Impact of DNA methylation on trophoblast function

The influence of epigenetics is evident in many fields of medicine today. This is also true in placentology, where versatile epigenetic mechanisms that regulate expression of genes have shown to have important influence on trophoblast implantation and placentation. Such gene regulation can be established in different ways and on different molecular levels, the most common being the DNA methylation. DNA methylation has been shown today as an important predictive component in assessing clinical prognosis of certain malignant tumors; in addition, it opens up new possibilities for non-invasive prenatal diagnosis utilizing cell-free fetal DNA methods. By using a well known demethylating agent 5-azacytidine in pregnant rat model, we have been able to change gene expression and, consequently, the processes of trophoblast differentiation and placental development. In this review, we describe how changes in gene methylation effect trophoblast development and placentation and offer our perspective on use of trophoblast epigenetic research for better understanding of not only placenta development but cancer cell growth and invasion as well

Modification of conservative treatment of heterotopic cervical pregnancy by Foley catheter balloon fixation with cerclage sutures at the level of the external cervical os: a case report

<p>Abstract</p> <p>Introduction</p> <p>Conservative treatment of a heterotopic cervical pregnancy was performed with a modification of the fixation of a Foley catheter at the level of the external cervical os, followed by the ligature of the descending cervical branches of the uterine arteries and systemic methotrexate application.</p> <p>Case presentation</p> <p>A 34-year-old Caucasian woman was diagnosed with double gestation after 6 weeks of <it>in vitro </it>fertilization treatment. A gynecological examination and color Doppler ultrasound scan revealed intra-uterine and cervical gestational sacs both containing live fetuses. A Foley catheter balloon was inserted into the cervical canal, inflated and fixed by a cerclage suture at the level of the external cervical os, followed by ligation of the descending cervical branches of the uterine arteries. Systemic methotrexate was applied. Three days after removal of the Foley catheter, an evacuation of the intra-uterine gestational sac was performed. Hemorrhage from the implantation site was controlled immediately and a pregnancy termination was successfully performed. The procedure was uneventful and our patient was discharged with a preserved uterus.</p> <p>Conclusions</p> <p>Conservative treatment of cervical pregnancy using a Foley catheter balloon is more efficacious if the Foley catheter balloon is attached in the correct position with a cerclage suture at the level of the external os, followed by ligation of the descending cervical branches of the uterine arteries, thereby exerting maximal pressure on the bleeding vessels.</p

Discovery of DNA methylation markers in cervical cancer using relaxation ranking

<p>Abstract</p> <p>Background</p> <p>To discover cancer specific DNA methylation markers, large-scale screening methods are widely used. The pharmacological unmasking expression microarray approach is an elegant method to enrich for genes that are silenced and re-expressed during functional reversal of DNA methylation upon treatment with demethylation agents. However, such experiments are performed in <it>in vitro </it>(cancer) cell lines, mostly with poor relevance when extrapolating to primary cancers. To overcome this problem, we incorporated data from primary cancer samples in the experimental design. A strategy to combine and rank data from these different data sources is essential to minimize the experimental work in the validation steps.</p> <p>Aim</p> <p>To apply a new relaxation ranking algorithm to enrich DNA methylation markers in cervical cancer.</p> <p>Results</p> <p>The application of a new sorting methodology allowed us to sort high-throughput microarray data from both cervical cancer cell lines and primary cervical cancer samples. The performance of the sorting was analyzed <it>in silico</it>. Pathway and gene ontology analysis was performed on the top-selection and gives a strong indication that the ranking methodology is able to enrich towards genes that might be methylated. Terms like regulation of progression through cell cycle, positive regulation of programmed cell death as well as organ development and embryonic development are overrepresented. Combined with the highly enriched number of imprinted and X-chromosome located genes, and increased prevalence of known methylation markers selected from cervical (the highest-ranking known gene is <it>CCNA1</it>) as well as from other cancer types, the use of the ranking algorithm seems to be powerful in enriching towards methylated genes.</p> <p>Verification of the DNA methylation state of the 10 highest-ranking genes revealed that 7/9 (78%) gene promoters showed DNA methylation in cervical carcinomas. Of these 7 genes, 3 (<it>SST</it>, <it>HTRA3 </it>and <it>NPTX1</it>) are not methylated in normal cervix tissue.</p> <p>Conclusion</p> <p>The application of this new relaxation ranking methodology allowed us to significantly enrich towards methylation genes in cancer. This enrichment is both shown <it>in silico </it>and by experimental validation, and revealed novel methylation markers as proof-of-concept that might be useful in early cancer detection in cervical scrapings.</p

Gene deletion of P-Selectin and ICAM-1 does not inhibit neutrophil infiltration into peritoneal cavity following cecal ligation-puncture

BACKGROUND: Neutrophil infiltration is one of the critical cellular components of an inflammatory response during peritonitis. The adhesion molecules, P-selectin and intercellular adhesion molecule (ICAM)-1, mediate neutrophil-endothelial cell interactions and the subsequent neutrophil transendothelial migration during the inflammatory response. Despite very strong preclinical data, recent clinical trials failed to show a protective effect of anti-adhesion therapy, suggesting that the length of injury might be a critical factor in neutrophil infiltration. Therefore, the objective of this study was to determine the role of P-selectin and ICAM-1 in neutrophil infiltration into the peritoneal cavity during early and late phases of peritonitis. METHODS: Peritonitis was induced in both male wild-type and P-selectin/ICAM-1 double deficient (P/I null) mice by cecal ligation-puncture (CLP). Peripheral blood and peritoneal lavage were collected at 6 and 24 hours after CLP. The total leukocyte and neutrophil contents were determined, and neutrophils were identified with the aid of in situ immunohistochemical staining. Comparisons between groups were made by applying ANOVA and student t-test analysis. RESULTS: CLP induced a severe inflammatory response associated with a significant leukopenia in both wild-type and P/I null mice. Additionally, CLP caused a significant neutrophil infiltration into the peritoneal cavity that was detected in both groups of mice. However, neutrophil infiltration in the P/I null mice at 6 hours of CLP was significantly lower than the corresponding wild-type mice, which reached a similar magnitude at 24 hours of CLP. In contrast, in peritonitis induced by intraperitoneal inoculation of 2% glycogen, no significant difference in neutrophil infiltration was observed between the P/I null and wild-type mice at 6 hours of peritonitis. CONCLUSIONS: The data suggest that alternative adhesion pathway(s) independent of P-selectin and ICAM-1 can participate in neutrophil migration during peritonitis and that the mode of stimuli and duration of the injury modulate the neutrophil infiltration

Performance of an endcap prototype of the ATLAS accordion electromagnetic calorimeter

The design and construction of a lead-liquid argon endcap calorimeter prototype using an accordion geometry and conceived as a sector of the inner wheel of the endcap calorimeter of the future ATLAS experiment at the LHC is described. The performance obtained using electron beam data is presented. The main results are an energy resolution with a sampling term below $11\%/\sqrt{E(\rm GeV)}$ and a small local constant term, a good linearity of the response with the incident energy and a global constant term of 0.8\% over an extended area in the rapidity range of $2.2 < \eta <2.9$. These properties make the design suitable for the ATLAS electromagnetic endcap calorimeter

Insulin-like growth factor axis in pregnancies affected by fetal growth disorders

Background: Insulin-like growth factors 1 and 2 (IGF1 and IGF2) and their binding proteins (IGFBPs) are expressed in the placenta and known to regulate fetal growth. DNA methylation is an epigenetic mechanism which involves addition of methyl group to a cytosine base in the DNA forming a methylated cytosine-phosphate-guanine (CpG) dinucleotide which is known to silence gene expression. This silences gene expression, potentially altering the expression of IGFs and their binding proteins. This study investigates the relationship between DNA methylation of components of the IGF axis in the placenta and disorders in fetal growth. Placental samples were obtained from cord insertions immediately after delivery from appropriate, small (defined as birthweight the 90th percentile for the gestation [LGA]) neonates. Placental DNA methylation, mRNA expression and protein levels of components of the IGF axis were determined by pyrosequencing, rtPCR and Western blotting. Results: In the placenta from small for gestational age (SGA) neonates (n = 16), mRNA and protein levels of IGF1 were lower and of IGFBPs (1, 2, 3, 4 and 7) were higher (p < 0.05) compared to appropriately grown neonates (n = 37). In contrast, in the placenta from large for gestational age (LGA) neonates (n = 20), mRNA and protein levels of IGF1 was not different and those of IGFBPs (1, 2, 3 and 4) were lower (p < 0.05) compared to appropriately grown neonates. Compared to appropriately grown neonates, CpG methylation of the promoter regions of IGF1 was higher in SGA neonates. The CpG methylation of the promoter regions of IGFBP1, IGFBP2, IGFBP3, IGFBP4 and IGFBP7 was lower in the placenta from SGA neonates as compared to appropriately grown neonates, but was unchanged in the placenta from LGA neonates. Conclusions: Our results suggest that changes in CpG methylation contribute to the changes in gene expression of components of the IGF axis in fetal growth disorders. Differential methylation of the IGF1 gene and its binding proteins is likely to play a role in the pathogenesis of SGA neonates

Impact of 5-Azacytidine on placental weight, glycoprotein pattern and proliferating cell nuclear antigen expression in rat placenta

During the placentation process, the expression of various glycoproteins plays an important role in embryonal development. Alterations in DNA methylation caused by 5-azacytidine (5azaC) can disturb normal glycoprotein expression as well as the proliferative ability of trophoblast cells. In order to assess this, a single dose of 5azaC was injected intraperitoneally into pregnant rats during days 1-19 of gestation. Animals were euthanised on day 20 and placental weight, as well as glycoprotein composition, was analysed together with immunohistological assessment of the degree of proliferation of the trophoblast cells. The placental weight was found to be significantly smaller in animals treated by 5azaC during days 4 to 14 of gestation (p<0.01, Student's t-test). The treatment on days 4, 5, and 6 resulted in a lack of labyrinth with the strong proliferative activity of the cells in the basal layer. Expression of glycoproteins with molecular mass smaller than 60kDa was reduced with treatment on day 6. The 5azaC administered from days 7 to 10 completely disturbed the placental structure and the proliferation of trophoblast cells was poor. During these days GP70 exhibited stronger expression in treated animals, contrary to GP40, which was stronger in controls. A natural border between the labyrinth and the basal layer was established on days 11 and 12. The basal layer was dominant with a lower proliferation of trophoblast cells compared with the controls. With the establishment of the labyrinth on day 13, the expression of GP40 was restored. Proliferation of the trophoblast cells from days 13 to 15 was higher compared with the controls. The changes in placental mass and the proliferative ability of trophoblast cells in rat placenta exposed to 5azaC represent more proof of the importance of epigenetics in the regulation of placental development

A frost regime microclimatological study in southern Argentina (Añelo, province of Neuquén)

Frosts constitute a main hazard for most of the Argentine olive growing area. As a component of a feasibility assessment, financed by the Argentine Federal Investment Council (CFI) to introduce olive growing in southern Argentina (Neuquén), a microclimatological study of the frost regime in 3760 ha was conducted. The field occupies a band, with moderate gradient, which runs between the limit of the Patagonian plateau and the Neuquén River that flows in the lower part of a narrow valley, thus allowing cold air to drain, mitigating frost intensity. To evaluate frost hazard, a meteorological automatic station with capacity to transmit data, and 30 registering temperature sensors were installed, making observations from March to November 2010. Those registers were correlated with data from two neighbor climate stations, reconstructing the local series for the 2001-2010 decade. Those reconstructed data were introduced in a model developed taking as a basis the NCEP (National Centers for Environmental Prediction of NOAA) Climate Forecast System Reanalysis a downscaling process was carried on, for a 2.5 by 2.5 m grid, covering the area in study to a high degree of resolution. Maps of mean value, beginning, end and duration of the annual periods with temperatures equal or less than 0, -1, -3, -7, -10 and -12°C, were elaborated. For all thermal levels, it was observed that the lower portions of the fields are prone to suffer longer lasting and more intense frosts, making evident the importance of two main microclimate factors: (a) upper portion of the area, higher wind speed, impedes air stratification avoiding the thermal inversion necessary for radiation frost development; and (b) lower portions of the field, cold air drainage ads to thermal loss increasing cooling. Those criteria allowed dividing the field in sub-areas with different levels of frost hazard.Fil: Vita Serman, F.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Mendoza - San Juan. Estación Experimental Agropecuaria San Juan; Argentina. Universidad Nacional de San Juan. Facultad de Ingeniería; ArgentinaFil: Babelis, G.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Mendoza - San Juan. Estación Experimental Agropecuaria San Juan; Argentina. Universidad Nacional de San Juan. Facultad de Ingeniería; ArgentinaFil: Gallego, M. E.. Consejo Federal de Inversiones; ArgentinaFil: Kiessling, J.. Instituto Nacional de Tecnología Agropecuaria. Centro Regional Patagonia Norte. Estación Experimental Agropecuaria Alto Valle; ArgentinaFil: Sierra, Eduardo Mario. Universidad de Buenos Aires. Facultad de Agronomia; Argentina. Consejo Nacional de Investigaciones Científicas y Técnicas; ArgentinaFil: López, E. Universidad de Buenos Aires. Facultad de Agronomia; Argentin