During the pre-implantation development, bovine embryos require and/or are susceptible to different sources of energy substrates according to their stage. More specifically, the embryo from the zygote to the 8–16 cell stage is mainly dependent on pyruvate and lactate. Later, it acquires the capacity to use glucose, which will be efficiently metabolized from the blastocyst stage on [1]. In natural conditions, the organ responsible for creating the ideal environment for early embryo development is the oviduct. Specifically in bovine species, this early embryo development period in the oviduct takes about 4–5 days after fertilization, which contains the embryonic critical transition moment from pyruvate/lactate to glucose as the main energy source [1]. The oviductal lumen holds a complex fluid, which is composed essentially of transudate from blood plasma and components produced by the oviductal cells [2]. This oviductal fluid (OF) is constantly renewed in an estrous cycle stage and oviductal segment-dependent ways. In cattle, the greatest OF secretion rate occurs during the peri-ovulatory stage in the ampulla segment [5]. By using the approach of in vivo cannulation of the oviduct, Hugentobler et al. [3] showed that the oviduct is capable of creating a specifically controlled environment for the embryo necessities, by decreasing glucose concentration in 70 % and increasing lactate concentration to 900 % in the OF compared to the blood plasma [3]. Even though the mechanisms and/or factors that might interfere with the OF energy substrate composition are still poorly known, yet, it is clear that the oviductal cells have a specific mechanism for controlling glucose metabolism and its metabolites in the OF.

Not enough studies have been able to perform cannulation procedures of the oviduct to expand the knowledge related to OF formation and the processes involved in this [3]. Moreover, some concerns are involved with this procedure, such as the possibility of increasing inflammatory processes in the oviduct as well as the ethical use of animal welfare, which is also a principle that restricts in vivo/ex vivo studies. Analyzing the oviduct in vivo faces some limitations, mainly due to its difficult in situ access and its highly tortuous anatomy [4], resulting in the slaughter of the animal to access the oviduct. Altogether, the study of the oviduct under in vitro conditions is highly practiced. However, when considering the culture media composition for oviductal cells, we noticed that a wide and not standardized range of glucose levels has been applied in the studies. Compared to the glucose levels in the circulatory blood conditions (6.1–7.7 mM glucose [3]), many studies are submitting the oviductal cells to higher concentrations of glucose (for instance, when using DMEM/Ham's F12 media, which has 17.5 mM glucose [5,6] or DMEM high glucose, which has 25 mM glucose [7]). Moreover, compared to the local oviductal glucose level, the scenario becomes even more discrepant. As demonstrated by Hugentobler et al. [3], the glucose concentration in the oviductal lumen is lower than the plasmatic blood (2.7 mM glucose [3]). Therefore, we consider that the glucose metabolism should be better understood in the oviductal cell in vitro culture.

Glucose can be metabolized through different metabolic pathways. After entering the cellular cytoplasm, it is converted by hexokinases to glucose-6P, which can follow the glycolysis pathway, resulting in pyruvate production, or it can be driven to nucleotides production through the pentose phosphate pathway (PPP). The pyruvate generated by the glycolytic pathway can either be converted to lactate, in the cellular cytoplasm or to acetyl-coenzyme A (acetyl-CoA). The latter can occur in the cytoplasm or by entering the mitochondria, which can follow the tricarboxylic acid (TCA) cycle for energy production [8]. Besides energy production, cellular metabolism has been exponentially associated with cellular epigenetic control changes. Known as metaboloepigenetic, this crosstalk has been explored in more dynamic tissues such as embryonic development (reviewed by Ref. [9]), tumoral cells, and stem cells (reviewed by Ref. [10]). Among the epigenetics modifications, the most studied is DNA methylation, however, the histone modifications are the most variable [9]. The addition of the methyl group to the DNA cytosine molecules results in the formation of 5-methylcytosines (5 mC, [11]), which, in general, is associated with transcription repression, genomic imprinting, and post-translational histone modifications [12,13]. This epigenetic modification interacts, for example, with the metabolism of the one-carbon, which generates as a product the S-Adenosyl methionine (SAM), the primary methyl donor for DNA methylation [14]. On the contrary, some histone modification, such as the histone H3 acetylation at lysine 9 (H3K9ac), is associated with active gene transcription status [15,16]. The insertion and removal of the acetyl group are done by enzymes named histone acetyltransferases (HATs) and histone deacetylases (HDACs), respectively [17]. For that, the acetyl group present in acetyl-coenzyme A (acetyl-CoA) is a central metabolic intermediate, which is generated in mitochondria as a byproduct of both glucose and lipid metabolism (β-oxidation) [18]. Even though acetyl-CoA level is a determinant for histone acetylation, it remains controversial if this is a specific or a global response [9].

All things considered, we hypothesized that the metaboloepigenetic mechanism might be related to the oviductal cell functionality and, consequently, the OF energy substrate composition. Therefore, our objective was to identify the existence of a relationship between metabolism and epigenetic alterations in the oviductal epithelial cells under in vitro culture conditions. To do that, we selected glucose as a metabolic disturbed factor to evaluate two epigenetic markers (5 mC and H3K9ac). Because the oviductal metabolism might be affect by a cycle stage-dependent way, the oviductal epithelial cells were harvested at two stages: follicular stage (high estradiol – E2 – and low progesterone – P4) and mid-luteal stage (low E2 and high P4).