Expression and regulation of vascular endothelial growth factor ligands and receptors during menstruation and post-menstrual repair of human endometrium

Regeneration and growth of the human endometrium after shedding of the functional layer during menstruation depends on an adequate angiogenic response. We analysed the mRNA expression levels of all known vascular endothelial growth factor (VEGF) ligands and receptors in human endometrium collected in the menstrual and proliferative phases of the menstrual cycle. In addition, we evaluated the expression of VEGF-A, VEGF-R2 and NRP-1 at the protein level. Two periods of elevated mRNA expression of ligands and receptors were observed, separated by a distinct drop at cycle days (CDs) 9 and 10. Immunohistochemical staining showed that VEGF and VEGF-R2 were expressed in epithelial, stromal and endothelial cells. NRP-1 was mainly confined to stroma and blood vessels; only in late-proliferative endometrium, epithelial staining was also observed. Except for endothelial VEGF-R2 expression in CDs 6-8, there were no significant differences in the expression of VEGF, VEGF-R2 or NRP-1 in any of the cell compartments. In contrast, VEGF release by cultured human endometrium explants decreased during the proliferative phase. This output was significantly reduced in menstrual and early-proliferative endometrium by estradiol (E2) treatment. Western blot analysis indicated that part of the VEGF-A was trapped in the extracellular matrix (ECM). Changes in VEGF ligands and receptors were associated with elevated expression of the hypoxia markers HIF1 alpha and CA-IX in the menstrual and early proliferative phases. HIF1 alpha was also detected in late-proliferative phase endometrium. Our findings indicate that VEGF-A exerts its actions mostly during the first half of the proliferative phase. Furthermore, VEGF-A production appears to be triggered by hypoxia in the menstrual phase and subsequently suppressed toy estrogen during the late proliferative phase.

Menstrual effluent induces epithelial-mesenchymal transitions in mesothelial cells

Background Menstrual effluent affects mesothelial cell (MC) morphology. We evaluated whether these changes were consistent with epithelial-mesenchymal transitions (EMT). Methods Monolayer cultures of MC were incubated overnight in conditioned media, prepared from cells isolated form menstrual effluent, with or without kinase and ATP inhibitors. Changes in cell morphology were monitored using time-lapse video microscopy and immunohistochemistry. Effects on the expression of EMT-associated molecules were evaluated using real-time RT-PCR and/or Western blot analysis. Results Incubation in conditioned media disrupted cell-cell contacts, and increased MC motility. The changes were reversible. During the changes the distribution of cytokeratins, fibrillar actin and α-tubulin changed. Sodium azide, an inhibitor of ATP production, and Genistein, a general tyrosine kinase inhibitor, antagonized these effects. Wortmannin, a phosphatidylinositol 3-kinase inhibitor, and SU6656, an Src tyrosine kinase inhibitor, only partially antagonized the effect. The expression of Snail and vimentin was markedly up-regulated, whereas the expression of E-cadherin was decreased and cytokeratins were altered. Conclusions In MC, menstrual effluent initiates a reversible, energy-dependent transition process from an epithelial to a mesenchymal phenotype. Involvement of the (Src) tyrosine kinase signalling pathway and the changes in the expression of cytokeratins, Snail, vimentin and E-cadherin demonstrate that the morphological changes are EMT.

Oestrogen-modulated gene expression in the human endometrium

To identify key regulatory mechanisms in the growth and development of the human endometrium, microarray analysis was performed on uncultured human endometrium collected during menstruation (M) and the late-proliferative (LATE-P)-phase of the menstrual cycle, as well as after 24 h incubation in the presence of oestradiol (17beta-E2). We demonstrate the expression of novel gene transcripts in the human endometrium. i.e. mucin-9, novel oestrogen-responsive gene transcripts, i.e. gelsolin and flotillin-1, and genes known to be expressed in human endometrium but not yet shown to be oestrogen responsive, i.e. connexin-37 and TFF1/pS2. Genes reported to be expressed during the implantation window and implicated in progesterone action, i.e. secretoglobin family 2A, member 2 (mammaglobin) and homeobox-containing proteins, were up-regulated in uncultured LATE-P-phase endometrium compared to M-phase endometrium. Some gene transcripts are regulated directly by 17beta-E2 alone, others are influenced by the in vivo environment as well. These observations emphasise that the regulation of endometrium maturation by oestrogen entails more then just stimulation of cell proliferation.

The effect of ongoing blood loss on human serum concentrations of perfluorinated acids

Perfluorinated alkyl acids (PFAAs) have been detected in serum at low concentrations in background populations. Higher concentrations haven been observed in adult males compared to females, with a possible explanation that menstruation offers females an additional elimination route. In this study, we examined the significance of blood loss as an elimination route of PFAAs. Pooled serum samples were collected from individuals undergoing a medical procedure involving ongoing blood withdrawal called venesection. Concentrations from male venesection patients were approximately 40% lower than males in the general population for perfluorohexane sulfonate (PFHxS), perfluorooctane sulfonate (PFOS) and perfluorooctanoic acid (PFOA). A simple pharmacokinetic model was used to test the hypothesis that blood loss could explain why adult males have higher concentrations of PFAAs than females, and why males undergoing venesections had lower concentrations compared to males in the general population. The model application generally supported these hypotheses showing that venesection might reduce blood serum concentrations by 37% (PFOA) and 53% (PFOS) compared to the observed difference of 44% and 37%. Menstruation was modeled to show a 22% reduction in PFOA serum concentrations compared to a 24% difference in concentrations between males and females in the background population. Uncertainties in the modeling and the data are identified and discussed.