Effect of Aromatase Inhibitor Letrozole on the Placenta of Adult Albino Rats: A Histopathological, Immunohistochemical, and Biochemical Study.

Background: Letrozole, an aromatase inhibitor, has recently been introduced as the preferred treatment option for ectopic pregnancy. To date, no study has investigated the effect of letrozole alone on placental tissue. The present study aimed to evaluate the effect of different doses of letrozole on the placenta of rats and to clarify the underlying mechanism. Methods: Sixty pregnant female rats were equally divided into three groups, namely the control group (GI), low-dose (0.5 mg/Kg/day) letrozole group (GII), which is equivalent to the human daily dose (HED) of 5 mg, and high-dose (1 mg/Kg/day) letrozole group (GIII), equivalent to the HED of 10 mg. Letrozole was administered by oral gavage daily from day 6 to 16 of gestation. Data were analyzed using a one-way analysis of variance followed by Tukey's post hoc test and Chi square test. P<0.05 was considered statistically significant. Results: Compared to the GI and GII groups, high-dose letrozole significantly increased embryonic mortality with a high post-implantation loss rate (P<0.001) and significantly reduced the number of viable fetuses (P<0.001) and placental weight (P<0.001) of pregnant rats. Moreover, it significantly reduced placental estrogen receptor (ER) and progesterone receptor (PR) (P<0.001) and the expression of vascular endothelial growth factor (P<0.001), while increasing the apoptotic index of cleaved caspase-3 (P<0.001). Conclusion: Letrozole inhibited the expression of ER and PR in rat placenta. It interrupted stimulatory vascular signals causing significant apoptosis and placental vascular dysfunction. Letrozole in an equivalent human daily dose of 10 mg caused a high post-implantation loss rate without imposing severe side effects.

Gestational exposure to 1-NP induces ferroptosis in placental trophoblasts via CYP1B1/ERK signaling pathway leading to fetal growth restriction.

Fetal growth restriction (FGR) is a prevalent complication in obstetrics, yet its exact aetiology remains unknown. Numerous studies suggest that the degradation of the living environment is a significant risk factor for FGR. 1-Nitropyrene (1-NP) is a widespread environmental pollutant as a representative substance of nitro-polycyclic aromatic hydrocarbons. In this study, we revealed that 1-NP induced FGR in fetal mice by constructing 1-NP exposed pregnant mice models. Intriguingly, we found that placental trophoblasts of 1-NP exposed mice exhibited significant ferroptosis, which was similarly detected in placental trophoblasts from human FGR patients. In this regard, we established a 1-NP exposed cell model in vitro using two human trophoblast cell lines, HTR8/SVneo and JEG-3. We found that 1-NP not only impaired the proliferation, migration, invasion and angiogenesis of trophoblasts, but also induced severe cellular ferroptosis. Meanwhile, the ferroptosis inhibitor ferrostatin-1 (Fer-1) effectively rescued 1-NP-induced trophoblast biological function impairment. Mechanistically, we revealed that 1-NP regulated ferroptosis by activating the ERK signaling pathway. Moreover, we innovatively revealed that CYP1B1 was essential for the activation of ERK signaling pathway induced by 1-NP. Overall, our study innovatively identified ferroptosis as a significant contributor to 1-NP induced trophoblastic functional impairment leading to FGR and clarified the specific mechanism by which 1-NP induced ferroptosis via the CYP1B1/ERK signaling pathway. Our study provided novel insights into the aetiology of FGR and revealed new mechanisms of reproductive toxicity of environmental pollutants.

Cannabis Exposure and Adverse Pregnancy Outcomes Related to Placental Function.

Importance: Cannabis use is increasing among reproductive-age individuals and the risks associated with cannabis exposure during pregnancy remain uncertain. Objective: To evaluate the association between maternal cannabis use and adverse pregnancy outcomes known to be related to placental function. Design, Setting, and Participants: Ancillary analysis of nulliparous individuals treated at 8 US medical centers with stored urine samples and abstracted pregnancy outcome data available. Participants in the Nulliparous Pregnancy Outcomes Study: Monitoring Mothers-to-Be cohort were recruited from 2010 through 2013; the drug assays and analyses for this ancillary project were completed from June 2020 through April 2023. Exposure: Cannabis exposure was ascertained by urine immunoassay for 11-nor-9-carboxy-Δ9-tetrahydrocannabinol using frozen stored urine samples from study visits during the pregnancy gestational age windows of 6 weeks and 0 days to 13 weeks and 6 days (visit 1); 16 weeks and 0 days to 21 weeks and 6 days (visit 2); and 22 weeks and 0 days to 29 weeks and 6 days (visit 3). Positive results were confirmed with liquid chromatography tandem mass spectrometry. The timing of cannabis exposure was defined as only during the first trimester or ongoing exposure beyond the first trimester. Main Outcome and Measure: The dichotomous primary composite outcome included small-for-gestational-age birth, medically indicated preterm birth, stillbirth, or hypertensive disorders of pregnancy ascertained by medical record abstraction by trained perinatal research staff with adjudication of outcomes by site investigators. Results: Of 10 038 participants, 9257 were eligible for this analysis. Of the 610 participants (6.6%) with cannabis use, 32.4% (n = 197) had cannabis exposure only during the first trimester and 67.6% (n = 413) had ongoing exposure beyond the first trimester. Cannabis exposure was associated with the primary composite outcome (25.9% in the cannabis exposure group vs 17.4% in the no exposure group; adjusted relative risk, 1.27 [95% CI, 1.07-1.49]) in the propensity score-weighted analyses after adjustment for sociodemographic characteristics, body mass index, medical comorbidities, and active nicotine use ascertained via urine cotinine assays. In a 3-category cannabis exposure model (no exposure, exposure only during the first trimester, or ongoing exposure), cannabis use during the first trimester only was not associated with the primary composite outcome; however, ongoing cannabis use was associated with the primary composite outcome (adjusted relative risk, 1.32 [95% CI, 1.09-1.60]). Conclusions and Relevance: In this multicenter cohort, maternal cannabis use ascertained by biological sampling was associated with adverse pregnancy outcomes related to placental dysfunction.

Oxycodone does not affect placental circulatory physiology during the early first stage of labor-A randomized trial.

INTRODUCTION: Opioids are used for pain relief during the first stage of labor. Oxycodone can cause maternal hypotension that may modify utero- and fetoplacental circulatory physiology. We hypothesized that maternal intravenous (i.v.) oxycodone has no detrimental effect on utero- and fetoplacental hemodynamics during the early first stage of labor. MATERIAL AND METHODS: Twenty-two parturients requiring pain relief during the first stage of labor were randomized in a double-blinded and placebo-controlled study. By Doppler ultrasonography, both uterine artery (Ut) and umbilical vein (UV) volume blood flows (Q), Ut pulsatility index (PI), and Ut vascular resistance (RUt) were calculated. Blood flow velocity waveforms were obtained between uterine contractions. After baseline measurements, women received oxycodone 0.05 mg/kg or a placebo intravenous. Doppler ultrasonography was repeated up to 120 min after the first drug administration. The second dose of oxycodone 0.05 mg/kg was allowed at 60 min to all parturients with contraction pain ≥5/10. Maternal plasma samples were collected at each study phase and after delivery with umbilical cord plasma samples, to measure oxycodone concentrations. CLINICALTRIALS: gov identifier (NCT no. NCT02573831). RESULTS: At baseline, mean QUt and QUV did not differ significantly between the placebo-first (478 mL/min and 57 mL/min/kg) and the oxycodone-first (561 mL/min and 71 mL/min/kg) groups. In addition, RUt and Ut PI were comparable between the groups. Following oxycodone at 60 min, mean QUt and QUV (714 mL/min and 52 mL/min/kg) were similar to the placebo-first (520 mL/min and 55 mL/min/kg) group. Furthermore, all the measured parameters were comparable to the baseline values. At 60 min after the first study drug administration, all the parturients in the placebo-first group needed intravenous oxycodone 0.05 mg/kg. At 120 min, we found no statistically significant change in any of the measured parameters. No significant correlation was found between maternal oxycodone concentration and QUt or QUV. Furthermore, newborn oxycodone concentration did not correlate with QUV. CONCLUSIONS: Oxycodone did not have any detrimental effect on either utero- or fetoplacental circulatory physiology during the early first stage of labor. Maternal plasma oxycodone did not correlate with utero- and fetoplacental hemodynamics. No correlation was found between newborn oxycodone concentration and fetoplacental hemodynamics.

Toxicological effects of the Morinda citrifolia L. fruit extract on maternal reproduction and fetal development in rats.

Morinda citrifolia L., also known as Noni, is widely used plant in folk medicine for various therapeutic purposes. However, reports on its effects during pregnancy are limited. Therefore, the objective of this study was to evaluate the effects of the M. citrifolia fruit extract on maternal performance and fetal development during pregnancy in rats. Pregnant Wistar rats (n = 12/group) were treated from gestational days (GD) 0-21 with water (control group) or the aqueous extract of M. citrifolia fruit at doses of 200, 400, or 750 mg/kg, orally. During pregnancy, clinical signs of toxicity, maternal weight, feed intake, and water consumption were noted. On GD 21, the rats were anesthetized and blood was collected to evaluate various biochemical parameters. During laparotomy, reproductive performance parameters were recorded, and fetuses were weighed and the anomalies analyzed. Reduced placental efficiency and fetal growth restriction were observed in the group treated with 400 mg/kg of M. citrifolia extract. The highest dose (750 mg/kg) augmented aspartate aminotransferase concentration and preimplantation losses, while reducing the number of live fetuses. Furthermore, both doses (400 and 750 mg/kg) of the plant extract caused fetal anomalies. In conclusion, consumption of high doses of the M. citrifolia aqueous extrac during pregnancy leads to maternal hepatotoxicity, anti-implantation effects, intrauterine growth restriction and fetal abnormalities, indicating that the plant fruit extract can be harmful to both the mother and the fetus.

Role of progesterone on dexamethasone-induced alterations in placental vascularization and progesterone receptors in rats†.

BACKGROUND: Intrauterine growth restriction (IUGR) is manifested by lower maternal progesterone levels, smaller placental size, and decreased placental vascularity indicated by lower expression of vascular endothelial growth factor (VEGF). Studies showed that progesterone increases angiogenesis and induces VEGF expression in different tissues. Therefore, the aim of the present study is to evaluate the effect of progesterone on placental vascular bed and VEGF expression and the modulation of nuclear and membranous progesterone receptors (PR) in dexamethasone-induced rat IUGR model. METHODS: Pregnant Sprague-Dawley rats were allocated into four groups and given intraperitoneal injections of either saline, dexamethasone, dexamethasone, and progesterone or progesterone. Injections started on gestation day (DG) 15 and lasted until the days of euthanization (19 and 21 DG). Enzyme-linked immunosorbent assay was used to evaluate plasma progesterone levels. Real-time PCR and western blotting were used to evaluate gene and protein expressions of VEGF, and PR in labyrinth and basal placental zones. Immunohistochemistry was used to locate VEGF and different PRs in placental cells. Immunofluorescence was used to monitor the expression of blood vessel marker (αSMA). RESULTS: Dexamethasone decreased the vascular bed fraction and the expression of VEGF in both placental zones. Progesterone co-treatment with dexamethasone prevented this reduction. Nuclear and membrane PRs showed tissue-specific expression in different placental zones and responded differently to both dexamethasone and progesterone. CONCLUSIONS: Progesterone treatment improves the outcomes in IUGR pregnancy. Progesterone alleviated DEX-induced IUGR probably by promoting placental VEGF and angiogenesis.

The impact of pre-pregnancy folic acid intake on placental DNA methylation in a fortified cohort.

Folate plays an important role in the modulation of one-carbon metabolism and DNA methylation through a complex biosynthesis pathway. Folate deficiency during pregnancy has been associated with an increased risk for birth defects. This study investigates the extent to which the availability of folate and S-Adenosylmethionine (SAM) affects placental DNA methylation. We hypothesized that maintaining sufficient levels of folate and SAM is particularly important in individuals carrying the MTHFR C677T polymorphism. Maternal- and cord blood was analyzed to genotype the MTHFR rs1801133 SNP. Red blood cell (RBC) folate, vitamin B12, SAM, and S-Adenosylhomocysteine (SAH) were analyzed in cord blood. Epigenome-wide methylation analyses were performed on 90 placenta tissue samples isolated from the fetal side of the placenta; 45 originating from mother-infant dyads homozygous for the MTHFR C677T variant and 45 originating from mother-infant dyads with the homozygous wild type MTHFR677 genotype. Verification of the results was performed using pyrosequencing assays. Genome-wide placental DNA methylation patterns were relatively stable and not significantly affected by levels of one-carbon metabolites. MTHFR genotype was associated with DNA methylation of several loci, including a locus in the MTHFR region. RBC folate and particularly the SAM:SAH ratio did affect overall CpG DNA methylation in some CpG regions when the loci were split according to their CpG island relation. This was most evident in participants carrying the MTHFR C677T variant suggesting a stronger influence of the biosynthesis pathway on the overall placental DNA methylation in MTHFR TT individuals than in MTHFR CC individuals.

Alcohol induced increases in sperm Histone H3 lysine 4 trimethylation correlate with increased placental CTCF occupancy and altered developmental programming.

Using a mouse model, studies by our group reveal that paternal preconception alcohol intake affects offspring fetal-placental growth, with long-lasting consequences on adult metabolism. Here, we tested the hypothesis that chronic preconception male alcohol exposure impacts histone enrichment in sperm and that these changes are associated with altered developmental programming in the placenta. Using chromatin immunoprecipitation, we find alcohol-induced increases in sperm histone H3 lysine 4 trimethylation (H3K4me3) that map to promoters and presumptive enhancer regions enriched in genes driving neurogenesis and craniofacial development. Given the colocalization of H3K4me3 with the chromatin binding factor CTCF across both sperm and embryos, we next examined CTCF localization in the placenta. We find global changes in CTCF binding within placentae derived from the male offspring of alcohol-exposed sires. Furthermore, altered CTCF localization correlates with dysregulated gene expression across multiple gene clusters; however, these transcriptional changes only occur in male offspring. Finally, we identified a correlation between genomic regions exhibiting alcohol-induced increases in sperm H3K4me3 and increased CTCF binding in male placentae. Collectively, our analysis demonstrates that the chromatin landscape of sperm is sensitive to chronic alcohol exposure and that a subset of these affected regions exhibits increased placental CTCF enrichment.

Oxidative Stress and DNA Damage Effect of Dioscorea hispida Dennst. on Placental Tissues of Rats.

Dioscorea hispida Dennst. locally known as "ubi gadung" has been used as a traditional remedy and source of carbohydrate among Malaysians. To assess the effect of Dioscorea hispida aqueous extract (DHAE) on the production of reactive oxygen species (ROS) and their effects on DNA damage in Sprague Dawley rat's placental tissues, pregnant rats were randomly divided into four groups. The animals were orally treated with distilled water (negative control) and three different concentrations of DHAE (250, 500 and 1000 mg/kg body weight (BW)) from gestation day 6 until 20. The oxidative stress in placental tissues was evaluated at day 21 by measuring the level of ROS, superoxide dismutase (SOD) and lipid peroxidation biomarker, malondialdehyde (MDA) while comet assay was used for DNA damage. There was no significant production of ROS and SOD activities in all groups. Significant changes were observed in the MDA level at 1000 mg/kg BW DHAE. Comet assay revealed a significant increase (p < 0.05) of DNA damage on animals treated with 250 and 500 mg/kg BW DHAE but not at the highest concentration. It was postulated that the placental cells could have undergone necrosis which destroys all components including DNA. This occurrence simultaneously reduces the levels of DNA damage which can be represented by lower level of tail moments. This finding correlates with our histopathological examination where necrotic cells of spongiotrophoblast were observed in the basal zone of placental tissue. The high amount of hydrogen cyanide and other compounds in 1000 mg/kg BW DHAE could elevate the lipid peroxidation and directly induce cell necrosis which requires further investigation.

Leucine supplementation during late gestation globally alters placental metabolism and nutrient transport via modulation of the PI3K/AKT/mTOR signaling pathway in sows.

In a previously published study we reported that sow dietary leucine supplementation during late pregnancy significantly improved newborn piglet birth weight by stimulating protein synthesis in the longissimus dorsi muscle. However, there is still limited knowledge as to whether leucine can exert its effects on the placenta, one of the most important temporal organs during pregnancy, to promote maternal-fetal nutrient supply and thus contribute to fetal intrauterine development. Therefore, we tested this hypothesis in the present study. In total, 150 sows at day 90 of gestation were divided into three groups and fed with either a control diet (CON), CON + 0.4% Leu or CON + 0.8% Leu, respectively, until parturition. Placental metabolomics, full spectrum amino acids and nutrient transporters were systematically analyzed after sample collection. The results indicated that Leu supplementation led to an altered placental metabolism with an increased number of metabolites related to glycolysis and the oxidation of fatty acids, as well as elevated levels of amino acid accumulation in the placenta. In addition, nutrient transporters of amino acids, glucose and fatty acids in the placenta were globally up-regulated and several enzymes related to energy metabolism, including hexokinase, succinate dehydrogenase, lactated hydrogenase, glycogen phosphorylase and hydroxyacyl-CoA-dehydrogenase, were also significantly increased with no change observed in the antioxidative status of those groups with Leu supplementation. Furthermore, the phosphorylation of PI3K, Akt, and mTOR was enhanced in the placenta of sows undergoing Leu treatment. Collectively, we concluded that supplementing the diets of sows with Leu during late gestation globally altered placental metabolism and promoted maternal-fetus nutrient transport (amino acids, glucose, and fatty acids) via modulation of the PI3K/Akt/mTOR signaling pathway.

Role of JNK, ERK, and p38 MAPK signaling pathway in protective effect of sildenafil in cyclophosphamide-induced placental injury in rats.

AIMS: Chemotherapeutic agents; cyclophosphamide (CYC) is used for treatment of cancer and autoimmune diseases. Grievously, CYC is non-selective as it affects both tumor and healthy cells resulting in systemic toxicity including placenta. The present study aimed to evaluate the effect of phosphodiesterase 5 inhibitor, sildenafil (Sild) on CYC-induced placental injury in rats. MATERIALS AND METHODS: Thirty-two female Wister rats were randomly divided into 4 experimental groups. Group 1: control pregnant group; Group 2: Sild-treated pregnant rats; Group 3: pregnant rats received CYC; Group 4: pregnant rats received Sild and CYC. Placental malondialdehyde (MDA), total nitrite/nitrate (NOx), reduced glutathione (GSH), tumor necrosis factor-α (TNF-α), platelet growth factor (PlGF), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinase (p38MAPK), extracellular signal-regulated kinase (ERK) and cleaved caspase-3 were measured. Histological changes, Nuclear Factor kappa-light-chain-enhancer of activated B (NF-κB), Connexin 43 (GJA1) and proliferating cell nuclear antigen (PCNA) immuno-expressions were also evaluated. KEY FINDINGS: CYC showed significant decrease in placental GSH, NOx, PlGF, GJA1 and PCNA immuno-expressions but significant increase in placental MDA, TNF-α, JNK, P38MAPK, ERK, caspase-3 and NF-kB immuno-expression. Sild showed significant improvement in all oxidative, inflammatory and apoptotic parameters. SIGNIFICANCE: Sild is a promising protective drug against placental injury induced by CYC through antagonizing MAPK (JNK, ERK, and p38) signaling pathway with anti-oxidant, anti-inflammatory and anti-apoptotic effects.

Extended Prophylactic Effect of N-tert-Butyl-α-phenylnitron against Oxidative/Nitrosative Damage Caused by the DNA-Hypomethylating Drug 5-Azacytidine in the Rat Placenta.

Antioxidant N-tert-Butyl-α-phenylnitron (PBN) partly protected embryos from the negative effects of a DNA demethylating drug 5-azacytidine during pregnancy. Our aim was to investigate PBN's impact on the placenta. Fischer rat dams were treated on gestation days (GD) 12 and 13 by PBN (40 mg/kg), followed by 5azaC (5 mg/kg) after one hour. Global methylation was assessed by pyrosequencing. Numerical density was calculated from immunohistochemical expression in single cells for proliferating (PCNA), oxidative (oxoguanosine) and nitrosative (nitrotyrosine) activity. Results were compared with the PBN-treated and control rats. PBN-pretreatment significantly increased placental weight at GD15 and GD20, diminished by 5azaC, and diminished apoptosis in GD 20 placentas caused by 5azaC. Oxoguanosine expression in placentas of 5azaC-treated dams was especially high in the placental labyrinth on GD 15, while PBN-pretreatment lowered its expression on GD 15 and GD 20 in both the labyrinth and basal layer. 5azaC enhanced nitrotyrosine level in the labyrinth of both gestational stages, while PBN-pretreatment lowered it. We conclude that PBN exerted its prophylactic activity against DNA hypomethylating agent 5azaC in the placenta through free radical scavenging, especially in the labyrinthine part of the placenta until the last day of pregnancy.

Effects of an ω3 fatty acid-biased diet on luteolysis, parturition, and uterine prostanoid synthesis in pregnant mice.

The ω3 polyunsaturated fatty acids (PUFAs) are known to have beneficial effects on health and diseases, and hence their intake is encouraged. However, it remains unknown as to how ω3 PUFAs affect female reproduction processes, in which ω6 PUFA-derived prostaglandin (PG) E2 and PGF2α play crucial roles. We therefore compared female reproductive performance between ω3 PUFA-biased linseed oil diet-fed (Lin) mice and ω6 PUFA-biased soybean oil diet-fed (Soy) mice. In Lin mice, the uterine levels of arachidonic acid (AA) and eicosapentaenoic acid (EPA) were 0.42 fold and 16 fold of those in Soy mice, respectively, with the EPA/AA ratio being 0.7 (vs 0.02 in Soy mice). Lin mice showed no alterations in any of the fertility indexes, including luteolysis and parturition. The uterine PG synthesis profiles of Lin mice were similar to those of Soy mice, but the levels of PGF2α and PGE2 were 50% of those in Soy mice, as a result of the increased EPA/AA ratio. PGF3α and PGE3 were undetectable in the uterine tissues of Soy and Lin mice. Interestingly, in Lin mice, 'luteolytic' PGF2α synthesis was considerably maintained even in the ω6 PUFA-reduced condition. These results suggest the existence of an elaborate mechanism securing PGF2α synthesis to a level that is sufficient for triggering luteolysis and parturition, even under ω6 PUFA-reduced conditions.

Potential roles of imprinted genes in the teratogenic effects of alcohol on the placenta, somatic growth, and the developing brain.

Despite several decades of research and prevention efforts, fetal alcohol spectrum disorders (FASD) remain the most common preventable cause of neurodevelopmental disabilities worldwide. Animal and human studies have implicated fetal alcohol-induced alterations in epigenetic programming as a chief mechanism in FASD. Several studies have demonstrated fetal alcohol-related alterations in methylation and expression of imprinted genes in placental, brain, and embryonic tissue. Imprinted genes are epigenetically regulated in a parent-of-origin-specific manner, in which only the maternal or paternal allele is expressed, and the other allele is silenced. The chief functions of imprinted genes are in placental development, somatic growth, and neurobehavior-three domains characteristically affected in FASD. In this review, we summarize the growing body of literature characterizing prenatal alcohol-related alterations in imprinted gene methylation and/or expression and discuss potential mechanistic roles for these alterations in the teratogenic effects of prenatal alcohol exposure. Future research is needed to examine potential physiologic mechanisms by which alterations in imprinted genes disrupt development in FASD, which may, in turn, elucidate novel targets for intervention. Furthermore, mechanistic alterations in imprinted gene expression and/or methylation in FASD may inform screening assays that identify individuals with FASD neurobehavioral deficits who may benefit from early interventions.

Conjugation to PEG as a Strategy to Limit the Uptake of Drugs by the Placenta: Potential Applications for Drug Administration in Pregnancy.

Here, we evaluated the feasibility of non-prodrug PEG-drug conjugates to decrease the accumulation of drugs within the placental tissues. The results showed that PEG was biocompatible with the human placenta with no alteration of the basal rate of proliferation or apoptosis in term placental explants. No significant changes in the released levels of lactate dehydrogenase and the human chorionic gonadotropin were observed after PEG treatment. The cellular uptake studies revealed that conjugating Cy5.5 and haloperidol to PEG significantly reduced (by up to ∼40-fold) their uptake by the placenta. These findings highlight the viability of novel non-prodrug polymer-drug conjugates to avoid the accumulation of drugs within the placenta.

The selective progesterone receptor modulator-promegestone-delays term parturition and prevents systemic inflammation-mediated preterm birth in mice.

BACKGROUND: Progesterone, acting via its nuclear receptors called progesterone receptors, promotes myometrial relaxation during pregnancy, and suspension of this activity triggers labor. We previously found that 20α-hydroxysteroid dehydrogenase causes a local withdrawal of progesterone in the term and preterm myometrium by converting the progesterone into an inactive form before it accesses the progesterone receptors. OBJECTIVE: We hypothesized that a selective progesterone receptor modulator called promegestone, which is not metabolized by 20α-hydroxysteroid dehydrogenase, would sustain progesterone receptor signaling and prevent/delay term labor and preterm labor in mice. STUDY DESIGN: In the term labor mouse model, promegestone (0.2 mg/dam) or a vehicle were administered subcutaneously in timed-pregnant CD-1 mice at gestational days 15, 16, and 17 (term gestational days, 19.5). In the inflammation preterm labor model, pregnant mice received promegestone or a vehicle on gestational days 15, 16, and 17, which was 24 hours before, immediately before, and 24 hours after systemic bacterial endotoxin (50 µg intraperitoneal; lipopolysaccharide group) or vehicle (saline) administration. The maternal and fetal tissues were collected on gestational day 16 6 hours after lipopolysaccharide±promegestone injection and at term gestational day 18.75. The protein levels of 10 cytokines were measured by multiplex immunoassay in maternal plasma and amniotic fluid. Myometrial, decidual, and placental messenger RNA levels of multiple cytokines and procontractile proteins were evaluated by real-time polymerase chain reaction and confirmed by immunoblotting. RESULTS: Promegestone prevented term labor and maintained mice pregnancy postterm >24 hours. The litter size and fetal weights were not different from the controls. Promegestone prevented systemic bacterial-endotoxin-induced preterm labor in 100% of the mice, blocked uterine contractions, significantly inhibited all systemic inflammation-induced myometrial cytokines, and partially inhibited decidual and placental inflammation. Promegestone did not prevent bacterial-endotoxin-induced fetal toxicity. CONCLUSION: Promegestone a selective progesterone receptor modulator that binds progesterone receptors with high affinity and is not metabolized by 20α-hydroxysteroid dehydrogenase could completely suppress term parturition and systemic bacterial-endotoxin-induced preterm birth in mice. We suggest that such selective progesterone receptor modulators may represent a potential therapeutic approach to the prevention of preterm labor in women at high risk of preterm birth.

Effects of early daily alcohol exposure on placental function and fetal growth in a rhesus macaque model.

BACKGROUND: Prenatal alcohol exposure is the most common cause of birth defects and intellectual disabilities and can increase the risk of stillbirth and negatively impact fetal growth. OBJECTIVE: To determine the effect of early prenatal alcohol exposure on nonhuman primate placental function and fetal growth. We hypothesized that early chronic prenatal alcohol would alter placental perfusion and oxygen availability that adversely affects fetal growth. STUDY DESIGN: Rhesus macaques self-administered 1.5 g/kg/d of ethanol (n=12) or isocaloric maltose-dextrin (n=12) daily before conception through the first 60 days of gestation (term is approximately 168 days). All animals were serially imaged with Doppler ultrasound to measure fetal biometry, uterine artery volume blood flow, and placental volume blood flow. Following Doppler ultrasound, all animals underwent both blood oxygenation level-dependent magnetic resonance imaging to characterize placental blood oxygenation and dynamic contrast-enhanced magnetic resonance imaging to quantify maternal placental perfusion. Animals were delivered by cesarean delivery for placental collection and fetal necropsy at gestational days 85 (n=8), 110 (n=8), or 135 (n=8). Histologic and RNA-sequencing analyses were performed on collected placental tissue. RESULTS: Placental volume blood flow was decreased at all gestational time points in ethanol-exposed vs control animals, but most significantly at gestational day 110 by Doppler ultrasound (P<.05). A significant decrease in total volumetric blood flow occurred in ethanol-exposed vs control animals on dynamic contrast-enhanced magnetic resonance imaging at both gestation days 110 and 135 (P<.05); moreover, a global reduction in T2∗, high blood deoxyhemoglobin concentration, occurred throughout gestation (P<.05). Similarly, evidence of placental ischemic injury was notable by histologic analysis, which revealed a significant increase in microscopic infarctions in ethanol-exposed, not control, animals, largely present at middle to late gestation. Fetal biometry and weight were decreased in ethanol-exposed vs control animals, but the decrease was not significant. Analysis with RNA sequencing suggested the involvement of the inflammatory and extracellular matrix response pathways. CONCLUSION: Early chronic prenatal alcohol exposure significantly diminished placental perfusion at mid to late gestation and also significantly decreased the oxygen supply to the fetal vasculature throughout pregnancy, these findings were associated with the presence of microscopic placental infarctions in the nonhuman primate. Although placental adaptations may compensate for early environmental perturbations to fetal growth, placental blood flow and oxygenation were reduced, consistent with the evidence of placental ischemic injury.

Elevated Serum SFRP5 Levels During Preeclampsia and Its Potential Association with Trophoblast Dysfunction via Wnt/ß-Catenin Suppression.

Preeclampsia (PE) is a life-threatening pregnancy complication associated with diminished trophoblast migration and invasion. Wnt signalling is one of the most important regulators of placentation. Secreted frizzled-related protein 5 (SFRP5) is an anti-inflammatory adipokine that may inhibit Wnt signalling. In this study, we aimed to investigate the relationship between SFRP5 and PE and its effect on trophoblast function, as well as the underlying signalling pathways. SFRP5 levels in the serum and placental tissues were detected using enzyme-linked immunosorbent assay and immunohistochemistry, respectively. To evaluate the effect of SFRP5 on Wnt signalling, the human trophoblast cell line HTR8/SVneo was treated with recombinant human SFRP5 and Dickkopf-related protein 1 (Dkk-1, canonical Wnt inhibitor) proteins and lithium chloride (LiCl, canonical Wnt agonist). The migration and invasion ability of HTR8/SVneo cells was evaluated using wound-healing and Matrigel Transwell assays. The activities of multiple matrix metalloproteinases (MMP)-2/9 were detected using gelatin zymography. Expression of glycogen synthase kinase-3 beta (GSK3ß) and ß-catenin proteins was investigated using western blotting. The serum SFRP5 levels were elevated in patients with PE, but SFRP5 expression was not detected in the placental tissues. Furthermore, SFRP5 inhibited the migration and invasion of HTR8/SVneo cells in vitro, increased GSK3ß, and decreased ß-catenin expression and MMP-2/9 activity in HTR8/SVneo cells. In conclusion, this study suggests that SFRP5 inhibits trophoblast migration and invasion potentially via the inhibition of Wnt/ß-catenin signalling, which might be involved in the development of PE. However, the primary cause of the increased SFRP5 levels needs to be investigated.

Acetone Ingestion Mimics a Fasting State to Improve Glucose Tolerance in a Mouse Model of Gestational Hyperglycemia.

Gestational diabetes mellitus results, in part, from a sub-optimal ß-cell mass (BCM) during pregnancy. Artemisinins were reported to increase BCM in models of diabetes by α- to ß-cell conversion leading to enhanced glucose tolerance. We used a mouse model of gestational glucose intolerance to compare the effects of an artemisinin (artesunate) on glycemia of pregnant mice with vehicle treatment (acetone) or no treatment. Animals were treated daily from gestational days (GD) 0.5 to 6.5. An intraperitoneal glucose tolerance test was performed prior to euthanasia at GD18.5 or post-partum. Glucose tolerance was significantly improved in both pregnant and non-pregnant mice with both artesunate and vehicle-alone treatment, suggesting the outcome was primarily due to the acetone vehicle. In non-pregnant, acetone-treated animals, improved glucose tolerance was associated with a higher BCM and a significant increase in bihormonal insulin and glucagon-containing pancreatic islet cells, suggesting α- to ß-cell conversion. BCM did not differ with treatment during pregnancy or post-partum. However, placental weight was higher in acetone-treated animals and was associated with an upregulation of apelinergic genes. Acetone-treated animals had reduced weight gain during treatment despite comparable food consumption to non-treated mice, suggesting transient effects on nutrient uptake. The mean duodenal and ileum villus height was reduced following exposure to acetone. We conclude that acetone treatment may mimic transient fasting, resulting in a subsequent improvement in glucose tolerance during pregnancy.

Suppression of uterine and placental ferroptosis by N-acetylcysteine in a rat model of polycystic ovary syndrome.

The mechanisms that link hyperandrogenism and insulin (INS) resistance (HAIR) to the increased miscarriage rate in women with polycystic ovary syndrome (PCOS) remain elusive. Previous studies demonstrate that increased uterine and placental ferroptosis is associated with oxidative stress-induced fetal loss in a pre-clinical PCOS-like rat model. Here, we investigated the efficacy and molecular mechanism of action of the antioxidant N-acetylcysteine (NAC) in reversing gravid uterine and placental ferroptosis in pregnant rats exposed to 5α-dihydrotestosterone (DHT) and INS. Molecular and histological analyses showed that NAC attenuated DHT and INS-induced uterine ferroptosis, including dose-dependent increases in anti-ferroptosis gene content. Changes in other molecular factors after NAC treatment were also observed in the placenta exposed to DHT and INS, such as increased glutathione peroxidase 4 protein level. Furthermore, increased apoptosis-inducing factor mitochondria-associated 2 mRNA expression was seen in the placenta but not in the uterus. Additionally, NAC was not sufficient to rescue DHT + INS-induced mitochondria-morphological abnormalities in the uterus, whereas the same treatment partially reversed such abnormalities in the placenta. Finally, we demonstrated that NAC selectively normalized uterine leukemia inhibitory factor, osteopontin/secreted phosphoprotein 1, progesterone receptor, homeobox A11 mRNA expression and placental estrogen-related receptor beta and trophoblast-specific protein alpha mRNA expression. Collectively, our data provide insight into how NAC exerts beneficial effects on differentially attenuating gravid uterine and placental ferroptosis in a PCOS-like rat model with fetal loss. These results indicate that exogenous administration of NAC represents a potential therapeutic strategy in the treatment of HAIR-induced uterine and placental dysfunction.