Successful pregnancy is dependent on a number of essential events, including embryo implantation, decidualization, and placentation. Failure of the above process may lead to pregnancy-related complications, including preeclampsia, gestational diabetes mellitus, preterm birth, and fetal growth restriction, may affect 15% of pregnancies, and lead to increased mortality and morbidity of pregnant women and perinatal infants, as well as the occurrence of short-term and long-term diseases. These complications have distinct etiology and pathogenesis, and the present comprehension is still lacking. Post-translational modifications are important events in epigenetics, altering the properties of proteins through protein hydrolysis or the addition of modification groups to one or more amino acids, with different modification states regulating subcellular localization, protein degradation, protein–protein interaction, signal transduction, and gene transcription. In this review, we focus on the impact of various post-translational modifications on the progress of embryo and placenta development and pregnancy-related complications, which will provide important experimental bases for exploring new insights into the physiology of pregnancy and pathogenesis associated with pregnancy complications.

Summary Sentence

An in-depth understanding of post-translational modifications associated with normal pregnancy and related complications will provide an important experimental basis for further insights of physiological and pathological pregnancy.

Extracellular vesicles, particularly exosomes, play a pivotal role in the cellular mechanisms underlying cancer. This review explores the various functions of exosomes in the progression, growth, and metastasis of cancers affecting the male and female reproductive systems. Exosomes are identified as key mediators in intercellular communication, capable of transferring bioactive molecules such as microRNAs, proteins, and other nucleic acids that influence cancer cell behavior and tumor microenvironment interactions. It has been shown that non-coding RNAs transported by exosomes play an important role in tumor growth processes. Significant molecules that may serve as biomarkers in the development and progression of male reproductive cancers include miR-125a-5p, miR-21, miR-375, the miR-371 ∼ 373 cluster, and miR-145-5p. For female reproductive cancers, significant microRNAs include miR-26a-5p, miR-148b, miR-205, and miRNA-423-3p. This review highlights the potential of these noncoding RNAs as biomarkers and prognostics in tumor diagnostics. Understanding the diverse roles of exosomes may hold promise for developing new therapeutic strategies and improving treatment outcomes for cancer patients.

Summary Sentence

Research into exosomal ncRNAs in reproductive cancers is crucial because it combines fundamental biological insights with immediate clinical applications. It holds the potential to revolutionize early detection, treatment personalization, and the understanding of metastatic and resistance mechanisms, ultimately improving outcomes for patients with these challenging malignancies.

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Bovine viral diarrhea virus (BVDV) infection during pregnancy is a significant contributor to reproductive failures in cattle. The bovine receptor for BVDV (CD46) was previously edited with a six amino acid substitution (G₈₂QVLAL to A₈₂LPTFS) and shown to have significantly reduced BVDV susceptibility in a Gir heifer calf. Since a role for CD46 has been proposed in mammalian fertilization, our objective was to assess the edited heifer's fertilization rates, early embryonic development, and germline transmission conformation of the edit. Cumulus oocyte complexes were collected from the edited heifer and unedited females, fertilized with semen from an unedited bull and cultured until the blastocyst stage. Ultrasound examinations and serum progesterone concentration were also monitored to confirm estrous cyclicity in the CD46-edited heifer. Estrous cyclicity was normal with visualization of a corpus luteum and elevated progesterone concentrations. Fertilization rates and blastocyst development were not different in oocytes from edited and unedited controls. Genome sequence analysis of blastocysts confirmed germline transmission of either edited allele from the heifer. Subsequently, the CD46-edited heifer was artificially inseminated with semen from an unedited Gir bull and fertility status was confirmed with a diagnosed conception at Day 35 of gestation. Thus, a six amino acid substitution in CD46 did not negatively affect fertilization of edited oocytes or early embryonic development when fertilized with semen from an unedited bull. An edited bull is still needed to similarly evaluate reproductive function of sperm cells carrying this CD46 edit.

Summary Sentence

Fertility is not affected in the CD46-edited heifer and the CD46 edit is shown to be heritable.

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Developing embryos are susceptible to fluctuations in the nutrients and metabolites concentrations within the reproductive tract, which can lead to alterations in their developmental trajectory. Ketotic dairy cows have diminished fertility, and elevated levels of the ketone body beta-hydroxybutyrate (BHB) have been associated with poor embryonic development. We used an in vitro model based on either in vitro fertilization (IVF) or parthenogenesis to investigate the effects of BHB on the preimplantation bovine embryo development, epigenome, and transcriptome. Embryo culture medium was supplemented with BHB at a similar concentration to that present in the blood of cows suffering with severe ketosis, followed by analysis of blastocysts formation rate, diameter, total number of cells, levels of H3K9 beta-hydroxybutyrylation (H3K9bhb), apoptosis, and transcriptional alterations. As a result, we observed that BHB reduced the blastocysts rates, the diameter and the total number of cells in both parthenotes and IVF embryos. Exposure to BHB for either 3 or 7 days greatly increased the H3K9bhb levels in parthenotes at the 8-cells and blastocyst stages, and affected the expression of HDAC1, TET1, DNMT1, KDM6B, NANOG, and MTHFD2 genes. Additionally, culture of IVF embryos with BHB for 7 days dramatically increased H3K9bhb and reduced NANOG in blastocysts. RNA-seq analysis of IVF blastocysts revealed that BHB modulated the expression of 118 genes, which were involved with biological processes such as embryonic development, implantation, reproduction, proliferation, and metabolism. These findings provided valuable insights into the mechanisms through which BHB disrupts preimplantation embryonic development and affects the fertility in dairy cows.

Summary Sentence

Beta-hydroxybutyrate induces histone lysine β-hydroxybutyrylation in parthenotes and IVF-embryos, decrease their development to blastocyst, and alters the transcription of genes involved in implantation, reproduction, proliferation, and metabolism.

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Intrauterine adhesions (IUA) represent a prevalent uterine endometrial disorder frequently correlated with menstrual irregularities and infertility. Some members of the secretoglobin(SCGB) family have demonstrated anti-fibrotic effects, however, the specific role of SCGB1D4, one of the family members, in anti-fibrosis remains unclear. This study aimed to investigate the expression of SCGB1D4 in IUA tissues, validate the role of SCGB1D4 in endometrial fibrosis, and assess its potential therapeutic significance by analyzing clinical features and constructing rat and cell models. Clinical characteristics of patients with intrauterine adhesions (IUA) were compared and analyzed against control subjects. Additionally, a rat uterine adhesion model was successfully established using a combination of mechanical injury and infection. The expression levels of SCGB1D4 in patient tissues and animal models were detected through immunohistochemistry, Western blot, and real-time fluorescence quantitative PCR, and the changes in fibrosis markers COL1A1 and α-SMA were also evaluated. Furthermore, human endometrial stromal cell lines (HESCs) induced by transforming growth factor-β-1 conversion were differentiated into myofibroblasts to establish cell models of intrauterine adhesion. We detected the expression of SCGB1D4 and fibrosis-related factors by real-time fluorescence quantitative PCR and Western blot. Cell proliferation and cell cycle changes were assessed using flow cytometry and CCK8. IUA patients showed increased miscarriage rates and decreased endometrial thickness. Clinical tissue specimens revealed significantly lower expression of SCGB1D4 in the endometrial tissues of IUA patients, accompanied by a notable increase in COL1A1 and α-SMA. The established rat model of intrauterine adhesion exhibited decreased expression of SCGB1D4 and a significant increase in fibrosis. After overexpression of SCGB1D4 on the IUA cell model, SCGB1D4 expression was elevated, while COL1A1 and α-SMA expression was significantly reduced. Cell proliferation was inhibited and cell cycle distribution was altered. This study has confirmed the low expression of SCGB1D4 in patients with IUA, as well as in animal and cell models. Furthermore, the overexpression of SCGB1D4 in a cell model of IUA demonstrates that it may play a key role in inhibiting fibrosis. SCGB1D4 holds promise as a potential therapeutic target for IUA, providing a new avenue for overcoming fertility issues caused by IUA.

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Altered peristaltic and ciliary dysfunction is a feature of females with endometriosis. To further explore this premise, we examined the ampulla of rhesus macaques (Macaca mulatta) with and without spontaneous endometriosis for the expression of adenylate kinase 7 (AK7), a mitochondrial-dwelling nucleotide converting enzyme with critical roles in cellular kinesis, forkhead protein box J1 (FOXJ1), a marker of cilia abundance, and Anoctamin 1 (ANO1) as a marker of both smooth muscle contraction and ciliogenesis. We further performed an in vitro experiment that treated ampullary segments with peritoneal fluid from animals with and without endometriosis. We report significantly downregulated expression of ANO1 in the ampulla of monkeys with endometriosis (in vivo), and in the ampullary segments exposed to peritoneal fluid of animals with endometriosis. We did not observe statistically significant differences in the expression of AK7 or FOXJ1 both in vivo and in vitro. This highlights potentially essential roles of ANO1 in the oviduct, the dampening of which may lead to a specific subtype of endometriosis-caused subfertility.

Summary Sentence

Anoctamin1 is a calcium-activated chloride channel that orchestrates smooth muscle contractions, ciliary beating, and epithelial secretion in the respiratory, gastrointestinal, and reproductive organs. We observed the inhibition of this channel in the ampulla of monkeys with spontaneous endometriosis, and were able to replicate this downregulation in vitro by using peritoneal fluid collected from endometriosis cases.

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Endometriosis is an estrogen dependent disease, which is related to infertility. Decidualization is a prerequisite for successful implantation of human embryos, and endometriosis affects the occurrence of decidualization. However, the mechanism that affects decidualization in endometriosis is not fully understood. Here, we find that Aurora kinase A (AURKA) is upregulated in the eutopic endometrium of endometriosis. AURKA inhibits the decidualization of stromal cells in the eutopic endometrium of endometriosis. Furthermore, in animal experiments, AURKA promotes endometriosis and inhibits decidualization in mice with endometriosis, leading to decreased expression of decidualization markers, such as prolactin, insulin-like growth factor-binding protein-1, and desmin. Afterwards, we find that nuclear factor-κB (NF-κB) p65 is a new substrate of AURKA. AURKA interacts with p65 to promote its phosphorylation and nuclear translocation. Meanwhile, AURKA enhances the protein stability of p65 by prolonging its half-life. In summary, AURKA inhibits the decidualization of the eutopic endometrium in patients with endometriosis by regulating p65, which may provide new ideas for improving decidualization defect in patients with endometriosis.

Summary Sentence

AURKA causes endometrial associated decidualization damage through phosphorylation of p65.

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Intraflagellar transport 25 is a component of the intraflagellar transport 25-B complex. In mice, even though this intraflagellar transport component is not required for cilia formation in somatic cells, it is essential for sperm formation. However, the intracellular localization of this protein in male germ cells is not known given no reliable antibodies are available for histologic studies, and the dynamic trafficking in the developing sperm flagella is not clear. To examine localization of the protein in male germ cells and further investigate the mechanism of intraflagellar transport in sperm formation, particularly to look into the dynamic trafficking of the protein, we generated a mouse intraflagellar transport 25–green fluorescent protein knock-in mouse model using the clustered regularly interspaced short palindromic repeats/clustered regularly interspaced short palindromic repeats associated protein 9 system, with the mouse intraflagellar transport 25 protein fused with a green fluorescent protein tag in the C-terminus. Three independent lines were analyzed. Western blotting using both anti-intraflagellar transport 25 and anti-green fluorescent protein antibodies showed that the intraflagellar transport 25–green fluorescent protein fusion protein was highly abundant only in the testis, which is consistent with the endogenous intraflagellar transport 25 protein. Examination of localization of the intraflagellar transport 25–green fluorescent protein in isolated germ cells revealed that the fusion protein was present in the cytoplasm of spermatocytes and round spermatids and a strong signal was present in the developing sperm flagellar. The homozygous knock-in mice had normal spermatogenesis, fertility and sperm parameters. Diffusion analysis of intraflagellar transport 25 within the developing flagellar revealed the presence of both mobile and immobile fractions as revealed by fluorescence recovery after photobleaching. Kymograph and fluorescence recovery after photobleaching analyses demonstrate the transport of intraflagellar transport 25–green fluorescent protein within the developing tail demonstrate no apparent preference for trafficking toward and away from the cell body. The speed of trafficking depends on the stage of sperm development, ranging from highly mobile unrestricted diffusion initially, mobile punctate structures in developing sperm, and immobile punctate structures in mature sperm. Our studies demonstrate that mouse intraflagellar transport 25 travels along the developing sperm flagella in two directions that might be essential for functional sperm formation.

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Developmental exposure to environmental chemicals perturbs establishment and maintenance of the ovarian reserve across the reproductive lifetime, leading to premature follicle depletion and ovarian aging. Considering humans are exposed to a complex mixture of environmental chemicals, real-life models assessing their cumulative impact on the ovarian reserve are needed. Biosolids are a source of a real-life mixture of environmental chemicals. While earlier studies demonstrated that grazing pregnant sheep on biosolids-treated pastures did not influence establishment of the ovarian reserve in fetal life, its impact on subsequent depletion of ovarian reserve during reproductive life of offspring is unknown. We hypothesized that developmental exposure to biosolids accelerates depletion of ovarian reserve. Ovaries were collected from F1 juveniles (9.5 weeks) and adults (2.5 years) born to F0 ewes grazed on control inorganic fertilizer pastures or biosolids-treated pastures from before conception and throughout gestation. The impact on follicular density, activation rate, and anti-Müllerian hormone (mediator of activation) expression by immunohistochemistry was determined. Activation rate was increased in F1 biosolids-treated pastures juveniles with a corresponding reduction in primordial follicle density. In contrast, activation rate and ovarian reserve were similar between control and F1 biosolids-treated pastures adults. The density of anti-Müllerian hormone-positive antral follicles was lower in biosolids-treated pastures juveniles, whereas anti-Müllerian hormone expression tended to be higher in antral follicles of biosolids-treated pastures adults, consistent with the changes in the ovarian reserve. These findings of detrimental effects of developmental exposure to biosolids during juvenile life that normalizes in adults is supportive of a shift in activation rate likely related to peripubertal hormonal changes.

Summary Sentence

Maternal preconceptional and gestational exposure to biosolids programs increased activation rate and reduced ovarian reserve in juvenile offspring that is normalized by adult life, which is likely related to changes during peripuberty.

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Natural ovarian aging is one of the major causes for declining fertility in female animals, which has become an insurmountable issue in human reproduction clinics and assisted reproductive technology procedures. Nevertheless, the molecular basis of oocyte aging remains poorly understood, and feasible improvement strategies are unavailable. In the present study, in vivo supplementation of pyrroloquinoline-quinone effectively elevated the fecundity of reproductively aged mice by balancing hormonal secretion, harmonizing the estrus cycle, and eliminating ovarian fibrosis. Moreover, oocyte quality also increased in aged mice after pyrroloquinoline-quinone administration from various aspects, including nuclear and cytoplasmic maturation competency, fertilization capacity, and pre-implantation embryonic development potential. Transcriptomic analysis identified target pathways that might mediate pyrroloquinoline-quinone's effects in aged oocytes. Specifically, it was demonstrated that pyrroloquinoline-quinone supplementation restored the mitochondrial dynamics and lysosomal function to remove excessive reactive oxygen species and suppress apoptosis in aged oocytes. Jointly, these findings demonstrate pyrroloquinoline-quinone administration is an efficacious method to restore the compromised ovary function and damaged oocyte quality in reproductively aged mice, which might be a potential clinical therapy for women of advanced maternal age with infertility.

Summary Sentence

Jointly, these findings demonstrate PQQ administration is an efficacious method to restore the compromised ovary function and damaged oocyte quality in reproductively aged mice, which might be a potential clinical therapy for women of advanced maternal age with infertility.

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Preeclampsia (PE) is a condition of pregnancy in which symptoms of hypertension develop after 20 weeks of gestation. it can lead to placental dysfunction, maternal and perinatal mortality and morbidity. The incidence of PE is increasing, posing a serious threat to the lives of pregnant women and their unborn children. Currently, most of the research on the pathogenesis of PE has focused on placenta, However, maternal decidualization is the basis for placental formation and growth. Chondroitin sulfate proteoglycan 4 (CSPG4) is a transmembrane protein that plays a role in cell proliferation, invasion, and migration. However, its function during decidualization is not yet understood. In this study, we investigated the role of CSPG4 and found that its expression was significantly down-regulated in the decidual tissue of patients with severe PE compared to normal pregnant women. During artificially induced decidualization, CSPG4 expression was significantly increased. Knockdown of CSPG4 by small interfering RNA inhibited decidualization, which, in turn, inhibited the invasion of trophoblast cells. In both pseudopregnant and pregnant mice, endometrial stromal cells proliferated rapidly and Cspg4 expression increased during decidualization. Therefore, we believe that CSPG4 plays a crucial role in the process of decidualization. The defect in decidualization caused by abnormal CSPG4 expression could lead to insufficient trophoblast invasion, ultimately contributing to the occurrence of PE.

Summary Sentence

CSPG4 plays a crucial role in decidualization, the defect caused by abnormal CSPG4 expression could lead to insufficient trophoblast invasion, ultimately contributing to the occurrence of preeclampsia.

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Visfatin regulates energy homeostasis, metabolism, inflammation, and reproduction via the hypothalamus-pituitary-ovary axis. Our previous study showed the visfatin gene and protein expression in the human placenta. This study aimed to investigate the in vitro effect of visfatin on the proliferation and apoptosis of placental JEG-3 and BeWo cells but also in villous explants collected from normal pregnancies and complicated by intrauterine growth restriction (IUGR), preeclampsia (PE), and gestational diabetes mellitus (GDM). We studied placenta cells viability, proliferation, cell cycle, proliferation/apoptotic factors and insulin receptor (INSR) expression, DNA fragmentation, CASP3/7 activity, and phosphorylation of ERK1/2, AKT, AMPKα, STAT3 with their involvement after pharmacological inhibition in visfatin action on proliferation and apoptosis. Visfatin (1, 10, 100 ng/mL) decreased the viability and proliferation of JEG-3 after 48 h, and a similar effect was observed via co-administration of visfatin (10 ng/mL) and insulin (10 ng/mL) in JEG-3 and BeWo after 48 h and 72 h, respectively. Visfatin reduced the transition from the G2/M phase, and expression of PCNA or cyclins D, E, A, and B in JEG-3 and PCNA in normal, IUGR, PE, and GDM placentas. It increased DNA fragmentation, CASP3/7 activity, P53, BAX/BCL2, CASP9, CASP 8, CASP3 levels in BeWo, and CASP3 expression in tested placentas. Furthermore, visfatin modulated INSR, ERK1/2, AKT, AMPKα, and STAT3 expression in JEG-3 and BeWo, and its anti-proliferative and pro-apoptotic effects occurred via mentioned factors. In conclusion, visfatin, by affecting the proliferation and apoptosis of human placenta cells, may be an important factor in the development and function of the organ.

Summary Sentence

Visfatin exerts an anti-proliferative and pro-apoptotic effect in the human placenta via insulin receptor, ERK1/2, AKT, AMPKα, and STAT3 signaling pathways.

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Anti-Müllerian hormone (AMH) is widely used in the clinic as a biomarker for ovarian reserve and to predict ovarian response to gonadotropin stimulation. Patients with higher AMH levels tend to yield more oocytes and have better outcomes from assisted reproductive technology procedures. The goal of this study is to determine if AMH can be used to predict the outcome of controlled ovarian stimulation in rhesus macaques, which are commonly used in biomedical research, to refine animal use while maximizing oocyte yield. We hypothesized that pre-stimulation AMH values can be used to predict oocyte yield and quality. Regularly cycling adult macaques underwent controlled ovarian stimulation and baseline (pre-stimulation) plasma AMH levels were determined using an AMH-specific enzyme-linked immunoassay. Oocytes were collected by laparoscopic or ultrasound-guided aspiration, then counted and evaluated for quality and stage of meiosis. Sperm from established fertile males were used to inseminate the oocytes in vitro with fertilization success checked 14–16 h later. Females were grouped by oocyte yield: low ≤17; mid = 18–41; high ≥42. We found that high and mid yielders had significantly higher AMH than low yielders (p < 0.0001) and the percent of mature oocytes was greater in the high and mid yielders. There were no significant differences in oocyte quality or ova fertilization rate. These data suggest that AMH is a useful measure for controlled ovarian stimulation success in rhesus macaques and can be used to identify suitable animals for oocyte donation before entering them into a stimulation protocol.

Summary Sentence

Measurement of serum Anti-Müllerian hormone prior to a controlled ovarian stimulation can be used to predict oocyte yield and the recovered proportion of mature ova in rhesus macaques.

Graphical Abstract

Anti-Mullerian hormone is a useful measure for controlled ovarian stimulation success in rhesus macaques and can be used to refine animal use while maximizing oocyte yield. Created in https://BioRender.com.

Bisphenols are a family of chemicals used in the manufacture of consumer products containing polycarbonate plastics and epoxy resins. Studies have shown that exposure to bisphenol A (BPA) may disrupt steroidogenesis and induce adverse effects on male and female reproduction, but little is known about BPA replacements. We determined the effects of six bisphenols on the steroidogenic function of MA-10 Leydig cells and KGN granulosa cells by measuring the levels of progesterone and estradiol produced by these cells as well as the expression of transcripts involved in steroid and cholesterol biosynthesis. MA-10 and KGN cells were exposed for 48 h to one of six bisphenols (0.01–50 µM): BPA, bisphenol F, bisphenol S, bisphenol AF, bisphenol M, or bisphenol TMC, under both basal and dibutyryl cAMP (Bu₂cAMP)-stimulated conditions. In MA-10 cells, most bisphenols increased the Bu₂cAMP-stimulated production of progesterone. In KGN cells, there was a general decrease in progesterone production, while estradiol levels were increased following exposure to many bisphenols. Quantitative real-time polymerase chain reaction analyses revealed that all six bisphenols (≥1 µM) upregulated the expression of STAR, a cholesterol transporter, in both cell lines after stimulation. Key transcripts directly involved in steroid and cholesterol biosynthesis were significantly altered in a cell line, chemical, and concentration-dependent manner. Thus, BPA and five of its analogs can disrupt steroid production in two steroidogenic cell lines and alter the levels of transcripts involved in this process. Importantly, BPA replacements do not appear to have fewer effects than BPA.

Summary Sentence

Bisphenol A and its structural analogs affect progesterone and estradiol production and differentially alter the expression of genes involved in steroidogenesis and cholesterol biosynthesis in MA-10 mouse Leydig cells and KGN human granulosa cells.

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The embryonic diapause of the giant panda (Ailuropoda melanoleuca) has caused great difficulties in monitoring pregnancy in this vulnerable species. The secretion of prolactin (PRL) from anterior pituitary glandular lactotropic cells is an important signal for the termination of embryonic dormancy. Currently, the mechanism by which PRL affects embryonic diapause in giant pandas and methods for detecting PRL in this species is poorly understood. In this study, the first sandwich enzyme immunoassay for detecting PRL in giant panda urine was established by using two antigiant panda PRL antibodies prepared as coating and labeling antibodies, and PRL recombinant proteins prepared via the prokaryotic system as standards. The established method was used to detect the levels of PRL in the urine of giant pandas during pregnancy. At the same time, the changes in PRL levels in giant pandas and the relationship between PRL and progestagen levels were analyzed during the luteal phase. The results showed that in female giant pandas, PRL levels significantly increased before the progestagen peak, and during the luteal phase, the PRL level was significantly greater in giant pandas that gave birth than in those that did not give birth and those in the nonestrus group. To the best of our knowledge, this is the first study to preliminarily explore the mode of action of PRL in the gestation period of giant pandas and lays a foundation for further study of the regulatory mechanisms of endocrine hormones in the giant panda.

Summary Sentence

Prolactin has an important effect on embryonic diapause, but its role in embryonic diapause in the giant panda has not been studied.

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