Synthesis of prostaglandins by pig blastocysts cultured in medium containing estradiol or catechol estrogen

Two experiments were conducted to determine the effects of 2-hydroxy-estradiol-17β (2---OH---E₂; 0, 50 and 100 μM) and estradiol-17β (E₂; 0, 25 and 50 μM) on prostaglandin (PG) E and PGF₂α synthesis by day-10 pig blastocysts (day 0 is first day of estrus). Blastocysts were incubated in a modified Krebs-Ringer bicarbonate medium, supplemented with bovine serum albumin (4 mg/ml) and the vitamins and amino acids (essential and nonessential) in Minimum Essential Medium (without phenol red or antibiotics). The incubations were conducted at 39°C for three 2-h periods; the second and third periods included an E₂ or catechol estrogen treatment. Release of PGF₂α into the culture medium decreased (p<0.001) linearly with increasing concentrations of 2---H---E₂ in both periods. Release of PGE was not affected by 2---OH---E₂, therefore 2---OH---E₂ increased (p<0.06) the PGE:PGF₂α. When E₂ was added to the medium, release of PGE was decreased (p<0.01) during the second and third periods. Release of PGF₂α also was decreased (p<0.05) by E₂ during period 2, but E₂ did not alter the PGE:PGF₂α. Content of PGs in blastocysts at recovery was less than 10% of the PGs released in vitro. Therefore, these studies demonstrate effects of both the primary and catechol forms of E₂ on the synthesis of PGE and PGF₂α. Catechol estrogens and E₂ may inhibit PG synthesis and modify the PGE:PGF₂α during the establishment of pregnancy in pigs.     

Free Glycogen in Vaginal Fluids Is Associated with Lactobacillus Colonization and Low Vaginal pH

Objective

Lactobacillus dominates the lower genital tract microbiota of many women, producing a low vaginal pH, and is important for healthy pregnancy outcomes and protection against several sexually transmitted pathogens. Yet, factors that promote Lactobacillus remain poorly understood. We hypothesized that the amount of free glycogen in the lumen of the lower genital tract is an important determinant of Lactobacillus colonization and a low vaginal pH.

Methods

Free glycogen in lavage samples was quantified. Pyrosequencing of the 16S rRNA gene was used to identify microbiota from 21 African American women collected over 8–11 years.

Results

Free glycogen levels varied greatly between women and even in the same woman. Samples with the highest free glycogen had a corresponding median genital pH that was significantly lower (pH 4.4) than those with low glycogen (pH 5.8; p<0.001). The fraction of the microbiota consisting of Lactobacillus was highest in samples with high glycogen versus those with low glycogen (median = 0.97 vs. 0.05, p<0.001). In multivariable analysis, having 1 vs. 0 male sexual partner in the past 6 months was negatively associated, while BMI ≥30 was positively associated with glycogen. High concentrations of glycogen corresponded to higher levels of L. crispatus and L. jensenii, but not L. iners.

Conclusion

These findings show that free glycogen in genital fluid is associated with a genital microbiota dominated by Lactobacillus, suggesting glycogen is important for maintaining genital health. Treatments aimed at increasing genital free glycogen might impact Lactobacillus colonization.     

Putative therapeutic impacts of cardiac CTRP9 in ischaemia/reperfusion injury

Recently, cytokines belonging to C1q/tumour necrosis factor‐related proteins (CTRPs) superfamily have attracted increasing attention due to multiple metabolic functions and desirable anti‐inflammatory effects. These various molecular effectors exhibit key roles upon the onset of cardiovascular diseases, making them novel adipo/cardiokines. This review article aimed to highlight recent findings correlated with therapeutic effects and additional mechanisms specific to the CTRP9, particularly in cardiac ischaemia/reperfusion injury (IRI). Besides, the network of the CTPR9 signalling pathway and its possible relationship with IRI were discussed. Together, the discovery of all involved underlying mechanisms could shed light to alleviate the pathological sequelae after the occurrence of IRI.     

Global gene expression analysis to identify molecular markers of uterine receptivity and embryo implantation

Infertility and spontaneous pregnancy losses are an enduring problem to women's health. The establishment of pregnancy depends on successful implantation, where a complex series of interactions occurs between the heterogeneous cell types of the uterus and blastocyst. Although a number of genes are implicated in embryo-uterine interactions during implantation, genetic evidence suggests that only a small number of them are critical to this process. To obtain a global view and identify novel pathways of implantation, we used a dual screening strategy to analyze the expression of nearly 10,000 mouse genes by microarray analysis. Comparison of implantation and interimplantation sites by a conservative statistical approach revealed 36 up-regulated genes and 27 down-regulated genes at the implantation site. We also compared the uterine gene expression profile of progesterone-treated, delayed implanting mice to that of mice in which delayed implantation was terminated by estrogen. The results show up-regulation of 128 genes and down-regulation of 101 genes after termination of the delayed implantation. A combined analysis of these experiments showed specific up-regulation of 27 genes both at the implantation site and during uterine activation, representing a broad diversity of molecular functions. In contrast, the majority of genes that were decreased in the combined analysis were related to host immunity or the immune response, suggesting the importance of these genes in regulating the uterine environment for the implanting blastocyst. Collectively, we identified genes with recognized roles in implantation, genes with potential roles in this process, and genes whose functions have yet to be defined in this event. The identification of unique genetic markers for the onset of implantation signifies that genome-wide analysis coupled with functional assays is a promising approach to resolve the molecular pathways required for successful implantation.     

Ontogenetic changes in spot configuration (numbers,circularity, size and asymmetry) and lateral line in Neurergus microspilotus (Caudata: Salamandridae)

Coloration in three of four species of the genus Neurergus including N. microspilotus is characterized by the presence of yellow spots on a dark skin, but there is no available information about changes in spot configuration, speed of development and degree of association between melanophore‐free region and the lateral line. In this study, spot numbers, spot circularity, spot size and spot asymmetry were studied during larval to adult growth in N. microspilotus during July 2012 to June 2015. The mean numbers of spots increased during the late larval stage till postmetamorphic period from 13.33 ± 3.77 to 22.53 ± 4.09 and reached 42.62 ± 4.06 in adults. At the same time, the extent of spots gradually decreased in size from 5.80 ± 1.00 to 3.57 ± 0.97 mm² and reached 3.55 ± 1.42 mm² in adults, but the spot circularity increased from 0.48 ± 0.23 to 0.78 ± 0.49 and reached 0.80 ± 0.15 in adults. In adults, the numbers, circularity, size and asymmetry of spots remain stable with little but non‐significant changes during the study period. Histological study shows that formation of a melanophore‐free region correlates with the development of the lateral line receptors. This study demonstrates that the effects of lateral line on chromatophores persist through middle larval stages but diminish as metamorphosis completes.     

Reversal of indomethacin-induced inhibition of implantation in the mouse by epidermal growth factor.

In the mouse, estriol (E3) can induce implantation as a phase I of estrogen action. E3-induced implantation was attenuated by indomethacin(INDO), an inhibitor of prostaglandin (PG) synthesis. The inhibitory effect of INDO was reversed by administration of epidermal growth factor (EGF), and this EGF effect was dose-dependent. These results suggest that one of the functions of estrogen could be to activate the EGF ligand-receptor signalling in the uterus in generating PGs required for initiation of implantation. This is consistent with the results of EGF stimulation of synthesis of PGs in uterine stromal cells in culture.     

Epidermal growth factor-specific protein tyrosine phosphorylation in preimplantation embryo development.

We examined whether epidermal growth factor (EGF)-induced preimplantation mouse embryo development and function are mediated by EGF-specific protein tyrosine phosphorylation (PTP). In situ cross-linking and autophosphorylation studies showed that EGF receptor (EGF-R) in Day 4 mouse blastocysts is a protein of approximately 170 kDa that is phosphorylated when exposed to EGF and ATP. Furthermore, EGF induced about a twofold increase in protein tyrosine kinase (PTK) activity in Day 4 blastocysts when incubated in the presence of a peptide substrate with a tyrosine moiety and ATP. RG 50864, a specific inhibitor of EGF-dependent PTK, diminished autophosphorylation of the 170-kDa protein and completely blocked PTK activity in the blastocyst induced by EGF. However, this inhibitor did not affect EGF binding to the embryonic cell surface. In contrast, an inactive tyrphostin compound, RG 50862, did not alter EGF-induced PTK activity in the blastocyst. These findings led us to examine the effects of these tyrphostin compounds on preimplantation mouse embryo development and blastocyst hatching in vitro. RG 50864, in a dose-dependent manner, inhibited EGF-dependent development of 2-cell embryos to blastocysts and the number of cells per blastocyst. This inhibitor also antagonized EGF-induced zona-hatching of blastocysts formed from 8-cell embryos in culture. However, the inhibitor was not effective in deterring transforming growth factor-beta 1-induced blastocyst formation. The inactive compound, RG 50862, had no effects on EGF-dependent blastocyst formation or zona-hatching. The data show that the effects of RG 50864 are specific and mediated by inhibition of EGF-specific PTK activity.(ABSTRACT TRUNCATED AT 250 WORDS)     

Alpha5beta1, alphaVbeta3 and the platelet-associated integrin alphaIIbbeta3 coordinately regulate adhesion and migration of differentiating mouse trophoblast cells

During blastocyst implantation, interaction between integrins on the apical surface of the trophoblast and extracellular matrix (ECM) in the endometrium anchors the embryo to the uterine wall. Strong adhesion of the blastocyst to fibronectin (FN) requires integrin signaling initiated by exogenous fibronectin. However, it is not known how integrin signaling enhances blastocyst adhesion. We present new evidence that the integrin, alphaIIbbeta3, plays a key role in trophoblast adhesion to fibronectin during mouse peri-implantation development. Trafficking of alphaIIb to the apical surface of the trophoblast increased dramatically after blastocysts were exposed to fibronectin, whereas other fibronectin-binding integrins, alpha5beta1 and alphaVbeta3, were resident at the apical surface before ligand exposure. Functional comparisons among the three integrins revealed that ligation of alpha5beta1 most efficiently strengthened blastocyst fibronectin-binding activity, while subsequent trophoblast cell migration was dependent primarily on the beta3-class integrins. In vivo, alphaIIb was highly expressed by invasive trophoblast cells in the ectoplacental cone and trophoblast giant cells of the parietal yolk sac. These data demonstrate that trafficking of alphaIIb regulates adhesion between trophoblast cells and fibronectin as invasion of the endometrium commences.