Metabolic Checkpoint for Gametes and Embryo ReproductiveCompetence

Selectivity is a main feature of animal and human reproductiveprocesses. In male gametogenesis, hundreds of millions ofsperm are released in a single ejaculate whereas only one isnecessary to achieve a fertilization. In females, a wide cohort ofprimordial follicles is activated at each ovarian cycle with onlya few or a single one (e.g. humans) reaching the final maturationand reproductive competence. In human assis ted reproduction,the in vitro cultured embryos undergo a s trict selection anda large majority of them arres t their development before reachingthe blas tocys t s tage. Mechanisms explaining these arres tsinclude a failure of the activation of the zygote genome, excessof oxygen radicals and endoplasmic reticulum s tress and aneuploidy.These failures can be explained by a deficit of metabolismin supporting the chromatin re-modeling, the bioenergeticsand the redox balance, all of them dependent on the function ofthe one carbon metabolism (OCM).The OCM is the epicenter of cellular metabolism as providingthe activated carbon units for transmethylations and the reducingequivalents (glutathione – GSH) for the redox balance. Activatedmethyl groups are necessary for the epigenetic regulationof DNA and his tones, to regulate protein functions and tosus tain key cellular syntheses including carnitine, creatine andCoQ۱۰. GSH, besides dis tributing reducing power to the wholeendogenous antioxidant sys tem provides ROS neutralizationwithin mitochondria and is s trictly related to the energetic efficiency.In turn, the OCM is s trictly dependent on the feedof micronutrients from the diet and other environment signals,which fits with its role in epigenetics.In sperms, a massive and timed methylation of DNA and histonesis necessary for the protamine transition and for the finalnuclear compaction allowing resis tance to ROS generatedat time of swimming and acrosomal reaction. The adminis trationof methyl donors known to feed the OCM to ART-resis tantinfertile men resulted in a significant improvement of spermnuclear maturation, which correlated with the achieved pregnancies.In vitro, we showed that micronutrients in support tothe OCM supported the mitochondrial function of alive humansperms. Oocytes necessitate a huge reserve of methylationcapacity to exert the DNA repair function and the epigeneticre-methylation of the whole genome of the zygote. Theadminis tration of micronutrients to ladies with very low ovarianreserve appeared to re-activate the release of AMH and togenerate spontaneous pregnancies. These effects vouched foran improved oocyte quality as well as to a better priming totheir pos t-fertilization job.To further elucidate the underlying mechanisms, more recentlywe used an in vitro maturation model of bovine oocytes to investigate the effect of a micronutrient support on their developmentalpotential. Sibling bovine oocytes were randomlyassigned to ۲۴-hour in vitro maturation with or without the additionto the culture medium of micronutrients at physiologicconcentrations. These micronutrients included methyl donors(methylfolate, methylcobalamin, betaine), a cys teine donor (Lcystine) and cofactors for the involved enzymes (vit.s B۲, B۳,B۶ and zinc). There were no differences in the MII rate betweenthe groups of oocytes, however the supplemented oocytesshowed lower DNA fragmentation, and higher mitochondrialmass and DNMT۳a protein expression. The fertilization rateswere as well overlapping, but zygotes from supplemented oocytesexerted a far larger methylation of the female pro-nuclei,confirming the boos t to the methylation metabolism from micronutrients.Thereafter, the rate of maturation to blas tocys t thatwas doubled in zygotes from supplemented oocytes. In summary,the short (۲۴ hours) exposure of oocytes to the metabolicenhancers within the critical time window of final maturation,although exerting no effect on their maturation and fertilizationrate, was already enough to dramatically improve their potentialto develop.Based on these evidences, we propose the occurrence of anOCM-based metabolic check point for the selection of the bes tgametes and embryos. In the context of evolution, this allowsthe mothers-to-be to further inves t in reproductive efforts onlyon those gametes and embryos exerting the genetic/metabolicpattern better fitting with the available environment/feed. In thecase of our bovine embryos the conditioning environment wasthe culture medium and its enrichment allowed a larger rate ofoocytes to produce competent zygotes and viable blas tocys ts.The same is likely to happen in gametogenesis: Only the bes tsuited follicle/oocyte(s) will be ovulated after an ovarian cycleand only the bes t suited sperm will be able to swim fas ter and inthe right direction to achieve the fertilization.These concepts are of paramount importance to unders tand themetabolic demand of reproduction and to clarify the role of dietand of dietary integration is support to fertility as well as to improvethe yield from in vitro generation of embryos.