ReviewMitochondrial function in the human oocyte and embryo and their role in developmental competence
Introduction
In the context of the likelihood that a fertilized egg will develop to term, it often comes as a surprise to those not involved in the study of early human development that reproduction in our species is an inherently inefficient process (Edwards, 1986). However, evidence of high frequencies of embryo demise during the pre- and early post-implantation stages of development, and fetal demise during the early months of pregnancy, have long suggested that over half of normally fertilized oocytes will never progress to birth and for women of advanced reproductive age (>~ 39), this frequency is significantly higher (Boue et al., 1975, Burgoyne et al., 1991, Wilcox et al., 1988, Van Blerkom, 1994). Cytogenetic analyses of tens of thousands of human oocytes and preimplantation stage embryos obtained by in vitro fertilization (IVF) over that the last 30 years consistently demonstrate high frequencies of developmentally lethal aneuploidies that arise during the preovulatory resumption of meiosis, and severe chromosomal segregation disorders that occur during the initial cleavage (mitotic) divisions. Current estimates suggest that ~ 40% of meiotically mature (metaphase II, MII) oocytes obtained for IVF from women ~ 35 years of age or younger may be aneuploid, and that this frequency increases progressively and markedly thereafter (Angell, 1994, Munné et al., 1995, Kushnir and Frattarelli, 2009).
In clinical IVF, management of the menstrual cycle (termed, controlled ovarian hyperstimulation) typically involves downregulation of endogenous gonadotropin production followed by the administration of gonadotropins (FSH, LH) at supraphysiological levels to stimulate the growth of multiple follicles, with ovulation induction initiated when follicles are deemed fully developed. It is commonly believed that the frequency of meiotically immature oocytes, mature oocytes (metaphase II, MII) with developmentally lethal aneuploidies, and a large proportion of developmentally incompetent embryos with major chromosomal defects, is increased with ovarian hyperstimulation when compared to levels observed in natural (mono-ovulatory) cycles. While this increase is usually attributed to adverse intrafollicular physiology or biochemistry associated with exogenous stimulation, the identification of defects in specific molecular processes or signaling pathways that adversely compromise developmental competence has remained elusive (for reviews: Van Blerkom and Trout, 2007, Van Blerkom, 2009a, Van Blerkom, in press).
The development of selection schemes to identify competent human oocytes and embryos has been a common theme in IVF research since it was first introduced for infertility treatment over 30 years ago, and until recently, morphological characteristics of embryo performance in vitro have been the primary determinants. More recently, various types of high-resolution spectrophometric methods that provide a molecular profile of the follicular fluid to which the oocyte is exposed prior to ovulation, and molecular signatures of metabolites in spent embryo culture medium, have been introduced as potential noninvasive means for competence assessment. However, the power of these analytical methods has yet to be demonstrated by significant improvements in outcome (Van Blerkom, in press). The need for a more detailed understanding of the etiologies of maturation and fertilization failure for the oocyte, and early demise for the embryo is not merely one of academic curiosity. Indeed, it has become a necessity in some counties owing to legal limitations on the number of oocytes that can be inseminated and embryos returned to the patient, and in some instances, the stage of embryogenesis at which transfers can be performed (Benagiano and Gianaroli, 2004, Gerris and DeSutter, 2009). Where no legal mandates exist, many IVF programs have imposed similar limits on embryo transfers. These limitations are part of a collective, worldwide consensus in the clinical IVF field that the potential for higher order gestations has to be reduced. One of the foremost trends in this regard has been a transition from the transfer of multiple preimplantation stage embryos to the replacement of a single embryo. However, morphological, behavioral and biochemical characteristics can be of limited predictive value even when culture is extended, as evidenced by multiple gestations with embryos characterized as stage-inappropriate at the time of transfer, or where performance in vitro was considered suboptimal and likely inconsistent with an ongoing pregnancy. The limited predictive value of current competence assessments has stimulated new avenues of investigation based on the cell biology of the human oocyte and its relationship to the heterogeneous developmental ability of the human embryo. It is in this context that mitochondria have emerged as a major focus of research.
Section snippets
Mitochondria as determinants of developmental competence for the human oocyte and embryo
Recent studies of the origins of developmental competence in the human oocyte and its maintenance during early embryogenesis have taken a very different research direction from those which consider intrafollicular influences at the antral stage to be primary factors. These investigations are testing the assumption that idiopathic fertilization failure and embryo demise during the preimplantation stages have a common etiology that can be traced to intrinsic defects in the oocyte. A current theme
Mitochondrial “ageing” in human oocytes
Mitochondrial swelling and disruption of cristae appear to be a common feature of the oocytes of women of advanced reproductive age (≥ 40 years of age; Muller-Hocker et al., 1996). In these instances, swelling may be caused by loss of control of volume homeostasis related to the inability of mitochondria to maintain a ΔΨm that is consistent with normal levels of ATP generation, as discussed below. High frequencies of premature meiotic arrest (i.e., prior to MII) after ovulation induction,
Mitochondrial genetics, reactive oxygen species production and developmental competence
In clinical IVF, the notion that mitochondria have a central role in the normality of oocyte and early embryonic development has been generally considered in terms of their universal respiratory function as the primary source of ATP and therefore, as drivers of morphodynamic processes and stage-specific bioactivities during oogenesis and embryogenesis. This is not an unexpected assumption given that the mitochondrial complement present in the oocyte at fertilization is derived from a progenitor
Lifestyle factors and mitochondrial function in oocytes
The possibility that some proportion of infertility or chronic embryo/fetal demise is associated with mtDNA defects that are lifestyle-related, such as tobacco smoking, has been an underlying concern in evaluation of the efficacy of clinical treatments when negative outcomes occur repeatedly. For men, tobacco smoking can be a very significant factor with subfertility resulting from (i) the depressive effects of reduced (sperm) mitochondrial respiration on motility (Chohan and Badawy, 2010),
Mitochondrial complement size and oocyte bioenergetics
Despite growing evidence of a critical role as a primary driver of developmental competence for the oocyte and early embryo, it is surprising that certain fundamental issues concerning mitochondria and their activity during these stages remain to be resolved. This is especially true for the human, where bioenergetic deficiencies have been suggested to correlate with oocyte-specific mtDNA copy number and may be a central factor in failures of oocyte maturation and fertilization, and embryo
ATP thresholds, mtDNA degradation and disproportionate mitochondrial inheritance during early development
That mitochondrial respiration is a central determinant of developmental competence in the mature oocyte and newly fertilized egg is based on the notion that bioenergetic deficits that drop net cytoplasmic ATP levels below a stage specific threshold likely compromise the progression of oocyte maturation and preimplantation embryogenesis. It should be noted that few studies have examined the association between stage specific ATP thresholds and clinical outcomes with respect to the normality of
ΔΨm and cytoplasmic microzonation
As discussed above, the current emphasis of mitochondrial research in early human development has focused primarily on characterizations of mtDNA copy number and bioenergetic levels that are consistent with normal development at different stages of oocyte maturation and preimplantation embryogenesis in vitro. However, an awareness of other important functions and roles mitchonndria have in somatic cells, such signal transduction, maintenance of calcium homeostasis, and apoptosis (Pozzan et al.,
Stage specific spatial remodeling of mitochondria
In certain species, such as the mouse and pig, preovulatory maturation of the oocyte is clearly accompanied by stage-specific cytoplasmic remodeling. In these species, mitochondria not associated with the subplasmalemmal domain form small aggregates that, between the germinal vesicle breakdown and circular bivalent stages, translocate to the nuclear region (Van Blerkom and Runner, 1985, Tokura et al., 1993; Van Blerkom et al., 2002; Sun et al., 2001, Van Blerkom, 2009a, Van Blerkom, 2009b). In
Clinical correlates: mitochondria and developmental competence
While microzonation, functional compartmentalization, and dynamic stage specific changes in mitochondrial distribution and activity are not new concepts in cell biology, there is an emerging view that they may be important forces in the developmental biology of the early mammalian embryo that could have important clinical implications if regulatory functions are perturbed. For example, defects in the organization or function of the SER could perturb the dynamic regulation of the ATP
Therapeutic strategies in clinical IVF to overcome presumed mitochondrial dysfunctions and known pathogenic mtDNA mutations
The potential contribution of mitochondria to the normality of early development has had direct clinical implications in the derivation of novel treatments intended to improve outcomes and eliminate the inheritance of pathogenic mtDNA mutations. A clinical therapy in which a small volume of ooplasm from presumably normal oocytes was transferred to the oocytes of infertile women whose fertilized eggs, on previous IVF attempts, repeatedly experienced high levels of lethal fragmentation during the
Mitochondria regulation of redox state
Unlike the oocytes of other species such as the sea urchin, where mitochondrial activity has a regulatory function in development by virtue of their influence redox-sensitive signal transduction pathways, such as the determination axial polarities and cell fate (Coffman et al., 2004, Coffman, 2009), a similar role has not been considered for the mammal oocyte and early embryo, until recently. Mitochondrial regulation of the redox state (Cao and Chen, 2009) that effect redox-sensitive
Conclusion
Despite over 30 years of experience with human IVF, contemporary practice is far from an exact science and formidable challenges to improved outcome remain. Current clinical and experimental evidence indicate the central importance of mitochondria in the developmental biology of the mammalian oocyte and preimplantation stage embryo. However, the specific means by which mitochondrial defects or bioenergetic deficiencies influence outcome has only recently become the subject of investigation,
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