rom A Quantitative Tri-fluorescent Yeast Two-hybrid System: From Flow Cytometry to In cellula Affinities [Technological Innovation and Resources] By feedproxy.google.com Published On :: 2020-04-01T00:05:32-07:00 We present a technological advancement for the estimation of the affinities of Protein-Protein Interactions (PPIs) in living cells. A novel set of vectors is introduced that enables a quantitative yeast two-hybrid system based on fluorescent fusion proteins. The vectors allow simultaneous quantification of the reaction partners (Bait and Prey) and the reporter at the single-cell level by flow cytometry. We validate the applicability of this system on a small but diverse set of PPIs (eleven protein families from six organisms) with different affinities; the dissociation constants range from 117 pm to 17 μm. After only two hours of reaction, expression of the reporter can be detected even for the weakest PPI. Through a simple gating analysis, it is possible to select only cells with identical expression levels of the reaction partners. As a result of this standardization of expression levels, the mean reporter levels directly reflect the affinities of the studied PPIs. With a set of PPIs with known affinities, it is straightforward to construct an affinity ladder that permits rapid classification of PPIs with thus far unknown affinities. Conventional software can be used for this analysis. To permit automated analysis, we provide a graphical user interface for the Python-based FlowCytometryTools package. Full Article
rom Arginine in C9ORF72 Dipolypeptides Mediates Promiscuous Proteome Binding and Multiple Modes of Toxicity [Research] By feedproxy.google.com Published On :: 2020-04-01T00:05:32-07:00 C9ORF72-associated Motor Neuron Disease patients feature abnormal expression of 5 dipeptide repeat (DPR) polymers. Here we used quantitative proteomics in a mouse neuronal-like cell line (Neuro2a) to demonstrate that the Arg residues in the most toxic DPRS, PR and GR, leads to a promiscuous binding to the proteome compared with a relative sparse binding of the more inert AP and GA. Notable targets included ribosomal proteins, translation initiation factors and translation elongation factors. PR and GR comprising more than 10 repeats appeared to robustly stall on ribosomes during translation suggesting Arg-rich peptide domains can electrostatically jam the ribosome exit tunnel during synthesis. Poly-GR also recruited arginine methylases, induced hypomethylation of endogenous proteins, and induced a profound destabilization of the actin cytoskeleton. Our findings point to arginine in GR and PR polymers as multivalent toxins to translation as well as arginine methylation that may explain the dysfunction of biological processes including ribosome biogenesis, mRNA splicing and cytoskeleton assembly. Full Article
rom Improving Identification of In-organello Protein-Protein Interactions Using an Affinity-enrichable, Isotopically Coded, and Mass Spectrometry-cleavable Chemical Crosslinker [Research] By feedproxy.google.com Published On :: 2020-04-01T00:05:32-07:00 An experimental and computational approach for identification of protein-protein interactions by ex vivo chemical crosslinking and mass spectrometry (CLMS) has been developed that takes advantage of the specific characteristics of cyanurbiotindipropionylsuccinimide (CBDPS), an affinity-tagged isotopically coded mass spectrometry (MS)-cleavable crosslinking reagent. Utilizing this reagent in combination with a crosslinker-specific data-dependent acquisition strategy based on MS2 scans, and a software pipeline designed for integrating crosslinker-specific mass spectral information led to demonstrated improvements in the application of the CLMS technique, in terms of the detection, acquisition, and identification of crosslinker-modified peptides. This approach was evaluated on intact yeast mitochondria, and the results showed that hundreds of unique protein-protein interactions could be identified on an organelle proteome-wide scale. Both known and previously unknown protein-protein interactions were identified. These interactions were assessed based on their known sub-compartmental localizations. Additionally, the identified crosslinking distance constraints are in good agreement with existing structural models of protein complexes involved in the mitochondrial electron transport chain. Full Article
rom Quantitative Profiling of the Human Substantia Nigra Proteome from Laser-capture Microdissected FFPE Tissue [Research] By feedproxy.google.com Published On :: 2020-05-01T00:05:26-07:00 Laser-capture microdissection (LCM) allows the visualization and isolation of morphologically distinct subpopulations of cells from heterogeneous tissue specimens. In combination with formalin-fixed and paraffin-embedded (FFPE) tissue it provides a powerful tool for retrospective and clinically relevant studies of tissue proteins in a healthy and diseased context. We first optimized the protocol for efficient LCM analysis of FFPE tissue specimens. The use of SDS containing extraction buffer in combination with the single-pot solid-phase-enhanced sample preparation (SP3) digest method gave the best results regarding protein yield and protein/peptide identifications. Microdissected FFPE human substantia nigra tissue samples (~3,000 cells) were then analyzed, using tandem mass tag (TMT) labeling and LC-MS/MS, resulting in the quantification of >5,600 protein groups. Nigral proteins were classified and analyzed by abundance, showing an enrichment of extracellular exosome and neuron-specific gene ontology (GO) terms among the higher abundance proteins. Comparison of microdissected samples with intact tissue sections, using a label-free shotgun approach, revealed an enrichment of neuronal cell type markers, such as tyrosine hydroxylase and alpha-synuclein, as well as proteins annotated with neuron-specific GO terms. Overall, this study provides a detailed protocol for laser-capture proteomics using FFPE tissue and demonstrates the efficiency of LCM analysis of distinct cell subpopulations for proteomic analysis using low sample amounts. Full Article
rom Australian Federal Police walk away from $145 million Israeli crime-fighting software deal By www.smh.com.au Published On :: Wed, 19 Aug 2015 06:25:36 GMT Police walk away from deal with contractor, conceding numerous issues have put project beyond rescue. Full Article
rom From making scarves to building a $165 million tech start-up: Canva's Melanie Perkins By www.smh.com.au Published On :: Tue, 06 Oct 2015 10:42:33 GMT To say it has been a wild ride for Canva founder and CEO Melanie Perkins would be an understatement. Full Article
rom From AFL star to Big Apple start-up, Swift's Joel MacDonald is kicking goals By www.smh.com.au Published On :: Fri, 13 Nov 2015 05:53:09 GMT Two years ago Joel MacDonald was in Melbourne playing in the AFL; now he's kicking goals in New York. Full Article
rom Troubled myGov website to be taken from Human Services and given to Digital Transformation Office for streamlining By www.smh.com.au Published On :: Thu, 28 Jan 2016 12:30:00 GMT Malcolm Turnbull's DTO has been critical of myGov, now it has the chance to show it can do better. Full Article
rom Malcolm Turnbull promises $50 million reboot for troubled myGov By www.smh.com.au Published On :: Tue, 21 Jun 2016 22:06:13 GMT Takeover of troubled portal by Digital Transformation Office confirmed Full Article
rom Tax time in danger from ATO's tech wreck By www.smh.com.au Published On :: Tue, 07 Feb 2017 13:15:00 GMT IT projects thrown overboard as ATO orders all hands to keep tax time afloat. Full Article
rom Cyber security threat: Is Australia's power grid safe from hackers? By www.smh.com.au Published On :: Fri, 10 Nov 2017 13:15:00 GMT Cyber attacks have labelled the number one threat to power and utility companies worldwide, a new EY report has found. Full Article
rom FIFA 18 and Harry Potter play rescued from bots by Queensland start-up By www.smh.com.au Published On :: Sun, 31 Dec 2017 02:16:04 GMT The FIFA soccer game and Harry Potter and the Cursed Child have been protected from bots thanks to a Queensland start-up. Full Article
rom How to Fight the Economic Fallout From the Coronavirus By feedproxy.google.com Published On :: Wed, 04 Mar 2020 03:56:03 +0000 4 March 2020 Creon Butler Research Director, Trade, Investment & New Governance Models: Director, Global Economy and Finance Programme LinkedIn Finance ministries and central banks have a critical role to play to mitigate the threat Covid-19 poses to the global economy. 2020-03-03-TokyoCV.jpg A pedestrian wearing a face mask walks past stock prices in Tokyo on 25 February. Photo: Getty Images. Epidemics, of the size of Covid-19, have huge economic impacts – not just from the costs of managing the health of people, but stopping them, and keeping the economy working. The 10% fall in global stock markets since it became clear that Covid-19 would not be limited to China has boldly highlighted this.Suppressing the epidemic, but allowing the economy to still function, requires key decisions, in which central banks and finance ministries play a part.The role of fiscal and monetary authorities in managing an epidemic economyThe scope to use monetary policy to manage the economic impact of Covid-19 is limited. The fact that the underlying cause of the shock is an infectious disease outbreak (rather than a banking crisis, as in 2008-09) and nominal interest rates are currently close to zero in most major advanced economies reduces the effectiveness of monetary policy.Since 2010, reductions in fiscal deficits mean there is more scope for supportive fiscal action. But even here, high public debt levels and the desire not to underwrite ‘zombie’ companies that may have been sustained by a decade of ultra-low interest rates remain constraints. However, outside broad based fiscal and monetary policies there are six ways in which finance ministries and central banks will play a critical role in responding to the crisis.A first crucial role for finance ministries and central banks is in helping provide the best possible economic evaluation of strict containment measures (trying to isolate each potential case) versus managing the epidemic (delaying the spread of the virus, protecting the most vulnerable and treating the sick, while enabling the majority of people to get on with daily life). Given the economic consequences, they must play a full part, alongside health experts, in advising political leaders on this key decision.Second, if large numbers of staff are required to work from home to manage the epidemic, they have the lead role in doing whatever is necessary to ensure that financial markets – and thus the wider economy – will continue to function smoothly.Third, they need to ensure adequate funding for the public health response. Steps that can make an enormous difference to the success of containment strategies, such as strengthening surveillance, and guaranteeing the availability of testing kits and protective equipment for front line health workers, must not fail because of a lack of funding. Fourth, they have a lead role in designing targeted economic interventions for the wider economy. Some of these are needed immediately to re-enforce and incentivize strict containment strategies, such as ensuring that employees without full or adequate sick leave cover have the financial support to enable them to report and self-isolate when they get sick. Other interventions may help improve the resilience of the economy in accommodating moderate ‘social distancing’ measures; for example, by providing assistance to small firms to help them gear up for home working.Yet others are needed, as a contingency, to safeguard the most vulnerable sectors (such as tourism, retail and transport) in circumstances where there is a prolonged downturn. The latter may include schemes to allow deferral of tax payments by SMEs, or steps to encourage loan extensions and other forms of liquidity support from the banking system, or by moves to underwrite continued provision of business insurance.Fifth, national economic authorities will need to play their part in combatting ‘fake news’ through providing transparent and high-quality analysis. This includes providing forecasts on the likely economic impact of the virus under different scenarios, but also detailed information on the support and contingency measures they are considering, so they can be improved and refined through feedback. Sixth, they will need to ensure that there is generous international support for poor countries, by ensuring the available multilateral support facilities from the international financial institutions and multilateral development banks are adequately funded and fit for purpose. The World Bank has already announced an initial $12 billion financing package, but much more is likely to be needed.They also need to support coordinated bilateral aid where this is more effective, as well as special measures to support particularly vulnerable groups, for example, in refugee camps and prisons. Given the importance of distributing sophisticated medical equipment and expertise quickly, it is also important that every effort is made to avoid delays due to customs and migration checks.Managing the futureThe response to the immediate crisis will rightly take priority now, but economic authorities must also play their part in ensuring the world finally takes decisive steps to prevent a repeat of Covid-19 in future.The experience with SARS, H1N1 and Ebola shows that, while some progress is made after each outbreak, this is often not sustained. This epidemic shows that managing diseases is absolutely critical to the long-term health of global economy, and doubly so in circumstances where traditional central bank and finance ministry tools for dealing with major global economic shocks are limited.Finance ministries and central banks therefore need to push hard within government to ensure sustained long-term funding of research on prevention and strengthening of public health systems. They also need to ensure that the right lessons are drawn by the private sector on making international supply chains more robust.Critical to the overall success of the economic effort will be effective international coordination. The G20 was established as the premier economic forum for international economic cooperation in 2010, and global health issues have been a substantive part of the G20 agenda since the 2017 Hamburg Summit. At the same time, G7 finance ministers and deputies remain one of the most effective bodies for managing economic crises on a day-to-day basis and should continue this within the framework provided by the G20.However, to be effective, the US, as current president of the G7, will need to put aside its reservations on multilateral economic cooperation and working with China to provide strong leadership. Full Article
rom Emerging Lessons From COVID-19 By feedproxy.google.com Published On :: Thu, 02 Apr 2020 14:49:54 +0000 2 April 2020 Jim O'Neill Chair, Chatham House Exploring what lessons can be learned from the crisis to improve society and the functioning of our economic model going forward. 2020-04-02-COVID-Italy A man with a protective mask by the Coliseum in Rome during the height of Italy's COVID-19 epidemic. Photo by ALBERTO PIZZOLI/AFP via Getty Images. As tentative evidence emerges that Italy and Spain may have reached - or are close to - the peak of the curve, this could demonstrate that not only can Asian countries get to grips with COVID-19, but so can western democracies. And, if so, this offers a path for the rest of us.The last few weeks does demonstrate there is a role for governments to intervene in society, whether it be health, finance or any walk of life, as they have had to implement social distancing. Some have been forced, and the interventions are almost definitely only temporary, but perhaps some others may be less so.Governments of all kinds now realise there is a connection between our health system quality and our economic capability. On an index of global economic sustainability that I presided over creating when I was at Goldman Sachs, the top ten best performing countries on growth environment scores includes eight of the best performing ten countries - so far- in handling the crisis in terms of deaths relative to their population.Health system qualityThe top three on the index (last calculated in 2014) were Singapore, Hong Kong and South Korea, all of which are exemplary to the rest of us on how to deal with this mess. This suggests that once we are through this crisis, a number of larger populated countries - and their international advisors such as the IMF - might treat the quality of countries' health systems just as importantly as many of the other more standard indicators in assessing ability to deal with shocks.Policymakers have also been given a rather stark warning about other looming health disasters, especially antimicrobial resistance, of which antibiotic resistance lies at the heart. An independent review I chaired recommended 29 interventions, requiring $42 bn worth of investment, essentially peanuts compared to the costs of no solution, and the current economic collapse from COVID-19. It would seem highly likely to me that policymakers are going to treat this more seriously now.As a clear consequence of the - hopefully, temporary - global economic collapse, our environment suddenly seems to be cleaner and fresher and, in this regard, we have bought some time in the battle against climate change. Surely governments are going to be able to have a bigger influence on fossil fuel extractors and intense users as we emerge from this crisis?For any industries requiring government support, the government can make it clear this is dependent on certain criteria. And surely the days of excessive use of share buy backs and extreme maximisation of profit at the expense of other goals, are over?It seems to me an era of 'optimisation' of a number of business goals is likely to be the mantra, including profits but other things too such as national equality especially as it relates to income. Here in the UK, the government has offered its strongest fiscal support to the lower end of the income earning range group and, in a single swoop, has presided over its most dramatic step towards narrowing income inequality for a long time.This comes on top of a period of strong initiatives to support higher levels of minimum earnings, meaning we will emerge later in 2020, into 2021, and beyond, with lower levels of income inequality.The geographic issue of rural versus urban is also key. COVID-19 has spread more easily in more tightly packed cities such as London, New York and many others. More geographically remote places, by definition, are better protected. Perhaps now there will be some more thought given by policymakers to the quality and purpose of life outside our big metropolitan areas.Lastly, will China emerge from this crisis by offering a mammoth genuine gesture to the rest of the world, and come up, with, unlike, in 2008, a fiscal stimulus to its own consumers, that is geared towards importing a lot of things from the rest of the world? Now that would be good way of bringing the world back together again.This is a version of an article originally published in The Article Full Article
rom A Direct Fluorometric Activity Assay for Lipid Kinases and Phosphatases By feedproxy.google.com Published On :: 2020-04-27 Jiachen SunApr 27, 2020; 0:jlr.D120000794v1-jlr.D120000794Methods Full Article
rom An LC/MS/MS method for analyzing the steroid metabolome with high accuracy and from small serum samples By feedproxy.google.com Published On :: 2020-04-01 Teng-Fei YuanApr 1, 2020; 61:580-586Methods Full Article
rom Serum non-esterified fatty acids have utility as dietary biomarkers of fat intake from fish, fish oil and dairy in women By feedproxy.google.com Published On :: 2020-03-31 Sandi M. AzabMar 31, 2020; 0:jlr.D120000630v1-jlr.D120000630Methods Full Article
rom Metabolic phospholipid labeling of intact bacteria enables a fluorescence assay that detects compromised outer membranes By feedproxy.google.com Published On :: 2020-03-10 Inga NilssonMar 10, 2020; 0:jlr.RA120000654v1-jlr.RA120000654Research Articles Full Article
rom The fatty acids from LPL-mediated processing of triglyceride-rich lipoproteins are taken up rapidly by cardiomyocytes By feedproxy.google.com Published On :: 2020-04-02 Haibo JiangApr 2, 2020; 0:jlr.ILR120000783v1-jlr.ILR120000783Images in Lipid Research Full Article
rom WITHDRAWN: Structural and mechanistic studies of hydroperoxide conversions catalyzed by a CYP74 clan epoxy alcohol synthase from amphioxus (Branchiostoma floridae) [Research Articles] By feedproxy.google.com Published On :: 2014-03-04T09:59:12-08:00 This manuscript has been withdrawn by the Author. Full Article
rom Metabolic phospholipid labeling of intact bacteria enables a fluorescence assay that detects compromised outer membranes [Research Articles] By feedproxy.google.com Published On :: 2020-03-10T09:30:25-07:00 Gram-negative bacteria possess an asymmetric outer membrane (OM) composed primarily of lipopolysaccharides (LPS) on the outer leaflet and phospholipids (PLs) on the inner leaflet. Loss of this asymmetry due to mutations in the lipopolysaccharide (LPS) biosynthesis or transport pathways causes externalization of PLs to the outer leaflet of the OM and leads to OM permeability defects. Here, we employed metabolic labeling to detect a compromised OM in intact bacteria. Phosphatidylcholine synthase (Pcs) expression in Escherichia coli allowed for incorporation of exogenous propargylcholine (PCho) into phosphatidyl(propargyl)choline (PPC) and for incorporation of exogenous 1-azidoethyl-choline (AECho) into phosphatidyl(azidoethyl)choline (AEPC) as confirmed by LC-MS analyses. A fluorescent copper-free click reagent poorly labeled AEPC in intact wild-type cells, but readily labeled AEPC from lysed cells. Fluorescence microscopy and flow cytometry analyses confirmed the absence of significant AEPC labeling from intact wild-type E. coli strains, and revealed significant AEPC labeling in an E. coli LPS transport mutant (lptD4213) and an LPS biosynthesis mutant (E. coli lpxC101). Our results suggest that metabolic PL labeling with AECho is a promising tool to detect a compromised bacterial OM, reveal aberrant PL externalization, and identify or characterize novel cell-active inhibitors of LPS biosynthesis or transport. Full Article
rom Serum non-esterified fatty acids have utility as dietary biomarkers of fat intake from fish, fish oil and dairy in women [Methods] By feedproxy.google.com Published On :: 2020-03-31T09:30:24-07:00 Nutritional studies rely on various biological specimens for fatty acid (FA) determination, yet it is unclear how levels of serum non-esterified FA (NEFAs) correlate with other circulating lipid pools. Here, we used a high throughput method (< 4 min/sample) based on multisegment injection-non-aqueous-capillary electrophoresis–mass spectrometry (MSI-NACE-MS) to investigate whether specific serum NEFAs have utility as biomarkers of dietary fat intake in women. We first identified circulating NEFAs correlated with long-term/habitual food intake among pregnant women with contrasting dietary patterns (n=50). Acute changes in serum NEFA trajectories were also studied in non-pregnant women (n=18) following high-dose (5 g/day) fish oil (FO) supplementation or isoenergetic sunflower oil placebo over 56 days. In the cross-sectional study, serum omega-3 (-3) FA correlated with self-reported total -3 daily intake, notably eicosapentaenoic acid (EPA) as its NEFA (r=0.46; p=0.001), whereas pentadecanoic acid was associated with full-fat dairy intake (r=0.43; p=0.002), outcomes consistent with results from total FA serum hydrolysates. In the intervention cohort, serum -3 NEFAs increased 2.5-fold from baseline within 28 days following FO supplementation, and this increase was most pronounced for EPA (p=0.0004). Unlike for docosahexaenoic acid, circulating EPA as its NEFA also strongly correlated to EPA concentrations measured from erythrocyte phospholipid hydrolysates (r=0.66; p=4.6 x 10-10), and was better suited to detect dietary non-adherence. We conclude that MSI-NACE-MS offers a rapid method to quantify serum NEFAs and objectively monitor dietary fat intake in women that is complementary to diet records or food frequency questionnaires. Full Article
rom The fatty acids from LPL-mediated processing of triglyceride-rich lipoproteins are taken up rapidly by cardiomyocytes [Images in Lipid Research] By feedproxy.google.com Published On :: 2020-04-02T09:30:25-07:00 Full Article
rom A Direct Fluorometric Activity Assay for Lipid Kinases and Phosphatases [Methods] By feedproxy.google.com Published On :: 2020-04-27T13:30:26-07:00 Lipid kinases and phosphatases play key roles in cell signaling and regulation, and are implicated in many human diseases, and are hence thus attractive targets for drug development. Currently, no direct in vitro activity assay is available for these important enzymes, which hampers mechanistic studies as well as high-throughput screening of small molecule modulators. Here we report a highly sensitive and quantitative assay employing a ratiometric fluorescence sensor that directly and specifically monitors the real-time concentration change of a single lipid species. Due Because of to its modular design, the assay system can be applied to a wide variety of lipid kinases and phosphatases, including Class I phosphoinositide 3-kinase (PI3K) and phosphatase and tensin homolog (PTEN). When applied to PI3K, the assay provided the newdetailed mechanistic information about the product inhibition and substrate acyl acyl-chain selectivity of PI3K and allowed enabled rapid evaluation of its small molecule inhibitors. We also used this assay to quantitatively determine the substrate specificity of PTEN, providing new insight into its physiological functionThe assay also quantitatively determined the substrate specificity of PTEN, thereby providing new insight into its physiological function. In summary, we have developed a fluorescence-based real-time assay for PI3K and PTEN that we anticipate could be adapted to measure the activities of other lipid kinases and phosphatases with high sensitivity and accuracy. Full Article
rom Mass spectrometry imaging and LC-MS reveal decreased cerebellar phosphoinositides in Niemann-Pick type C1-null mice [Research Articles] By feedproxy.google.com Published On :: 2020-05-05T13:30:26-07:00 Niemann-Pick disease, type C1 (NPC1) is a lipid storage disorder in which cholesterol and glycosphingolipids accumulate in late endosomal/lysosomal compartments because of mutations in the NPC1 gene. A hallmark of NPC1 is progressive neurodegeneration of the cerebellum as well as visceral organ damage; however, the mechanisms driving this disease pathology are not fully understood. Phosphoinositides are phospholipids that play distinct roles in signal transduction and vesicle trafficking. Here, we utilized consensus spectra analysis of MS imaging datasets and orthogonal LC–MS analyses to evaluate the spatial distribution of phosphoinositides and quantify them in cerebellar tissue from Npc1-null mice. Our results suggest significant depletion of multiple phosphoinositide species, including phosphatidylinositol (PI), phosphatidylinositol monophosphate (PIP), and bisphosphate (PIP2), in the cerebellum of the Npc1-null mice in both whole-tissue lysates and myelin-enriched fractions. Additionally, we observed altered levels of the regulatory enzyme phosphatidylinositol 4-kinase type 2 α (PI4K2A) in Npc1-null mice. In contrast, the levels of related kinases, phosphatases, and transfer proteins were unaltered in the Npc1-null mouse model as observed by Western blot analysis. Our discovery of phosphoinositide lipid biomarkers for NPC1 opens new perspectives on the pathophysiology underlying this fatal neurodegenerative disease. Full Article
rom Let's Emerge From COVID-19 with Stronger Health Systems By feedproxy.google.com Published On :: Thu, 26 Mar 2020 09:33:28 +0000 26 March 2020 Robert Yates Director, Global Health Programme; Executive Director, Centre for Universal Health @yates_rob Heads of state should grasp the opportunity to become universal health heroes to strengthen global health security 2020-03-26-Health-Protest A "Big Insurance: Sick of It" rally in New York City. Photo by Mario Tama/Getty Images. As the COVID-19 pandemic presents the greatest threat to human health in over a century, people turn to their states to resolve the crisis and protect their health, their livelihoods and their future well-being.How leaders perform and respond to the pandemic is likely to define their premiership - and this therefore presents a tremendous opportunity to write themselves into the history books as a great leader, rescuing their people from a crisis. Just as Winston Churchill did in World War Two.Following Churchill’s advice to “never let a good crisis go to waste”, if leaders take decisive action now, they may emerge from the COVID-19 crisis as a national hero. What leaders must do quickly is to mitigate the crisis in a way which has a demonstrable impact on people’s lives.Given the massive shock caused by the pandemic to economies across the world, it is not surprising that heads of state and treasury ministers have implemented enormous economic stimulus packages to protect businesses and jobs – this was to be expected and has been welcome.National heroes can be madeBut, in essence, this remains primarily a health crisis. And one obvious area for leaders to act rapidly is strengthening their nation’s health system to stop the spread of the virus and successfully treat those who have fallen sick. It is perhaps here that leaders have the most to gain - or lose - and where national heroes can be made.This is particularly the case in countries with weak and inequitable health systems, where the poor and vulnerable often fail to access the services they need. One major practical action that leaders can implement immediately is to launch truly universal, publicly-financed health reforms to cover their entire population – not only for COVID-19 services but for all services.This would cost around 1-2% GDP in the short-term but is perfectly affordable in the current economic climate, given some of the massive fiscal stimuluses already being planned (for example, the UK is spending 15% GDP to tackle COVID-19).Within one to two years, this financing would enable governments to implement radical supply side reforms including scaling up health workforces, increasing the supply of essential medicines, diagnostics and vaccines and building new infrastructure. It would also enable them to remove health service user fees which currently exclude hundreds of millions of people worldwide from essential healthcare. Worldwide these policies have proven to be effective, efficient, equitable and extremely popular.And there is plenty of precedent for such a move. Universal health reform is exactly what political leaders did in the UK, France and Japan as post-conflict states emerging from World War Two. It is also the policy President Kagame launched in the aftermath of the genocide in Rwanda, as did Prime Minister Thaksin in Thailand after the Asian Financial Crisis in 2002, and the Chinese leadership did following the SARS crisis, also in 2003.In China’s case, reform involved re-socialising the health financing system using around 2% GDP in tax financing to increase health insurance coverage from a low level of one-third right up to 96% of the population.All these universal health coverage (UHC) reforms delivered massive health and economic benefits to the people - just what is needed now to tackle COVID-19 - and tremendous political benefits to the leaders that implemented them.When considering the current COVID-19 crisis, this strategy would be particularly relevant for countries underperforming on health coverage and whose health systems are more likely to be overwhelmed if flooded with a surge of patients, such as India, Pakistan, Bangladesh, Myanmar, Indonesia and most of sub-Saharan Africa, where many governments spend less than 1% of their GDP on health and most people have to buy services over the counter.But also the two OECD countries without a universal health system – the United States and Ireland – are seeing the threat of COVID-19 already fuelling the debate about the need to create national, publicly-financed health system. And the presidents of South Africa, Kenya and Indonesia have already committed their governments to eventually reach full population coverage anyway, and so may use this crisis to accelerate their own universal reforms. Although difficult to predict which leaders are likely to grasp the opportunity, if some of these countries now fast-track nationwide UHC, at least something good will be coming from the crisis, something which will benefit their people forever. And ensuring everyone accesses the services they need, including public health and preventive services, also provides the best protection against any future outbreaks becoming epidemics.Every night large audiences are tuning in to press briefings fronted by their heads of state hungry for the latest update on the crisis and to get reassurance that their government’s strategy will bring the salvation they desperately need. To truly improve health security for people across the world, becoming UHC heroes could be the best strategic decision political leaders ever make. Full Article
rom Emerging Lessons From COVID-19 By feedproxy.google.com Published On :: Thu, 02 Apr 2020 14:49:54 +0000 2 April 2020 Jim O'Neill Chair, Chatham House Exploring what lessons can be learned from the crisis to improve society and the functioning of our economic model going forward. 2020-04-02-COVID-Italy A man with a protective mask by the Coliseum in Rome during the height of Italy's COVID-19 epidemic. Photo by ALBERTO PIZZOLI/AFP via Getty Images. As tentative evidence emerges that Italy and Spain may have reached - or are close to - the peak of the curve, this could demonstrate that not only can Asian countries get to grips with COVID-19, but so can western democracies. And, if so, this offers a path for the rest of us.The last few weeks does demonstrate there is a role for governments to intervene in society, whether it be health, finance or any walk of life, as they have had to implement social distancing. Some have been forced, and the interventions are almost definitely only temporary, but perhaps some others may be less so.Governments of all kinds now realise there is a connection between our health system quality and our economic capability. On an index of global economic sustainability that I presided over creating when I was at Goldman Sachs, the top ten best performing countries on growth environment scores includes eight of the best performing ten countries - so far- in handling the crisis in terms of deaths relative to their population.Health system qualityThe top three on the index (last calculated in 2014) were Singapore, Hong Kong and South Korea, all of which are exemplary to the rest of us on how to deal with this mess. This suggests that once we are through this crisis, a number of larger populated countries - and their international advisors such as the IMF - might treat the quality of countries' health systems just as importantly as many of the other more standard indicators in assessing ability to deal with shocks.Policymakers have also been given a rather stark warning about other looming health disasters, especially antimicrobial resistance, of which antibiotic resistance lies at the heart. An independent review I chaired recommended 29 interventions, requiring $42 bn worth of investment, essentially peanuts compared to the costs of no solution, and the current economic collapse from COVID-19. It would seem highly likely to me that policymakers are going to treat this more seriously now.As a clear consequence of the - hopefully, temporary - global economic collapse, our environment suddenly seems to be cleaner and fresher and, in this regard, we have bought some time in the battle against climate change. Surely governments are going to be able to have a bigger influence on fossil fuel extractors and intense users as we emerge from this crisis?For any industries requiring government support, the government can make it clear this is dependent on certain criteria. And surely the days of excessive use of share buy backs and extreme maximisation of profit at the expense of other goals, are over?It seems to me an era of 'optimisation' of a number of business goals is likely to be the mantra, including profits but other things too such as national equality especially as it relates to income. Here in the UK, the government has offered its strongest fiscal support to the lower end of the income earning range group and, in a single swoop, has presided over its most dramatic step towards narrowing income inequality for a long time.This comes on top of a period of strong initiatives to support higher levels of minimum earnings, meaning we will emerge later in 2020, into 2021, and beyond, with lower levels of income inequality.The geographic issue of rural versus urban is also key. COVID-19 has spread more easily in more tightly packed cities such as London, New York and many others. More geographically remote places, by definition, are better protected. Perhaps now there will be some more thought given by policymakers to the quality and purpose of life outside our big metropolitan areas.Lastly, will China emerge from this crisis by offering a mammoth genuine gesture to the rest of the world, and come up, with, unlike, in 2008, a fiscal stimulus to its own consumers, that is geared towards importing a lot of things from the rest of the world? Now that would be good way of bringing the world back together again.This is a version of an article originally published in The Article Full Article
rom Justice for the Rohingya: Lessons from the Khmer Rouge Tribunal By feedproxy.google.com Published On :: Wed, 08 Apr 2020 12:22:46 +0000 8 April 2020 Sandra Smits Programme Manager, Asia-Pacific Programme The Cambodian case study illustrates the challenges of ensuring justice and accountability for the Rohingya in Myanmar. 2020-04-08-Rohingya.jpg Coast guards escort Rohingya refugees following a boat capsizing accident in Teknaf on 11 February 2020. Photo: Getty Images. International criminal justice provides a stark reminder that state sovereignty is not an absolute, and that the world’s most heinous crimes should be prosecuted at an international level, particularly where domestic systems lack the capacity or will to hold perpetrators to account. The post-Cold War period witnessed a dramatic rise in the number of international tribunals with jurisdiction over war crimes and serious human rights abuses in countries including Cambodia, East Timor, Rwanda, Liberia, Sierra Leone and Yugoslavia. With these processes approaching, or having reached the end of their dockets, many have called for the creation of new tribunals to address more recent conflicts, including the army crackdown in Myanmar in 2017 that resulted in evidence of crimes against humanity against the Rohingya. In January this year, the International Court of Justice (ICJ) imposed emergency provisional measures on Myanmar, instructing it to prevent genocidal violence against its Rohingya minority. But a final judgement is expected to take years and the ICJ has no way of enforcing these interim measures. Myanmar has already responded defiantly to international criticism. Model for justiceMyanmar is not the first country to face scrutiny for such crimes in Southeast Asia. The Extraordinary Chambers in the Courts of Cambodia (ECCC), more commonly known as the Khmer Rouge Tribunal was established in 1997 to prosecute Khmer Rouge leaders for alleged violations of international law and serious crimes perpetrated during the Cambodian genocide. This provides an opportunity to consider whether the Tribunal can act as a ‘hybrid’ model for justice in the region. The first lesson that can be taken from the Cambodian context is that the state must have the political will and commitment to pursue accountability. It was indeed the Cambodian government itself, who requested international assistance from the United Nations (UN), to organize a process for holding trials. The initial recommendation of the UN-commissioned Group of Experts was for the trial to be held under UN control, in light of misgivings about Cambodia’s judicial system. Prime Minister Hun Sen rejected this assessment and in prolonged negotiations, continued to spearhead the need for domestic involvement (arguably, in order to circumscribe the search for justice). This eventually resulted in the creation of a hybrid body consisting of parallel international and Cambodian judges and prosecutors with supermajority decision-making rules. It is worth noting that the Hun Sen government initially chose to do business with former Khmer Rouge leaders, until it became more advantageous to embrace a policy of putting them on trial. It is possible to infer from this that there will be no impetus for action in Myanmar until it is domestically advantageous to do so. At present, this appetite is clearly lacking, demonstrated by de-facto leader Aung San Suu Kyi shying away from accountability and instead defending the government’s actions before the ICJ.One unique aspect of the Khmer Rouge Tribunal has been the vast participation by the Cambodian people in witnessing the trials as well as widespread support for the tribunal. This speaks to the pent-up demand in Cambodia for accountability and the importance of local participation. While international moral pressure is clear, external actors cannot simply impose justice for the Rohingya when there is no domestic incentive or support to pursue this. The reality is that the anti-Rohingya campaign has galvanized popular support from the country’s Buddhist majority. What is more, the Rohingya are not even seen as part of Myanmar so there is an additional level of disenfranchisement.Secondly, the Cambodian Tribunal illustrates the need for safeguards against local political interference. The ECCC was designed as national court with international participation. There was an agreement to act in accordance with international standards of independence and impartiality, but no safeguards in place against serious deficiencies in the Cambodian judicial system. Close alliances between judges and the ruling Cambodian People’s Party, as well as high levels of corruption meant the tribunal effectively gave Hun Sen’s government veto power over the court at key junctures. Despite the guise of a hybrid structure, the Cambodian government ultimately retained the ability to block further prosecutions and prevent witnesses from being called. In Myanmar, political interference could be a concern, but given there is no popular support for justice and accountability for crimes committed against the Rohingya, the prospects of a domestic or hybrid process remain unlikely. However, there are still international options. The investigation by the International Criminal Court (ICC) into crimes that may have taken place on the Myanmar–Bangladesh border represents a potential route for justice and accountability. The UN Human Rights Council has also recently established the Independent Investigative Mechanism for Myanmar (IIMM), mandated to collect and preserve evidence, as well as to prepare files for future cases before criminal courts.Finally, the Cambodian case illustrates the culture of impunity in the region. The ECCC was conceived partly as a showcase for international standards of justice, which would have a ‘contagion effect’ upon the wider Cambodian and regional justice systems. Cambodia was notorious for incidents in which well-connected and powerful people flouted the law. This culture of impunity was rooted in the failure of the government to arrest, try and punish the Khmer Rouge leadership. The Tribunal, in holding perpetrators of the worst crimes to account, sought to send a clear signal that lesser violations would not be tolerated in the same way. Arguably, it did not achieve this in practice as Cambodia still has a highly politicized judicial system with high levels of corruption and clear limits to judicial independence. What this illustrates is that the first step towards accountability is strengthening domestic institutions. The United Nation’s Special Rapporteur on the situation of human rights in Myanmar has urged domestic authorities to embrace democracy and human rights, highlighting the need to reform the judicial system in order to ensure judicial independence, remove systemic barriers to accountability and build judicial and investigatory capacity in accordance with international standards. Based on this assessment, it is clear that domestic institutions are currently insufficiently independent to pursue accountability.The ECCC, despite its shortcomings, does stand as proof that crimes against humanity will not go completely unpunished. However, a process does not necessarily equal justice. The region is littered with justice processes that never went anywhere: Indonesia, Nepal, and Sri Lanka. International recourse is also challenging in a region with low ratification of the ICC, and the absence of regional mechanisms like the Inter-American Court of Human Rights, the European Court of Human Rights, and the African Court on Human and Peoples’ Rights (although their remit is not mass atrocity prosecutions). The Cambodian case study illustrates the challenges of ensuring justice and accountability within the region. The end of impunity is critical to ensure peaceful societies, but a purely legalistic approach will fail unless it is supported by wider measures and safeguards. It is these challenges, that undermine the prospects for ensuring justice for the Rohingya within Myanmar. Full Article
rom Fourier transform mass spectrometry [Invited] By feedproxy.google.com Published On :: 2011-05-09T12:10:35-07:00 This article provides an introduction to Fourier transform-based mass spectrometry (FTMS). The key performance characteristics of FTMS, mass accuracy and resolution, are presented in the view of how they impact the interpretation of measurements in proteomic applications. The theory and principles of operation of two types of mass analyzer, Fourier transform ion cyclotron resonance and Orbitrap, are described. Major benefits as well as limitations of FTMS technology are discussed in the context of practical sample analysis, and illustrated with examples included as figures in this text and in the accompanying slide set. Comparisons highlighting the performance differences between the two mass analyzers are made where deemed useful in assisting the user with choosing the most appropriate technology for his/her application. Recent developments of these high-performing mass spectrometers are mentioned to provide a future outlook. Full Article
rom WITHDRAWN: Quantitative mass spectrometry analysis of PD-L1 protein expression, N-glycosylation and expression stoichiometry with PD-1 and PD-L2 in human melanoma [Research] By feedproxy.google.com Published On :: 2017-04-28T07:30:39-07:00 This article has been withdrawn by the authors. We discovered an error after this manuscript was published as a Paper in Press. Specifically, we learned that the structures of glycans presented for the PD-L1 peptide were drawn and labeled incorrectly. We wish to withdraw this article and submit a corrected version for review. Full Article
rom Selection of features with consistent profiles improves relative protein quantification in mass spectrometry experiments [Research] By feedproxy.google.com Published On :: 2020-03-31T13:35:14-07:00 In bottom-up mass spectrometry-based proteomics, relative protein quantification is often achieved with data-dependent acquisition (DDA), data-independent acquisition (DIA), or selected reaction monitoring (SRM). These workflows quantify proteins by summarizing the abundances of all the spectral features of the protein (e.g., precursor ions, transitions or fragments) in a single value per protein per run. When abundances of some features are inconsistent with the overall protein profile (for technological reasons such as interferences, or for biological reasons such as post-translational modifications), the protein-level summaries and the downstream conclusions are undermined. We propose a statistical approach that automatically detects spectral features with such inconsistent patterns. The detected features can be separately investigated, and if necessary removed from the dataset. We evaluated the proposed approach on a series of benchmark controlled mixtures and biological investigations with DDA, DIA and SRM data acquisitions. The results demonstrated that it can facilitate and complement manual curation of the data. Moreover, it can improve the estimation accuracy, sensitivity and specificity of detecting differentially abundant proteins, and reproducibility of conclusions across different data processing tools. The approach is implemented as an option in the open-source R-based software MSstats. Full Article
rom A cross-linking mass spectrometry approach defines protein interactions in yeast mitochondria [Research] By feedproxy.google.com Published On :: 2020-04-24T09:36:17-07:00 Protein cross-linking and the analysis of cross-linked peptides by mass spectrometry is currently receiving much attention. Not only is this approach applied to isolated complexes to provide information about spatial arrangements of proteins but it is also increasingly applied to entire cells and their organelles. As in quantitative proteomics, the application of isotopic labelling further makes it possible to monitor quantitative changes in the protein-protein interactions between different states of a system. Here, we cross-linked mitochondria from Saccharomyces cerevisiae grown on either glycerol- or glucose-containing medium to monitor protein-protein interactions under non-fermentative and fermentative conditions. We investigated qualitatively the protein-protein interactions of the 400 most abundant proteins applying stringent data-filtering criteria, i.e. a minimum of two cross-linked peptide spectrum matches and a cut-off in the spectrum scoring of the used search engine. The cross-linker BS3 proved to be equally suited for connecting proteins in all compartments of mitochondria when compared with its water-insoluble but membrane-permeable derivative DSS. We also applied quantitative cross-linking to mitochondria of both the growth conditions using stable-isotope labelled BS3. Significant differences of cross-linked proteins under glycerol and glucose conditions were detected, however, mainly due to the different copy numbers of these proteins in mitochondria under both the conditions. Results obtained from the glycerol condition indicate that the internal NADH:ubiquinone oxidoreductase Ndi1 is part of an electron transport chain supercomplex. We have also detected several hitherto uncharacterized proteins and identified their interaction partners. Among those, Min8 was found to be associated with cytochrome c oxidase. BN-PAGE analyses of min8 mitochondria suggest that Min8 promotes the incorporation of Cox12 into cytochrome c oxidase. Full Article
rom Immunopeptidomic analysis reveals that deamidated HLA-bound peptides arise predominantly from deglycosylated precursors [Technological Innovation and Resources] By feedproxy.google.com Published On :: 2020-05-01T11:35:42-07:00 The presentation of post-translationally modified (PTM) peptides by cell surface HLA molecules has the potential to increase the diversity of targets for surveilling T cells. Whilst immunopeptidomics studies routinely identify thousands of HLA-bound peptides from cell lines and tissue samples, in-depth analyses of the proportion and nature of peptides bearing one or more PTMs remains challenging. Here we have analyzed HLA-bound peptides from a variety of allotypes and assessed the distribution of mass spectrometry-detected PTMs, finding deamidation of asparagine or glutamine to be highly prevalent. Given that asparagine deamidation may arise either spontaneously or through enzymatic reaction, we assessed allele-specific and global motifs flanking the modified residues. Notably, we found that the N-linked glycosylation motif NX(S/T) was highly abundant across asparagine-deamidated HLA-bound peptides. This finding, demonstrated previously for a handful of deamidated T cell epitopes, implicates a more global role for the retrograde transport of nascently N-glycosylated polypeptides from the ER and their subsequent degradation within the cytosol to form HLA-ligand precursors. Chemical inhibition of Peptide:N-Glycanase (PNGase), the endoglycosidase responsible for the removal of glycans from misfolded and retrotranslocated glycoproteins, greatly reduced presentation of this subset of deamidated HLA-bound peptides. Importantly, there was no impact of PNGase inhibition on peptides not containing a consensus NX(S/T) motif. This indicates that a large proportion of HLA-I bound asparagine deamidated peptides are generated from formerly glycosylated proteins that have undergone deglycosylation via the ER-associated protein degradation (ERAD) pathway. The information herein will help train deamidation prediction models for HLA-peptide repertoires and aid in the design of novel T cell therapeutic targets derived from glycoprotein antigens. Full Article
rom Promoting a Just Transition to an Inclusive Circular Economy By feedproxy.google.com Published On :: Wed, 01 Apr 2020 11:19:03 +0000 1 April 2020 Considerations of justice and social equity are as important for the circular economy transition as they are in the contexts of low-carbon transitions and digitalization of the economy. This paper sets out the just transition approach, and its relevance in climate change and energy transition debates. Read online Download PDF Patrick Schröder Senior Research Fellow, Energy, Environment and Resources Programme @patricks_CH Google Scholar 2020-04-01-circular-economy.jpg Residents of Mount Ijen take sulphur at Ijen Crater, Banyuwangi, East Java, on 2 July 2018. Photo: Getty Images. Summary Many social and political issues have so far been neglected in planning for the circular economy transition. This paper aims to redress this by considering how ‘just transition’ and social equity may be achieved through policy and practice.The prevailing economic model is linear, in that resources are extracted, transformed into products, used, and finally discarded. In contrast, the circular economy recognizes that natural resources are finite, and aims to keep the materials in products in circulation for as long as possible: reusing, repairing, remanufacturing, sharing and recycling. While the concept of the circular economy is largely focused on developing new technologies and businesses to enable keeping materials in circulation, it also includes the notions of ‘designing out’ waste, substituting renewable materials for non-renewable ones, and restoring natural systems.The UN 2030 Agenda demonstrates that environmental, social and economic sustainability objectives cannot be separated. As the links between the environmental issues of climate change, overconsumption of resources and waste generation, and social issues of inequality and the future of work become increasingly obvious, the urgency to connect environmental with social justice is gaining in significance. The language of ‘just transition’ – a transition that ensures environmental sustainability, decent work, social inclusion and poverty eradication – has started to penetrate debates and research on sustainability policy, particularly in the contexts of climate change and low-carbon energy transition.A just transition framework for the circular economy can identify opportunities that reduce waste and stimulate product innovation, while at the same time contributing positively to sustainable human development. And a just transition is needed to reduce inequalities within and between countries, and to ensure that the commitment of the UN Sustainable Development Goals to leave no one behind is fulfilled.It is important to identify the likely impacts on employment as a result of digitalization and industrial restructuring. Combining circular economy policies with social protection measures will be important in order to ensure that the burden of efforts to promote circularity will not fall on the poor through worsening working conditions and health impacts, reduced livelihoods, or job losses. Identifying potential winners and losers through participatory ‘roadmapping’ can help shape effective cooperation mechanisms and partnerships nationally and internationally.Many low- and middle-income countries that rely heavily on ‘linear’ sectors such as mining, manufacturing of non-repairable fast-moving consumer goods, textiles and agriculture, and the export of these commodities to higher-income countries, are likely to be negatively affected by the shift to circularity. These countries will need support from the international community through targeted assistance programmes if international trade in established commodities and manufactures declines in the medium to long term. International cooperation to create effective and fair governance mechanisms, and policy coordination at regional, national and local levels will play an important role in shaping a just transition. Multilateral technical assistance programmes will need to be designed and implemented, in particular to support low- and middle-income countries.Governments, international development finance institutions and banks are among the bodies beginning to establish circular economy investment funds and programmes. Just transition principles are yet to be applied to many of these new finance mechanisms, and will need to be integrated into development finance to support the circular economy transition.New international cooperation programmes, and a global mechanism to mobilize dedicated support funds for countries in need, will be critical to successful implementation across global value chains. Transparent and accountable institutions will also be important in ensuring that just transition funds reach those affected as intended. Department/project Energy, Environment and Resources Programme Full Article
rom Letter from David Hannay By feedproxy.google.com Published On :: Mon, 06 Apr 2020 12:03:37 +0000 1 February 2008 , Number 5 David Hannay House of Lords, London v-p-lb-e-00262h.jpg Full Article
rom Three Takeaways From the Belarusian Parliamentary Elections By feedproxy.google.com Published On :: Thu, 28 Nov 2019 10:45:08 +0000 28 November 2019 Ryhor Astapenia Robert Bosch Stiftung Academy Fellow, Russia and Eurasia Programme @ryhorastapenia LinkedIn Lukashenka’s domestic support is waning and he is not willing to make concessions to the West. Instead, he is trying to appease the ruling cadre. 2019-11-28-Luk.jpg Alexander Lukashenka leaves a voting booth on 17 November. Photo: Getty Images. Belarus’s parliamentary elections, held on 17 November, were predictably non-transparent, with numerous violations. The regime of Alexander Lukasheka allowed no opposition candidates as members of parliament – in contrast to the previous parliament, in which there were two opposition MPs. While this might seem to be a return to ‘business as usual’, three key takeaways from the elections highlight a shifting political and social landscape.1. Lukashenka is appeasing his ruling cadre by promising to increase their role in the political system.With several influential officials becoming new MPs, it is more likely that parliament will be more involved in any forthcoming discussion of a new constitution. Lukashenka has been promising constitutional reform for several years; he has said publicly that it will lead to an increased significance of government agencies as well as parliament. The aim of this is to keep them more engaged and on Lukashenka’s side.In terms of the composition of the new parliament itself, there are some key differences with previous years. It is no longer a comfortable place for officials to while away their pre-retirement: many MPs are now in their fifties or younger, and have plans for careers beyond parliament.It also looks as if small steps are being taken towards the emergence of a party system in Belarus. The leader of Belaya Rus, a pro-government association of Belarusian officials, got a seat in parliament for the first time, increasing the likelihood of it becoming a political party. The number of MPs from different parties has increased to 21 (out of 110 in total). Although these still all broadly support Lukashenka, they can differ from the president in policy positions. For example, the Labour and Justice Party, with 6 seats in parliament, supported the annexation of Crimea in 2014. Change, of course, may not necessarily be in a pro-Western direction.Moreover, the newly elected members of parliament look more like real politicians. They go to debates, speak to independent journalists and have their own social media channels. Some have even felt able to criticize the actions of the authorities.Similar changes have been taking place in other institutions in Belarus. The government is now more competent than it has ever been. The National Bank has managed to carry out macroeconomic stabilization on one of the most unstable currencies in Europe, while the ministries responsible for the economic development have implemented certain small-scale reforms. The Minister of the Interior has even acknowledged mistakes made by his department (under his predecessor), and undertaken to make improvements.This has resulted in a near-comical situation, whereby the Belarusian non-state media outlets have an increasingly positive view of some state officials, such as Prime Minister Siarhei Rumas, while the state media has been scaling back its coverage of him to ensure he does not become too popular.2. Belarus has less need for the West and is reluctant to make even small concessions.Since the slight warming of Belarusian relations with the West in 2014, Lukashenka has been having more meetings with prominent Western officials. Western institutions began trying to cooperate more closely with Belarus, but soon saw that it was not very interested. In 2018, the European Bank of Reconstruction and Development allocated €360 million to Belarus. However, it is now considering a reduction, as reforms in Belarus have not, in its view, gone far enough. The European Union has also committed considerable time and money to regional authorities, but this has not brought any significant changes to Belarusian local government.The absence of opposition candidates also demonstrates that the Belarusian authorities are prepared for a new deterioration of relations with the West. The authorities could certainly have afforded some opposition in parliament, especially since they themselves choose whom to appoint. Given that they did not, they either do not consider worsened relations a problem or are confident that the West will continue to cooperate with Belarus in order to limit Russian power, regardless of its violations of the rule of law.3. There is growing popular dissatisfaction with the current regime, but the state has no good plan for how to deal with it.Parliamentary election campaigns in Belarus are traditionally low-key, but this year they were particularly muted. The authorities tried to ensure that people knew as little about the election as possible. Campaign posters appeared on the streets just two weeks before polling day. It seems the authorities were reluctant to politicize society, as further resentment at autocratic rule is brewing.Many Belarusians who previously supported Lukashenka now have a very critical opinion of him. Take political blogging: the most popular political blogger in Belarus is a 22-year-old man who goes by the name of NEXTA. He produces low-quality videos which are highly critical of the authorities. A film by him about Lukashenka, released a month ago, has already received 1.8 million views, even though there are only 9.5 million people in Belarus.The authorities are not in a concessionary mood. The presidential elections in 2020 will also likely be a sham. If the authorities’ grip over the country is weakened, they will fear an outbreak of anger, resulting in widespread protests which the regime might once again have to meet with violence. Full Article
rom Quantification of bile acids: a mass spectrometry platform for studying gut microbe connection to metabolic diseases [Research Articles] By feedproxy.google.com Published On :: 2020-02-01T00:05:23-08:00 Bile acids (BAs) serve multiple biological functions, ranging from the absorption of lipids and fat-soluble vitamins to serving as signaling molecules through the direct activation of dedicated cellular receptors. Synthesized by both host and microbial pathways, BAs are increasingly understood as participating in the regulation of numerous pathways relevant to metabolic diseases, including lipid and glucose metabolism, energy expenditure, and inflammation. Quantitative analyses of BAs in biological matrices can be problematic due to their unusual and diverse physicochemical properties, making optimization of a method that shows good accuracy, precision, efficiency of extraction, and minimized matrix effects across structurally distinct human and murine BAs challenging. Herein we develop and clinically validate a stable-isotope-dilution LC/MS/MS method for the quantitative analysis of numerous primary and secondary BAs in both human and mouse biological matrices. We also utilize this tool to investigate gut microbiota participation in the generation of structurally specific BAs in both humans and mice. We examine circulating levels of specific BAs and in a clinical case-control study of age- and gender-matched type 2 diabetes mellitus (T2DM) versus nondiabetics. BAs whose circulating levels are associated with T2DM include numerous 12α-hydroxyl BAs (taurocholic acid, taurodeoxycholic acid, glycodeoxycholic acid, deoxycholic acid, and 3-ketodeoxycholic acid), while taurohyodeoxycholic acid was negatively associated with diabetes. The LC/MS/MS-based platform described should serve as a robust, high-throughput investigative tool for studying the potential involvement of structurally specific BAs and the gut microbiome on both physiological and disease processes. Full Article
rom Exon 9-deleted CETP inhibits full length-CETP synthesis and promotes cellular triglyceride storage [Research Articles] By feedproxy.google.com Published On :: 2020-03-01T00:06:33-08:00 Cholesteryl ester transfer protein (CETP) exists as full-length (FL) and exon 9 (E9)-deleted isoforms. The function of E9-deleted CETP is poorly understood. Here, we investigated the role of E9-deleted CETP in regulating the secretion of FL-CETP by cells and explored its possible role in intracellular lipid metabolism. CETP overexpression in cells that naturally express CETP confirmed that E9-deleted CETP is not secreted, and showed that cellular FL- and E9-deleted CETP form an isolatable complex. Coexpression of CETP isoforms lowered cellular levels of both proteins and impaired FL-CETP secretion. These effects were due to reduced synthesis of both isoforms; however, the predominate consequence of FL- and E9-deleted CETP coexpression is impaired FL-CETP synthesis. We reported previously that reducing both CETP isoforms or overexpressing FL-CETP impairs cellular triglyceride (TG) storage. To investigate this further, E9-deleted CETP was expressed in SW872 cells that naturally synthesize CETP and in mouse 3T3-L1 cells that do not. E9-deleted CETP overexpression stimulated SW872 triglyceride synthesis and increased stored TG 2-fold. Expression of E9-deleted CETP in mouse 3T3-L1 cells produced a similar lipid phenotype. In vitro, FL-CETP promotes the transfer of TG from ER-enriched membranes to lipid droplets. E9-deleted CETP also promoted this transfer, although less effectively, and it inhibited the transfer driven by FL-CETP. We conclude that FL- and E9-deleted CETP isoforms interact to mutually decrease their intracellular levels and impair FL-CETP secretion by reducing CETP biosynthesis. E9-deleted CETP, like FL-CETP, alters cellular TG metabolism and storage but in a contrary manner. Full Article
rom The citrus flavonoid nobiletin confers protection from metabolic dysregulation in high-fat-fed mice independent of AMPK [Research Articles] By feedproxy.google.com Published On :: 2020-03-01T00:06:33-08:00 Obesity, dyslipidemia, and insulin resistance, the increasingly common metabolic syndrome, are risk factors for CVD and type 2 diabetes that warrant novel therapeutic interventions. The flavonoid nobiletin displays potent lipid-lowering and insulin-sensitizing properties in mice with metabolic dysfunction. However, the mechanisms by which nobiletin mediates metabolic protection are not clearly established. The central role of AMP-activated protein kinase (AMPK) as an energy sensor suggests that AMPK is a target of nobiletin. We tested the hypothesis that metabolic protection by nobiletin required phosphorylation of AMPK and acetyl-CoA carboxylase (ACC) in mouse hepatocytes, in mice deficient in hepatic AMPK (Ampkβ1–/–), in mice incapable of inhibitory phosphorylation of ACC (AccDKI), and in mice with adipocyte-specific AMPK deficiency (iβ1β2AKO). We fed mice a high-fat/high-cholesterol diet with or without nobiletin. Nobiletin increased phosphorylation of AMPK and ACC in primary mouse hepatocytes, which was associated with increased FA oxidation and attenuated FA synthesis. Despite loss of ACC phosphorylation in Ampkβ1–/– hepatocytes, nobiletin suppressed FA synthesis and enhanced FA oxidation. Acute injection of nobiletin into mice did not increase phosphorylation of either AMPK or ACC in liver. In mice fed a high-fat diet, nobiletin robustly prevented obesity, hepatic steatosis, dyslipidemia, and insulin resistance, and it improved energy expenditure in Ampkβ1–/–, AccDKI, and iβ1β2AKO mice to the same extent as in WT controls. Thus, the beneficial metabolic effects of nobiletin in vivo are conferred independently of hepatic or adipocyte AMPK activation. These studies further underscore the therapeutic potential of nobiletin and begin to clarify possible mechanisms. Full Article
rom Hepatic PLIN5 signals via SIRT1 to promote autophagy and prevent inflammation during fasting [Research Articles] By feedproxy.google.com Published On :: 2020-03-01T00:06:33-08:00 Lipid droplets (LDs) are energy-storage organelles that are coated with hundreds of proteins, including members of the perilipin (PLIN) family. PLIN5 is highly expressed in oxidative tissues, including the liver, and is thought to play a key role in uncoupling LD accumulation from lipotoxicity; however, the mechanisms behind this action are incompletely defined. We investigated the role of hepatic PLIN5 in inflammation and lipotoxicity in a murine model under both fasting and refeeding conditions and in hepatocyte cultures. PLIN5 ablation with antisense oligonucleotides triggered a pro-inflammatory response in livers from mice only under fasting conditions. Similarly, PLIN5 mitigated lipopolysaccharide- or palmitic acid-induced inflammatory responses in hepatocytes. During fasting, PLIN5 was also required for the induction of autophagy, which contributed to its anti-inflammatory effects. The ability of PLIN5 to promote autophagy and prevent inflammation were dependent upon signaling through sirtuin 1 (SIRT1), which is known to be activated in response to nuclear PLIN5 under fasting conditions. Taken together, these data show that PLIN5 signals via SIRT1 to promote autophagy and prevent FA-induced inflammation as a means to maintain hepatocyte homeostasis during periods of fasting and FA mobilization. Full Article
rom Erratum: Unequivocal evidence for endogenous geranylgeranoic acid biosynthesized from mevalonate in mammalian cells [Errata] By feedproxy.google.com Published On :: 2020-04-01T00:05:29-07:00 Full Article
rom An LC/MS/MS method for analyzing the steroid metabolome with high accuracy and from small serum samples [Methods] By feedproxy.google.com Published On :: 2020-04-01T00:05:29-07:00 Analyzing global steroid metabolism in humans can shed light on the etiologies of steroid-related diseases. However, existing methods require large amounts of serum and lack the evaluation of accuracy. Here, we developed an LC/MS/MS method for the simultaneous quantification of 12 steroid hormones: testosterone, pregnenolone, progesterone, androstenedione, corticosterone, 11-deoxycortisol, cortisol, 17-hydroxypregnenolone, 17-hydroxyprogesterone, dehydroepiandrosterone, estriol, and estradiol. Steroids and spiked internal standards in 100 μl serum were extracted by protein precipitation and liquid-liquid extraction. The organic phase was dried by evaporation, and isonicotinoyl chloride was added for steroid derivatization, followed by evaporation under nitrogen and redissolution in 50% methanol. Chromatographic separation was performed on a reverse-phase PFP column, and analytes were detected on a triple quadrupole mass spectrometer with ESI. The lower limits of quantification ranged from 0.005 ng/ml for estradiol to 1 ng/ml for cortisol. Apparent recoveries of steroids at high, medium, and low concentrations in quality control samples were between 86.4% and 115.0%. There were limited biases (–10.7% to 10.5%) between the measured values and the authentic values, indicating that the method has excellent reliability. An analysis of the steroid metabolome in pregnant women highlighted the applicability of the method in clinical serum samples. We conclude that the LC/MS/MS method reported here enables steroid metabolome analysis with high accuracy and reduced serum consumption, indicating that it may be a useful tool in both clinical and scientific laboratory research. Full Article
rom Problem Notes for SAS®9 - 65939: "ERROR: Unable to transcode data to/from UCS-2 encoding" occurs when you run an SQL query using SAS/ACCESS Interface to ODBC on SAS 9.4M5 with UTF-8 By feedproxy.google.com Published On :: Fri, 8 May 2020 11:20:02 EST When you run an SQL query using SAS/ACCESS Interface to ODBC under the following conditions, you might receive an error: You run SAS 9.4M5 (TS1M5) or SAS 9.4M6 (TS1M6) i Full Article ODBC+SAS/ACCESS+Interface+to+ODBC
rom Problem Notes for SAS®9 - 65899: "ERROR: ORACLE disconnect error: ORA-03135" occurs when SAS disconnects from the Oracle database server By feedproxy.google.com Published On :: Thu, 7 May 2020 08:04:08 EST When you run 32-bit SAS on Windows and disconnect from the Oracle database server, you might see the error: "ERROR: ORACLE disconnect error: ORA-03135: connection lost contact." Full Article ORACLE+SAS/ACCESS+Interface+to+Oracle
rom Rays' 2019 mantra: Prepare to win from within By mlb.mlb.com Published On :: Fri, 8 Feb 2019 16:05:02 EDT Over the past couple seasons, the Rays have preached patience as the organization provided time for the top prospects to make it up to the Majors. Now, the focus has been primarily in remaining flexible and keeping positions open for the young talent arriving from the Minors. Full Article
rom A little rain can't keep eager Rays from field By mlb.mlb.com Published On :: Wed, 13 Feb 2019 15:17:29 EDT A rainy morning didn't stop the Rays' pitchers and catchers from officially taking the field Wednesday. "Other than the pitchers getting out there and getting in their legs a little bit and running some of the more casual [pitchers' fielding practice drills], everything is fine," manager Kevin Cash said. Full Article
rom ADAM10 and ADAM17 proteases mediate proinflammatory cytokine-induced and constitutive cleavage of endomucin from the endothelial surface [Membrane Biology] By feedproxy.google.com Published On :: 2020-05-08T03:41:14-07:00 Contact between inflammatory cells and endothelial cells (ECs) is a crucial step in vascular inflammation. Recently, we demonstrated that the cell-surface level of endomucin (EMCN), a heavily O-glycosylated single-transmembrane sialomucin, interferes with the interactions between inflammatory cells and ECs. We have also shown that, in response to an inflammatory stimulus, EMCN is cleared from the cell surface by an unknown mechanism. In this study, using adenovirus-mediated overexpression of a tagged EMCN in human umbilical vein ECs, we found that treatment with tumor necrosis factor α (TNF-α) or the strong oxidant pervanadate leads to loss of cell-surface EMCN and increases the levels of the C-terminal fragment of EMCN 3- to 4-fold. Furthermore, treatment with the broad-spectrum matrix metalloproteinase inhibitor batimastat (BB94) or inhibition of ADAM metallopeptidase domain 10 (ADAM10) and ADAM17 with two small-molecule inhibitors, GW280264X and GI254023X, or with siRNA significantly reduced basal and TNFα-induced cell-surface EMCN cleavage. Release of the C-terminal fragment of EMCN by TNF-α treatment was blocked by chemical inhibition of ADAM10 alone or in combination with ADAM17. These results indicate that cell-surface EMCN undergoes constitutive cleavage and that TNF-α treatment dramatically increases this cleavage, which is mediated predominantly by ADAM10 and ADAM17. As endothelial cell-surface EMCN attenuates leukocyte–EC interactions during inflammation, we propose that EMCN is a potential therapeutic target to manage vascular inflammation. Full Article
rom Dopamine and Early Retinal Dysfunction in Diabetes: Insights From a Phase 1 Study By diabetes.diabetesjournals.org Published On :: 2020-04-20T15:26:16-07:00 Full Article
rom Coregulator Sin3a Promotes Postnatal Murine {beta}-Cell Fitness by Regulating Genes in Ca2+ Homeostasis, Cell Survival, Vesicle Biosynthesis, Glucose Metabolism, and Stress Response By diabetes.diabetesjournals.org Published On :: 2020-04-21T12:16:29-07:00 Swi-independent 3a and 3b (Sin3a and Sin3b) are paralogous transcriptional coregulators that direct cellular differentiation, survival, and function. Here, we report that mouse Sin3a and Sin3b are co-produced in most pancreatic cells during embryogenesis but become much more enriched in endocrine cells in adults, implying continued essential roles in mature endocrine-cell function. Mice with loss of Sin3a in endocrine progenitors were normal during early postnatal stages but gradually developed diabetes before weaning. These physiological defects were preceded by the compromised survival, insulin-vesicle packaging, insulin secretion, and nutrient-induced Ca2+ influx of Sin3a-deficient β-cells. RNA-seq coupled with candidate chromatin-immunoprecipitation assays revealed several genes that could be directly regulated by Sin3a in β-cells, which modulate Ca2+/ion transport, cell survival, vesicle/membrane trafficking, glucose metabolism, and stress responses. Lastly, mice with loss of both Sin3a and Sin3b in multipotent embryonic pancreatic progenitors had significantly reduced islet-cell mass at birth, caused by decreased endocrine-progenitor production and increased β-cell death. These findings highlight the stage-specific requirements for the presumed "general" coregulators Sin3a and Sin3b in islet β-cells, with Sin3a being dispensable for differentiation but required for postnatal function and survival. Full Article
rom Hyperuricemia Predisposes to the Onset of Diabetes via Promoting Pancreatic {beta}-Cell Death in Uricase Deficiency Male Mice By diabetes.diabetesjournals.org Published On :: 2020-04-24T13:05:31-07:00 Clinical studies have shown a link between hyperuricemia (HU) and diabetes, while the exact effect of soluble serum urate on glucose metabolism remains elusive. This study aims to characterize the glucose metabolic phenotypes and investigate the underlying molecular mechanisms using a novel spontaneous HU mouse model in which the Uricase (Uox) gene is absent. In an attempt to study the role of HU in glycometabolism, we implemented external stimulation on Uox-knockout (KO) and wild-type (WT) males with a high-fat diet (HFD) and/or injections of multiple low-dose streptozotocin (MLD-STZ) to provoke the potential role of urate. Notably, while Uox-KO mice developed glucose intolerance in the basal condition, no mice spontaneously developed diabetes, even with aging. HFD-fed Uox-KO mice manifested similar insulin sensitivity compared with WT controls. HU augmented the existing glycometabolism abnormality induced by MLD-STZ and eventually led to diabetes, as evidenced by the increased random glucose. Reduced β-cell masses and increased terminal deoxynucleotidyl TUNEL-positive β-cells suggested that HU-mediated diabetes was cell death dependent. However, urate-lowering treatment (ULT) cannot ameliorate the diabetes incidence or reverse β-cell apoptosis with significance. ULT displayed a significant therapeutic effect of HU-crystal– associated kidney injury and tubulointerstitial damage in diabetes. Moreover, we present transcriptomic analysis of isolated islets, using Uox-KO versus WT mice and streptozotocin-induced diabetic WT (STZ-WT) versus diabetic Uox-KO (STZ-KO) mice. Shared differentially expressed genes of HU primacy revealed Stk17β is a possible target gene in HU-related β-cell death. Together, this study suggests that HU accelerates but does not cause diabetes by inhibiting islet β-cell survival. Full Article