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CD3 + T cell proliferation monitored using CFSE labeling. CFSE labeled splenocytes (2610 6 cells/well) were treated with or without 2 mmol/L of sodium selenite in the absence or presence of anti-CD3, ConA or PHA, and proliferation was analyzed at 48 h of culture. Cells were stained with anti-CD3-PEcy5. CD3 + T cells from the entire well were analyzed for proliferation by flow cytometry. The percentage of proliferating cells for each culture is indicated. A representative experiment from two separate experiments is shown. doi:10.1371/journal.pone.0035375.g002
Source publication
There is controversy in the literature over whether the selenium (Se) influences cellular immune responses, and the mechanisms possibly underlying these effects are unclear. In this study, the effects of Se on T-cell proliferation and IL-2 production were studied in primary porcine splenocytes. Splenocytes were treated with different mitogens in th...
Contexts in source publication
Context 1
... pattern to that of water soluble tetrazolium salt-8 (WST-8, containing [2-(2-methoxy-4-nitrophenyl)-3-(4-nitrophenyl) 5-(2,4- disulfophenyl)-2H-tetrazolium, monosodium salt]). Total CD3 + T- cells proliferated more in the selenium treated group than in the control group in TCR or ConA stimulated splenocytes (53% versus 69%; 51% versus 65%, Fig. 2), but the percentage of proliferating cells did not differ in the unstimulated and PHA- stimulated groups (2.4% versus 2.5%; 53% versus 52%, Fig. 2). . Cells without any stimulus were used as control (CON). GPx1 mRNA was measured by quantitative real-time RT-PCR, and the ratio of the level of GPx1 mRNA to that of the b-actin internal ...
Context 2
... monosodium salt]). Total CD3 + T- cells proliferated more in the selenium treated group than in the control group in TCR or ConA stimulated splenocytes (53% versus 69%; 51% versus 65%, Fig. 2), but the percentage of proliferating cells did not differ in the unstimulated and PHA- stimulated groups (2.4% versus 2.5%; 53% versus 52%, Fig. 2). . Cells without any stimulus were used as control (CON). GPx1 mRNA was measured by quantitative real-time RT-PCR, and the ratio of the level of GPx1 mRNA to that of the b-actin internal control was used for statistical comparison. Data represent mean 6 S.E. of two independent experiments, each measured in triplicate. Mean values ...
Citations
... CD3/CD28 antibodies, PMA/Ionomycin, and Concanavalin A were used to maintain the cell viability and facilitate the differentiation and proliferation of the cells as previously described (Wang and Ai 2021;Wang et al. 2019). CD3/CD28 antibodies activate the TCR signaling pathway along with the co-stimulatory signals, and PMA/Ionomycin and Concanavalin A are known to amplify those cellular responses (Li and Kurlander 2010;Crawford et al. 2014;Ren et al. 2012;Oh-hora 2009;Boilard and Surette 2001). For Th1 differentiation, 10 ng/ ml recombinant murine (rm) IL-12p70 (BioLegend) was added instead of retinoic acid, TGFβ, and IFNγ. ...
Background
In sepsis, intestinal barrier dysfunction is often caused by the uncontrolled death of intestinal epithelial cells (IECs). CD4CD8αα intraepithelial lymphocytes (IELs), a subtype of CD4 ⁺ T cells residing within the intestinal epithelium, exert cytotoxicity by producing granzyme B (GrB) and perforin (Prf). Extracellular cold-inducible RNA-binding protein (eCIRP) is a recently identified alarmin which stimulates TLR4 on immune cells to induce proinflammatory responses. Here, we hypothesized that eCIRP enhances CD4CD8αα IEL cytotoxicity and induces IEC death in sepsis.
Methods
We subjected wild-type (WT) and CIRP −/− mice to sepsis by cecal ligation and puncture (CLP) and collected the small intestines to isolate IELs. The expression of GrB and Prf in CD4CD8αα IELs was assessed by flow cytometry. IELs isolated from WT and TLR4 −/− mice were challenged with recombinant mouse CIRP (eCIRP) and assessed the expression of GrB and Prf in CD4CD8αα by flow cytometry. Organoid-derived IECs were co-cultured with eCIRP-treated CD4CD8αα cells in the presence/absence of GrB and Prf inhibitors and assessed IEC death by flow cytometry.
Results
We found a significant increase in the expression of GrB and Prf in CD4CD8αα IELs of septic mice compared to sham mice. We found that GrB and Prf levels in CD4CD8αα IELs were increased in the small intestines of WT septic mice, while CD4CD8αα IELs of CIRP −/− mice did not show an increase in those cytotoxic granules after sepsis. We found that eCIRP upregulated GrB and Prf in CD4CD8αα IELs isolated from WT mice but not from TLR4 −/− mice. Furthermore, we also revealed that eCIRP-treated CD4CD8αα cells induced organoid-derived IEC death, which was mitigated by GrB and Prf inhibitors. Finally, histological analysis of septic mice revealed that CIRP −/− mice were protected from tissue injury and cell death in the small intestines compared to WT mice.
Conclusion
In sepsis, the cytotoxicity initiated by the eCIRP/TLR4 axis in CD4CD8αα IELs is associated with intestinal epithelial cell (IEC) death, which could lead to gut injury.
... To characterize the influence of immunomodulatory substances on porcine PBMC cultures, we used PBMCs of six GMs for in-vitro studies and compared the observed effects to human PBMC cultures. We assessed the in-vitro stimulation capacity for porcine and human PBMCs using three different well-established T-cell stimuli: the Tcell mitogens concanavalin A (ConA) and phytohemagglutinin-L (PHA) and the superantigen staphylococcal enterotoxin B (SEB) (21)(22)(23). Furthermore, we tested the in-vitro effect of four immune suppressive drugs used in human medicine for the treatment of rheumatoid arthritis and rejection in transplant recipients (24)(25)(26). ...
Considering the similarities between swine and humans, it is a logical consequence to use swine as a translational model in research and drug development, including non-clinical safety. Here, we compared the reactivity of peripheral blood mononuclear cells (PBMCs) from humans and minipigs under the influence of different compounds in vitro. We conducted a flow cytometry-based proliferation assay that focused on the T-cell response to three different stimuli: concanavalin A (ConA), phytohemagglutinin-L (PHA-L), and staphylococcal Enterotoxin B (SEB). Furthermore, four approved immunosuppressive drugs—abatacept, belatacept, rapamycin, and tofacitinib—which are used for the treatment of rheumatoid arthritis or rejection in transplant recipients, were combined with the different stimuli. This allowed us to study the effect of suppressive drugs in comparison with the different stimuli in both species. We examined proliferating T cells (CD3⁺) and investigated the presence of TCR-αβ⁺ and TCR-γδ⁺ T cells. Differences in the response of T cells of the two species under these various conditions were evident. CD4⁺ T cells were more activated within humans, whereas CD8⁺ T cells were generally more abundant in swine. The effectiveness of the used humanized antibodies is most likely related to the conserved structure of CTLA-4 as abatacept induced a much stronger reduction in swine compared with belatacept. The reduction of proliferation of rapamycin and tofacitinib was highly dependent on the used stimuli. We further investigated the effect of the immunosuppressive compounds on antigen-specific restimulation of pigs immunized against porcine circovirus 2 (PCV2). Treatment with all four compounds resulted in a clear reduction of the proliferative response, with rapamycin showing the strongest effect. In conclusion, our findings indicate that the effectiveness of suppressive compounds is highly dependent on the stimuli used and must be carefully selected to ensure accurate results. The results highlight the importance of considering the response of T cells in different species when evaluating the potential of an immunomodulatory drug.
... The levels of CD4+ T-cell-related cytokines and transcription factors in the [26,38]mouse thyroid were determined by enzyme-linked immunosorbent assay. T-Bet is a transcription factor of Th1 cells, and TNF-α and IFN-γ are the cytokines produced by them. ...
... This can be explained by the fact that Se is an essential micronutrient for biological survival, thus the Se from the nanocomposite was absorbed by the normal cell and promoted cell proliferation. Studies show that Se plays a key role in terminal differentiation, cell growth and development by controlling the cell cycle (Ren et al., 2012;Hassan et al., 2014). Yan et al. found that Se can stimulate proliferation of normal chondrogenic cell ATDC5 by accelerating cell cycle progression (Yan et al., 2012). ...
... A physiologically relevant approach used beads coated with anti-CD3 and anti-CD28 to stimulate T cells in a manner that partially mimics antigenic stimulation [43]. In a related study on the primary porcine spleen, both PHA and Con A effectively stimulated T cell proliferation and IL-2 production [44]. Stimulating PBMCs with anti-CD3/ CD28 coated beads provides an alternative method for driving T cell expansion that may be very useful in immunotherapy [45]. ...
Background:
Sheep are an important livestock species worldwide and an essential large-animal model for animal husbandry and veterinary research. Understanding fundamental immune indicators, especially T-lymphocyte parameters, is necessary for research on sheep diseases and vaccines, to better understand the immune response to bacteria and viruses for reducing the use of antibiotics and improving the welfare of sheep. We randomly selected 36 sheep of similar ages to analyze cell-related immune indicators in peripheral blood mononuclear cells (PBMCs). The proportions of CD4+ and CD8+ T cells in PBMCs were detected by flow cytometry. We used Concanavalin A (Con A) and Phorbol-12-myristate-13-acetate (PMA)/Ionomycin to stimulate PBMCs, and measured the expression of IFN-γ, IL-4, and IL-17A using enzyme-linked immunosorbent assay (ELISA) and enzyme-linked immunospot assay (ELISpot). Simultaneously, PMA/Ionomycin/brefeldin A (BFA) was added to PBMCs, then the expression of IFN-γ, IL-4, and IL-17A was detected by flow cytometry after 4 h of culturing. In addition, we observed the proliferation of PBMCs stimulated with Con A for 3, 4, and 5 days.
Results:
The proportions of CD4+ T lymphocytes (18.70 ± 4.21%) and CD8+ T lymphocytes (8.70 ± 3.65%) were generally consistent among individuals, with a CD4/CD8 ratio of 2.40 ± 0.79. PBMCs produced high levels of IFN-γ, IL-4, and IL-17A after stimulation with PMA/Ionomycin and Con A. Furthermore, PMA/Ionomycin stimulation of PBMC yielded significantly higher cytokine levels than Con A stimulation. Flow cytometry showed that the level of IFN-γ (51.49 ± 11.54%) in CD8+ T lymphocytes was significantly (p < 0.001) higher than that in CD4+ T lymphocytes (14.29 ± 3.26%); IL-4 (16.13 ± 6.81%) in CD4+ T lymphocytes was significantly (p < 0.001) higher than that in CD8+ T lymphocytes (1.84 ± 1.33%), There was no difference in IL-17A between CD4+ (2.83 ± 0.98%) and CD8+ T lymphocytes (1.34 ± 0.67%). The proliferation of total lymphocytes, CD4+ T lymphocytes, and CD8+ T lymphocytes continued to increase between days 3 and 5; however, there were no significant differences in proliferation between the cell types during the stimulation period.
Conclusions:
Evaluating primary sheep immune indicators, especially T lymphocytes, is significant for studying cellular immunity. This study provided valuable data and theoretical support for assessing the immune response of sheep to pathogens and improving sheep welfare.
... Selenoproteins play essential roles in biological functions such as antioxidant defense system, reproduction, DNA synthesis, fertility, formation of thyroid hormones, and cancer prevention (Mehdi et al. 2013). Selenoproteins (i.e., GPX1 and TR1) and Se metabolites are involved in inflammatory and immune responses (Ren et al. 2012). Also, Se plays an important role in cellular activities and biosynthesis of several selenoproteins such as thioredoxin reductase (TrxR), glutathione peroxidase, and deiodinases which are known to participate in redox homeostasis maintenance and prevention of oxidative stress-induced apoptosis (Labunskyy et al. 2014). ...
Excessive exposure to manganese (Mn) is associated with neurotoxicity characterized by oxidative stress, inflammation, and apoptosis induction. Selenium (Se) has been shown to possess antioxidant, anti-inflammatory, and anti-apoptotic properties in humans and animals. The present study investigated the neuroprotective mechanism of Se in rats sub-chronically treated with Mn at 30 mg/kg body weight or orally co-treated with Se at 0.2 and 0.4 mg/kg body weight for 35 consecutive days. Locomotive and exploratory profiles were recorded and computed with the aid of ANY-Maze (a video-tracking software) for 5-min trial, in a novel apparatus. The ANY-Maze analysis showed that Se significantly (p < 0.05) abated Mn-induced locomotive impairment evidenced by increased in maximum speed, total time traveled, absolute turn angle, number of line crossing, rotation and forelimb grip and decreased total time immobile, grooming, and negative geotaxis as verified by the enhanced track plot density. Furthermore, the striatum and hippocampus of the rats were excised and the levels of Mn and Se, oxidative stress markers, proinflammatory cytokines including acetylcholinesterase and caspase-3 activities were assayed. The result shows that Se abates Mn-mediated accumulation of Mn. Also, Se ameliorated Mn-induced decrease in antioxidant enzymes as well as glutathione level and increase in acetylcholinesterase activity, lipid peroxidation, proinflammatory cytokines (i.e., interleukin (IL)-6, IL-1β, tumor necrosis factor alpha), and caspase-3 activation in the striatum and hippocampus of the rats. Collectively, Se abated Mn-induced striatal and hippocampal toxicity via abrogation of neurobehavioral deficits, biometal accumulation, oxidative stress, inflammation, and caspase-3 activation in rats. Se may serve as a neuroprotective agent against Mn-mediated neurotoxicity.
... IL-2 is mainly produced by type 1 T helper (Th1) cells and plays a significant role in macrophage activation and in promoting a cell-mediated immune response (32). Previous studies have reported consistent results, that Se promotes glutathione peroxidase (GPx1) and thioredoxin reductase 1 (TR1) expression, and enhances ConA induced T-cell activation and secretion of IL-2 in porcine splenocytes (33). The ability of Se to induce augmented expression of IL-2 appears to occur through the increased expression of the IL-2 receptor (34). ...
The purpose of this study was to investigate the effects of dietary Selenohomolanthionine (SeHLan) on antioxidant status and immune response in canine parvovirus (CPV) vaccinated puppies. In this study, 30 weaned puppies were randomly divided into six groups: control group (–Se/–Vacc), immunization group (–Se/+Vacc), supplementation of sodium selenite group (SS/+Vacc, 0.35 mg/kg DM), low-dose SeHLan group (SeHLan-L/+Vacc, 0.35 mg/kg DM), mid-dose SeHLan group (SeHLan-M/+Vacc, 1.0 mg/kg DM), and high-dose SeHLan group (SeHLan-H/+Vacc, 2.0 mg/kg DM). The puppies were fed for 42 days and vaccinated with Vanguard Plus 5 on day 0 and day 21. Blood samples were collected on 7, 14, 21, 28, 35, 42 days post-immunization (PI) for determination of antioxidant indicators, lymphocyte proliferation index, serum cytokine concentration (IL-2, IL-4), canine polymorphonuclear neutrophils (PMN) phagocytic function, and the level of CPV antibody titers. The results showed that SeHLan supplementation raised the serum Se concentration and glutathione peroxidase (GSH-Px) activity in a dose-dependent manner (P < 0.05). It also increased the activity of serum superoxide dismutase (SOD) and decreased serum malondialdehyde (MDA) content, especially in SeHLan-M/+Vacc group (1.0 mg/kg DM) (P < 0.01). SeHLan supplementation significantly increased lymphocyte proliferation, IL-2, and IL-4 levels in canine serum, and enhanced phagocytosis of PMN in vaccinated puppies (P < 0.05). Moreover, SeHLan supplementation shortened the CPV antibody production time and increased the CPV antibody titers (P < 0.05). Of note, the beneficial effects of SeHLan were superior to those of SS. In conclusion, dietary SeHLan supplementation improved antioxidant activity, increased CPV antibody titers, and enhanced immune function in puppies after weaning. An appropriate dosage of SeHLan (1~2 mg/kg DM) may confer nutritional benefits in puppies.
... The study revealed that Se increased the expression of T cell receptors (TCR), proliferation of T cells and the synthesis of IL-2 by a statistically significant margin. Additionally, Se increased the concentration of GPX1 and the mRNA of thioredoxin reductase, increased the activity of GPX1 and the concentration of reduced glutathione in splenocytes [51]. Another study conducted on dairy cows revealed that selenium could aid the immune system in mitigating oxidative stress and decrease the intensity of inflammatory disorders affecting the production efficiency of dairy cows such as mastitis or uterine inflammation [52]. ...
Introduction. Selenium (Se) is a trace element found mainly in meat, seafood, nuts and grains. Se is found in selenoproteins such as selenocystein or selenomethionin. A well balanced diet provides enough Se. Many regulatory and metabolic enzymes contain Se as their component, which is why Se supplementation is used in the treatment as well as prevention of multiple disorders. Se may, however, be toxic if overdosed.
Aim. The aim of this review is to summarize the data about functions of Se in human body and to discuss its use in treatment and prevention of diseases.
Materials and methods. The search was conducted using the PubMed and Google Scholar databases in March and April 2020. The key words used were: ‘selenium’, ‘cardiovascular disease’, ‘selenium supplementation’, ‘Keshan disease’, ‘source of selenium’. A total of 68 articles were analysed.
Results. The first cases of chronic Se deficiency cases were documented 85 years ago in China. The patients with cardiomyopathy, extensive fibrosis and degenerative changes in the heart were diagnosed with Keshan disease. Human selenoproteonome consists of at least 25 selenoproteins. Se plays a role in immunity and metabolism via its role in functioning of numerous enzymes: glutathione peroxidase, thioredoxine and methionine sulfoxide reductase, methionine-sulfoxide reductase B1. Se plays a role in glucose homeostasis, Alzheimer’s disease, thyroid disorders, infectious, inflammatory diseases, vascular diseases and fertility.
Conclusion. Se deficiency increases the risk of Keshan disease, but there is not enough evidence to recommend its supplementation for prevention of cardiovascular disease. However, Se status is important part of health assessment. Se supplementation should not exceed the dose of 55μg/day.
... Later, the thymic-derived lymphocytes were cultured in DMEM-H modified with 10% FBS containing 5 µg/mL of phytohemagglutinin (PHA) and 10 µg/mL of concanavalin A (ConA) for stimulation. 17,18 The thymic-derived lymphocytes incubated in 10% FBS DMEM-H were used as the control group. We observed the cell morphology of these two groups at 0 hours, 24 hours, and 72 hours. ...
Background:
Renal tubulointerstitial fibrosis is known to occur as a result of epithelial cell transformation into myofibroblasts via the epithelial-to-mesenchymal transition (EMT) process. It has been reported that macrophages, regulatory T (Treg) cells, and gamma delta T (γδ T) cells can promote fibrosis via EMT in vivo.
Objective:
Our study intended to detect whether thymocytes can induce renal tubular cells to undergo the EMT.
Methods:
Rat thymocytes were activated by phytohemagglutinin and concanavalin A. The rat renal tubular epithelial cells (NRK-52E) were incubated in a conditioned medium harvested from activated thymocytes or co-cultured with freshly isolated thymocytes for 48 hours. Real-time reverse transcription-polymerase chain reaction, immunofluorescence, and western blotting analysis were used to test the expression of the epithelial and mesenchymal markers in NRK-52E cells. Scratch assay was designed to test the cell migration abilities of NRK-52E cells. Student's t test and oneway analysis of variance test were used for statistical analysis.
Results:
The combined stimulation with phytohemagglutinin and concanavalin A activated the primary isolated rat thymocytes. After treatment with conditioned medium or freshly isolated thymocytes, the expression levels of cytokeratin 19 and E-cadherin were downregulated in NRK-52E cells, while the mRNA and protein expression levels of alpha-smooth muscle actin, desmin, and vimentin were upregulated (P < 0.05). We found that the cell migration abilities of the induced NRK-52E cells were significantly improved.
Conclusions:
Both activated rat thymocytes (more percentage of CD8+ T cells) and freshly isolated thymocytes have promoting effects on the EMT of NRK-52E cells.
... Likewise, selenium promotes human T-cell activation through T-cell receptors. Increased expression of GPX1 and TXNRD1 is involved in this effect [77]. While selenium enhances T cell response when needed, it prevents the uncontrolled activation of immune system in autoimmune diseases [78,79]. ...
Management of IBD (inflammatory bowel disease) has always been a challenge for physicians. Current treatment protocols may cause numerous adverse effects. Selenium is known for its putative anti-inflammatory properties. Selenium is needed for the biosynthesis of enzymatically active selenoproteins, which contribute to antioxidative defence, and effective function of immune systems. Several studies showed that IBD patients have a lower selenium level compared to healthy subjects. Hence, experimental studies mimicking ulcerative colitis and Crohn's disease investigated the effect of selenium supplementation on IBD.
Previous studies indicated that: i) Selenoproteins can curb the inflammatory response and attenuate oxidative stress. This anti-inflammatory property caused remission in animal models of colitis. ii) Selenium supports protective gut microbiota which indirectly improves management of IBD. iii) Selenium may block some of the predominant tumorigenesis pathways proposed in colitis-associated colorectal cancer. iv) Selenium supplementation showed promising results in preliminary clinical studies particularly in patients with selenium deficiency.
While selenium supplementation seems to be beneficial for IBD, clinical studies remained too preliminary in this regard. Randomized clinical trials are needed to measure the short-term and long-term effects of selenium on both active and quiescent IBD particularly in IBD patients whom have documented selenium deficiency.
