Clinically Relevant Oxygraphic Assay to Assess Mitochondrial Energy Metabolism in Acute Myeloid Leukemia Patients
Abstract
:Simple Summary
Abstract
1. Introduction
2. Materials and Methods
2.1. Chemicals
2.2. Blast Collection and Cell Culture Conditions
2.3. Oxygen Consumption Rate (OCR) Measurements
2.4. High-Content Imaging
2.5. Cytofluorometric Analysis
2.6. Statistical Analysis
3. Results
3.1. Minimum Number of Blasts Required to Measure OXPHOS
3.2. Sequential Injections of Inhibitors to Measure OXPHOS
3.3. Optimization of FCCP Concentrations Required to Reach the Maximal OCR
3.4. Cytokine Supplementation Is Necessary to Maintain Mitochondrial Metabolism during Culture
3.5. Cryopreservation Slightly Alters Mitochondrial Metabolism of Blasts
3.6. Assessment of OXPHOS in Blasts from Bone Marrow
4. Discussion
5. Conclusions
Supplementary Materials
Author Contributions
Funding
Institutional Review Board Statement
Informed Consent Statement
Data Availability Statement
Acknowledgments
Conflicts of Interest
References
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Patient | Sex | Age | FAB | ELN | Cytogenetic Karyotype |
---|---|---|---|---|---|
AML#1 | F | 88 | M5a | 3 | 47, XX, +6, +8, del(9)(q21), −12, +13, del(17)(p11) |
AML#2 | M | 65 | M2 | N.D. | 46, XY, t(3;3)(q21;q26.2), der(16) t(1;16)(q21;q24) |
AML#3 | F | 75 | M1 | 2 | 46, XX, add(9)(q3?) |
AML#4 | M | 69 | M0 | 3 | 46XY, Isochomosomy 11, del(17), t(17,21) |
AML##5 | M | 78 | M2 | 3 | 45, XY, t(3;3)(q21;q26), −7 |
AML#6 | M | 65 | N.D. | 3 | 46,XY,del(7)(q12q36)/47,XY,+21/46,XY |
AML#7 | M | 70 | M1 | 2 | 46,XY |
AML#8 | M | 50 | M1 | 2 | 46,XY |
AML#9 | M | 71 | M5a | 3 | 47,XY,t(1;14)(p32;q32),del(7)(q22q34),der(7)t(7;11)(q34;q22),+13 |
AML#10 | F | 49 | N.D. | N.D. | N.D. |
AML#11 | M | 38 | s-AML | NA | N.D. |
AML#12 | F | 46 | M2 | 1 | 46,XX,t(8;21)(q22;q22)/45,sl,-X,del(9)(q22) |
AML#13 | M | 54 | M1 | 1 | 46,XY |
AML#14 | F | 67 | M1 | 2 | 46,XX |
AML#15 | F | 58 | N.D. | 2 | 46,XX |
AML#16 | M | 34 | N.D. | 3 | 45,XY,−7,−12,+mar |
AML#17 | F | 86 | M4 | 3 | 44,X,-X,del(4)(q21),−8,add(9)(p24),add(11)(p15),−16,−17,−12,+19,−20,add(21)(p13),+mar |
AML#18 | F | 67 | N.D. | 2 | 46,XX,i(7)(p10)/46,XX |
AML#19 | M | 69 | M4 | 3 | 46,XY,t(7;21), RUNX1 |
AML#20 | F | 86 | N.D. | 3 | 46,XX,−6,+8, del(7p) add17p |
AML#21 | M | 30 | N.D. | N.D. | N.D. |
AML#22 | M | 67 | M3 | 1 | 46,XY,t(15;17)(q24;q21)/46,XY, PML-RARα |
AML#23 | M | 59 | M4 | 2 | 46,XY, EVI1 overexpression |
AML#24 | F | 90 | N.D. | N.D. | 46,XX |
AML#25 | M | 72 | M4 | 1 | 46,XY,inv(16)(p13q22) |
AML#26 | F | 81 | M4 | 2 | 46,XX |
AML#27 | F | 63 | M5a | 3 | 48,XX,+8,t(9;11)(p22;q23),+ider(9)(p10).ish t(9;11)(3’KMT2A+;5’KMT2A+),ider(9)(3’KMT2Ax2), KMT2A-MLLT3, EVI1 overexpression |
Settings | Cycles | Mix | Wait | Measure | Total Duration |
---|---|---|---|---|---|
Basal | 3 | 2 min 40 s | 2 min | 2 min | 20 min |
Oligomycin (2 µM) | 2 | 1 min 40 s | 2 min | 2 min | 11 min 20 s |
FCCP1 (0.27–1.1 µM) or BAM15 (0.06–2 µM) | 2 | 1 min 40 s | 2 min | 2 min | 11 min 20 s |
FCCP2 (0.55–2.2 µM) or BAM15 (0.13–2.5 µM) | 2 | 1 min 40 s | 2 min | 2 min | 11 min 20 s |
Antimycin A + Rotenone (1 µM each) | 2 | 1 min 40 s | 1 min 20 s | 2 min | 10 min |
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Fovez, Q.; Laine, W.; Goursaud, L.; Berthon, C.; Germain, N.; Degand, C.; Sarry, J.-E.; Quesnel, B.; Marchetti, P.; Kluza, J. Clinically Relevant Oxygraphic Assay to Assess Mitochondrial Energy Metabolism in Acute Myeloid Leukemia Patients. Cancers 2021, 13, 6353. https://doi.org/10.3390/cancers13246353
Fovez Q, Laine W, Goursaud L, Berthon C, Germain N, Degand C, Sarry J-E, Quesnel B, Marchetti P, Kluza J. Clinically Relevant Oxygraphic Assay to Assess Mitochondrial Energy Metabolism in Acute Myeloid Leukemia Patients. Cancers. 2021; 13(24):6353. https://doi.org/10.3390/cancers13246353
Chicago/Turabian StyleFovez, Quentin, William Laine, Laure Goursaud, Celine Berthon, Nicolas Germain, Claire Degand, Jean-Emmanuel Sarry, Bruno Quesnel, Philippe Marchetti, and Jerome Kluza. 2021. "Clinically Relevant Oxygraphic Assay to Assess Mitochondrial Energy Metabolism in Acute Myeloid Leukemia Patients" Cancers 13, no. 24: 6353. https://doi.org/10.3390/cancers13246353