pH-dependent structural transitions in cationic ionizable lipid mesophases are critical for lipid nanoparticle function.

Lipid nanoparticles (LNPs) are advanced core-shell particles for messenger RNA (mRNA) based therapies that are made of polyethylene glycol (PEG) lipid, distearoylphosphatidylcholine (DSPC), cationic ionizable lipid (CIL), cholesterol (chol), and mRNA. Yet the mechanism of pH-dependent response that is believed to cause endosomal release of LNPs is not well understood. Here, we show that eGFP (enhanced green fluorescent protein) protein expression in the mouse liver mediated by the ionizable lipids DLin-MC3-DMA (MC3), DLin-KC2-DMA (KC2), and DLinDMA (DD) ranks MC3 ≥ KC2 > DD despite similar delivery of mRNA per cell in all cell fractions isolated. We hypothesize that the three CIL-LNPs react differently to pH changes and hence study the structure of CIL/chol bulk phases in water. Using synchrotron X-ray scattering a sequence of ordered CIL/chol mesophases with lowering pH values are observed. These phases show isotropic inverse micellar, cubic Fd3m inverse micellar, inverse hexagonal [Formula: see text] and bicontinuous cubic Pn3m symmetry. If polyadenylic acid, as mRNA surrogate, is added to CIL/chol, excess lipid coexists with a condensed nucleic acid lipid [Formula: see text] phase. The next-neighbor distance in the excess phase shows a discontinuity at the Fd3m inverse micellar to inverse hexagonal [Formula: see text] transition occurring at pH 6 with distinctly larger spacing and hydration for DD vs. MC3 and KC2. In mRNA LNPs, DD showed larger internal spacing, as well as retarded onset and reduced level of DD-LNP-mediated eGFP expression in vitro compared to MC3 and KC2. Our data suggest that the pH-driven Fd3m-[Formula: see text] transition in bulk phases is a hallmark of CIL-specific differences in mRNA LNP efficacy.

Reduction of Paraoxonase Expression Followed by Inactivation across Independent Semiaquatic Mammals Suggests Stepwise Path to Pseudogenization.

Convergent adaptation to the same environment by multiple lineages frequently involves rapid evolutionary change at the same genes, implicating these genes as important for environmental adaptation. Such adaptive molecular changes may yield either change or loss of protein function; loss of function can eliminate newly deleterious proteins or reduce energy necessary for protein production. We previously found a striking case of recurrent pseudogenization of the Paraoxonase 1 (Pon1) gene among aquatic mammal lineages-Pon1 became a pseudogene with genetic lesions, such as stop codons and frameshifts, at least four times independently in aquatic and semiaquatic mammals. Here, we assess the landscape and pace of pseudogenization by studying Pon1 sequences, expression levels, and enzymatic activity across four aquatic and semiaquatic mammal lineages: pinnipeds, cetaceans, otters, and beavers. We observe in beavers and pinnipeds an unexpected reduction in expression of Pon3, a paralog with similar expression patterns but different substrate preferences. Ultimately, in all lineages with aquatic/semiaquatic members, we find that preceding any coding-level pseudogenization events in Pon1, there is a drastic decrease in expression, followed by relaxed selection, thus allowing accumulation of disrupting mutations. The recurrent loss of Pon1 function in aquatic/semiaquatic lineages is consistent with a benefit to Pon1 functional loss in aquatic environments. Accordingly, we examine diving and dietary traits across pinniped species as potential driving forces of Pon1 functional loss. We find that loss is best associated with diving activity and likely results from changes in selective pressures associated with hypoxia and hypoxia-induced inflammation.

Facilitative and competitive interactions between mycorrhizal and nonmycorrhizal plants in an extremely phosphorus-impoverished environment: role of ectomycorrhizal fungi and native oomycete pathogens in shaping species coexistence.

Nonmycorrhizal cluster root-forming species enhance the phosphorus (P) acquisition of mycorrhizal neighbours in P-impoverished megadiverse systems. However, whether mycorrhizal plants facilitate the defence of nonmycorrhizal plants against soil-borne pathogens, in return and via their symbiosis, remains unknown. We characterised growth and defence-related compounds in Banksia menziesii (nonmycorrhizal) and Eucalyptus todtiana (ectomycorrhizal, ECM) seedlings grown either in monoculture or mixture in a multifactorial glasshouse experiment involving ECM fungi and native oomycete pathogens. Roots of B. menziesii had higher levels of phytohormones (salicylic and jasmonic acids, jasmonoyl-isoleucine and 12-oxo-phytodienoic acid) than E. todtiana which further activated a salicylic acid-mediated defence response in roots of B. menziesii, but only in the presence of ECM fungi. We also found that B. menziesii induced a shift in the defence strategy of E. todtiana, from defence-related secondary metabolites (phenolic and flavonoid) towards induced phytohormone response pathways. We conclude that ECM fungi play a vital role in the interactions between mycorrhizal and nonmycorrhizal plants in a severely P-impoverished environment, by introducing a competitive component within the facilitation interaction between the two plant species with contrasting nutrient-acquisition strategies. This study sheds light on the interplay between beneficial and detrimental soil microbes that shape plant-plant interaction in severely nutrient-impoverished ecosystems.

Phosphorus fractions in leaves.

Leaf phosphorus (P) comprises four major fractions: inorganic phosphate (Pi ), nucleic acids, phospholipids, P-containing metabolites and a residual fraction. In this review paper, we investigated whether allocation of P fractions varies among groups of terrestrial vascular plants, and is indicative of a species' strategy to use P efficiently. We found that as leaf total P concentration increases, the Pi fraction increases the most, without a plateau, while other fractions plateau. Variability of the concentrations of leaf P fractions is greatest among families > species(family) > regions > plant life forms. The percentage of total P allocated to nucleic acid-P (20-35%) and lipid-P (14-34%) varies less among families/species. High photosynthetic P-use efficiency is associated with low concentrations of all P fractions, and preferential allocation of P to metabolite-P and mesophyll cells. Sequential resorption of P from senescing leaves starts with Pi , followed by metabolite-P, and then other organic P fractions. Allocation of P to leaf P fractions varies with season. Leaf phytate concentrations vary considerably among species, associated with variation in photosynthesis and defence. Plasticity of P allocation to its fractions is important for acclimation to low soil P availability, and species-specific P allocation is needed for co-occurrence with other species.

Differences in foliar phosphorus fractions, rather than in cell-specific phosphorus allocation, underlie contrasting photosynthetic phosphorus use efficiency among chickpea genotypes.

Although significant intraspecific variation in photosynthetic phosphorus (P) use efficiency (PPUE) has been shown in numerous species, we still know little about the biochemical basis for differences in PPUE among genotypes within a species. Here, we grew two high PPUE and two low PPUE chickpea (Cicer arietinum) genotypes with low P supply in a glasshouse to compare their photosynthesis-related traits, total foliar P concentration ([P]) and chemical P fractions (i.e. inorganic P (Pi), metabolite P, lipid P, nucleic acid P, and residual P). Foliar cell-specific nutrient concentrations including P were characterized using elemental X-ray microanalysis. Genotypes with high PPUE showed lower total foliar [P] without slower photosynthetic rates. No consistent differences in cellular [P] between the epidermis and mesophyll cells occurred across the four genotypes. In contrast, high PPUE was associated with lower allocation to Pi and metabolite P, with PPUE being negatively correlated with the percentage of these two fractions. Furthermore, a lower allocation to Pi and metabolite P was correlated with a greater allocation to nucleic acid P, but not to lipid P. Collectively, our results suggest that a different allocation to foliar P fractions, rather than preferential P allocation to specific leaf tissues, underlies the contrasting PPUE among chickpea genotypes.

High-Precision Spectroscopy of

The excited states of unstable ^{20}O were investigated via γ-ray spectroscopy following the ^{19}O(d,p)^{20}O reaction at 8 AMeV. By exploiting the Doppler shift attenuation method, the lifetimes of the 2_{2}^{+} and 3_{1}^{+} states were firmly established. From the γ-ray branching and E2/M1 mixing ratios for transitions deexciting the 2_{2}^{+} and 3_{1}^{+} states, the B(E2) and B(M1) were determined. Various chiral effective field theory Hamiltonians, describing the nuclear properties beyond ground states, along with a standard USDB interaction, were compared with the experimentally obtained data. Such a comparison for a large set of γ-ray transition probabilities with the valence space in medium similarity renormalization group ab initio calculations was performed for the first time in a nucleus far from stability. It was shown that the ab initio approaches using chiral effective field theory forces are challenged by detailed high-precision spectroscopic properties of nuclei. The reduced transition probabilities were found to be a very constraining test of the performance of the ab initio models.

Health consequences of exposure to aircraft contaminated air and fume events: a narrative review and medical protocol for the investigation of exposed aircrew and passengers.

Thermally degraded engine oil and hydraulic fluid fumes contaminating aircraft cabin air conditioning systems have been well documented since the 1950s. Whilst organophosphates have been the main subject of interest, oil and hydraulic fumes in the air supply also contain ultrafine particles, numerous volatile organic hydrocarbons and thermally degraded products. We review the literature on the effects of fume events on aircrew health. Inhalation of these potentially toxic fumes is increasingly recognised to cause acute and long-term neurological, respiratory, cardiological and other symptoms. Cumulative exposure to regular small doses of toxic fumes is potentially damaging to health and may be exacerbated by a single higher-level exposure. Assessment is complex because of the limitations of considering the toxicity of individual substances in complex heated mixtures.There is a need for a systematic and consistent approach to diagnosis and treatment of persons who have been exposed to toxic fumes in aircraft cabins. The medical protocol presented in this paper has been written by internationally recognised experts and presents a consensus approach to the recognition, investigation and management of persons suffering from the toxic effects of inhaling thermally degraded engine oil and other fluids contaminating the air conditioning systems in aircraft, and includes actions and investigations for in-flight, immediately post-flight and late subsequent follow up.

Examining the role of paraoxonase 2 in the dopaminergic system of the mouse brain.

BACKGROUND: Paraoxonase 2 (PON2) is an intracellular antioxidant enzyme located at the inner mitochondrial membrane. Previous studies have found PON2 to be an important antioxidant in a variety of cellular systems, such as the cardiovascular and renal system. Recent work has also suggested that PON2 plays an important role in the central nervous system (CNS), as decreased PON2 expression in the CNS leads to higher oxidative stress and subsequent cell toxicity. However, the precise role of PON2 in the CNS is still largely unknown, and what role it may play in specific regions of the brain remains unexamined. Dopamine metabolism generates considerable oxidative stress and antioxidant function is critical to the survival of dopaminergic neurons, providing a potential mechanism for PON2 in the dopaminergic system. METHODS: In this study, we investigated the role of PON2 in the dopaminergic system of the mouse brain by comparing transcript and protein expression of dopaminergic-related genes in wildtype (WT) and PON2 deficient (PON2-def) mouse striatum, and exposing WT cultured primary neurons to dopamine receptor agonists. RESULTS: We found alterations in multiple key dopaminergic genes at the transcript level, however many of these changes were not observed at the protein level. In cultured neurons, PON2 mRNA and protein were increased upon exposure to quinpirole, a dopamine receptor 2/3 (DRD2/3) agonist, but not fenoldopam, a dopamine receptor 1/5 (DRD1/5) agonist, suggesting a receptor-specific role in dopamine signaling. CONCLUSIONS: Our findings suggest PON2 deficiency significantly impacts the dopaminergic system at the transcript level and may play a role in mitigating oxidative stress in this system further downstream through dopamine receptor signaling.

Evidence of Partial Seniority Conservation in the πg_{9/2} Shell for the N=50 Isotones.

The reduced transition probabilities for the 4_{1}^{+}→2_{1}^{+} and 2_{1}^{+}→0_{1}^{+} transitions in ^{92}Mo and ^{94}Ru and for the 4_{1}^{+}→2_{1}^{+} and 6_{1}^{+}→4_{1}^{+} transitions in ^{90}Zr have been determined in this experiment making use of a multinucleon transfer reaction. These results have been interpreted on the basis of realistic shell-model calculations in the f_{5/2}, p_{3/2}, p_{1/2}, and g_{9/2} proton valence space. Only the combination of extensive lifetime information and large scale shell-model calculations allowed the extent of the seniority conservation in the N=50 g_{9/2} orbital to be understood. The conclusion is that seniority is largely conserved in the first πg_{9/2} orbital.

The Conformation of the N-Terminal Tails of Deinococcus grandis Dps Is Modulated by the Ionic Strength.

DNA-binding proteins from starved cells (Dps) are homododecameric nanocages, with N- and C-terminal tail extensions of variable length and amino acid composition. They accumulate iron in the form of a ferrihydrite mineral core and are capable of binding to and compacting DNA, forming low- and high-order condensates. This dual activity is designed to protect DNA from oxidative stress, resulting from Fenton chemistry or radiation exposure. In most Dps proteins, the DNA-binding properties stem from the N-terminal tail extensions. We explored the structural characteristics of a Dps from Deinococcus grandis that exhibits an atypically long N-terminal tail composed of 52 residues and probed the impact of the ionic strength on protein conformation using size exclusion chromatography, dynamic light scattering, synchrotron radiation circular dichroism and small-angle X-ray scattering. A novel high-spin ferrous iron-binding site was identified in the N-terminal tails, using Mössbauer spectroscopy. Our data reveals that the N-terminal tails are structurally dynamic and alter between compact and extended conformations, depending on the ionic strength of the buffer. This prompts the search for other physiologically relevant modulators of tail conformation and hints that the DNA-binding properties of Dps proteins may be affected by external factors.

Anomalous SAXS at P12 beamline EMBL Hamburg: instrumentation and applications.

Small-angle X-ray scattering (SAXS) is an established method for studying nanostructured systems and in particular biological macromolecules in solution. To obtain element-specific information about the sample, anomalous SAXS (ASAXS) exploits changes of the scattering properties of selected atoms when the energy of the incident X-rays is close to the binding energy of their electrons. While ASAXS is widely applied to condensed matter and inorganic systems, its use for biological macromolecules is challenging because of the weak anomalous effect. Biological objects are often only available in small quantities and are prone to radiation damage, which makes biological ASAXS measurements very challenging. The BioSAXS beamline P12 operated by the European Molecular Biology Laboratory (EMBL) at the PETRA III storage ring (DESY, Hamburg) is dedicated to studies of weakly scattering objects. Here, recent developments at P12 allowing for ASAXS measurements are presented. The beamline control, data acquisition and data reduction pipeline of the beamline were adapted to conduct ASAXS experiments. Modelling tools were developed to compute ASAXS patterns from atomic models, which can be used to analyze the data and to help designing appropriate data collection strategies. These developments are illustrated with ASAXS experiments on different model systems performed at the P12 beamline.

Experimental Evidence for Common Driving Effects in Low-Energy Fission from Sublead to Actinides.

Isotopic distributions of fragments from fission of the neutron-deficient ^{178}Hg nuclide are reported. This experimental observable is obtained for the first time in the region around lead using an innovative approach based on inverse kinematics and the coincidence between the large acceptance magnetic spectrometer VAMOS++ and a new detection arm close to the target. The average fragment N/Z ratio and prompt neutron M_{n} multiplicity are derived and compared with current knowledge from actinide fission. A striking consistency emerges, revealing the unexpected dominant role of the proton subsystem with atomic number between the Z=28 and 50 magic numbers. The origin of nuclear charge polarization in fission and fragment deformation at scission are discussed.

Isospin Properties of Nuclear Pair Correlations from the Level Structure of the Self-Conjugate Nucleus

The low-lying energy spectrum of the extremely neutron-deficient self-conjugate (N=Z) nuclide _{44}^{88}Ru_{44} has been measured using the combination of the Advanced Gamma Tracking Array (AGATA) spectrometer, the NEDA and Neutron Wall neutron detector arrays, and the DIAMANT charged particle detector array. Excited states in ^{88}Ru were populated via the ^{54}Fe(^{36}Ar,2nγ)^{88}Ru^{*} fusion-evaporation reaction at the Grand Accélérateur National d'Ions Lourds (GANIL) accelerator complex. The observed γ-ray cascade is assigned to ^{88}Ru using clean prompt γ-γ-2-neutron coincidences in anticoincidence with the detection of charged particles, confirming and extending the previously assigned sequence of low-lying excited states. It is consistent with a moderately deformed rotating system exhibiting a band crossing at a rotational frequency that is significantly higher than standard theoretical predictions with isovector pairing, as well as observations in neighboring N>Z nuclides. The direct observation of such a "delayed" rotational alignment in a deformed N=Z nucleus is in agreement with theoretical predictions related to the presence of strong isoscalar neutron-proton pair correlations.

Carbon nanotube-based thin-film resistive sensor for point-of-care screening of tuberculosis.

For point-of-care diagnosis of tuberculosis (TB), current TB diagnostic approaches need to be further improved for achieving an accurate diagnosis that is rapid and low-cost. This paper presents an immuno-resistive sensor on a plastic film for inexpensive, simple TB screening. The sensor is composed of single-walled carbon nanotubes (SWCNTs) functionalized with polyclonal antibodies raised against the MPT64 surface antigen from Mycobacterium tuberculosis (MTB). The target analyte of either MTB or MPT64 is spiked in tongue swab and sputum samples. Under optimized conditions, targets are directly detected from tongue swab samples by resistive measurement. Target analytes spiked into human sputa are enriched with a magnetic bead protocol followed by resistive detection. This highly sensitive film sensor will facilitate rapid TB screening with the added benefits of a small form factor, simple operation, low power requirement, and low cost.

Investigation of pH-Responsiveness inside Lipid Nanoparticles for Parenteral mRNA Application Using Small-Angle X-ray Scattering.

Messenger ribonucleic acid (mRNA)-based nanomedicines have shown to be a promising new lead in a broad field of potential applications such as tumor immunotherapy. Of these nanomedicines, lipid-based mRNA nanoparticles comprising ionizable lipids are gaining increasing attention as versatile technologies for fine-tuning toward a given application, with proven potential for successful development up to clinical practice. Still, several hurdles have to be overcome to obtain a drug product that shows adequate mRNA delivery and clinical efficacy. In this study, pH-induced changes in internal molecular organization and overall physicochemical characteristics of lipoplexes comprising ionizable lipids were investigated using small-angle X-ray scattering and supplementary techniques. These changes were determined for different types of ionizable lipids, present at various molar fractions and N/P ratios inside the phospholipid membranes. The investigated systems showed a lamellar organization, allowing an accurate determination of pH-dependent structural changes. The differences in the pH responsiveness of the systems comprising different ionizable lipids and mRNA fractions could be clearly revealed from their structural evolution. Measurements of the degree of ionization and pH-dependent mRNA loading into the systems by fluorescence assays supported the findings from the structural investigation. Our approach allows for direct in situ determination of the structural response of the lipoplex systems to changes of the environmental pH similar to that observed for endosomal uptake. These data therefore provide valuable complementary information for understanding and fine-tuning of tailored mRNA delivery systems toward improved cellular uptake and endosomal processing.

[Postoperative complications and long-term outcomes of transplantation with expended criteria donors transplants]. / Complications chirurgicales et résultats à long terme des transplantations rénales avec transplants à critères élargis.

OBJECTIVE: The objective of this study was to analyze early, late complications and outcomes following expended criteria donors (ECD) kidney transplantation compared to standard donors. MATERIALS AND METHODS: We performed a retrospective study including 470 patients who received a kidney transplant between 2005 and 2016. Expended criteria donors were defined following the United Network of Organ Sharing criteria. In each group, length of stay, delayed graft function, surgical site infection, acute rejection, surgical complications by type and according to Clavien and Dindo classification were analyzed in univariate and multivariate analysis. The impact of ECD transplant on transplant and patient survival was assessed using a Cox proportional regression model. RESULTS: One hundred and ninety seven (41.9%) patients received ECD kidney. The mean follow-up was 61,4 months (22.4-93.89). Patients with ECD transplant presented more delayed graft function (HR=2.1 (1.1-3.68), P=0.008) but the rate of complications including surgical complications was not different. Patients and transplant survival were decreased in ECD transplant group (P=0.005 et 0.001 respectively). In multivariate analysis ECD kidney was an independent factor only associated with decreased transplant survival (HR=1.81 (1.1-2.98), P=0.029) but not with patient survival. CONCLUSION: ECD kidney transplantation was not associated with increased postoperative complications but a higher rate of delayed graft function. Nevertheless, it was associated with a decreased transplant survival. The use of pulsatile perfusion machine for explanted criteria transplant should be evaluated to improve these results.

First Direct Measurement of Isotopic Fission-Fragment Yields of

A direct and complete measurement of isotopic fission-fragment yields of ^{239}U has been performed for the first time. The ^{239}U fissioning system was produced with an average excitation energy of 8.3 MeV in one-neutron transfer reactions between a ^{238}U beam and a ^{9}Be target at Coulomb barrier energies. The fission fragments were detected and isotopically identified using the VAMOS++ spectrometer at the GANIL facility. The measurement allows us to directly evaluate the fission models at excitation energies of fast neutrons, which are relevant for next-generation nuclear reactors. The present data, in agreement with model calculations, do not support the recently reported anomaly in the fission-fragment yields of ^{239}U, and they confirm the persistence of spherical shell effects in the Sn region at excitation energies exceeding the fission barrier by a few mega-electron volts.

Smaller capillaries improve the small-angle X-ray scattering signal and sample consumption for biomacromolecular solutions.

Radiation damage by intense X-ray beams at modern synchrotron facilities is one of the major complications for biological small-angle X-ray scattering (SAXS) investigations of macromolecules in solution. To limit the damage, samples are typically measured under a laminar flow through a cell (typically a capillary) such that fresh solution is continuously exposed to the beam during measurement. The diameter of the capillary that optimizes the scattering-to-absorption ratio at a given X-ray wavelength can be calculated a priori based on fundamental physical properties. However, these well established scattering and absorption principles do not take into account the radiation susceptibility of the sample or the often very limited amounts of precious biological material available for an experiment. Here it is shown that, for biological solution SAXS, capillaries with smaller diameters than those calculated from simple scattering/absorption criteria allow for a better utilization of the available volumes of radiation-sensitive samples. This is demonstrated by comparing two capillary diameters di (di = 1.7 mm, close to optimal for 10 keV; and di = 0.9 mm, which is nominally sub-optimal) applied to study different protein solutions at various flow rates. The use of the smaller capillaries ultimately allows one to collect higher-quality SAXS data from the limited amounts of purified biological macromolecules.

Neutron Skin Effects in Mirror Energy Differences: The Case of

Energy differences between analogue states in the T=1/2 ^{23}Mg-^{23}Na mirror nuclei have been measured along the rotational yrast bands. This allows us to search for effects arising from isospin-symmetry-breaking interactions (ISB) and/or shape changes. Data are interpreted in the shell model framework following the method successfully applied to nuclei in the f_{7/2} shell. It is shown that the introduction of a schematic ISB interaction of the same type of that used in the f_{7/2} shell is needed to reproduce the data. An alternative novel description, applied here for the first time, relies on the use of an effective interaction deduced from a realistic charge-dependent chiral nucleon-nucleon potential. This analysis provides two important results: (i) The mirror energy differences give direct insight into the nuclear skin; (ii) the skin changes along the rotational bands are strongly correlated with the difference between the neutron and proton occupations of the s_{1/2} "halo" orbit.

Pseudospin Symmetry and Microscopic Origin of Shape Coexistence in the

Lifetime measurements of excited states of the light N=52 isotones ^{88}Kr, ^{86}Se, and ^{84}Ge have been performed, using the recoil distance Doppler shift method and VAMOS and AGATA spectrometers for particle identification and gamma spectroscopy, respectively. The reduced electric quadrupole transition probabilities B(E2;2^{+}→0^{+}) and B(E2;4^{+}→2^{+}) were obtained for the first time for the hard-to-reach ^{84}Ge. While the B(E2;2^{+}→0^{+}) values of ^{88}Kr, ^{86}Se saturate the maximum quadrupole collectivity offered by the natural valence (3s, 2d, 1g_{7/2}, 1h_{11/2}) space of an inert ^{78}Ni core, the value obtained for ^{84}Ge largely exceeds it, suggesting that shape coexistence phenomena, previously reported at N≲49, extend beyond N=50. The onset of collectivity at Z=32 is understood as due to a pseudo-SU(3) organization of the proton single-particle sequence reflecting a clear manifestation of pseudospin symmetry. It is realized that the latter provides actually reliable guidance for understanding the observed proton and neutron single particle structure in the whole medium-mass region, from Ni to Sn, pointing towards the important role of the isovector-vector ρ field in shell-structure evolution.