET capacity in may cell types.
- + Application of the cytochrome c test early in the protocol ensures comparability of all states in case of any effect of c.
- + Reasonable duration of the experiment.
- - SRot(E) may be underestimated if S is not saturating.
- - CIV activity is not measured, to save experimental time.
- - GM and PM yield typically identical fluxes in human skeletal muscle fibres. However, PM is the superior alternative to GM: the fraction of the N-pathway is lower and of the S-pathway is higher with GM compared to PM (GMP is inhibited by the CII inhibitor malonic acid to a larger extent than PMP). PM, therefore, yields a more sensitive assay for the diagnosis of injuries in the N-pathway, since an impairment of N-pathway capacity can be compensated partially by activation of the S-pathway. This is a disadvantage compared to SUIT-004 and SUIT-008 for diagnosis of N-capacity.
- - To detect an additive effect of P after GMP, pyruvate would have to be added as step 3 (before S). However, inhibition of respiration was observed after titration of P (5 mM) in horse skeletal muscle fibres (Votion et al 2012), which was not the case when P was titrated in steps of 1 mM.
- - When evaluating the additive effect of the N- and S-pathway, it has to be considered that NSP- and NSE-capacities can only be compared with NP- and SE-capacities. This is not a problem when NSP = NSE (Gnaiger 2009). Otherwise, it may be assumed that SP = SE (Votion et al 2012), such that NSP can be compared with NP + SP. SUIT-004 should be chosen for the additive effect in the ET-state.
- - Rox may be lower in substrate states earlier in the SUIT protocol. Therefore, this Rox measurement is frequently taken as a methodological control rather than as the final basis of Rox correction of mitochondrial respiration (mt).
- - Careful washing is required after the experiment to avoid carry-over of inhibitors and uncoupler.
Compare SUIT protocols
- GM and PM yield typically identical fluxes in human skeletal muscle fibres.
- SUIT-004 1PM;2D;3U;4S;5Rot-
- SUIT-008 1PM;2D;3G;4S;5U;6Rot-
- 1PM;2D;3U;4G;5S;6Oct;7Rot;8Gp-
- 1PGM;2D;3S;4U;5Rot-
MitoPedia concepts: SUIT protocol, SUIT A