The use of ICP-MS to measure metal ion content in biological tissues offers a highly sensitive means to study metal-dependent physiological processes. Here we describe the application of ICP-MS to measure membrane transport of Rb and K ions by the Na,K-ATPase in mouse skeletal muscles and human red blood cells. The ICP-MS method provides greater precision and statistical power than possible with conventional tracer flux methods. The method is widely applicable to studies of other metal ion transporters and metal-dependent processes in a range of cell types and conditions.

BACKGROUND. Clear cell renal cell carcinoma (ccRCC) is the most common histologically defined renal cancer. However, it is not a uniform disease and includes several genetic subtypes with different prognoses. ccRCC is also characterized by distinctive metabolic reprogramming. Tobacco smoking (TS) is an established risk factor for ccRCC, with unknown effects on tumor pathobiology. METHODS. We investigated the landscape of ccRCCs and paired normal kidney tissues using integrated transcriptomic, metabolomic, and metallomic approaches in a cohort of white males who were long-term current smokers (LTS) or were never smokers (NS). RESULTS. All 3 Omics domains consistently identified a distinct metabolic subtype of ccRCCs in LTS, characterized by activation of oxidative phosphorylation (OXPHOS) coupled with reprogramming of the malate-aspartate shuttle and metabolism of aspartate, glutamate, glutamine, and histidine. Cadmium, copper, and inorganic arsenic accumulated in LTS tumors, showing redistribution among intracellular pools, including relocation of copper into the cytochrome c oxidase complex. A gene expression signature based on the LTS metabolic subtype provided prognostic stratification of The Cancer Genome Atlas ccRCC tumors that was independent of genomic alterations. CONCLUSION. The work identified the TS-related metabolic subtype of ccRCC with vulnerabilities that can be exploited for precision medicine approaches targeting metabolic pathways. The results provided rationale for the development of metabolic biomarkers with diagnostic and prognostic applications using evaluation of OXPHOS status. The metallomic analysis revealed the role of disrupted metal homeostasis in ccRCC, highlighting the importance of studying effects of metals from e-cigarettes and environmental exposures. FUNDING. Department of Defense, Veteran Administration, NIH, ACS, and University of Cincinnati Cancer Institute.

Copper (Cu) is a cofactor of cytochrome c oxidase (CuCOX), indispensable for aerobic mitochondrial respiration. This study reveals that advanced clear cell renal cell carcinomas (ccRCC) accumulate Cu, allocating it to CuCOX. Using a range of orthogonal approaches, including metabolomics, lipidomics, isotope-labeled glucose and glutamine flux analysis, and transcriptomics across tumor samples, cell lines, xenografts, and patient-derived xenograft models, combined with genetic and pharmacologic interventions, we explored the role of Cu in ccRCC. Elevated Cu levels stimulate CuCOX biogenesis, providing bioenergetic and biosynthetic benefits that promote tumor growth. This effect is complemented by glucose-dependent glutathione production, which facilitates detoxification and mitigates Cu–H2O2 toxicity. Single-cell RNA sequencing and spatial transcriptomics reveal increased oxidative metabolism, altered glutathione and Cu metabolism, and diminished hypoxia-inducible transcription factor activity during ccRCC progression. Thus, Cu drives an integrated oncogenic remodeling of bioenergetics, biosynthesis, and redox homeostasis, fueling ccRCC growth, which can be targeted for new therapeutic approaches. Significance: The work establishes a requirement for glucose-dependent coordination between energy production and redox homeostasis, which is fundamental for the survival of cancer cells that accumulate Cu and contributes to tumor growth.

This work focuses on phytochelatins (PCs) in plants chronically exposed to low or moderate metal concentrations in soil, which commonly occurs in natural environments. The original approach was to evaluate PCs in nopal (Opuntia ficus) in relation to the plant and soil levels of Cd, Pb, Cu and Ag. These four metals were selected because of their known capability to induce PCs in plants and also because of their abundance in the region of Guanajuato (Mexico), where the nopal plants were collected. ICP-MS determination of metals in nopal roots, stems and in soil fractions provided information relevant to their bioavailability in soil as well as their uptake by nopal. In search of a possible relationship between the parameters measured, correlation analysis was carried out (Cd was not considered owing to its low levels found in the plant tissues). The results obtained indicate that sequential metal leaching from soil does not always provide direct information on metal bioavailability. The uptake of metals by nopal roots correlated with metals in the organic matter and sulfides fraction. Since low levels of humic substances were found in soil collected at cactus roots and because of the low solubility of Pb, Cu and Ag sulfides, the participation of rizosphere processes was suggested in facilitating metal uptake. Metal distribution in PC molecular mass fractions were studied by SEC-ICP-MS and the results compared with the total phytochelatin pool in plants as determined by reversed phase HPLC with spectrofluorimetric detection. The two procedures provide complementary analytical information when the global effect of all metals/metalloids present in soil on PC production is to be studied. Correlation analysis revealed the strong association of Pb with PCs both in roots and in stems, which demonstrates that Pb is important in nopal PCs and suggests that Pb bound to PCs might be transported from roots to stems. For Cu, the correlation between metal-free PCs and metal bound to high molecular mass fraction of plant PC-rich extract was observed only in roots, suggesting differences in the transport of Pb and Cu. In contrast to these metals, no correlation was found for silver, suggesting no association with PCs, even though its elution with a high molecular mass fraction was observed in PC-rich plant extracts. The results obtained from this study indicate the complex role environmental conditions play in the accumulation of heavy metals and in phytochelatin production in the Opuntia genus.

The promise of personalized medicine is a therapeutic advance where tumor signatures obtained from different omics platforms, such as genomics, transcriptomics, proteomics, and metabolomics, in addition to environmental factors including metals and metalloids, are used to guide the treatments. Clear cell renal carcinoma (ccRCC), the most common type of kidney cancer, can be sporadic (frequently) or genetic (rare), both characterized by loss of the von Hippel-Lindau (VHL) gene that controls hypoxia inducible factors. Recently, several genomic subtypes were identified with different prognoses. Transcriptomics, proteomics, metabolomics and metallomic data converge on altered metabolism as the principal feature of the disease. However, in view of multiple biochemical alterations and high level of tumor heterogeneity, identification of clearly defined subtypes is necessary for further improvement of treatments. In the future, single-cell combined multi-omics approaches will be the next generation of analyses gaining deeper insights into ccRCC progression and allowing for design of specific signatures, with better prognostic/predictive clinical applications.

Macrophages deploy numerous strategies to combat invasion by microbes. One tactic is to restrict acquisition of diverse nutrients, including trace metals, a process termed nutritional immunity. Intracellular pathogens adapt to a resource-poor environment by marshaling mechanisms to harvest nutrients. Carbon acquisition is crucial for pathogen survival; compounds that reduce availability are a potential strategy to control intracellular replication. Treatment of macrophages with the glucose analog 2-deoxy-D-glucose (2-DG) armed phagocytes to eliminate the intracellular fungal pathogen Histoplasma capsulatum in vitro and in vivo. Killing did not rely on altering access to carbon-containing molecules or changes in ATP, ER stress, or autophagy. Unexpectedly, 2-DG undermined import of exogenous zinc into macrophages, decreasing the quantity of cytosolic and phagosomal zinc. The fungus perished as a result of zinc starvation. This change in metal ingress was not ascribed to a defect in a single importer; rather, there was a collective impairment in transporter activity. This effect promoted the antifungal machinery of macrophages and expanded the complexity of 2-DG activities far beyond manipulating glycolysis. Mechanistic metabolic studies employing 2-DG will have to consider its effect on zinc transport. Our preclinical data support consideration of this agent as a possible adjunctive therapy for histoplasmosis.