Search Results (61)

Background/Objectives: The corpus luteum (CL) in the dog is the sole source of progesterone (P4) during diestrus and pregnancy, making it a key regulator of reproductive function. However, robust and functionally validated in vitro models of canine luteal cells remain limited. This study aimed to establish and characterize a reproducible primary culture system of canine luteal cells with preserved steroidogenic activity and regulatory responsiveness. Methods: Ovaries containing CLs were collected from five clinically healthy bitches undergoing routine ovariohysterectomy (OHE). Luteal tissue was mechanically fragmented and enzymatically digested using collagenase type II. Primary cultures were established using an explant-based approach and maintained in Dulbecco’s Modified Eagle Medium/Ham’s F-12 nutrient mixture (DMEM/F12) or Roswell Park Memorial Institute medium 1640 (RPMI 1640) supplemented with 20% fetal bovine serum (FBS). Cellular morphology, proliferation, expression of steroidogenic markers—steroidogenic acute regulatory protein (STAR) and 3β-hydroxysteroid dehydrogenase type 1 (HSD3B1), P4 secretion, and responsiveness to forskolin stimulation were evaluated. Results: Cultured luteal cells exhibited stable attachment, proliferation, and a predominantly spindle-shaped morphology. Both media supported maintenance of a steroidogenic phenotype, while RPMI 1640 enabled enhanced proliferation, allowing expansion up to passage three and efficient cryobanking. Cells remained functionally active, secreting progesterone for up to 28 days in vitro. Forskolin stimulation increased progesterone secretion up to 2.7-fold, confirming preserved cyclic AMP-dependent steroidogenic responsiveness. Conclusions: The canine CL is a reliable source of functionally competent luteal cells, and the established culture system represents a physiologically relevant in vitro model. To our knowledge, this is the first functionally validated in vitro model of the canine CL. This platform enables controlled investigations of luteal function, endocrine regulation, and mechanisms of P4 synthesis, supporting its application in mechanistic and translational reproductive research. Full article

The corpus luteum number in sows is a key reproductive indicator for measuring ovulation rate and reproductive efficiency. Its formation is precisely regulated by a complex gene regulatory network composed of multi-level molecular interactions. To systematically elucidate the molecular basis of this trait, this study comprehensively analyzed genes located within QTL for corpus luteum number. This approach identified a series of key regulatory molecules specifically expressed in the ovary, including transcription factors (TFs), long non-coding RNAs (lncRNAs), and microRNAs (miRNAs). Using bioinformatics methods to predict the target genes of candidate miRNAs, combined with functional enrichment analysis, revealed that these target genes were significantly enriched in multiple core reproductive pathways closely related to cell proliferation, differentiation, and hormone regulation, including the ErbB signaling pathway, PI3K-Akt signaling pathway, and TGF-beta signaling pathway. Based on the above findings, this study ultimately constructed a TF-lncRNA-miRNA-mRNA network, which is associated with QTL for corpus luteum number. Furthermore, key genes were validated via quantitative real-time PCR (qRT-PCR). Significant positive correlations were identified between the transcription factor NEUROG2 and lncRNA LOC102167554, along with its potential target gene ESRP1, as well as between transcription factor SNAI2 and lncRNAs (LOC102167554, LOC102167796) and their potential target genes (FXR, ERBB4). In addition, the functional validation results showed that the interference of LOC102167554 significantly reduced the proliferation ability of sGCs. These key genes represent potential targets for genetic improvement of sow reproduction. Full article

Paroxetine, a selective serotonin reuptake inhibitor commonly used to treat various psychiatric disorders, may adversely affect female reproductive function. Although apigenin has been shown to ameliorate reproductive abnormalities and ovarian dysfunction, its effect on paroxetine-induced reproductive toxicity in females remains unclear. Therefore, this study investigated the potential protective effects of apigenin against paroxetine-induced reproductive alterations in female rats. Female rats with regular estrous cycles were randomly assigned to four groups (n = 9 per group): control, apigenin, paroxetine, and paroxetine + apigenin. The rats received saline, apigenin (20 mg/kg), paroxetine (10 mg/kg), or their combination by oral gavage once daily for about 29 consecutive days. Compared with paroxetine treatment alone, apigenin co-administration restored decreased serum anti-Müllerian hormone (AMH) levels, enhanced PAS reactivity in the zona pellucida, reduced ovarian iNOS immunoreactivity, increased follicle and corpus luteum numbers, and increased ovarian VEGF immunoreactivity. However, apigenin administration alone was associated with reduced testosterone levels and alterations in certain ovarian and uterine histological features in female rats. In conclusion, the findings suggest that apigenin may ameliorate paroxetine-induced reproductive alterations in female rats by modulating AMH levels, follicle and corpus luteum numbers, and ovarian histochemical and molecular parameters. Full article

Ovarian function relies on a network of well-coordinated molecular mechanisms that regulate follicular development, oocyte maturation, ovulation, and corpus luteum function. When these processes are disrupted, infertility can result. Extracellular matrix (ECM) remodeling represents a central regulatory component in these processes and is essential for follicle rupture and oocyte release. This mechanism involves metalloproteinases (MMPs), mainly MMP-2 and MMP-9, which degrade the ECM and allow the necessary structural changes. Other ECM-modulating proteases, such as ADAM and ADAMTS families, also contribute to this process. Their activity is tightly regulated by tissue inhibitors of metalloproteinases (TIMPs), ensuring that tissue remodeling occurs in a controlled manner. Disruption of the balance between MMPs and TIMPs increases the risk of infertility-related conditions such as polycystic ovary syndrome (PCOS), endometriosis, luteinizing hormone (LH) deficiency syndrome, and ovarian aging. In addition to the ECM, other factors, including intracellular signaling pathways, oxidative stress (OS), and mitochondrial function, contribute to ovarian physiology and directly affect oocyte quality and viability. This narrative review focuses on the molecular mechanisms governing ovarian function, with particular emphasis on the remodeling of the ECM by MMPs during ovulation, and examines how their disorders contribute to infertility. A deeper understanding of these mechanisms may lead to the identification of new therapeutic targets and the improvement of assisted reproduction outcomes. Full article

Heat stress (HS) occurs when animals are unable to effectively dissipate excess body heat, leading to increased core temperature and physiological imbalance. In mammals, HS negatively affects female reproduction. Infertility associated with HS is well documented in swine and is increasingly recognized in other mammals, including humans. HS disrupts several systemic processes that are essential for normal reproductive function, including endocrine regulation, nutrient metabolism, immune activity, and intestinal barrier integrity. Reduced feed intake and changes in metabolic hormones such as insulin and prolactin can impair ovarian function. Increased intestinal permeability during HS may allow bacterial endotoxins to enter the bloodstream, triggering inflammation that further compromises reproductive physiology. At the ovarian level, HS alters key cellular pathways involved in cell survival and metabolism, including Janus Kinase/Signal Transducer and Activator of Transcription (JAK–STAT), Phosphoinositide 3-Kinase/Protein Kinase B (PI3K/AKT), oxidative stress responses, autophagy, apoptosis, and heat shock protein expression. These changes disrupt follicular development, hormone production, oocyte quality, and corpus luteum function, resulting in reduced conception rates and increased embryonic loss. This review summarizes current knowledge of systemic and ovarian mechanisms by which HS impairs female reproduction in pigs and identifies areas requiring further investigation to improve fertility under increasing environmental temperatures. Full article

Persistent corpus luteum (PCL) and ovarian quiescence (OQ) are key manifestations of ovarian dysfunction (OD) that lead to reduced reproductive capacity in beef cattle, posing a serious challenge to the industry. Anthocyanins (ACNs) are known for their antioxidant properties. This study aimed to investigate the regulatory effects of ACNs on PCL and OQ and to explore the underlying mechanisms. Forty-eight beef cows diagnosed with both OQ and PCL were selected and continuously fed ACNs for 60 days. The results showed that the regulatory effects of ACNs were dose-dependent. A high dose of ACNs (ACNH) significantly increased the number of large follicles and reduced the occurrence of PCL. ACNH treatment significantly decreased serum progesterone (P4) levels and increased estradiol (E2) levels. Furthermore, ACNH reduced microbial diversity in OD cows but significantly increased the abundance of Patescibacteria, Actinobacteriota, Proteobacteria, and Chloroflexi, while decreasing the abundance of Desulfobactera, indicating that ACNs may affect ovarian function by regulating the gut microbial environment. In an ovarian granulosa cell model of oxidative damage, ACN intervention could reduce oxidative stress levels and mitigate oxidative damage. ACNs downregulated various pro-apoptotic genes, such as P53, Fas, and Bax, while upregulating anti-apoptotic genes Bcl-2 and Bcl-xL, suggesting that ACNs significantly inhibit cell apoptosis. To conclude, these results demonstrate that ACNs improve the ovarian function of beef cows by regulating gut microbiota and reducing oxidative stress-induced apoptosis in ovarian granulosa cells, thereby enhancing the reproductive capacity of beef cattle that show reproductive disorders. These findings provide a theoretical basis for the application of ACNs in the cattle industry and showcase their potential value as natural antioxidants. Full article

The aim of this study was to comparatively evaluate echotextural and hemodynamic changes in the corpus luteum (CL), uterus, and uterine artery, together with serum progesterone (P4) concentrations, using B-mode and Doppler ultrasonography between days 5 and 21 post-insemination in pregnant and non-pregnant cows. Twelve clinically healthy Brown Swiss cows were enrolled and allocated into a cyclic non-inseminated group (n = 6) and an inseminated group (n = 6). Ultrasonographic examinations and progesterone measurements were performed daily during the post-insemination period, and pregnancy was confirmed on day 30. Echotextural parameters (mean gray value and homogeneity) were obtained from the corpus luteum and uterus using B-mode ultrasonography. Doppler ultrasonography was used to assess corpus luteum vascular parameters and uterine artery blood flow, and serum progesterone concentrations were measured at each examination. Corpus luteum mean gray value showed a significant time effect (p < 0.001). For Corpus luteum area and perfusion area, both the time effect and the group × time interaction were significant (p < 0.001), and marked differences between pregnant and non-pregnant cows were observed on days 19, 20, and 21 (p < 0.05). Serum progesterone concentrations also differed significantly between groups on days 20 and 21. In conclusion, changes in corpus luteum area and perfusion area were associated with early pregnancy-related differences and may represent earlier functional ultrasonographic indicators compared with uterine artery Doppler parameters and progesterone concentrations alone. These findings may have practical implications for herd management by potentially enabling differentiation between pregnant and non-pregnant cows approximately 1–2 days earlier than serum progesterone measurements. Full article

The aim of this study was to investigate the effects of dietary supplementation with 0.11% N-carbamylglutamate (NCG) during early pregnancy (0–90 days) on reproductive performance and fetal development, and to elucidate the underlying placental regulatory mechanisms in primiparous Hu sheep. Twenty-two 10-month-old sexually mature primiparous Hu sheep meeting the mating criteria were randomly assigned to two groups. The control group was fed a basal diet, while the NCG group received the basal diet supplemented with 0.11% NCG, with both feeding regimens maintained for 90 days. By measuring uterine and fetal growth indices, maternal plasma biochemical parameters, and amino acid levels, as well as assessing cotyledon indices and observing cotyledon morphology and histological structure, basic data related to placental function and fetal growth in pregnant ewes was collected. Combined with transcriptomic sequencing of maternal placental tissue, the mechanism by which NCG influences placental function and fetal growth and development in pregnant ewes was further investigated. The supplementation of NCG could increase the number of fetuses, total weight of fetuses, the number of corpus luteum and the ratio of fetuses to corpus luteum, but the difference was not significant (p > 0.05). The levels of plasma NO, inducible Nitric Oxide Synthase (iNOS) and several amino acids were significantly increased (p < 0.05). In ewes’ uteri, the average uterine weight, number of uterine glands, total cotyledon weight, and average weight per cotyledon were significantly increased (p < 0.05), whereas uterine mucosal thickness was markedly decreased. The Quantitative Real-time PCR (q-PCR) results for differentially expressed genes were consistent with those of transcriptomic analysis, showing significant changes in the expression levels of certain differentially expressed genes in maternal placental tissues. These changes regulated pathways such as vascular endothelial growth factor (VEGF), insulin-like growth factor (IGF), phosphatidylinositol 3-kinase–protein kinase B (PI3K–AKT) signaling pathways and Mitogen-Activated Protein Kinase (MAPK) pathway, which are involved in angiogenesis, energy supply and metabolism, and somatic growth and development. Dietary supplementation with NCG during early pregnancy can significantly improve the reproductive performance of primiparous Hu sheep, optimize the intrauterine environment and nutrient supply, and thereby facilitate pregnancy maintenance and fetal development. The underlying mechanism may involve promoting endogenous arginine synthesis in ewes, increasing plasma levels of NO, arginine, and certain amino acids, which collectively validate the positive effects of NCG on the reproductive performance and growth of Hu sheep during early pregnancy at the molecular level. Full article

Effective luteal support is crucial for successful pregnancy following embryo transfer in dromedary camels, highlighting the importance of maintaining adequate luteal function. This study compared three luteal support protocols for pregnancy rates after embryo transfer (ET) in dromedary camels. Recipients with a single ovarian follicle (14~16 mm) were assigned into: (1) untreated control, (2) Phenylbutazone (PBZ) at 3.7 mg/kg IV prior to ET, or (3) human chorionic gonadotropin (hCG) 2000 IU plus 3.7 mg/kg IV PBZ prior to ET. Recipients that showed ovarian inactivity were treated with an injection of 1800 IU of equine chorionic gonadotropin (eCG) after progesterone (P₄) priming. Pregnancy was assessed on days 10 (P₄ assay) and 60 (transrectal ultrasonography) post-ET. Pregnancy rates on day 10 were higher in the hCG + PBZ (73.3%) and eCG (68.0%) groups than in the PBZ (44.0%) and control (35.0%) groups (p < 0.05). By day 60, the hCG + PBZ group exhibited the highest pregnancy maintenance rate (81.8%) and the lowest embryonic loss rate (18.2%), compared with eCG (53.0%; 47.0%), PBZ (54.5%; 45.5%), and control (57.1%; 42.9%) groups, respectively (p < 0.05). However, there was no significant difference among the eCG, PBZ, and control groups in embryonic loss rates. In conclusion, hCG synergizes with PBZ to enhance corpus luteum function, uterine receptivity, and pregnancy maintenance more effectively than PBZ or eCG. Full article

This study was designed, for the first time, to fully characterize the corpus luteum’s (CL) dynamics, echotexture, and ovarian blood flow on the ipsilateral side of the CL during different stages of the follicular wave in spontaneously non-mated camels (Camelus dromedarius) and to correlate the CL’s size echotexture with Doppler parameters. Of 20 non-mated camels, only 7 exhibited spontaneous ovulation. B- and color-mode analyses of the CL were estimated; CL frozen image echotextures [CL echogenicity (CLE) and CL heterogeneity (CLH)] and ovarian artery (OV. A.) dynamics were recorded, and ultrasound scanning was performed. Blood sampling and progesterone (P4) levels were measured after ovulation. CL diameter and echotexture were elevated (p = 0.025 and p = 0.037) at the mid-maturation stage and compared to the early growth and late regression stages (1.03 ± 0.45/cm and 82.65 ± 2.87 for CLE and 33.65 ± 1.83 for CLH vs. 1.98 ± 0.88 cm; 66.52 ± 4.32 for CLE and 15.66 ± 0.25 for CLH vs. 1.02 ± 0.02 cm, 65.12 ± 2.66 for CLE, and 19.32 ± 1.33 for CLH), as those parameters are critical in the determination of CL activity. Ipsilateral OV. A. diameter increased (p = 0.021) in the mid-maturation and regression stages, with a significant elevation in Doppler velocities (p = 0.025) in the maturation stage, with a decline in Doppler indices (p = 0.013), while the contralateral side was not affected. Ipsilateral mean velocity (Vm; cm/s) and blood flow volume (BFV; mL/min) were increased in the mid-maturation stage (23.55 ± 0.66 cm/s and 25.62 ± 0.32 mL/min). CL diameter was positively correlated with the CL’s total colored area/pixels (r = 0.81; p = 0.001), total colored area % (r = 0.93; p = 0.001), and OV. A. velocities (r = 0.96; p = 0.001). In addition, there was a positive correlation between CLH and OV. A.BFV (r = 0.89; p = 0.001). After spontaneous ovulation, the CL increases in diameter and reaches its peak on day 12, with an elevation in the P4 level at day 10, and the total colored area of the CL continues to elevate until day 14. Ipsilateral OV. A. blood flow is elevated and linked to changes that occur in the CL’s total coloration %. Evaluating luteal function in camels presents several challenges due to the species’ unique reproductive physiology. Full article

The ovarian corpus luteum has functional mechanisms for formation and regression in the reproductive endocrine system. The main functional events of the corpus luteum are angiogenesis and apoptosis mechanisms. The development of the corpus luteum involves homogeneous physiological mechanisms, including cellular functions and reproductive hormones. Angiogenesis is controlled by pro-angiogenic and anti-angiogenic factors. The microenvironment involves various signaling molecules and pathways that may play a potential role in angiogenic response during corpus luteum growth. In luteolysis, the corpus luteum undergoes degeneration, notably induced by reproductive hormones that promote programmed cell death in luteal cells through the apoptosis mechanism. In this sudy, we discuss the mechanisms and functional roles of angiogenesis and apoptosis in the endocrine microenvironment during corpus luteum formation and regression, based on the interrelationship of physiological events in the ovary. Full article

With estrus confined to three winter months, early pregnancy detection is essential for reproductive management in farmed sika deer. However, the development of reliable non-invasive early pregnancy detection techniques has been hindered by limited understanding of their reproductive physiology. To identify pregnancy-specific biomarkers in sika deer, we performed RNA-sequencing (RNA-Seq) on maternal peripheral blood collected on days 0, 7, 15, and 20 after artificial insemination. Using time-series clustering analysis and weighted gene co-expression network analysis (WGCNA), we identified key genes and pathways at each stage. Notably, maternal-fetal recognition-related interferon-stimulated genes (ISGs; IFNAR1/2, STAT1/2, MX1/2, and RSAD2), anti-apoptotic and immune-regulatory genes (BCL2, XIAP, and IL10), and cysteine metabolism genes (CTH, CBS, GCLC, and GCLM) were upregulated by day 7, suggesting their role in supporting corpus luteum development through immune regulation and redox homeostasis. By days 15–20, upregulated genes were enriched in pathways related to mitochondrial function, cell adhesion, and cell cycle regulation, indicating their involvement in embryo adhesion and syndesmochorial placentation. In conclusion, this study demonstrates that ISGs, immune-regulatory genes and cysteine metabolism genes are detectable as early as day 7 post-insemination, highlighting their promise as early pregnancy biomarkers and providing a molecular basis for non-invasive diagnostic development in sika deer. Full article

Mammals exhibit unique and highly variable mechanisms for the establishment and maintenance of pregnancy. Ruminants (e.g., sheep, cows, and goats) have novel mechanisms whereby the conceptus (embryo and its extra-embryonic membranes) signals for the establishment of pregnancy and exhibits unique metabolic pathways favoring conceptus development. Embryos of ruminants reach the spherical blastocyst stage at 5 to 10 mm in diameter and then elongate rapidly to elongated filamentous conceptuses of greater than 250 mm as they make contact with the uterine luminal epithelium (LE) for implantation. During conceptus elongation the trophectoderm cells secrete interferon tau (IFNT), a novel pregnancy recognition signal for ruminants to ensure maintenance of a functional corpus luteum (CL) to secrete progesterone (P4) required for pregnancy. P4 induces uterine epithelia cells to express the endogenous Jaagsiekte Retrovirus (enJSRV) that may transactivate toll-like receptors 7 and 8 in the conceptus trophectoderm to induce secretion of IFNT, a classical viral–antiviral mechanism. IFNT silences expression of receptors for estradiol (E2) and oxytocin (OXTR), which abrogates the mechanism whereby oxytocin from CL and posterior pituitary would otherwise induce large pulses of prostaglandin F_2α (PGF) by uterine epithelia to cause regression of the CL and its secretion of P4. IFNT has another novel role in silencing expression of not only ESR1 and OXTR, but all classical interferon-stimulated genes in the uterine LE and superficial glandular epithelium (sGE), but with P4 increasing expression of genes for transport of nutrients such as glucose and arginine into the uterine lumen to support conceptus development. Ruminant conceptuses convert glucose to fructose, a novel hexose sugar that cannot be transported back to the maternal circulation. Fructose is converted to fructose-1-PO4 for metabolism, not via the pathway for glycolysis but via the novel fructolysis pathway uninhibited by low pH, citrate, or ATP as is the case for glycolysis. Thus, fructose and its metabolites support the pentose cycle, hexosamine biosynthesis pathway, one-carbon metabolism, and the citric acid cycle for all cells of the conceptus. Arginine is another key nutrient transported into the uterine lumen by the uterine LE/sGE in response to P4 and IFNT. Arginine is metabolized to generate nitric oxide, polyamines, and creatine, essential for conceptus growth and development, while enhancing production of IFNT as a novel pregnancy recognition signal, and upregulating expression of genes in the uterine LE/sGE for transport of nutrients. Fructose is the major hexose sugar supporting major metabolic pathways required for conceptus growth and development in ruminants. Full article

This study aimed to evaluate the effect of combined supplementation of MSG with glycerin, a glucogenic precursor, on ovarian function in sheep. Twenty-four ewes had estrus and follicular waves synchronized using three prostaglandin injections at 7-day intervals. The ewes were grouped: baseline TMR diet (Control; n = 8); glutamate diet (MSG; n = 8), receiving MSG (1 g/kg of body weight/day) for 16 days; and MSG plus glycerin (MSGLY; n = 8), which received MSG plus 150 mL of glycerin during the eight days prior to ovulation induction. MSG showed lower dry matter intake, while the MSGLY group showed increased heart and respiratory rates and skin temperature. Rectal temperature was higher in MSG and MSGLY. MSGLY also showed reduced triglyceride and urea levels. MSG and MSGLY exhibited decreased cholesterol and creatinine. MSGLY exhibited a higher number of large follicles and greater intraovarian blood perfusion after ovulation induction and larger corpus luteum perfusion. Ovulation rate increased by 64% in the supplemented groups vs. control. MSG supplementation led to greater SCL1A1, GRIA1, and GLUD1 genes expression. Thus, the combined supplementation of MSG and glycerin effectively enhances ovarian function in sheep, representing a viable nutritional strategy to improve reproductive outcomes. Full article

Early embryonic loss is a major cause of reproductive inefficiency in cattle, primarily due to premature luteolysis. Interferon tau (IFN-τ), secreted by the trophoblast, plays a critical role in maternal recognition of pregnancy by maintaining corpus luteum function. However, its practical application has been limited by its rapid degradation and short half-life in vivo. Here, we developed a novel formulation of recombinant bovine IFN-τ, combining chitosan-based microencapsulation with starch–chitosan hydrogel delivery, enabling sustained intrauterine release. This dual-delivery strategy offers a significant improvement over conventional IFN-τ administration methods that rely on repeated intrauterine infusions of soluble protein. The rbIFN-τ was expressed in Pichia pastoris, purified to 90.1% homogeneity, and structurally validated via homology modeling and molecular docking, confirming its interaction with type I interferon receptors. The encapsulated formulation retained antiviral activity, stimulated transcription of interferon-stimulated genes (PKR, OAS1, OAS2), and showed sustained release in vitro for up to 26 days. In vivo evaluation demonstrated safety and biological efficacy, with treated cattle showing inhibited luteolysis, sustained serum progesterone levels, and preserved corpus luteum integrity. This formulation represents a promising biotechnological approach to improve reproductive efficiency through a long-acting, species-specific IFN-τ delivery system. Full article