Vitamin D Receptor Modulates the Neoplastic Phenotype Through Antagonistic Growth Regulatory Signals

Vitamin D receptor (VDR) can modulate functionally antagonistic growth regulatory pathways, involving beta-catenin/E-cadherin on one hand and osteopontin (OPN) on the other. This study investigates effects of VDR ligand treatment on the balance of these discordant signals and on associated cell behavior. Treatment of Rama 37 or SW480 cells by 1 alpha,25-(OH)(2) D-3 or analogs suppressed beta-catenin/Lef-1/Tcf signaling and upregulated E-cadherin, consistent with a cancer-inhibitory action. Conversely, treatment also increased transcription of OPN that may be implicated in tumor progression. Molecular crosstalk was observed between the antagonistic VDR-dependent signals, in that beta-catenin/Lef-1/Tcf molecules modulated VDR activation of OPN. Treatment effects on cell growth were related to a constitutive balance of OPN and E-cadherin expression. No growth effects were observed in Rama 37 cells that have low OPN and high E-cadherin expression. Conversely, treatment of Rama 37 stably transfected subclones that had high OPN and/or low level E-cadherin induced small but significant increases of cell attachment to fibronectin, anchorage-independent growth or invasion. This study shows that relative expression levels of key VDR downstream genes may influence growth regulation by 1 alpha,25-(OH)(2) D-3 or analogs. These findings may be relevant to the cell- or tissue-specificity of vitamin D growth regulation. (C) 2009 Wiley-Liss, Inc.

Langerhans cell (LC) proliferation mediates neonatal development, homeostasis, and inflammation-associated expansion of the epidermal LC network.

Most tissues develop from stem cells and precursors that undergo differentiation as their proliferative potential decreases. Mature differentiated cells rarely proliferate and are replaced at the end of their life by new cells derived from precursors. Langerhans cells (LCs) of the epidermis, although of myeloid origin, were shown to renew in tissues independently from the bone marrow, suggesting the existence of a dermal or epidermal progenitor. We investigated the mechanisms involved in LC development and homeostasis. We observed that a single wave of LC precursors was recruited in the epidermis of mice around embryonic day 18 and acquired a dendritic morphology, major histocompatibility complex II, CD11c, and langerin expression immediately after birth. Langerin+ cells then undergo a massive burst of proliferation between postnatal day 2 (P2) and P7, expanding their numbers by 10–20-fold. After the first week of life, we observed low-level proliferation of langerin+ cells within the epidermis. However, in a mouse model of atopic dermatitis (AD), a keratinocyte signal triggered increased epidermal LC proliferation. Similar findings were observed in epidermis from human patients with AD. Therefore, proliferation of differentiated resident cells represents an alternative pathway for development in the newborn, homeostasis, and expansion in adults of selected myeloid cell populations such as LCs. This mechanism may be relevant in locations where leukocyte trafficking is limited.

Persistent measles virus infection of mouse neural cells lacking known human entry receptors

Aims: Infection of the mouse central nervous system with wild type (WT) and vaccine strains of measles virus (MV) results in lack of clinical signs and limited antigen detection. It is considered that cell entry receptors for these viruses are not present on murine neural cells and infection is restricted at cell entry.

Methods: To examine this hypothesis, virus antigen and caspase 3 expression (for apoptosis) was compared in primary mixed, neural cell cultures infected in vitro or prepared from mice infected intracerebrally with WT, vaccine or rodent neuroadapted viruses. Viral RNA levels were examined in mouse brain by nested and real-time reverse transcriptase polymerase chain reaction.

Results: WT and vaccine strains were demonstrated for the first time to infect murine oligodendrocytes in addition to neurones despite a lack of the known MV cell receptors. Unexpectedly, the percentage of cells positive for viral antigen was higher for WT MV than neuroadapted virus in both in vitro and ex vivo cultures. In the latter the percentage of positive cells increased with time after mouse infection. Viral RNA (total and mRNA) was detected in brain for up to 20 days, while cultures were negative for caspase 3 in WT and vaccine virus infections.

Conclusions: WT and vaccine MV strains can use an endogenous cell entry receptor(s) or alternative virus uptake mechanism in murine neural cells. However, viral replication occurs at a low level and is associated with limited apoptosis. WT MV mouse infection may provide a model for the initial stages of persistent MV human central nervous system infections.

Effects of breed and production system on lameness parameters in dairy cattle

The aim of the present study was to assess the effects of Holstein-Friesian (HF) and Norwegian (N) dairy cattle genotypes on lameness parameters in dairy cattle within different production systems over the first 2 lactations. Following calving, HF (n = 39) and N (n = 45) heifers were allocated to 1 of 3 systems of production (high level of concentrate, low level of concentrate, and grass-based). High-and low-concentrate animals were continuously housed indoors on a rotational system so that they spent similar amounts of time on slatted and solid concrete floors. Animals on the grass treatment grazed from spring to autumn in both years of the study, so that most animals on this treatment grazed from around peak to late lactation. Claw health was recorded in both hind claws of each animal at 4 observation periods during each lactation as follows: 1) -8 to 70 d postcalving, 2) 71 to 150 d postcalving, 3) 151 to 225 d postcalving, and 4) 226 to 364 d postcalving. Sole lesions, heel erosion, axial wall deviation, sole length of the right lateral hind claw (claw length), right heel width, and right lateral hind heel height were recorded as well as the presence of digital dermatitis. The N cows had lower (better) white line and total lesion scores than HF cows. Cows on the high-and low-concentrate treatments had better sole and total lesion scores than cows on the grass treatment. The HF cows had better locomotion scores than N cows. Breed and production system differences were observed with respect to claw conformation, including claw length, heel width, and heel height. Digital dermatitis was associated with worse sole lesion scores and interacted with production system to influence white line lesion scores and maximum heel erosion scores. This study shows that genetic, environmental, and infectious factors are associated with hoof pathologies in dairy cows.

Detection of glucocorticoid bioactivity in bovine urine samples using a reporter gene assay.

The illegal use of anabolic substances in the meat producing industry is an ongoing problem due to the continual production of new synthetic compounds and/or the practice of low-level cocktail administration to avoid detection by the surveillance schemes of EU member states National Plan surveillance systems.

We present a highly sensitive reporter gene assay and sample extraction procedure based on a two step solid phase extraction and high performance liquid chromatography, developed for the detection of glucocorticoid abuse in bovine urine. The assay is capable of detecting compounds with glucocorticoid bioactivity and is extremely sensitive with an EC50 of 0.79 ng mL-1 for dexamethasone. New or unknown compounds with glucocorticoid bioactivity and low-level cocktail mixtures are detectable by this assay.

Cross-reactivity data for a range of 11ß-hydroxyglucocorticoids has been provided. This assay shows low interference from the 11-keto prohormones and other steroidal hormones. The assay may be suitable for application in other matrices such as hair. In conclusion this screening assay offers advantages over existing analytical techniques.

On-line Emotion Recognition in a 3-D Activation-Valence-Time Continuum using Acoustic and Linguistic Cues

For many applications of emotion recognition, such as virtual agents, the system must select responses while the user is speaking. This requires reliable on-line recognition of the user’s affect. However most emotion recognition systems are based on turnwise processing. We present a novel approach to on-line emotion recognition from speech using Long Short-Term Memory Recurrent Neural Networks. Emotion is recognised frame-wise in a two-dimensional valence-activation continuum. In contrast to current state-of-the-art approaches, recognition is performed on low-level signal frames, similar to those used for speech recognition. No statistical functionals are applied to low-level feature contours. Framing at a higher level is therefore unnecessary and regression outputs can be produced in real-time for every low-level input frame. We also investigate the benefits of including linguistic features on the signal frame level obtained by a keyword spotter.

Multimodal Biometric Human Recognition for Perceptual Human-Computer Interaction

In this paper, a novel video-based multimodal biometric verification scheme using the subspace-based low-level feature fusion of face and speech is developed for specific speaker recognition for perceptual human--computer interaction (HCI). In the proposed scheme, human face is tracked and face pose is estimated to weight the detected facelike regions in successive frames, where ill-posed faces and false-positive detections are assigned with lower credit to enhance the accuracy. In the audio modality, mel-frequency cepstral coefficients are extracted for voice-based biometric verification. In the fusion step, features from both modalities are projected into nonlinear Laplacian Eigenmap subspace for multimodal speaker recognition and combined at low level. The proposed approach is tested on the video database of ten human subjects, and the results show that the proposed scheme can attain better accuracy in comparison with the conventional multimodal fusion using latent semantic analysis as well as the single-modality verifications. The experiment on MATLAB shows the potential of the proposed scheme to attain the real-time performance for perceptual HCI applications.

The application of Reporter Gene Assays for the detection of endocrine disruptors in sport supplements.

The increasing availability and use of sports supplements is of concern as highlighted by a number of studies reporting endocrine disruptor contamination in such products. The health food supplement market, including sport supplements, is growing across the Developed World. Therefore, the need to ensure the quality and safety of sport supplements for the consumer is essential. The development and validation of two reporter gene assays coupled with solid phase sample preparation enabling the detection of estrogenic and androgenic constituents in sport supplements is reported. Both assays were shown to be of high sensitivity with the estrogen and androgen reporter gene assays having an EC50 of 0.01 ng mL-1 and 0.16 ng mL-1 respectively. The developed assays were applied in a survey of 63 sport supplements samples obtained across the Island of Ireland with an additional seven reference samples previously investigated using LC–MS/MS. Androgen and estrogen bio-activity was found in 71% of the investigated samples. Bio-activity profiling was further broken down into agonists, partial agonists and antagonists. Supplements (13) with the strongest estrogenic bio-activity were chosen for further investigation. LC–MS/MS analysis of these samples determined the presence of phytoestrogens in seven of them. Supplements (38) with androgen bio-activity were also selected for further investigation. Androgen agonist bio-activity was detected in 12 supplements, antagonistic bio-activity was detected in 16 and partial antagonistic bio-activity was detected in 10. A further group of supplements (7) did not present androgenic bio-activity when tested alone but enhanced the androgenic agonist bio-activity of dihydrotestosterone when combined. The developed assays offer advantages in detection of known, unknown and low-level mixtures of endocrine disruptors over existing analytical screening techniques. For the detection and identification of constituent hormonally active compounds the combination of biological and physio-chemical techniques is optimal.

‘The ‘Complexity of Imprisonment: The Northern Ireland Experience'

This article will examine the thesis that Northern Ireland experiences a relatively low level of crime. It will explore the possible reasons why crime in the North has not witnessed a dramatic increase. In light of this, the article will highlight the difficulties surrounding the current prison system and illustrate that once again Northern Ireland is experiencing a very different criminal justice system in comparison to Great Britain. Although the prisons are now being used predominately to deal with “ordinary’ crime”, they are still part of the political process.

Glycoprotein VI/Fc receptor gamma chain-independent tyrosine phosphorylation and activation of murine platelets by collagen

We have investigated the ability of collagen to induce signalling and functional responses in suspensions of murine platelets deficient in the FcRgamma (Fc receptor gamma) chain, which lack the collagen receptor GPVI (glycoprotein VI). In the absence of the FcRgamma chain, collagen induced a unique pattern of tyrosine phosphorylation which was potentiated by the thromboxane analogue U46619. Immunoprecipitation studies indicated that neither collagen alone nor the combination of collagen plus U46619 induced phosphorylation of the GPVI-regulated proteins Syk and SLP76 (Src homology 2-containing leucocyte protein of 76 kDa). A low level of tyrosine phosphorylation of phospholipase Cgamma2 was observed, which was increased in the presence of U46619, although the degree of phosphorylation remained well below that observed in wild-type platelets (similar to 10%). By contrast, collagen-induced phosphorylation of the adapter ADAP (adhesion- and degranulation-promoting adapter protein) was substantially potentiated by U46619 to levels equivalent to those observed in wild-type platelets. Collagen plus U46619 also induced significant phosphorylation of FAK (focal adhesion kinase). The functional significance of collagen-induced non-GPVI signals was highlighted by the ability of U46619 and collagen to induce the secretion of ATP in FcRgamma chain-deficient platelets, even though neither agonist was effective alone. Protein tyrosine phosphorylation and the release of ATP were abolished by the anti(alpha2 integrin) antibodies Ha1/29 and HMalpha2, but not by blockade of alphaIIbbeta3. These results illustrate a novel mechanism of platelet activation by collagen which is independent of the GPVI-FcRgamma chain complex, and is facilitated by binding of collagen to integrin alpha2beta1.

Nitride-strengthened reduced activation ferritic/martensitic steels

Two nitride-strengthened reduced activation ferritic/martensitic (RAFM) steels with different Mn contents were investigated. The experimental steels were designed based on the chemical composition of Eurofer 97 steel but the C content was reduced to an extremely low level. Microstructure observation and hardness tests showed that the steel with low Mn content (0.47 wt.%) could not obtain a full martensitic microstructure due to the inevitable δ-ferrite independent of cooling rate after soaking. This steel showed similar room temperature strength and higher strength at 600 °C, but lower impact toughness, compared with Eurofer 97 steel. Fractography of the Charpy impact specimen revealed that the low room temperature toughness should be related to the Ta-rich inclusions initiating the cleavage fracture. The larger amount of V-rich nitrides and more dissolved Cr in the matrix could be responsible for the strength being similar to Eurofer 97 steel. In the second steel developed from the first steel by increasing the Mn content from 0.47 wt.% to 3.73 wt.%, a microstructure of full martensite could be obtained.

Conflict, Transformation, and Former Loyalist Paramilitary Prisoners in Northern Ireland

Following the 1998 Belfast Agreement in Northern Ireland, levels of paramilitary violence have declined substantially. Among loyalists, the Ulster Volunteer Force (UVF) and associated Red Hand Commando (RHC) have formally renounced violence, and dissolved their 'military structures', and perhaps the most reticent of all of the major paramilitary groupings, the Ulster Defence Association (UDA), has taken on board the central tenets of conflict transformation, and 'stood down' all of its 'active service units' in the Ulster Freedom Fighters (UFF). Thus, paramilitary violence now is mainly confined to the activities of 'dissident' republican groups, notably the Real and Continuity IRAs, although low-level sectarian violence remains a problem. Such dramatic societal and political change has resulted in a focus on the roles of formal party political leadership as agents of social change. This gaze, however, tends to obscure other important events such as the efforts, structures and approaches taken at the grassroots level to uphold and sustain conflict transformation and to maintain a reduction in violence. This article provides analysis of the role played by former loyalist paramilitary combatants in conflict transformation, and draws on material obtained through significant access to those former paramilitaries engaged in processes of societal shifts. In both personal and structural terms there is evidence of former combatants working to diminish the political tensions that remain as a result of the long-term inter-communal hostility developed across decades of violence and conflict.

Surface Plasmon Resonance Biosensor Screening Method for Paralytic Shellfish Poisoning Toxins: A Pilot Interlaboratory Study

A surface plasmon resonance (SPR) optical biosensor method was developed for the detection of paralytic shellfish poisoning (PSP) toxins in shellfish. This application was transferred in the form of a prototype kit to seven laboratories using Biacore QSPR optical biosensor instrumentation for interlaboratory evaluation. Each laboratory received 20 shellfish samples across a range of species including blind duplicates for analysis. The samples consisted of 4 noncontaminated samples spiked in duplicate with a low level of PSP toxins (240 mu g STXcliHCl equivalents/kg), a high level of saxitoxin (825 mu g STXdiHCl/kg), 2 noncontarninated, and 14 naturally contaminated samples. All 7 participating laboratories completed the study, and HorRat values obtained were

Characterisation of the photocatalyst Pilkington Activ (TM): a reference film photocatalyst?

Pilkington Glass Activ(TM) represents a possible suitable successor to P25 TiO2, especially as a benchmark photocatalyst film for comparing other photocatalyst or PSH self-cleaning films. Activ(TM) is a glass product with a clear, colourless, effectively invisible, photocatalytic coating of titania that also exhibits PSH. Although not as active as a film of P25 TiO2, Activ(TM) vastly superior mechanical stability, very reproducible activity and widespread commercial availability makes it highly attractive as a reference photocatalytic film. The photocatalytic and photo-induced superhydrophilitic (PSH) properties of Activ(TM) are studied in some detail and the results reported. Thus, the kinetics of stearic acid destruction (a 104 electron process) are zero order over the stearic acid range 4-129 monolayers and exhibit formal quantum efficiencies (FQE) of 0.7 X 10(-5) and 10.2 x 10(-5) molecules per photon when irradiated with light of 365 +/- 20 and 254 nm, respectively; the latter appears also to be the quantum yield for Activ(TM) at 254 nm. The kinetics of stearic acid destruction exhibit Langmuir-Hinshelwood-like saturation type kinetics as a function of oxygen partial pressure, with no destruction occurring in the absence of oxygen and the rate of destruction appearing the same in air and oxygen atmospheres. Further kinetic work revealed a Langmuir adsorption type constant for oxygen of 0.45 +/- 0.16 kPa(-1) and an activation energy of 19 +/- 1 Kj mol(-1). A study of the PSH properties of Activ(TM) reveals a high water contact angle (67) before ultra-bandgap irradiation reduced to 0degrees after prolonged irradiation. The kinetics of PSH are similar to those reported by others for sol-gel films using a low level of UV light. The kinetics of contact angle recovery in the dark appear monophasic and different to the biphasic kinetics reported recently by others for sol-gel films [J. Phys. Chem. B 107 (2003) 1028]. Overall, Activ(TM) appears a very suitable reference material for semiconductor film photocatalysis. (C) 2003 Elsevier Science B.V All rights reserved.

The activity of a putative polyisoprenol-linked sugar translocase (Wzx) involved in Escherichia coli O antigen assembly is independent of the chemical structure of the O repeat

During O antigen lipopolysaccharide (LPS) synthesis in bacteria, transmembrane migration of undecaprenylpyrophosphate (Und-P-P)-bound O antigen subunits occurs before their polymerization and ligation to the rest of the LPS molecule. Despite the general nature of the translocation process, putative O-antigen translocases display a low level of amino acid sequence similarity. In this work, we investigated whether complete O antigen subunits are required for translocation. We demonstrate that a single sugar, GlcNAc, can be incorporated to LPS of Escherichia coli K-12. This incorporation required the functions of two O antigen synthesis genes, wecA (UDP-GlcNAc:Und-P GlcNAc-1-P transferase) and wzx (O-antigen translocase). Complementation experiments with putative O-antigen translocases from E. coli O7 and Salmonella enterica indicated that translocation of O antigen subunits is independent of the chemical structure of the saccharide moiety. Furthermore, complementation with putative translocases involved in synthesis of exopolysaccharides demonstrated that these proteins could not participate in O antigen assembly. Our data indicate that recognition of a complete Und-P-P-bound O antigen subunit is not required for translocation and suggest a model for O antigen synthesis involving recognition of Und-P-P-linked sugars by a putative complex made of Wzx translocase and other proteins involved in the processing of O antigen.