Mitochondrial function, quantity and DNA integrity in human oocytes at different stages of maturation

Abstract

OBJECTIVE: Human oocytes significantly vary in pregnancy potential. Differences in energy metabolism may contribute to this variation. We investigated ATP content and mitochondrial DNA (mtDNA) copy number and integrity in individual human oocytes at different stages of maturation.DESIGN: Experimental study.MATERIALS AND METHODS: A total of 41 oocytes obtained from 12 women 25-38 years of age during IVF cycles were examined. 22 were at prophase 1 (P1), 11 were at metaphase 1 (M1) and 6 were at the M2 stage, whereas 2 were degenerating. The main outcome parameters were ATP content and mtDNA copy number; a secondary outcome was mtDNA integrity, as assessed by the existence of the 5 Kb “common” mtDNA deletion. A chemiluminsecent luciferase/luciferin reaction was utilized to measure ATP content, whereas real-time PCR was used to quantify the total mtDNA copy number. Nested PCR was applied to detect the 5 Kb mtDNA deletion.RESULTS: The mean ATP content was 774 ± 418 fmol 977 ± 364 fmol , and 1957 ± 516 fmol for P1, M1 and M2 oocytes respectively (p=0.003 for P1-M2, p=0.025 for M1-M2, p=1.0 for P1-M1). Total mtDNA copy numbers were similar among the various stages of oocyte maturation (57,116 ± 83,834, 323,372 ± 521,503 and 77,834 ± 115,755 for P1, M1 and M2 oocytes, respectively). mtDNA copy number correlated positively with the duration of incubation between oocyte retrieval and lysis for analysis (r=0.43, p=0.01); whereas ATP content did not. There was no effect of woman's age on ATP content and mtDNA copy number. However, we observed the presence of the common deletion in the mtDNA in 6 (4 M1, 2 degenerating) of the 22 oocytes tested.CONCLUSIONS: ATP content increased with oocyte developmental progress from the P1 to M2 stage. In addition, mitochondrial number increased in parallel to the duration of in vitro culture. We speculate that these changes prepare the oocyte for an optimized mitochondrial redistribution during early embryogenesis. OBJECTIVE: Human oocytes significantly vary in pregnancy potential. Differences in energy metabolism may contribute to this variation. We investigated ATP content and mitochondrial DNA (mtDNA) copy number and integrity in individual human oocytes at different stages of maturation. DESIGN: Experimental study. MATERIALS AND METHODS: A total of 41 oocytes obtained from 12 women 25-38 years of age during IVF cycles were examined. 22 were at prophase 1 (P1), 11 were at metaphase 1 (M1) and 6 were at the M2 stage, whereas 2 were degenerating. The main outcome parameters were ATP content and mtDNA copy number; a secondary outcome was mtDNA integrity, as assessed by the existence of the 5 Kb “common” mtDNA deletion. A chemiluminsecent luciferase/luciferin reaction was utilized to measure ATP content, whereas real-time PCR was used to quantify the total mtDNA copy number. Nested PCR was applied to detect the 5 Kb mtDNA deletion. RESULTS: The mean ATP content was 774 ± 418 fmol 977 ± 364 fmol , and 1957 ± 516 fmol for P1, M1 and M2 oocytes respectively (p=0.003 for P1-M2, p=0.025 for M1-M2, p=1.0 for P1-M1). Total mtDNA copy numbers were similar among the various stages of oocyte maturation (57,116 ± 83,834, 323,372 ± 521,503 and 77,834 ± 115,755 for P1, M1 and M2 oocytes, respectively). mtDNA copy number correlated positively with the duration of incubation between oocyte retrieval and lysis for analysis (r=0.43, p=0.01); whereas ATP content did not. There was no effect of woman's age on ATP content and mtDNA copy number. However, we observed the presence of the common deletion in the mtDNA in 6 (4 M1, 2 degenerating) of the 22 oocytes tested. CONCLUSIONS: ATP content increased with oocyte developmental progress from the P1 to M2 stage. In addition, mitochondrial number increased in parallel to the duration of in vitro culture. We speculate that these changes prepare the oocyte for an optimized mitochondrial redistribution during early embryogenesis.