Two-photon excitation with pico-second fluorescence lifetime imaging to detect nuclear association of flavanols

Two-photon excitation enabled for the first time the observation and measurement of excited state fluorescence lifetimes from three flavanols in solution, which were ∼1.0 ns for catechin and epicatechin, but <45 ps for epigallocatechin gallate (EGCG). The shorter lifetime for EGCG is in line with a lower fluorescence quantum yield of 0.003 compared to catechin (0.015) and epicatechin (0.018). In vivo experiments with onion cells demonstrated that tryptophan and quercetin, which tend to be major contributors of background fluorescence in plant cells, have sufficiently low cross sections for two-photon excitation at 630 nm and therefore do not interfere with detection of externally added or endogenous flavanols in Allium cepa or Taxus baccata cells. Applying two-photon excitation to flavanols enabled 3-D fluorescence lifetime imaging microscopy and showed that added EGCG penetrated the whole nucleus of onion cells. Interestingly, EGCG and catechin showed different lifetime behaviour when bound to the nucleus: EGCG lifetime increased from <45 to 200 ps, whilst catechin lifetime decreased from 1.0 ns to 500 ps. Semi-quantitative measurements revealed that the relative ratios of EGCG concentrations in nucleoli associated vesicles: nucleus: cytoplasm were ca. 100:10:1. Solution experiments with catechin, epicatechin and histone proteins provided preliminary evidence, via the appearance of a second lifetime (τ2 = 1.9–3.1 ns), that both flavanols may be interacting with histone proteins. We conclude that there is significant nuclear absorption of flavanols. This advanced imaging using two-photon excitation and biophysical techniques described here will prove valuable for probing the intracellular trafficking and functions of flavanols, such as EGCG, which is the major flavanol of green tea.

Synthesis and evaluation of lipophilic BTBP ligands for An/Ln separation in nuclear waste treatment: the effect of alkyl substitution on extraction properties and implications for ligand design

Four new 6,6′-bis(1,2,4-triazin-3-yl)-2,2′-bipyridine (BTBP) ligands, which contain either additional alkyl groups on the pyridine rings or seven-membered aliphatic rings attached to the triazine rings, have been synthesized, and the effects of the additional alkyl substitution in the 4- and 4′-positions of the pyridine rings on their extraction properties with LnIII and AnIII cations in simulated nuclear waste solutions have been studied. The speciation of ligand 13 with some trivalent lanthanide nitrates was elucidated by 1H NMR spectroscopic titrations and ESI-MS. Although 13 formed both 1:1 and 1:2 complexes with LaIII and YIII, only 1:2 complexes were observed with EuIII and CeIII. Quite unexpectedly, both alkyl-substituted ligands 12 and 13 showed lower solubilities in certain diluents than the unsubstituted ligand CyMe4-BTBP. Compared to CyMe4-BTBP, alkyl-substitution was found to decrease the rates of metal-ion extraction of the ligands in both 1-octanol and cyclohexanone. A highly efficient (DAm > 10) and selective (SFAm/Eu > 90) extraction was observed for 12 and 13 in cyclohexanone and for 13 in 1-octanol in the presence of a phase-transfer agent. The implications of these results for the design of improved extractants for radioactive waste treatment are discussed.

Supranutritional selenium induces alterations in molecular targets related to energy metabolism in skeletal muscle and visceral adipose tissue of pigs

While selenium (Se) is an essential micronutrient for humans, epidemiological studies have raised concern that supranutritional Se intake may increase the risk to develop Type 2 diabetes mellitus (T2DM). We aimed to determine the impact of Se at a dose and source frequently ingested by humans on markers of insulin sensitivity and signalling. Male pigs were fed either a Se-adequate (0.17 mg Se/kg) or a Se-supranutritional (0.50 mg Se/kg; high-Se) diet. After 16 weeks of intervention, fasting plasma insulin and cholesterol levels were non-significantly increased in the high-Se pigs, whereas fasting glucose concentrations did not differ between the two groups. In skeletal muscle of high-Se pigs, glutathione peroxidase activity was increased, gene expression of forkhead box O1 transcription factor and peroxisomal proliferator-activated receptor- coactivator 1 were increased and gene expression of the glycolytic enzyme pyruvate kinase was decreased. In visceral adipose tissue of high-Se pigs, mRNA levels of sterol regulatory element-binding transcription factor 1 were increased, and the phosphorylation of Akt, AMP-activated kinase and mitogen-activated protein kinases was affected. In conclusion, dietary Se oversupply may affect expression and activity of proteins involved in energy metabolism in major insulin target tissues, though this is probably not sufficient to induce diabetes.

Neuroprotective effects of phenolic antioxidant tBHQ associate with inhibition of FoxO3a nuclear translocation and activity

The Forkhead transcription factor, FoxO3a induces genomic death responses in neurones following translocation from the cytosol to the nucleus. Nuclear translocation of FoxO3a is triggered by trophic factor withdrawal, oxidative stress and the stimulation of extrasynaptic NMDA receptors. Receptor activation of phosphatidylinositol 3-kinase (PI3K) – Akt signalling pathways retains FoxO3a in the cytoplasm thereby inhibiting the transcriptional activation of death promoting genes. We hypothesised that phenolic antioxidants such as tert-Butylhydroquinone (tBHQ), which is known to stimulate PI3K-Akt signalling, would inhibit FoxO3a translocation and activity. Treatment of cultured cortical neurones with NMDA increased the nuclear localisation of FoxO3a, reduced the phosphorylation of FoxO3a, increased caspase activity and upregulated Fas ligand expression. In contrast the phenolic antioxidant tBHQ caused retention of FoxO3a in the cytosol coincident with enhanced PI3K- dependent phosphorylation of FoxO3a. tBHQ-induced nuclear exclusion of FoxO3a was associated with reduced FoxO-mediated transcriptional activity. Exposure of neurones to tBHQ inhibited NMDA-induced nuclear translocation of FoxO3a prevented NMDA-induced upregulation of FoxO-mediated transcriptional activity, blocked caspase activation and protected neurones from NMDA-induced excitotoxic death. Collectively, these data suggest that phenolic antioxidants such as tBHQ oppose stress-induced activation of FoxO3a and therefore have potential neuroprotective utility in neurodegeneration.

Nuclear fuel cycles: interfaces with the environment

The waste materials generated in the nuclear fuel cycle are very varied,ranging from the tailings arising from mining and processing uranium ore, depleted uranium in a range of chemical forms, to a range of process wastes of differing activities and properties. Indeed, the wastes generated are intimately linked to the options selected in operating the nuclear fuel cycle, most obviously to the management of spent fuel. An open fuel cycle implies the disposal of highly radioactive spent fuel, whereas a closed fuel cycle generates a complex array of waste streams. On the other hand, a closed fuel cycle offers options for waste management, for example reduction in highly active waste volume, decreased radiotoxicity, and removal of fissile material. Many technological options have been proposed or explored, and each brings its own particular mix of wastes and environmental challenges.

The Bomb: Nuclear Weapons in their Historical, Strategic and Ethical Context

This book examines to what extent the invention and first use of nuclear weapons was a turning point in the history of warfare and strategy(to what extent was it a mere continuation or perfection of air power strategy? Were the casualty numbers really unprecedented?), the ethics of war (was this form of war against civilians unprecedented?), and it asks whether it was an expression of total war or did it create total war

Nuclear Mentalities? Strategies and Beliefs in Britain, France and the FRG

This is volume II of my two-volume study of the nuclear strategies/strategy preferences of Britain, France and West Germany in the Cold War. It shows that the great variable is the mentalities, culture, and particular interpretations of particular historical experiences of these three countries. These mental frameworks are reconstructed, in the tradition of the French Annales/Mentalite schools (that in turn is rooted in anthropology) by looking at a very large nunber of public texts in the context of the public discourse about nuclear strategy in each country.

NATO, Britain, France and the FRG: Nuclear Strategies and Forces for Europe, 1949-2000

This is vol. I of my two-volume study of the nuclear strategies/strategy preferences of NATO collectively, and individually of Britain, France, and West Germany in the Cold War. It shows that NATO strategy was a fragile compromise, and that these three countries, all within range of Soviet medium/intermediate range nuclear missiles and thus with less geostrategic difference than in previous military threat contexts, had wildly divergent strategies/preferences which cannot be explained merely by geography. It raises the question of what made them so different, addressed in Volume II "Nuclear Mentalities" (q.v.)

‘Victory in a Nuclear War? A Comparison of NATO and WTO War Aims and Strategies’

Given the danger of the unbearable catastrophe of nuclear war, NATO in the mid-1950s abandoned any war aim of achieving victory in an all-out war with the Soviet Union or the Warsaw Treaty Organisation. Instead, it adopted the war aim of a cease-fire or war termination. By contrast, the WTO clung on to the war aim of victory - expressed even in terms of the survival of slightly more Soviet citizens than NATO citizens, after unprecedented losses of life in nuclear exchanges - until the mid-1980s when under Gorbachev the concept of victory in nuclear war was abandoned.

Beliefs, cultures, proliferation and use of nuclear weapons

This article demonstrates the centrality of mentality, culture, beliefs and historical lessons for nuclear prolifertion. Using the historical casestudies of Britain, France, and West Germany, it encourages researchers to look at the mentality/culture of potential proliferators rather than apply a culture-less "Realist" IR theory approach that assumes that desicion-makers the world over think like Bismarck.

‘What Price Solidarity? Nuclear Dependence in NATO, 1949-1967’

Archive-based study of the dependence of NATO member states on the USA as nuclear guarantor, and the problems this entailed.

‘European strategists and European identity in the nuclear age’

European writers on strategy (in French: strategistes, as opposed to practitioners, stratèges) developed their thoughts on the best strategies and postures of nuclear deterrence against their own beliefs in the identities of their own countries - were they seen as "Europesn" or as "nation-states" who must under no condition surrender their sovereignty?

‘Containing Uncertainty: Options for British Nuclear Strategy’

If Britain wants to stem the tide of nuclear proliferation, it must continue to assume "the nuclear man's burden" and guarantee the security of non-nuclear allies, as it did in the Cold War.