Sirolimus Stimulates Vascular Stem/Progenitor Cell Migration and Differentiation Into Smooth Muscle Cells via Epidermal Growth Factor Receptor/Extracellular Signal-Regulated Kinase/β-Catenin Signaling Pathway

Sirolimus-eluting stent therapy has achieved considerable success in overcoming coronary artery restenosis. However, there remain a large number of patients presenting with restenosis after the treatment, and the source of its persistence remains unclarified. Although recent evidence supports the contribution of vascular stem/progenitor cells in restenosis formation, their functional and molecular responses to sirolimus are largely unknown.

Queering the Family: Attitudes towards Lesbian and Gay Relationships and Families in Northern Ireland

The Northern Ireland Life andTimes (NILT) Survey has asked questions on lesbian, gay, bisexual and transgender (LGBT) issues since 1998. To date survey data have focused primarily on issues relating to prejudice, discrimination and tolerance. In 2012 a range of questions focussing more specifically on LGBT1 issues was included. This collected information on knowledge and perceptions of the LGBT population; personal prejudice; attitudes on equality issues; the visibility of LGBT people and family-related issues.

This update provides an overview of some of the information emerging from this data. It discusses attitudes towards same-sex relations and notable changes over time. Given recent political debate the primary focus of this paper is on attitudes relating to ‘queer’ marriage, family and parenting. We use the term ‘queer’ here to refer to ‘the diverse family structures formed by those with non-normative gender behaviours or sexual orientations’ (Bernstein and Reimann, 2001: 3). As previous updates have noted, there have been significant legislative and policy changes in this area (Jarman, 2010) and this continues with ongoing discussions regarding the development of a Sexual Orientation Strategy for Northern Ireland (Gray et al, 2013).

A Loop-mediated Isothermal Amplification Method for Rapid Direct Detection and Differentiation of Non-pathogenic and Verocytotoxigenic Escherichia coli in Beef and Bovine Faeces

The aim of this study was to develop a multiplex loop-mediated isothermal amplification (LAMP) method capable of detecting Escherichia coli generally and verocytotoxigenic E. coli (VTEC) specifically in beef and bovine faeces. The LAMP assay developed was highly specific (100%) and able to distinguish between E. coli and VTEC based on the amplification of the phoA, and stx1 and/or stx2 genes, respectively. In the absence of an enrichment step, the limit of detection 50% (LOD50) of the LAMP assay was determined to be 2.83, 3.17 and 2.83-3.17 log CFU/g for E. coli with phoA, stx1 and stx2 genes, respectively, when artificially inoculated minced beef and bovine faeces were tested. The LAMP calibration curves generated with pure cultures, and spiked beef and faeces, suggested that the assay had good quantification capability. Validation of the assay, performed using retail beef and bovine faeces samples, demonstrated good correlation between counts obtained by the LAMP assay and by a conventional culture method, but suggested the possibility of false negative LAMP results for 12.5-14.7% of samples tested. The multiplex LAMP assay developed potentially represents a rapid alternative to culture for monitoring E.coli levels in beef or faeces and it would provide additional information on the presence of VTEC. However, some further optimisation is needed to improve detection sensitivity.

Association of gene expression with sequential proliferation, differentiation and tumor formation in murine skin

Differential gene expression in two established initiation and promotion skin carcinogenesis models during promotion and tumor formation was determined by microarray technology with the purpose of distinguishing the genes more associated with neoplastic transformation from those linked with proliferation and differentiation. The first model utilized dimethylbenz[a]anthracene initiation and 12-O-tetradecanoylphorbol 13-acetate (TPA) promotion in the FVB/N mouse, and the second TPA promotion of the Tg.Ac mouse, which is endogenously initiated by virtue of an activated Ha-ras transgene. Comparison of gene expression profiles across the two models identified genes whose altered expression was associated with papilloma formation rather than TPA-induced proliferation and differentiation. DMBA suppressed TPA-induced differentiation which allowed identification of those genes associated more specifically with differentiation rather than proliferation. EASE (Expression Analysis Systemic Explorer) indicated a correlation between muscle-associated genes and skin differentiation, whereas genes involved with protein biosynthesis were strongly correlated with proliferation. For verification the altered expression of selected genes were confirmed by RT-PCR; Carbonic anhydrase 2, Thioredoxin 1 and Glutathione S-transferase omega 1 associated with papilloma formation and Enolase 3, Cystatin 6 and Filaggrin associated with TPA-induced proliferation and differentiation. In situ analysis located the papillomas Glutathione S-transferase omega 1 expression to the proliferating areas of the papillomas. Thus we have identified profiles of differential gene expression associated with the tumorigenesis and promotion stages for skin carcinogenesis in the mouse.

miR-7 and miR-214 are specifically expressed during neuroblastoma differentiation, cortical development and embryonic stem cells differentiation, and control neurite outgrowth in vitro

The mammalian nervous system exerts essential control on many physiological processes in the organism and is itself controlled extensively by a variety of genetic regulatory mechanisms. microRNA (miR), an abundant class of small non-coding RNA, are emerging as important post-transcriptional regulators of gene expression in the brain. Increasing evidence indicates that miR regulate both the development and function of the nervous system. Moreover, deficiency in miR function has also been implicated in a number of neurological disorders. Expression profile analysis of miR is necessary to understand their complex role in the regulation of gene expression during the development and differentiation of cells. Here we present a comparative study of miR expression profiles in neuroblastoma, in cortical development, and in neuronal differentiation of embryonic stem (ES) cells. By microarray profiling in combination with real time PCR we show that miR-7 and miR-214 are modulated in neuronal differentiation (as compared to miR-1, -16 and -133a), and control neurite outgrowth in vitro. These findings provide an important step toward further elucidation of miR function and miR-related gene regulatory networks in the mammalian central nervous system. (C) 2010 Elsevier Inc. All rights reserved.

Inhibition of histone deacetylase 2 increases apoptosis and p21(Cip1/WAF1) expression, independent of histone deacetylase 1

Histone deacetylases ( HDACs) 1 and 2 share a high degree of homology and coexist within the same protein complexes. Despite their close association, each possesses unique functions. We show that the upregulation of HDAC2 in colorectal cancer occurred early at the polyp stage, was more robust and occurred more frequently than HDAC1. Similarly, while the expression of HDACs1 and 2 were increased in cervical dysplasia and invasive carcinoma, HDAC2 expression showed a clear demarcation of high-intensity staining at the transition region of dysplasia compared to HDAC1. Upon HDAC2 knockdown, cells displayed an increased number of cellular extensions reminiscent of cell differentiation. There was also an increase in apoptosis, associated with increased p21(Cip1/WAF1) expression that was independent of p53. These results suggest that HDACs, especially HDAC2, are important enzymes involved in the early events of carcinogenesis, making them candidate markers for tumor progression and targets for cancer therapy.

FUS expression alters the differentiation response to all-trans retinoic acid in NB4 and NB4R2 cells

The FUS gene is overexpressed in acute myeloid leukaemia (AML) patients and has roles in transcription and mRNA processing. We used ectopic expression of FUS and FUS antisense sequences to assess the effect of modulation of FUS expression in all-trans retinoic acid (ATRA)-sensitive (NB4) and insensitive (NB4R2) human acute promyelocytic (APL) cell lines which express the t(15:17) translocation. Growth, viability and differentiation patterns were maintained, but the expression of the FUS antisense construct in both the cell lines altered the response to ATRA: the previously ATRA-sensitive NB4 cells exhibited resistance; whilst the previously resistant NB4R2 cells showed a differentiation response to treatment.

The role of p63 in epidermal morphogenesis and neoplasia

The p53 family of transcription factors is made up of p53, p63 and p73, which share significant structural homology. In particular, transcriptional complexity and the expression of multiple protein isoforms are an emergent trait of all family members. p63 is the evolutionarily eldest member of the p53 family and the various isoforms have critical roles in the development of stratifying epithelia. Recent results have uncovered additional splice variants, adding to the complexity of the transcriptional architecture of p63. These observations and the emerging extensive interplay between p63 and p53 in development, proliferation and differentiation underline the importance of considering all isoforms and family members in studies of the function of p53 family members.

HDAC3 is crucial in shear- and VEGF-induced stem cell differentiation toward endothelial cells

Reendothelialization involves endothelial progenitor cell (EPC) homing, proliferation, and differentiation, which may be influenced by fluid shear stress and local flow pattern. This study aims to elucidate the role of laminar flow on embryonic stem (ES) cell differentiation and the underlying mechanism. We demonstrated that laminar flow enhanced ES cell-derived progenitor cell proliferation and differentiation into endothelial cells (ECs). Laminar flow stabilized and activated histone deacetylase 3 (HDAC3) through the Flk-1-PI3K-Akt pathway, which in turn deacetylated p53, leading to p21 activation. A similar signal pathway was detected in vascular endothelial growth factor-induced EC differentiation. HDAC3 and p21 were detected in blood vessels during embryogenesis. Local transfer of ES cell-derived EPC incorporated into injured femoral artery and reduced neointima formation in a mouse model. These data suggest that shear stress is a key regulator for stem cell differentiation into EC, especially in EPC differentiation, which can be used for vascular repair, and that the Flk-1-PI3K-Akt-HDAC3-p53-p21 pathway is crucial in such a process.

Changes of 5-hydroxymethylcytosine distribution during myeloid and lymphoid differentiation of CD34+ cells

BACKGROUND: Hematopoietic stem cell renewal and differentiation are regulated through epigenetic processes. The conversion of 5-methylcytosine into 5-hydroxymethylcytosine (5hmC) by ten-eleven-translocation enzymes provides new insights into the epigenetic regulation of gene expression during development. Here, we studied the potential gene regulatory role of 5hmC during human hematopoiesis.

RESULTS: We used reduced representation of 5-hydroxymethylcytosine profiling (RRHP) to characterize 5hmC distribution in CD34+ cells, CD4+ T cells, CD19+ B cells, CD14+ monocytes and granulocytes. In all analyzed blood cell types, the presence of 5hmC at gene bodies correlates positively with gene expression, and highest 5hmC levels are found around transcription start sites of highly expressed genes. In CD34+ cells, 5hmC primes for the expression of genes regulating myeloid and lymphoid lineage commitment. Throughout blood cell differentiation, intragenic 5hmC is maintained at genes that are highly expressed and required for acquisition of the mature blood cell phenotype. Moreover, in CD34+ cells, the presence of 5hmC at enhancers associates with increased binding of RUNX1 and FLI1, transcription factors essential for hematopoiesis.

CONCLUSIONS: Our study provides a comprehensive genome-wide overview of 5hmC distribution in human hematopoietic cells and new insights into the epigenetic regulation of gene expression during human hematopoiesis.

Relationship between biomedical catheter surface properties and lubricity as determined using textural analysis and multiple regression analysis

In this study, the surface properties of and work required to remove 12 commercially available and developmental catheters from a model biological medium (agar), a measure of catheter lubricity, were characterised and the relationships between these properties were examined using multiple regression and correlation analysis. The work required for removal of catheter sections (7 cm) from a model biological medium (1% w/w agar) were examined using tensile analysis. The water wettability of the catheters were characterised using dynamic contact angle analysis, whereas surface roughness was determined using atomic force microscopy. Significant differences in the ease of removal were observed between the various catheters, with the silicone-based materials generally exhibiting the greatest ease of removal. Similarly, the catheters exhibited a range of advancing and receding contact angles that were dependent on the chemical nature of each catheter. Finally, whilst the microrugosities of the various catheters differed, no specific relationship to the chemical nature of the biomaterial was apparent. Using multiple regression analysis, the relationship between ease of removal, receding contact angle and surface roughness was defined as: Work done (N mm) 17.18 + 0.055 Rugosity (nm)-0.52 Receding contact angle (degrees) (r = 0.49). Interestingly, whilst the relationship between ease of removal and surface roughness was significant (r = 0.48, p = 0.0005), in which catheter lubricity increased as the surface roughness decreased, this was not the case with the relationship between ease of removal and receding contact angle (r = -0.18, p > 0.05). This study has therefore uniquely defined the contributions of each of these surface properties to catheter lubricity. Accordingly, in the design of urethral catheters. it is recommended that due consideration should be directed towards biomaterial surface roughness to ensure maximal ease of catheter removal. Furthermore, using the method described in this study, differences in the lubricity of the various catheters were observed that may be apparent in their clinical use. (C) 2003 Elsevier Ltd. All rights reserved.