Stages Of Human Pregnancy Research Articles

A discussion on the 'dispensable' amino acids.

Recently, the concept of dietary dispensable amino acids has been increasingly challenged, as more indirect and direct (experimental) evidence has pointed to the need for them in the diet during specific life stages or disease states. Here, we discuss the classification of amino acids, methods to assess the needs for dispensable amino acids with experimental evidence from our recent studies, and highlight the role of specific dispensable amino acids in metabolism and health. There exist differences among the dispensable amino acids to act as effective nitrogen sources in humans. Glycine, a dispensable amino acid is conditionally indispensable in later stages of human pregnancy. The so-called 'dispensable' amino acids are quantitatively nearly 75% of the daily protein needs in humans. In certain life-stages and diseases, there is a dietary demand for the dispensable amino acids. Future well-designed studies are required to identify the dietary demand for these amino acids, which will certainly be useful for dietary management in specific diseases and to maintain health across all life-stages.

Modeling human peri-implantation placental development and function†.

It is very difficult to gain a better understanding of the events in human pregnancy that occur during and just after implantation because such pregnancies are not yet clinically detectable. Animal models of human placentation are inadequate. In vitro models that utilize immortalized cell lines and cells derived from trophoblast cancers have multiple limitations. Primary cell and tissue cultures often have limited lifespans and cannot be obtained from the peri-implantation period. We present here two contemporary models of human peri-implantation placental development: extended blastocyst culture and stem-cell derived trophoblast culture. We discuss current research efforts that employ these models and how such models might be used in the future to study the "black box" stage of human pregnancy.

Histopathological lesions of congenital Zika syndrome in newborn squirrel monkeys

The absence of an adequate animal model for studies has limited the understanding of congenital Zika syndrome (CZS) in humans during the outbreak in America. In this study, we used squirrel monkeys (Saimiri collinsi), a neotropical primate (which mimics the stages of human pregnancy), as a model of Zika virus (ZIKV) infection. Seven pregnant female squirrel monkeys were experimentally infected at three different gestational stages, and we were able reproduce a broad range of clinical manifestations of ZIKV lesions observed in newborn humans. Histopathological and immunohistochemical analyses of early-infected newborns (2/4) revealed damage to various areas of the brain and ZIKV antigens in the cytoplasm of neurons and glial cells, indicative of CZS. The changes caused by ZIKV infection were intrauterine developmental delay, ventriculomegaly, simplified brain gyri, vascular impairment and neuroprogenitor cell dysfunction. Our data show that the ZIKV infection outcome in squirrel monkeys is similar to that in humans, indicating that this model can be used to help answer questions about the effect of ZIKV infection on neuroembryonic development and the morphological changes induced by CZS.

Estrogen-enhanced apical and basolateral secretion of apolipoprotein B-100 by polarized trophoblast-derived BeWo cells

Cholesterol is an important nutrient for fetal development and transplacental transport occurs at all stages of human pregnancy. Furthermore, cholesterol is required for membrane building as well as steroid hormone synthesis. Therefore, all placental cell types require cholesterol for proper function. In human term placenta, the syncytiotrophoblast (STB) faces the maternal circulation. Uptake of maternal-derived cholesterol at the apical membrane of the STB is well understood, but the route by which cholesterol exits at the basal side for subsequent transfer across the fetal endothelial cells (FEC) or to other placental cell types remains not well characterized. Our aim was to provide evidence for basal secretion of apolipoprotein B-100 (apoB) containing lipoproteins. Furthermore, we investigated the placental localization of apolipoprotein receptors (LRP2, LDLR and LRP1) to identify cell targets of lipoprotein particles secreted in a polarized fashion by the STB. In trophoblast-derived BeWo cells grown on permeable filter supports, we demonstrate by immunoprecipitation apical as well as basolateral apoB secretion, which was significantly upregulated by estrogen-treatment for 24 or 48 h. Furthermore, we showed by immunofluorescence microscopy apoB and microsomal triglyceride transfer protein subunits localization in the STB and placental stromal cells in situ. All investigated receptors were detected by RT-qPCR and western blot in BeWo cells, but only expression of LRP2 was estrogen-inducible. In situ, the multi-ligand receptor LRP2 was expressed exclusively in the cytotrophoblast (CTB), the STB precursor cell type. LDLR and LRP1 localized to trophoblasts as well as stromal cells in situ. In summary, basal apoB secretion by BeWo cells supports the concept of basal lipoprotein particle secretion by placental STB. These lipoprotein particles may serve as cholesterol source for STB precursor cells, the CTBs, as well as all stromal cells of the chorionic villi including FECs, which were herein demonstrated to express apoB receptors, LRP2 and LDLR, respectively.

2 CIRCULATING microRNA SIGNATURES ASSOCIATED WITH EARLY BOVINE PREGNANCY

High-producing dairy cattle are characterised by low fertility, which extends calving intervals and reduces annual milk production. Early pregnancy diagnosis before Day 21 of gestation can shorten calving intervals, improve milk production, and increase overall profits from modern dairy herds. At present, accurate diagnosis is only possible after the third week of pregnancy. Circulating microRNA (miRNA) have been proposed as diagnostic biomarkers of numerous human conditions such as cancer and diabetes. Moreover, distinct circulating miRNA profiles have been associated with mid to late stages of human pregnancy. The aim of this study was to determine whether specific circulating miRNA signatures that occur during early pregnancy in cattle could be used for diagnostic purposes. To do this, we have compared miRNA levels in plasma samples from cycling, nonpregnant (n = 24) and cycling, early pregnant (D16, n = 11; D24, n = 11) Holstein heifers using next-generation sequencing and PCR arrays. Using small-RNA sequencing on 46 individual samples, we generated 2.8 million miRNA reads, and up to 328 miRNAs were detected in each sample. There were no significant differences (false discovery rate <0.1) in miRNA levels between nonpregnant and pregnant (D16, D24) heifers. In addition, 12 pooled samples (n = 9 for nonpregnant, n = 3 for D24) were analysed for the expression of 176 miRNA using PCR arrays; 16 miRNA had differentially expressed levels (2- to 3-fold; false discovery rate <0.1) in pregnant heifers. The QPCR validation of miRNA candidates obtained from the high-throughput analyses identified 6 miRNA that were differentially expressed in pregnant heifers. These included bta-miR-26a, bta-miR-29c, bta-miR-138, and bta-miR-204, which were differentially expressed at D16, bta-miR-1249, which was differentially expressed at D24, and hsa-miR-4532, which was differentially expressed at both D16 and D24 of pregnancy (P < 0.05, fold-change ≤2.4). The miRNA miR-26a and miR-204 have been previously associated with early (D20) and late pregnancy in pig and human respectively. Furthermore, miR-1249 has been associated with the regulation of ovarian function, and miR-26a is involved in immune regulation, suggesting a potential role for this miRNA as an immuno-suppressor during early pregnancy. In summary, we identified a miRNA signature associated with early pregnancy in cattle. We are currently using next-generation sequencing to identify differentially expressed miRNA at other stages of pregnancy, including Day-60 heifers, and this data will also be presented.This study was funded by Zoetis and BBSRC.

103 CIRCULATING microRNAs FOR EARLY DIAGNOSIS OF BOVINE PREGNANCY

Early diagnosis of pregnancy can shorten calving intervals, improve annual milk production and increase overall profits from modern dairy herds. At present, accurate diagnosis is only possible after the third week of pregnancy. Circulating microRNAs (miRNAs) have been proposed as diagnostic biomarkers for numerous human conditions such as cancer and diabetes. Moreover, distinct circulating miRNA profiles have been associated with different stages of human pregnancy. The objective of this study was to determine whether differential miRNA profiles occur in circulation during early pregnancy (Day 24 or earlier) in cattle that could be used for diagnostic purposes. Holstein-cross heifers were oestrous-synchronised and artificially inseminated (AI, n = 11) or sham-inseminated (control, n = 8) at first detected oestrus. Plasma samples were collected on Days 0, 8, 16 and 24 after insemination. Circulating miRNA levels were independently determined in pooled plasma samples (n = 3 pools for each of pregnant Day 24 and nonpregnant Days 0, 8, and 16) using Qiagen qPCR arrays (Qiagen, Valencia, CA, USA) and in individual samples (n = 11 samples for each pregnant Days 16 and 24, and 8 samples for each of nonpregnant Days 0, 8, and 16, respectively) using Illumina miRNA sequencing. The qPCR array data were analysed using the ΔΔCq method. The miRNA sequencing data were normalised using EdgeR. Differential expression between pregnant and nonpregnant groups was determined using 2-sample t-tests with false discovery rate (FDR) adjustment. Differences in miRNA expression were validated by RT-qPCR. Out of a total of 191 miRNAs analysed in pooled samples using qPCR arrays, 8 were differentially expressed (<3-fold, FDR <0.1) in Day 24 pregnant heifers relative to nonpregnant heifers (Days 0, 8, and 16 combined). No miRNAs were differentially expressed (FDR >0.1) between nonpregnant time-points. Changes in levels of 11 miRNAs were validated by RT-qPCR in individual plasma samples; although expression trends for these miRNAs were the same as in pooled samples, none of the changes in individual samples were significant after FDR adjustment (P > 0.1). Deep sequencing (96 million miRNA reads) identified 231 miRNAs in bovine plasma. There were no significant differences (FDR >0.1) in the expression of any miRNAs between pregnant heifers (Days 16 or 24) and nonpregnant (Days 0, 8, and 16 individually or combined). In addition, no significant differences were identified among nonpregnant time-points. In summary, we successfully performed miRNA profiling of bovine plasma using both deep sequencing and qPCR; however, we did not detect differences in miRNA expression between early pregnant (Day 16 or 24) and nonpregnant heifers. Changes in circulating miRNA levels may involve low abundance miRNAs that cannot be accurately quantified using current technology. Alternatively, changes in circulating miRNA levels may only occur later during pregnancy in cattle.

The expression of fucose isoforms of amniotic and plasma alpha-1-acid glycoprotein derived from 2nd and 3rd trimester normal pregnancies

Objectives To analyse modifications in AGP fucosylation in relation to different stages of human pregnancy. Design and methods The relative amounts of three fucosyl-glycotopes on AGP were analysed by lectin-ELISA using fucose-specific biotinylated lectins in 169 plasma and 178 amniotic fluid samples from normal pregnancies with gestational ages of 14 to 42 weeks. Results The plasma AGPs of all the pregnant women and amniotic AGPs from the 2nd trimester lacked fucoses. In contrast, in the 3rd trimester the amniotic AGPs were highly decorated by the innermost α1,6-fucose as well as α1,2- and α1,3-fucoses of the outer arms, reaching the highest expression around the perinatal period. At delivery the relative amounts of the α1,3- and α1,2-AGP isoforms, but not the α1,6 isoform, significantly decreased. Conclusions The highly fucosylated amniotic AGP isoforms could be implicated in regulatory processes to ensure homeostasis during pregnancy and to protect the fetus. They have the potential of becoming laboratory markers in obstetrics to monitor pregnancy.

Labour pains: giving birth to new mechanisms for the regulation of myometrial contractility

Labour pains: giving birth to new mechanisms for the regulation of myometrial contractility

Oxidative stress and antioxidants: exposure and impact on female fertility

Reproductive failure is a significant public health concern. Although relatively little is known about factors affecting fertility and early pregnancy loss, a growing body of literature suggests that environmental and lifestyle factors play an important role. There is sufficient evidence to hypothesize that diet, particularly its constituent antioxidants, and oxidative stress (OS) may influence the timing and maintenance of a viable pregnancy. We hypothesize that conditions leading to OS in the female affect time-to-pregnancy and early pregnancy loss. We review the epidemiology of female infertility related to antioxidant defenses and oxidation and examine potential sources of OS from the ovarian germ cell through the stages of human pregnancy and pregnancy complications related to infertility. Articles were identified through a search of the PubMed database. Female OS is a likely mediator of conception and threshold levels for OS exist, dependent on anatomic location and stage of preconception. Prospective pregnancy studies with dietary assessment and collection of biological samples prior to conception with endpoints of time-to-pregnancy and early pregnancy loss are needed.

Alterations of N-glycan branching and expression of sialic acid on amniotic fluid alpha-1-acid glycoprotein derived from second and third trimesters of normal and prolonged pregnancies

Background Alterations in the AGP glycoform pattern are not only disease related, but also can occur during physiological processes such as pregnancy. In this paper, possible changes in human amniotic fluid AGP glycan branching and in the type of sialic acid glycosidic attachment to glycans were analyzed with regard to different stages of human pregnancy. Methods Crossed-affinity immunoelectrophoresis with concanavalin A was used to study AGP branching and lectin–ELISA with two agglutinins from Maackia amurensis and Sambucus nigra was applied to differentiate α2,3 and α2,6 type sialic acid attachments. Results Despite almost unchanged levels of total amniotic fluid AGP during pregnancy, alterations in N-glycan branching and in the expression of sialic acid linkage on AGP were found to be associated with different stages of normal pregnancy. Amniotic fluid AGP glycans derived from third trimester compared with those from the second trimester had a higher percentage of tri- and tetra-antennary sialylated N-glycans. In the second trimester, sialic acid α2,6 linkage occurred twice as frequently as α2,3 linkage, while during the third trimester α2,3 linkage increased and both types of linkage appeared in equal proportion. Conclusions Branched and α2,3-sialylated AGP glycoforms in amniotic fluid could contribute to natural innate fetomaternal defense.

The relationship between decidualization and prolactin mRNA and production at different stages of human pregnancy

Within the human utero-placental unit only decidualized stromal cells express mRNA for prolactin. However, it is not clear if the level of prolactin production is related to the number of decidualized cells or the capacity of individual decidual cells to synthesize prolactin, either or both of which parameters may change during pregnancy. In the present study, prolactin production at different stages of human pregnancy was examined using quantitative in situ hybridization to assess decidual prolactin mRNA abundance, immunocytochemistry to examine the prolactin content inside decidual cells and RIA to measure decidual prolactin output into amniotic fluid. Throughout pregnancy the proportion of stromal cells showing positive immunostaining and mRNA for prolactin increased. There was a parallel increase in decidual cell size which was correlated with an increase in prolactin gene expression and intensity of immunostaining for prolactin in individual decidual cells. These changes in decidual cells were consistent with the changes in the concentration of prolactin in amniotic fluid. These results suggest that there is a close link between the level of prolactin gene expression and production of prolactin by individual decidual cells, which in turn is directly related to the process of decidualization that continues throughout human pregnancy.

Comparison Between Four Radioimmunoassays for Plasma Estriol

Abstract We have compared the results obtained for plasma estriol (E3) when four radioimmunoassay methods were used to measure this steroid during various stages of human pregnancy. The first method used a nonspecific antiserum combined with a chromatographic step. The second method utilizes as binding reagent a specific antibody against estriol combined with the same chromatographic step. The last two methods involve solvent extraction only, combined with the specific antiserum. Dichloromethane and ether respectively are used as solvent. The lowest levels of E3 were obtained with methods I and II. With dichloromethane extraction the E3 levels were comparable to those obtained with methods I and II. When using ether extraction the E3 levels were in most cases two to four times higher. Even with highly specific anti- E3 sera, chromatography is still required to achieve specificity.