The mammalian oviduct is the place where life begins as it is the site of fertilization and preimplantation embryo development. Recent research has highlighted the important role played by the oviduct both in sperm selection for natural fertilization and in the genetic and epigenetic reprogramming of preimplantation embryo development. This review examines oviduct fluid composition with a special emphasis on exosomes and the role played by the oviduct in sperm selection, early embryo development, and in reshaping the epigenetic landscape of the embryo. In addition, the implications of data obtained for improving assisted reproductive technologies are discussed.

Summary Sentence

Herein we review the influence of the oviduct on spermatozoa and embryo with special emphasis on the composition of the oviductal flow, sperm selection and remodelling of embryo epigenetics.

Decidualization is regulated by crosstalk of progesterone and the cAMP pathway. It involves extensive reprogramming of gene expression and includes a wide range of functions. To investigate how cell cycle regulatory genes drive the human endometrial stromal cell (ESC) exit cell cycle and enter differentiation, primary cultured ESC was treated with 8-Br-cAMP and MPA and cell cycle distribution was investigated by flow cytometry. High-throughput cell cycle regulatory gene expression was also studied by microarray. To validate the results of microarray chip, immunohistochemistry and semi-quantitative method of optical density were used to analyze the expression of cell cycle regulator proteins in proliferative phase of endometrium (n = 6) and early pregnancy decidua (n = 6). In addition, we selected cyclin-dependent kinase inhibitor 1c (CDKN1C, also known as P57) and cyclin-dependent kinase inhibitor 2b (CDKN2B, also known as P15) in order to study their role in the process of decidualization by the RNAi method. ESC was arrested at G0/G1 checkpoints during decidualization. Cell cycle regulatory genes P57 and P15 were upregulated, while cyclin D1 (CCND1), cyclin-dependent kinase 2 (CDK2), and cell division cycle protein 2 homolog (CDC2) were downregulated during ESC differentiation both in vitro and vivo. P57 siRNA impaired ESC decidualization and caused differentmorphological and ultrastructural changes as well as a relatively low secretion of prolactin, but P15 siRNA had no effects. We concluded that P15, CCND1, CDK2, and CDC2 may participate in ESC withdraw from the cell cycle and go into differentiation both in vitro and in vivo. P57 is one of the key determinants of ESC differentiation due to its effect on the cell cycle distribution, but its association with the decidua-specific transcription factor needs further investigation.

Summary Sentence

Cell cycle regulatory genes P57 and P15 are upregulated, while Cyclin D1, CDK2, and CDC2 are downregulated during the ESC differentiation and P57 is one of the key regulators that modulates ESC differentiation by controlling cell cycle distribution.

In 2011, DEHP (plasticizer) was reported to illegally be added in food and beverage products in Taiwan, which caused great concerns about food safety worldwide. DHEP has multiple toxic effects to human and animals such as endocrine disruption, cardiotoxicity, reproductive function, and development defects. However, the toxic effects of DEHP on mammalian oocyte quality are still unclear. SinceMEHP is the active metabolite of DEHP in vivo, in this study we used porcine oocyte asmodel to explore the effects of MEHP on oocyte maturation and we also studied the effects of melatonin administration on MEHP exposure-induced meiosis defects. Our results showed that exposure to MEHP significantly decreased the polar body extrusion rate in porcine oocytes. Further study showed that cell cycle progression, meiotic spindle organization, and actin assembly were all disturbed after MEHP exposure. Moreover, the DNA and histone methylation levels were also affected, showing with altered 5mC and H3K4me2 levels. These results indicated thatMEHP affected porcine oocytematuration, while MEHP exposure-induced meiotic defects were all remarkably ameliorated by the administration of melatonin in porcine oocytes. We further tried to explore the causes of MEHP toxicity on oocytes, and we found that MEHP exposure resulted in significant elevations of oxidative stress and induced early apoptosis as well as elevated autophagy, while melatonin administration could reduce these. Taken together, our results indicated that MEHP exposure induced deterioration of oocyte quality, whereas melatonin supplement showed amelioration on oocyte maturation through its rescue effects on oocyte oxidative stress-mediated apoptosis and autophagy.

Summary Sentence

Melatonin supplement rescued cell cycle and cytoskeleton defects through ameliorating oxidative stress-mediated apoptosis/autophagy in MEHP-exposed porcine oocytes.

Preterm birth is widespread and causes 35% of all neonatal deaths. Infants who survive face potential long-term complications. A major contributing factor of preterm birth is infection. We investigated the role of interleukin 22 (IL22) as a potential clinically relevant cytokine during gestational infection. IL22 is an effector molecule secreted by immune cells. While the expression of IL22 was reported in normal nonpregnant endometrium and early pregnancy decidua, little is known about uterine IL22 expression during mid or late gestational stages of pregnancy. Since IL22 has been shown to be an essential mediator in epithelial regeneration and wound repair, we investigated the potential role of IL22 during defense against an inflammatory response at the maternal–fetal interface. We used a well-established model to study infection and infection-associated inflammation during preterm birth in the mouse. We have shown that IL22 is upregulated to respond to an intrauterine lipopolysaccharide administration and plays an important role in controlling the risk of inflammation-induced preterm birth. This paper proposes IL22 as a treatment method to combat infection and prevent preterm birth in susceptible patients.

Summary Sentence

IL22 is upregulated in uterine tissue in response to bacterial endotoxin and contributes to defense against inflammatory reaction at the maternal-fetal interface

Gestational choriocarcinomas are derived from placental trophoblast cells, with HLA-C being the only class I polymorphic molecule expressed. However, choriocarcinomas have not been profiled for endoplasmic reticulum aminopeptidase 2 (ERAP2) expression. ERAP2 trims peptides presented by human leukocyte antigens (HLA) that have shown to modulate immune response. Over 50% of choriocarcinomas we screened lack ERAP2 expression, which suggests that the absence of ERAP2 expression allows immune evasion of choriocarcinoma cells. We demonstrate that the ability of choriocarcinoma cells to activate lymphocytes was lowest with cells lacking ERAP2 (JEG-3) or HLA-C (JAr). This observation suggests that activation is dependent on expression of both ERAP2 and HLA-C molecules. In addition, an ERAP2 variant in which lysine is changed to asparagine (K392N) results in increased trimming activity (165-fold) for hydrophobic peptides and biologically never been detected. We hypothesize that homozygosity for the N392 ERAP2 variant is prohibited because it modulates the immune recognition of placental trophoblasts. We demonstrate that NK-cell activation and killing were significantly dependent on forced expression of the N392 ERAP2 isoform in JEG-3 cells. Cytotoxicity was confirmed by 7AAD killing assays showing that N392 ERAP2-isoform expressing JEG-3 cells had the highest percentage of apoptotic cells independent of the expression level of CD11a on lymphocytes. This is the first report showing that N392 ERAP2 promotes an immune clearance pathway for choriocarcinoma cells, and provides an explanation for why embryonic homozygosity for the N392 ERAP2 variant is not detected in any population.

Summary Sentence

The N392 ERAP2 isoform expression promotes immune cell activation and apoptotic immune clearance of choriocarcinoma cells.

A major gene for bovine ovulation rate has been mapped to a 1.2 Mb region of chromosome 10. Screening of coding regions of positional candidate genes within this region failed to reveal a causative polymorphism, leading to the hypothesis that the phenotype results from differences in candidate gene expression rather than alteration of gene structure. This study tested differences in expression of positional candidate genes in granulosa cells between carriers and noncarriers of the high fecundity allele, as well as characterizing differences in the transcriptomic profile between genotypes. Five carriers and five noncarriers, female descendants of “Trio,” a carrier of the high fecundity allele were initially used in an RNA-seq analysis of gene expression. Four of ten samples were contaminated with theca cells, so that six samples were used in the final analysis (three of each genotype). Of 14 973 genes expressed, 143 were differentially expressed (false discovery rate P < 0.05) in carriers versus noncarriers. Among the positional candidate genes, SMAD6 was 6.6-fold overexpressed in the carriers compared to noncarriers (P < 5 × 10^-5). This result was replicated in an independent group of 12 females (7 carriers and 5 noncarriers) using quantitative real-time PCR; SMAD6 was 9.3-fold overexpressed in carriers versus noncarriers (P = 1.17 × 10^-6). Association of overexpression of SMAD6, an inhibitor of the BMP/SMAD signaling pathway, with high ovulation rate corresponds well with disabling mutations in ligands (BMP15 and GDF9) and a receptor (BMPR1B) of this pathway that cause increased ovulation rate in sheep.

Summary Sentence

Carriers of an allele for high ovulation rate exhibit a six-fold overexpression of the positional candidate gene SMAD6 compared to noncarriers, strongly implicating SMAD6 as the gene responsible for the high ovulation rate phenotype.

The newly discovered Trio high-fecundity allele produces multiple ovulations in cattle. This study evaluated (1) size and growth rates of follicles in Trio carriers during a synchronized follicular wave, induced by follicle aspiration; (2) follicle-stimulating hormone (FSH) patterns associated with the follicular wave; (3) size of corpora lutea (CL) and circulating progesterone; and (4) intrafollicular estradiol concentrations prior to normal deviation. Trio carriers had mean dominant follicles that were significantly smaller in diameter and volume than noncarriers. Onset of diameter deviation occurred at ∼3 days after the last follicle aspiration in both genotypes despite Trio carriers having much smaller individual follicles. Follicles of Trio carriers grew at a slower rate than noncarrier follicles (∼65% in mm/day or ∼30% in mm³/day) resulting in much smaller individual dominant follicles (∼25% volume). However, total dominant follicle volume, calculated as the sum of all dominant follicles in each animal, was similar in carriers and noncarriers of Trio throughout the entire follicular wave. Circulating FSH was greater in Trio carriers during the 24 h encompassing deviation. Trio carriers had significantly more ovulations than noncarriers, and individual CL volume was smaller, although total luteal tissue volume and circulating P4 were not different. Thus, increased ovulation rate in Trio carriers relates to smaller individual follicles (one-third the volume) near the time of deviation due to slower follicle growth rate, although time of deviation is similar, with increased circulating FSH near deviation leading to selection of multiple dominant follicles in Trio carriers with similar total follicle volume.

Summary Sentence

Results reveal that high ovulation rate in carriers of the Trio allele is related to smaller individual follicles due to slower growth rate but similar total follicle volume and time of follicle deviation accompanied by greater FSH near deviation.

The acquisition of dominance and ovulatory capacity was evaluated in follicles from cows that were carriers or half-sibling noncarriers of the Trio allele. Follicle size at acquisition of follicular dominance was determined by evaluating whether follicles ovulate after GnRH challenge (ovulatory capacity—experiment 1) and by determination of intrafollicular concentrations of estradiol and free insulin like growth factor 1 (IGF1) and relative mRNA expression of cytochrome P450 family 19 subfamily A member 1 (CYP19A1), luteinizing hormone/choriogonadotropin receptor (LHCGR), and pappalysin 1 (PAPPA, previously known as pregnancy-associated plasma protein A, pappalysin 1) in granulosa cells from follicles of different sizes (experiment 2). Ovulatory capacity developed in follicles at 8.3 mm (50% ovulatory capacity) in noncarriers but at smaller sizes (5.5 mm) in Trio carriers. Similarly, in experiment 2, follicles of Trio carriers acquired a dominant phenotype, as determined by intrafollicular estradiol and CYP19A1, LHCGR, and PAPPA mRNA expression in granulosa cells, at significantly smaller sizes but at a similar time after wave emergence. Overall, dominance/ovulatory capacity was acquired when follicles of Trio carriers were ∼30% the size (volume basis) of follicles in noncarriers. In addition, follicles in Trio carriers appear to acquire dominance in a hierarchal manner, as demonstrated by the progressively greater number of follicles with a dominant phenotype between days 2 and 4 after wave emergence. Thus, results from this study provide further support for a physiological model in which selection of multiple follicles in Trio allele carriers is characterized by acquisition of dominance at a smaller follicle size but at a similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence.

Summary Sentence

Results support a model for selection of multiple follicles in Trio allele carriers due to acquisition of dominance at a smaller follicle size but similar time in the follicular wave with multiple follicles acquiring dominance in a hierarchal sequence.

Thrombospondin-1 (THBS1) affects corpus luteum (CL) regression. Highly induced during luteolysis, it acts as a natural anti-angiogenic, proapoptotic compound. THBS1 expression is regulated in bovine luteal endothelial cells (LECs) by fibroblast growth factor-2 (FGF2) and transforming growth factor-beta1 (TGFB1) acting in an opposite manner. Here we sought to identify specific microRNAs (miRNAs) targeting THBS1 and investigate their possible involvement in FGF2 and TGFB1-mediated THBS1 expression. Several miRNAs predicted to target THBS1 mRNA (miR-1, miR-18a, miR-144, miR-194, and miR-221) were experimentally tested. Of these, miR-221 was shown to efficiently target THBS1 expression and function in LECs. We found that this miRNA is highly expressed in luteal cells and in mid-cycle CL. Consistent with the inhibition of THBS1 function, miR-221 also reduced Serpin Family E Member 1 [SERPINE1] in LECs and promoted angiogenic characteristics of LECs. Plasminogen activator inhibitor-1 (PAI-1), the gene product of SERPINE1, inhibited cell adhesion, suggesting that PAI-1, like THBS1, has anti-angiogenic properties. Importantly, FGF2, which negatively regulates THBS1, elevates miR-221. Conversely, TGFB1 that stimulates THBS1, significantly reduces miR-221. Furthermore, FGF2 enhances the suppression of THBS1 caused by miR-221 mimic, and prevents the increase in THBS1 induced by miR-221 inhibitor. In contrast, TGFB1 reverses the inhibitory effect of miR-221 mimic on THBS1, and enhances the upregulation of THBS1 induced by miR-221 inhibitor. These data support the contention that FGF2 and TGFB1 modulate THBS1 via miR-221. These in vitro data propose that dynamic regulation of miR-221 throughout the cycle, affecting THBS1 and SERPINE1, can modulate vascular function in the CL.

Summary Sentence

Dynamic regulation ofmiR-221 throughout the cycle, affecting THBS1 and subsequently SERPINE1, can modulate vascular function in the corpus luteum.

The role of progesterone (P4) in the regulation of the local (uterine) and systemic innate immune system, myometrial expression of connexin 43 (Cx-43) and cyclooxygenase 2 (COX-2), and the onset of parturition was examined in (i) naïve mice delivering at term; (ii) E16mice treated with RU486 (P4-antagonist) to induce preterm parturition; and (iii) in mice treated with P4 to prevent term parturition. In naïve mice, myometrial neutrophil and monocyte numbers peaked at E18 and declined with the onset of parturition. In contrast, circulating monocytes did not change and although neutrophils were increased with pregnancy, they did not change across gestation. The myometrial mRNA and protein levels of most chemokines/cytokines, Cx-43, and COX-2 increased with, but not before, parturition. With RU486-induced parturition, myometrial and systemic neutrophil numbers increased before and myometrial monocyte numbers increased with parturition only. Myometrial chemokine/cytokine mRNA abundance increased with parturition, but protein levels peaked earlier at between 4.5 and 9 h post-RU486. Cx-43, but not COX-2, mRNA expression and protein levels increased prior to the onset of parturition. In mice treated with P4, the gestation-linked increase in myometrial monocyte, but not neutrophil, numbers was prevented, and expression of Cx-43 and COX-2 was reduced. On E20 of P4 supplementation,myometrial chemokine/cytokine and leukocyte numbers, but not Cx-43 and COX-2 expression, increased. These data show that during pregnancy P4 controls myometrial monocyte infiltration, cytokine and prolabor factor synthesis via mRNAdependent and independent mechanisms and, with prolonged P4 supplementation, P4 action is repressed resulting in increased myometrial inflammation.

Summary Sentence

Progesterone independently regulates the maternal immune system and the onset of parturition in the mouse.

Streptococcus agalactiae (group B streptococcus [GBS]) infection in pregnant women is the leading cause of infectious neonatal morbidity and mortality in the United States. Although inflammation during infection has been associated with preterm birth, the contribution of GBS to preterm birth is less certain. Moreover, the early mechanisms by which GBS interacts with the gestational tissue to affect adverse pregnancy outcomes are poorly understood. We hypothesized that short-term GBS inoculation activates pathways related to inflammation and premature birth in human extraplacental membranes. We tested this hypothesis using GBS-inoculated human extraplacental membranes in vitro. In agreement with our hypothesis, a microarray-based transcriptomics analysis of gene expression changes in GBS-inoculated membranes revealed that GBS activated pathways related to inflammation and preterm birth with significant gene expression changes occurring as early as 4 h postinoculation. In addition, pathways related to DNA replication and repair were downregulated with GBS treatment. Conclusions based on our transcriptomics data were further supported by responses of prostaglandin E2 (PGE2), and matrix metalloproteinases 1 (MMP1) and 3 (MMP3), all of which are known to be involved in parturition and premature rupture of membranes. These results support our initial hypothesis and provide new information on molecular targets of GBS infection in human extraplacental membranes.

Summary Sentence

Group B Streptococcus (GBS) activates premature birth and inflammation pathways in human extraplacental membranes in vitro, providing insight into early molecular responses underpinning GBS contributions to adverse birth outcomes.

With half a million babies born preterm each year in the USA and about 15 million worldwide, preterm birth (PTB) remains a global health issue. Preterm birth is a primary cause of infant morbidity and mortality and can impact lives long past infancy. The fact that there are numerous, and many currently unidentified, etiologies of PTB has hindered development of tools for risk evaluation and preventative therapies. Infection is estimated to be involved in nearly 40% of PTBs of known etiology; therefore, understanding how infection-mediated inflammation alters the cervical milieu and leads to preterm tissue biomechanical changes are questions of interest. Using RNA-seq, we identified enrichment of components involved in inflammasome activation and unique proteases in the mouse cervix during lipopolysaccharide (LPS)-mediated PTB and not physiologically at term before labor. Despite transcriptional induction of inflammasome components, there was no evidence of functional activation based on assessment of mature IL1B and IL18 proteins. The increased transcription of proteases that target both elastic fibers and collagen and concentration of myeloid-derived cells capable of protease synthesis in the cervical stroma support the structural disruption of elastic fibers as a functional output of protease activity. The recent demonstration that elastic fibers contribute to the biomechanical function of the pregnant cervix suggests their protease-induced disruption in the infection model of LPS-mediated PTB and may contribute to premature loss of mechanical competency and preterm delivery. Collectively, the transcriptomics and ultrastructural data provide new insights into the distinct mechanisms of premature cervical remodeling in response to infection.

Summary Sentence

A distinct transcriptome signature and pattern of ECM ultrastructure distinguish inflammationmediated premature cervical ripening from term ripening.

Preterm deliveries remain the leading cause of neonatal morbidity and mortality. Current therapies target only myometrial contractions and are largely ineffective. As labor involves multiple coordinated events across maternal and fetal tissues, identifying fundamental regulatory pathways of normal term labor is vital to understanding successful parturition and consequently labor pathologies. We aimed to identify transcriptomic signatures of human normal term labor of two tissues: in the fetal-facing choriodecidua and the maternalmyometrium. Microarray transcriptomic data from choriodecidua and myometrium following term labor were analyzed for functional hierarchical networks, using Cytoscape 2.8.3. Hierarchically high candidates were analyzed for their regulatory casual relationships using Ingenuity Pathway Analysis. Selectedmaster regulators were then chemically inhibited and effects on downstream targets were assessed using real-time quantitative PCR (RT-qPCR). Unbiased network analysis identified upstream molecular components in choriodecidua including vimentin, TLR4, and TNFSF13B. In the myometrium, candidates included metallothionein 2 (MT2A), TLR2, and RELB. These master regulators had significant differential gene expression during labor, hierarchically high centrality in community cluster networks, interactions amongst the labor gene set, and strong causal relationships with multiple downstream effects. In vitro experiments highlighted MT2A as an effective regulator of labor-associated genes. We have identified unique potential regulators of the term labor transcriptome in uterine tissues using a robust sequence of unbiased mathematical and literature-based in silico analyses. These findings encourage further investigation into the efficacy of predicted master regulators in blocking multiple pathways of labor processes across maternal and fetal tissues, and their potential as therapeutic approaches.

Summary Sentence

Combined in silico and in vitro analyses have identified novel transcriptomic regulators of term labor in human choriodecidua and myometrium with further therapeutic potential.