The chronic metabolic dysfunction-associated steatotic liver disease (MASLD) is characterized by hepatic accumulation of lipids, which results in gradually progressing loss of liver function with concomitant oxidative stress and hepatic inflammation. While effects of the macronutrient intake have been widely studied, less is known about the contribution of micronutrients to MASLD development. The trace element selenium is of particular interest as it is highly abundant in the liver, is incorporated into antioxidant selenoproteins, and supports the immune response, thus targeting key aspects of MASLD pathogenesis. In the European population, the selenium intake is usually suboptimal meaning that not all selenoproteins are fully expressed. Accordingly, we asked the question if a selenium-deficient diet would have an impact on the development and severity of high-fat/high-fructose (Hf/Hf)-induced MASLD in mice. Male C57BL/6Jrj mice were fed a Hf/Hf diet for 8, 12, 16, and 24 weeks, which either contained adequate or deficient amounts of selenium. Even though the selenium deficiency substantially reduced expression of selenoproteins, we observed overall only moderate changes on liver parameters. During the development of MASLD, selenium deficiency increased hepatic inflammation, which however, was not detectable after 24 weeks anymore. Intestinal barrier integrity was reduced in the selenium-deficient group, which might have contributed to the observed pro-inflammatory effect in the liver. Overall, suboptimal selenium levels appear to accelerate the progression of MASLD in its early stages, particularly because of elevated inflammation. Thus, adequate selenium intake might be specifically important in early-stage MASLD patients with suboptimal selenium status to slow disease progression.

Selenium-binding protein 1 (SELENBP1) catalyzes the oxidative conversion of methanethiol to hydrogen sulfide, hydrogen peroxide and formaldehyde. Methanethiol oxidase (MTO) activity of recombinant human SELENBP1 was shown to require copper (Cu) ions, whereas selenium (Se) was largely dispensable. However, the impact of Cu and Se on MTO activity in mammals has not been explored so far. To investigate whether SELENBP1 levels and MTO activity in mice are affected by dietary availability of trace elements, liver samples of C57BL/6JRj mice held on diets deficient in Cu and/or Se for eight weeks were analyzed, as compared to mice fed the control diet with adequate trace element supply. Dietary Cu deficiency always resulted in diminished hepatic MTO activity, as detected by methanethiol-derived production of hydrogen sulfide. The effect of Cu deficiency was more pronounced in male than in female mice. Se deficiency only slightly, and solely in males, lowered MTO activity. Another sex-specific effect was observed with hepatic SELENBP1 levels: whereas diets deficient in Cu and/or Se increased SELENBP1 levels in males, they were decreased in female mice under Cu-deficient conditions. In addition to dietary Cu deficiency, MTO activity and SELENBP1 levels were explored in a rat model of Wilson’s disease, which is characterized by hepatic Cu accumulation. Hepatic SELENBP1 levels were decreased in male but not in female rats under conditions of excess intrahepatic Cu. However, hepatic MTO activity was not altered in both sexes, if the liver was not severely damaged. Notwithstanding the relatively low number of 5-6 animals included in each group, our data allow for the conclusion that hepatic MTO activity depends on adequate dietary Cu supply, whereas excess Cu does not further enhance the MTO activity of SELENBP1.

Older age combined with chronic disease increases the risk of malnutrition and frailty, impacting disease recovery and overall clinical outcomes. Serum concentrations of several trace elements and their respective biomarkers have not yet been investigated with regard to frailty in older adults with disease, although these patients most likely have an altered trace element profile owing to both inflammatory conditions and inadequate dietary intake. This cross-sectional study investigated trace element profiles in relation to age, disease, and frailty status in geriatric patients (n = 198) as well as in old (n = 80) and young (n = 60) healthy controls. Serum concentrations of iron, zinc, selenium, iodine, copper, and manganese were quantified via ICP-MS/MS, alongside inflammatory markers and functional biomarkers. Analysis revealed distinct trace element profiles in patients compared to healthy controls, with lower manganese, iron, zinc, and selenium, but higher copper and iodine (p ≤ 0.001). Trace element concentrations were similar in young and older healthy controls. Principal component (PC) analysis identified two profiles. PC1 was negatively associated with age, number of drugs, sex, and inflammation. Only PC2 was associated with anorexia (β = 0.053 ± 0.020; 95

As animal-derived foods are the main source of selenium, zinc, and copper, children and adolescents on vegetarian or vegan dietary patterns are at risk of an inadequate supply. Among 342 children and adolescents (6–18 years) with different dietary patterns (86 vegans, 120 vegetarians, 118 omnivores) from the cross-sectional VeChi Youth study serum concentrations of selenium, zinc, and copper and functional biomarkers such as glutathione peroxidase-3 activity (GPX3) and selenoprotein P (SELENOP) for selenium and ceruloplasmin oxidase activity (CPO) for copper were measured. Dietary intake of these trace elements was estimated using a 3-day weighed food record. Group differences were assessed by analysis of covariance, adjusted for age, sex, puberty status, and further covariates. Trace element intake differed across dietary patterns with lower selenium intake in vegans compared to omnivores (p < 0.0001), and lower zinc but higher copper intake in vegans (p = 0.0487 and p < 0.0001) and vegetarians (p = 0.0354 and p < 0.0001) than in omnivores. Lower serum selenium as well as SELENOP concentrations were observed in vegans (p < 0.0001 and p < 0.0001) and vegetarians (p < 0.0001 and p < 0.0001) in comparison to omnivores, but no difference in GPX3 activity across the dietary groups was observed. Similarly, serum zinc concentrations were lower in vegans (p = 0.0122) and vegetarians (p = 0.0016) compared to omnivores while serum copper concentrations and CPO did not differ between the dietary patterns. Vegetarian and vegan dietary patterns are associated with lower intake and serum biomarkers of selenium and zinc and should be monitored in children and adolescents on vegan or vegetarian dietary patterns. Trial registration number and date of registration DRKS00012835, 11.07.2018.

Trace elements such as copper, zinc, and selenium are essential micronutrients that play crucial roles in various physiological processes, mainly through their involvement in enzymes and regulatory proteins. A deficiency of any of these elements can impair physiological functions and lead to a range of symptoms. While copper deficiency is rare, e.g., vegans are particularly susceptible to inadequate intake of zinc and selenium. To investigate the effects of multiple simultaneous deficiencies, a feeding study was conducted in adult male and female C57BL/6Jrj mice receiving diets low in copper, zinc, and selenium. This approach enabled us to explore potential interactions between trace elements and to identify organ-specific effects based on their distribution profiles. We observed a substantial depletion of copper and selenium concentrations in the circulation and in almost all organs although to a varying extent. In contrast, zinc levels were well maintained and only declined in serum and bone. In line with the well-known antagonistic relationship between copper and zinc, our findings revealed that zinc deficiency mitigated symptoms of copper deficiency, which was most pronounced in female mice. Moreover, copper deficiency led to increased selenium concentrations in various organs, which, however, was not accompanied by generally higher selenoprotein expression. Therefore, it is essential to consider potential effects of single trace element deficiencies on other trace elements taking also combined effects into account.

A vegan diet is associated with health benefits but may also lead to inadequate intake of essential nutrients. Due to the lower selenium content in plant-based compared to animal-based foods, many vegans do not reach the recommended selenium intake in Europe. The only plant-based food with high selenium content is the Brazil nut, even though there is also a high variability. Therefore, we investigated the effectiveness of Brazil nut butter compared to a dietary supplement as selenium source to improve the selenium status of vegans and omnivores. 44 vegans and 42 omnivores were randomly assigned to one of three intervention groups, either receiving placebo or consuming additional 55 µg of selenium daily as Brazil nut butter or supplement for two weeks. Serum selenium concentrations, glutathione peroxidase 3 (GPX3), and selenoprotein P (SELENOP) were measured at baseline and after intervention. Additionally, dietary selenium intake was estimated using a five-day dietary protocol. The estimated selenium intake was significantly lower in vegans compared to omnivores and correlated with all three selenium biomarkers. Independent of the dietary pattern (vegan or omnivore), Brazil nut butter as well as supplement significantly increased serum selenium and SELENOP concentrations, while there were no changes in the placebo groups. Both interventions were equally effective in increasing selenium levels, but the upregulation of SELENOP was more pronounced in vegans than in omnivores. Brazil nuts are a plant-based source of selenium suitable for vegans and omnivores to improve their selenium status when consumed once in a while. Clinical trials registration number: NCT05814874, April 18 2023.

The ageing process is associated with alterations of systemic trace element (TE) homeostasis increasing the risk, e.g. neurodegenerative diseases. Here, the impact of long-term modulation of dietary intake of copper, iron, selenium, and zinc was investigated in murine cerebellum. Four- and 40-wk-old mice of both sexes were supplied with different amounts of those TEs for 26 wk. In an adequate supply group, TE concentrations were in accordance with recommendations for laboratory mice while suboptimally supplied animals received only limited amounts of copper, iron, selenium, and zinc. An additional age-adjusted group was fed selenium and zinc in amounts exceeding recommendations. Cerebellar TE concentrations were measured by inductively coupled plasma-tandem mass spectrometry. Furthermore, the expression of genes involved in TE transport, DNA damage response, and DNA repair as well as selected markers of genomic stability [8-oxoguanine, incision efficiency toward 8-oxoguanine, 5-hydroxyuracil, and apurinic/apyrimidinic sites and global DNA (hydroxy)methylation] were analysed. Ageing resulted in a mild increase of iron and copper concentrations in the cerebellum, which was most pronounced in the suboptimally supplied groups. Thus, TE changes in the cerebellum were predominantly driven by age and less by nutritional intervention. Interestingly, deviation from adequate TE supply resulted in higher manganese concentrations of female mice even though the manganese supply itself was not modulated. Parameters of genomic stability were neither affected by age, sex, nor diet. Overall, this study revealed that suboptimal dietary TE supply does not substantially affect TE homeostasis in the murine cerebellum. Graphical Abstract Ageing is associated with an altered TE status. In the cerebellum of mice, TE homeostasis was most strongly influenced by age whereas both suboptimal and age-adjusted dietary TE supply did not have substantial impact. Parameters of genomic stability were neither affected by age, sex nor diet. This emphasizes the limitations in counteracting age- and sex-specific changes of TE concentrations in the murine brain via dietary supply.

Selenium homeostasis depends on hepatic biosynthesis of selenoprotein P (SELENOP) and SELENOP-mediated transport from the liver to e.g. the brain. In addition, the liver maintains copper homeostasis. Selenium and copper metabolism are inversely regulated, as increasing copper and decreasing selenium levels are observed in blood during aging and inflammation. Here we show that copper treatment increased intracellular selenium and SELENOP in hepatocytes and decreased extracellular SELENOP levels. Hepatic accumulation of copper is a characteristic of Wilson's disease. Accordingly, SELENOP levels were low in serum of Wilson's disease patients and Wilson's rats. Mechanistically, drugs targeting protein transport in the Golgi complex mimicked some of the effects observed, indicating a disrupting effect of excessive copper on intracellular SELENOP transport resulting in its accumulation in the late Golgi. Our data suggest that hepatic copper levels determine SELENOP release from the liver and may affect selenium transport to peripheral organs such as the brain.

Mitochondria play multifaceted roles in cellular function, and impairments across domains of mitochondrial biology are known to promote cellular integrated stress response (ISR) pathways as well as systemic metabolic adaptations. However, the temporal dynamics of specific mitochondrial ISR related to physiological variations in tissue-specific energy demands remains unknown. Here, we conducted a comprehensive 24-hour muscle and plasma profiling of male and female mice with ectopic mitochondrial respiratory uncoupling in skeletal muscle (mUcp1-transgenic, TG). TG mice are characterized by increased muscle ISR, elevated oxidative stress defense, and increased secretion of FGF21 and GDF15 as ISR-induced myokines. We observed a temporal signature of both cell-autonomous and systemic ISR in the context of endocrine myokine signaling and cellular redox balance, but not of ferroptotic signature which was also increased in TG muscle. We show a progressive increase of muscle ISR on transcriptional level during the active phase (night time), with a subsequent peak in circulating FGF21 and GDF15 in the early resting phase. Moreover, we found highest levels of muscle oxidative defense (GPX and NQO1 activity) between the late active to early resting phase, which could aim to counteract excessive iron-dependent lipid peroxidation and ferroptosis in muscle of TG mice. These findings highlight the temporal dynamics of cell-autonomous and endocrine ISR signaling under skeletal muscle mitochondrial uncoupling, emphasizing the importance of considering such dissociation in translational strategies and sample collection for diagnostic biomarker analysis.

BACKGROUND:Trace elements exhibit essential functions in many physiological processes. Thus, for research focusing on trace element homeostasis and metabolism analytical methods allowing for multi-element analyses are fundamental. Small sample amounts may be a big challenge in trace element analyses especially if also other end points want to be addressed in the same sample. Therefore, the aim of the present study was to examine trace elements (iron, copper, zinc, and selenium) in murine liver tissue prepared by a RIPA buffer-based lyses method.METHODS AND RESULTS:After centrifugation, lysates and pellets were obtained and trace elements were analyzed with TXRF in liver lysates. The results were compared to that obtained by a standard microwave-assisted acidic digestion with subsequent ICP-MS/MS analysis of the same liver tissue, liver lysates, and remaining pellets. In addition, trace element concentrations, determined in murine serum with both methods, were compared. For serum samples, both TXRF and ICP-MS/MS provide similar and highly correlating results. Furthermore, in liver lysate samples prepared with RIPA buffer, comparable trace element concentrations were measured by TXRF as with the standard digestion technique and ICP-MS/MS. Only marginal amounts of trace elements were detected in the pellets.CONCLUSION:Taken together, the results obtained by the present study indicate that the RIPA buffer-based method is suitable for sample preparation for trace element analyses via TXRF, at least for the here investigated murine liver samples.

Alterations in reduced and oxidized glutathione (GSH/GSSG) levels represent an important marker for oxidative stress and potential disease progression in toxicological research. Since GSH can be oxidized rapidly, using a stable and reliable method for sample preparation and GSH/GSSG quantification is essential to obtain reproducible data. Here we describe an optimised sample processing combined with a liquid chromatography-tandem mass spectrometry (LC-MS/MS) method, validated for different biological matrices (lysates from HepG2 cells, C. elegans, and mouse liver tissue). To avoid autoxidation of GSH, samples were treated with the thiol-masking agent N-ethylmaleimide (NEM) and sulfosalicylic acid (SSA) in a single step. With an analysis time of 5 min, the developed LC-MS/MS method offers simultaneous determination of GSH and GSSG at high sample throughput with high sensitivity. This is especially interesting with respect of screening for oxidative and protective properties of substances in in vitro and in vivo models, e.g. C. elegans. In addition to method validation parameters (linearity, limit of detection (LOD), limit of quantification (LOQ), recovery, interday, intraday), we verified the method by using menadione and L-buthionine-(S,R)-sulfoximine (BSO) as well established modulators of cellular GSH and GSSG concentrations. Thereby menadione proved to be a reliable positive control also in C. elegans.

Despite advances in cancer research, cancer is still one of the leading causes of death worldwide. An early diagnosis substantially increases the survival rate and treatment success. Thus, it is important to establish biomarkers which could reliably identify cancer patients. As cancer is associated with changes in the systemic trace element status and distribution, serum concentrations of selenium, iron, copper, and zinc could contribute to an early diagnosis. To test this hypothesis, case control studies measuring trace elements in cancer patients vs. matched controls were selected and discussed focusing on lung, prostate, breast, and colorectal cancer. Overall, cancer patients had elevated serum copper and diminished zinc levels, while selenium and iron did not show consistent changes for all four cancer types. Within the tumor tissue, mainly copper and selenium are accumulating. Whether these concentrations also predict the survival probability of cancer patients needs to be further investigated.

Selenium and iodine are the two central trace elements for the homeostasis of thyroid hormones but additional trace elements such as iron, zinc, and copper are also involved. To compare the primary effects of inadequate intake of selenium and iodine on the thyroid gland, as well as the target organs of thyroid hormones such as liver and kidney, mice were subjected to an eight-week dietary intervention with low versus adequate selenium and iodine supply. Analysis of trace element levels in serum, liver, and kidney demonstrated a successful intervention. Markers of the selenium status were unaffected by the iodine supply. The thyroid gland was able to maintain serum thyroxine levels even under selenium-deficient conditions, despite reduced selenoprotein expression in liver and kidney, including deiodinase type 1. Thyroid hormone target genes responded to the altered selenium and iodine supply, whereas the iron, zinc, and copper homeostasis remained unaffected. There was a notable interaction between thyroid hormones and copper, which requires further clarification. Overall, the effects of an altered selenium and iodine supply were pronounced in thyroid hormone target tissues, but not in the thyroid gland.

Trace elements (TEs) are essential for diverse processes maintaining body function and health status. The complex regulation of the TE homeostasis depends among others on age, sex, and nutritional status. If the TE homeostasis is disturbed, negative health consequences can result, e.g., caused by impaired redox homeostasis and genome stability maintenance. Based on age-related shifts in TEs which have been described in mice well-supplied with TEs, we aimed to understand effects of a long-term feeding with adequate or suboptimal amounts of four TEs in parallel. As an additional intervention, we studied mice which received an age-adapted diet with higher concentrations of selenium and zinc to counteract the age-related decline of both TEs. We conducted comprehensive analysis of diverse endpoints indicative for the TE and redox status, complemented by analysis of DNA (hydroxy)methylation and markers denoting genomic stability maintenance. TE concentrations showed age-specific alterations which were relatively stable and independent of their nutritional supply. In addition, hepatic DNA hydroxymethylation was significantly increased in the elderly mice and markers indicative for the redox status were modulated. The reduced nutritional supply with TEs inconsistently affected their status, with most severe effects regarding Fe deficiency. This may have contributed to the sex-specific differences observed in the alterations related to the redox status and DNA repair activity. Overall, our results highlight the complexity of factors impacting on the TE status and its physiological consequences. Alterations in TE supply, age, and sex proved to be important determinants that need to be taken into account when considering TE interventions for improving general health and supporting convalescence in the clinics.