Operation and Utilisation of the High Flux Reactor: Annual Report 2013

The High Flux Reactor (HFR) at Petten is managed by the Institute for Energy and Transport (IET) of the European Commission's Joint Research Centre (JRC) and operated by the Nuclear Research and consultancy Group (NRG) which is also the licence holder and responsible for its commercial activities. The High Flux Reactor (HFR) operates at 45 MW and is of the tank-in-pool type, light water cooled and moderated. It is one of the most powerful multi-purpose materials testing reactors in the world and one of the world's leaders in target irradiation for the production of medical radioisotopes.JRC.F.4-Innovative Technologies for Nuclear Reactor Safet

Rab11 and phosphoinositides: A synergy of signal transducers in the control of vesicular trafficking

AbstractRab11 and phosphoinositides are signal transducers able to direct the delivery of membrane components to the cell surface. Rab11 is a small GTPase that, by cycling from an active to an inactive state, controls key events of vesicular transport, while phosphoinositides are major determinants of membrane identity, modulating compartmentalized small GTPase function. By sharing common effectors, these two signal transducers synergistically direct vesicular traffic to specific intracellular membranes. This review focuses on the latest advances regarding the mechanisms that ensure the compartmentalized regulation of Rab11 function through its interaction with phosphoinositides

Effect of angiotensin-converting enzyme inhibitor and angiotensin receptor blocker initiation on organ support-free days in patients hospitalized with COVID-19

IMPORTANCE Overactivation of the renin-angiotensin system (RAS) may contribute to poor clinical outcomes in patients with COVID-19. Objective To determine whether angiotensin-converting enzyme (ACE) inhibitor or angiotensin receptor blocker (ARB) initiation improves outcomes in patients hospitalized for COVID-19. DESIGN, SETTING, AND PARTICIPANTS In an ongoing, adaptive platform randomized clinical trial, 721 critically ill and 58 non–critically ill hospitalized adults were randomized to receive an RAS inhibitor or control between March 16, 2021, and February 25, 2022, at 69 sites in 7 countries (final follow-up on June 1, 2022). INTERVENTIONS Patients were randomized to receive open-label initiation of an ACE inhibitor (n = 257), ARB (n = 248), ARB in combination with DMX-200 (a chemokine receptor-2 inhibitor; n = 10), or no RAS inhibitor (control; n = 264) for up to 10 days. MAIN OUTCOMES AND MEASURES The primary outcome was organ support–free days, a composite of hospital survival and days alive without cardiovascular or respiratory organ support through 21 days. The primary analysis was a bayesian cumulative logistic model. Odds ratios (ORs) greater than 1 represent improved outcomes. RESULTS On February 25, 2022, enrollment was discontinued due to safety concerns. Among 679 critically ill patients with available primary outcome data, the median age was 56 years and 239 participants (35.2%) were women. Median (IQR) organ support–free days among critically ill patients was 10 (–1 to 16) in the ACE inhibitor group (n = 231), 8 (–1 to 17) in the ARB group (n = 217), and 12 (0 to 17) in the control group (n = 231) (median adjusted odds ratios of 0.77 [95% bayesian credible interval, 0.58-1.06] for improvement for ACE inhibitor and 0.76 [95% credible interval, 0.56-1.05] for ARB compared with control). The posterior probabilities that ACE inhibitors and ARBs worsened organ support–free days compared with control were 94.9% and 95.4%, respectively. Hospital survival occurred in 166 of 231 critically ill participants (71.9%) in the ACE inhibitor group, 152 of 217 (70.0%) in the ARB group, and 182 of 231 (78.8%) in the control group (posterior probabilities that ACE inhibitor and ARB worsened hospital survival compared with control were 95.3% and 98.1%, respectively). CONCLUSIONS AND RELEVANCE In this trial, among critically ill adults with COVID-19, initiation of an ACE inhibitor or ARB did not improve, and likely worsened, clinical outcomes. TRIAL REGISTRATION ClinicalTrials.gov Identifier: NCT0273570

Ceroid-lipofuscinosis (Batten disease) : a thesis presented in partial fulfilment of the requirements for the degree of Doctor of Philosophy at Massey University

The ceroid-lipofuscinoses are a group of inherited neurodegenerative diseases occurring in human beings and animals. Histologically there is generalised accumulation of a fluorescent lipopigment within the cytoplasm of many cell types and selective necrosis of some populations of neurons. Clinical signs include loss of vision, seizures and mental retardation with premature death being the eventual outcome. Systematic analyses of isolated storage material have previously resulted in the identification of subunit c of mitochondrial ATP synthase as being a major constituent in the late infantile human, ovine, bovine and three canine forms of ceroid-lipofuscinosis. Saposins A and D have been identified as being stored in the infantile human disease. Brain biopsy has been routinely used to diagnose ceroid-lipofuscinosis in sheep. The efficacy of this technique was investigated and it was concluded that it was a safe and reliable method for diagnosis in lambs aged 2 1/2 months. In contrast, diagnosis of the disease by clinical examination could only be made comparatively late in the disease course at 9-12 months. Repeated neurological examinations enabled "clinical staging" of the progression of the disease. Haematopoietic cell transplantation was carried out in foetal lambs with ceroid-lipofuscinosis. Stable engraftment of an average of 9% blood cells was achieved but under the conditions of the experiment there was no alteration in the disease course or severity of lesions in transplanted lambs with ceroid-lipofuscinosis as compared to affected lambs without transplanted cells. The observation of autofluorescence from storage cytosomes in the ceroid-lipofuscinoses has lead to the assumption that they contain a fluorophore of critical significance to explanations of pathogenenesis. Studies on the nature of the fluorescence from storage bodies and isolated storage body components allowed the conclusion that no single significant fluorophore other than protein was present. Antibodies to subunit c of mitochondrial ATP synthase were produced. These and similar antibodies from other sources were used for immunocytochemistry. The staining pattern observed varied, depending on the fixation regime, the antibody used and the form of disease. It was concluded that different epitopes were exposed in different forms of the ceroid-lipofuscinoses. Positive immunostaining of storage material in muscle and cartilage from the ear depended on the age of the patient and could assist but not replace current methods of diagnosis. Storage cytosomes were also labelled using immunogold staining at the ultrastructural level. The ceroid-lipofuscinoses are a genetically diverse group of diseases which appear to have complex biochemical systems underlying them. The hypothesis was developed that the defect may lie in the disassembly of the F0 complex rather than in proteolysis per se. The aggregation of subunit c with lipids could result in a complex structure resistant to catabolism by proteases

Filling a hole in cytochrome P450 BM3 improves substrate binding and catalytic efficiency

Cytochrome P450BM3 (CYP102A1) from Bacillus megaterium, a fatty acid hydroxylase, is a member of a very large superfamily of monooxygenase enzymes. The available crystal structures of the enzyme show non-productive binding of substrates with their ω-end distant from the iron in a hydrophobic pocket at one side of the active site. We have constructed and characterised mutants in which this pocket is filled by large hydrophobic sidechains replacing alanine at position 82. The mutants having phenylalanine or tryptophan at this position have very much (~800-fold) greater affinity for substrate, with a greater conversion of the haem iron to the high-spin state, and similarly increased catalytic efficiency. The enzyme as isolated contains bound palmitate, reflecting this much higher affinity. We have determined the crystal structure of the haem domain of the Ala82Phe mutant with bound palmitate; this shows that the substrate is binding differently from the wild-type enzyme but still distant from the haem iron. Detailed analysis of the structure indicates that the tighter binding in the mutant reflects a shift in the conformational equilibrium of the substrate-free enzyme towards the conformation seen in the substrate complex rather than differences in the enzyme-substrate interactions. On this basis, we outline a sequence of events for the initial stages of the catalytic cycle. The Ala82Phe and Ala82Trp mutants are also very much more effective catalysts of indole hydroxylation than the wild-type enzyme, suggesting that they will be valuable starting points for the design of mutants to catalyse synthetically useful hydroxylation reactions

Three C-terminal residues from the sulphonylurea receptor contribute to the functional coupling between the KATP channel subunits SUR2A and Kir6.2

Cardiac ATP-sensitive potassium (KATP) channels are metabolic sensors formed by the association of the inward rectifier potassium channel Kir6.2 and the sulphonylurea receptor SUR2A. SUR2A adjusts channel gating as a function of intracellular ATP and ADP and is the target of pharmaceutical openers and blockers which, respectively, up- and down-regulate Kir6.2. In an effort to understand how effector binding to SUR2A translates into Kir6.2 gating modulation, we examined the role of a 65-residue SUR2A fragment linking transmembrane domain TMD2 and nucleotide-binding domain NBD2 that has been shown to interact with Kir6.2. This fragment of SUR2A was replaced by the equivalent residues of its close homologue, the multidrug resistance protein MRP1. The chimeric construct was expressed in Xenopus oocytes and characterized using the patch-clamp technique. We found that activation by MgADP and synthetic openers was greatly attenuated although apparent affinities were unchanged. Further chimeragenetic and mutagenetic studies showed that mutation of three residues, E1305, I1310 and L1313 (rat numbering), was sufficient to confer this defective phenotype. The same mutations had no effects on channel block by the sulphonylurea glibenclamide or by ATP, suggesting a role for these residues in activatory – but not inhibitory – transduction processes. These results indicate that, within the KATP channel complex, the proximal C-terminal of SUR2A is a critical link between ligand binding to SUR2A and Kir6.2 up-regulation