Development of single-cell ICP-TOFMS to measure nanoplastics association with human cells† † Electronic supplementary information (ESI) available. See DOI: https://doi.org/10.1039/d3en00681f
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ABSTRACT:
SUBMITTER: Hendriks L
PROVIDER: S-EPMC10702630 | biostudies-literature | 2023 Nov
REPOSITORIES: biostudies-literature
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Project description:A synthetic route to producing gold-doped environmentally relevant nanoplastics and a method for the rapid and high-throughput qualitative investigation of their cellular interactions have been developed. Polyethylene (PE) and polyvinyl chloride (PVC) nanoparticles, doped with ultrasmall gold nanoparticles, were synthesized via an oil-in-water emulsion technique as models for floating and sedimenting nanoplastics, respectively. Gold nanoparticles were chosen as a dopant as they are considered to be chemically stable, relatively easy to obtain, interference-free for elemental analysis, and suitable for bio-applications. The suitability of the doped particles for quick detection via inductively coupled plasma mass spectrometry (ICP-MS), operating in single-cell mode (scICP-MS), was demonstrated. Specifically, the method was applied to the analysis of nanoplastics in sizes ranging from 50 to 350 nm, taking advantage of the low limit of detection of single-cell ICP-MS for gold nanoparticles. As an initial proof of concept, gold-doped PVC and PE nanoplastics were employed to quantify the interaction and uptake of nanoplastics by the RAW 264.7 mouse macrophage cell line, using scICP-MS and electron microscopy. Macrophages were chosen because their natural biological functions would make them likely to internalize nanoplastics and, thus, would produce samples to verify the test methodology. Finally, the method was applied to assess the uptake by CaCo-2 human intestinal cells, this being a more relevant model for humanexposure to those nanoplastics that are potentially available in the food chain. For both case studies, two concentrations of nanoplastics were employed to simulate both standard environmental conditions and exceptional circumstances, such as pollution hotspot areas.
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Project description:Recently, spICP-MS analyses of microplastics have demonstrated that the detection capabilities of ICP-MS are sufficient to determine the size and composition of such materials. However, solution nebulization or microdroplet generation limits the sizes of droplets, microparticles, or cells that can be efficiently introduced into commonly used horizontal ICP-MS configurations. Therefore, we introduced the development of a downward-pointing ICP coupled to a time-of-flight mass spectrometer (ICP-TOFMS), which enables quantitative transport of large microdroplets (diameters up to 90 µm) into the ICP. Here, we report the capabilities of downward ICP-TOFMS for the quantitative analysis of single cells and microplastic particles. For calibration of element mass amount per particle or cell, microdroplets (70 µm diameter) composed of multielement solutions were measured by ICP-TOFMS. Microplastic beads (polystyrene) and spleenocyte cells were then also embedded in microdroplets and measured by ICP-TOFMS with ion optics optimized to determine the signals from 12C+ and other isotopes of interest, including 140Ce, 153Eu, 165Ho, and 175Lu from the REE beads and 31P for the cells. The results achieved using the prototype instrument of a vertical downward-pointing ICP-TOFMS demonstrate that such a plasma configuration is well suited to analyze microplastics and single cells. For single microbead and cell analyses, the critical mass for carbon was 4.8 pg, and the mean determined carbon mass amounts were 14 and 23 pg, respectively. For the microbead analysis, the detected carbon mass corresponds to a particle diameter of 2.93 ± 0.24 µm, which is consistent with the scanning (transmission) electron microscopy-determined diameter of 2.97 ± 0.04 µm. Furthermore, in the analysis of spleenocyte cells, carbon and phosphorus masses were determined to be correlated.
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Project description:A high-throughput laser ablation-inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS) workflow was implemented for quantitative single-cell analysis following cytospin preparation of cells. For the first time, in vitro studies on cisplatin exposure addressed human monocytes and monocyte-derived macrophages (undifferentiated THP-1 monocytic cells, differentiated M0 macrophages, as well as further polarized M1 and M2 phenotypes) at the single-cell level. The models are of particular interest as macrophages comprise the biggest part of immune cells present in the tumor microenvironment and play an important role in modulating tumor growth and progression. The introduced bioimaging workflow proved to be universally applicable to adherent and suspension cell cultures and fit-for-purpose for the quantitative analysis of several hundreds of cells within minutes. Both, cross-validation of the method with single-cell analysis in suspension for THP-1 cells and with LA-ICP-TOFMS analysis of adherent M0 cells grown on chambered glass coverslips, revealed agreeing platinum concentrations at the single-cell level. A high incorporation of cisplatin was observed in M2 macrophages compared to the M0 and M1 macrophage subtypes and the monocyte model, THP-1. The combination with bright-field images and monitoring of highly abundant endogenous elements such as phosphorus and sodium at a high spatial resolution allowed assessing cell size and important morphological cell parameters and thus straightforward control over several cell conditions. This way, apoptotic cells and cell debris as well as doublets or cell clusters could be easily excluded prior to data evaluation without additional staining.
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Project description:More than a decade ago, studies on cellular cisplatin accumulation via active membrane transport established the role of the high affinity copper uptake protein 1 (CTR1) as a main uptake route besides passive diffusion. In this work, CTR1 expression, cisplatin accumulation and intracellular copper concentration was assessed for single cells revisiting the case of CTR1 in the context of acquired cisplatin resistance. The single-cell workflow designed for in vitro experiments enabled quantitative imaging at resolutions down to 1 µm by laser ablation-inductively coupled plasma-time-of-flight mass spectrometry (LA-ICP-TOFMS). Cisplatin-sensitive ovarian carcinoma cells A2780 as compared to the cisplatin-resistant subline A2780cis were investigated. Intracellular cisplatin and copper levels were absolutely quantified for thousands of individual cells, while for CTR1, relative differences of total CTR1 versus plasma membrane-bound CTR1 were determined. A markedly decreased intracellular cisplatin concentration accompanied by reduced copper concentrations was observed for single A2780cis cells, along with a distinctly reduced (total) CTR1 level as compared to the parental cell model. Interestingly, a significantly different proportion of plasma membrane-bound versus total CTR1 in untreated A2780 as compared to A2780cis cells was observed. This proportion changed in both models upon cisplatin exposure. Statistical analysis revealed a significant correlation between total and plasma membrane-bound CTR1 expression and cisplatin accumulation at the single-cell level in both A2780 and A2780cis cells. Thus, our study recapitulates the crosstalk of copper homeostasis and cisplatin uptake, and also indicates a complex interplay between subcellular CTR1 localization and cellular cisplatin accumulation as a driver for acquired resistance development.
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Project description:Nanogeochemistry is an emerging focus area recognizing the role of nanoparticles in Earth systems. Engineered nanotechnology has cultivated advanced analytical techniques that are also applicable to nanogeochemistry. Single particle inductively coupled plasma ICP-time-of-flight-mass spectrometry (ICP-TOF-MS) promises a significant step forward, as time-of-flight mass analyzers enable simultaneous quantification of the entire atomic mass spectrum (∼7-250 m/z +). To demonstrate the utility of this approach, samples were collected and analyzed from a large, boreal river, and its surrounding tributaries. These samples provided us with a diversity of particle compositions and morphologies, while their interconnected nature allowed for an examination of the various nanogeochemical processes present in this system. To further expand on this effort, we combined this high-throughput technique with AF4-ICPMS, focusing on major carriers of trace elements. Using spICP-TOF-MS, Al, Si, and Fe were grouped into classes having all combinations of one or more of these elements. Particle-by-particle ICP-TOF-MS analysis found chemically heterogeneous populations, indicating the predominance of diverse mineralogy or heteroaggregates. The importance of suspended Fe and Mn for the speciation of Pb was observed by single particle ICP-TOF-MS and complemented by AF4-ICPMS analysis of dissolved organic matter and nanoparticulate Fe/Mn. Our study exploits the combination of spICP-TOF-MS and AF4-ICP-MS for studying isotopic and elemental ratios (mineralogy) of individual nanoparticles, which opens the door to further explore the mechanisms of colloid facilitated transport of trace elements.
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Project description:While spICP-MS has been used mainly to measure nanoparticles, it can also be used to measure microparticles. The transport efficiency of nanoparticles is typically independent of their size. However, the transport efficiency of microparticles can be particle size (mass) dependent as well as being dependent on the sample uptake rate and sample introduction system used. To measure both nanoparticles and microparticles a very large linear dynamic range (where signal intensity is linearly proportional to the measured analyte(s) mass within a very short measurement time (∼300 to 500 µs, the width of signals produced by an individual particle)) is needed. Deviations from linearity could occur due to incomplete particle vaporization or from signals that are beyond the instrument's ion detection system linear dynamic range. To characterize and determine the cause of nonlinearity we measured sets of nearly monodisperse engineered SiO2 particles with diameters from 500 to 5000 nm and Au particles with diameters from 60 to 1500 nm. We found that by reducing the sensitivity (up to a factor of 269×) the upper end of the linear dynamic range, in particle size that produced signal intensities that were linearly proportional to the particle (analyte) mass, could be greatly extended. Not surprisingly, reducing the sensitivity increased the minimum size detectable particle. The results are consistent with SiO2 particles as large as 5000 nm being completely vaporized in the ICP.
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Project description:This is the first report of the use of laser ablation-inductively coupled plasma time-of-flight mass spectrometry (LA-ICP-TOFMS) to analyze human malignant pleural mesothelioma (MPM) samples at the cellular level. MPM is an aggressive, incurable cancer associated with asbestos exposure, with a long latency and poor overall survival. Following careful optimization of the laser fluence, the simultaneous ablation of soft biological tissue and hard mineral fibers was possible, allowing the spatial detection of elements such as Si, Mg, Ca, and Fe, which are also present in the glass substrate. A low-dispersion LA setup was employed, which provided the high spatial resolution necessary to identify the asbestos fibers and fiber fragments in the tissue and to characterize the metallome at the cellular level (a pixel size of 2 μm), with a high speed (at 250 Hz). The multielement LA-ICP-TOFMS imaging approach enabled (i) the detection of asbestos fibers/mineral impurities within the MPM tissue samples of patients, (ii) the visualization of the tissue structure with the endogenous elemental pattern at high spatial resolution, and (iii) obtaining insights into the metallome of MPM patients with different pathologies in a single analysis run. Asbestos and other mineral fibers were detected in the lung and pleura tissue of MPM patients, respectively, based on their multielement pattern (Si, Mg, Ca, Fe, and Sr). Interestingly, strontium was detected in asbestos fibers, suggesting a link between this potential toxic element and MPM pathogenesis. Furthermore, monitoring the metallome around the talc deposit regions (characterized by elevated levels of Al, Mg, and Si) revealed significant tissue damage and inflammation caused by talc pleurodesis. LA-ICP-TOFMS results correlated to Perls' Prussian blue and histological staining of the corresponding serial sections. Ultimately, the ultra-high-speed and high-spatial-resolution capabilities of this novel LA-ICP-TOFMS setup may become an important clinical tool for simultaneous asbestos detection, metallome monitoring, and biomarker identification.
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Project description:We developed an inductively coupled plasma mass spectrometry (ICP-MS) method using Universal Cell Technology (UCT) with a PerkinElmer NexION ICP-MS, to measure arsenic (As), chromium (Cr), and nickel (Ni) in human urine samples. The advancements of the UCT allowed us to expand the calibration range to make the method applicable for both low concentrations of biomonitoring applications and high concentrations that may be observed from acute exposures and emergency response. Our method analyzes As and Ni in kinetic energy discrimination (KED) mode with helium (He) gas, and Cr in dynamic reaction cell (DRC) mode with ammonia (NH3) gas. The combination of these elements is challenging because a carbon source, ethanol (EtOH), is required for normalization of As ionization in urine samples, which creates a spectral overlap (40Ar12C+) on 52Cr. This method additionally improved lab efficiency by combining elements from two of our previously published methods(Jarrett et al., 2007; Quarles et al., 2014) allowing us to measure Cr and Ni concentrations in urine samples collected as part of the National Health and Nutrition Examination Survey (NHANES) beginning with the 2017-2018 survey cycle. We present our rigorous validation of the method selectivity and accuracy using National Institute of Standards and Technology (NIST) Standard Reference Materials (SRM), precision using in-house prepared quality control materials, and a discussion of the use of a modified UCT, a BioUCell, to address an ion transmission phenomenon we observed on the NexION 300 platform when using higher elemental concentrations and high cell gas pressures. The rugged method detection limits, calculated from measurements in more than 60 runs, for As, Cr, and Ni are 0.23 μg L-1, 0.19 μg L-1, and 0.31 μg L-1, respectively.
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