scholarly journals Activation of ChTLR15/ChNF-κB-ChNLRP3/ChIL-1β signaling transduction pathway mediated inflammatory responses to E. tenella infection

2021 ◽  
Vol 52 (1) ◽  
Author(s):  
Jian Li ◽  
Xuelian Yang ◽  
Zhipeng Jia ◽  
Chunli Ma ◽  
Xinghui Pan ◽  
...  
Keyword(s):  
Inflammatory Responses ◽  
Chicken Embryo Fibroblast ◽  
Economic Losses ◽  
Fibroblast Cells ◽  
Mrna Levels ◽  
Transduction Pathway ◽  
Toll Like Receptor ◽  
Detailed Mechanism ◽  
Protein Levels ◽  
Signaling Transduction Pathway

AbstractAvian coccidiosis caused by Eimeria leads to severe economic losses in the global poultry industry. Although chicken Toll-like receptor 15 (ChTLR15) was reported to be involved in Eimeria infection, the detailed mechanism underlying its role in the inflammatory response remains to be discovered. The present study demonstrated that the mRNA expression levels of ChTLR15, ChMyD88, ChNF-κB, ChNLRP3, ChCaspase-1, ChIL-18 and ChIL-1β and the protein levels of ChTLR15 and ChNLRP3 in cecal tissues of Eimeria-infected chickens were significantly elevated at 4, 12, and 24 h compared with those in noninfected control chickens (p < 0.01). Moreover, the mRNA levels of molecules in the ChTLR15/ChNF-κB and ChNLRP3/ChIL-1β pathways and the protein levels of ChTLR15 and ChNLRP3 in chicken embryo fibroblast cells (DF-1) stimulated by E. tenella sporozoites were consistent with those in Eimeria-infected chickens. Furthermore, overexpression of ChTLR15 in DF1 cells augmented activation of the ChTLR15/ChNF-κB and ChNLRP3/ChIL-1β pathways when stimulated with E. tenella sporozoites, while knockdown of ChTLR15 in DF1 cells showed inverse effects. Taken together, the present study provides evidence that E. tenella sporozoites specifically activate ChTLR15 and then trigger activation of the ChNLRP3/ChIL-1β pathway, which partially mediates inflammatory responses to Eimeria infection.

scholarly journals Kyungheechunggan-Tang-01, a New Herbal Medication, Suppresses LPS-Induced Inflammatory Responses through JAK/STAT Signaling Pathway in RAW 264.7 Macrophages

2017 ◽  
Vol 2017 ◽  
pp. 1-15 ◽  
Author(s):  
Hee-Soo Han ◽  
Eungyeong Jang ◽  
Ji-Sun Shin ◽  
Kyung-Soo Inn ◽  
Jang-Hoon Lee ◽  
...  
Keyword(s):  
Signaling Pathway ◽  
Inflammatory Responses ◽  
Inflammatory Diseases ◽  
Molecular Data ◽  
Mrna Levels ◽  
Protein Levels ◽  
Stat3 Phosphorylation ◽  
Stat Signaling ◽  
Cox 2 ◽  
Anti Inflammatory

Medicinal plants have been used as alternative therapeutic tools to alleviate inflammatory diseases. The objective of this study was to evaluate anti-inflammatory properties of Kyungheechunggan-tang- (KCT-) 01, KCT-02, and Injinchunggan-tang (IJCGT) as newly developed decoctions containing 3–11 herbs in LPS-induced macrophages. KCT-01 showed the most potent inhibitory effects on LPS-induced NO, PGE2, TNF-α, and IL-6 production among those three herbal formulas. In addition, KCT-01 significantly inhibited LPS-induced iNOS and COX-2 at protein levels and expression of iNOS, COX-2, TNF-α, and IL-6 at mRNA levels. Molecular data revealed that KCT-01 attenuated the activation of JAK/STAT signaling cascade without affecting NF-κB or AP-1 activation. In ear inflammation induced by croton oil, KCT-01 significantly reduced edema, MPO activity, expression levels of iNOS and COX-2, and STAT3 phosphorylation in ear tissues. Taken together, our findings suggest that KCT-01 can downregulate the expression of proinflammatory genes by inhibiting JAK/STAT signaling pathway under inflammatory conditions. This study provides useful data for further exploration and application of KCT-01 as a potential anti-inflammatory medicine.

scholarly journals A Novel Role of Numb as A Regulator of Pro-inflammatory Cytokine Production in Macrophages in Response to Toll-like Receptor 4

2015 ◽  
Vol 5 (1) ◽  
Author(s):  
Patipark Kueanjinda ◽  
Sittiruk Roytrakul ◽  
Tanapat Palaga
Keyword(s):  
Notch Signaling ◽  
P38 Mapk ◽  
Inflammatory Responses ◽  
Notch Signaling Pathway ◽  
Negative Regulator ◽  
Mrna Levels ◽  
Toll Like Receptor ◽  
Toll Like Receptor 4 ◽  
Murine Bone Marrow

Abstract Activation of macrophages triggers the release of pro-inflammatory cytokines leading to inflammation. Numb is a negative regulator of Notch signaling, but the role of Numb in macrophages is not fully understood. In this study, the role of Numb as a regulator of inflammatory responses in macrophages was investigated. Murine bone marrow-derived macrophages, in which expression of Numb was silenced, secreted significantly less TNFα, IL-6 and IL-12 and more IL-10 upon activation by lipopolysaccharide (LPS), a ligand for Toll-like receptor 4 (TLR4), despite increased Notch signaling. The Tnfα mRNA levels both in Numb-deficient and wild-type macrophages were not significantly different, unlike those of Il6 and Il12-p40. In Numb-deficient macrophages, the Tnfα mRNAs were degraded at faster rate, compared to those in control macrophages. Activation of p38 MAPK and NF-κΒ p65 were compromised in activated Numb deficient macrophages. Numb was found to interact with the E3 ubiquitin ligase, Itch, which reportedly regulates p38 MAPK. In addition, blocking the Notch signaling pathway in activated, Numb-deficient macrophages did not further reduce TNFα levels, suggesting a Notch-independent role for Numb. A proteomics approach revealed a novel funciton for Numb in regulating complex signaling cascades downstream of TLRs, partially involving Akt/NF-κB p65/p38 MAPK in macrophages.

scholarly journals Differential release and deposition of S100A8/A9 proteins in inflamed upper airway tissue

2015 ◽  
Vol 47 (1) ◽  
pp. 264-274 ◽  
Author(s):  
Koen Van Crombruggen ◽  
Thomas Vogl ◽  
Claudina Pérez-Novo ◽  
Gabriele Holtappels ◽  
Claus Bachert
Keyword(s):  
Inflammatory Mediators ◽  
Inflammatory Responses ◽  
Upper Airway ◽  
Toll Like Receptor ◽  
Functional Responses ◽  
Protein Levels ◽  
Glycation End Products ◽  
Polyp Tissue ◽  
Intracellular Ca ◽  
Blocking Antibodies

Intracellular Ca2+-binding S100A8/A9 proteins gain novel functions when released during inflammation. The exact outcome of their extracellular function depends on the local tissue environment in which they are released; both anti-inflammatory and pro-inflammatory responses are described, modulating the immune system by binding Toll-like receptor (TLR)-4 or the receptor for advanced glycation end-products (RAGE). However, the contribution of the proteins in the pathophysiology of chronic rhinosinusitis (CRS) remains unclear.Homomeric S100A8 and S100A9, and heteromeric S100A8/A9 proteins were evaluated in CRS with/without nasal polyps (CRSw/sNP) and controls. Functional responses were assessed in polyp tissue stimulated with S100 proteins in the presence of TLR-4 and RAGE blocking antibodies.S100A8, S100A9 and S100A8/A9 protein levels were significantly higher in CRSwNP patients, showing increased deposition on extracellular matrix (ECM) structures of CRSwNP tissue in contrast to CRSsNP and controls. In the presence ofStaphylococcus aureus, S100A8/A9 is released from neutrophils and from the ECM. Extracellular S100A8 and S100A9 proteins induced increased levels of diverse inflammatory mediatorsviaTLR-4 engagement.The inflammatory/remodelling characteristics of CRSwNP specifically allow increased retention of S100A8, S100A9 and S100A8/A9 proteins in the ECM of CRSwNP tissue. Upon release, homodimeric proteins act as a local danger signal inducing inflammatory mediators, predominantlyviaTLR-4 activation.

scholarly journals Pro-inflammatory AGE-RAGE signaling is activated during arousal from hibernation in ground squirrel adipose

PeerJ ◽  
2018 ◽  
Vol 6 ◽  
pp. e4911 ◽  
Author(s):  
Samantha M. Logan ◽  
Kenneth B. Storey
Keyword(s):  
Gene Expression ◽  
Ground Squirrel ◽  
Inflammatory Responses ◽  
Mrna Levels ◽  
Protein Levels ◽  
Ictidomys Tridecemlineatus ◽  
Molecular Pattern ◽  
Brown Adipose ◽  
Dying Cells ◽  
Relative Gene

Background Inflammation is generally suppressed during hibernation, but select tissues (e.g. lung) have been shown to activate both antioxidant and pro-inflammatory pathways, particularly during arousal from torpor when breathing rates increase and oxidative metabolism fueling the rewarming process produces more reactive oxygen species. Brown and white adipose tissues are now understood to be major hubs for the regulation of immune and inflammatory responses, yet how these potentially damaging processes are regulated by fat tissues during hibernation has hardly been studied. The advanced glycation end-product receptor (RAGE) can induce pro-inflammatory responses when bound by AGEs (which are glycated and oxidized proteins, lipids, or nucleic acids) or damage associated molecular pattern molecules (DAMPs, which are released from dying cells). Methods Since gene expression and protein synthesis are largely suppressed during torpor, increases in AGE-RAGE pathway proteins relative to a euthermic control could suggest some role for these pro-inflammatory mediators during hibernation. This study determined how the pro-inflammatory AGE-RAGE signaling pathway is regulated at six major time points of the torpor-arousal cycle in brown and white adipose from a model hibernator, Ictidomys tridecemlineatus. Immunoblotting, RT-qPCR, and a competitive ELISA were used to assess the relative gene expression and protein levels of key regulators of the AGE-RAGE pathway during a hibernation bout. Results The results of this study revealed that RAGE is upregulated as animals arouse from torpor in both types of fat, but AGE and DAMP levels either remain unchanged or decrease. Downstream of the AGE-RAGE cascade, nfat5 was more highly expressed during arousal in brown adipose. Discussion An increase in RAGE protein levels and elevated mRNA levels of the downstream transcription factor nfat5 during arousal suggest the pro-inflammatory response is upregulated in adipose tissue of the hibernating ground squirrel. It is unlikely that this cascade is activated by AGEs or DAMPs. This research sheds light on how a fat-but-fit organism with highly regulated metabolism may control the pro-inflammatory AGE-RAGE pathway, a signaling cascade that is often dysregulated in other obese organisms.

scholarly journals Glucan phosphate inhibits HMGB-1 release from rat myocardial H9C2 cells in sepsis via TLR4/NF-кB signal pathway

2017 ◽  
Vol 40 (2) ◽  
pp. 66 ◽  
Author(s):  
Haizhu Wang ◽  
Zhifei Cui ◽  
Fei Sun ◽  
Huayong Ding
Keyword(s):  
Cultured Cells ◽  
Mrna Levels ◽  
H9c2 Cells ◽  
Toll Like Receptor ◽  
Mobility Shift ◽  
Toll Like Receptor 4 ◽  
Protein Levels ◽  
Elevated Activity ◽  
Underlying Mechanisms ◽  
Electrophoretic Mobility Shift Assays

Purpose: The effect of glucan phosphate (GP) on the release of HMGB-1 from rat myocardial cells (H9C2) during lipopolysaccharide-induced sepsis, and the underlying mechanisms, were investigated. Methods: H9C2 cells were divided into three groups: normal; lipopolysaccharide (LPS) (1 mg/ml LPS); and, LPS+GP (2 mg/ml GP). Western blot was used to determine toll-like receptor 4 (TLR4) levels, and electrophoretic mobility-shift assays (EMSA) was used to determine nuclear factor-кB (NF-кB) activity 3, 6 and 9 h after treatment. HMGB-1 mRNA levels in cultured cells were determined by real-time PCR and supernatant HMGB-1 protein levels were evaluated by ELISA at 12, 24, 36 and 48 h after treatment. Following the transfection of H9C2 cells with Ad5-IкBα, which inhibits NF-кB activity, TLR4, NF-кB and HMGB-1 levels were determined. Results: Intracellular TLR4 levels and NF-кB activity in LPS and LPS+GP groups increased 3-9 h after stimulation, but the increased levels of TLR4 and elevated activity of NF-кB were significantly lower in the LPS+GP group vs. the LPS group. HMGB-1 mRNA levels in both LPS and LPS+GP groups, increased gradually from 24 h after stimulation, but the increase was more obvious in the LPS group vs. the LPS+GP group. Supernatant HMGB-1 levels in the LPS and LPS+GP groups increased gradually from 9 h after stimulation, and also increased markedly in the LPS group. After the inhibition of NF-кB activity, LPS-induced HMGB-1 release decreased significantly (p

scholarly journals Skin Improvement Effects of Gardeniae fructus Extract in HaCaT Keratinocytes, B16F10 Melanocytes, and CCD-986sk Fibroblast Cells

Cosmetics ◽  
2019 ◽  
Vol 6 (3) ◽  
pp. 48 ◽  
Author(s):  
Se Hyang Hong ◽  
Jin Mo Ku ◽  
Seung Hwan Lee ◽  
Ho Jong Shim ◽  
Dong Sun Park ◽  
...  
Keyword(s):  
Free Radical ◽  
Inhibitory Activity ◽  
Radical Scavenging ◽  
Fibroblast Cells ◽  
Mrna Levels ◽  
Protein Levels ◽  
Free Radical Scavenging Assay ◽  
Free Radical Activity ◽  
Dose Dependent ◽  
Whitening Activity

The development of functional cosmetics with skin improvement effects from natural sources is necessary. In this study, the antioxidant, antiwrinkling, moisturizing, and whitening effects of Gardeniae fructus extract (GF) were investigated in keratinocytes, melanocytes, and fibroblast cells. Antioxidant activity was determined by a DPPH free radical scavenging assay. MMP-1, MMP-9, HAS1, and filaggrin mRNA levels were measured by RT-PCR in keratinocytes and fibroblast cells. MITF and tyrosinase protein levels were evaluated by blotting analysis in melanocytes. DPPH free radical activity was investigated to determine whether GF showed dose-dependent inhibitory activity. GF induced the upregulation of HAS1 and filaggrin mRNA expression in keratinocytes and fibroblast cells. GF led to the downregulation of MMP mRNA levels in keratinocytes and fibroblast cells. Western blotting was performed to confirm the whitening-related protein (MITF and tyrosinase) levels induced by GF in melanocytes, and the inhibitory activity was superior to that of the α-MSH used for the comparison test. GF showed marked antioxidant, antiwrinkling, skin moisturizing, and whitening activity in keratinocytes, melanocytes, and fibroblast cells. Through the results of these experiments, the applicability of GF as a natural and functional cosmetic material was verified.

Abstract 578: Na/K-ATPase Forms a Signaling Complex with CD36 and Toll-like Receptor 4 to Mediate NF-kappaB Activation in Macrophages

2016 ◽  
Vol 36 (suppl_1) ◽  
Author(s):  
Yiliang Chen ◽  
Wenxin Huang ◽  
Moua Yang ◽  
Roy Silverstein
Keyword(s):  
Inflammatory Responses ◽  
Culture Media ◽  
Mrna Level ◽  
Specific Inhibitor ◽  
Adaptor Protein ◽  
Human Monocyte ◽  
Mrna Levels ◽  
Signaling Complex ◽  
Protein Levels ◽  
Plasma Membrane Receptor

Cardiotonic steroids such as ouabain, digoxin, and marinobufagenin are known ligands for the plasma membrane receptor, Na/K-ATPase (NKA). These ligands stimulate complex formation of NKA with other membrane and cytosolic proteins leading to cellular events such as activation of kinase cascades and gene transcription independent of the ion transporting function. Endogenous cardiotonic steroids have been detected within human circulation and their levels are elevated in patients with diseases associated with chronic systemic inflammation. However, the role of cardiotonic steroids and NKA-mediated signaling in inflammation has not been well studied. We now report that ouabain activates the NF-κB pathway in human monocyte-derived macrophages, leading to pro-inflammatory responses. Using Western blot and densitometry analysis, we found that physiological concentrations of ouabain stimulated IκBα degradation (n=3). Using quantitative RT-PCR (n=4), we found that ouabain increased mRNA levels of pro-inflammatory genes MCP-1 (3.2±1.1 fold), TNF-α (59.7±35.6 fold), CXCL-10 (2.8±1.6 fold), ICAM1 (2.9±0.5 fold), and PTGS2 (3.1±0.1 fold). Consistent with the increase in mRNA level, MCP-1 protein levels in post-culture media assessed by ELISA were elevated by 1.4±0.1 fold (n=4). Mechanistically, when macrophages were incubated with ouabain for 20 min and then subjected to IP, we found that antibodies to NKA co-precipitated more TLR4 (2.2±0.8 fold) compared to unstimulated cells. Additionally, antibodies to CD36 co-precipitated more NKA (1.8±0.6 fold) and more TLR4 (2.2±0.5 fold) (n=3). Blockade of TLR4 signaling using a specific inhibitor, CLI-095, abolished ouabain-induced MCP-1 production as assessed by ELISA (n=4). Next, we used siRNA to specifically knock down either NKA α1, CD36, TLR4 or MyD88 (the adaptor protein downstream of TLR4) in THP-1 cells stably transfected with a NF-κB reporter construct. After siRNA transfection for 48h, cells were treated with ouabain for additional 24h. Knockdown of any one of these proteins fully suppressed ouabain-induced NF-κB activation by NF-κB reporter assay (n=4). In conclusion, ouabain activates NF-κB through an NKA/CD36/TLR4 complex leading to pro-inflammatory responses in human macrophages.

The Effects of Vitamin K1 and Warfarin on Prothrombin Expression in Human Hepatoblastoma (HepG2) Cells

1992 ◽  
Vol 68 (01) ◽  
pp. 040-047 ◽  
Author(s):  
C Scott Jamison ◽  
Bryan F Burkey ◽  
Sandra J Friezner Degen
Keyword(s):  
Total Protein ◽  
Hepg2 Cells ◽  
Enzyme Linked Immunosorbent Assay ◽  
Response Analysis ◽  
Secreted Protein ◽  
Mrna Levels ◽  
Vitamin K1 ◽  
Maximal Increase ◽  
Protein Levels ◽  
Warfarin Treatment

SummaryCultures of human hepatoblastoma (HepG2) cells were treated with vitamin K1 or warfarin and prothrombin antigen and mRNA levels were determined. With 3 and 6 h of 10 µg vitamin K1 treatment secreted prothrombin antigen levels, relative to total secreted protein levels, were increased 1.5-fold and 2.1-fold, respectively, over ethanol-treated control levels as determined by an enzyme-linked immunosorbent assay. Dose-response analysis with 3 h of 25 µg/ml vitamin K1 treatment demonstrated a maximal increase of 2.0-fold in secreted prothrombin antigen levels, relative to total secreted protein levels, over ethanol-treated control levels. Pulse-chase analysis with 35S-methionine and immunoprecipitation of 35S-labelled prothrombin demonstrated that, with vitamin K1 treatment (25 µg/ml, 3 h), the rate of prothrombin secretion increased approximately 2-fold and the total amount (intra- and extracellular) of prothrombin synthesized increased approximately 50% over ethanol-treated control levels. Warfarin treatment (1, 5, or 10 µg/ml, 24 h) resulted in decreases in secreted prothrombin antigen levels, relative to total protein levels to approximately 85%, 87% or 81% of ethanol-treated control levels. Analysis of total RNA isolated from these cultures by Northern and solution hybridization techniques demonstrated that prothrombin mRNA was approximately 2.1 kb and that neither vitamin K1 nor warfarin treatment affected the quantity of prothrombin mRNA (ranging from 240–350 prothrombin mRNA molecules per cell). These results demonstrate that vitamin K1 and warfarin, in addition to effects on γ-carboxylation, affect prothrombin synthesis post-transcriptionally, perhaps influencing translation, post-translational processing and/or secretion mechanisms.

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