New insights into globoids of protein storage vacuoles in wheat aleurone using synchrotron soft X-ray microscopy

Mature developed seeds are physiologically and biochemically committed to store nutrients, principally as starch, protein, oils, and minerals. The composition and distribution of elements inside the aleurone cell layer reflect their biogenesis, structural characteristics, and physiological functions. It is therefore of primary importance to understand the mechanisms underlying metal ion accumulation, distribution, storage, and bioavailability in aleurone subcellular organelles for seed fortification purposes. Synchrotron radiation soft X-ray full-field imaging mode (FFIM) and low-energy X-ray fluorescence (LEXRF) spectromicroscopy were applied to characterize major structural features and the subcellular distribution of physiologically important elements (Zn, Fe, Na, Mg, Al, Si, and P). These direct imaging methods reveal the accumulation patterns between the apoplast and symplast, and highlight the importance of globoids with phytic acid mineral salts and walls as preferential storage structures. C, N, and O chemical topographies are directly linked to the structural backbone of plant substructures. Zn, Fe, Na, Mg, Al, and P were linked to globoid structures within protein storage vacuoles with variable levels of co-localization. Si distribution was atypical, being contained in the aleurone apoplast and symplast, supporting a physiological role for Si in addition to its structural function. These results reveal that the immobilization of metals within the observed endomembrane structures presents a structural and functional barrier and affects bioavailability. The combination of high spatial and chemical X-ray microscopy techniques highlights how in situ analysis can yield new insights into the complexity of the wheat aleurone layer, whose precise biochemical composition, morphology, and structural characteristics are still not unequivocally resolved

The Conservation of VIT1-Dependent Iron Distribution in Seeds

One third of people suffer from anemia, with iron (Fe) deficiency being the most common reason. The human diet includes seeds of staple crops, which contain Fe that is poorly bioavailable. One reason for low bioavailability is that these seeds store Fe in cellular compartments that also contain antinutrients, such as phytate. Thus, several studies have focused on decreasing phytate concentrations. In theory, as an alternative approach, Fe reserves might be directed to cellular compartments that are free of phytate, such as plastids. However, it is not known if seed plastid can represent a major Fe storage compartment in nature. To discover distinct types of Fe storage in nature, we investigated metal localizations in the seeds of more than twenty species using histochemical or X-ray based techniques. Results showed that in Rosids, the largest clade of eudicots, Fe reserves were primarily confined to the embryo of the seeds. Furthermore, inside the embryos, Fe accumulated specifically in the endodermal cell layer, a well-known feature that is mediated by VACUOLAR IRON TRANSPORTER1 (VIT1) in model plant Arabidopsis thaliana. In rice, Fe enrichment is lost around the provasculature in the mutants of VIT1 orthologs. Finally, in Carica papaya, Fe accumulated in numerous organelles resembling plastids; however, these organelles accumulated reserve proteins but not ferritin, failing to prove to be plastids. By investigating Fe distribution in distinct plant lineages, this study failed to discover distinct Fe storage patterns that can be useful for biofortification. However, it revealed Fe enrichment is widely conserved in the endodermal cell layer in a VIT1-dependent manner in the plant kingdom

Gadolinium tissue deposition in the periodontal ligament of mice with reduced renal function exposed to Gd-based contrast agents

Gadolinium deposition in tissue is linked to nephrogenic systemic fibrosis (NSF): a rare disorder occurring in patients with severe chronic kidney disease and associated with administration of Gd-based contrast agents (GBCAs) for Magnetic Resonance Imaging (MRI). It is suggested that the GBCAs prolonged permanence in blood in these patients may result in a Gd precipitation in peripheral or central organs, where it initiates a fibrotic process. In this study we investigated new sites of retention/precipitation of Gd in a mouse model of renal disease (5/6 nephrectomy) receiving two doses (closely after each other) of a linear GBCA. Two commercial GBCAs (Omniscan\uae and Magnevist\uae) were administered at doses slightly higher than those used in clinical practice (0.7 mmol/kg body weight, each). The animals were sacrificed one month after the last administration and the explanted organs (kidney, liver, femur, dorsal skin, teeth) were analysed by X-ray fluorescence (XRF) at two synchrotron facilities. The XRF analysis with a millimetre-sized beam at the SYRMEP beamline (Elettra, Italy) produced no detectable levels of Gd in the examined tissues, with the notable exception of the incisors of the nephrectomised mice. The XRF analyses at sub-micron resolution performed at ID21 (ESRF, France) allowed to clearly localize Gd in the periodontal ligaments of teeth both from Omniscan\uae and Magnevist\uae treated nephrectomised mice. The latter results were further confirmed by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). The study prompts that prolonged permanence of GBCAs in blood may result in Gd retention in this particular muscular tissue, opening possibilities for diagnostic applications at this level when investigating Gd-related toxicities

Detention and mapping of iron and toxic environmental elements in human ovarian endometriosis: A suggested combined role

Background: Endometriosis is a disease affecting 10-15 % of women worldwide, consisting in the ectopic growth of endometrial cells outside the uterine cavity. Whist the pathogenetic mechanisms of endometriosis remain elusive and contemplating even environmental causes, iron deposits are common in endometrial lesions, indicating an altered iron metabolism at this level. This study was undertaken to reveal a possible relationship between iron dysmetabolism and accumulation of environmental metals. Methods: By combining histological and histochemical analysis (H&E and Perl's staining) with μ- and nano- synchrotron-based (SR-based) X-ray Fluorescence (XRF) microscopy, we investigated the distribution of iron and other elements in the ovarian endometriomas of 12 endometriosis patients and in 7 healthy endometrium samples. Results: XRF microscopy expanded the findings obtained by Perl's staining, revealing with an exceptional sensitivity intracellular features of iron accumulation in the epithelial endometrium, stroma and macrophages of the endometriotic lesions. XRF evidenced that iron was specifically accumulated in multiple micro aggregates, reaching concentrations up to 10-20 % p/p. Moreover, by XRF analysis we revealed for the first time the retention of a number of exogenous and potentially toxic metals such as Pb, Br, Ti, Al Cr, Si and Rb partially or totally co-localizing with iron. Conclusion: μXRF reveals accumulation and colocalization of iron and environmental metals in human ovarian endometriosis, suggesting a role in the pathogenesis of endometriosis

Antiinflammatory Therapy with Canakinumab for Atherosclerotic Disease

Background: Experimental and clinical data suggest that reducing inflammation without affecting lipid levels may reduce the risk of cardiovascular disease. Yet, the inflammatory hypothesis of atherothrombosis has remained unproved. Methods: We conducted a randomized, double-blind trial of canakinumab, a therapeutic monoclonal antibody targeting interleukin-1β, involving 10,061 patients with previous myocardial infarction and a high-sensitivity C-reactive protein level of 2 mg or more per liter. The trial compared three doses of canakinumab (50 mg, 150 mg, and 300 mg, administered subcutaneously every 3 months) with placebo. The primary efficacy end point was nonfatal myocardial infarction, nonfatal stroke, or cardiovascular death. RESULTS: At 48 months, the median reduction from baseline in the high-sensitivity C-reactive protein level was 26 percentage points greater in the group that received the 50-mg dose of canakinumab, 37 percentage points greater in the 150-mg group, and 41 percentage points greater in the 300-mg group than in the placebo group. Canakinumab did not reduce lipid levels from baseline. At a median follow-up of 3.7 years, the incidence rate for the primary end point was 4.50 events per 100 person-years in the placebo group, 4.11 events per 100 person-years in the 50-mg group, 3.86 events per 100 person-years in the 150-mg group, and 3.90 events per 100 person-years in the 300-mg group. The hazard ratios as compared with placebo were as follows: in the 50-mg group, 0.93 (95% confidence interval [CI], 0.80 to 1.07; P = 0.30); in the 150-mg group, 0.85 (95% CI, 0.74 to 0.98; P = 0.021); and in the 300-mg group, 0.86 (95% CI, 0.75 to 0.99; P = 0.031). The 150-mg dose, but not the other doses, met the prespecified multiplicity-adjusted threshold for statistical significance for the primary end point and the secondary end point that additionally included hospitalization for unstable angina that led to urgent revascularization (hazard ratio vs. placebo, 0.83; 95% CI, 0.73 to 0.95; P = 0.005). Canakinumab was associated with a higher incidence of fatal infection than was placebo. There was no significant difference in all-cause mortality (hazard ratio for all canakinumab doses vs. placebo, 0.94; 95% CI, 0.83 to 1.06; P = 0.31). Conclusions: Antiinflammatory therapy targeting the interleukin-1β innate immunity pathway with canakinumab at a dose of 150 mg every 3 months led to a significantly lower rate of recurrent cardiovascular events than placebo, independent of lipid-level lowering. (Funded by Novartis; CANTOS ClinicalTrials.gov number, NCT01327846.

The Changing Landscape for Stroke\ua0Prevention in AF: Findings From the GLORIA-AF Registry Phase 2

Background GLORIA-AF (Global Registry on Long-Term Oral Antithrombotic Treatment in Patients with Atrial Fibrillation) is a prospective, global registry program describing antithrombotic treatment patterns in patients with newly diagnosed nonvalvular atrial fibrillation at risk of stroke. Phase 2 began when dabigatran, the first non\u2013vitamin K antagonist oral anticoagulant (NOAC), became available. Objectives This study sought to describe phase 2 baseline data and compare these with the pre-NOAC era collected during phase 1. Methods During phase 2, 15,641 consenting patients were enrolled (November 2011 to December 2014); 15,092 were eligible. This pre-specified cross-sectional analysis describes eligible patients\u2019 baseline characteristics. Atrial fibrillation disease characteristics, medical outcomes, and concomitant diseases and medications were collected. Data were analyzed using descriptive statistics. Results Of the total patients, 45.5% were female; median age was 71 (interquartile range: 64, 78) years. Patients were from Europe (47.1%), North America (22.5%), Asia (20.3%), Latin America (6.0%), and the Middle East/Africa (4.0%). Most had high stroke risk (CHA2DS2-VASc [Congestive heart failure, Hypertension, Age  6575 years, Diabetes mellitus, previous Stroke, Vascular disease, Age 65 to 74 years, Sex category] score  652; 86.1%); 13.9% had moderate risk (CHA2DS2-VASc = 1). Overall, 79.9% received oral anticoagulants, of whom 47.6% received NOAC and 32.3% vitamin K antagonists (VKA); 12.1% received antiplatelet agents; 7.8% received no antithrombotic treatment. For comparison, the proportion of phase 1 patients (of N = 1,063 all eligible) prescribed VKA was 32.8%, acetylsalicylic acid 41.7%, and no therapy 20.2%. In Europe in phase 2, treatment with NOAC was more common than VKA (52.3% and 37.8%, respectively); 6.0% of patients received antiplatelet treatment; and 3.8% received no antithrombotic treatment. In North America, 52.1%, 26.2%, and 14.0% of patients received NOAC, VKA, and antiplatelet drugs, respectively; 7.5% received no antithrombotic treatment. NOAC use was less common in Asia (27.7%), where 27.5% of patients received VKA, 25.0% antiplatelet drugs, and 19.8% no antithrombotic treatment. Conclusions The baseline data from GLORIA-AF phase 2 demonstrate that in newly diagnosed nonvalvular atrial fibrillation patients, NOAC have been highly adopted into practice, becoming more frequently prescribed than VKA in Europe and North America. Worldwide, however, a large proportion of patients remain undertreated, particularly in Asia and North America. (Global Registry on Long-Term Oral Antithrombotic Treatment in Patients With Atrial Fibrillation [GLORIA-AF]; NCT01468701

Micro-XANES analysis of metal accumulation in plants on sub-cellular level

There is a growing need to develop powerful analytical tools for monitoring concentrations and chemical state of trace element in the biosphere and its abiotic environment, due to pollution and degradation of ecosystems worldwide. Increased metal concentrations present in the environment pose a threat to all living organisms from microorganisms and plants to animals and humans, because they interfere with vital biological processes. The goal is to efficiently assess metal bioavailability and toxicity, and gain more knowledge on the mechanisms of metal uptake, accumulation and detoxification in living organisms [1]. In this work we demonstrate that a combination of micro-XRF imaging and micro-XANES and EXAFS analysis represents a powerful and indispensable tool for characterization of metal pollutants on subcellular level. The methodological approaches for efficient micro-XAS experiments are presented, the limitations and sources of potential systematic errors in XANES and EXAFS analysis (especially at low energies) due to self-absorption effects and strong energy dependent penetration depth of X-ray beam in the sample are discussed. Some typical examples of such combined micro-spectroscopy analysis are selected from the following research fields: Cd/Zn hyper-accumulating plants, that can be used for phytoremediation of Cd/Zn polluted and degraded ecosystems, including investigation of the role of externally supplied sulphur compounds in the nutrient solution of the Cd/Zn hyperaccumulator Thlaspi praecox, that may alter leaf Cd distribution and Cd ligand environment [2]; biofortification, which aims to increase essential elements (Fe) concentrations in the edible plant parts [3]; microbial regulation of metal (Fe, Pd) uptake and formation of Fe-oxide and Pd nanoparticles, encapsulated in exopolysaccharide to avoid iron toxicity under anaerobic conditions, discovered on a strain of Klebsiella oxytoca, isolated from acid pyrite-mine drainage[4]

Nickel coordination in hyperaccumulator plants studied by XANES and EXAFS

Soils derived from ultramafic or serpentine rocks are characterized by elevated concentrations of Ni, Cr and Co. About 2% of plants on ultramafic soils accumulate metals in their shoots: a phenomenon known as hyperaccumulation. Ni coordination in plant organs of a Ni hyperaccumulator (HA), Berkheya zeyheri subsp. rehmannii var. rogersiana, and its non-hyperaccumulating (NHA) counterpart Berkheya zeyheri subsp. rehmannii var. rehmannii was studied by EXAFS and XANES. Both varieties originate from the vicinity of Barberton Mpumalanga, South Africa. Ni K-edge spectra of HA and NHA leaves, veins, stems and roots were measured at BM23 beamline of ESRF synchrotron facility, Grenoble, France and at P64 beamline of PETRA III, Hamburg, Germany. Samples were rapidly frozen against a metal plate cooled with liquid nitrogen, put into a windowless cryostat and evacuated. The spectra were collected in fluorescence mode. For comparison, spectra of soil and Ni standards i.e. nickel bound in citrate, malate, methionine, proline, catecholate and histidine complexes were measured in transmission mode. IFEFFIT program package [1] was used for data evaluation.The shape and position of the Ni K edge strongly indicate that nickel is in the form of Ni2+ ion in all samples. Spectra of HA leaves, veins and stem are almost identical and consistent with spectra of Ni bound to organic acid complexes, while Principal Component Analysis of the roots spectra exhibits a mixture of Ni coordinated as in the soil structure, to the organic acids and to histidine. NHA plants accumulate much lower amount of Ni, from which only the roots spectra give enough signal for analysis: there, Ni is predominantly bound as in the soil, and in a lesser amount to histidine. Results show that - in comparison with the NHA plant - HA roots not only uptake more Ni but they transform its chemical environment to a higher degree, enabling transport of the metal ions to higher parts of the plan