Spermatogenesis is a complex and highly regulated process involving the proliferation, differentiation, and apoptosis of germ cells. This process is controlled by various hormonal, genetic, and environmental factors, including temperature. In hormonal regulation, follicle-stimulating hormone (FSH), luteinizing hormone (LH), and testosterone (T) are essential for correct spermatogenesis development from the early stages and spermatogonia proliferation to germ cell maturation. Other hormones, like inhibin and activin, finely participate tuning the process of spermatogenesis. Genetic regulation involves various transcription factors, such as SOX9, SRY, and DMRT1, which are crucial for the development and maintenance of the testis and germ cells. MicroRNAs (miRNAs) play a significant role by regulating gene expression post-transcriptionally. Epigenetic modifications, including DNA methylation, histone modifications, and chromatin remodelling, are also vital. Temperature regulation is another critical aspect, with the testicular temperature maintained around 2–4 °C below body temperature, essential for efficient spermatogenesis. Heat shock proteins (HSPs) protect germ cells from heat-induced damage by acting as molecular chaperones, ensuring proper protein folding and preventing the aggregation of misfolded proteins during thermal stress. Elevated testicular temperature can impair spermatogenesis, increasing germ cell apoptosis and inducing oxidative stress, DNA damage, and the disruption of the blood–testis barrier, leading to germ cell death and impaired differentiation. The cellular mechanisms of germ cell proliferation, differentiation, and death include the mitotic divisions of spermatogonia to maintain the germ cell pool and produce spermatocytes. Spermatocytes undergo meiosis to produce haploid spermatids, which then differentiate into mature spermatozoa. Apoptosis, or programmed cell death, ensures the removal of defective germ cells and regulates the germ cell population. Hormonal imbalance, genetic defects, and environmental stress can trigger apoptosis during spermatogenesis. Understanding these mechanisms is crucial for addressing male infertility and developing therapeutic interventions. Advances in molecular biology and genetics continue to uncover the intricate details of how spermatogenesis is regulated at multiple levels, providing new insights and potential targets for treatment.

Spermatogenesis is a complex and highly regulated process involving the proliferation, differ-entiation, and apoptosis of germ cells. This process is controlled by various hormonal, genetic, and environmental factors, including temperature. In Hormonal regulation, Follicle-Stimulating Hormone (FSH), Luteinizing Hormone (LH) and testosterone (T), are essential for a correct spermatogenesis development from early stages and spermatogonia proliferation to germ cells maturation. Other hormones, like inhibin and activin, finely participate tuning the process of spermatogenesis. Genetic regulation involves various transcription factors such as SOX9, SRY, and DMRT1, which are crucial for the development and maintenance of the testis and germ cells. MicroRNAs (miRNAs) play a significant role by regulating gene expression post-transcriptionally. Epigenetic modifications, including DNA methylation, histone modifica-tions, and chromatin remodelling, are also vital. Temperature regulation is another critical as-pect, with the testicular temperature maintained around 2-4°C below body temperature, essen-tial for efficient spermatogenesis. Heat shock proteins (HSPs) protect germ cells from heat-induced damage by acting as molecular chaperones, ensuring proper protein folding and preventing the aggregation of misfolded proteins during thermal stress. Elevated testicular temperature can impair spermatogenesis, increasing germ cell apoptosis, and inducing oxidative stress, DNA damage, and disruption of the blood-testis barrier, leading to germ cell death and impaired differentiation. Cellular mechanisms of germ cell proliferation, differentiation, and death include the mitotic divisions of spermatogonia to maintain the germ cell pool and pro-duce spermatocytes. Spermatocytes undergo meiosis to produce haploid spermatids, which then differentiate into mature spermatozoa. Apoptosis, or programmed cell death, ensures the re-moval of defective germ cells and regulates the germ cell population. Hormonal imbalance, ge-netic defects, and environmental stress can trigger apoptosis during spermatogenesis. Under-standing these mechanisms is crucial for addressing male infertility and developing therapeutic interventions. Advances in molecular biology and genetics continue to uncover the intricate de-tails of how spermatogenesis is regulated at multiple levels, providing new insights and poten-tial targets for treatment.

Background: In vitro embryo production is a highly demanded reproductive technology in horses, which requires the recovery (in vivo or post-mortem) and in vitro maturation (IVM) of oocytes. Oocytes subjected to IVM exhibit poor developmental competence compared to their in vivo counterparts, being this related to a suboptimal composition of commercial maturation media. The objective of this work was to study the effect of different concentrations of secretome obtained from equine preovulatory follicular fluid (FF) on cumulus-oocyte complexes (COCs) during IVM. COCs retrieved in vivo by ovum pick up (OPU) or post-mortem from a slaughterhouse (SLA) were subjected to IVM in the presence or absence of secretome (Control: 0 µg/ml, S20: 20 µg/ml or S40: 40 µg/ml). After IVM, the metabolome of the medium used for oocyte maturation prior (Pre-IVM) and after IVM (Post-IVM), COCs mRNA expression, and oocyte meiotic competence were analysed. Results: IVM leads to lactic acid production and an acetic acid consumption in COCs obtained from OPU and SLA. However, glucose consumption after IVM was higher in COCs from OPU when S40 was added (Control Pre-IVM vs. S40 Post-IVM: 117.24 ± 7.72 vs. 82.69 ± 4.24; Mean µM ± SEM; p < 0.05), while this was not observed in COCs from SLA. Likewise, secretome enhanced uptake of threonine (Control Pre-IVM vs. S20 Post-IVM vs. S40 Post-IVM: 4.93 ± 0.33 vs. 3.04 ± 0.25 vs. 2.84 ± 0.27; Mean µM ± SEM; p < 0.05) in COCs recovered by OPU. Regarding the relative mRNA expression of candidate genes related to metabolism, Lactate dehydrogenase A (LDHA) expression was significantly downregulated when secretome was added during IVM at 20–40 µg/ml in OPU-derived COCs (Control vs. S20 vs. S40: 1.77 ± 0.14 vs. 1 ± 0.25 vs. 1.23 ± 0.14; fold change ± SEM; p < 0.05), but not in SLA COCs. Conclusions: The addition of secretome during in vitro maturation (IVM) affects the gene expression of LDHA, glucose metabolism, and amino acid turnover in equine cumulus-oocyte complexes (COCs), with diverging outcomes observed between COCs retrieved using ovum pick up (OPU) and slaughterhouse-derived COCs (SLA).

Maternal undernutrition during pregnancy in mammals can induce long-term effects in offspring health through molecular programming of the gametes. Using the rabbit model, this study investigates whether maternal food restriction (FR) during the first two-thirds of gestation induces effects on ovarian follicular, oocyte, and early embryo developmental markers in F1 female offspring at the onset of reproductive life. Additionally, body composition, metabolic profile, and growth trajectory from birth to sexual maturity (16 weeks) were assessed to evaluate potential impacts on overall health. Pregnant females (F0) were fed either ad libitum (Control group) or a restricted diet covering 60% of nutritional requirements (FR group). Offspring from both groups were fed ad libitum . Maternal FR had no significant effects on birth weight and survival of progeny, growth trajectory, feed intake or glycemic profile during the juvenile phase. Body weight, body composition, lipid, and glycemic profiles in F1 sexually mature females were similar. However, serum aminotransferase levels were significantly elevated in the F1 females from FR group (P < 0.05), indicating potential hepatic stress. In FR group, F2 oocytes showed a significant upregulation of SOD2, G6PD , and FABP4 mRNA expression levels (P < 0.05), while cumulus cells (CCs) exhibited increased TP53 and decreased CASP3 transcripts levels (P < 0.05). At ovulation time, the progesterone/estradiol ratio was significantly higher (P < 0.001), coinciding with an increased proportion of F2 expanded blastocysts (P < 0.005) and total embryo cell counts (P < 0.05). Serum anti-Müllerian hormone levels, ovulation rate, apoptosis rate, and in vitro embryo development did not differ between groups. These findings suggest that re-feeding after maternal food restriction can attenuate adverse long-term effects on offspring growth and metabolism, while modulating the expression of genes related to oxidative stress, apoptosis and fatty acid uptake in the oocytes of F1 females. This modulation may reflect the activation of compensatory intracellular mechanisms that support early embryonic development in juvenile females at the onset of their reproductive life.

Understanding how extant physiological landscapes arise from novel genetic interactions is key to elucidating phenotypic evolution. Sperm cells exemplify a striking case of functional compartmentalization shaped by molecular adjustments, notably regarding energy metabolism. Here, we examine the impact of gene duplication and loss on the evolution of sperm energetics in mammals. Our findings reveal that the acquisition of an exclusive mechanism controlling the sperm plasma membrane Na+ gradient, critical for glucose uptake, emerged in the ancestor of mammals through gene duplication, which originated the Na+/K+ ATPase transporting subunit alpha 4 transporter (Atp1a4). Furthermore, we demonstrate that testis-specific expression of Atp1a4 was acquired after the divergence from monotremes. Notably, we identify three independent pseudogenization events of Atp1a4, including in pangolins, the naked mole-rat (Heterocephalus glaber) and toothed whales. The recurrent loss of function in Atp1a4 coincides with the erosion of the testis-specific glycolytic pathway in these lineages. Furthermore, enrichment analysis of striped dolphin (Stenella coeruleoalba) and naked mole-rat testis transcriptomes also suggests significant alterations in sperm capacitation processes. Overall, we show that the elaboration of a sodium-dependent glucose uptake wiring was a key innovation in the energetic landscape governing mammalian spermatozoa, with secondary gene loss in three separate lineages pointing to drastic alterations in motility and capacitation processes. Our findings illustrate how metabolic pathways co-shaped by gene duplication and erosion define extant physiological phenotypes.