Transcription Factor NF Research Articles

Particulate matters increase epithelial-mesenchymal transition and lung fibrosis through the ETS-1/NF-\u03baB-dependent pathway in lung epithelial cells

BackgroundParticulate matters (PMs) in ambient air pollution are closely related to the incidence of respiratory diseases and decreased lung function. Our previous report demonstrated that PMs-induced oxidative stress increased the expression of proinflammatory intracellular adhesion molecule-1 (ICAM-1) through the IL-6/AKT/STAT3/NF-κB pathway in A549 cells. However, the role of O-PMs in epithelial-mesenchymal transition (EMT) development and pulmonary fibrosis and the related mechanisms have not been determined. The aim of this study was to investigate the effects of O-PMs on the pathogenesis of EMT and pulmonary fibrosis as well as the expression of ETS-1 and NF-κB p65, in vitro and in vivo.ResultsO-PMs treatment induced EMT development, fibronectin expression, and cell migration. O-PMs affected the expression of the EMT-related transcription factors NF-κB p65 and ETS-1. Interference with NF-κB p65 significantly decreased O-PMs-induced fibronectin expression. In addition, O-PMs affected the expression of fibronectin, E-cadherin, and vimentin through modulating ETS-1 expression. ATN-161, an antagonist of integrin α5β1, decreased the expression of fibronectin and ETS-1 and EMT development. EMT development and the expression of fibronectin and ETS-1 were increased in the lung tissue of mice after exposure to PMs for 7 and 14 days. There was a significant correlation between fibronectin and ETS-1 expression in human pulmonary fibrosis tissue.ConclusionO-PMs can induce EMT and fibronectin expression through the activation of transcription factors ETS-1 and NF-κB in A549 cells. PMs can induce EMT development and the expression of fibronectin and ETS-1 in mouse lung tissues. These findings suggest that the ETS-1 pathway could be a novel and alternative mechanism for EMT development and pulmonary fibrosis.

Ethanolic extract of Artemisia campestris subsp. glutinosa (Besser) Batt. inhibits HIV–1 replication in vitro through the activity of terpenes and flavonoids on viral entry and NF–κB pathway

Ethno–pharmacological relevanceThe genus Artemisia spp. is well known for its anti–infectious properties and its high content in anti–infectious compounds, like the well–known sweet wormwood (Artemisia annua L.). Another Artemisia species, Artemisia campestris subsp. glutinosa (Besser) Batt., field wormwood, has been traditionally used as medicinal plant in the Mediterranean region. Aim of the studyThe aim of this study is to investigate the anti–HIV activity of field wormwood, to identify the compounds responsible for this activity and their structure and mechanism of action. Materials and methodsAntiviral activity of isolated compounds and extracts was evaluated in HIV–1 infections of lymphoblastoid cells. We also evaluated the mechanism of action of isolated compounds. Viral entry was studied comparing the inhibitory effect of isolated compounds on wild type HIV–1 and VSV pseudotyped HIV–1. To assess the viral transcriptional effect, plasmids encoding luciferase reporter genes under the control of the whole genome of HIV–1 or NF–κB or Sp1 transcription factors were transfected in the presence of the compounds under evaluation. Finally, antioxidant activity was assessed by quantitation of reduced and total glutathione in treated cell cultures. ResultsEthanolic and aqueous extracts of Artemisia campestris subsp. glutinosa (Besser) Batt. subsp. glutinosa displayed anti–HIV activity in vitro, although ethanolic extract was more powerful (IC50 14.62 μg/mL). Bio–guided ethanolic extract fractionation leads to the isolation and characterization of two terpenes, damsin and canrenone, and four flavonoids, 6, 2′, 4′–trimethoxyflavone, acerosin, cardamonin and xanthomicrol. All the isolated compounds inhibited HIV–1 replication in vitro with IC50 values between the middle nanomolar and the low micromolar range. Their anti–HIV mechanism of action is due to the bloking of viral entry and/or transcription inhibition, without correlation with the antioxidant activity, through interference with the cellular transcription factors NF–κB and Sp1, which are targets that are not currently reached by antiretroviral therapy. ConclusionWe describe here the anti–HIV activity of field wormwood, Artemisia campestris subsp. glutinosa (Besser) Batt., and the isolation and study of the mechanism of action of two terpenes and four flavonoids, responsible, at least in part, for its activity, through the inhibition of two different cellular targets affecting the HIV replication cycle. The activity of these compounds in cellular targets could explain why plant extracts can be used in the treatment of different diseases. Besides, the presence of several compounds with dual and different mechanisms of action could prove useful in the treatment of HIV–1 infection, since it could aid to overcome drug resistances and simplify drug therapy. This work is a further step in understanding the anti–infectious activity of wormwood species and their use in treating infectious diseases.

HIV-1 Tat \u2013 TLR4/MD2 interaction drives the expression of IDO-1 in monocytes derived dendritic cells through NF-\u03baB dependent pathway

In the present study we showed that HIV-1 Tat protein stimulated the expression of Indoleamine 2,3 dioxygenase (IDO) -1 in human monocytes derived dendritic cells (MoDC) but not IDO-2 by acting directly at the cell membrane level. This induction of IDO-1 is dependent on the secondary structure of Tat protein, since stimulation with a chemically oxidized Tat protein loses its capacity to induce the production of IDO-1. Among the variety of candidate receptors described for Tat, we demonstrated that Tat protein interacted physically with TLR4/MD2 complex. Strikingly, blockade of Tat-TLR4 interaction by anti-TLR4 antibodies (clone HTA125), LPS-RS, a known TLR4 antagonist, or by soluble recombinant TLR4/MD2 complex inhibited strongly or totally the capacity of Tat to induce IDO-1 in MoDC while such treatments had no effect on IFN-γ-induced IDO-1. Furthermore, we showed that the activation of the transcription factor NF-κB by Tat is essential for the production of IDO-1 by human MoDC. Indeed, Tat activated NF-κB pathway in MoDC as demonstrated by the phosphorylation of p65 in Tat-treated MoDC. Further, we demonstrate that the stimulation of IDO-1 by Tat or by IFN-γ was totally or partially inhibited in the presence of NF-κB inhibitor respectively. These results suggest that Tat and IFN-γ act probably by two distinct mechanisms to induce the production of IDO-1. Our results clearly demonstrated that, although TLR4 pathway is necessary for Tat-induced IDO-1 in MoDC, it seems not to be sufficient since stable transfection of a functional TLR4/MD2 pathway in HEK or HeLa cell lines which are endogenously defectives for TLR4, did not restore the capacity of Tat to induce IDO-1 while IFN-γ treatment induces IDO-1 in HeLa cells independently of TLR4 pathway. These results suggest the involvement of additional stimuli in addition to TLR4 pathway which remain to be identified. Altogether our results demonstrated that, in human MoDC, HIV-1 Tat protein induced IDO-1 expression and activity in a NF-κB dependent-manner by recruiting TLR4 pathway.

INHIBITION OF TRANSCRIPTION FACTORS NF KAPPA B AND AP-1 LIMITS THE PROGRESSION OF OXIDATIVE-NITROSATIVE STRESS IN THE TISSUE OF CEREBRAL HEMISPHERES IN RATS AFTER MODELLED TRAUMATIC BRAIN INJURY

The study was aimed as investigating the effects of inhibitors of transcription factors NF kappa B and AP-1 activation on the development of oxidative-nitrosative stress in rat cerebral hemispheres following experimental traumatic brain injury (TBI). The study included 60 white Wistar male rats weighing 180-220 g, divided into 4 groups of 7 animals in each: the 1st group included pseudo-injured animals subjected to the same manipulations (ether anaesthesia, fixation) as the animals in the experimental groups, except for TBI modelling, the 2nd group included the animals exposed to modelled TBI, the 3rd and 4th groups involved the rats who received ammonium pyrolidine dithiocarbamate, the nuclear translocation inhibitor NF kappa B in a dose of 76 mg/kg and the inhibitor AP-1 SR 11302 in a dose of 1 mg/kg, respectively for 7 days following the TBI modelling. On the 7th day after the simulation of a moderate TBI in rat cerebral hemisphere tissue, the following signs of oxidative-nitrosative stress have been detected: increased production of superoxide radical anion by NADPH and NADH-dependent electron transport chains, increased activity of nitric oxide synthase (total and inducible), a decrease and impairment of the coupling of its constitutive isoform, growth in the concentration of peroxynitrite, the development of decompensated lipid peroxidation. The application of transcription factor inhibitors NF kappa B (PDTC) and AP-1 (SR 11302) significantly reduces the signs of oxidative-nitrosative stress in the tissue of rat cerebral hemispheres on the 7th day of the experiment, reduces the production of superoxide anion radical and the activity of nitric oxide synthase (total and inducible), improves the coupling of its constitutive isoform, limits the peroxynitrite concentration, and enhances the antioxidant potential.

Inhibition of effector B cells by ibrutinib in systemic sclerosis

ObjectiveSystemic sclerosis (SSc) is a connective tissue disease with a significant morbidity and reduced survival of patients. Effective treatment and clinical control of the disease remain challenging. In particular, the development of pulmonary and cardiac fibrosis and pulmonary hypertension are severe complications responsible for excessive mortality. Currently available treatment strategies only alleviate symptoms and slow disease progression. Here, we investigated the therapeutic potential of ibrutinib, a Bruton’s tyrosine kinase (BTK) inhibitor used in B cell malignancies, to alter B cell pathology in SSc in an in vitro model of autoimmunity.MethodsPBMCs and sorted B cells of 24 patients with SSc were used for functional testing after stimulation with hypomethylated DNA fragments (CpG) to induce an innate immune response. The effects of ibrutinib on cytokine production, autoantibody release, and activation of the transcription factor NFκB were evaluated.ResultsIbrutinib was able to reduce the production of the profibrotic hallmark cytokines IL-6 and TNF-α mainly from the effector B cell population in patients with SSc. Importantly, small doses of ibrutinib (0.1 μM) preserved the production of immunoregulatory IL-10 while effectively inhibiting hyperactivated, profibrotic effector B cells. In a flow cytometry analysis of phosphorylated NFκB, an important transcription factor in the induction of innate immune responses in B cells, significantly less activation was observed with ibrutinib treatment.ConclusionOur data could pave the avenue for a clinical application of ibrutinib for patients with SSc as a novel treatment option for the underlying pathogenetic immune imbalance contributing to disease onset and progression.

PUMA-mediated epithelial cell apoptosis promotes Helicobacter pylori infection-mediated gastritis

The molecular mechanism responsible for Helicobacter pylori infection-mediated gastritis and carcinogenesis is not yet clear. Increased evidence suggests that chronic gastritis and elevated gastric epithelial cell (GEC) apoptosis are crucial events during stomach carcinoma transformation. PUMA is a potent proapoptotic Bcl-2 protein and mediates acute tissue injury. In this study, we aimed to investigate the role of PUMA in GEC apoptosis and inflammation induced by H. pylori infection. As a result, we found that PUMA expression was elevated in gastritis tissues compared with uninvolved tissues, and it was correlated with the severity of apoptosis and gastritis. In mice, PUMA mRNA and protein were markedly induced in GECs upon induction of gastritis by H. pylori. PUMA-deficient mice were highly resistant to apoptosis and gastritis induced by H. pylori. Furthermore, the transcription factor NF-κB p65 binds to PUMA promoter to activate PUMA transcription after H. pylori infection. In addition, NF-κB inhibitor could rescue H. pylori-induced apoptosis and gastritis. Finally, H. pylori-induced activation of p-p65 and PUMA was mediated via Toll-like receptor 2 (TLR2) and blocked in TLR2 knockout mice. Taken together, these results verified the pro-inflammatory effect of PUMA in H. pylori-infected gastric tissue. Moreover, TLR2/NF-κB-mediated transcriptional regulation of PUMA contributes to the pathogenesis of H. pylori-infected gastritis.

Epigenetic regulation of IFITM1 expression in lipopolysaccharide-stimulated human mesenchymal stromal cells

BackgroundToll-like receptor 4 (TLR4) ligands such as lipopolysaccharide (LPS) activate immunomodulatory functions and the migration of human mesenchymal stromal cells (hMSCs). Here, we study the migration-related gene expression of LPS-stimulated hMSCs and the role and regulation of one of the upregulated genes, encoding the interferon-induced transmembrane protein 1 (IFITM1).MethodsGene expression profiles were determined by whole-transcriptome analysis (RNA-seq) and quantitative real-time PCR (qRT-PCR). Bioinformatics approaches were used to perform network and pathway analyses. The cell migration-related genes were identified with an in vitro wound healing assay. RNA interference (RNAi) was used to suppress the IFITM1 gene expression. The IFITM1 gene enhancer was analyzed by chromatin immunoprecipitation (ChIP) sequencing, ChIP-to-PCR, luciferase reporter assays, and qRT-PCR for enhancer RNAs (eRNAs).ResultsRNA-seq confirmed IFITM1 as an LPS-stimulated gene, and RNAi demonstrated its importance for the LPS-stimulated migration. LPS treatment increased the eRNA expression in enhancer region R2 (2 kb upstream) of the IFITM1 gene and enriched R2 for H3K27ac. Bioinformatics implicated the transcription factors NF-κB and IRF1, ChIP assays revealed their binding to R2, and chemical inhibition of NF-κB and RNAi directed against IRF1 prevented R2 eRNA and IFITM1 gene expression.ConclusionsIncreased expression of the IFITM1 gene is required for LPS-stimulated hMSC migration. We described several underlying changes in the IFITM1 gene enhancer, most notably the NF-κB-mediated activation of enhancer region R2.

Inhibitory Effects of Myrtucommuacetalone 1 (MCA-1) from Myrtus communis on Inflammatory Response in Mouse Macrophages.

(1) Introduction: Reactive oxygen species (ROS) and nitric oxide (NO) are key signaling molecules that play important roles in the progression of inflammatory disorders. The objective of this study was to explore the use of myrtucommuacetalone-1 (MCA-1), as a novel compound of natural origin and a potential anti-inflammatory agent. (2) Methodology: The anti-inflammatory potential of MCA-1, which was isolated from Myrthus communis Linn, was determined by assaying superoxide, hydrogen peroxide, and nitric oxide production in macrophages. Furthermore, the effects of the compound were analyzed via phosphorylation and translocation of the transcription factor NF kappa B, which is a key regulator of iNOS activation. The effect of MCA-1 on the inducible nitric oxide synthase (iNOS) enzyme was also examined using in silico docking studies. The anticancer potential for MCA-1 was evaluated with an MTT cytotoxic assay. (3) Results: In stimulated macrophages, MCA-1 inhibited superoxide production by 48%, hydrogen peroxide by 53%, and nitric oxide (NO) with an IC50 of <1 µg/mL. MCA-1 also showed a very strong binding pattern within the active site of the inducible nitric oxide synthase enzyme. Furthermore, 25 µg/mL of MCA-1 inhibited inducible nitric oxide synthase expression and abolished transcription factor (NFκB) phosphorylation and translocation to the nucleus. Cytotoxicity analyses of MCA-1 on 3T3 mouse fibroblasts, CC1 liver cell line, J774.2, macrophages and MDBK bovine kidney epithelial cell, yielded IC50 values of 6.53 ± 1.2, 4.6 ± 0.7, 5 ± 0.8, and 4.6 ± 0.7, µg/mL, respectively. (4) Conclusion: Our results suggest that MCA-1, a major phloroglucinol-type compound, shows strong anti-inflammatory activity and has a potential to be a leading therapeutic agent in the future.

DNA methylation of a NF-\u03baB binding site in the aquaporin 5 promoter impacts on mortality in sepsis

Altered aquaporin 5 (AQP5) expression in immune cells impacts on key mechanisms of inflammation and is associated with sepsis survival. Since epigenetic regulation via DNA methylation might contribute to a differential AQP5 expression in sepsis, we tested the hypotheses that DNA methylation of the AQP5 promotor (1) influences AQP5 expression, (2) is associated with the 30-day survival of septic patients, and (3) alters the nuclear transcription factor NF-κB binding. AQP5 mRNA expression was quantified by real-time PCR in whole blood samples of 135 septic patients. In silico computer analysis of the AQP5 promoter (nt-567 to nt-975) revealed seven putative inflammatory transcription factor binding sites and methylation of these sites was analyzed. Electrophoretic mobility shift assays were performed to assess the binding of nuclear NF-κB to the AQP5 promoter region nt-937. After adjustment for multiple testing, a greater methylation rate was found at cytosine site nt-937 in the AQP5 promoter linked to NF-κB binding in non-survivors compared to survivors (p = 0.002, padj = 0.014). This was associated with greater AQP5 mRNA expression in non-survivors (p = 0.037). Greater (≥16%) promoter methylation at nt-937 was also associated with an independently increased risk of death within 30 days (HR: 3.31; 95% CI: 1.54–6.23; p = 0.002). We detected a functionally important AQP5 promoter cytosine site (nt-937) linked to the binding of the inflammatorily acting nuclear transcription factor NF-κB, with increased methylation in sepsis non-survivors. Thus, nt-937 APQ5 promoter methylation, presumably related to NF-κB binding, is prognostically relevant in sepsis and demonstrates that epigenetic changes impact on sepsis outcome.

Loss-of-function phenotype of a PKC\u03b8T219A knockin mouse strain

BackgroundProtein kinase C θ has been established as an important signaling intermediate in T-effector-cell activation and survival pathways by controlling activity of the key transcription factors NF-κB and NFAT. Previous studies identified an activation-induced auto-phosphorylation site at Thr-219, located between the tandem C1 domains of the regulatory fragment in PKCθ, as a structural requirement for its correct membrane translocation and the subsequent transactivation of downstream signals leading to IL-2 production in a human T cell line.MethodsThe present work aimed to define the role of this phosphorylation switch on PKCθ in a physiological context through a homozygous T219A knockin mouse strain. T cell activation was analyzed by H3-thymidine uptake (proliferative response), qRT-PCR and luminex measurements (cytokine production). NFAT and NF-κB transactivation responses were estimated by Gel mobility shift and Alpha Screen assays. Frequencies of T cell subsets were analyzed by flow cytometry.ResultsDespite a normal T cell development, in vitro activated effector T cells clearly revealed a requirement of Thr-219 phosphorylation site on PKCθ for a transactivation of NF-κB and NFAT transcription factors and, subsequently, robust IL-2 and IFN-γ expression.ConclusionThis phenotype is reminiscent of the PKCθ knockout T cells, physiologically validating that this (p) Thr-219 auto-phosphorylation site indeed critically regulates PKCθ function in primary mouse T cells.

Cerium oxide nanoparticles improve liver regeneration after acetaminophen-induced liver injury and partial hepatectomy in rats

Background and aimsCerium oxide nanoparticles are effective scavengers of reactive oxygen species and have been proposed as a treatment for oxidative stress-related diseases. Consequently, we aimed to investigate the effect of these nanoparticles on hepatic regeneration after liver injury by partial hepatectomy and acetaminophen overdose.MethodsAll the in vitro experiments were performed in HepG2 cells. For the acetaminophen and partial hepatectomy experimental models, male Wistar rats were divided into three groups: (1) nanoparticles group, which received 0.1 mg/kg cerium nanoparticles i.v. twice a week for 2 weeks before 1 g/kg acetaminophen treatment, (2) N-acetyl-cysteine group, which received 300 mg/kg of N-acetyl-cysteine i.p. 1 h after APAP treatment and (3) partial hepatectomy group, which received the same nanoparticles treatment before partial hepatectomy. Each group was matched with vehicle-controlled rats.ResultsIn the partial hepatectomy model, rats treated with cerium oxide nanoparticles showed a significant increase in liver regeneration, compared with control rats. In the acetaminophen experimental model, nanoparticles and N-acetyl-cysteine treatments decreased early liver damage in hepatic tissue. However, only the effect of cerium oxide nanoparticles was associated with a significant increment in hepatocellular proliferation. This treatment also reduced stress markers and increased cell cycle progression in hepatocytes and the activation of the transcription factor NF-κB in vitro and in vivo.ConclusionsOur results demonstrate that the nanomaterial cerium oxide, besides their known antioxidant capacities, can enhance hepatocellular proliferation in experimental models of liver regeneration and drug-induced hepatotoxicity.

Investigation of the variants at the binding site of inflammatory transcription factor NF-\u03baB in patients with end-stage renal disease

BackgroundA chronic inflammatory state is a prominent feature in patients with end-stage renal disease (ESRD). Nuclear factor-kappa B (NF-κB) is a transcription factor that regulates the expression of genes involved in inflammation. Some genetic studies have demonstrated that the NF-κB genetic mutation could cause kidney injury and kidney disease progression. However, the association of a gene polymorphism in the transcription factor binding site of NF-κB with kidney disease is not clear.MethodsWe used the Taiwan Biobank database, the University of California, Santa Cruz, reference genome, and a chromatin immunoprecipitation sequencing database to find single nucleotide polymorphisms (SNPs) at potential binding sites of NF-κB. In addition, we performed a case–control study and genotyped 847 patients with ESRD and 846 healthy controls at Tri-Service General Hospital from 2015 to 2016. Furthermore, we used the ChIP assay to identify the binding activity of different genotypes and used Luciferase reporter assay to examine the function of the rs9395890 polymorphism.ResultThe results of biometric screening in the databases revealed 15 SNPs with the potential binding site of NF-κB. Genotype distributions of rs9395890 were significantly different in ESRD cases and healthy controls (P = 0.049). The ChIP assay revealed an approximately 1.49-fold enrichment of NF-κB of the variant type TT when compared to that of the wild-type GG in rs9395890 (P = 0.027; TT = 3.20 ± 0.16, GT = 2.81 ± 0.20, GG = 1.71 ± 0.18). The luciferase reporter assay showed that the NF-κB binding site activity in T allele was slightly higher than that in G allele, though it is not significant.ConclusionsOur findings indicate that rs9395890 is associated with susceptibility to ESRD in Taiwan population.

SIRT1 inhibits monocyte adhesion to the vascular endothelium by suppressing Mac-1 expression on monocytes

SIRT1 signaling pathways modulate vascular inflammation; however, the precise role of SIRT1 in monocyte adhesion to the vascular endothelium, a key event initiating vascular inflammation, is unclear. Thus, this study investigated the roles and molecular interaction of SIRT1 and TLR2 in regulating monocyte adhesion to the vascular endothelium. In vitro, both Mac-1 expression and the endothelial adhesion of THP-1 cells stimulated with Pam3CSK4, a TLR2 ligand, were markedly increased in association with a decreased expression of SIRT1. In THP-1 cells stimulated with Pam3CSK4, the promoter activity and expression of SIRT1 were decreased. The TLR2-dependent suppression of SIRT1 expression in THP-1 cells was mediated by the transcription factors NF-κB and CREB, suggesting that the TLR2-mediated NF-κB and CREB signaling downregulated SIRT1 expression in monocytes. In peripheral blood monocytes (PBMCs) isolated from SIRT1 transgenic (TG) mice and THP-1 cells treated with recombinant SIRT1, both the increased Mac-1 expression and endothelial adhesion induced by Pam3CSK4 were significantly attenuated. In addition, the en face immunohistochemical study showed a marked increase in monocyte adhesion to the aortic endothelium of WT mice treated with Pam3CSK4, which was significantly attenuated in Pam3CSK4-treated SIRT1 TG mice. Moreover, a greater number of atherosclerotic plaques formed in WT mice fed a high-fat diet than in SIRT1 TG mice, indicating a pivotal role for SIRT1 in preventing vascular inflammation. Based on these results, SIRT1 might be a potential target for researchers aiming to develop therapeutic interventions for vascular inflammation, including atherosclerosis.

In vivo, Extract from Withania somnifera Root Ameliorates Arthritis via Regulation of Key Immune Mediators of Inflammation in Experimental Model of Arthritis.

BackgroundRheumatoid Arthritis (RA) is a devastating disease characterized by continual addition of leukocytes and T cells within the articular cavity causing inflammation and cartilage destruction. Withania somnifera is one of the most precious medicinal herbs, reported to have antioxidant, anti‐inflammatory, and immunomodulatory properties.ObjectiveThe purpose of this study was to evaluate anti-inflammatory activity of aqueous extract of Withania somnifera roots (WSAq) in Collagen Induced Arthritic (CIA) rats.MethodsTo achieve this, we assessed the level of inflammatory cytokines such as Tumor Necrosis Factor (TNF)-α, IL-1β, IL-6 and IL-10 in CIA rats. Further, transcription factor, oxidative stress parameters and CD+8 expressions were also analyzed in CIA rats.ResultsArthritic rats showed a greater increase in the levels of pro inflammatory cytokines such as TNF-α, IL-1β, IL-6, transcription factor NF-κB and a decrease in IL-10 concentration than controls rats. Oral administration of WSAq at a dose of 300mg/kg.wt. (WSAq300) appreciably attenuated the production of these pro inflammatory cytokines. This anti-inflammatory activity of WSAq300 might be partly mediated through an increase in the secretion of IL-10 and inhibition of NF-κB activity. Further, arthritic rats also show increased oxidative stress as compared to control rats. This increased oxidative stress in the arthritic rats appears to be the outcome of both an activated pro-oxidant and a poor antioxidant defense system. Treatment with WSAq300 strongly ameliorates all these ROS parameters significantly to near normal. Additional, metalloproteinase MMP-8 levels were also measured and found to be increased in CIA rats, which after treatment with WSAq300 came down to near normal.ConclusionFrom the above results, it can be concluded that the use of WSAq300 may be a valuable supplement which can improve human arthritis.

USP15 inhibits multiple myeloma cell apoptosis through activating a feedback loop with the transcription factor NF-\u03baBp65

USP15 has been shown to stabilize transcription factors, to be amplified in many cancers and to mediate cancer cell survival. However, the underlying mechanism by which USP15 regulates multiple myeloma (MM) cell proliferation and apoptosis has not been established. Here, our results showed that USP15 mRNA expression was upregulated in MM patients. USP15 silencing induced MM cell proliferation inhibition, apoptosis, and the expression of nuclear and cytoplasmic NF-κBp65, while USP15 overexpression exhibited an inverse effect. Moreover, in vivo experiments indicated that USP15 silencing inhibited MM tumor growth and NF-κBp65 expression. PDTC treatment significantly inhibited USP15 overexpression-induced cell proliferation, apoptosis inhibition, and NF-κBp65 expression. USP15 overexpression promoted NF-κBp65 expression through inhibition of its ubiquitination, whereas NF-κBp65 promoted USP15 expression as a positive regulator. Taken together, the USP15-NF-κBp65 loop is involved in MM tumorigenesis and may be a potential therapeutic target for MM.

The collective dynamics of NF − κB in cellular ensembles

The transcription factor NF − κB is a crucial component in inflammatory signalling. Its dynamics is known to be oscillatory and has been extensively studied. Using a recently developed model of NF − κB regulation, we examine the collective dynamics of a network of NF − κB oscillators that are coupled exogenously by a common drive (in this case a periodically varying cytokine signal corresponding to the TNF molecule concentration). There is multistability owing to the overlapping of Arnol’d tongues in each of the oscillators, and thus the collective dynamics exhibit a variety of complex dynamical states. We also study the case of a globally (mean field) coupled network and observe that the ensemble can display global synchronisation, cluster synchronisation and splay states. In addition, there can be dynamical chimeras, namely coexisting synchronised and desynchronized clusters. The basins of attraction of these different collective states are studied and the parametric dependence in the basin uncertainty is examined.

Ipatasertib, a novel Akt inhibitor, induces transcription factor FoxO3a and NF-\u03baB directly regulates PUMA-dependent apoptosis

Colon cancer is one of the three common malignant tumors, with a lower survival rate. Ipatasertib, a novel highly selective ATP-competitive pan-Akt inhibitor, shows a strong antitumor effect in a variety of carcinoma, including colon cancer. However, there is a lack of knowledge about the precise underlying mechanism of clinical therapy for colon cancer. We conducted this study to determine that ipatasertib prevented colon cancer growth through PUMA-dependent apoptosis. Ipatasertib led to p53-independent PUMA activation by inhibiting Akt, thereby activating both FoxO3a and NF-κB synchronously that will directly bind to PUMA promoter, up-regulating PUMA transcription and Bax-mediated intrinsic mitochondrial apoptosis. Remarkably, Akt/FoxO3a/PUMA is the major pathway while Akt/NF-κB/PUMA is the secondary pathway of PUMA activation induced by ipatasertib in colon cancer. Knocking out PUMA eliminated ipatasertib-induced apoptosis both in vitro and in vivo (xenografts). Furthermore, PUMA is also indispensable in combinational therapies of ipatasertib with some conventional or novel drugs. Collectively, our study demonstrated that PUMA induction by FoxO3a and NF-κB is a critical step to suppress the growth of colon cancer under the therapy with ipatasertib, which provides some theoretical basis for clinical assessment.

Lactic acid Bacteria isolated from European badgers (Meles meles) reduce the viability and survival of Bacillus Calmette-Guerin (BCG) vaccine and influence the immune response to BCG in a human macrophage model

BackgroundBovine tuberculosis (bTB) caused by Mycobacterium bovis is the most serious endemic disease affecting livestock in the UK. The European badger (Meles meles) is the most important wildlife reservoir of bTB transmission to cattle, making eradication particularly difficult. In this respect, oral vaccination with the attenuated M. bovis vaccine Bacillus Calmette-Guerin (BCG) has been suggested as a wide-scale intervention to reduce bTB infection in badgers. However, experimental studies show variable protection. Among the possibilities for this variation is that the resident gut bacteria may influence the success of oral vaccination in badgers; either through competitive exclusion and/or inhibition, or via effects on the host immune system. In order to explore this possibility, we have tested whether typical gut commensals such as Lactic Acid Bacteria (LAB) have the capacity to impact on the viability and survival rate of BCG and to modulate the immune response to BCG using an in vitro model.ResultsTwelve LAB isolated from badger faeces displayed inhibitory activity to BCG that was species-dependent. Weissella had a bacteriostatic effect, whereas isolates of enterococci, lactobacilli and pediococci had a more bactericidal activity. Furthermore, BCG-induced activation of the pro-inflammatory transcription factor NF-κB in human THP-1 macrophages was modulated by LAB in a strain-dependent manner. Most pediococci enhanced NF-κB activation but one strain had the opposite effect. Interestingly, isolates of enterococci, lactobacilli and weissella had different effects as immunomodulators of BCG-induced macrophage responses as some had no significant influence on NF-κB activation, but others increased it significantly.ConclusionsOur in vitro results show that LAB isolated from badgers exhibit significant inhibitory activity against BCG and influence the immune activation mediated by BCG in a human macrophage assay. These findings suggest that gut commensal bacteria could play a role in influencing the outcome of oral BCG vaccination. Inactivated cells of LAB, or LAB that are bacteriostatic but have a synergistic immunostimulatory effect with BCG, could be potential adjuvants to be used for oral vaccination in badgers. Further work is needed to take into account the complex nature of the gut microbiome, specific immunity of the badger and the in vivo context.

Endoplasmic reticulum stress in perivascular adipose tissue promotes destabilization of atherosclerotic plaque by regulating GM-CSF paracrine

BackgroundPerivascular adipose tissue (PVAT) accelerates plaque progression and increases cardiovascular risk. We tested the hypothesis that PVAT contributed to plaque vulnerability and investigated whether endoplasmic reticulum stress (ER stress) in PVAT played an important role in vulnerable plaque.MethodsWe transplanted thoracic aortic PVAT or subcutaneous adipose tissue as a control, from donor mice to carotid arteries of recipient apolipoprotein E deficient (apoE−/−) mice after removing carotid artery collar placed for 6 weeks. Two weeks after transplantation, ER stress inhibitor 4-phenyl butyric acid (4-PBA) was locally administrated to the transplanted PVAT and then animals were euthanized after 4 weeks. Immunohistochemistry was performed to quantify plaque composition and neovascularization. Mouse angiogenesis antibody array kit was used to test the angiogenic factors produced by transplanted adipose tissue. In vitro tube formation assay, scratch wound migration assay and mouse aortic ring assay were used to assess the angiogenic capacity of supernatant of transplanted PVAT.ResultsUltrastructural detection by transmission electron microscopy showed transplanted PVAT was a mixed population of white and brown adipocytes with abundant mitochondria. Transplanted PVAT increased the intraplaque macrophage infiltration, lipid core, intimal and vasa vasorum neovascularization and MMP2/9 expression in plaque while decreased smooth muscle cells and collagen in atherosclerotic plaque, which were restored by local 4-PBA-treatment. Antibody array analysis showed that 4-PBA reduced several angiogenic factors [Granulocyte Macrophage Colony Stimulating Factor (GM-CSF), MCP-1, IL-6] secreted by PVAT. Besides, conditioned medium from 4-PBA treated-PVAT inhibited tube formation and migration capacity of endothelial cells and ex vivo mouse aortic ring angiogenesis compared to conditioned medium from transplanted PVAT. mRNA expression and protein levels of GM-CSF were markedly elevated in adipocytes under ER stress which would be suppressed by 4-PBA. In addition, ER stress enhanced NF-κB binding to the promoter of the mouse GM-CSF gene in adipocytes confirmed by Chromatin immunoprecipitation analyses.ConclusionsOur findings demonstrate that ER stress in PVAT destabilizes atherosclerotic plaque, in part through increasing GM-CSF paracrine via transcription factor NF-κB.

Subunit-Specific Role of NF-κB in Cancer.

The transcription factor NF-κB is a key player in inflammation, cancer development, and progression. NF-κB stimulates cell proliferation, prevents apoptosis, and could promote tumor angiogenesis as well as metastasis. Extending the commonly accepted role of NF-κB in cancer formation and progression, different NF-κB subunits have been shown to be active and of particular importance in distinct types of cancer. Here, we summarize overexpression data of the NF-κB subunits RELA, RELB, and c-REL (referring to the v-REL, which is the oncogene of Reticuloendotheliosis virus strain T) as well as of their upstream kinase inhibitor, namely inhibitor of κB kinases (IKK), in different human cancers, assessed by database mining. These data argue against a universal mechanism of cancer-mediated activation of NF-κB, and suggest a much more elaborated mode of NF-κB regulation, indicating a tumor type-specific upregulation of the NF-κB subunits. We further discuss recent findings showing the diverse roles of NF-κB signaling in cancer development and metastasis in a subunit-specific manner, emphasizing their specific transcriptional activity and the role of autoregulation. While non-canonical NF-κB RELB signaling is described to be mostly present in hematological cancers, solid cancers reveal constitutive canonical NF-κB RELA or c-REL activity. Providing a linkage to cancer therapy, we discuss the recently described pivotal role of NF-κB c-REL in regulating cancer-targeting immune responses. In addition, current strategies and ongoing clinical trials are summarized, which utilize genome editing or drugs to inhibit the NF-κB subunits for cancer treatment.