Role of hypoxia and hypoxia-inducible factor-1 in tumour immune escape

Previous studies revealed that, upon exposure to hypoxia, tumour cells acquire resistance to the cytolytic activity of IL-2-activated lymphocytes. The MHC class I chain-related (MIC) molecules – comprised of MICA and MICB – are ligands for the activating NKG2D receptor on Natural Killer (NK) and CD8+ T cells. MIC-NKG2D interactions lead to the activation of NK and CD8+ T cells and the subsequent lysis of the tumour cells. The study also showed that the mechanism of the hypoxia-mediated immune escape involves the shedding of MIC, specifically MICA, from the tumour cell surface. The objective of the present study was to determine whether the shedding of MICA requires the expression of hypoxia inducible factor-1 (HIF-1), a transcription factor that regulates cellular adaptations to hypoxia. Exposure to hypoxia (0.5% O2 vs. 20% O2) led to the shedding of MIC from the surface of MDA-MB-231 human breast cancer cells and DU-145 human prostate cancer cells as determined by flow cytometry. Knockdown of HIF-1α mRNA using siRNA technology resulted in inhibition of HIF-1α accumulation under hypoxic conditions as determined by Western blot analysis. Parallel study revealed that knockdown of HIF-1α also blocked the shedding of MICA from the surface of MDA-MB-231 cells exposed to hypoxia. These results indicate that HIF-1 is required for the hypoxia-mediated shedding of MICA and, consequently, that HIF-1 may play an important role in tumour immune escape. Ongoing studies aim to determine the HIF-1 target genes involved in the shedding of MICA under hypoxia.

ROLE OF CAVEOLIN-3 IN THE MODULATION OF HERG POTASSIUM CHANNEL

The human ether-a-go-go-related gene (hERG) encodes the pore-forming subunit of the rapidly activating delayed rectifier potassium channel (IKr) that is important for cardiac repolarization. Previously, we have discovered that hERG channels rapidly internalize in low extracellular K+ ([K+]o). In cell culture, this process is driven by the endocytic protein, caveolin-1 (Cav1), which is an integral player in the caveolae-dependant endocytosis pathway. However, in the heart, Caveolin-3 (Cav3) is, in fact, the predominant form in the myocyte, and thus may play a direct role in regulating hERG expression in the heart. Thus, I hypothesize that this reduction of hERG conductance in cardiac myocytes derives from the presence of Cav3, which is integral regulator of hERG homeostasis innately in the heart. To investigate the effect of Cav3 on hERG, I overexpressed Cav3 in human embryonic kidney 293 (HEK-293) cells stably expressing hERG channels. Cav3 overexpression significantly and specifically decreased both the hERG current amplitude and the mature channel expression in normal culture conditions. Co-immunoprecipitation analysis and confocal imaging demonstrated an association between hERG and Cav3 in HEK cells as well as rat and rabbit cardiomyocytes. Mechanistically, I discovered that Cav3 possesses a faster turnover rate compared to Cav1, and can enhance hERG degradation through up-regulating mature channel ubiquitination via the ubiquitin ligase, NEDD4-2. Knockdown of Cav3 in neonatal cardiac myocytes also enhanced hERG expression. My data indicate that Cav3 participates in hERG trafficking, and is an important regulator of hERG channel homeostasis in cardiac myocytes. This information provides a platform for future intervention of the hERG-induced type-2 long QT syndrome (LQTS).

Teachers' and teacher educators' lives: the role of emotion

This paper reports research which focuses on ways of enhancing understandings by teachers of the key role that emotions play in their personal professional growth. It combines the narrative, autobiographical accounts of teachers attending part-time masters degree programmes in England (Continuing Professional Development and School Improvement) and Northern Ireland (Personal and Social Development) with an interrogation of the underlying values which affect the practices of their tutors. It reveals the effects of powerful and often unacknowledged interaction between personal biography and professional and social contexts upon teachers in schools and higher education.

Amyloid B-peptide 1-40 increases neuronal membrane fluidity: role of cholesterol and brain region

There is increasing evidence of an interaction between cholesterol dynamics and Alzheimer's disease (AD), and amyloid ß-peptide may play an important role in this interaction. Aß destabilizes brain membranes and this action of Aß may be dependent on the amount of membrane cholesterol. We tested this hypothesis by examining effects of Aß1-40 on the annular fluidity (i.e., lipid environment adjacent to proteins) and bulk fluidity of rat synaptic plasma membranes (SPM) of the cerebral cortex, cerebellum, and hippocampus using the fluorescent probe pyrene and energy transfer. Amounts of cholesterol and phospholipid of SPM from each brain region were determined. SPM of the cerebellum were significantly more fluid as compared with SPM of the cerebral cortex and hippocampus. Aß significantly increased (P 0.01) annular and bulk fluidity in SPM of cerebral cortex and hippocampus. In contrast, Aß had no effect on annular fluidity and bulk fluidity of SPM of cerebellum. The amounts of cholesterol in SPM of cerebral cortex and hippocampus were significantly higher (P 0.05) than amount of cholesterol in SPM of cerebellum. There was significantly less (P 0.05) total phospholipid in cerebellar SPM as compared with SPM of cerebral cortex. Neuronal membranes enriched in cholesterol may promote accumulation of Aß by hydrophobic interaction, and such an interpretation is consistent with recent studies showing that soluble Aß can act as a seed for fibrillogenesis in the presence of cholesterol.

Effects of trauma on bipolar disorder: the mediational role of interpersonal difficulties and alcohol dependence

Objectives: This study examined: (i) the prevalence of trauma in a bipolar disorder (BD) sample, and (ii) how trauma histories mediated by interpersonal difficulties and alcohol dependence impact on the severity of BD. The prevalence of posttraumatic stress disorder (PTSD) and its relationship to outcomes in BD were also examined.

Methods: Sixty participants were recruited from a geographically well-defined mental health service in Northern Ireland. Self-reported trauma histories, PTSD, interpersonal difficulties and alcohol dependence and were examined in relation to illness severity.

Results: A high prevalence of trauma was found. Trauma predicted the frequency of hospital admissions (R-2 = 0.08), quality of life (R-2 = 0.23) and inter-episode depressive symptoms (R-2 = 0.13). Interpersonal difficulties, but not alcohol dependence, appeared to play an important role in mediating these adverse effects. While only 8% of the sample met criteria for active PTSD, this comorbid disorder was associated with BD severity.

Conclusions: This study indicates that awareness of trauma is important in understanding individual differences in bipolar presentations. The theoretical and clinical implications of evidence that trauma is related to more adverse outcomes in BD are discussed. The finding that interpersonal difficulties mediate the relationship between trauma and BD severity is novel. The need for adjunctive evidence-based treatments targeting interpersonal difficulties is considered.

The role of advanced glycation end products in retinal microvascular leukostasis

PURPOSE: A critical event in the pathogenesis of diabetic retinopathy is the inappropriate adherence of leukocytes to the retinal capillaries. Advanced glycation end-products (AGEs) are known to play a role in chronic inflammatory processes, and the authors postulated that these adducts may play a role in promoting pathogenic increases in proinflammatory pathways within the retinal microvasculature. METHODS: Retinal microvascular endothelial cells (RMECs) were treated with glycoaldehyde-modified albumin (AGE-Alb) or unmodified albumin (Alb). NFkappaB DNA binding was measured by electromobility shift assay (EMSA) and quantified with an ELISA: In addition, the effect of AGEs on leukocyte adhesion to endothelial cell monolayers was investigated. Further studies were performed in an attempt to confirm that this was AGE-induced adhesion by co-incubation of AGE-treated cells with soluble receptor for AGE (sRAGE). Parallel in vivo studies of nondiabetic mice assessed the effect of intraperitoneal delivery of AGE-Alb on ICAM-1 mRNA expression, NFkappaB DNA-binding activity, leukostasis, and blood-retinal barrier breakdown. RESULTS: Treatment with AGE-Alb significantly enhanced the DNA-binding activity of NFkappaB (P = 0.0045) in retinal endothelial cells (RMECs) and increased the adhesion of leukocytes to RMEC monolayers (P = 0.04). The latter was significantly reduced by co-incubation with sRAGE (P <0.01). Mice infused with AGE-Alb demonstrated a 1.8-fold increase in ICAM-1 mRNA when compared with control animals (P <0.001, n = 20) as early as 48 hours, and this response remained for 7 days of treatment. Quantification of retinal NFkappaB demonstrated a threefold increase with AGE-Alb infusion in comparison to control levels (AGE Alb versus Alb, 0.23 vs. 0.076, P <0.001, n = 10 mice). AGE-Alb treatment of mice also caused a significant increase in leukostasis in the retina (AGE-Alb versus Alb, 6.89 vs. 2.53, n = 12, P <0.05) and a statistically significant increase in breakdown of the blood-retinal barrier (AGE Alb versus Alb, 8.2 vs. 1.6 n = 10, P <0.001). CONCLUSIONS: AGEs caused upregulation of NFkappaB in the retinal microvascular endothelium and an AGE-specific increase in leukocyte adhesion in vitro was also observed. In addition, increased leukocyte adherence in vivo was demonstrated that was accompanied by blood-retinal barrier dysfunction. These findings add further evidence to the thinking that AGEs may play an important role in the pathogenesis of diabetic retinopathy.

New ways to break an old bond: the bacterial carbon-phosphorus hydrolases and their role in biogeochemical phosphorus cycling

Phosphonates are organophosphorus molecules that contain the highly stable C-P bond, rather than the more common, and more labile, C-O-P phosphate ester bond. They have ancient origins but their biosynthesis is widespread among more primitive organisms and their importance in the contemporary biosphere is increasingly recognized; for example phosphonate-P is believed to play a particularly significant role in the productivity of the oceans. The microbial degradation of phosphonates was originally thought to occur only under conditions of phosphate limitation, mediated exclusively by the poorly characterized C-P lyase multienzyme system, under Pho regulon control. However, more recent studies have demonstrated the Pho-independent mineralization by environmental bacteria of three of the most widely distributed biogenic phosphonates: 2-aminoethylphosphonic acid (ciliatine), phosphonoacetic acid, and 2-amino-3-phosphonopropionic acid (phosphonoalanine). The three phosphonohydrolases responsible have unique specificities and are members of separate enzyme superfamilies; their expression is regulated by distinct members of the LysR family of bacterial transcriptional regulators, for each of which the phosphonate substrate of the respective degradative operon serves as coinducer. Previously no organophosphorus compound was known to induce the enzymes required for its own degradation. Whole-genome and metagenome sequence analysis indicates that the genes encoding these newly described C-P hydrolases are distributed widely among prokaryotes. As they are able to function under conditions in which C-P lyases are inactive, the three enzymes may play a hitherto-unrecognized role in phosphonate breakdown in the environment and hence make a significant contribution to global biogeochemical P-cycling.

The functional role of Gammarus (Crustacea, Amphipoda): shredders, predators, or both?

Gammarus spp. are traditionally viewed under the functional feeding group (FFG) concept as herbivorous 'shredders'. Although recent studies suggest that Gammarus should also be viewed as predators, this latter role remains contentious. Here, in a laboratory experiment, we objectively examine the balance between shredder and predator roles in a common freshwater species. Gammarus pulex preyed significantly on mayfly nymph, Baetis rhodani, in both the presence and absence of excess leaf material. There was no significant difference in predation where the alternative food, that is, leaf material, was present as compared to absent. Also, G. pulex shredded leaf material in the presence and absence of B. rhodani. However, shredding was significantly reduced where alternative food, that is, B. rhodani prey, was present as compared to absent. Further, G. pulex had a clear leaf species preference. Our results suggest that Gammarus function as both predators and shredders, with the balance of the two roles perhaps depending on food availability and quality. We discuss implications for the use of the FFG concept in assessing freshwater processes, and the role that Gammarus predation may play in structuring macroinvertebrate communities.

The catalytic role of water in CO oxidation

Water, one of the most popular species in our planet, can play a catalytic role in many reactions, including reactions in heterogeneous catalysis. In a recent experimental work, Bergeld, Kasemo, and Chakarov demonstrated that water is able to promote CO oxidation under low temperatures (similar to200 K). In this study, we choose CO oxidation on Pt(111) in the presence of water as a model system to address the catalytic role of water for surface reactions in general using density functional theory. Many elementary steps possibly involved in the CO oxidation on Pt(111) at low temperatures have been investigated. We find the following. First, in the presence of water, the CO oxidation barrier is reduced to 0.33 eV (without water the barrier is 0.80 eV). This barrier reduction is mainly due to the H-bonding between the H in the H2O and the O at the transition state (TS), which stabilizes the TS. Second, CO can readily react with OH with a barrier of 0.44 eV, while COOH dissociation to produce CO2 is not easy (the barrier is 1.02 eV). Third, in the H2O+OH mixed phase, CO can be easily converted into CO2. It occurs through two steps: CO reacts with OH, forming COOH; and COOH transfers the H to a nearby H2O and, at the same time, an H in the H2O transfers to a OH, leading to CO2 formation. The reaction barrier of this process is 0.60 eV under CO coverage of 1/6 ML and 0.33 eV under CO coverage of 1/3 ML. The mechanism of CO oxidation at low temperatures is discussed. On the basis of our calculations, we propose that the water promotion effect can in general be divided into two classes: (i) By H-bonding between the H of H2O and an electron negative species such as the O in the reaction of CO+O+H2O-->CO2+H2O, H2O can stabilize the TS of the reaction and hence reduce the barrier. (ii) H2O first dissociates into H and OH and then OH or H participates directly in the reaction to induce new reaction mechanism with more favorable routes, in which OH or H can act as an intermediate. (C) 2003 American Institute of Physics.

A Dual Role for Lecithin:Cholesterol Acyltransferase (EC 2.3.1.43) in Lipoprotein Oxidation

Lecithin:cholesterol acyltransferase (LCAT) is a key enzyme involved in lipoprotein metabolism. It mediates the transesterification of free cholesterol to cholesteryl ester in an apoprotein A-I-dependent process. We have isolated purified LCAT from human plasma using anion-exchange chromatography and characterized the extracted LCAT in terms of its molecular weight, molar absorption coefficient, and enzymatic activity. The participation of LCAT in the oxidation of very low density lipoproteins (VLDL) and low-density lipoproteins (LDL) was examined by supplementing lipoproteins with exogenous LCAT over a range of protein concentrations. LCAT-depleted lipoproteins were also prepared and their oxidation kinetics examined. Our results provide evidence for a dual role for LCAT in lipoprotein oxidation, whereby it acts in a dose-responsive manner as a potent pro-oxidant during VLDL oxidation, but as an antioxidant during LDL oxidation. We believe this novel pro-oxidant effect may be attributable to the LCAT-mediated formation of oxidized cholesteryl ester in VLDL, whereas the antioxidant effect is similar to that of chain-breaking antioxidants. Thus, we have demonstrated that the high-density lipoprotein-associated enzyme LCAT may have a significant role to play in lipoprotein modification and hence atherogenesis. (C) 2007 Elsevier Inc. All rights reserved.

The Role of Dignity in Equality Law: Lessons from Canada and South Africa

This paper examines the link made on occasion between the concept of dignity and substantive equality; it is further noted that dignity can have very different meanings in different contexts. While the notion of dignity does not often play a substantive role in the resolution of decisions, sometimes the underlying understanding of dignity does matter. However, in all cases, judges should avoid the temptation to rely on unarticulated value judgments or subjective notions of dignity. When judges make reference to dignity, they should articulate the values underpinning their conception of it.

Lanthanide(III) nitrobenzenesulfonates as new nitration catalysts: The role of the metal and of the counterion in the catalytic efficiency

Lanthanide(III) complexes of p-nitrobenzenesulfonic acid, Ln(p-NBSA)(3), m-nitrobenzenesulfonic acid, Ln(m-NBSA)(3), and 2,4-nitrobenzenesulfonic acid, Ln(2,4-NBSA)(3), were prepared, characterized and examined as catalyst for the nitration of benzene, toluene, xylenes, naphthalene, bromobenzene and chlorobenzene. The initial screening of the catalysts showed that lanthanum(III) complexes were more effective than the corresponding ytterbium(III) complexes, and that catalysts containing the bulky 2,4-NBSA ligand were less effective than the catalyst containing p-NBSA (nosylate) or m-NBSA ligands. Examination of a series of Ln(p-NBSA)(3) and Ln(m-NBSA)(3) catalysts revealed that there is a clear correlation between the ionic radii of the lanthanide(III) ions and the yields of nitration, with the lighter lanthanides being more effective. The X-ray single crystal structure of Yb(m-NBSA)(3).6H(2)O shows that two m-NBSA ligands are directly bound to the metal centre while the third ligand is not located in the first coordination sphere, but it is hydrogen bonded to one of the water molecules which is coordinated to ytterbium(III). NMR studies suggest that this structure is preserved under the conditions used in the nitration reaction. The structure of Yb(m-NBSA)(3) is markedly different from the structure of the well-known ytterbium(III) triflate catalyst. The coordination of the nitrobenzenesulfonate counterion to the lanthanide(III) ion suggests that steric effects might play an important role in determining the efficiency of these novel nitration catalysts. ((C) Wiley-VCH Verlag GmbH & Co. KGaA, 69451 Weinheim, Germany, 2004).

The Role of Process Parameters in Determining Wall Thickness Distribution in Plug-Assisted Thermoforming

This article describes the results of a comprehensive investigation to determine the link between process parameters and observed wall thickness output for the plug-assisted thermoforming process. The overall objective of the work was to systematically investigate the process parameters that may be adjusted during production to control the wall thickness distribution of parts manufactured by plug-assisted thermoforming. The parameters investigated were the sheet temperature, plug temperature, plug speed, plug displacement, plug shape, and air pressure. As well as quantifying the effects of each parameter on the wall thickness distribution, a further aim of the work was to improve the understanding of the physical mechanisms of deformation of the sheet during the different stages of the process. The process parameters shown to have the greatest effect on experimentally determined wall thickness distribution were the plug displacement, sheet temperature, plug temperature, and plug shape. It is proposed that during the plug-assisted thermoforming of polystyrene the temperature dependent friction between the plug and sheet surface was the most important factor in determining product wall thickness distribution, whereas heat transfer was shown to play a less important role. POLYM. ENG. SCI., 2010. © 2010 Society of Plastics Engineers

Regulation of OPA1-mediated mitochondrial fusion by leucine zipper/EF-hand-containing transmembrane protein-1 plays a role in apoptosis

Carboxyl-terminal modulator protein (CTMP) is a tumor suppressor-like binding partner of Protein kinase B (PKB/Akt) that negative regulates this kinase. In the course of our recent work, we identified that CTMP is consistently associated with leucine zipper/EF-hand-containing transmembrane-1 (LETM1). Here, we report that adenovirus-LETM1 increased the sensitivity of HeLa cells to apoptosis, induced by either staurosporine or actinomycin D. As shown previously, LETM1 localized to the inner mitochondrial membrane. Electron-microscopy analysis of adenovirus-LETM1 transduced cells revealed that mitochondrial cristae were swollen in these cells, a phenotype similar to that observed in optic atrophy type-1 (OPA1)-ablated cells. OPA1 cleavage was increased in LETM1-overexpressing cells, and this phenotype was reversed by overexpression of OPA1 variant-7, a cleavage resistant form of OPA1. Taken together, these data suggest that LETM1 is a novel binding partner for CTMP that may play an important role in mitochondrial fragmentation via OPA1-cleavage. (C) 2009 Elsevier Inc. All rights reserved

Differential expression of steroid 5 alpha-reductase isozymes and association with disease severity and angiogenic genes predict their biological role in prostate cancer

The biological role of steroid 5 alpha-reductase isozymes (encoded by the SRD5A1 and SRD5A2 genes) and angiogenic factors that play important roles in the pathogenesis and vascularization of prostate cancer (PC) is poorly understood. The sub-cellular expression of these isozymes and vascular endothelial growth factor (VEGF) in PC tissue microarrays (n=62) was examined using immunohistochemistry. The effect of SRD5A inhibition on the angiogenesis pathway genes in PC was also examined in prostate cell lines, LNCaP, PC3, and RWPE-1, by treating them with the SRD5A inhibitors finasteride and dutasteride, followed by western blot, quantitative PCR, and ELISA chip array techniques. In PC tissues, nuclear SRD5A1 expression was strongly associated with higher cancer Gleason scores (P=0.02), higher cancer stage (P=0.01), and higher serum prostate specific antigen (PSA) levels (P=0.01), whereas nuclear SRD5A2 expression was correlated with VEGF expression (P=0.01). Prostate tumor cell viability was significantly reduced in dutasteride-treated PC3 and RWPE-1 cells compared with finasteride-treated groups. Expression of the angiogenesis pathway genes transforming growth factor beta 1 (TGFB1), endothelin (EDN1), TGF alpha (TGFA), and VEGFR1 was upregulated in LNCaP cells, and at least 7 out of 21 genes were upregulated in PC3 cells treated with finasteride (25 mu M). Our findings suggest that SRD5A1 expression predominates in advanced PC, and that inhibition of SRD5A1 and SRD5A2 together was more effective in reducing cell numbers than inhibition of SRD5A2 alone. However, these inhibitors did not show any significant difference in prostate cell angiogenic response. Interestingly, some angiogenic genes remained activated after treatment, possibly due to the duration of treatment and tumor resistance to inhibitors. Endocrine-Related Cancer (2010) 17 757-770