•Dopaminergic neurons dominate effective transverse relaxation in nigrosome 1.•Ion beam microscopy reveals highest iron concentrations in dopaminergic neurons.•Developed biophysical model links MRI ...parameters to cellular iron content.•Ferritin- and neuromelanin-bound iron impact MRI parameters differently.•Quantitative MRI provides a potential biomarker of iron in dopaminergic neurons.
In Parkinson’s disease, the depletion of iron-rich dopaminergic neurons in nigrosome 1 of the substantia nigra precedes motor symptoms by two decades. Methods capable of monitoring this neuronal depletion, at an early disease stage, are needed for early diagnosis and treatment monitoring. Magnetic resonance imaging (MRI) is particularly suitable for this task due to its sensitivity to tissue microstructure and in particular, to iron. However, the exact mechanisms of MRI contrast in the substantia nigra are not well understood, hindering the development of powerful biomarkers. In the present report, we illuminate the contrast mechanisms in gradient and spin echo MR images in human nigrosome 1 by combining quantitative 3D iron histology and biophysical modeling with quantitative MRI on post mortem human brain tissue. We show that the dominant contribution to the effective transverse relaxation rate (R2*) in nigrosome 1 originates from iron accumulated in the neuromelanin of dopaminergic neurons. This contribution is appropriately described by a static dephasing approximation of the MRI signal. We demonstrate that the R2* contribution from dopaminergic neurons reflects the product of cell density and cellular iron concentration. These results demonstrate that the in vivo monitoring of neuronal density and iron in nigrosome 1 may be feasible with MRI and provide directions for the development of biomarkers for an early detection of dopaminergic neuron depletion in Parkinson’s disease.
Information about the surface topography is very important to achieve the correct elemental concentrations from the 2D X-ray intensity maps in micro-PIXE. Previously, an innovative approach of ...stereo-PIXE has been introduced to reconstruct the surface topography using two X-ray spectrometers in the lateral direction of the detector positions. Here, we developed the stereo-PIXE method to the four-segment Silicon drift detector (SDD), mounted at the Jožef Stefan Institute (JSI) microprobe, by considering a set of stereovision maps acquired by orthogonal segments to reconstruct the surface topography both in the lateral and longitudinal directions of the scanned area. To this aim, X-ray intensity disparity from the opposing segments was simulated for an ideal flat inclined model of the sample to infer the inclination angle across the opposing segments. Then, the obtained two gradient components of the surface topography from the inclination angles, along the axes that connect the two opposing segments, were reoriented along the scanned area with a proper rotation angle. Thus, to reconstruct the 3-D sample surface topography profile, the integration of the surface topography gradient was numerically performed in the two dimensions along the sample surface. The feasibility of the method is demonstrated on the pure Ti sample with scratched structures.
The increased modularity in the total hip arthroplasty (THA) allows adapting the prosthesis to the specific anatomical characteristics of each patient. However, the advantages of the modular THA are ...shadowed by the increased number of prosthesis failures observed in patients. The presence of junction in modular THA may increase its risk of mechanical failure. Moreover, the micro movements between neck and stem could lead to the production of metallic debris which may cause tissue inflammation and unsealing of prosthesis due to osseous dissolution. It is necessary to understand the mechanism of dispersion of the metal particles from the prosthesis into the tissue. Techniques currently applied in hospitals, such as X-ray scans or optical tissue microscopies, are able to distinguish metal particles, but unable to identify their specific metallic origin. In this work, within the TissueMaps project, we have proved that Proton Induced X-ray Emission (PIXE) is able to provide the distribution and elemental composition of particles from the prosthesis into the pseudo capsular tissue samples (near the femoral head) and identify the features observed under optical microscopy, in a case of broken neck prosthesis.
Porous tantalum has been extensively used in orthopaedic surgery, including uncemented total knee arthroplasty (TKA). Favourable results were reported with earlier monobloc tibial components and the ...design evolved to modular implants. We aimed to analyse possible causes for extensive medial tibia bone loss, resulting in modular porous tantalum tibia baseplate fracture after primary TKA. Retrieved tissue samples were scanned with 3 MeV focused proton beam for Proton-Induced X-ray Emission (micro-PIXE) elemental analysis. Fractographic and microstructural analysis were performed by stereomicroscopy. A full 3D finite-element model was made for numerical analysis of stress-strain conditions of the tibial baseplate. Histological examination of tissue underneath the broken part of the tibial baseplate revealed dark-stained metal debris, which was confirmed by micro-PIXE to consist of tantalum and titanium. Fractographic analysis and tensile testing showed that the failure of the tibial baseplate fulfilled the criteria of a typical fatigue fracture. Microstructural analysis of the contact surface revealed signs of bone ingrowth in 22.5% of the surface only and was even less pronounced in the medial half of the tibial baseplate. Further studies are needed to confirm the responsibility of metal debris for an increased bone absorption leading to catastrophic tibial tray failure.
MeV-SIMS is an emerging mass spectrometry imaging method that employs fast, heavy ions to desorb secondary molecules from the analyzed sample. High yields and low fragmentation rates of large ...molecules, associated with the dominating electronic sputtering process, make it particularly useful in biomedical research, where insight into the distribution of organic molecules is vital. Both yield and fragmentation of desorbed molecules in MeV-SIMS rely on characteristics of the primary ion but may also be impaired by poor instrumental settings. After utilizing secondary ion optics in the linear mass spectrometer at the micro-analytical center of the Jožef Stefan Institute, we demonstrate very efficient detection of secondary ions. As a result, the secondary ion yield, using such settings, solely depends on the species and the characteristics of the primary ion. In order to analyze the yield dependence on the primary ion energy, and the corresponding stopping power within the electronic excitation regime, we used a continuous electron multiplier detector to measure the primary ion current during each measurement of the mass spectra. Secondary ion yield as a function of the primary ion energy and charge is presented as well as fragmentation rates of organic molecules arginine and leu-enkephalin. Other influential instrumental drawbacks are also studied, and their effect on the results is discussed.
Breeding and engineering of biofortified crops will benefit from a better understanding of bottlenecks controlling micronutrient loading within the seeds. However, few studies have addressed the ...changes in micronutrient concentrations, localization, and speciation occurring over time. Therefore, we studied spatial patterns of zinc and iron accumulation during grain development in two barley lines with contrasting grain zinc concentrations. Microparticle-induced-X-ray emission and laser ablation-inductively coupled plasma mass spectrometry were used to determine tissue-specific accumulation of zinc, iron, phosphorus, and sulfur. Differences in zinc accumulation between the lines were most evident in the endosperm and aleurone. A gradual decrease in zinc concentrations from the aleurone to the underlying endosperm was observed, while iron and phosphorus concentrations decreased sharply. Iron co-localized with phosphorus in the aleurone, whereas zinc co-localized with sulfur in the sub-aleurone. We hypothesize that differences in grain zinc are largely explained by the endosperm storage capacity. Engineering attempts should be targeted accordingly.
Genetic biofortification requires knowledge on natural variation and the underlying mechanisms of micronutrient accumulation. We therefore studied diversity in grain micronutrient concentrations and ...spatial distribution in barley (Hordeum vulgare), a genetically tractable model cereal and an important crop with widespread cultivation.
We assembled a diverse collection of barley cultivars and landraces and analysed grain micronutrient profiles in genebank material and after three independent cultivations. Lines with contrasting grain zinc (Zn) accumulation were selected for in-depth analysis of micronutrient distribution within the grain by micro-proton-induced X-ray emission (l-PIXE). Also, we addressed association with grain cadmium (Cd) accumulation.
The analysis of > 120 lines revealed substantial variation, especially in grain Zn concentrations. A large fraction of this variation is due to genetic differences. Grain dissection and l-PIXE analysis of contrasting lines showed that differences in grain Zn accumulation apply to all parts of the grain including the endosperm. Cd concentrations exceeded the Codex Alimentarius threshold in most of the representative barley lines after cultivation in a Cd-contaminated agricultural soil.
Two important conclusions for biofortification are: first, high-Zn grains contain more Zn also in the consumed parts of the grain; and second, higher micronutrient concentrations are strongly associated with higher Cd accumulation.
• Lead (Pb) ranks among the most problematic environmental pollutants. Background contamination of soils is nearly ubiquitous, yet plant Pb accumulation is barely understood. In a survey covering 165 ...European populations of the metallophyte Arabidopsis halleri, several field samples had indicated Pb hyperaccumulation, offering a chance to dissect plant Pb accumulation.
• Accumulation of Pb was analysed in A. halleri individuals from contrasting habitats under controlled conditions to rule out aerial deposition as a source of apparent Pb accumulation. Several elemental imaging techniques were employed to study the spatial distribution and ligand environment of Pb.
• Regardless of genetic background, A. halleri individuals showed higher shoot Pb accumulation than A. thaliana. However, dose–response curves revealed indicator rather than hyperaccumulator behaviour. Xylem sap data and elemental imaging unequivocally demonstrated the in planta mobility of Pb. Highest Pb concentrations were found in epidermal and vascular tissues. Distribution of Pb was distinct from that of the hyperaccumulated metal zinc. Most Pb was bound by oxygen ligands in bidentate coordination.
• A. halleri accumulates Pb whenever soil conditions render Pb phytoavailable. Considerable Pb accumulation under such circumstances, even in leaves of A. thaliana, strongly suggests that Pb can enter food webs and may pose a food safety risk.
Phosphorus (P) deficiency limits crop production worldwide. Crops differ in their ability to acquire and utilise the P available. The aim of this study was to determine root traits (root exudates, ...root system architecture (RSA), tissue-specific allocation of P, and gene expression in roots) that (a) play a role in P-use efficiency and (b) contribute to large shoot zinc (Zn) concentration in Brassica oleracea.
Two B. oleracea accessions (var. sabellica C6, a kale, and var. italica F103, a broccoli) were grown in a hydroponic system or in a high-throughput-root phenotyping (HTRP) system where they received Low P (0.025 mM) or High P (0.25 mM) supply for 2 weeks. In hydroponics, root and shoot P and Zn concentrations were measured, root exudates were profiled using both Fourier-Transform-Infrared spectroscopy and gas-chromatography-mass spectrometry and previously published RNAseq data from roots was re-examined. In HTRP experiments, RSA (main and lateral root number and lateral root length) was assessed and the tissue-specific distribution of P was determined using micro-particle-induced-X-ray emission. The C6 accession had greater root and shoot biomass than the F103 accession, but the latter had a larger shoot P concentration than the C6 accession, regardless of the P supply in the hydroponic system. The F103 accession had a larger shoot Zn concentration than the C6 accession in the High P treatment. Although the F103 accession had a larger number of lateral roots, which were also longer than in the C6 accession, the C6 accession released a larger quantity and number of polar compounds than the F103 accession. A larger number of P-responsive genes were found in the Low P treatment in roots of the F103 accession than in roots of the C6 accession. Expression of genes linked with "phosphate starvation" was up-regulated, while those linked with iron homeostasis were down-regulated in the Low P treatment.
The results illustrate large within-species variability in root acclimatory responses to P supply in the composition of root exudates, RSA and gene expression, but not in P distribution in root cross sections, enabling P sufficiency in the two B. oleracea accessions studied.
In the present work we have investigated the chemical sensitivity of a mass spectrometry imaging method MeV – SIMS. Primary ion beam within the MeV energy range domain was employed to bombard samples ...of two organic compounds; amino acid arginine, and peptide hormone angiotensin II (human), with average molecular weights of 174.2 u and 1046.2 u respectively. Secondary ion yield was measured as a function of number of molecules per area unit, and the detection limit was determined. For both molecular compounds and two different energies of primary 35Cl ion beam, the secondary ion yield exhibited a significant decrease below the area density of 1015 molecules/cm2, while the density of 1013 molecules/cm2 resulted in molecular peak / background ratio being lesser than 3, which is below the commonly used sensitivity threshold in other techniques. Other experiment related drawbacks to sensitivity were also discussed.
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