Quantification of human ooplasmic mitochondria

Abstract

It is likely that there is an association between the fitness of mitochondria and their ability to support normal cellular function. Oocytes are greatly enriched in the number of mitochondria as they support essential developmental processes such as oocyte maturation and embryonic development, while their replication is deferred until gastrulation. The mitochondrial DNA (mtDNA) copy number in 87 human oocytes from 29 patients was evaluated after DNA extraction and real-time quantitative polymerase chain reaction (PCR). The average mtDNA copy number was 795,000 (+/- 243,000) in metaphase II oocytes. mtDNA content varied considerably between oocytes, even within the same patient. No relationship was found between mtDNA copy number and maternal age. The findings suggest that mtDNA replication is fully accomplished by the germinal vesicle stage in the fully developed oocyte.

Characterization of mitochondrial replication and transcription control during rat early development in vivo and in vitro

“…Ovarian hyperstimulation does not change the mtDNA copy number in metaphase II oocytes and the resultant embryos. This is a natural outcome, because fully grown germinal vesicle oocytes, responsible for eCG, have already accomplished mtDNA replication (Barritt et al 2002). In bovine oocytes, we observed a slight decrease in the mtDNA copy number during the in vitro maturation process .…”

Lost in the zygote: the dilution of paternal mtDNA upon fertilization

“…With an average of 3.67 Â 10 5 mtDNAs per mammalian oocyte (Steuerwald et al, 2000;Reynier et al, 2001;Barritt et al, 2002;Chan et al, 2005;May-Panloup et al, 2005a, b;Santos et al, 2006), the estimated mtDNA content per salmon oocyte is approximately three to four orders of magnitude higher than that reported for mammals. This finding is in agreement with stereological analyses estimating the number of mitochondria in toad and frog oocytes, indicating the mitochondrial DNA content of external fertilizers to exceed that of mammalian oocytes by at least three orders of magnitude (Romek and Krzysztofowicz, 2005;Shoubridge and Wai, 2007).…”

“…Gametes of mice were estimated to contain 10-75 and 1.6 Â 10 5 mtDNA molecules per single sperm and oocyte, respectively (Hecht et al, 1984;Shitara et al, 2000;Steuerwald et al, 2000). Human sperm were estimated to contain 1.4-6.8 molecules (MayPanloup et al, 2003;Amaral et al, 2007) and oocytes 1.93-7.95 Â 10 5 mtDNAs (Steuerwald et al, 2000;Reynier et al, 2001;Barritt et al, 2002;Chan et al, 2005;May-Panloup et al, 2005a;Santos et al, 2006). These estimates lead to ratios of 1:5.7 Â 10 5 -1:2.8 Â 10 4 of paternal-to-maternal mtDNA in human and 1:1.6 Â 10 4 -1:2.1 Â 10 3 in mouse zygotes if paternal mtDNA enters the egg upon fertilization.…”

Mitochondria in human oogenesis and preimplantation embryogenesis: engines of metabolism, ionic regulation and developmental competence

“…With this method, between 120 000 and 350 000 mitochondria have been estimated to occur in MII human oocytes (Jansen 2000b, Cummins 2002. However, when mtDNA copy numbers are determined by polymerase chain reaction methodology using probes for specific genes such as ATPase 6 (Van Blerkom 2004) or directed to specific sequences (Brenner 2004), the number of human mitochondrial genomes in MII oocytes from the same or different cohort(s) has been reported to differ by well over an order of magnitude, ranging from a low of approximately 20 000 to well over 800 000 , Steuerwald et al 2000, Reynier et al 2001, Barrit et al 2002, Van Blerkom 2004. If the current consensus that each oocyte mitochondrion contains a single genome is accurate (Cummins 2002), this rather astonishing and unexpected variation in mtDNA numbers between similarly appearing oocytes raises some fundamental questions about how competence may be determined well before fertilization.…”