The Influence of Exercise-Associated Small Extracellular Vesicles on Trophoblasts In Vitro.

Exercise induces the release of small extracellular vesicles (sEVs) into circulation that are postulated to mediate tissue cross-talk during exercise. We previously reported that pregnant individuals released greater levels of sEVs into circulation after exercise compared to matched non-pregnant controls, but their biological functions remain unknown. In this study, sEVs isolated from the plasma of healthy pregnant and non-pregnant participants after a single bout of moderate-intensity exercise were evaluated for their impact on trophoblasts in vitro. Exercise-associated sEVs were found localized within the cytoplasm of BeWo choriocarcinoma cells, used to model trophoblasts in vitro. Exposure to exercise-associated sEVs did not significantly alter BeWo cell proliferation, gene expression of angiogenic growth factors VEGF and PLGF, or the release of the hormone human chorionic gonadotropin. The results from this pilot study support that exercise-associated sEVs could interact with trophoblasts in vitro, and warrant further investigation to reveal their potential role in communicating the effects of exercise to the maternal-fetal interface.

Elucidating the interaction between maternal physical activity and circulating myokines throughout gestation: A scoping review.

Physical activity (PA) during pregnancy provides both maternal and fetal health benefits. It has been theorized that myokines, peptides secreted by contracting skeletal muscle, may play an important mechanistic role in facilitating the health benefits obtained from prenatal exercise. The objective of this review was to synthesize the current literature on the relationship between maternal PA and myokine response. A search strategy was developed using the terms pregnancy, PA, IL-6, IL-10, IL-13, and TNF-α. A systematic search was completed in July 2020, in Medline, SPORTDiscus, EMBASE, CENTRAL, and in November 2020 for unpublished dissertations (grey literature; Proquest). Both human- and animal-based studies of any design were included, while commentaries and editorial articles were excluded. Data were extracted by two independent reviewers and summarized narratively. Data were thematically summarized based on the myokine and whether findings were from human or animal studies. Ten studies were included in this review. Findings from studies that examined IL-6, IL-10, and TNF-α suggest a trimester-specific interaction between PA and myokine levels; no studies evaluated IL-13. Future research should investigate the PA-myokine relationship throughout all stages of gestation.

Circulating small extracellular vesicles increase after an acute bout of moderate-intensity exercise in pregnant compared to non-pregnant women.

The physiological and molecular mechanisms linking prenatal physical activity and improvements in maternal-fetal health are unknown. It is hypothesized that small extracellular vesicles (EVs, ~ 10-120 nm) are involved in tissue cross-talk during exercise. We aimed to characterize the circulating small EV profile of pregnant versus non-pregnant women after an acute bout of moderate-intensity exercise. Pregnant (N = 10) and non-pregnant control (N = 9) women performed a single session of moderate-intensity treadmill walking for 30 min. Plasma was collected immediately pre- and post-exercise, and small EVs were isolated by differential ultracentrifugation. EV presence was confirmed by western blotting for the small EV proteins TSG-101 and flottilin-1. Small EVs were quantified by size and concentration using nanoparticle tracking analysis and transmission electron microscopy. All EV fractions were positive for TSG-101 and flotillin-1, and negative for calnexin. Mean vesicle size at baseline and percent change in size post-exercise were not different between groups. At baseline, pregnant women had higher levels of small EVs compared to controls (1.83E+10 ± 1.25E+10 particles/mL vs. 8.11E+09 ± 4.04E+09 particles/mL, respectively; p = 0.032). Post-exercise, small EVs increased significantly in the circulation of pregnant compared to non-pregnant women after correcting for baseline values (64.7 ± 24.6% vs. - 23.3 ± 26.1%, respectively; F = 5.305, p = 0.035). Further research is needed to assess the functional roles of exercise-induced small EVs in pregnancy.

Does exercise during pregnancy impact organs or structures of the maternal-fetal interface?

Exercise during pregnancy has been shown to be associated with improved health outcomes both during and after pregnancy for mother and fetus across the lifespan. Increasing physical activity and reducing sedentary behaviour during pregnancy have been recommended by many researchers and clinicians-alike. It is thought that the placenta plays a central role in mediating any positive or negative pregnancy outcomes. The positive outcomes obtained through prenatal exercise are postulated to result from exercise-induced regulation of maternal physiology and placental development. Considerable research has been performed to understand the placenta's role in pregnancy-related diseases, such as preeclampsia, fetal growth restriction, and gestational diabetes mellitus. However, little research has examined the potential for healthy lifestyle and behavioural changes to improve placental growth, development, and function. While the placenta represents the critical maternal-fetal interface responsible for all gas, nutrient, and waste exchange between the mother and fetus, the impact of exercise during pregnancy on placental biology and function is not well known. This review will focus on prenatal exercise and its promising influence on the structures of the maternal-fetal interface, with particular emphasis on the placenta. Potential molecular mechanistic hypotheses are presented to aid future investigations of prenatal exercise and placental health.

Placental superoxide dismutase 3 mediates benefits of maternal exercise on offspring health.

Poor maternal diet increases the risk of obesity and type 2 diabetes in offspring, adding to the ever-increasing prevalence of these diseases. In contrast, we find that maternal exercise improves the metabolic health of offspring, and here, we demonstrate that this occurs through a vitamin D receptor-mediated increase in placental superoxide dismutase 3 (SOD3) expression and secretion. SOD3 activates an AMPK/TET signaling axis in fetal offspring liver, resulting in DNA demethylation at the promoters of glucose metabolic genes, enhancing liver function, and improving glucose tolerance. In humans, SOD3 is upregulated in serum and placenta from physically active pregnant women. The discovery of maternal exercise-induced cross talk between placenta-derived SOD3 and offspring liver provides a central mechanism for improved offspring metabolic health. These findings may lead to novel therapeutic approaches to limit the transmission of metabolic disease to the next generation.

Physical activity differentially regulates VEGF, PlGF, and their receptors in the human placenta.

Physical activity (PA) has beneficial effects on the function of many organs by modulating their vascular development. Regular PA during pregnancy is associated with favorable short- and long-term outcomes for both mother and fetus. During pregnancy, appropriate vascularization of the placenta is crucial for adequate maternal-fetal nutrient and gas exchange. How PA modulates angiogenic factors, VEGF, and its receptors in the human placenta, is as of yet, unknown. We objectively measured the PA of women at 24-28 and 34-38 weeks of gestation. Participants were considered "active" if they had met or exceeded 150 min of moderate-intensity PA per week during their 2nd trimester. Term placenta tissues were collected from active (n = 23) or inactive (n = 22) women immediately after delivery. We examined the expression of the angiogenic factors VEGF, PlGF, VEGFR-1, and VEGFR-2 in the placenta. Western blot analysis showed VEGF and its receptor, VEGFR-1 was significantly (p < 0.05) higher both at the protein and mRNA levels in placenta from physically active compared to inactive women. No difference in VEGFR-2 was observed. Furthermore, immunohistochemistry showed differential staining patterns of VEGF and its receptors in placental endothelial, stromal, and trophoblast cells and in the syncytial brush border. In comparison, PlGF expression did not differ either at the protein or mRNA level in the placenta from physically active or inactive women. The expression and localization pattern of VEGF and its receptors suggest that PA during pregnancy may support a pro-angiogenic milieu to the placental vascular network.

Physical activity may be an adjuvant treatment option for substance use disorders during pregnancy: A scoping review.

BACKGROUND: Substance abuse in pregnancy increases the chance of physical and neurobehavioral disabilities as well as many other undesirable fetal outcomes. In nonpregnant populations, physical exercise has shown to be an effective adjunctive therapy option for substance use disorders. Given the known positive maternal and fetal physiological and mental health benefits associated with prenatal exercise, perhaps exercise during pregnancy may also be a viable adjuvant therapy option for women with substance use disorders. The purpose of this scoping review was to summarize the available literature that has assessed the relationship between prenatal exercise and substance use disorders. METHODS: A search strategy was developed combining the terms pregnancy, exercise/physical activity, and substance use. A systematic search was completed in the following databases: Medline/PubMed, SPORTDiscus, and ProQuest. Substances eligible for inclusion included illicit drugs, alcohol, and cannabis. Retrieved data were categorized as animal or human model studies, and were summarized narratively. RESULTS: Eight studies were included in this review (five human studies, three animal model studies). Studies in humans suggest that pregnant women with substance use disorders are interested in engaging in physical activity interventions; however, known acute metabolic and physiological responses to prenatal exercise may be impaired in this population. Rodent models show preliminary evidence for improved mental health outcomes following prenatal exercise for substance use disorders. CONCLUSION: The findings from this review may inform the development of future clinical trials to test the effect of structured exercise programs as an adjunctive treatment option for pregnant women with substance use disorders.

The Effect of Maternal Physical Activity and Gestational Weight Gain on Placental Efficiency.

INTRODUCTION: Adherence to physical activity (PA) and gestational weight gain (GWG) recommendations during pregnancy has been shown to improve maternal and fetal health outcomes, including reducing the risk for chronic diseases. Limited research has evaluated the effect of meeting PA in combination with GWG recommendations on placental efficiency (Pl-E), a surrogate marker of the placenta's ability to exchange nutrients and gas based on surface area. The purpose of this study was to measure and compare Pl-E based on meeting PA and GWG recommendations. METHOD: Healthy pregnant women (n = 61) wore accelerometers in their second and third trimesters to objectively measure PA. Women were classified as active or inactive at each time point based on meeting the 2019 Canadian prenatal PA guidelines. Total GWG was calculated as weight measured in the third trimester minus self-reported prepregnancy weight, and were categorized as insufficient (n = 19), adequate (n = 22), and excessive (n = 20) according to the 2009 Institute of Medicine guidelines. Placental weight (PW) and birth weight (BW) were measured within 30 min of delivery and 24-48 h postdelivery, respectively. Pl-E was determined in three ways: BW:PW ratio, residual BW, and measured BW, with a higher value indicating better Pl-E. Pl-E was compared by PA and GWG status using a two-way ANOVA. RESULTS: No differences were found in the BW:PW ratio or residual BW corresponding to PA and GWG status. Measured BW was significantly higher in newborns of women who gained weight excessively compared with those who gained insufficient weight (P < 0.05). CONCLUSION: These findings suggest that prenatal PA does not compromise Pl-E; however, further research is required to evaluate the potential mechanistic benefits of meeting PA and GWG guidelines on the placenta.

Does "Sitting" Stand Alone? A Brief Report Evaluating the Effects of Prenatal Sedentary Time on Maternal and Newborn Anthropometric Outcomes.

BACKGROUND: Research on sedentary behavior and effects on maternal and newborn outcomes has been inconclusive. The objective of this report was to correlate sedentary time with maternal and fetal anthropometric measurements and compare the effect on sedentary time based on meeting prenatal activity guidelines. METHODS: Healthy pregnant women (N = 61) in their second trimester (24-28 wk gestation) provided 7-day accelerometry data. Outcomes, including neonatal weight, length, and body fat percentage, were collected 24 to 48 hours after delivery. Placenta weight was measured immediately after delivery. Gestational weight gain was calculated by subtracting self-reported prepregnancy weight from measured weight at 38 weeks gestation. Correlations between sedentary time and outcomes were tested with Spearman and Pearson coefficient of correlations in all women separately and in accordance with the 2019 Canadian prenatal exercise guidelines. RESULTS: No significant associations were found between sedentary time and the selected outcomes, even when compared by prenatal exercise level. There was no difference in total time spent sedentary between active (576.7 [52.8] min) and inactive women (599.3 [51.6] min). CONCLUSIONS: Meeting exercise recommendations during pregnancy does not significantly decrease total sedentary time. Future studies should aim to evaluate the health effects of both decreasing sedentary time and meeting prenatal exercise guidelines.

Physical Activity During Pregnancy Is Associated with Increased Placental FATP4 Protein Expression.

Placental function is of utmost importance to ensure proper fetal development in utero. Among the placenta's many roles includes the passage of sufficient macronutrients, such as glucose, amino acids, and fatty acids, to the fetus. Macronutrients are carried from maternal circulation to the fetus across transporters within the placenta. The objective of this study was to examine the impact of (i) an acute bout of exercise and (ii) chronic exercise participation on placenta nutrient transporter expression and localization. To investigate the effect of acute exercise, pre- and post-exercise serum was collected from pregnant (n = 5) and non-pregnant (n = 5) women who underwent a moderate-intensity exercise session and used to treat BeWo cells. To assess chronic physical activity, we analyzed term placenta from women categorized as active (n = 10) versus non-active (n = 10). Protein expression and localization for the transporters GLUT1, SNAT1, and FATP4 were examined for both groups. GLUT1 expression in BeWo cells treated with serum from pregnant women was higher compared with that from non-pregnant, independent of exercise. FATP4 protein expression was elevated in the term placenta of active women. Immunohistochemistry analysis of term placenta illustrated increased staining of FATP4 in placental tissue from active women and differential staining pattern of GLUT1 depending on physical activity status. Chronic exercise during pregnancy increases the expression of placental FATP4 in vivo, suggesting greater metabolism and usage of fatty acids. Additionally, serum from pregnant women could contain factors that increase GLUT1 protein expression in vitro. BeWo cells treated with pre- and post-exercise serum from pregnant women resulted in greater GLUT1 expression compared with those treated with pre- and post-exercise serum from non-pregnant women. Physical activity appears to differentially impact key placental transporters involved in the transfer and availability of nutrients from mother to fetus. Future research ought to examine the mechanisms involved in regulating these changes and their impact on fetal growth and health.

Serum glucose, urea nitrogen, cholesterol, and total proteins in crossbred repeat breeder and normally cyclic cows.

OBJECTIVE: This study was designed to determine and compare the serum glucose, urea nitrogen, cholesterol, and total protein (TP) level in crossbred repeat breeder (RB) and normally cyclic cows to find out the relationship of these metabolic factors with repeat breeding syndrome (RBS). MATERIALS AND METHODS: A total of 592 breedable cows from 34 farms were examined from Mymensingh and Chittagong districts. Seventy cows were identified as RB cows and another 10 cows were randomly selected as normally cyclic control cows for this study. Blood sample from each cow was collected and the serum was separated. The serum samples were analyzed by auto blood analyzer. RESULTS: Cows affected with RBS had significant variation in their glucose, urea, and cholesterol levels. Serum glucose (43.00 gm/dl) level was significantly lower than in normally cyclic cows. On the other hand, serum urea and cholesterol levels were significantly higher than in normally cyclic cows. However, the TP concentrations did not differ between RB and cyclic cows. RB cows had a lower trend (6.815 mg/dl) of serum TP than in normally cyclic cows. CONCLUSION: This research might help scientists and veterinarians to understand that the high serum urea and cholesterol level along with low glucose and TP level could have some effect in the development of RBS in crossbred cows. It will potentially help veterinary practitioners and farmers to take preventive measures against RBS of crossbred cows.

Factor inhibiting HIF1-A novel target of SUMOylation in the human placenta.

Adaptations to changes in oxygen are critical to ensure proper placental development, and impairments in oxygen sensing mechanisms characterize placental pathologies such as preeclampsia. In this study, we examined the involvement of SUMOylation, a reversible posttranslational modification, in the regulation of the asparaginyl hydroxylase Factor Inhibiting Hypoxia Inducible Factor 1 (FIH1) in the human placenta in development and in disease status. FIH1 protein abundance and spatial distribution in the developing placenta directly correlated with oxygen tension in vivo. Immunofluorescence analysis showed that early on FIH1 primarily localized to nuclei of cytotrophoblast cells, while after 10 weeks of gestation it was present in nuclei and cytoplasm of both cytotrophoblast and syncytiotrophoblast cells. Exposure of choriocarcinoma JEG-3 cells to hypoxia induced FIH1 SUMOylation by promoting its association to SUMO2/3. Transfection of JEG-3 cells with FIH1 constructs containing SUMO-mutated sites revealed that SUMOylation of FIH1 by SUMO2/3 targeted it for proteasomal degradation, particularly in hypoxia. SUMOylation of FIH1 directly impacted on HIF1A activity as determined by HIF-responsive luciferase assay. Co-immunoprecipitation analyses revealed enhanced FIH1-SUMO2/3 associations early in development, when FIH1 levels are low, while deSUMOylation of FIH1 by SENP3 increased later in gestation, when FIH1 levels are rising. In preeclampsia, decreased FIH1 protein expression associated with impaired deSUMOylation by SENP3 and increased association with the ubiquitin ligase RNF4. We propose a novel mode of regulation of FIH1 stability by dynamic SUMOylation and deSUMOylation in the human placenta in response to changing oxygen tension, thereby mediating HIF1A transcriptional activity in physiological and pathological conditions.

Dynamic regulation of HIF1Α stability by SUMO2/3 and SENP3 in the human placenta.

INTRODUCTION: Hypoxia-inducible factor 1A (HIF1A) stability is tightly regulated by hydroxylation and ubiquitination. Emerging evidence indicates that HIF1A is also controlled by the interplay between SUMO-specific ligases, which execute protein SUMOylation, and Sentrin/SUMO-specific proteases that de-SUMOylate target proteins. Given the significance of HIF1A in the human placenta, we investigated whether placental HIF1A is subject to SUMOylation in physiological and pathological conditions. METHODS: Placentae were obtained from normal and pregnancies complicated by preeclampsia. Human choriocarcinoma JEG3 cells were maintained at either 21% or 3% oxygen or exposed to sodium nitroprusside (SNP). Cells were transfected with SUMO2/3 constructs with and without Mg132, a proteasome inhibitor. Expression, distribution and associations of SUMO/SENPs and HIF1A were evaluated by Western blotting, immunohistochemistry and co-immunoprecipitation. RESULTS: HIF1A-SUMO2/3 associations peaked at 9-10 weeks, while its deSUMOylation by SENP3 was greatest at 10-12 weeks. In E-PE, HIF1A deSUMOylation by SENP3 was significantly elevated, while HIF1A-SUMO2/3 associations remained constant. In vitro, overexpression of SUMO2/3 de-stabilized HIF1A in hypoxia, and abrogated HIF1A expression following Mg132 treatment in normoxia. Hypoxia and SNP treatments promoted SENP3 nuclear redistribution from nucleoli to the nucleoplasm where it associates with HIF1A. CONCLUSION: During early placental development, SUMOylation events control HIF1A stability in an oxygen-dependent manner. In E-PE, enhanced deSUMOylation of HIF1A by SENP3 may in part contribute to increased HIF1A activity and stability found in this pathology.

Oxygen governs Galß1-3GalNAc epitope in human placenta.

It is becoming increasingly apparent that the dynamics of glycans reflect the physiological state of cells involved in several cell functions including growth, response to signal molecules, migration, as well as adhesion to, interaction with, and recognition of other cells. The presence of glycoconjugates in human placenta suggests their major role in maternal-fetal exchanges, intercellular adhesion, cellular metabolism, and villous vessel branching. Although several studies have described glycoconjugate distribution in the human placenta descriptions of their physiological function and control mechanisms during placental development are lacking. In this study we investigated the developmental distribution and regulation of placental core 1 O- and N-glycans focusing on early and late first trimester human pregnancy. To define the control mechanisms of the oligosaccharide chains during early placentation process, chorionic villous explants and human trophoblast cell lines were exposed to various oxygen levels. We found that oxygen tension regulates changes in core-1 O-glycan (the disaccharide Galß1-3GalNAc) epitope expression levels. Moreover, by double affinity chromatography and subsequent analysis with mass spectrometry, we identified in the heat shock protein 90-α (HSP90α) a good candidate as carrier of the Galß1-3GalNAc epitope at low oxygen tension. Our results support a fundamental role of oxygen tension in modulating glycosylation of proteins during placental development.

Milk progesterone enzyme-linked immunosorbent assay as a tool to investigate ovarian cyclicity of water buffaloes in relation to body condition score and milk production.

BACKGROUND: Application of assisted reproductive technologies in buffaloes is limited to some extent by farmers' inability to detect oestrus because of its poor expression. The present study aimed at investigating reliability of a milk progesterone enzyme-linked immunosorbent assay (ELISA) to assess the ovarian cyclicity during post partum, oestrus and post-breeding periods in water buffaloes. METHODS: Progesterone concentrations were measured by an ELISA in milk of 23 postpartum buffaloes in relation to oestrus, pregnancy, body condition score (BCS) and milk production. Two milk samples were taken at 10 days intervals, every month starting from day 30 and continued to day 150 post partum. BCS and milk production were recorded during sample collection. Milk samples from bred buffaloes were collected at Day 0 (day of breeding), Days 10-12 and Days 22-24. Defatted milk was preserved at -80°C until analysis. Pregnancy was confirmed by palpation per rectum on Days 70-90. RESULTS: Seventeen buffaloes had 47 ovulatory cycles, one to four in each, 13 were detected in oestrus once (28 % oestrus detection rate). Progesterone concentration ≥1 ng/ml in one of the two 10-day-interval milk samples reflected ovulation and corpus luteum formation. The intervals between calving to first luteal activity and to first detected oestrus varied from 41 to 123 days (n = 17) and 83 to 135 (n = 13) days, respectively. Eight buffaloes were bred in the course of the study and seven were found pregnant. These buffaloes had a progesterone profile of low (<1 ng/ml), high (≥ 1 ng/ml) and high (≥ 1 ng/ml) on Day 0, Days 10-12 and Days 22-24, respectively. Buffaloes cycling later in the postpartum period had fewer missed oestruses (P < 0.05). Buffaloes with a superior BCS had a shorter calving to oestrus interval and produced more milk (P < 0.05). CONCLUSIONS: Milk progesterone ELISA is a reliable tool for monitoring ovarian cyclicity and good BCS may be an indicator of resuming cyclicity in water buffalo.

Pro-inflammatory cytokines in animal and human gestation.

The story of cytokines in pregnancy began about 30 years ago, approximately in concomitance with the understanding that cytokines are autocrine-paracrine regulators of physiological processes. Pro-inflammatory cytokines are predominant in the early and late events of gestation, e.g. pregnancy establishment and parturition, both of which have been described as inflammatory-like events. Pro-inflammatory cytokines are also produced in response to microbes constantly in contact with the female reproductive tract. While a pro-inflammatory response is beneficial to successful pregnancy, an exaggerated response, as may occur for an unresolved infection, could result in an unfavorable pregnancy outcome in animals and humans. Therapeutic strategies are required to avoid the risks to the health of fetus and mother. In this review, we discuss the involvement of pro-inflammatory cytokines in pregnancy at implantation and parturition, including the pathologies which might be related to an alteration of the cytokine levels. We also deal with the use of anti-cytokines and/or anti-inflammatory mediators to antagonize the action of pro-inflammatory cytokines. Finally we discuss the potential of animal models to evaluate the association of cytokines in the establishment and maintenance of pregnancy.

17{beta}-Estradiol modulates the macrophage migration inhibitory factor secretory pathway by regulating ABCA1 expression in human first-trimester placenta.

Successful pregnancy involves a series of events, most of them mediated by hormones and cytokines. Estrogens, besides being important for placental growth and embryo development, have a marked effect on the immune system exerting either pro- or anti-inflammatory properties. Numerous studies suggest that estrogens directly affect cellular function, including cytokine production. Macrophage migration inhibitory factor (MIF) is a proinflammatory cytokine involved in pregnancy, particularly during the earlier stages of placentation. Since reports on mice have shown that estrogens modulate MIF, herein we investigated the effect of estrogens on human placental MIF. By using an in vitro model of first-trimester chorionic villous explants, we found that 17beta-estradiol (E(2)) was able to modulate the release of MIF in a dose-dependent manner (10(-12) vs. 10(-9) M, P < 0.05; 10(-9) vs. 10(-5) M, P < 0.05; 10(-12) vs. 10(-5) M, P < 0.001). Unlike MIF release, no significant change in tissue MIF protein or MIF mRNA was observed. We showed evidence that E(2) concentrations (10(-9) and 10(-5) M) act on placental tissue downregulating the mRNA and protein expression of the ATP-binding cassette transporter protein A1, a membrane transporter involved in MIF secretion. These findings emphasize the mutual cooperation between hormones and cytokines and suggest that increasing estrogen levels with advancing gestation may have a major role in regulating placental MIF secretion.