Preimplantation genomic selection (PGS) using single nucleotide polymorphism (SNP) genotyping can be used to accelerate genetic improvement in cattle, but the technical components for in vitro-produced (IVP) embryos remain unvalidated in Japanese Black cattle; we aimed to address this gap in the present study. In the technical validation experiments, biopsies of approximately 15 cells and 5 cells achieved median call rates of 97.8% and 95.2%, respectively, with a median SNP genotype concordance of 99.9% with the corresponding embryos in both groups. Among biopsied IVP blastocysts, vitrification preserved a significantly higher cell number than slow freezing. In Japanese Black IVP embryos, call rates ≥ 90% were achieved in 88.5%, variation in genomic estimated breeding values (GEBVs) was observed among full-sibling embryos, and the conception rate after vitrification and transfer was 24.0%. At biopsied-cell call rates ≥ 90%, GEBV ranking was preserved between biopsied cells and the corresponding calves. These findings support the potential utility of PGS using IVP embryos as a breeding tool for Japanese Black cattle.

In mice, zygotic gene activation (ZGA) occurs in two waves: a minor wave at the 1-cell stage and a major wave at the 2-cell stage. At the 1-cell stage, chromatin is highly decondensed and transcriptionally permissive; however, global transcription is low. At the 2-cell stage, global transcription increases with chromatin compaction. Thus, the early embryo departs from the usual association between open chromatin and active transcription. Open chromatin in the 1-cell embryo is regarded as a hallmark of totipotency; however, its contribution to transcription remains unclear. To address this issue, whether increasing accessibility is sufficient to enhance global transcription and whether the response depends on the mechanism of chromatin opening were investigated. Therefore, global chromatin accessibility was increased at each developmental stage using two mechanistically distinct perturbations, KDM4D-mediated removal of H3K9me3 and trichostatin A (TSA)-mediated histone hyperacetylation, and the resulting accessibility and transcription were evaluated by DNase-TUNEL and 5-ethynyl uridine incorporation, respectively. At the 1-cell stage, both perturbations increased accessibility but not global transcription. KDM4D decreased transcription in the maternal pronucleus, whereas TSA reduced transcription in both pronuclei. At the 2-cell stage, KDM4D increased accessibility without altering global transcription. The simultaneous removal of H3K9me3 and H3K27me3 did not increase global transcription. In contrast, TSA increased global accessibility and global transcription. Thus, increasing global chromatin accessibility alone was insufficient to enhance global transcription and could even reduce it. The transcriptional response depended on the mechanism of chromatin perturbation. Furthermore, maternally enriched H3K9me3 did not account for the lower transcriptional activity in the maternal pronucleus.

Sperm freezability varies considerably among pigs owing to genetic, husbandry, and management differences. Boar sperm cryopreservation remains underutilized owing to its inconsistent freezability. Semen with good freezability (GF) increases sow litter size and reduces breeding costs. Previous studies identified freezability biomarkers in cattle, sheep, and pigs. To comprehensively investigate these biomarkers, we conducted proteomic profiling of sperm with poor freezability (PF) and GF in Wannan Black pigs. We detected 2,485 proteins, identifying 337 significantly differentially expressed proteins (DEPs). Gene Ontology annotation revealed the involvement of DEPs in crucial processes, such as protein homeostasis maintenance, zona pellucida binding, polyspermy blockade, and ATPase activation regulation. Its molecular functions include immune signaling and oxidative stress defense. KEGG pathway analysis highlighted DEPs enrichment in seminal adhesion complexes, nucleic acid helicase families, and mitochondrial membrane protein clusters. Subcellular localization showed predominant DEPs distribution in the cytoplasm, extracellular matrix, nucleus, mitochondria, and plasma membranes. Western blotting validation of differentially expressed proteins (HSP90AA1, ACE, ANXA2, DEFB1, and S100A8) demonstrated results consistent with the proteomic sequencing data. This study delineates the key proteomic differences underlying boar sperm freezability, providing insights into refining cryopreservation protocols and augmenting swine reproductive efficiency.

Seasonal reproductive dynamics were investigated in two adult male Pacific white-sided dolphins (Aethalodelphis obliquidens, formerly Lagenorhynchus obliquidens) maintained at the Enoshima Aquarium over a two-year period (December 2018–November 2020). Serum testosterone (T), testicular cross-sectional area (TS), and pixel intensity (PI), an index of testicular echogenicity, were measured along with rectal body temperature at two depths (shallow, 15 cm; deep, 30 cm). Untargeted serum metabolomic profiling was performed using gas chromatography–mass spectrometry (GC-MS). In both individuals, T showed a clear seasonal pattern, with elevations lasting approximately one month during spring and summer, followed by prolonged increases in TS (approximately three months) and PI (three–four months). This temporal sequence was consistent across both years of observation. The T, TS, PI, and photoperiod were strongly and positively correlated with one another (Spearman’s rs > 0.74, P < 0.01). Shallow-body temperature varied seasonally with water temperature, whereas deep-body temperature remained relatively stable. GC-MS detected 131 metabolites, and partial least squares discriminant analysis revealed a separation between high-T (> 10 ng/ml) and low-T (< 1 ng/ml) states. The four metabolites with the highest variable importance in projection scores were myoinositol, palmitoleic acid, L-tyrosine, and L-phenylalanine. Myoinositol and palmitoleic acid were positively correlated with the reproductive indices, whereas L-tyrosine and L-phenylalanine were negatively correlated, suggesting coordinated shifts in inositol, lipid, and amino acid metabolism during reproductive activation. To the best of our knowledge, this is the first study to integrate endocrine, ultrasonographic, thermal, and metabolomic approaches to characterize male reproductive seasonality in this species.

Fertilization depends on the proper formation and function of the acrosome, a specialized organelle essential for sperm-oocyte interaction. Defects in acrosome biogenesis impair fertilization and cause severe male infertility, including globozoospermia. However, the molecular mechanisms underlying acrosome formation and maintenance remain poorly understood. Here, we investigated the roles of the testis-enriched proteins LRRC37 and LRRC37A using knockout (KO) mouse models. While Lrrc37 KO males were fertile with only mild sperm head abnormalities, Lrrc37a KO males were completely infertile and produced round-headed spermatozoa characteristic of globozoospermia. LRRC37A localized to the acrosome, and its loss resulted in abnormal acrosome enlargement accompanied by the accumulation of Golgi- and vesicle-like structures. Acrosome formation in Lrrc37a KO mice appeared normal during early spermiogenesis but became abnormal at later stages, indicating a requirement for LRRC37A in acrosomal structural integrity. These defects were associated with impaired sperm head shaping during spermiogenesis, leading to globozoospermia. Together, these findings establish LRRC37A as an essential factor in maintaining acrosome integrity and sperm head morphogenesis, highlighting its distinct function from LRRC37. This work provides new insight into the molecular basis of acrosome-related infertility and has important implications for understanding the etiology of globozoospermia.

Prosaposin (PSAP), a secreted glycoprotein with neurotrophic activity, and protectin D1, a docosahexaenoic acid-derived lipid mediator, are endogenous ligands of G protein-coupled receptor 37 (GPR37). Although PSAP is expressed in the pituitary gland, the cellular localization and functional significance of PSAP–GPR37 signaling and protectin D1-associated pathways remain unclear. In this study, we investigated the localization and potential functions of PSAP, GPR37, and protectin D1-related factors in the rat pituitary gland, focusing on adult pituitary stem/progenitor cells (CD9/SRY-box transcription factor 2 [SOX2]-positive cells) in the marginal cell layer (MCL) facing the residual lumen of Rathke’s pouch. In situ hybridization and immunohistochemistry revealed that PSAP was localized in CD9/SOX2-positive cells in the intermediate lobe-side MCL (IL-MCL) and in subsets of adrenocorticotropic hormone (ACTH)- and growth hormone-producing cells in the anterior lobe (AL) of the pituitary gland. Gpr37 was predominantly detected in CD9/SOX2-positive cells in the IL-MCL and in some ACTH-producing cells. The protectin D1-associated factors, fatty acid transport protein 1 (FATP1/SLC27A1) and arachidonate 15-lipoxygenase (ALOX15), colocalized with CD9/SOX2-positive cells in the IL-MCL. In a pharmacologically induced prolactinoma model, Gpr37, Slc27a1, and Alox15 expression level was upregulated in IL-derived CD9-positive cells. Functional analyses showed that PSAP treatment increased proopiomelanocortin expression in AL-derived CD9-negative cells, whereas protectin D1 treatment increased Sox2 expression and decreased Gpr37 expression in CD9/SOX2-positive cells. These findings suggest that ligand-dependent GPR37 signaling may have distinct roles in the pituitary gland: PSAP–GPR37 signaling may regulate ACTH production, whereas protectin D1–GPR37 signaling may contribute to Sox2 expression in CD9/SOX2-positive cells.

The homogenous, compact or non-cavitary corpus luteum (CL) is an ovarian structure appearing after ovulation and responsible of progesterone synthesis during the estrous cycle and pregnancy. The cavitary CL is similar to the homogeneous but it presents a more or less developed central cavity. The prevalence of a cavitary CL ranges from 18 to 79% during the period of 6 to 12 days post-ovulation. Most cavities regress throughout the cycle. This review aims to present the state of art of the prevalence, physio-morphological characteristics, diagnostic methods and effects on fertility of the cavitary corpus luteum, most of the aspects remaining a topic of debate. From a clinical perspective, the presence of a CL with a large cavity with a very thin wall, resembling an ovarian follicular/luteal cyst, in absence of a further CL in pregnant cows is also described.

Cervical ripening promotes dilation of the birth canal and placental maturation facilitates the expulsion of fetal membranes after parturition, both of which involve coordinated inflammatory processes. Understanding the mechanisms that regulate sterile inflammation in uterine tissues is crucial for preventing periparturient complications in cattle, including dystocia, stillbirth, and retained placenta. Although we previously showed that attenuated upregulation of interferon (IFN) signaling in placentomes at calving is associated with retained placenta, the role of IFN signaling in sterile inflammation remains poorly understood. This study examined whether changes in IFN activity in the cervical mucosa during preparturition indicate inflammation during normal calving. Transcriptome analysis revealed increased expression of the IFN-stimulated genes ISG20 and IFIT1 at calving, similar to that observed in the placentomes. Real-time polymerase chain reaction revealed that IFNA, IFNG, and IFNT expression in the cervical mucosa decreased significantly at calving, whereas IFN-stimulated gene expression increased. Furthermore, ISG20 and MX2 levels showed highly consistent increases among individuals. Changes in IFN-related gene expression in peripheral blood leukocytes were less distinct than those in the cervical mucosa. Furthermore, IL1A, IL1B, IL8, and TNFA expression increased in the cervical mucosa, whereas IL6 expression decreased. Among these cytokine changes, IL1B and IL8 were consistent among individuals. These findings highlight IFN signaling as a common immune regulator in the cervix and placentomes at calving, indicating its potential utility in the development of predictive and preventive interventions for periparturient disorders associated with dysregulated sterile inflammation.

Selenium (Se) is an essential micronutrient involved in male reproductive physiology through its integration into selenoproteins that orchestrate antioxidant defense and spermatogenesis. This study compared the effects of inorganic and yeast- and bacteria-derived Se on the bioavailability, testicular development, sperm functional traits, epididymal plasma biochemistry, and testicular glycogen reserves in growing lambs. Twenty-four lambs (6–7 months old) were randomly assigned to four dietary groups (n = 6) for 90 days: control and sodium selenite (0.5 mg/kg), Se-yeast (0.5 mg/kg), and bacteria-derived Se (0.5 mg/kg) supplemented groups. Se supplementation significantly increased Se concentrations in serum, epididymal plasma, and testicular tissue (P < 0.0001), with bacteria-derived Se exhibiting the highest tissue deposition. Improved bioavailability enhanced testicular development, as evidenced by paired testicular weight, gonadosomatic index, and other morphometric parameters (P < 0.0001). Se also fortified the antioxidant architecture, upregulating glutathione peroxidase, superoxide dismutase, and catalase activities, while markedly reducing malondialdehyde levels (P ≤ 0.0004). Epididymal sperm quality improved significantly, including motility, concentration, membrane integrity, viability, and reduced abnormalities (P < 0.0001), without affecting the recovered fluid volume (P = 0.6306). Additionally, Se enhanced epididymal plasma biochemical profiles and increased testicular glycogen reserves (P < 0.0001). Overall, bacteria-derived Se resulted in superior tissue Se deposition and improved reproductive parameters compared to that from other sources. While these responses are likely associated with an enhanced antioxidant status, the specific Se species and underlying molecular mechanisms require further investigation. Consequently, bacteria-derived Se represents a promising alternative source for supporting male reproductive development in small ruminants.