Over the years, the utilization of in vitro fertilization (IVF) in bovine embryo production has increased globally to accelerate the selection of cows with high genetic values. The selection of ...embryos with high implantation potential is a critical factor in establishing pregnancy. Time-lapse monitoring (TLM) has emerged as a new technique that allows frequent and non-invasive imaging of developing embryos. TLM is considered to have several advantages over the conventional morphological evaluation of embryos, which has been widely used in bovine embryo production. Establishing a novel embryo selection algorithm specifically for bovine IVF embryos is a critical challenge, but information on the association between morphokinetic data obtained using TLM and the implantation potential of embryos is still limited. This review outlines the potential application of TLM technology to improve the fertility of bovine IVF embryos, focusing on the results of human and bovine TLM studies that can be applied to select bovine embryos with high implantation potential. First, the progress of the TLM technology in bovine embryo production is summarized. The association between kinetic and morphological parameters and the developmental and implantation potential of human and bovine embryos is outlined. Finally, the benefits of evaluating blastocyst collapse and re-expansion as indicators of bovine embryo viability and the possible application of TLM to detect chromosomal abnormalities and determine embryo sex will be discussed.
Uterine inflammatory diseases commonly occur in postpartum dairy cows, resulting in reduced reproductive performance due to aberrant uterine and ovarian activity. Infection of the uterus with ...gram-negative bacteria results in the detection of lipopolysaccharide (LPS) in the plasma and follicular fluid of cows along with uterine inflammation. LPS acts on follicular components such as theca cells, granulosa cells, and follicle-enclosed oocytes, leading to impaired follicular activity. Follicles with a high LPS environment exhibit reduced follicular steroidogenesis due to the inhibition of steroidogenic enzyme transcription. Primary cell cultures of bovine granulosa and theca cells have shown that LPS acts on follicular cells to impair steroid production, which may disturb follicle growth and/or reduce their ability to ovulate. Even if ovulation occurs, cows with uterine inflammation are less likely to conceive because in addition to uterine damage, LPS also impairs the developmental competence of oocytes. LPS perturbs the nuclear and cytoplasmic maturation of bovine oocytes. Moreover, oocytes matured using LPS treatment are less likely to develop into the blastocyst stage. Such oocytes also have a reduced number of trophoblast cells in blastocysts. Therefore, the detrimental effects of LPS on ovarian activity may be partly responsible for infertility in cows with uterine inflammation. Novel treatment and prevention strategies for uterine inflammatory diseases can be developed by advancing our knowledge of the pathophysiology underlying ovarian dysfunction, and this can only be achieved by further research. The present review outlines the molecular pathogenesis of LPS-induced ovarian dysfunction.
Inflammatory diseases attenuate reproductive functions in humans and domestic animals. Lipopolysaccharide (LPS), an endotoxin released by bacteria, is known to disrupt female reproductive functions ...in various inflammatory diseases. LPS administration has been used to elucidate the impact of pathophysiological activation of the immune system on reproduction. Hypothalamic kisspeptin neurons are the master regulators of mammalian reproduction, mediating direct stimulation of hypothalamic gonadotropin-releasing hormone (GnRH) release and consequent release of gonadotropins, such as luteinizing hormone (LH) and follicle-stimulating hormone from the pituitary. The discovery of kisspeptin neurons in the mammalian hypothalamus has drastically advanced our understanding of how inflammatory stress causes reproductive dysfunction in both humans and domestic animals. Inflammation-induced ovarian dysfunction could be caused, at least partly, by aberrant GnRH and LH secretion, which is regulated by kisspeptin signaling. In this review, we focus on the effects of LPS on hypothalamic kisspeptin neurons to outline the impact of inflammatory stress on neuroendocrine regulation of mammalian reproductive systems. First, we summarize the attenuation of female reproduction by LPS during inflammation and the effects of LPS on ovarian and pituitary function. Second, we outline the inhibitory effects of LPS on pulsatile- and surge-mode GnRH/LH release. Third, we discuss the LPS-responsive hypothalamic-pituitary-adrenal axis and hypothalamic neural systems in terms of the cytokine-mediated pathway and the possible direct action of LPS via its hypothalamic receptors. This article describes the impact of LPS on hypothalamic kisspeptin neurons and the possible mechanisms underlying LPS-mediated disruption of LH pulses/surge via kisspeptin neurons.
•Systemic LPS increases during inflammation and disrupts female reproduction.•Hypothalamic ARC, AVPV, and ME express TLR4 genes and may directly respond to LPS.•LPS disrupts the pulsatile- and surge-mode of GnRH/LH release in various animals.•LPS suppresses GnRH pulse generator activity and ARC kisspeptin gene expression.
Abstract
Lowered glucose availability, sensed by the hindbrain, has been suggested to enhance gluconeogenesis and food intake as well as suppress reproductive function. In fact, our previous ...histological and in vitro studies suggest that hindbrain ependymal cells function as a glucose sensor. The present study aimed to clarify the hindbrain glucose sensor-hypothalamic neural pathway activated in response to hindbrain glucoprivation to mediate counterregulatory physiological responses. Administration of 2-deoxy-D-glucose (2DG), an inhibitor of glucose utilization, into the fourth ventricle (4V) of male rats for 0.5 hour induced messenger RNA (mRNA) expression of c-fos, a marker for cellular activation, in ependymal cells in the 4V, but not in the lateral ventricle, the third ventricle or the central canal without a significant change in blood glucose and testosterone levels. Administration of 2DG into the 4V for 1 hour significantly increased blood glucose levels, food intake, and decreased blood testosterone levels. Simultaneously, the expression of c-Fos protein was detected in the 4V ependymal cells; dopamine β-hydroxylase-immunoreactive cells in the C1, C2, and A6 regions; neuropeptide Y (NPY) mRNA-positive cells in the C2; corticotropin-releasing hormone (CRH) mRNA-positive cells in the hypothalamic paraventricular nucleus (PVN); and NPY mRNA-positive cells in the arcuate nucleus (ARC). Taken together, these results suggest that lowered glucose availability, sensed by 4V ependymal cells, activates hindbrain catecholaminergic and/or NPY neurons followed by CRH neurons in the PVN and NPY neurons in the ARC, thereby leading to counterregulatory responses, such as an enhancement of gluconeogenesis, increased food intake, and suppression of sex steroid secretion.
Metabolic stress and subsequent hepatic dysfunction in high-producing dairy cows are associated with inflammatory diseases and declining fertility. Lipopolysaccharide (LPS)-binding protein (LBP) is ...produced by hepatocytes and controls the immune response, suggesting that it is involved in the pathophysiology of inflammation-related attenuation of reproductive functions during metabolic stress. This study investigated the effect of LBP on the inflammatory status, oocyte quality, and steroidogenesis in the follicular microenvironment of dairy cows. Using bovine ovaries obtained from a slaughterhouse, follicular fluid and granulosa cells were collected from large follicles to evaluate the follicular status of metabolism, inflammation, and steroidogenesis. Cumulus-oocyte complexes were aspirated from small follicles and subjected to in vitro embryo production. The results showed that follicular fluid LBP concentrations were significantly higher in cows with fatty livers and hepatitis than in those with healthy livers. Follicular fluid LBP and LPS concentrations were negatively correlated, whereas LPS concentration showed a positive correlation with the concentrations of non-esterified fatty acids (NEFA) and β-hydroxybutyric acid in follicular fluid. The blastulation rate of oocytes after in vitro fertilization was impaired in cows in which coexisting large follicles had high NEFA levels. Follicular fluid NEFA concentration was negatively correlated with granulosa cell expression of the estradiol (E2) synthesis-related gene (CYP19A1). Follicular fluid LBP concentration was positively correlated with follicular fluid E2 concentration and granulosa cell CYP19A1 expression. In conclusion, follicular fluid LBP may be associated with favorable conditions in the follicular microenvironment, including low LPS levels and high E2 production by granulosa cells.
The brain mechanism responsible for the pulsatile secretion of gonadotropin-releasing hormone (GnRH) is important for maintaining reproductive function in mammals. Accumulating evidence suggests that ...kisspeptin/neurokinin B/dynorphin A (KNDy) neurons in the hypothalamic arcuate nucleus (ARC) play a critical role in the regulation of pulsatile GnRH and subsequent gonadotropin secretion. Dynorphin A (Dyn) and its receptor, kappa-opioid receptor (KOR, encoded by Oprk1), have been shown to be involved in the suppression of pulsatile GnRH/luteinizing hormone (LH) release. On the other hand, it is still unclear whether the inhibitory Dyn signaling affects KNDy neurons or KOR-expressing non-KNDy cells in the ARC or other brain regions. We therefore aimed to clarify the role of ARC-specific Dyn-KOR signaling in the regulation of pulsatile GnRH/LH release by the ARC specific cell deletion of KOR-expressing cells using Dyn-conjugated-saporin (Dyn-SAP). Estrogen-primed ovariectomized female rats were administered Dyn-SAP to the ARC. In situ hybridization of Oprk1 showed that ARC Dyn-SAP administration significantly decreased the number of Oprk1-expressing cells in the ARC, but not in the ventromedial hypothalamic nucleus and paraventricular nucleus. The frequency of LH pulses significantly increased in animals bearing the ARC Dyn-SAP administration. The number of Kiss1-expressing cells in the ARC was not affected by ARC Dyn-SAP treatment. Dyn-KOR signaling within the ARC seems to mediate the suppression of the frequency of pulsatile GnRH/LH release, and ARC non-KNDy KOR neurons may be involved in the mechanism modulating GnRH/LH pulse generation.
Reduction in oocyte quality is a major factor responsible for declining fertility associated with maternal aging in cows. The objective of the present study was to determine whether intracytoplasmic ...sperm injection (ICSI) could increase the efficiency of embryo production in older cows. We used cows aged 30 to 50 months or >120 months, which were defined as young or aged, respectively. The distribution of cortical granules in oocytes was affected by age as older cows had lower proportion of oocytes with mature cytoplasm containing evenly dispersed cortical granules compared to young cows. Although fertilization rates did not differ significantly between the two groups after in vitro fertilization (IVF), the rate of abnormal fertilization was higher, and the numbers of total and diploid blastocysts were lower for aged cows compared to young cows. However, in the embryos produced by ICSI, there was no significant difference in these parameters between young and aged cows. Although ICSI did not improve the blastocyst development rate, ICSI increased the proportion of diploid blastocysts in aged cows compared to IVF. In conclusion, maternal aging may negatively affect cytoplasmic maturation of bovine oocytes, which could be associated with abnormal fertilization or low developmental competence of oocytes. Our data also suggests beneficial effects of ICSI on the production of chromosomally normal embryos in aged cows.
In postpartum dairy cows, lipopolysaccharide (LPS) derived from gram-negative bacteria such as Escherichia coli causes uterine inflammation and leads to ovarian dysfunction. The aim of this study was ...to determine the effect of LPS on steroid production in bovine theca cells at different stages of follicular development. Theca cells isolated from pre- and post-selection follicles (PRFs, <8.5 mm in diameter, and POFs, >8.5 mm in diameter, respectively) of bovine ovaries were exposed to LPS under luteinizing hormone (LH) conditions, estradiol (E2) conditions or both conditions in vitro. Bovine theca cells expressed the LPS receptor gene complex: Toll-like receptor 4 (TLR4), CD14 and MD2. LPS suppressed progesterone (P4) and androstenedione (A4) production with downregulation of steroidogenic enzyme transcripts when theca cells were stimulated with LH. By contrast, LPS did not affect P4 or A4 production when theca cells were stimulated with E2. P4 and A4 production in theca cells from PRFs was suppressed by LPS as early as at 48 h of culture, whereas the effect of LPS on theca cells from POFs was observed at 96 h of culture. The results demonstrate that LPS inhibits steroid production in theca cells under LH conditions. Moreover, theca cells from POFs showed a slower response to LPS compared with that of theca cells from PRFs, which might imply a distinct effect of LPS on follicles at different developmental stages. These findings suggest a possible mechanism of ovarian dysfunction and subsequent infertility in cows with endometritis.
Uterine inflammatory diseases are a major cause of infertility in humans and domestic animals. The current findings that intrauterine lipopolysaccharide is absorbed in systemic circulation and ...attenuates ovarian cyclic activities could provide a basis for developing novel treatments to improve fertility.
Uterine inflammatory diseases are a major cause of infertility in humans and domestic animals. Circulating lipopolysaccharide (LPS), a bacterial endotoxin causing uterine inflammation, reportedly downregulates the hypothalamic-pituitary-gonadal axis to mediate ovarian dysfunction. In contrast, the mechanism whereby intrauterine LPS affects ovarian function has not been fully clarified. This study aimed to elucidate whether uterine exposure to LPS downregulates hypothalamic kisspeptin gene (Kiss1) expression, gonadotropin release, and ovarian function. Uterine inflammation was induced by intrauterine LPS administration to ovary-intact and ovariectomized female rats. As a result, plasma LPS concentrations were substantially higher in control rats until 48 h post injection, and the estrous cyclicity was disrupted with a prolonged diestrous phase. Three days post injection, the number of Graafian follicles and plasma estradiol concentration were reduced in LPS-treated rats, while numbers of Kiss1-expressing cells in the anteroventral periventricular nucleus and arcuate nucleus (ARC) were comparable in ovary-intact rats. Four days post injection, ovulation rate and plasma progesterone levels reduced significantly while gene expression of interleukin1β and tumor necrosis factor α was upregulated in the ovaries of LPS-treated rats that failed to ovulate. Furthermore, the number of Kiss1-expressing cells in the ARC and pulsatile luteinizing hormone (LH) release were significantly reduced in ovariectomized rats 24 h post injection. In conclusion, these results indicate that intrauterine LPS is absorbed in systemic circulation and attenuates ovarian function. This detrimental effect might be caused, at least partly, by the inhibition of ARC Kiss1 expression and LH pulses along with an induction of ovarian inflammatory response.
We investigated the association among endometrial hyperemia, uterine bacterial infection, and features of the large ovarian follicles in dairy cows. Genital organs were collected in a complete set at ...a slaughterhouse, and the degree of endometrial hyperemia was examined for the direct evaluation of uterine inflammation. The rate of bacterial infection in the uterus was higher in cows with endometrial hyperemia regardless of the severity of hyperemia, compared with cows without hyperemia. Moreover, the characteristics of the follicular fluid were changed in cows with uterine bacterial infection and included high concentrations of lipopolysaccharide and malondialdehyde (lipid peroxidation marker). These findings can be utilized as the basic information for the direct evaluation of the uterine inflammatory status in dairy cows.
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