Maternal obesity is well known to have negative impacts on fertility. We have identified a number of reproductive developmental defects that arise from high fat diet induced obesity in the mouse system, which are summarized in . Of embryos expected to be at the two-cell stage from obese mothers, ~65% are abnormal being either degraded or remaining at the 1 cell stage. Of the two-cell stage embryos of normal morphology, only 50% proceed to the blastocyst stage. Thus there is an early embryonic loss of ~80% in obese mothers, suggesting an oocyte maternal effect. Analysis of oocytes at the MII stage indicates that ~45% show significant spindle or chromosome misalignment defects, which are likely to generate embryos with massive aneuploidy. However, the ~45% of MII oocytes with meiotic abnormalities does not fully account for the ~80% of zygotes that fail to reach the blastocyst stage, suggesting that there is an additional defective process that results in early embryonic developmental abnormalities. The nature of this additional likely oocyte maternal effect awaits further investigation.
Summary of developmental phenotypes observed in oocytes and embryos from HFD obese mice.
Previously we found that mice fed a HFD prior to and throughout pregnancy had fetuses that were growth retarded, with the live-born pups significantly smaller at birth 
. Here we used blastocyst transfers to demonstrate that growth retardation, as well as brain developmental abnormalities, were not a result of an obese uterine environment. Instead, the defect must occur prior to the blastocyst stage, possibly also as early as the oocyte stage. Importantly the blastocysts used in the transfer assay had normal morphology and implanted and established pregnancy similar to controls, suggesting that the defective processes that leads to the fetal growth retardation and brain abnormalities is distinct from the defects that lead to the early embryonic abnormalities. Thus we propose that the HFD obese maternal environment leads to at least three abnormal reproductive developmental processes which are likely oocyte maternal effects: meiotic aneuploidy that contributes to early embryonic loss; a defect distinct from meiotic aneuploidy that leads to early embryonic loss; and fetal growth retardation and brain developmental abnormalities, which must originate from defects prior to the blastocyst stage. Determining if the fetal abnormalities are due to an oocyte maternal effect or a problem that arises during cleavage divisions will require transfers of one-cell zygotes from HFD mothers into control diet mothers; however this is technically challenging given the high rate of pre-blastocyst lethality. The basis of the fetal abnormalities are not known, although we note that a failure to maintain paternal imprints in the early embryo could result in growth retardation phenotypes.
Obese women commonly deliver macrosomic infants, and thus our model would appear to be contradictory. Two points, however, need to be clarified. First, mice are delivered at a much less mature stage than humans. This has been shown in the development of the immune system, respiratory system as well as neural-sensory system. Thus, birth weight in a mouse may reflect an earlier developmental timepoint than in human neonates. We have found that these smaller fetuses from mothers fed a high fat diet throughout pregnancy catch up within 21 days of birth and exceed their aged match controls, perhaps reflecting the larger size human newborn. Secondly, among women with preexisting severe and morbid obesity, a significant percentile delivers growth retarded or small for gestation age [SGA, birthweight below the 10th percentile] infants 
. These women are believed to have more central adiposity and the prognosis for these infants is much more grave than the standard obese patients 
. A recent Dutch study reported an overall incidence of SGA at 18% in obese women, significantly higher than 10% in the general population 
The brain abnormalities we observed are especially significant in light of results from several recent human studies suggesting that maternal obesity predisposes their infants to a greater risk of neurodevelopmental delay 
and atypical neurodevelopment 
than those born to healthy, lean women. Furthermore, a recent review examining 12 studies that explored the connection between maternal obesity and cognitive, behavioral, and emotional problems in offspring and supported the conclusion that the offspring of obese women may be at increased risk of behavioral and cognitive deficits in childhood, as well as eating disorders in adolescence and psychotic disorders in adulthood 
. In response to such findings, the Institute of Medicine has highlighted neurodevelopment as an important potential long-term consequence of gestational weight gain that needs further investigation 
. Our study suggests that preconceptional weight gain adversely affects pregnancy outcomes and fetal development. Further studies into the behavioral phenotypes of these offspring will strengthen this hypothesis. In light of our findings, preconceptional counseling may be indicated as the preferable, earlier target for intervention in obese women desiring pregnancy and healthy outcomes.
We observed that oocytes from mice on a high fat diet displayed abnormal mitochondrial metabolism as assessed by citrate levels in individual denuded oocytes. Oocytes from HFD mice also had significantly higher numbers of abnormal appearing mitochondria, characterized by intra-organelle vacuoles, swollen cristae and less dense matrix as compared to control diet fed mice. Moreover, as has been shown to occur in other mtDNA rich cells such as cardiomyocytes and skeletal muscle, increased reactive oxygen species led to induction of specific transcription factors in order to increase mitochondrial biogenesis, evident by elevated mtDNA copy number and increased accumulation of proteins responsible for mitochondrial biogenesis and fission, consistent with results reported by Igosheva et al 
. Cumulus cells from the same high fat fed mice also demonstrated abnormal mitochondrial ultrastructure. It has been established in several systems that high fat feeding results in changes in mitochondrial membrane fatty acid composition, and it has been suggested that these changes lead to significant alterations in uncoupling proteins and energy metabolism 
. By extension, our study would be the first to suggest that altered mitochondrial membrane fatty acid composition may occur in the mammalian oocyte - cumulus complex and that this perturbation can manifest as early as after one month of the diet. It will be interesting to revisit the maternal HFD model to examine redox state in the mitochondria and for how long these metabolic changes persist following fertilization and embryonic development in a non-obese environment.
Skeletal muscle is a major model for study the effects of HFD and diabetes on physiology, cellular and mitochondrial function 
. Similar to oocytes, the HFD induces abnormal mitochondrial function and ultrastructural abnormalities in skeletal muscle including swelling, disarrayed cristae and reduced matrix 
. However, unlike the oocyte, in skeletal muscle there is decrease in mitochondria number and decreased expression of mitochondrial gene products, including PGC1α 
. This distinct mitochondrial biogenesis response to the HFD likely reflects the different biological functions of the two cell types.
Previously, we found disordered spindles and misaligned chromosomes in MII oocytes from mouse models of type 1 diabetes 
, which also display abnormal mitochondrial metabolism and ultrastructure. It is thus tempting to speculate that the deficits in mitochondrial function are the cause of the meiotic spindle and chromosome alignment defects. Although the mechanistic basis might be similar, the proportions of oocytes exhibiting abnormal chromosome alignment, aberrant spindle formation and aneuploidy were much higher in this model of HFD compared to the hyperglycemic type1 diabetic model 
. This difference may be caused by hormonal or environmental differences, such as the level of oxidative stress, which could be more severe in this HFD model of obesity and type 2 diabetes 
Our finding that there are three abnormal reproductive developmental processes in a HFD mouse model of obesity informs future studies on reduced fertility and congenital abnormalities observed in obese woman, which is an increasing proportion of the population. Our proposal that the reproductive developmental defects may arise from an oocyte maternal effect is consistent with IVF studies where the increased pregnancy failure rate in obese women returns to the normal rate if donor oocytes are used instead of autologous oocytes 
. Our four-week HFD obesity model suggests that the oocyte defects arise in the antral follicles that are recruited in the latest cycle, exposed to the highest effect of the HFD environment. This suggests potential therapeutic routes for reversing the oocyte reproductive obesity effects through diet and exercise.