Sperm microRNA content is altered in a mouse model of male obesity, but the same suite of microRNAs are not altered in offspring's sperm

The prevalence of obesity is increasing worldwide and has tripled in men of reproductive age since the 1970s. Concerningly, obesity is not only comorbid with other chronic diseases, but there is mounting evidence that it increases the non-communicable disease load in their children (eg mortality, obesity, autism). Animal studies have demonstrated that paternal obesity increases the risk of metabolic (eg glucose metabolism defects, obesity) and reproductive disorders in offspring. Epigenetic changes within sperm are clear mechanistic candidates that are associated with both changes to the father’s environment and offspring phenotype. Specifically there is emerging evidence that a father’s sperm microRNA content both responds to paternal environmental cues and alters the gene expression profile and subsequent development of the early embryo. We used a mouse model of high fat diet (HFD) induced obesity to investigate whether male obesity could modulate sperm microRNA content. We also investigated whether this alteration to a father’s sperm microRNA content lead to a similar change in the sperm of male offspring. Our investigations were initially guided by a Taqman PCR array, which indicated the differential abundance of 28 sperm borne microRNAs in HFD mice. qPCR confirmation in a much larger cohort of founder males demonstrated that 13 of these microRNAs were differentially abundant (11 up-regulated; 2 down-regulated) due to HFD feeding. Despite metabolic and reproductive phenotypes also being observed in grand-offspring fathered via the male offspring lineage, there was no evidence that any of the 13 microRNAs were also dysregulated in male offspring sperm. This was presumably due to the variation seen within both groups of offspring and suggests other mechanisms might act between offspring and grand-offspring. Thus 13 sperm borne microRNAs are modulated by a father’s HFD and the presumed transfer of this altered microRNA payload to the embryo at fertilisation potentially acts to alter the embryonic molecular makeup post-fertilisation, altering its growth trajectory, ultimately affecting adult offspring phenotype and may contribute to paternal programming.Tod Fullston, E. Maria C. Ohlsson-Teague, Cristin G. Print, Lauren Y. Sandeman, Michelle Lan

Civic education in Spain: a critical review of policy

The international revival in civic education over the past decade did not have a pervading impact on the Spanish education system. Unlike efforts in Britain, many former communist countries and democracies in other parts of the world such as Australia, Spanish efforts to invigorate education for democratic citizenship have been modest. Nevertheless there has been a growing interest and an increasing realization that Spain is part of a new, more democratic Europe which needs civic education for its young people to enhance democratic citizenship. For example, the 1990 General Law for the Regulation of the Spanish Educational System (LOGSE) included Civic Education as part of the planned educational reform for Spain at the end of the Twentieth century. Our objective is to critically analyze the Civic and Moral Education program proposed by the Spanish Ministry of Education, in 1990, from its earliest formulations, including the modifications that have been incorporated until 2001. The legal texts and orders issued by the ministry (MEC: Ministerio Español de Educación y Ciencia) during the specified period were analyzed. Published bibliography presenting the theoretical basis of the proposal, as well as the references concerning its implementation, were also studied. We conclude that despite these efforts in educational reform, if Spain wants to see itself, educationally, as part of the new Europe, it will need to engage a more vigorous program in civic education for democratic citizenship. Currently young people in Spain are not well prepared for active, democratic citizenship in the new Europe and an increasingly globalized worl

The relationship between seminal leukocytes, oxidative status in the ejaculate, and apoptotic markers in human spermatozoa

The aim of this study was to investigate the relationship between seminal leukocytes, reactive oxygen species (ROS) production in the ejaculate, and markers of apoptosis in human spermatozoa. Semen samples were collected from 60 patients attending fertility clinics at the Reproductive Biology Unit at Tygerberg Academic Hospital and Vincent Pallotti Hospital, Cape Town, South Africa. The concentration of seminal leukocytes was determined and was correlated with ROS production in the ejaculate, the percentage of superoxide (·O2 )- and hydrogen peroxide (H2O2)-positive spermatozoa, glutathione activation in the ejaculate, and with markers of apoptosis in spermatozoa, namely cysteine-dependent aspartate-directed proteases (caspase)-3/7 activation, mitochondrial membrane potential (ΔΨm), and the percentage of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL)-positive sperm. Significant correlations with the concentration of seminal leukocytes were found for ROS production in the ejaculate, the percentage of ·O2 -positive spermatozoa, and caspase-3/7 activation in the ejaculate. Leukocytospermic samples showed significantly higher ROS production, percentage of ·O2 -positive sperm, GSH activation, and caspase-3/7 activation compared to non-leukocytospermic samples. The percentage of ·O2 -positive sperm was significantly correlated with sperm ΔΨm and caspase-3/7 activation in the ejaculate. Sperm ΔΨm and TUNEL-positive sperm did not correlate with seminal leukocyte concentration. Data demonstrate that high seminal leukocyte concentrations that leads to increased seminal ROS production, and is also associated with caspase activation in the male germ cell and increased mitochondrial ROS production. The latter could possibly be a result of disturbed ΔΨm. The activation of caspase-3/7 could then follow the increased intrinsic superoxide levels due to depleted intrinsic glutathione (GSH). These cellular events might not directly and immediately lead to DNA fragmentation as an endpoint of apoptosis because of topological hindrances.Web of Scienc

A Study of the Ceramicisation of Allylhydridopolycarbosilane by Thermal Volatilisation Analysis and Solid-State Nuclear Magnetic Resonance

AHPCS is a pre-ceramic polymer utilised as a precursor to SiC. An initial polymerisation to a cross-linked network is followed by a complex sequence of processes ultimately leading to amorphous SiC. Using thermal volatilisation analysis (TVA) accompanied with solid-state NMR (SSNMR), FTIR, MS, DSC and TGA the complete thermal profile was identified. Between 160 – 300 °C, AHPCS cross-links through the allyl group and undergoes some carbon-silicon rearrangement, with a volatilisation of low mass oligomeric material and significant volumes of hydrogen released from dehydrocoupling of SiH moieties. By 300 °C the allyl group is completely cross-linked but the polymer starts to undergo pyrolytic degradation of the network, with the release of chain fragments and low molar mass species such as methane, ethane, methanol, propane, propene and silane species. Hydrogen once again becomes the major volatile product above 400 °C due to higher proportion of dehydrocoupling forming Si–C and Si–Si bonds. Small chain fragments are seen in the form of larger alkyl silanes. These fragments come from the chain scission of the polymer at weaker parts of the network. The process of side group scission leads to further radical recombination reactions of silicon and carbon atoms to build the SiC network. By 500 °C higher proportion of dehydrocoupling occurs with recombination of Si–Si and Si–C species. The Si–H bonds in -SiH3 groups have completely cleaved along with C-H bonds in the CH3 and CH2 groups leaving SiC, -SiH and HCSi3 present in the material. This bond cleavage leads the silicon and carbon radical species to undergo radical recombination in the network with the volatile release being dominated by H2. By 650 °C the cleavage and recombination of remaining -SiH2-, -SiH- and HCSi3 groups ultimately form amorphous SiC. The volatiles released are mostly hydrogen with very few condensable products seen. Finally, SiC is then crystallised at higher temperatures forming β-SiC at 1100 °C and then subsequently α-SiC above 1500 °C

Zinc Finger Nuclease mediated knockout of ADP dependent Glucokinase in Cancer cell lines: Effects on cell survival and Mitochondrial Oxidative Metabolism

<div><p>Zinc finger nucleases (ZFN) are powerful tools for editing genes in cells. Here we use ZFNs to interrogate the biological function of <i>ADPGK</i>, which encodes an ADP-dependent glucokinase (ADPGK), in human tumour cell lines. The hypothesis we tested is that ADPGK utilises ADP to phosphorylate glucose under conditions where ATP becomes limiting, such as hypoxia. We characterised two ZFN knockout clones in each of two lines (H460 and HCT116). All four clones had frameshift mutations in all alleles at the target site in exon 1 of <i>ADPGK,</i> and were ADPGK-null by immunoblotting. <i>ADPGK</i> knockout had little or no effect on cell proliferation, but compromised the ability of H460 cells to survive siRNA silencing of hexokinase-2 under oxic conditions, with clonogenic survival falling from 21±3% for the parental line to 6.4±0.8% (p = 0.002) and 4.3±0.8% (p = 0.001) for the two knockouts. A similar increased sensitivity to clonogenic cell killing was observed under anoxia. No such changes were found when <i>ADPGK</i> was knocked out in HCT116 cells, for which the parental line was less sensitive than H460 to anoxia and to hexokinase-2 silencing. While knockout of <i>ADPGK</i> in HCT116 cells caused few changes in global gene expression, knockout of <i>ADPGK</i> in H460 cells caused notable up-regulation of mRNAs encoding cell adhesion proteins. Surprisingly, we could discern no consistent effect on glycolysis as measured by glucose consumption or lactate formation under anoxia, or extracellular acidification rate (Seahorse XF analyser) under oxic conditions in a variety of media. However, oxygen consumption rates were generally lower in the <i>ADPGK</i> knockouts, in some cases markedly so. Collectively, the results demonstrate that <i>ADPGK</i> can contribute to tumour cell survival under conditions of high glycolytic dependence, but the phenotype resulting from knockout of <i>ADPGK</i> is cell line dependent and appears to be unrelated to priming of glycolysis in these lines.</p></div

The Development of Inclusive Learning Relationships in Mainstream Settings: A Multimodal Perspective

The debate regarding the inclusion of children with Special Educational Needs and Disabilities (SEND) in mainstream education in the UK partly revolves around what makes the classroom environment inclusive. Through the potential offered by the specific qualitative methodologies employed, this study aimed to explore the development of teachers’ pedagogical practices and learning relationships upon the inclusive education of children with special educational needs and disabilities in two primary school classes. The study considered the views and behaviours of primary school pupils with and without special educational needs, primary school teachers and teaching assistants (TAs) in one mainstream school. Drawing on a multimodal approach to discourse analysis to account for the complex relationships between symbolic and non-verbal modes of classroom signification, the study explored how meaning is produced in classrooms and children’s modes of communication, as well as in teachers’ practices. The two classes are compared on the basis of teaching observations, interviews, transcripts of dialogues, and analyses of classroom organisation and decoration. This paper suggests that the greatest influence on the educational and social outcomes of students with special educational needs is the behaviour and practices of the classroom teacher

A Gene Expression Signature of Invasive Potential in Metastatic Melanoma Cells

BACKGROUND: We are investigating the molecular basis of melanoma by defining genomic characteristics that correlate with tumour phenotype in a novel panel of metastatic melanoma cell lines. The aim of this study is to identify new prognostic markers and therapeutic targets that might aid clinical cancer diagnosis and management. PRINCIPAL FINDINGS: Global transcript profiling identified a signature featuring decreased expression of developmental and lineage specification genes including MITF, EDNRB, DCT, and TYR, and increased expression of genes involved in interaction with the extracellular environment, such as PLAUR, VCAN, and HIF1a. Migration assays showed that the gene signature correlated with the invasive potential of the cell lines, and external validation by using publicly available data indicated that tumours with the invasive gene signature were less melanocytic and may be more aggressive. The invasion signature could be detected in both primary and metastatic tumours suggesting that gene expression conferring increased invasive potential in melanoma may occur independently of tumour stage. CONCLUSIONS: Our data supports the hypothesis that differential developmental gene expression may drive invasive potential in metastatic melanoma, and that melanoma heterogeneity may be explained by the differing capacity of melanoma cells to both withstand decreased expression of lineage specification genes and to respond to the tumour microenvironment. The invasion signature may provide new possibilities for predicting which primary tumours are more likely to metastasize, and which metastatic tumours might show a more aggressive clinical course

PDX1 DNA methylation distinguishes two subtypes of pancreatic neuroendocrine neoplasms with a different prognosis

DNA methylation is a crucial epigenetic mechanism for gene expression regulation and cell differentiation. Furthermore, it was found to play a major role in multiple pathological processes, including cancer. In pancreatic neuroendocrine neoplasms (PNENs), epigenetic deregulation is also considered to be of significance, as the most frequently mutated genes have an important function in epigenetic regulation. However, the exact changes in DNA methylation between PNENs and the endocrine cells of the pancreas, their likely cell-of-origin, remain largely unknown. Recently, two subtypes of PNENs have been described which were linked to cell-of-origin and have a different prognosis. A difference in the expression of the transcription factor PDX1 was one of the key molecular differences. In this study, we performed an exploratory genome-wide DNA methylation analysis using Infinium Methylation EPIC arrays (Illumina) on 26 PNENs and pancreatic islets of five healthy donors. In addition, the methylation profile of the PDX1 region was used to perform subtyping in a global cohort of 83 PNEN, 2 healthy alpha cell and 3 healthy beta cell samples. In our exploratory analysis, we identified 26,759 differentially methylated CpGs and 79 differentially methylated regions. The gene set enrichment analysis highlighted several interesting pathways targeted by altered DNA methylation, including MAPK, platelet-related and immune system-related pathways. Using the PDX1 methylation in 83 PNEN, 2 healthy alpha cell and 3 healthy beta cell samples, two subtypes were identified, subtypes A and B, which were similar to alpha and beta cells, respectively. These subtypes had different clinicopathological characteristics, a different pattern of chromosomal alterations and a different prognosis, with subtype A having a significantly worse prognosis compared with subtype B (HR 0.22 [95% CI: 0.051–0.95], p = 0.043). Hence, this study demonstrates that several cancer-related pathways are differently methylated between PNENs and normal islet cells. In addition, we validated the use of the PDX1 methylation status for the subtyping of PNENs and its prognostic importance