Environmentally Relevant Level of Aflatoxin B1 Dysregulates Human Dendritic Cells Through Signaling on Key Toll-Like Receptors

2014 ◽  
Vol 33 (3) ◽  
pp. 175-186 ◽  
Author(s):  
Azam Mohammadi ◽  
Jalil Mehrzad ◽  
Mahmoud Mahmoudi ◽  
Marion Schneider
Keyword(s):  
Dendritic Cells ◽  
Messenger Rna ◽  
Molecular Mechanisms ◽  
Quantitative Polymerase Chain Reaction ◽  
Immune Surveillance ◽  
Toll Like Receptors ◽  
Myeloid Dendritic Cells ◽  
Protein Levels ◽  
Tlr4 Mrna ◽  
Relevant Level

Aflatoxins (AFs) are highly hazardous fungal biometabolites usually present in feeds and foods. Aflatoxin B1 (AFB1) is the most toxic and a known carcinogen. Toll-like receptors (TLRs), highly expressed by myeloid dendritic cells (DC), are key innate immune-surveillance molecules. Toll-like receptors not only sense pathogen-associated molecular patterns but also contribute to infections and cancer. To assess AFB1–TLR interactions on human myeloid DC, pure CD11c+ DC were generated from monocytes isolated from healthy individuals and then exposed to relevant level of AFB1 for 2 hours. Both quantitative polymerase chain reaction and flow cytometric assays were used to quantify, respectively, expression of TLR2 and TLR4 at the messenger RNA (mRNA) and protein levels in these DC. Levels of interleukin (IL) 1β, IL-6, and IL-10 were also analyzed in AFB1- and mock-treated DC. Compared to nontreated CD11c+ DC, expression levels of both TLR2 and TLR4 mRNA and proteins were significantly upregulated in AFB1-treated cells. Further, although IL-10 levels in AFB1-treated DC were similar to those in the mock-treated DC, the AFB1-exposed DC secreted higher amounts of IL-1β and IL-6. Dendritic cells are sensitive to environmentally relevant level of AFB1, and TLR2 and TLR4 are involved in sensing AFB1. Considering the broad roles of TLR2, TLR4, and DC in immunity and infections, our novel findings open a new door to understanding the molecular mechanisms and functional consequences of AFB1 in inducing immunodysregulation, immunotoxicity, and thus (non)infectious diseases in humans.

scholarly journals PKM2 Is Required to Activate Myeloid Dendritic Cells from Patients with Severe Aplastic Anemia

2018 ◽  
Vol 2018 ◽  
pp. 1-9 ◽  
Author(s):  
Chunyan Liu ◽  
Mengying Zheng ◽  
Ting Wang ◽  
Huijuan Jiang ◽  
Rong Fu ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Aplastic Anemia ◽  
Immune Status ◽  
Bone Marrow Failure ◽  
Severe Aplastic Anemia ◽  
Myeloid Dendritic Cells ◽  
Protein Levels ◽  
Immune Mediated ◽  
Normal Controls

Severe aplastic anemia (SAA) is an autoimmune disease in which bone marrow failure is mediated by activated myeloid dendritic cells (mDCs) and T lymphocytes. Recent research has identified a strong immunomodulatory effect of pyruvate kinase M2 (PKM2) on dendritic cells in immune-mediated diseases. In this study, we aimed to explore the role of PKM2 in the activation of mDCs in SAA. We observed conspicuously higher levels of PKM2 in mDCs from SAA patients compared to normal controls at both the gene and protein levels. Concurrently, we unexpectedly discovered that after the mDC-specific downregulation of PKM2, mDCs from patients with SAA exhibited weakened phagocytic activity and significantly decreased and shortened dendrites relative to their counterparts from normal controls. The expression levels of the costimulatory molecules CD86 and CD80 were also reduced on mDCs. Our results also suggested that PKM2 knockdown in mDCs reduced the abilities of these cells to promote the activation of CD8+ T cells (CTLs), leading to the decreased secretion of cytotoxic factors by the latter cell type. These findings demonstrate that mDC activation requires an elevated intrinsic PKM2 level and that PKM2 improves the immune status of patients with SAA by enhancing the functions of mDCs and, consequently, CTLs.

scholarly journals Increase of E3 ubiquitin ligase NEDD4 expression leads to degradation of its target proteins PTEN/IGF1R during the formation of goose fatty liver

2020 ◽  
Vol 98 (9) ◽  
Author(s):  
Chunchi Yan ◽  
Minmeng Zhao ◽  
Shuo Li ◽  
Tongjun Liu ◽  
Cheng Xu ◽  
...  
Keyword(s):  
Fatty Liver ◽  
Mrna Expression ◽  
Messenger Rna ◽  
Quantitative Polymerase Chain Reaction ◽  
Normal Liver ◽  
Protective Mechanism ◽  
Nonalcoholic Fatty Liver ◽  
Target Proteins ◽  
Protein Levels ◽  
Gene 4

Abstract Goose fatty liver may have a unique protective mechanism as it does not show a pathological injury even in the case of severe steatosis. Although neural precursor cell-expressed developmentally downregulated gene 4 (NEDD4) participates in repair and regeneration of injured liver through its target proteins, its role in nonalcoholic fatty liver disease remains unknown. Using quantitative polymerase chain reaction (PCR) and immunoblot analyses, here, we found that the messenger RNA (mRNA) and protein expressions of NEDD4 were induced in goose fatty liver compared with normal liver. The mRNA expression of the gene of phosphate and tension homology deleted on chromosome ten (PTEN) and insulin-like growth factor 1 receptor (IGF1R) was also induced in goose fatty liver; however, their protein expression was or tended to be suppressed. Moreover, the co-immunoprecipitation analysis indicated that there was a physical association between NEDD4 and PTEN in goose liver, which was consistent with the ubiquitination of PTEN in goose fatty liver. Furthermore, NEDD4 overexpression in goose primary hepatocytes suppressed the PTEN and IGF1R protein levels without a significant effect on their mRNA expression. In conclusion, the increased expression of NEDD4 leads to the degradation of PTEN and IGF1R proteins through ubiquitination in goose fatty liver, suggesting that NEDD4 may protect goose fatty liver from severe steatosis-associated injury via its target proteins during the development of goose fatty liver.

scholarly journals Schistosome-derived omega-1 drives Th2 polarization by suppressing protein synthesis following internalization by the mannose receptor

2012 ◽  
Vol 209 (10) ◽  
pp. 1753-1767 ◽  
Author(s):  
Bart Everts ◽  
Leonie Hussaarts ◽  
Nicole N. Driessen ◽  
Moniek H.J. Meevissen ◽  
Gabriele Schramm ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Protein Synthesis ◽  
Messenger Rna ◽  
Molecular Mechanisms ◽  
Mannose Receptor ◽  
Site Directed Mutagenesis ◽  
Directed Mutagenesis ◽  
Th2 Polarization ◽  
Th2 Responses ◽  
Egg Antigen

Omega-1, a glycosylated T2 ribonuclease (RNase) secreted by Schistosoma mansoni eggs and abundantly present in soluble egg antigen, has recently been shown to condition dendritic cells (DCs) to prime Th2 responses. However, the molecular mechanisms underlying this effect remain unknown. We show in this study by site-directed mutagenesis of omega-1 that both the glycosylation and the RNase activity are essential to condition DCs for Th2 polarization. Mechanistically, we demonstrate that omega-1 is bound and internalized via its glycans by the mannose receptor (MR) and subsequently impairs protein synthesis by degrading both ribosomal and messenger RNA. These experiments reveal an unrecognized pathway involving MR and interference with protein synthesis that conditions DCs for Th2 priming.

scholarly journals AB0062 CHARACTERIZATION OF IL-12 AND IL-23 REDUCTION BY TOFACITINIB IN MDCS

2020 ◽  
Vol 79 (Suppl 1) ◽  
pp. 1332.2-1333
Author(s):  
N. Vincken ◽  
C. Angiolilli ◽  
S. Cardoso ◽  
A. Lopes ◽  
M. Olde-Nordkamp ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Psoriatic Arthritis ◽  
Mrna Expression ◽  
Intracellular Signaling ◽  
Magnetic Beads ◽  
Mononuclear Cells ◽  
Quantitative Polymerase Chain Reaction ◽  
Enzyme Linked Immunosorbent Assay ◽  
Myeloid Dendritic Cells ◽  
Ifn Γ

Background:Psoriatic arthritis (PsA) is a chronic inflammatory auto-immune disease characterized by an excessive production of pathogenic mediators that cause inflammation of the skin, peripheral joints, entheses and the spine. Among these, interleukin (IL)-23, IL-12, the IL-17 family and TNF constitute key players in PsA pathogenesis.1,2IL-23, consisting of IL23A (IL-23p19) and IL12B (IL-12p40) subunits, is predominantly produced by myeloid dendritic cells (mDCs). While the p19 subunit is unique to IL-23, the p40 subunit is shared with IL-12. Together, IL-12 and IL-23 play a crucial role in promoting the differentiation of naïve T lymphocytes into T helper (Th) interferon (IFN)-γ-producing Th1 or IL17-producing Th17 cells, respectively.3Small-molecule inhibitors, such as the JAK/STAT inhibitor Tofacitinib, have recently shown promising therapeutic potential in PsA clinical trials.4The inhibition of JAKs by Tofacitinib results in the direct suppression of multiple intracellular signaling pathways which constitute key hubs in the cytokine network.5However, whether Tofacitinib is able directly target IL-12/IL-23 production by mDCs has not yet been documented. Suppression of these canonical inflammatory pathways would provide further evidence that Tofacitinib is an effective drug in halting both innate and adaptive immune responses.Objectives:To evaluate the transcriptional and molecular events underlining IL-12 and IL-23 regulation by Tofacitinib in mDCs.Methods:Peripheral blood mononuclear cells from healthy donors were isolated by Ficoll gradient. Monocytes and myeloid dendritic cells (mDCs) were isolated by using magnetic beads on autoMACS. Monocytes were cultured for 6 days in the presence of IL-4 and granulocyte-macrophage colony-stimulating factor (GM-CSF) to generate monocyte-derived dendritic cells (moDCs). MoDCs were harvested, washed and put to rest for 1 day prior to stimulation, while mDCs were stimulated on the same day of isolation. Both moDCs and mDCs were pre-treated with Tofacitinib and then stimulated with either lipopolysaccharide (LPS) or combination of LPS with IFN-γ for 4 hours. Cytokines were measured using enzyme-linked immunosorbent assay (ELISA) and gene expression was assessed using quantitative polymerase chain reaction (qPCR).Results:Treatment of both mDCs and moDCs with Tofacitinib led to a decreased mRNA expression of IL-12 p40 (IL12B) in the presence of TLR4 and IFNγ co-stimulation. The decreasedIL12BmRNA expression also resulted in lower production of IL-12 p40 and IL-23 proteins in mDCs.Conclusion:In this work, we demonstrated for the first time that Tofacitinib can suppress the production of IL-23/IL-12 p40 subunit in mDCs, upon the condition that an active type II IFN signalling is also present in these cells. This observation indicates that specific factors, such as endogenous IFN-γ levels in the serum of PsA patients, can possibly predict differential responses to Tofacitinib treatment.References:[1]Gaffen SL. et al. The IL-23-IL-17 immune axis: from mechanisms to therapeutic testing. Nat Rev Immunol. 2014 Sep;14(9):585-600[2]Bravo A, Kavanaugh A. Bedside to bench: defining the immunopathogenesis of psoriatic arthritis. Nat Rev Rheumatol. 2019 Nov;15(11):645-656[3]Floss DM. et al. Insights into IL-23 biology: From structure to function. Cytokine Growth Factor Rev. 2015 Oct;26(5):569-78[4]Berekmeri A. et al. Tofacitinib for the treatment of psoriasis and psoriatic arthritis. Expert Rev Clin Immunol. 2018 Sep;14(9):719-730[5]T Virtanen A. et al. Selective JAKinibs: Prospects in Inflammatory and Autoimmune Diseases. BioDrugs. 2019 Feb;33(1):15-32Disclosure of Interests:None declared

scholarly journals Loss of STAT5 causes liver fibrosis and cancer development through increased TGF-β and STAT3 activation

2009 ◽  
Vol 206 (4) ◽  
pp. 819-831 ◽  
Author(s):  
Atsushi Hosui ◽  
Akiko Kimura ◽  
Daisuke Yamaji ◽  
Bing-mei Zhu ◽  
Risu Na ◽  
...  
Keyword(s):  
Liver Fibrosis ◽  
Messenger Rna ◽  
Molecular Mechanisms ◽  
Transforming Growth Factor ◽  
Null Mouse ◽  
Protein Levels ◽  
Β Protein ◽  
Ccl4 Treatment ◽  
The Stability

The molecular mechanisms underlying the development of hepatocellular carcinoma are not fully understood. Liver-specific signal transducer and activator of transcription (STAT) 5A/B–null mice (STAT5-LKO) were treated with carbon tetrachloride (CCl4), and histological analyses revealed liver fibrosis and tumors. Transforming growth factor (TGF)–β levels and STAT3 activity were elevated in liver tissue from STAT5-LKO mice upon CCl4 treatment. To define the molecular link between STAT5 silencing and TGF-β up-regulation, as well as STAT3 activation, we examined STAT5-null mouse embryonic fibroblasts and primary hepatocytes. These cells displayed elevated TGF-β protein levels, whereas messenger RNA levels remained almost unchanged. Protease inhibitor studies revealed that STAT5 deficiency enhanced the stability of mature TGF-β. Immunoprecipitation and immunohistochemistry analyses demonstrated that STAT5, through its N-terminal sequences, could bind to TGF-β and that retroviral-mediated overexpression of STAT5 decreased TGF-β levels. To confirm the in vivo significance of the N-terminal domain of STAT5, we treated mice that expressed STAT5 lacking the N terminus (STAT5-ΔN) with CCl4. STAT5-ΔN mice developed CCl4-induced liver fibrosis but no tumors. In conclusion, loss of STAT5 results in elevated TGF-β levels and enhanced growth hormone–induced STAT3 activity. We propose that a deregulated STAT5–TGF-β–STAT3 network contributes to the development of chronic liver disease.

Cytokine production by mouse myeloid dendritic cells in relation to differentiation and terminal maturation induced by lipopolysaccharide or CD40 ligation

Blood ◽  
2001 ◽  
Vol 98 (5) ◽  
pp. 1512-1523 ◽  
Author(s):  
Adrian E. Morelli ◽  
Alan F. Zahorchak ◽  
Adriana T. Larregina ◽  
Bridget L. Colvin ◽  
Alison J. Logar ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Transforming Growth Factor ◽  
De Novo ◽  
Transforming Growth Factor Β1 ◽  
Th2 Cells ◽  
Myeloid Dendritic Cells ◽  
Cd40 Ligation ◽  
Murine Bone Marrow ◽  
Protein Levels ◽  
Tnf Α

Although it is known that dendritic cells (DCs) produce cytokines, there is little information about how cytokine synthesis is regulated during DC development. A range of cytokine mRNA/proteins was analyzed in immature (CD86−) or mature (CD86+) murine bone marrow (BM)- derived DCs. Highly purified, flow-sorted, immature DCs exhibited higher amounts of interleukin-1α (IL-1α), IL-1β, tumor necrosis factor-α (TNF-α), transforming growth factor β1 (TGF-β1), and macrophage migration inhibitory factor (MIF) mRNA/protein than mature DCs. After differentiation, DC up-regulated the levels of IL-6 and IL-15 mRNA/protein and synthesized de novo mRNA/protein for IL-12p35, IL-12p40, and IL-18. Although immature BM-derived DCs did not stimulate naive allogeneic T cells, mature DCs elicited a mixed population of T helper (Th) 1 (mainly) and Th2 cells in 3d-mixed leukocyte reactions. CD86+ BM DCs switched to different cytokine patterns according to whether they were terminally differentiated by lipopolysaccharide (LPS) or CD40 ligation. Although both stimuli increased IL-6, IL-12p40, IL-15, and TNF-α mRNA/protein levels, only LPS up-regulated transcription of IL-1α, IL-1β, IL-12p35, and MIF genes. Although LPS and CD40 cross-linking increased the T-cell allostimulatory function of BM DCs, only LPS stimulation shifted the balance of naive Th differentiation to Th1 cells, a mechanism dependent on the up-regulation of IL-12p35 and not of IL-23. These results demonstrate that, depending on the stimuli used to terminally mature BM DCs, DCs synthesize a different pattern of cytokines and exhibit distinct Th cell–driving potential.

scholarly journals Pathways Involved in Interleukin-1β–Mediated Murine Cardiomyocyte Apoptosis

2015 ◽  
Vol 42 (2) ◽  
pp. 109-116 ◽  
Author(s):  
Yi Shen ◽  
Jie Qin ◽  
Peili Bu
Keyword(s):  
Heart Failure ◽  
Messenger Rna ◽  
Molecular Mechanisms ◽  
Quantitative Polymerase Chain Reaction ◽  
Interleukin 1 ◽  
Cardiomyocyte Apoptosis ◽  
Enzyme Linked Immunosorbent Assay ◽  
Endonuclease G ◽  
Apoptosis Protein

Accumulating evidence suggests that interleukin-1 (IL-1) signaling plays an essential role in the pathogenesis of heart failure by inducing cardiomyocyte apoptosis, but the mechanisms of this process are poorly defined. We further explored these molecular pathways. We isolated cardiomyocytes from neonatal mice and then cultured and stimulated them with murine IL-1β in vitro. Cell apoptotic ratios were measured by means of flow cytometry. Expression of effector molecules was analyzed by means of enzyme-linked immunosorbent assay, Western blotting, and real-time quantitative polymerase chain reaction. The results showed that IL-1β induced murine cardiomyocyte apoptosis through a release of cytochrome c into cytoplasm and through caspase 3 activation. Simultaneously, IL-1β signaling promoted expression of endonuclease G and high-temperature requirement protein A2 messenger RNA. Survivin and X-linked inhibitors of apoptosis protein (IAP), members of the IAP family, were inhibited on the messenger RNA level during IL-1β–mediated cardiomyocyte apoptosis. We found that IL-1β signaling during cardiomyocyte apoptosis in vitro induced the activation of caspase-dependent and caspase-independent pathways, and inhibited IAPs. Understanding the molecular mechanisms involved in IL-1β–mediated cardiomyocyte apoptosis might assist in the design of therapeutic approaches to protect cardiomyocyte function and prevent heart failure.

scholarly journals HMGB1 Activates Myeloid Dendritic Cells by Up-Regulating mTOR Pathway in Systemic Lupus Erythematosus

2021 ◽  
Vol 8 ◽  
Author(s):  
Xinghui Song ◽  
Hui Zhang ◽  
Yun Zhao ◽  
Yuanzhen Lin ◽  
Qiya Tang ◽  
...  
Keyword(s):  
Dendritic Cells ◽  
Lupus Erythematosus ◽  
Molecular Mechanisms ◽  
Mtor Pathway ◽  
Systemic Lupus ◽  
Myeloid Dendritic Cells ◽  
Hla Dr ◽  
Tnf Α ◽  
Hmgb1 Expression ◽  
Pro Inflammatory Cytokines

Research has shown that HMGB1 can activate dendritic cells (DCs), but its molecular mechanisms are not clear. In this study, we reported that the myeloid dendritic cells (mDCs) were activated in the peripheral blood of SLE patients, and the activation of mDCs was associated with the up-regulation of HMGB1 and mTOR. After stimulated by HMGB1, expression of mTOR and its substrates P70S6K and 4EBP1 in dendritic cells increased considerably (P < 0.01). The expression of HLA-DR, CD40, and CD86 on dendritic cells also significantly increased following these stimuli (P < 0.01). In addition, stimulation with HMGB1 enhanced cytokine (IL-1β, IL-6, and TNF-a) production in dendritic cells. In contrast, the HMGB1-mediated expression of HLA-DR, CD40, and CD86 on dendritic cells and production of IL-1β, IL-6, and TNF-α were reduced by rapamycin. Rapamycin can inhibit HMGB1-induced activation of mDCs and secretion of pro-inflammatory cytokines. These findings indicated that HMGB1activates mDCs by up-regulating the mTOR pathway in SLE.

scholarly journals Effect of 8-Day Fasting on Leukocytes Expression of Genes and Proteins Involved in Iron Metabolism in Healthy Men

2021 ◽  
Vol 22 (6) ◽  
pp. 3248
Author(s):  
Andżelika Borkowska ◽  
Maja Tomczyk ◽  
Małgorzata Żychowska ◽  
Wiesław Pilis ◽  
Michał Zych ◽  
...  
Keyword(s):  
Exercise Capacity ◽  
Molecular Mechanisms ◽  
Iron Acquisition ◽  
Quantitative Polymerase Chain Reaction ◽  
Essential Element ◽  
The Body ◽  
Iron Storage ◽  
Healthy Men ◽  
Protein Levels ◽  
Expression Of Genes

The popularity of fasting and restricted food intake is increasing. While the body’s adaptability to dietary insufficiency is crucial for health, molecular mechanisms of adaptive changes are not well understood. Here, we compared the effects of fasting and exercise on the expression of leukocyte genes and proteins involved in the storage, export, and acquisition of iron, an essential element with physiological roles. Healthy men participated in the study (age, 30–70 years; body weight, 60–100 kg; body mass index, 20–29.9 kg/m2). The participants performed an exercise test with a gradually increasing intensity until the individual maximum exercise capacity was reached, before and after 8-d fast. Blood samples were collected before, immediately after, and 3 h after exercise. Gene expression was analyzed by reverse-transcription quantitative polymerase chain reaction and protein levels were analyzed by immunobloting. Eight days of total starvation diet affected the body composition and decreased exercise capacity. Further, fasting decreased the expression of genes associated with iron storage and export, and increased the expression of genes involved in iron acquisition. Conversely, only PCBP2 protein increased after fasting; however, an upward trend was apparent for all proteins. In conclusion, the body adapts to starvation by adjusting iron economy.

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