Despite advances in our understanding of fertility, implantation failure remains a significant problem for both spontaneous and assisted pregnancies. Most research efforts concerning the process of implantation are embryo-centric, with a dearth of studies on endometrial factors. Currently, there are no practical and effective diagnostic tools available to precisely predict endometrial receptivity. Transcriptomics, a field based on microarray technology, has a number of procedures for clinical applications, although the functional relevance of most identified genes remains unclear. Importantly, RNA sequencing will further improve the precision and broaden the clinical use of the transcriptome by detecting previously undiscovered genes, which could be used to further our understanding of endometrial receptivity. In this review, potential biomarkers based on endometrium gene expression profiles of human endometrial receptivity were described and compared in natural and stimulated cycles toward discovering future prospects for personalized medical approaches. The intent of this synthesis is to provide researchers, doctors, and clinicians in the field with a better understanding of endometrium receptivity, promote further study in the transcriptome in embryo implantation, and ultimately, improve pregnancy outcome.

Summary Sentence

Transcriptomics is emerging as a powerful tool to identify potential molecular biomarkers for endometrial receptivity, thereby improving forecasts of pregnancy outcome during in vitro fertilization treatment.

Understanding factors that regulate zygotic genome activation (ZGA) is critical for determining how cells are reprogrammed to become totipotent or pluripotent. There is limited information regarding how this process occurs physiologically in early mammalian embryos. Here, we identify a mediator complex subunit, MED13, as translated duringmouse oocytematuration and transcribed early from the zygotic genome. Knockdown and conditional knockout approaches demonstrate that MED13 is essential for ZGA in the mouse, in part by regulating expression of the embryospecific chromatin remodeling complex, esBAF. The role of MED13 in ZGA is mediated in part by interactions with E2F transcription factors. In addition to MED13, its paralog, MED13L, is required for successful preimplantation embryo development. MED13L partially compensates for loss of MED13 function in preimplantation knockout embryos, but postimplantation development is not rescued by MED13L. Our data demonstrate an essential role for MED13 in supporting chromatin reprogramming and directed transcription of essential genes during ZGA.

Summary Sentence

MED13 is required for activating transcription of essential genes during the maternal to zygotic transition in the mouse.

In ruminants, uterine pulses of prostaglandin (PG) F_2α characterize luteolysis, while increased PGE₂/PGE₁ distinguish early pregnancy. This study evaluated intrauterine (IU) infusions of PGF_2α and PGE₁ pulses on corpus luteum (CL) function and gene expression. Cows on day 10 of estrous cycle received 4 IU infusions (every 6 h; n = 5/treatment) of saline, PGE₁ (2 mg PGE₁), PGF_2α (0.25 mg PGF_2α), or PGE₁ + PGF_2α. A luteal biopsy was collected at 30 min after third infusion for determination of gene expression by RNA-Seq. As expected, IU pulses of PGF_2α decreased (P < 0.01) P4 luteal volume. However, there were no differences in circulating P4 or luteal volume between saline, PGE₁, and PGE₁ + PGF_2α, indicating inhibition of PGF_2α-induced luteolysis by IU pulses of PGE₁. After third pulse of PGF_2α, luteal expression of 955 genes were altered (false discovery rate [FDR] < 0.01), representing both typical and novel luteolytic transcriptomic changes. Surprisingly, after third pulse of PGE₁ or PGE₁ + PGF_2α, there were no significant changes in luteal gene expression (FDR > 0.10) compared to saline cows. Increased circulating concentrations of the metabolite of PGF_2α (PGFM; after PGF_2α and PGE₁ + PGF_2α) and the metabolite PGE (PGEM; after PGE₁ and PGE₁ + PGF_2α) demonstrated that PGF_2α and PGE₁ are entering bloodstream after IU infusions. Thus, IU pulses of PGF_2α and PGE₁ allow determination of changes in luteal gene expression that could be relevant to understanding luteolysis and pregnancy. Unexpectedly, by third pulse of PGE₁, there is complete blockade of either PGF_2α transport to the CL or PGF_2α action by PGE₁ resulting in complete inhibition of transcriptomic changes following IU PGF_2α pulses.

Summary Sentence

Treatment with PGF_2α induced dramatic changes in expression of 955 genes, based on RNA-Seq after the third PGF_2α pulse (FDR < 0.01), whereas simultaneous treatment with pulses of PGE₁ blocked luteolysis and gene expression induced by PGF_2α.

Intrauterine adhesion (IUA) is characterized by endometrial fibrosis, which ultimately leads to menstrual abnormalities, infertility, and recurrent miscarriages. The Shh/Gli2 pathway plays a critical role in tissue fibrogenesis and regeneration; Gli2 activation induces profibrogenic effects in various tissues, such as the liver and kidney. However, the role of Gli2 in endometrial fibrosis remains unknown. The purpose of this study was to test the hypothesis that activated Gli2 promotes endometrial fibrosis. Endometrial samples from moderate and severe IUA patients exhibited significantly enhanced expression of Gli2 compared with normal endometrial samples and mild IUA samples. Transfection with overactive Gli2 plasmids induced higher fibrosis-related protein expression, while blocking Gli2 signaling with cyclopamine caused the opposite effect in endometriotic stromal cells (ESCs), including inducing cell-cycle arrest. Menstrual-derived stem cell conditioned medium (MenSCs-CM) reduced endometrial fibrosis by reducing Gli2 protein levels and causing cell-cycle arrest in ESCs through granulocyte-colony stimulating factor (G-CSF). The effect was weakened after neutralization with a G-CSF antibody. Gli2 overexpression reduced the effects of MenSC-CM and G-CSF on fibrosis and cell-cycle progression in vitro. The antifibrotic effect of G-CSF was also observed in murine model. These findings demonstrate that Gli2 signaling promotes endometrial fibrosis, and the inhibition of Gli2 through MenSCs-secreted G-CSF may be of therapeutic value for managing endometrial fibrosis.

Summary Sentence

G-CSF secreted by endometrial stem cell could attenuate endometrial fibrosis via Gli2 protein.

Human female reproductive aging features declining ovarian follicle reserve and oocyte quality, and rising levels of circulating follicle-stimulating hormone (FSH).We determined the effects of elevated FSH on oocyte–embryo development in mature mice exhibiting premature infertility caused by progressively rising transgenic human FSH (TgFSH) levels. Oocyte–embryo developmental competence and quality were examined using oocyte maturation and aneuploidy rates, biomarkers of oocyte quality, and reciprocal embryo transfers assessed for implantation and pregnancy. In vitro maturation suggested that TgFSH exposure only hindered oocyte developmental competence in old females, as significantly more oocytes from ≥12-month-old TgFSH females remained at germinal vesicle stage compared with age-matched control oocytes. Aneuploidy rates were equivalent in oocytes from aging TgFSH compared with wildtype females. Cumulus cell expression levels of candidate biomarker Inhba, Egfr, and Rgs2 transcripts were elevated in associated aneuploid vs euploid oocytes from both TgFSH and wildtype females. In vivo, embryos transferred from subfertile 6-month-old TgFSH females to wildtype recipients yielded normal implantation rates and more pups born compared with controls. Transfer of wildtype embryos rescued the fertility of 6-month-old TgFSH-recipient females, although pup birth weight was reduced in TgFSH vs wildtype recipients. Our current findings show that elevated FSH had minimal disruption of either embryo developmental capacity or uterine function when examined in isolation, and the subfertility of TgFSH female mice was not caused by altered oocyte aneuploidy or quality.

Summary Sentence

Elevated FSH causes minimal disruption of either embryo developmental capacity or uterine function when examined in isolation, and the subfertility of TgFSH female mice is not caused by altered oocyte aneuploidy or quality.

Exportin 1 (XPO1) is a nuclear transport receptor involved in the nuclear export ofmajority proteins in somatic cells. In mammalian oocytes, however, only the presence of XPO1 has been reported at mRNA and protein levels, and the definitive functions of XPO1 and its effects on the meiotic maturation of oocytes have never been directly examined. In the present study, the expression state and the nuclear-export function of porcine XPO1 were analyzed in porcine oocytes. In addition, we investigated the effects of the overexpression and inhibition of XPO1 on meiotic regulation in fullgrown and growing oocytes by mRNA injection and inhibitor treatment. Endogenous XPO1 was stably expressed in porcine oocytes during the germinal vesicle (GV) stage, and the expression of exogenous XPO1 significantly decreased the nuclear localization of XPO1 cargos, snurportin 1, and WEE1B. Inhibition of XPO1 by a specific inhibitor, leptomycin B, delayed the GV breakdown (GVBD), whereas the overexpression of XPO1 by mRNA injection accelerated the GVBD. XPO1 overexpression overcame the meiotic arrest induced by WEE1B expression in full-grown oocytes. Surprisingly, the GVBD of porcine growing oocytes, which could not resume meiosis by the maturation culture in vitro, was induced by the expression of exogenous XPO1. These results showed the presence of XPO1 and its function as a nuclear export receptor in mammalian oocytes, including growing oocytes, and they suggest that the regulation of nuclear transport has a large influence on the GV maintenance and meiotic resumption of oocytes.

Summary Sentence

Exportin 1 functions as a nuclear export receptor and is involved in meiotic resumption of porcine full-grown and growing oocytes via its nuclear-export function.

The Ca²⁺ entry mechanism that sustains the Ca²⁺ oscillations in fertilized pig oocytes was investigated. Stromal interaction molecule 1 (STIM1) and ORAI1 proteins tagged with various fluorophores were expressed in the oocytes. In some cells, the Ca²⁺ stores were depleted using cyclopiazonic acid (CPA); others were inseminated. Changes in the oocytes' cytosolic free Ca²⁺ concentration were monitored, while interaction between the expressed fusion proteins was investigated using fluorescence resonance energy transfer (FRET). Store depletion led to an increase of the FRET signal in oocytes co-expressing mVenus-STIM1 and mTurquoise2-ORAI1, indicating that Ca²⁺ release was followed by an interaction between these proteins. A similar FRET increase in response to CPA was also detected in oocytes co-expressing mVenus-STIM1 and mTurquoise2-STIM1, which is consistent with STIM1 forming punctae after store depletion. ML-9, an inhibitor that can interfere with STIM1 puncta formation, blocked store-operated Ca²⁺ entry (SOCE) induced by Ca²⁺ add-back after a CPA treatment; it also disrupted the Ca²⁺ oscillations in fertilized oocytes. In addition, oocytes overexpressing mVenus-STIM1 showed high-frequency Ca²⁺ oscillations when fertilized, arguing for an active role of the protein. High-frequency Ca²⁺ oscillations were also detected in fertilized oocytes co-expressing mVenus-STIM1 and mTurquoise2-ORAI1, and both of these high-frequency Ca²⁺ oscillations could be stopped by inhibitors of SOCE. Importantly, in oocytes co-expressing mVenus-STIM1 and mTurquoise2-ORAI1, we were also able to detect cyclic increases of the FRET signal indicating repetitive interactions between STIM1 and ORAI1. The results confirm the notion that in pig oocytes, SOCE is involved in the maintenance of the repetitive Ca²⁺ transients at fertilization.

Summary Sentence

The sperm-induced Ca²⁺ oscillations in pig oocytes are sustained by store-operated Ca²⁺ entry, which is mediated by repetitive interactions between STIM1 and ORAI1 proteins

Hyperandrogenism is associated with hyperinsulinemia and insulin resistance in adult females. We tested whether androgens dysregulate pancreatic beta cell function to induce hyperinsulinemia through transcriptional regulation of insulin gene (Ins) in the islets. Adult female Wistar rats implanted with dihydrotestosterone (DHT; 7.5-mg, 90-d release) or placebo pellets were examined after 10 weeks. DHT exposure increased plasma DHT levels by 2-fold similar to that in polycystic ovary syndrome in women. DHT exposure induced hyperinsulinemia with increased HOMA-IR index in fasting state and glucose intolerance and exaggerated insulin responses following glucose tolerance test. DHT females had no change in islet number, size and beta cell proliferation/apoptosis but exhibited significant mitochondrial dysfunction (higher ADP/ATP ratio, decreased mtDNA copy number, increased reactive oxygen production and downregulation of mitochondrial biogenesis) and enhanced glucose-stimulated insulin secretion. Ins expression was increased in DHT islets. In vitro incubation of control islets with DHT dose dependently stimulated Ins transcription. Analysis of Ins1 gene revealed a putative androgen responsive element in the promoter. Chromatinimmunoprecipitation assays showed that androgen receptors bind to this element in response to DHT stimulation. Furthermore, reporter assays showed that the promoter element is highly responsive to androgens. Insulin-stimulated glucose uptake in skeletal muscle was decreased with associated decrease in IRβ expression in DHT females. Our studies identified a novel androgenmediated mechanism for the control of Ins expression via transcriptional regulation providing a molecular mechanism linking elevated androgens and hyperinsulemia. Decreased IRβ expression in the skeletal muscles may contribute, in part, to glucose intolerance in this model.

Summary Sentence

Elevated androgen levels lead to hyperinsulinemia despitemitochondrial dysfunction in pancreatic islets; hyperandrogenism directly controls the expression of Ins1 in pancreatic islets by positively regulating Ins1 transcription which might play an underlying role in hyperinsulinemia.

Following proliferation of oogonia inmammals, great numbers of germ cells are discarded, primarily by apoptosis, while the remainder form primordial follicles (the ovarian reserve) that determine fertility and reproductive lifespan. More massive, rapid, and essentially total loss of oocytes, however, occurs when the transcription factor Lhx8 is ablated—though the cause and mechanism of germ cell loss from the Lhx8-/- ovaries has been unknown. We found that Lhx8-/- ovaries maintain the same number of germ cells throughout embryonic development; rapid decrease in the pool of oocytes starts shortly before birth. The loss results from activation of autophagy, which becomes overwhelming within the first postnatal week, with extracellular matrix proteins filling the space previously occupied by follicles to produce a fibrotic ovary. Associated with this process, as early as a few days before birth, Lhx8-/- oocytes failed to repair DNA damage—which normally occurs when meiosis is initiated during embryonic development; and DNA damage repair genes were downregulated throughout the oocyte short lifespan. Based on gene expression analyses and morphological changes, we propose a model in which lineage-restricted failure of DNA repair triggers germ cell autophagy, causing premature depletion of the ovarian reserve in Lhx8-/- mice.

Summary Sentence

Ablation of Lhx8 causes premature loss of germ cells by autophagy associated with impairment of DNA damage repair during meiosis.

Progesterone is a steroid hormone secreted from the corpus luteum (CL), which is responsible for establishment and maintenance of pregnancy. Early embryonic mortality often occurs due to inadequate regulation of uterine prostaglandin (PG) F2α secretion, leading to a decrease in progesterone and loss of pregnancy. The objective of the current study was to determine the effects of fish meal supplementation on luteal sensitivity to intrauterine infusions of PGF_2α. Nonlactating beef cows received corn glutenmeal or fishmeal supplementation for 60 days. Cows were administered four intrauterine infusions of 0.25 mL saline at 6-h intervals (n = 6 corn gluten meal; n = 5 fish meal) or two doses of 0.5 mg PGF_2α in 0.25 mL saline at 12-h intervals (n = 11 corn gluten meal; n = 11 fishmeal) commencing on days 10 to 12 of the estrous cycle. At time of each infusion, luteal biopsies were collected to determine the effects of supplementation on expression of immediate early and steroidogenic genes involved in cholesterol transport and progesterone biosynthesis. Transrectal ultrasonography was performed to measure diameter of CL, and blood samples were collected to determine serum progesterone. Intrauterine infusion of PGF_2α resulted in upregulation or no change in FOS, NR4A1, and 3BHSD and downregulation in LDLR, STARD1, and CYP11A1. Although CL diameter decreased, infusion of PGF_2α resulted in functional regression in 91% of cows supplemented with corn gluten meal, and only 46% for fish meal supplemented animals. Results demonstrate that fish meal supplementation alters luteal sensitivity to PGF_2α, which may affect fertility.

Summary Sentence

Fish meal supplementation alters luteal sensitivity to prostaglandin F2α in the bovine.

Porcine parvovirus (PPV) is a major virus that leads to fetal death in swine. However, the effects of PPV infection on sows are poorly understood. The aim of this study was to investigate the effects of PPV on porcine steroidogenic luteal cells (SLCs) survival and functions and underlying mechanisms. In vivo experiment results showed that artificial infection of PPV significantly reduced the concentration of serum progesterone and induced histopathological lesions and SLCs apoptosis in porcine corpora luteum. In in vitro cultured primary porcine SLCs, PPV could infect and replicate in SLCs and induced SLCs apoptosis through mitochondria, but not the death receptor, mediated apoptosis pathway. Meanwhile, PPV infection also decreased progesterone production in SLCs. Moreover, PPV infection could increase active p53 transcriptional activity and protein expression as well as promoting p53 translocation to nucleus. Using the p53-specific pharmacological inhibitor (pifithrin-α) and siRNA could partially attenuate PPV-induced Bax upregulation, caspase-3 activation, apoptosis, and the reduction of progesterone production in primary porcine SLCs. Furthermore, the phosphorylation of p38 mitogen-activated protein kinase (MAPK) was also increased in PPV-infected SLCs. Pretreatment with p38 MAPK inhibitor (SB203580) suppressed PPV-induced p53 accumulation and translocation, SLCs apoptosis, and progesterone production reduction. In summary, these findings indicate that PPV could induce SLCs apoptosis and a decrease of progesterone production in vivo and in vitro via p38 MAPK signaling and p53-dependent mitochondrial pathway, which provides the potential clinical therapy methods for PPV infection.

Summary Sentence

Porcine parvovirus induced porcine steroidogenesis luteal cells apoptosis through the activation of p38MAPK, p53, andmitochondria-mediated death pathway, resulting in a decrease of progesterone production in vivo and in vitro.

Seipin is an integral endoplasmic reticulum (ER) membrane protein encoded by Berardinelli– Seip congenital lipodystrophy type 2 (BSCL2/Bscl2) gene. Most litters (59%) from Bscl2^-/- dams mated with wild type (WT) (Bscl2^+/+) males did not survive postnatal day 5 (PND5) and pups (Bscl2^+/-) lacked milk in their stomachs. The survived litters had reduced pup survival rate at PND21. It was hypothesized that seipin was critical for lactation. Bscl2 was upregulated and highly detected in the lactation day 1 (LD1) WT mammary gland alveolar epithelial cells. LD1 Bscl2^-/- mammary glands lacked adipocytes and alveolar clusters and had varied alveolar morphology: from interconnected mammary gland alveoli with dilated lumen and sloughed epithelial cells to undifferentiated mammary gland alveoli with unexpanded lumen. Comparable levels of whey acidic protein (WAP, a major component in rodent milk) staining and Nile Red lipid droplet staining between WT and Bscl2^-/- LD1 alveolar epithelial cells indicated normal milk protein synthesis and lipid syntheses in LD1 Bscl2^-/- mammary glands. Significantly reduced percentage of larger lipid droplets was detected in LD1 Bscl2^-/- alveoli with unexpanded lumen. There was no obviously impaired proliferation detected by PCNA staining but increased apoptosis detected by cleaved caspase-3 staining in LD1 Bscl2^-/- alveolar epithelial cells. Increased expression of protein disulfide isomerase and binding immunoglobulin protein in the LD1 Bscl2^-/- mammary gland alveolar epithelial cells indicated increased ER stress. This study demonstrates increased ER stress and apoptosis in LD1 Bscl2^-/- mammary gland alveolar epithelial cells and reveals a novel in vivo function of seipin in lactation.

Summary Sentence

Our findings that pups from Bscl2^-/- dams lacked milk and had reduced survival rate as well as that LD1 Bscl2^-/- mammary gland alveolar epithelial cells had increased ER stress and apoptosis reveal a novel in vivo function of seipin in lactation.

Malnutrition is a global threat to pregnancy health and impacts offspring development. Establishing an optimal pregnancy environment requires the coordination of maternal metabolic and immune pathways, which converge at the gut. Diet, metabolic, and immune dysfunctions have been associated with gut dysbiosis in the nonpregnant individual. In pregnancy, these states are associated with poor pregnancy outcomes and offspring development. However, the impact of malnutrition on maternal gut microbes, and their relationships with maternal metabolic and immune status, has been largely underexplored. To determine the impact of undernutrition and overnutrition on maternal metabolic status, inflammation, and the microbiome, and whether relationships exist between these systems, pregnant mice were fed either a normal, calorically restricted (CR), or a high fat (HF) diet. In late pregnancy, maternal inflammatory and metabolic biomarkers were measured and the cecal microbiome was characterized. Microbial richness was reduced in HF mothers although they did not gain more weight than controls. First trimester weight gain was associated with differences in the microbiome. Microbial abundance was associated with altered plasma and gut inflammatory phenotypes and peripheral leptin levels. Taxa potentially protective against elevated maternal leptin, without the requirement of a CR diet, were identified. Suboptimal dietary conditions common during pregnancy adversely impact maternal metabolic and immune status and the microbiome. HF nutrition exerts the greatest pressures on maternal microbial dynamics and inflammation. Key gut bacteria may mediate local and peripheral inflammatory events in response to maternal nutrient and metabolic status, with implications for maternal and offspring health.

Summary Sentence

Malnutrition alters immune and metabolic systems in pregnancy, and gut microbes may mediate the maternal response to these suboptimal dietary exposures.

The reproductive consequences of global warming are not currently understood. In order to address this issue, we have examined the reproductive consequences of exposingmalemice to a mild heat stress. For this purpose, adultmalemice were exposed to an elevated ambient temperature of 35°C under two exposure models. The first involved acute exposure for 24 h, followed by recovery periods between 1 day and 6 weeks. The alternative heating regimen involved a daily exposure of 8 h for periods of 1 or 2 weeks. In our acute model, we identified elevated sperm mitochondrial ROS generation (P < 0.05), increased sperm membrane fluidity (P < 0.05), DNA damage in the form of single-strand breaks (P<0.001), and oxidative DNAdamage (P<0.05), characteristic of an oxidative stress cascade. This DNA damage was detected in pachytene spermatocytes (P < 0.001) and round spermatids (P < 0.001) isolated from testes after 1 day heat recovery. Despite these lesions, the spermatozoa of heat-treated mice exhibited no differences in their ability to achieve hallmarks of capacitation or to fertilize the oocyte and support development of embryos to the blastocyst stage (all P>0.05). Collectively, our acute heat stress model supports the existence of heat susceptible stages of germ cell development, with the round spermatids being most perturbed and spermatogonial stem cells exhibiting resistance to this insult. Such findings were complemented by our chronic heat stress model, which further supported the vulnerability of the round spermatid population.

Summary Sentence

Environmental heating induces a state of oxidative stress in the male germ line, affecting multiple germ cell types; primarily the round spermatid and pachytene spermatocyte populations.