Sequential removal of cation/H+ exchangers reveals their additive role in elemental distribution, calcium depletion and anoxia tolerance.

Multiple Arabidopsis H+ /Cation exchangers (CAXs) participate in high-capacity transport into the vacuole. Previous studies have analysed single and double mutants that marginally reduced transport; however, assessing phenotypes caused by transport loss has proven enigmatic. Here, we generated quadruple mutants (cax1-4: qKO) that exhibited growth inhibition, an 85% reduction in tonoplast-localised H+ /Ca transport, and enhanced tolerance to anoxic conditions compared to CAX1 mutants. Leveraging inductively coupled plasma mass spectrometry (ICP-MS) and synchrotron X-ray fluorescence (SXRF), we demonstrate CAX transporters work together to regulate leaf elemental content: ICP-MS analysis showed that the elemental concentrations in leaves strongly correlated with the number of CAX mutations; SXRF imaging showed changes in element partitioning not present in single CAX mutants and qKO had a 40% reduction in calcium (Ca) abundance. Reduced endogenous Ca may promote anoxia tolerance; wild-type plants grown in Ca-limited conditions were anoxia tolerant. Sequential reduction of CAXs increased mRNA expression and protein abundance changes associated with reactive oxygen species and stress signalling pathways. Multiple CAXs participate in postanoxia recovery as their concerted removal heightened changes in postanoxia Ca signalling. This work showcases the integrated and diverse function of H+ /Cation transporters and demonstrates the ability to improve anoxia tolerance through diminishing endogenous Ca levels.

Deciduous teeth from the New Hampshire birth cohort study: Early life environmental and dietary predictors of dentin elements.

BACKGROUND: Sparse research exists on predictors of element concentrations measured in deciduous teeth. OBJECTIVE: To estimate associations between maternal/child characteristics, elements measured in home tap water during pregnancy and element concentrations in the dentin of shed deciduous teeth. METHODS: Our analysis included 152 pregnant person-infant dyads followed from the second trimester through the end of the first postnatal year from the New Hampshire Birth Cohort Study. During pregnancy and early infancy, we collected dietary and sociodemographic information via surveys, measured elements in home tap water, and later collected naturally exfoliated teeth from child participants. We measured longitudinal deposition of elements in dentin using LA-ICP-MS. Multivariable linear mixed models were used to estimate associations between predictors and dentin element concentrations. RESULTS: We measured 12 elements in dentin including those previously reported (Ba, Mn, Pb, Sr, Zn) and less frequently reported (Al, As, Cd, Cu, Hg, Li, and W). A doubling of Pb or Sr concentrations in water was associated with higher dentin Pb or Sr respectively in prenatally formed [9% (95%CI: 3%, 15%); 3% (1%, 6%)] and postnatally formed [10% (2%, 19%); 6% (2%, 10%)] dentin. Formula feeding from birth to 6 weeks or 6 weeks to 4 months was associated with higher element concentrations in postnatal dentin within the given time period as compared to exclusive human milk feeding: Sr: 6 weeks: 61% (36%, 90%) and 4 months: 85% (54%, 121%); Ba: 6 weeks: 35% (3.3%, 77%) and 4 months: 42% (10%, 83%); and Li: 6 weeks: 61% (33%, 95%) and 4 months: 58% (31%, 90%). SIGNIFICANCE: These findings offer insights into predictors of dentin elements and potential confounders in exposure-health outcome relationships during critical developmental periods.

Human placental microRNAs dysregulated by cadmium exposure predict neurobehavioral outcomes at birth.

BACKGROUND: Prenatal cadmium (Cd) exposure has been implicated in both placental toxicity and adverse neurobehavioral outcomes. Placental microRNAs (miRNAs) may function to developmentally program adverse pregnancy and newborn health outcomes in response to gestational Cd exposure. METHODS: In a subset of the Rhode Island Child Health Study (RICHS, n = 115) and the New Hampshire Birth Cohort Study (NHBCS, = 281), we used small RNA sequencing and trace metal analysis to identify Cd-associated expression of placental miRNAs using negative binomial generalized linear models. We predicted mRNAs targeted by Cd-associated miRNAs and relate them to neurobehavioral outcomes at birth through the integration of transcriptomic data and summary scores from the NICU Network Neurobehavioral Scale (NNNS). RESULTS: Placental Cd concentrations are significantly associated with the expression level of five placental miRNAs in NHBCS, with similar effect sizes in RICHS. These miRNA target genes overrepresented in nervous system development, and their expression is correlated with NNNS metrics suggestive of atypical neurobehavioral outcomes at birth. CONCLUSIONS: Gestational Cd exposure is associated with the expression of placental miRNAs. Predicted targets of these miRNAs are involved in nervous system development and may also regulate placental physiology, allowing their dysregulation to modify developmental programming of early life health outcomes. IMPACT: This research aims to address the poor understanding of the molecular mechanisms governing adverse pregnancy and newborn health outcomes in response to Gestational cadmium (Cd) exposure. Our results outline a robust relationship between Cd-associated placental microRNA expression and NICU Network Neurobehavioral Scales (NNNS) at birth indicative of atypical neurobehavior. This study utilized healthy mother-infant cohorts to describe the role of Cd-associated dysregulation of placental microRNAs as a potential mechanism by which adverse neurobehavioral outcomes are developmentally programmed.

Redundant roles of four ZIP family members in zinc homeostasis and seed development in Arabidopsis thaliana.

Zinc (Zn) is essential for normal plant growth and development. The Zn-regulated transporter, iron-regulated transporter (IRT)-like protein (ZIP) family members are involved in Zn transport and cellular Zn homeostasis throughout the domains of life. In this study, we have characterized four ZIP transporters from Arabidopsis thaliana (IRT3, ZIP4, ZIP6, and ZIP9) to better understand their functional roles. The four ZIP proteins can restore the growth defect of a yeast Zn uptake mutant and are upregulated under Zn deficiency. Single and double mutants show no phenotypes under Zn-sufficient or Zn-limited growth conditions. In contrast, triple and quadruple mutants show impaired growth irrespective of external Zn supply due to reduced Zn translocation from root to shoot. All four ZIP genes are highly expressed during seed development, and siliques from all single and higher-order mutants exhibited an increased number of abnormal seeds and decreased Zn levels in mature seeds relative to wild type. The seed phenotypes could be reversed by supplementing the soil with Zn. Our data demonstrate that IRT3, ZIP4, ZIP6, and ZIP9 function redundantly in maintaining Zn homeostasis and seed development in A. thaliana.

Association between placental toxic metal exposure and NICU Network Neurobehavioral Scales (NNNS) profiles in the Rhode Island Child Health Study (RICHS).

BACKGROUND: Prenatal exposure to heavy metals has been linked to a variety of adverse outcomes in newborn health and later life. Toxic metals such as cadmium (Cd), manganese (Mn) and lead (Pb) have been implicated to negatively affect newborn neurobehavior. Placental levels of these metals may provide additional understandings on the link between prenatal toxic metal exposures and neurobehavioral performances in newborns. OBJECTIVE: To evaluate associations between placental concentrations of toxic metals and newborn neurobehavioral performance indicated through the NICU Network Neurobehavioral Scales (NNNS) latent profiles. METHOD: In the Rhode Island Child Health Study cohort (n = 625), newborn neurobehavioral performance was assessed with NNNS, and a latent profile analysis was used to define five discrete neurobehavioral profiles based on summary scales. Using multinomial logistic regression, we determined whether increased levels of placental toxic metals Cd, Mn and Pb associated with newborns assigned to the profile demonstrating atypical neurobehavioral performances. RESULTS: Every doubling in placenta Cd concentration was associated with increased odds of newborns belonging to the atypical neurobehavior profile (OR: 2.72, 95% CI [1.09, 6.79]). Detectable placental Pb also demonstrated an increased odds of newborns assignment to the atypical profile (OR: 3.71, 95% CI [0.97, 13.96]) compared to being in the typical neurobehavioral profile. CONCLUSIONS: Toxic metals Cd and Pb measured in placental tissue may adversely impact newborn neurobehavior. Utilizing the placenta as a prenatal toxic metal exposure biomarker is useful in elucidating the associated impacts of toxic metals on newborn health.

Distribution and accumulation of mercury in pot-grown African rice cultivars (Oryza glaberrima Steud. and Oryza sativa L.) determined via LA-ICP-MS.

There is limited information concerning the distribution of mercury in rice, particularly in African rice. The objective was to compare the distribution of total mercury (THg) and methylmercury (MeHg) in African rice (Oryza glaberrima Steud.) and Asian rice (O. sativa L.). It is hypothesized that increased mineral accumulation and greater stress tolerance in O. glaberrima will affect the uptake and distribution of THg and MeHg, compared to O. sativa. Rice varieties from the Republic of Mali, including O. glaberrima (n =1) and O. sativa (n = 2), were cultivated in a greenhouse, in mercury-spiked soil (50 mg/kg) (n =3 replicates/variety). THg and MeHg concentrations were analyzed in the grain (brown rice), and the THg distribution was analyzed using laser ablation inductively coupled-plasma mass spectrometry (LA-ICP-MS). THg and MeHg concentrations did not differ between O. glaberrima and O. sativa grain. However, in both O. sativa varieties, THg was highly concentrated in the scutellum, which surrounds the embryo and is removed during polishing. Conversely, in O. glaberrima grain, THg was widely distributed throughout the endosperm, the edible portion of the grain. Differences in the THg distribution in O. glaberrima grain, compared to O. sativa, may elevate the risk of mercury exposure through ingestion of polished rice. The novelty of this study includes the investigation of a less-studied rice species (O. glaberrima), the use of a highly sensitive elemental imaging technique (LA-ICP-MS), and its finding of a different grain THg distribution in O. glaberrima than has been observed in O. sativa.

Functional overlap of two major facilitator superfamily transporter, ZIF1, and ZIFL1 in zinc and iron homeostasis.

Zinc and iron are essential micronutrients for plant growth, and their homeostasis must be tightly regulated. Previously, it has been shown that Zinc-Induced Facilitator 1 (ZIF1) is involved in basal Zn tolerance by controlling the vacuolar storage of nicotianamine (NA). However, knowledge of the functional roles of two ZIF1 paralogs, ZIF-LIKE1 (ZIFL1) and ZIFL2, in metal homeostasis remains limited. Here, we functionally characterized the roles of ZIF1, ZIFL1, and ZIFL2 in Zn and Fe homeostasis. Expression of ZIF1 and ZIFL1 was induced by both excess Zn and Fe-deficiency, and their loss-of-function led to hypersensitivity under excess Zn and Fe-deficiency, suggesting functional overlap between ZIF1 and ZIFL1. By contrast, the disruption of ZIFL2 resulted in no obvious phenotypic alteration under both conditions. Additionally, the expression of ZIFL1, but not that of ZIFL2, in the zif1 mutant partially restored the phenotype under excess Zn, suggesting that ZIF1 and ZIFL1 perform functionally redundant roles in Zn homeostasis.

Synchrotron-Based X-Ray Fluorescence Microscopy as a Technique for Imaging of Elements in Plants.

Understanding the distribution of elements within plant tissues is important across a range of fields in plant science. In this review, we examine synchrotron-based x-ray fluorescence microscopy (XFM) as an elemental imaging technique in plant sciences, considering both its historical and current uses as well as discussing emerging approaches. XFM offers several unique capabilities of interest to plant scientists, including in vivo analyses at room temperature and pressure, good detection limits (approximately 1-100 mg kg-1), and excellent resolution (down to 50 nm). This has permitted its use in a range of studies, including for functional characterization in molecular biology, examining the distribution of nutrients in food products, understanding the movement of foliar fertilizers, investigating the behavior of engineered nanoparticles, elucidating the toxic effects of metal(loid)s in agronomic plant species, and studying the unique properties of hyperaccumulating plants. We anticipate that continuing technological advances at XFM beamlines also will provide new opportunities moving into the future, such as for high-throughput screening in molecular biology, the use of exotic metal tags for protein localization, and enabling time-resolved, in vivo analyses of living plants. By examining current and potential future applications, we hope to encourage further XFM studies in plant sciences by highlighting the versatility of this approach.

Elemental distribution in developing rice grains and the effect of flag-leaf arsenate exposure.

We measured the bulk grain concentrations of arsenic (As), along with rubidium (Rb) and strontium (Sr) as indicators of phloem and xylem transport respectively, in rice (Oryza sativa cv. Italica Carolina) pulsed with arsenate at two exposure levels for 5 day periods at progressively later stages of grain fill, between anthesis and maturity, through the cut flag leaf. We compared these to unexposed (negative) controls and positive controls; pulsed with dimethylarsinic acid (DMA). We collected elemental maps of As and micronutrient elements (Fe, Zn, Mn, Cu and Ni) from developing grains of rice. Exposures were either 25 or 100 µg/ml arsenate (As(V)) at various stages of grain development, compared to 25 µg/ml dimethylarsinic acid (DMA); the most efficiently transported As species identified in rice. We used the spatial distribution of arsenic in the grain to infer the presence of As transporters. By exposing grains through the flag leaf rather than via the roots, we were able to measure arsenic transport into the grain during filling under controlled conditions. Exposure to 100 µg/ml As(V) resulted in widespread As localization in both embryo and endosperm, especially in grains exposed to As at later stages of panicle development. This suggests loss of selective transport, likely to be the result of As toxicity. At 25 µg/ml As(V), As colocalized with Mn in the ovular vascular trace (OVT). Exposure to either As(V) or DMA reduced grain Fe, an effect more pronounced when exposure occurred earlier in grain development. The abundance of Cu and Zn were also reduced by As. Arsenic exposure later in grain development caused higher grain As concentrations, indicating the existence of As transporters whose efficiency increases during grain fill. We conclude that localization of As in the grain is a product of both As species and exposure concentration, and that high As(V) translocation from the flag leaf can result in high As concentrations in the endosperm.

OPT3 Is a Phloem-Specific Iron Transporter That Is Essential for Systemic Iron Signaling and Redistribution of Iron and Cadmium in Arabidopsis.

Iron is essential for both plant growth and human health and nutrition. Knowledge of the signaling mechanisms that communicate iron demand from shoots to roots to regulate iron uptake as well as the transport systems mediating iron partitioning into edible plant tissues is critical for the development of crop biofortification strategies. Here, we report that OPT3, previously classified as an oligopeptide transporter, is a plasma membrane transporter capable of transporting transition ions in vitro. Studies in Arabidopsis thaliana show that OPT3 loads iron into the phloem, facilitates iron recirculation from the xylem to the phloem, and regulates both shoot-to-root iron signaling and iron redistribution from mature to developing tissues. We also uncovered an aspect of crosstalk between iron homeostasis and cadmium partitioning that is mediated by OPT3. Together, these discoveries provide promising avenues for targeted strategies directed at increasing iron while decreasing cadmium density in the edible portions of crops and improving agricultural productivity in iron deficient soils.

Maternal exposure to selenium and cadmium, fetal growth, and placental expression of steroidogenic and apoptotic genes.

BACKGROUND: Cadmium (Cd) and selenium (Se) antagonistically influence redox balance and apoptotic signaling, with Cd potentially promoting and Se inhibiting oxidative stress and apoptosis. Alterations to placental redox and apoptotic functions by maternal exposure to Cd and Se during pregnancy may explain some of the Cd and Se associations with fetal development. OBJECTIVES: Investigate associations between Cd and Se concentrations in maternal toenails with placental expression patterns of tumor necrosis factor (TNF) and steroidogenic genes involved in redox reactions and test associations with fetal growth. METHODS: In a sub-sample from the Rhode Island Child Health Study (n = 173), we investigated the relationships between: (1) maternal toenail Cd and Se concentrations and fetal growth using logistic regression, (2) Cd and Se interactions with factor scores from placental TNF and steroidogenic expression patterns (RNAseq) using linear models, and (3) TNF and steroidogenic expression factors with fetal growth via analysis of covariance. RESULTS: Se was associated with decreased odds of intrauterine growth restriction (IUGR) (OR = 0.27, p-value = 0.045). Cd was associated with increased odds of IUGR (OR = 1.95, p-value = 0.13) and small for gestational age (SGA) births (OR = 1.46, p-value = 0.11), though not statistically significant. Cd and Se concentrations were antagonistically associated with placental TNF and steroidogenic expression patterns, which also differed by birth size. CONCLUSIONS: Se may act as an antagonist to Cd and as a modifiable protective factor in fetal growth restriction, and these data suggest these effects may be due to associated variations in the regulation of genes involved in placental redox balance and/or apoptotic signaling.

Bypassing Iron Storage in Endodermal Vacuoles Rescues the Iron Mobilization Defect in the natural resistance associated-macrophage protein3natural resistance associated-macrophage protein4 Double Mutant.

To improve seed iron (Fe) content and bioavailability, it is crucial to decipher the mechanisms that control Fe storage during seed development. In Arabidopsis (Arabidopsis thaliana) seeds, most Fe is concentrated in insoluble precipitates, with phytate in the vacuoles of cells surrounding the vasculature of the embryo. NATURAL RESISTANCE ASSOCIATED-MACROPHAGE PROTEIN3 (AtNRAMP3) and AtNRAMP4 function redundantly in Fe retrieval from vacuoles during germination. When germinated under Fe-deficient conditions, development of the nramp3nramp4 double mutant is arrested as a consequence of impaired Fe mobilization. To identify novel genes involved in seed Fe homeostasis, we screened an ethyl methanesulfonate-mutagenized population of nramp3nramp4 seedlings for mutations suppressing their phenotypes on low Fe. Here, we report that, among the suppressors, two independent mutations in the VACUOLAR IRON TRANSPORTER1 (AtVIT1) gene caused the suppressor phenotype. The AtVIT1 transporter is involved in Fe influx into vacuoles of endodermal and bundle sheath cells. This result establishes a functional link between Fe loading in vacuoles by AtVIT1 and its remobilization by AtNRAMP3 and AtNRAMP4. Moreover, analysis of subcellular Fe localization indicates that simultaneous disruption of AtVIT1, AtNRAMP3, and AtNRAMP4 limits Fe accumulation in vacuolar globoids.

Placental Metal Concentrations in Relation to Maternal and Infant Toenails in a U.S. Cohort.

Metal contaminants cross the placenta, presenting a heightened risk of perturbing fetal development. Information about placental concentrations and transfer of multiple potentially toxic metals from low to moderate exposure is lacking. We measured concentrations of Cd, Pb, Hg, Mn, Se, and Zn in 750 placentas collected from women enrolled in the New Hampshire Birth Cohort Study and examined the correlation between elements, and profiles of potentially toxic metals (Cd, Pb, Hg, and Mn) stratified by nutrient concentrations (Zn and Se) using principal components analyses. We further examined the indirect effects of maternal metal concentrations on infant metal concentrations through placental metal concentrations using structural equation models. Placental metal concentrations were all correlated, particularly Zn and Mn, and Zn and Cd, and the principal component of metals differed by stratum of high versus low Zn and Se. Associations were observed between placenta and maternal toenail Se (ß = 63.49; P < 0.0001) and Pb (ß = 0.90; P < 0.0001) but not other metals. Structural equation models did not indicate any statistically significant indirect effects through placental metal concentrations. Placental metal concentrations may represent a distinct biomarker of metal exposure and adverse health impacts to the fetus, particularly those stemming from the placenta.

High-resolution elemental mapping of human placental chorionic villi using synchrotron X-ray fluorescence spectroscopy.

The placenta is the organ that mediates transport of nutrients and waste materials between mother and fetus. Synchrotron X-ray fluorescence (SXRF) microanalysis is a tool for imaging the distribution and quantity of elements in biological tissue, which can be used to study metal transport across biological membranes. Our aims were to pilot placental biopsy specimen preparation techniques that could be integrated into an ongoing epidemiology birth cohort study without harming rates of sample acquisition. We studied the effects of fixative (formalin or glutaraldehyde) and storage duration (30 days or immediate processing) on metal distribution and abundance and investigated a thaw-fixation protocol for archived specimens stored at -80 °C. We measured fixative elemental composition with and without a placental biopsy via inductively coupled plasma mass spectrometry (ICP-MS) to quantify fixative-induced elemental changes. Formalin-fixed specimens showed hemolysis of erythrocytes. The glutaraldehyde-paraformaldehyde solution in HEPES buffer (GTA-HEPES) had superior anatomical preservation, avoided hemolysis, and minimized elemental loss, although some cross-linking of exogenous Zn was evident. Elemental loss from tissue stored in fixative for 1 month showed variable losses (≈40 % with GTA-HEPES), suggesting storage duration be controlled for. Thawing of tissue held at -80 °C in a GTA-HEPES solution provided high-quality visual images and elemental images.

Contrasting calcium localization and speciation in leaves of the Medicago truncatula mutant cod5 analyzed via synchrotron X-ray techniques.

Oxalate-producing plants accumulate calcium oxalate crystals (CaOx(c)) in the range of 3-80% w/w of their dry weight, reducing calcium (Ca) bioavailability. The calcium oxalate deficient 5 (cod5) mutant of Medicago truncatula has been previously shown to contain similar Ca concentrations to wild-type (WT) plants, but lower oxalate and CaOx(c) concentrations. We imaged the Ca distribution in WT and cod5 leaflets via synchrotron X-ray fluorescence mapping (SXRF). We observed a difference in the Ca distribution between cod5 and WT leaflets, manifested as an abundance of Ca in the interveinal areas and a lack of Ca along the secondary veins in cod5, i.e. the opposite of what is observed in WT. X-ray microdiffraction (µXRD) of M. truncatula leaves confirmed that crystalline CaOx(c) (whewellite; CaC2 O4 · H2 O) was present in the WT only, in cells sheathing the secondary veins. Together with µXRD, microbeam Ca K-edge X-ray absorption near-edge structure spectroscopy (µXANES) indicated that, among the forms of CaOx, i.e. crystalline or amorphous, only amorphous CaOx was present in cod5. These results demonstrate that deletion of COD5 changes both Ca localization and the form of CaOx within leaflets.

Rice consumption contributes to arsenic exposure in US women.

Emerging data indicate that rice consumption may lead to potentially harmful arsenic exposure. However, few human data are available, and virtually none exist for vulnerable periods such as pregnancy. Here we document a positive association between rice consumption and urinary arsenic excretion, a biomarker of recent arsenic exposure, in 229 pregnant women. At a 6-mo prenatal visit, we collected a urine sample and 3-d dietary record for water, fish/seafood, and rice. We also tested women's home tap water for arsenic, which we combined with tap water consumption to estimate arsenic exposure through water. Women who reported rice intake (n = 73) consumed a median of 28.3 g/d, which is ∼0.5 cup of cooked rice each day. In general linear models adjusted for age and urinary dilution, both rice consumption (g, dry mass/d) and arsenic exposure through water (µg/d) were significantly associated with natural log-transformed total urinary arsenic (ßrice = 0.009, ßwater = 0.028, both P < 0.0001), as well as inorganic arsenic, monomethylarsonic acid, and dimethylarsinic acid (each P < 0.005). Based on total arsenic, consumption of 0.56 cup/d of cooked rice was comparable to drinking 1 L/d of 10 µg As/L water, the current US maximum contaminant limit. US rice consumption varies, averaging ∼0.5 cup/d, with Asian Americans consuming an average of >2 cups/d. Rice arsenic content and speciation also vary, with some strains predominated by dimethylarsinic acid, particularly those grown in the United States. Our findings along with others indicate that rice consumption should be considered when designing arsenic reduction strategies in the United States.

Urinary biomarkers of exposure to toxic and essential elements: A comparison of infants fed with human milk or formula.

Background: Early-life exposure to nonessential (toxic) and essential trace elements can influence child development. Although infant formula powders and the water used to reconstitute them can contain higher concentrations of many elements compared with human milk, the influence of feeding mode on reliable biomarkers of infant exposure has rarely been demonstrated. Methods: We evaluated associations between urinary biomarkers and feeding mode (exclusively human milk, exclusively formula, or combination-fed) for four toxic (arsenic, cadmium, nickel, and uranium) and three essential elements (cobalt, molybdenum, and selenium) using general linear models. Results: A total of 462 participants from the rural New Hampshire Birth Cohort Study were on average 6 weeks old between July 2012 and March 2019 and had urine samples, 3-day food diaries, and relevant covariate data available. In adjusted models, urinary arsenic was 5.15 (95% confidence interval = 4.04, 6.58), molybdenum was 19.02 (14.13-25.59), and selenium was 1.51 (1.35-1.68) times higher in infants fed exclusively with formula compared with infants fed exclusively with human milk. By contrast, urinary uranium was 0.59 (0.46-0.75) and cobalt was 0.78 (0.65-0.95) times lower with formula feeding than human milk feeding. Conclusion: Our findings suggest that infant exposure to several potentially toxic elements varies by feeding mode, as concentrations of reliable urinary biomarkers were higher with formula or human milk, depending on the element. Importantly, exposure to arsenic increased with household tap water arsenic regardless of feeding mode, suggesting that all infants could be at risk in populations with high concentrations of arsenic in drinking water.

Integrative Co-Registration of Elemental Imaging and Histopathology for Enhanced Spatial Multimodal Analysis of Tissue Sections through TRACE.

Summary: Elemental imaging provides detailed profiling of metal bioaccumulation, offering more precision than bulk analysis by targeting specific tissue areas. However, accurately identifying comparable tissue regions from elemental maps is challenging, requiring the integration of hematoxylin and eosin (H&E) slides for effective comparison. Facilitating the streamlined co-registration of Whole Slide Images (WSI) and elemental maps, TRACE enhances the analysis of tissue regions and elemental abundance in various pathological conditions. Through an interactive containerized web application, TRACE features real-time annotation editing, advanced statistical tools, and data export, supporting comprehensive spatial analysis. Notably, it allows for comparison of elemental abundances across annotated tissue structures and enables integration with other spatial data types through WSI co-registration. Availability and Implementation: Available on the following platforms- GitHub: jlevy44/trace_app , PyPI: trace_app , Docker: joshualevy44/trace_app , Singularity: joshualevy44/trace_app . Contact: joshua.levy@cshs.org. Supplementary information: Supplementary data are available.

Lead exposure across the life course and age at death.

BACKGROUND: Lead (Pb) exposure has been associated with an increased risk of all-cause mortality, even at low levels. Little is known about how the timing of Pb exposure throughout life may influence these relationships. Quantifying the amount of Pb present in various tissues of the body provides measurements of exposure from different periods of life. These include bone, tooth enamel, which is the hard outer layer of the crown, and tooth cementum, which is the calcified connective tissue covering the tooth root. The purpose of the study was to examine Pb exposure at multiple periods throughout life, including childhood (enamel), adulthood (cementum), and later life (bone), and to estimate their associations with age at death. METHODS: 208 skeleton donors (born 1910-1960) from an ongoing case-control study were included in this study. Pb was measured in tibia (shin), bone using X-Ray Florescence and in teeth using Laser-Ablation Inductively Coupled Plasma Mass Spectroscopy. After excluding unusually high measurements (>2sd), this resulted in a final sample of 111 with all exposure measures. Correlations across measures were determined using partial Spearman correlations. Associations between Pb exposure and age at death were estimated using Multivariable Linear Regression. RESULTS: Pb measures across exposure periods were all significantly correlated, with the highest correlation between cementum and tibia measures (r = 0.61). Donors were largely female (63.0 %), White (97.3 %), and attended some college (49.5 %). Single exposure models found that higher tooth cementum Pb (-1.27; 95 % CI: -2.48, -0.06) and tibia bone Pb (-0.91; 95 % CI: -1.67, -0.15) were significantly associated with an earlier age at death. When considered simultaneously, only cementum Pb remained significant (-1.51; 95 % CI: -2.92, -0.11). Secondary analyses suggest that the outer cementum Pb may be especially associated with an earlier age at death. CONCLUSION: Results suggest that higher Pb exposure is associated with an earlier age at death, with adulthood as the life period of most relevance. Additional studies using Pb exposure measures from different life stages should be conducted.

The role of CAX1 and CAX3 in elemental distribution and abundance in Arabidopsis seed.

The ability to alter nutrient partitioning within plants cells is poorly understood. In Arabidopsis (Arabidopsis thaliana), a family of endomembrane cation exchangers (CAXs) transports Ca(2+) and other cations. However, experiments have not focused on how the distribution and partitioning of calcium (Ca) and other elements within seeds are altered by perturbed CAX activity. Here, we investigate Ca distribution and abundance in Arabidopsis seed from cax1 and cax3 loss-of-function lines and lines expressing deregulated CAX1 using synchrotron x-ray fluorescence microscopy. We conducted 7- to 10-µm resolution in vivo x-ray microtomography on dry mature seed and 0.2-µm resolution x-ray microscopy on embryos from lines overexpressing deregulated CAX1 (35S-sCAX1) and cax1cax3 double mutants only. Tomograms showed an increased concentration of Ca in both the seed coat and the embryo in cax1, cax3, and cax1cax3 lines compared with the wild type. High-resolution elemental images of the mutants showed that perturbed CAX activity altered Ca partitioning within cells, reducing Ca partitioning into organelles and/or increasing Ca in the cytosol and abolishing tissue-level Ca gradients. In comparison with traditional volume-averaged metal analysis, which confirmed subtle changes in seed elemental composition, the collection of spatially resolved data at varying resolutions provides insight into the impact of altered CAX activity on seed metal distribution and indicates a cell type-specific function of CAX1 and CAX3 in partitioning Ca into organelles. This work highlights a powerful technology for inferring transport function and quantifying nutrient changes.