Resilience of preimplanta0on bovine embryos to the availability of energy substrates

Exposure to metabolic stress during fetal life increases the suscep0bility to metabolic diseases in adulthood. This no0on is supported by epidemiological and experimental evidences that led to the theoriza0on of the Developmental Origins of Health and Disease. The adap0ve mechanisms the embryo/fetus puts in place to cope with intra-uterine stressors is defined developmental plas0city and entails the capacity of one genotype to generate several phenotypes in response to a different environment. To capture the main molecular events of developmental plas0city, we monitored the gene expression of bovine blastocysts exposed in vitro to a mild metabolic challenge. With preliminary experiments we standardized the experimental model to remove sources of variability, such as serum, embryos of different sex, and blastocyst at different stages. Then metabolic challenges were administered by varying the content in energe0c substrates of the culture medium. Three energe0c levels, containing 0.5, 1.0, and 1.5-fold increase in energe0c substrates, were selected based on the absence of apparent changes in preimplanta0on embryo development, evaluated by blastocyst rate, distribu0on of blastocyst morphology (early, expanded, hatch-ed/ing), number of cells composing the blastocyst, and paWern of cell lineage specifica0on. Genome-wide analysis revealed minimal differences in gene expression among the three groups, likely exposing key regulatory genes whose differen0al expression allowed the adapta0on to the changing metabolic environment. Two dis0nct expression paWerns were observed for these genes: progressive upregula0on and progressive downregula0on along with the increasing energe0c availability. The lack of substan0al differences seems in line with the general observa0on that offspring born aZer a mild intra-uterine exposure to metabolic stress have normal physiological and biochemical parameters un0l later in life, while a generalized disrup0on of gene expression would probably impact embryo/fetal/early post-natal life rather than induce a late onset of the disease. Nevertheless, if metabolic stress experienced during preimplanta0on development were to commit a late phenotype, some kind of mark shall be established at this stage. A possible answer to this ques0on came from the analysis of transcript isoforms. Using a specific bioinforma0c pipeline, the presence of two or more transcript isoforms of genes related to epigene0c changes and nuclear reprogramming were detected, indica0ng that, even in absence of obvious changes in gene expression, the metabolic challenge induced biological effects that can be epigene0cally encoded in the embryo. As a proof of concept, acetyla0on of histone proteins increased when the energe0c substrates were higher. These findings shed light on the mechanisms at the onset of developmental plas0city, whereby the ac0va0on/repression of few key genes and usage of transcript isoforms confer resilience to metabolic stressors and provide a direct link between changes in the availability of energe0c substrates and epigene0c reprogramming. Funded by: MUR PRIN2017, No. 20172N2WL3_002; Piano di Sostegno alla Ricerca UNIMI: Linea 2 – Azione A