Renal localization and quantification of uranium in rodent exposed to uranyl nitrate by LA-ICP-MS

International audienceThe over-exposure to uranium results in renal toxicity, which is derived form an excessive accumulation of the element in the kidney, characterized by reduced glomerular filtration, exerting its toxic effect by chemical and radiological action [1-3]. In addition, its tissue distribution is heterogeneous mostly accumulating and producing pathological lesions in the S2 and S3 segments of the proximal tubule located in the cortical zone (100-fold above mean renal concentration) [4-6]. Chronic exposure (occupational exposure) to uranium, is linked to its bioaccumulation in kidney and could be associated with renal dysfunction, an increased risk of cancer mortality and kidney failure [7-9]. Up to day, few quantitative studies have been carried out employing high-energy synchrotron radiation X-ray fluorescence analysis (SR-XRF) and X-ray absorption fine structure (XAFS) [10, 11]. Unfortunately, the limited access to these analytical techniques limits considerably its daily use for routine analysis. Therefore, this works presents the development of an analytical methodology based on mass spectrometry imaging (MSI) using laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) for mapping and quantifying uranium in histological tissue sections of mouse kidney. To the author’s knowledge a single work has been published recently for the semi-quantitative analysis of uranium in mice kidneys due to the lack of an appropriate internal standard [12]. In this study, the uranium exposure of mice was performed using a direct nasal instillation of few µL of uranyl nitrate solutions at different uranium concentrations. Total uranium analysis was done by conventional liquid ICP-MS after acid digestion using one kidney from each pair. In parallel, the other kidney was employed for LA-ICP-MS analysis. In order to perform quantitative LA-ICPMS bio-imaging analysis, synthetic matrix-matched laboratory standards and a normalisation strategy based on internal standard spiked gelatine were developed. Preliminary quantitative images of cryo-sections revealed heterogeneous distribution of uranium within the renal tissue, being the cortical concentration 60-fold higher than the medullary concentration

Renal localization and quantification of uranium in rodent exposed to uranyl nitrate by LA-ICP-MS

International audienceThe over-exposure to uranium results in renal toxicity, which is derived form an excessive accumulation of the element in the kidney, characterized by reduced glomerular filtration, exerting its toxic effect by chemical and radiological action [1-3]. In addition, its tissue distribution is heterogeneous mostly accumulating and producing pathological lesions in the S2 and S3 segments of the proximal tubule located in the cortical zone (100-fold above mean renal concentration) [4-6]. Chronic exposure (occupational exposure) to uranium, is linked to its bioaccumulation in kidney and could be associated with renal dysfunction, an increased risk of cancer mortality and kidney failure [7-9]. Up to day, few quantitative studies have been carried out employing high-energy synchrotron radiation X-ray fluorescence analysis (SR-XRF) and X-ray absorption fine structure (XAFS) [10, 11]. Unfortunately, the limited access to these analytical techniques limits considerably its daily use for routine analysis. Therefore, this works presents the development of an analytical methodology based on mass spectrometry imaging (MSI) using laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) for mapping and quantifying uranium in histological tissue sections of mouse kidney. To the author’s knowledge a single work has been published recently for the semi-quantitative analysis of uranium in mice kidneys due to the lack of an appropriate internal standard [12]. In this study, the uranium exposure of mice was performed using a direct nasal instillation of few µL of uranyl nitrate solutions at different uranium concentrations. Total uranium analysis was done by conventional liquid ICP-MS after acid digestion using one kidney from each pair. In parallel, the other kidney was employed for LA-ICP-MS analysis. In order to perform quantitative LA-ICPMS bio-imaging analysis, synthetic matrix-matched laboratory standards and a normalisation strategy based on internal standard spiked gelatine were developed. Preliminary quantitative images of cryo-sections revealed heterogeneous distribution of uranium within the renal tissue, being the cortical concentration 60-fold higher than the medullary concentration

LOCALISATION ET QUANTIFICATION DE L'URANIUM DANS LES STRUCTURES RENALES DE RONGEURS EXPOSES AU NITRATE D'URANYLE PAR LA TECHNIQUE LA-ICP-MS

International audienceLa surexposition à l'uranium entraîne un dysfonctionnement rénal suite à son effet toxique par des actions chimique et radiologique. De plus, la distribution tissulaire de l’uranium est hétérogène, s'accumulant et induisant principalement des lésions pathologiques dans les segments S2 et S3 du tubule proximal situés dans la zone corticale (100 fois la concentration rénale moyenne). Ce travail présente le développement d'une méthodologie analytique basée sur la spectrométrie de masse à couplage inductif et la technique d’ablation laser (LA-ICP-MS) pour la cartographie et la quantification de l'uranium dans des coupes de tissus histologiques de reins de souris. À ce jour, une seule équipe a récemment publié une analyse de l’uranium dans les reins de souris qui reste semi-quantitative à cause de l’absence d’un étalon interne approprié. Afin de réaliser des analyses quantitatives de bio-imagerie LA-ICPMS, nous avons développé des standards de laboratoire adaptés à la matrice rénale et une stratégie de normalisation basée sur la gélatine dopée avec un standard interne. Les premiers résultats quantitatifs obtenus à partir d’images préliminaires de cryo-sections de rein ont mis en évidence une distribution hétérogène de l'uranium dans le tissu rénal, avec une concentration corticale 60 fois supérieure à la concentration médullaire

A novel calibration strategy based on internal standard spiked gelatine for quantitative bio-imaging by LA-ICP-MS:Application to renal localization and quantification of uranium

International audienceMass spectrometry imaging (MSI) using laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has been employed for the elemental bio-distribution and quantification of uranium (U) in histological tissue sections of rodent kidneys. Samples were immediately immersed into 4% paraformaldehyde (PFA) solution during 24h, Tissue Tek OCT embedded and stored at -80°C until cutting in cryostat and mounting in gel covered glass slides. Sample preparation and laser conditions were carefully optimized to assure the complete ablation of the sample matrix. This works presents the development of internal standard correction methodology based on doped gelatine. In order to correct matrix effects, lack of tissue homogeneity and instrumental drift, a thulium (Tm, internal standard) spiked 10% (m/v) gel was prepared and deposited on the top of glass slides. For quantification purposes, matrix-matched laboratory standards were prepared from a pool of rat kidneys by dosing each level with different concentrations of uranium. The quantitative images of cryo-sections revealed heterogeneous distribution of uranium within the renal tissue, being the cortical concentration up to 120-fold higher than the medullary concentration

LOCALISATION ET QUANTIFICATION DE L'URANIUM DANS LES STRUCTURES RENALES DE RONGEURS EXPOSES AU NITRATE D'URANYLE PAR LA TECHNIQUE LA-ICP-MS

International audienceLa surexposition à l'uranium entraîne un dysfonctionnement rénal suite à son effet toxique par des actions chimique et radiologique. De plus, la distribution tissulaire de l’uranium est hétérogène, s'accumulant et induisant principalement des lésions pathologiques dans les segments S2 et S3 du tubule proximal situés dans la zone corticale (100 fois la concentration rénale moyenne). Ce travail présente le développement d'une méthodologie analytique basée sur la spectrométrie de masse à couplage inductif et la technique d’ablation laser (LA-ICP-MS) pour la cartographie et la quantification de l'uranium dans des coupes de tissus histologiques de reins de souris. À ce jour, une seule équipe a récemment publié une analyse de l’uranium dans les reins de souris qui reste semi-quantitative à cause de l’absence d’un étalon interne approprié. Afin de réaliser des analyses quantitatives de bio-imagerie LA-ICPMS, nous avons développé des standards de laboratoire adaptés à la matrice rénale et une stratégie de normalisation basée sur la gélatine dopée avec un standard interne. Les premiers résultats quantitatifs obtenus à partir d’images préliminaires de cryo-sections de rein ont mis en évidence une distribution hétérogène de l'uranium dans le tissu rénal, avec une concentration corticale 60 fois supérieure à la concentration médullaire

Thulium spiked gel for internal standardisation in LA-ICP-MS bio-imaging: quantitative elemental distribution of uranium in kidney tissue

International audienceThe quantitative analysis of trace metals in different organs or cellular structures is a topic of emerging interest for the assessment of toxicological risk. The kidney is recognized as a major site for uranium accumulation able to induce renal toxicity. Due to its heterogeneous distribution within the tissue, the quantification appears to be of significant interest. Mass spectrometry imaging (MSI) using laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS) has been employed for mapping and quantifying uranium in histological tissue sections of mouse kidney. Samples were immediately immersed into a PFA solution during 24h, OCT embedded and stored at -80°C until cutting in cryostat and mounting in gel covered glass slides. Sample preparation (tissue section thickness) and laser conditions were carefully optimized to assure the complete ablation of the sample matrix. This works presents the development of internal standard correction methodology based on doped gelatine. In order to correct matrix effects, lack of tissue homogeneity and instrumental drift, a thulium (Tm, internal standard) spiked 10% (m/v) gel was prepared and deposited on the top of glass microscope slides. The reproducibility and homogeneity of the deposited gel was assessed by ablating several areas (0.5 x 0.5 mm, n=20) in which Tm was recorded. For quantification purposes, matrix-matched laboratory standards were prepared from a pool of mouse kidneys by dosing each level with different concentration of uranium. In the same way, several quality control (QC) samples were synthetized to prove the accuracy and precision of the analytical method. The unspiked tissue was used for blank control. The concentrations were verified by conventional liquid ICP-MS after acid digestion. These standards were cut and mounted onto glass slides in the same way as the samples.Thulium spiked gel was demonstrated to be adequate for compensation of instrumental drifts during measuring time and matrix effects, which improves the quantification of elemental distributions and allows a trustworthy comparison of experiments performed at different times. The proposed analytical bio-imaging approach was successfully applied for quantification of uranium of kidney samples. The comparison of the calculated average concentration obtained by LA-ICP-MS and the results obtained after liquid ICP-MS analysis were in good accordance

DEVELOPPEMENT D’UNE TECHNIQUE DE CALIBRATION POUR L’IMAGERIE QUANTITATIVE D'ECHANTILLONS BIOLOGIQUES PAR ABLATION AU LASER COMPATIBLE AVEC L’ANALYSE SIMS : APPLICATION A LA LOCALISATION DE L'URANIUM DANS LE TISSU RENAL

International audienceLa spectrométrie de masse à plasma à couplage inductif par ablation laser a évolué pour devenir une technique analytique performante pour l'analyse quantitative in situ d'échantillons solides. Cependant, les progrès dans les applications des sciences biologiques sont toujours dépendants de la disponibilité d’étalons externes et internes appropriés. En outre, il serait intéressant que les échantillons et les étalons soient compatibles entre différentes techniques d'imagerie. Le but de ce travail est donc de trouver une méthodologie de préparation d’échantillons / étalons compatible avec les analyses par LA-ICP-MS et SIMS afin de rendre ces deux techniques complémentaires pour l’étude de la distribution de l’uranium dans les reins. D'une part, des étalons solides internes ont été préparés à partir d'homogénat de rein enrichi d'uranium. D’autre part, une nouvelle approche basée sur une résine dopée au nitrate d’uranyle (en poudre) a également été testée. De plus un standard interne (thulium) est incorporé directement dans la résine dopée. L'homogénéité de l'étalon interne et de l’uranium a été évaluée. Les avantages de cette méthodologie sont 1) pas besoin des homogénats, 2) une préparation d’échantillon compatible avec plusieurs techniques d’imagerie, 3) la facilité de préparation et 4) la conservation des étalons à température ambiante