Drosophila Evi5 is a critical regulator of intracellular iron transport via transferrin and ferritin interactions.

Vesicular transport is essential for delivering cargo to intracellular destinations. Evi5 is a Rab11-GTPase-activating protein involved in endosome recycling. In humans, Evi5 is a high-risk locus for multiple sclerosis, a debilitating disease that also presents with excess iron in the CNS. In insects, the prothoracic gland (PG) requires entry of extracellular iron to synthesize steroidogenic enzyme cofactors. The mechanism of peripheral iron uptake in insect cells remains controversial. We show that Evi5-depletion in the Drosophila PG affected vesicle morphology and density, blocked endosome recycling and impaired trafficking of transferrin-1, thus disrupting heme synthesis due to reduced cellular iron concentrations. We show that ferritin delivers iron to the PG as well, and interacts physically with Evi5. Further, ferritin-injection rescued developmental delays associated with Evi5-depletion. To summarize, our findings show that Evi5 is critical for intracellular iron trafficking via transferrin-1 and ferritin, and implicate altered iron homeostasis in the etiology of multiple sclerosis.

Interpreting Neural Network Models for Toxicity Prediction by Extracting Learned Chemical Features.

Neural network models have become a popular machine-learning technique for the toxicity prediction of chemicals. However, due to their complex structure, it is difficult to understand predictions made by these models which limits confidence. Current techniques to tackle this problem such as SHAP or integrated gradients provide insights by attributing importance to the input features of individual compounds. While these methods have produced promising results in some cases, they do not shed light on how representations of compounds are transformed in hidden layers, which constitute how neural networks learn. We present a novel technique to interpret neural networks which identifies chemical substructures in training data found to be responsible for the activation of hidden neurons. For individual test compounds, the importance of hidden neurons is determined, and the associated substructures are leveraged to explain the model prediction. Using structural alerts for mutagenicity from the Derek Nexus expert system as ground truth, we demonstrate the validity of the approach and show that model explanations are competitive with and complementary to explanations obtained from an established feature attribution method.

Preservation of corneous ß-proteins in Mesozoic feathers.

Fossil proteins are valuable tools in evolutionary biology. Recent technological advances and better integration of experimental methods have confirmed the feasibility of biomolecular preservation in deep time, yielding new insights into the timing of key evolutionary transitions. Keratins (formerly α-keratins) and corneous ß-proteins (CBPs, formerly ß-keratins) are of particular interest as they define tissue structures that underpin fundamental physiological and ecological strategies and have the potential to inform on the molecular evolution of the vertebrate integument. Reports of CBPs in Mesozoic fossils, however, appear to conflict with experimental evidence for CBP degradation during fossilization. Further, the recent model for molecular modification of feather chemistry during the dinosaur-bird transition does not consider the relative preservation potential of different feather proteins. Here we use controlled taphonomic experiments coupled with infrared and sulfur X-ray spectroscopy to show that the dominant ß-sheet structure of CBPs is progressively altered to α-helices with increasing temperature, suggesting that (α-)keratins and α-helices in fossil feathers are most likely artefacts of fossilization. Our analyses of fossil feathers shows that this process is independent of geological age, as even Cenozoic feathers can comprise primarily α-helices and disordered structures. Critically, our experiments show that feather CBPs can survive moderate thermal maturation. As predicted by our experiments, analyses of Mesozoic feathers confirm that evidence of feather CBPs can persist through deep time.

Modular Functionalization of Metal-Organic Frameworks for Nitrogen Recovery from Fresh Urine.

Nitrogen recovery from wastewater represents a sustainable route to recycle reactive nitrogen (Nr). It can reduce the demand of producing Nr from the energy-extensive Haber-Bosch process and lower the risk of causing eutrophication simultaneously. In this aspect, source-separated fresh urine is an ideal source for nitrogen recovery given its ubiquity and high nitrogen contents. However, current techniques for nitrogen recovery from fresh urine require high energy input and are of low efficiencies because the recovery target, urea, is a challenge to separate. In this work, we developed a novel fresh urine nitrogen recovery treatment process based on modular functionalized metal-organic frameworks (MOFs). Specifically, we employed three distinct modification methods to MOF-808 and developed robust functional materials for urea hydrolysis, ammonium adsorption, and ammonia monitoring. By integrating these functional materials into our newly developed nitrogen recovery treatment process, we achieved an average of 75 % total nitrogen reduction and 45 % nitrogen recovery with a 30-minute treatment of synthetic fresh urine. The nitrogen recovery process developed in this work can serve as a sustainable and efficient nutrient management that is suitable for decentralized wastewater treatment. This work also provides a new perspective of implementing versatile advanced materials for water and wastewater treatment.

Metal ion content of internal organs in the calorically restricted Wistar rat.

BACKGROUND: Despite the agreed principle that access to food is a human right, undernourishment and metal ion deficiencies are public health problems worldwide, exacerbated in impoverished or war-affected areas. It is known that maternal malnutrition causes growth retardation and affects behavioral and cognitive development of the newborn. Here we ask whether severe caloric restriction leads per se to disrupted metal accumulation in different organs of the Wistar rat. METHODS: Inductively coupled plasma optical emission spectroscopy was used to determine the concentration of multiple elements in the small and large intestine, heart, lung, liver, kidney, pancreas, spleen, brain, spinal cord, and three skeletal muscles from control and calorically restricted Wistar rats. The caloric restriction protocol was initiated from the mothers prior to mating and continued throughout gestation, lactation, and post-weaning up to sixty days of age. RESULTS: Both sexes were analyzed but dimorphism was rare. The pancreas was the most affected organ presenting a higher concentration of all the elements analyzed. Copper concentration decreased in the kidney and increased in the liver. Each skeletal muscle responded to the treatment differentially: Extensor Digitorum Longus accumulated calcium and manganese, gastrocnemius decreased copper and manganese, whereas soleus decreased iron concentrations. Differences were also observed in the concentration of elements between organs independently of treatment: The soleus muscle presents a higher concentration of Zn compared to the other muscles and the rest of the organs. Notably, the spinal cord showed large accumulations of calcium and half the concentration of zinc compared to brain. X-ray fluorescence imaging suggests that the extra calcium is attributable to the presence of ossifications whereas the latter finding is attributable to the low abundance of zinc synapses in the spinal cord. CONCLUSION: Severe caloric restriction did not lead to systemic metal deficiencies but caused instead specific metal responses in few organs.

Identification, transformations and mobility of hazardous arsenic-based pigments on 19th century bookbindings in accessible library collections.

This study focused on the non-destructive characterization of potentially hazardous Victorian-era books found in the Northwestern University Libraries. XRF, Raman and FTIR were used to identify and isolate hazardous books containing As-based pigments. These techniques also permitted, on selected books, to characterize the pigment as being Emerald green. However, none allowed for the identification of equally hazardous degradation products or potential transfer to adjacent books. These analytical gaps create limits in thoroughly identifying the level of risks associated with these books for library users and hampered the application of effective risk mitigation measures. Such limitations were overcome with synchrotron radiation (SR) techniques. Through SR-XRF, Cu/As distributions were mapped across covers and spines of green and neighboring books, whereas SR-X-ray absorption near edge structure (SR-XANES) was used to characterize the As oxidation state, leading to the identification of arsenates as degradation products. Besides successfully identifying hazardous books, this study demonstrated that hazards extend beyond As-containing green books to innocuous, long-standing neighboring books and non-colored pages due to migration and transfer of pigment and degradation products. Aside from helping to implement workplace health and safety measures, this study also informs how other libraries can identify and characterize potentially hazardous items in their collections.

Response to comment on "Reexamination of 2.5-Ga 'whiff' of oxygen interval points to anoxic ocean before GOE".

Sedimentological, textural, and microscale analyses of the Mount McRae Shale revealed a complex postdepositional history, previously unrecognized in bulk geochemical studies. We found that metal enrichments in the shale do not reside with depositional organic carbon, as previously proposed by Anbar et al., but with late-stage pyrite, compromising claims for a "whiff" of oxygen ~50 million years before the Great Oxygenation Event.

Two Pathways for the Degradation of Orpiment Pigment (As2S3) Found in Paintings.

Paintings are complex objects containing many different chemical compounds that can react over time. The degradation of arsenic sulfide pigments causes optical changes in paintings. The main degradation product was thought to be white arsenolite (As2O3), but previous research also showed the abundant presence of As(V) species. In this study, we investigate the influence of the presence of a medium on the degradation mechanism of orpiment (As2S3) using synchrotron radiation (SR)-based tomographic transmission X-ray microscopy, SR-based micro-X-ray fluorescence, and X-ray absorption near edge structure spectroscopy. Upon direct illumination of dry orpiment powder using UV-visible light, only the formation of As2O3 was observed. When As2S3 was surrounded by a medium and illuminated, As2O3 was only observed in the area directly exposed to light, while As(V) degradation species were found elsewhere in the medium. Without accelerated artificial light aging, As(V)(aq) species are formed and migrate throughout the medium within weeks after preparation. In both scenarios, the As(V) species form via intermediate As(III)(aq) species and the presence of a medium is necessary. As(V)(aq) species can react with available cations to form insoluble metal arsenates, which induces stress within the paint layers (leading to, e.g., cracks and delamination) or can lead to a visual change of the image of the painting.

Evaluation of quantitative synchrotron radiation micro-X-ray fluorescence in rice grain.

Concentrations of nutrients and contaminants in rice grain affect human health, specifically through the localization and chemical form of elements. Methods to spatially quantify the concentration and speciation of elements are needed to protect human health and characterize elemental homeostasis in plants. Here, an evaluation was carried out using quantitative synchrotron radiation microprobe X-ray fluorescence (SR-µXRF) imaging by comparing average rice grain concentrations of As, Cu, K, Mn, P, S and Zn measured with rice grain concentrations from acid digestion and ICP-MS analysis for 50 grain samples. Better agreement was found between the two methods for high-Z elements. Regression fits between the two methods allowed quantitative concentration maps of the measured elements. These maps revealed that most elements were concentrated in the bran, although S and Zn permeated into the endosperm. Arsenic was highest in the ovular vascular trace (OVT), with concentrations approaching 100 mg kg-1 in the OVT of a grain from a rice plant grown in As-contaminated soil. Quantitative SR-µXRF is a useful approach for comparison across multiple studies but requires careful consideration of sample preparation and beamline characteristics.

Organic preservation of vase-shaped microfossils from the late Tonian Chuar Group, Grand Canyon, Arizona, USA.

Vase-shaped microfossils (VSMs) are found globally in middle Neoproterozoic (800-730 Ma) marine strata and represent the earliest evidence for testate (shell-forming) amoebozoans. VSM tests are hypothesized to have been originally organic in life but are most commonly preserved as secondary mineralized casts and molds. A few reports, however, suggest possible organic preservation. Here, we test the hypothesis that VSMs from shales of the lower Walcott Member of the Chuar Group, Grand Canyon, Arizona, contain original organic material, as reported by B. Bloeser in her pioneering studies of Chuar VSMs. We identified VSMs from two thin section samples of Walcott Member black shales in transmitted light microscopy and used scanning electron microscopy to image VSMs. Carbonaceous material is found within the internal cavity of all VSM tests from both samples and is interpreted as bitumen mobilized from Walcott shales likely during the Cretaceous. Energy dispersive X-ray spectroscopy (EDS) and wavelength dispersive X-ray spectroscopy (WDS) reveal that VSM test walls contain mostly carbon, iron, and sulfur, while silica is present only in the surrounding matrix. Raman spectroscopy was used to compare the thermal maturity of carbonaceous material within the samples and indicated the presence of pyrite and jarosite within fossil material. X-ray absorption spectroscopy revealed the presence of reduced organic sulfur species within the carbonaceous test walls, the carbonaceous material found within test cavities, and in the sedimentary matrix, suggesting that organic matter sulfurization occurred within the Walcott shales. Our suite of spectroscopic analyses reveals that Walcott VSM test walls are organic and sometimes secondarily pyritized (with the pyrite variably oxidized to jarosite). Both preservation modes can occur at a millimeter spatial scale within sample material, and at times even within a single specimen. We propose that sulfurization within the Walcott Shales promoted organic preservation, and furthermore, the ratio of iron to labile VSM organic material controlled the extent of pyrite replacement. Based on our evidence, we conclude that the VSMs are preserved with original organic test material, and speculate that organic VSMs may often go unrecognized, given their light-colored, translucent appearance in transmitted light.

A new µ-high energy resolution fluorescence detection microprobe imaging spectrometer at the Stanford Synchrotron Radiation Lightsource beamline 6-2.

Here, we describe a new synchrotron X-ray Fluorescence (XRF) imaging instrument with an integrated High Energy Fluorescence Detection X-ray Absorption Spectroscopy (HERFD-XAS) spectrometer at the Stanford Synchrotron Radiation Lightsource at beamline 6-2. The X-ray beam size on the sample can be defined via a range of pinhole apertures or focusing optics. XRF imaging is performed using a continuous rapid scan system with sample stages covering a travel range of 250 × 200 mm2, allowing for multiple samples and/or large samples to be mounted. The HERFD spectrometer is a Johann-type with seven spherically bent 100 mm diameter crystals arranged on intersecting Rowland circles of 1 m diameter with a total solid angle of about 0.44% of 4π sr. A wide range of emission lines can be studied with the available Bragg angle range of ∼64.5°-82.6°. With this instrument, elements in a sample can be rapidly mapped via XRF and then selected features targeted for HERFD-XAS analysis. Furthermore, utilizing the higher spectral resolution of HERFD for XRF imaging provides better separation of interfering emission lines, and it can be used to select a much narrower emission bandwidth, resulting in increased image contrast for imaging specific element species, i.e., sparse excitation energy XAS imaging. This combination of features and characteristics provides a highly adaptable and valuable tool in the study of a wide range of materials.

Sulfur cycling at natural hydrocarbon and sulfur seeps in Santa Paula Creek, CA.

Biogeochemical cycling of sulfur is relatively understudied in terrestrial environments compared to marine environments. However, the comparative ease of access, observation, and sampling of terrestrial settings can expand our understanding of organisms and processes important in the modern sulfur cycle. Furthermore, these sites may allow for the discovery of useful process analogs for ancient sulfur-metabolizing microbial communities at times in Earth's past when atmospheric O2 concentrations were lower and sulfide was more prevalent in Earth surface environments. We identified a new site at Santa Paula Creek (SPC) in Ventura County, CA-a remarkable freshwater, gravel-bedded mountain stream charged with a range of oxidized and reduced sulfur species and heavy hydrocarbons from the emergence of subsurface fluids within the underlying sulfur- and organic-rich Miocene-age Monterey Formation. SPC hosts a suite of morphologically distinct microbial biofacies that form in association with the naturally occurring hydrocarbon seeps and sulfur springs. We characterized the geology, stream geochemistry, and microbial facies and diversity of the Santa Paula Creek ecosystem. Using geochemical analyses and 16S rRNA gene sequencing, we found that SPC supports a dynamic sulfur cycle that is largely driven by sulfide-oxidizing microbial taxa, with contributions from smaller populations of sulfate-reducing and sulfur-disproportionating taxa. This preliminary characterization of SPC revealed an intriguing site in which to study geological and geochemical controls on microbial community composition and to expand our understanding of sulfur cycling in terrestrial environments.

Analysis of the benefits of imputation models over traditional QSAR models for toxicity prediction.

Recently, imputation techniques have been adapted to predict activity values among sparse bioactivity matrices, showing improvements in predictive performance over traditional QSAR models. These models are able to use experimental activity values for auxiliary assays when predicting the activity of a test compound on a specific assay. In this study, we tested three different multi-task imputation techniques on three classification-based toxicity datasets: two of small scale (12 assays each) and one large scale with 417 assays. Moreover, we analyzed in detail the improvements shown by the imputation models. We found that test compounds that were dissimilar to training compounds, as well as test compounds with a large number of experimental values for other assays, showed the largest improvements. We also investigated the impact of sparsity on the improvements seen as well as the relatedness of the assays being considered. Our results show that even a small amount of additional information can provide imputation methods with a strong boost in predictive performance over traditional single task and multi-task predictive models.

X-ray fluorescence microscopy methods for biological tissues.

Synchrotron-based X-ray fluorescence microscopy is a flexible tool for identifying the distribution of trace elements in biological specimens across a broad range of sample sizes. The technique is not particularly limited by sample type and can be performed on ancient fossils, fixed or fresh tissue specimens, and in some cases even live tissue and live cells can be studied. The technique can also be expanded to provide chemical specificity to elemental maps, either at individual points of interest in a map or across a large field of view. While virtually any sample type can be characterized with X-ray fluorescence microscopy, common biological sample preparation methods (often borrowed from other fields, such as histology) can lead to unforeseen pitfalls, resulting in altered element distributions and concentrations. A general overview of sample preparation and data-acquisition methods for X-ray fluorescence microscopy is presented, along with outlining the general approach for applying this technique to a new field of investigation for prospective new users. Considerations for improving data acquisition and quality are reviewed as well as the effects of sample preparation, with a particular focus on soft tissues. The effects of common sample pretreatment steps as well as the underlying factors that govern which, and to what extent, specific elements are likely to be altered are reviewed along with common artifacts observed in X-ray fluorescence microscopy data.

Trace Impurities Identified as Forensic Signatures in CMX-5 Fuel Pellets Using X-ray Spectroscopic Techniques.

Small-particle analysis is a highly promising emerging forensic tool for analysis of interdicted special nuclear materials. Integration of microstructural, morphological, compositional, and molecular impurity signatures could provide significant advancements in forensic capabilities. We have applied rapid, high-sensitivity, hard X-ray synchrotron chemical imaging to analyze impurity signatures in two differently fabricated fuel pellets from the 5th Collaborative Materials Exercise (CMX5) of the IAEA Nuclear Forensics International Working Group. The spatial distributions, chemical compositions, and morphological and molecular characteristics of impurities were evaluated using X-ray absorption near-edge structure (XANES) and X-ray fluorescence chemical imaging to discover principal impurities, their granularity, particle sizes, modes of occurrence (distinct grains vs incorporation in the UO2 lattice), and sources and mechanisms of incorporation. Differences in UO2+x stoichiometry were detected at the microscale in nominally identical UO2 ceramics (CMX5-A and CMX5-B), implying the presence of multiple UO2 host phases with characteristic microstructures and feedstock compositions. Al, Fe, Ni, W, and Zr impurities and integrated impurity signature analysis identified distinctly different pellet synthesis and processing methods. For example, two different Al, W, and Zr populations in the CMX5-B sample indicated a more complex processing history than the CMX5-A sample. K-edge XANES measurements reveal both metallic and oxide forms of Fe and Ni but with different proportions between each sample. Altogether, these observations suggest multiple sources of impurities, including fabrication (e.g., force-sieving) and feedstock (mineral oxides). This study demonstrates the potential of synchrotron techniques to integrate different signatures across length scales (angstrom to micrometer) to detect and differentiate between contrasting UO2 fuel fabrication techniques.

Reexamination of 2.5-Ga "whiff" of oxygen interval points to anoxic ocean before GOE.

Transient appearances of oxygen have been inferred before the Great Oxygenation Event (GOE) [∼2.3 billion years (Ga) ago] based on redox-sensitive elements such as Mo and S­most prominently from the ∼2.5-Ga Mount McRae Shale in Western Australia. We present new spatially resolved data including synchrotron-based x-ray spectroscopy and secondary ion mass spectrometry to characterize the petrogenesis of the Mount McRae Shale. Sediments were primarily composed of organic matter and volcanic ash (a potential source of Mo), with U-Pb ages revealing extremely low sedimentation rates. Catagenesis created bedding-parallel microfractures, which subsequently acted as fluid pathways for metasomatic alteration and recent oxidative weathering. Our collective observations suggest that the bulk chemical datasets pointing toward a "whiff" of oxygen developed during postdepositional events. Nonzero Δ33S in trace-metal­poor, early diagenetic pyrite and the unusually enriched organic carbon at low sedimentation rates instead suggest that environmental oxygen levels were negligible ∼150 million years before the GOE.

Functional Adaptation of LPS-affected Dentoalveolar Fibrous Joints in Rats.

INTRODUCTION: The functional interplay between cementum of the root and alveolar bone of the socket is tuned by a uniquely positioned 70-80 µm wide fibrous and lubricious ligament in a dentoalveolar joint (DAJ). In this study, structural and biomechanical properties of the DAJ, periodontal ligament space (PDL-space also known as the joint space), alveolar bone of the socket, and cementum of the tooth root that govern the biomechanics of a lipopolysaccharide (LPS)-affected DAJ were mapped both in space and time. METHODS: The hemi-maxillae from 20 rats (4 control at 6 weeks of age, 4 control and 4 LPS-affected at 12 weeks of age, 4 control and 4 LPS-affected at 16 weeks of age) were investigated using a hybrid technique; micro-X-ray computed tomography (5 µm resolution) in combination with biomechanical testing in situ. Temporal variations in bone and cementum volume fractions were evaluated. Trends in mineral apposition rates (MAR) in additional six Sprague Dawley rats (3 controls, 3 LPS-affected) were revealed by transforming spatial fluorochrome signals to functional growth rates (linearity factor - RW) of bone, dentin, and cementum using a fast Fourier transform on fluorochrome signals from 100-µm hemi-maxillae sections. RESULTS: An overall change in LPS-affected DAJ biomechanics (a 2.5-4.5X increase in tooth displacement and 2X tooth rotation at 6 weeks, no increase in displacement and a 7X increase in rotation at 12 weeks; 27% increase in bone effective strain at 6 weeks and 11% at 12 weeks relative to control) was associated with structural changes in the coronal regions of the DAJ (15% increase in PDL-space from 0 to 6 weeks but only 5% from 6 to 12 weeks compared to control). A significant increase (p < 0.05) in PDL-space between ligated and age-matched control was observed. The bone fraction of ligated at 12 weeks was significantly lower than its age-matched control, and no significant differences (p > 0.05) between groups were observed at 6 weeks. Cementum in the apical regions grew faster but nonlinearly (11% and 20% increase in cementum fraction (CF) at 6 and 12 weeks) compared to control. Alveolar bone revealed site-specific nonlinear growth with an overall increase in MAR (108.5 µm/week to 126.7 µm/week after LPS treatment) compared to dentin (28.3 µm/week in control vs. 26.1 µm/week in LPS-affected) and cementum (126.5 µm/week in control vs. 119.9 µm/week in LPS-affected). A significant increase in CF (p < 0.05) in ligated specimens was observed at 6 weeks of age. CONCLUSIONS: Anatomy-specific responses of cementum and bone to the mechano-chemo stimuli, and their collective temporal contribution to observed changes in PDL-space were perpetuated by altered tooth movement. Data highlight the "resilience" of DAJ function through the predominance of nonlinear growth response of cementum, changes in PDL-space, and bone architecture. Despite the significant differences in bone and cementum architectures, data provided insights into the reactionary effects of cementum as a built-in compensatory mechanism to reestablish functional competence of the DAJ. The spatial shifts in architectures of alveolar bone and cementum, and consequently ligament space, highlight adaptations farther away from the site of insult, which also is another novel insight from this study. These adaptations when correlated within the context of joint function (biomechanics) illustrate that they are indeed necessary to sustain DAJ function albeit being pathological.

Electrochemically induced metal- vs. ligand-based redox changes in mackinawite: identification of a Fe3+- and polysulfide-containing intermediate.

Under anaerobic conditions, ferrous iron reacts with sulfide producing FeS, which can then undergo a temperature, redox potential, and pH dependent maturation process resulting in the formation of oxidized mineral phases, such as greigite or pyrite. A greater understanding of this maturation process holds promise for the development of iron-sulfide catalysts, which are known to promote diverse chemical reactions, such as H+, CO2 and NO3- reduction processes. Hampering the full realization of the catalytic potential of FeS, however, is an incomplete knowledge of the molecular and redox processess ocurring between mineral and nanoparticulate phases. Here, we investigated the chemical properties of iron-sulfide by cyclic voltammetry, Raman and X-ray absorption spectroscopic techniques. Tracing oxidative maturation pathways by varying electrode potential, nanoparticulate n(Fe2+S2-)(s) was found to oxidize to a Fe3+ containing FeS phase at -0.5 V vs. Ag/AgCl (pH = 7). In a subsequent oxidation, polysulfides are proposed to give a material that is composed of Fe2+, Fe3+, S2- and polysulfide (Sn2-) species, with its composition described as Fe2+1-3xFe3+2xS2-1-y(Sn2-)y. Thermodynamic properties of model compounds calculated by density functional theory indicate that ligand oxidation occurs in conjunction with structural rearrangements, whereas metal oxidation may occur prior to structural rearrangement. These findings together point to the existence of a metastable FeS phase located at the junction of a metal-based oxidation path between FeS and greigite (Fe2+Fe3+2S2-4) and a ligand-based oxidation path between FeS and pyrite (Fe2+(S2)2-).

Prodromal neuroinflammatory, cholinergic and metabolite dysfunction detected by PET and MRS in the TgF344-AD transgenic rat model of AD: a collaborative multi-modal study.

Mouse models of Alzheimer's disease (AD) are valuable but do not fully recapitulate human AD pathology, such as spontaneous Tau fibril accumulation and neuronal loss, necessitating the development of new AD models. The transgenic (TG) TgF344-AD rat has been reported to develop age-dependent AD features including neuronal loss and neurofibrillary tangles, despite only expressing APP and PSEN1 mutations, suggesting an improved modelling of AD hallmarks. Alterations in neuronal networks as well as learning performance and cognition tasks have been reported in this model, but none have combined a longitudinal, multimodal approach across multiple centres, which mimics the approaches commonly taken in clinical studies. We therefore aimed to further characterise the progression of AD-like pathology and cognition in the TgF344-AD rat from young-adults (6 months (m)) to mid- (12 m) and advanced-stage (18 m, 25 m) of the disease. Methods: TgF344-AD rats and wild-type (WT) littermates were imaged at 6 m, 12 m and 18 m with [18F]DPA-714 (TSPO, neuroinflammation), [18F]Florbetaben (Aß) and [18F]ASEM (α7-nicotinic acetylcholine receptor) and with magnetic resonance spectroscopy (MRS) and with (S)-[18F]THK5117 (Tau) at 15 and 25 m. Behaviour tests were also performed at 6 m, 12 m and 18 m. Immunohistochemistry (CD11b, GFAP, Aß, NeuN, NeuroChrom) and Tau (S)-[18F]THK5117 autoradiography, immunohistochemistry and Western blot were also performed. Results: [18F]DPA-714 positron emission tomography (PET) showed an increase in neuroinflammation in TG vs wildtype animals from 12 m in the hippocampus (+11%), and at the advanced-stage AD in the hippocampus (+12%), the thalamus (+11%) and frontal cortex (+14%). This finding coincided with strong increases in brain microgliosis (CD11b) and astrogliosis (GFAP) at these time-points as assessed by immunohistochemistry. In vivo [18F]ASEM PET revealed an age-dependent increase uptake in the striatum and pallidum/nucleus basalis of Meynert in WT only, similar to that observed with this tracer in humans, resulting in TG being significantly lower than WT by 18 m. In vivo [18F]Florbetaben PET scanning detected Aß accumulation at 18 m, and (S)-[18F]THK5117 PET revealed subsequent Tau accumulation at 25m in hippocampal and cortical regions. Aß plaques were low but detectable by immunohistochemistry from 6 m, increasing further at 12 and 18 m with Tau-positive neurons adjacent to Aß plaques at 18 m. NeuroChrom (a pan neuronal marker) immunohistochemistry revealed a loss of neuronal staining at the Aß plaques locations, while NeuN labelling revealed an age-dependent decrease in hippocampal neuron number in both genotypes. Behavioural assessment using the novel object recognition task revealed that both WT & TgF344-AD animals discriminated the novel from familiar object at 3 m and 6 m of age. However, low levels of exploration observed in both genotypes at later time-points resulted in neither genotype successfully completing the task. Deficits in social interaction were only observed at 3 m in the TgF344-AD animals. By in vivo MRS, we showed a decrease in neuronal marker N-acetyl-aspartate in the hippocampus at 18 m (-18% vs age-matched WT, and -31% vs 6 m TG) and increased Taurine in the cortex of TG (+35% vs age-matched WT, and +55% vs 6 m TG). Conclusions: This multi-centre multi-modal study demonstrates, for the first time, alterations in brain metabolites, cholinergic receptors and neuroinflammation in vivo in this model, validated by robust ex vivo approaches. Our data confirm that, unlike mouse models, the TgF344-AD express Tau pathology that can be detected via PET, albeit later than by ex vivo techniques, and is a useful model to assess and longitudinally monitor early neurotransmission dysfunction and neuroinflammation in AD.

An ecophysiological explanation for manganese enrichment in rock varnish.

Desert varnish is a dark rock coating that forms in arid environments worldwide. It is highly and selectively enriched in manganese, the mechanism for which has been a long-standing geological mystery. We collected varnish samples from diverse sites across the western United States, examined them in petrographic thin section using microscale chemical imaging techniques, and investigated the associated microbial communities using 16S amplicon and shotgun metagenomic DNA sequencing. Our analyses described a material governed by sunlight, water, and manganese redox cycling that hosts an unusually aerobic microbial ecosystem characterized by a remarkable abundance of photosynthetic Cyanobacteria in the genus Chroococcidiopsis as the major autotrophic constituent. We then showed that diverse Cyanobacteria, including the relevant Chroococcidiopsis taxon, accumulate extraordinary amounts of intracellular manganese-over two orders of magnitude higher manganese content than other cells. The speciation of this manganese determined by advanced paramagnetic resonance techniques suggested that the Cyanobacteria use it as a catalytic antioxidant-a valuable adaptation for coping with the substantial oxidative stress present in this environment. Taken together, these results indicated that the manganese enrichment in varnish is related to its specific uptake and use by likely founding members of varnish microbial communities.