Nothing Special   »   [go: up one dir, main page]

You seem to have javascript disabled. Please note that many of the page functionalities won't work as expected without javascript enabled.
 
 
Sign in to use this feature.

Years

Between: -

Subjects

remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline
remove_circle_outline

Journals

Article Types

Countries / Regions

remove_circle_outline
remove_circle_outline

Search Results (334)

Search Parameters:
Keywords = selenoproteins

Order results
Result details
Results per page
Select all
Export citation of selected articles as:
15 pages, 2976 KiB  
Article
Foliar Biofortification of Maize (Zea mays L.) with Selenium: Effects of Compound Type, Application Rate, and Growth Stage
by Tomáš Mrština, Lukáš Praus, Jiřina Száková, Lukáš Kaplan and Pavel Tlustoš
Agriculture 2024, 14(12), 2105; https://doi.org/10.3390/agriculture14122105 - 21 Nov 2024
Viewed by 202
Abstract
Nowadays, attention is focused on the lack of selenium in the average diet, which is a highly valued element in the body’s antioxidant system. The major metabolites of selenium are selenoproteins, which have an irreplaceable function in the body. This study focused on [...] Read more.
Nowadays, attention is focused on the lack of selenium in the average diet, which is a highly valued element in the body’s antioxidant system. The major metabolites of selenium are selenoproteins, which have an irreplaceable function in the body. This study focused on optimizing conditions for the biofortification of maize (Zea mays L.) with selenium (Se). Three separate pot experiments were conducted to identify the key factors influencing the efficacy of foliar selenium application. The experiments were designed to investigate the effects of different forms of selenium (selenite, selenate, and selenium nanoparticles) on maize development, the influence of the phenological stage of maize at the time of foliar Se application, and the optimal application rate of Se (100, 150, 200, or 250 µg). The results indicated that sodium selenate without a wetting agent was the most effective form for enhancing total Se content in maize, with the greatest accumulation being in leaves (3.01 mg/kg dry matter). Phenological stages (BBCH) 51 and 60 were identified as the most suitable phenological stages for Se application in terms of total Se content about 1 mg/kg in leaves and about 0.4 mg/kg in grain and the presence of organic Se compounds (mostly selenate ion and selenomethionine). We concluded from the study that a foliar application of 200 µg of sodium selenate per pot during these stages resulted in maximum Se uptake without adversely affecting plant yield. Further research is recommended to validate these findings under field conditions, paving the way for improved agricultural practices in selenium biofortification. Full article
(This article belongs to the Section Crop Production)
Show Figures

Figure 1

Figure 1
<p>Yield of maize plant parts after application of different Se forms. Different lowercase letters indicate a statistically significant differences among the treatments according to a one-way analysis of variance (<span class="html-italic">p</span> &lt; 0.05, <span class="html-italic">n</span> = 4).</p>
Full article ">Figure 2
<p>Total selenium content in parts of maize after application of different Se forms. Different lowercase letters indicate statistically significant differences among the treatments by a one-way analysis of variance (<span class="html-italic">p</span> &lt; 0.05, <span class="html-italic">n</span> = 4).</p>
Full article ">Figure 3
<p>Yields of maize parts after the application of Se in different phenological phases. Different lowercase letters indicate statistically significant differences among the treatments by a one-way analysis of variance (<span class="html-italic">p</span> &lt; 0.05, <span class="html-italic">n</span> = 4).</p>
Full article ">Figure 4
<p>Contents of total selenium in maize parts (<span class="html-italic">Zea mays</span> L.) after application of Se in different phenological phases. Different lowercase letters indicate statistically significant differences among treatments according to a one-way analysis of variance (<span class="html-italic">p</span> &lt; 0.05, <span class="html-italic">n</span> = 4).</p>
Full article ">Figure 5
<p>Yields of maize (<span class="html-italic">Zea mays</span> L.) parts after the application of different Se concentrations. Different lowercase letters indicate statistically significant differences among the treatments according to a one-way analysis of variance (<span class="html-italic">p</span> &lt; 0.05, <span class="html-italic">n</span> = 4).</p>
Full article ">Figure 6
<p>Content of total selenium in maize parts (<span class="html-italic">Zea mays</span> L.) after the application of different concentrations of Se. Different lowercase letters indicate a statistically significant differences among the treatments according to the one-way analysis of variance (<span class="html-italic">p</span> &lt; 0.05, <span class="html-italic">n</span> = 4).</p>
Full article ">Figure 7
<p>Regression curve slopes modelling the relationship between the applied Se dose and Se content in maize leaf.</p>
Full article ">Figure 8
<p>Regression curve slopes modelling the relationship between the applied Se dose and Se content in maize grain.</p>
Full article ">Figure 9
<p>Regression curve slopes modelling the relationship between the applied Se dose and Se content in maize stover.</p>
Full article ">
20 pages, 640 KiB  
Review
Selenium, Immunity, and Inflammatory Bowel Disease
by James A. Sousa, Derek M. McKay and Maitreyi Raman
Nutrients 2024, 16(21), 3620; https://doi.org/10.3390/nu16213620 - 25 Oct 2024
Viewed by 863
Abstract
Dietary intervention is a subject of growing interest in the management of inflammatory bowel disease (IBD), as new incident cases across the globe are rapidly rising, suggesting environmental factors as contributing elements. Dietary components and micronutrients have been associated with IBD pathogenesis or [...] Read more.
Dietary intervention is a subject of growing interest in the management of inflammatory bowel disease (IBD), as new incident cases across the globe are rapidly rising, suggesting environmental factors as contributing elements. Dietary components and micronutrients have been associated with IBD pathogenesis or reductions in disease severity. Selenium, a diet-derived essential micronutrient that is important for proper immune system function, has received limited attention in the context of IBD. Selenium deficiency is a common finding in patients with IBD, but few clinical trials have been published to address the consequences of this deficiency. Here, we review the physiological and immunological roles of selenium and its putative role in IBD, and draw attention to knowledge gaps and unresolved issues, with the goal of stimulating more research on selenium in IBD. Full article
Show Figures

Figure 1

Figure 1
<p>Selenium absorption and metabolism. The consumption of selenium-rich food leads to the absorption of organic and inorganic forms of Se in the gastrointestinal tract through transporters, depending on their molecular form. The absorbed Se is converted into H<sub>2</sub>Se and can be used by enterocytes for the production of selenoproteins or transported to the liver for the production of SelenoP. The transport of Se to other tissues is largely mediated through SelenoP that is then degraded and used in the synthesis of selenoproteins. The catabolism of SelenoP results in the release of selenocysteine that is subsequently converted by SCLY into H<sub>2</sub>Se, which, in turn, is converted by SEPHS2 into selenophosphate, which can then be incorporated into new selenoproteins. H<sub>2</sub>Se, hydrogen selenide; SEPHS2, Selenophosphate Synthetase 2; Sec-tRNA, Selenocysteine transfer RNA; GPx, Glutathione Peroxidase; LDL, low-density lipoprotein; VLDL, very low-density lipoprotein; SelenoP, Selenoprotein P; SCLY, Selenocysteine Lyase. Created in BioRender. Mckay, D. (2024) <a href="http://BioRender.com/o96h565" target="_blank">BioRender.com/o96h565</a> (accessed on 1 October 2024).</p>
Full article ">
17 pages, 3292 KiB  
Article
NRF2 and Thioredoxin Reductase 1 as Modulators of Interactions between Zinc and Selenium
by Alina Löser, Maria Schwarz and Anna Patricia Kipp
Antioxidants 2024, 13(10), 1211; https://doi.org/10.3390/antiox13101211 - 8 Oct 2024
Viewed by 843
Abstract
Background: Selenium and zinc are essential trace elements known to regulate cellular processes including redox homeostasis. During inflammation, circulating selenium and zinc concentrations are reduced in parallel, but underlying mechanisms are unknown. Accordingly, we modulated the zinc and selenium supply of HepG2 cells [...] Read more.
Background: Selenium and zinc are essential trace elements known to regulate cellular processes including redox homeostasis. During inflammation, circulating selenium and zinc concentrations are reduced in parallel, but underlying mechanisms are unknown. Accordingly, we modulated the zinc and selenium supply of HepG2 cells to study their relationship. Methods: HepG2 cells were supplied with selenite in combination with a short- or long-term zinc treatment to investigate intracellular concentrations of selenium and zinc together with biomarkers describing their status. In addition, the activation of the redox-sensitive transcription factor NRF2 was analyzed. Results: Zinc not only increased the nuclear translocation of NRF2 after 2 to 6 h but also enhanced the intracellular selenium content after 72 h, when the cells were exposed to both trace elements. In parallel, the activity and expression of the selenoprotein thioredoxin reductase 1 (TXNRD1) increased, while the gene expression of other selenoproteins remained unaffected or was even downregulated. The zinc effects on the selenium concentration and TXNRD activity were reduced in cells with stable NRF2 knockdown in comparison to control cells. Conclusions: This indicates a functional role of NRF2 in mediating the zinc/selenium crosstalk and provides an explanation for the observed unidirectional behavior of selenium and zinc. Full article
(This article belongs to the Special Issue Oxidative Stress and NRF2 in Health and Disease—2nd Edition)
Show Figures

Figure 1

Figure 1
<p>Establishment of culture conditions with 2.5% FBS. HepG2 cells were treated with or without 50 nM sodium selenite in combination with or without 100 μM zinc sulfate for 72 h in media containing 10% or 2.5% FCS. The relative cell count (<b>a</b>) was determined by trypan blue exclusion test. Intracellular trace element concentrations of Se (<b>b</b>), and Zn (<b>c</b>) were determined by total reflection X-ray fluorescence spectrometry (TXRF). The measurement was performed for 1000 s with 1 mg/L yttrium as an internal standard. Protein expression of MT (<b>d</b>) was determined by Western blot, normalized to Ponceau staining. Either the −Se/−Zn 10% FBS group (<b>a</b>) or the +Se/+Zn group 10% FBS (<b>b</b>–<b>d</b>) were set as 1. Results are presented as mean + SD (n = 3–4). *** <span class="html-italic">p</span> &lt; 0.001 vs. −Se, # <span class="html-italic">p</span> &lt; 0.05, ## <span class="html-italic">p</span> &lt; 0.01, ### <span class="html-italic">p</span> &lt; 0.001 vs. −Zn, <span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.01, <span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.001 vs. 10% FBS, calculated by two-factorial ANOVA with Bonferroni’s post-test.</p>
Full article ">Figure 2
<p>Zinc increased the activity of TXNRD but not of GPX. HepG2 cells were treated with or without 50 nM sodium selenite in combination with or without 100 µM zinc sulfate for up to 72 h (<b>a</b>–<b>e</b>,<b>g</b>,<b>h</b>) or with indicated Zn concentrations (<b>f</b>) in media containing 2.5% FCS. Enzyme activities of GPX (a) and TXNRD (<b>e</b>,<b>f</b>) and the protein expression levels of GPX1 (<b>b</b>), GPX2 (<b>c</b>), GPX4 (<b>d</b>), TXNRD1 (<b>g</b>), and TXNRD2 (<b>h</b>) were determined photometrically (<b>a</b>,<b>e</b>,<b>f</b>) or by Western blot (<b>b</b>–<b>d</b>,<b>g</b>,<b>h</b>). Protein expression was normalized to Ponceau staining and presented relative to +Se/+Zn treatment. Results are presented as mean + SD (n = 3–4). * <span class="html-italic">p</span> &lt; 0.05, *** <span class="html-italic">p</span> &lt; 0.001 vs. −Se, # <span class="html-italic">p</span> &lt; 0.05, ## <span class="html-italic">p</span> &lt; 0.01, ### <span class="html-italic">p</span> &lt; 0.001 vs. −Zn, calculated by two-factorial ANOVA (<b>a</b>–<b>e</b>,<b>g</b>,<b>h</b>) or with one-factorial ANOVA (<b>f</b>) with Bonferroni’s post-test.</p>
Full article ">Figure 3
<p>Zinc enhanced the nuclear translocation of NRF2 and MTF1 and increased the expression of their target genes. HepG2 cells were treated with or without 50 nM sodium selenite in combination with or without 100 μM zinc sulfate up to 72 h (<b>c</b>,<b>d</b>) or treated with selenite for 72 h in combination with or without zinc for the time as indicated (<b>a</b>,<b>b</b>,<b>e</b>–<b>j</b>) in media containing 2.5% FCS. Nuclear protein levels of MTF1 (<b>a</b>), NRF2 (<b>b</b>) and SELENOH (<b>g</b>), enzyme activity of NQO1 (<b>c</b>), and protein expression of NQO1 (<b>d</b>) were determined photometrically (<b>c</b>) or by Western blot (<b>a</b>,<b>b</b>,<b>d</b>,<b>g</b>). Protein expression was normalized to Ponceau staining and presented relative to samples with selenium treatment in combination with 2 h zinc treatment (<b>a</b>,<b>b</b>) or to samples with selenium treatment in combination with 6 h zinc treatment (<b>g</b>) or samples with selenium treatment (<b>d</b>). The mRNA expression levels of NRF2 and MTF1 target genes (<b>e</b>–<b>f</b>,<b>h</b>–<b>j</b>) were analyzed by qPCR. Gene expression was normalized to the normalization factor of the reference genes HPRT, RPL13a, and GAPDH and presented relative to samples with selenium treatment and 6 h zinc treatment. Results are presented as mean + SD (n = 3–4). * <span class="html-italic">p</span> &lt; 0.05, ** <span class="html-italic">p</span> &lt; 0.01, *** <span class="html-italic">p</span> &lt; 0.001 vs. −Se, # <span class="html-italic">p</span> &lt; 0.05, ## <span class="html-italic">p</span> &lt; 0.01, ### <span class="html-italic">p</span> &lt; 0.001 vs. −Zn, <span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.001 vs. 6 h Zn, calculated by two-factorial ANOVA with Bonferroni’s post-test.</p>
Full article ">Figure 4
<p>NRF2 partially mediated the zinc effects on selenium homeostasis. HepG2 cells with a stable small hairpin RNA-mediated NRF2 knockdown (NRF2-KD) and scramble (Scr) control cells were treated with 50 nM sodium selenite for 72 h and 100 μM zinc sulfate for the indicated time (<b>a</b>–<b>c</b>) or for 72 h (<b>d</b>,<b>e</b>) in media containing 2.5% FCS. The nuclear protein levels of NRF2 (<b>a</b>,<b>c</b>) and MTF1 (<b>b</b>,<b>c</b>) were analyzed by Western blot. Protein expression was normalized to the Ponceau staining. Enzyme activities of TNXRD (<b>d</b>) and intracellular Se concentrations (<b>e</b>) were determined photometrically or by total reflection X-ray fluorescence spectrometry, respectively. The measurement was performed for 1000 s with 1 mg/L yttrium as an internal standard. The results are presented as mean + SD (n = 3). ### <span class="html-italic">p</span> &lt; 0.001 vs. –Zn, *** <span class="html-italic">p</span> &lt; 0.001 vs. 6 h Zn, <span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.001 vs. Scr (<b>a</b>,<b>b</b>) or *** <span class="html-italic">p</span> &lt; 0.001 vs. –Se, ### <span class="html-italic">p</span> &lt; 0.001 vs. –Zn, <span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.01, <span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.001 vs. Scr (<b>d</b>,<b>e</b>) calculated by two-factorial ANOVA with Bonferroni’s post-test.</p>
Full article ">Figure 5
<p>Trace element transporters were increased by zinc. HepG2 cells were treated with or without 50 nM sodium selenite for 72 h in combination with or without 100 μM zinc sulfate for indicated time (up to 72 h) in media containing 2.5% FCS. The mRNA expression levels of ZIP8 (<b>a</b>), XCT (<b>b</b>), and APOER2 (<b>d</b>) were analyzed by qPCR. Gene expression was normalized to the normalization factor of the reference genes HPRT, RPL13a, and GAPDH and was presented relative to samples with selenium and 6 h zinc treatment. XCT (<b>c</b>) and APOER2 (<b>e</b>) were analyzed by Western blot. Protein expression was normalized to Ponceau staining and presented relative to samples with selenium and 48 h (<b>c</b>) or 72 h zinc treatment (<b>e</b>). Results are presented as mean + SD (n = 3–4). * <span class="html-italic">p</span> &lt; 0.05 vs. −Se; # <span class="html-italic">p</span> &lt; 0.05, ## <span class="html-italic">p</span> &lt; 0.01, ### <span class="html-italic">p</span> &lt; 0.001 vs. −Zn, <span>$</span><span>$</span><span>$</span> <span class="html-italic">p</span> &lt; 0.001 vs. 6 h Zn, calculated by two-factorial ANOVA (<b>a</b>,<b>b</b>,<b>d</b>) or with one-factorial ANOVA (<b>c</b>,<b>e</b>) with Bonferroni’s post-test.</p>
Full article ">
28 pages, 1521 KiB  
Review
The Immunomodulatory Effects of Selenium: A Journey from the Environment to the Human Immune System
by Rebecka A. Sadler, Bonnie A. Mallard, Umesh K. Shandilya, Mohammed A. Hachemi and Niel A. Karrow
Nutrients 2024, 16(19), 3324; https://doi.org/10.3390/nu16193324 - 30 Sep 2024
Viewed by 1771
Abstract
Selenium (Se) is an essential nutrient that has gained attention for its impact on the human immune system. The purpose of this review is to explore Se’s immunomodulatory properties and to make up-to-date information available so novel therapeutic applications may emerge. People acquire [...] Read more.
Selenium (Se) is an essential nutrient that has gained attention for its impact on the human immune system. The purpose of this review is to explore Se’s immunomodulatory properties and to make up-to-date information available so novel therapeutic applications may emerge. People acquire Se through dietary ingestion, supplementation, or nanoparticle applications. These forms of Se can beneficially modulate the immune system by enhancing antioxidant activity, optimizing the innate immune response, improving the adaptive immune response, and promoting healthy gut microbiota. Because of these many actions, Se supplementation can help prevent and treat pathogenic diseases, autoimmune diseases, and cancers. This review will discuss Se as a key micronutrient with versatile applications that supports disease management due to its beneficial immunomodulatory effects. Further research is warranted to determine safe dosing guidelines to avoid toxicity and refine the application of Se in medical treatments. Full article
(This article belongs to the Section Micronutrients and Human Health)
Show Figures

Figure 1

Figure 1
<p>Selenium sources and uses. Se: selenium; SeMeth: selenomethionine; SeCys: selenocysteine [<a href="#B4-nutrients-16-03324" class="html-bibr">4</a>,<a href="#B5-nutrients-16-03324" class="html-bibr">5</a>,<a href="#B6-nutrients-16-03324" class="html-bibr">6</a>].</p>
Full article ">Figure 2
<p>Absorption, distribution, and metabolism of selenium. Se: selenium; SeMeth: selenomethionine; SeCys: selenocysteine; SelP: selenoprotein P [<a href="#B21-nutrients-16-03324" class="html-bibr">21</a>,<a href="#B28-nutrients-16-03324" class="html-bibr">28</a>,<a href="#B29-nutrients-16-03324" class="html-bibr">29</a>].</p>
Full article ">Figure 3
<p>Antioxidant activity of selected selenoproteins [<a href="#B52-nutrients-16-03324" class="html-bibr">52</a>].</p>
Full article ">
23 pages, 5143 KiB  
Article
Modular Polymerase Synthesis and Internal Protein Domain Swapping via Dual Opposed Frameshifts in the Ebola Virus L Gene
by David B. Stubbs, Jan A. Ruzicka and Ethan W. Taylor
Pathogens 2024, 13(10), 829; https://doi.org/10.3390/pathogens13100829 - 25 Sep 2024
Viewed by 694
Abstract
Sequence analysis of the Zaire ebolavirus (EBOV) polymerase (L gene) mRNA, using online tools, identified a highly ranked −1 programmed ribosomal frameshift (FS) signal including an ideal slippery sequence heptamer (UUUAAAA), with an overlapping coding region featuring two tandem UGA codons, immediately followed [...] Read more.
Sequence analysis of the Zaire ebolavirus (EBOV) polymerase (L gene) mRNA, using online tools, identified a highly ranked −1 programmed ribosomal frameshift (FS) signal including an ideal slippery sequence heptamer (UUUAAAA), with an overlapping coding region featuring two tandem UGA codons, immediately followed by an RNA region that is the inverse complement (antisense) to a region of the mRNA of the selenoprotein iodothyronine deiodinase II (DIO2). This antisense interaction was confirmed in vitro via electrophoretic gel shift assay, using cDNAs at the EBOV and DIO2 segments. The formation of a duplex between the two mRNAs could trigger the ribosomal frameshift, by mimicking the enhancing role of a pseudoknot structure, while providing access to the selenocysteine insertion sequence (SECIS) element contained in the DIO2 mRNA. This process would allow the −1 frame UGA codons to be recoded as selenocysteine, forming part of a C-terminal module in a low abundance truncated isoform of the viral polymerase, potentially functioning in a redox role. Remarkably, 90 bases downstream of the −1 FS site, an active +1 FS site can be demonstrated, which, via a return to the zero frame, would enable the attachment of the entire C-terminal of the polymerase protein. Using a construct with upstream and downstream reporter genes, spanning a wildtype or mutated viral insert, we show significant +1 ribosomal frameshifting at this site. Acting singly or together, frameshifting at these sites (both of which are highly conserved in EBOV strains) could enable the expression of several modified isoforms of the polymerase. The 3D modeling of the predicted EBOV polymerase FS variants using the AI tool, AlphaFold, reveals a peroxiredoxin-like active site with arginine and threonine residues adjacent to a putative UGA-encoded selenocysteine, located on the back of the polymerase “hand”. This module could serve to protect the viral RNA from peroxidative damage. Full article
(This article belongs to the Special Issue New Insights in Viral Diseases and Computational Biology)
Show Figures

Figure 1

Figure 1
<p>Schematic of the proposed mechanism for viral mRNA capturing the SECIS element of a host selenoprotein mRNA. As proposed by Taylor et al. [<a href="#B6-pathogens-13-00829" class="html-bibr">6</a>], viral mRNA (green) is bound to the ribosome and is tethered downstream to a host selenoprotein mRNA (red) by one or more antisense tethering interactions (ATIs), which can potentially be in either a protein coding region or in a 3′ untranslated region. The captured host mRNA SECIS element (red hairpin) can then enable the viral mRNA to recode its UGA stop codon as selenocysteine (Sec), adding it to the growing peptide chain (colored circles) instead of terminating translation. Abbreviations: EF = elongation factor; SBP2 = SECIS binding protein 2.</p>
Full article ">Figure 2
<p>Previously predicted FS site in the L gene of Ebola Zaire. The slippery sequence heptamer (UUUAAAA, in blue highlight) is a perfect match to the X XXY YYZ pattern that is observed in most −1 FS sites. Downstream, after a few bases, is a region that can form a pseudoknot, a structure formed from two interlocking stem–loops. The first stem is formed by the pairing of sequences A and A′ (red) and the second stem is formed from the B and B’ sequences (green). These can form the secondary structure at bottom left. In the alternative base pairing shown at bottom right, the same region of the L gene mRNA is also an antisense match to a region of the human deiodinase II (DIO2) mRNA (blue). The host mRNA in this pair includes an SECIS element that could facilitate recoding (as selenocysteine) of the tandem UGA codons (*) in the overlapping region of the L gene accessed by the frameshift. Thus, DIO2 mRNA binding could be a trigger for a FS event enabling the expression of a viral selenoprotein module [<a href="#B6-pathogens-13-00829" class="html-bibr">6</a>,<a href="#B20-pathogens-13-00829" class="html-bibr">20</a>].</p>
Full article ">Figure 3
<p>Schematic of the protein expression cassettes in the dual reporter FS assay construct. Translation begins with a green fluorescent protein (GFP) reporter gene (green) upstream of the viral L gene insert containing the putative FS site; downstream of the viral insert is the luciferase (LUC) reporter gene (yellow). With appropriately designed inserts and controls, this approach enables assessment of the ratio of the proteins translated in the zero vs. either the −1 or +1 reading frames. Shown here are the designs used to assay for a predicted +1 FS site.</p>
Full article ">Figure 4
<p>Predicted antisense interaction between regions of human DIO2 and Ebov L gene mRNAs. The interaction was initially identified using BLAST, then assessed further using RNAHybrid 2.2 [<a href="#B20-pathogens-13-00829" class="html-bibr">20</a>]. The predicted interaction (<span class="html-italic">p</span> &lt; 0.005) is shown as both a sequence alignment and in secondary structure format; the binding free energy was calculated as −28.4 kcal/mol (<span class="html-italic">p</span> &lt; 0.005). For comparison, the Ebola Pol MDE of −47.6 kcal/mol represents the lowest (best) possible binding energy for that fragment, calculated by binding the Ebola region shown to its Watson–Crick inverse complement.</p>
Full article ">Figure 5
<p>Gel mobility shift assay validating the antisense tethering interaction between regions of human iodothyronine deiodinase II cDNA and Ebola polymerase (L) cDNA. The red “hot pixels” are an artifact of the intense light signal at those points.</p>
Full article ">Figure 6
<p>Potential +1 FS site in EBOV L gene at the 3′ end of overlapping coding region in the −1 frame. The primary FS signal is the shifty proline codon followed by the stop codon, CCUUAA. The RNA stem A:A’ may be an additional enhancer of frameshifting and is predicted by RNA folding, along with a second stem that has the potential to form the B:B’ stem in the pseudoknotted structure shown here, as a kissing loop between stem A:A’ and a downstream stem (see <a href="#app1-pathogens-13-00829" class="html-app">Figure S2</a>). The entire RNA region shown in <a href="#app1-pathogens-13-00829" class="html-app">Figure S2</a> was included in the L gene insert used in the FS assay to validate +1 frameshifting at this site.</p>
Full article ">Figure 7
<p>Evidence for an active +1 frameshift site in the EBOV L gene. The bars correspond to the ratio of luciferase (LUC) to GFP expression in cells transfected with wildtype or mutant dual reporter assay constructs, or not expressing LUC as a baseline control (Zero-L). Data are the mean of 3 independent experiments ± SEM, each of which was conducted with 5 or 6 replicates. The ratio of LUC/GFP signals was normalized to Mutant = 1 in each separate experiment, as that represents 100% readthrough from GFP into the LUC regions. In the mutant control construct, this results in a significantly higher LUC production over the wildtype sequence (<span class="html-italic">p</span> = 0.003 by unpaired <span class="html-italic">t</span>-test). The wildtype construct still produced significant LUC relative to the Zero-L control (<span class="html-italic">p</span> = 0.005), confirming that +1 ribosomal frameshifting is induced by this L gene fragment, at about 10% efficiency.</p>
Full article ">Figure 8
<p>Predicted L protein isoforms resulting from single or dual opposed frameshifts with and without UGA stop codon readthrough. See text for details. Despite substantial truncation, isoforms B and C have the potential to possess functional RNA polymerase activity. Calculated molecular masses of the truncated isoforms B and C are 90 and 94 kDa, respectively. Isoforms C and D include a small 28-residue module with potential redox-associated function (hatched blue region), that is encoded in the −1 reading frame of the L gene. Panel E is modified from Figure 2A of Liang [<a href="#B38-pathogens-13-00829" class="html-bibr">38</a>], CC by 4.0.</p>
Full article ">Figure 9
<p>Sequence and overlapped structures of the frameshifted region from 5 AlphaFold2 models. (<b>A</b>). Protein sequence encoded in the −1 reading frame between the two predicted FS sites (<span class="html-italic">Sequence 1</span>), compared to the zero-frame sequence of the EBOV polymerase (<span class="html-italic">Sequence 2</span>). The U represents potential selenocysteine residues, encoded by the UGA codon. (<b>B</b>). Overlap of five different models generated by AlphaFold2, with the UGA codons modeled as cysteine (sulfur atoms shown in yellow). All five show a similar fold of four short beta strands. The two most highly ranked models (blue and red ribbons) are essentially identical (average Cα RMS = 0.68 Å) and differ only slightly from the other three models at the C-terminal end (right). Average Cα RMS of model 1 vs. model 5 was still only 2.4 Å. The sidechains of some important residues discussed in the text are shown.</p>
Full article ">Figure 10
<p>AlphaFold2 model of truncated frameshift isoform C of EBOV polymerase. (<b>A</b>). The truncated RdRp domain is rendered in ribbon (colored by position), terminating in the 28-residue module encoded in the −1 frame, shown in all-atom spacefill with CPK colors. The top left view is down into the palm of the polymerase hand, with the fingers at the top (green and cyan ribbon). Note that despite the deletion of the C-terminal thumb and downstream domains, the base of the “thumb” remains, as that region is encoded near the N-terminus of the protein (orange and yellow ribbon). The frameshifted domain is predicted by AlphaFold to occupy the same space and structure corresponding to part of polymerase Motif D and all of Motif E in the native structure, at the C-terminal end of the palm domain. (<b>B</b>). Orthogonal views of the same structure entirely in ribbon rendition, with a few key residues shown as follows: <span class="html-italic">A</span>, a cluster of acidic residues at the polymerase active site (Motifs A and C); <span class="html-italic">B</span>: a conserved pair of basic lysine residues that is identical in both the zero frame and −1 frame sequences, due to a run of seven A nucleotides in the coding sequence; and <span class="html-italic">R</span>, a putative redox module where two possible selenocysteine residues (in spacefill) protrude at the back of the polymerase hand.</p>
Full article ">Figure 11
<p>Putative −1 FS redox module compared to human peroxiredoxin 1 active site<b>.</b> Closeup of key residues in the EBOV frameshifted region (<b>left</b>) shown as R in <a href="#pathogens-13-00829-f010" class="html-fig">Figure 10</a>B, compared to that of a typical peroxiredoxin active site, from 1prx.pdb (<b>right</b>). The three universally conserved peroxiredoxin active site residues are threonine, cysteine, and arginine, with the cysteine located between the other two residues [<a href="#B44-pathogens-13-00829" class="html-bibr">44</a>]. In human peroxiredoxin 1, a glutamate (E50) also participates; in EBOV, glutamine Q322 could play a similar role, and is notably also an essential active site residue of glutathione peroxidases. Note that in the EBOV redox module, R318 and Q322 are contributed to by an upstream domain that is not part of the frameshift-encoded structure, as opposed to residues 790 and 791, encoded by tandem UGA codons in the −1 frame. These are shown here as selenocysteine, but could also be decoded as cysteine under conditions of Se deficiency via a known mechanism [<a href="#B42-pathogens-13-00829" class="html-bibr">42</a>]. The second selenocysteine or cysteine (residue 791) could play the role of the second, “resolving” cysteine of peroxiredoxins [<a href="#B44-pathogens-13-00829" class="html-bibr">44</a>].</p>
Full article ">Figure 12
<p>Opening of the active site cleft via a hinge motion in both the EBOV polymerase and in the swapped domain of isoform D. (<b>A</b>). <span class="html-italic">Left:</span> The EBOV polymerase structure from 1yer.pdb with G809 shown in spacefill, circled. <span class="html-italic">Right:</span> The same structure after rotation of a single backbone dihedral angle of G809 (<span class="html-italic">psi</span>) by 30°. (<b>B</b>). <span class="html-italic">Left:</span> The complete isoform D AlphaFold2 model with the GlyPro pair in the swapped domain shown in spacefill, circled. The two lysine residues that are conserved in this region in both the native and frameshifted proteins are displayed just above the circled GP pair, and the two putative selenium atoms of the frameshift-swapped redox domain are displayed as orange spheres. <span class="html-italic">Right:</span> The same structure after rotation of the four glycine and proline backbone dihedral angles by −6° and +34° for G810 <span class="html-italic">phi</span> and <span class="html-italic">psi</span>, and −14° and +7° for P811 <span class="html-italic">phi</span> and <span class="html-italic">psi</span>, respectively.</p>
Full article ">
15 pages, 3338 KiB  
Article
Alleviation of Lipid Disorder and Liver Damage in High-Fat Diet-Induced Obese Mice by Selenium-Enriched Cardamine violifolia with Cadmium Accumulation
by Junying Zhu, Qingqing Lv, Fengna Li, Ping Xu, Ziyu Han, Aolin Yang, Zhan Shi, Chao Wang, Jie Jiang, Yunfen Zhu, Xiaofei Chen, Lvhui Sun, Xin Gen Lei and Ji-Chang Zhou
Nutrients 2024, 16(18), 3208; https://doi.org/10.3390/nu16183208 - 22 Sep 2024
Viewed by 1424
Abstract
Background/Objectives: As a hyperaccumulator of selenium (Se), Cardamine violifolia (Cv) and its peptide extract could ameliorate the negative effects of a high-fat diet (HFD). However, the effects of the coaccumulation of cadmium (Cd) in Se-enriched Cv (Cv2) and the [...] Read more.
Background/Objectives: As a hyperaccumulator of selenium (Se), Cardamine violifolia (Cv) and its peptide extract could ameliorate the negative effects of a high-fat diet (HFD). However, the effects of the coaccumulation of cadmium (Cd) in Se-enriched Cv (Cv2) and the potential confounding effect on the roles of enriched Se remain unknown. We aimed to investigate whether Cv2 could alleviate HFD-induced lipid disorder and liver damage. Methods: Three groups of 31-week-old female mice were fed for 41 weeks (n = 10–12) with a control Cv-supplemented diet (Cv1D, 0.15 mg Se/kg, 30 µg Cd/kg, and 10% fat calories), a control Cv-supplemented HFD (Cv1HFD, 45% fat calories), and a Cv2-supplemented HFD (Cv2HFD, 1.5 mg Se/kg, 0.29 mg Cd/kg, and 45% fat calories). Liver and serum were collected to determine the element concentrations, markers of liver injury and lipid disorder, and mRNA and/or protein expression of lipid metabolism factors, heavy metal detoxification factors, and selenoproteins. Results: Both Cv1HFD and Cv2HFD induced obesity, and Cv2HFD downregulated Selenoi and upregulated Dio3 compared with Cv1D. When comparing Cv2HFD against Cv1HFD, Cv2 increased the liver Se and Cd, the protein abundance of Selenoh, and the mRNA abundance of 10 selenoproteins; reduced the serum TG, TC, and AST; reduced the liver TG, lipid droplets, malondialdehyde, and mRNA abundance of Mtf1 and Mt2; and differentially regulated the mRNA levels of lipid metabolism factors. Conclusions: Cv2 alleviated HFD-induced lipid dysregulation and liver damage, which was probably associated with its unique Se speciation. However, further research is needed to explore the interaction of plant-coenriched Se and Cd and its effects on health. Full article
Show Figures

Figure 1

Figure 1
<p>Growth performance in adult female mice during the 41-week experiment. (<b>A</b>) Body weight. (<b>B</b>) Total weight gain. Data are means ± SEM (<span class="html-italic">n</span> = 10–12) and differ with ****, <span class="html-italic">p</span> &lt; 0.0001. <span class="html-italic">Cv1</span>D, control diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv</span>, <span class="html-italic">Cardamine violifolia</span>; <span class="html-italic">Cv1</span>HFD, high-fat diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv2</span>HFD, high-fat diet with 1.5 mg Se and 0.29 mg Cd (in the form of Se-enriched <span class="html-italic">Cv</span>)/kg.</p>
Full article ">Figure 2
<p>Accumulation of selenium (Se) (<b>A</b>) and cadmium (Cd) (<b>B</b>) and their correlation (<b>C</b>) in the livers of adult female mice. Data are means ± SEM (<span class="html-italic">n</span> = 9–12) and differ with ****, <span class="html-italic">p</span> &lt; 0.0001. <span class="html-italic">Cv1</span>D, control diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv</span>, <span class="html-italic">Cardamine violifolia</span>; <span class="html-italic">Cv1</span>HFD, high-fat diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv2</span>HFD, high-fat diet with 1.5 mg Se and 0.29 mg Cd (in the form of Se-enriched <span class="html-italic">Cv</span>)/kg.</p>
Full article ">Figure 3
<p>Lipid levels in the serum and liver and hepatic injury in adult female mice. (<b>A</b>–<b>C</b>) Serum concentrations of TG, TC, and LDL-C. (<b>D</b>–<b>F</b>) Liver concentrations of TG, TC, and LDL-C. (<b>G</b>,<b>H</b>) Serum ALT and AST activity. Data are means ± SEM (<span class="html-italic">n</span> = 8–10) and differ with *, <span class="html-italic">p</span> &lt; 0.05; **, <span class="html-italic">p</span> &lt; 0.01; ***, <span class="html-italic">p</span> &lt; 0.001; and ****, <span class="html-italic">p</span> &lt; 0.0001. ALT, alanine aminotransferase; AST, aspartate transaminase; <span class="html-italic">Cv1</span>D, control diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv</span>, <span class="html-italic">Cardamine violifolia</span>; <span class="html-italic">Cv1</span>HFD, high-fat diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv2</span>HFD, high-fat diet with 1.5 mg Se and 0.29 mg Cd (in the form of Se-enriched <span class="html-italic">Cv</span>)/kg; LDL-C, low-density lipoprotein cholesterol; Se, selenium; TC, total cholesterol; TG, triglyceride.</p>
Full article ">Figure 4
<p>Improvement due to selenium (Se)-enriched <span class="html-italic">Cardamine violifolia</span> (<span class="html-italic">Cv</span>) in hepatic lipid accumulation and related molecular mechanisms. (<b>A</b>) H&amp;E staining of the liver tissue with scale bar = 100 μM; the average number and area of hepatic lipid droplets were quantified with three fields per slide by using the “Analyze particles” function of ImageJ, and the mean values were calculated. (<b>B</b>) Total liver weight. (<b>C</b>) Expression of metabolic genes of triglycerides. (<b>D</b>) Expression of metabolic genes of cholesterol. Data are means ± SEM (<span class="html-italic">n</span> = 6–12) and differ with *, <span class="html-italic">p</span> &lt; 0.05; **, <span class="html-italic">p</span> &lt; 0.01; and ***, <span class="html-italic">p</span> &lt; 0.001. <span class="html-italic">Abca1</span>, ATP-binding cassette subfamily A member 1; <span class="html-italic">Abcg1</span>/<span class="html-italic">5</span>/<span class="html-italic">8</span>, ATP-binding cassette subfamily G member 1/5/8; <span class="html-italic">Atgl</span>, adipose triglyceride lipase; ATP, adenosine triphosphate; <span class="html-italic">Cv1</span>D, control diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Dgat1</span>/<span class="html-italic">2</span>, diacylglycerol O-acyltransferase 1/2; <span class="html-italic">Gpat1</span>/<span class="html-italic">2</span>, glycerol-3-phosphate acyltransferase 1/2; <span class="html-italic">Cv1</span>HFD, high-fat diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv2</span>HFD, high-fat diet with 1.5 mg Se and 0.29 mg Cd (in the form of Se-enriched <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Hmgcr</span>, 3-hydroxy-3-methylglutaryl coenzyme A reductase; <span class="html-italic">Hsl</span>, hormone-sensitive lipase; <span class="html-italic">Ldlr</span>, low-density lipoprotein receptor; <span class="html-italic">Lipc</span>, lipase C hepatic type; <span class="html-italic">Lpl</span>, lipoprotein lipase; No., number; <span class="html-italic">Sqle</span>, squalene epoxidase.</p>
Full article ">Figure 5
<p>Oxidative stress and antioxidant levels in the livers of adult female mice. (<b>A</b>–<b>C</b>) The relative mRNA expression of <span class="html-italic">Mtf1</span>, <span class="html-italic">Mt1</span>, and <span class="html-italic">Mt2</span>. (<b>D</b>) A representative immunoblot image for Mtf1, Mt, and Sod1. (<b>E</b>–<b>G</b>) The relative protein levels of Mtf1, Mt, and Sod1. (<b>H</b>) The T-AOC level. (<b>I</b>) The level of MDA in the liver. Data are means ± SEM (<span class="html-italic">n</span> = 8–12) and differ with *, <span class="html-italic">p</span> &lt; 0.05; **, <span class="html-italic">p</span> &lt; 0.01; and ***, <span class="html-italic">p</span> &lt; 0.001. <span class="html-italic">Cv1</span>D, control diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv</span>, <span class="html-italic">Cardamine violifolia</span>; <span class="html-italic">Cv1</span>HFD, high-fat diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv2</span>HFD, high-fat diet with 1.5 mg Se and 0.29 mg Cd (in the form of Se-enriched <span class="html-italic">Cv</span>)/kg; MDA, malondialdehyde; <span class="html-italic">Mt1</span>/<span class="html-italic">2</span>, metallothionein 1/2; <span class="html-italic">Mtf1</span>, metal regulatory transcription factor 1; Se, selenium; Sod1, superoxide dismutase 1; T-AOC, total antioxidant capacity.</p>
Full article ">Figure 6
<p>Expression of selenoproteins in the livers of adult female mice. (<b>A</b>) The relative mRNA expression levels of <span class="html-italic">Selenof</span>, <span class="html-italic">Selenoi</span>, and <span class="html-italic">Selenos</span>. (<b>B</b>) The relative mRNA expression levels of <span class="html-italic">Dio1</span>, <span class="html-italic">Dio3</span>, <span class="html-italic">Gpx4</span>, <span class="html-italic">Msrb1</span>, <span class="html-italic">Selenoo</span>, <span class="html-italic">Selenow</span>, <span class="html-italic">Sephs2</span>, <span class="html-italic">Txnrd1</span>, <span class="html-italic">Txnrd2</span>, and <span class="html-italic">Txnrd3</span>. (<b>C</b>) A representative immunoblot image for Selenof, Selenoi, and Selenos. (<b>D</b>) A representative immunoblot image for Dio1, Dio3, Gpx4, Selenoo, Txnrd1, and Txnrd2. (<b>E</b>) The relative protein levels of Selenof, Selenoi, and Selenos. (<b>F</b>) The relative protein levels of Dio1, Dio3, Gpx4, Selenoo, Txnrd1, and Txnrd2. Data are means ± SEM (<span class="html-italic">n</span> = 8–12) and differ with *, <span class="html-italic">p</span> &lt; 0.05 and **, <span class="html-italic">p</span> &lt; 0.01. <span class="html-italic">Cv1</span>D, control diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv</span>, <span class="html-italic">Cardamine violifolia</span>; <span class="html-italic">Dio1/3</span>, iodothyronine deiodinase 1/3; ER, endoplasmic reticulum; <span class="html-italic">Gpx4</span>, glutathione peroxidase 4; <span class="html-italic">Cv1</span>HFD, high-fat diet with 0.15 mg Se and 0.03 mg Cd (in the form of control <span class="html-italic">Cv</span>)/kg; <span class="html-italic">Cv2</span>HFD, high-fat diet with 1.5 mg Se and 0.29 mg Cd (in the form of Se-enriched <span class="html-italic">Cv</span>)/kg; <span class="html-italic">MsrB1</span>, methionine sulfoxide reductase B1; Se, selenium; <span class="html-italic">Selenof/i/o/s/w</span>, selenoprotein F/I/O/S/W; <span class="html-italic">Sephs2</span>, selenophosphate synthetase 2; <span class="html-italic">Txnrd1/2/3</span>, thioredoxin reductase 1/2/3.</p>
Full article ">
18 pages, 3012 KiB  
Review
Overcoming Challenges with Biochemical Studies of Selenocysteine and Selenoproteins
by Antavius Cain and Natalie Krahn
Int. J. Mol. Sci. 2024, 25(18), 10101; https://doi.org/10.3390/ijms251810101 - 20 Sep 2024
Viewed by 784
Abstract
Selenocysteine (Sec) is an essential amino acid that distinguishes itself from cysteine by a selenium atom in place of a sulfur atom. This single change imparts distinct chemical properties to Sec which are crucial for selenoprotein (Sec-containing protein) function. These properties include a [...] Read more.
Selenocysteine (Sec) is an essential amino acid that distinguishes itself from cysteine by a selenium atom in place of a sulfur atom. This single change imparts distinct chemical properties to Sec which are crucial for selenoprotein (Sec-containing protein) function. These properties include a lower pKa, enhanced nucleophilicity, and reversible oxidation. However, studying Sec incorporation in proteins is a complex process. While we find Sec in all domains of life, each domain has distinct translation mechanisms. These mechanisms are unique to canonical translation and are composed of Sec-specific enzymes and an mRNA hairpin to drive recoding of the UGA stop codon with Sec. In this review, we highlight the obstacles that arise when investigating Sec insertion, and the role that Sec has in proteins. We discuss the strategic methods implemented in this field to address these challenges. Though the Sec translation system is complex, a remarkable amount of information has been obtained and specialized tools have been developed. Continued studies in this area will provide a deeper understanding on the role of Sec in the context of proteins, and the necessity that we have for maintaining this complex translation machinery to make selenoproteins. Full article
(This article belongs to the Special Issue Molecular Research of Selenocysteine in Selenoproteins)
Show Figures

Figure 1

Figure 1
<p>Natural insertion of selenocysteine (Sec) into proteins. (<b>A</b>) Sec is synthesized on its tRNA (tRNA<sup>Sec</sup>) because of the labile nature of the free amino acid (red letters). This requires tRNA<sup>Sec</sup> to first be aminoacylated with serine (Ser) by seryl-tRNA synthetase (SerRS). Ser is then converted to Sec in a single step in bacteria by selenocysteine synthase (SelA) or two steps in archaea and eukaryotes by <span class="html-italic">O</span>-phosphoseryl-tRNA<sup>Sec</sup> kinase (PSTK) and <span class="html-italic">O</span>-phosphoserine tRNA<sup>Sec</sup>: selenocysteine synthase (SepSecS). Selenophosphate (SePO<sub>3</sub><sup>3−</sup>) is the donor molecule required for converting Ser to Sec, but this is also labile (red letters) and therefore is generated by a separate enzyme (SelD or SPS2) from elemental selenium (HSe<sup>−</sup>). (<b>B</b>) The process of bringing Sec-tRNA<sup>Sec</sup> to the ribosome for translation requires multiple factors that differ depending on the domain of life. Briefly, Sec-tRNA<sup>Sec</sup> is recognized by a specialized elongation factor (SelB, aSelB or EFSec) and is directed to a UGA codon by an mRNA hairpin (SECIS element) required in the translated (bacteria) or untranslated region (archaea and eukaryotes) of the mRNA.</p>
Full article ">Figure 2
<p>Schematic of experimental requirements required to study various stages of (<b>A</b>) aminoacylation and (<b>B</b>) translation for selenoproteins. Molecules are color-coordinated to follow when they are needed.</p>
Full article ">Figure 3
<p>Strategies to express selenoproteins that (<b>A</b>) utilize the SECIS element or (<b>B</b>) are SECIS independent. (<b>A</b>) In bacteria, when the Sec to be inserted is near the C-terminal end of the protein, a SECIS element is placed right after the stop codon (UAA) in the 3′UTR. In eukaryotes, a SECIS element from <span class="html-italic">Toxoplasma gondii</span> is inserted downstream of the designated stop codon in the 3′UTR. (<b>B</b>) Removing the SECIS element in <span class="html-italic">Escherichia coli</span> and <span class="html-italic">Saccharomyces cerevisiae</span> requires the use of an engineered tRNA (allo-tRNA or <span class="html-italic">Sc</span>tRNA<sup>Sec</sup>) that is recognized by <span class="html-italic">Aeromonas salmonicida</span> SelA for conversion of Ser to Sec and the endogenous elongation factor (EF-Tu or EF-1α).</p>
Full article ">Figure 4
<p>Cell-free methods for selenoprotein expression involving (<b>A</b>) in vitro protein expression and (<b>B</b>) chemical synthesis. (<b>A</b>) Selenoproteins can be expressed using in vitro translation systems based on the previously described in vivo methods. Here we show one method that cannot be replicated in vivo, which involves acylating tRNA<sup>Cys</sup> with Sec using CysRS. Under the correct reducing conditions (DTT in an anaerobic environment), selenocystine is reduced to Sec and can be used as a substrate for aminoacylation. In the absence of Cys, all codons encoding for Cys will contain Sec. (<b>B</b>) Native chemical ligation involves the fusion of two peptide fragments together. These can be expressed in a host or chemically synthesized. For selenoproteins, Sec is an active amino acid in the ligation reaction and, with chemical synthesis into a peptide, can facilitate the formation of a selenoprotein.</p>
Full article ">
17 pages, 1305 KiB  
Review
An Updated Review of Emerging Sources of Selenium in Weaned Piglet Nutrition
by Wenyue Zhou, Zheng Yang, Jiajun Han, Xingping Chen, Tiande Zou, Jinming You and Jun Chen
Animals 2024, 14(17), 2599; https://doi.org/10.3390/ani14172599 - 6 Sep 2024
Viewed by 1032
Abstract
The antioxidant and immune systems of weaned piglets are not fully mature and are also subjected to serious stress challenges related to oxidative stress and inflammation. Selenium (Se) is an essential element for pigs, with documented roles encompassing antioxidative and anti-inflammatory properties via [...] Read more.
The antioxidant and immune systems of weaned piglets are not fully mature and are also subjected to serious stress challenges related to oxidative stress and inflammation. Selenium (Se) is an essential element for pigs, with documented roles encompassing antioxidative and anti-inflammatory properties via selenoproteins. Sodium selenite and Se-enriched yeast are commonly acknowledged as conventional sources of Se for piglets. In the past decade, several novel Se sources have emerged in the field of weaned piglet nutrition. In this review, we will initially outline the historical timeline of Se sources as reported in weaned piglet nutrition. Afterwards, our attention will turn towards the nutritional regulation of Se sources in relation to the antioxidant and anti-inflammatory aspects of healthy weaned piglets. Ultimately, we will provide a detailed review highlighting the potential of emerging Se sources in alleviating various adverse effects of stress challenges faced by weaned piglets. These challenges include oxidative stress, enterotoxigenic Escherichia coli infection, lipopolysaccharide-induced inflammation, heat stress, and exposure to feed mycotoxins. The output of this review will emphasize the fundamental importance of incorporating emerging Se sources in the diet of weaned piglets. Full article
(This article belongs to the Special Issue Impact of Genetics and Feeding on Growth Performance of Pigs)
Show Figures

Figure 1

Figure 1
<p>Historical timeline of selenium (Se) sources reported in the field of weaned piglet nutrition for the first time [<a href="#B14-animals-14-02599" class="html-bibr">14</a>,<a href="#B27-animals-14-02599" class="html-bibr">27</a>,<a href="#B28-animals-14-02599" class="html-bibr">28</a>,<a href="#B29-animals-14-02599" class="html-bibr">29</a>,<a href="#B30-animals-14-02599" class="html-bibr">30</a>,<a href="#B31-animals-14-02599" class="html-bibr">31</a>,<a href="#B33-animals-14-02599" class="html-bibr">33</a>,<a href="#B34-animals-14-02599" class="html-bibr">34</a>,<a href="#B37-animals-14-02599" class="html-bibr">37</a>,<a href="#B38-animals-14-02599" class="html-bibr">38</a>,<a href="#B39-animals-14-02599" class="html-bibr">39</a>,<a href="#B45-animals-14-02599" class="html-bibr">45</a>].</p>
Full article ">Figure 2
<p>The schematic diagram of emerging selenium (Se) sources for weaned piglets to combat external stress challenges based on current knowledge [<a href="#B14-animals-14-02599" class="html-bibr">14</a>,<a href="#B38-animals-14-02599" class="html-bibr">38</a>,<a href="#B39-animals-14-02599" class="html-bibr">39</a>,<a href="#B40-animals-14-02599" class="html-bibr">40</a>,<a href="#B41-animals-14-02599" class="html-bibr">41</a>,<a href="#B42-animals-14-02599" class="html-bibr">42</a>,<a href="#B43-animals-14-02599" class="html-bibr">43</a>,<a href="#B44-animals-14-02599" class="html-bibr">44</a>,<a href="#B45-animals-14-02599" class="html-bibr">45</a>,<a href="#B46-animals-14-02599" class="html-bibr">46</a>].</p>
Full article ">
18 pages, 3650 KiB  
Article
Impaired Upper Airway Muscle Function with Excessive or Deficient Dietary Intake of Selenium in Rats
by David P. Burns, Sarah E. Drummond, Stefanie Wölfel, Kevin H. Murphy, Joanna Szpunar, Ken D. O’Halloran and John J. Mackrill
Antioxidants 2024, 13(9), 1080; https://doi.org/10.3390/antiox13091080 - 4 Sep 2024
Viewed by 798
Abstract
Obstructive sleep apnoea (OSA) involves impaired upper airway muscle function and is linked to several pathologies including systemic hypertension, daytime somnolence and cognitive decline. Selenium is an essential micronutrient that exerts many of its effects through selenoproteins. Evidence indicates that either deficient or [...] Read more.
Obstructive sleep apnoea (OSA) involves impaired upper airway muscle function and is linked to several pathologies including systemic hypertension, daytime somnolence and cognitive decline. Selenium is an essential micronutrient that exerts many of its effects through selenoproteins. Evidence indicates that either deficient or excessive dietary selenium intake can result in impaired muscle function, termed nutritional myopathy. To investigate the effects of selenium on an upper airway muscle, the sternohyoid, rats were fed on diets containing deficient, normal (0.5 ppm sodium selenite) or excessive (5 ppm selenite) selenium for a period of two weeks. Sternohyoid contractile function was assessed ex vivo. Serum selenium levels and activity of the glutathione antioxidant system were determined by biochemical assays. The abundance of three key muscle selenoproteins (selenoproteins -N, -S and -W (SELENON, SELENOS and SELENOW)) in sternohyoid muscle were quantified by immunoblotting. Levels of these selenoproteins were also compared between rats exposed to chronic intermittent hypoxia, a model of OSA, and sham treated animals. Although having no detectable effect on selected organ masses and whole-body weight, either selenium-deficient or -excessive diets severely impaired sternohyoid contractile function. These changes did not involve altered fibre size distribution. These dietary interventions resulted in corresponding changes in serum selenium concentrations but did not alter the activity of glutathione-dependent antioxidant systems in sternohyoid muscle. Excess dietary selenium increased the abundance of SELENOW protein in sternohyoid muscles but had no effect on SELENON or SELENOS. In contrast, chronic intermittent hypoxia increased SELENON, decreased SELENOW and had no significant effect on SELENOS in sternohyoid muscle. These findings indicate that two-week exposure to selenium-deficient or -excessive diets drastically impaired upper airway muscle function. In the sternohyoid, SELENON, SELENOS and SELENOW proteins show distinct alterations in level following exposure to different dietary selenium intakes, or to chronic intermittent hypoxia. Understanding how alterations in Se and selenoproteins impact sternohyoid muscle function has the potential to be translated into new therapies for prevention or treatment of OSA. Full article
(This article belongs to the Section Antioxidant Enzyme Systems)
Show Figures

Figure 1

Figure 1
<p>Impact of dietary Se intake on serum Se levels in rats. For two weeks, three groups of adult male Wistar rats were fed with either Se-deficient chow (nominally Se free), or that supplemented with either 0.5 ppm or 5 ppm sodium selenite (n = 10 for each group). The box-and-whisker plot shows the effect of these dietary interventions on serum selenium levels. Levels of Se were significantly different between groups (<span class="html-italic">p</span> &lt; 0.0001 by one-way ANOVA; 0.5 ppm versus 5 ppm <span class="html-italic">p</span> &lt; 0.05 and deficient versus 5 ppm <span class="html-italic">p</span> &lt; 0.01, using Dunnett’s post hoc test).</p>
Full article ">Figure 2
<p>Dietary Se deficiency or excess has a detrimental impact on ex vivo contractile performance of rat sternohyoid muscle. The performance of SH muscle from rats fed with diets either deficient, adequate (0.5 ppm) or excessive (5 ppm) in selenium (n = 8 for each condition) was investigated ex vivo. For twitch kinetics, the time to peak (CT) was significantly reduced in deficient rats and significantly increased in 5 ppm Se diets, (panel <b>A</b>) and <a href="#antioxidants-13-01080-t002" class="html-table">Table 2</a>. The peak twitch tension (P<sub>t</sub>) was also significantly reduced in deficient rat SH relative to the 0.5 ppm controls, <a href="#antioxidants-13-01080-t002" class="html-table">Table 2</a>. Half-relaxation time was not significantly different between the three groups, (panel <b>B</b>). For specific force–stimulation frequency relationships, Se deficiency resulted in a significant reduction in peak isometric tetanic force relative to the control chow diet. Force–frequency relationships (original traces, panel <b>C</b>) and graphical presentation of mean data with SD (panel <b>D</b>) were compared using repeated measures two-way ANOVA; deficient versus 0.5 ppm, frequency: <span class="html-italic">p</span> &lt; 0.0001, deficient: <span class="html-italic">p</span> &lt; 0.0001, interaction: <span class="html-italic">p</span> &lt; 0.0001 (80 Hz <span class="html-italic">p</span> &lt; 0.05 (*), 100 Hz <span class="html-italic">p</span> &lt; 0.01 (**), 120 Hz <span class="html-italic">p</span> &lt; 0.01 (**), 140 Hz <span class="html-italic">p</span> &lt; 0.01 (**), 160 Hz <span class="html-italic">p</span> &lt; 0.01(**)); deficient versus 5 ppm, frequency: <span class="html-italic">p</span> &lt; 0.0001, 5 ppm: <span class="html-italic">p</span> = 0.1262, interaction: <span class="html-italic">p</span> = 0.0801; 0.5 ppm versus 5 ppm, frequency: <span class="html-italic">p</span> &lt; 0.0001, 5 ppm: <span class="html-italic">p</span> = 0.2491, interaction: <span class="html-italic">p</span> = 0.0322.</p>
Full article ">Figure 3
<p>Effect of dietary Se deficiency or excess on rat sternohyoid contractile response to repeated stimulation. (Panel <b>A</b>) shows representative original traces of force production by SH muscle from deficient-, 0.5 ppm- or 5 ppm-Se fed rats, in response to repeat bouts of electrical stimulation. (Panel <b>B</b>) shows mean specific force generation versus time of stimulation (n = 8), including SD. By two-way ANOVA, time: <span class="html-italic">p</span> &lt; 0.0001, deficient versus 0.5 ppm: <span class="html-italic">p</span> = 0.0042, 0.5 ppm versus 5 ppm: <span class="html-italic">p</span> = 0.0609, deficient versus 5 ppm: <span class="html-italic">p</span> = 0.2042; * <span class="html-italic">p</span> &lt; 0.05, *** <span class="html-italic">p</span> &lt; 0.001.</p>
Full article ">Figure 4
<p>Se deficiency or excess does not affect rat sternohyoid muscle fibre size. Feeding rats with deficient, 0.5 ppm (adequate) or 5 ppm (excessive) Se diets for two weeks did not significantly alter SH muscle fibre size. (<b>A</b>) shows representative images of sternohyoid sections immunofluorescently labelled for laminin to show individual muscle fibres. (<b>B</b>) Data are shown as mean (SD) for Se-deficient (n = 7), control (0.5 ppm, n = 8) and Se-excess (5 ppm, n = 9). Data were statistically compared by one-way ANOVA for mean minimal Feret’s diameter (above) and mean cross-sectional area (CSA) Horizontal scale bars represent 200 μm.</p>
Full article ">Figure 5
<p>Effect of Se deficiency or excess on glutathione-dependent anti-oxidant systems in rat sternohyoid muscle. Feeding rats with deficient, 0.5 ppm (adequate) or 5 ppm (excessive) Se diets for two weeks did not significantly alter sternohyoid muscle glutathione reductase activity, glutathione peroxidase activity, total glutathione (all per mg protein), or the ratio between oxidized and reduced glutathione (GSSG:GSH). The box-and-whisker plots display mean values (n = 8) and SD. <span class="html-italic">p</span> values were determined by one-way ANOVA.</p>
Full article ">Figure 6
<p>Effect of dietary selenium levels on rat sternohyoid muscle SELENON, SELENOS and SELENOW protein abundance. Sternohyoid muscle lysates (100 μg protein/lane) from the three dietary groups were resolved by 7.5% (<b>A</b>) or 15% (<b>B</b>) and (<b>C</b>) SDS-PAGE, transferred onto nitrocellulose and probed with rabbit antibodies against SELENON (<b>A</b>), SELENOS (<b>B</b>) or SELENOW (<b>C</b>). Immunodecorated proteins, indicated by arrows, were detected as described in <a href="#sec2dot6dot1-antioxidants-13-01080" class="html-sec">Section 2.6.1</a>. Protein densities were expressed as a ratio of the total protein loading (Ponceau S staining) and were normalized to the mean 0.5 ppm ratio. The graphs on the left-hand side of each panel show these normalized values (with SD, n = 10). One-way ANOVA indicated no significant differences between groups for SELENON and SELENOS, but in the 5 ppm Se group, SELENOW protein levels were significantly enhanced relative to Se-deficient (<span class="html-italic">a</span>, <span class="html-italic">p</span> &lt; 0.001) or 0.5 ppm Se (<span class="html-italic">b</span>, <span class="html-italic">p</span> &lt; 0.05) groups.</p>
Full article ">Figure 7
<p>Effects of chronic intermittent hypoxia (CIH) on SELENON, SELENOS and SELENOW protein abundance in rat sternohyoid and diaphragm muscles. Western blots of sternohyoid (SH) and diaphragm (DIA) muscle lysates (100 μg protein/lane), from sham- or CIH-treated rats, were probed with antisera against SELENON (<b>A</b>), SELENOS (<b>B</b>) or SELENOW (<b>C</b>), as described in <a href="#antioxidants-13-01080-f006" class="html-fig">Figure 6</a>. Immunoreactive band densities were divided by total protein densities (Ponceau S) and the mean values (n = 3) were graphed as mean values (with SD), normalized to the mean SH sham ratio. By one-way ANOVA and Tukey’s multiple comparison test, for SELENON, <span class="html-italic">a</span>, <span class="html-italic">p</span> &lt; 0.001 versus SH sham; and for SELENOS, <span class="html-italic">b</span>, <span class="html-italic">p</span> &lt; 0.01 versus SH sham. Note that SELENOW protein was not detectable in diaphragm; by Student’s <span class="html-italic">t</span>-test, <span class="html-italic">c</span>, <span class="html-italic">p</span> &lt; 0.01 for SH sham versus SH CIH.</p>
Full article ">
14 pages, 915 KiB  
Review
Effect of Selenium and Selenoproteins on Radiation Resistance
by Shidi Zhang, Guowei Zhang, Pengjie Wang, Lianshun Wang, Bing Fang and Jiaqiang Huang
Nutrients 2024, 16(17), 2902; https://doi.org/10.3390/nu16172902 - 30 Aug 2024
Viewed by 976
Abstract
With the advancement of radiological medicine and nuclear industry technology, radiation is increasingly used to diagnose human health disorders. However, large-scale nuclear leakage has heightened concerns about the impact on human organs and tissues. Selenium is an essential trace element that functions in [...] Read more.
With the advancement of radiological medicine and nuclear industry technology, radiation is increasingly used to diagnose human health disorders. However, large-scale nuclear leakage has heightened concerns about the impact on human organs and tissues. Selenium is an essential trace element that functions in the body mainly in the form of selenoproteins. Selenium and selenoproteins can protect against radiation by stimulating antioxidant actions, DNA repair functions, and immune enhancement. While studies on reducing radiation through antioxidants have been conducted for many years, the underlying mechanisms of selenium and selenoproteins as significant antioxidants in radiation damage mitigation remain incompletely understood. Therefore, this paper aims to provide new insights into developing safe and effective radiation protection agents by summarizing the anti-radiation mechanism of selenium and selenoproteins. Full article
Show Figures

Figure 1

Figure 1
<p>Radiation resistance mechanism of different selenoproteins.</p>
Full article ">
15 pages, 461 KiB  
Article
Is Maternal Selenium Status Associated with Pregnancy Outcomes in Physiological and Complicated Pregnancy?
by Joanna Pieczyńska, Sylwia Płaczkowska, Rafał Sozański and Halina Grajeta
Nutrients 2024, 16(17), 2873; https://doi.org/10.3390/nu16172873 - 27 Aug 2024
Viewed by 964
Abstract
Selenium is essential for the synthesis and function of various selenoenzymes, such as glutathione peroxidases, selenoprotein P, and thioredoxin reductase. These enzymes play a critical role in both antioxidant defense and in limiting oxidative damage. Numerous studies have reported associations between serum selenium [...] Read more.
Selenium is essential for the synthesis and function of various selenoenzymes, such as glutathione peroxidases, selenoprotein P, and thioredoxin reductase. These enzymes play a critical role in both antioxidant defense and in limiting oxidative damage. Numerous studies have reported associations between serum selenium concentration, obstetric complications and pregnancy outcomes. The aim of this study was to determine whether the dietary intake of selenium, its serum concentration, and the activity of glutathione peroxidase in subsequent trimesters of pregnancy affect the birth condition of newborns. This was assessed based on the APGAR score in the 1st and 5th minute of life, birth weight, body length and head and chest circumference in both physiological and complicated pregnancy courses. Twenty-seven pregnant women, with a mean age of 29.6 ± 4.8 years from the Lower Silesia region of Poland, participated in the study. Fifty-five percent of the study group experienced pregnancy complications. The median reported selenium intake and serum selenium content for Polish pregnant women in the first trimester was 56.30 μg/day and 43.89 μg/L, respectively. These figures changed in the second trimester to 58.31 μg/day and 41.97 μg/L and in the third trimester to 55.60 μg/day and 41.90 μg/L. In the subgroup of pregnant women with a physiological pregnancy course, a weak, positive correlation was observed in the first trimester between Se intake and the length (R = 0.48, p = 0.019) and the birth weight of newborns (R = 0.472, p = 0.022). In the second trimester, a positive correlation was noted with the APGAR score at the 1st (R = 0.680, p = 0.005) and 5th minutes (R = 0.55, p = 0.033), and in the third trimester with the APGAR score at the 1st minute (R = 0.658, p = 0.019). The glutathione peroxidase activity had a strong positive correlation with the APGAR score at the 1st min (R = 0.650, p = 0.008) in the second trimester and with the birth weight of the newborns (R = 0.598, p = 0.039) in the third trimester. No correlation was found between newborns’ birth measurements and serum selenium concentration. In the subgroup of pregnant women with complications, a strong, negative correlation was found between Se intake in the second trimester and gestational age (R = −0.618, p = 0.032). In the third trimester, a positive correlation was noted between Se concentration in serum and head circumference (R = 0.587, p = 0.021). The results indicate that maternal selenium status during pregnancy, including dietary intake, serum concentration, and glutathione peroxidase activity, correlates with anthropometric parameters of the newborn, such as birth weight, length, and APGAR score, especially in pregnancies with a physiological course. However, these relationships diminish in importance when pregnancy complications occur. Full article
(This article belongs to the Special Issue Maternal Nutrition and Its Effect on Offspring Growth)
Show Figures

Figure 1

Figure 1
<p>Participant flow chart.</p>
Full article ">
20 pages, 3986 KiB  
Review
An Examination of Chemical Tools for Hydrogen Selenide Donation and Detection
by Rynne A. Hankins and John C. Lukesh
Molecules 2024, 29(16), 3863; https://doi.org/10.3390/molecules29163863 - 15 Aug 2024
Viewed by 975
Abstract
Hydrogen selenide (H2Se) is an emerging biomolecule of interest with similar properties to that of other gaseous signaling molecules (i.e., gasotransmitters that include nitric oxide, carbon monoxide, and hydrogen sulfide). H2Se is enzymatically generated in humans where it serves [...] Read more.
Hydrogen selenide (H2Se) is an emerging biomolecule of interest with similar properties to that of other gaseous signaling molecules (i.e., gasotransmitters that include nitric oxide, carbon monoxide, and hydrogen sulfide). H2Se is enzymatically generated in humans where it serves as a key metabolic intermediate in the production of selenoproteins and other selenium-containing biomolecules. However, beyond its participation in biosynthetic pathways, its involvement in cellular signaling or other biological mechanisms remains unclear. To uncover its true biological significance, H2Se-specific chemical tools capable of functioning under physiological conditions are required but lacking in comparison to those that exist for other gasotransmitters. Recently, researchers have begun to fill this unmet need by developing new H2Se-releasing compounds, along with pioneering methods for selenide detection and quantification. In combination, the chemical tools highlighted in this review have the potential to spark groundbreaking explorations into the chemical biology of H2Se, which may lead to its branding as the fourth official gasotransmitter. Full article
(This article belongs to the Special Issue Organosulfur and Organoselenium Chemistry)
Show Figures

Graphical abstract

Graphical abstract
Full article ">Figure 1
<p>A simplified schematic of H<sub>2</sub>Se production in mammalian systems. Glutathione (GSH), glutathione disulfide (GSSG), glutaredoxin (Grx), glutathione reductase (GR), thioredoxin (Trx), thioredoxin reductase (TrxR), transsulfuration pathway (TSP), selenocysteine lyase (SCLY), selenophosphate synthetase (SPS), and selenocysteine synthase (SecS).</p>
Full article ">Figure 2
<p>(<b>A</b>) Selenotrisulfides as H<sub>2</sub>Se donors. (<b>B</b>) Peripheral delivery of H<sub>2</sub>Se via human serum albumin. Glutathione (GSH), glutaredoxin (Grx), glutathione reductase (GR), thioredoxin (Trx), thioredoxin reductase (TrxR), and human serum albumin (HSA).</p>
Full article ">Figure 3
<p>(<b>A</b>) Chalcogen anhydrides (R-Se, R-S, and R-O) under examination; (<b>B</b>) Proposed fragmentation pattern for R-Se in 50% methanol/water under ESI conditions.</p>
Full article ">Figure 4
<p>Proposed hydrolytic pathway of TDN1042, which results in the release of two equivalents of H<sub>2</sub>Se and the formation of phenylphosphonic acid.</p>
Full article ">Figure 5
<p>(<b>A</b>) Cyclic-PSe donors arranged in order of decreasing rates of hydrolysis; (<b>B</b>) Proposed hydrolytic pathway of cyclic-PSe donors, which results in the release of one equivalent of H<sub>2</sub>Se and the formation of phenylphosphonic acid.</p>
Full article ">Figure 6
<p>(<b>A</b>) Rapid hydrolysis of SelenoMichler’s ketone generates H<sub>2</sub>Se in PBS (pH 7.4). (<b>B</b>) Proposed mechanism for cysteine/thiol-triggered H<sub>2</sub>Se donation from selenocyclopropenones. (<b>C</b>) Proposed mechanism for cysteine-triggered H<sub>2</sub>Se donation from arylselenoamides.</p>
Full article ">Figure 7
<p>(<b>A</b>) General strategy for generating stimuli-responsive COS/H<sub>2</sub>S donors. (<b>B</b>) Proposed mechanism for the direct release of H<sub>2</sub>Se from PhotoSeCM upon irradiation at 365 nm. (<b>C</b>) Control compound used to highlight the stability of <span class="html-italic">γ</span>-ketoselenocarbamates in water. (<b>D</b>) Proposed mechanism for the direct release of H<sub>2</sub>Se from <span class="html-italic">γ</span>-ketoselenocarbamates containing a deprotonatable hydrogen at the β position.</p>
Full article ">Figure 8
<p>(<b>A</b>) A library of <span class="html-italic">γ</span>-ketoselenides that undergo base-promoted α-deprotonation/β-elimination to release H<sub>2</sub>Se. (<b>B</b>) Cell growth inhibition of HeLa and HCT116 cells in culture. IC<sub>50</sub> values were determined after a 24 h incubation period with donor.</p>
Full article ">Figure 9
<p>5′-<span class="html-italic">O</span>-Selenophosphates (<b>dGMPSe</b>) undergo HINT1-catalyzed hydrolysis to generate H<sub>2</sub>Se.</p>
Full article ">Figure 10
<p>Use of nonspecific electrophilic traps to confirm H<sub>2</sub>Se release from donors ((<b>A</b>) TDN1042, (<b>B</b>) 1, (<b>C</b>) 10) by forming stable selenide/diselenide products that can be easily identified by spectroscopic methods.</p>
Full article ">Figure 11
<p>(<b>A</b>) Reaction-based fluorescent sensor with selectivity towards H<sub>2</sub>Se based on benzoselenadiazole Se–N bond cleavage. (<b>B</b>) A lysosomal-targeting fluorescent sensor for H<sub>2</sub>Se bioimaging.</p>
Full article ">Figure 12
<p>Proposed mechanism for Hcy-H<sub>2</sub>Se turn-on fluorescence initiated by H<sub>2</sub>Se-promoted disulfide reduction.</p>
Full article ">Figure 13
<p>(<b>A</b>,<b>B</b>) Commercially available electrophilic traps that provide a colorimetric readout for monitoring H<sub>2</sub>Se donor progress. (<b>C</b>) Cautioning researchers that complementary H<sub>2</sub>Se measurements should be employed to avoid confusing donor alkylation with the alkylation of released H<sub>2</sub>Se in solution.</p>
Full article ">Figure 14
<p>(<b>A</b>) Aryl azide reduction for H<sub>2</sub>S-initiated turn-on fluorescence. (<b>B</b>) The use of aryl azides as fluorescent probes for tracking H<sub>2</sub>Se liberation from donor compounds.</p>
Full article ">
28 pages, 6856 KiB  
Article
Regulatory Role and Cytoprotective Effects of Exogenous Recombinant SELENOM under Ischemia-like Conditions and Glutamate Excitotoxicity in Cortical Cells In Vitro
by Egor A. Turovsky, Egor Y. Plotnikov and Elena G. Varlamova
Biomedicines 2024, 12(8), 1756; https://doi.org/10.3390/biomedicines12081756 - 5 Aug 2024
Viewed by 930
Abstract
Despite the successes in the prevention and treatment of strokes, it is still necessary to search for effective cytoprotectors that can suppress the damaging factors of cerebral ischemia. Among the known neuroprotectors, there are a number of drugs with a protein nature. In [...] Read more.
Despite the successes in the prevention and treatment of strokes, it is still necessary to search for effective cytoprotectors that can suppress the damaging factors of cerebral ischemia. Among the known neuroprotectors, there are a number of drugs with a protein nature. In the present study, we were able to obtain recombinant SELENOM, a resident of the endoplasmic reticulum that exhibits antioxidant properties in its structure and functions. The resulting SELENOM was tested in two brain injury (in vitro) models: under ischemia-like conditions (oxygen-glucose deprivation/reoxygenation, OGD/R) and glutamate excitotoxicity (GluTox). Using molecular biology methods, fluorescence microscopy, and immunocytochemistry, recombinant SELENOM was shown to dose-dependently suppress ROS production in cortical cells in toxic models, reduce the global increase in cytosolic calcium ([Ca2+]i), and suppress necrosis and late stages of apoptosis. Activation of SELENOM’s cytoprotective properties occurs due to its penetration into cortical cells through actin-dependent transport and activation of the Ca2+ signaling system. The use of SELENOM resulted in increased antioxidant protection of cortical cells and suppression of the proinflammatory factors and cytokines expression. Full article
(This article belongs to the Special Issue Advanced Research in Neuroprotection)
Show Figures

Figure 1

Figure 1
<p>Effect of 24 h pre-incubation of neuroglial cortical cultures with 5 or 20 μg/mL exogenous SELENOM on GluTox-induced cell death. Double staining of cells with Hoechst 33342 (HO342) and propidium iodide (PI). GluTox—induction of the excitotoxic effect of glutamate (300 µM glutamate for 24 h) without prior incubation with SELENOM. SELENOM 5 µg and SELENOM 20 µg—24 h pre-incubation of cells with SELENOM and then 300 µM glutamate (GluTox) was added to the culture medium for 24 h.</p>
Full article ">Figure 2
<p>Effect of 24 h pre-incubation of cortical cells with various concentrations of exogenous SELENOM on GluTox-induced cell death. (<b>A</b>) Cytogram demonstrating the viability of cortical cells after 24 h of exposure to GluTox depending on the concentration of SELENOM. <span class="html-italic">X</span>-axis—PI fluorescence intensity. The <span class="html-italic">Y</span>-axis is the fluorescence intensity of Hoechst 33342. Cells were stained with probes 24 h after GluTox. (<b>B</b>) Effect of 24 h pre-incubation with 5 or 20 μg of SELENOM on the induction of necrosis and apoptosis. Black asterisks indicate the differences between the experimental groups comparable to the Control group. Differences between experimental groups are marked with asterisks of different colors. n/s—data not significant (<span class="html-italic">p</span> &gt; 0.05), *** <span class="html-italic">p</span> &lt; 0.001, ** <span class="html-italic">p</span> &lt; 0.01, and * <span class="html-italic">p</span> &lt; 0.05. Cell images are presented in <a href="#biomedicines-12-01756-f001" class="html-fig">Figure 1</a>.</p>
Full article ">Figure 3
<p>Effect of 24 h pre-incubation of neuroglial cortical cultures with 5, 20, or 50 μg/mL exogenous SELENOM on OGD/R-induced cell death. Double staining of cells with Hoechst 33342 (HO342) and propidium iodide (PI). OGD/R—induction of ischemia-like conditions (2 h) and reoxygenation for 24 h without pre-incubation with SELENOM. SELENOM 5 µg + OGD/R, SELENOM 20 µg + OGD/R, and SELENOM 50 µg + OGD/R—24 h pre-incubation of cells with SELENOM followed by creation of OGD/R.</p>
Full article ">Figure 4
<p>Effect of 24 h pre-incubation of cortical cells with various concentrations of exogenous SELENOM on OGD/R-induced cell death. (<b>A</b>) Cytogram demonstrating the viability of cortical cells after 24 h of exposure to OGD/R depending on the concentration of SELENOM. <span class="html-italic">X</span>-axis—PI fluorescence intensity. The <span class="html-italic">Y</span>-axis is the fluorescence intensity of Hoechst 33342. Cells were stained with probes 24 h after OGD/R. (<b>B</b>) Effect of 24 h pre-incubation with 5, 20, or 50 μg/mL SELENOM on the induction of necrosis and apoptosis in OGD/R. Designations: OGD/R—induction of ischemia-like conditions (2 h of oxygen-glucose deprivation) and reoxygenation for 24 h without prior incubation with SELENOM. SELENOM 5 µg + OGD/R, SELENOM 20 µg + OGD/R, and SELENOM 50 µg + OGD/R—24 h pre-incubation of cells with SELENOM followed by creation of OGD/R. Results are presented as mean ± SEM. Black asterisks indicate the differences between the experimental groups comparable to the OGD group. Differences between experimental groups are marked with asterisks of different colors. n/s—data not significant (<span class="html-italic">p</span> &gt; 0.05), *** <span class="html-italic">p</span> &lt; 0.001, ** <span class="html-italic">p</span> &lt; 0.01, and * <span class="html-italic">p</span> &lt; 0.05. Cell images are presented in <a href="#biomedicines-12-01756-f003" class="html-fig">Figure 3</a>.</p>
Full article ">Figure 5
<p>Effect of 24 hours’ pre-incubation of neuroglial cortical cultures with 5, 20, or 50 μg/mL exogenous SELENOM on OGD/R-induced cell death. Double staining of cells with Hoechst 33342 (HO342) and propidium iodide (PI). OGD/R—induction of ischemia-like conditions (2 h) and reoxygenation for 24 h without pre-incubation with SELENOM. SELENOM 5 µg + OGD/R, SELENOM 20 µg + OGD/R, and SELENOM 50 µg + OGD/R—24 h pre-incubation of cells with SELENOM followed by creation of OGD/R. Effect of 24 h pre-incubation of cortical neuroglial cultures with 50 μg/mL exogenous SELENOM on basal (<b>A</b>), GluTox-induced (<b>B</b>), and OGD/R-induced (<b>C</b>) expression of genes encoding proteins involved in redox status regulation. The dotted line indicates protein expression in cortical cells treated with solvent buffer for panel (<b>A</b>), in cells after 24 h of GluTox exposure without pre-incubation with SELENOM for panel (<b>B</b>), and ischemia-like conditions (OGD/R) without pre-incubation with SELENOM for panel (<b>C</b>). Results are presented as mean ± SEM. Black asterisks indicate the differences between the experimental groups comparable to the Buffer-treated group (<b>A</b>), GluTox-treated group (<b>B</b>), and OGD/R-treated group (<b>C</b>) without pre-incubation with SELENOM. n/s—data not significant (<span class="html-italic">p</span> &gt; 0.05), *** <span class="html-italic">p</span> &lt; 0.001, * <span class="html-italic">p</span> &lt; 0.05.</p>
Full article ">Figure 6
<p>Effect of 24 h pre-incubation of cortical neuroglial cultures with 50 μg/mL exogenous SELENOM on basal (<b>A</b>), GluTox-induced (<b>B</b>), and OGD/R-induced (<b>C</b>) expression of genes encoding proteins involved in cell death. The dotted line indicates protein expression in cortical cells treated with solvent buffer for panel (<b>A</b>), in cells after 24 h of GluTox exposure without pre-incubation with SELENOM for panel (<b>B</b>), and ischemia-like conditions (OGD/R) without pre-incubation with SELENOM for panel (<b>C</b>). Results are presented as mean ± SEM. Black asterisks indicate the differences between the experimental groups comparable to the Buffer-treated group (<b>A</b>), GluTox-treated group (<b>B</b>), and OGD/R-treated group (<b>C</b>) without pre-incubation with SELENOM. n/s—data not significant (<span class="html-italic">p</span> &gt; 0.05), *** <span class="html-italic">p</span> &lt; 0.001, ** <span class="html-italic">p</span> &lt; 0.01, and * <span class="html-italic">p</span> &lt; 0.05.</p>
Full article ">Figure 7
<p>Dose-dependent effect of acute (<b>A</b>,<b>C</b>,<b>D</b>) recombinant SELENOM action and 24 h pre-incubation (<b>B</b>) of cortical astrocytes with different concentrations of SELENOM on ROS production. (<b>A</b>) ROS production in cortical astrocytes on the application of various SELENOM concentrations in an acute experiment after 3 h of recording DCF fluorescence. (<b>B</b>) ROS production in cortical astrocytes after 24 h pre-incubation with various SELENOM concentrations. Data obtained with an automated multiplate reader (Spark™ 10M multimode microplate reader) are presented. Data are shown as the mean of fluorescence intensity, arb.units ± S.E.M. Statistical analysis of experimental groups versus Control (dashed line) was performed with a paired <span class="html-italic">t</span>-test. *** <span class="html-italic">p</span> &lt; 0.001, and n/s—insignificant differences (<span class="html-italic">p</span> &gt; 0.05). (<b>C</b>,<b>D</b>) Predominantly cytosolic ROS production (<b>C</b>) and mitochondrial ROS production (<b>D</b>) in cortical astrocytes in response to different concentrations of SELENOM and 100 μM H<sub>2</sub>O<sub>2</sub>. The curves of ROS production averaged over several dozens of cells are presented.</p>
Full article ">Figure 8
<p>The effect of 24 h pre-incubation of astrocytes from a pure cortical culture with 50 μg/mL recombinant SELENOM on the level of gene expression encoding redox status proteins. The dotted line indicates expression in cortical astrocytes treated with protein solvent buffer. The results are presented as mean ± SEM. Black asterisks indicate the differences between the experimental groups comparable to the Buffer-treated group. n/s—data not significant (<span class="html-italic">p</span> &gt; 0.05), *** <span class="html-italic">p</span> &lt; 0.001, ** <span class="html-italic">p</span> &lt; 0.01.</p>
Full article ">Figure 9
<p>Effect of exogenous SELENOM on Ca<sup>2+</sup> signals of neurons (<b>A</b>) and astrocytes (<b>B</b>) and their death in a mixed cortical neuroglial culture when exposed to glutamate excitotoxicity (GluTox) for ~40 min. (<b>A</b>,<b>B</b>) Ca<sup>2+</sup> signals of neurons (<b>A</b>) and astrocytes (<b>B</b>) under GluTox, depending on pre-incubation for 24 h with 50 µg/mL SELENOM. The Ca<sup>2+</sup> signals of neurons and astrocytes averaged over several dozen cells in one experiment are presented. Neurons were distinguished from astrocytes using a short-term (30 s) application of 35 mM KCl, to which only neurons responded by generating Ca<sup>2+</sup> signals. Conversely, astrocytes responded with Ca<sup>2+</sup> signals to the addition of 10 µM ATP. (<b>C</b>,<b>D</b>) Propidium iodide (PI) staining of cell cultures after ~40 min of GluTox without pre-incubation with SELENOM (<b>C</b>) and after 24 h of pre-incubation with SELENOM (50 µg/mL). The appearance of PI fluorescence (red cell nuclei) reflects necrotic cell death.</p>
Full article ">Figure 10
<p>Effect of exogenous SELENOM on Ca<sup>2+</sup> signals of neurons (<b>A</b>) and astrocytes (<b>B</b>) and their death after ischemia-like conditions (OGD, oxygen-glucose deprivation) for ~40 min. (<b>A</b>,<b>B</b>) Ca<sup>2+</sup> signals of neurons (<b>A</b>) and astrocytes (<b>B</b>) under the OGD, depending on pre-incubation for 24 h with 50 µg/mL SELENOM. The Ca<sup>2+</sup> signals of neurons and astrocytes averaged over several dozen cells in one experiment are presented. Neurons were distinguished from astrocytes using a short-term (30 s) application of 35 mM KCl, to which only neurons responded by generating Ca<sup>2+</sup> signals. Conversely, astrocytes responded with Ca<sup>2+</sup> signals to the application of 10 µM ATP. (<b>C</b>,<b>D</b>) Propidium iodide (PI) staining of cell cultures after ~40 min OGD without pre-incubation with SELENOM (<b>C</b>) and after 24 h pre-incubation with SELENOM (50 µg/mL). The appearance of PI fluorescence (red cell nuclei) reflects necrotic cell death.</p>
Full article ">Figure 11
<p>Putative signaling pathways of recombinant SELENOM-induced [Ca<sup>2+</sup>]<sub>i</sub> increase in cortical neurons and astrocytes. (<b>A</b>) Application of 100 μL solvent buffer for SELENOM did not cause Ca<sup>2+</sup> signal generation in neurons (black curve) and astrocytes (red curve). (<b>B</b>,<b>C</b>) Application of various exogenous SELENOM concentrations to neurons (<b>B</b>) and astrocytes (<b>C</b>) in a cortical neuroglial culture. (<b>D</b>) Application of 50 μg exogenous SELENOM to a cortical neuroglial culture after 1 h pre-incubation with 10 μM of the actin-dependent endocytosis blocker Cytochalasin D (Cyto D). (<b>E</b>) Application of 50 μg exogenous SELENOM to a neuroglial culture in a nominally calcium-free medium (Ca<sup>2+</sup>-free) with the addition of 0.5 mM of the calcium chelator EGTA. (<b>F</b>) Application of 50 μg exogenous SELENOM to a neuroglial culture after depletion of the Ca<sup>2+</sup> store of the endoplasmic reticulum using the SERCA blocker thapsigargin (TG, 10 μM).</p>
Full article ">Figure 12
<p>Western blot analysis of cortical cell lysate samples after incubation with 50 μg of exogenous SELENOM or buffer. (<b>A</b>) Immunoblotting results obtained using antibodies against hexahistidine-labeled SELENOM at 30 min, 2 h, and 24 h after application of SELENOM to the culture medium. (<b>B</b>) Quantification of the studied proteins in the samples obtained using ImageJ software, presented as mean ± standard deviation of three independent experiments. GAPDH was used as a control for normalization. The expression level in the control (without treatment) was taken as 1 (dotted line). Statistical comparisons were made relative to the control group using the <span class="html-italic">t</span>-test. *** <span class="html-italic">p</span> &lt; 0.001.</p>
Full article ">Figure 13
<p>Immunocytochemical staining of cortical cells after pre-incubation with 50 μg/mL recombinant SELENOM for 30 min and 2 h. (<b>A</b>) Images of astrocytes (GFAP<sup>+</sup> cells) and non-astrocytes (GFAP<sup>–</sup> cells) stained with antibodies against histidine (His), reflecting the presence of recombinant SELENOM in the cells. HO342—cell nuclei stained with Hoechst 33342. GFAP + His—merged images of GFAP<sup>+</sup> cells and histidine<sup>+</sup> (His) cells, reflecting the presence of recombinant SELENOM in cortical astrocytes. (<b>B</b>) Intensity levels of histidine were determined by confocal imaging. We analyzed individual cells that had fluorescence of secondary antibodies. The quantitative data reflecting the level of histidine expression in GFAP<sup>+</sup>-cells and GFAP<sup>—</sup>-cells are presented as fluorescence intensity values in summary bar charts (mean ± SEM). The values were averaged by 100 cells for each column. The results obtained after immunostaining agree with the data of fluorescence presented in (<b>A</b>). Statistical significance was assessed using a paired <span class="html-italic">t</span>-test. Comparison with Buffer is marked by black asterisk. Comparisons between experimental groups are marked in red. n/s—data not significant (<span class="html-italic">p</span> &gt; 0.05), *** <span class="html-italic">p</span> &lt; 0.001.</p>
Full article ">
21 pages, 6595 KiB  
Article
Proteomics Analysis of the Protective Effect of Polydeoxyribonucleotide Extracted from Sea Cucumber (Apostichopus japonicus) Sperm in a Hydrogen Peroxide-Induced RAW264.7 Cell Injury Model
by Zhiqiang Shu, Yizhi Ji, Fang Liu, Yuexin Jing, Chunna Jiao, Yue Li, Yunping Zhao, Gongming Wang and Jian Zhang
Mar. Drugs 2024, 22(7), 325; https://doi.org/10.3390/md22070325 - 21 Jul 2024
Cited by 1 | Viewed by 1356
Abstract
Sea cucumber viscera contain various naturally occurring active substances, but they are often underutilized during sea cucumber processing. Polydeoxyribonucleotide (PDRN) is an adenosine A2A receptor agonist that activates the A2A receptor to produce various biological effects. Currently, most studies on the [...] Read more.
Sea cucumber viscera contain various naturally occurring active substances, but they are often underutilized during sea cucumber processing. Polydeoxyribonucleotide (PDRN) is an adenosine A2A receptor agonist that activates the A2A receptor to produce various biological effects. Currently, most studies on the activity of PDRN have focused on its anti-inflammatory, anti-apoptotic, and tissue repair properties, yet relatively few studies have investigated its antioxidant activity. In this study, we reported for the first time that PDRN was extracted from the sperm of Apostichopus japonicus (AJS-PDRN), and we evaluated its antioxidant activity using 2,2-diphenyl-1-picrylhydrazyl (DPPH), 2,2′-azino-bis-3-ethylbenzothiazoline-6-sulphonic acid (ABTS), and hydroxyl radical scavenging assays. An in vitro injury model was established using H2O2-induced oxidative damage in RAW264.7 cells, and we investigated the protective effect of AJS-PDRN on these cells. Additionally, we explored the potential mechanism by which AJS-PDRN protects RAW264.7 cells from damage using iTRAQ proteomics analysis. The results showed that AJS-PDRN possessed excellent antioxidant activity and could significantly scavenge DPPH, ABTS, and hydroxyl radicals. In vitro antioxidant assays demonstrated that AJS-PDRN was cytoprotective and significantly enhanced the antioxidant capacity of RAW264.7 cells. The results of GO enrichment and KEGG pathway analysis indicate that the protective effects of AJS-PDRN pretreatment on RAW264.7 cells are primarily achieved through the regulation of immune and inflammatory responses, modulation of the extracellular matrix and signal transduction pathways, promotion of membrane repair, and enhancement of cellular antioxidant capacity. The results of a protein–protein interaction (PPI) network analysis indicate that AJS-PDRN reduces cellular oxidative damage by upregulating the expression of intracellular selenoprotein family members. In summary, our findings reveal that AJS-PDRN mitigates H2O2-induced oxidative damage through multiple pathways, underscoring its significant potential in the prevention and treatment of diseases caused by oxidative stress. Full article
(This article belongs to the Special Issue Marine Anti-Inflammatory and Antioxidant Agents, 4th Edition)
Show Figures

Figure 1

Figure 1
<p>AJS-PDRN scavenging assays for DPPH, ABTS, and hydroxyl radicals. (<b>A</b>) AJS-PDRN scavenging assay of DPPH radicals; (<b>B</b>) AJS-PDRN scavenging assay of ABTS radicals; and (<b>C</b>) AJS-PDRN scavenging assay of hydroxyl radicals. <sup>a,b</sup> IC<sub>50</sub> data with different alphabets show significantly different values (<span class="html-italic">p</span> &lt; 0.05).</p>
Full article ">Figure 2
<p>Effects of AJS-PDRN on RAW264.7 cell viability with/without H<sub>2</sub>O<sub>2</sub> induction. (<b>A</b>) Cells treated with the indicated concentrations of H<sub>2</sub>O<sub>2</sub> for 4 h; (<b>B</b>) cells treated with the indicated concentrations of AJS-PDRN for 6 h; and (<b>C</b>) cells pretreated with the indicated concentrations of AJS-PDRN for 6 h and then stimulated with 400 μM H<sub>2</sub>O<sub>2</sub> for 4 h. The results are presented as the mean ± SD of three replicates. Different letters at the top of the columns indicate statistically significant differences.</p>
Full article ">Figure 3
<p>Effects of AJS-PDRN on the levels of oxidative stress biomarkers in H<sub>2</sub>O<sub>2</sub>-induced RAW264.7 cells. (<b>A</b>) SOD activity; (<b>B</b>) CAT activity; (<b>C</b>) GSH content; and (<b>D</b>) MDA content. The results are presented as the mean ± SD of three replicates. Different letters at the top of the columns indicate statistically significant differences.</p>
Full article ">Figure 4
<p>Effect of AJS-PDRN on H<sub>2</sub>O<sub>2</sub>-induced protein expression profile in RAW264.7 cells. (<b>A</b>) PCA based on quantitative data of the selected proteins in each group (<span class="html-italic">n</span> = 3 for each group); (<b>B</b>) Venn diagram of DEPs in the H<sub>2</sub>O<sub>2</sub> group and the AJS-PDRN group; and (<b>C</b>) Venn diagram demonstrating the number of DEPs identified in each comparison.</p>
Full article ">Figure 5
<p>GO enrichment analysis of DEPs. (<b>A</b>) The H<sub>2</sub>O<sub>2</sub> group vs. the control group; (<b>B</b>) the AJS-PDRN group vs. the H<sub>2</sub>O<sub>2</sub> group. The number after each term represents the number of differential proteins annotated to that term.</p>
Full article ">Figure 6
<p>The bubble diagram of KEGG pathway analysis of DEPs. (<b>A</b>) The H<sub>2</sub>O<sub>2</sub> group vs. the control group; (<b>B</b>) the AJS-PDRN group vs. the H<sub>2</sub>O<sub>2</sub> group. The larger the bubble the greater the number of differential proteins contained in the entry. The color of the bubbles changes from blue to red, indicating that the smaller the enrichment <span class="html-italic">p</span>-value, the greater the degree of significance.</p>
Full article ">Figure 7
<p>PPI network analysis of DEPs. (<b>A</b>) PPI network diagram of all DEPs with interactions in the H<sub>2</sub>O<sub>2</sub> group vs. the control group. (<b>B</b>) The highest-scoring sub-network in the H<sub>2</sub>O<sub>2</sub> group was selected using the MCODE algorithm. (<b>C</b>) The major GO BP terms were significantly enriched by 11 proteins in the sub-network, as well as the related DEPs. (<b>D</b>) PPI network diagram of all DEPs with interactions in the AJS-PDRN group vs. the H<sub>2</sub>O<sub>2</sub> group. (<b>E</b>) The highest-scoring sub-network in the AJS-PDRN group was selected using the MCODE algorithm. (<b>F</b>) The major GO BP terms and KEGG pathway were significantly enriched by 9 proteins in the sub-network, as well as the related DEPs.</p>
Full article ">
14 pages, 1549 KiB  
Article
Selenium Discrepancies in Fetal Bovine Serum: Impact on Cellular Selenoprotein Expression
by François Parant, Fabrice Mure, Julien Maurin, Léana Beauvilliers, Chaïma Chorfa, Chaymae El Jamali, Théophile Ohlmann and Laurent Chavatte
Int. J. Mol. Sci. 2024, 25(13), 7261; https://doi.org/10.3390/ijms25137261 - 1 Jul 2024
Cited by 1 | Viewed by 1185
Abstract
Selenium is an essential trace element in our diet, crucial for the composition of human selenoproteins, which include 25 genes such as glutathione peroxidases and thioredoxin reductases. The regulation of the selenoproteome primarily hinges on the bioavailability of selenium, either from dietary sources [...] Read more.
Selenium is an essential trace element in our diet, crucial for the composition of human selenoproteins, which include 25 genes such as glutathione peroxidases and thioredoxin reductases. The regulation of the selenoproteome primarily hinges on the bioavailability of selenium, either from dietary sources or cell culture media. This selenium-dependent control follows a specific hierarchy, with “housekeeping” selenoproteins maintaining constant expression while “stress-regulated” counterparts respond to selenium level fluctuations. This study investigates the variability in fetal bovine serum (FBS) selenium concentrations among commercial batches and its effects on the expression of specific stress-related cellular selenoproteins. Despite the limitations of our study, which exclusively used HEK293 cells and focused on a subset of selenoproteins, our findings highlight the substantial impact of serum selenium levels on selenoprotein expression, particularly for GPX1 and GPX4. The luciferase reporter assay emerged as a sensitive and precise method for evaluating selenium levels in cell culture environments. While not exhaustive, this analysis provides valuable insights into selenium-mediated selenoprotein regulation, emphasizing the importance of serum composition in cellular responses and offering guidance for researchers in the selenoprotein field. Full article
(This article belongs to the Special Issue Molecular Research of Selenocysteine in Selenoproteins)
Show Figures

Figure 1

Figure 1
<p>Elemental- and selenium-related composition of the serum as documented in the literature. (<b>a</b>) Concentration levels of toxic, neutral, and essential elements measured in the serum of lactating cows, detectable by multielement ICP-MS, adapted from [<a href="#B36-ijms-25-07261" class="html-bibr">36</a>]. The data are depicted as mean values ± standard error, utilizing a logarithmic scale, obtained from a random selection of 20 cows out of the 188 present on the farm situated in Northern Spain [<a href="#B36-ijms-25-07261" class="html-bibr">36</a>]. Essential elements are depicted in green bars, while the remaining elements are shown in yellow. Phosphorus (P), calcium (Ca), magnesium (Mg), iron (Fe), zinc (Zn), copper (Cu), boron (B), molybdenum (Mo), strontium (Sr), selenium (Se), lithium (Li), barium (Ba), chromium (Cr), arsenic (As), manganese (Mn), antimony (Sb), nickel (Ni), lead (Pb), cobalt (Co), mercury (Hg), and cadmium (Cd). (<b>b</b>) Pie charts depict selenium distribution among serum proteins in various contexts, with total selenium concentration indicated above each chart in blue. The various studies concerned human-certified reference materials [<a href="#B43-ijms-25-07261" class="html-bibr">43</a>], healthy human adults [<a href="#B41-ijms-25-07261" class="html-bibr">41</a>], human serum at birth [<a href="#B44-ijms-25-07261" class="html-bibr">44</a>], and young calves under different selenium diets [<a href="#B42-ijms-25-07261" class="html-bibr">42</a>]. Selenium distribution was assessed using ICP-MS coupled with various separation methods. Total selenium concentration is denoted above each chart in blue, and the percentage of each part is indicated in white on the pie chart.</p>
Full article ">Figure 2
<p>Variation in selenium concentration across the 12 different FBS samples analyzed in this study and listed in <a href="#ijms-25-07261-t001" class="html-table">Table 1</a>. TQ ICP MS was used in mass shift mode using O<sub>2</sub> as a reaction gas to determine the levels of selenium. Each sample was analyzed in triplicate and represented as means ± S.D. The difference between the lowest and highest values is indicated by a blue bracket, with the corresponding fold-change factor beside it.</p>
Full article ">Figure 3
<p>Variation of intracellular selenoprotein expression levels depending on the serum used, visualized via Western blot analysis. HEK293 cells were cultured in the presence of each FBS variant listed in <a href="#ijms-25-07261-t001" class="html-table">Table 1</a> and harvested after three days of growth. Cellular extracts were subsequently separated on protein gels and subjected to Western blot analysis to determine the levels of GPX1 (<b>a</b>), GPX4 (<b>b</b>), and TXNRD1 (<b>c</b>). (<b>d</b>) One gel was stained with InstantBlue Coomassie staining solution and used for signal intensity normalization. For GPX1 and GPX4, an additional, more exposed gel is also presented to confirm the presence of signal at a lower level. The arrows indicate the migration of the selenoproteins. Non-specific bands are indicated by stars. The relative protein level intensity is indicated as a percentage of the strongest signal across the entire blot at the bottom of each image. On the right side of each blot, a graph depicting the Western blot signal intensity as a function of serum selenium concentration is displayed. A linear regression line is shown along with its Pearson linear determination coefficient (R<sup>2</sup>). For each graph, the differences between the lowest and highest values in the Western blot signal are indicated by a blue bracket, with the corresponding fold-change factor beside it.</p>
Full article ">Figure 4
<p>Evolution of intracellular GPX and TXNRD activity as a function of the FBS used. Cellular extracts from HEK293 cells cultured in the presence of each FBS variant were assessed for their GPX (<b>a</b>) and TXNRD (<b>b</b>) activities. The enzymatic activities are presented in histograms as the means of triplicates ± standard deviation. On the right side of each histogram, a graph illustrating the enzymatic activities in relation to serum selenium concentration is provided. The differences between the lowest and highest values of enzymatic activities are delineated by a blue bracket, accompanied by the corresponding fold-change factor.</p>
Full article ">Figure 5
<p>Use of stable HEK cell lines expressing luciferase reporters to assess the selenium content of FBS. (<b>a</b>) Schematic representation of the UGA-SECIS luciferase reporters. This system relies on the expression of a luciferase gene containing an in-frame UGA codon at position 258, downstream of which a minimal SECIS element is inserted. Two variants of this construct, featuring either GPX1 or GPX4 SECIS, were employed to establish stable cell lines [<a href="#B25-ijms-25-07261" class="html-bibr">25</a>,<a href="#B49-ijms-25-07261" class="html-bibr">49</a>]. Active luciferase enzyme is produced only when the UGA codon is recoded as selenocysteine and not when it is read as a stop codon. Cellular extracts from HEK293 cell lines expressing either Luc UGA—GPX1 SECIS (<b>b</b>) or Luc UGA—GPX4 SECIS (<b>c</b>) were cultured in the presence of each FBS variant and evaluated for luciferase activity. The luciferase enzymatic activities were normalized to protein concentration and then expressed relative to the strongest signal obtained with FBS#11, set as 100%. Data are presented in histograms as means of triplicates ± standard deviation. On the right side of each histogram, a graph illustrating the enzymatic activities in relation to serum selenium concentration is provided. The differences between the lowest and highest values of luciferase activities are depicted by a blue bracket, accompanied by the corresponding fold-change factor.</p>
Full article ">
Back to TopTop