lactoferrin activates nf-kb: Topics by Science.gov

Sample records for lactoferrin activates nf-kb

Lactoferrin from Camelus dromedarius Inhibits Nuclear Transcription Factor-kappa B Activation, Cyclooxygenase-2 Expression and Prostaglandin E2 Production in Stimulated Human Chondrocytes

PubMed Central

Rasheed, Naila; Alghasham, Abdullah; Rasheed, Zafar

2016-01-01

Background: Osteoarthritis (OA) is a progressive joint disorder, which remains the leading cause of chronic disability in aged people. Nuclear factor-kappa B (NF)-κB is a major cellular event in OA and its activation by interleukin-1β (IL-1β) plays a critical role in cartilage breakdown in these patients. Objective: In this study, we examined the effect of lactoferrin on NF-κB activation, cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) production in stimulated human articular chondrocytes. Materials and Methods: Human chondrocytes were derived from OA articular cartilage and treated with camel lactoferrin and then stimulated with IL-1β. Gene expression was determined by TaqMan assays and protein expression was studied by Western immunoblotting. NF-κB activity and PGE2 levels were determined by ELISA based assays. NF-κB activity was also determined by treatment of chondrocytes with NF-κB specific inhibitor Bay 11–7082. Results: Lactoferrin inhibited IL-1β-induced activation and nuclear translocation of NF-κB p65 in human OA chondrocytes. Lactoferrin also inhibited mRNA/protein expression of COX-2 and production of PGE2. Moreover, Bay 11–7082 also inhibited IL-1β-induced expression of COX-2 and production of PGE2. The inhibitory effect of lactoferrin on the IL-1β induced expression of COX-2 or production of PGE2 was mediated at least in part via suppression of NF-κB activation. Conclusions: Our data determine camel lactoferrin as a novel inhibitor of IL-1β-induced activation of NF-κB signaling events and production of cartilage-degrading molecule PGE2 via inhibition of COX-2 expressions. These results may have important implications for the development of novel therapeutic strategies for the prevention/treatment of OA and other degenerative/inflammatory diseases. SUMMARY Lactoferrin shows anti-arthritic activity in IL-1β stimulated primary human chondrocytes.Lactoferrin inhibits IL-1β-induced NF-κB activation.Lactoferrin inhibits
Lactoferrin from Camelus dromedarius Inhibits Nuclear Transcription Factor-kappa B Activation, Cyclooxygenase-2 Expression and Prostaglandin E2 Production in Stimulated Human Chondrocytes.

PubMed

Rasheed, Naila; Alghasham, Abdullah; Rasheed, Zafar

2016-01-01

Osteoarthritis (OA) is a progressive joint disorder, which remains the leading cause of chronic disability in aged people. Nuclear factor-kappa B (NF)-κB is a major cellular event in OA and its activation by interleukin-1β (IL-1β) plays a critical role in cartilage breakdown in these patients. In this study, we examined the effect of lactoferrin on NF-κB activation, cyclooxygenase-2 (COX-2) expression and prostaglandin E2 (PGE2) production in stimulated human articular chondrocytes. Human chondrocytes were derived from OA articular cartilage and treated with camel lactoferrin and then stimulated with IL-1β. Gene expression was determined by TaqMan assays and protein expression was studied by Western immunoblotting. NF-κB activity and PGE2 levels were determined by ELISA based assays. NF-κB activity was also determined by treatment of chondrocytes with NF-κB specific inhibitor Bay 11-7082. Lactoferrin inhibited IL-1β-induced activation and nuclear translocation of NF-κB p65 in human OA chondrocytes. Lactoferrin also inhibited mRNA/protein expression of COX-2 and production of PGE2. Moreover, Bay 11-7082 also inhibited IL-1β-induced expression of COX-2 and production of PGE2. The inhibitory effect of lactoferrin on the IL-1β induced expression of COX-2 or production of PGE2 was mediated at least in part via suppression of NF-κB activation. Our data determine camel lactoferrin as a novel inhibitor of IL-1β-induced activation of NF-κB signaling events and production of cartilage-degrading molecule PGE2 via inhibition of COX-2 expressions. These results may have important implications for the development of novel therapeutic strategies for the prevention/treatment of OA and other degenerative/inflammatory diseases. Lactoferrin shows anti-arthritic activity in IL-1β stimulated primary human chondrocytes.Lactoferrin inhibits IL-1β-induced NF-κB activation.Lactoferrin inhibits production of cartilage degrading PGE2 via inhibition of COX-2 expression
A Role for the NF-kb/Rel Transcription Factors in Human Breast Cancer

DTIC Science & Technology

1998-07-01

binding proteins present in a series of nuclear extracts from cell lines and from breast tumor tissues as well as normal mammary epithelium. Finally, we...RelA is nuclear in several examples. Our recent data on nuclear extracts of breast tumors shows that there is a significant increase in NF-KB binding...Figure 2 in the appendix). Additionally, immunoblotting of nuclear extracts versus adjacent tissue controls showed that NF-KB p50, p52 and c-Rel were
Role of TGR-B1-Mediated Down Regulation of NF-kB/Rel Activity During Growth Arrest of Breast Cancer Cells

DTIC Science & Technology

2001-05-01

gallate ( EGCG ), which has been shown to inhibit the induction of NF-KB and growth of breast cancer cell lines in vitro. EGCG reduced NF-KB levels in the...demonstrated activation of NF-KB is induced upon over-expression of Her-2/neu. Thus, studies were initiated with green tea pholyphenol, epigallocatechin -3...NF639 cell line derived from an MMTV-Her-2/neu mouse tumor. NF639 clonal isolates resistant to EGCG appear to display elevated levels of NF-KB. Overall
Molecular characterization of the breakpoints of a 12-kb deletion in the NF1 gene in a family showing germ-line mosaicism.

PubMed Central

Lázaro, C; Gaona, A; Lynch, M; Kruyer, H; Ravella, A; Estivill, X

1995-01-01

Neurofibromatosis type 1 (NF1) is caused by deletions, insertions, translocations, and point mutations in the NF1 gene, which spans 350 kb on the long arm of human chromosome 17. Although several point mutations have been described, large molecular abnormalities have rarely been characterized in detail. We describe here the molecular breakpoints of a 12-kb deletion of the NF1 gene, which is responsible for the NF1 phenotype in a kindred with two children affected because of germline mosaicism in the unaffected father, who has the mutation in 10% of his spermatozoa. The mutation spans introns 31-39, removing 12,021 nt and inserting 30 bp, of which 19 bp are a direct repetition of a sequence located in intron 31, just 4 bp before the 5' breakpoint. The 5' and 3' breakpoints contain the sequence TATTTTA, which could be involved in the generation of the deletion. The most plausible explanation for the mechanism involved in the generation of this 12-kb deletion is homologous/nonhomologous recombination. Since sperm of the father does not contain the corresponding insertion of the 12-kb deleted sequence, this deletion could have occurred within the NF1 chromosome through loop formation. RNA from lymphocytes of one of the NF1 patients showed similar levels of the mutated and normal transcripts, suggesting that the NF1-mRNA from mutations causing frame shifts of the reading frame or stop codons in this gene is not degraded during its processing. The mutation was not detected in fresh lymphocytes from the unaffected father by PCR analysis, supporting the case for true germ-line mosaicism. Images Figure 1 Figure 3 PMID:7485153
Antimicrobial activity of immobilized lactoferrin and lactoferricin.

PubMed

Chen, Renxun; Cole, Nerida; Dutta, Debarun; Kumar, Naresh; Willcox, Mark D P

2017-11-01

Lactoferrin and lactoferricin were immobilized on glass surfaces via two linkers, 4-azidobenzoic acid (ABA) or 4-fluoro-3-nitrophenyl azide (FNA). The resulting surfaces were characterized by X-ray photoelectron spectroscopy (XPS) and contact angle measurements. The antimicrobial activity of the surfaces was determined using Pseudomonas aeruginosa and Staphylococcus aureus strains by fluorescence microscopy. Lactoferrin and lactoferricin immobilization was confirmed by XPS showing significant increases (p < 0.05) in nitrogen on the glass surface. The immobilization of both proteins slightly increased the overall hydrophobicity of the glass. Both lactoferrin and lactoferricin immobilized on glass significantly (p < 0.05) reduced the numbers of viable bacterial cells adherent to the glass. For P. aeruginosa, the immobilized proteins consistently increased the percentage of dead cells compared to the total cells adherent to the glass surfaces (p < 0.03). Lactoferrin and lactoferricin were successfully immobilized on glass surfaces and showed promising antimicrobial activity against pathogenic bacteria. © 2016 Wiley Periodicals, Inc. J Biomed Mater Res Part B: Appl Biomater, 105B: 2612-2617, 2017. © 2016 Wiley Periodicals, Inc.
Exploring a Link Between NF-KB and G2/M Cell Cycle Arrest in Breast Cancer Cells

DTIC Science & Technology

2005-04-01

studies with esophageal squamous cell carcinom a lines have shown that IR induced p21waf1/ ciP ’ and a G2 cell cycle arrest that could als o be...i AD Award Number : DAMD17-02-1-062 3 TITLE : Exploring a Link Between NF-KB and G 2 /M Cell Cycle Arres t in Breast Cancer Cell s PRINCIPAL...Mar 2005 ) 4 . TITLE AND SUBTITL E Exploring a Link Between NF-kB and G 2 /M Cell Cycle Arres t in Breast Cancer Cells 5. FUND/NG NUMBERS DAMD17-02-1
Studies on anticancer activities of lactoferrin and lactoferricin.

PubMed

Yin, Cui Ming; Wong, Jack Ho; Xia, Jiang; Ng, Tzi Bun

2013-09-01

This review mainly summarizes results of recent studies on the anticancer activity of the multifunctional protein lactoferrin (Lf) and its derived peptide lactoferricin (Lfcin). The basic information on Lf and Lfcin, such as their sources, structures, and biological properties which favor their antitumor activity is introduced. The major anticancer mechanisms of Lf and Lfcin including cell cycle arrest, apoptosis, anti-angiogenesis, antimetastasis, immune modulation and necrosis are discussed. Other information from in vivo studies employing a mouse model is also provided. In addition, the roles of talatoferrin and delta lactoferrin, as well as improvement in drug delivery will be covered.
PPARγ and NF-κB regulate the gene promoter activity of their shared repressor, TNIP1

PubMed Central

Gurevich, Igor; Zhang, Carmen; Encarnacao, Priscilla C.; Struzynski, Charles P.; Livings, Sarah E.; Aneskievich, Brian J.

2011-01-01

Human TNFAIP3 interacting protein 1 (TNIP1) has diverse functions including support of HIV replication through its interaction with viral Nef and matrix proteins, reduction of TNFα-induced signaling through its interaction with NF-κB pathway proteins, and corepression of agonist-bound retinoic acid receptors and peroxisome proliferator-activated receptors (PPAR). The wide tissue distribution of TNIP1 provides the opportunity to influence numerous cellular responses in these roles and defining control of TNIP1 expression would be central to improved understanding of its impact on cell function. We cloned 6kb of the human TNIP1 promoter and performed predictive and functional analyses to identify regulatory elements. The promoter region proximal to the transcription start site is GC-rich without a recognizable TATA box. In contrast to this proximal ~500bp region, 6kb of the promoter increased reporter construct constitutive activity over five-fold. Throughout the 6kb length, in silico analysis identified several potential binding sites for both constitutive and inducible transcription factors; among the latter were candidate NF-κB binding sequences and peroxisome proliferator response elements (PPREs). We tested NF-κB and PPAR regulation of the endogenous TNIP1 gene and cloned promoter by expression studies, electrophoretic mobility shift assays, and chromatin immunoprecipitations. We validated NF-κB sites in the TNIP1 promoter proximal and distal regions as well as one PPRE in the distal region. The ultimate control of the TNIP1 promoter is likely to be a combination of constitutive transcription factors and those subject to activation such as NF-κB and PPAR. PMID:22001530
Quality control of commercial bovine lactoferrin.

PubMed

Wakabayashi, Hiroyuki; Yamauchi, Koji; Abe, Fumiaki

2018-06-01

Herein we review commercial bovine lactoferrin quality issues by describing an example of industrial production, the current status of global quality standardization, and quality-activity concerns for further discussion. Morinaga Milk Industry has been industrially producing bovine lactoferrin in Milei GmbH, Germany, since 1989. We delineate its production and quality as an example of safe and high-quality manufacturing. Currently, global standardization in the quality of bovine lactoferrin is progressing through Novel Food and GRAS in the EU and USA, respectively. Novel Food was applied or notified to seven lactoferrin manufacturers and GRAS was notified to three manufacturers, two of which are for infant use and one is for adult use, by the end of 2017. The specifications of these regulations are relatively high, including more than 95% lactoferrin purity in protein, which means that such companies can supply relatively high-grade lactoferrin. There appear to be several concerns regarding lactoferrin quality affecting activities, including contamination of lipopolysaccharide (LPS) and angiogenin, purity, and degradation of lactoferrin sample. Although LPS is immunologically toxic when invading the body, it is distributed normally in foods and the gut. However, an industrial lactoferrin sample may contain LPS at a maximum LPS/lactoferrin molecule ratio = 1/1724, which means 99.9% of the lactoferrin molecule is LPS-free. It is difficult to speculate that LPS contained in a lactoferrin sample affects its activities. Finally in order to achieve good and reproducible results, we make proposals to researchers a use of high-grade lactoferrin, careful storage, and indication the manufacturers' names and specifications in the paper.
Lactoferrin from Milk: Nutraceutical and Pharmacological Properties

PubMed Central

Giansanti, Francesco; Panella, Gloria; Leboffe, Loris; Antonini, Giovanni

2016-01-01

Lactoferrin is an iron-binding protein present in large quantities in colostrum and in breast milk, in external secretions and in polymorphonuclear leukocytes. Lactoferrin’s main function is non-immune protection. Among several protective activities shown by lactoferrin, those displayed by orally administered lactoferrin are: (i) antimicrobial activity, which has been presumed due to iron deprivation, but more recently attributed also to a specific interaction with the bacterial cell wall and extended to viruses and parasites; (ii) immunomodulatory activity, with a direct effect on the development of the immune system in the newborn, together with a specific antinflammatory effects; (iii) a more recently discovered anticancer activity. It is worth noting that most of the protective activities of lactoferrin have been found, sometimes to a greater extent, also in peptides derived from limited proteolysis of lactoferrin that could be generated after lactoferrin ingestion. Lactoferrin could therefore be considered an ideal nutraceutic product because of its relatively cheap production from bovine milk and of its widely recognized tolerance after ingestion, along with its well demonstrated protective activities. The most important protective activities shown by orally administered bovine lactoferrin are reviewed in this article. PMID:27690059
The dominant expression of functional human lactoferrin in transgenic cloned goats using a hybrid lactoferrin expression construct.

PubMed

Yu, Huiqing; Chen, Jianquan; Sun, Wei; Liu, Siguo; Zhang, Aimin; Xu, Xujun; Wang, Xuebin; He, Zhuzi; Liu, Guohui; Cheng, Guoxiang

2012-10-31

Human Lactoferrin (hLF) is an iron-binding protein with multiple physiological functions. As the availability of natural hLF is limited, alternative means of producing this biopharmaceutical protein have been extensively studied. Here we report on the dominant expression of recombinant human lactoferrin (rhLF) in transgenic cloned goats using a novel optimised construct made by fusing a 3.3 kb hLF minigene to the regulatory elements of the β-casein gene. The transgenic goat produced more than 30 mg/ml rhLF in its milk, and rhLF expression was stable during the entire lactation cycle. The rhLF purification efficiency from whole goat milk is approximately 70%, and its purity is above 98%. Compared with natural hLF, the rhLF from transgenic goats has similar biological characteristics including molecular mass, N-terminal sequence, isoelectric point, immunoreactivity and digestive stability. More importantly, the purified rhLF showed specific anti-tumour activity in the mouse model of melanoma experimental metastasis. Therefore, our study shows that the large-scale production of functional rhLF in transgenic goat milk could be an economical and promising source of human therapeutic use in the future. Copyright © 2012. Published by Elsevier B.V.
Lactoferrin – A Novel Bone Growth Factor

PubMed Central

Naot, Dorit; Grey, Andrew; Reid, Ian R; Cornish, Jillian

2005-01-01

Lactoferrin is an iron-binding glycoprotein that belongs to the transferrin family. It is present in breast milk, in epithelial secretions, and in the secondary granules of neutrophils. In healthy subjects lactoferrin circulates at concentrations of 2–7 x 10−6 g/ml. Lactoferrin is a pleiotropic factor with potent antimicrobial and immunomodulatory activities. Recently, we have shown that lactoferrin can also promote bone growth. At physiological concentrations, lactoferrin potently stimulates the proliferation and differentiation of primary osteoblasts and also acts as a survival factor inhibiting apoptosis induced by serum withdrawal. Lactoferrin also affects osteoclast formation and, in murine bone marrow culture, lactoferrin potently inhibits osteoclastogenesis. In vivo, local injection of lactoferrin above the hemicalvaria of adult mice results in substantial increases in the dynamic histomorphometric indices of bone formation and bone area. The mitogenic effect of lactoferrin in osteoblast-like cells is mediated mainly through LRP1, a member of the family of low-density lipoprotein receptor-related proteins that are primarily known as endocytic receptors. Using confocal laser scanning microscopy, we demonstrated that fluorescently labeled lactoferrin is endocytosed and can be visualized in the cytoplasm of primary osteoblastic cells. Lactoferrin also induces activation of p42/44 MAPK signaling in primary osteoblasts, but the two pathways seem to operate independently as activation of MAPK signaling, but not endocytosis, is necessary for the mitogenic effect of lactoferrin. We conclude that lactoferrin may have a physiological role in bone growth and healing, and a potential therapeutic role as an anabolic factor in osteoporosis. PMID:16012127
Iron uptake and increased intracellular enzyme activity follow host lactoferrin binding by Trichomonas vaginalis receptors

PubMed Central

1984-01-01

Lactoferrin acquisition and iron uptake by pathogenic Trichomonas vaginalis was examined. Saturation binding kinetics were obtained for trichomonads using increasing amounts of radioiodinated lactoferrin, while no significant binding by transferrin under similar conditions was achieved. Only unlabeled lactoferrin successfully and stoichiometrically competed with 125I-labeled lactoferrin binding. Time course studies showed maximal lactoferrin binding by 30 min at 37 degrees C. Data suggest no internalization of bound lactoferrin. The accumulation of radioactivity in supernatants after incubation of T. vaginalis with 125I-labeled lactoferrin and washing in PBS suggested the presence of low affinity sites for this host macromolecule. Scatchard analysis indicated the presence of 90,000 receptors per trichomonad with an apparent Kd of 1.0 microM. Two trichomonad lactoferrin binding proteins were identified by affinity chromatography and immunoprecipitation of receptor-ligand complexes. A 30-fold accumulation of iron was achieved using 59Fe-lactoferrin when compared to the steady state concentration of bound lactoferrin. The activity of pyruvate/ferrodoxin oxidoreductase, an enzyme involved in trichomonal energy metabolism, increased more than sixfold following exposure of the parasites to lactoferrin, demonstrating a biologic response to the receptor-mediated binding of lactoferrin. These data suggest that T. vaginalis possesses specific receptors for biologically relevant host proteins and that these receptors contribute to the metabolic processes of the parasites. PMID:6088662
Lactoferrin-derived Peptides Active towards Influenza: Identification of Three Potent Tetrapeptide Inhibitors.

PubMed

Scala, Maria Carmina; Sala, Marina; Pietrantoni, Agostina; Spensiero, Antonia; Di Micco, Simone; Agamennone, Mariangela; Bertamino, Alessia; Novellino, Ettore; Bifulco, Giuseppe; Gomez-Monterrey, Isabel M; Superti, Fabiana; Campiglia, Pietro

2017-09-06

Bovine lactoferrin is a biglobular multifunctional iron binding glycoprotein that plays an important role in innate immunity against infections. We have previously demonstrated that selected peptides from bovine lactoferrin C-lobe are able to prevent both Influenza virus hemagglutination and cell infection. To deeper investigate the ability of lactoferrin derived peptides to inhibit Influenza virus infection, in this study we identified new bovine lactoferrin C-lobe derived sequences and corresponding synthetic peptides were synthesized and assayed to check their ability to prevent viral hemagglutination and infection. We identified three tetrapeptides endowed with broad anti-Influenza activity and able to inhibit viral infection in a concentration range femto- to picomolar. Our data indicate that these peptides may constitute a non-toxic tool for potential applications as anti-Influenza therapeutics.
Kinetics of Competitive Adsorption between Lysozyme and Lactoferrin on Silicone Hydrogel Contact Lenses and the Effect on Lysozyme Activity.

PubMed

Hall, Brad; Jones, Lyndon; Forrest, James A

2015-05-01

To determine the effect of competitive adsorption between lysozyme and lactoferrin on silicone hydrogel contact lenses and the effect on lysozyme activity. Three commercially available silicone hydrogel contact lens materials (senofilcon A, lotrafilcon B and balafilcon A) were examined, for time points ranging from 10 s to 2 h. Total protein deposition was determined by I(125) radiolabeling of lysozyme and lactoferrin, while the activity of lysozyme was determined by a micrococcal activity assay. Senofilcon A and balafilcon A did not show any relevant competitive adsorption between lysozyme and lactoferrin. Lotrafilcon B showed reduced protein deposition due to competitive adsorption for lactoferrin at all time points and lysozyme after 7.5 min. Co-adsorption of lactoferrin and lysozyme decreased the activity of lysozyme in solution for senofilcon A and lotrafilcon B, but co-adsorption had no effect on the surface activity of lysozyme for all lens types investigated. Competition between lysozyme and lactoferrin is material specific. Co-adsorption of lysozyme and lactoferrin does not affect the activity of surface-bound lysozyme but can reduce the activity of subsequently desorbed lysozyme.
Antibacterial activity of recombinant human lactoferrin from rice: effect of heat treatment.

PubMed

Conesa, Celia; Rota, Carmen; Castillo, Eduardo; Pérez, María-Dolores; Calvo, Miguel; Sánchez, Lourdes

2009-06-01

The antibacterial activity of recombinant human lactoferrin from rice (rhLF) compared with that of human milk lactoferrin (hLF) was evaluated against Escherichia coli O157:H7, Salmonella Enteritidis and Listeria monocytogenes. The hydrolysates of rhLF and hLF were found to be more active than native proteins against E. coli O157:H7, and their activity was independent of their iron saturation. The effect of different heat treatments on the antibacterial activity of apo-rhLF was studied and compared with hLF. We observed that an HTST pasteurization treatment did not affect the antimicrobial activity of lactoferrin against the pathogens studied. Furthermore, the activity of apo-rhLF and hLF against E. coli O157:H7 and L. monocytogenes in UHT milk and whey was assayed, finding a decrease in the number of bacteria, although lower than that observed in a broth medium. This study shows the similar antibacterial activity of rhLF and hLF which is important in order to consider the addition of rhLF as a supplement in special products.
Advances in lactoferrin research concerning bovine mastitis.

PubMed

Shimazaki, Kei-Ichi; Kawai, Kazuhiro

2017-02-01

Lactoferrin is a multifunctional, iron-binding glycoprotein found in milk and other exocrine secretions. Lactoferrin in milk plays vital roles in the healthy development of newborn mammals, and is also an innate resistance factor involved in the prevention of mammary gland infection by microorganisms. Inflammation of the udder because of bacterial infection is referred to as mastitis. There have been many investigations into the relationships between lactoferrin and mastitis, which fall into several categories. The main categories are fluctuations in the lactoferrin concentration of milk, lactoferrin activity against mastitis pathogens, elucidation of the processes underlying the onset of mastitis, participation of lactoferrin in the immune system, and utilization of lactoferrin in mastitis treatment and prevention. This minireview describes lactoferrin research concerning bovine mastitis. In the 1970s, many researchers reported that the lactoferrin concentration fluctuates in milk from cows with mastitis. From the late 1980s, many studies clarified the infection-defense mechanism in the udder and the contribution of lactoferrin to the immune system. After the year 2000, the processes underlying the onset of mastitis were elucidated in vivo and in vitro, and lactoferrin was applied for the treatment and prevention of mastitis.
Amoebicidal Activity of Milk, Apo-lactoferrin, sIgA and Lysozyme

PubMed Central

León-Sicairos, Nidia; López-Soto, Fernando; Reyes-López, Magda; Godínez-Vargas, Delfino; Ordaz-Pichardo, Cynthia; de la Garza, Mireya

2006-01-01

Objectives: To identify amoebicidal components in human milk and the effect of iron on the amoebicidal activity. Design: Investigation in axenic cultures of Entamoeba histolytica trophozoites. Methods: Amoebas were treated with 5%–20% of human, bovine and swine milk, with 10% of human milk fractions (i.e., casein, proteins except casein and fat) or with 1 mg/ml of human milk apo-lactoferrin, human secretory immunoglobulin type A (sIgA) and chicken egg-white lysozyme (i.e., purified proteins). Milk proteins were detected using immunoblot. Confocal microscopy was used to define the interaction of milk proteins (100 μM each) and amoebas. Experiments were done at least three times in triplicate, and mean and standard deviations were calculated. Results: Human and bovine milk were amoebicidal showing a concentration-dependent effect. The amoebicidal effect was increased in the absence of iron. Milk protein fractions, with the exception of casein, were the components responsible for the amoebicidal activity found. Apo-lactoferrin, sIgA and lysozyme were identified in the amoebicidal milk protein fraction. Apo-lactoferrin showed the major amoebicidal effect. These proteins, either alone or in combination, showed a killing effect on the trophozoites. They bound to the amoebic membrane causing cell rounding, lipid disruption and damage. Conclusions: Milk proteins such as apo-lactoferrin, sIgA and lysozyme are able to kill Entamoeba histolytica trophozoites. This study confirms the importance of feeding breast milk to newborns. PMID:16809402
Lactoferrin for prevention of common viral infections.

PubMed

Wakabayashi, Hiroyuki; Oda, Hirotsugu; Yamauchi, Koji; Abe, Fumiaki

2014-11-01

Although lactoferrin has many biological functions, the host-protective effects against pathogenic microorganisms including bacteria, fungi, and viruses are regarded as one of the most important. Here, we review research on the protective role of lactoferrin administration against common viral infections. Many studies have shown the in vitro antiviral activity of lactoferrin against viral pathogens that cause common infections such as the common cold, influenza, gastroenteritis, summer cold, and herpes, where lactoferrin inhibits mainly viral attachment to the target cells. Recently, studies indicating the in vivo protective effects of lactoferrin by oral administration against common viral infections have been increasing. For instance, norovirus is an extremely important emerging human pathogen that causes a majority of gastroenteritis outbreaks worldwide that may be a target candidate for lactoferrin. Lactoferrin consumption reduced the incidence of noroviral gastroenteritis in children and a similar effect was observed in a wide range of ages in a preliminary survey. A recent in vitro study reported that lactoferrin inhibits both cellular attachment of the murine norovirus, a virus closely-related to the human norovirus, and viral replication in the cells by inducing antiviral cytokines interferon (IFN)-α/β. Lactoferrin administration also enhances NK cell activity and Th1 cytokine responses, which lead to protection against viral infections. In conclusion, lactoferrin consumption may protect the host from viral infections through inhibiting the attachment of a virus to the cells, replication of the virus in the cells, and enhancement of systemic immune functions. Copyright © 2014 Japanese Society of Chemotherapy and The Japanese Association for Infectious Diseases. Published by Elsevier Ltd. All rights reserved.

Titanium Surface Roughing Treatments contribute to Higher Interaction with Salivary Proteins MG2 and Lactoferrin.

PubMed

Cavalcanti, Yuri Wanderley; Soare, Rodrigo Villamarim; Leite Assis, Marina Araújo; Zenóbio, Elton Gonçalves; Girundi, Francisco Mauro da Silva

2015-02-01

Some surface treatments performed on titanium can alter the composition of salivary pellicle formed on this abiotic surface. Such treatments modify the titanium's surface properties and can promote higher adsorption of proteins, which allow better integration of titanium to the biotic system. This study aimed to evaluate the interactions between salivary proteins and titanium disks with different surface treatments. Machined titanium disks (n = 48) were divided into four experimental groups (n = 12), according to their surface treatments: surface polishing (SP); acid etching (A); spot-blasting plus acid etching (SB-A); spot-blasting followed by acid etching and nano-functionalization (SB-A-NF). Titanium surfaces were characterized by surface roughness and scanning electron microscopy (SEM). Specimens were incubated with human saliva extracted from submandibular and sublingual glands. Total salivary protein adsorbed to titanium was quantified and samples were submitted to western blotting for mucin glycoprotein 2 (MG2) and lactoferrin identification. Surface roughness was statistically higher for SB-A and SB-A-NF groups. Scanning electron microscopy images confirmed that titanium surface treatments increased surface roughness with higher number of porous and scratches for SB-A and SB-A-NF groups. Total protein adsorption was significantly higher for SB-A and SB-A-NF groups (p < 0.05), which also presented higher interactions with MG2 and lactoferrin proteins. The roughing of titanium surface by spot-blasting plus acid etching treatments contribute to higher interaction with salivary proteins, such as MG2 and lactoferrin. Titanium surface roughing increases the interactions of the substratum with salivary proteins, which can influence the integration of dental implants and their components to the oral environment. However, those treatments should be used carefully intraorally, avoiding increase biofilm formation.
Bactericidal effect of lactoferrin and lactoferrin chimera against halophilic Vibrio parahaemolyticus.

PubMed

Leon-Sicairos, Nidia; Canizalez-Roman, Adrian; de la Garza, Mireya; Reyes-Lopez, Magda; Zazueta-Beltran, Jorge; Nazmi, Kamran; Gomez-Gil, Bruno; Bolscher, Jan G

2009-01-01

Infections caused by Vibrio parahaemolyticus, an halophilic member of the genus Vibrio, have increased globally in the last 5 years. Diarrhea caused by V. parahaemolyticus results from eating raw or undercooked seafood. The aim of this work was to investigate whether lactoferrin and some lactoferrin-peptides have bactericidal activity against Vibrio parahaemolyticus ATCC 17802, the pandemic strain O3:K6, and the multidrug resistant isolate 727, as well as against Vibrio cholerae strains O1 and non-O1. Whereas both peptides lactoferricin (17-30) and lactoferrampin (265-284) did not have bactericidal activity, 40 microM of lactoferrin chimera (a fusion of the two peptides) inhibited the growth of all Vibrio tested to the same extent as the antibiotic gentamicin. The cidal effect of LFchimera showed a clear concentration response in contrast to bovine lactoferrin which showed higher inhibition at 10 microM than at 40 microM. FITC-labeled LFchimera bound to the bacterial membranes. Moreover LFchimera permeabilized bacterial cells and membranes were seriously damaged. Finally, in experiments with the multidrug resistant isolate 727, sub-lethal doses of LFchimera strongly reduced the concentrations of ampicillin, gentamicin or kanamicin needed to reach more than 95% growth inhibition, suggesting synergistic effects. These data indicate that LFchimera is a potential candidate to combat the multidrug resistant pathogenic Vibrio species.
Deleterious effect of salusin-β in paraventricular nucleus on sympathetic activity and blood pressure via NF-κB signaling in a rat model of obesity hypertension.

PubMed

Huang, Xiaodong; Wang, Yanchun; Ren, Kuang

2015-08-01

The paraventricular nucleus (PVN) has been shown to play a critical role in regulating blood pressure and sympathetic activity in obesity hypertension (OH). Salusin-β is a bioactive peptide with potential roles in mediating cardiovascular activity. The study was designed to test the hypothesis that salusin-β in the PVN can modulate sympathetic activity and blood pressure in OH. Male Sprague-Dawley rats were used to induce OH by a 12-week feeding of a high-fat diet (42% kcal as fat). Microinjection of salusin-β into the PVN increased the renal sympathetic nerve activity (RSNA), mean arterial pressure (MAP) and heart rate (HR) in a dose-dependent manner, whereas salusin-β antibody elicited significant decreases in RSNA, MAP and HR, and abolished the effects of salusin-β only in the OH rats. As expected, the OH rats had a higher norepinephrine level, which was further increased by salusin-β. Furthermore, salusin-β in the PVN accelerated the nuclear translocation of the p65 subunit of nuclear factor kappa B (NF-KB) and the degradation of IKB-α (an endogenous inhibitor of NF-KB). Pretreatment with pyrrolidine dithiocarbamate (an exogenous inhibitor of NF-KB) decreased RSNA, MAP and HR, and abolished the effects of salusin-β in the PVN in the OH rats. We concluded that salusin-β in the PVN markedly increased sympathetic outflow and blood pressure in diet-induced OH rats via NF-κB signaling.
Investigation of the enhanced antimicrobial activity of combination dry powder inhaler formulations of lactoferrin.

PubMed

Marshall, Lindsay J; Oguejiofor, Wilson; Price, Robert; Shur, Jagdeep

2016-12-05

The airways of most people with cystic fibrosis are colonized with biofilms of the Gram-negative, opportunistic pathogen Pseudomonas aeruginosa. Delivery of antibiotics directly to the lung in the form of dry powder aerosols offers the potential to achieve high local concentrations directly to the biofilms. Unfortunately, current aerosolised antibiotic regimes are unable to efficiently eradicate these biofilms from the airways. We investigated the ability of the innate antimicrobial, lactoferrin, to enhance the activity of two aminoglycoside antibiotics (tobramycin and gentamicin) against biofilms of P. aeruginosa strain PAO1. Biofilms were prepared in 96 well polystyrene plates. Combinations of the antibiotics and various lactoferrin preparations were spray dried. The bacterial cell viability of the various spray dried combinations was determined. Iron-free lactoferrin (apo lactoferrin) induced a 3-log reduction in the killing of planktonic cell by the aminoglycoside antibiotics (p<0.01) and also reduced both the formation and persistence of P. aeruginosa biofilms (p<0.01). Combinations of lactoferrin and an aminoglycoside displays potential as an effective new therapeutic strategy in the treatment of P. aeruginosa biofilms infections such as those typical of the CF lungs. Copyright © 2016 Elsevier B.V. All rights reserved.
A retinoic acid response element that overlaps an estrogen response element mediates multihormonal sensitivity in transcriptional activation of the lactoferrin gene.

PubMed

Lee, M O; Liu, Y; Zhang, X K

1995-08-01

The lactoferrin gene is highly expressed in many different tissues, and its expression is controlled by different regulators. In this report, we have defined a retinoic acid response element (RARE) in the 5'-flanking region of the lactoferrin gene promoter. The lactoferrin-RARE is composed of two AGGTCA-like motifs arranged as a direct repeat with 1-bp spacing (DR-1). A gel retardation assay demonstrated that it bound strongly with retinoid X receptor (RXR) homodimers and RXR-retinoic acid receptor (RAR) heterodimers as well as chicken ovalbumin upstream promoter transcription factor (COUP-TF) orphan receptor. In CV-1 cells, the lactoferrin-RARE linked with a heterologous thymidine kinase promoter was strongly activated by RXR homodimers in response to 9-cis-retinoic acid (9-cis-RA) but not to all-trans-RA. When the COUP-TF orphan receptor was cotransfected, the 9-cis-RA-induced RXR homodimer activity was strongly repressed. A unique feature of the lactoferrin-RARE is that it has an AGGTCA-like motif in common with an estrogen-responsive element (ERE). The composite RARE/ERE contributes to the functional interaction between retinoid receptors and the estrogen receptor (ER) and their ligands. In CV-1 cells, cotransfection of the retinoid and estrogen receptors led to mutual inhibition of the other's activity, while an RA-dependent inhibition of ER activity was observed in breast cancer cells. Furthermore, the lactoferrin-RARE/ERE showed differential transactivation activity in different cell types. RAs could activate the lactoferrin-RARE/ERE in human leukemia HL-60 cells and U937 cells but not in human breast cancer cells. By gel retardation analyses, we demonstrated that strong binding of the endogenous COUP-TF in breast cancer cells to the composite element contributed to diminished RA response in these cells. Thus, the lactoferrin-RARE/ERE functions as a signaling switch module that mediates multihormonal responsiveness in the regulation of lactoferrin gene
Lactoferrin modulation of BCG-infected dendritic cell functions

PubMed Central

Hwang, Shen-An

2009-01-01

Lactoferrin, an 80-kDa iron-binding protein with immune modulating properties, is a unique adjuvant component able to enhance efficacy of the existing Mycobacterium bovis Bacillus Calmette Guerin (BCG) vaccine to protect against murine model of tuberculosis. Although identified as having effects on macrophage presentation events, lactoferrin's capability to modulate dendritic cells (DCs) function when loaded with BCG antigens has not been previously recognized. In this study, the potential of lactoferrin to modulate surface expression of MHC II, CD80, CD86 and CD40 from bone marrow-derived dendritic cells (BMDCs) was examined. Generally, lactoferrin decreased pro-inflammatory cytokines [tumor necrosis factor (TNF)-α, IL-6 and IL-12p40] and chemokines [macrophage inflammatory protein (MIP)-1α and MIP-2] and increased regulatory cytokine, transforming growth factor-β1 and a T-cell chemotatic factor, monocyte chemotactic protein-1, from uninfected or BCG-infected BMDCs. Culturing BCG-infected BMDCs with lactoferrin also enhanced their ability to respond to IFN-γ activation through up-regulation of maturation markers: MHC I, MHC II and the ratio of CD86:CD80 surface expression. Furthermore, lactoferrin-exposed BCG-infected DCs increased stimulation of BCG-specific CD3+CD4+ splenocytes, as defined by increasing IFN-γ production. Finally, BCG-/lactoferrin-vaccinated mice possessed an increased pool of BCG antigen-specific IFN-γ producing CD3+CD4+CD62L− splenocytes. These studies suggest a mechanism in which lactoferrin may exert adjuvant activity by enhancing DC function to promote generation of antigen-specific T cells. PMID:19692539
Isolation of lactoferrin from milk of different species: calorimetric and antimicrobial studies.

PubMed

Conesa, Celia; Sánchez, Lourdes; Rota, Carmen; Pérez, María-Dolores; Calvo, Miguel; Farnaud, Sebastien; Evans, Robert W

2008-05-01

Lactoferrin (LF) is an iron-binding glycoprotein found in different biological fluids of mammals and in neutrophils. It has been proposed to be involved in many functions, including protection from pathogens. In this work, purification of lactoferrin using an ion-exchange chromatography (SP-Sepharose) was attempted for the milk of the following animals: sheep (Ovis aries), goat (Capra hircus), camel (Camelus bactrianus), alpaca (Lama pacos), elephant (Elephas maximus) and grey seal (Halichoerus grypus), as well as human (Homo sapiens). Lactoferrin was identified in all the milks apart from that from grey seal. The thermal stability of the purified lactoferrins, in their native and iron-saturated forms, was studied by differential scanning calorimetry (DSC). Maximum temperature, onset temperature and enthalpy change of denaturation were higher when lactoferrins were saturated with iron than in their native form, indicating an increase in the stability of the protein structure upon iron-binding. Human lactoferrin was found to be the most heat-resistant and the other lactoferrins presented different degrees of thermoresistance, that of elephant being the least resistant. The antimicrobial activity of the different isolated lactoferrins was investigated against Escherichia coli 0157:H7. The minimal inhibitory concentrations (MICs) were determined by measuring the absorbance at 620 nm. The minimum bactericidal concentrations (MBCs) were also measured and it was found that camel lactoferrin was the most active lactoferrin against E. coli 0157:H7, whereas alpaca and human lactoferrins were the least active.
Protective effect of human lactoferrin in the gastrointestinal tract.

PubMed

Queiroz, Valterlinda Alves de O; Assis, Ana Marlúcia O; R Júnior, Hugo da Costa

2013-01-01

To describe mechanisms of action of human lactoferrin to protect gastrointestinal morbidities. Nonsystematic literature review using the following databases: SciELO, Lilacs and Medline from 1990 to 2011. The key-words used were lactoferrin, human milk/breastfeeding, gastrointestinal, and immunity, in Portuguese and English. Lactoferrin is the second predominant protein in the human milk, with higher concentrations in the colostrum (5.0 to 6.7mg/mL) if compared to mature milk (0.2 to 2.6mg/mL.) In contrast, cow's milk has lower levels, with 0.83mg/mL in the colostrum and 0.09mg/mL in the mature milk. Lactoferrin has several physiological functions to protect the gastrointestinal tract. The antimicrobial activity is related to the ability to sequester iron from biological fluids and/or to destruct the membrane of microorganisms. Lactoferrin also has the ability to stimulate cell proliferation. The anti-inflammatory action exercised by lactoferrin is associated with its ability to penetrate the core of the leukocyte and to block the Kappa B nuclear factor transcription. Given the importance of lactoferrin to prevent infectious diseases for breastfed children, the industry is using genetic engineering techniques to develop the expression of recombinant human lactoferrin in animals and plants, attempting to adjust the composition of infant formulas to that of human milk. Human lactoferrin is a peptide with great potential for preventing morbidity, especially in the gastrointestinal tract. Scientific evidence of the protective effects of human lactoferrin strengthens even more the recommendation for breastfeeding.
Antimicrobial Functions of Lactoferrin Promote Genetic Conflicts in Ancient Primates and Modern Humans.

PubMed

Barber, Matthew F; Kronenberg, Zev; Yandell, Mark; Elde, Nels C

2016-05-01

Lactoferrin is a multifunctional mammalian immunity protein that limits microbial growth through sequestration of nutrient iron. Additionally, lactoferrin possesses cationic protein domains that directly bind and inhibit diverse microbes. The implications for these dual functions on lactoferrin evolution and genetic conflicts with microbes remain unclear. Here we show that lactoferrin has been subject to recurrent episodes of positive selection during primate divergence predominately at antimicrobial peptide surfaces consistent with long-term antagonism by bacteria. An abundant lactoferrin polymorphism in human populations and Neanderthals also exhibits signatures of positive selection across primates, linking ancient host-microbe conflicts to modern human genetic variation. Rapidly evolving sites in lactoferrin further correspond to molecular interfaces with opportunistic bacterial pathogens causing meningitis, pneumonia, and sepsis. Because microbes actively target lactoferrin to acquire iron, we propose that the emergence of antimicrobial activity provided a pivotal mechanism of adaptation sparking evolutionary conflicts via acquisition of new protein functions.
Telomere erosion in NF1 tumorigenesis.

PubMed

Jones, Rhiannon E; Grimstead, Julia W; Sedani, Ashni; Baird, Duncan; Upadhyaya, Meena

2017-06-20

Neurofibromatosis type 1 (NF1; MIM# 162200) is a familial cancer syndrome that affects 1 in 3,500 individuals worldwide and is inherited in an autosomal dominant fashion. Malignant Peripheral Nerve Sheath Tumors (MPNSTs) represent a significant cause of morbidity and mortality in NF1 and currently there is no treatment or definite prognostic biomarkers for these tumors. Telomere shortening has been documented in numerous tumor types. Short dysfunctional telomeres are capable of fusion and it is considered that the ensuing genomic instability may facilitate clonal evolution and the progression to malignancy. To evaluate the potential role of telomere dysfunction in NF1-associated tumors, we undertook a comparative analysis of telomere length in samples derived from 10 cutaneous and 10 diffused plexiform neurofibromas, and 19 MPNSTs. Telomere length was determined using high-resolution Single Telomere Length Analysis (STELA). The mean Xp/Yp telomere length detected in MPNSTs, at 3.282 kb, was significantly shorter than that observed in both plexiform neurofibromas (5.793 kb; [p = 0.0006]) and cutaneous neurofibromas (6.141 kb; [p = 0.0007]). The telomere length distributions of MPNSTs were within the length-ranges in which telomere fusion is detected and that confer a poor prognosis in other tumor types. These data indicate that telomere length may play a role in driving genomic instability and clonal progression in NF1-associated MPNSTs.
Noncanonical NF-κB Signaling Is Limited by Classical NF-κB Activity

PubMed Central

Gray, Carolyn M.; Remouchamps, Caroline; McCorkell, Kelly A.; Solt, Laura A.; Dejardin, Emmanuel; Orange, Jordan S.; May, Michael J.

2014-01-01

Precise regulation of nuclear factor κB (NF-κB) signaling is crucial for normal immune responses, and defective NF-κB activity underlies a range of immunodeficiencies. NF-κB is activated through two signaling cascades: the classical and noncanonical pathways. The classical pathway requires inhibitor of κB kinase β (IKKβ) and NF-κB essential modulator (NEMO), and hypomorphic mutations in the gene encoding NEMO (ikbkg) lead to inherited immunodeficiencies, collectively termed NEMO-ID. Noncanonical NF-κB activation requires NF-κB–inducing kinase (NIK) and IKKα, but not NEMO. We found that noncanonical NF-κB was basally active in peripheral blood mononuclear cells from NEMO-ID patients, and that noncanonical NF-κB signaling was similarly enhanced in cell lines lacking functional NEMO. NIK, which normally undergoes constitutive degradation, was aberrantly present in resting NEMO-deficient cells, and regulation of its abundance was rescued by reconstitution with full-length NEMO, but not a mutant NEMO protein unable to physically associate with IKKα or IKKβ. Binding of NEMO to IKKα was not required for ligand-dependent stabilization of NIK or noncanonical NF-κB signaling. Rather, an intact and functional IKK complex was essential to suppress basal NIK activity in unstimulated cells. Despite interacting with IKKα and IKKβ to form an IKK complex, NEMO mutants associated with immunodeficiency failed to rescue classical NF-κB signaling or reverse the accumulation of NIK. Together, these findings identify a crucial role for classical NF-κB activity in the suppression of basal noncanonical NF-κB signaling. PMID:24497610
Microbicidal effect of the lactoferrin peptides lactoferricin17-30, lactoferrampin265-284, and lactoferrin chimera on the parasite Entamoeba histolytica.

PubMed

López-Soto, Fernando; León-Sicairos, Nidia; Nazmi, Kamran; Bolscher, Jan G; de la Garza, Mireya

2010-06-01

Entamoeba histolytica is a parasitic protozoan that produces amoebiasis, an intestinal disease characterized by ulcerative colitis and dysentery. In some cases, trophozoites can travel to the liver leading to hepatic abscesses and death. Recently, lactoferrin and lactoferricin B have been shown to be amoebicidal in axenic cultures. The aim of this work was to determine whether the lactoferrin-peptides lactoferricin amino acids 17-30, lactoferrampin amino acids 265-284, and lactoferrin chimera which is a fusion product of the two peptides, are capable of producing a microbicidal effect to trophozoites of E. histolytica. We evaluated the killing effect of these peptides in growth kinetics carried out in axenic culture medium to which different concentrations of peptides were added. At 50 muM of peptide concentration, lactoferricin and lactoferrampin had a moderate amoebicidal effect, since a 45-50% of trophozoites remained viable at 24 h culture. However, at 50 microM of the lactoferrin chimera 75% amoeba were killed whereas at 100 microM all cells died. These data indicate that of lactoferrin-peptides mainly the chimera have amoebicidal activity in a time- and concentration-dependent manner. The lactoferrin-peptides might be useful as therapeutic agents against amoebiasis and thereby diminish the use of metronidazole, which is extremely toxic for the host.
Lactoferrin derived resistance against plant pathogen in transgenic plants

USDA-ARS?s Scientific Manuscript database

Lactoferrin (LF) is a ubiquitous cationic iron-binding milk glycoprotein and it is known to exert a broad-spectrum primary defense activity against bacteria, fungi, protozoa and viruses in mammals. The Bovine lactoferrin gene was introduced to tobacco (Nicotiana tabacum var Xanthi), Arabidopsis (A. ...
Biological activity of lactoferrin-functionalized biomimetic hydroxyapatite nanocrystals

PubMed Central

Nocerino, Nunzia; Fulgione, Andrea; Iannaccone, Marco; Tomasetta, Laura; Ianniello, Flora; Martora, Francesca; Lelli, Marco; Roveri, Norberto; Capuano, Federico; Capparelli, Rosanna

2014-01-01

The emergence of bacterial strains resistant to antibiotics is a general public health problem. Progress in developing new molecules with antimicrobial properties has been made. In this study, we evaluated the biological activity of a hybrid nanocomposite composed of synthetic biomimetic hydroxyapatite surface-functionalized by lactoferrin (LF-HA). We evaluated the antimicrobial, anti-inflammatory, and antioxidant properties of LF-HA and found that the composite was active against both Gram-positive and Gram-negative bacteria, and that it modulated proinflammatory and anti-inflammatory responses and enhanced antioxidant properties as compared with LF alone. These results indicate the possibility of using LF-HA as an antimicrobial system and biomimetic hydroxyapatite as a candidate for innovative biomedical applications. PMID:24623976
Isolation and Characterization of Lactoferrin Peptides with Stimulatory Effect on Osteoblast Proliferation.

PubMed

Fan, Fengjiao; Tu, Maolin; Liu, Meng; Shi, Pujie; Wang, Yun; Wu, Di; Du, Ming

2017-08-23

Lactoferrin is reported to be a potential food protein with osteogenic activity. However, the activity of lactoferrin peptides is questionable. In the present study, we isolated and characterized peptides from lactoferrin with stimulatory effect on osteoblast proliferation. Peptides from the lactoferrin pepsin hydrolysate were purified using cation-exchange and gel-filtration chromatography. Effects of different hydrolysates and peptides on the proliferation of osteoblast MC3T3-E1 cells were compared by MTT assay. Results showed that fraction P5-a from Superdex Peptide 10/300 GL gel chromatography showed better activity. Tricine-sodium dodecyl sulfate polyacrylamide gel electrophoresis and high-performance liquid chromatography coupled to electrospray ionization tandem mass spectrometry confirmed that two peptides components of P5-a corresponded to fractions of 20-78 and 191-277 amino acids in Bos taurus lactoferrin molecule (GI: 221706349). These results will provide some theoretical and practical data for the preparation and application of osteogenic peptides in functional food industry.
Lactoferrin expression and secretion in the stallion epididymis.

PubMed

Pearl, Christopher A; Roser, Janet F

2014-04-01

Lactoferrin is one of the most abundant proteins secreted by the stallion epididymis, but its cellular localization and regulation remain unknown. This study was designed to address the following objectives: (1) identify the epididymal cell types producing lactoferrin in pre-pubertal, peri-pubertal and post-pubertal animals; (2) demonstrate that lactoferrin binds to stallion sperm; and (3) determine if testosterone and estradiol regulate lactoferrin secretion in vitro. Using an immunohistochemical method, lactoferrin was localized in the cytoplasm of principal cells in the corpus and cauda of peri- and post-pubertal animals. The epididymis of pre-pubertal animals did not express lactoferrin. Immunolabeling of lactoferrin was also observed on the mid-piece and tail of the sperm. The role of estradiol and testosterone in regulating secretion of lactoferrin in the post-pubertal epididymis was investigated using tissue culture methods. Lactoferrin concentration in the culture media was determined by validated enzyme-linked immunosorbent assays (ELISA). Testosterone did not increase the concentration of lactoferrin in the media in any epididymal region. In contrast, estradiol-17β significantly increased the concentration of lactoferrin in the media containing tissue from the cauda. In conclusion, the expression of lactoferrin was found in the cytoplasm of principal cells in the corpus and cauda of the epididymis in peri- and post-pubertal stallions but not pre-pubertal stallions. Furthermore, lactoferrin binds to sperm, suggesting a biological role for protection or regulation of sperm in the corpus and cauda. In addition, estrogen appears to regulate lactoferrin secretion in the cauda of the epididymis in post-pubertal stallions. Copyright © 2014 Society for Biology of Reproduction & the Institute of Animal Reproduction and Food Research of Polish Academy of Sciences in Olsztyn. Published by Elsevier Urban & Partner Sp. z o.o. All rights reserved.
Genomic organization of the neurofibromatosis 1 gene (NF1)

DOE Office of Scientific and Technical Information (OSTI.GOV)

Li, Y.; O`Connell, P.; Huntsman Breidenbach, H.

Neurofibromatosis 1 maps to chromosome band 17q11.2, and the NF1 locus has been partially characterized. Even though the full-length NF1 cDNA has been sequenced, the complete genomic structure of the NF1 gene has not been elucidated. The 5{prime} end of NF1 is embedded in a CpG island containing a NotI restriction site, and the remainder of the gene lies in the adjacent 350-kb NotI fragment. In our efforts to develop a comprehensive screen for NF1 mutations, we have isolated genomic DNA clones that together harbor the entire NF1 cDNA sequence. We have identified all intron-exon boundaries of the coding regionmore » and established that it is composed of 59 exons. Furthermore, we have defined the 3{prime}-untranslated region (3{prime}-UTR) of the NF1 gene; it spans approximately 3.5 kb of genomic DNA sequence and is continuous with the stop codon. Oligonucleotide primer pairs synthesized from exon-flanking DNA sequences were used in the polymerase chain reaction with cloned, chromosome 17-specific genomic DNA as template to amplify NF1 exons 1 through 27b and the exon containing the 3{prime}-UTR separately. This information should be useful for implementing a comprehensive NF1 mutation screen using genomic DNA as template. 41 refs., 3 figs., 2 tabs.« less
NF-κB Activation Protects Oligodendrocytes against Inflammation

PubMed Central

Stone, Sarrabeth; Jamison, Stephanie; Yue, Yuan; Durose, Wilaiwan

2017-01-01

NF-κB is a key player in inflammatory diseases, including multiple sclerosis (MS) and its animal model, experimental autoimmune encephalomyelitis (EAE). However, the effects of NF-κB activation on oligodendrocytes in MS and EAE remain unknown. We generated a mouse model that expresses IκBαΔN, a super-suppressor of NF-κB, specifically in oligodendrocytes and demonstrated that IκBαΔN expression had no effect on oligodendrocytes under normal conditions (both sexes). Interestingly, we showed that oligodendrocyte-specific expression of IκBαΔN blocked NF-κB activation in oligodendrocytes and resulted in exacerbated oligodendrocyte death and hypomyelination in young, developing mice that express IFN-γ ectopically in the CNS (both sexes). We also showed that NF-κB inactivation in oligodendrocytes aggravated IFN-γ-induced remyelinating oligodendrocyte death and remyelination failure in the cuprizone model (male mice). Moreover, we found that NF-κB inactivation in oligodendrocytes increased the susceptibility of mice to EAE (female mice). These findings imply the cytoprotective effects of NF-κB activation on oligodendrocytes in MS and EAE. SIGNIFICANCE STATEMENT Multiple sclerosis (MS) is an inflammatory demyelinating disease of the CNS. NF-κB is a major player in inflammatory diseases that acts by regulating inflammation and cell viability. Data indicate that NF-κB activation in inflammatory cells facilitates the development of MS. However, to date, attempts to understand the role of NF-κB activation in oligodendrocytes in MS have been unsuccessful. Herein, we generated a mouse model that allows for inactivation of NF-κB specifically in oligodendrocytes and then used this model to determine the precise role of NF-κB activation in oligodendrocytes in models of MS. The results presented in this study represent the first demonstration that NF-κB activation acts cell autonomously to protect oligodendrocytes against inflammation in animal models of MS
NF45/ILF2 tissue expression, promoter analysis, and interleukin-2 transactivating function

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhao Guohua; Shi Lingfang; Qiu Daoming

2005-05-01

NF45/ILF2 associates with NF90/ILF3 in the nucleus and regulates IL-2 gene transcription at the antigen receptor response element (ARRE)/NF-AT DNA target sequence (P.N. Kao, L. Chen, G. Brock, J. Ng, A.J. Smith, B. Corthesy, J. Biol. Chem. 269 (1994) 20691-20699). NF45 is widely expressed in normal tissues, especially testis, brain, and kidney, with a predominantly nuclear distribution. NF45 mRNA expression is increased in lymphoma and leukemia cell lines. The human and murine NF45 proteins differ only by substitution of valine by isoleucine at amino acid 142. Fluorescence in situ hybridization localized the human NF45 gene to chromosome 1q21.3, and mousemore » NF45 gene to chromosome 3F1. Promoter analysis of 2.5 kB of the murine NF45 gene reveals that significant activation is conferred by factors, possible including NF-Y, that bind to the CCAAT-box sequence. The function of human NF45 in regulating IL-2 gene expression was characterized in Jurkat T-cells stably transfected with plasmids directing expression of NF45 cDNA in sense or antisense orientations. NF45 sense expression increased IL-2 luciferase reporter gene activity 120-fold, and IL-2 protein expression 2-fold compared to control cells. NF45 is a highly conserved, regulated transcriptional activator, and one target gene is IL-2.« less
Synergistic Fungistatic Effects of Lactoferrin in Combination with Antifungal Drugs against Clinical Candida Isolates

PubMed Central

Kuipers, M. E.; de Vries, H. G.; Eikelboom, M. C.; Meijer, D. K. F.; Swart, P. J.

1999-01-01

Because of the rising incidence of failures in the treatment of oropharyngeal candidosis in the case of severely immunosuppressed patients (mostly human immunodeficiency virus [HIV]-infected patients), there is need for the development of new, more effective agents and/or compounds that support the activity of the common antifungal agents. Since lactoferrin is one of the nonspecific host defense factors present in saliva that exhibit antifungal activity, we studied the antifungal effects of human, bovine, and iron-depleted lactoferrin in combination with fluconazole, amphotericin B, and 5-fluorocytosine in vitro against clinical isolates of Candida species. Distinct antifungal activities of lactoferrin were observed against clinical isolates of Candida. The MICs generally were determined to be in the range of 0.5 to 100 mg · ml−1. Interestingly, in the combination experiments we observed pronounced cooperative activity against the growth of Candida by using lactoferrin and the three antifungals tested. Only in a limited concentration range was minor antagonism detected. The use of lactoferrin and fluconazole appeared to be the most successful combination. Significant reductions in the minimal effective concentrations of fluconazole were found when it was combined with a relatively low lactoferrin concentration (1 mg/ml). Such combinations still resulted in complete growth inhibition, while synergy of up to 50% against several Candida species was observed. It is concluded that the combined use of lactoferrin and antifungals against severe infections with Candida is an attractive therapeutic option. Since fluconazole-resistant Candida species have frequently been reported, especially in HIV-infected patients, the addition of lactoferrin to the existing fluconazole therapy could postpone the occurrence of species resistance against fluconazole. Clinical studies to further elucidate the potential utility of this combination therapy have been initiated. PMID:10543740

Synergistic fungistatic effects of lactoferrin in combination with antifungal drugs against clinical Candida isolates.

PubMed

Kuipers, M E; de Vries, H G; Eikelboom, M C; Meijer, D K; Swart, P J

1999-11-01

Because of the rising incidence of failures in the treatment of oropharyngeal candidosis in the case of severely immunosuppressed patients (mostly human immunodeficiency virus [HIV]-infected patients), there is need for the development of new, more effective agents and/or compounds that support the activity of the common antifungal agents. Since lactoferrin is one of the nonspecific host defense factors present in saliva that exhibit antifungal activity, we studied the antifungal effects of human, bovine, and iron-depleted lactoferrin in combination with fluconazole, amphotericin B, and 5-fluorocytosine in vitro against clinical isolates of Candida species. Distinct antifungal activities of lactoferrin were observed against clinical isolates of Candida. The MICs generally were determined to be in the range of 0.5 to 100 mg. ml(-1). Interestingly, in the combination experiments we observed pronounced cooperative activity against the growth of Candida by using lactoferrin and the three antifungals tested. Only in a limited concentration range was minor antagonism detected. The use of lactoferrin and fluconazole appeared to be the most successful combination. Significant reductions in the minimal effective concentrations of fluconazole were found when it was combined with a relatively low lactoferrin concentration (1 mg/ml). Such combinations still resulted in complete growth inhibition, while synergy of up to 50% against several Candida species was observed. It is concluded that the combined use of lactoferrin and antifungals against severe infections with Candida is an attractive therapeutic option. Since fluconazole-resistant Candida species have frequently been reported, especially in HIV-infected patients, the addition of lactoferrin to the existing fluconazole therapy could postpone the occurrence of species resistance against fluconazole. Clinical studies to further elucidate the potential utility of this combination therapy have been initiated.
High-Level Expression of Recombinant Bovine Lactoferrin in Pichia pastoris with Antimicrobial Activity

PubMed Central

Iglesias-Figueroa, Blanca; Valdiviezo-Godina, Norberto; Siqueiros-Cendón, Tania; Sinagawa-García, Sugey; Arévalo-Gallegos, Sigifredo; Rascón-Cruz, Quintín

2016-01-01

In this study, bovine lactoferrin (bLf), an iron-binding glycoprotein considered an important nutraceutical protein because of its several properties, was expressed in Pichia pastoris KM71-H under AOX1 promoter control, using pJ902 as the recombinant plasmid. Dot blotting analysis revealed the expression of recombinant bovine lactoferrin (rbLf) in Pichia pastoris. After Bach fermentation and purification by molecular exclusion, we obtained an expression yield of 3.5 g/L of rbLf. rbLf and predominantly pepsin-digested rbLf (rbLfcin) demonstrated antibacterial activity against Escherichia coli (E. coli) BL21DE3, Staphylococcus aureus (S. aureus) FRI137, and, in a smaller percentage, Pseudomonas aeruginosa (Ps. Aeruginosa) ATCC 27833. The successful expression and characterization of functional rbLf expressed in Pichia pastoris opens a prospect for the development of natural antimicrobial agents produced recombinantly. PMID:27294912
Lactoferrin Levels in Tears are Increased by the Topical Application of Diadenosine Tetraphosphate.

PubMed

Loma, Patricia; Guzman-Aranguez, Ana; Perez de Lara, Maria J; Pintor, Jesus

2016-09-01

This study was undertaken to determine the effect of the topical application of diadenosine tetraphosphate on lactoferrin levels in rabbit tears. Diadenosine tetraphosphate was topically instilled in a single-dose, tear samples were collected by micropipette and lactoferrin was measured by Enzyme-Linked ImmunoSorbent Assay (ELISA). The concentration of lactoferrin in rabbit tears was significantly increased 1 h after diadenosine tetraphosphate application, remaining elevated for 3 h more. This effect was blocked by P2 receptors antagonists. Topical application of diadenosine tetraphosphate stimulates the secretion of lactoferrin in rabbit tears through P2 receptor activation.
Bovine Lactoferrampin, Human Lactoferricin, and Lactoferrin 1-11 Inhibit Nuclear Translocation of HIV Integrase.

PubMed

Wang, Winston Yan; Wong, Jack Ho; Ip, Denis Tsz Ming; Wan, David Chi Cheong; Cheung, Randy Chifai; Ng, Tzi Bun

2016-08-01

This study aimed to investigate fragments derived from human and bovine lactoferrins for ability to inhibit nuclear translocation of HIV-1 integrase. It was shown that human lactoferricin, human lactoferrin 1-11, and bovine lactoferrampin reduced nuclear distribution of HIV-1 integrase. Bovine lactoferrampin could inhibit both the activity and nuclear translocation of HIV-1 integrase. Human lactoferrampin, bovine lactoferricin, and bovine lactoferrin 1-11 had no effect on HIV-1 integrase nuclear translocation. Human lactoferrampin which inhibited the activity of integrase did not prevent its nuclear translocation. Human lactoferricin and lactoferrin 1-11 did not inhibit HIV-1 integrase nuclear translocation despite their ability to attenuate the enzyme activity. The discrepancy between the findings on reduction of HIV-1 activity and inhibition of nuclear translocation of HIV-1 integrase was due to the different mechanisms involved. A similar reasoning can also be applied to the different inhibitory potencies of the milk peptides on different HIV enzymes, i.e., nuclear translocation.
Antimicrobial activity of buttermilk and lactoferrin peptide extracts on poultry pathogens.

PubMed

Jean, Catherine; Boulianne, Martine; Britten, Michel; Robitaille, Gilles

2016-11-01

Antibiotics are commonly used in poultry feed as growth promoters. This practice is questioned given the arising importance of antibiotic resistance. Antimicrobial peptides can be used as food additives for a potent alternative to synthetic or semi-synthetic antibiotics. The objective of this study was to develop a peptide production method based on membrane adsorption chromatography in order to produce extracts with antimicrobial activity against avian pathogens (Salmonella enterica var. Enteritidis, Salmonella enterica var. Typhimurium, and two Escherichia coli strains, O78:H80 and TK3 O1:K1) as well as Staphylococcus aureus. To achieve this, buttermilk powder and purified lactoferrin were digested with pepsin. The peptide extracts (<10 kDa) were fractionated depending on their charges through high-capacity cation-exchange and anion-exchange adsorptive membranes. The yields of cationic peptide extracts were 6·3 and 15·4% from buttermilk and lactoferrin total peptide extracts, respectively. Antimicrobial activity was assessed using the microdilution technique on microplates. Our results indicate that the buttermilk cationic peptide extracts were bactericidal at less than 5 mg/ml against the selected avian strains, with losses of 1·7 log CFU/ml (Salm. Typhimurium) to 3 log CFU/ml (E. coli O78:H80); viability decreased by 1·5 log CFU/ml for Staph. aureus, a Gram-positive bacterium. Anionic and non-adsorbed peptide extracts were inactive at 5 mg/ml. These results demonstrate that membrane adsorption chromatography is an effective way to prepare a cationic peptide extract from buttermilk that is active against avian pathogens.
Human lactoferrin induces asthmatic symptoms in NC/Nga mice.

PubMed

Nagaoka, Kenjiro; Ito, Tatsuo; Ogino, Keiki; Eguchi, Eri; Fujikura, Yoshihisa

2017-08-01

Lactoferrin in commercial supplements is known to exert anti-viral and anti-allergic effects. However, this is the first study to evaluate the induction of allergic airway inflammation in NC/Nga mice. Human lactoferrin was administered intraperitoneally with aluminum oxide for sensitization. Five days later, lactoferrin was inoculated intranasally for 5 days, and then on the 12th day, the single inoculation of lactoferrin intranasally was performed as a challenge. On the 13th day, airway hypersensitivity was assessed (AHR), a bronchoalveolar fluid (BALF) cell analysis was conducted, serum IgE and serum lactoferrin-specific IgG and IgE levels as well as the mRNA expression levels of cytokines and chemokines in the lung were measured, and a histopathological analysis of the lung was performed. Human lactoferrin increased AHR, the number of eosinophils in BALF, serum lactoferrin-specific IgG levels, and the mRNA levels of IL-13, eotaxin 1, and eotaxin 2. Moreover, the accumulation of inflammatory cells around the bronchus and the immunohistochemical localization of arginase I and human lactoferrin were detected. Collectively, these results indicate that human lactoferrin induced allergic airway inflammation in mice. Therefore, the commercial use of human lactoferrin in supplements warrants more intensive study. © 2017 The Authors. Physiological Reports published by Wiley Periodicals, Inc. on behalf of The Physiological Society and the American Physiological Society.
Lactobacilli-lactoferrin interplay in Chlamydia trachomatis infection.

PubMed

Sessa, Rosa; Di Pietro, Marisa; Filardo, Simone; Bressan, Alessia; Mastromarino, Paola; Biasucci, Alessandra Vittoria; Rosa, Luigi; Cutone, Antimo; Berlutti, Francesca; Paesano, Rosalba; Valenti, Piera

2017-07-31

In the cervicovaginal microenvironment, lactobacilli are known to protect against genital infections and, amongst the host defence compounds, lactoferrin has recently acquired importance for its anti-microbial and anti-inflammatory properties. An abnormal genital microenvironment facilitates the acquisition of pathogens like Chlamydia trachomatis, the leading cause of bacterial sexually transmitted infections worldwide. The aim of our study is to investigate the effects of Lactobacillus crispatus, Lactobacillus brevis and bovine lactoferrin on chlamydial infection, in order to shed light on the complex interplay between host defence mechanisms and C. trachomatis. We have also evaluated the effect of these defence factors to modulate the chlamydia-mediated inflammatory state. To this purpose, we have determined the infectivity and progeny production of C. trachomatis as well as interleukin-8 and interleukin-6 synthesis. The main result of our study is that the combination of L. brevis and bovine lactoferrin is the most effective in inhibiting the early phases (adhesion and invasion) of C. trachomatis infection of cervical epithelial cells and in decreasing the levels of both cytokines. In conclusion, the interaction between L. brevis and lactoferrin seems to play a role in the protection against C. trachomatis, reducing the infection and regulating the immunomodulatory activity, thus decreasing the risk of severe complications. © FEMS 2017. All rights reserved. For permissions, please e-mail: journals.permissions@oup.com.
21 CFR 866.5570 - Lactoferrin immunological test system.

Code of Federal Regulations, 2010 CFR

2010-04-01

... (CONTINUED) MEDICAL DEVICES IMMUNOLOGY AND MICROBIOLOGY DEVICES Immunological Test Systems § 866.5570 Lactoferrin immunological test system. (a) Identification. A lactoferrin immunological test system is a device... 21 Food and Drugs 8 2010-04-01 2010-04-01 false Lactoferrin immunological test system. 866.5570...
The amiodarone derivative KB130015 activates hERG1 potassium channels via a novel mechanism

PubMed Central

Gessner, Guido; Macianskiene, Regina; Starkus, John G.; Schönherr, Roland; Heinemann, Stefan H.

2010-01-01

Human ether à go-go related gene (hERG1) potassium channels underlie the repolarizing IKr current in the heart. Since they are targets of various drugs with cardiac side effects we tested whether the amiodarone derivative 2-methyl-3-(3,5-diiodo-4-carboxymethoxybenzyl)benzofuran (KB130015) blocks hERG1 channels like its parent compound. Using patch-clamp and two-electrode voltage-clamp techniques we found that KB130015 blocks native and recombinant hERG1 channels at high voltages, but it activates them at low voltages. The activating effect has an apparent EC50 value of 12 μM and is brought about by an about 4-fold acceleration of activation kinetics and a shift in voltage-dependent activation by −16 mV. Channel activation was not use-dependent and was independent of inactivation gating. KB130015 presumably binds to the hERG1 pore from the cytosolic side and functionally competes with hERG1 block by amiodarone, E4031 (N-[4-[[1-[2-(6-methyl-2-pyridinyl)ethyl] -4-piperidinyl] carbonyl] phenyl] methanesulfonamide dihydrochloride), and sertindole. Vice versa, amiodarone attenuates hERG1 activation by KB130015. Based on synergic channel activation by mallotoxin and KB130015 we conclude that the hERG1 pore contains at least two sites for activators that are functionally coupled among each other and to the cavity-blocker site. KB130015 and amiodarone may serve as lead structures for the identification of hERG1 pore-interacting drugs favoring channel activation vs. block. PMID:20097192
Effects of in vitro lactoferricin and lactoferrin on the head kidney cells of European sea bass (Dicentrarchus labrax, L.).

PubMed

Henry, Morgane A; Alexis, Maria N

2009-08-15

Antimicrobial, anti-inflammatory and immunomodulating properties of lactoferrin have been demonstrated in mammals and in fish. However, in vivo, lactoferrin is digested by gastric pepsin treatment into the N-terminal derived peptide named lactoferricin. This has been so far overlooked in fish in vitro studies. The aim of the present study was to assess in vitro the effects of both lactoferricin and lactoferrin on the head kidney cells of European sea bass (Dicentrarchus labrax, L.) in order to determine their potential as dietary additives and to get some insight into their mode of action. In vitro lactoferricin decreased significantly the chemiluminescent response of head kidney cells but did not affect the zymosan-triggered chemiluminescence activity. On the other hand, a high concentration of lactoferrin directly stimulated chemiluminescence but reduced the zymosan-triggered chemiluminescence. The bactericidal activity of head kidney cells was also significantly diminished by pre-incubation with lactoferrin in a dose-dependent manner. Although no significant effect of lactoferricin or lactoferrin was evidenced on head kidney cellular viability, absent or negative effect on the priming of respiratory burst activity suggested that care should be taken when using lactoferrin in the diet of sea bass and high doses should be avoided. Hypotheses about the mechanisms of action of lactoferricin and lactoferrin are presented.
Effect of technological treatments on bovine lactoferrin: An overview.

PubMed

Franco, Indira; Pérez, María Dolores; Conesa, Celia; Calvo, Miguel; Sánchez, Lourdes

2018-04-01

Lactoferrin (LF) is a multifunctional protein that exerts important activities in the neonate through its presence in milk, and also in other external mucosas, acting as a defense protein of innate immunity. The addition of bovine LF to infant formula and also to other functional products and cosmetics has increased during the last decades. Consequently, it is essential to know the effect that the technological processes, necessary to elaborate those products, have on LF activity. In this study, we have revised the effect of classical treatments on lactoferrin structure and activity, such as heat treatment or drying, and also of emerging technologies, like high pressure or pulsed electric field. The results of the studies included in this review indicate that LF stability is dependent on its level of iron-saturation and on the characteristics of the treatment media. Furthermore, the studies revised here reveal that the non-thermal treatments are interesting alternatives to the traditional ones, as they protect better the structure and activity of lactoferrin. It is also clear the need for research on LF encapsulation by different ways, to protect its properties before it reaches the intestine. All this knowledge would allow designing processes less harmful for LF, thus maintaining all its functionality. Copyright © 2017 Elsevier Ltd. All rights reserved.
Kinetic and thermodynamic parameters for thermal denaturation of ovine milk lactoferrin determined by its loss of immunoreactivity.

PubMed

Navarro, F; Harouna, S; Calvo, M; Pérez, M D; Sánchez, L

2015-07-01

Lactoferrin is a protein with important biological functions that can be obtained from milk and by-products derived from the dairy industry, such as whey. Although bovine lactoferrin has been extensively studied, ovine lactoferrin is not quite as well known. In the present study, the effect of several heat treatments in 3 different media, over a temperature range from 66 to 75°C, has been studied on lactoferrin isolated from sheep milk. Denaturation of lactoferrin was determined by measuring its immunoreactivity with specific polyclonal antibodies. Kinetic and thermodynamic parameters obtained indicate that lactoferrin denatures by heat more rapidly in whey than in phosphate buffer or milk. The value of activation energy found for the denaturation process of lactoferrin when treated in whey is higher (390kJ/mol) than that obtained in milk (194kJ/mol) or phosphate buffer (179kJ/mol). This indicates that a great amount of energy is necessary to start denaturation of ovine lactoferrin, probably due to the interaction of this protein with other whey proteins. The changes in the hydrophobicity of lactoferrin after heat treatments were determined by fluorescence measurement using acrylamide. The decrease in the hydrophobicity constant was very small for the treatments from 66 to 75°C, up to 20min, which indicates that lactoferrin conformation did not experienced a great change. The results obtained in this study permit the prediction of behavior of ovine lactoferrin under several heat treatments and show that high-temperature, short-time pasteurization (72°C, 15 s) does not cause loss of its immunoreactivity and, consequently, would not affect its conformation and biological activity. Copyright © 2015 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
Bovine Lactoferrin and Lactoferricin, a Peptide Derived from Bovine Lactoferrin, Inhibit Tumor Metastasis in Mice

PubMed Central

Watanabe, Shikiko; Watanabe, Ryosuke; Hata, Katsusuke; Shimazaki, Kei–ichi; Azuma, Ichiro

1997-01-01

We investigated the effect of a bovine milk protein, lactoferrin (LF–B), and a pepsin–generated peptide of LF–B, lactoferricin (Lfcin–B), on inhibition of tumor metastasis produced by highly metastatic murine tumor cells, B16–BL6 melanoma and L5178Y–ML25 lymphoma cells, using experimental and spontaneous metastasis models in syngeneic mice. The subcutaneous (s.c.) administration of bovine apo–lactoferrin (apo–LF–B, 1 mg/mouse) and Lfcin–B (0.5 mg/monse) 1 day after tumor inoculation significantly inhibited liver and lung metastasis of L5178Y–ML25 cells. However, human apo–lactoferrin (apo–LF–H) and bovine holo–lactoferrin (holo–LF–B) at the dose of 1 mg/mouse failed to inhibit tumor metastasis of L5178Y–ML25 cells. Similarly, the s.c. administration of apo–LF–B as well as Lfcin–B, but not apo–LF–H and holo–LF–B, 1 day after tumor inoculation resulted in significant inhibition of lung metastasis of B16–BL6 cells in an experimental metastasis model. Furthermore, in in vivo analysis for tumor–induced angiogenesis, both apo–LF–B and Lfcin–B inhibited the number of tumor–induced blood vessels and suppressed tumor growth on day 8 after tumor inoculation. However, in a long–term analysis of tumor growth for up to 21 days after tumor inoculation, single administration of apo–LF–B significantly suppressed the growth of B16–BL6 cells throughout the examination period, whereas Lfcin–B showed inhibitory activity only during the early period (8 days). In spontaneous metastasis of B16–BL6 melanoma cells, multiple administration of both apo–LF–B and Lfcin–B into tumor–bearing mice significantly inhibited lung metastasis produced by B16–BL6 cells, though only apo–LF–B exhibited an inhibitory effect on tumor growth at the time of primary tumor amputation (on day 21) after tumor inoculation. These results suggest that apo–LF–B and Lfcin–B inhibit tumor metastasis through different
NF-kappa B activation correlates with disease phenotype in Crohn’s disease

PubMed Central

Han, Yoo Min; Koh, Jaemoon; Lee, Changhyun; Koh, Seong-Joon; Kim, ByeongGwan; Lee, Kook Lae; Im, Jong Pil; Kim, Joo Sung

2017-01-01

Background/Aims Unregulated activation of nuclear factor-κB (NF-κB) plays a critical role in the pathogenesis of Crohn’s disease. In this study, we investigated the clinical characteristics and disease outcome of Crohn’s disease patients with varying levels of the NF-κB activation. Methods Crohn’s disease patients who underwent surgical bowel resection were divided into two groups, based on the activation status of NF-κB. NF-κB activation was assessed by the immunoreactivity of nuclear NF-κB during immunohistochemical staining of bowel resection specimens. We compared the demographic, clinical and histologic characteristics between groups. Furthermore, the occurrence of reoperation, readmission, and medication change due to disease flare-up were investigated according to NF-κB activation status. Results Among 83 Crohn’s disease patients, 47 (56%) showed high NF-κB activity and 36 (44%) showed low NF-κB activity. Patients with high NF-κB activity had higher frequency of ileocolonic involvement (P = 0.028) and lower frequency of perianal involvement (P = 0.042) relative to those with low NF-κB activity. Total histologic scores were significantly higher in patients with high NF-κB activity than those with low NF-κB activity (P = 0.044). There was no significant difference in the frequency of reoperation, readmission, and medication change in relation to NF-κB activation status. Conclusions Crohn’s disease patients with high NF-κB activation showed specific clinical manifestations of higher frequency of ileocolonic involvement and lower frequency of perianal involvement relative to those with low NF-κB activation. High NF-κB activity was associated with higher histologic scores. However, the NF-κB activity did not affect the outcome and disease course after surgery. PMID:28753650
Tubular Epithelial NF-κB Activity Regulates Ischemic AKI

PubMed Central

Vigolo, Emilia; Hinze, Christian; Park, Joon-Keun; Roël, Giulietta; Balogh, András; Choi, Mira; Wübken, Anne; Cording, Jimmi; Blasig, Ingolf E.; Luft, Friedrich C.; Scheidereit, Claus; Schmidt-Ott, Kai M.; Schmidt-Ullrich, Ruth; Müller, Dominik N.

2016-01-01

NF-κB is a key regulator of innate and adaptive immunity and is implicated in the pathogenesis of AKI. The cell type–specific functions of NF-κB in the kidney are unknown; however, the pathway serves distinct functions in immune and tissue parenchymal cells. We analyzed tubular epithelial-specific NF-κB signaling in a mouse model of ischemia-reperfusion injury (IRI)–induced AKI. NF-κB reporter activity and nuclear localization of phosphorylated NF-κB subunit p65 analyses in mice revealed that IRI induced widespread NF-κB activation in renal tubular epithelia and in interstitial cells that peaked 2–3 days after injury. To genetically antagonize tubular epithelial NF-κB activity, we generated mice expressing the human NF-κB super-repressor IκBαΔN in renal proximal, distal, and collecting duct epithelial cells. Compared with control mice, these mice exhibited improved renal function, reduced tubular apoptosis, and attenuated neutrophil and macrophage infiltration after IRI-induced AKI. Furthermore, tubular NF-κB–dependent gene expression profiles revealed temporally distinct functional gene clusters for apoptosis, chemotaxis, and morphogenesis. Primary proximal tubular cells isolated from IκBαΔN-expressing mice and exposed to hypoxia-mimetic agent cobalt chloride exhibited less apoptosis and expressed lower levels of chemokines than cells from control mice did. Our results indicate that postischemic NF-κB activation in renal tubular epithelia aggravates tubular injury and exacerbates a maladaptive inflammatory response. PMID:26823548
Identification of polymorphism in exons 7 and 12 of lactoferrin gene and its association with incidence of clinical mastitis in Murrah buffalo.

PubMed

Dinesh, Krishanender; Verma, Archana; Das Gupta, Ishwar; Thakur, Yash Pal; Verma, Nishant; Arya, Ashwani

2015-04-01

Lactoferrin gene is one of the important candidate genes for mastitis resistance. The gene is located on chromosome BTA 22 and consists of 17 exons spanning over 34.5 kb of genomic DNA. The present study was undertaken with the objectives to identify allelic variants in exons 7 and 12 of lactoferrin gene and to analyze association between its genetic variants and incidence of clinical mastitis in Murrah buffalo. The amplification of exons 7 and 12 of lactoferrin gene yielded amplicons of 232- and 461-bp sizes. PCR-restriction fragment length polymorphism (RFLP) analysis of 232-bp amplicon using BccI restriction enzyme revealed three genotypes (AA, AB, and BB) with frequencies of 0.62, 0.22, and 0.16, respectively. The frequencies of two alleles, A and B, were estimated as 0.73 and 0.27. Hpy188I-RFLP for 461-bp amplicon revealed polymorphism with three genotypes, CC, CD, and DD, with respective frequencies of 0.06, 0.39, and 0.56, whereas frequencies for C and D alleles were 0.25 and 0.75. The chi-square (χ(2)) analysis revealed a significant association between incidence of clinical mastitis and genetic variants of exon 7, and animals of AA genotype of exon 7 were found to be least susceptible to mastitis. The findings indicate potential scope for incorporation of lactoferrin gene in selection and breeding of Murrah buffaloes for improved genetic resistance to mastitis.
Human milk lactoferrin inactivates two putative colonization factors expressed by Haemophilus influenzae.

PubMed

Qiu, J; Hendrixson, D R; Baker, E N; Murphy, T F; St Geme, J W; Plaut, A G

1998-10-13

Haemophilus influenzae is a major cause of otitis media and other respiratory tract disease in children. The pathogenesis of disease begins with colonization of the upper respiratory mucosa, a process that involves evasion of local immune mechanisms and adherence to epithelial cells. Several studies have demonstrated that human milk is protective against H. influenzae colonization and disease. In the present study, we examined the effect of human milk on the H. influenzae IgA1 protease and Hap adhesin, two autotransported proteins that are presumed to facilitate colonization. Our results demonstrated that human milk lactoferrin efficiently extracted the IgA1 protease preprotein from the bacterial outer membrane. In addition, lactoferrin specifically degraded the Hap adhesin and abolished Hap-mediated adherence. Extraction of IgA1 protease and degradation of Hap were localized to the N-lobe of the bilobed lactoferrin molecule and were inhibited by serine protease inhibitors, suggesting that the lactoferrin N-lobe may contain serine protease activity. Additional experiments revealed no effect of lactoferrin on the H. influenzae P2, P5, and P6 outer-membrane proteins, which are distinguished from IgA1 protease and Hap by the lack of an N-terminal passenger domain or an extracellular linker region. These results suggest that human milk lactoferrin may attenuate the pathogenic potential of H. influenzae by selectively inactivating IgA1 protease and Hap, thereby interfering with colonization. Future studies should examine the therapeutic potential of lactoferrin, perhaps as a supplement in infant formulas.
Lactoferrin Isolation Using Monolithic Column Coupled with Spectrometric or Micro-Amperometric Detector

PubMed Central

Adam, Vojtech; Zitka, Ondrej; Dolezal, Petr; Zeman, Ladislav; Horna, Ales; Hubalek, Jaromir; Sileny, Jan; Krizkova, Sona; Trnkova, Libuse; Kizek, Rene

2008-01-01

Lactoferrin is a multifunctional protein with antimicrobial activity and others to health beneficial properties. The main aim of this work was to propose easy to use technique for lactoferrin isolation from cow colostrum samples. Primarily we utilized sodium dodecyl sulphate – polyacrylamide gel electrophoresis for isolation of lactoferrin from the real samples. Moreover we tested automated microfluidic Experion electrophoresis system to isolate lactoferrin from the collostrum sample. The well developed signal of lactoferrin was determined with detection limit (3 S/N) of 20 ng/ml. In spite of the fact that Experion is faster than SDS-PAGE both separation techniques cannot be used in routine analysis. Therefore we have tested third separation technique, ion exchange chromatography, using monolithic column coupled with UV-VIS detector (LC-UV-VIS). We optimized wave length (280 nm), ionic strength of the elution solution (1.5 M NaCl) and flow rate of the retention and elution solutions (0.25 ml/min and 0.75 ml/min. respectively). Under the optimal conditions the detection limit was estimated as 0.1 μg/ml of lactoferrin measured. Using LC-UV-VIS we determined that lactoferrin concentration varied from 0.5 g/l to 1.1 g/l in cow colostrums collected in the certain time interval up to 72 hours after birth. Further we focused on miniaturization of detection device. We tested amperometric detection at carbon electrode. The results encouraged us to attempt to miniaturise whole detection system and to test it on analysis of real samples of human faeces, because lactoferrin level in faeces is closely associated with the inflammations of intestine mucous membrane. For the purpose of miniaturization we employed the technology of printed electrodes. The detection limit of lactoferrin was estimated as 10 μg/ml measured by the screen-printed electrodes fabricated by us. The fabricated electrodes were compared with commercially available ones. It follows from the obtained
Oral lactoferrin protects against experimental candidiasis in mice

PubMed Central

Velliyagounder, Kabilan; Alsaedi, Wijdan; Alabdulmohsen, Waad; Markowitz, Kenneth; Fine, Daniel H.

2015-01-01

Aims To determine the role of lactoferrin in protecting the oral cavities of mice against Candida albicans infection in lactoferrin knockout (LFKO−/−) mice were compared to wild-type (WT) mice. We also determine the protective role of human lactoferrin in the LFKO−/− mice. Methods and Results Antibiotic treated immunosuppressed mice were inoculated with C. albicans (or sham infection) by oral swab and evaluated for the severity of infection after 7 days of infection. To determine the protective role of hLF, we added 0.3% solution of hLF to the drinking water given to some of the mice. CFU count, scoring of lesions and microscopic observations were carried out to determine the severity of infection. LFKO−/−I mice showed a 2 log (P=0.001) higher CFUs of C. albicans in the oral cavity compared to the WTI mice. LFKO−/−I mice given hLF had a 3 log (P=0.001) reduction in CFUs in the oral cavity compared to untreated LFKO−/−I mice. The severity of infection, observed by light microscopy revealed that the tongue of the LFKO−/−I mice showed more white patches compared to WTI and LFKO−/−I+hLF mice. Scanning electron microscopic observation revealed that more filiform papillae were destroyed in LFKO−/−I mice when compared to WTI or LFKO−/−I +hLF mice. Conclusions Human lactoferrin is important in protecting mice from oral C. albicans infection. Administered hLF may be used to prevent C. albicans infection. Significance and Impact of the Study Human lactoferrin, a multifunctional iron-binding glycoprotein can be used as a therapeutic active ingredient in oral health care products against C. albicans. PMID:25319508
Whey Protein Components - Lactalbumin and Lactoferrin - Improve Energy Balance and Metabolism.

PubMed

Zapata, Rizaldy C; Singh, Arashdeep; Pezeshki, Adel; Nibber, Traj; Chelikani, Prasanth K

2017-08-30

Whey protein promotes weight loss and improves diabetic control, however, less is known of its bioactive components that produce such benefits. We compared the effects of normal protein (control) diet with high protein diets containing whey, or its fractions lactalbumin and lactoferrin, on energy balance and metabolism. Diet-induced obese rats were randomized to isocaloric diets: Control, Whey, Lactalbumin, Lactoferrin, or pair-fed to lactoferrin. Whey and lactalbumin produced transient hypophagia, whereas lactoferrin caused prolonged hypophagia; the hypophagia was likely due to decreased preference. Lactalbumin decreased weight and fat gain. Notably, lactoferrin produced sustained weight and fat loss, and attenuated the reduction in energy expenditure associated with calorie restriction. Lactalbumin and lactoferrin decreased plasma leptin and insulin, and lactalbumin increased peptide YY. Whey, lactalbumin and lactoferrin improved glucose clearance partly through differential upregulation of glucoregulatory transcripts in the liver and skeletal muscle. Interestingly, lactalbumin and lactoferrin decreased hepatic lipidosis partly through downregulation of lipogenic and/or upregulation of β-oxidation transcripts, and differentially modulated cecal bacterial populations. Our findings demonstrate that protein quantity and quality are important for improving energy balance. Dietary lactalbumin and lactoferrin improved energy balance and metabolism, and decreased adiposity, with the effects of lactoferrin being partly independent of caloric intake.

Lactoferrin-Immobilized Surfaces onto Functionalized PLA Assisted by the Gamma-Rays and Nitrogen Plasma to Create Materials with Multifunctional Properties.

PubMed

Stoleru, Elena; Zaharescu, Traian; Hitruc, Elena Gabriela; Vesel, Alenka; Ioanid, Emil G; Coroaba, Adina; Safrany, Agnes; Pricope, Gina; Lungu, Maria; Schick, Christoph; Vasile, Cornelia

2016-11-23

Both cold nitrogen radiofrequency plasma and gamma irradiation have been applied to activate and functionalize the polylactic acid (PLA) surface and the subsequent lactoferrin immobilization. Modified films were comparatively characterized with respect to the procedure of activation and also with unmodified sample by water contact angle measurements, mass loss, X-ray photoelectron spectroscopy (XPS), attenuated total reflectance-Fourier transform infrared spectroscopy (ATR-FTIR), atomic force microscopy (AFM), and chemiluminescence measurements. All modified samples exhibit enhanced surface properties mainly those concerning biocompatibility, antimicrobial, and antioxidant properties, and furthermore, they are biodegradable and environmentally friendly. Lactoferrin deposited layer by covalent coupling using carbodiimide chemistry showed a good stability. It was found that the lactoferrin-modified PLA materials present significantly increased oxidative stability. Gamma-irradiated samples and lactoferrin-functionalized samples show higher antioxidant, antimicrobial, and cell proliferation activity than plasma-activated and lactoferrin-functionalized ones. The multifunctional materials thus obtained could find application as biomaterials or as bioactive packaging films.
Angiogenin: a novel inhibitor of neutrophil lactoferrin release during extracorporeal circulation.

PubMed

Schmaldienst, Sabine; Oberpichler, André; Tschesche, Harald; Hörl, Walter H

2003-01-01

Degranulation of polymorphonuclear leukocytes (PMNL) occurs during extracorporeal circulation. A degranulation-inhibiting protein identical to angiogenin was recently isolated from high-flux dialyzer ultrafiltrate. This protein inhibits the release of lactoferrin and metalloproteinases from PMNL in vitro. In the present study, we investigated end-stage renal disease patients undergoing regular hemodialysis treatment with either high-flux dialyzers (n = 51) or low-flux dialyzers (n = 44), and chronically uremic patients undergoing hemodiafiltration (n = 30). Hemodialysis therapy with low-flux polysulfone or cellulose triacetate membranes caused no or only minimal reduction (lactoferrin release from PMNL. Hemodialysis therapy with high-flux membranes (e.g. cellulose triacetate, polymethylmethacrylate) or hemodiafiltration resulted in a reduction of plasma angiogenin levels by 20-40% after 2 h associated with a nearly 4-fold PMNL lactoferrin release. The release of PMNL elastase was not affected by the different treatment modalities used. We conclude that high angiogenin plasma levels protect against lactoferrin release from PMNL during extracorporeal circulation in chronically uremic patients. A decrease of plasma angiogenin between 20 and 40% during extracorporeal circulation, however, results in marked PMNL lactoferrin release. This novel mechanism may explain, at least in part, PMNL degranulation also in non complement activating high-flux membranes. Copyright 2003 S. Karger AG, Basel
21 CFR 866.5570 - Lactoferrin immunological test system.

Code of Federal Regulations, 2011 CFR

2011-04-01

... that consists of the reagents used to measure by immunochemical techniques the lactoferrin (an iron... fluids, and tissues. Measurement of lactoferrin may aid in the diagnosis of an inherited deficiency of...
21 CFR 866.5570 - Lactoferrin immunological test system.

Code of Federal Regulations, 2014 CFR

2014-04-01

... that consists of the reagents used to measure by immunochemical techniques the lactoferrin (an iron... fluids, and tissues. Measurement of lactoferrin may aid in the diagnosis of an inherited deficiency of...
21 CFR 866.5570 - Lactoferrin immunological test system.

Code of Federal Regulations, 2012 CFR

2012-04-01

... that consists of the reagents used to measure by immunochemical techniques the lactoferrin (an iron... fluids, and tissues. Measurement of lactoferrin may aid in the diagnosis of an inherited deficiency of...
21 CFR 866.5570 - Lactoferrin immunological test system.

Code of Federal Regulations, 2013 CFR

2013-04-01

... that consists of the reagents used to measure by immunochemical techniques the lactoferrin (an iron... fluids, and tissues. Measurement of lactoferrin may aid in the diagnosis of an inherited deficiency of...
Amorfrutin A inhibits TNF-α-induced NF-κB activation and NF-κB-regulated target gene products.

PubMed

Shi, Hui; Ma, Juan; Mi, Chunliu; Li, Jing; Wang, Fei; Lee, Jung Joon; Jin, Xuejun

2014-07-01

The nuclear factor-κB (NF-κB) transcription factors control many physiological processes including inflammation, immunity, apoptosis, and angiogenesis. In our search for NF-κB inhibitors from natural resources, we identified amorfrutin A as an inhibitor of NF-κB activation from the fruits of Amorpha fruticosa L. In present study, this compound significantly inhibited the TNF-α-induced expression of NF-κB reporter gene. Further analysis revealed that amorfrutin A was a potent inhibitor of NF-κB activation by the suppression of TNF-α-induced inhibitor of κBα (IκBα) degradation, p65 nuclear translocation, and DNA-binding activity of NF-κB. We also demonstrated that pretreatment of cells with this compound prevented the TNF-α-induced expression of NF-κB target genes, such as antiapoptosis (cIAP-1 and FLIP), proliferation (COX-2 and cyclinD1), invasion (MMP-9), angiogenesis (VEGF), and major inflammatory cytokines (TNF-α, IL-8, and MCP1). Furthermore, our results suggest that amorfrutin A potentiates TNF-α-induced apoptosis. Taken together, amorfrutin A could be a valuable candidate for the intervention of NF-κB-dependent pathological conditions such as inflammation. Copyright © 2014 Elsevier B.V. All rights reserved.
Role of Lactoferrin in Neonates and Infants: An Update.

PubMed

Manzoni, Paolo; Dall'Agnola, Alberto; Tomé, Daniel; Kaufman, David A; Tavella, Elena; Pieretto, Marta; Messina, Alessandro; De Luca, Daniele; Bellaiche, Marc; Mosca, Alexis; Piloquet, Hugues; Simeoni, Umberto; Picaud, Jean-Charles; Del Vecchio, Antonio

2018-05-01

Lactoferrin is one of the most represented and important bioactive proteins in human and mammal milk. In humans, lactoferrin is responsible for several actions targeting anti-infective, immunological, and gastrointestinal domains in neonates, infants, and young children. Evidence-based data vouch for the ability of supplemented lactoferrin to prevent sepsis and necrotizing enterocolitis in preterm infants and to reduce the burden of morbidity related to gastrointestinal and respiratory pathogens in young children. However, several issues remain pending regarding answers and clarification related to quality control, correct intakes, optimal schedules and schemes of supplementations, interactions with probiotics, and different types of milk and formulas. This review summarizes the current evidence regarding lactoferrin and discusses the areas in need of further guidance prior to the adoption of strategies that include a routine use of lactoferrin in neonates and young children. Thieme Medical Publishers 333 Seventh Avenue, New York, NY 10001, USA.
MicroRNA-22 and microRNA-140 suppress NF-{kappa}B activity by regulating the expression of NF-{kappa}B coactivators

DOE Office of Scientific and Technical Information (OSTI.GOV)

Takata, Akemi; Otsuka, Motoyuki, E-mail: otsukamo-tky@umin.ac.jp; Kojima, Kentaro

2011-08-12

Highlights: {yields} miRNAs were screened for their ability to regulate NF-{kappa}B activity. {yields} miRNA-22 and miRNA-140-3p suppress NF-{kappa}B activity by regulating coactivators. {yields} miRNA-22 targets nuclear receptor coactivator 1 (NCOA1). {yields} miRNA-140-3p targets nuclear receptor-interacting protein 1 (NRIP1). -- Abstract: Nuclear factor {kappa}B (NF-{kappa}B) is a transcription factor that regulates a set of genes that are critical to many biological phenomena, including liver tumorigenesis. To identify microRNAs (miRNAs) that regulate NF-{kappa}B activity in the liver, we screened 60 miRNAs expressed in hepatocytes for their ability to modulate NF-{kappa}B activity. We found that miRNA-22 and miRNA-140-3p significantly suppressed NF-{kappa}B activity bymore » regulating the expression of nuclear receptor coactivator 1 (NCOA1) and nuclear receptor-interacting protein 1 (NRIP1), both of which are NF-{kappa}B coactivators. Our results provide new information about the roles of miRNAs in the regulation of NF-{kappa}B activity.« less
A preliminary approach to creating an overview of lactoferrin multi-functionality utilizing a text mining method.

PubMed

Shimazaki, Kei-ichi; Kushida, Tatsuya

2010-06-01

Lactoferrin is a multi-functional metal-binding glycoprotein that exhibits many biological functions of interest to many researchers from the fields of clinical medicine, dentistry, pharmacology, veterinary medicine, nutrition and milk science. To date, a number of academic reports concerning the biological activities of lactoferrin have been published and are easily accessible through public data repositories. However, as the literature is expanding daily, this presents challenges in understanding the larger picture of lactoferrin function and mechanisms. In order to overcome the "analysis paralysis" associated with lactoferrin information, we attempted to apply a text mining method to the accumulated lactoferrin literature. To this end, we used the information extraction system GENPAC (provided by Nalapro Technologies Inc., Tokyo). This information extraction system uses natural language processing and text mining technology. This system analyzes the sentences and titles from abstracts stored in the PubMed database, and can automatically extract binary relations that consist of interactions between genes/proteins, chemicals and diseases/functions. We expect that such information visualization analysis will be useful in determining novel relationships among a multitude of lactoferrin functions and mechanisms. We have demonstrated the utilization of this method to find pathways of lactoferrin participation in neovascularization, Helicobacter pylori attack on gastric mucosa, atopic dermatitis and lipid metabolism.
Bovine lactoferrin and lactoferricin exert antitumor activities on human colorectal cancer cells (HT-29) by activating various signaling pathways.

PubMed

Jiang, Rulan; Lönnerdal, Bo

2017-02-01

Lactoferrin (Lf) is an iron-binding glycoprotein that is present at high concentrations in milk. Bovine lactoferricin (LfcinB) is a peptide fragment generated by pepsin proteolysis of bovine lactoferrin (bLf). LfcinB consists of amino acid residues 17-41 proximal to the N-terminus of bLf and a disulfide bond between residues 19 and 36, forming a loop. Both bLf and LfcinB have been demonstrated to have antitumor activities. Colorectal cancer is the second most common cause of cancer death in developed countries. We hypothesized that bLf and LfcinB exert antitumor activities on colon cancer cells (HT-29) by triggering various signaling pathways. bLf and LfcinB significantly induced apoptosis in HT-29 cells but not in normal human intestinal epithelial cells, as revealed by the ApoTox-Glo Triplex Assay. The LIVE/DEAD cell viability assay showed that both bLf and LfcinB reduced the viability of HT-29 cells. Transcriptome analysis indicated that bLf, cyclic LfcinB, and linear LfcinB exerted antitumor activities by differentially activating diverse signaling pathways, including p53, apoptosis, and angiopoietin signaling. Immunoblotting results confirmed that both bLf and LfcinBs increased expression of caspase-8, p53, and p21, critical proteins in tumor suppression. These results provide valuable information regarding bLf and LfcinB for potential clinical applications in colon cancer therapy.
Acetaminophen-induced hepatotoxicity is associated with early changes in NF-kB and NF-IL6 DNA binding activity.

PubMed

Blazka, M E; Germolec, D R; Simeonova, P; Bruccoleri, A; Pennypacker, K R; Luster, M I

Nuclear transcription factors, such as NF-kB and NF-IL6, are believed to play an important role in regulating the expression of genes that encode for products involved in tissue damage and inflammation and, thus, may represent early biomarkers for chemical toxicities. In the present study changes in DNA binding activity of these factors were examined in livers of mice administered hepatotoxic doses of acetaminophen (APAP). NF-kB and NF-IL6 DNA binding occurred constitutively in control mouse liver. However, within 4 hr following administration of hepatotoxic doses of APAP, their binding activities were transiently lost and is in contrast to AP-1 transcription factor where activation occurs under similar conditions. These changes corresponded with increased release of inflammatory mediators (IL-6, serum amyloid A) and increased levels of enzymatic markers of hepatocyte damage. Similarly, treatment of mice with gadolinium chloride, an inhibitor of Kupffer cell activation and known to protect against APAP-induced hepatotoxicity, reduced the observed pathophysiological response in the liver while altering the APAP-associated changes in NF-kB DNA binding activity. NF-kB was found predominantly in parenchymal and endothelial cells and was composed primarily of relatively inactive p50 homodimer subunits in control liver. Taken together, these studies suggest that hepatotoxicity is associated with early and complex changes in DNA binding activities of specific transcription factors. In particular, NF-kB and NF-IL6 may serve as negative regulators of hepatocyte-derived inflammatory mediators and is analogous to that previously observed in certain other cell systems such as B lymphocytes.
Antibacterial activity in bovine lactoferrin-derived peptides.

PubMed Central

Hoek, K S; Milne, J M; Grieve, P A; Dionysius, D A; Smith, R

1997-01-01

Several peptides sharing high sequence homology with lactoferricin B (Lf-cin B) were generated from bovine lactoferrin (Lf) with recombinant chymosin. Two peptides were copurified, one identical to Lf-cin B and another differing from Lf-cin B by the inclusion of a C-terminal alanine (lactoferricin). Two other peptides were copurified from chymosin-hydrolyzed Lf, one differing from Lf-cin B by the inclusion of C-terminal alanyl-leucine and the other being a heterodimer linked by a disulfide bond. These peptides were isolated in a single step from chymosin-hydrolyzed Lf by membrane ion-exchange chromatography and were purified by reverse-phase high-pressure liquid chromatography (HPLC). They were characterized by N-terminal Edman sequencing, mass spectrometry, and antibacterial activity determination. Pure lactoferricin, prepared from pepsin-hydrolyzed Lf, was purified by standard chromatography techniques. This peptide was analyzed against a number of gram-positive and gram-negative bacteria before and after reduction of its disulfide bond or cleavage after its single methionine residue and was found to inhibit the growth of all the test bacteria at a concentration of 8 microM or less. Subfragments of lactoferricin were isolated from reduced and cleaved peptide by reverse-phase HPLC. Subfragment 1 (residues 1 to 10) was active against most of the test microorganisms at concentrations of 10 to 50 microM. Subfragment 2 (residues 11 to 26) was active against only a few microorganisms at concentrations up to 100 microM. These antibacterial studies indicate that the activity of lactoferricin is mainly, but not wholly, due to its N-terminal region. PMID:8980754
Porcine arterivirus activates the NF-{kappa}B pathway through I{kappa}B degradation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lee, Sang-Myeong; Kleiboeker, Steven B.

2005-11-10

Nuclear factor-kappaB (NF-{kappa}B) is a critical regulator of innate and adaptive immune function as well as cell proliferation and survival. The present study demonstrated for the first time that a virus belonging to the Arteriviridae family activates NF-{kappa}B in MARC-145 cells and alveolar macrophages. In porcine reproductive and respiratory syndrome virus (PRRSV)-infected cells, NF-{kappa}B activation was characterized by translocation of NF-{kappa}B from the cytoplasm to the nucleus, increased DNA binding activity, and NF-{kappa}B-regulated gene expression. NF-{kappa}B activation was increased as PRRSV infection progressed and in a viral dose-dependent manner. UV-inactivation of PRRSV significantly reduced the level of NF-{kappa}B activation. Degradationmore » of I{kappa}B protein was detected late in PRRSV infection, and overexpression of the dominant negative form of I{kappa}B{alpha} (I{kappa}B{alpha}DN) significantly suppressed NF-{kappa}B activation induced by PRRSV. However, I{kappa}B{alpha}DN did not affect viral replication and viral cytopathic effect. PRRSV infection induced oxidative stress in cells by generating reactive oxygen species (ROS), and antioxidants inhibited NF-{kappa}B DNA binding activity in PRRSV-infected cells, suggesting ROS as a mechanism by which NF-{kappa}B was activated by PRRSV infection. Moreover, NF-{kappa}B-dependent expression of matrix metalloproteinase (MMP)-2 and MMP-9 was observed in PRRSV-infected cells, an observation which implies that NF-{kappa}B activation is a biologically significant aspect of PRRSV pathogenesis. The results presented here provide a basis for understanding molecular pathways of pathology and immune evasion associated with disease caused by PRRSV.« less
Lactoferrin Dampens High-Fructose Corn Syrup-Induced Hepatic Manifestations of the Metabolic Syndrome in a Murine Model

PubMed Central

Li, Yi-Chieh; Hsieh, Chang-Chi

2014-01-01

Hepatic manifestations of the metabolic syndrome are related obesity, type 2 diabetes/insulin resistance and non-alcoholic fatty liver disease. Here we investigated how the anti-inflammatory properties of lactoferrin can protect against the onset of hepatic manifestations of the metabolic syndrome by using a murine model administered with high-fructose corn syrup. Our results show that a high-fructose diet stimulates intestinal bacterial overgrowth and increases intestinal permeability, leading to the introduction of endotoxin into blood circulation and liver. Immunohistochemical staining of Toll-like receptor-4 and thymic stromal lymphopoietin indicated that lactoferrin can modulate lipopolysaccharide-mediated inflammatory cascade. The important regulatory roles are played by adipokines including interleukin-1β, interleukin-6, tumor necrosis factor-α, monocyte chemotactic protein-1, and adiponectin, ultimately reducing hepatitis and decreasing serum alanine aminotransferase release. These beneficial effects of lactoferrin related to the downregulation of the lipopolysaccharide-induced inflammatory cascade in the liver. Furthermore, lactoferrin reduced serum and hepatic triglycerides to prevent lipid accumulation in the liver, and reduced lipid peroxidation, resulting in 4-hydroxynonenal accumulation. Lactoferrin reduced oral glucose tolerance test and homeostasis model assessment-insulin resistance. Lactoferrin administration thus significantly lowered liver weight, resulting from a decrease in the triglyceride and cholesterol synthesis that activates hepatic steatosis. Taken together, these results suggest that lactoferrin protected against high-fructose corn syrup induced hepatic manifestations of the metabolic syndrome. PMID:24816278
Lactoferrin dampens high-fructose corn syrup-induced hepatic manifestations of the metabolic syndrome in a murine model.

PubMed

Li, Yi-Chieh; Hsieh, Chang-Chi

2014-01-01

Hepatic manifestations of the metabolic syndrome are related obesity, type 2 diabetes/insulin resistance and non-alcoholic fatty liver disease. Here we investigated how the anti-inflammatory properties of lactoferrin can protect against the onset of hepatic manifestations of the metabolic syndrome by using a murine model administered with high-fructose corn syrup. Our results show that a high-fructose diet stimulates intestinal bacterial overgrowth and increases intestinal permeability, leading to the introduction of endotoxin into blood circulation and liver. Immunohistochemical staining of Toll-like receptor-4 and thymic stromal lymphopoietin indicated that lactoferrin can modulate lipopolysaccharide-mediated inflammatory cascade. The important regulatory roles are played by adipokines including interleukin-1β, interleukin-6, tumor necrosis factor-α, monocyte chemotactic protein-1, and adiponectin, ultimately reducing hepatitis and decreasing serum alanine aminotransferase release. These beneficial effects of lactoferrin related to the downregulation of the lipopolysaccharide-induced inflammatory cascade in the liver. Furthermore, lactoferrin reduced serum and hepatic triglycerides to prevent lipid accumulation in the liver, and reduced lipid peroxidation, resulting in 4-hydroxynonenal accumulation. Lactoferrin reduced oral glucose tolerance test and homeostasis model assessment-insulin resistance. Lactoferrin administration thus significantly lowered liver weight, resulting from a decrease in the triglyceride and cholesterol synthesis that activates hepatic steatosis. Taken together, these results suggest that lactoferrin protected against high-fructose corn syrup induced hepatic manifestations of the metabolic syndrome.
Lactoferrin: A bioinspired, anti-biofilm therapeutic

PubMed Central

Ammons, M.C.; Copié, V.

2013-01-01

Medically relevant biofilms have gained a significant level of interest, in part because of the epidemic rise in obesity and an aging population in the developed world. The associated comorbidities of chronic wounds such as pressure ulcers, venous leg ulcers, and diabetic foot wounds remain recalcitrant to the therapies available currently. Development of chronicity in the wound is due primarily to an inability to complete the wound healing process owing to the presence of a bioburden, specifically bacterial biofilms. New therapies are clearly needed which specifically target biofilms. Lactoferrin is a multifaceted molecule of the innate immune system found primarily in milk. While further investigation is warranted to elucidate mechanisms of action, in vitro analyses of lactoferrin and its derivatives have demonstrated that these complex molecules are structurally and functionally well suited to address the heterogeneity of bacterial biofilms. In addition, use of lactoferrin and its derivatives has proven promising in the clinic. PMID:23574002
NF-{kappa}B regulates Lef1 gene expression in chondrocytes

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yun, Kangsun; Choi, Yoo Duk; Nam, Jong Hee

The relation of Wnt/{beta}-catenin signaling to osteoarthritis progression has been revealed with little information on the underlying molecular mechanism. In this study we found overexpression of Lef1 in cartilage tissue of osteoarthritic patients and elucidated molecular mechanism of NF-{kappa}B-mediated Lef1 gene regulation in chondrocytes. Treatment of IL-1{beta} augmented Lef1 upregulation and nuclear translocation of NF-{kappa}B in chondrocytes. Under IL-1{beta} signaling, treatment of NF-{kappa}B nuclear translocation inhibitor SN-50 reduced Lef1 expression. A conserved NF-{kappa}B-binding site between mouse and human was selected through bioinformatic analysis and mapped at the 14 kb upstream of Lef1 transcription initiation site. NF-{kappa}B binding to the sitemore » was confirmed by chromatin immunoprecipitation assay. Lef1 expression was synergistically upregulated by interactions of NF-{kappa}B with Lef1/{beta}-catenin in chondrocytes. Our results suggest a pivotal role of NF-{kappa}B in Lef1 expression in arthritic chondrocytes or cartilage degeneration.« less
Lactoferrin reduces mortality in preweaned calves with diarrhea.

PubMed

Habing, G; Harris, K; Schuenemann, G M; Piñeiro, J M; Lakritz, J; Clavijo, X Alcaraz

2017-05-01

Calf diarrhea is the most common reason for mortality and antimicrobial therapy in preweaned calves on dairy farms in the United States. Conventional and organic livestock producers require alternative therapies for calf diarrhea to reduce the necessity of conventional antimicrobials. Alternatives administered for mild cases or early in the disease course may be useful to mitigate disease progression and reduce the likelihood of septicemia and negative sequelae. Lactoferrin is a bioactive protein naturally found in colostrum that has been shown to prevent septicemia in high-risk infants. Among organic producers, garlic extract is widely used for the treatment of disease and perceived to be efficacious. The objectives of the study were to determine the effectiveness of lactoferrin and garlic extract to reduce mortality and culling, improve weight gain, and reduce the duration of disease in preweaned calves with the first diagnosis of diarrhea. In total, 628 calves with diarrhea from a single commercial dairy were enrolled in a blinded, randomized field trial. Calves diagnosed with diarrhea (fecal score ≥3), were randomized to 3 consecutive days of oral garlic extract, lactoferrin, or water (control). Calves were clinically evaluated for up to 10 d. Body weight was measured at enrollment and 10 d later. For calves receiving garlic extract, the risk of death or culling was not significantly different than calves in the control group; however, calves that received lactoferrin had approximately half the risk of death or culling in the 120 d following diagnosis. Additionally, the relative risk of death or culling in the 60 d following diagnosis was significantly lower for the subset of calves with severe diarrhea at enrollment. Neither garlic nor lactoferrin had a significant effect on disease duration or average weight gain during the 10-d period. Lactoferrin significantly reduced mortality and culling when administered to preweaned calves with the first diagnosis of
Apoptotic effects of bovine apo-lactoferrin on HeLa tumor cells.

PubMed

Luzi, Carla; Brisdelli, Fabrizia; Iorio, Roberto; Bozzi, Argante; Carnicelli, Veronica; Di Giulio, Antonio; Lizzi, Anna Rita

2017-01-01

Lactoferrin (Lf), a cationic iron-binding glycoprotein of 80 kDa present in body secretions, is known as a compound with marked antimicrobial activity. In the present study, the apoptotic effect of iron-free bovine lactoferrin (apo-bLf) on human epithelial cancer (HeLa) cells was examined in association with reactive oxygen species and glutathione (GSH) levels. Apoptotic effect of iron-free bovine lactoferrin inhibited the growth of HeLa cells after 48 hours of treatment while the diferric-bLf was ineffective in the concentration range tested (from 1 to 12.5 μM). Western blot analysis showed that key apoptotic regulators including Bax, Bcl-2, Sirt1, Mcl-1, and PARP-1 were modulated by 1.25 μM of apo-bLf. In the same cell line, apo-bLf induced apoptosis together with poly (ADP-ribose) polymerase cleavage, caspase activation, and a significant drop of NAD + . In addition, apo-bLf-treated HeLa cells showed a marked increase of reactive oxygen species level and a significant GSH depletion. On the whole, apo-bLf triggered apoptosis of HeLa cells upon oxygen radicals burst and GSH decrease. Copyright © 2017 John Wiley & Sons, Ltd.

Neuroprotective effect of curcumin-loaded lactoferrin nano particles against rotenone induced neurotoxicity.

PubMed

Bollimpelli, V Satish; Kumar, Prashant; Kumari, Sonali; Kondapi, Anand K

2016-05-01

Curcumin is known to have neuroprotective role and possess antioxidant, anti-inflammatory activities. Rotenone, a flavonoid induced neurotoxicity in dopaminergic cells is being widely studied in Parkinson's Disease (PD) research. In the present study, curcumin loaded lactoferrin nano particles prepared by sol-oil chemistry were used to protect dopaminergic cell line SK-N-SH against rotenone induced neurotoxicity. These curcumin loaded nano particles were of 43-60 nm diameter size and around 100 nm hydrodynamic size as assessed by transmission electron microscopy, atomic force microscopy and dynamic light scattering analysis respectively. The encapsulation efficiency was 61.3% ± 2.4%. Cellular uptake of curcumin through these nano particles was confirmed by confocal imaging and spectrofluorimetric analysis. The curcumin loaded lactoferrin nanoparticles showed greater intracellular drug uptake, sustained retention and greater neuroprotection than soluble counterpart. Neuroprotective activity was characterized through viability assays and by estimating ROS levels. Furthermore rotenone induced PD like features were characterized by decrease in tyrosine hydroxylase expression and increase in α-synuclein expression. Taken together curcumin loaded lactoferrin nanoparticles could be a promising drug delivery strategy against neurotoxicity in dopaminergic neurons. Copyright © 2016 Elsevier Ltd. All rights reserved.
Molecular evidence of stereo-specific lactoferrin dimers in solution.

PubMed

Persson, Björn A; Lund, Mikael; Forsman, Jan; Chatterton, Dereck E W; Akesson, Torbjörn

2010-10-01

Gathering experimental evidence suggests that bovine as well as human lactoferrin self-associate in aqueous solution. Still, a molecular level explanation is unavailable. Using force field based molecular modeling of the protein-protein interaction free energy we demonstrate (1) that lactoferrin forms highly stereo-specific dimers at neutral pH and (2) that the self-association is driven by a high charge complementarity across the contact surface of the proteins. Our theoretical predictions of dimer formation are verified by electrophoretic mobility and N-terminal sequence analysis on bovine lactoferrin. 2010 Elsevier B.V. All rights reserved.
[Presence of lactoferrin (LF) in lymphocutaneous sporotrichosis. Yeast-bound antimicrobial peptide].

PubMed

Palma-Ramos, Alejandro; Castrillón-Rivera, Laura E; Vega-Mémije, María Elisa; Arenas-Guzmán, Roberto; Rangel-Gamboa, Lucía

Sporotrichosis is a common subcutaneous mycosis in Latin America, produced by dimorphic fungi belong to Sporothrix schenckii complex of cryptic species. Infection is acquired by traumatic inoculation with contaminated organic material. Host immune response includes polymorphonuclear neutrophils chemotaxis and release of granular components. Lactoferrin is a protein member of the transferrin family of iron-binding proteins, present inside polymorphonuclear granular structure, and has been reported to affect growth and development of infectious agents, including fungal organisms. Nevertheless, lactoferrin expression in sporotrichosis infections has not been reported yet. To determine the expression of lactoferrin using immunohistochemical staining in sporotrichosis human infection. The dermatology department's files during a period of five years were reviewed; cases with a diagnosis of sporotrichosis were selected and lactoferrin immunostaining was performed when enough biological material was available. Three cases with a diagnosis of sporotrichosis and adequate biological material on paraffin block were identified. In all cases, lactoferrin immunostaining was positive around yeast cell.
New Epigenetic Therapeutic Intervention for Metastatic Breast Cancer

DTIC Science & Technology

2016-04-01

also discovered that constitutively activated NF -kB in TNBC sustains prolonged activation of pro-inflammatory cytokines, enabling rapid spread...metastasis) of TNBC tumors. Notably, the functions of both transcription factors Twist and NF -kB in gene activation require lysine acetylation, which signs...proteins including Twist, NF -kB and STAT3. b. Define the molecular basis of the BET BrDs’ selective interactions with effector proteins through structure
Supplemental lactoferrin improves health and growth of Holstein calves during the preweaning phase.

PubMed

Robblee, E D; Erickson, P S; Whitehouse, N L; McLaughlin, A M; Schwab, C G; Rejman, J J; Rompala, R E

2003-04-01

Lactoferrin is a milk protein that exhibits broad-spectrum antimicrobial properties. Previous studies indicated that supplemental lactoferrin may alter the microbial populations in the gut of nonruminants and increase preweaning weight gains in calves. In the present study, 40 Holstein calves were used to examine the effects of supplemental lactoferrin (0, 1, 2, or 3 g/d) on health, growth, and feed intake from 3 d of age to 2 wk postweaning. Lactoferrin was mixed and fed with a nonmedicated milk replacer. Calves were housed in individual pens and offered a textured, nonmedicated starter and water for ad libitum consumption. Body weight and heart girth were measuredweekly. Intakes of milk replacer and starter were determined daily. Fecal consistency was monitored three times per week. Calves were weaned when they met certain criteria based on body weight gain and starter intake. Preweaning fecal score responded quadratically, with the group fed 1 g/d of lactoferrin having the lowest score. Overall and preweaning number of days medicated responded in the same manner as fecal score. Preweaning average daily gain and gain-to-feed ratio increased linearly with lactoferrin supplementation, whereas postweaning gain-to-feed ratio decreased linearly with lactoferrin. Overall average daily heart girth gain increased linearly with lactoferrin. Body weight, weaning age, and dry matter intake were not different among treatments. Based on the observed improved gain-to-feed ratios, increased average daily gains, improved fecal scores, and reduced morbidity in preweaned calves, it appears that lactoferrin may be a beneficial supplement in the diets of neonatal calves prior to weaning.
Novel recombinant human lactoferrin: differential activation of oxidative stress related gene expression.

PubMed

Kruzel, Marian L; Actor, Jeffrey K; Zimecki, Michał; Wise, Jasen; Płoszaj, Paulina; Mirza, Shaper; Kruzel, Mark; Hwang, Shen-An; Ba, Xueqing; Boldogh, Istvan

2013-12-01

Lactoferrin, an iron-binding protein found in high concentrations in mammalian exocrine secretions, is an important component of the host defense system. It is also a major protein of the secondary granules of neutrophils from which is released upon activation. Due to its potential clinical utility, recombinant human lactoferrin (rhLF) has been produced in various eukaryotic expression systems; however, none of these are fully compatible with humans. Most of the biopharmaceuticals approved by the FDA for use in humans are produced in mammalian expression systems. The Chinese hamster ovary cells (CHO) have become the system of choice for proteins that require post-translational modifications, such as glycoproteins. The aim of this study was to scale-up expression and purification of rhLF in a CHO expression system, verify its glycan primary structure, and assess its biological properties in cell culture models. A stable CHO cell line producing >200mg/L of rhLF was developed and established. rhLF was purified by a single-step cation-exchange chromatography procedure. The highly homogenous rhLF has a molecular weight of approximately 80 kDa. MALDI-TOF mass spectrometric analysis revealed N-linked, partially sialylated glycans at two glycosylation sites, typical for human milk LF. This novel rhLF showed a protective effect against oxidative stress in a similar manner to its natural counterpart. In addition, rhLF revealed a modulatory effect on cellular redox via upregulation of key antioxidant enzymes. These data imply that the CHO-derived rhLF is fully compatible with the native molecule, thus it has promise for human therapeutic applications. Copyright © 2013 Elsevier B.V. All rights reserved.
NF-kappaB activation in cancer: a challenge for ubiquitination- and proteasome-based therapeutic approach.

PubMed

Amit, Sharon; Ben-Neriah, Yinon

2003-02-01

Nuclear factor-kappa B (NF-kappaB) activation relies primarily on ubiquitin-mediated degradation of its inhibitor IkappaB. NF-kappaB plays an important role in many aspects of tumor development, progression, and therapy. Some types of cancer are characterized by constitutive NF-kappaB activity, whereas in others such activity is induced following chemotherapy. NF-kappaB-harboring tumors are generally resistant to chemotherapy and their eradication requires NF-kappaB inhibition. Here we describe the mechanisms of NF-kappaB activation in normal and tumor cells, review prevalent notions regarding the factor's contribution to tumorigenicity and discuss present and future options for NF-kappaB inhibition in cancer. The ubiquitination-mediated activation of NF-kappaB is intersected by another cancer-associated protein, beta-catenin. We, therefore, compare the related activation pathways and discuss the possibility of differential targeting of the involved ubiquitination machinery. Copyright 2002 Elsevier Science Ltd.
[Effect of NF-κB activation on the radiation response of esophageal cancer cells].

PubMed

Li, Baozhong; Chen, Zhaoli; Zhou, Fang; He, Jie

2014-07-01

To investigate the effect of NF-κB activation on radiation response of esophageal carcinoma. The expression of NF-κB was detected in pretreatment and posttreatment specimens of patients with ESCC by immunohistochemistry. Electrophoretic mobility shift assay (EMSA) and Western blot were used to detect the activation of NF-κB in esophageal cancer cell line KYSE150 cells. SN50, a specific NF-κB inhibitor, was applied to inhibit the activation of NF-κB. Clone formation test was used to detect the radiosensitivity of esophageal cancer cells. The median survival time of patients with activated and inactivated NF-κB in the pretreatment specimens were 16 and 19 months, respectively, with a non-significant difference between the two groups (P > 0.05). As to the patients with activated and inactivated NF-κB in posttreatment specimens, the median survival times were 13 and 35 months, respectively, with a significant difference (P < 0.01) between them. Western blot showed that the cytoplasmic expression of NF-κB was reduced with increasing radiation dose at 1.5 and 3 hours after radiation treatment. However, the expression of NF-κB in the cell nuclei was increased under the same condition, showing a trend of increased nucleus/cytoplasm ratio. The clone number in SN50 group was 96.66, 64.66, 76.66 and 10.00 under 0, 2, 4 and 12 Gy irradiation, which demonstrated a significant difference compared with the control groups (P < 0.001). Our results show that activation of NF-κB is induced by radiotherapy. Activation of NF-κB reduces the outcome of radiation treatment of esophageal cancer patients.
Houttuynia cordata blocks HSV infection through inhibition of NF-κB activation.

PubMed

Chen, Xiaoqing; Wang, Zhongxia; Yang, Ziying; Wang, Jingjing; Xu, Yunxia; Tan, Ren-Xiang; Li, Erguang

2011-11-01

Houttuynia cordata Thunb. is a medicinal plant widely used in folk medicine in several Asian countries. It has been reported that a water extract of H. cordata exhibits activity against herpes simplex virus (HSV) and the virus of severe acute respiratory syndrome (SARS), although the mechanisms are not fully understood yet. Previous studies have demonstrated absolute requirement of NF-κB activation for efficient replication of HSV-1 and HSV-2 and inhibition of NF-κB activation has been shown to suppress HSV infection. Here we show that a hot water extract of H. cordata (HCWE) inhibits HSV-2 infection through inhibition of NF-κB activation. The IC(50) was estimated at 50 μg/ml of lyophilized HCWE powder. At 150 and 450 μg/ml, HCWE blocked infectious HSV-2 production by more than 3 and 4 logs, respectively. The inhibitory activity was concomitant with an inhibition of NF-κB activation by HSV-2 infection. Although activation of NF-κB and Erk MAPK has been implicated for HSV replication and growth, HCWE showed no effect on HSV-2-induced Erk activation. Furthermore, we show that treatment with quercetin, quercitrin or isoquercitrin, major water extractable flavonoids from H. cordata, significantly blocked HSV-2 infection. These results together demonstrated that H. cordata blocks HSV-2 infection through inhibition of NF-κB activation. Copyright © 2011 Elsevier B.V. All rights reserved.
New Epigenetic Therapeutic Intervention for Metastatic Breast Cancer

DTIC Science & Technology

2016-04-01

transcription factor Twist are markedly over-expressed in TNBC but not luminal breast cancer cells. We also discovered that constitutively activated NF -kB in...transcription factors Twist and NF -kB in gene activation require lysine acetylation, which signs to activate the transcriptional machinery in chromatin...including Twist, NF -kB and STAT3. b. Define the molecular basis of the BET BrDs’ selective interactions with effector proteins through structure-guided
Activation of the classical pathway of complement by binding of bovine lactoferrin to unencapsulated Streptococcus agalactiae.

PubMed Central

Rainard, P

1993-01-01

The ability of lactoferrin (Lf) bound to Streptococcus agalactiae to interfere with the deposition of complement components on the bacterial surface was investigated by enzyme-linked immunosorbent assay (ELISA). By using a strain of S. agalactiae which activates the alternative pathway of complement in the absence of antibodies, it was found that pretreatment of bacteria with Lf shortened the lag phase preceding the deposition of C3 on bacteria. The kinetics of C3 deposition was comparable to that obtained by adding antibodies against S. agalactiae to agammaglobulinaemic precolostral calf serum (PCS) heated at 56 degrees for 3 min to inactivate the alternative pathway. Accelerated C3 deposition did not occur in the absence of Ca2+ ions. Deposition of C4 on bacteria occurred only when either antibodies or Lf were added to PCS. These results demonstrate that the interaction of lactoferrin with bacteria activated the classical pathway of complement in the absence of antibodies. The binding of purified C1q to bacteria was promoted in a dose-dependent manner by Lf, suggesting that recruitment of classical pathway of complement resulted from the interaction of C1q with Lf adsorbed to the bacterial surface. Phagocytosis of bacteria opsonized with heated PCS (at 56 degrees for 3 min) and Lf was comparable to that occurring in the presence of heated PCS and antibodies. In conclusion, Lf was able to substitute for antibodies in order to activate the classical pathway of complement and to opsonize unencapsulated S. agalactiae efficiently. PMID:8406591
Inhibiting NF-κB Activation by Small Molecules As a Therapeutic Strategy

PubMed Central

Gupta, Subash C; Sundaram, Chitra; Reuter, Simone; Aggarwal, Bharat B

2010-01-01

Because nuclear factor-κB (NF-κB) is a ubiquitously expressed proinflammatory transcription factor that regulates the expression of over 500 genes involved in cellular transformation, survival, proliferation, invasion, angiogenesis, metastasis, and inflammation, the NF-κB signaling pathway has become a potential target for pharmacological intervention. A wide variety of agents can activate NF-κB through canonical and noncanonical pathways. Canonical pathway involves various steps including the phosphorylation, ubiquitnation, and degradation of the inhibitor of NF-κB (IκBα), which leads to the nuclear translocation of the p50- p65 subunits of NF-κB followed by p65 phosphorylation, acetylation and methylation, DNA binding, and gene transcription. Thus, agents that can inhibit protein kinases, protein phosphatases, proteasomes, ubiquitnation, acetylation, methylation, and DNA binding steps have been identified as NF-κB inhibitors. Here, we review the small molecules that suppress NF-κB activation and thus may have therapeutic potential. PMID:20493977
NF90–NF45 is a selective RNA chaperone that rearranges viral and cellular riboswitches: biochemical analysis of a virus host factor activity

PubMed Central

Friedrich, Susann; Golbik, Ralph Peter

2017-01-01

Abstract The heterodimer NF90–NF45 is an RNA-binding protein complex that modulates the expression of various cellular mRNAs on the post-transcriptional level. Furthermore, it acts as a host factor that supports the replication of several RNA viruses. The molecular mechanisms underlying these activities have yet to be elucidated. Recently, we showed that the RNA-binding capabilities and binding specificity of NF90 considerably improves when it forms a complex with NF45. Here, we demonstrate that NF90 has a substrate-selective RNA chaperone activity (RCA) involving RNA annealing and strand displacement activities. The mechanism of the NF90-catalyzed RNA annealing was elucidated to comprise a combination of ‘matchmaking’ and compensation of repulsive charges, which finally results in the population of dsRNA products. Heterodimer formation with NF45 enhances ‘matchmaking’ of complementary ssRNAs and substantially increases the efficiency of NF90’s RCA. During investigations of the relevance of the NF90–NF45 RCA, the complex was shown to stimulate the first step in the RNA replication process of hepatitis C virus (HCV) in vitro and to stabilize a regulatory element within the mRNA of vascular endothelial growth factor (VEGF) by protein-guided changes of the RNAs’ structures. Thus, our study reveals how the intrinsic properties of an RNA-binding protein determine its biological activities. PMID:29040738
Neutrophils induce macrophage anti-inflammatory reprogramming by suppressing NF-κB activation.

PubMed

Marwick, John A; Mills, Ross; Kay, Oliver; Michail, Kyriakos; Stephen, Jillian; Rossi, Adriano G; Dransfield, Ian; Hirani, Nikhil

2018-06-04

Apoptotic cells modulate the function of macrophages to control and resolve inflammation. Here, we show that neutrophils induce a rapid and sustained suppression of NF-κB signalling in the macrophage through a unique regulatory relationship which is independent of apoptosis. The reduction of macrophage NF-κB activation occurs through a blockade in transforming growth factor β-activated kinase 1 (TAK1) and IKKβ activation. As a consequence, NF-κB (p65) phosphorylation is reduced, its translocation to the nucleus is inhibited and NF-κB-mediated inflammatory cytokine transcription is suppressed. Gene Set Enrichment Analysis reveals that this suppression of NF-κB activation is not restricted to post-translational modifications of the canonical NF-κB pathway, but is also imprinted at the transcriptional level. Thus neutrophils exert a sustained anti-inflammatory phenotypic reprogramming of the macrophage, which is reflected by the sustained reduction in the release of pro- but not anti- inflammatory cytokines from the macrophage. Together, our findings identify a novel apoptosis-independent mechanism by which neutrophils regulate the mediator profile and reprogramming of monocytes/macrophages, representing an important nodal point for inflammatory control.
Norepinephrine activates NF-κB transcription factor in cultured rat pineal gland.

PubMed

Villela, Darine; de Sá Lima, Larissa; Peres, Rafael; Peliciari-Garcia, Rodrigo Antonio; do Amaral, Fernanda Gaspar; Cipolla-Neto, José; Scavone, Cristóforo; Afeche, Solange Castro

2014-01-17

The circadian rhythm in mammalian pineal melatonin secretion is modulated by norepinephrine (NE) released at night. NE interaction with β1-adrenoceptors activates PKA that phosphorylates the transcription factor CREB, leading to the transcription and translation of the arylalkylamine-N-acetyltransferase (AANAT) enzyme. Several studies have reported the interplay between CREB and the nuclear factor-κB (NF-κB) and a circadian rhythm for this transcription factor was recently described in the rat pineal gland. In this work we studied a direct effect of NE on NF-κB activation and the role played by this factor on melatonin synthesis and Aanat transcription and activity. Cultured rat pineal glands were incubated in the presence of two different NF-κB inhibitors, pyrrolidine-dithiocarbamate or sodium salicylate, and stimulated with NE. Melatonin content was quantified by HPLC with electrochemical detection. AANAT activity was measured by a radiometric assay and the expression of Aanat mRNA was analyzed by real-time PCR. Gel shift assay was performed to study the NF-κB activation in cultured rat pineal glands stimulated by NE. Our results showed that the p50/p50 homodimer of NF-κB is activated by NE and that it has a role in melatonin synthesis, acting on Aanat transcription and activity. Here we present evidence that NF-κB is an important transcription factor that acts, directly or indirectly, on Aanat transcription and activity leading to a modulation of melatonin synthesis. NE plays a role in the translocation of NF-κB p50/p50 homodimer to the nucleus of pinealocytes, thus probably influencing the nocturnal pineal melatonin synthesis. Copyright © 2013 Elsevier Inc. All rights reserved.
Oral recombinant human or mouse lactoferrin reduces Mycobacterium tuberculosis TDM induced granulomatous lung pathology.

PubMed

Hwang, Shen-An; Kruzel, Marian L; Actor, Jeffrey K

2017-02-01

Trehalose 6'6-dimycolate (TDM) is the most abundant glycolipid on the cell wall of Mycobacterium tuberculosis (MTB). TDM is capable of inducing granulomatous pathology in mouse models that resembles those induced by MTB infection. Using the acute TDM model, this work investigates the effect of recombinant human and mouse lactoferrin to reduce granulomatous pathology. C57BL/6 mice were injected intravenously with TDM at a dose of 25 μg·mouse -1 . At day 4 and 6, recombinant human or mouse lactoferrin (1 mg·(100 μL) -1 ·mouse -1 ) were delivered by gavage. At day 7 after TDM injection, mice were evaluated for lung pathology, cytokine production, and leukocyte populations. Mice given human or mouse lactoferrin had reduced production of IL-12p40 in their lungs. Mouse lactoferrin increased IL-6 and KC (CXCL1) in lung tissue. Increased numbers of macrophages were observed in TDM-injected mice given human or mouse lactoferrin. Granulomatous pathology, composed of mainly migrated leukocytes, was visually reduced in mice that received human or mouse lactoferrin. Quantitation of granulomatous pathology demonstrated a significant decrease in mice given human or mouse lactoferrin compared with TDM control mice. This report is the first to directly compare the immune modulatory effects of both heterologous recombinant human and homologous mouse lactoferrin on the development of TDM-induced granulomas.
Thioredoxin-mediated denitrosylation regulates cytokine-induced nuclear factor κB (NF-κB) activation.

PubMed

Kelleher, Zachary T; Sha, Yonggang; Foster, Matthew W; Foster, W Michael; Forrester, Michael T; Marshall, Harvey E

2014-01-31

S-nitrosylation of nuclear factor κB (NF-κB) on the p65 subunit of the p50/p65 heterodimer inhibits NF-κB DNA binding activity. We have recently shown that p65 is constitutively S-nitrosylated in the lung and that LPS-induced injury elicits a decrease in SNO-p65 levels concomitant with NF-κB activation in the respiratory epithelium and initiation of the inflammatory response. Here, we demonstrate that TNFα-mediated activation of NF-κB in the respiratory epithelium similarly induces p65 denitrosylation. This process is mediated by the denitrosylase thioredoxin (Trx), which becomes activated upon cytokine-induced degradation of thioredoxin-interacting protein (Txnip). Similarly, inhibition of Trx activity in the lung attenuates LPS-induced SNO-p65 denitrosylation, NF-κB activation, and airway inflammation, supporting a pathophysiological role for this mechanism in lung injury. These data thus link stimulus-coupled activation of NF-κB to a specific, protein-targeted denitrosylation mechanism and further highlight the importance of S-nitrosylation in the regulation of the immune response.
Heteroaggregation of lipid droplets coated with sodium caseinate and lactoferrin.

PubMed

de Figueiredo Furtado, Guilherme; Michelon, Mariano; de Oliveira, Davi Rocha Bernardes; da Cunha, Rosiane Lopes

2016-11-01

Formation and characterization of droplet heteroaggregates were investigated by mixing two emulsions previously stabilized by proteins oppositely charged. Emulsions were composed of 5vol.% of sunflower oil and 95vol.% of sodium caseinate or lactoferrin aqueous dispersions. They were produced using ultrasound with fixed power (300W) and sonication time (6min). Different volume ratios (0-100%) of sodium caseinate-stabilized emulsion (droplet diameter around 1.75μm) to lactoferrin-stabilized emulsion (droplet diameter around 1.55μm) were mixed under conditions that both proteins showed opposite charges (pH7). Influence of ionic strength (0-400mM NaCl) on the heteroaggregates stability was also evaluated. Creaming stability, zeta potential, microstructure, mean particle diameter and rheological properties of the heteroaggregates were measured. These properties depended on the volume ratio (0-100%) of sodium caseinate to lactoferrin-stabilized emulsion (C:L) and the ionic strength. In the absence of salt, different zeta potential values were obtained, rheological properties (viscosity and elastic moduli) were improved and the largest heteroaggregates were formed at higher content of lactoferrin-stabilized emulsion (60-80%). The system containing 40 and 60vol.% of sodium caseinate and lactoferrin stabilized emulsion, respectively, presented good stability against phase separation besides showing enhanced rheological and size properties due to extensive droplets aggregation. Phase separation was observed only in the absence of sodium caseinate, demonstrating the higher susceptibility of lactoferrin to NaCl. The heteroaggregates produced may be useful functional agents for texture modification and controlled release since different rheological properties and sizes can be achieved depending on protein concentrations. Copyright © 2016 Elsevier Ltd. All rights reserved.
Antimicrobial Activity of Lactoferrin-Related Peptides and Applications in Human and Veterinary Medicine.

PubMed

Bruni, Natascia; Capucchio, Maria Teresa; Biasibetti, Elena; Pessione, Enrica; Cirrincione, Simona; Giraudo, Leonardo; Corona, Antonio; Dosio, Franco

2016-06-11

Antimicrobial peptides (AMPs) represent a vast array of molecules produced by virtually all living organisms as natural barriers against infection. Among AMP sources, an interesting class regards the food-derived bioactive agents. The whey protein lactoferrin (Lf) is an iron-binding glycoprotein that plays a significant role in the innate immune system, and is considered as an important host defense molecule. In search for novel antimicrobial agents, Lf offers a new source with potential pharmaceutical applications. The Lf-derived peptides Lf(1-11), lactoferricin (Lfcin) and lactoferrampin exhibit interesting and more potent antimicrobial actions than intact protein. Particularly, Lfcin has demonstrated strong antibacterial, anti-fungal and antiparasitic activity with promising applications both in human and veterinary diseases (from ocular infections to osteo-articular, gastrointestinal and dermatological diseases).
NF-κB DNA-binding activity in embryos responding to a teratogen, cyclophosphamide

PubMed Central

Torchinsky, Arkady; Lishanski, Lucy; Wolstein, Orit; Shepshelovich, Jeanne; Orenstein, Hasida; Savion, Shoshana; Zaslavsky, Zeev; Carp, Howard; Brill, Alexander; Dikstein, Rivka; Toder, Vladimir; Fein, Amos

2002-01-01

Background The Rel/NF-κB transcription factors have been shown to regulate apoptosis in different cell types, acting as inducers or blockers in a stimuli- and cell type-dependent fashion. One of the Rel/NF-κB subunits, RelA, has been shown to be crucial for normal embryonic development, in which it functions in the embryonic liver as a protector against TNFα-induced physiological apoptosis. This study assesses whether NF-κB may be involved in the embryo's response to teratogens. Fot this, we evaluated how NF-KappaB DNA binding activity in embryonic organs demonstraiting differential sensitivity to a reference teratogen, cyclophosphamide, correlates with dysmorphic events induced by the teratogen at the cellular level (excessive apoptosis) and at the organ level (structural anomalies). Results The embryonic brain and liver were used as target organs. We observed that the Cyclophosphamide-induced excessive apoptosis in the brain, followed by the formation of severe craniofacial structural anomalies, was accompanied by suppression of NF-κB DNA-binding activity as well as by a significant and lasting increase in the activity of caspases 3 and 8. However, in the liver, in which cyclophosphamide induced transient apoptosis was not followed by dysmorphogenesis, no suppression of NF-κB DNA-binding activity was registered and the level of active caspases 3 and 8 was significantly lower than in the brain. It has also been observed that both the brain and liver became much more sensitive to the CP-induced teratogenic insult if the embryos were exposed to a combined treatment with the teratogen and sodium salicylate that suppressed NF-κB DNA-binding activity in these organs. Conclusion The results of this study demonstrate that suppression of NF-κB DNA-binding activity in embryos responding to the teratogenic insult may be associated with their decreased resistance to this insult. They also suggest that teratogens may suppress NF-κB DNA-binding activity in the

NF-κB-Induced IL-6 Ensures STAT3 Activation and Tumor Aggressiveness in Glioblastoma

PubMed Central

McFarland, Braden C.; Hong, Suk W.; Rajbhandari, Rajani; Twitty, George B.; Gray, G. Kenneth; Yu, Hao; Benveniste, Etty N.; Nozell, Susan E.

2013-01-01

Glioblastoma (GBM) is the most aggressive, neurologically destructive and deadly tumor of the central nervous system (CNS). In GBM, the transcription factors NF-κB and STAT3 are aberrantly activated and associated with tumor cell proliferation, survival, invasion and chemoresistance. In addition, common activators of NF-κB and STAT3, including TNF-α and IL-6, respectively, are abundantly expressed in GBM tumors. Herein, we sought to elucidate the signaling crosstalk that occurs between the NF-κB and STAT3 pathways in GBM tumors. Using cultured GBM cell lines as well as primary human GBM xenografts, we elucidated the signaling crosstalk between the NF-κB and STAT3 pathways utilizing approaches that either a) reduce NF-κB p65 expression, b) inhibit NF-κB activation, c) interfere with IL-6 signaling, or d) inhibit STAT3 activation. Using the clinically relevant human GBM xenograft model, we assessed the efficacy of inhibiting NF-κB and/or STAT3 alone or in combination in mice bearing intracranial xenograft tumors in vivo. We demonstrate that TNF-α-induced activation of NF-κB is sufficient to induce IL-6 expression, activate STAT3, and elevate STAT3 target gene expression in GBM cell lines and human GBM xenografts in vitro. Moreover, the combined inhibition of NF-κB and STAT3 signaling significantly increases survival of mice bearing intracranial tumors. We propose that in GBM, the activation of NF-κB ensures subsequent STAT3 activation through the expression of IL-6. These data verify that pharmacological interventions to effectively inhibit the activity of both NF-κB and STAT3 transcription factors must be used in order to reduce glioma size and aggressiveness. PMID:24244348
NF-κB-induced IL-6 ensures STAT3 activation and tumor aggressiveness in glioblastoma.

PubMed

McFarland, Braden C; Hong, Suk W; Rajbhandari, Rajani; Twitty, George B; Gray, G Kenneth; Yu, Hao; Benveniste, Etty N; Nozell, Susan E

2013-01-01

Glioblastoma (GBM) is the most aggressive, neurologically destructive and deadly tumor of the central nervous system (CNS). In GBM, the transcription factors NF-κB and STAT3 are aberrantly activated and associated with tumor cell proliferation, survival, invasion and chemoresistance. In addition, common activators of NF-κB and STAT3, including TNF-α and IL-6, respectively, are abundantly expressed in GBM tumors. Herein, we sought to elucidate the signaling crosstalk that occurs between the NF-κB and STAT3 pathways in GBM tumors. Using cultured GBM cell lines as well as primary human GBM xenografts, we elucidated the signaling crosstalk between the NF-κB and STAT3 pathways utilizing approaches that either a) reduce NF-κB p65 expression, b) inhibit NF-κB activation, c) interfere with IL-6 signaling, or d) inhibit STAT3 activation. Using the clinically relevant human GBM xenograft model, we assessed the efficacy of inhibiting NF-κB and/or STAT3 alone or in combination in mice bearing intracranial xenograft tumors in vivo. We demonstrate that TNF-α-induced activation of NF-κB is sufficient to induce IL-6 expression, activate STAT3, and elevate STAT3 target gene expression in GBM cell lines and human GBM xenografts in vitro. Moreover, the combined inhibition of NF-κB and STAT3 signaling significantly increases survival of mice bearing intracranial tumors. We propose that in GBM, the activation of NF-κB ensures subsequent STAT3 activation through the expression of IL-6. These data verify that pharmacological interventions to effectively inhibit the activity of both NF-κB and STAT3 transcription factors must be used in order to reduce glioma size and aggressiveness.
Potent Anti-Inflammatory Activity of Ursolic Acid, a Triterpenoid Antioxidant, Is Mediated through Suppression of NF-κB, AP-1 and NF-AT

PubMed Central

Checker, Rahul; Sandur, Santosh K.; Sharma, Deepak; Patwardhan, Raghavendra S.; Jayakumar, S.; Kohli, Vineet; Sethi, Gautam; Aggarwal, Bharat B.; Sainis, Krishna B.

2012-01-01

Background Ursolic acid (UA), a pentacyclic triterpenoid carboxylic acid, is the major component of many plants including apples, basil, cranberries, peppermint, rosemary, oregano and prunes and has been reported to possess antioxidant and anti-tumor properties. These properties of UA have been attributed to its ability to suppress NF-κB (nuclear factor kappa B) activation. Since NF-κB, in co-ordination with NF-AT (nuclear factor of activated T cells) and AP-1(activator protein-1), is known to regulate inflammatory genes, we hypothesized that UA might exhibit potent anti-inflammatory effects. Methodology/Principal Findings The anti-inflammatory effects of UA were assessed in activated T cells, B cells and macrophages. Effects of UA on ERK, JNK, NF-κB, AP-1 and NF-AT were studied to elucidate its mechanism of action. In vivo efficacy of UA was studied using mouse model of graft-versus-host disease. UA inhibited activation, proliferation and cytokine secretion in T cells, B cells and macrophages. UA inhibited mitogen-induced up-regulation of activation markers and co-stimulatory molecules in T and B cells. It inhibited mitogen-induced phosphorylation of ERK and JNK and suppressed the activation of immunoregulatory transcription factors NF-κB, NF-AT and AP-1 in lymphocytes. Treatment of cells with UA prior to allogenic transplantation significantly delayed induction of acute graft-versus-host disease in mice and also significantly reduced the serum levels of pro-inflammatory cytokines IL-6 and IFN-γ. UA treatment inhibited T cell activation even when added post-mitogenic stimulation demonstrating its therapeutic utility as an anti-inflammatory agent. Conclusions/Significance The present study describes the detailed mechanism of anti-inflammatory activity of UA. Further, UA may find application in the treatment of inflammatory disorders. PMID:22363615
Eupatilin inhibits T-cell activation by modulation of intracellular calcium flux and NF-kappaB and NF-AT activity.

PubMed

Kim, Young-Dae; Choi, Suck-Chei; Oh, Tae-Young; Chun, Jang-Soo; Jun, Chang-Duk

2009-09-01

Eupatilin, one of the pharmacologically active ingredients of Artemisia princeps, exhibits a potent anti-ulcer activity, but its effects on T-cell immunity have not been investigated. Here, we show that eupatilin has a profound inhibitory effect on IL-2 production in Jurkat T cells as well as in human peripheral blood leukocytes. Eupatilin neither influenced clustering of CD3 and LFA-1 to the immunological synapse nor inhibited conjugate formation between T cells and B cells in the presence or absence of superantigen (SEE). Eupatilin also failed to inhibit T-cell receptor (TCR) internalization, thereby, suggesting that eupatilin does not interfere with TCR-mediated signals on the membrane proximal region. In unstimulated T cells, eupatilin significantly induced apoptotic cell death, as evidenced by an increased population of annexin V(+)/PI(+) cells and cleavage of caspase-3 and PARP. To our surprise, however, once cells were activated, eupatilin had little effect on apoptosis, and instead slightly protected cells from activation-induced cell death, suggesting that apoptosis also is not a mechanism for eupatilin-induced T-cell suppression. On the contrary, eupatilin dramatically inhibited I-kappaBalpha degradation and NF-AT dephosphorylation and, consequently, inhibited NF-kappaB and NF-AT promoter activities in PMA/A23187-stimulated T cells. Interestingly, intracellular calcium flux was significantly perturbed in cells pre-treated with eupatilin, suggesting that calcium-dependent cascades might be targets for eupatilin action. Collectively, our results provide evidence for dual regulatory functions of eupatilin: (1) a pro-apoptotic effect on resting T cells and (2) an immunosuppressive effect on activated T cells, presumably through modulation of Ca(2+) flux. (c) 2009 Wiley-Liss, Inc.
Polymorphisms in the promoter region of the bovine lactoferrin gene influence milk somatic cell score and milk production traits in Chinese Holstein cows.

PubMed

Mao, Yongjiang; Zhu, Xiaorui; Xing, Shiyu; Zhang, Meirong; Zhang, Huimin; Wang, Xiaolong; Karrow, Niel; Yang, Liguo; Yang, Zhangping

2015-12-01

Lactoferrin is an iron-binding protein found in cow's milk that plays an important role in preventing mastitis caused by intramammary infection. In this study, 20 Chinese Holstein cows were selected randomly for PCR amplification and sequencing of the bovine lactoferrin gene promoter region and used for SNP discovery in the region between nucleotide positions -461 to -132. Three SNPs (-270T>C, -190G>A and -156A>G) were identified in bovine lactoferrin, then Chinese Holstein cows (n=866) were genotyped using Sequenom MassARRAY (Sequenom Inc., San Diego, CA) based on the previous SNP information in this study, and the associations between SNPs or haplotype and milk somatic cell score (SCS) and production traits were analyzed by the least squares method in the GLM procedure of SAS. SNPs -270T>C and -156A>G showed close linkage disequilibrium (r(2)=0.76). The SNP -190G>A showed a significant association with SCS, and individuals with genotype GG had higher SCS than genotypes AG and AA. Associations were found between the SNPs -270T>C and -190G>A with SCS and the milk composition. The software MatInspector revealed that these SNPs were located within several potential transcription factor binding sites, including NF-κB p50, KLF7 and SP1, and may alter gene expression, but further investigation will be required to elucidate the biological and practical relevance of these SNPs. Copyright © 2015 Elsevier Ltd. All rights reserved.
Simultaneous imaging of temporal changes of NF-κB activity and viable tumor cells in Huh7/NF-κB-tk-luc2/rfp tumor-bearing mice.

PubMed

Wang, Wei-Hsun; Chiang, I-Tsang; Liu, Yu-Chang; Hsu, Fei-Ting; Chen, Hong-Wen; Chen, Chuan-Lin; Lee, Yi-Jang; Lin, Wuu-Jyh; Hwang, Jeng-Jong

2013-01-01

Few studies have reported that the effect of sorafenib on advanced human hepatocellular carcinoma (HCC) is taking place via the inhibition of NF-κB signal transduction. Here we constructed a human HCC Huh7 stable clone with NF-κB-responsive element to drive dual reporter genes, herpes simplex virus thymidine kinase (tk) and firefly luciferase (luc2), and co-transfected with a third red fluorescent protein (rfp) gene, renamed as Huh7/NF-κB-tk-luc2/rfp cells, and combined with bioluminescent imaging (BLI) and red fluorescent protein imaging (RFPI) to monitor the effect of sorafenib on NF-κB activation and tumor inhibition. The results show that sorafenib could suppress the NF-κB-DNA binding activity, and the expression of downstream effector proteins. Notably, the relative photon fluxes obtained from RFPI and BLI, which represent the viable tumor cells and cells with NF-κB activation, decreased after sorafenib treatment by 50 to 65%, and 87.5 to >90%, respectively, suggesting that NF-κB activation is suppressed in viable HCC cells by sorafenib. Simultaneous molecular imaging of the temporal change of NF-κB activity and of viable cells in the same Huh7/NF-κB-tk-luc2/rfp tumors of the animal may reflect the real status of NF-κB activity and the viable tumor cells at the time of imaging.
Porcine reproductive and respiratory syndrome virus nonstructural protein 2 contributes to NF-κB activation.

PubMed

Fang, Ying; Fang, Liurong; Wang, Yang; Lei, Yingying; Luo, Rui; Wang, Dang; Chen, Huanchun; Xiao, Shaobo

2012-04-30

Nuclear factor-kappaB (NF-κB) is an inducible transcription factor that plays a key role in inflammation and immune responses, as well as in the regulation of cell proliferation and survival. Previous studies by our group and others have demonstrated that porcine reproductive and respiratory syndrome virus (PRRSV) infection could activate NF-κB in MARC-145 cells and alveolar macrophages. The nucleocapsid (N) protein was identified as an NF-κB activator among the structural proteins encoded by PRRSV; however, it remains unclear whether the nonstructural proteins (Nsps) contribute to NF-κB activation. In this study, we identified which Nsps can activate NF-κB and investigated the potential mechanism(s) by which they act. By screening the individual Nsps of PRRSV strain WUH3, Nsp2 exhibited great potential to activate NF-κB in MARC-145 and HeLa cells. Overexpression of Nsp2 induced IκBα degradation and nuclear translocation of NF-κB. Furthermore, Nsp2 also induced NF-κB-dependent inflammatory factors, including interleukin (IL)-6, IL-8, COX-2, and RANTES. Compared with the Nsp2 of the classical PRRSV strain, the Nsp2 of highly pathogenic PRRSV (HP-PRRSV) strains that possess a 30 amino acid (aa) deletion in Nsp2 displayed greater NF-κB activation. However, the 30-aa deletion was demonstrated to not be associated with NF-κB activation. Further functional domain analyses revealed that the hypervariable region (HV) of Nsp2 was essential for NF-κB activation. Taken together, these data indicate that PRRSV Nsp2 is a multifunctional protein participating in the modulation of host inflammatory response, which suggests an important role of Nsp2 in pathogenesis and disease outcomes.
Myocardial NF-κB activation is essential for zebrafish heart regeneration

PubMed Central

Karra, Ravi; Knecht, Anne K.; Kikuchi, Kazu; Poss, Kenneth D.

2015-01-01

Heart regeneration offers a novel therapeutic strategy for heart failure. Unlike mammals, lower vertebrates such as zebrafish mount a strong regenerative response following cardiac injury. Heart regeneration in zebrafish occurs by cardiomyocyte proliferation and reactivation of a cardiac developmental program, as evidenced by induction of gata4 regulatory sequences in regenerating cardiomyocytes. Although many of the cellular determinants of heart regeneration have been elucidated, how injury triggers a regenerative program through dedifferentiation and epicardial activation is a critical outstanding question. Here, we show that NF-κB signaling is induced in cardiomyocytes following injury. Myocardial inhibition of NF-κB activity blocks heart regeneration with pleiotropic effects, decreasing both cardiomyocyte proliferation and epicardial responses. Activation of gata4 regulatory sequences is also prevented by NF-κB signaling antagonism, suggesting an underlying defect in cardiomyocyte dedifferentiation. Our results implicate NF-κB signaling as a key node between cardiac injury and tissue regeneration. PMID:26472034
Myocardial NF-κB activation is essential for zebrafish heart regeneration.

PubMed

Karra, Ravi; Knecht, Anne K; Kikuchi, Kazu; Poss, Kenneth D

2015-10-27

Heart regeneration offers a novel therapeutic strategy for heart failure. Unlike mammals, lower vertebrates such as zebrafish mount a strong regenerative response following cardiac injury. Heart regeneration in zebrafish occurs by cardiomyocyte proliferation and reactivation of a cardiac developmental program, as evidenced by induction of gata4 regulatory sequences in regenerating cardiomyocytes. Although many of the cellular determinants of heart regeneration have been elucidated, how injury triggers a regenerative program through dedifferentiation and epicardial activation is a critical outstanding question. Here, we show that NF-κB signaling is induced in cardiomyocytes following injury. Myocardial inhibition of NF-κB activity blocks heart regeneration with pleiotropic effects, decreasing both cardiomyocyte proliferation and epicardial responses. Activation of gata4 regulatory sequences is also prevented by NF-κB signaling antagonism, suggesting an underlying defect in cardiomyocyte dedifferentiation. Our results implicate NF-κB signaling as a key node between cardiac injury and tissue regeneration.
Rig-I regulates NF-κB activity through binding to Nf-κb1 3′-UTR mRNA

PubMed Central

Zhang, Hong-Xin; Liu, Zi-Xing; Sun, Yue-Ping; Lu, Shun-Yuan; Liu, Xue-Song; Huang, Qiu-Hua; Xie, Yin-Yin; Dang, Su-Ying; Zheng, Guang-Yong; Li, Yi-Xue; Kuang, Ying; Fei, Jian; Chen, Zhu; Wang, Zhu-Gang

2013-01-01

Retinoic acid inducible gene I (RIG-I) senses viral RNAs and triggers innate antiviral responses through induction of type I IFNs and inflammatory cytokines. However, whether RIG-I interacts with host cellular RNA remains undetermined. Here we report that Rig-I interacts with multiple cellular mRNAs, especially Nf-κb1. Rig-I is required for NF-κB activity via regulating Nf-κb1 expression at posttranscriptional levels. It interacts with the multiple binding sites within 3′-UTR of Nf-κb1 mRNA. Further analyses reveal that three distinct tandem motifs enriched in the 3′-UTR fragments can be recognized by Rig-I. The 3′-UTR binding with Rig-I plays a critical role in normal translation of Nf-κb1 by recruiting the ribosomal proteins [ribosomal protein L13 (Rpl13) and Rpl8] and rRNAs (18S and 28S). Down-regulation of Rig-I or Rpl13 significantly reduces Nf-κb1 and 3′-UTR–mediated luciferase expression levels. These findings indicate that Rig-I functions as a positive regulator for NF-κB signaling and is involved in multiple biological processes in addition to host antivirus immunity. PMID:23553835
INTERFERON α ACTIVATES NF-κ B IN JAK1-DEFICIENT CELLS THROUGH A TYK2-DEPENDENT PATHWAY

PubMed Central

Yang, Chuan He; Murti, Aruna; Valentine, William J.; Du, Ziyun; Pfeffer, Lawrence M.

2005-01-01

In addition to activating members of the STAT transcription factor family, IFN α/β activates the NF-κ B transcription factor. To determine the role of the JAK-STAT pathway in NF-κ B activation by IFN, we examined NF-κ B activation in JAK1-deficient mutant human fibrosarcoma cells. In wild-type fibrosarcoma cells (2fTGH) IFN activates STAT1, STAT2 and STAT3, as well as NF-κB complexes comprised of p50 and p65. In contrast, in JAK1-deficient cells IFN induces NF-κB activation and NF-κB dependent gene transcription, but does not activate these STAT proteins and has no effect on STAT-dependent gene transcription. Expression of a catalytically-inactive TYK2 tyrosine kinase in JAK1-deficient cells, as well as in the highly IFN-sensitive Daudi lymphoblastoid cell line, abrogates NF-κB activation by IFN. Moreover, IFN does not promote NF-κB activation in TYK2-deficient mutant fibrosarcoma cells. Our results demonstrate a dichotomy between the classical JAK-STAT pathway and the NF-κB signaling pathway. In the IFN signaling pathway leading to STAT activation both JAK1 and TYK2 are essential, while NF-κB activation requires only TYK2. PMID:15883164
Activation of NF-kappa B Signaling Promotes Growth of Prostate Cancer Cells in Bone

PubMed Central

Jin, Renjie; Sterling, Julie A.; Edwards, James R.; DeGraff, David J.; Lee, Changki; Park, Serk In; Matusik, Robert J.

2013-01-01

Patients with advanced prostate cancer almost invariably develop osseous metastasis. Although many studies indicate that the activation of NF-κB signaling appears to be correlated with advanced cancer and promotes tumor metastasis by influencing tumor cell migration and angiogenesis, the influence of altered NF-κB signaling in prostate cancer cells within boney metastatic lesions is not clearly understood. While C4-2B and PC3 prostate cancer cells grow well in the bone, LNCaP cells are difficult to grow in murine bone following intraskeletal injection. Our studies show that when compared to LNCaP, NF-κB activity is significantly higher in C4-2B and PC3, and that the activation of NF-κB signaling in prostate cancer cells resulted in the increased expression of the osteoclast inducing genes PTHrP and RANKL. Further, conditioned medium derived from NF-κB activated LNCaP cells induce osteoclast differentiation. In addition, inactivation of NF-κB signaling in prostate cancer cells inhibited tumor formation in the bone, both in the osteolytic PC3 and osteoblastic/osteoclastic mixed C4-2B cells; while the activation of NF-κB signaling in LNCaP cells promoted tumor establishment and proliferation in the bone. The activation of NF-κB in LNCaP cells resulted in the formation of an osteoblastic/osteoclastic mixed tumor with increased osteoclasts surrounding the new formed bone, similar to metastases commonly seen in patients with prostate cancer. These results indicate that osteoclastic reaction is required even in the osteoblastic cancer cells and the activation of NF-κB signaling in prostate cancer cells increases osteoclastogenesis by up-regulating osteoclastogenic genes, thereby contributing to bone metastatic formation. PMID:23577181
Lactoferrin affects the adherence and invasion of Streptococcus dysgalactiae ssp. dysgalactiae in mammary epithelial cells.

PubMed

O'Halloran, Fiona; Beecher, Christine; Chaurin, Valerie; Sweeney, Torres; Giblin, Linda

2016-06-01

Streptococcus dysgalactiae ssp. dysgalactiae is an important causative agent of bovine mastitis worldwide. Lactoferrin is an innate immune protein that is associated with many functions including immunomodulatory, antiproliferative, and antimicrobial properties. This study aimed to investigate the interactions between lactoferrin and a clinical bovine mastitis isolate, Strep. dysgalactiae ssp. dysgalactiae DPC5345. Initially a deliberate in vivo bovine intramammary challenge was performed with Strep. dysgalactiae DPC5345. Results demonstrated a significant difference in lactoferrin mRNA levels in milk cells between the control and infused quarters 7h postinfusion. Milk lactoferrin levels in the Strep. dysgalactiae DPC5345 infused quarters were significantly increased compared with control quarters at 48h postinfusion. In vitro studies demonstrated that lactoferrin had a bacteriostatic effect on the growth of Strep. dysgalactiae DPC5345 and significantly decreased the ability of the bacteria to internalize into HC-11 mammary epithelial cells. Confocal microscopy images of HC-11 cells exposed to Strep. dysgalactiae and lactoferrin further supported this effect by demonstrating reduced invasion of bacteria to HC-11 cells. The combined data suggest that a bovine immune response to Strep. dysgalactiae infection includes a significant increase in lactoferrin expression in vivo, and based on in vitro data, lactoferrin limits mammary cell invasion of this pathogen by binding to the bacteria and preventing its adherence. Copyright © 2016 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
Significant antibacterial activity and synergistic effects of camel lactoferrin with antibiotics against methicillin-resistant Staphylococcus aureus (MRSA).

PubMed

Redwan, Elrashdy M; El-Baky, Nawal Abd; Al-Hejin, Ahmed M; Baeshen, Mohammed N; Almehdar, Hussein A; Elsaway, Abdulrahman; Gomaa, Abu-Bakr M; Al-Masaudi, Saad Berki; Al-Fassi, Fahad A; AbuZeid, Isam Eldin; Uversky, Vladimir N

2016-01-01

Methicillin-resistant Staphylococcus aureus (MRSA) causes major healthcare problems in many countries, as it is present as several hospital- and community-associated strains. Hospital-associated MRSA is one of the most prevalent nosocomial pathogens throughout the world and infections caused by community-acquired MRSA are rising. This emphasizes the need for new and efficient anti-MRSA agents. We evaluated the antibacterial effects of camel lactoferrin (cLf) and human lactoferrin (hLf) alone and in combination with several antibiotics against MRSA. Antimicrobials were tested against MRSA and an S. aureus control strain by the agar disc diffusion method. The minimum inhibitory concentration (MIC) was determined for antimicrobials by the broth microdilution method. Synergy between cLf or hLf and antibiotics was examined by checkerboard and time-kill assays. The agar disc diffusion assay showed that MRSA growth was inhibited by cLf at 0.25-3 mg/ml and hLf at 1-3 mg/ml. cLf demonstrated 3 times higher inhibitory activity against MRSA than hLf in terms of MIC values (250 vs. 750 μg/ml, respectively). Biotinylated cLf was recognized by two membrane proteins of MRSA, 66-67 KDa. Combinations of cLf or hLf and oxacillin or vancomycin at sub-MIC levels enhanced in vitro antibacterial activity against MRSA compared with each agent alone. Copyright © 2016 Institut Pasteur. Published by Elsevier Masson SAS. All rights reserved.
In vitro and in vivo anticandidal activities of alginate-enclosed chitosan–calcium phosphate-loaded Fe-bovine lactoferrin nanocapsules

PubMed Central

Leng, Khoo Miew; Vijayarathna, Soundararajan; Jothy, Subramanion L; Sasidharan, Sreenivasan; Kanwar, Jagat R

2018-01-01

Aim: To study the in vitro and in vivo anticandidal activity of nanocapsulated bovine lactoferrin. Materials & methods: In vitro and in vivo antimicrobial activities were conducted to study the anticandidal activities of nanocapsules (NCs). Results: The NCs showed good anticandidal activities. The disruption of cell wall and cell membrane was noted via microscopy studies. The NCs changed the normal growth profile of Candida albicans. NCs reduced the colony forming unit in kidney and blood samples. Histopathological examination showed better cell structure and coordination compared with untreated mice kidney. NCs also enhanced the natural killing properties of C. albicans by epithelial cells. Conclusion: NCs have effective anticandidal properties and have the potential as a therapeutic agent against candidiasis. PMID:29379633
A Non-SUMOylated Tax Protein Is Still Functional for NF-κB Pathway Activation

PubMed Central

Pène, Sabrina; Waast, Laetitia; Bonnet, Amandine; Bénit, Laurence

2014-01-01

ABSTRACT Whether NF-κB promoter transactivation by the human T-cell leukemia virus type 1 (HTLV-1) Tax protein requires Tax SUMOylation is still a matter of debate. In this study, we revisited the role of Tax SUMOylation using a strategy based on the targeting of Ubc9, the unique E2 SUMO-conjugating enzyme. We show that either a catalytically inactive form of Ubc9 (Ubc9-C93S) or Ubc9 small interfering RNA (siRNA) dramatically reduces Tax conjugation to endogenous SUMO-1 or SUMO-2/3, demonstrating that as expected, Tax SUMOylation is under the control of the catalytic activity of Ubc9. We further report that a non-SUMOylated Tax protein produced in 293T cells is still able to activate either a transfected or an integrated NF-κB reporter promoter and to induce expression of an NF-κB-regulated endogenous gene. Importantly, blocking Ubc9 activity in T cells also results in the production of a non-SUMOylated Tax that is still fully functional for the activation of a NF-κB promoter. These results provide the definitive evidence that Tax SUMOylation is not required for NF-κB-driven gene induction. IMPORTANCE Human T-cell leukemia virus type 1 is able to transform CD4+ T lymphocytes. The viral oncoprotein Tax plays a key role in this process by promoting cell proliferation and survival, mainly through permanent activation of the NF-κB pathway. Elucidating the molecular mechanisms involved in NF-κB pathway activation by Tax is therefore a key issue to understand HTLV-1-mediated transformation. Tax SUMOylation was initially proposed to be critical for Tax-induced NF-κB promoter activation, which was challenged by our later observation that a low-level-SUMOylated Tax mutant was still functional for activation of NF-κB promoters. To clarify the role of Tax SUMOylation, we set up a new approach based on the inhibition of the SUMOylation machinery in Tax-expressing cells. We show that blocking the SUMO-conjugating enzyme Ubc9 abolishes Tax SUMOylation and that a non
A non-SUMOylated tax protein is still functional for NF-κB pathway activation.

PubMed

Pène, Sabrina; Waast, Laetitia; Bonnet, Amandine; Bénit, Laurence; Pique, Claudine

2014-09-01

Whether NF-κB promoter transactivation by the human T-cell leukemia virus type 1 (HTLV-1) Tax protein requires Tax SUMOylation is still a matter of debate. In this study, we revisited the role of Tax SUMOylation using a strategy based on the targeting of Ubc9, the unique E2 SUMO-conjugating enzyme. We show that either a catalytically inactive form of Ubc9 (Ubc9-C93S) or Ubc9 small interfering RNA (siRNA) dramatically reduces Tax conjugation to endogenous SUMO-1 or SUMO-2/3, demonstrating that as expected, Tax SUMOylation is under the control of the catalytic activity of Ubc9. We further report that a non-SUMOylated Tax protein produced in 293T cells is still able to activate either a transfected or an integrated NF-κB reporter promoter and to induce expression of an NF-κB-regulated endogenous gene. Importantly, blocking Ubc9 activity in T cells also results in the production of a non-SUMOylated Tax that is still fully functional for the activation of a NF-κB promoter. These results provide the definitive evidence that Tax SUMOylation is not required for NF-κB-driven gene induction. Human T-cell leukemia virus type 1 is able to transform CD4(+) T lymphocytes. The viral oncoprotein Tax plays a key role in this process by promoting cell proliferation and survival, mainly through permanent activation of the NF-κB pathway. Elucidating the molecular mechanisms involved in NF-κB pathway activation by Tax is therefore a key issue to understand HTLV-1-mediated transformation. Tax SUMOylation was initially proposed to be critical for Tax-induced NF-κB promoter activation, which was challenged by our later observation that a low-level-SUMOylated Tax mutant was still functional for activation of NF-κB promoters. To clarify the role of Tax SUMOylation, we set up a new approach based on the inhibition of the SUMOylation machinery in Tax-expressing cells. We show that blocking the SUMO-conjugating enzyme Ubc9 abolishes Tax SUMOylation and that a non-SUMOylated Tax still
BFV activates the NF-kappaB pathway through its transactivator (BTas) to enhance viral transcription

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang Jian; Tan Juan; Zhang Xihui

2010-05-10

Multiple families of viruses have evolved sophisticated strategies to regulate nuclear factor-kappaB (NF-kappaB) signaling, which plays a pivotal role in diverse cellular events, including virus-host interactions. In this study, we report that bovine foamy virus (BFV) is able to activate the NF-kappaB pathway through the action of its transactivator, BTas. Both cellular IKKbeta and IkappaBalpha also participate in this activation. In addition, we demonstrate that BTas induces the processing of p100, which implies that BTas can activate NF-kappaB through a noncanonical pathway as well. Co-immunoprecipitation analysis shows that BTas interacts with IKK catalytic subunits (IKKalpha and IKKbeta), which may bemore » responsible for regulation of IKK kinase activity and persistent NF-kappaB activation. Furthermore, our results indicate that the level of BTas-mediated LTR transcription correlates with the activity of cellular NF-kappaB. Together, this study suggests that BFV activates the NF-kappaB pathway through BTas to enhance viral transcription.« less
BFV activates the NF-kappaB pathway through its transactivator (BTas) to enhance viral transcription.

PubMed

Wang, Jian; Tan, Juan; Zhang, Xihui; Guo, Hongyan; Zhang, Qicheng; Guo, Tingting; Geng, Yunqi; Qiao, Wentao

2010-05-10

Multiple families of viruses have evolved sophisticated strategies to regulate nuclear factor-kappaB (NF-kappaB) signaling, which plays a pivotal role in diverse cellular events, including virus-host interactions. In this study, we report that bovine foamy virus (BFV) is able to activate the NF-kappaB pathway through the action of its transactivator, BTas. Both cellular IKKbeta and IkappaBalpha also participate in this activation. In addition, we demonstrate that BTas induces the processing of p100, which implies that BTas can activate NF-kappaB through a noncanonical pathway as well. Co-immunoprecipitation analysis shows that BTas interacts with IKK catalytic subunits (IKKalpha and IKKbeta), which may be responsible for regulation of IKK kinase activity and persistent NF-kappaB activation. Furthermore, our results indicate that the level of BTas-mediated LTR transcription correlates with the activity of cellular NF-kappaB. Together, this study suggests that BFV activates the NF-kappaB pathway through BTas to enhance viral transcription. Copyright 2010 Elsevier Inc. All rights reserved.
XEDAR activates the non-canonical NF-κB pathway

DOE Office of Scientific and Technical Information (OSTI.GOV)

Verhelst, Kelly, E-mail: Kelly.Verhelst@irc.VIB-UGent.be; Department of Biomedical Molecular Biology, Ghent University, Ghent; Gardam, Sandra, E-mail: s.gardam@garvan.org.au

2015-09-18

Members of the tumor necrosis factor receptor (TNFR) superfamily are involved in a number of physiological and pathological responses by activating a wide variety of intracellular signaling pathways. The X-linked ectodermal dysplasia receptor (XEDAR; also known as EDA2R or TNFRSF27) is a member of the TNFR superfamily that is highly expressed in ectodermal derivatives during embryonic development and binds to ectodysplasin-A2 (EDA-A2), a member of the TNF family that is encoded by the anhidrotic ectodermal dysplasia (EDA) gene. Although XEDAR was first described in the year 2000, its function and molecular mechanism of action is still largely unclear. XEDAR hasmore » been reported to activate canonical nuclear factor κB (NF-κB) signaling and mitogen-activated protein (MAP) kinases. Here we report that XEDAR is also able to trigger the non-canonical NF-κB pathway, characterized by the processing of p100 (NF-κB2) into p52, followed by nuclear translocation of p52 and RelB. We provide evidence that XEDAR-induced p100 processing relies on the binding of XEDAR to TRAF3 and TRAF6, and requires the kinase activity of NIK and IKKα. We also show that XEDAR stimulation results in NIK accumulation and that p100 processing is negatively regulated by TRAF3, cIAP1 and A20. - Highlights: • XEDAR activates the non-canonical NF-κB pathway. • XEDAR-induced processing of p100 depends on XEDAR interaction with TRAF3 and TRAF6. • XEDAR-induced processing of p100 depends on NIK and IKKα activity. • Overexpression of XEDAR leads to NIK accumulation. • XEDAR-induced processing of p100 is negatively regulated by TRAF3 cIAP1 and A20.« less

Anti and Androgenic Activities in MDA-KB2 Cells: A Comparison of Performance in 96 Well Versus HTS Assays

EPA Science Inventory

We developed the MDA-kb2 cell line to screen androgen agonists/antagonists (Wilson et al., ToxSci 66:69, 2002). MDA-kb2 has been used to quantify anti- and androgenic activities of chemicals, mixtures, combustion by-products, oil dispersants and waste, source and drinking water s...
Lactoferricin but not lactoferrin inhibit herpes simplex virus type 2 infection in mice.

PubMed

Shestakov, Andrey; Jenssen, Håvard; Nordström, Inger; Eriksson, Kristina

2012-03-01

We have evaluated the potential of bovine lactoferrin and lactoferricin for their ability to prevent and/or treat genital HSV-2 infection in mice. We confirm previous data showing that both lactoferrin and lactoferricin have antiviral properties in vitro and can inhibit HSV-2 infection of GMK cells in a dose-dependent manner. When tested in vivo, lactoferricin but not lactoferrin was also a potent inhibitor of HSV-2 infection. When admixed with virus prior to inoculation, lactoferricin inhibited disease development and significantly reduced the viral load in a genital model of HSV-2 infection in mice. Lactoferrin and lactoferricin were also tested for their ability to stimulate the production of chemokines. Neither of the compounds induced the production of CCL3, CCL5, CXCL1 or CXCL2 by mouse splenocytes in vitro. However, when tested in vivo, both lactoferrin and lactoferricin were able to induce local vaginal production of CCL5. Lactoferrin also induced CXCL2 production. The prophylactic and/or therapeutic effects of lactoferrin or lactoferricin were also tested. But none of the compounds were efficient in blocking HSV-2 infection when given 24h prior to HSV-2 infection. Lactoferricin however showed promising results as a therapeutic agent and delayed both disease onset by 3days as well as reducing the viral load almost 15-fold when given as a single dose 24h post-infection. These data show that lactoferricin can block genital herpes infection in mice, and perhaps also be used for post-infection treatment. Copyright © 2012 Elsevier B.V. All rights reserved.
A role for NF-κB activity in skin hyperplasia and the development of keratoacanthomata in mice.

PubMed

Poligone, Brian; Hayden, Matthew S; Chen, Luojing; Pentland, Alice P; Jimi, Eijiro; Ghosh, Sankar

2013-01-01

Previous studies have implicated NF-κB signaling in both cutaneous development and oncogenesis. However, these studies have been limited in part by the lethality that results from extreme over- or under-expression of NF-κB in available mouse models. Even cre-driven tissue specific expression of transgenes, or targeted deletion of NF-κB can cause cell death. Therefore, the present study was undertaken to evaluate a novel mouse model of enhanced NF-κB activity in the skin. A knock-in homologous recombination technique was utilized to develop a mouse model (referred to as PD mice) with increased NF-κB activity. The data show that increased NF-κB activity leads to hyperproliferation and dysplasia of the mouse epidermis. Chemical carcinogenesis in the context of enhanced NF-κB activity promotes the development of keratoacanthomata. Our findings support an important role for NF-κB in keratinocyte dysplasia. We have found that enhanced NF-κB activity renders keratinocytes susceptible to hyperproliferation and keratoacanthoma (KA) development but is not sufficient for transformation and SCC development. We therefore propose that NF-κB activation in the absence of additional oncogenic events can promote TNF-dependent, actinic keratosis-like dysplasia and TNF-independent, KAs upon chemical carcinogensis. These studies suggest that resolution of KA cannot occur when NF-κB activation is constitutively enforced.
Effect of low extracellular pH on NF-κB activation in macrophages.

PubMed

Gerry, A B; Leake, D S

2014-04-01

Many diseases, including atherosclerosis, involve chronic inflammation. The master transcription factor for inflammation is NF-κB. Inflammatory sites have a low extracellular pH. Our objective was to demonstrate the effect of pH on NF-κB activation and cytokine secretion. Mouse J774 macrophages or human THP-1 or monocyte-derived macrophages were incubated at pH 7.0-7.4 and inflammatory cytokine secretion and NF-κB activity were measured. A pH of 7.0 greatly decreased pro-inflammatory cytokine secretion (TNF or IL-6) by J774 macrophages, but not THP-1 or human monocyte-derived macrophages. Upon stimulation of mouse macrophages, the levels of IκBα, which inhibits NF-κB, fell but low pH prevented its later increase, which normally restores the baseline activity of NF-κB, even though the levels of mRNA for IκBα were increased. pH 7.0 greatly increased and prolonged NF-κB binding to its consensus promoter sequence, especially the anti-inflammatory p50:p50 homodimers. Human p50 was overexpressed using adenovirus in THP-1 macrophages and monocyte-derived macrophages to see if it would confer pH sensitivity to NF-κB activity in human cells. Overexpression of p50 increased p50:p50 DNA-binding and in THP-1 macrophages inhibited considerably TNF and IL-6 secretion, but there was still no effect of pH on p50:p50 DNA binding or cytokine secretion. A modest decrease in pH can sometimes have marked effects on NF-κB activation and cytokine secretion and might be one reason to explain why mice normally develop less atherosclerosis than do humans. Copyright © 2014 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.
p38 mitogen-activated protein kinase up-regulates NF-{kappa}B transcriptional activation through RelA phosphorylation during stretch-induced myogenesis

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ji, Guoping; Liu, Dongxu; Liu, Jing

2010-01-01

p38 MAPK and nuclear factor-B (NF-B) signaling pathways play an indispensable role in the control of skeletal myogenesis. The specific contribution of these signaling pathways to the response of myoblast to the mechanical stimulation and the molecular mechanisms underlying this response remain unresolved. Using an established in vitro model, we now show that p38 MAP kinase activity regulates the transcriptional activation of NF-{kappa}B in response to mechanical stimulation of myoblasts. Furthermore, SB203580 blocked stretch-induced NF-{kappa}B activation during myogenesis, not through down-regulation of degradation of I{kappa}B-{alpha}, and consequent translocation of the p65 subunit of NF-{kappa}B to the nucleus. It is likelymore » that stretch-induced NF-{kappa}B activation by phosphorylation of p65 NF-{kappa}B. Moreover, depletion of p38{alpha} using siRNA significantly reduces stretch-induced phosphorylation of RelA and NF-{kappa}B activity. These results provides the first evidence of a cross-talk between p38 MAPK and NF-{kappa}B signaling pathways during stretch-induced myogenesis, with phosphorylation of RelA being one of the effectors of this promyogenic mechanism. The {alpha} isoform of p38MAP kinase regulates the transcriptional activation of NF-{kappa}B following stimulation with cyclic stretch.« less
Protein composition of different sized casein micelles in milk after the binding of lactoferrin or lysozyme.

PubMed

Anema, Skelte G; de Kruif, C G Kees

2013-07-24

Casein micelles with bound lactoferrin or lysozyme were fractionated into sizes ranging in radius from ∼50 to 100 nm. The κ-casein content decreased markedly and the αS-casein/β-casein content increased slightly as micelle size increased. For lactoferrin, higher levels were bound to smaller micelles. The lactoferrin/κ-casein ratio was constant for all micelle sizes, whereas the lactoferrin/αS-casein and lactoferrin/β-casein ratio decreased with increasing micelle size. This indicates that the lactoferrin was binding to the surface of the casein micelles. For lysozyme, higher levels bound to larger casein micelles. The lysozyme/αS-casein and lysozyme/β-casein ratios were nearly constant, whereas the lysozyme/κ-casein ratio increased with increasing micelle size, indicating that lysozyme bound to αS-casein and β-casein in the micelle core. Lactoferrin is a large protein that cannot enter the casein protein mesh; therefore, it binds to the micelle surface. The smaller lysozyme can enter the protein mesh and therefore binds to the more charged αS-casein and β-casein.
Regulation Of Nf=kb And Mnsod In Low Dose Radiation Induced Adaptive Protection Of Mouse And Human Skin Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Jian Li

2012-11-07

A sampling of publications resulting from this grant is provided. One is on the subject of NF-ÎºB-Mediated HER2 Overexpression in Radiation-Adaptive Resistance. Another is on NF-ÎºB-mediated adaptive resistance to ionizing radiation.
Thalidomide suppresses NF-kappa B activation induced by TNF and H2O2, but not that activated by ceramide, lipopolysaccharides, or phorbol ester.

PubMed

Majumdar, Sekhar; Lamothe, Betty; Aggarwal, Bharat B

2002-03-15

Thalidomide ([+]-alpha-phthalimidoglutarimide), a psychoactive drug that readily crosses the blood-brain barrier, has been shown to exhibit anti-inflammatory, antiangiogenic, and immunosuppressive properties through a mechanism that is not fully established. Due to the central role of NF-kappaB in these responses, we postulated that thalidomide mediates its effects through suppression of NF-kappaB activation. We investigated the effects of thalidomide on NF-kappaB activation induced by various inflammatory agents in Jurkat cells. The treatment of these cells with thalidomide suppressed TNF-induced NF-kappaB activation, with optimum effect occurring at 50 microg/ml thalidomide. These effects were not restricted to T cells, as other hematopoietic and epithelial cell types were also inhibited. Thalidomide suppressed H(2)O(2)-induced NF-kappaB activation but had no effect on NF-kappaB activation induced by PMA, LPS, okadaic acid, or ceramide, suggesting selectivity in suppression of NF-kappaB. The suppression of TNF-induced NF-kappaB activation by thalidomide correlated with partial inhibition of TNF-induced degradation of an inhibitory subunit of NF-kappaB (IkappaBalpha), abrogation of IkappaBalpha kinase activation, and inhibition of NF-kappaB-dependent reporter gene expression. Thalidomide abolished the NF-kappaB-dependent reporter gene expression activated by overexpression of TNFR1, TNFR-associated factor-2, and NF-kappaB-inducing kinase, but not that activated by the p65 subunit of NF-kappaB. Overall, our results clearly demonstrate that thalidomide suppresses NF-kappaB activation specifically induced by TNF and H(2)O(2) and that this may contribute to its role in suppression of proliferation, inflammation, angiogenesis, and the immune system.
Vitamin K2 stimulates osteoblastogenesis and suppresses osteoclastogenesis by suppressing NF-κB activation.

PubMed

Yamaguchi, Masayoshi; Weitzmann, M Neale

2011-01-01

Several bone protective factors are reported to exhibit stimulatory activities on bone formation coupled with inhibitory effects on bone resorption; one such factor is vitamin K2. Vitamin K species [K1 (phylloquinone) and K2 (menaquinone)] have long been associated with bone protective activities and are receiving intense interest as nutritional supplements for the prevention or amelioration of bone disease in humans. However, the mechanisms of vitamin K action on the skeleton are poorly defined. Activation of the nuclear factor κB (NF-κB) signal transduction pathway is essential for osteoclast formation and resorption. By contrast, NF-κB signaling potently antagonizes osteoblast differentiation and function, prompting us to speculate that NF-κB antagonists may represent a novel class of dual anti-catabolic and pro-anabolic agents. We now show that vitamin K2 action on osteoblast and osteoclast formation and activity is accomplished by down-regulating basal and cytokine-induced NF-κB activation, by increasing IκB mRNA, in a γ-carboxylation-independent manner. Furthermore, vitamin K2 prevented repression by tumor necrosis factor α (TNFα) of SMAD signaling induced by either transforming growth factor ß (TGFß) or bone morphogenetic protein-2 (BMP-2). Vitamin K2 further antagonized receptor activator of NF-κB (RANK) ligand (RANKL)-induced NF-κB activation in osteoclast precursors. Our data provide a novel mechanism to explain the dual pro-anabolic and anti-catabolic activities of vitamin K2, and may further support the concept that pharmacological modulation of NF-κB signal transduction may constitute an effective mechanism for ameliorating pathological bone loss and for promoting bone health.
Bactericidal effect of bovine lactoferrin and synthetic peptide lactoferrin chimera in Streptococcus pneumoniae and the decrease in luxS gene expression by lactoferrin.

PubMed

León-Sicairos, Nidia; Angulo-Zamudio, Uriel A; Vidal, Jorge E; López-Torres, Cynthia A; Bolscher, Jan G M; Nazmi, Kamran; Reyes-Cortes, Ruth; Reyes-López, Magda; de la Garza, Mireya; Canizalez-Román, Adrian

2014-10-01

Streptococcus pneumoniae (pneumococcus) is responsible for nearly one million child deaths annually. Pneumococcus causes infections such as pneumonia, otitis media, meningitis, and sepsis. The human immune system includes antibacterial peptides and proteins such as lactoferrin (LF), but its activity against pneumococcus is not fully understood. The aim of this work was to evaluate the bactericidal effect of bovine lactoferrin (bLF) and the synthetic LF-peptides lactoferricin (LFcin17-30), lactoferrampin (LFampin265-284), and LFchimera against S. pneumoniae planktonic cells. The mechanism of damage was also investigated, as well as the impact of these peptides on the transcription levels of genes known to encode important virulence factors. S. pneumoniae planktonic cells were treated with bLF, LFcin17-30, LFampin265-284 and LFchimera at different time points. The viability of treated planktonic cells was assessed by dilution and plating (in CFU/ml). The interaction between LF and LF-peptides coupled to fluorescein was visualized using a confocal microscope and flow cytometry, whereas the damage at structural levels was observed by electron microscopy. Damage to bacterial membranes was further evaluated by membrane permeabilization by use of propidium iodide and flow cytometry, and finally, the expression of pneumococcal genes was evaluated by qRT-PCR. bLF and LFchimera were the best bactericidal agents. bLF and peptides interacted with bacteria causing changes in the shape and size of the cell and membrane permeabilization. Moreover, the luxS gene was down-regulated in bacteria treated with LF. In conclusion, LF and LFchimera have a bactericidal effect, and LF down-regulates genes involved in the pathogenicity of pneumococcus, thus demonstrating potential as new agents for the treatment of pneumococcal infections.
Inhibition of the aggregation of lactoferrin and (-)-epigallocatechin gallate in the presence of polyphenols, oligosaccharides, and collagen peptide.

PubMed

Yang, Wei; Liu, Fuguo; Xu, Chenqi; Sun, Cuixia; Yuan, Fang; Gao, Yanxiang

2015-05-27

The aggregation of lactoferrin and (-)-epigallocatechin gallate (EGCG) was inhibited by polyphenols, oligosaccharides, and collagen peptide in this study. Polyphenols, oligosaccharides, or collagen peptide can effectively prevent the formation of lactoferrin-EGCG aggregates, respectively. The addition sequence of lactoferrin, polyphenols (oligosaccharides or collagen peptide) and EGCG can affect the turbidity and particle size of the ternary complexes in the buffer solution; however, it hardly affected the ζ-potential and fluorescence characteristics. With either positive or negative charge, polyphenols and collagen peptide disrupted the formation of lactoferrin-EGCG aggregate mainly through the mechanism of its competition with EGCG molecules which surrounded the lactoferrin molecule surface with weaker binding affinities, forming polyphenols or a collagen peptide-lactoferrin-EGCG ternary complex; for neutral oligosaccharides, the ternary complex was generated mainly through steric effects, accompanied by a change in the lactoferrin secondary structure induced by gallic acid, chlorogenic acid, and xylo-oligosaccharide. Polyphenols, oligosaccharides, or collagen peptide restraining the formation of lactoferrin-EGCG aggregate could be applied in the design of clear products in the food, pharmaceutical, and cosmetic industries.
[NF-kappaB-induced gp96 up-regulation promotes hepatocyte growth, cell cycle progression and transition].

PubMed

Feng, Cong; Wu, Bo; Fan, Hongxia; Li, Changfei; Meng, Songdong

2014-10-04

To investigate the mechanism of gp96 raised during hepatitis B virus (HBV) infection and the pathological mechanism. The mechanism of NF-KB activating gp96 expression was determined by bioinformatics analysis, luciferase reporter assay, real-time PCR and Western blot. The effect of over-expression and knockdown gp96 expression by transfection or RNA interference on hepatocyte proliferation, apoptosis and cell cycle was examined by CCK-8 and flow cytometry. The role of gp96 for HCC development was determined by epithelial-mesenchymal transition (EMT) and colony formation assay. NF-kB significantly increased the gp96 expression by binding to the NF-kappaB binding site. Over-expression and knockdown studies both show that gp96 promoted hepatocyte proliferation, inhibited apoptosis, and induced G0/G1 to S phase cell cycle progression. Moreover, gp96 induced epithelial-mesenchymal transition and increased colony formation ability of hepatocytes. Our results therefore provide insights in chronic HBV infection-induced gp96 expression, and indicate that elevated gp96 may contribute to HCC development during chronic inflammation.
NF-kappaB mediates mitogen-activated protein kinase pathway-dependent iNOS expression in human melanoma.

PubMed

Uffort, Deon G; Grimm, Elizabeth A; Ellerhorst, Julie A

2009-01-01

Tumor expression of inducible nitric oxide synthase (iNOS) predicts poor outcomes for melanoma patients. We have reported the regulation of melanoma iNOS by the mitogen-activated protein kinase (MAPK) pathway. In this study, we test the hypothesis that NF-kappaB mediates this regulation. Western blotting of melanoma cell lysates confirmed the constitutive expression of iNOS. Western blot detected baseline levels of activated nuclear extracellular signal-regulated kinase and NF-kappaB. Indirect immunofluorescence confirmed the presence of NF-kappaB p50 and p65 in melanoma cell nuclei, with p50 being more prevalent. Electrophoretic mobility shift assay demonstrated baseline NF-kappaB activity, the findings confirmed by supershift analysis. Treatment of melanoma cells with the MEK inhibitor U0126 decreased NF-kappaB binding to its DNA recognition sequence, implicating the MAPK pathway in NF-kappaB activation. Two specific NF-kappaB inhibitors suppressed iNOS expression, demonstrating regulation of iNOS by NF-kappaB. Several experiments indicated the presence of p50 homodimers, which lack a transactivation domain and rely on the transcriptional coactivator Bcl-3 to carry out this function. Bcl-3 was detected in melanoma cells and co-immunoprecipitated with p50. These data suggest that the constitutively activated melanoma MAPK pathway stimulates activation of NF-kappaB hetero- and homodimers, which, in turn, drive iNOS expression and support melanoma tumorigenesis.
NF-κB signaling participates in both Receptor Activator of NF-κB Ligand- (RANKL) and interleukin-4- (IL-4) induced macrophage fusion: Receptor cross-talk leads to alterations in NF-κB pathways

PubMed Central

Yu, Minjun; Qi, Xiulan; Moreno, Jose L.; Farber, Donna L.; Keegan, Achsah D.

2011-01-01

NF-κB activation is essential for RANKL-induced osteoclast formation. IL-4 is known to inhibit the RANKL-induced osteoclast differentiation, while at the same time promote macrophage fusion to form multinucleated giant cells (MNG). Several groups have proposed that IL-4 inhibition of osteoclastogenesis is mediated by suppressing the RANKL-induced activation of NF-κB. However, we found that IL-4 did not block proximal, canonical NF-κB signaling. Instead, we found that IL-4 inhibited alternative NF-κB signaling and induced p105/50 expression. Interestingly, in nfκb1−/− bone marrow macrophages (BMM), the formation of both multinucleated osteoclast and MNG induced by RANKL or IL-4 respectively was impaired. This suggests that NF-κB signaling also plays an important role in IL-4-induced macrophage fusion. Indeed, we found that the RANKL-induced and IL-4-induced macrophage fusion were both inhibited by the NF-κB inhibitors IKK2 inhibitor, and NEMO inhibitory peptide. Furthermore, overexpression of p50, p65, p52 and RelB individually in nfκb1−/− or nfκb1+/+ BMM enhanced both giant osteoclast and MNG formation. Interestingly, knockdown of nfκb2 in wild type BMM dramatically enhanced both osteoclast and MNG formation. In addition, both RANKL- and IL-4-induced macrophage fusion were impaired in NIK−/− BMM. These results suggest IL-4 influences NF-κB pathways by increasing p105/p50 and suppressing RANKL-induced p52 translocation, and that NF-κB pathways participate in both RANKL- and IL-4- induced giant cell formation. PMID:21734075
Effect of human milk prostaglandins and lactoferrin on respiratory syncytial virus and rotavirus.

PubMed

Grover, M; Giouzeppos, O; Schnagl, R D; May, J T

1997-03-01

The effect of lactoferrin and prostaglandins E and F2 alpha on the growth of rotavirus and respiratory syncytial virus in cell culture was investigated. Lactoferrin inhibited the growth of respiratory syncytial virus at a concentration tenfold lower than that normally present in human milk. The prostaglandins had no effect on either virus growth, even at a concentration of 100-fold more than that found in human milk. Lactoferrin may have some antiviral properties in human milk in addition to its known antibacterial functions.
Polycystin-1 promotes PKC{alpha}-mediated NF-{kappa}B activation in kidney cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Banzi, Manuela; Aguiari, Gianluca; Trimi, Viky

2006-11-17

Polycystin-1 (PC1), the PKD1 gene product, is a membrane receptor which regulates many cell functions, including cell proliferation and apoptosis, both typically increased in cyst lining cells in autosomal dominant polycystic kidney disease. Here we show that PC1 upregulates the NF-{kappa}B signalling pathway in kidney cells to prevent cell death. Human embryonic kidney cell lines (HEK293{sup CTT}), stably expressing a PC1 cytoplasmic terminal tail (CTT), presented increased NF-{kappa}B nuclear levels and NF-{kappa}B-mediated luciferase promoter activity. This, consistently, was reduced in HEK293 cells in which the endogenous PC1 was depleted by RNA interference. CTT-dependent NF-{kappa}B promoter activation was mediated by PKC{alpha}more » because it was blocked by its specific inhibitor Ro-320432. Furthermore, it was observed that apoptosis, which was increased in PC1-depleted cells, was reduced in HEK293{sup CTT} cells and in porcine kidney LtTA cells expressing a doxycycline-regulated CTT. Staurosporine, a PKC inhibitor, and parthenolide, a NF-{kappa}B inhibitor, significantly reduced the CTT-dependent antiapoptotic effect. These data reveal, therefore, a novel pathway by which polycystin-1 activates a PKC{alpha}-mediated NF-{kappa}B signalling and cell survival.« less
Berberine induces FasL-related apoptosis through p38 activation in KB human oral cancer cells

PubMed Central

KIM, JAE-SUNG; OH, DAHYE; YIM, MIN-JI; PARK, JIN-JU; KANG, KYEONG-ROK; CHO, IN-A; MOON, SUNG-MIN; OH, JI-SU; YOU, JAE-SEEK; KIM, CHUN SUNG; KIM, DO KYUNG; LEE, SOOK-YOUNG; LEE, GYEONG-JE; IM, HEE-JEONG; KIM, SU-GWAN

2015-01-01

In the present study, we examined the anticancer properties of berberine in KB oral cancer cells with a specific focus on its cellular mechanism. Berberine did not affect the cell viability of the primary human normal oral keratinocytes that were used as a control. However, the viability of KB cells was found to decrease significantly in the presence of berberine in a dose-dependent manner. Furthermore, in KB cells, berberine induced the fragmentation of genomic DNA, changes in cell morphology, and nuclear condensation. In addition, caspase-3 and -7 activation, and an increase in apoptosis were observed. Berberine was also found to upregulate significantly the expression of the death receptor ligand, FasL. In turn, this upregulation triggered the activation of pro-apoptotic factors such as caspase-8, -9 and -3 and poly(ADP-ribose) polymerase (PARP). Furthermore, pro-apoptotic factors such as Bax, Bad and Apaf-1 were also significantly upregulated by berberine. Anti-apoptotic factors such as Bcl-2 and Bcl-xL were downregulated. Z-VAD-FMK, a cell-permeable pan-caspase inhibitor, suppressed the activation of caspase-3 and PARP. These results clearly indicate that berberine-induced cell death of KB oral cancer cells was mediated by both extrinsic death receptor-dependent and intrinsic mitochondrial-dependent apoptotic signaling pathways. In addition, berberine-induced upregulation of FasL was shown to be mediated by the p38 MAPK signaling pathway. We also found that berberine-induced migration suppression was mediated by downregulation of MMP-2 and MMP-9 through phosphorylation of p38 MAPK. In summary, berberine has the potential to be used as a chemotherapeutic agent, with limited side-effects, for the management of oral cancer. PMID:25634589
NF-κB activity in muscle from obese and type 2 diabetic subjects under basal and exercise-stimulated conditions

PubMed Central

Tantiwong, Puntip; Shanmugasundaram, Karthigayan; Monroy, Adriana; Ghosh, Sangeeta; Li, Mengyao; DeFronzo, Ralph A.; Cersosimo, Eugenio; Sriwijitkamol, Apiradee; Mohan, Sumathy

2010-01-01

NF-κB is a transcription factor that controls the gene expression of several proinflammatory proteins. Cell culture and animal studies have implicated increased NF-κB activity in the pathogenesis of insulin resistance and muscle atrophy. However, it is unclear whether insulin-resistant human subjects have abnormal NF-κB activity in muscle. The effect that exercise has on NF-κB activity/signaling also is not clear. We measured NF-κB DNA-binding activity and the mRNA level of putative NF-κB-regulated myokines interleukin (IL)-6 and monocyte chemotactic protein-1 (MCP-1) in muscle samples from T2DM, obese, and lean subjects immediately before, during (40 min), and after (210 min) a bout of moderate-intensity cycle exercise. At baseline, NF-κB activity was elevated 2.1- and 2.7-fold in obese nondiabetic and T2DM subjects, respectively. NF-κB activity was increased significantly at 210 min following exercise in lean (1.9-fold) and obese (2.6-fold) subjects, but NF-κB activity did not change in T2DM. Exercise increased MCP-1 mRNA levels significantly in the three groups, whereas IL-6 gene expression increased significantly only in lean and obese subjects. MCP-1 and IL-6 gene expression peaked at the 40-min exercise time point. We conclude that insulin-resistant subjects have increased basal NF-κB activity in muscle. Acute exercise stimulates NF-κB in muscle from nondiabetic subjects. In T2DM subjects, exercise had no effect on NF-κB activity, which could be explained by the already elevated NF-κB activity at baseline. Exercise-induced MCP-1 and IL-6 gene expression precedes increases in NF-κB activity, suggesting that other factors promote gene expression of these cytokines during exercise. PMID:20739506
NF-κB activity in muscle from obese and type 2 diabetic subjects under basal and exercise-stimulated conditions.

PubMed

Tantiwong, Puntip; Shanmugasundaram, Karthigayan; Monroy, Adriana; Ghosh, Sangeeta; Li, Mengyao; DeFronzo, Ralph A; Cersosimo, Eugenio; Sriwijitkamol, Apiradee; Mohan, Sumathy; Musi, Nicolas

2010-11-01

NF-κB is a transcription factor that controls the gene expression of several proinflammatory proteins. Cell culture and animal studies have implicated increased NF-κB activity in the pathogenesis of insulin resistance and muscle atrophy. However, it is unclear whether insulin-resistant human subjects have abnormal NF-κB activity in muscle. The effect that exercise has on NF-κB activity/signaling also is not clear. We measured NF-κB DNA-binding activity and the mRNA level of putative NF-κB-regulated myokines interleukin (IL)-6 and monocyte chemotactic protein-1 (MCP-1) in muscle samples from T2DM, obese, and lean subjects immediately before, during (40 min), and after (210 min) a bout of moderate-intensity cycle exercise. At baseline, NF-κB activity was elevated 2.1- and 2.7-fold in obese nondiabetic and T2DM subjects, respectively. NF-κB activity was increased significantly at 210 min following exercise in lean (1.9-fold) and obese (2.6-fold) subjects, but NF-κB activity did not change in T2DM. Exercise increased MCP-1 mRNA levels significantly in the three groups, whereas IL-6 gene expression increased significantly only in lean and obese subjects. MCP-1 and IL-6 gene expression peaked at the 40-min exercise time point. We conclude that insulin-resistant subjects have increased basal NF-κB activity in muscle. Acute exercise stimulates NF-κB in muscle from nondiabetic subjects. In T2DM subjects, exercise had no effect on NF-κB activity, which could be explained by the already elevated NF-κB activity at baseline. Exercise-induced MCP-1 and IL-6 gene expression precedes increases in NF-κB activity, suggesting that other factors promote gene expression of these cytokines during exercise.
Antimicrobial Lactoferrin Peptides: The Hidden Players in the Protective Function of a Multifunctional Protein

PubMed Central

Sinha, Mau; Kaushik, Sanket; Kaur, Punit; Singh, Tej P.

2013-01-01

Lactoferrin is a multifunctional, iron-binding glycoprotein which displays a wide array of modes of action to execute its primary antimicrobial function. It contains various antimicrobial peptides which are released upon its hydrolysis by proteases. These peptides display a similarity with the antimicrobial cationic peptides found in nature. In the current scenario of increasing resistance to antibiotics, there is a need for the discovery of novel antimicrobial drugs. In this context, the structural and functional perspectives on some of the antimicrobial peptides found in N-lobe of lactoferrin have been reviewed. This paper provides the comparison of lactoferrin peptides with other antimicrobial peptides found in nature as well as interspecies comparison of the structural properties of these peptides within the native lactoferrin. PMID:23554820

Orientia tsutsugamushi uses two Ank effectors to modulate NF-κB p65 nuclear transport and inhibit NF-κB transcriptional activation.

PubMed

Evans, Sean M; Rodino, Kyle G; Adcox, Haley E; Carlyon, Jason A

2018-05-01

Orientia tsutsugamushi causes scrub typhus, a potentially fatal infection that threatens over one billion people. Nuclear translocation of the transcription factor, NF-κB, is the central initiating cellular event in the antimicrobial response. Here, we report that NF-κB p65 nuclear accumulation and NF-κB-dependent transcription are inhibited in O. tsutsugamushi infected HeLa cells and/or primary macrophages, even in the presence of TNFα. The bacterium modulates p65 subcellular localization by neither degrading it nor inhibiting IκBα degradation. Rather, it exploits host exportin 1 to mediate p65 nuclear export, as this phenomenon is leptomycin B-sensitive. O. tsutsugamushi antagonizes NF-κB-activated transcription even when exportin 1 is inhibited and NF-κB consequently remains in the nucleus. Two ankyrin repeat-containing effectors (Anks), Ank1 and Ank6, each of which possess a C-terminal F-box and exhibit 58.5% amino acid identity, are linked to the pathogen's ability to modulate NF-κB. When ectopically expressed, both translocate to the nucleus, abrogate NF-κB-activated transcription in an exportin 1-independent manner, and pronouncedly reduce TNFα-induced p65 nuclear levels by exportin 1-dependent means. Flag-tagged Ank 1 and Ank6 co-immunoprecipitate p65 and exportin 1. Both also bind importin β1, a host protein that is essential for the classical nuclear import pathway. Importazole, which blocks importin β1 activity, abrogates Ank1 and Ank6 nuclear translocation. The Ank1 and Ank6 regions that bind importin β1 also mediate their transport into the nucleus. Yet, these regions are distinct from those that bind p65/exportin 1. The Ank1 and Ank6 F-box and the region that lies between it and the ankyrin repeat domain are essential for blocking p65 nuclear accumulation. These data reveal a novel mechanism by which O. tsutsugamushi modulates the activity and nuclear transport of NF-κB p65 and identify the first microbial proteins that co-opt both
A Novel Murine Anti-Lactoferrin Monoclonal Antibody Activates Human Polymorphonuclear Leukocytes through Membrane-Bound Lactoferrin and TLR4

PubMed Central

Hu, Xiao-Min; Xu, Yan-Rui; Yan, Ru; Sun, Shu-Liang; Dong, Hong-Liang; Wang, Jun; Gao, Xiao-Ming

2015-01-01

Soluble lactoferrin (LTF) is a versatile molecule that not only regulates the iron homeostasis, but also harbors direct microbicidal and immunomodulating abilities in mammalian body fluids. In contrast, little is known about the function of membrane-bound LTF (mbLTF), although its expression on human polymorphonuclear leukocytes (huPMNs) has been reported for decades. Given that LTF/anti-LTF antibodies represent a potential diagnostic/prognostic biomarker and a therapeutic target in patients with immune disorders, we wished, in the present study, to generate a novel human LTF- (huLTF-) specific mAb suitable for detailed analyses on the expression and function of mbLTF as well as for deciphering the underlying mechanisms. By using the traditional hybridoma cell fusion technology, we obtained a murine IgG1 (kappa) mAb, M-860, against huLTF. M-860 recognizes a conformational epitope of huLTF as it binds to natural, but not denatured, huLTF in ELISA. Moreover, M-860 detects mbLTF by FACS and captures endogenous huLTF in total cell lysates of huPMNs. Functionally, M-860 induces the activation of huPMNs partially through TLR4 but independently of phagocytosis. M-860 is thus a powerful tool to analyze the expression and function of human mbLTF, which will further our understanding of the roles of LTF in health and disease. PMID:26649297
Adsorption Analysis of Lactoferrin to Titanium, Stainless Steel, Zirconia, and Polymethyl Methacrylate Using the Quartz Crystal Microbalance Method

PubMed Central

Yoshida, Eiji; Hayakawa, Tohru

2016-01-01

It is postulated that biofilm formation in the oral cavity causes some oral diseases. Lactoferrin is an antibacterial protein in saliva and an important defense factor against biofilm development. We analyzed the adsorbed amount of lactoferrin and the dissociation constant (K d) of lactoferrin to the surface of different dental materials using an equilibrium analysis technique in a 27 MHz quartz crystal microbalance (QCM) measurement. Four different materials, titanium (Ti), stainless steel (SUS), zirconia (ZrO2) and polymethyl methacrylate (PMMA), were evaluated. These materials were coated onto QCM sensors and the surfaces characterized by atomic force microscopic observation, measurements of surface roughness, contact angles of water, and zeta potential. QCM measurements revealed that Ti and SUS showed a greater amount of lactoferrin adsorption than ZrO2 and PMMA. Surface roughness and zeta potential influenced the lactoferrin adsorption. On the contrary, the K d value analysis indicated that the adsorbed lactoferrin bound less tightly to the Ti and SUS surfaces than to the ZrO2 and PMMA surfaces. The hydrophobic interaction between lactoferrin and ZrO2 and PMMA is presumed to participate in better binding of lactoferrin to ZrO2 and PMMA surfaces. It was revealed that lactoferrin adsorption behavior was influenced by the characteristics of the material surface. PMID:26998486
Primate lentiviruses use at least three alternative strategies to suppress NF-κB-mediated immune activation

PubMed Central

Gawanbacht, Ali; Van Driessche, Benoît; Van Lint, Carine; Peeters, Martine; Kirchhoff, Frank

2017-01-01

Primate lentiviruses have evolved sophisticated strategies to suppress the immune response of their host species. For example, HIV-2 and most simian immunodeficiency viruses (SIVs) use their accessory protein Nef to prevent T cell activation and antiviral gene expression by downmodulating the T cell receptor CD3. This Nef function was lost in HIV-1 and other vpu-encoding viruses suggesting that the acquisition of Vpu-mediated NF-κB inhibition reduced the selection pressure for inhibition of T cell activation by Nef. To obtain further insights into the modulation of NF-κB activity by primate lentiviral accessory factors, we analyzed 32 Vpr proteins from a large panel of divergent primate lentiviruses. We found that those of SIVcol and SIVolc infecting Colobinae monkeys showed the highest efficacy in suppressing NF-κB activation. Vpr-mediated inhibition of NF-κB resulted in decreased IFNβ promoter activity and suppressed type I IFN induction in virally infected primary cells. Interestingly, SIVcol and SIVolc differ from all other primate lentiviruses investigated by the lack of both, a vpu gene and efficient Nef-mediated downmodulation of CD3. Thus, primate lentiviruses have evolved at least three alternative strategies to inhibit NF-κB-dependent immune activation. Functional analyses showed that the inhibitory activity of SIVolc and SIVcol Vprs is independent of DCAF1 and the induction of cell cycle arrest. While both Vprs target the IKK complex or a factor further downstream in the NF-κB signaling cascade, only SIVolc Vpr stabilizes IκBα and inhibits p65 phosphorylation. Notably, only de-novo synthesized but not virion-associated Vpr suppressed the activation of NF-κB, thus enabling NF-κB-dependent initiation of viral gene transcription during early stages of the replication cycle, while minimizing antiviral gene expression at later stages. Our findings highlight the key role of NF-κB in antiviral immunity and demonstrate that primate lentiviruses
Effect of indomethacin and lactoferrin on human tenocyte proliferation and collagen formation in vitro

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Yaonan; Department of Orthopaedic, Beijing Hospital of Ministry of Public Health, Beijing, China 100730; Wang, Xiao

Highlights: • Indomethacin, a classic NSAID, inhibited human tenocyte proliferation at high concentration (100 µM). • Lactoferrin at 50-100 µg/ml promoted human tenocyte survival, proliferation and collagen synthesis. • Lactoferrin is anabolic to human tenocytes in vitro and reverses potential inhibitory effects of NSAIDs on human tenocytes. - Abstract: Non-steroidal anti-inflammatory drugs (NSAIDs) are widely used in patients with injuries and inflammation of tendon and ligament, and as post-surgical analgesics. The aim of this study is to investigate the effect of indomethacin, a classic NSAID and its combinational effect with an anabolic agent of skeletal tissue, lactoferrin, on the proliferationmore » and collagen formation of human tenocytes in vitro. A factorial experimental design was employed to study the dose-dependent effect of the combination of indomethacin and lactoferrin. The results showed that indomethacin at high concentration (100 μM) inhibited human tenocyte proliferation in culture medium with 1–10% fetal bovine serum (FBS) in vitro. Also, high dose of indomethacin inhibited the collagen formation of human tenocytes in 1% FBS culture medium. Lactoferrin at 50–100 μg/ml promoted human tenocyte survival in serum-free culture medium and enhanced proliferation and collagen synthesis of human tenocytes in 1% FBS culture medium. When 50–100 μg/ml lactoferrin was used in combination with 100–200 μM indomethacin, it partially rescued the inhibitory effects of indomethacin on human tenocyte proliferation, viability and collagen formation. To our knowledge, it is the first evidence that lactoferrin is anabolic to human tenocytes in vitro and reverses potential inhibitory effects of NSAIDs on human tenocytes.« less
Lactoferrin gene promoter variants and their association with clinical and subclinical mastitis in indigenous and crossbred cattle.

PubMed

Chopra, A; Gupta, I D; Verma, A; Chakravarty, A K; Vohra, V

2015-01-01

Lactoferrin (Lf) gene promoter was screened for the presence of single nucleotide polymphism in indigenous and crossbred cattle from North India and to evaluate its association with Mastitis. Study revealed the presence of genetic variation in regulatory region of bovine Lactoferrin gene using PCR-RFLP technique. Three genotypes namely GG, GH and HH were identified. A single nucleotide change, from guanine to adenine at 25th position was found to be significantly associated (p<0.05) with clinical mastitis in indigenous Sahiwal and crossbred Karan Fries cattle maintained at organised herd of National Dairy Research Institute, Karnal. A non-significant association was observed between subclinical mastitis, somatic cell score (SCS), and GG genotype in Karan Fries cattle, however, a lower SCS was observed in animals having GG genotype. Overall a lower incidence of clinical mastitis was recorded in those animals having GG genotype of Lf in Sahiwal and Karan Fries (KF) cattle. The SNP identified in the promoter region may effect expression lactoferrin protein, which may lead to different levels of antibacterial and anti-inflammatory activity of Lf gene. Results from this study indicated the probable role played by Lactoferrin promoter to serve as candidate gene for mastitis susceptibility among indigenous and crossbred milch cattle.
Relationship between carbachol hyperstimulation-induced pancreatic intracellular trypsinogen and NF-kappa B activation in rats in vitro.

PubMed

Jiang, Chunfang; Zheng, Hai; Liu, Sunan; Fang, Kaifeng

2008-02-01

The relationship between intracellular trypsinogen activation and NF-kappa B activation in rat pancreatic acinar cells induced by M3 cholinergic receptor agonist (carbachol) hyperstimulation was studied. Rat pancreatic acinar cells were isolated, cultured and treated with carbachol, the active protease inhibitor (pefabloc) and NF-kappa B inhibitor (PDTC) in vitro. Intracellular trypsin activity was measured by using a fluorogenic substrate. The activity of NF-kappa B was monitored by using electrophoretic mobility shift assay. The results showed that after pretreatment with 2 mmol/L pefabloc, the activities of trypsin and NF-kappa B in pancreatic acinar cells treated with high concentrations of carbachol (10(-3) mol/L) in vitro was significantly decreased as compared with control group (P<0.01). The addition of 10(-2) mol/L PDTC resulted in a significant decrease of NF-kappa B activities in pancreatic acinar cells after treated with high concentrations of carbachol (10(-3) mol/L) in vitro, but the intracellular trypsinogen activity was not obviously inhibited (P>0.05). It was concluded that intracellular trypsinogen activation is likely involved in the regulation of high concentrations of carbachol-induced NF-kappa B activation in pancreatic acinar cells in vitro. NF-kappa B activation is likely not necessary for high concentrations of carbachol-induced trypsinogen activation in pancreatic acinar cells in vitro.
The high-level expression of human tissue plasminogen activator in the milk of transgenic mice with hybrid gene locus strategy.

PubMed

Zhou, Yanrong; Lin, Yanli; Wu, Xiaojie; Xiong, Fuyin; Lv, Yuemeng; Zheng, Tao; Huang, Peitang; Chen, Hongxing

2012-02-01

Transgene expression for the mammary gland bioreactor aimed at producing recombinant proteins requires optimized expression vector construction. Previously we presented a hybrid gene locus strategy, which was originally tested with human lactoferrin (hLF) as target transgene, and an extremely high-level expression of rhLF ever been achieved as to 29.8 g/l in mice milk. Here to demonstrate the broad application of this strategy, another 38.4 kb mWAP-htPA hybrid gene locus was constructed, in which the 3-kb genomic coding sequence in the 24-kb mouse whey acidic protein (mWAP) gene locus was substituted by the 17.4-kb genomic coding sequence of human tissue plasminogen activator (htPA), exactly from the start codon to the end codon. Corresponding five transgenic mice lines were generated and the highest expression level of rhtPA in the milk attained as to 3.3 g/l. Our strategy will provide a universal way for the large-scale production of pharmaceutical proteins in the mammary gland of transgenic animals.
NF-κB activation impairs somatic cell reprogramming in ageing.

PubMed

Soria-Valles, Clara; Osorio, Fernando G; Gutiérrez-Fernández, Ana; De Los Angeles, Alejandro; Bueno, Clara; Menéndez, Pablo; Martín-Subero, José I; Daley, George Q; Freije, José M P; López-Otín, Carlos

2015-08-01

Ageing constitutes a critical impediment to somatic cell reprogramming. We have explored the regulatory mechanisms that constitute age-associated barriers, through derivation of induced pluripotent stem cells (iPSCs) from individuals with premature or physiological ageing. We demonstrate that NF-κB activation blocks the generation of iPSCs in ageing. We also show that NF-κB repression occurs during cell reprogramming towards a pluripotent state. Conversely, ageing-associated NF-κB hyperactivation impairs the generation of iPSCs by eliciting the reprogramming repressor DOT1L, which reinforces senescence signals and downregulates pluripotency genes. Genetic and pharmacological NF-κB inhibitory strategies significantly increase the reprogramming efficiency of fibroblasts from Néstor-Guillermo progeria syndrome and Hutchinson-Gilford progeria syndrome patients, as well as from normal aged donors. Finally, we demonstrate that DOT1L inhibition in vivo extends lifespan and ameliorates the accelerated ageing phenotype of progeroid mice, supporting the interest of studying age-associated molecular impairments to identify targets of rejuvenation strategies.
Regulation of NF-{kappa}B activity in astrocytes: effects of flavonoids at dietary-relevant concentrations

DOE Office of Scientific and Technical Information (OSTI.GOV)

Spilsbury, Alison; Vauzour, David; Spencer, Jeremy P.E.

Highlights: Black-Right-Pointing-Pointer We tested the hypothesis that low concentrations of flavonoids inhibit NF-{kappa}B in astrocytes. Black-Right-Pointing-Pointer Primary cultured astrocytes possess a functional {kappa}B-system, measured using luciferase assays. Black-Right-Pointing-Pointer Seven flavonoids (100 nM-1 {mu}M) failed to reduce NF-{kappa}B activity in astrocytes. Black-Right-Pointing-Pointer Four flavonoids (100 nM-1 {mu}M) failed to reduce TNFa-stimulated NF-{kappa}B activity in astrocytes. Black-Right-Pointing-Pointer (-)-Epicatechin did not regulate nuclear translocation of the NF-{kappa}B subunit, p65. -- Abstract: Neuroinflammation plays an important role in the progression of neurodegenerative disorders such as Alzheimer's disease and Parkinson's disease. Sustained activation of nuclear transcription factor {kappa}B (NF-{kappa}B) is thought to play an importantmore » role in the pathogenesis of neurodegenerative disorders. Flavonoids have been shown to possess antioxidant and anti-inflammatory properties and we investigated whether flavonoids, at submicromolar concentrations relevant to their bioavailability from the diet, were able to modulate NF-{kappa}B signalling in astrocytes. Using luciferase reporter assays, we found that tumour necrosis factor (TNF{alpha}, 150 ng/ml) increased NF-{kappa}B-mediated transcription in primary cultures of mouse cortical astrocytes, which was abolished on co-transfection of a dominant-negative I{kappa}B{alpha} construct. In addition, TNF{alpha} increased nuclear localisation of p65 as shown by immunocytochemistry. To investigate potential flavonoid modulation of NF-{kappa}B activity, astrocytes were treated with flavonoids from different classes; flavan-3-ols ((-)-epicatechin and (+)-catechin), flavones (luteolin and chrysin), a flavonol (kaempferol) or the flavanones (naringenin and hesperetin) at dietary-relevant concentrations (0.1-1 {mu}M) for 18 h. None of the flavonoids modulated constitutive or
NF-kB activation and its downstream target genes expression after heavy ions exposure

NASA Astrophysics Data System (ADS)

Chishti, Arif Ali; Baumstark-Khan, Christa; Hellweg, Christine; Schmitz, Claudia; Koch, Kristina; Feles, Sebastian

2016-07-01

To enable long-term human space flight cellular radiation response to densely ionizing radiation needs to be better understood for developing appropriate countermeasures to mitigate acute effects and late radiation risks for the astronaut. The biological effectiveness of accelerated heavy ions (which constitute the most important radiation type in space) with high linear energy transfer (LET) for effecting DNA damage response pathways as a gateway to cell death or survival is of major concern not only for space missions but also for new regimes of tumor radiotherapy. In the current research study, the contribution of NF-κB in response to space-relevant radiation qualities was determined by a NF-κB reporter cell line (HEK-pNF-κB-d2EGFP/Neo L2). The NF-κB dependent reporter gene expression (d2EGFP) after ionizing radiation (X-rays and heavy ions) exposure was evaluated by flow cytometry. Because of differences in the extent of NF-κB activation after X-irradiation and heavy ions exposure, it was expected that radiation quality (LET) might play an important role in the cellular radiation response. In addition, the biological effectiveness (RBE) of NF-κB activation and reduction of cellular survival was examined for heavy ions having a broad range of LET (˜0.3 - 9674 keV/µm). Furthermore, the effect of LET on NF-κB target gene expression was analyzed by real time reverse transcriptase quantitative PCR (RT-qPCR). In this study it was proven that NF-κB activation and NF-κB dependent gene expression comprises an early step in cellular radiation response. Taken together, this study clearly demonstrates that NF-κB activation and NF-κB-dependent gene expression by heavy ions are highest in the LET range of ˜50-200 keV/μupm. The up-regulated chemokines and cytokines (CXCL1, CXCL2, CXCL10, IL-8 and TNF) might be important for cell-cell communication among hit as well as unhit cells (bystander effect). The results obtained suggest the NF-κB pathway to be a
MUB40 Binds to Lactoferrin and Stands as a Specific Neutrophil Marker.

PubMed

Anderson, Mark C; Chaze, Thibault; Coïc, Yves-Marie; Injarabian, Louise; Jonsson, Friederike; Lombion, Naelle; Selimoglu-Buet, Dorothée; Souphron, Judith; Ridley, Caroline; Vonaesch, Pascale; Baron, Bruno; Arena, Ellen T; Tinevez, Jean-Yves; Nigro, Giulia; Nothelfer, Katharina; Solary, Eric; Lapierre, Valérie; Lazure, Thierry; Matondo, Mariette; Thornton, David; Sansonetti, Philippe J; Baleux, Françoise; Marteyn, Benoit S

2018-04-19

Neutrophils represent the most abundant immune cells recruited to inflamed tissues. A lack of dedicated tools has hampered their detection and study. We show that a synthesized peptide, MUB 40 , binds to lactoferrin, the most abundant protein stored in neutrophil-specific and tertiary granules. Lactoferrin is specifically produced by neutrophils among other leukocytes, making MUB 40 a specific neutrophil marker. Naive mammalian neutrophils (human, guinea pig, mouse, rabbit) were labeled by fluorescent MUB 40 conjugates (-Cy5, Dylight405). A peptidase-resistant retro-inverso MUB 40 (RI-MUB 40 ) was synthesized and its lactoferrin-binding property validated. Neutrophil lactoferrin secretion during in vitro Shigella infection was assessed with RI-MUB 40 -Cy5 using live cell microscopy. Systemically administered RI-MUB 40 -Cy5 accumulated at sites of inflammation in a mouse arthritis inflammation model in vivo and showed usefulness as a potential tool for inflammation detection using non-invasive imaging. Improving neutrophil detection with the universal and specific MUB 40 marker will aid the study of broad ranges of inflammatory diseases. Copyright © 2018 Elsevier Ltd. All rights reserved.
Regulation of CYBB Gene Expression in Human Phagocytes by a Distant Upstream NF-κB Binding Site.

PubMed

Frazão, Josias B; Thain, Alison; Zhu, Zhiqing; Luengo, Marcos; Condino-Neto, Antonio; Newburger, Peter E

2015-09-01

The human CYBB gene encodes the gp91-phox component of the phagocyte oxidase enzyme complex, which is responsible for generating superoxide and other downstream reactive oxygen species essential to microbial killing. In the present study, we have identified by sequence analysis a putative NF-κB binding site in a DNase I hypersensitive site, termed HS-II, located in the distant 5' flanking region of the CYBB gene. Electrophoretic mobility assays showed binding of the sequence element by recombinant NF-κB protein p50 and by proteins in nuclear extract from the HL-60 myeloid leukemia cell line corresponding to p50 and to p50/p65 heterodimers. Chromatin immunoprecipitation demonstrated NF-κB binding to the site in intact HL-60 cells. Chromosome conformation capture (3C) assays demonstrated physical interaction between the NF-κB binding site and the CYBB promoter region. Inhibition of NF-κB activity by salicylate reduced CYBB expression in peripheral blood neutrophils and differentiated U937 monocytic leukemia cells. U937 cells transfected with a mutant inhibitor of κB "super-repressor" showed markedly diminished CYBB expression. Luciferase reporter analysis of the NF-κB site linked to the CYBB 5' flanking promoter region revealed enhanced expression, augmented by treatment with interferon-γ. These studies indicate a role for this distant, 15 kb upstream, binding site in NF-κB regulation of the CYBB gene, an essential component of phagocyte-mediated host defense. © 2015 Wiley Periodicals, Inc.
Late-phase synthesis of IκBα insulates the TLR4-activated canonical NF-κB pathway from noncanonical NF-κB signaling in macrophages

PubMed Central

Mukherjee, Tapas; Taye, Nandaraj; Vijayaragavan, Bharath; Chattopadhyay, Samit; Gomes, James; Basak, Soumen

2017-01-01

The nuclear factor κB (NF-κB) transcription factors coordinate the inflammatory immune response during microbial infection. Pathogenic substances engage canonical NF-κB signaling through the heterodimer RelA:p50, which is subjected to rapid negative feedback by inhibitor of κBα (IκBα). The noncanonical NF-κB pathway is required for the differentiation of immune cells; however, crosstalk between both pathways can occur. Concomitantly activated noncanonical signaling generates p52 from the p100 precursor. The synthesis of p100 is induced by canonical signaling, leading to formation of the late-acting RelA:p52 heterodimer. This crosstalk prolongs inflammatory RelA activity in epithelial cells to ensure pathogen clearance. We found that the Toll-like receptor 4 (TLR4)–activated canonical NF-κB signaling pathway is insulated from lymphotoxin β receptor (LTβR)–induced noncanonical signaling in mouse macrophage cell lines. Combined computational and biochemical studies indicated that the extent of NF-κB–responsive expression of Nfkbia, which encodes IκBα, inversely correlated with crosstalk. The Nfkbia promoter showed enhanced responsiveness to NF-κB activation in macrophages compared to that in fibroblasts. We found that this hyperresponsive promoter engaged the RelA:p52 dimer generated during costimulation of macrophages through TLR4 and LTβR to trigger synthesis of IκBα at late time points, which prevented the late-acting RelA crosstalk response. Together, these data suggest that despite the presence of identical signaling networks in cells of diverse lineages, emergent crosstalk between signaling pathways is subject to cell type–specific regulation. We propose that the insulation of canonical and noncanonical NF-κB pathways limits the deleterious effects of macrophage-mediated inflammation. PMID:27923915
Oral lactoferrin for the prevention of sepsis and necrotizing enterocolitis in preterm infants

USDA-ARS?s Scientific Manuscript database

Lactoferrin, a normal component of human colostrum, milk, tears, and saliva can enhance host defence and may be effective in the prevention of sepsis and necrotizing enterocolitis (NEC) in preterm neonates. To assess the safety and effectiveness of oral lactoferrin in the prevention of sepsis and NE...
Oral lactoferrin for the prevention of sepsis and necrotizing enterocolotis in preterm infants

USDA-ARS?s Scientific Manuscript database

Lactoferrin, a normal component of human colostrum, milk, tears and saliva can enhance host defence and may be effective in the prevention of sepsis and necrotizing enterocolitis (NEC) in preterm neonates. To assess the safety and effectiveness of oral lactoferrin in the prevention of sepsis and NEC...
Lactoferrin Adsorbed onto Biomimetic Hydroxyapatite Nanocrystals Controlling - In Vivo - the Helicobacter pylori Infection

PubMed Central

Fulgione, Andrea; Nocerino, Nunzia; Iannaccone, Marco; Roperto, Sante; Capuano, Federico; Roveri, Norberto; Lelli, Marco; Crasto, Antonio; Calogero, Armando; Pilloni, Argenia Paola; Capparelli, Rosanna

2016-01-01

Background The resistance of Helicobacter pylori to the antibiotic therapy poses the problem to discover new therapeutic approaches. Recently it has been stated that antibacterial, immunomodulatory, and antioxidant properties of lactoferrin are increased when this protein is surface-linked to biomimetic hydroxyapatite nanocrystals. Objective Based on these knowledge, the aim of the study was to investigate the efficacy of lactoferrin delivered by biomimetic hydroxyapatite nanoparticles with cell free supernatant from probiotic Lactobacillus paracasei as an alternative therapy against Helicobacter pylori infection. Methods Antibacterial and antinflammatory properties, humoral antibody induction, histopathological analysis and absence of side effects were evaluated in both in vitro and in vivo studies. Results The tests carried out have been demonstrated better performance of lactoferrin delivered by biomimetic hydroxyapatite nanoparticles combined with cell free supernatant from probiotic Lactobacillus paracasei compared to both lactoferrin and probiotic alone or pooled. Conclusion These findings indicate the effectiveness and safety of our proposed therapy as alternative treatment for Helicobacter pylori infection. PMID:27384186
Coadsorption of Human Milk Lactoferrin into the Dipalmitoylglycerolphosphatidylcholine Phospholipid Monolayer Spread at the Air/Water Interface

PubMed Central

Miano, Fausto; Zhao, Xiubo; Lu, Jian R.; Penfold, Jeff

2007-01-01

The coadsorption of human milk lactoferrin into a spread monolayer of dipalmitoylglycerol phosphatidylcholine (DPPC) at the air/water interface has been studied by neutron reflection. The system is a good model of the preocular tear film outer interface, which was the motivation for the study. The association of the protein with the surface was indicated by an increase of the surface pressure exerted by the DPPC monolayer. The extent of lactoferrin coadsorption was found to decrease with increasing surface pressure in the lipid monolayer, a trend consistent with the observation reported for other proteins, such as lysozyme and β-lactoglobulin. The neutron reflectivity measurements were subsequently carried out at the three surface pressures of 8, 15, and 35 mN/m to examine the structure and composition of lactoferrin coadsorbed at the interface. Whereas the DPPC monolayer effectively prevented lactoferrin insertion at the high surface pressure, a measurable amount of lactoferrin was found at the air/water interface at the two lower surface pressures. At 15 mN/m it was difficult to identify the distribution of lactoferrin with respect to the DPPC monolayer, due to its relatively low adsorbed amount and much broader distribution. At the lowest surface pressure of 8 mN/m, the lactoferrin coadsorption was found to increase with time over the first few hours. After 5 h the distribution of the lactoferrin layer became similar to, though quantitatively lower than, that adsorbed in the absence of the DPPC monolayer. It is characterized by a top dense sublayer of 15 Å with a bottom diffuse sublayer of 60 Å, indicating structural unfolding induced by surface adsorption under these conditions. PMID:17114223
SIRT1 Activators Suppress Inflammatory Responses through Promotion of p65 Deacetylation and Inhibition of NF-κB Activity

PubMed Central

Yang, Hongying; Zhang, Wei; Pan, Heng; Feldser, Heidi G.; Lainez, Elden; Miller, Christine; Leung, Stewart; Zhong, Zhong; Zhao, Huizhen; Sweitzer, Sharon; Considine, Thomas; Riera, Thomas; Suri, Vipin; White, Brian; Ellis, James L.; Vlasuk, George P.; Loh, Christine

2012-01-01

Chronic inflammation is a major contributing factor in the pathogenesis of many age-associated diseases. One central protein that regulates inflammation is NF-κB, the activity of which is modulated by post-translational modifications as well as by association with co-activator and co-repressor proteins. SIRT1, an NAD+-dependent protein deacetylase, has been shown to suppress NF-κB signaling through deacetylation of the p65 subunit of NF-κB resulting in the reduction of the inflammatory responses mediated by this transcription factor. The role of SIRT1 in the regulation of NF-κB provides the necessary validation for the development of pharmacological strategies for activating SIRT1 as an approach for the development of a new class of anti-inflammatory therapeutics. We report herein the development of a quantitative assay to assess compound effects on acetylated p65 protein in the cell. We demonstrate that small molecule activators of SIRT1 (STACs) enhance deacetylation of cellular p65 protein, which results in the suppression of TNFα-induced NF-κB transcriptional activation and reduction of LPS-stimulated TNFα secretion in a SIRT1-dependent manner. In an acute mouse model of LPS-induced inflammation, the STAC SRTCX1003 decreased the production of the proinflammatory cytokines TNFα and IL-12. Our studies indicate that increasing SIRT1-mediated NF-κB deacetylation using small molecule activating compounds is a novel approach to the development of a new class of therapeutic anti-inflammatory agents. PMID:23029496
Synergistic antifungal effect of lactoferrin with azole antifungals against Candida albicans and a proposal for a new treatment method for invasive candidiasis.

PubMed

Kobayashi, Tsutomu; Kakeya, Hiroshi; Miyazaki, Taiga; Izumikawa, Koichi; Yanagihara, Katsunori; Ohno, Hideaki; Yamamoto, Yoshihiro; Tashiro, Takayoshi; Kohno, Shigeru

2011-01-01

The combination of lactoferrin with fluconazole has been reported to synergistically enhance the antifungal activity of fluconazole against Candida spp. and inhibit the hyphal formation in fluconazole-resistant strains of Candida albicans. In this study, we investigated the association between the therapeutic effects of this combination and the pharmacological characteristics of fluconazole and itraconazole and the variation in these effects with differences among the strains in terms of the susceptibility and resistance mechanisms. Lactoferrin enhanced the growth-inhibitory activity of fluconazole against two different ergosterol mutants but not againt pump mutants or an azole-susceptible strain; but increased the activity of itraconazole against all the strains tested in this study. Exogenous iron cancelled the synergistic effect, which suggests that the iron-chelating function of lactoferrin may contribute to the synergism. Besides, radiolabeled fluconazole assays revealed that lactoferrin did not affect the intracellular concentrations of fluconazole, thereby indicating that these synergistic effects were not due to the alteration of the intracellular uptake of the drug. The development of new clinical treatments and therapeutic method against resistant Candida will depend on our understanding of the resistance mechanisms and methods to overcome them by the application of suitable drug combinations with synergistic effects. The results of this study might contribute to the improvement of our understand of the mechanisms underlying the resistance of Candida strains.

NF-κB Activation in Hypothalamic Pro-opiomelanocortin Neurons Is Essential in Illness- and Leptin-induced Anorexia*

PubMed Central

Jang, Pil-Geum; Namkoong, Cherl; Kang, Gil Myoung; Hur, Man-Wook; Kim, Seung-Whan; Kim, Geun Hyang; Kang, Yeoungsup; Jeon, Min-Jae; Kim, Eun Hee; Lee, Myung-Shik; Karin, Michael; Baik, Ja-Hyun; Park, Joong-Yeol; Lee, Ki-Up; Kim, Young-Bum; Kim, Min-Seon

2010-01-01

Anorexia and weight loss are prevalent in infectious diseases. To investigate the molecular mechanisms underlying these phenomena, we established animal models of infection-associated anorexia by administrating bacterial and viral products, lipopolysaccharide (LPS) and human immunodeficiency virus-1 transactivator protein (Tat). In these models, we found that the nuclear factor-κB (NF-κB), a pivotal transcription factor for inflammation-related proteins, was activated in the hypothalamus. In parallel, administration of LPS and Tat increased hypothalamic pro-inflammatory cytokine production, which was abrogated by inhibition of hypothalamic NF-κB. In vitro, NF-κB activation directly stimulated the transcriptional activity of pro-opiomelanocortin (POMC), a precursor of anorexigenic melanocortin, and mediated the stimulatory effects of LPS, Tat, and pro-inflammatory cytokines on POMC transcription, implying the involvement of NF-κB in controlling feeding behavior. Consistently, hypothalamic injection of LPS and Tat caused a significant reduction in food intake and body weight, which was prevented by blockade of NF-κB and melanocortin. Furthermore, disruption of IκB kinase-β, an upstream kinase of NF-κB, in POMC neurons attenuated LPS- and Tat-induced anorexia. These findings suggest that infection-associated anorexia and weight loss are mediated via NF-κB activation in hypothalamic POMC neurons. In addition, hypothalamic NF-κB was activated by leptin, an important anorexigenic hormone, and mediates leptin-stimulated POMC transcription, indicating that hypothalamic NF-κB also serves as a downstream signaling pathway of leptin. PMID:20097762
NF-kappaB activation in hypothalamic pro-opiomelanocortin neurons is essential in illness- and leptin-induced anorexia.

PubMed

Jang, Pil-Geum; Namkoong, Cherl; Kang, Gil Myoung; Hur, Man-Wook; Kim, Seung-Whan; Kim, Geun Hyang; Kang, Yeoungsup; Jeon, Min-Jae; Kim, Eun Hee; Lee, Myung-Shik; Karin, Michael; Baik, Ja-Hyun; Park, Joong-Yeol; Lee, Ki-Up; Kim, Young-Bum; Kim, Min-Seon

2010-03-26

Anorexia and weight loss are prevalent in infectious diseases. To investigate the molecular mechanisms underlying these phenomena, we established animal models of infection-associated anorexia by administrating bacterial and viral products, lipopolysaccharide (LPS) and human immunodeficiency virus-1 transactivator protein (Tat). In these models, we found that the nuclear factor-kappaB (NF-kappaB), a pivotal transcription factor for inflammation-related proteins, was activated in the hypothalamus. In parallel, administration of LPS and Tat increased hypothalamic pro-inflammatory cytokine production, which was abrogated by inhibition of hypothalamic NF-kappaB. In vitro, NF-kappaB activation directly stimulated the transcriptional activity of pro-opiomelanocortin (POMC), a precursor of anorexigenic melanocortin, and mediated the stimulatory effects of LPS, Tat, and pro-inflammatory cytokines on POMC transcription, implying the involvement of NF-kappaB in controlling feeding behavior. Consistently, hypothalamic injection of LPS and Tat caused a significant reduction in food intake and body weight, which was prevented by blockade of NF-kappaB and melanocortin. Furthermore, disruption of I kappaB kinase-beta, an upstream kinase of NF-kappaB, in POMC neurons attenuated LPS- and Tat-induced anorexia. These findings suggest that infection-associated anorexia and weight loss are mediated via NF-kappaB activation in hypothalamic POMC neurons. In addition, hypothalamic NF-kappaB was activated by leptin, an important anorexigenic hormone, and mediates leptin-stimulated POMC transcription, indicating that hypothalamic NF-kappaB also serves as a downstream signaling pathway of leptin.
A role for NRAGE in NF-κB activation through the non-canonical BMP pathway

PubMed Central

2010-01-01

Background Previous studies have linked neurotrophin receptor-interacting MAGE protein to the bone morphogenic protein signaling pathway and its effect on p38 mediated apoptosis of neural progenitor cells via the XIAP-Tak1-Tab1 complex. Its effect on NF-κB has yet to be explored. Results Herein we report that NRAGE, via the same XIAP-Tak1-Tab1 complex, is required for the phosphorylation of IKK -α/β and subsequent transcriptional activation of the p65 subunit of NF-κB. Ablation of endogenous NRAGE by siRNA inhibited NF-κB pathway activation, while ablation of Tak1 and Tab1 by morpholino inhibited overexpression of NRAGE from activating NF-κB. Finally, cytokine profiling of an NRAGE over-expressing stable line revealed the expression of macrophage migration inhibitory factor. Conclusion Modulation of NRAGE expression revealed novel roles in regulating NF-κB activity in the non-canonical bone morphogenic protein signaling pathway. The expression of macrophage migration inhibitory factor by bone morphogenic protein -4 reveals novel crosstalk between an immune cytokine and a developmental pathway. PMID:20100315
Lactoferrin and prematurity: a promising milk protein?

PubMed

Ochoa, Theresa J; Sizonenko, Stéphane V

2017-02-01

Lactoferrin (Lf) is the major whey protein in milk, with multiple beneficial health effects including direct antimicrobial activities, anti-inflammatory effects, and iron homeostasis. Oral Lf supplementation in human preterm infants has been shown to reduce the incidence of sepsis and necrotizing enterocolitis. In preclinical models of antenatal stress and perinatal brain injury, bovine Lf protected the developing brain from neuronal loss, improved connectivity, increased neurotrophic factors, and decreased inflammation. It also supported brain development and cognition. Further, Lf can prevent preterm delivery by reducing proinflammatory factors and inhibiting premature cervix maturation. We review here the latest research on Lf in the field of neonatology.
Low nuclear body formation and tax SUMOylation do not prevent NF-kappaB promoter activation.

PubMed

Bonnet, Amandine; Randrianarison-Huetz, Voahangy; Nzounza, Patrycja; Nedelec, Martine; Chazal, Maxime; Waast, Laetitia; Pene, Sabrina; Bazarbachi, Ali; Mahieux, Renaud; Bénit, Laurence; Pique, Claudine

2012-09-25

The Tax protein encoded by Human T-lymphotropic virus type 1 (HTLV-1) is a powerful activator of the NF-κB pathway, a property critical for HTLV-1-induced immortalization of CD4⁺ T lymphocytes. Tax permanently stimulates this pathway at a cytoplasmic level by activating the IκB kinase (IKK) complex and at a nuclear level by enhancing the binding of the NF-κB factor RelA to its cognate promoters and by forming nuclear bodies, believed to represent transcriptionally active structures. In previous studies, we reported that Tax ubiquitination and SUMOylation play a critical role in Tax localization and NF-κB activation. Indeed, analysis of lysine Tax mutants fused or not to ubiquitin or SUMO led us to propose a two-step model in which Tax ubiquitination first intervenes to activate IKK while Tax SUMOylation is subsequently required for promoter activation within Tax nuclear bodies. However, recent studies showing that ubiquitin or SUMO can modulate Tax activities in either the nucleus or the cytoplasm and that SUMOylated Tax can serve as substrate for ubiquitination suggested that Tax ubiquitination and SUMOylation may mediate redundant rather than successive functions. In this study, we analyzed the properties of a new Tax mutant that is properly ubiquitinated, but defective for both nuclear body formation and SUMOylation. We report that reducing Tax SUMOylation and nuclear body formation do not alter the ability of Tax to activate IKK, induce RelA nuclear translocation, and trigger gene expression from a NF-κB promoter. Importantly, potent NF-κB promoter activation by Tax despite low SUMOylation and nuclear body formation is also observed in T cells, including CD4⁺ primary T lymphocytes. Moreover, we show that Tax nuclear bodies are hardly observed in HTLV-1-infected T cells. Finally, we provide direct evidence that the degree of NF-κB activation by Tax correlates with the level of Tax ubiquitination, but not SUMOylation. These data reveal that the
Low nuclear body formation and tax SUMOylation do not prevent NF-kappaB promoter activation

PubMed Central

2012-01-01

Background The Tax protein encoded by Human T-lymphotropic virus type 1 (HTLV-1) is a powerful activator of the NF-κB pathway, a property critical for HTLV-1-induced immortalization of CD4+ T lymphocytes. Tax permanently stimulates this pathway at a cytoplasmic level by activating the IκB kinase (IKK) complex and at a nuclear level by enhancing the binding of the NF-κB factor RelA to its cognate promoters and by forming nuclear bodies, believed to represent transcriptionally active structures. In previous studies, we reported that Tax ubiquitination and SUMOylation play a critical role in Tax localization and NF-κB activation. Indeed, analysis of lysine Tax mutants fused or not to ubiquitin or SUMO led us to propose a two-step model in which Tax ubiquitination first intervenes to activate IKK while Tax SUMOylation is subsequently required for promoter activation within Tax nuclear bodies. However, recent studies showing that ubiquitin or SUMO can modulate Tax activities in either the nucleus or the cytoplasm and that SUMOylated Tax can serve as substrate for ubiquitination suggested that Tax ubiquitination and SUMOylation may mediate redundant rather than successive functions. Results In this study, we analyzed the properties of a new Tax mutant that is properly ubiquitinated, but defective for both nuclear body formation and SUMOylation. We report that reducing Tax SUMOylation and nuclear body formation do not alter the ability of Tax to activate IKK, induce RelA nuclear translocation, and trigger gene expression from a NF-κB promoter. Importantly, potent NF-κB promoter activation by Tax despite low SUMOylation and nuclear body formation is also observed in T cells, including CD4+ primary T lymphocytes. Moreover, we show that Tax nuclear bodies are hardly observed in HTLV-1-infected T cells. Finally, we provide direct evidence that the degree of NF-κB activation by Tax correlates with the level of Tax ubiquitination, but not SUMOylation. Conclusions These
Sustained hyperoxia-induced NF-κB activation improves survival and preserves lung development in neonatal mice

PubMed Central

McKenna, Sarah; Michaelis, Katherine A.; Agboke, Fadeke; Liu, Thanh; Han, Kristie; Yang, Guang; Dennery, Phyllis A.

2014-01-01

Oxygen toxicity contributes to the pathogenesis of bronchopulmonary dysplasia (BPD). Neonatal mice exposed to hyperoxia develop a simplified lung structure that resembles BPD. Sustained activation of the transcription factor NF-κB and increased expression of protective target genes attenuate hyperoxia-induced mortality in adults. However, the effect of enhancing hyperoxia-induced NF-κB activity on lung injury and development in neonatal animals is unknown. We performed this study to determine whether sustained NF-κB activation, mediated through IκBβ overexpression, preserves lung development in neonatal animals exposed to hyperoxia. Newborn wild-type (WT) and IκBβ-overexpressing (AKBI) mice were exposed to hyperoxia (>95%) or room air from day of life (DOL) 0–14, after which all animals were kept in room air. Survival curves were generated through DOL 14. Lung development was assessed using radial alveolar count (RAC) and mean linear intercept (MLI) at DOL 3 and 28 and pulmonary vessel density at DOL 28. Lung tissue was collected, and NF-κB activity was assessed using Western blot for IκB degradation and NF-κB nuclear translocation. WT mice demonstrated 80% mortality through 14 days of exposure. In contrast, AKBI mice demonstrated 60% survival. Decreased RAC, increased MLI, and pulmonary vessel density caused by hyperoxia in WT mice were significantly attenuated in AKBI mice. These findings were associated with early and sustained NF-κB activation and expression of cytoprotective target genes, including vascular endothelial growth factor receptor 2. We conclude that sustained hyperoxia-induced NF-κB activation improves neonatal survival and preserves lung development. Potentiating early NF-κB activity after hyperoxic exposure may represent a therapeutic intervention to prevent BPD. PMID:24748603
Uric Acid Induces Renal Inflammation via Activating Tubular NF-κB Signaling Pathway

PubMed Central

Zhou, Yang; Fang, Li; Jiang, Lei; Wen, Ping; Cao, Hongdi; He, Weichun; Dai, Chunsun; Yang, Junwei

2012-01-01

Inflammation is a pathologic feature of hyperuricemia in clinical settings. However, the underlying mechanism remains unknown. Here, infiltration of T cells and macrophages were significantly increased in hyperuricemia mice kidneys. This infiltration of inflammatory cells was accompanied by an up-regulation of TNF-α, MCP-1 and RANTES expression. Further, infiltration was largely located in tubular interstitial spaces, suggesting a role for tubular cells in hyperuricemia-induced inflammation. In cultured tubular epithelial cells (NRK-52E), uric acid, probably transported via urate transporter, induced TNF-α, MCP-1 and RANTES mRNA as well as RANTES protein expression. Culture media of NRK-52E cells incubated with uric acid showed a chemo-attractive ability to recruit macrophage. Moreover uric acid activated NF-κB signaling. The uric acid-induced up-regulation of RANTES was blocked by SN 50, a specific NF-κB inhibitor. Activation of NF-κB signaling was also observed in tubule of hyperuricemia mice. These results suggest that uric acid induces renal inflammation via activation of NF-κB signaling. PMID:22761883
NF-{kappa}B p65 represses {beta}-catenin-activated transcription of cyclin D1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hwang, Injoo; Choi, Yong Seok; Jeon, Mi-Ya

2010-12-03

Research highlights: {yields} Cyclin D1 transcription is directly activated by {beta}-catenin; however, {beta}-catenin-induced cyclin D1 transcription is reduced by NF-{kappa}B p65. {yields} Protein-protein interaction between NF-{kappa}B p65 and {beta}-catenin might be responsible for p65-mediated repression of cyclin D1. {yields} One of five putative binding sites, located further upstream of other sites, is the major {beta}-catenin binding site in the cyclin D1 promoter. {yields} NF-{kappa}B binding site in cyclin D1 is occupied not only by p65 but also by {beta}-catenin, which is dynamically regulated by the signal. -- Abstract: Signaling crosstalk between the {beta}-catenin and NF-{kappa}B pathways represents a functional network.more » To test whether the crosstalk also occurs on their common target genes, the cyclin D1 promoter was used as a model because it contains binding sites for both proteins. {beta}-catenin activated transcription from the cyclin D1 promoter, while co-expression of NF-{kappa}B p65 reduced {beta}-catenin-induced transcription. Chromatin immunoprecipitation revealed lithium chloride-induced binding of {beta}-catenin on one of the T-cell activating factor binding sites. More interestingly, {beta}-catenin binding was greatly reduced by NF-{kappa}B p65, possibly by the protein-protein interaction between the two proteins. Such a dynamic and complex binding of {beta}-catenin and NF-{kappa}B on promoters might contribute to the regulated expression of their target genes.« less
Effect of Oral Lactoferrin on Cataract Surgery Induced Dry Eye: A Randomised Controlled Trial.

PubMed

Devendra, Jaya; Singh, Sneha

2015-10-01

Cataract surgery is one of the most frequently performed intra-ocular surgeries, of these manual Small Incision Cataract Surgery (SICS) is a time tested technique of cataract removal. Any corneal incisional surgery, including cataract surgery, can induce dry eye postoperatively. Various factors have been implicated, of which oneis the inflammation induced by the surgery. Lactoferrin, a glycoprotein present in tears is said to have anti-inflammatory effects, and promotes cell growth. It has been used orally in patients of immune mediated dry eye to alleviate symptoms. This study was aimed to evaluate the dry eyes induced by manual Small Incision Cataract Surgery, and the effect if any, of oral lactoferrin on the dry eyes. A single centre, prospective randomised controlled trial with a concurrent parallel design. The study was carried out on patients presenting in the OPD of Rohilkhand Medical College hospital for cataract surgery. Sixty four patients of cataract surgery were included in the study. Patients with pre-existing dry eyes, ocular disease or systemic disease predisposing to dry eyes were excluded from the study. The selected patients were assigned into two groups by simple randomisation-Control Group A-32 patients that did not receive oral lactoferrin postoperatively. Group B-32 patients that received oral lactoferrin 350 gm postoperatively from day 1 after SICS. All patients were operated for cataract and their pre and postoperative (on days 7, 14, 30 and 60) dry eye status was assessed using the mean tear film break-up time (tBUT) and Schirmer test 1 (ST 1) as the evaluating parameters. Subjective evaluation of dry eye was done using Ocular Surface Disease Index (OSDI) scoring. Data was analysed for 58 patients, as 6 did not complete the follow up. Unpaired t-test was used to calculate the p-values. There was a statistically significant difference between the tBUT values of the Control and Lactoferrin group from day 14 onwards. The tBUT of control group
Glutathione S-transferase pi modulates NF-κB activation and pro-inflammatory responses in lung epithelial cells.

PubMed

Jones, Jane T; Qian, Xi; van der Velden, Jos L J; Chia, Shi Biao; McMillan, David H; Flemer, Stevenson; Hoffman, Sidra M; Lahue, Karolyn G; Schneider, Robert W; Nolin, James D; Anathy, Vikas; van der Vliet, Albert; Townsend, Danyelle M; Tew, Kenneth D; Janssen-Heininger, Yvonne M W

2016-08-01

Nuclear Factor kappa B (NF-κB) is a transcription factor family critical in the activation of pro- inflammatory responses. The NF-κB pathway is regulated by oxidant-induced post-translational modifications. Protein S-glutathionylation, or the conjugation of the antioxidant molecule, glutathione to reactive cysteines inhibits the activity of inhibitory kappa B kinase beta (IKKβ), among other NF-κB proteins. Glutathione S-transferase Pi (GSTP) is an enzyme that has been shown to catalyze protein S-glutathionylation (PSSG) under conditions of oxidative stress. The objective of the present study was to determine whether GSTP regulates NF-κB signaling, S-glutathionylation of IKK, and subsequent pro-inflammatory signaling. We demonstrated that, in unstimulated cells, GSTP associated with the inhibitor of NF-κB, IκBα. However, exposure to LPS resulted in a rapid loss of association between IκBα and GSTP, and instead led to a protracted association between IKKβ and GSTP. LPS exposure also led to increases in the S-glutathionylation of IKKβ. SiRNA-mediated knockdown of GSTP decreased IKKβ-SSG, and enhanced NF-κB nuclear translocation, transcriptional activity, and pro-inflammatory cytokine production in response to lipopolysaccharide (LPS). TLK117, an isotype-selective inhibitor of GSTP, also enhanced LPS-induced NF-κB transcriptional activity and pro-inflammatory cytokine production, suggesting that the catalytic activity of GSTP is important in repressing NF-κB activation. Expression of both wild-type and catalytically-inactive Y7F mutant GSTP significantly attenuated LPS- or IKKβ-induced production of GM-CSF. These studies indicate a complex role for GSTP in modulating NF-κB, which may involve S-glutathionylation of IKK proteins, and interaction with NF-κB family members. Our findings suggest that targeting GSTP is a potential avenue for regulating the activity of this prominent pro-inflammatory and immunomodulatory transcription factor. Copyright �
Encephalomyocarditis Virus 3C Protease Relieves TRAF Family Member-associated NF-κB Activator (TANK) Inhibitory Effect on TRAF6-mediated NF-κB Signaling through Cleavage of TANK.

PubMed

Huang, Li; Liu, Qinfang; Zhang, Lijie; Zhang, Quan; Hu, Liang; Li, Changyao; Wang, Shengnan; Li, Jiangnan; Zhang, Yuanfeng; Yu, Huibin; Wang, Yan; Zhong, Zhaohua; Xiong, Tao; Xia, Xueshan; Wang, Xiaojun; Yu, Li; Deng, Guohua; Cai, Xuehui; Cui, Shangjin; Weng, Changjiang

2015-11-13

TRAF family member-associated NF-κB activator (TANK) is a negative regulator of canonical NF-κB signaling in the Toll-like receptor- and B-cell receptor-mediated signaling pathways. However, functions of TANK in viral infection-mediated NF-κB activation remain unclear. Here, we reported that TANK was cleaved by encephalomyocarditis virus 3C at the 197 and 291 glutamine residues, which depends on its cysteine protease activity. In addition, encephalomyocarditis virus 3C impaired the ability of TANK to inhibit TRAF6-mediated NF-κB signaling. Interestingly, we found that several viral proteases encoded by the foot and mouth disease virus, porcine reproductive and respiratory syndrome virus, and equine arteritis virus also cleaved TANK. Our results suggest that TANK is a novel target of some viral proteases, indicating that some positive RNA viruses have evolved to utilize their major proteases to regulate NF-κB activation. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.
Encephalomyocarditis Virus 3C Protease Relieves TRAF Family Member-associated NF-κB Activator (TANK) Inhibitory Effect on TRAF6-mediated NF-κB Signaling through Cleavage of TANK*

PubMed Central

Huang, Li; Liu, Qinfang; Zhang, Lijie; Zhang, Quan; Hu, Liang; Li, Changyao; Wang, Shengnan; Li, Jiangnan; Zhang, Yuanfeng; Yu, Huibin; Wang, Yan; Zhong, Zhaohua; Xiong, Tao; Xia, Xueshan; Wang, Xiaojun; Yu, Li; Deng, Guohua; Cai, Xuehui; Cui, Shangjin; Weng, Changjiang

2015-01-01

TRAF family member-associated NF-κB activator (TANK) is a negative regulator of canonical NF-κB signaling in the Toll-like receptor- and B-cell receptor-mediated signaling pathways. However, functions of TANK in viral infection-mediated NF-κB activation remain unclear. Here, we reported that TANK was cleaved by encephalomyocarditis virus 3C at the 197 and 291 glutamine residues, which depends on its cysteine protease activity. In addition, encephalomyocarditis virus 3C impaired the ability of TANK to inhibit TRAF6-mediated NF-κB signaling. Interestingly, we found that several viral proteases encoded by the foot and mouth disease virus, porcine reproductive and respiratory syndrome virus, and equine arteritis virus also cleaved TANK. Our results suggest that TANK is a novel target of some viral proteases, indicating that some positive RNA viruses have evolved to utilize their major proteases to regulate NF-κB activation. PMID:26363073
Apoptotic death of Listeria monocytogenes-infected human macrophages induced by lactoferricin B, a bovine lactoferrin-derived peptide.

PubMed

Longhi, C; Conte, M P; Ranaldi, S; Penta, M; Valenti, P; Tinari, A; Superti, F; Seganti, L

2005-01-01

Listeria monocytogenes, an intracellular facultative food-borne pathogen, was reported to induce apoptosis in vitro and in vivo in a variety of cell types with the exception of murine macrophages. These cells represent the predominant compartment of bacterial multiplication and die as a result of necrosis. In this study we showed that human non-activated and IFN-gamma-activated macrophagic-like (THP-1) cells infected with L. monocytogenes, mainly die by necrosis rather than by an apoptotic process. Two natural products derived from bovine milk, lactoferrin and its derivative peptide lactoferricin B, are capable of regulating the fate of infected human macrophages. Bovine lactoferrin treatment of macrophages protects them from L. monocytogenes-induced death whereas lactoferricin B, its derivative peptide, determines a shifting of the equilibrium from necrosis to apoptosis.
Arsenic Promotes NF-Kb-Mediated Fibroblast Dysfunction and Matrix Remodeling to Impair Muscle Stem Cell Function

PubMed Central

Zhang, Changqing; Ferrari, Ricardo; Beezhold, Kevin; Stearns-Reider, Kristen; D’Amore, Antonio; Haschak, Martin; Stolz, Donna; Robbins, Paul D.; Barchowsky, Aaron; Ambrosio, Fabrisia

2016-01-01

Arsenic is a global health hazard that impacts over 140 million individuals worldwide. Epidemiological studies reveal prominent muscle dysfunction and mobility declines following arsenic exposure; yet, mechanisms underlying such declines are unknown. The objective of this study was to test the novel hypothesis that arsenic drives a maladaptive fibroblast phenotype to promote pathogenic myomatrix remodeling and compromise the muscle stem (satellite) cell (MuSC) niche. Mice were exposed to environmentally relevant levels of arsenic in drinking water before receiving a local muscle injury. Arsenic-exposed muscles displayed pathogenic matrix remodeling, defective myofiber regeneration and impaired functional recovery, relative to controls. When naïve human MuSCs were seeded onto three-dimensional decellularized muscle constructs derived from arsenic-exposed muscles, cells displayed an increased fibrogenic conversion and decreased myogenicity, compared with cells seeded onto control constructs. Consistent with myomatrix alterations, fibroblasts isolated from arsenic-exposed muscle displayed sustained expression of matrix remodeling genes, the majority of which were mediated by NF-κB. Inhibition of NF-κB during arsenic exposure preserved normal myofiber structure and functional recovery after injury, suggesting that NF-κB signaling serves as an important mechanism of action for the deleterious effects of arsenic on tissue healing. Taken together, the results from this study implicate myomatrix biophysical and/or biochemical characteristics as culprits in arsenic-induced MuSC dysfunction and impaired muscle regeneration. It is anticipated that these findings may aid in the development of strategies to prevent or revert the effects of arsenic on tissue healing and, more broadly, provide insight into the influence of the native myomatrix on stem cell behavior. PMID:26537186
Systemic Inhibition of NF-κB Activation Protects from Silicosis

PubMed Central

Di Giuseppe, Michelangelo; Gambelli, Federica; Hoyle, Gary W.; Lungarella, Giuseppe; Studer, Sean M.; Richards, Thomas; Yousem, Sam; McCurry, Ken; Dauber, James; Kaminski, Naftali; Leikauf, George; Ortiz, Luis A.

2009-01-01

Background Silicosis is a complex lung disease for which no successful treatment is available and therefore lung transplantation is a potential alternative. Tumor necrosis factor alpha (TNFα) plays a central role in the pathogenesis of silicosis. TNFα signaling is mediated by the transcription factor, Nuclear Factor (NF)-κB, which regulates genes controlling several physiological processes including the innate immune responses, cell death, and inflammation. Therefore, inhibition of NF-κB activation represents a potential therapeutic strategy for silicosis. Methods/Findings In the present work we evaluated the lung transplant database (May 1986–July 2007) at the University of Pittsburgh to study the efficacy of lung transplantation in patients with silicosis (n = 11). We contrasted the overall survival and rate of graft rejection in these patients to that of patients with idiopathic pulmonary fibrosis (IPF, n = 79) that was selected as a control group because survival benefit of lung transplantation has been identified for these patients. At the time of lung transplantation, we found the lungs of silica-exposed subjects to contain multiple foci of inflammatory cells and silicotic nodules with proximal TNFα expressing macrophage and NF-κB activation in epithelial cells. Patients with silicosis had poor survival (median survival 2.4 yr; confidence interval (CI): 0.16–7.88 yr) compared to IPF patients (5.3 yr; CI: 2.8–15 yr; p = 0.07), and experienced early rejection of their lung grafts (0.9 yr; CI: 0.22–0.9 yr) following lung transplantation (2.4 yr; CI:1.5–3.6 yr; p<0.05). Using a mouse experimental model in which the endotracheal instillation of silica reproduces the silica-induced lung injury observed in humans we found that systemic inhibition of NF-κB activation with a pharmacologic inhibitor (BAY 11-7085) of IκBα phosphorylation decreased silica-induced inflammation and collagen deposition. In contrast, transgenic mice expressing
Tumor regression following intravenous administration of lactoferrin- and lactoferricin-bearing dendriplexes

PubMed Central

Lim, Li Ying; Koh, Pei Yin; Somani, Sukrut; Al Robaian, Majed; Karim, Reatul; Yean, Yi Lyn; Mitchell, Jennifer; Tate, Rothwelle J.; Edrada-Ebel, RuAngelie; Blatchford, David R.; Mullin, Margaret; Dufès, Christine

2015-01-01

The possibility of using gene therapy for the treatment of cancer is limited by the lack of safe, intravenously administered delivery systems able to selectively deliver therapeutic genes to tumors. In this study, we investigated if the conjugation of the polypropylenimine dendrimer to lactoferrin and lactoferricin, whose receptors are overexpressed on cancer cells, could result in a selective gene delivery to tumors and a subsequently enhanced therapeutic efficacy. The conjugation of lactoferrin and lactoferricin to the dendrimer significantly increased the gene expression in the tumor while decreasing the non-specific gene expression in the liver. Consequently, the intravenous administration of the targeted dendriplexes encoding TNFα led to the complete suppression of 60% of A431 tumors and up to 50% of B16-F10 tumors over one month. The treatment was well tolerated by the animals. These results suggest that these novel lactoferrin- and lactoferricin-bearing dendrimers are promising gene delivery systems for cancer therapy. From the Clinical Editor Specific targeting of cancer cells should enhance the delivery of chemotherapeutic agents. This is especially true for gene delivery. In this article, the authors utilized a dendrimer-based system and conjugated this with lactoferrin and lactoferricin to deliver anti-tumor genes. The positive findings in animal studies should provide the basis for further clinical studies. PMID:25933695
Tumor regression following intravenous administration of lactoferrin- and lactoferricin-bearing dendriplexes.

PubMed

Lim, Li Ying; Koh, Pei Yin; Somani, Sukrut; Al Robaian, Majed; Karim, Reatul; Yean, Yi Lyn; Mitchell, Jennifer; Tate, Rothwelle J; Edrada-Ebel, RuAngelie; Blatchford, David R; Mullin, Margaret; Dufès, Christine

2015-08-01

The possibility of using gene therapy for the treatment of cancer is limited by the lack of safe, intravenously administered delivery systems able to selectively deliver therapeutic genes to tumors. In this study, we investigated if the conjugation of the polypropylenimine dendrimer to lactoferrin and lactoferricin, whose receptors are overexpressed on cancer cells, could result in a selective gene delivery to tumors and a subsequently enhanced therapeutic efficacy. The conjugation of lactoferrin and lactoferricin to the dendrimer significantly increased the gene expression in the tumor while decreasing the non-specific gene expression in the liver. Consequently, the intravenous administration of the targeted dendriplexes encoding TNFα led to the complete suppression of 60% of A431 tumors and up to 50% of B16-F10 tumors over one month. The treatment was well tolerated by the animals. These results suggest that these novel lactoferrin- and lactoferricin-bearing dendrimers are promising gene delivery systems for cancer therapy. Specific targeting of cancer cells should enhance the delivery of chemotherapeutic agents. This is especially true for gene delivery. In this article, the authors utilized a dendrimer-based system and conjugated this with lactoferrin and lactoferricin to deliver anti-tumor genes. The positive findings in animal studies should provide the basis for further clinical studies. Copyright © 2015 The Authors. Published by Elsevier Inc. All rights reserved.
HTLV-1 Tax-Induced Rapid Senescence Is Driven by the Transcriptional Activity of NF-κB and Depends on Chronically Activated IKKα and p65/RelA

PubMed Central

Ho, Yik-Khuan; Zhi, Huijun; DeBiaso, Dominic; Philip, Subha; Shih, Hsiu-Ming

2012-01-01

The HTLV-1 oncoprotein Tax is a potent activator of classical and alternative NF-κB pathways and is thought to promote cell proliferation and transformation via NF-κB activation. We showed recently that hyperactivation of NF-κB by Tax triggers a cellular senescence response (H. Zhi et al., PLoS Pathog. 7:e1002025, 2011). Inhibition of NF-κB activation by expression of I-κBα superrepressor or by small hairpin RNA (shRNA)-mediated knockdown of p65/RelA rescues cells from Tax-induced rapid senescence (Tax-IRS). Here we demonstrate that Tax-IRS is driven by the transcriptional activity of NF-κB. Knockdown of IKKγ, the primary Tax target, by shRNAs abrogated Tax-mediated activation of both classical and alternative NF-κB pathways and rendered knockdown cells resistant to Tax-IRS. Consistent with a critical role of IKKα in the transcriptional activity of NF-κB, IKKα deficiency drastically decreased NF-κB trans-activation by Tax, although it only modestly reduced Tax-mediated I-κBα degradation and NF-κB nuclear localization. In contrast, although IKKβ knockdown attenuated Tax-induced NF-κB transcriptional activation, the residual NF-κB activation in IKKβ-deficient cells was sufficient to trigger Tax-IRS. Importantly, the phenotypes of NIK and TAK1 knockdown were similar to those of IKKα and IKKβ knockdown, respectively. Finally, double knockdown of RelB and p100 had a minor effect on senescence induction by Tax. These data suggest that Tax, through its interaction with IKKγ, helps recruit NIK and TAK1 for IKKα and IKKβ activation, respectively. In the presence of Tax, the delineation between the classical and alternative NF-κB pathways becomes obscured. The senescence checkpoint triggered by Tax is driven by the transcriptional activity of NF-κB, which depends on activated IKKα and p65/RelA. PMID:22740410
The nuclear factor kappa B (NF-κB) activation is required for phagocytosis of staphylococcus aureus by RAW 264.7 cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhu, Fei, E-mail: zhufei@zju.edu.cn; Yue, Wanfu; Wang, Yongxia

Nuclear factor kappa B (NF-κB) is a ubiquitous transcription factor which controls the expression of various genes involved in immune responses. However, it is not clear whether NF-κB activation is critical for phagocytosis when Staphylococcus aureus is the pathogen. Using oligonucleotide microarrays, we investigated whether NF-κB cascade genes are altered in a mouse leukemic monocyte macrophage cell line (RAW 264.7) when the cells were stimulated to activate a host innate immune response against live S. aureus or heat-inactivated S. aureus (HISA). NF-κB cascade genes such as Nfκb1, Nfκbiz, Nfκbie, Rel, Traf1 and Tnfaip3 were up-regulated by all treatments at onemore » hour after incubation. NF-κB play an important role in activating phagocytosis in RAW 264.7 cells infected with S. aureus. Inhibition of NF-κB significantly blocked phagocytosis of fluorescently labeled S. aureus and decreased the expression of NFκB1, IL1α, IL1β and TLR2 in this cell line. Our results demonstrate that S. aureus may activate the NF-κB pathway and that NF-κB activation is required for phagocytosis of S. aureus by macrophages. - Highlights: • NF-κB cascade genes such as Nfκb1 and Traf1 were up-regulated by heat-inactivated S. aureus. • Inhibition of NF-κB significantly blocked phagocytosis of fluorescently labeled S. aureus. • NF-κB activation is required for phagocytosis of S. aureus by macrophages.« less

4-Hydroxy-17-methylincisterol from Agaricus blazei Decreased Cytokine Production and Cell Proliferation in Human Peripheral Blood Mononuclear Cells via Inhibition of NF-AT and NF-κB Activation

PubMed Central

Tsai, Wei-Jern; Yang, Shih-Chien; Huang, Yu-Ling; Chen, Chien-Chih; Chuang, Kai-An; Kuo, Yuh-Chi

2013-01-01

Agaricus blazei Murill is an edible and medicinal mushroom. In the previous study, we have proved that extracts of A. blazei inhibit human peripheral blood mononuclear cell (PBMC) proliferation activated with phytohemagglutinin (PHA). Currently, we purified 4-hydroxy-17-methylincisterol (4-HM; C21H33O3) from A. blazei investigated its regulatory effects on cytokine productions and cell proliferation of PBMC induced by PHA. The results indicated that 4-HM suppressed, in activated PBMC, the production and mRNA expression of interleukin-2 (IL-2), IL-4, tumor necrosis factor-α, and interferon-γ in a concentration-dependent manner. This inhibition was not related to cell viability. While 4-HM did not affect ERK phosphorylation and its downstream c-fos gene expression in PBMC induced by PHA, it decreased both NF-AT and NF-κB activation. The upstream signaling of NF-AT and NF-κB, intracellular calcium concentrations ([Ca2+]i), and protein kinase C theta (PKC θ) activation in PHA-treated PBMC were reduced by 4-HM. The data demonstrated that the suppressant effects of 4-HM on cell proliferation in PBMC activated by PHA appeared to be mediated, at least in part, through inhibition of Ca2+ mobilization and PKC θ activation, NF-AT and NF-κB activation, and cytokine transcripts and productions of PBMC. We suggested that A. blazei contained a potential immunomodulator 4-HM. PMID:23533483
RRM2 induces NF-{kappa}B-dependent MMP-9 activation and enhances cellular invasiveness

DOE Office of Scientific and Technical Information (OSTI.GOV)

Duxbury, Mark S.; Whang, Edward E.

2007-03-02

Ribonucleotide reductase is a dimeric enzyme that catalyzes conversion of ribonucleotide 5'-diphosphates to their 2'-deoxynucleotide forms, a rate-limiting step in the production of 2'-deoxyribonucleoside 5'-triphosphates required for DNA synthesis. The ribonucleotide reductase M2 subunit (RRM2) is a determinant of malignant cellular behavior in a range of human cancers. We examined the effect of RRM2 overexpression on pancreatic adenocarcinoma cellular invasiveness and nuclear factor-{kappa}B (NF-{kappa}B) transcription factor activity. RRM2 overexpression increases pancreatic adenocarcinoma cellular invasiveness and MMP-9 expression in a NF-{kappa}B-dependent manner. RNA interference (RNAi)-mediated silencing of RRM2 expression attenuates cellular invasiveness and NF-{kappa}B activity. NF-{kappa}B is a key mediator ofmore » the invasive phenotypic changes induced by RRM2 overexpression.« less
NF-κB in Hematological Malignancies

PubMed Central

Imbert, Véronique; Peyron, Jean-François

2017-01-01

NF-κB (Nuclear Factor Κ-light-chain-enhancer of activated B cells) transcription factors are critical regulators of immunity, stress response, apoptosis, and differentiation. Molecular defects promoting the constitutive activation of canonical and non-canonical NF-κB signaling pathways contribute to many diseases, including cancer, diabetes, chronic inflammation, and autoimmunity. In the present review, we focus our attention on the mechanisms of NF-κB deregulation in hematological malignancies. Key positive regulators of NF-κB signaling can act as oncogenes that are often prone to chromosomal translocation, amplifications, or activating mutations. Negative regulators of NF-κB have tumor suppressor functions, and are frequently inactivated either by genomic deletions or point mutations. NF-κB activation in tumoral cells is also driven by the microenvironment or chronic signaling that does not rely on genetic alterations. PMID:28561798
β-Arrestin2 functions as a phosphorylation-regulated suppressor of UV-induced NF-κB activation

PubMed Central

Luan, Bing; Zhang, Zhenning; Wu, Yalan; Kang, Jiuhong; Pei, Gang

2005-01-01

NF-κB activation is an important mechanism of mammalian UV response to protect cells. UV-induced NF-κB activation depends on the casein kinase II (CK2) phosphorylation of IκBα at a cluster of C-terminal sites, but how it is regulated remains unclear. Here we demonstrate that β-arrestin2 can function as an effective suppressor of UV-induced NF-κB activation through its direct interaction with IκBα. CK2 phosphorylation of β-arrestin2 blocks its interaction with IκBα and abolishes its suppression of NF-κB activation, indicating that the β-arrestin2 phosphorylation is critical. Moreover, stimulation of β2-adrenergic receptors, a representative of G-protein-coupled receptors in epidermal cells, promotes dephosphorylation of β-arrestin2 and its suppression of NF-κB activation. Consequently, the β-arrestin2 suppression leads to promotion of UV-induced cell death, which is also under regulation of β-arrestin2 phosphorylation. Thus, β-arrestin2 is identified as a phosphorylation-regulated suppressor of UV response and this may play a functional role in the response of epidermal cells to UV. PMID:16308565
Glycyrrhetinic acid suppressed NF-κB activation in TNF-α-induced hepatocytes.

PubMed

Chen, Hong-Jhang; Kang, Shih-Pei; Lee, I-Jung; Lin, Yun-Lian

2014-01-22

Tumor necrosis factor-alpha (TNF-α) is a crucial inflammatory cytokine when hepatocytes are damaged. Glycyrrhiza uralensis Fisch. (Chinese licorice) has been widely used in Chinese herbal prescriptions for the treatment of liver diseases and as a food additive. Nuclear factor-kappa B (NF-κB) reporter gene assay in TNF-α-induced HepG2 was used as a screening platform. IκBα phosphorylation and p65 translocation were measured by Western blotting, and nitric oxide (NO) production and inducible nitric oxide synthase (iNOS) gene expression were further confirmed in rat primary hepatocytes. Results showed that TNF-α enhanced NF-κB activity was significantly attenuated by glycyrrhetinic acid in a concentration-dependent manner in the NF-κB reporter gene assay. Glycyrrhetinic acid decreased the gene expression of iNOS through inhibited IκBα phosphorylation and p65 translocation in protein level. Furthermore, NO production and iNOS expression were reduced by glycyrrhetinic acid in TNF-α-induced rat primary hepatocytes. These results suggest that glycyrrhetinic acid may provide hepatoprotection against chronic liver inflammation through attenuating NF-κB activation to alleviate the inflammation.
TAK1 regulates NF-{Kappa}B and AP-1 activation in airway epithelial cells following RSV infection

DOE Office of Scientific and Technical Information (OSTI.GOV)

Dey, Nilay; Liu Tianshuang; Garofalo, Roberto P.

2011-09-30

Respiratory syncytial virus (RSV) is the most common cause of epidemic respiratory diseases in infants and young children. RSV infection of airway epithelial cells induces the expression of immune/inflammatory genes through the activation of a subset of transcription factors, including Nuclear Factor-{kappa}B (NF-{kappa}B) and AP-1. In this study, we have investigated the signaling pathway leading to activation of these two transcription factors in response to RSV infection. Our results show that IKK{beta} plays a key role in viral-induced NF-{kappa}B activation, while JNK regulates AP-1-dependent gene transcription, as demonstrated by using kinase inactive proteins and chemical inhibitors of the two kinases.more » Inhibition of TAK1 activation, by overexpression of kinase inactive TAK1 or using cells lacking TAK1 expression, significantly reduced RSV-induced NF-{kappa}B and AP-1 nuclear translocation and DNA-binding activity, as well as NF-{kappa}B-dependent gene expression, identifying TAK1 as an important upstream signaling molecule regulating RSV-induced NF-{kappa}B and AP-1 activation. - Highlights: > IKK{beta} is a major kinase involved in RSV-induced NF-{kappa}B activation. > JNK regulates AP-1-dependent gene transcription in RSV infection. > TAK1 is a critical upstream signaling molecule for both pathways in infected cells.« less
The structure of lactoferrin-binding protein B from Neisseria meningitidis suggests roles in iron acquisition and neutralization of host defences

PubMed Central

Brooks, Cory L.; Arutyunova, Elena; Lemieux, M. Joanne

2014-01-01

Pathogens have evolved a range of mechanisms to acquire iron from the host during infection. Several Gram-negative pathogens including members of the genera Neisseria and Moraxella have evolved two-component systems that can extract iron from the host glycoproteins lactoferrin and transferrin. The homologous iron-transport systems consist of a membrane-bound transporter and an accessory lipoprotein. While the mechanism behind iron acquisition from transferrin is well understood, relatively little is known regarding how iron is extracted from lactoferrin. Here, the crystal structure of the N-terminal domain (N-lobe) of the accessory lipoprotein lactoferrin-binding protein B (LbpB) from the pathogen Neisseria meningitidis is reported. The structure is highly homologous to the previously determined structures of the accessory lipoprotein transferrin-binding protein B (TbpB) and LbpB from the bovine pathogen Moraxella bovis. Docking the LbpB structure with lactoferrin reveals extensive binding interactions with the N1 subdomain of lactoferrin. The nature of the interaction precludes apolactoferrin from binding LbpB, ensuring the specificity of iron-loaded lactoferrin. The specificity of LbpB safeguards proper delivery of iron-bound lactoferrin to the transporter lactoferrin-binding protein A (LbpA). The structure also reveals a possible secondary role for LbpB in protecting the bacteria from host defences. Following proteolytic digestion of lactoferrin, a cationic peptide derived from the N-terminus is released. This peptide, called lactoferricin, exhibits potent antimicrobial effects. The docked model of LbpB with lactoferrin reveals that LbpB interacts extensively with the N-terminal lactoferricin region. This may provide a venue for preventing the production of the peptide by proteolysis, or directly sequestering the peptide, protecting the bacteria from the toxic effects of lactoferricin. PMID:25286931
Vitamin E δ-Tocotrienol Augments the Anti-tumor Activity of Gemcitabine and Suppresses Constitutive NF-κB Activation in Pancreatic Cancer

PubMed Central

Husain, Kazim; Francois, Rony A.; Yamauchi, Teruo; Perez, Marta; Sebti, Said M.; Malafa, Mokenge P.

2011-01-01

The nuclear factor-κB (NF-κB) transcription factor functions as a crucial regulator of cell survival and chemoresistance in pancreatic cancer. Recent studies suggest that tocotrienols, which are the unsaturated forms of vitamin E, are a promising class of anti-cancer compounds that inhibit the growth and survival of many cancer cells, including pancreatic cancer. Here, we show that tocotrienols inhibited NF-κB activity and the survival of human pancreatic cancer cells in vitro and in vivo. Importantly, we found the bioactivity of the 4 natural tocotrienol compounds (α-, β-, δ-, and γ-tocotrienol) to be directly related to their ability to suppress NF-κB activity in vitro and in vivo. The most bioactive tocotrienol for pancreatic cancer, δ-tocotrienol, significantly enhanced the efficacy of gemcitabine to inhibit pancreatic cancer growth and survival in vitro and in vivo. Moreover, we found that δ-tocotrienol augmentation of gemcitabine activity in pancreatic cancer cells and tumors is associated with significant suppression of NF-κB activity and the expression of NF-κB transcriptional targets [Bcl-XL, X-linked inhibitor of apoptosis (XIAP), and survivin]. Our study represents the first comprehensive pre-clinical evaluation of the activity of natural vitamin E compounds in pancreatic cancer. Given these results, we are conducting a phase I trial of δ-tocotrienol in patients with pancreatic cancer utilizing pancreatic tumor cell survival and NF-κB signaling components as intermediate biomarkers. Our data also support future clinical investigation of δ-tocotrienol to augment gemcitabine activity in pancreatic cancer. PMID:21971120
Gastroesophageal reflux activates the NF-κB pathway and impairs esophageal barrier function in mice

PubMed Central

Fang, Yu; Chen, Hao; Hu, Yuhui; Djukic, Zorka; Tevebaugh, Whitney; Shaheen, Nicholas J.; Orlando, Roy C.; Hu, Jianguo

2013-01-01

The barrier function of the esophageal epithelium is a major defense against gastroesophageal reflux disease. Previous studies have shown that reflux damage is reflected in a decrease in transepithelial electrical resistance associated with tight junction alterations in the esophageal epithelium. To develop novel therapies, it is critical to understand the molecular mechanisms whereby contact with a refluxate impairs esophageal barrier function. In this study, surgical models of duodenal and mixed reflux were developed in mice. Mouse esophageal epithelium was analyzed by gene microarray. Gene set enrichment analysis showed upregulation of inflammation-related gene sets and the NF-κB pathway due to reflux. Significance analysis of microarrays revealed upregulation of NF-κB target genes. Overexpression of NF-κB subunits (p50 and p65) and NF-κB target genes (matrix metalloproteinases-3 and -9, IL-1β, IL-6, and IL-8) confirmed activation of the NF-κB pathway in the esophageal epithelium. In addition, real-time PCR, Western blotting, and immunohistochemical staining also showed downregulation and mislocalization of claudins-1 and -4. In a second animal experiment, treatment with an NF-κB inhibitor, BAY 11-7085 (20 mg·kg−1·day−1 ip for 10 days), counteracted the effects of duodenal and mixed reflux on epithelial resistance and NF-κB-regulated cytokines. We conclude that gastroesophageal reflux activates the NF-κB pathway and impairs esophageal barrier function in mice and that targeting the NF-κB pathway may strengthen esophageal barrier function against reflux. PMID:23639809
Faecal lactoferrin and calprotectin in patients with Clostridium difficile infection: a case-control study.

PubMed

Barbut, F; Gouot, C; Lapidus, N; Suzon, L; Syed-Zaidi, R; Lalande, V; Eckert, C

2017-12-01

Calprotectin and lactoferrin are released by the gastrointestinal tract in response to infection and mucosal inflammation. Our objective was to assess the usefulness of quantifying faecal lactoferrin and calprotectin concentrations in Clostridium difficile infection (CDI) patients with or without free toxins in the stools. We conducted a single-centre 22-month case-control study. Patients with a positive CDI diagnosis were compared to two control groups: group 1 = diarrhoeic patients negative for C. difficile and matched (1:1) to CDI cases on the ward location and age, and group 2 = diarrhoeic patients colonised with a non-toxigenic strain of C. difficile. Faecal lactoferrin and calprotectin concentrations in faeces were determined for patients with CDI and controls. Of 135 patients with CDI, 87 (64.4%) had a positive stool cytotoxicity assay (free toxin) and 48 (35.6%) had a positive toxigenic culture without detectable toxins in the stools. The median lactoferrin values were 26.8 μg/g, 8.0 μg/g and 15.8 μg/g in CDI patients and groups 1 and 2, respectively. The median calprotectin values were 218.0 μg/g, 111.5 μg/g and 111.3 μg/g, respectively. Among patients with CDI, faecal lactoferrin and calprotectin levels were higher in those with free toxins in their stools (39.2 vs. 10.2 μg/g, p = 0.003 and 274.0 vs. 166.0 μg/g, p = 0.051, respectively). Both faecal calprotectin and lactoferrin were higher in patients with CDI, especially in those with detectable toxin in faeces, suggesting a correlation between intestinal inflammation and toxins in stools.
Systemic activation of NF-κB driven luciferase activity in transgenic mice fed advanced glycation end products modified albumin.

PubMed

Nass, Norbert; Bayreuther, Kristina; Simm, Andreas

2017-04-01

Advanced glycation end products (AGEs) are stable end products of the Maillard reaction and accumulate with progressing ageing and degenerative diseases. Significant amounts of AGE-modified peptides are also consumed with processed food. AGEs bind to specific receptors, especially the receptor of AGEs (RAGE). Activation of RAGE then evokes intracellular signalling, finally resulting in the activation of the NF-κB transcription factor and therefore a proinflammatory state. We here analysed, whether NF-κB is activated in short term upon feeding an AGE-modified protein in-vivo. Transgenic mice expressing firefly luciferase under the control of an NF-κB responsive promoter were intraperitoneally injected or fed with AGE-modified- or control albumin and luciferase expression was analysed by in-vivo imaging and by in-vitro by determination of luciferase enzyme activity in heart, lung, gut, spleen, liver and kidney. In all organs, an activation of the luciferase reporter gene was observed in response to AGE-BSA feeding, however with different intensity and timing. The gut exhibited highest luciferase activity and this activity peaked 6-8 h post AGE-feeding. In heart and kidney, luciferase activity increased for up to 12 h post feeding. All other organs tested, exhibited highest activity at 10 h after AGE-consumption. Altogether, these data demonstrate that feeding AGE-modified protein resulted in a transient and systemic activation of the NF-κB reporter.
Lactoferrin and necrotizing enterocolitis.

PubMed

Sherman, Michael P

2013-03-01

Lactoferrin (LF) is a multifunctional protein and a member of the transferrin family. LF and lysozyme in breast milk kill bacteria. In the stomach, pepsin digests and releases a potent peptide antibiotic called lactoferricin from native LF. The antimicrobial characteristics of LF may facilitate a healthy intestinal microbiome. LF is the major whey in human milk; its highest concentration is in colostrum. This fact highlights early feeding of colostrum and also fresh mature milk as a way to prevent necrotizing enterocolitis. Copyright © 2013 Elsevier Inc. All rights reserved.
HTLV-I Tax protein binds to MEKK1 to stimulate IkappaB kinase activity and NF-kappaB activation.

PubMed

Yin, M J; Christerson, L B; Yamamoto, Y; Kwak, Y T; Xu, S; Mercurio, F; Barbosa, M; Cobb, M H; Gaynor, R B

1998-05-29

NF-kappaB, a key regulator of the cellular inflammatory and immune response, is activated by the HTLV-I transforming and transactivating protein Tax. We show that Tax binds to the amino terminus of the protein kinase MEKK1, a component of an IkappaB kinase complex, and stimulates MEKK1 kinase activity. Tax expression increases the activity of IkappaB kinase beta (IKKbeta) to enhance phosphorylation of serine residues in IkappaB alpha that lead to its degradation. Dominant negative mutants of both IKKbeta and MEKK1 prevent Tax activation of the NF-kappaB pathway. Furthermore, recombinant MEKK1 stimulates IKKbeta phosphorylation of IkappaB alpha. Thus, Tax-mediated increases in NF-kappaB nuclear translocation result from direct interactions of Tax and MEKK1 leading to enhanced IKKbeta phosphorylation of IkappaB alpha.
Prevalence of mutations in hepatitis C virus core protein associated with alteration of NF-kappaB activation.

PubMed

Mann, Elizabeth A; Stanford, Sandra; Sherman, Kenneth E

2006-10-01

The hepatitis C virus (HCV) core protein is a key structural element of the virion but also affects a number of cellular pathways, including nuclear factor kappaB (NF-kappaB) signaling. NF-kappaB is a transcription factor that regulates both anti-apoptotic and pro-inflammatory genes and its activation may contribute to HCV-mediated pathogenesis. Amino acid sequence divergence in core is seen at the genotype level as well as within patient isolates. Recent work has implicated amino acids 9-11 of core in the modulation of NF-kappaB activation. We report that the sequence RKT is highly conserved (93%) at this position across all HCV genotypes, based on sequences collected in the Los Alamos HCV database. Of the 13 types of variants present in the database, the two most prevalent substitutions are RQT and RKP. We further show that core encoding RKP fails to activate NF-kappaB signaling in vitro while NF-kappaB activation by core encoding RQT does not differ from control RKT core. The effect of RKP core is specific to NF-kappaB signaling as activator protein 1 (AP-1) activity is not altered. Further studies are needed to assess potential associations between specific amino acid substitutions at positions 9-11 and liver disease progression and/or response to treatment in individual patients.
Reconstitution of the NF1 GAP-related domain in NF1-deficient human Schwann cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Thomas, Stacey L.; Neuroscience Program, Loyola University Medical Center, Maywood, IL; Department of Anatomy and Cell Biology, University of Illinois Chicago, Chicago, IL

Schwann cells derived from peripheral nerve sheath tumors from individuals with Neurofibromatosis Type 1 (NF1) are deficient for the protein neurofibromin, which contains a GAP-related domain (NF1-GRD). Neurofibromin-deficient Schwann cells have increased Ras activation, increased proliferation in response to certain growth stimuli, increased angiogenic potential, and altered cell morphology. This study examined whether expression of functional NF1-GRD can reverse the transformed phenotype of neurofibromin-deficient Schwann cells from both benign and malignant peripheral nerve sheath tumors. We reconstituted the NF1-GRD using retroviral transduction and examined the effects on cell morphology, growth potential, and angiogenic potential. NF1-GRD reconstitution resulted in morphologic changes,more » a 16-33% reduction in Ras activation, and a 53% decrease in proliferation in neurofibromin-deficient Schwann cells. However, NF1-GRD reconstitution was not sufficient to decrease the in vitro angiogenic potential of the cells. This study demonstrates that reconstitution of the NF1-GRD can at least partially reverse the transformation of human NF1 tumor-derived Schwann cells.« less
ANKRD1 modulates inflammatory responses in C2C12 myoblasts through feedback inhibition of NF-κB signaling activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Xin-Hua; Department of Medicine, Icahn School of Medicine at Mount Sinai, New York, NY 10029; Bauman, William A.

2015-08-14

Transcription factors of the nuclear factor-kappa B (NF-κB) family play a pivotal role in inflammation, immunity and cell survival responses. Recent studies revealed that NF-κB also regulates the processes of muscle atrophy. NF-κB activity is regulated by various factors, including ankyrin repeat domain 2 (AnkrD2), which belongs to the muscle ankyrin repeat protein family. Another member of this family, AnkrD1 is also a transcriptional effector. The expression levels of AnkrD1 are highly upregulated in denervated skeletal muscle, suggesting an involvement of AnkrD1 in NF-κB mediated cellular responses to paralysis. However, the molecular mechanism underlying the interactive role of AnkrD1 inmore » NF-κB mediated cellular responses is not well understood. In the current study, we examined the effect of AnkrD1 on NF-κB activity and determined the interactions between AnkrD1 expression and NF-κB signaling induced by TNFα in differentiating C2C12 myoblasts. TNFα upregulated AnkrD1 mRNA and protein levels. AnkrD1-siRNA significantly increased TNFα-induced transcriptional activation of NF-κB, whereas overexpression of AnkrD1 inhibited TNFα-induced NF-κB activity. Co-immunoprecipitation studies demonstrated that AnkrD1 was able to bind p50 subunit of NF-κB and vice versa. Finally, CHIP assays revealed that AnkrD1 bound chromatin at a NF-κB binding site in the AnrkD2 promoter and required NF-κB to do so. These results provide evidence of signaling integration between AnkrD1 and NF-κB pathways, and suggest a novel anti-inflammatory role of AnkrD1 through feedback inhibition of NF-κB transcriptional activity by which AnkrD1 modulates the balance between physiological and pathological inflammatory responses in skeletal muscle. - Highlights: • AnkrD1 is upregulated by TNFα and represses NF-κB-induced transcriptional activity. • AnkrD1 binds to p50 subunit of NF-κB and is recruited to NF-κB bound to chromatin. • AnkrD1 mediates a feed
NF-κB dynamics show digital activation and analog information processing in cells

NASA Astrophysics Data System (ADS)

Tay, Savas; Hughey, Jake; Lee, Timothy; Lipniacki, Tomasz; Covert, Markus; Quake, Stephen

2010-03-01

Cells operate in ever changing environments using extraordinary communication capabilities. Cell-to-cell communication is mediated by signaling molecules that form spatiotemporal concentration gradients, which requires cells to respond to a wide range of signal intensities. We used high-throughput microfluidic cell culture, quantitative gene expression analysis and mathematical modeling to investigate how single mammalian cells respond to different concentrations of the signaling molecule TNF-α via the transcription factor NF-κB. We measured NF-κB activity in thousands of live cells under TNF-α doses covering four orders of magnitude. In contrast to population studies, the activation is a stochastic, switch-like process at the single cell level with fewer cells responding at lower doses. The activated cells respond fully and express early genes independent of the TNF-α concentration, while only high dose stimulation results in the expression of late genes. Cells also encode a set of analog parameters such as the NF-κB peak intensity, response time and number of oscillations to modulate the outcome. We developed a stochastic model that reproduces both the digital and analog dynamics as well as the gene expression profiles at all measured conditions, constituting a broadly applicable model for TNF-α induced NF-κB signaling in various types of cells.
Spatio-temporal modelling of the NF-κB intracellular signalling pathway: the roles of diffusion, active transport, and cell geometry.

PubMed

Terry, Alan J; Chaplain, Mark A J

2011-12-07

The nuclear factor kappa B (NF-κB) intracellular signalling pathway is central to many stressful, inflammatory, and innate immune responses. NF-κB proteins themselves are transcription factors for hundreds of genes. Experiments have shown that the NF-κB pathway can exhibit oscillatory dynamics-a negative feedback loop causes oscillatory nuclear-cytoplasmic translocation of NF-κB. Given that cell size and shape are known to influence intracellular signal transduction, we consider a spatio-temporal model of partial differential equations for the NF-κB pathway, where we model molecular movement by diffusion and, for several key species including NF-κB, by active transport as well. Through numerical simulations we find values for model parameters such that sustained oscillatory dynamics occur. Our spatial profiles and animations bear a striking resemblance to experimental images and movie clips employing fluorescent fusion proteins. We discover that oscillations in nuclear NF-κB may occur when active transport is across the nuclear membrane only, or when no species are subject to active transport. However, when active transport is across the nuclear membrane and NF-κB is additionally actively transported through the cytoplasm, oscillations are lost. Hence transport mechanisms in a cell will influence its response to activation of its NF-κB pathway. We also demonstrate that sustained oscillations in nuclear NF-κB are somewhat robust to changes in the shape of the cell, or the shape, location, and size of its nucleus, or the location of ribosomes. Yet if the cell is particularly flat or the nucleus sufficiently small, then oscillations are lost. Thus the geometry of a cell may partly determine its response to NF-κB activation. The NF-κB pathway is known to be constitutively active in several human cancers. Our spatially explicit modelling approach will allow us, in future work, to investigate targeted drug therapy of tumours. Copyright © 2011 Elsevier Ltd
Oridonin stabilizes retinoic acid receptor alpha through ROS-activated NF-κB signaling.

PubMed

Cao, Yang; Wei, Wei; Zhang, Nan; Yu, Qing; Xu, Wen-Bin; Yu, Wen-Jun; Chen, Guo-Qiang; Wu, Ying-Li; Yan, Hua

2015-04-10

Retinoic acid receptor alpha (RARα) plays an essential role in the regulation of many biological processes, such as hematopoietic cell differentiation, while abnormal RARα function contributes to the pathogenesis of certain diseases including cancers, especially acute promyelocytic leukemia (APL). Recently, oridonin, a natural diterpenoid isolated from Rabdosia rubescens, was demonstrated to regulate RARα by increasing its protein level. However, the underlying molecular mechanism for this action has not been fully elucidated. In the APL cell line, NB4, the effect of oridonin on RARα protein was analyzed by western blot and real-time quantitative RT-PCR analyses. Flow cytometry was performed to detect intracellular levels of reactive oxygen species (ROS). The association between nuclear factor-kappa B (NF-κB) signaling and the effect of oridonin was assessed using specific inhibitors, shRNA gene knockdown, and immunofluorescence assays. In addition, primary leukemia cells were treated with oridonin and analyzed by western blot in this study. RARα possesses transcriptional activity in the presence of its ligand, all-trans retinoic acid (ATRA). Oridonin remarkably stabilized the RARα protein, which retained transcriptional activity. Oridonin also moderately increased intracellular ROS levels, while pretreatment with the ROS scavenger, N-acetyl-l-cysteine (NAC), dramatically abrogated RARα stabilization by oridonin. More intriguingly, direct exposure to low concentrations of H2O2 also increased RARα protein but not mRNA levels, suggesting a role for ROS in oridonin stabilization of RARα protein. Further investigations showed that NAC antagonized oridonin-induced activation of NF-κB signaling, while the NF-κB signaling inhibitor, Bay 11-7082, effectively blocked the oridonin increase in RARα protein levels. In line with this, over-expression of IκΒα (A32/36), a super-repressor form of IκΒα, or NF-κB-p65 knockdown inhibited oridonin or H2O2-induced
Methylation-Dependent Activation of CDX1 through NF-κB

PubMed Central

Rau, Tilman T.; Rogler, Anja; Frischauf, Myrjam; Jung, Andreas; Konturek, Peter C.; Dimmler, Arno; Faller, Gerhard; Sehnert, Bettina; El-Rifai, Wael; Hartmann, Arndt; Voll, Reinhard E.; Schneider-Stock, Regine

2013-01-01

The caudal homeobox factor 1 (CDX1) is an essential transcription factor for intestinal differentiation. Its aberrant expression in intestinal metaplasia of the upper gastrointestinal tract is a hallmark within the gastritis-metaplasia-carcinoma sequence. CDX1 expression is influenced by certain pathways, such as Wnt, Ras, or NF-κB signaling; however, these pathways alone cannot explain the transient expression of CDX1 in intestinal metaplasia or the molecular inactivation mechanism of its loss in cases of advanced gastric cancer. In this study, we investigated the epigenetic inactivation of CDX1 by promoter methylation, as well as the functional link of CDX1 promoter methylation to the inflammatory NF-κB signaling pathway. We identified methylation-dependent NF-κB binding to the CDX1 promoter and quantified it using competitive electrophoretic mobility shift assays and chromatin immunoprecipitation. A methylated CDX1 promoter was associated with closed chromatin structure, reduced NF-κB binding, and transcriptional silencing. Along the gastritis-metaplasia-carcinoma sequence, we observed a biphasic pattern of tumor necrosis factor-α (TNF-α) protein expression and an inverse biphasic pattern of CDX1 promoter methylation; both are highly consistent with CDX1 protein expression. The stages of hyper-, hypo-, and hyper-methylation patterns of the CDX1 promoter were inversely correlated with the NF-κB signaling activity along this sequence. In conclusion, these functionally interacting events drive CDX1 expression and contribute to intestinal metaplasia, epithelial dedifferentiation, and carcinogenesis in the human stomach. PMID:22749770

NF-κB deregulation in Hodgkin lymphoma.

PubMed

Weniger, Marc A; Küppers, Ralf

2016-08-01

Hodgkin and Reed/Sternberg (HRS) cells in classical Hodgkin lymphoma (HL) show constitutive activity of both the canonical and non-canonical NF-κB signaling pathways. The central pathogenetic role of this activity is indicated from studies with HL cell lines, which undergo apoptosis upon NF-κB inhibition. Multiple factors contribute to the strong NF-κB activity of HRS cells. This includes interaction with other cells in the lymphoma microenvironment through CD30, CD40, BCMA and other receptors, but also recurrent somatic genetic lesions in various factors of the NF-κB pathway, including destructive mutations in negative regulators of NF-κB signaling (e.g. TNFAIP3, NFKBIA), and copy number gains of genes encoding positive regulators (e.g. REL, MAP3K14). In Epstein-Barr virus-positive cases of classical HL, the virus-encoded latent membrane protein 1 causes NF-κB activation by mimicking an active CD40 receptor. NF-κB activity is also seen in the tumor cells of the rare nodular lymphocyte predominant form of HL, but the causes for this activity are largely unclear. Copyright © 2016 Elsevier Ltd. All rights reserved.
MicroRNA-146 inhibits thrombin-induced NF-κB activation and subsequent inflammatory responses in human retinal endothelial cells.

PubMed

Cowan, Colleen; Muraleedharan, Chithra K; O'Donnell, James J; Singh, Pawan K; Lum, Hazel; Kumar, Ashok; Xu, Shunbin

2014-07-01

Nuclear factor-κB (NF-κB), a key regulator of immune and inflammatory responses, plays important roles in diabetes-induced microvascular complications including diabetic retinopathy (DR). Thrombin activates NF-κB through protease-activated receptor (PAR)-1, a member of the G-protein-coupled receptor (GPCR) superfamily, and contributes to DR. The current study is to uncover the roles of microRNA (miRNA) in thrombin-induced NF-κB activation and retinal endothelial functions. Target prediction was performed using the TargetScan algorithm. Predicted target was experimentally validated by luciferase reporter assays. Human retinal endothelial cells (HRECs) were transfected with miRNA mimics or antimiRs and treated with thrombin. Expression levels of miR-146 and related protein-coding genes were analyzed by quantitative (q)RT-PCR. Functional changes of HRECs were analyzed by leukocyte adhesion assays. We identified that caspase-recruitment domain (CARD)-containing protein 10 (CARD10), an essential scaffold/adaptor protein of GPCR-mediated NF-κB activation pathway, is a direct target of miR-146. Thrombin treatment resulted in NF-κB-dependent upregulation of miR-146 in HRECs; while transfection of miR-146 mimics resulted in significant downregulation of CARD10 and prevented thrombin-induced NF-κB activation, suggest that a negative feedback regulation of miR-146 on thrombin-induced NF-κB through targeting CARD10. Furthermore, overexpression of miR-146 prevented thrombin-induced increased leukocyte adhesion to HRECs. We uncovered a novel negative feedback regulatory mechanism on thrombin-induced GPCR-mediated NF-κB activation by miR-146. In combination with the negative feedback regulation of miR-146 on the IL-1R/toll-like receptor (TLR)-mediated NF-κB activation in RECs that we reported previously, our results underscore a pivotal, negative regulatory role of miR-146 on multiple NF-κB activation pathways and related inflammatory processes in DR. Copyright 2014
Inhibition of spontaneous activity of rabbit atrioventricular node cells by KB-R7943 and inhibitors of sarcoplasmic reticulum Ca2+ ATPase

PubMed Central

Cheng, Hongwei; Smith, Godfrey L.; Hancox, Jules C.; Orchard, Clive H.

2011-01-01

The atrioventricular node (AVN) can act as a subsidiary cardiac pacemaker if the sinoatrial node fails. In this study, we investigated the effects of the Na–Ca exchange (NCX) inhibitor KB-R7943, and inhibition of the sarcoplasmic reticulum calcium ATPase (SERCA), using thapsigargin or cyclopiazonic acid (CPA), on spontaneous action potentials (APs) and [Ca2+]i transients from cells isolated from the rabbit AVN. Spontaneous [Ca2+]i transients were monitored from undialysed AVN cells at 37 °C using Fluo-4. In separate experiments, spontaneous APs and ionic currents were recorded using the whole-cell patch clamp technique. Rapid application of 5 μM KB-R7943 slowed or stopped spontaneous APs and [Ca2+]i transients. However, in voltage clamp experiments in addition to blocking NCX current (INCX) KB-R7943 partially inhibited L-type calcium current (ICa,L). Rapid reduction of external [Na+] also abolished spontaneous activity. Inhibition of SERCA (using 2.5 μM thapsigargin or 30 μM CPA) also slowed or stopped spontaneous APs and [Ca2+]i transients. Our findings are consistent with the hypothesis that sarcoplasmic reticulum (SR) Ca2+ release influences spontaneous activity in AVN cells, and that this occurs via [Ca2+]i-activated INCX; however, the inhibitory action of KB-R7943 on ICa,L means that care is required in the interpretation of data obtained using this compound. PMID:21163524
Identification of milk proteins enhancing the antimicrobial activity of lactoferrin and lactoferricin.

PubMed

Murata, M; Wakabayashi, H; Yamauchi, K; Abe, F

2013-08-01

Lactoferrin (LF) is known as an iron-binding antimicrobial protein present in exocrine secretions such as milk and releases the potent antimicrobial peptide lactoferricin (LFcin) by hydrolysis with pepsin. The antimicrobial activity of LF and LFcin has been studied well; however, their cooperative action with other milk proteins remains to be elucidated. In this study, we identified milk proteins enhancing the antimicrobial activity of bovine LF and LFcin against gram-negative bacteria, gram-positive bacteria, and fungi. As the target fraction, we isolated a minor milk protein fraction around 15 kDa, which was identified as bovine RNase 5 (angiogenin-1), RNase 4, and angiogenin-2 by matrix-assisted laser desorption/ionization-time-of-flight mass spectrometry. As these proteins are collectively known as the RNase A family, we referred to the target protein fraction as milk RNase of 15 kDa (MR15). The number of colony-forming units of Escherichia coli and other pathogenic microorganisms with the addition of MR15 to LF (MR15:LF ratio=16:1,000) was dramatically lowered than that with LF alone. On the other hand, MR15 itself did not show any reductions in the number of colony-forming units at the concentrations tested. Similarly, the antimicrobial activities of LFcin against various microorganisms were significantly enhanced by the addition of MR15. These results suggest that LF and MR15 may be concomitantly acting antimicrobial agents in milk. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
NF-κB Transcriptional Activity Is Modulated by FK506-binding Proteins FKBP51 and FKBP52

PubMed Central

Erlejman, Alejandra G.; De Leo, Sonia A.; Mazaira, Gisela I.; Molinari, Alejandro M.; Camisay, María Fernanda; Fontana, Vanina; Cox, Marc B.; Piwien-Pilipuk, Graciela; Galigniana, Mario D.

2014-01-01

Hsp90 binding immunophilins FKBP51 and FKBP52 modulate steroid receptor trafficking and hormone-dependent biological responses. With the purpose to expand this model to other nuclear factors that are also subject to nuclear-cytoplasmic shuttling, we analyzed whether these immunophilins modulate NF-κB signaling. It is demonstrated that FKBP51 impairs both the nuclear translocation rate of NF-κB and its transcriptional activity. The inhibitory action of FKBP51 requires neither the peptidylprolyl-isomerase activity of the immunophilin nor its association with Hsp90. The TPR domain of FKBP51 is essential. On the other hand, FKBP52 favors the nuclear retention time of RelA, its association to a DNA consensus binding sequence, and NF-κB transcriptional activity, the latter effect being strongly dependent on the peptidylprolyl-isomerase activity and also on the TPR domain of FKBP52, but its interaction with Hsp90 is not required. In unstimulated cells, FKBP51 forms endogenous complexes with cytoplasmic RelA. Upon cell stimulation with phorbol ester, the NF-κB soluble complex exchanges FKBP51 for FKBP52, and the NF-κB biological effect is triggered. Importantly, FKBP52 is functionally recruited to the promoter region of NF-κB target genes, whereas FKBP51 is released. Competition assays demonstrated that both immunophilins antagonize one another, and binding assays with purified proteins suggest that the association of RelA and immunophilins could be direct. These observations suggest that the biological action of NF-κB in different cell types could be positively regulated by a high FKBP52/FKBP51 expression ratio by favoring NF-κB nuclear retention, recruitment to the promoter regions of target genes, and transcriptional activity. PMID:25104352
Analysis and Quantitation of NF-[kappa]B Nuclear Translocation in Tumor Necrosis Factor Alpha (TNF-[alpha]) Activated Vascular Endothelial Cells

NASA Astrophysics Data System (ADS)

Fuseler, John W.; Merrill, Dana M.; Rogers, Jennifer A.; Grisham, Matthew B.; Wolf, Robert E.

2006-07-01

Nuclear factor kappa B (NF-[kappa]B) is a heterodimeric transcription factor typically composed of p50 and p65 subunits and is a pleiotropic regulator of various inflammatory and immune responses. In quiescent cells, p50/p65 dimers are sequestered in the cytoplasm bound to its inhibitors, the I-[kappa]Bs, which prevent entry into the nucleus. Following cellular stimulation, the I-[kappa]Bs are rapidly degraded, activating NF-[kappa]B. The active form of NF-[kappa]B rapidly translocates into the nucleus, binding to consensus sequences in the promoter/enhancer region of various genes, promoting their transcription. In human vascular endothelial cells activated with tumor necrosis factor-alpha, the activation and translocation of NF-[kappa]B is rapid, reaching maximal nuclear localization by 30 min. In this study, the appearance of NF-[kappa]B (p65 subunit, p65-NF-[kappa]B) in the nucleus visualized by immunofluorescence and quantified by morphometric image analysis (integrated optical density, IOD) is compared to the appearance of activated p65-NF-[kappa]B protein in the nucleus determined biochemically. The appearance of p65-NF-[kappa]B in the nucleus measured by fluorescence image analysis and biochemically express a linear correlation (R2 = 0.9477). These data suggest that localization and relative protein concentrations of NF-[kappa]B can be reliably determined from IOD measurements of the immunofluorescent labeled protein.
DAPk1 inhibits NF-κB activation through TNF-α and INF-γ-induced apoptosis.

PubMed

Yoo, Heon Jong; Byun, Hyun-Jung; Kim, Boh-Ram; Lee, Ki Hwan; Park, Sang-Yoon; Rho, Seung Bae

2012-07-01

Recent studies have shown DAPk as a molecular modulator induced by the second messenger, responsible for controlling cell destiny decisions, but the detailed mechanism mediating the role of DAPk1 during cell death is still not fully understood. In this present report, we attempted to characterize the effects of TNF-α and INF-γ on DAPk1 in human ovarian carcinoma cell lines, OVCAR-3. Both TNF-α and INF-γ significantly induce DAPk1 levels in a time-dependent manner. At the same time, they both arrested cell cycle progression in the G(0)-G(1) and G2/M phase, down-regulated cyclin D1, CDK4 and NF-κB expression, while also up-regulating p27 and p16 expression. Subsequently, the efficacy of the combined treatment with DAPk1 was investigated. In the presence of DAPk1, TNF-α or INF-γ-induced apoptosis was additively increased, while TNF-α or INF-γ-induced NF-κB activity was inhibited. Conversely, TNF-α or INF-γ-dependent NF-κB activity was further enhanced by the inhibition of DAPk1 with its specific siRNA. The activity of NF-κB was dependent on the level of DAPk1, indicating the requirement of DAPk1 for the activation of NF-κB. Low levels of DAPk1 expression were frequently observed in different human patient's tissue and cancer cell lines compared to normal samples. In addition, over-expression of DAPk1 from either TNF-α or INF-γ-treatment cells suppressed the anti-apoptosis protein XIAP as well as COX-2 and ICAM-1, more than control. Taken together, our data findings suggest that DAPk1 can mediate the pro-apoptotic activity of TNF-α and INF-γ via the NF-κB signaling pathways. Copyright © 2012 Elsevier Inc. All rights reserved.
Effect of the Various Steps in the Processing of Human Milk in the Concentrations of IgA, IgM, and Lactoferrin.

PubMed

Arroyo, Gerardo; Ortiz Barrientos, Kevin Alexander; Lange, Karla; Nave, Federico; Miss Mas, Gabriela; Lam Aguilar, Pamela; Soto Galindo, Miguel Angel

2017-09-01

Human milk immune components are unique and important for the development of the newborn. Milk processing at the Human Milk Banks (HMB), however, causes partial destruction of immune proteins. The objective of this study was to determine the effects that heating during the milk processing procedure at the HMB had on the concentrations of IgA, IgM, and lactoferrin at three critical points in time. Fifty milk samples (150 mL) were collected from voluntary donors at the HMB at the Hospital Nacional Pedro de Bethancourt, located in Antigua Guatemala. Samples from three critical points in time during the milk processing procedure were selected for analysis: freezing/thawing I, freezing/thawing II, and pasteurization. IgA, IgM, and lactoferrin concentrations were determined during each critical point and compared with a baseline concentration. After milk processing, IgA, IgM, and lactoferrin mean concentrations were reduced by 30.0%, 36.0%, and 70.0%, respectively (p < 0.001). Reduction of biological activity was mainly attributed to pasteurization for IgA and lactoferrin (p < 0.001); the first freezing/thawing processes before pasteurization showed no significant reduction difference between mean concentrations of IgA (p = 0.160) and lactoferrin (p = 0.345) but showed a significant effect on IgM concentration (p = 0.016), and the second freezing/thawing procedure only showed a significant effect on IgA (p < 0.001). The effects of milk processing on the immune proteins that were evaluated in this study demonstrated a significant reduction.
[Do lactoferrin, lysozyme and the lactoperoxidase-thiocyanate-hydrogen peroxide-system cause negative microbiological results in mastitis secretions?].

PubMed

Schmedt Auf Der Günne, H; Tenhagen, B A; Kutzer, P; Forderung, D; Heuwieser, W

2002-07-01

Lactoferrin, lysozyme and the lactoperoxidase-thiocyanate-peroxide-system are naturally occurring antimicrobial components of milk. The objective of this study was to examine, whether these components were responsible for negative results, when mastitis milk is cultured microbiologically. Quarter milk samples from 75 cows with clinical mastitis on a dairy farm in Brandenburg were submitted for microbiological culture and analysed for the content and the activities of the three components. Animals from all stages of lactation with clinical mastitis were included in the study. Animals were examined clinically and milk samples were collected prior to first treatment. Secretions from quarters with clinical mastitis were compared to those of neighbouring quarters without clinical mastitis. Secretions with positive cultural results were compared to those with negative results. The concentrations or activities of the three factors were significantly higher in the diseased quarters than in the quarters without clinical signs of mastitis. The concentration of lysozyme increased with severity of the clinical signs (local swelling and changes in secretion). The concentration of lactoferrin was significantly higher in quarters with slight alterations of glandular tissue than in quarters with medium or severe alterations (P < 0.05). LPS-activities did not correlate with the severity of clinical signs. No differences in the concentration of lactoferrin or LPS-activities were seen between mastitis with positive and negative culture results. The concentration of lysozyme was even higher in culturally positive samples than in negative samples (P < 0.05). Results from this study indicate that the three factors examined did not impair the results of microbiological culture of milk samples from quarters with clinical mastitis.
Pim-2 activates API-5 to inhibit the apoptosis of hepatocellular carcinoma cells through NF-kappaB pathway.

PubMed

Ren, Ke; Zhang, Wei; Shi, Yujun; Gong, Jianping

2010-06-01

Pim-2 is proved to be relevant to the tumorigenesis of hepatocellular carcinoma (HCC), but the mechanism is unclear. We studied the relationship among Pim-2, NF-kappaB and API-5. In our experiment, expression level of the three factors and phosphorylation level of API-5, as well as NF-kappaB activity, were detected in HCC tissues and the nontumorous controls. Then Pim-2 gene was transfected into nontumorous liver cells L02, and Pim-2 SiRNA was transfected into hepatoblastoma cell line HepG2. Parthenolide was added as NF-kappaB inhibitor. The same detections as above were repeated in the cells, along with the apoptosis analysis. We found the levels of Pim-2, NF-kappaB and API-5, as well as NF-kappaB activity, were significantly higher in HCC tissues. Pim-2 level was increased in L02 cells after the transfection of Pim-2 gene, but decreased in HepG2 cells after the transfection of Pim-2 SiRNA. The levels of NF-kappaB and API-5, as well as NF-kappaB activity and API-5 phosphorylation level, were in accordance with Pim-2 level, but could be reversed by Parthenolide. Cell apoptosis rates were negatively correlated with API-5 phosphorylation level. Therefore, we infer that Pim-2 could activate API-5 to inhibit the apoptosis of liver cells, and NF-kappaB is the key regulator.
Production of human lactoferrin in animal milk.

PubMed

Goldman, I L; Georgieva, S G; Gurskiy, Ya G; Krasnov, A N; Deykin, A V; Popov, A N; Ermolkevich, T G; Budzevich, A I; Chernousov, A D; Sadchikova, E R

2012-06-01

Genetic constructs containing the human lactoferrin (hLf) gene were created within a joint program of Russian and Belorussian scientists. Using these constructs, transgenic mice were bred (the maximum hLf concentration in their milk was 160 g/L), and transgenic goats were also generated (up to 10 g/L hLf in their milk). Experimental goatherds that produced hLf in their milk were also bred, and the recombinant hLf was found to be identical to the natural protein in its physical and chemical properties. These properties included electrophoretic mobility, isoelectric point, recognition by polyclonal and monoclonal antibodies, circular dichroic spectra, interaction with natural ligands (DNA, lipopolysaccharides, and heparin), the binding of iron ions, the sequence of the 7 terminal amino acids, and its biological activity. The latter was assessed by the agglutination of Micrococcus luteus protoplasts, bactericidal activity against Escherichia coli and Listeria monocytogenes , and fungicidal activity against Candida albicans . We also demonstrated a significant increase in the activity of antibiotics when used in combination with Lf.
The Scaffold Protein TANK/I-TRAF Inhibits NF-κB Activation by Recruiting Polo-like Kinase 1

PubMed Central

Zhang, Wanqiao; Zhang, Ying; Yuan, Yanzhi; Guan, Wei; Jin, Chaozhi; Chen, Hui; Wang, Xiaohui

2010-01-01

TANK/I-TRAF is a TRAF-binding protein that negatively regulates NF-κB activation. The underlying mechanism of this activity remains unclear. Here we show that TANK directly interacts with PLK1, a conserved cell cycle–regulated kinase. PLK1 inhibits NF-κB transcriptional activation induced by TNF-α, IL-1β, or several activators, but not by nuclear transcription factor p65. PLK1 expression reduces the DNA-binding activity of NF-κB induced by TNF-α. Moreover, endogenous activation of PLK1 reduces the TNF-induced phosphorylation of endogenous IκBα. PLK1 is bound to NEMO (IKKγ) through TANK to form a ternary complex in vivo. We describe a new regulatory mechanism for PLK1: PLK1 negatively regulates TNF-induced IKK activation by inhibiting the ubiquitination of NEMO. These findings reveal that the scaffold protein TANK recruits PLK1 to negatively regulate NF-κB activation and provide direct evidence that PLK1 is required for the repression function of TANK. PMID:20484576
Lactoferrin-binding proteins in Shigella flexneri.

PubMed Central

Tigyi, Z; Kishore, A R; Maeland, J A; Forsgren, A; Naidu, A S

1992-01-01

The ability of Shigella flexneri to interact with lactoferrin (Lf) was examined with a 125I-labeled protein-binding assay. The percent binding of human lactoferrin (HLf) and bovine lactoferrin (BLf) to 45 S. flexneri strains was 19 +/- 3 and 21 +/- 3 (mean +/- standard error of the mean), respectively. 125I-labeled HLf and BLf binding to strain M90T reached an equilibrium within 2 h. Unlabeled HLf and BLf displaced the 125I-HLf-bacteria interaction in a dose-dependent manner. The Lf-bacterium complex was uncoupled by KSCN or urea, but not by NaCl. The interaction was specific, and approximately 4,800 HLf binding sites (affinity constant [Ka], 690 nM) or approximately 5,700 BLf binding sites (Ka, 104 nM) per cell were estimated in strain M90T by a Scatchard plot analysis. The native cell envelope (CE) and outer membrane (OM) did not reveal Lf-binding components in sodium dodecyl sulfate-polyacrylamide gel electrophoresis. However, after being boiled, the CE and OM preparations showed three distinct horseradish peroxidase-Lf reactive bands of about 39, 22, and 16 kDa. The 39-kDa component was also reactive to a monoclonal antibody specific for porin (PoI) proteins of members of the family Enterobacteriaceae. The Lf-binding protein pattern was similar with BLf or HLf, for Crb+ and Crb- strains. The protein-Lf complex was dissociable by KSCN or urea and was stable after treatment with NaCl. Variation (loss) in the O chain of lipopolysaccharide (LPS) markedly enhanced the Lf-binding capacity in the isogenic rough strain SFL1070-15 compared with its smooth parent strain, SFL1070. These data establish that Lf binds to specific components in the bacterial OM; the heat-modifiable, anti-PoI-reactive, and LPS-associated properties suggested that the Lf-binding proteins are porins in S. flexneri. Images PMID:1319403
Constitutive NF-κB activation and tumor-growth promotion by Romo1-mediated reactive oxygen species production

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chung, Jin Sil; Lee, Sora; Yoo, Young Do, E-mail: ydy1130@korea.ac.kr

2014-08-08

Highlights: • Romo1 expression is required for constitutive nuclear DNA-binding activity of NF-κB. • Romo1 depletion suppresses tumor growth in vivo. • Romo1 presents a potential therapeutic target for diseases. - Abstract: Deregulation of nuclear factor-κB (NF-κB) and related pathways contribute to tumor cell proliferation and invasion. Mechanisms for constitutive NF-κB activation are not fully explained; however, the underlying defects appear to generate and maintain pro-oxidative conditions. In hepatocellular carcinoma (HCC) tissues, up-regulation of reactive oxygen species modulator 1 (Romo1) correlates positively with tumor size. In the present study, we showed that Romo1 expression is required to maintain constitutive nuclearmore » DNA-binding activity of NF-κB and transcriptional activity through constitutive IκBα phosphorylation. Overexpression of Romo1 promoted p65 nuclear translocation and DNA-binding activity. We also show that Romo1 depletion suppressed anchorage-independent colony formation by HCC cells and suppressed tumor growth in vivo. Based on these findings, Romo1 may be a principal regulatory factor in the maintenance of constitutive NF-κB activation in tumor cells. In the interest of anti-proliferative treatments for cancer, Romo1 may also present a productive target for drug development.« less
NF-κB Signalling in Glioblastoma

PubMed Central

Soubannier, Vincent; Stifani, Stefano

2017-01-01

Nuclear factor-κB (NF-κB) is a transcription factor regulating a wide array of genes mediating numerous cellular processes such as proliferation, differentiation, motility and survival, to name a few. Aberrant activation of NF-κB is a frequent event in numerous cancers, including glioblastoma, the most common and lethal form of brain tumours of glial cell origin (collectively termed gliomas). Glioblastoma is characterized by high cellular heterogeneity, resistance to therapy and almost inevitable recurrence after surgery and treatment. NF-κB is aberrantly activated in response to a variety of stimuli in glioblastoma, where its activity has been implicated in processes ranging from maintenance of cancer stem-like cells, stimulation of cancer cell invasion, promotion of mesenchymal identity, and resistance to radiotherapy. This review examines the mechanisms of NF-κB activation in glioblastoma, the involvement of NF-κB in several mechanisms underlying glioblastoma propagation, and discusses some of the important questions of future research into the roles of NF-κB in glioblastoma. PMID:28598356
Lactoferrin-derived peptides and Lactoferricin chimera inhibit virulence factor production and biofilm formation in Pseudomonas aeruginosa.

PubMed

Xu, G; Xiong, W; Hu, Q; Zuo, P; Shao, B; Lan, F; Lu, X; Xu, Y; Xiong, S

2010-10-01

To investigate the bactericidal activity of lactoferrin-derived peptides and a new LF-derived peptides chimera (LFchimera) against P. aeruginosa and the influence on virulence factors of P. aeruginosa. Lactoferricin (LFcin) and lactoferrampin (LFampin) are highly bioactive peptides isolated from the N-terminal region of lactoferrin (LF) by pepsin digestion. In this study, we designed LFchimera containing LFcin amino acids 17-30 and LFampin amino acids 268-284. Pseudomonas aeruginosa cells were incubated in medium with peptides at different concentrations, and then the assays of viability, pyocyanin, elastase activity and biofilm formation of P. aeruginosa were performed. We found that the concentration-dependent antibactericidal activity and down-regulating pyocyanin, elastase and biofilm formation of LFchimera were significantly stronger than those of LF, LFcin, LFampin or LFcin plus LFampin. Our results indicated that LF, LFcin, LFampin and LFchimera were potential candidates to combat P. aeruginosa, and LFchimera was the most effective in them. The new LFchimera has better activity against P. aeruginosa than LF, LFcin and LFampin and may be a promising new compound for treatment of P. aeruginosa infection. © 2010 The Authors. Journal compilation © 2010 The Society for Applied Microbiology.
Cocoa consumption reduces NF-κB activation in peripheral blood mononuclear cells in humans.

PubMed

Vázquez-Agell, M; Urpi-Sarda, M; Sacanella, E; Camino-López, S; Chiva-Blanch, G; Llorente-Cortés, V; Tobias, E; Roura, E; Andres-Lacueva, C; Lamuela-Raventós, R M; Badimon, L; Estruch, R

2013-03-01

Epidemiological studies have demonstrated an association between high-polyphenol intake and reduced incidence of atherosclerosis. The healthy effects of cocoa-polyphenols may be due to their antioxidant and anti-inflammatory actions, although the exact mechanisms are unknown and depend on the matrix in which cocoa-polyphenols are delivered. Nuclear factor κB (NF-κB) is a key molecule in the pathophysiology of atherosclerosis involved in the regulation of adhesion molecules(AM) and cytokine expression and its activation is the first step in triggering the inflammatory process. The aim of this study was to evaluate the effect of acute cocoa consumption in different matrices related to the bioavailability of cocoa-polyphenols in NF-κB activation and the expression of AM. Eighteen healthy volunteers randomly received 3 interventions: 40g of cocoa powder with milk (CM), with water (CW), and only milk (M). NF-κB activation in leukocytes and AM (sICAM, sVCAM, E-selectin) were measured before and 6h after each intervention. Consumption of CW significantly decreased NF-κB activation compared to baseline and to CM (P < 0.05, both), did not change after CM intervention, and significantly increased after M intervention (P = 0.014). sICAM-1 concentrations significantly decreased after 6h of CW and CM interventions (P ≤ 0.026; both) and E-selectin only decreased after CW intervention (P = 0.028). No significant changes were observed in sVCAM-1 concentrations. The anti-inflammatory effect of cocoa intake may depend on the bioavailability of bioactive compounds and may be mediated at least in part by the modulation of NF-κB activation and downstream molecules reinforcing the link between cocoa intake and health. Copyright © 2011 Elsevier B.V. All rights reserved.
Hydroxyapatite nanorod and microsphere functionalized with bioactive lactoferrin as a new biomaterial for enhancement bone regeneration.

PubMed

Shi, Pujie; Wang, Qun; Yu, Cuiping; Fan, Fengjiao; Liu, Meng; Tu, Maolin; Lu, Weihong; Du, Ming

2017-07-01

Lactoferrin (LF) has been recently recognized as a promising new novel bone growth factor for the beneficial effects on bone cells and promotion of bone growth. Currently, it has been attracted wide attention in bone regeneration as functional food additives or a potential bioactive protein in bone tissue engineering. The present study investigated the possibility that hydroxyapatite (HAP) particles, a widely used bone substitute material for high biocompatibility and osteoconductivity, functionalized with lactoferrin as a composite material are applied to bone tissue engineering. Two kinds of hydroxyapatite samples with different sizes, including nanorods and microspheres particles, were functionalized with lactoferrin molecules, respectively. A detailed characterization of as-prepared HAP-LF complex is presented, combining thermal gravimetric analysis (TGA) and Fourier Transform Infrared Spectroscopy (FT-IR). Zeta potential and the analysis of electrostatic surface potential of lactoferrin were carried to reveal the mechanism of adsorption. The effects of HAP-LF complex on MC3T3-E1 osteoblast proliferation and morphology were systematically evaluated at different culture time. Interestingly, results showed that cell viability of HAP-LF group was significantly higher than HAP group indicating that the HAP-LF can improve the biocompatibility of HAP, which mainly originated from a combination of HAP-LF interaction. These results indicated that hydroxyapatite particles can work as a controlled releasing carrier of lactoferrin successfully, and lactoferrin showed better potentiality on using in the field of bone regeneration by coupling with hydroxyapatite. This study would provide a new biomaterial and might offer a new insight for enhancement of bone regeneration. Copyright © 2017 Elsevier B.V. All rights reserved.
Destruction of single-species biofilms of Escherichia coli or Klebsiella pneumoniae subsp. pneumoniae by dextranase, lactoferrin, and lysozyme

USDA-ARS?s Scientific Manuscript database

The activity of dextranase, lactoferrin, lysozyme, and nisin against biofilms composed of either Klebsiella pneumonia or Escherichia coli was examined using the MBEC Assay™. Mature biofilms were treated and then sonicated to remove the adherent biofilm. This material was quantified using a lumines...
[NF-κB signaling pathways and the future perspectives of bone disease therapy using selective inhibitors of NF-κB].

PubMed

Jimi, Eijiro; Fukushima, Hidefumi

2016-02-01

The transcriptional factor nuclear factor κB(NF-κB)regulates the expression of a wide variety of genes that are involved in immune and inflammatory responses, proliferation, and tumorigenesis. NF-κB consists of five members, such as p65(RelA), RelB, c-Rel, p50/p105(NF-κB1), and p52/p100(NF-κB2). There are two distinct NF-κB activation pathways, termed the classical and alternative NF-κB signaling pathways. Since mice lacking both p50 and p52 subunits developed typical osteopetrosis, due to total lack of osteoclasts, NF-κB is also important osteoclast differentiation. A selective NF-κB inhibitor blocked receptor activator of NF-κB ligand(RANKL)-induced osteoclastogenesis both in vitro and in vivo. Recent findings have shown that inactivation of NF-κB enhances osteoblast differentiation in vitro and bone formation in vivo. NF-κB is constitutively activated in many cancers including oral squamous cell carcinoma(OSCC), and is involved in the invasive characteristics of OSCC. A selective NF-κB inhibitor also prevented jaw bone destruction by OSCC by reduced osteoclast numbers in animal model. Thus the inhibition of NF-κB might useful for the treatment of bone diseases, such as arthritis, osteoporosis, periodontitis, and bone invasion by OSCC by inhibiting bone resorption and by stimulating bone formation.

ARSENITE ACTIVATES KB-DEPENDENT IL-8 GENE EXPRESSION IN AIRWAY EPITHELIM IN THE ABSENCE OF NUCLEAR TRANSLOCATION OF NF-KB

EPA Science Inventory

Airway epithelial cells respond to certain environmental stresses by mounting a proinflammatory response, which is characterized by enhanced synthesis and release of the neutrophil chemotactic and activating factor interleukin-8 (IL-8). IL-8 expression is regulated at the transcr...
EBV induces persistent NF-κB activation and contributes to survival of EBV-positive neoplastic T- or NK-cells.

PubMed

Takada, Honami; Imadome, Ken-Ichi; Shibayama, Haruna; Yoshimori, Mayumi; Wang, Ludan; Saitoh, Yasunori; Uota, Shin; Yamaoka, Shoji; Koyama, Takatoshi; Shimizu, Norio; Yamamoto, Kouhei; Fujiwara, Shigeyoshi; Miura, Osamu; Arai, Ayako

2017-01-01

Epstein-Barr virus (EBV) has been detected in several T- and NK-cell neoplasms such as extranodal NK/T-cell lymphoma nasal type, aggressive NK-cell leukemia, EBV-positive peripheral T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood, and chronic active EBV infection (CAEBV). However, how this virus contributes to lymphomagenesis in T or NK cells remains largely unknown. Here, we examined NF-κB activation in EBV-positive T or NK cell lines, SNT8, SNT15, SNT16, SNK6, and primary EBV-positive and clonally proliferating T/NK cells obtained from the peripheral blood of patients with CAEBV. Western blotting, electrophoretic mobility shift assays, and immunofluorescent staining revealed persistent NF-κB activation in EBV-infected cell lines and primary cells from patients. Furthermore, we investigated the role of EBV in infected T cells. We performed an in vitro infection assay using MOLT4 cells infected with EBV. The infection directly induced NF-κB activation, promoted survival, and inhibited etoposide-induced apoptosis in MOLT4 cells. The luciferase assay suggested that LMP1 mediated NF-κB activation in MOLT4 cells. IMD-0354, a specific inhibitor of NF-κB that suppresses NF-κB activation in cell lines, inhibited cell survival and induced apoptosis. These results indicate that EBV induces NF-κB-mediated survival signals in T and NK cells, and therefore, may contribute to the lymphomagenesis of these cells.
EBV induces persistent NF-κB activation and contributes to survival of EBV-positive neoplastic T- or NK-cells

PubMed Central

Shibayama, Haruna; Yoshimori, Mayumi; Wang, Ludan; Saitoh, Yasunori; Uota, Shin; Yamaoka, Shoji; Koyama, Takatoshi; Shimizu, Norio; Yamamoto, Kouhei; Fujiwara, Shigeyoshi; Miura, Osamu

2017-01-01

Epstein–Barr virus (EBV) has been detected in several T- and NK-cell neoplasms such as extranodal NK/T-cell lymphoma nasal type, aggressive NK-cell leukemia, EBV-positive peripheral T-cell lymphoma, systemic EBV-positive T-cell lymphoma of childhood, and chronic active EBV infection (CAEBV). However, how this virus contributes to lymphomagenesis in T or NK cells remains largely unknown. Here, we examined NF-κB activation in EBV-positive T or NK cell lines, SNT8, SNT15, SNT16, SNK6, and primary EBV-positive and clonally proliferating T/NK cells obtained from the peripheral blood of patients with CAEBV. Western blotting, electrophoretic mobility shift assays, and immunofluorescent staining revealed persistent NF-κB activation in EBV-infected cell lines and primary cells from patients. Furthermore, we investigated the role of EBV in infected T cells. We performed an in vitro infection assay using MOLT4 cells infected with EBV. The infection directly induced NF-κB activation, promoted survival, and inhibited etoposide-induced apoptosis in MOLT4 cells. The luciferase assay suggested that LMP1 mediated NF-κB activation in MOLT4 cells. IMD-0354, a specific inhibitor of NF-κB that suppresses NF-κB activation in cell lines, inhibited cell survival and induced apoptosis. These results indicate that EBV induces NF-κB-mediated survival signals in T and NK cells, and therefore, may contribute to the lymphomagenesis of these cells. PMID:28346502
Curcumin suppresses colon cancer cell invasion via AMPK-induced inhibition of NF-κB, uPA activator and MMP9.

PubMed

Tong, Weihua; Wang, Quan; Sun, Donghui; Suo, Jian

2016-11-01

Curcumin, an active nontoxic ingredient of turmeric, possesses potent anti-inflammatory, antioxidant and anti-cancer properties; however, the molecular mechanisms of curcumin are not fully understood. The transcription factor nuclear factor-κB (NF-κB) is key in cellular processes, and the expression/activation of urokinase-type plasminogen activator (uPA) and matrix metalloproteinase-9 (MMP9) are crucial for cell invasion. The present study investigated the hypothesis that curcumin inhibits colon cancer cell invasion by modulating NF-κB-mediated expression and activation of uPA and MMP9. Human colon cancer SW480 and LoVo cells were treated with various concentrations of curcumin. Curcumin was demonstrated to dose-dependently inhibit the adhesion and proliferation ability of LoVo and SW480 cells using Transwell and MTT assays, respectively. In addition, curcumin activated 5' AMP-activated protein kinase (AMPK) and suppressed p65 NF-κB phosphorylation, as shown by western blot analysis. Compound C, a potent AMPK inhibitor, abolished curcumin-induced inhibition of NF-κB, uPA and MMP9, suggesting that AMPK activation is responsible for curcumin-mediated NF-κB, uPA and MMP9 inhibition. The binding activity of NF-κB to DNA was examined and western blotting and quantitative polymerase reaction was performed to detect the effect of curcumin on the expression of uPA and MMP9. The present results revealed that curcumin significantly decreased the expression of uPA and MMP9 and NF-κB DNA binding activity. Furthermore, curcumin decreased the level of the p65 subunit of NF-κB binding to the promoter of the gene encoding uPA and MMP9, which suppressed transcriptional activation of uPA and MMP9. Overall, the present data suggest that curcumin inhibits colon cancer cell invasion via AMPK activation and subsequent inhibition of p65 NF-κB, uPA and MMP9. The therapeutic potential of curcumin for colon cancer metastasis required additional study.
NF-κB as a target for oncogenic viruses

PubMed Central

Sun, Shao-Cong; Cesarman, Ethel

2013-01-01

NF-κB is a pivotal transcription factor that controls cell survival and proliferation in diverse physiological processes. The activity of NF-κB is tightly controlled through its cytoplasmic sequestration by specific inhibitors, IκBs. Various cellular stimuli induce the activation of an IκB kinase (IKK), which phosphorylates IκBs and triggers their proteasomal degradation, causing nuclear translocation of activated NF-κB. Under normal conditions, the activation of NF-κB occurs transiently, thus ensuring rapid but temporary induction of target genes. Deregulated NF-κB activation contributes to the development of various diseases, including cancers and immunological disorders. Accumulated studies demonstrate that the NF-κB signaling pathway is a target of several human oncogenic viruses, including the human T-cell leukemia virus type 1 (HTLV1), the Kaposi sarcoma-associated herpesvirus (KSHV), and the Epstein bar virus (EBV). These viruses encode specific oncoproteins that target different signaling components of the NF-κB pathway, leading to persistent activation of NF-κB. This chapter will discuss the molecular mechanisms by which NF-κB is activated by the viral oncoproteins. PMID:20845110
NF-kB activity-dependent P-selectin involved in ox-LDL-induced foam cell formation in U937 cell

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Yi, E-mail: wangyi2004a@126.com; Wang, Xiang; Sun, Minghui

Highlights: {yields} Ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. {yields} Ox-LDL induced expression of P-selectin through degradation of IkBa and augment of NF-kB activity and protein level during macrophage-derived foam cell formation. {yields} P-selectin and NF-kB may be identified as pivotal regulators of ox-LDL-induced foam cell formation. {yields} Therapy based on the inhibition of P-selectin and NF-kB may complement conventional treatments to prevent atherosclerosis. -- Abstract: Oxidized low-density lipoprotein (ox-LDL) plays a critical role in regulation of atherosclerosis. However, little is known about the role of Nuclear factor kBmore » (NF-kB) activity-dependent P-selectin in ox-LDL-induced foam cell formation during atherosclerosis. In this study, we first investigated ox-LDL induced foam cell formation in the human U937 promonocytic cell line in a dose- and time-dependent manner. Treatment of U937 cells with ox-LDL increased lipid accumulation as well as intracellular cholesterol content. Next, a comparative analysis of gene expression profiling using cDNA microarray and Real-time-PCR indicated that ox-LDL exposure induced, in three treated groups, an extremely marked increase in the mRNA level of P-selectin. Protein levels of P-selectin and its upstream regulators IkBa and NF-kB showed that NF-kB pathway is involved in the ox-LDL-induced foam cell formation. Finally, overexpression of NF-kB significantly accelerated, whereas, inhibition of NF-kB with siRNA remarkably attenuated ox-LDL-induced macrophage-derived foam cell formation. It was concluded that the activity of NF-kB is augmented during macrophage-derived foam cell formation. Activation of NF-kB increased, whereas, inhibition of NF-kB decreased ox-LDL-induced P-selectin expression and lipid accumulation in macrophages, suggesting ox-LDL induced expression of P-selectin through degradation of IkBa and activation of NF
TRAF Family Member-associated NF-κB Activator (TANK) Inhibits Genotoxic Nuclear Factor κB Activation by Facilitating Deubiquitinase USP10-dependent Deubiquitination of TRAF6 Ligase*

PubMed Central

Wang, Wei; Huang, Xuan; Xin, Hong-Bo; Fu, Mingui; Xue, Aimin; Wu, Zhao-Hui

2015-01-01

DNA damage-induced NF-κB activation plays a critical role in regulating cellular response to genotoxic stress. However, the molecular mechanisms controlling the magnitude and duration of this genotoxic NF-κB signaling cascade are poorly understood. We recently demonstrated that genotoxic NF-κB activation is regulated by reversible ubiquitination of several essential mediators involved in this signaling pathway. Here we show that TRAF family member-associated NF-κB activator (TANK) negatively regulates NF-κB activation by DNA damage via inhibiting ubiquitination of TRAF6. Despite the lack of a deubiquitination enzyme domain, TANK has been shown to negatively regulate the ubiquitination of TRAF proteins. We found TANK formed a complex with MCPIP1 (also known as ZC3H12A) and a deubiquitinase, USP10, which was essential for the USP10-dependent deubiquitination of TRAF6 and the resolution of genotoxic NF-κB activation upon DNA damage. Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-mediated deletion of TANK in human cells significantly enhanced NF-κB activation by genotoxic treatment, resulting in enhanced cell survival and increased inflammatory cytokine production. Furthermore, we found that the TANK-MCPIP1-USP10 complex also decreased TRAF6 ubiquitination in cells treated with IL-1β or LPS. In accordance, depletion of USP10 enhanced NF-κB activation induced by IL-1β or LPS. Collectively, our data demonstrate that TANK serves as an important negative regulator of NF-κB signaling cascades induced by genotoxic stress and IL-1R/Toll-like receptor stimulation in a manner dependent on MCPIP1/USP10-mediated TRAF6 deubiquitination. PMID:25861989
Transgenic expression of lactoferrin imparts enhanced resistance to head blight of wheat caused by Fusarium graminearum.

PubMed

Han, Jigang; Lakshman, Dilip K; Galvez, Leny C; Mitra, Sharmila; Baenziger, Peter Stephen; Mitra, Amitava

2012-03-09

The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Using the tools of plant genetic engineering, a broad-spectrum antimicrobial gene was tested for resistance against head blight caused by Fusarium graminearum Schwabe, a devastating disease of wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) that reduces both grain yield and quality. A construct containing a bovine lactoferrin cDNA was used to transform wheat using an Agrobacterium-mediated DNA transfer system to express this antimicrobial protein in transgenic wheat. Transformants were analyzed by Northern and Western blots to determine lactoferrin gene expression levels and were inoculated with the head blight disease fungus F. graminearum. Transgenic wheat showed a significant reduction of disease incidence caused by F. graminearum compared to control wheat plants. The level of resistance in the highly susceptible wheat cultivar Bobwhite was significantly higher in transgenic plants compared to control Bobwhite and two untransformed commercial wheat cultivars, susceptible Wheaton and tolerant ND 2710. Quantification of the expressed lactoferrin protein by ELISA in transgenic wheat indicated a positive correlation between the lactoferrin gene expression levels and the levels of disease resistance. Introgression of the lactoferrin gene into elite commercial wheat, barley and other susceptible cereals may enhance resistance to F. graminearum.
Primate Lentiviruses Modulate NF-κB Activity by Multiple Mechanisms to Fine-Tune Viral and Cellular Gene Expression

PubMed Central

Heusinger, Elena; Kirchhoff, Frank

2017-01-01

The transcription factor nuclear factor kappa-light-chain-enhancer of activated B cells (NF-κB) plays a complex role during the replication of primate lentiviruses. On the one hand, NF-κB is essential for induction of efficient proviral gene expression. On the other hand, this transcription factor contributes to the innate immune response and induces expression of numerous cellular antiviral genes. Recent data suggest that primate lentiviruses cope with this challenge by boosting NF-κB activity early during the replication cycle to initiate Tat-driven viral transcription and suppressing it at later stages to minimize antiviral gene expression. Human and simian immunodeficiency viruses (HIV and SIV, respectively) initially exploit their accessory Nef protein to increase the responsiveness of infected CD4+ T cells to stimulation. Increased NF-κB activity initiates Tat expression and productive replication. These events happen quickly after infection since Nef is rapidly expressed at high levels. Later during infection, Nef proteins of HIV-2 and most SIVs exert a very different effect: by down-modulating the CD3 receptor, an essential factor for T cell receptor (TCR) signaling, they prevent stimulation of CD4+ T cells via antigen-presenting cells and hence suppress further induction of NF-κB and an effective antiviral immune response. Efficient LTR-driven viral transcription is maintained because it is largely independent of NF-κB in the presence of Tat. In contrast, human immunodeficiency virus type 1 (HIV-1) and its simian precursors have lost the CD3 down-modulation function of Nef and use the late viral protein U (Vpu) to inhibit NF-κB activity by suppressing its nuclear translocation. In this review, we discuss how HIV-1 and other primate lentiviruses might balance viral and antiviral gene expression through a tight temporal regulation of NF-κB activity throughout their replication cycle. PMID:28261165
Retinoic Acid Protects Cardiomyocytes from High Glucose-Induced Apoptosis via Inhibition of Sustained Activation of NF-κB Signaling

PubMed Central

Nizamutdinova, Irina T.; Guleria, Rakeshwar S.; Singh, Amar B.; Kendall, Jonathan A.; Baker, Kenneth M.; Pan, Jing

2012-01-01

We have previously shown that retinoic acid (RA) has protective effects on high glucose (HG)-induced cardiomyocyte apoptosis. To further elucidate the molecular mechanisms of RA effects, we determined the interaction between nuclear factor (NF)-κB and RA signaling. HG induced a sustained phosphorylation of IKK/IκBα and transcriptional activation of NF-κB in cardiomyocytes. Activated NF-κB signaling has an important role in HG-induced cardiomyocyte apoptosis and gene expression of interleukin-6 (IL-6), tumor necrosis factor (TNF)-α and monocyte chemoattractant protein-1 (MCP-1). All-trans RA (ATRA) and LGD1069, through activation of RAR/RXR-mediated signaling, inhibited the HG-mediated effects in cardiomyocytes. The inhibitory effect of RA on NF-κB activation was mediated through inhibition of IKK/IκBα phosphorylation. ATRA and LGD1069 treatment promoted protein phosphatase 2A (PP2A) activity, which was significantly suppressed by HG stimulation. The RA effects on IKK and IκBα were blocked by okadaic acid or silencing the expression of PP2Ac-subunit, indicating that the inhibitory effect of RA on NF-κB is regulated through activation of PP2A and subsequent dephosphorylation of IKK/IκBα. Moreover, ATRA and LGD1069 reversed the decreased PP2A activity and inhibited the activation of IKK/IκBα and gene expression of MCP-1, IL-6 and TNF-α in the hearts of Zucker diabetic fatty rats. In summary, our findings suggest that the suppressed activation of PP2A contributed to sustained activation of NF-κB in HG-stimulated cardiomyocytes; and that the protective effect of RA on hyperglycemia-induced cardiomyocyte apoptosis and inflammatory responses is partially regulated through activation of PP2A and suppression of NF-κB-mediated signaling and downstream targets. PMID:22718360
NF-κB transcriptional activity is modulated by FK506-binding proteins FKBP51 and FKBP52: a role for peptidyl-prolyl isomerase activity.

PubMed

Erlejman, Alejandra G; De Leo, Sonia A; Mazaira, Gisela I; Molinari, Alejandro M; Camisay, María Fernanda; Fontana, Vanina; Cox, Marc B; Piwien-Pilipuk, Graciela; Galigniana, Mario D

2014-09-19

Hsp90 binding immunophilins FKBP51 and FKBP52 modulate steroid receptor trafficking and hormone-dependent biological responses. With the purpose to expand this model to other nuclear factors that are also subject to nuclear-cytoplasmic shuttling, we analyzed whether these immunophilins modulate NF-κB signaling. It is demonstrated that FKBP51 impairs both the nuclear translocation rate of NF-κB and its transcriptional activity. The inhibitory action of FKBP51 requires neither the peptidylprolyl-isomerase activity of the immunophilin nor its association with Hsp90. The TPR domain of FKBP51 is essential. On the other hand, FKBP52 favors the nuclear retention time of RelA, its association to a DNA consensus binding sequence, and NF-κB transcriptional activity, the latter effect being strongly dependent on the peptidylprolyl-isomerase activity and also on the TPR domain of FKBP52, but its interaction with Hsp90 is not required. In unstimulated cells, FKBP51 forms endogenous complexes with cytoplasmic RelA. Upon cell stimulation with phorbol ester, the NF-κB soluble complex exchanges FKBP51 for FKBP52, and the NF-κB biological effect is triggered. Importantly, FKBP52 is functionally recruited to the promoter region of NF-κB target genes, whereas FKBP51 is released. Competition assays demonstrated that both immunophilins antagonize one another, and binding assays with purified proteins suggest that the association of RelA and immunophilins could be direct. These observations suggest that the biological action of NF-κB in different cell types could be positively regulated by a high FKBP52/FKBP51 expression ratio by favoring NF-κB nuclear retention, recruitment to the promoter regions of target genes, and transcriptional activity. © 2014 by The American Society for Biochemistry and Molecular Biology, Inc.
Lactoferrin conjugated iron oxide nanoparticles for targeting brain glioma cells in magnetic particle imaging

NASA Astrophysics Data System (ADS)

Tomitaka, Asahi; Arami, Hamed; Gandhi, Sonu; Krishnan, Kannan M.

2015-10-01

Magnetic Particle Imaging (MPI) is a new real-time imaging modality, which promises high tracer mass sensitivity and spatial resolution directly generated from iron oxide nanoparticles. In this study, monodisperse iron oxide nanoparticles with median core diameters ranging from 14 to 26 nm were synthesized and their surface was conjugated with lactoferrin to convert them into brain glioma targeting agents. The conjugation was confirmed with the increase of the hydrodynamic diameters, change of zeta potential, and Bradford assay. Magnetic particle spectrometry (MPS), performed to evaluate the MPI performance of these nanoparticles, showed no change in signal after lactoferrin conjugation to nanoparticles for all core diameters, suggesting that the MPI signal is dominated by Néel relaxation and thus independent of hydrodynamic size difference or presence of coating molecules before and after conjugations. For this range of core sizes (14-26 nm), both MPS signal intensity and spatial resolution improved with increasing core diameter of nanoparticles. The lactoferrin conjugated iron oxide nanoparticles (Lf-IONPs) showed specific cellular internalization into C6 cells with a 5-fold increase in MPS signal compared to IONPs without lactoferrin, both after 24 h incubation. These results suggest that Lf-IONPs can be used as tracers for targeted brain glioma imaging using MPI.
Relationship between salivary immunoglobulin a, lactoferrin and lysozyme flow rates and lifestyle factors in Japanese children: a cross-sectional study.

PubMed

Ide, Momo; Saruta, Juri; To, Masahiro; Yamamoto, Yuko; Sugimoto, Masahiro; Fuchida, Shinya; Yokoyama, Mina; Kimoto, Shigenari; Tsukinoki, Keiichi

2016-10-01

The antimicrobial substances in saliva contribute to the maintenance of both oral health and overall health of the body. Therefore, the associations among immunoglobulin A (IgA), lactoferrin and lysozyme flow rates in the saliva of children, and their relationships with the physical attributes and lifestyle factors of children, were examined. Saliva was collected from 90 children who visited the Kanagawa Dental University Hospital Pediatric Dentistry, and questionnaires were completed by guardians. IgA, lactoferrin and lysozyme concentrations were measured in the saliva samples using enzyme-linked immunosorbent assays (ELISAs). The IgA flow rate in saliva increased as age, height and weight increased. A correlation was found between lactoferrin and lysozyme flow rates. When the antimicrobial substance flow rates in the saliva were divided into two groups of 22 children each based on the highest and lowest quartiles, children with either a low or high IgA flow rate also had a high or low lactoferrin flow rate, respectively. The same pattern was observed for lactoferrin and lysozyme flow rates. There is a high probability that the IgA flow rate in the saliva of children reflects and corresponds to the developmental status of immune function as the child ages and increases in height and weight. The flow rates of lactoferrin and lysozyme were correlated in children. In addition, regarding lifestyle factors, the duration of sleep and lactoferrin flow rate were also related.
Analysis of Nuclear Factor-κB (NF-κB) Essential Modulator (NEMO) Binding to Linear and Lysine-linked Ubiquitin Chains and Its Role in the Activation of NF-κB*

PubMed Central

Kensche, Tobias; Tokunaga, Fuminori; Ikeda, Fumiyo; Goto, Eiji; Iwai, Kazuhiro; Dikic, Ivan

2012-01-01

Nuclear factor-κB (NF-κB) essential modulator (NEMO), a component of the inhibitor of κB kinase (IKK) complex, controls NF-κB signaling by binding to ubiquitin chains. Structural studies of NEMO provided a rationale for the specific binding between the UBAN (ubiquitin binding in ABIN and NEMO) domain of NEMO and linear (Met-1-linked) di-ubiquitin chains. Full-length NEMO can also interact with Lys-11-, Lys-48-, and Lys-63-linked ubiquitin chains of varying length in cells. Here, we show that purified full-length NEMO binds preferentially to linear ubiquitin chains in competition with lysine-linked ubiquitin chains of defined length, including long Lys-63-linked deca-ubiquitins. Linear di-ubiquitins were sufficient to activate both the IKK complex in vitro and to trigger maximal NF-κB activation in cells. In TNFα-stimulated cells, NEMO chimeras engineered to bind exclusively to Lys-63-linked ubiquitin chains mediated partial NF-κB activation compared with cells expressing NEMO that binds to linear ubiquitin chains. We propose that NEMO functions as a high affinity receptor for linear ubiquitin chains and a low affinity receptor for long lysine-linked ubiquitin chains. This phenomenon could explain quantitatively distinct NF-κB activation patterns in response to numerous cell stimuli. PMID:22605335
Regulation of MDA5-MAVS Antiviral Signaling Axis by TRIM25 through TRAF6-Mediated NF-κB Activation.

PubMed

Lee, Na-Rae; Kim, Hye-In; Choi, Myung-Soo; Yi, Chae-Min; Inn, Kyung-Soo

2015-09-01

Tripartite motif protein 25 (TRIM25), mediates K63-linked polyubiquitination of Retinoic acid inducible gene I (RIG-I) that is crucial for downstream antiviral interferon signaling. Here, we demonstrate that TRIM25 is required for melanoma differentiation-associated gene 5 (MDA5) and MAVS mediated activation of NF-κB and interferon production. TRIM25 is required for the full activation of NF-κB at the downstream of MAVS, while it is not involved in IRF3 nuclear translocation. Mechanical studies showed that TRIM25 is involved in TRAF6-mediated NF-κB activation. These collectively indicate that TRIM25 plays an additional role in RIG-I/MDA5 signaling other than RIG-I ubiquitination via activation of NF-κB.
Regulation of MDA5-MAVS Antiviral Signaling Axis by TRIM25 through TRAF6-Mediated NF-κB Activation

PubMed Central

Lee, Na-Rae; Kim, Hye-In; Choi, Myung-Soo; Yi, Chae-Min; Inn, Kyung-Soo

2015-01-01

Tripartite motif protein 25 (TRIM25), mediates K63-linked polyubiquitination of Retinoic acid inducible gene I (RIG-I) that is crucial for downstream antiviral interferon signaling. Here, we demonstrate that TRIM25 is required for melanoma differentiation-associated gene 5 (MDA5) and MAVS mediated activation of NF-κB and interferon production. TRIM25 is required for the full activation of NF-κB at the downstream of MAVS, while it is not involved in IRF3 nuclear translocation. Mechanical studies showed that TRIM25 is involved in TRAF6-mediated NF-κB activation. These collectively indicate that TRIM25 plays an additional role in RIG-I/MDA5 signaling other than RIG-I ubiquitination via activation of NF-κB. PMID:26299329
The effect of bovine lactoferrin and lactoferricin B on the ability of feline calicivirus (a norovirus surrogate) and poliovirus to infect cell cultures.

PubMed

McCann, K B; Lee, A; Wan, J; Roginski, H; Coventry, M J

2003-01-01

To characterize the effect of bovine lactoferrin and lactoferricin B against feline calicivirus (FCV), a norovirus surrogate and poliovirus (PV), as models for enteric viruses. Crandell-Reese feline kidney (CRFK) cells were used for the propagation of FCV and monkey embryo kidney (MEK) cells for PV. The assays included visual assessment of cell lines for cytopathic effects and determination of the percentage cell death using MTT [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium] dye reduction assay. Incubation of bovine lactoferrin with CRFK cells either prior to or together with FCV inoculation substantially reduced FCV infection. In contrast, the interference of lactoferrin with the infection of cells with PV was demonstrated only when lactoferrin was present with cell lines and virus for the entire assay period. Using indirect immunofluorescence, lactoferrin was detected on the surface of both CRFK and MEK cells, suggesting that the interference of viral infection may be attributed to lactoferrin binding to the surfaces of susceptible cells, thereby preventing the attachment of the virus particles. Lactoferricin B, a cationic antimicrobial peptide derived from the N-terminal domain of bovine lactoferrin, reduced FCV but not PV infection. Lactoferrin was shown to interfere with the infection of cells for both FCV and PV. However, lactoferricin B showed no interference of infection with PV and interference with infection for FCV required the presence of lactoferricin B together with the cell line and virus. An in vitro basis is provided for the effects of bovine lactoferrin and lactoferricin B in moderating food-borne infections of enteric viruses.
TRAF Family Member-associated NF-κB Activator (TANK) Inhibits Genotoxic Nuclear Factor κB Activation by Facilitating Deubiquitinase USP10-dependent Deubiquitination of TRAF6 Ligase.

PubMed

Wang, Wei; Huang, Xuan; Xin, Hong-Bo; Fu, Mingui; Xue, Aimin; Wu, Zhao-Hui

2015-05-22

DNA damage-induced NF-κB activation plays a critical role in regulating cellular response to genotoxic stress. However, the molecular mechanisms controlling the magnitude and duration of this genotoxic NF-κB signaling cascade are poorly understood. We recently demonstrated that genotoxic NF-κB activation is regulated by reversible ubiquitination of several essential mediators involved in this signaling pathway. Here we show that TRAF family member-associated NF-κB activator (TANK) negatively regulates NF-κB activation by DNA damage via inhibiting ubiquitination of TRAF6. Despite the lack of a deubiquitination enzyme domain, TANK has been shown to negatively regulate the ubiquitination of TRAF proteins. We found TANK formed a complex with MCPIP1 (also known as ZC3H12A) and a deubiquitinase, USP10, which was essential for the USP10-dependent deubiquitination of TRAF6 and the resolution of genotoxic NF-κB activation upon DNA damage. Clustered regularly interspaced short palindromic repeats (CRISPR)/Cas9-mediated deletion of TANK in human cells significantly enhanced NF-κB activation by genotoxic treatment, resulting in enhanced cell survival and increased inflammatory cytokine production. Furthermore, we found that the TANK-MCPIP1-USP10 complex also decreased TRAF6 ubiquitination in cells treated with IL-1β or LPS. In accordance, depletion of USP10 enhanced NF-κB activation induced by IL-1β or LPS. Collectively, our data demonstrate that TANK serves as an important negative regulator of NF-κB signaling cascades induced by genotoxic stress and IL-1R/Toll-like receptor stimulation in a manner dependent on MCPIP1/USP10-mediated TRAF6 deubiquitination. © 2015 by The American Society for Biochemistry and Molecular Biology, Inc.
KB425796-A, a novel antifungal antibiotic produced by Paenibacillus sp. 530603.

PubMed

Kai, Hirohito; Yamashita, Midori; Takase, Shigehiro; Hashimoto, Michizane; Muramatsu, Hideyuki; Nakamura, Ikuko; Yoshikawa, Koji; Ezaki, Masami; Nitta, Kumiko; Watanabe, Masato; Inamura, Noriaki; Fujie, Akihiko

2013-08-01

The novel antifungal macrocyclic lipopeptidolactone, KB425796-A (1), was isolated from the fermentation broth of bacterial strain 530603, which was identified as a new Paenibacillus species based on morphological and physiological characteristics, and 16S rRNA sequences. KB425796-A (1) was isolated as white powder by solvent extraction, HP-20 and ODS-B column chromatography, and lyophilization, and was determined to have the molecular formula C79H115N19O18. KB425796-A (1) showed antifungal activities against Aspergillus fumigatus and the micafungin-resistant infectious fungi Trichosporon asahii, Rhizopus oryzae, Pseudallescheria boydii and Cryptococcus neoformans.
Nonylphenol regulates cyclooxygenase-2 expression via Ros-activated NF-κB pathway in sertoli TM4 cells.

PubMed

Liu, Xiaozhen; Nie, Shaoping; Huang, Danfei; Xie, Mingyong

2015-09-01

The aim of this study was to investigate the signaling pathways involved in the cyclooxygenase (COX)-2 regulation induced by nonylphenol (NP) in mouse testis Sertoli TM4 cells. Our results showed that treatment of TM4 cells with NP increased COX-2 protein expression and interleukin-6 (IL)-6 and prostaglandin E2 (PGE2) secretion in a dose-dependent manner. Pretreatment with reactive oxygen species (ROS) scavenger, N-acetylcysteine (NAC), attenuated NP-induced ROS production, COX-2 expression, and IL-6 and PGE2 release in TM4 cells. Exposure to NP stimulated activation of NF-κB, whereas the NF-κB inhibitor, pyrrolidine dithiocarbamate, attenuated NP-enhanced COX-2 expression and IL-6 and PGE2 release in TM4 cells in a dose-dependent manner. Furthermore, NAC blocked NP-induced activation of NF-κB. In addition, inhibition of COX-2 mitigated NP-induced IL-6 release. In conclusion, NP induced ROS generation, activation of NF-κB pathway, COX-2 upregulation, and IL-6 and PGE2 secretion in TM4 cells. NP may regulate COX-2 expression via ROS-activated NF-κB pathway in Sertoli TM4 cells. © 2014 Wiley Periodicals, Inc.

Antiproliferative activity of flower hexane extract obtained from Mentha spicata associated with Mentha rotundifolia against the MCF7, KB, and NIH/3T3 cell lines.

PubMed

Nedel, Fernanda; Begnini, Karine; Carvalho, Pedro Henrique de Azambuja; Lund, Rafael Guerra; Beira, Fátima T A; Del Pino, Francisco Augusto B

2012-11-01

This study assessed the antiproliferative effect in vitro of the flower hexane extract obtained from Mentha spicata associated with Mentha rotundifolia against the human breast adenocarcinoma (MCF-7), human mouth epidermal carcinoma (KB), and mouse embryonic fibroblast (NIH 3T3) cell lines, using sulforhodamine B (SRB) assay. A cell density of 2×10(4)/well was seeded in 96-well plates, and samples at different concentrations ranging from 10 to 500 mg/mL were tested. The optical density was determined in an ELISA multiplate reader (Thermo Plate TP-Reader). Results demonstrated that the hexane extract presented antiproliferative activity against both the tumor cell lines KB and MCF-7, presenting a GI(50) (MCF-7=13.09 mg/mL), TGI (KB=37.76 mg/mL), and IL(50) (KB=291.07 mg/mL). Also, the hexane extract presented antiproliferative activity toward NIH 3T3 cells GI(50) (183.65 mg/mL), TGI (280.54 mg/mL), and IL(50) (384.59 mg/mL). The results indicate that the flower hexane extract obtained from M. spicata associated with M. rotundifolia presents an antineoplastic activity against KB and MCF-7, although an antiproliferative effect at a high concentration of the extract was observed toward NIH 3T3.
SIRT1 counteracted the activation of STAT3 and NF-κB to repress the gastric cancer growth.

PubMed

Lu, Juanjuan; Zhang, Liping; Chen, Xiang; Lu, Qiming; Yang, Yuxia; Liu, Jingping; Ma, Xin

2014-01-01

Sirtuin-1 (SIRT1) possesses apparently dual roles in regulation of tumor. Previous reports have documented the crosstalk between SIRT1 with signal transducer and activator of transcription 3 (STAT3) and nuclear factor kappa-B (NF-κB) signaling in leukemia, lymphoma and myeloma. In this study, the purpose was to survey the regulatory effects of SIRT1 on gastric cancer (GC) cells (AGS and MKN-45) and the relationships between SIRT1 and activation of STAT3 and NF-κB in GC cells. We found the SIRT1 activator (resveratrol RSV) contributed to the repression of viability and increase of senescence, which were rescued by SIRT1 inhibitor (nicotinamide NA) and SIRT1 depletion by CCK-8 assay and SA-β-gal assay respectively. Further study found SIRT1 activation (RSV supplement) not only inhibited the activation of STAT3 including STAT3 mRNA level, c-myc mRNA level phosphorylated STAT3 (pSTAT3) proteins and acetylizad STAT3 (acSTAT3) proteins, but also repression of pNF-κB p65 and acNF-κB p65. NA reversed the effects of RSV. In addition, either RSV or NA application could not change the cellular viability and senescence in MKN-45 cells with STAT3 knockdown or NF-κB knockdown. Overall, our findings suggested SIRT1 activation could induced the loss of viability and increases of senescence in GC in vitro. Moreover, our observations revealed SIRT1 displayed growth inhibitory activity in GC cells highly associated with causing repression of activation of STAT3 and NF-κB proteins via deacetylation.
Corneal NF-kappaB activity is necessary for the retention of transparency in the cornea of UV-B-exposed transgenic reporter mice.

PubMed

Alexander, George; Carlsen, Harald; Blomhoff, Rune

2006-04-01

To determine the dynamics of Nuclear Factor-kappaB (NF-kappaB) in murine corneal pathology and the role of NF-kappaB in maintaining corneal clarity after ultraviolet B radiation insult, transgenic mice containing NF-kappaB-luciferase reporter were exposed to LPS (bacterial lipopolysaccharide), TNF-alpha (Tumor Necrosis Factor-alpha) or 4 kJ m(-2) UV-B radiation. NF-kappaB decoy oligonucleotides were also administered in some of the UV-B experiments. Following various exposure times, the mice were sacrificed and whole eyes or corneal tissues were obtained. Whole eyes were examined for scattering using a point-source optical imaging technique. Tissue homogenates were examined for luciferase activity using a luminometer. TNF-alpha and LPS-injected NF-kappaB-luciferase transgenic mice demonstrated 3-10-fold increases in cornea NF-kappaB with peak activities at 4 and 6 hr post-injection, respectively. Mice exposed to 4 kJ m(-2) UV-B exhibited a 3-fold increase in NF-kappaB activity 4 hr post-exposure. The administration of NF-kappaB-decoy oligonucleotides to mice had the effect of reducing UV-B-induced NF-kappaB activity in the cornea and significantly increasing the amount of light scattering in UV-B exposed corneas 7 days post-UV-B exposure when compared to sham injected mice. These results indicate that NF-kappaB is activated in cornea in pathologies that involves increased plasma levels of LPS and TNF-alpha, as well as direct UV-B exposure, and suggest that NF-kappaB activation play an essential part in the corneal healing process.
Curcumin Regulates Low-Linear Energy Transfer {gamma}-Radiation-Induced NF{kappa}B-Dependent Telomerase Activity in Human Neuroblastoma Cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Aravindan, Natarajan, E-mail: naravind@ouhsc.ed; Veeraraghavan, Jamunarani; Madhusoodhanan, Rakhesh

2011-03-15

Purpose: We recently reported that curcumin attenuates ionizing radiation (IR)-induced survival signaling and proliferation in human neuroblastoma cells. Also, in the endothelial system, we have demonstrated that NF{kappa}B regulates IR-induced telomerase activity (TA). Accordingly, we investigated the effect of curcumin in inhibiting IR-induced NF{kappa}B-dependent hTERT transcription, TA, and cell survival in neuroblastoma cells. Methods and Materials: SK-N-MC or SH-SY5Y cells exposed to IR and treated with curcumin (10-100 nM) with or without IR were harvested after 1 h through 24 h. NF{kappa}B-dependent regulation was investigated either by luciferase reporter assays using pNF{kappa}B-, pGL3-354-, pGL3-347-, or pUSE-I{kappa}B{alpha}-Luc, p50/p65, or RelA siRNA-transfectedmore » cells. NF{kappa}B activity was analyzed using an electrophoretic mobility shift assay and hTERT expression using the quantitative polymerase chain reaction. TA was determined using the telomerase repeat amplification protocol assay and cell survival using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltertrazolium bromide and clonogenic assay. Results: Curcumin profoundly inhibited IR-induced NF{kappa}B. Consequently, curcumin significantly inhibited IR-induced TA and hTERT mRNA at all points investigated. Furthermore, IR-induced TA is regulated at the transcriptional level by triggering telomerase reverse transcriptase (TERT) promoter activation. Moreover, NF{kappa}B becomes functionally activated after IR and mediates TA upregulation by binding to the {kappa}B-binding region in the promoter region of the TERT gene. Consistently, elimination of the NF{kappa}B-recognition site on the telomerase promoter or inhibition of NF{kappa}B by the I{kappa}B{alpha} mutant compromises IR-induced telomerase promoter activation. Significantly, curcumin inhibited IR-induced TERT transcription. Consequently, curcumin inhibited hTERT mRNA and TA in NF{kappa}B overexpressed cells. Furthermore, curcumin
Mapping PDB chains to UniProtKB entries.

PubMed

Martin, Andrew C R

2005-12-01

UniProtKB/SwissProt is the main resource for detailed annotations of protein sequences. This database provides a jumping-off point to many other resources through the links it provides. Among others, these include other primary databases, secondary databases, the Gene Ontology and OMIM. While a large number of links are provided to Protein Data Bank (PDB) files, obtaining a regularly updated mapping between UniProtKB entries and PDB entries at the chain or residue level is not straightforward. In particular, there is no regularly updated resource which allows a UniProtKB/SwissProt entry to be identified for a given residue of a PDB file. We have created a completely automatically maintained database which maps PDB residues to residues in UniProtKB/SwissProt and UniProtKB/trEMBL entries. The protocol uses links from PDB to UniProtKB, from UniProtKB to PDB and a brute-force sequence scan to resolve PDB chains for which no annotated link is available. Finally the sequences from PDB and UniProtKB are aligned to obtain a residue-level mapping. The resource may be queried interactively or downloaded from http://www.bioinf.org.uk/pdbsws/.
Lactoferrin or ferrous salts for iron deficiency anemia in pregnancy: A meta-analysis of randomized trials.

PubMed

Abu Hashim, Hatem; Foda, Osama; Ghayaty, Essam

2017-12-01

This systematic review and meta-analysis aimed to evaluate the efficacy of daily oral bovine lactoferrin versus daily oral ferrous iron preparations for treatment of iron deficiency anemia (IDA) during pregnancy. Searches were conducted on PubMed, ScienceDirect, ClinicalTrials.gov and CENTRAL databases from inception to February 2017 and the bibliographies of retrieved articles were screened. The PRISMA Statement was followed. Published English language randomized trials comparing lactoferrin with oral ferrous iron preparations in pregnant women with iron deficiency anemia were included. Quasi-randomized, non- randomized or studies including other known cause of anemia, gestational or pre-existent maternal diseases were excluded. Accordingly, 4 eligible trials (600 women) were analyzed. Primary outcome was change in hemoglobin level at 4 weeks of treatment. Secondary outcomes were; change in serum ferritin and iron, rates of gastrointestinal side effects, preterm birth, low birthweight, neonatal death and mean birthweight. Quality assessment was performed by the Cochrane risk of bias tool. Odds ratio and mean difference were used to integrate dichotomous and continuous outcomes respectively. Pooled estimates for change in hemoglobin levels at four weeks favored daily oral lactoferrin over daily oral ferrous sulphate (mean difference 0.77; 95% confidence interval [CI] 0.04-1.55; P=0.04, 4 trials, 600 women). However, after subgroup analysis (degree of anemia), no significant difference in hemoglobin levels were found between both groups in mild anemia (mean difference 0.80; 95% CI -0.21 to 1.82, 3 trials, 372 women), but a significant increase favoring lactoferrin was reported in moderate anemia (mean difference 0.68; 95% CI 0.53-0.83; P<0.00001, one trial, 228 women). Significantly less gastrointestinal side effects were reported with lactoferrin treatment. No significant differences existed with regard to other outcomes. In conclusion, for pregnant women with IDA
Three-dimensional solution structure of lactoferricin B, an antimicrobial peptide derived from bovine lactoferrin.

PubMed

Hwang, P M; Zhou, N; Shan, X; Arrowsmith, C H; Vogel, H J

1998-03-24

The solution structure of bovine lactoferricin (LfcinB) has been determined using 2D 1H NMR spectroscopy. LfcinB is a 25-residue antimicrobial peptide released by pepsin cleavage of lactoferrin, an 80 kDa iron-binding glycoprotein with many immunologically important functions. The NMR structure of LfcinB reveals a somewhat distorted antiparallel beta-sheet. This contrasts with the X-ray structure of bovine lactoferrin, in which residues 1-13 (of LfcinB) form an alpha-helix. Hence, this region of lactoferricin B appears able to adopt a helical or sheetlike conformation, similar to what has been proposed for the amyloidogenic prion proteins and Alzheimer's beta-peptides. LfcinB has an extended hydrophobic surface comprised of residues Phe1, Cys3, Trp6, Trp8, Pro16, Ile18, and Cys20. The side chains of these residues are well-defined in the NMR structure. Many hydrophilic and positively charged residues surround the hydrophobic surface, giving LfcinB an amphipathic character. LfcinB bears numerous similarities to a vast number of cationic peptides which exert their antimicrobial activities through membrane disruption. The structures of many of these peptides have been well characterized, and models of their membrane-permeabilizing mechanisms have been proposed. The NMR solution structure of LfcinB may be more relevant to membrane interaction than that suggested by the X-ray structure of intact lactoferrin. Based on the solution structure, it is now possible to propose potential mechanisms for the antimicrobial action of LfcinB.
Downregulation of STAT3/NF-κB potentiates gemcitabine activity in pancreatic cancer cells.

PubMed

Gong, Jingjing; Muñoz, Amanda R; Pingali, Subramanya; Payton-Stewart, Florastina; Chan, Daniel E; Freeman, James W; Ghosh, Rita; Kumar, Addanki P

2017-02-01

There is an unmet need to develop new agents or strategies against therapy resistant pancreatic cancer (PanCA). Recent studies from our laboratory showed that STAT3 negatively regulates NF-κB and that inhibition of this crosstalk using Nexrutine® (Nx) reduces transcriptional activity of COX-2. Inhibition of these molecular interactions impedes pancreatic cancer cell growth as well as reduces fibrosis in a preclinical animal model. Nx is an extract derived from the bark of Phellodendron amurense and has been utilized in traditional Chinese medicine as antidiarrheal, astringent, and anti-inflammatory agent for centuries. We hypothesized that "Nx-mediated inhibition of survival molecules like STAT3 and NF-κB in pancreatic cancer cells will improve the efficacy of the conventional chemotherapeutic agent, gemcitabine (GEM)." Therefore, we explored the utility of Nx, one of its active constituents berberine and its derivatives, to enhance the effects of GEM. Using multiple human pancreatic cancer cells we found that combination treatment with Nx and GEM resulted in significant alterations of proteins in the STAT3/NF-κB signaling axis culminating in growth inhibition in a synergistic manner. Furthermore, GEM resistant cells were more sensitive to Nx treatment than their parental GEM-sensitive cells. Interestingly, although berberine, the Nx active component used, and its derivatives were biologically active in GEM sensitive cells they did not potentiate GEM activity when used in combination. Taken together, these results suggest that the natural extract, Nx, but not its active component, berberine, has the potential to improve GEM sensitivity, perhaps by down regulating STAT3/NF-κB signaling. © 2016 Wiley Periodicals, Inc. © 2016 Wiley Periodicals, Inc.
Lactoferrin release and interleukin-1, interleukin-6, and tumor necrosis factor production by human polymorphonuclear cells stimulated by various lipopolysaccharides: relationship to growth inhibition of Candida albicans.

PubMed

Palma, C; Cassone, A; Serbousek, D; Pearson, C A; Djeu, J Y

1992-11-01

Lipopolysaccharides (LPSs) from Escherichia coli, Serratia marcescens, and Salmonella typhimurium, at doses from 1 to 100 ng/ml, strongly enhanced growth inhibition of Candida albicans by human polymorphonuclear leukocytes (PMN) in vitro. Flow cytometry analysis demonstrated that LPS markedly augmented phagocytosis of Candida cells by increasing the number of yeasts ingested per neutrophil as well as the number of neutrophils capable of ingesting fungal cells. LPS activation caused augmented release of lactoferrin, an iron-binding protein which itself could inhibit the growth of C. albicans in vitro. Antibodies against lactoferrin effectively and specifically reduced the anti-C. albicans activity of both LPS-stimulated and unstimulated PMN. Northern (RNA blot) analysis showed enhanced production of mRNAs for interleukin-1 beta, tumor necrosis factor alpha, and interleukin-6 and in neutrophils within 1 h of stimulation with LPS. The cytokines were also detected in the supernatant of the activated PMN, and their synthesis was prevented by pretreatment of LPS-stimulated PMN with protein synthesis inhibitors, such as emetine and cycloheximide. These inhibitors, however, did not block either lactoferrin release or the anti-Candida activity of LPS-stimulated PMN. These results demonstrate the ability of various bacterial LPSs to augment neutrophil function against C. albicans and suggest that the release of a candidastatic, iron-binding protein, lactoferrin, may contribute to the antifungal effect of PMN. Moreover, the ability to produce cytokines upon stimulation by ubiquitous microbial products such as the endotoxins points to an extraphagocytic, immunomodulatory role of PMN during infection.
Transgenic expression of lactoferrin imparts enhanced resistance to head blight of wheat caused by Fusarium graminearum

PubMed Central

2012-01-01

Background The development of plant gene transfer systems has allowed for the introgression of alien genes into plant genomes for novel disease control strategies, thus providing a mechanism for broadening the genetic resources available to plant breeders. Using the tools of plant genetic engineering, a broad-spectrum antimicrobial gene was tested for resistance against head blight caused by Fusarium graminearum Schwabe, a devastating disease of wheat (Triticum aestivum L.) and barley (Hordeum vulgare L.) that reduces both grain yield and quality. Results A construct containing a bovine lactoferrin cDNA was used to transform wheat using an Agrobacterium-mediated DNA transfer system to express this antimicrobial protein in transgenic wheat. Transformants were analyzed by Northern and Western blots to determine lactoferrin gene expression levels and were inoculated with the head blight disease fungus F. graminearum. Transgenic wheat showed a significant reduction of disease incidence caused by F. graminearum compared to control wheat plants. The level of resistance in the highly susceptible wheat cultivar Bobwhite was significantly higher in transgenic plants compared to control Bobwhite and two untransformed commercial wheat cultivars, susceptible Wheaton and tolerant ND 2710. Quantification of the expressed lactoferrin protein by ELISA in transgenic wheat indicated a positive correlation between the lactoferrin gene expression levels and the levels of disease resistance. Conclusions Introgression of the lactoferrin gene into elite commercial wheat, barley and other susceptible cereals may enhance resistance to F. graminearum. PMID:22405032
Oxaliplatin antagonizes HIV-1 latency by activating NF-κB without causing global T cell activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhu, Xiaoli; Liu, Sijie; Wang, Pengfei

Highlights: • The chemotherapeutic drug oxaliplatin reactivates latent HIV-1 in this cell line model of HIV-1 latency. • Reactivation is synergized when oxaliplatin is used in combination with valproic acid. • Oxaliplatin reactivates latent HIV-1 through activation of NF-kB and does not induce T cell activation. - Abstract: Reactivation of latent HIV-1 is a promising strategy for the clearance of the viral reservoirs. Because of the limitations of current agents, identification of new latency activators is urgently required. Using an established model of HIV-1 latency, we examined the effect of Oxaliplatin on latent HIV-1 reactivation. We showed that Oxaliplatin, alonemore » or in combination with valproic acid (VPA), was able to reactivate HIV-1 without inducing global T cell activation. We also provided evidence that Oxaliplatin reactivated HIV-1 expression by inducing nuclear factor kappa B (NF-κB) nuclear translocation. Our results indicated that Oxaliplatin could be a potential drug candidate for anti-latency therapies.« less
NF-κB-dependent transcriptional upregulation of cyclin D1 exerts cytoprotection against hypoxic injury upon EGFR activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Chen, Zhi-Dong; Xu, Liang; Tang, Kan-Kai

Apoptosis of neural cells is one of the main pathological features in hypoxic/ischemic brain injury. Nuclear factor-κB (NF-κB) might be a potential therapeutic target for hypoxic/ischemic brain injury since NF-κB has been found to be inactivated after hypoxia exposure, yet the underlying molecular mechanisms of NF-κB inactivation are largely unknown. Here we report that epidermal growth factor receptor (EGFR) activation prevents neuron-like PC12 cells apoptosis in response to hypoxia via restoring NF-κB-dependent transcriptional upregulation of cyclin D1. Functionally, EGFR activation by EGF stimulation mitigates hypoxia-induced PC12 cells apoptosis in both dose- and time-dependent manner. Of note, EGFR activation elevates IKKβmore » phosphorylation, increases IκBα ubiquitination, promotes P65 nuclear translocation and recruitment at cyclin D1 gene promoter as well as upregulates cyclin D1 expression. EGFR activation also abrogates the decrease of IKKβ phosphorylation, reduction of IκBα ubiquitination, blockade of P65 nuclear translocation and recruitment at cyclin D1 gene promoter as well as downregulation of cyclin D1 expression induced by hypoxia. Furthermore, NF-κB-dependent upregulation of cyclin D1 is instrumental for the EGFR-mediated cytoprotection against hypoxic apoptosis. In addition, the dephosphorylation of EGFR induced by either EGF siRNA transfection or anti-HB-EGF neutralization antibody treatment enhances hypoxic cytotoxicity, which are attenuated by EGF administration. Our results highlight the essential role of NF-κB-dependent transcriptional upregulation of cyclin D1 in EGFR-mediated cytoprotective effects under hypoxic preconditioning and support further investigation of EGF in clinical trials of patients with hypoxic/ischemic brain injury. - Highlights: • EGFR activation significantly decreases hypoxia-induced PC12 cells injury. • EGFR activation abrogates the transcriptional repression of cyclin D1 induced by hypoxia in a
Glaucarubinone sensitizes KB cells to paclitaxel by inhibiting ABC transporters via ROS-dependent and p53-mediated activation of apoptotic signaling pathways

PubMed Central

Karthikeyan, Subburayan; Hoti, Sugeerappa Laxmanappa; Nazeer, Yasin; Hegde, Harsha Vasudev

2016-01-01

Multidrug resistance (MDR) is considered to be the major contributor to failure of chemotherapy in oral squamous cell carcinoma (SCC). This study was aimed to explore the effects and mechanisms of glaucarubinone (GLU), one of the major quassinoids from Simarouba glauca DC, in potentiating cytotoxicity of paclitaxel (PTX), an anticancer drug in KB cells. Our data showed that the administration of GLU pre-treatment significantly enhanced PTX anti-proliferative effect in ABCB1 over-expressing KB cells. The Rh 123 drug efflux studies revealed that there was a significant transport function inhibition by GLU-PTX treatment. Interestingly, it was also found that this enhanced anticancer efficacy of GLU was associated with PTX-induced cell arrest in the G2/M phase of cell cycle. Further, the combined treatment of GLU-PTX had significant decrease in the expression levels of P-gp, MRPs, and BCRP in resistant KB cells at both mRNA and protein levels. Furthermore, the combination treatments showed significant reactive oxygen species (ROS) production, chromatin condensation and reduced mitochondrial membrane potential in resistant KB cells. The results from DNA fragmentation analysis also demonstrated the GLU induced apoptosis in KB cells and its synergy with PTX. Importantly, GLU and/or PTX triggered apoptosis through the activation of pro-apoptotic proteins such as p53, Bax, and caspase-9. Our findings demonstrated for the first time that GLU causes cell death in human oral cancer cells via the ROS-dependent suppression of MDR transporters and p53-mediated activation of the intrinsic mitochondrial pathway of apoptosis. Additionally, the present study also focussed on investigation of the protective effect of GLU and combination drugs in human normal blood lymphocytes. Normal blood lymphocytes assay indicated that GLU is able to induce selective toxicity in cancer cells and in silico molecular docking studies support the choice of GLU as ABC inhibitor to enhance PTX efficacy
Characterizing Myeloid Cell Activation in NF1 Vasculopathy

DTIC Science & Technology

2016-07-01

mechanistic insight and develop therapeutic targets for the prevention/treatment of neurofibromatosis type 1 (NF1) related cardiovascular disease ...therapeutic targets for the prevention/treatment of neurofibromatosis type 1 (NF1) related cardiovascular diseases . Cardiovascular disease affects upwards...superoxide; macrophages; monocytes; arteries; cardiovascular disease Major Goals and Accomplishments: Significant progress toward accomplishing
Characterization of the 101-Kilobase-Pair Megaplasmid pKB1, Isolated from the Rubber-Degrading Bacterium Gordonia westfalica Kb1

PubMed Central

Bröker, Daniel; Arenskötter, Matthias; Legatzki, Antje; Nies, Dietrich H.; Steinbüchel, Alexander

2004-01-01

The complete sequence of the circular 101,016-bp megaplasmid pKB1 from the cis-1,4-polyisoprene-degrading bacterium Gordonia westfalica Kb1, which represents the first described extrachromosomal DNA of a member of this genus, was determined. Plasmid pKB1 harbors 105 open reading frames. The predicted products of 46 of these are significantly related to proteins of known function. Plasmid pKB1 is organized into three functional regions that are flanked by insertion sequence (IS) elements: (i) a replication and putative partitioning region, (ii) a putative metabolic region, and (iii) a large putative conjugative transfer region, which is interrupted by an additional IS element. Southern hybridization experiments revealed the presence of another copy of this conjugational transfer region on the bacterial chromosome. The origin of replication (oriV) of pKB1 was identified and used for construction of Escherichia coli-Gordonia shuttle vectors, which was also suitable for several other Gordonia species and related genera. The metabolic region included the heavy-metal resistance gene cadA, encoding a P-type ATPase. Expression of cadA in E. coli mediated resistance to cadmium, but not to zinc, and decreased the cellular content of cadmium in this host. When G. westfalica strain Kb1 was cured of plasmid pKB1, the resulting derivative strains exhibited slightly decreased cadmium resistance. Furthermore, they had lost the ability to use isoprene rubber as a sole source of carbon and energy, suggesting that genes essential for rubber degradation are encoded by pKB1. PMID:14679241
Influence of exogenous lactoferrin on the oxidant/antioxidant balance and molecular profile of hormone receptor-positive and -negative human breast cancer cells in vitro.

PubMed

Zalutskii, I V; Lukianova, N Y; Storchai, D M; Burlaka, A P; Shvets, Y V; Borikun, T V; Todor, I M; Lukashevich, V S; Rudnichenko, Y A; Chekhun, V F

2017-07-01

To investigate the mechanisms of cytotoxic activity and pro-/antioxidant effect of lactoferrin on hormone receptor-positive and receptor-negative breast cancer cells in vitro. The study was performed on receptor-positive (MCF-7, T47D) and receptor-negative (MDA-MB-231, MDA-MB-468) human breast cancer cell lines. Immunocytochemical staining, flow cytometry, low-temperature electron paramagnetic resonance, and the Comet assay were used. Upon treatment with lactoferrin, the increased levels of reactive oxygen species (ROS) (p < 0.05), NO generation rate by inducible NO-synthase (p < 0.05) and the level of "free" iron (p < 0.05) were observed. Moreover, the effects of lactoferrin were more pronounced in receptor-negative MDA-MB-231 and MDA-MB-468 cells. These changes resulted in increased expression of proapoptotic Bax protein (p < 0.05), reduced expression of the antiapoptotic Bcl-2 protein (p < 0.05) and level of not-oxidized mitochondrial cardiolipin (1.4-1.7-fold, p < 0.05). This, in turn, caused an increase in the percentage of apoptotic cells (by 14-24%, p < 0.05). Cytotoxic effects of lactoferrin were accompanied by an increase in the percentage of DNA in the comet tail and blocking cell cycle at G2/M phase, especially in receptor-negative cell lines. The study showed that exogenous lactoferrin causes a violation of an antioxidant balance by increasing the level of ROS, "free" iron and NO generation rate, resalting in the blocking of cell cycle at G2/M-phase and apoptosis of malignant cells.
A heterodimer comprised of two bovine lactoferrin antimicrobial peptides exhibits powerful bactericidal activity against Burkholderia pseudomallei.

PubMed

Puknun, Aekkalak; Bolscher, Jan G M; Nazmi, Kamran; Veerman, Enno C I; Tungpradabkul, Sumalee; Wongratanacheewin, Surasakdi; Kanthawong, Sakawrat; Taweechaisupapong, Suwimol

2013-07-01

Melioidosis is a severe infectious disease that is endemic in Southeast Asia and Northern Australia. Burkholderia pseudomallei, the causative agent of this disease, has developed resistance to an increasing list of antibiotics, demanding a search for novel agents. Lactoferricin and lactoferrampin are two antimicrobial domains of lactoferrin with a broad spectrum of antimicrobial activity. A hybrid peptide (LFchimera) containing lactoferrampin (LFampin265-284) and a part of lactoferricin (LFcin17-30) has strikingly higher antimicrobial activities compared to the individual peptides. In this study, the antimicrobial activities of this chimeric construct (LFchimera1), as well as of another one containing LFcin17-30 and LFampin268-284, a shorter fragment of LFampin265-284 (LFchimera2), and the constituent peptides were tested against 7 isolates of B. pseudomallei and compared to the preferential antibiotic ceftazidime (CAZ). All isolates including B. pseudomallei 979b shown to be resistant to CAZ, at a density of 10(5) CFU/ml, could be killed by 5-10 μM of LFchimera1 within 2 h, while the other peptides as well as the antibiotic CAZ only inhibited the B. pseudomallei strains resulting in an overgrowth in 24 h. These data indicate that LFchimera1 could be considered for development of therapeutic agents against B. pseudomallei.
NF-κB and the link between inflammation and cancer.

PubMed

DiDonato, Joseph A; Mercurio, Frank; Karin, Michael

2012-03-01

The nuclear factor-κB (NF-κB) transcription factor family has been considered the central mediator of the inflammatory process and a key participant in innate and adaptive immune responses. Coincident with the molecular cloning of NF-κB/RelA and identification of its kinship to the v-Rel oncogene, it was anticipated that NF-κB itself would be involved in cancer development. Oncogenic activating mutations in NF-κB genes are rare and have been identified only in some lymphoid malignancies, while most NF-κB activating mutations in lymphoid malignancies occur in upstream signaling components that feed into NF-κB. NF-κB activation is also prevalent in carcinomas, in which NF-κB activation is mainly driven by inflammatory cytokines within the tumor microenvironment. Importantly, however, in all malignancies, NF-κB acts in a cell type-specific manner: activating survival genes within cancer cells and inflammation-promoting genes in components of the tumor microenvironment. Yet, the complex biological functions of NF-κB have made its therapeutic targeting a challenge. © 2012 John Wiley & Sons A/S.
Sulforaphane inhibits phorbol ester-stimulated IKK-NF-κB signaling and COX-2 expression in human mammary epithelial cells by targeting NF-κB activating kinase and ERK.

PubMed

Kim, Ha-Na; Kim, Do-Hee; Kim, Eun-Hee; Lee, Mee-Hyun; Kundu, Joydeb Kumar; Na, Hye-Kyung; Cha, Young-Nam; Surh, Young-Joon

2014-08-28

Sulforaphane, an isothiocyanate present in cruciferous vegetables, has been reported to possess anti-inflammatory and cancer chemopreventive properties. However, the molecular mechanisms by which sulforaphane suppresses inflammation and carcinogenesis are yet to be fully elucidated. Since the aberrant expression of cyclooxygenase-2 (COX-2) links inflammation and cancer, the present study was aimed to elucidate the mechanisms by which sulforaphane modulates COX-2 overexpression in human mammary epithelial (MCF-10A) cells stimulated with a prototypic tumor promoter 12-O-tetradecanoylphorbol-13-acetate (TPA). Treatment of MCF-10A cells with sulforaphane significantly inhibited TPA-induced expression of COX-2 protein and its mRNA transcript. Transient transfection of cells with deletion mutant constructs of COX-2 promoter revealed that the transcription factor nuclear factor-kappaB (NF-κB) plays a key role in TPA-induced COX-2 expression in MCF-10A cells. Pretreatment with sulforaphane significantly attenuated nuclear localization, DNA binding and the transcriptional activity of NF-κB through inhibition of phosphorylation and subsequent degradation of IκBα in MCF-10A cells stimulated with TPA. Sulforaphane also attenuated TPA-induced activation of IκB kinases (IKK), NF-κB-activating kinase (NAK) and extracellular signal-regulated kinase-1/2 (ERK1/2). Pharmacological inhibition of IKK or transient transfection of cells with dominant-negative mutant forms of this kinase abrogated TPA-induced NF-κB activation and COX-2 expression. In addition, the blockade of ERK1/2 activation negated the catalytic activity of IKKα, but not that of IKKβ, whereas silencing NAK by specific siRNA abrogated the IKKβ activity in TPA-treated cells. Taken together, sulforaphane inhibits TPA-induced NF-κB activation and COX-2 expression in MCF-10A cells by blocking two distinct signaling pathways mediated by ERK1/2-IKKα and NAK-IKKβ. Copyright © 2014 Elsevier Ireland Ltd. All rights
Extracellular HSP27 acts as a signaling molecule to activate NF-κB in macrophages.

PubMed

Salari, Samira; Seibert, Tara; Chen, Yong-Xiang; Hu, Tieqiang; Shi, Chunhua; Zhao, Xiaoling; Cuerrier, Charles M; Raizman, Joshua E; O'Brien, Edward R

2013-01-01

Heat shock protein 27 (HSP27) shows attenuated expression in human coronary arteries as the extent of atherosclerosis progresses. In mice, overexpression of HSP27 reduces atherogenesis, yet the precise mechanism(s) are incompletely understood. Inflammation plays a central role in atherogenesis, and of particular interest is the balance of pro- and anti-inflammatory factors produced by macrophages. As nuclear factor-kappa B (NF-κB) is a key immune signaling modulator in atherogenesis, and macrophages are known to secrete HSP27, we sought to determine if recombinant HSP27 (rHSP27) alters NF-κB signaling in macrophages. Treatment of THP-1 macrophages with rHSP27 resulted in the degradation of an inhibitor of NF-κB, IκBα, nuclear translocation of the NF-κB p65 subunit, and increased NF-κB transcriptional activity. Treatment of THP-1 macrophages with rHSP27 yielded increased expression of a variety of genes, including the pro-inflammatory factors, IL-1β, and TNF-α. However, rHSP27 also increased the expression of the anti-inflammatory factors IL-10 and GM-CSF both at the mRNA and protein levels. Our study suggests that in macrophages, activation of NF-κB signaling by rHSP27 is associated with upregulated expression and secretion of key pro- and anti-inflammatory cytokines. Moreover, we surmise that it is the balance in expression of these mediators and antagonists of inflammation, and hence atherogenesis, that yields a favorable net effect of HSP27 on the vessel wall.

Exosomes derived from gastric cancer cells activate NF-κB pathway in macrophages to promote cancer progression.

PubMed

Wu, Lijun; Zhang, Xu; Zhang, Bin; Shi, Hui; Yuan, Xiao; Sun, Yaoxiang; Pan, Zhaoji; Qian, Hui; Xu, Wenrong

2016-09-01

Exosomes are nano-sized membrane vesicles secreted by both normal and cancer cells. Emerging evidence indicates that cancer cells derived exosomes contribute to cancer progression through the modulation of tumor microenvironment. However, the effects of exosomes derived from gastric cancer cells on macrophages are not well understood. In this study, we investigated the biological role of gastric cancer cells derived exosomes in the activation of macrophages. We demonstrated that gastric cancer cells derived exosomes activated macrophages to express increased levels of proinflammatory factors, which in turn promoted tumor cell proliferation and migration. In addition, gastric cancer cells derived exosomes remarkably upregulated the phosphorylation of NF-κB in macrophages. Inhibiting the activation of NF-κB reversed the upregulation of proinflammatory factors in macrophages and blocked their promoting effects on gastric cancer cells. Moreover, we found that gastric cancer cells derived exosomes could also activate macrophages from human peripheral blood monocytes through the activation of NF-κB. In conclusion, our results suggest that gastric cancer cells derived exosomes stimulate the activation of NF-κB pathway in macrophages to promote cancer progression, which provides a potential therapeutic approach for gastric cancer by interfering with the interaction between exosomes and macrophages in tumor microenvironment.
Detection of recombinant human lactoferrin and lysozyme produced in a bitransgenic cow.

PubMed

Kaiser, Germán G; Mucci, Nicolás C; González, Vega; Sánchez, Lourdes; Parrón, José A; Pérez, María D; Calvo, Miguel; Aller, Juan F; Hozbor, Federico A; Mutto, Adrián A

2017-03-01

Lactoferrin and lysozyme are 2 glycoproteins with great antimicrobial activity, being part of the nonspecific defensive system of human milk, though their use in commercial products is difficult because human milk is a limited source. Therefore, many investigations have been carried out to produce those proteins in biological systems, such as bacteria, yeasts, or plants. Mammals seem to be more suitable as expression systems for human proteins, however, especially for those that are glycosylated. In the present study, we developed a bicistronic commercial vector containing a goat β-casein promoter and an internal ribosome entry site fragment between the human lactoferrin and human lysozyme genes to allow the introduction of both genes into bovine adult fibroblasts in a single transfection. Embryos were obtained by somatic cell nuclear transfer, and, after 6 transferences to recipients, 3 pregnancies and 1 viable bitransgenic calf were obtained. The presence of the vector was confirmed by fluorescent in situ hybridization of skin cells. At 13 mo of life and after artificial induction of lactation, both recombinant proteins were found in the colostrum and milk of the bitransgenic calf. Human lactoferrin concentration in the colostrum was 0.0098 mg/mL and that in milk was 0.011 mg/mL; human lysozyme concentration in the colostrum was 0.0022 mg/mL and that in milk was 0.0024 mg/mL. The molar concentration of both human proteins revealed no differences in protein production of the internal ribosome entry site upstream and downstream protein. The enzymatic activity of lysozyme in the transgenic milk was comparable to that of human milk, being 6 and 10 times higher than that of bovine lysozyme present in milk. This work represents an important step to obtain multiple proteins or enhance single protein production by using animal pharming and fewer regulatory and antibiotic-resistant foreign sequences, allowing the design of humanized milk with added biological value for
Immunosuppressive effects of fisetin in ovalbumin-induced asthma through inhibition of NF-κB activity.

PubMed

Wu, Mei-Yao; Hung, Shih-Kai; Fu, Shu-Ling

2011-10-12

Fisetin, a flavonoid compound commonly present in fruits and vegetables, can exert anti-inflammation activities via inhibition of the NF-κB-signaling pathway. This study aims to evaluate the antiasthma activity of fisetin and investigate its possible molecular mechanisms. We found that fisetin attenuated lung inflammation, goblet cell hyperplasia, and airway hyperresponsiveness in ovalbumin-induced asthma and decreased eosinophils and lymphocytes in bronchoalveolar lavage fluid. Fisetin treatment reduced expression of the key initiators of allergic airway inflammation (eotaxin-1 and TSLP), Th2-associated cytokines (IL-4, IL-5, and IL-13) in lungs, and Th2-predominant transcription factor GATA-3 and cytokines in thoracic lymph node cells and splenocytes. Notably, fisetin treatment impaired NF-κB activation in OVA-stimulated lung tissues and TNF-α-stimulated bronchial epithelial cells. Collectively, this study demonstrated the beneficial effect of fisetin in the amelioration of asthmatic phenotypes. The antiasthma activity of fisetin is associated with reduction of Th2 responses as well as suppression of NF-κB and its downstream chemokines.
Curcumin Modulates the Radiosensitivity of Colorectal Cancer Cells by Suppressing Constitutive and Inducible NF-{kappa}B Activity

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sandur, Santosh K.; Deorukhkar, Amit; Pandey, Manoj K.

2009-10-01

Purpose: Radiation therapy is an integral part of the preoperative treatment of rectal cancers. However, only a minority of patients achieve a complete pathologic response to therapy because of resistance of these tumors to radiation therapy. This resistance may be mediated by constitutively active pro-survival signaling pathways or by inducible/acquired mechanisms in response to radiation therapy. Simultaneous inhibition of these pathways can sensitize these tumors to radiation therapy. Methods and Materials: Human colorectal cancer cells were exposed to clinically relevant doses of gamma rays, and the mechanism of their radioresistance was investigated. We characterized the transcription factor nuclear factor-{kappa}B (NF-{kappa}B)more » activation as a mechanism of inducible radioresistance in colorectal cancer and used curcumin, the active ingredient in the yellow spice turmeric, to overcome this resistance. Results: Curcumin inhibited the proliferation and the post-irradiation clonogenic survival of multiple colorectal cancer cell lines. Radiation stimulated NF-{kappa}B activity in a dose- and time-dependent manner, whereas curcumin suppressed this radiation-induced NF-{kappa}B activation via inhibition of radiation-induced phosphorylation and degradation of inhibitor of {kappa}B alpha, inhibition of inhibitor of {kappa}B kinase activity, and inhibition of Akt phosphorylation. Curcumin also suppressed NF-{kappa}B-regulated gene products (Bcl-2, Bcl-x{sub L}, inhibitor of apoptosis protein-2, cyclooxygenase-2, and cyclin D1). Conclusions: Our results suggest that transient inducible NF-{kappa}B activation provides a prosurvival response to radiation that may account for development of radioresistance. Curcumin blocks this signaling pathway and potentiates the antitumor effects of radiation therapy.« less
Loss of NF1 in cutaneous melanoma is associated with RAS activation and MEK dependence.

PubMed

Nissan, Moriah H; Pratilas, Christine A; Jones, Alexis M; Ramirez, Ricardo; Won, Helen; Liu, Cailian; Tiwari, Shakuntala; Kong, Li; Hanrahan, Aphrothiti J; Yao, Zhan; Merghoub, Taha; Ribas, Antoni; Chapman, Paul B; Yaeger, Rona; Taylor, Barry S; Schultz, Nikolaus; Berger, Michael F; Rosen, Neal; Solit, David B

2014-04-15

Melanoma is a disease characterized by lesions that activate ERK. Although 70% of cutaneous melanomas harbor activating mutations in the BRAF and NRAS genes, the alterations that drive tumor progression in the remaining 30% are largely undefined. Vemurafenib, a selective inhibitor of RAF kinases, has clinical utility restricted to BRAF-mutant tumors. MEK inhibitors, which have shown clinical activity in NRAS-mutant melanoma, may be effective in other ERK pathway-dependent settings. Here, we investigated a panel of melanoma cell lines wild type for BRAF and NRAS to determine the genetic alteration driving their transformation and their dependence on ERK signaling in order to elucidate a candidate set for MEK inhibitor treatment. A cohort of the BRAF/RAS wild type cell lines with high levels of RAS-GTP had loss of NF1, a RAS GTPase activating protein. In these cell lines, the MEK inhibitor PD0325901 inhibited ERK phosphorylation, but also relieved feedback inhibition of RAS, resulting in induction of pMEK and a rapid rebound in ERK signaling. In contrast, the MEK inhibitor trametinib impaired the adaptive response of cells to ERK inhibition, leading to sustained suppression of ERK signaling and significant antitumor effects. Notably, alterations in NF1 frequently co-occurred with RAS and BRAF alterations in melanoma. In the setting of BRAF(V600E), NF1 loss abrogated negative feedback on RAS activation, resulting in elevated activation of RAS-GTP and resistance to RAF, but not MEK, inhibitors. We conclude that loss of NF1 is common in cutaneous melanoma and is associated with RAS activation, MEK-dependence, and resistance to RAF inhibition. ©2014 AACR.
Identification of a gene encoding a membrane-anchored toll-like receptor 5 (TLR5M) in Oplegnathus fasciatus that responds to flagellin challenge and activates NF-κB.

PubMed

Umasuthan, Navaneethaiyer; Bathige, S D N K; Thulasitha, William Shanthakumar; Jayasooriya, R G P T; Shin, Younhee; Lee, Jehee

2017-03-01

Toll-like receptor 5 (TLR5) recognizes bacterial flagellin and induces the downstream signaling through the myeloid differentiation primary response gene 88 (MyD88) protein to produce proinflammatory cytokines. In this study, we describe a TLR5 membrane form (OfTLR5M) and its adaptor protein MyD88 (OfMyD88) in rock bream, Oplegnathus fasciatus. Both Oftlr5m (6.7 kb) and Ofmyd88 (3.7 kb) genes displayed a quinquepartite structure with five exons and four introns. Protein structure of OfTLR5M revealed the conventional architecture of TLRs featured by an extracellular domain with 22 leucine rich repeats (LRR), a transmembrane domain and an endodomain with TIR motif. Primary OfTLR5M sequence shared a higher homology with teleost TLR5M. The evolutional analysis confirmed that TLR5 identified in the current study is a membrane receptor and the data further suggested the co-evolution of the membrane-anchored and soluble forms of TLR5 in teleosts. Inter-lineage comparison of gene structures in vertebrates indicated that the tlr5m gene has evolved with extensive rearrangement; whereas, the myd88 gene has maintained a stable structure throughout the evolution. Inspection of 5' flanking region of these genes disclosed the presence of several transcription factor binding sites including NF-κB. Quantitative real-time PCR (qPCR) detected Oftlr5m mRNA in eleven tissues with the highest abundance in liver. In vivo flagellin administration strongly induced the transcripts of both Oftlr5m and Ofmyd88 in gills and head kidney tissues suggesting their ligand-mediated upregulation. In a luciferase assay, HEK293T cells transiently transfected with Oftlr5m and Ofmyd88 demonstrated a higher NF-κB activity than the mock control, and the luciferase activity was intensified when cells were stimulated with flagellin. Collectively, our study represents the genomic, evolutional, expressional and functional insights into a receptor and adaptor molecules of teleost origin that are involved
Glycyrrhetinic acid inhibits ICAM-1 expression via blocking JNK and NF-κB pathways in TNF-α-activated endothelial cells

PubMed Central

Chang, Ying-ling; Chen, Chien-lin; Kuo, Chao-Lin; Chen, Bor-chyuan; You, Jyh-sheng

2010-01-01

Aim: To investigate the effects of glycyrrhetinic acid (GA), an active component extracted from the root of Glycyrrhizae glabra, on the expression of intercellular adhesion molecule-1 (ICAM-1) in tumor necrosis factor-α (TNF-α)-activated human umbilical vein endothelial cells (HUVEC). Methods: ICAM-1 mRNA and protein levels were detected using RT-PCR and cell enzyme-linked immunosorbent assays. The adherence of human monocytic THP-1 cells labeled with [3H]thymidine to HUVEC was determined by counting radioactivity with a scintillation counter. The activation of mitogen-activated protein kinases as well as the degradation of IκB and nuclear factor-κB (NF-κB) or phospho-c-Jun in the nucleus were detected by western blots. NF-κB binding activity was detected using electrophoretic mobility shift assay. Results: GA (50 and 100 μmol/L) significantly inhibits TNF-α-induced ICAM-1 mRNA and protein expressions, as well as THP-1 cell adhesiveness in HUVEC. GA selectively inhibited TNF-α-activated signal pathway of c-Jun N-terminal kinase (JNK), without affecting extracellular signal-regulated kinase 1/2 and p38. Furthermore, GA apparently inhibited IκB/NF-κB signaling system by preventing IκB degradation, NF-κB translocation, and NF-κB/DNA binding activity. Finally, pretreatment with GA or the inhibitors of NF-κB, JNK, and p38 reduced the ICAM-1 protein expression induced by TNF-α. Conclusion: GA inhibits TNF-α-stimulated ICAM-1 expression, leading to a decrease in adherent monocytes to HUVEC. This inhibition is attributed to GA interruption of both JNK/c-Jun and IκB/NF-κB signaling pathways, which decrease activator protein-1 (AP-1) and NF-κB mediated ICAM-1 expressions. The results suggest that GA may provide a beneficial effect in treating vascular diseases associated with inflammation, such as atherosclerosis. PMID:20418897
Phytochemicals and botanical extracts regulate NF-κB and Nrf2/ARE reporter activities in DI TNC1 astrocytes

PubMed Central

Ajit, Deepa; Simonyi, Agnes; Li, Runting; Chen, Zihong; Hannink, Mark; Fritsche, Kevin L.; Mossine, Valeri V.; Smith, Robert E.; Dobbs, Thomas K.; Luo, Rensheng; Folk, William R.; Gu, Zezong; Lubahn, Dennis B.; Weisman, Gary A.; Sun, Grace Y.

2016-01-01

The increase in oxidative stress and inflammatory responses associated with neurodegenerative diseases has drawn considerable attention towards understanding the transcriptional signaling pathways involving NF-κB (nuclear factor kappa-light-chain-enhancer of activated B cells) and Nrf2 (Nuclear Factor Erythroid 2-like 2). Our recent studies with immortalized murine microglial cells (BV-2) demonstrated effects of botanical polyphenols to inhibit lipopolysaccharide (LPS)-induced nitric oxide (NO) and enhance Nrf2-mediated antioxidant responses (Sun et al., 2015). In this study, an immortalized rat astrocyte (DI TNC1) cell line expressing a luciferase reporter driven by the NF-κB or the Nrf2/Antioxidant Response Element (ARE) promoter was used to assess regulation of these two pathways by phytochemiscals such as quercetin, rutin, cyanidin, cyanidin-3-O-glucoside, as well as botanical extracts from Withania somnifera (Ashwagandha), Sutherlandia frutescens (Sutherlandia) and Euterpe oleracea (Açaí). Quercetin effectively inhibited LPS-induced NF-κB reporter activity and stimulated Nrf2/ARE reporter activity in DI TNC1 astrocytes. Cyanidin and the glycosides showed similar effects but only at much higher concentrations. All three botanical extracts effectively inhibited LPS-induced NF-κB reporter activity. These extracts were capable of enhancing ARE activity by themselves and further enhanced ARE activity in the presence of LPS. Quercetin and botanical extracts induced Nrf2 and HO-1 protein expression. Interestingly, Ashwagandha extract was more active in inducing Nrf2 and HO-1 expression in DI TNC1 astrocytes as compared to Sutherlandia and Açaí extracts. In summary, this study demonstrated NF-kB and Nrf2/ARE promotor activities in DI TNC1 astrocytes, and further showed differences in ability for specific botanical polyphenols and extracts to down-regulate LPS-induced NF-kB and up-regulate the NRF2/ARE activities in these cells. PMID:27166148
Activation of the NF-κB pathway by the STAT3 inhibitor JSI-124 in human glioblastoma cells

PubMed Central

McFarland, Braden C.; Gray, G. Kenneth; Nozell, Susan E.; Hong, Suk W.; Benveniste, Etty N.

2013-01-01

Glioblastoma tumors are characterized by their invasiveness and resistance to therapies. The transcription factor STAT3 was recently identified as a master transcriptional regulator in the mesenchymal subtype of GBM, which has generated an increased interest in targeting STAT3. We have evaluated more closely the mechanism of action of one particular STAT3 inhibitor, JSI-124 (cucurbitacin I). In this study, we confirmed that JSI-124 inhibits both constitutive and stimulus-induced JAK2 and STAT3 phosphorylation, and decreases cell proliferation while inducing apoptosis in cultured GBM cells. However, we discovered that prior to the inhibition of STAT3, JSI-124 activates the NF-κB pathway, via NF-κB p65 phosphorylation and nuclear translocation. In addition, JSI-124 treatment induces the expression of IL-6, IL-8 and SOCS3 mRNA, which leads to a corresponding increase in IL-6, IL-8 and SOCS3 protein expression. Moreover, the NF-κB driven SOCS3 expression acts as a negative regulator of STAT3, abrogating any subsequent STAT3 activation and provides a mechanism of STAT3 inhibition following JSI-124 treatment. Chromatin immunoprecipitation analysis confirms that NF-κB p65 in addition to other activating co-factors are found at the promoters of IL-6, IL-8 and SOCS3, following JSI-124 treatment. Using pharmacological inhibition of NF-κB and inducible knockdown of NF-κB p65, we found that JSI-124-induced expression of IL-6, IL-8 and SOCS3 was significantly inhibited, demonstrating an NF-κB dependent mechanism. Our data indicate that although JSI-124 may demonstrate potential anti-tumor effects through inhibition of STAT3, other off-target pro-inflammatory pathways are activated, emphasizing that more careful and thorough pre-clinical investigations must be implemented to prevent potential harmful effects. PMID:23386688
Expression of lactoferrin receptors is increased in the mesencephalon of patients with Parkinson disease.

PubMed Central

Faucheux, B A; Nillesse, N; Damier, P; Spik, G; Mouatt-Prigent, A; Pierce, A; Leveugle, B; Kubis, N; Hauw, J J; Agid, Y

1995-01-01

The degeneration of nigral dopaminergic neurons in Parkinson disease is believed to be associated with oxidative stress. Since iron levels are increased in the substantia nigra of parkinsonian patients and this metal catalyzes the formation of free radicals, it may be involved in the mechanisms of nerve cell death. The cause of nigral iron increase is not understood. Iron acquisition by neurons may occur from iron-transferrin complexes with a direct interaction with specific membrane receptors, but recent results have shown a low density of transferrin receptors in the substantia nigra. To investigate whether neuronal death in Parkinson disease may be associated with changes in a pathway supplementary to that of transferrin, lactoferrin (lactotransferrin) receptor expression was studied in the mesencephalon. In this report we present evidence from immunohistochemical staining of postmortem human brain tissue that lactoferrin receptors are localized on neurons (perikarya, dendrites, axons), cerebral microvasculature, and, in some cases, glial cells. In parkinsonian patients, lactoferrin receptor immunoreactivity on neurons and microvessels was increased and more pronounced in those regions of the mesencephalon where the loss of dopaminergic neurons is severe. Moreover, in the substantia nigra, the intensity of immunoreactivity on neurons and microvessels was higher for patients with higher nigral dopaminergic loss. These data suggest that lactoferrin receptors on vulnerable neurons may increase intraneuronal iron levels and contribute to the degeneration of nigral dopaminergic neurons in Parkinson disease. Images Fig. 1 Fig. 2 PMID:7568181
TRAF2-binding BIR1 domain of c-IAP2/MALT1 fusion protein is essential for activation of NF-kappaB.

PubMed

Garrison, J B; Samuel, T; Reed, J C

2009-04-02

Marginal zone mucosa-associated lymphoid tissue (MALT) B-cell lymphoma is the most common extranodal non-Hodgkin lymphoma. The t(11;18)(q21;q21) translocation occurs frequently in MALT lymphomas and creates a chimeric NF-kappaB-activating protein containing the baculoviral IAP repeat (BIR) domains of c-IAP2 (inhibitor of apoptosis protein 2) fused with portions of the MALT1 protein. The BIR1 domain of c-IAP2 interacts directly with TRAF2 (TNFalpha-receptor-associated factor-2), but its role in NF-kappaB activation is still unclear. Here, we investigated the role of TRAF2 in c-IAP2/MALT1-induced NF-kappaB activation. We show the BIR1 domain of c-IAP2 is essential for NF-kappaB activation, whereas BIR2 and BIR3 domains are not. Studies of c-IAP2/MALT1 BIR1 mutant (E47A/R48A) that fails to activate NF-kappaB showed loss of TRAF2 binding, but retention of TRAF6 binding, suggesting that interaction of c-IAP2/MALT1 with TRAF6 is insufficient for NF-kappaB induction. In addition, a dominant-negative TRAF2 mutant or downregulation of TRAF2 achieved by small interfering RNA inhibited NF-kappaB activation by c-IAP2/MALT1 showing that TRAF2 is indispensable. Comparisons of the bioactivity of intact c-IAP2/MALT1 oncoprotein and BIR1 E47A/R48A c-IAP2/MALT1 mutant that cannot bind TRAF2 in a lymphoid cell line provided evidence that TRAF2 interaction is critical for c-IAP2/MALT1-mediated increases in the NF-kappaB activity, increased expression of endogenous NF-kappaB target genes (c-FLIP, TRAF1), and resistance to apoptosis.
Benefits of Lactoferrin, Osteopontin and Milk Fat Globule Membranes for Infants.

PubMed

Demmelmair, Hans; Prell, Christine; Timby, Niklas; Lönnerdal, Bo

2017-07-28

The provision of essential and non-essential amino acids for breast-fed infants is the major function of milk proteins. In addition, breast-fed infants might benefit from bioactivities of milk proteins, which are exhibited in the intestine during the digestive phase and by absorption of intact proteins or derived peptides. For lactoferrin, osteopontin and milk fat globule membrane proteins/lipids, which have not until recently been included in substantial amounts in infant formulas, in vitro experiments and animal models provide a convincing base of evidence for bioactivities, which contribute to the protection of the infant from pathogens, improve nutrient absorption, support the development of the immune system and provide components for optimal neurodevelopment. Technologies have become available to obtain these compounds from cow´s milk and the bovine compounds also exhibit bioactivities in humans. Randomized clinical trials with experimental infant formulas incorporating lactoferrin, osteopontin, or milk fat globule membranes have already provided some evidence for clinical benefits. This review aims to compare findings from laboratory and animal experiments with outcomes of clinical studies. There is good justification from basic science and there are promising results from clinical studies for beneficial effects of lactoferrin, osteopontin and the milk fat globule membrane complex of proteins and lipids. Further studies should ideally be adequately powered to investigate effects on clinically relevant endpoints in healthy term infants.
Benefits of Lactoferrin, Osteopontin and Milk Fat Globule Membranes for Infants

PubMed Central

Prell, Christine; Timby, Niklas; Lönnerdal, Bo

2017-01-01

The provision of essential and non-essential amino acids for breast-fed infants is the major function of milk proteins. In addition, breast-fed infants might benefit from bioactivities of milk proteins, which are exhibited in the intestine during the digestive phase and by absorption of intact proteins or derived peptides. For lactoferrin, osteopontin and milk fat globule membrane proteins/lipids, which have not until recently been included in substantial amounts in infant formulas, in vitro experiments and animal models provide a convincing base of evidence for bioactivities, which contribute to the protection of the infant from pathogens, improve nutrient absorption, support the development of the immune system and provide components for optimal neurodevelopment. Technologies have become available to obtain these compounds from cow´s milk and the bovine compounds also exhibit bioactivities in humans. Randomized clinical trials with experimental infant formulas incorporating lactoferrin, osteopontin, or milk fat globule membranes have already provided some evidence for clinical benefits. This review aims to compare findings from laboratory and animal experiments with outcomes of clinical studies. There is good justification from basic science and there are promising results from clinical studies for beneficial effects of lactoferrin, osteopontin and the milk fat globule membrane complex of proteins and lipids. Further studies should ideally be adequately powered to investigate effects on clinically relevant endpoints in healthy term infants. PMID:28788066
TRAF family member-associated NF-κB activator (TANK) is a negative regulator of osteoclastogenesis and bone formation.

PubMed

Maruyama, Kenta; Kawagoe, Tatsukata; Kondo, Takeshi; Akira, Shizuo; Takeuchi, Osamu

2012-08-17

The differentiation of bone-resorbing osteoclasts is induced by RANKL signaling, and leads to the activation of NF-κB via TRAF6 activation. TRAF family member-associated NF-κB activator (TANK) acts as a negative regulator of Toll-like receptors (TLRs) and B-cell receptor (BCR) signaling by inhibiting TRAF6 activation. Tank(-/-) mice spontaneously develop autoimmune glomerular nephritis in an IL-6-dependent manner. Despite its importance in the TCRs and BCR-activated TRAF6 inhibition, the involvement of TANK in RANKL signaling is poorly understood. Here, we report that TANK is a negative regulator of osteoclast differentiation. The expression levels of TANK mRNA and protein were up-regulated during RANKL-induced osteoclastogenesis, and overexpression of TANK in vitro led to a decrease in osteoclast formation. The in vitro osteoclastogenesis of Tank(-/-) cells was significantly increased, accompanied by increased ubiquitination of TRAF6 and enhanced canonical NF-κB activation in response to RANKL stimulation. Tank(-/-) mice showed severe trabecular bone loss, but increased cortical bone mineral density, because of enhanced bone erosion and formation. TANK mRNA expression was induced during osteoblast differentiation and Tank(-/-) osteoblasts exhibited enhaced NF-κB activation, IL-11 expression, and bone nodule formation than wild-type control cells. Finally, wild-type mice transplanted with bone marrow cells from Tank(-/-) mice showed trabecular bone loss analogous to that in Tank(-/-) mice. These findings demonstrate that TANK is critical for osteoclastogenesis by regulating NF-κB, and is also important for proper bone remodeling.
TRAF Family Member-associated NF-κB Activator (TANK) Is a Negative Regulator of Osteoclastogenesis and Bone Formation*

PubMed Central

Maruyama, Kenta; Kawagoe, Tatsukata; Kondo, Takeshi; Akira, Shizuo; Takeuchi, Osamu

2012-01-01

The differentiation of bone-resorbing osteoclasts is induced by RANKL signaling, and leads to the activation of NF-κB via TRAF6 activation. TRAF family member-associated NF-κB activator (TANK) acts as a negative regulator of Toll-like receptors (TLRs) and B-cell receptor (BCR) signaling by inhibiting TRAF6 activation. Tank−/− mice spontaneously develop autoimmune glomerular nephritis in an IL-6-dependent manner. Despite its importance in the TCRs and BCR-activated TRAF6 inhibition, the involvement of TANK in RANKL signaling is poorly understood. Here, we report that TANK is a negative regulator of osteoclast differentiation. The expression levels of TANK mRNA and protein were up-regulated during RANKL-induced osteoclastogenesis, and overexpression of TANK in vitro led to a decrease in osteoclast formation. The in vitro osteoclastogenesis of Tank−/− cells was significantly increased, accompanied by increased ubiquitination of TRAF6 and enhanced canonical NF-κB activation in response to RANKL stimulation. Tank−/− mice showed severe trabecular bone loss, but increased cortical bone mineral density, because of enhanced bone erosion and formation. TANK mRNA expression was induced during osteoblast differentiation and Tank−/− osteoblasts exhibited enhaced NF-κB activation, IL-11 expression, and bone nodule formation than wild-type control cells. Finally, wild-type mice transplanted with bone marrow cells from Tank−/− mice showed trabecular bone loss analogous to that in Tank−/− mice. These findings demonstrate that TANK is critical for osteoclastogenesis by regulating NF-κB, and is also important for proper bone remodeling. PMID:22773835
Melatonin Inhibits Androgen Receptor Splice Variant-7 (AR-V7)-Induced Nuclear Factor-Kappa B (NF-κB) Activation and NF-κB Activator-Induced AR-V7 Expression in Prostate Cancer Cells: Potential Implications for the Use of Melatonin in Castration-Resistant Prostate Cancer (CRPC) Therapy

PubMed Central

Liu, Vincent Wing Sun; Yau, Wing Lung; Tam, Chun Wai; Yao, Kwok-Ming; Shiu, Stephen Yuen Wing

2017-01-01

A major current challenge in the treatment of advanced prostate cancer, which can be initially controlled by medical or surgical castration, is the development of effective, safe, and affordable therapies against progression of the disease to the stage of castration resistance. Here, we showed that in LNCaP and 22Rv1 prostate cancer cells transiently overexpressing androgen receptor splice variant-7 (AR-V7), nuclear factor-kappa B (NF-κB) was activated and could result in up-regulated interleukin (IL)-6 gene expression, indicating a positive interaction between AR-V7 expression and activated NF-κB/IL-6 signaling in castration-resistant prostate cancer (CRPC) pathogenesis. Importantly, both AR-V7-induced NF-κB activation and IL-6 gene transcription in LNCaP and 22Rv1 cells could be inhibited by melatonin. Furthermore, stimulation of AR-V7 mRNA expression in LNCaP cells by betulinic acid, a pharmacological NF-κB activator, was reduced by melatonin treatment. Our data support the presence of bi-directional positive interactions between AR-V7 expression and NF-κB activation in CRPC pathogenesis. Of note, melatonin, by inhibiting NF-κB activation via the previously-reported MT1 receptor-mediated antiproliferative pathway, can disrupt these bi-directional positive interactions between AR-V7 and NF-κB and thereby delay the development of castration resistance in advanced prostate cancer. Apparently, this therapeutic potential of melatonin in advanced prostate cancer/CRPC management is worth translation in the clinic via combined androgen depletion and melatonin repletion. PMID:28561752
Melatonin Inhibits Androgen Receptor Splice Variant-7 (AR-V7)-Induced Nuclear Factor-Kappa B (NF-κB) Activation and NF-κB Activator-Induced AR-V7 Expression in Prostate Cancer Cells: Potential Implications for the Use of Melatonin in Castration-Resistant Prostate Cancer (CRPC) Therapy.

PubMed

Liu, Vincent Wing Sun; Yau, Wing Lung; Tam, Chun Wai; Yao, Kwok-Ming; Shiu, Stephen Yuen Wing

2017-05-31

A major current challenge in the treatment of advanced prostate cancer, which can be initially controlled by medical or surgical castration, is the development of effective, safe, and affordable therapies against progression of the disease to the stage of castration resistance. Here, we showed that in LNCaP and 22Rv1 prostate cancer cells transiently overexpressing androgen receptor splice variant-7 (AR-V7), nuclear factor-kappa B (NF-κB) was activated and could result in up-regulated interleukin ( IL ) -6 gene expression, indicating a positive interaction between AR-V7 expression and activated NF-κB/IL-6 signaling in castration-resistant prostate cancer (CRPC) pathogenesis. Importantly, both AR-V7-induced NF-κB activation and IL-6 gene transcription in LNCaP and 22Rv1 cells could be inhibited by melatonin. Furthermore, stimulation of AR-V7 mRNA expression in LNCaP cells by betulinic acid, a pharmacological NF-κB activator, was reduced by melatonin treatment. Our data support the presence of bi-directional positive interactions between AR-V7 expression and NF-κB activation in CRPC pathogenesis. Of note, melatonin, by inhibiting NF-κB activation via the previously-reported MT₁ receptor-mediated antiproliferative pathway, can disrupt these bi-directional positive interactions between AR-V7 and NF-κB and thereby delay the development of castration resistance in advanced prostate cancer. Apparently, this therapeutic potential of melatonin in advanced prostate cancer/CRPC management is worth translation in the clinic via combined androgen depletion and melatonin repletion.
High-level replication of human immunodeficiency virus in thymocytes requires NF-kappaB activation through interaction with thymic epithelial cells.

PubMed

Chêne, L; Nugeyre, M T; Barré-Sinoussi, F; Israël, N

1999-03-01

We have previously demonstrated that interaction of infected thymocytes with autologous thymic epithelial cells (TEC) is a prerequisite for a high level of human immunodeficiency virus type 1 (HIV-1) replication in thymocytes (M. Rothe, L. Chêne, M. Nugeyre, F. Barré-Sinoussi, and N. Israël, J. Virol. 72:5852-5861, 1998). We report here that this activation of HIV replication takes place at the transcriptional level through activation of the Rel/NF-kappaB transcription factors. We first demonstrate that an HIV-1 provirus (SF-2 strain) very effectively replicates in thymocytes cocultured with TEC whereas this provirus, with kappaB sites deleted, fails to replicate. We provide evidence that several NF-kappaB complexes are constitutively found in the nuclei of thymocytes either freshly isolated from the thymus or maintained in coculture with autologous or heterologous TEC. The prevalent complex is the heterodimer p50-p65. NF-kappaB activity is tightly correlated with the transcriptional activity of a long terminal repeat (LTR) of HIV-1 transfected in thymocytes. The cotransfection of this LTR with a mutated IkappaBalpha molecule formally demonstrates that LTR transactivation is regulated by members of the Rel/NF-kappaB family in thymocytes. We also showed that tumor necrosis factor (TNF) and to a lesser extent interleukin-1 (IL-1), secreted within the coculture, induce NF-kappaB activity and a correlative LTR transactivation. However IL-7, a crucial factor for thymopoiesis that is secreted mainly by TEC, is a necessary cofactor for NF-kappaB activation elicited by TNF or IL-1. Together, these data indicate that NF-kappaB activation, required for a high level of HIV replication in thymocytes, is regulated in a specific manner in the thymic microenvironment which provides the necessary cytokines: TNF, IL-1, and IL-7.
Adsorption of pharmaceuticals onto isolated polyamide active layer of NF/RO membranes.

PubMed

Liu, Yan-Ling; Wang, Xiao-Mao; Yang, Hong-Wei; Xie, Yuefeng F

2018-06-01

Adsorption of trace organic compounds (TrOCs) onto the membrane materials has a great impact on their rejection by nanofiltration (NF) and reverse osmosis (RO) membranes. This study aimed to investigate the difference in adsorption of various pharmaceuticals (PhACs) onto different NF/RO membranes and to demonstrate the necessity of isolating the polyamide (PA) active layer from the polysulfone (PS) support layer for adsorption characterization and quantification. Both the isolated PA layers and the PA+PS layers of NF90 and ESPA1 membranes were used to conduct static adsorption tests. Results showed that apparent differences existed between the PA layer and the PA+PS layer in the adsorption capacity of PhACs as well as the time necessary to reach the adsorption equilibrium. PhACs with different physicochemical properties could be adsorbed to different extents by the isolated PA layer, which was mainly attributed to electrostatic attraction/repulsion and hydrophobic interactions. The PA layer of ESPA1 exhibited apparently higher adsorption capacities for the positively charged PhACs and similar adsorption capacities for the neutral PhACs although it had significantly less total interfacial area (per unit membrane surface area) for adsorption compared to the PA layer of NF90. The higher affinity of the PA layer of ESPA1 for the PhACs could be due to its higher capacity of forming hydrogen bonds with PhACs resulted from the modified chemistry with more -OH groups. This study provides a novel approach to determining the TrOC adsorption onto the active layer of membranes for the ease of investigating adsorption mechanisms. Copyright © 2018 Elsevier Ltd. All rights reserved.
FLICE-like inhibitory protein (FLIP) protects against apoptosis and suppresses NF-kappaB activation induced by bacterial lipopolysaccharide.

PubMed

Bannerman, Douglas D; Eiting, Kristine T; Winn, Robert K; Harlan, John M

2004-10-01

Bacterial lipopolysaccharide (LPS) via its activation of Toll-like receptor-4 contributes to much of the vascular injury/dysfunction associated with gram-negative sepsis. Inhibition of de novo gene expression has been shown to sensitize endothelial cells (EC) to LPS-induced apoptosis, the onset of which correlates with decreased expression of FLICE-like inhibitory protein (FLIP). We now have data that conclusively establish a role for FLIP in protecting EC against LPS-induced apoptosis. Overexpression of FLIP protected against LPS-induced apoptosis, whereas down-regulation of FLIP using antisense oligonucleotides sensitized EC to direct LPS killing. Interestingly, FLIP overexpression suppressed NF-kappaB activation induced by LPS, but not by phorbol ester, suggesting a specific role for FLIP in mediating LPS activation. Conversely, mouse embryo fibroblasts (MEF) obtained from FLIP -/- mice showed enhanced LPS-induced NF-kappaB activation relative to those obtained from wild-type mice. Reconstitution of FLIP-/- MEF with full-length FLIP reversed the enhanced NF-kappaB activity elicited by LPS in the FLIP -/- cells. Changes in the expression of FLIP had no demonstrable effect on other known LPS/Tlr-4-activated signaling pathways including the p38, Akt, and Jnk pathways. Together, these data support a dual role for FLIP in mediating LPS-induced apoptosis and NF-kappaB activation.

Altered ROS production, NF-κB activation and interleukin-6 gene expression induced by electrical stimulation in dystrophic mdx skeletal muscle cells.

PubMed

Henríquez-Olguín, Carlos; Altamirano, Francisco; Valladares, Denisse; López, José R; Allen, Paul D; Jaimovich, Enrique

2015-07-01

Duchenne muscular dystrophy is a fatal X-linked genetic disease, caused by mutations in the dystrophin gene, which cause functional loss of this protein. This pathology is associated with an increased production of reactive oxygen (ROS) and nitrogen species. The aim of this work was to study the alterations in NF-κB activation and interleukin-6 (IL-6) expression induced by membrane depolarization in dystrophic mdx myotubes. Membrane depolarization elicited by electrical stimulation increased p65 phosphorylation, NF-κB transcriptional activity and NF-κB-dependent IL-6 expression in wt myotubes, whereas in mdx myotubes it had the opposite effect. We have previously shown that depolarization-induced intracellular Ca2+ increases and ROS production are necessary for NF-κB activation and stimulation of gene expression in wt myotubes. Dystrophic myotubes showed a reduced amplitude and area under the curve of the Ca2+ transient elicited by electrical stimulation. On the other hand, electrical stimuli induced higher ROS production in mdx than wt myotubes, which were blocked by NOX2 inhibitors. Moreover, mRNA expression and protein levels of the NADPH oxidase subunits: p47phox and gp91phox were increased in mdx myotubes. Looking at ROS-dependence of NF-κB activation we found that in wt myotubes external administration of 50 μM H2O2 increased NF-κB activity; after administration of 100 and 200 μM H2O2 there was no effect. In mdx myotubes there was a dose-dependent reduction in NF-κB activity in response to external administration of H2O2, with a significant effect of 100 μM and 200 μM, suggesting that ROS levels are critical for NF-κB activity. Prior blockage with NOX2 inhibitors blunted the effects of electrical stimuli in both NF-κB activation and IL-6 expression. Finally, to ascertain whether stimulation of NF-κB and IL-6 gene expression by the inflammatory pathway is also impaired in mdx myotubes, we studied the effect of lipopolysaccharide on both NF
HSV-1-induced activation of NF-κB protects U937 monocytic cells against both virus replication and apoptosis

PubMed Central

Marino-Merlo, Francesca; Papaianni, Emanuela; Medici, Maria Antonietta; Macchi, Beatrice; Grelli, Sandro; Mosca, Claudia; Borner, Christoph; Mastino, Antonio

2016-01-01

The transcription factor nuclear factor-kappa B (NF-κB) is a crucial player of the antiviral innate response. Intriguingly, however, NF-κB activation is assumed to favour herpes simplex virus (HSV) infection rather than restrict it. Apoptosis, a form of innate response to viruses, is completely inhibited by HSV in fully permissive cells, but not in cells incapable to fully sustain HSV replication, such as immunocompetent cells. To resolve the intricate interplay among NF-κB signalling, apoptosis and permissiveness to HSV-1 in monocytic cells, we utilized U937 monocytic cells in which NF-κB activation was inhibited by expressing a dominant-negative IκBα. Surprisingly, viral production was increased in monocytic cells in which NF-κB was inhibited. Moreover, inhibition of NF-κB led to increased apoptosis following HSV-1 infection, associated with lysosomal membrane permeabilization. High expression of late viral proteins and induction of apoptosis occurred in distinct cells. Transcriptional analysis of known innate response genes by real-time quantitative reverse transcription-PCR excluded a contribution of the assayed genes to the observed phenomena. Thus, in monocytic cells NF-κB activation simultaneously serves as an innate process to restrict viral replication as well as a mechanism to limit the damage of an excessive apoptotic response to HSV-1 infection. This finding may clarify mechanisms controlling HSV-1 infection in monocytic cells. PMID:27584793
Effects of bovine lactoferrin in surgically created bone defects on bone regeneration around implants.

PubMed

Görmez, Ulaş; Kürkcü, Mehmet; E Benlidayi, Mehmet; Ulubayram, Kezban; Sertdemir, Yaşar; Dağlioğlu, Kenan

2015-03-01

The aim of this experimental study was to evaluate the effect of bovine lactoferrin (bLF)-loaded gelatin microspheres (GM) used in combination with anorganic bovine bone on bone regeneration in surgically created bone defects around tooth implants. Twenty-four uniform bone defects were created in the frontal bone via an extraoral approach in 12 domestic pigs. Twenty-four implants were placed at the center of the defects. In eight animals one of these defects was filled with 0.3 mL anorganic bovine bone while the other was left empty. In four animals, all defects were filled with 3 mg/defect bLF-loaded GM and anorganic bovine bone. All the defects were covered with collagen membranes. All animals were sacrificed after 10 weeks of healing, and the implants with the surrounding bone defects were removed en bloc. Undecalcified sections were prepared for histomorphometric analysis. The mean total area of hard tissue was 26.9 ± 6.0% in the empty defect group, 31.8 ± 8.4% in the graft group, and 47.6 ± 5.0% in the lactoferrin group (P < 0.001). The mean area of newly formed bone was 26.9 ± 6.0% in the empty defect group, 22.4 ± 8.2% in the graft group, and 46.1 ± 5.1% in the lactoferrin group (P < 0.001). The mean residual graft area was 9.4 ± 3.2% in the graft group and 1.5 ± 0.6% in the lactoferrin group (P < 0.001). The mean proportion of bone-implant contact in the defect region was 21.9 ± 8.4% in the empty defect group, 26.9 ± 10.1% in the graft group and 29.9 ± 10.3% in the lactoferrin group (P = 0.143). These data indicate that a combination of 3 mg bLF-loaded GM and bovine-derived HA promotes bone regeneration in defects around implants.
Gold nanoparticles induce transcriptional activity of NF-κB in a B-lymphocyte cell line

NASA Astrophysics Data System (ADS)

Sharma, Monita; Salisbury, Richard L.; Maurer, Elizabeth I.; Hussain, Saber M.; Sulentic, Courtney E. W.

2013-04-01

Gold nanoparticles (Au-NPs) have been designated as superior tools for biological applications owing to their characteristic surface plasmon absorption/scattering and amperometric (electron transfer) properties, in conjunction with low or no immediate toxicity towards biological systems. Many studies have shown the ease of designing application-based tools using Au-NPs but the interaction of this nanosized material with biomolecules in a physiological environment is an area requiring deeper investigation. Immune cells such as lymphocytes circulate through the blood and lymph and therefore are likely cellular components to come in contact with Au-NPs. The main aim of this study was to mechanistically determine the functional impact of Au-NPs on B-lymphocytes. Using a murine B-lymphocyte cell line (CH12.LX), treatment with citrate-stabilized 10 nm Au-NPs induced activation of an NF-κB-regulated luciferase reporter, which correlated with altered B lymphocyte function (i.e. increased antibody expression). TEM imaging demonstrated that Au-NPs can pass through the cellular membrane and therefore could interact with intracellular components of the NF-κB signaling pathway. Based on the inherent property of Au-NPs to bind to -thiol groups and the presence of cysteine residues on the NF-κB signal transduction proteins IκB kinases (IKK), proteins specifically bound to Au-NPs were extracted from CH12.LX cellular lysate exposed to 10 nm Au-NPs. Electrophoresis identified several bands, of which IKKα and IKKβ were immunoreactive. Further evaluation revealed activation of the canonical NF-κB signaling pathway as evidenced by IκBα phosphorylation at serine residues 32 and 36 followed by IκBα degradation and increased nuclear RelA. Additionally, expression of an IκBα super-repressor (resistant to proteasomal degradation) reversed Au-NP-induced NF-κB activation. Altered NF-κB signaling and cellular function in B-lymphocytes suggests a potential for off-target effects
Oral lactoferrin protects against experimental candidiasis in mice.

PubMed

Velliyagounder, K; Alsaedi, W; Alabdulmohsen, W; Markowitz, K; Fine, D H

2015-01-01

To determine the role of human lactoferrin (hLF) in protecting the oral cavities of mice against Candida albicans infection in lactoferrin knockout (LFKO(-/-)) mice was compared to wild-type (WT) mice. We also aim to determine the protective role of hLF in LFKO(-/-) mice. Antibiotic-treated immunosuppressed mice were inoculated with C. albicans (or sham infection) by oral swab and evaluated for the severity of infection after 7 days of infection. To determine the protective role of hLF, we added 0·3% solution of hLF to the drinking water given to some of the mice. CFU count, scoring of lesions and microscopic observations were carried out to determine the severity of infection. LFKO(-/-) I mice showed a 2 log (P = 0·001) higher CFUs of C. albicans in the oral cavity compared to the WT mice infected with C. albicans (WTI). LFKO(-/-) I mice given hLF had a 3 log (P = 0·001) reduction in CFUs in the oral cavity compared to untreated LFKO(-/-) I mice. The severity of infection, observed by light microscopy, revealed that the tongue of the LFKO(-/-) I mice showed more white patches compared to WTI and LFKO(-/-) I + hLF mice. Scanning electron microscopic observations revealed that more filiform papillae were destroyed in LFKO(-/-) I mice when compared to WTI or LFKO(-/-) I + hLF mice. Human LF is important in protecting mice from oral C. albicans infection. Administered hLF may be used to prevent C. albicans infection. Human LF, a multifunctional iron-binding glycoprotein can be used as a therapeutic active ingredient in oral healthcare products against C. albicans. © 2014 The Society for Applied Microbiology.
NF-κB Signaling in Gastric Cancer

PubMed Central

Sokolova, Olga; Naumann, Michael

2017-01-01

Gastric cancer is a leading cause of cancer death worldwide. Diet, obesity, smoking and chronic infections, especially with Helicobacter pylori, contribute to stomach cancer development. H. pylori possesses a variety of virulence factors including encoded factors from the cytotoxin-associated gene pathogenicity island (cagPAI) or vacuolating cytotoxin A (VacA). Most of the cagPAI-encoded products form a type 4 secretion system (T4SS), a pilus-like macromolecular transporter, which translocates CagA into the cytoplasm of the host cell. Only H. pylori strains carrying the cagPAI induce the transcription factor NF-κB, but CagA and VacA are dispensable for direct NF-κB activation. NF-κB-driven gene products include cytokines/chemokines, growth factors, anti-apoptotic factors, angiogenesis regulators and metalloproteinases. Many of the genes transcribed by NF-κB promote gastric carcinogenesis. Since it has been shown that chemotherapy-caused cellular stress could elicit activation of the survival factor NF-κB, which leads to acquisition of chemoresistance, the NF-κB system is recommended for therapeutic targeting. Research is motivated for further search of predisposing conditions, diagnostic markers and efficient drugs to improve significantly the overall survival of patients. In this review, we provide an overview about mechanisms and consequences of NF-κB activation in gastric mucosa in order to understand the role of NF-κB in gastric carcinogenesis. PMID:28350359
MafK/NF-E2 p18 is required for beta-globin genes activation by mediating the proximity of LCR and active beta-globin genes in MEL cell line.

PubMed

Du, Mei-Jun; Lv, Xiang; Hao, De-Long; Zhao, Guo-Wei; Wu, Xue-Song; Wu, Feng; Liu, De-Pei; Liang, Chih-Chuan

2008-01-01

Evidences indicate that locus control region (LCR) of beta-globin spatially closes to the downstream active gene promoter to mediate the transcriptional activation by looping. DNA binding proteins may play an important role in the looping formation. NF-E2 is one of the key transcription factors in beta-globin gene transcriptional activation. To shed light on whether NF-E2 is involved in this process, DS19MafKsiRNA cell pools were established by specifically knocked down the expression of MafK/NF-E2 p18, one subunit of NF-E2 heterodimer. In the above cell pools, it was observed that the occupancy efficiency of NF-E2 on beta-globin gene locus and the expression level of beta-globin genes were decreased. H3 acetylation, H3-K4 methylation and the deposition of RNA polymerase II, but not the recruitment of GATA-1, were also found reduced at the beta-globin gene cluster. Chromosome Conformation Capture (3C) assay showed that the cross-linking frequency between the main NF-E2 binding site HS2 and downstream structural genes was reduced compared to the normal cell. This result demonstrated that MafK/NF-E2 p18 recruitment was involved in the physical proximity of LCR and active beta-globin genes upon beta-globin gene transcriptional activation.
Lapatinib-induced NF-kappaB activation sensitizes triple-negative breast cancer cells to proteasome inhibitors.

PubMed

Chen, Yun-Ju; Yeh, Ming-Hsin; Yu, Meng-Chieh; Wei, Ya-Ling; Chen, Wen-Shu; Chen, Jhen-Yu; Shih, Chih-Yu; Tu, Chih-Yen; Chen, Chia-Hung; Hsia, Te-Chun; Chien, Pei-Hsuan; Liu, Shu-Hui; Yu, Yung-Luen; Huang, Wei-Chien

2013-11-12

Triple-negative breast cancer (TNBC), a subtype of breast cancer with negative expressions of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (HER2), is frequently diagnosed in younger women and has poor prognosis for disease-free and overall survival. Due to the lack of known oncogenic drivers for TNBC proliferation, clinical benefit from currently available targeted therapies is limited, and new therapeutic strategies are urgently needed. Triple-negative breast cancer cell lines were treated with proteasome inhibitors in combination with lapatinib (a dual epidermal growth factor receptor (EGFR)/HER2 tyrosine kinase inhibitor). Their in vitro and in vivo viability was examined by MTT assay, clonogenic analysis, and orthotopic xenograft mice model. Luciferase reporter gene, immunoblot, and RT-qPCR, immunoprecipitation assays were used to investigate the molecular mechanisms of action. Our data showed that nuclear factor (NF)-κB activation was elicited by lapatinib, independent of EGFR/HER2 inhibition, in TNBCs. Lapatinib-induced constitutive activation of NF-κB involved Src family kinase (SFK)-dependent p65 and IκBα phosphorylations, and rendered these cells more vulnerable to NF-κB inhibition by p65 small hairpin RNA. Lapatinib but not other EGFR inhibitors synergized the anti-tumor activity of proteasome inhibitors both in vitro and in vivo. Our results suggest that treatment of TNBCs with lapatinib may enhance their oncogene addiction to NF-κB, and thus augment the anti-tumor activity of proteasome inhibitors. These findings suggest that combination therapy of a proteasome inhibitor with lapatinib may benefit TNBC patients.
Magnesium sulfate provides neuroprotection in lipopolysaccharide-activated primary microglia by inhibiting NF-κB pathway.

PubMed

Gao, Feng; Ding, Baozhong; Zhou, Longan; Gao, Xueshan; Guo, Huiguang; Xu, Hong

2013-10-01

Magnesium sulfate has been used as an anticonvulsant in severe preeclamptic or eclamptic women prior to surgical trauma, but its effects on neuroinflammation is not well defined. In the present study, we investigated the neuroprotective effects of magnesium sulfate in lipopolysaccharide (LPS)-induced microglia and explored the underlying mechanism. Microglia was incubated with LPS in the presence or absence of various concentrations of magnesium sulfate, or L-type calcium channel activator BAY-K8644. The levels of inflammatory mediators, such as nitric oxide, prostaglandin E2, interleukin 1β, and tumor necrosis factor α, were measured using enzyme-linked immunosorbent assay. The expression of inducible nitric oxide synthase mRNA was detected by reverse-transcription polymerase chain reaction. Nuclear factor κB (NF-κB) activity in the nuclear extract of microglia was detected by NF-κB p50/p65 transcription factor assay kit. Magnesium sulfate at 5 and 10 mmol/L significantly inhibited the release of nitric oxide, prostaglandin E2, interleukin 1β, and tumor necrosis factor α, and the expression of inducible nitric oxide synthase mRNA in LPS-activated microglia. Furthermore, magnesium sulfate inhibited the translocation of NF-κB from the cytoplasm to the nucleus in a dose-dependent manner. Notably, these effects were significantly reversed by L-type calcium channel activator BAY-K8644. Magnesium sulfate protects microglia against LPS-induced release of inflammatory mediators, and these effects may be mediated by inhibiting L-type calcium channels and NF-κB signaling. Crown Copyright © 2013. Published by Elsevier Inc. All rights reserved.
Substances released from probiotic Lactobacillus rhamnosus GR-1 potentiate NF-κB activity in Escherichia coli-stimulated urinary bladder cells.

PubMed

Karlsson, Mattias; Scherbak, Nikolai; Khalaf, Hazem; Olsson, Per-Erik; Jass, Jana

2012-11-01

Lactobacillus rhamnosus GR-1 is a probiotic bacterium used to maintain urogenital health. The putative mechanism for its probiotic effect is by modulating the host immunity. Urinary tract infections (UTI) are often caused by uropathogenic Escherichia coli that frequently evade or suppress immune responses in the bladder and can target pathways, including nuclear factor-kappaB (NF-κB). We evaluated the role of L. rhamnosus GR-1 on NF-κB activation in E. coli-stimulated bladder cells. Viable L. rhamnosus GR-1 was found to potentiate NF-κB activity in E. coli-stimulated T24 bladder cells, whereas heat-killed lactobacilli demonstrated a marginal increase in NF-κB activity. Surface components released by trypsin- or LiCl treatment, or the resultant heat-killed shaved lactobacilli, had no effect on NF-κB activity. Isolation of released products from L. rhamnosus GR-1 demonstrated that the induction of NF-κB activity was owing to released product(s) with a relatively large native size. Several putative immunomodulatory proteins were identified, namely GroEL, elongation factor Tu and NLP/P60. GroEL and elongation factor Tu have previously been shown to elicit immune responses from human cells. Isolating and using immune-augmenting substances produced by lactobacilli is a novel strategy for the prevention or treatment of UTI caused by immune-evading E. coli. © 2012 Federation of European Microbiological Societies. Published by Blackwell Publishing Ltd. All rights reserved.
Polyubiquitination events mediate polymethylmethacrylate (PMMA) particle activation of NF-kappaB pathway.

PubMed

Yamanaka, Yasuhiro; Karuppaiah, Kannan; Abu-Amer, Yousef

2011-07-08

The pathologic response to implant wear-debris constitutes a major component of inflammatory osteolysis and remains under intense investigation. Polymethylmethacrylate (PMMA) particles, which are released during implant wear and loosening, constitute a major culprit by virtue of inducing inflammatory and osteolytic responses by macrophages and osteoclasts, respectively. Recent work by several groups has identified important cellular entities and secreted factors that contribute to inflammatory osteolysis. In previous work, we have shown that PMMA particles contribute to inflammatory osteolysis through stimulation of major pathways in monocytes/macrophages, primarily NF-κB and MAP kinases. The former pathway requires assembly of large IKK complex encompassing IKK1, IKK2, and IKKγ/NEMO. We have shown recently that interfering with the NF-κB and MAPK activation pathways, through introduction of inhibitors and decoy molecules, impedes PMMA-induced inflammation and osteolysis in mouse models of experimental calvarial osteolysis and inflammatory arthritis. In this study, we report that PMMA particles activate the upstream transforming growth factor β-activated kinase-1 (TAK1), which is a key regulator of signal transduction cascades leading to activation of NF-κB and AP-1 factors. More importantly, we found that PMMA particles induce TAK1 binding to NEMO and UBC13. In addition, we show that PMMA particles induce TRAF6 and UBC13 binding to NEMO and that lack of TRAF6 significantly attenuates NEMO ubiquitination. Altogether, these observations suggest that PMMA particles induce ubiquitination of NEMO, an event likely mediated by TRAF6, TAK1, and UBC13. Our findings provide important information for better understanding of the mechanisms underlying PMMA particle-induced inflammatory responses.
Learn about NF

MedlinePlus

... Understanding NF Neuroﬁbromatosis Intro to NF NF1 NF2 Schwannomatosis Genetics of NF Living with NF NF Registry ... of passing the condition on to their offspring. Schwannomatosis is less well understood, but the majority of ...
NEMO Binding Domain peptide inhibits constitutive NF-κB activity and reduces tumor burden in a canine model of relapsed, refractory Diffuse Large B-Cell Lymphoma

PubMed Central

Gaurnier-Hausser, Anita; Patel, Reema; Baldwin, Albert S.; May, Michael J.; Mason, Nicola J.

2011-01-01

Purpose Activated B-Cell Diffuse Large B-Cell Lymphoma (ABC-DLBCL) is an aggressive, poorly chemoresponsive lymphoid malignancy characterized by constitutive canonical NF-κB activity that promotes lymphomagenesis and chemotherapy resistance via over-expression of anti-apoptotic NF-κB target genes. Inhibition of the canonical NF-κB pathway may therefore have therapeutic relevance in ABC-DLBCL. Here we set out to determine whether dogs with spontaneous DLBCL have comparative aberrant constitutive NF-κB activity and to determine the therapeutic relevance of NF-κB inhibition in dogs with relapsed, resistant DLBCL. Experimental Design Canonical NF-κB activity was evaluated by electrophoretic mobility shift assays and immunoblot analyses, and NF-κB target gene expression was measured by qRT-PCR. Primary malignant canine B lymphocytes were treated with the selective IKK complex inhibitor Nemo Binding Domain (NBD) peptide, and evaluated for NF-κB activity and apoptosis. NBD peptide was administered intra-nodally to dogs with relapsed B-cell lymphoma and NF-κB target gene expression and tumor burden were evaluated pre and post treatment. Results Constitutive canonical NF-κB activity and increased NF-κB target gene expression was detected in primary DLBCL tissue. NBD peptide inhibited this activity and induced apoptosis of primary malignant B cells in vitro. Intra-tumoral injections of NBD peptide to dogs with relapsed DLBCL inhibited NF-κB target gene expression and reduced tumor burden. Conclusions This work shows that dogs with spontaneous DLBCL represent a clinically relevant, spontaneous, large animal model for human ABC-DLBCL and demonstrates the therapeutic relevance of NF-κB inhibition in the treatment of ABC-DLBCL. These results have important translational relevance for ABC-DLBCL treatment in human patients. PMID:21610150
Increased IL-2 production in T cells by xanthohumol through enhanced NF-AT and AP-1 activity.

PubMed

Choi, Jin Myung; Kim, Hyun Jung; Lee, Kwang Youl; Choi, Hyun Jin; Lee, Ik-Soo; Kang, Bok Yun

2009-01-01

Xanthohumol (XN) is a major chalcone found in hop, which is used to add bitterness and flavor to beer. In this study, we investigated the effects of XN on the production of interlukin-2 (IL-2), a potent T cell growth factor. Treatment with XN significantly increased IL-2 production in mouse EL-4 T cells activated with phorbol 12-myristate 13-acetate (PMA) plus ionomycin (Io) in a dose-dependent manner. To further characterize its regulatory mechanism of XN on increased IL-2 production, the effects of XN on IL-2 promoter activity and the activity of several transcription factors modulating IL-2 expression were analyzed. XN enhanced activity of the IL-2 promoter, which contains distal and proximal regulatory elements in PMA/Io-activated EL-4 T cells. Furthermore, the activity of NF-AT and AP-1 was enhanced but NF-kappaB activity was not influenced by XN in PMA/Io-activated EL-4 T cells. These results suggest that XN increased IL-2 production at the transcriptional levels via the up-regulation of NF-AT and AP-1 in PMA/Io-activated EL-4 T cells.
Vinpocetine inhibits amyloid-beta induced activation of NF-κB, NLRP3 inflammasome and cytokine production in retinal pigment epithelial cells

PubMed Central

Liu, Ruozhou Tom; Wang, Aikun; To, Eleanor; Gao, Jiangyuan; Cao, Sijia; Cui, Jing Z.; Matsubara, Joanne A.

2015-01-01

Chronic inflammation is a key pathogenic process in age-related macular degeneration (AMD). Amyloid-beta (Aβ) is a constituent of AMD drusen and promotes the activation of NLRP3 inflammasome which facilitates the production of cytokines. We investigated the role of transcription factor NF-κB in the activation of inflammasome in the RPE and the effect of vinpocetine, a dietary supplement with inhibitory effect on NF-κB. ARPE19/NF-κB-luciferase reporter cells treated with Aβ demonstrated enhanced NF-κB activation that was significantly suppressed by vinpocetine. Intraperitoneal injection of vinpocetine (15 mg/kg) inhibited NF-κB nuclear translocation and reduced the expression and activation of NLRP3, caspase-1, IL-1β, IL-18, and TNF-α in the RPE of adult rats that received intraocular Aβ, as measured by retinal immunohistochemistry and Western blot. Cytokine level in the vitreous was assayed using multiplex suspension arrays and revealed significantly lower concentration of MIP-3α, IL-6, IL-1α, IL-1β, IL-18, and TNF-α in vinpocetine treated animals. These results suggest that the NF-κB pathway is activated by Aβ in the RPE and signals the priming of NLRP3 inflammasome and the expression of pro-inflammatory cytokines including the inflammasome substrates IL-1β and IL-18. NF-κB inhibition may be an effective approach to stem the chronic inflammatory milieu that underlies the development of AMD. Vinpocetine is a potentially useful anti-inflammatory agent that is well-tolerated in long term use. PMID:25041941
Pasteurella haemolytica leukotoxin and endotoxin induced cytokine gene expression in bovine alveolar macrophages requires NF-kappaB activation and calcium elevation.

PubMed

Hsuan, S L; Kannan, M S; Jeyaseelan, S; Prakash, Y S; Malazdrewich, C; Abrahamsen, M S; Sieck, G C; Maheswaran, S K

1999-05-01

In bovine alveolar macrophages (BAMs), exposure to leukotoxin (Lkt) and endotoxin (LPS) from Pasteurella haemolytica results in expression of inflammatory cytokine genes and intracellular calcium ([Ca2+]i) elevation. Leukotoxin from P. haemolytica interacts only with leukocytes and platelets from ruminant species. Upregulation of cytokine genes in different cells by LPS involves activation of the transcription factor NF-kappaB (NF-kappaB), resulting in its translocation from the cytoplasm to the nucleus. Using immunocytochemical staining and confocal imaging, we studied whether NF-kappaB activation represents a common mechanism for the expression of multiple cytokine genes in BAMs (Lkt-susceptible cells) stimulated with Lkt and LPS. Bovine pulmonary artery endothelial cells and porcine alveolar macrophages were used as nonsusceptible cells. The role of Ca2+ and tyrosine kinases in NF-kappaB activation and inflammatory cytokine gene expression was studied, since an inhibitor of tyrosine kinases attenuates LPS-induced [Ca2+]i elevation in BAMs. The results are summarized as follows: (a) Lkt induced NF-kappaB activation and [Ca2+]i elevation only in BAMs, while LPS effects were demonstrable in all cell types; (b) chelation of [Ca2+]i blocked NF-kappaB activation and IL-1beta, TNFalpha, and IL-8 mRNA expression; and (c) tyrosine kinase inhibitor herbimycin A blocked expression of all three cytokine genes in BAMs stimulated with Lkt, while only the expression of IL-1beta was blocked in BAMs stimulated with LPS. We conclude that cytokine gene expression in BAMs requires NF-kappaB activation and [Ca2+]i elevation, and Lkt effects exhibit cell type- and species specificity. Copyright 1999 Academic Press.
Activation of IκB Kinase β and NF-κB Is Essential for Helicobacter pylori-Induced Chronic Gastritis in Mongolian Gerbils▿

PubMed Central

Yanai, Ayako; Maeda, Shin; Shibata, Wataru; Hikiba, Yohko; Sakamoto, Kei; Nakagawa, Hayato; Ohmae, Tomoya; Hirata, Yoshihiro; Ogura, Keiji; Muto, Susumu; Itai, Akiko; Omata, Masao

2008-01-01

The Mongolian gerbil model of Helicobacter pylori infection resembles human gastritis. In this study, we investigated the role of NF-κB activation in H. pylori-infected gerbils. Activated macrophages were significantly increased in H. pylori-infected gastric mucosa and were identified as being important cells with potent activation of NF-κB, which plays an important part in producing proinflammatory cytokines. Macrophage depletion by the administration of clodronate resulted in milder inflammation in gerbils infected with H. pylori. In macrophages, the inhibition of IκB kinase β (IKKβ), which is a critical kinase for NF-κB activation, resulted in lower proinflammatory cytokine expression caused by heat-killed H. pylori cells. Furthermore, treatment with IKKβ inhibitor resulted in milder inflammation in gerbils with H. pylori gastritis. Collectively, our data suggest that H. pylori-mediated gastric inflammation critically depends on the efficient recruitment and activation of macrophages, with sufficient NF-κB activation. PMID:18070894
Branches of the NF-κB signaling pathway regulate proliferation of oval cells in rat liver regeneration.

PubMed

Zhao, W M; Qin, Y L; Niu, Z P; Chang, C F; Yang, J; Li, M H; Zhou, Y; Xu, C S

2016-03-24

The NF-kB (nuclear factor kB) pathway is involved in the proliferation of many cell types. To explore the mechanism of the NF-kB signaling pathway underlying the oval cell proliferation during rat liver regeneration, the Rat Genome 230 2.0 Array was used to detect expression changes of NF-kB signaling pathway-related genes in oval cells. The results revealed that the expression levels of many genes in the NF-kB pathway were significantly changed. This included 48 known genes and 16 homologous genes, as well as 370 genes and 85 homologous genes related to cell proliferation. To further understand the biological significance of these changes, an expression profile function was used to analyze the potential biological processes. The results showed that the NF-kB pathway promoted oval cell proliferation mainly through three signaling branches; the tumor necrosis factor alpha branch (TNF-a pathway), the growth factor branch, and the chemokine branch. An integrated statistics method was used to define the key genes in the NF-kB pathway. Seven genes were identified to play vital roles in the NF-kB pathway. To confirm these results, the protein content, including two key genes (TNF and FGF11) and two non-key genes (CCL2 and TNFRSF12A), were analyzed using two-dimensional gel electrophoresis and MALDI-TOF/TOF mass spectrometry. The results were generally consistent with those of the array data. To conclude, three branches and seven key genes were involved in the NF-kB signaling pathway that regulates oval cell proliferation during rat liver regeneration.
Involvement of kinase PKC-zeta in the p62/p62(P392L)-driven activation of NF-κB in human osteoclasts.

PubMed

Chamoux, Estelle; McManus, Stephen; Laberge, Gino; Bisson, Martine; Roux, Sophie

2013-03-01

Mutations of the gene encoding sequestosome1 (SQSTM1/p62), clustering in or near the UBA domain, have been described in Paget's disease of bone (PDB); among these the P392L substitution is the most prevalent. Protein p62 mediates several cell functions, including the control of NF-κB signaling, and autophagy. This scaffolding protein interacts with atypical PKCζ in the RANKL-induced signaling complex. We have previously shown that osteoclasts (OCs) overexpressing the p62(P392L) variant were in a constitutively activated state, presenting activated kinase p-PKCζ/λ and activated NF-κB prior to RANKL stimulation. In the present study, we investigated the relationships between PKCζ and NF-κB activation in human OCs transfected with p62 variants. We showed that PKCζ and p-PKCζ/λ co-localize with p62, and that PKCζ is involved in the RANKL-induced NF-κB activation and in the RANKL-independent activation of NF-κB observed in p62(P392L)-transfected cells. We also observed a basal and RANKL-induced increase in IκBα levels in the presence of the p62(P392L) mutation that contrasted with the NF-κB activation. In this study we propose that PKCζ plays a role in the activation of NF-κB by acting as a p65 (RelA) kinase at Ser(536), independently of IκBα; this alternative pathway could be used preferentially in the presence of the p62(P392L) mutation, which may hinder the ubiquitin-proteasome pathway. Overall, our results highlight the importance of p62-associated PKCζ in the overactive state of pagetic OCs and in the activation of NF-κB, particularly in the presence of the p62(P392L) mutation. Copyright © 2012 Elsevier B.V. All rights reserved.
Shigella IpaH0722 E3 Ubiquitin Ligase Effector Targets TRAF2 to Inhibit PKC–NF-κB Activity in Invaded Epithelial Cells

PubMed Central

Ashida, Hiroshi; Nakano, Hiroyasu; Sasakawa, Chihiro

2013-01-01

NF-κB plays a central role in modulating innate immune responses to bacterial infections. Therefore, many bacterial pathogens deploy multiple mechanisms to counteract NF-κB activation. The invasion of and subsequent replication of Shigella within epithelial cells is recognized by various pathogen recognition receptors as pathogen-associated molecular patterns. These receptors trigger innate defense mechanisms via the activation of the NF-κB signaling pathway. Here, we show the inhibition of the NF-κB activation by the delivery of the IpaH E3 ubiquitin ligase family member IpaH0722 using Shigella's type III secretion system. IpaH0722 dampens the acute inflammatory response by preferentially inhibiting the PKC-mediated activation of NF-κB by ubiquitinating TRAF2, a molecule downstream of PKC, and by promoting its proteasome-dependent degradation. PMID:23754945

Soyasaponins Can Blunt Inflammation by Inhibiting the Reactive Oxygen Species-Mediated Activation of PI3K/Akt/NF-kB Pathway

PubMed Central

Zha, Longying; Chen, Jiading; Sun, Suxia; Mao, Limei; Chu, Xinwei; Deng, Hong; Cai, Junwei; Li, Xuefeng; Liu, Zhenqi; Cao, Wenhong

2014-01-01

We and others have recently shown that soyasaponins abundant in soybeans can decrease inflammation by suppressing the nuclear factor kappa B (NF-kB)-mediated inflammation. However, the exact molecular mechanisms by which soyasaponins inhibit the NF-kB pathway have not been established. In this study in macrophages, soyasaponins (A1, A2 and I) inhibited the lipopolysaccharide (LPS)-induced release of inflammatory marker prostaglandin E2 (PGE2) to a similar extent as the NF-kB inhibitor (BAY117082). Soyasaponins (A1, A2 and I) also suppressed the LPS-induced expression of cyclooxygenase 2 (COX-2), another inflammatory marker, in a dose-dependent manner by inhibiting NF-kB activation. In defining the associated mechanisms, we found that soyasaponins (A1, A2 and I) blunted the LPS-induced IKKα/β phosphorylation, IkB phosphorylation and degradation, and NF-kB p65 phosphorylation and nuclear translocation. In studying the upstream targets of soyasaponins on the NF-kB pathway, we found that soyasaponins (A1, A2 and I) suppressed the LPS-induced activation of PI3K/Akt similarly as the PI3K inhibitor LY294002, which alone blocked the LPS-induced activation of NF-kB. Additionally, soyasaponins (A1, A2 and I) reduced the LPS-induced production of reactive oxygen species (ROS) to the same extent as the anti-oxidant N-acetyl-L-cysteine, which alone inhibited the LPS-induced phosphorylation of Akt, IKKα/β, IkBα, and p65, transactivity of NF-kB, PGE2 production, and malondialdehyde production. Finally, our results show that soyasaponins (A1, A2 and I) elevated SOD activity and the GSH/GSSG ratio. Together, these results show that soyasaponins (A1, A2 and I) can blunt inflammation by inhibiting the ROS-mediated activation of the PI3K/Akt/NF-kB pathway. PMID:25233217
Biological Control Potential of Bacillus amyloliquefaciens KB3 Isolated from the Feces of Allomyrina dichotoma Larvae

PubMed Central

Nam, Hyo-Song; Yang, Hyun-Ju; Oh, Byung Jun; Anderson, Anne J.; Kim, Young Cheol

2016-01-01

Most biocontrol agents for plant diseases have been isolated from sources such as soils and plants. As an alternative source, we examined the feces of tertiary larvae of the herbivorous rhino beetle, Allomyrina dichotoma for presence of biocontrol-active microbes. The initial screen was performed to detect antifungal activity against two common fungal plant pathogens. The strain with strongest antifungal activity was identified as Bacillus amyloliquefaciens KB3. The inhibitory activity of this strain correlated with lipopeptide productions, including iturin A and surfactin. Production of these surfactants in the KB3 isolate varied with the culture phase and growth medium used. In planta biocontrol activities of cell-free culture filtrates of KB3 were similar to those of the commercial biocontrol agent, B. subtilis QST-713. These results support the presence of microbes with the potential to inhibit fungal growth, such as plant pathogens, in diverse ecological niches. PMID:27298603
Nickel(II) Complex of Polyhydroxybenzaldehyde N4-Thiosemicarbazone Exhibits Anti-Inflammatory Activity by Inhibiting NF-κB Transactivation

PubMed Central

Loh, Sheng Wei; Looi, Chung Yeng; Hassandarvish, Pouya; Phan, Alicia Yi Ling; Wong, Won Fen; Wang, Hao; Paterson, Ian C.; Ea, Chee Kwee; Mustafa, Mohd Rais; Maah, Mohd Jamil

2014-01-01

Background The biological properties of thiosemicarbazone have been widely reported. The incorporation of some transition metals such as Fe, Ni and Cu to thiosemicarbazone complexes is known to enhance its biological effects. In this study, we incorporated nickel(II) ions into thiosemicarbazone with N4-substitution groups H3L (H; H3L1, CH3; H3L2, C6H5; H3L3 and C2H5; H3L4) and examined its potential anti-inflammatory activity. Methodology/Principal Findings Four ligands (1–4) and their respective nickel-containing complexes (5–8) were synthesized and characterized. The compounds synthesized were tested for their effects on NF-κB nuclear translocation, pro-inflammatory cytokines secretion and NF-κB transactivation activity. The active compound was further evaluated on its ability to suppress carrageenan-induced acute inflammation in vivo. A potential binding target of the active compound was also predicted by molecular docking analysis. Conclusions/Significance Among all synthesized compounds tested, we found that complex [Ni(H2L1)(PPh3)]Cl (5) (complex 5), potently inhibited IκBα degradation and NF-κB p65 nuclear translocation in LPS-stimulated RAW264.7 cells as well as TNFα-stimulated HeLa S3 cells. In addition, complex 5 significantly down-regulated LPS- or TNFα-induced transcription of NF-κB target genes, including genes that encode the pro-inflammatory cytokines TNFα, IFNβ and IL6. Luciferase reporter assays confirmed that complex 5 inhibited the transactivation activity of NF-κB. Furthermore, the anti-inflammatory effect of complex 5 was also supported by its suppressive effect on carrageenan-induced paw edema formation in wild type C57BL/6 mice. Interestingly, molecular docking study showed that complex 5 potentially interact with the active site of IKKβ. Taken together, we suggest complex 5 as a novel NF-κB inhibitor with potent anti-inflammatory effects. PMID:24977407
Influence of lactoferrin in preventing preterm delivery: a pilot study.

PubMed

Giunta, Giuliana; Giuffrida, Lorena; Mangano, Katia; Fagone, Paolo; Cianci, Antonio

2012-01-01

Lactoferrin (Lf) is an approximately 80-kDa iron-binding glycoprotein, belonging to the transferrin family, with well-known bacteriostatic and bactericidal properties. It is produced and stored in specific (secondary) neutrophil granules and released during neutrophil activation and degranulation. Nowadays, Lf has a well-known therapeutic indication for combating iron deficiency anemia (IDA) in pregnant women. Studies suggest that Lf plays an important role against cervicovaginal infections by decreasing cytokines levels, such as interleukin (IL)-6, in cervicovaginal fluid. The aim of this preliminary trial was to evaluate the effectiveness of Lf in preventing preterm delivery caused by cervical infections and ripening. From November 2009 to August 2010, 21 pregnant women (26-32 weeks pregnant), aged between 22 and 36 years, suffering from IDA, at risk of preterm delivery, were prospectively enrolled in the study. One group (N=14) received 100 mg of recombinant human lactoferrin (bLf) [corrected] (lattoferrina; AG-pharma) twice a day before meals, for one month. The other group (N=7) received 520 mg of ferrous sulfate (Ferro-Grad; Abbott Laboratories, USA) once a day. The patients underwent transvaginal ultrasound to evaluate cervical length and funneling, and vaginal swabs were used to detect infections and cervicovaginal fluid sample collection to determine IL-6 levels. The results showed a correlation between the oral administration of 200 mg of bLf [corrected] with both the normalization of vaginal flora (vaginal infection disappearance) and the decrease in IL-6 cervicovaginal fluid levels in women at risk of preterm delivery.
Introducing the Forensic Research/Reference on Genetics knowledge base, FROG-kb.

PubMed

Rajeevan, Haseena; Soundararajan, Usha; Pakstis, Andrew J; Kidd, Kenneth K

2012-09-01

Online tools and databases based on multi-allelic short tandem repeat polymorphisms (STRPs) are actively used in forensic teaching, research, and investigations. The Fst value of each CODIS marker tends to be low across the populations of the world and most populations typically have all the common STRP alleles present diminishing the ability of these systems to discriminate ethnicity. Recently, considerable research is being conducted on single nucleotide polymorphisms (SNPs) to be considered for human identification and description. However, online tools and databases that can be used for forensic research and investigation are limited. The back end DBMS (Database Management System) for FROG-kb is Oracle version 10. The front end is implemented with specific code using technologies such as Java, Java Servlet, JSP, JQuery, and GoogleCharts. We present an open access web application, FROG-kb (Forensic Research/Reference on Genetics-knowledge base, http://frog.med.yale.edu), that is useful for teaching and research relevant to forensics and can serve as a tool facilitating forensic practice. The underlying data for FROG-kb are provided by the already extensively used and referenced ALlele FREquency Database, ALFRED (http://alfred.med.yale.edu). In addition to displaying data in an organized manner, computational tools that use the underlying allele frequencies with user-provided data are implemented in FROG-kb. These tools are organized by the different published SNP/marker panels available. This web tool currently has implemented general functions possible for two types of SNP panels, individual identification and ancestry inference, and a prediction function specific to a phenotype informative panel for eye color. The current online version of FROG-kb already provides new and useful functionality. We expect FROG-kb to grow and expand in capabilities and welcome input from the forensic community in identifying datasets and functionalities that will be most helpful
Introducing the Forensic Research/Reference on Genetics knowledge base, FROG-kb

PubMed Central

2012-01-01

Background Online tools and databases based on multi-allelic short tandem repeat polymorphisms (STRPs) are actively used in forensic teaching, research, and investigations. The Fst value of each CODIS marker tends to be low across the populations of the world and most populations typically have all the common STRP alleles present diminishing the ability of these systems to discriminate ethnicity. Recently, considerable research is being conducted on single nucleotide polymorphisms (SNPs) to be considered for human identification and description. However, online tools and databases that can be used for forensic research and investigation are limited. Methods The back end DBMS (Database Management System) for FROG-kb is Oracle version 10. The front end is implemented with specific code using technologies such as Java, Java Servlet, JSP, JQuery, and GoogleCharts. Results We present an open access web application, FROG-kb (Forensic Research/Reference on Genetics-knowledge base, http://frog.med.yale.edu), that is useful for teaching and research relevant to forensics and can serve as a tool facilitating forensic practice. The underlying data for FROG-kb are provided by the already extensively used and referenced ALlele FREquency Database, ALFRED (http://alfred.med.yale.edu). In addition to displaying data in an organized manner, computational tools that use the underlying allele frequencies with user-provided data are implemented in FROG-kb. These tools are organized by the different published SNP/marker panels available. This web tool currently has implemented general functions possible for two types of SNP panels, individual identification and ancestry inference, and a prediction function specific to a phenotype informative panel for eye color. Conclusion The current online version of FROG-kb already provides new and useful functionality. We expect FROG-kb to grow and expand in capabilities and welcome input from the forensic community in identifying datasets and
BCL11B enhances TCR/CD28-triggered NF-kappaB activation through up-regulation of Cot kinase gene expression in T-lymphocytes.

PubMed

Cismasiu, Valeriu B; Duque, Javier; Paskaleva, Elena; Califano, Danielle; Ghanta, Sailaja; Young, Howard A; Avram, Dorina

2009-01-15

BCL11B is a transcriptional regulator with an important role in T-cell development and leukaemogenesis. We demonstrated recently that BCL11B controls expression from the IL (interleukin)-2 promoter through direct binding to the US1 (upstream site 1). In the present study, we provide evidence that BCL11B also participates in the activation of IL-2 gene expression by enhancing NF-kappaB (nuclear factor kappaB) activity in the context of TCR (T-cell receptor)/CD28-triggered T-cell activation. Enhanced NF-kappaB activation is not a consequence of BCL11B binding to the NF-kappaB response elements or association with the NF-kappaB-DNA complexes, but rather the result of higher translocation of NF-kappaB to the nucleus caused by enhanced degradation of IkappaB (inhibitor of NF-kappaB). The enhanced IkappaB degradation in cells with increased levels of BCL11B was specific for T-cells activated through the TCR, but not for cells activated through TNFalpha (tumour necrosis factor alpha) or UV light, and was caused by increased activity of IkappaB kinase, as indicated by its increase in phosphorylation. As BCL11B is a transcription factor, we investigated whether the expression of genes upstream of IkappaB kinase in the TCR/CD28 signalling pathway was affected by increased BCL11B expression, and found that Cot (cancer Osaka thyroid oncogene) kinase mRNA levels were elevated. Cot kinase is known to promote enhanced IkappaB kinase activity, which results in the phosphorylation and degradation of IkappaB and activation of NF-kappaB. The implied involvement of Cot kinase in BCL11B-mediated NF-kappaB activation in response to TCR activation is supported by the fact that a Cot kinase dominant-negative mutant or Cot kinase siRNA (small interfering RNA) knockdown blocked BCL11B-mediated NF-kappaB activation. In support of our observations, in the present study we report that BCL11B enhances the expression of several other NF-kappaB target genes, in addition to IL-2. In addition, we
Regulation of nuclear factor κB (NF-κB) transcriptional activity via p65 acetylation by the chaperonin containing TCP1 (CCT).

PubMed

Pejanovic, Nadja; Hochrainer, Karin; Liu, Tao; Aerne, Birgit L; Soares, Miguel P; Anrather, Josef

2012-01-01

The NF-κB family member p65 is central to inflammation and immunity. The purpose of this study was to identify and characterize evolutionary conserved genes modulating p65 transcriptional activity. Using an RNAi screening approach, we identified chaperonin containing TCP1 subunit η (CCTη) as a regulator of Drosophila NF-κB proteins, Dorsal and Dorsal-related immunity factor (Dif). CCTη was also found to regulate NF-κB-driven transcription in mammalian cells, acting in a promoter-specific context, downstream of IκB kinase (IKK). CCTη knockdown repressed IκBα and CXCL2/MIP2 transcription during the early phase of NF-κB activation while impairing the termination of CCL5/RANTES and CXCL10/IP10 transcription. The latter effect was associated with increased DNA binding and reduced p65 acetylation, presumably by altering the activity of histone acetyltransferase CREB-binding protein (CBP). We identified p65 lysines (K) 122 and 123 as target residues mediating the CCTη-driven termination of NF-κB-dependent transcription. We propose that CCTη regulates NF-κB activity in a manner that resolves inflammation.
A new lactoferrin- and iron-dependent lysosomal death pathway is induced by benzo[a]pyrene in hepatic epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Gorria, Morgane; Tekpli, Xavier; Rissel, Mary

2008-04-15

While lysosomal disruption seems to be a late step of necrosis, a moderate lysosomal destabilization has been suggested to participate early in the apoptotic cascade. The origin of lysosomal dysfunction and its precise role in apoptosis or apoptosis-like process still needs to be clarified, especially upon carcinogen exposure. In this study, we focused on the implication of lysosomes in cell death induced by the prototype carcinogen benzo[a]pyrene (B[a]P; 50 nM) in rat hepatic epithelial F258 cells. We first demonstrated that B[a]P affected lysosomal morphology (increase in size) and pH (alkalinization), and that these changes were involved in caspase-3 activation andmore » cell death. Subsequently, we showed that lysosomal modifications were partly dependent on mitochondrial dysfunction, and that lysosomes together with mitochondria participate in B[a]P-induced oxidative stress. Using two iron chelators (desferrioxamine and deferiprone) and siRNA targeting the lysosomal iron-binding protease lactoferrin, we further demonstrated that both lysosomal iron content and lactoferrin were required for caspase-3 activation and apoptosis-like cell death.« less
Illness in breastfeeding infants relates to concentration of lactoferrin and secretory Immunoglobulin A in mother’s milk

PubMed Central

Breakey, Alicia A.; Hinde, Katie; Valeggia, Claudia R.; Sinofsky, Allison; Ellison, Peter T.

2015-01-01

Background and objectives: This study aims to better understand the relationship between immune compounds in human milk and infant health. We hypothesized that the concentration of immune compounds in milk would relate to infant illness symptoms according to two possible theoretical paradigms. In the ‘protective’ paradigm, high concentrations of immune compounds prevent infant illness. The converse, the ‘responsive’ framework, posits that concentrations of immune compounds are elevated in response to infection. Methodology: Milk samples (n = 110) and illness data were collected among the Toba of Argentina from 30 mother–infant dyads. Samples were assayed for two immune proteins, lactoferrin and secretory immunoglobulin A (sIgA). Generalized estimating equations were used to assess the relationship between immune composition of milk and symptoms of illness in infants. Results: Lactoferrin was positively associated with symptoms of illness in infants (odds ratios >1), both in the month preceding the sample collection and the subsequent month. sIgA was negatively associated with symptoms (odds ratios <1) in the preceding and subsequent months, an association which was particularly strong for gastrointestinal symptoms. Conclusions and implications: The two compounds investigated in our study had opposite relationships with symptoms of illness; the positive relationship between lactoferrin and illness lends support to our ‘responsive’ paradigm, and the negative relationship between sIgA and symptoms of illness was consistent with our ‘protective’ framework. That elevated lactoferrin is restricted to periods of illness suggests that there may be a cost to mother or infant associated with persistently elevated lactoferrin that is not incurred with elevated sIgA. PMID:25608691
Double NF1 Inactivation Affects Adrenocortical Function in NF1Prx1 Mice and a Human Patient

PubMed Central

Kobus, Karolina; Hartl, Daniela; Ott, Claus Eric; Osswald, Monika; Huebner, Angela; von der Hagen, Maja; Emmerich, Denise; Kühnisch, Jirko; Morreau, Hans; Hes, Frederik J.; Mautner, Victor F.; Harder, Anja; Tinschert, Sigrid; Mundlos, Stefan; Kolanczyk, Mateusz

2015-01-01

Background Neurofibromatosis type I (NF1, MIM#162200) is a relatively frequent genetic condition, which predisposes to tumor formation. Apart from tumors, individuals with NF1 often exhibit endocrine abnormalities such as precocious puberty (2,5–5% of NF1 patients) and some cases of hypertension (16% of NF1 patients). Several cases of adrenal cortex adenomas have been described in NF1 individuals supporting the notion that neurofibromin might play a role in adrenal cortex homeostasis. However, no experimental data were available to prove this hypothesis. Materials and Methods We analysed Nf1Prx1 mice and one case of adrenal cortical hyperplasia in a NF1patient. Results In Nf1Prx1 mice Nf1 is inactivated in the developing limbs, head mesenchyme as well as in the adrenal gland cortex, but not the adrenal medulla or brain. We show that adrenal gland size is increased in NF1Prx1 mice. Nf1Prx1 female mice showed corticosterone and aldosterone overproduction. Molecular analysis of Nf1 deficient adrenals revealed deregulation of multiple proteins, including steroidogenic acute regulatory protein (StAR), a vital mitochondrial factor promoting transfer of cholesterol into steroid making mitochondria. This was associated with a marked upregulation of MAPK pathway and a female specific increase of cAMP concentration in murine adrenal lysates. Complementarily, we characterized a patient with neurofibromatosis type I with macronodular adrenal hyperplasia with ACTH-independent cortisol overproduction. Comparison of normal control tissue- and adrenal hyperplasia- derived genomic DNA revealed loss of heterozygosity (LOH) of the wild type NF1 allele, showing that biallelic NF1 gene inactivation occurred in the hyperplastic adrenal gland. Conclusions Our data suggest that biallelic loss of Nf1 induces autonomous adrenal hyper-activity. We conclude that Nf1 is involved in the regulation of adrenal cortex function in mice and humans. PMID:25775093
NF-kappaB modulators from Valeriana officinalis.

PubMed

Jacobo-Herrera, Nadia J; Vartiainen, Nina; Bremner, Paul; Gibbons, Simon; Koistinaho, Jari; Heinrich, Michael

2006-10-01

Valeriana officinalis (Valerianaceae) has been of great interest for its therapeutic uses for treating mild nervous tension and temporary sleeping problems. In traditional European medicine it has been also reported as an antiinflammatory remedy. This study reports that the EtOAc extract of the underground parts of V. officinalis showed inhibitory activity against NF-kappaB at 100 microg/mL in the IL-6/Luc assay on HeLa cells and provided protection against excitotoxicity in primary brain cell cultures at micromolar concentrations. Bioassay-guided fractionation of the EtOAc extract led to the isolation of three known sesquiterpenes: acetylvalerenolic acid (1), valerenal (2) and valerenic acid (3), 1 and 3 were active as inhibitors of NF-kappaB at a concentration of 100 microg/mL. Acetylvalerenolic acid (1) reduced NF-kappaB activity to 4%, whereas valerenic acid (3) reduced NF-kappaB activity to 25%. Copyright 2006 John Wiley & Sons, Ltd.
Vinpocetine Ameliorates Acetic Acid-Induced Colitis by Inhibiting NF-κB Activation in Mice.

PubMed

Colombo, Bárbara B; Fattori, Victor; Guazelli, Carla F S; Zaninelli, Tiago H; Carvalho, Thacyana T; Ferraz, Camila R; Bussmann, Allan J C; Ruiz-Miyazawa, Kenji W; Baracat, Marcela M; Casagrande, Rúbia; Verri, Waldiceu A

2018-04-10

The idiopathic inflammatory bowel diseases (IBD) comprise two types of chronic intestinal disorders: Crohn's disease and ulcerative colitis. Recruited neutrophils and macrophages contribute to intestinal tissue damage via production of ROS and NF-κB-dependent pro-inflammatory cytokines. The introduction of anti-TNF-α therapies in the treatment of IBD patients was a seminal advance. This therapy is often limited by a loss of efficacy due to the development of adaptive immune response, underscoring the need for novel therapies targeting similar pathways. Vinpocetine is a nootropic drug and in addition to its antioxidant effect, it is known to have anti-inflammatory and analgesic properties, partly by inhibition of NF-κB and downstream cytokines. Therefore, the present study evaluated the effect of the vinpocetine in a model of acid acetic-induced colitis in mice. Treatment with vinpocetine reduced edema, MPO activity, microscopic score and macroscopic damage, and visceral mechanical hyperalgesia. Vinpocetine prevented the reduction of colonic levels of GSH, ABTS radical scavenging ability, and normalized levels of anti-inflammatory cytokine IL-10. Moreover, vinpocetine reduced NF-κB activation and thereby NF-κB-dependent pro-inflammatory cytokines IL-1β, TNF-α, and IL-33 in the colon. Thus, we demonstrate for the first time that vinpocetine has anti-inflammatory, antioxidant, and analgesic effects in a model of acid acetic-induced colitis in mice and deserves further screening to address its suitability as an approach for the treatment of IBD.
Resveratrol Protects against TNF-α-Induced Injury in Human Umbilical Endothelial Cells through Promoting Sirtuin-1-Induced Repression of NF-KB and p38 MAPK

PubMed Central

Huang, Shujie; Zhu, Pengli

2016-01-01

Inflammation and reactive oxygen species (ROS) play important roles in the pathogenesis of atherosclerosis. Resveratrol has been shown to possess anti-inflammatory and antioxidative stress activities, but the underlying mechanisms are not fully understood. In the present study, we investigated the molecular basis associated with the protective effects of resveratrol on tumor necrosis factor-alpha (TNF-α)-induced injury in human umbilical endothelial cells (HUVECs) using a variety of approaches including a cell viability assay, reverse transcription and quantitative polymerase chain reaction, western blot, and immunofluorescence staining. We showed that TNF-α induced CD40 expression and ROS production in cultured HUVECs, which were attenuated by resveratrol treatment. Also, resveratrol increased the expression of sirtuin 1 (SIRT1); and repression of SIRT1 by small-interfering RNA (siRNA) and the SIRT1 inhibitor Ex527 reduced the inhibitory effects of resveratrol on CD40 expression and ROS generation. In addition, resveratrol downregulated the levels of p65 and phospho-p38 MAPK, but this inhibitory effect was attenuated by the suppression of SIRT1 activity. Moreover, the p38 MAPK inhibitor SD203580 and the nuclear factor (NF)-κB inhibitor pyrrolidine dithiocarbamate (PDTC) achieved similar repressive effects as resveratrol on TNF-α-induced ROS generation and CD40 expression. Thus, our study provides a mechanistic link between resveratrol and the activation of SIRT1, the latter of which is involved in resveratrol-mediated repression of the p38 MAPK/NF-κB pathway and ROS production in TNF-α-treated HUVECs. PMID:26799794
NF-κB Hyper-Activation by HTLV-1 Tax Induces Cellular Senescence, but Can Be Alleviated by the Viral Anti-Sense Protein HBZ

PubMed Central

Zhi, Huijun; Yang, Liangpeng; Kuo, Yu-Liang; Ho, Yik-Khuan; Shih, Hsiu-Ming; Giam, Chou-Zen

2011-01-01

Activation of I-κB kinases (IKKs) and NF-κB by the human T lymphotropic virus type 1 (HTLV-1) trans-activator/oncoprotein, Tax, is thought to promote cell proliferation and transformation. Paradoxically, expression of Tax in most cells leads to drastic up-regulation of cyclin-dependent kinase inhibitors, p21CIP1/WAF1 and p27KIP1, which cause p53-/pRb-independent cellular senescence. Here we demonstrate that p21CIP1/WAF1-/p27KIP1-mediated senescence constitutes a checkpoint against IKK/NF-κB hyper-activation. Senescence induced by Tax in HeLa cells is attenuated by mutations in Tax that reduce IKK/NF-κB activation and prevented by blocking NF-κB using a degradation-resistant mutant of I-κBα despite constitutive IKK activation. Small hairpin RNA-mediated knockdown indicates that RelA induces this senescence program by acting upstream of the anaphase promoting complex and RelB to stabilize p27KIP1 protein and p21CIP1/WAF1 mRNA respectively. Finally, we show that down-regulation of NF-κB by the HTLV-1 anti-sense protein, HBZ, delay or prevent the onset of Tax-induced senescence. We propose that the balance between Tax and HBZ expression determines the outcome of HTLV-1 infection. Robust HTLV-1 replication and elevated Tax expression drive IKK/NF-κB hyper-activation and trigger senescence. HBZ, however, modulates Tax-mediated viral replication and NF-κB activation, thus allowing HTLV-1-infected cells to proliferate, persist, and evolve. Finally, inactivation of the senescence checkpoint can facilitate persistent NF-κB activation and leukemogenesis. PMID:21552325
Attenuation of liver pro-inflammatory responses by Zingiber officinale via inhibition of NF-kappa B activation in high-fat diet-fed rats.

PubMed

Li, Xiao-Hong; McGrath, Kristine C-Y; Nammi, Srinivas; Heather, Alison K; Roufogalis, Basil D

2012-03-01

The aim of this study was to investigate whether treatment with a ginger (Zingiber officinale) extract of high-fat diet (HFD)-fed rats suppresses Nuclear factor-kappa B (NF-κB)-driven hepatic inflammation and to subsequently explore the molecular mechanisms in vitro. Adult male Sprague-Dawley rats were treated with an ethanolic extract of Zingiber officinale (400 mg/kg) along with a HFD for 6 weeks. Hepatic cytokine mRNA levels, cytokine protein levels and NF-κB activation were measured by real-time PCR, Western blot and an NF-κB nuclear translocation assay, respectively. In vitro, cell culture studies were carried out in human hepatocyte (HuH-7) cells by treatment with Zingiber officinale (100 μg/mL) for 24 hr prior to interleukin-1β (IL-1β, 8 ng/mL)-induced inflammation. We showed that Zingiber officinale treatment decreased cytokine gene TNFα and IL-6 expression in HFD-fed rats, which was associated with suppression of NF-κB activation. In vitro, Zingiber officinale treatment decreased NF-κB-target inflammatory gene expression of IL-6, IL-8 and serum amyloid A1 (SAA1), while it suppressed NF-κB activity, IκBα degradation and IκB kinase (IKK) activity. In conclusion, Zingiber officinale suppressed markers of hepatic inflammation in HFD-fed rats, as demonstrated by decreased hepatic cytokine gene expression and decreased NF-κB activation. The study demonstrates that the anti-inflammatory effect of Zingiber officinale occurs at least in part through the NF-κB signalling pathway. © 2011 The Authors. Basic & Clinical Pharmacology & Toxicology © 2011 Nordic Pharmacological Society.
CFH Y402H polymorphism and the complement activation product C5a: effects on NF-κB activation and inflammasome gene regulation.

PubMed

Cao, Sijia; Wang, Jay Ching Chieh; Gao, Jiangyuan; Wong, Matthew; To, Elliott; White, Valerie A; Cui, Jing Z; Matsubara, Joanne A

2016-05-01

The Y402H polymorphism in the complement factor H (CFH) gene is an important risk factor for age-related macular degeneration (AMD). Complement activation products and proinflammatory cytokines are associated with this polymorphism at the systemic level, but less is known of the associations in the outer retina of the genotyped eye. Here we investigate complement activation products and their role in nuclear factor (NF)-κB activation and gene expression of the NLRP3 inflammasome pathway. Postmortem donor eyes were genotyped for the CFH Y402H polymorphism and assessed for complement C3a, C5a, interleukin (IL)-18 and tumour necrosis factor (TNF)-α. ARPE19 cells were stimulated basolaterally with C5a or TNF-α in polarised cultures. NF-κB activation was assessed with a reporter cell line. Gene expression of inflammasome-related (NLRP3, caspase-1, IL-1β and IL-18) and classic inflammatory (IL-6 and IL-8) genes was studied. The distribution of inflammasome products, IL-1β and IL-18, was studied in postmortem donor eyes with AMD pathologies. Eyes with the homozygous at-risk variant demonstrated higher levels of C5a, IL-18 and TNF-α in Bruch's membrane and choroid. C5a promoted NF-κB activation and upregulation of IL-18 in polarised ARPE19. TNF-α promoted NF-κB activation and gene expression of caspase-1, IL-1β, IL-18, IL-6 and IL-8, but downregulated NLRP3. In eyes with geographic atrophy, strong immunoreactivity was observed for inflammasome products IL-1β and IL-18 compared with age-matched controls. The at-risk polymorphism of the CFH Y402H may contribute to AMD disease process through increased complement and NF-κB activation, and the upregulation of IL-18, a product of inflammasome activation. Published by the BMJ Publishing Group Limited. For permission to use (where not already granted under a licence) please go to http://www.bmj.com/company/products-services/rights-and-licensing/
Lapatinib–induced NF-kappaB activation sensitizes triple-negative breast cancer cells to proteasome inhibitors

PubMed Central

2013-01-01

Introduction Triple-negative breast cancer (TNBC), a subtype of breast cancer with negative expressions of estrogen receptor, progesterone receptor, and human epidermal growth factor receptor 2 (HER2), is frequently diagnosed in younger women and has poor prognosis for disease-free and overall survival. Due to the lack of known oncogenic drivers for TNBC proliferation, clinical benefit from currently available targeted therapies is limited, and new therapeutic strategies are urgently needed. Methods Triple-negative breast cancer cell lines were treated with proteasome inhibitors in combination with lapatinib (a dual epidermal growth factor receptor (EGFR)/HER2 tyrosine kinase inhibitor). Their in vitro and in vivo viability was examined by MTT assay, clonogenic analysis, and orthotopic xenograft mice model. Luciferase reporter gene, immunoblot, and RT-qPCR, immunoprecipitation assays were used to investigate the molecular mechanisms of action. Results Our data showed that nuclear factor (NF)-κB activation was elicited by lapatinib, independent of EGFR/HER2 inhibition, in TNBCs. Lapatinib-induced constitutive activation of NF-κB involved Src family kinase (SFK)-dependent p65 and IκBα phosphorylations, and rendered these cells more vulnerable to NF-κB inhibition by p65 small hairpin RNA. Lapatinib but not other EGFR inhibitors synergized the anti-tumor activity of proteasome inhibitors both in vitro and in vivo. Our results suggest that treatment of TNBCs with lapatinib may enhance their oncogene addiction to NF-κB, and thus augment the anti-tumor activity of proteasome inhibitors. Conclusions These findings suggest that combination therapy of a proteasome inhibitor with lapatinib may benefit TNBC patients. PMID:24216290
Cooperative effects of hepatitis B virus and TNF may play important roles in the activation of metabolic pathways through the activation of NF-κB.

PubMed

Wu, Shuang; Kanda, Tatsuo; Nakamoto, Shingo; Jiang, Xia; Nakamura, Masato; Sasaki, Reina; Haga, Yuki; Shirasawa, Hiroshi; Yokosuka, Osamu

2016-08-01

Elevated levels of inflammatory cytokines such as tumor necrosis factor-α (TNF-α) and interleukin (IL)-1β are often observed in the sera of hepatitis B virus (HBV)-infected patients. It is well known that these cytokines activate nuclear factor-κB (NF-κB)-signaling, and are associated with endoplasmic reticulum (ER) stress. We investigated whether HBV or HBV X protein (HBx) enhanced the activation of NF-κB in the presence of TNF and/or IL-1β, and their effects on the expression of metabolic pathway‑associated genes. We examined whether HBV or HBx enhanced cytokine-induced activation of NF-κB in hepatocytes, using a reporter assay, in the presence or absence of TNF and/or IL-1β. The expression of insulin-like growth factor binding protein 1 (IGFBP1), one of the NF-κB target genes was also examined. The expression of metabolic pathway-associated genes in HepG2 and HepG2.2.15 cells in the presence or absence of TNF was evaluated by RT-qPCR. Human hepatocytes expressed TNF receptors and IL-1 receptors. NF-κB was activated by cooperation between HBx and TNF in human hepatocytes. We observed IGFBP1 expression in HBV infection and that a number of metabolic pathway-associated genes were upregulated in HepG2.2.15 cells, compared with HepG2 cells with or without TNF treatment. We observed the cooperative effects of HBV and TNF which enhanced the activation of NF-κB as well as upregulated the expression of metabolic pathway-associated genes in hepatocytes. These effects may be important in the development of HBV-associated metabolic syndrome.
Propofol inhibits NF-κB activation to ameliorate airway inflammation in ovalbumin (OVA)-induced allergic asthma mice.

PubMed

Zhang, Qiong; Wang, Liangrong; Chen, Baihui; Zhuo, Qian; Bao, Caiying; Lin, Lina

2017-10-01

Propofol, one of the most commonly used intravenous anesthetic agents, has been reported to have anti-inflammatory property. However, the anti-allergic inflammation effect of propofol and its underlying molecular mechanisms have not been elucidated. In the present study, we aim to investigate the roles of NF-kB activation in propofol anti-asthma effect on OVA-induced allergic airway inflammation in mice. In a standard experimental asthma model, Balb/c mice were sensitized with ovalbumin, treated with propofol (50,100,150mg/kg) or a vehicle control 1h before OVA challenge. Blood samples, bronchoalveolar lavage fluid (BALF) and lung tissues were harvested after measurement of airway hyperresponsiveness. Results revealed that propofol not only significantly inhibit airway hyperresponsiveness, but also inhibited the production of Th2 cytokines, NO, Ova-specific IgE and eotaxin. Histological studies indicated that propofol significantly attenuated OVA-induced inflammatory cell infiltration in the peribronchial areas and mucus hypersecretion. Meanwhile, our results indicated that propofol was found to inhibit NF-kB activation in OVA-Induced mice. Furthermore, propofol significantly reduced the TNF-α-induced NF-kB activation in A549 cells. In conclusion, our study suggested that propofol effectively reduced allergic airway inflammation by inhibiting NF-kB activation and could thus be used as a therapy for allergic asthma. Copyright © 2017. Published by Elsevier B.V.

Fungal-Induced Cell Cycle Impairment, Chromosome Instability and Apoptosis via Differential Activation of NF-κB

PubMed Central

Ben-Abdallah, Mariem; Sturny-Leclère, Aude; Avé, Patrick; Louise, Anne; Moyrand, Frédérique; Weih, Falk; Janbon, Guilhem; Mémet, Sylvie

2012-01-01

Microbial pathogens have developed efficient strategies to compromise host immune responses. Cryptococcus neoformans is a facultative intracellular pathogen, recognised as the most common cause of systemic fungal infections leading to severe meningoencephalitis, mainly in immunocompromised patients. This yeast is characterized by a polysaccharide capsule, which inhibits its phagocytosis. Whereas phagocytosis escape and macrophage intracellular survival have been intensively studied, extracellular survival of this yeast and restraint of host innate immune response are still poorly understood. In this study, we have investigated whether C. neoformans affected macrophage cell viability and whether NF-κB (nuclear factor-κB), a key regulator of cell growth, apoptosis and inflammation, was involved. Using wild-type (WT) as well as mutant strains of C. neoformans for the pathogen side, and WT and mutant cell lines with altered NF-κB activity or signalling as well as primary macrophages for the host side, we show that C. neoformans manipulated NF-κB-mediated signalling in a unique way to regulate macrophage cell fate and viability. On the one hand, serotype A strains reduced macrophage proliferation in a capsule-independent fashion. This growth decrease, which required a critical dosage of NF-κB activity, was caused by cell cycle disruption and aneuploidy, relying on fungal-induced modification of expression of several cell cycle checkpoint regulators in S and G2/M phases. On the other hand, C. neoformans infection induced macrophage apoptosis in a capsule-dependent manner with a differential requirement of the classical and alternative NF-κB signalling pathways, the latter one being essential. Together, these findings shed new light on fungal strategies to subvert host response through uncoupling of NF-κB activity in pathogen-controlled apoptosis and impairment of cell cycle progression. They also provide the first demonstration of induction of aneuploidy by a
Fungal-induced cell cycle impairment, chromosome instability and apoptosis via differential activation of NF-κB.

PubMed

Ben-Abdallah, Mariem; Sturny-Leclère, Aude; Avé, Patrick; Louise, Anne; Moyrand, Frédérique; Weih, Falk; Janbon, Guilhem; Mémet, Sylvie

2012-01-01

Microbial pathogens have developed efficient strategies to compromise host immune responses. Cryptococcus neoformans is a facultative intracellular pathogen, recognised as the most common cause of systemic fungal infections leading to severe meningoencephalitis, mainly in immunocompromised patients. This yeast is characterized by a polysaccharide capsule, which inhibits its phagocytosis. Whereas phagocytosis escape and macrophage intracellular survival have been intensively studied, extracellular survival of this yeast and restraint of host innate immune response are still poorly understood. In this study, we have investigated whether C. neoformans affected macrophage cell viability and whether NF-κB (nuclear factor-κB), a key regulator of cell growth, apoptosis and inflammation, was involved. Using wild-type (WT) as well as mutant strains of C. neoformans for the pathogen side, and WT and mutant cell lines with altered NF-κB activity or signalling as well as primary macrophages for the host side, we show that C. neoformans manipulated NF-κB-mediated signalling in a unique way to regulate macrophage cell fate and viability. On the one hand, serotype A strains reduced macrophage proliferation in a capsule-independent fashion. This growth decrease, which required a critical dosage of NF-κB activity, was caused by cell cycle disruption and aneuploidy, relying on fungal-induced modification of expression of several cell cycle checkpoint regulators in S and G2/M phases. On the other hand, C. neoformans infection induced macrophage apoptosis in a capsule-dependent manner with a differential requirement of the classical and alternative NF-κB signalling pathways, the latter one being essential. Together, these findings shed new light on fungal strategies to subvert host response through uncoupling of NF-κB activity in pathogen-controlled apoptosis and impairment of cell cycle progression. They also provide the first demonstration of induction of aneuploidy by a
Vinpocetine inhibits amyloid-beta induced activation of NF-κB, NLRP3 inflammasome and cytokine production in retinal pigment epithelial cells.

PubMed

Liu, Ruozhou Tom; Wang, Aikun; To, Eleanor; Gao, Jiangyuan; Cao, Sijia; Cui, Jing Z; Matsubara, Joanne A

2014-10-01

Chronic inflammation is a key pathogenic process in age-related macular degeneration (AMD). Amyloid-beta (Aβ) is a constituent of AMD drusen and promotes the activation of NLRP3 inflammasome which facilitates the production of cytokines. We investigated the role of transcription factor NF-κB in the activation of inflammasome in the RPE and the effect of vinpocetine, a dietary supplement with inhibitory effect on NF-κΒ. ARPE19/NF-κB-luciferase reporter cells treated with Aβ demonstrated enhanced NF-κB activation that was significantly suppressed by vinpocetine. Intraperitoneal injection of vinpocetine (15 mg/kg) inhibited NF-κB nuclear translocation and reduced the expression and activation of NLRP3, caspase-1, IL-1β, IL-18, and TNF-α in the RPE of adult rats that received intraocular Αβ, as measured by retinal immunohistochemistry and Western blot. Cytokine level in the vitreous was assayed using multiplex suspension arrays and revealed significantly lower concentration of MIP-3α, IL-6, IL-1α, IL-1β, IL-18, and TNF-α in vinpocetine treated animals. These results suggest that the NF-κB pathway is activated by Aβ in the RPE and signals the priming of NLRP3 inflammasome and the expression of pro-inflammatory cytokines including the inflammasome substrates IL-1β and IL-18. NF-κB inhibition may be an effective approach to stem the chronic inflammatory milieu that underlies the development of AMD. Vinpocetine is a potentially useful anti-inflammatory agent that is well-tolerated in long term use. Copyright © 2014 Elsevier Ltd. All rights reserved.
Serum albumin forms a lactoferrin-like soluble iron-binding complex in presence of hydrogen carbonate ions.

PubMed

Ueno, Hiroshi M; Urazono, Hiroshi; Kobayashi, Toshiya

2014-02-15

The iron-lactoferrin complex is a common food ingredient because of its iron-solubilizing capability in the presence of hydrogen carbonate ions. However, it is unclear whether the formation of a stable iron-binding complex is limited to lactoferrin. In this study, we investigated the effects of bovine serum albumin (BSA) on iron solubility and iron-catalyzed lipid oxidation in the presence of hydrogen carbonate ions. BSA could solubilize >100-fold molar equivalents of iron at neutral pH, exceeding the specific metal-binding property of BSA. This iron-solubilizing capability of BSA was impaired by thermally denaturing BSA at ≥ 70 °C for 10 min at pH 8.5. The resulting iron-BSA complex inhibited iron-catalyzed oxidation of soybean oil in a water-in-oil emulsion measured using the Rancimat test. Our study is the first to show that BSA, like lactoferrin, forms a soluble iron-binding complex in the presence of hydrogen carbonate ions. Copyright © 2013 Elsevier Ltd. All rights reserved.
Egg yolks inhibit activation of NF-κB and expression of its target genes in adipocytes after partial delipidation

PubMed Central

Shen, Qiwen; Riedl, Ken M.; Cole, Rachel M.; Lehman, Christopher; Xu, Lu; Alder, Hansjuerg; Belury, Martha A.; Schwartz, Steven J.; Ziouzenkova, Ouliana

2015-01-01

How composition of egg yolk (EY) influences NF-κB, a key transcription pathway in inflammation, remains unclear. We performed partial delipidation of EY that removed 20–30% of cholesterol and triglycerides. The resulting polar and non-polar fractions were termed EY-P and EY-NP. NF-κB activation in response to EY from different suppliers and their fractions was examined in 3T3-L1 adipocytes using a NF-κB response element reporter assay and by analyzing expression of 248 inflammatory genes. Although EY-P and EY contained similar level of vitamins, carotenoids, and fatty acids, only delipidated EY-P fraction suppressed NF-κB via down-regulation of toll like receptor-2 and up-regulation of inhibitory toll interacting protein (Tollip) and lymphocyte antigen 96 (Ly96). Our data suggest that anti-inflammatory activity of lutein and retinol were blunted by non-polar lipids in EY likely via crosstalk between SREBP and NF-κB pathways in adipocytes. Thus, moderate delipidation may improve their beneficial properties of regular eggs. PMID:25620076
Subneurotoxic copper(II)-induced NF-κB-dependent microglial activation is associated with mitochondrial ROS

DOE Office of Scientific and Technical Information (OSTI.GOV)

Hu, Zhuqin; Yu, Fengxiang; Gong, Ping

2014-04-15

Microglia-mediated neuroinflammation and the associated neuronal damage play critical roles in the pathogenesis of neurodegenerative disorders. Evidence shows an elevated concentration of extracellular copper(II) in the brains of these disorders, which may contribute to neuronal death through direct neurotoxicity. Here we explored whether extracellular copper(II) triggers microglial activation. Primary rat microglia and murine microglial cell line BV-2 cells were cultured and treated with copper(II). The content of tumor necrosis factor-α (TNF-α) and nitric oxide in the medium was determined. Extracellular hydrogen peroxide was quantified by a fluorometric assay with Amplex Red. Mitochondrial superoxide was measured by MitoSOX oxidation. At subneurotoxicmore » concentrations, copper(II) treatment induced a dose- and time-dependent release of TNF-α and nitric oxide from microglial cells, and caused an indirect, microglia-mediated neurotoxicity that was blocked by inhibition of TNF-α and nitric oxide production. Copper(II)-initiated microglial activation was accompanied with reduced IkB-α expression as well as phosphorylation and translocation of nuclear factor-κB (NF-κB) p65 and was blocked by NF-κB inhibitors (BAY11-7082 and SC-514). Moreover, copper(II) treatment evoked a rapid release of hydrogen peroxide from microglial cells, an effect that was not affected by NADPH oxidase inhibitors. N-acetyl-cysteine, a scavenger of reactive oxygen species (ROS), abrogated copper(II)-elicited microglial release of TNF-α and nitric oxide and subsequent neurotoxicity. Importantly, mitochondrial production of superoxide, paralleled to extracellular release of hydrogen peroxide, was induced after copper(II) stimulation. Our findings suggest that extracellular copper(II) at subneurotoxic concentrations could trigger NF-κB-dependent microglial activation and subsequent neurotoxicity. NADPH oxidase-independent, mitochondria-derived ROS may be involved in this activation
Increased p50/p50 NF-κB Activation in Human Papillomavirus Type 6- or Type 11-Induced Laryngeal Papilloma Tissue

PubMed Central

Vancurova, Ivana; Wu, Rong; Miskolci, Veronika; Sun, Shishinn

2002-01-01

We have observed elevated NF-κB DNA-binding activity in nuclear extracts from human papillomavirus type 6- and 11-infected laryngeal papilloma tissues. The predominant DNA-binding species is the p50/p50 homodimer. The elevated NF-κB activity could be correlated with a reduced level of cytoplasmic IκBβ and could be associated with the overexpression of p21CIP1/WAF1 in papilloma cells. Increased NF-κB activity and cytoplasmic accumulation of p21CIP1/WAF1 might counteract death-promoting effects elicited by overexpressed PTEN and reduced activation of Akt and STAT3 previously noted in these tissues. PMID:11773428
TNFα- and IKKβ-mediated TANK/I-TRAF phosphorylation: implications for interaction with NEMO/IKKγ and NF-κB activation

PubMed Central

Bonif, Marianne; Meuwis, Marie-Alice; Close, Pierre; Benoit, Valérie; Heyninck, Karen; Chapelle, Jean-Paul; Bours, Vincent; Merville, Marie-Paule; Piette, Jacques; Beyaert, Rudi; Chariot, Alain

2005-01-01

Pro-inflammatory cytokines trigger signalling cascades leading to NF-κB (nuclear factor-κB)-dependent gene expression through IKK [IκB (inhibitory κB) kinase]-dependent phosphorylation and subsequent degradation of the IκB proteins and via induced phosphorylation of p65. These signalling pathways rely on sequentially activated kinases which are assembled by essential and non-enzymatic scaffold proteins into functional complexes. Here, we show that the pro-inflammatory cytokine TNFα (tumour necrosis factor α) promotes TANK [TRAF (TNF receptor-associated factor) family member associated NF-κB activator] recruitment to the IKK complex via a newly characterized C-terminal zinc finger. Moreover, we show that TANK is phosphorylated by IKKβ upon TNFα stimulation and that this modification negatively regulates TANK binding to NEMO (NF-κB essential modulator). Interestingly, reduced TANK expression by RNA interference attenuates TNFα-mediated induction of a subset of NF-κB target genes through decreased p65 transactivation potential. Therefore the scaffold protein TANK is required for the cellular response to TNFα by connecting upstream signalling molecules to the IKKs and p65, and its subsequent IKKβ-mediated phosphorylation may be a mechanism to terminate the TANK-dependent wave of NF-κB activation. PMID:16336209
Lymphotoxin activation by human T-cell leukemia virus type I-infected cell lines: role for NF-kappa B.

PubMed

Paul, N L; Lenardo, M J; Novak, K D; Sarr, T; Tang, W L; Ruddle, N H

1990-11-01

Human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines constitutively produce high levels of biologically active lymphotoxin (LT; tumor necrosis factor-beta) protein and LT mRNA. To understand the regulation of LT transcription by HTLV-I, we analyzed the ability of a series of deletions of the LT promoter to drive the chloramphenicol acetyltransferase (CAT) reporter gene in HTLV-I-positive MT-2 cells. The smallest LT promoter fragment (-140 to +77) that was able to drive CAT activity contained a site that was similar to the immunoglobulin kappa-chain NF-kappa B-binding site. Since the HTLV-I tax gene activates the nuclear form of NF-kappa B, this finding suggested a possible means of HTLV-I activation of LT production. We found that the LT kappa B-like site specifically formed a complex with NF-kappa B-containing nuclear extract from MT-2, C81-66-45, and other activated T cells. Mutation of the LT kappa B site in the context of the LT promoter (-293 to +77) (mutant M1) reduced the ability of the promoter to drive the CAT gene in HTLV-I-infected and noninfected human T-cell lines. These data suggest a general role for NF-kappa B activation in the induction of LT gene transcription. Activation of LT in HTLV-I-infected cells may explain the pathology associated with HTLV-I infection, including the hypercalcemia that is prevalent in adult T-cell leukemia.
Growth characteristics of calves fed an intensified milk replacer regimen with additional lactoferrin.

PubMed

Cowles, K E; White, R A; Whitehouse, N L; Erickson, P S

2006-12-01

The objective of this study was to evaluate the effect of lactoferrin addition to milk replacer varying in crude protein (CP) on dry matter intake, growth, and days medicated. Thirty-four Holstein heifer calves were assigned to 4 treatments in a 2 x 2 factorial arrangement of treatments in a randomized complete block design. Treatments were as follows: 562 g daily of a nonmedicated conventional milk replacer (20% CP:20% fat) feeding regimen with or without 1 g of supplemental bovine lactoferrin (n = 9 for both treatments) or a nonmedicated intensified milk replacer feeding regimen (28% CP:20% fat) fed on a metabolizable energy basis (0.2 Mcal/kg BW(0.75)) from d 2 to 9, and at 0.27 Mcal/kg BW(0.75) from d 10 to 42 with or without 1g supplemental bovine lactoferrin (n = 8 for both treatments). Calves were fed pelleted starter (25% CP) in 227.5-g increments beginning on d 2 and had free access to water. Calves remained on the study for 14 d postweaning. Dry matter intake was determined daily. Growth measurements were taken weekly. Blood samples were taken twice weekly for determination of blood urea N. On d 10 of life, calves were subjected to a xylose challenge. Calves on conventional treatments ate more starter preweaning, during weaning, and postweaning. Preweaning, intensively fed calves had higher dry matter intakes. Weights of intensified-fed calves were greater at weaning. Intensified milk replacer-fed calves had greater average daily gain preweaning and overall and higher gain:feed ratios preweaning, but conventionally fed calves had higher gain:feed ratios during weaning. Intensified milk replacer-fed calves had greater hip heights during weaning and postweaning and greater heart girths preweaning, weaning, and postweaning. Days medicated were greater preweaning and overall for intensified-fed calves. There were no differences among treatments for xylose absorption. Calves on conventional treatments had increased blood urea nitrogen concentrations preweaning
Estrogen deficiency inhibits the odonto/osteogenic differentiation of dental pulp stem cells via activation of the NF-κB pathway.

PubMed

Wang, Yanping; Yan, Ming; Yu, Yan; Wu, Jintao; Yu, Jinhua; Fan, Zhipeng

2013-06-01

Various factors can affect the functions of dental pulp stem cells (DPSCs). However, little knowledge is available about the effects of estrogen deficiency on the differentiation of DPSCs. In this study, an estrogen-deficient rat model was constructed and multi-colony-derived DPSCs were obtained from the incisors of ovariectomized (OVX) or sham-operated rats. Odonto/osteogenic differentiation and the possible involvement of the nuclear factor kappa B (NF-κB) pathway in the OVX-DPSCs/Sham-DPSCs of these rats were then investigated. OVX-DPSCs presented decreased odonto/osteogenic capacity and an activated NF-κB pathway, as compared with Sham-DPSCs. When the cellular NF-κB pathway was specifically inhibited by BMS345541, the odonto/osteogenic potential in OVX-DPSCs was significantly upregulated. Thus, estrogen deficiency down-regulated the odonto/osteogenic differentiation of DPSCs by activating NF-κB signaling and inhibition of the NF-κB pathway effectively rescued the decreased differentiation potential of DPSCs.
The lymphotoxin promoter is stimulated by HTLV-I tax activation of NF-kappa B in human T-cell lines.

PubMed

Paul, N L; Millet, I; Ruddle, N H

1993-07-01

The HTLV-I transcriptional activator tax was used to gain insight into the mechanism of lymphotoxin (LT; TNF-beta) gene induction. Tax-expressing cell lines produce LT biologic activity. An LT promoter (LT-293) CAT construct that contained an NF-kappa B site was active in the LT-producing C81-66-45 cell line, which contains defective HTLV-I but expresses tax. The observation that a mutated LT-kappa B construct (M1-CAT) was inactive in C81-66-45, confirmed the importance of NF-kappa B in LT gene expression. Tax was transfected into HTLV-I-negative human T-cell lines. Jurkat T cells stably expressing tax contained elevated levels of NF-kappa B that directly bound to the LT-kappa B site. Tax co-transfected with reporter constructs into Jurkat cells maximally activated HTLV-I-LTR-CAT and kappa B-fos-CAT and also activated LT-293 to a lesser extent. In JM T cells, tax induced LT-293 activity by two- to four-fold, though there was no induction of M1-CAT. The increase in LT-293 CAT activity mirrored the increase in LT biologic activity seen under these conditions. These studies, the first to demonstrate induction of LT promoter activity over basal levels, indicate that HTLV-I tax causes low-level activation of both endogenous LT and the LT promoter, at least in part through activation of NF-kappa B.
A Novel Pathway Links Oxidative Stress to Loss of Insulin Growth Factor-2 (IGF2) Imprinting through NF-κB Activation

PubMed Central

Yang, Bing; Wagner, Jennifer; Damaschke, Nathan; Yao, Tianyu; Wuerzberger-Davis, Shelly M.; Lee, Moon-Hee; Svaren, John; Miyamoto, Shigeki; Jarrard, David F.

2014-01-01

Genomic imprinting is the allele-specific expression of a gene based on parental origin. Loss of imprinting(LOI) of Insulin-like Growth Factor 2 (IGF2) during aging is important in tumorigenesis, yet the regulatory mechanisms driving this event are largely unknown. In this study oxidative stress, measured by increased NF-κB activity, induces LOI in both cancerous and noncancerous human prostate cells. Decreased expression of the enhancer-blocking element CCCTC-binding factor(CTCF) results in reduced binding of CTCF to the H19-ICR (imprint control region), a major factor in the allelic silencing of IGF2. This ICR then develops increased DNA methylation. Assays identify a recruitment of the canonical pathway proteins NF-κB p65 and p50 to the CTCF promoter associated with the co-repressor HDAC1 explaining gene repression. An IκBα super-repressor blocks oxidative stress-induced activation of NF-κB and IGF2 imprinting is maintained. In vivo experiments using IκBα mutant mice with continuous NF-κB activation demonstrate increased IGF2 LOI further confirming a central role for canonical NF-κB signaling. We conclude CTCF plays a central role in mediating the effects of NF-κB activation that result in altered imprinting both in vitro and in vivo. This novel finding connects inflammation found in aging prostate tissues with the altered epigenetic landscape. PMID:24558376
Failure in activation of the canonical NF-κB pathway by human T-cell leukemia virus type 1 Tax in non-hematopoietic cell lines

DOE Office of Scientific and Technical Information (OSTI.GOV)

Mizukoshi, Terumi; Komori, Hideyuki; Mizuguchi, Mariko

2013-09-01

Human T-cell leukemia virus type 1 (HTLV-1) Tax (Tax1) plays crucial roles in leukemogenesis in part through activation of NF-κB. In this study, we demonstrated that Tax1 activated an NF-κB binding (gpκB) site of the gp34/OX40 ligand gene in a cell type-dependent manner. Our examination showed that the gpκΒ site and authentic NF-κB (IgκB) site were activated by Tax1 in hematopoietic cell lines. Non-hematopoietic cell lines including hepatoma and fibroblast cell lines were not permissive to Tax1-mediated activation of the gpκB site, while the IgκB site was activated in those cells in association with binding of RelB. However RelA bindingmore » was not observed in the gpκB and IgκB sites. Our results suggest that HTLV-1 Tax1 fails to activate the canonical pathway of NF-κB in non-hematopoietic cell lines. Cell type-dependent activation of NF-κB by Tax1 could be associated with pathogenesis by HTLV-1 infection. - Highlights: • HTLV-1 Tax1 does not activate RelA of NF-κB in non-hematopoietic cell lines. • Tax1 activates the NF-κB non-canonical pathway in non-hematopoietic cell lines. • Tax1 does not induce RelA nuclear translocation in those cell lines, unlike TNFα. • The OX40L promoter κB site is activated by ectopic, but not endogenous, RelA.« less
Trastuzumab-Resistant Luminal B Breast Cancer Cells Show Basal-Like Cell Growth Features Through NF-κB-Activation

PubMed Central

Kanzaki, Hirotaka; Mukhopadhya, Nishit K.; Cui, Xiaojiang; Ramanujan, V. Krishnan

2016-01-01

A major clinical problem in the treatment of breast cancer is mortality due to metastasis. Understanding the molecular mechanisms associated with metastasis should aid in designing new therapeutic approaches for breast cancer. Trastuzumab is the main therapeutic option for HER2+ breast cancer patients; however, the molecular basis for trastuzumab resistance (TZR) and subsequent metastasis is not known. Earlier, we found expression of basal-like molecular markers in TZR tissues from patients with invasive breast cancer.(1) The basal-like phenotype is a particularly aggressive form of breast cancer. This observation suggests that TZR might contribute to an aggressive phenotype. To understand if resistance to TZR can lead to basal-like phenotype, we generated a trastuzumab-resistant human breast cancer cell line (BT-474-R) that maintained human epidermal growth factor receptor 2 (HER2) overexpression and HER2 mediated signaling. Analysis showed that nuclear factor-kappa B (NF-κB) was constitutively activated in the BT-474-R cells, a feature similar to the basal-like tumor phenotype. Pharmacologic inhibition of NF-κB improved sensitivity of BT-474-R cells to trastuzumab. Interestingly, activation of HER2 independent NF-κB is not shown in luminal B breast cancer cells. Our study suggests that by activating the NF-κB pathway, luminal B cells may acquire a HER2+ basal-like phenotype in which NF-κB is constitutively activated; this notion is consistent with the recently proposed “progression through grade” or “evolution of resistance” hypothesis. Furthermore, we identified IKK-α/IKK-β and nuclear accumulation of RelA/p65 as the major determinants in the resistant cells. Thus our study additionally suggests that the nuclear accumulation of p65 may be a useful marker for identifying metastasis-initiating tumor cells and targeting RelA/p65 may limit metastasis of breast and other cancers associated with NF-κB activation. PMID:26871511
Development of novel NEMO-binding domain mimetics for inhibiting IKK/NF-κB activation.

PubMed

Zhao, Jing; Zhang, Lei; Mu, Xiaodong; Doebelin, Christelle; Nguyen, William; Wallace, Callen; Reay, Daniel P; McGowan, Sara J; Corbo, Lana; Clemens, Paula R; Wilson, Gabriela Mustata; Watkins, Simon C; Solt, Laura A; Cameron, Michael D; Huard, Johnny; Niedernhofer, Laura J; Kamenecka, Theodore M; Robbins, Paul D

2018-06-11

Nuclear factor κB (NF-κB) is a transcription factor important for regulating innate and adaptive immunity, cellular proliferation, apoptosis, and senescence. Dysregulation of NF-κB and its upstream regulator IκB kinase (IKK) contributes to the pathogenesis of multiple inflammatory and degenerative diseases as well as cancer. An 11-amino acid peptide containing the NF-κB essential modulator (NEMO)-binding domain (NBD) derived from the C-terminus of β subunit of IKK, functions as a highly selective inhibitor of the IKK complex by disrupting the association of IKKβ and the IKKγ subunit NEMO. A structure-based pharmacophore model was developed to identify NBD mimetics by in silico screening. Two optimized lead NBD mimetics, SR12343 and SR12460, inhibited tumor necrosis factor α (TNF-α)- and lipopolysaccharide (LPS)-induced NF-κB activation by blocking the interaction between IKKβ and NEMO and suppressed LPS-induced acute pulmonary inflammation in mice. Chronic treatment of a mouse model of Duchenne muscular dystrophy (DMD) with SR12343 and SR12460 attenuated inflammatory infiltration, necrosis and muscle degeneration, demonstrating that these small-molecule NBD mimetics are potential therapeutics for inflammatory and degenerative diseases.
Targeting Activation of Specific NF-κB Subunits Prevents Stress-Dependent Atherothrombotic Gene Expression

PubMed Central

Djuric, Zdenka; Kashif, Muhammed; Fleming, Thomas; Muhammad, Sajjad; Piel, David; von Bauer, Rüdiger; Bea, Florian; Herzig, Stephan; Zeier, Martin; Pizzi, Marina; Isermann, Berend; Hecker, Markus; Schwaninger, Markus; Bierhaus, Angelika; Nawroth, Peter P

2012-01-01

Psychosocial stress has been shown to be a contributing factor in the development of atherosclerosis. Although the underlying mechanisms have not been elucidated entirely, it has been shown previously that the transcription factor nuclear factor-κB (NF-κB) is an important component of stress-activated signaling pathway. In this study, we aimed to decipher the mechanisms of stress-induced NF-κB-mediated gene expression, using an in vitro and in vivo model of psychosocial stress. Induction of stress led to NF-κB-dependent expression of proinflammatory (tissue factor, intracellular adhesive molecule 1 [ICAM-1]) and protective genes (manganese superoxide dismutase [MnSOD]) via p50, p65 or cRel. Selective inhibition of the different subunits and the respective kinases showed that inhibition of cRel leads to the reduction of atherosclerotic lesions in apolipoprotein−/− (ApoE−/−) mice via suppression of proinflammatory gene expression. This observation may therefore provide a possible explanation for ineffectiveness of antioxidant therapies and suggests that selective targeting of cRel activation may provide a novel approach for the treatment of stress-related inflammatory vascular disease. PMID:23114885
Combination chemotherapy increases cytotoxicity of multiple myeloma cells by modification of nuclear factor (NF)-κB activity

PubMed Central

Salem, Kelley; Brown, Charles O.; Schibler, Jeanine; Goel, Apollina

2012-01-01

The NF-κB signaling pathway is critical in myeloma cell proliferation, inhibition of apoptosis, and emergence of therapy resistance. The chemotherapeutic drugs, dexamethasone (Dex) and bortezomib (BTZ), are widely used in clinical protocols for multiple myeloma (MM) and inhibit the NF-κB signaling pathway by distinct mechanisms. This study evaluates the efficacy of combination therapy with Dex and BTZ and investigates the mechanistic underpinning of endogenous and therapy-induced NF-κB activation in MM. Human myeloma cells and bone marrow stromal cells (BMSCs) were used in monocultures and co-cultures to determine the cytotoxic effects of Dex and/or BTZ. Our results show that combined treatment of Dex with BTZ enhanced direct apoptosis of drug-sensitive and drug-resistant myeloma cells. In the presence of BMSCs, Dex plus BTZ combination inhibited ionizing radiation (IR)-induced interleukin (IL)-6 secretion from BMSCs and induced myeloma cytotoxicity. Mechanistically, Dex treatment increased IκBα protein and mRNA expression and compensated for BTZ-induced IκBα degradation. Dex plus BTZ combination inhibited basal and therapy-induced NF-κB activity with cytotoxicity in myeloma cells resistant to BTZ. Furthermore, combination therapy down-regulated the NF-κB targeted gene expression of IL-6 and manganese superoxide dismutase (MnSOD), which can induce chemo- and radio-resistance in MM. This study provides mechanistic rationale for combining the NF-κB-targeting drugs Dex and BTZ in myeloma therapy and supports potential combinations of these drugs with radiotherapy and additional chemotherapeutic drugs, for clinical benefit in MM. PMID:23063726
Effect of iron saturation level of lactoferrin on osteogenic activity in vitro and in vivo.

PubMed

Wang, X Y; Guo, H Y; Zhang, W; Wen, P C; Zhang, H; Guo, Z R; Ren, F Z

2013-01-01

We studied the effect of iron saturation level on the osteogenic activity of lactoferrin (LF) in vitro and in vivo. Different iron saturation levels of LF (1.0, 9.0, 38, 58, and 96%) were prepared as the following samples: apo-LF, LF-9, LF-38, LF-58, and holo-LF. Using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay, we observed that the stimulating osteoblast proliferation activity of LF in vitro decreased with increasing iron saturation level at 100 and 1,000 μg/mL. In vivo, 4-wk-old ICR Swiss male mice were randomly divided into 4 groups: blank control (physiological saline), negative control (BSA), apo-LF, and holo-LF. Four groups of mice were injected subcutaneously with physiological saline, BSA, apo-LF, or holo-LF over the calvarial surface twice a day for 5 consecutive days at a dose of 4 mg/kg per day. Bone histomorphometry showed that new bone formation (assessed using tetracycline-HCl labels) tended to be stronger with apo-LF than with holo-LF. Using fluorescence spectroscopy and circular dichroism measurements, we found that exposure of tryptophan increased, α-helix content increased, but β-structure content decreased with increasing iron saturation level. These findings indicated that the osteogenic activity of LF decreases with increasing iron saturation level in vitro and in vivo, which may be related to conformational changes in LF. Copyright © 2013 American Dairy Science Association. Published by Elsevier Inc. All rights reserved.
Src-family-tyrosine kinase Lyn is critical for TLR2-mediated NF-κB activation through the PI 3-kinase signaling pathway.

PubMed

Toubiana, Julie; Rossi, Anne-Lise; Belaidouni, Nadia; Grimaldi, David; Pene, Frederic; Chafey, Philippe; Comba, Béatrice; Camoin, Luc; Bismuth, Georges; Claessens, Yann-Erick; Mira, Jean-Paul; Chiche, Jean-Daniel

2015-10-01

TLR2 has a prominent role in host defense against a wide variety of pathogens. Stimulation of TLR2 triggers MyD88-dependent signaling to induce NF-κB translocation, and activates a Rac1-PI 3-kinase dependent pathway that leads to transactivation of NF-κB through phosphorylation of the P65 NF-κB subunit. This transactivation pathway involves tyrosine phosphorylations. The role of the tyrosine kinases in TLR signaling is controversial, with discrepancies between studies using only chemical inhibitors and knockout mice. Here, we show the involvement of the tyrosine-kinase Lyn in TLR2-dependent activation of NF-κB in human cellular models, by using complementary inhibition strategies. Stimulation of TLR2 induces the formation of an activation cluster involving TLR2, CD14, PI 3-kinase and Lyn, and leads to the activation of AKT. Lyn-dependent phosphorylation of the p110 catalytic subunit of PI 3-kinase is essential to the control of PI 3-kinase biological activity upstream of AKT and thereby to the transactivation of NF-κB. Thus, Lyn kinase activity is crucial in TLR2-mediated activation of the innate immune response in human mononuclear cells. © The Author(s) 2015.

Sustained, neuron-specific IKK/NF-κB activation generates a selective neuroinflammatory response promoting local neurodegeneration with aging

PubMed Central

2013-01-01

Background Increasing evidence indicates that neuroinflammation is a critical factor contributing to the progression of various neurodegenerative diseases. The IKK/NF-κB signalling system is a central regulator of inflammation, but it also affects neuronal survival and differentiation. A complex interplay between different CNS resident cells and infiltrating immune cells, which produce and respond to various inflammatory mediators, determines whether neuroinflammation is beneficial or detrimental. The IKK/NF-κB system is involved in both production of and responses to these mediators, although the precise contribution depends on the cell type as well as the cellular context, and is only partially understood. Here we investigated the specific contribution of neuronal IKK/NF-κB signalling on the regulation of neuroinflammatory processes and its consequences. To address this issue, we established and analysed a conditional gain-of-function mouse model that expresses a constitutively active allele of IKK2 in principal forebrain neurons (IKK2nCA). Proinflammatory gene and growth factor expression, histopathology, microgliosis, astrogliosis, immune cell infiltration and spatial learning were assessed at different timepoints after persistent canonical IKK2/NF-κB activation. Results In contrast to other cell types and organ systems, chronic IKK2/NF-κB signalling in forebrain neurons of adult IKK2nCA animals did not cause a full-blown inflammatory response including infiltration of immune cells. Instead, we found a selective inflammatory response in the dentate gyrus characterized by astrogliosis, microgliosis and Tnf-α upregulation. Furthermore, downregulation of the neurotrophic factor Bdnf correlated with a selective and progressive atrophy of the dentate gyrus and a decline in hippocampus-dependent spatial learning. Neuronal degeneration was associated with increased Fluoro-jade staining, but lacked activation of apoptosis. Remarkably, neuronal loss could be
Suppression of NF-κB and NF-κB-Regulated Gene Expression by Apigenin through IκBα and IKK Pathway in TRAMP Mice

PubMed Central

Shukla, Sanjeev; Shankar, Eswar; Fu, Pingfu; MacLennan, Gregory T.; Gupta, Sanjay

2015-01-01

Aberrant Nuclear Factor-κappaB (NF-κB) activation due to rapid IκBα turnover and high basal IκBα kinase (IKK) activity has been frequently observed in prostate cancer. Apigenin, a naturally occurring plant flavone, exhibits anti-proliferative, anti-inflammatory and anti-carcinogenic activities by inhibiting NF-κB pathway, through a mechanism not fully understood. We found that apigenin feeding in microgram doses (bioavailable in humans) inhibited prostate tumorigenesis in TRAMP mice by interfering with NF-κB signaling. Apigenin feeding to TRAMP mice (20 and 50 μg/mouse/day, 6 days/week for 20 weeks) exhibited significant decrease in tumor volumes of the prostate and completely abolished metastasis, which correlated with inhibition of NF-κB activation and binding to the DNA. Apigenin intake blocked phosphorylation and degradation of IκBα by inhibiting IKK activation, which in turn led to suppression of NF-κB activation. The expression of NF-κB-regulated gene products involved in proliferation (cyclin D1, and COX-2), anti-apoptosis (Bcl-2 and Bcl-xL), and angiogenesis (vascular endothelial growth factor) were also downregulated after apigenin feeding. These events correlated with the induction of apoptosis in tumor cells, as evident by increased cleaved caspase-3 labeling index in the dorsolateral prostate. Our results provide convincing evidence that apigenin inhibits IKK activation and restores the expression of IκBα, preventing it’s phosphorylation in a fashion similar to that elicited by IKK and proteasomal inhibitors through suppression of NF-κB signaling pathway. PMID:26379052
Inhibition of NF-κB activity in the hypothalamic paraventricular nucleus attenuates hypertension and cardiac hypertrophy by modulating cytokines and attenuating oxidative stress

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu, Xiao-Jing; Zhang, Dong-Mei; Jia, Lin-Lin

We hypothesized that chronic inhibition of NF-κB activity in the hypothalamic paraventricular nucleus (PVN) delays the progression of hypertension and attenuates cardiac hypertrophy by up-regulating anti-inflammatory cytokines, reducing pro-inflammatory cytokines (PICs), attenuating nuclear factor-κB (NF-κB) p65 and NAD(P)H oxidase in the PVN of young spontaneously hypertensive rats (SHR). Young normotensive Wistar–Kyoto (WKY) and SHR rats received bilateral PVN infusions with NF–κB inhibitor pyrrolidine dithiocarbamate (PDTC) or vehicle for 4 weeks. SHR rats had higher mean arterial pressure and cardiac hypertrophy as indicated by increased whole heart weight/body weight ratio, whole heart weight/tibia length ratio, left ventricular weight/tibia length ratio, cardiomyocytemore » diameters of the left cardiac ventricle, and mRNA expressions of cardiac atrial natriuretic peptide (ANP) and beta-myosin heavy chain (β-MHC). These SHR rats had higher PVN levels of proinflammatory cytokines (PICs), reactive oxygen species (ROS), the chemokine monocyte chemoattractant protein-1 (MCP-1), NAD(P)H oxidase activity, mRNA expression of NOX-2 and NOX-4, and lower PVN IL-10, and higher plasma levels of PICs and NE, and lower plasma IL-10. PVN infusion of NF-κB inhibitor PDTC attenuated all these changes. These findings suggest that NF-κB activation in the PVN increases sympathoexcitation and hypertensive response, which are associated with the increases of PICs and oxidative stress in the PVN; PVN inhibition of NF-κB activity attenuates PICs and oxidative stress in the PVN, thereby attenuates hypertension and cardiac hypertrophy. - Highlights: • Spontaneously hypertensive rats exhibit neurohormonal excitation in the PVN. • PVN inhibition of NF-κB attenuates hypertension-induced cardiac hypertrophy. • PVN inhibition of NF-κB attenuates hypertension-induced neurohormonal excitation. • PVN inhibition of NF-κB attenuates hypertension-induced imbalance of
Influence of deoxynivalenol on NF-kappaB activation and IL-8 secretion in human intestinal Caco-2 cells.

PubMed

Van De Walle, Jacqueline; Romier, Béatrice; Larondelle, Yvan; Schneider, Yves-Jacques

2008-04-01

Deoxynivalenol (DON) is the most prevalent trichothecene mycotoxin in crops in Europe and North America. In human intestinal Caco-2 cells, DON activates the mitogen-activated protein kinases (MAPKs). We hypothesized a link between DON ingestion and intestinal inflammation, and used Caco-2 cells to assess the effects of DON, at plausible intestinal concentrations (250-10,000 ng/ml), on inflammatory mediators acting downstream the MAPKs cascade i.e. activation of nuclear factor-kappaB (NF-kappaB) and interleukin-8 (IL-8) secretion. In addition, Caco-2 cells were co-exposed to pro-inflammatory stimuli in order to mimic an inflamed intestinal epithelium. Dose-dependent increases in NF-kappaB activity and IL-8 secretion were observed, reaching 1.4- and 7.6-fold, respectively using DON at 10 microg/ml. Phosphorylation of inhibitor-kappaB (IkappaB) increased (1.6-fold) at DON levels <0.5 microg/ml. Exposure of Caco-2 cells to pro-inflammatory agents, i.e. 25 ng/ml interleukin-1beta, 100 ng/ml tumor necrosis factor-alpha or 10 microg/ml lipopolysaccharides, activated NF-kappaB and increased IL-8 secretion. Synergistic interactions between these stimuli and DON were observed. These data show that DON induces NF-kappaB activation and IL-8 secretion dose-dependently in Caco-2 cells, and this effect was accentuated upon pro-inflammatory stimulation, suggesting DON exposure could cause or exacerbate intestinal inflammation.
Assessment of the potential pathogenicity of missense mutations identified in the GTPase-activating protein (GAP)-related domain of the neurofibromatosis type-1 (NF1) gene.

PubMed

Thomas, Laura; Richards, Mark; Mort, Matthew; Dunlop, Elaine; Cooper, David N; Upadhyaya, Meena

2012-12-01

Neurofibromatosis type-1 (NF1) is caused by constitutional mutations of the NF1 tumor-suppressor gene. Although ∼85% of inherited NF1 microlesions constitute truncating mutations, the remaining ∼15% are missense mutations whose pathological relevance is often unclear. The GTPase-activating protein-related domain (GRD) of the NF1-encoded protein, neurofibromin, serves to define its major function as a negative regulator of the Ras-MAPK (mitogen-activated protein kinase) signaling pathway. We have established a functional assay to assess the potential pathogenicity of 15 constitutional nonsynonymous NF1 missense mutations (11 novel and 4 previously reported but not functionally characterized) identified in the NF1-GRD (p.R1204G, p.R1204W, p.R1276Q, p.L1301R, p.I1307V, p.T1324N, p.E1327G, p.Q1336R, p.E1356G, p.R1391G, p.V1398D, p.K1409E, p.P1412R, p.K1436Q, p.S1463F). Individual mutations were introduced into an NF1-GRD expression vector and activated Ras was assayed by an enzyme-linked immunosorbent assay (ELISA). Ten NF1-GRD variants were deemed to be potentially pathogenic by virtue of significantly elevated levels of activated GTP-bound Ras in comparison to wild-type NF1 protein. The remaining five NF1-GRD variants were deemed less likely to be of pathological significance as they exhibited similar levels of activated Ras to the wild-type protein. These conclusions received broad support from both bioinformatic analysis and molecular modeling and serve to improve our understanding of NF1-GRD structure and function. © 2012 Wiley Periodicals, Inc.
Loss of neurofibromatosis type 1 (NF1) gene expression in pheochromocytomas from patients without NF1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Geist, R.T.; Gutmann, D.H.; Moley, J.F.

The neurofibromatosis type 1 (NF1) gene encodes a tumor suppressor protein, termed neurofibromin. Loss of NF1 gene expression has been reported in Schwann cell tumors (neurofibrosarcomas) from patients with NF1 as well as malignant and neuroblastomas from patients without NF1. Previously, we demonstrated the lack of neurofibromin expression in six pheochromocytomas from patients with NF1, suggesting that neurofibromin loss is associated with the progression to neoplasia in pheochromocytomas in these patients. The lack of NF1 gene expression in NF1 patient pheochromocytomas supports the notion that neurofibromin might be an essential regulator of cell growth in these cells. To determine whethermore » NF1 gene expression is similarly altered in pheochromocytomas from patients without NF1, twenty pheochromocytomas were examined for the presence of NF1 RNA by reverse-transcribed PCR (RT-PCR). Lack of NF1 gene expression was documented in four of these twenty tumors (20%) which corresponds to previously reported numbers for malignant melanomas and neuroblastomas in non-NF1 patients. Of these twenty pheochromocytomas, one of four sporadic tumors, one of ten tumors from patients with MEN2A, one of four tumors from patients with MEN2B, and one of two tumors from patients with von Hippel-Lindau syndrome demonstrated loss of NF1 gene expression. In all cases, the quality and quantity of tumor RNA was determined by RT-PCR amplification using primers which amplify cyclophilin RNA. We previously demonstrated that these tumors do not harbor activating mutations of the N-ras, K-ras or H-ras proto-oncogenes. These results suggest that loss of NF1 gene expression is frequently associated with the progression to neoplasia in tumors derived from adrenal medullary tissue in patients without clinical manifestations of neurofibromatosis and supports the notion that neurofibromin is a tumor suppressor gene product involved in the pathogenesis of a wide variety of tumor types.« less
Lactoferrin-derived resistance against plant pathogens in transgenic plants.

PubMed

Lakshman, Dilip K; Natarajan, Savithiry; Mandal, Sudhamoy; Mitra, Amitava

2013-12-04

Lactoferrin (LF) is a ubiquitous cationic iron-binding milk glycoprotein that contributes to nutrition and exerts a broad-spectrum primary defense against bacteria, fungi, protozoa, and viruses in mammals. These qualities make lactoferrin protein and its antimicrobial motifs highly desirable candidates to be incorporated in plants to impart broad-based resistance against plant pathogens or to economically produce them in bulk quantities for pharmaceutical and nutritional purposes. This study introduced bovine LF (BLF) gene into tobacco ( Nicotiana tabacum var. Xanthi), Arabidopsis ( A. thaliana ) and wheat ( Triticum aestivum ) via Agrobacterium -mediated plant transformation. Transgenic plants or detached leaves exhibited high levels of resistance against the damping-off causing fungal pathogen Rhizoctonia solani and the head blight causing fungal pathogen Fusarium graminearum . LF also imparted resistance to tomato plants against a bacterial pathogen, Ralstonia solanacearum . Similarly, other researchers demonstrated expression of LF and LF-mediated high-quality resistance to several other aggressive fungal and bacterial plant pathogens in transgenic plants and against viral pathogens by foliar applications of LF or its derivatives. Taken together, these studies demonstrated the effectiveness of LF for improving crop quality and its biopharming potentials for pharmaceautical and nutritional applications.
Vaccinium bracteatum Thunb. Exerts Anti-Inflammatory Activity by Inhibiting NF-κB Activation in BV-2 Microglial Cells

PubMed Central

Kwon, Seung-Hwan; Ma, Shi-Xun; Ko, Yong-Hyun; Seo, Jee-Yeon; Lee, Bo-Ram; Lee, Taek Hwan; Kim, Sun Yeou; Lee, Seok-Yong; Jang, Choon-Gon

2016-01-01

This study was designed to evaluate the pharmacological effects of Vaccinium bracteatum Thunb. methanol extract (VBME) on microglial activation and to identify the underlying mechanisms of action of these effects. The anti-inflammatory properties of VBME were studied using lipopolysaccharide (LPS)-stimulated BV-2 microglial cells. We measured the production of nitric oxide (NO), inducible NO synthase (iNOS), cyclooxygenase (COX)-2, prostaglandin E2 (PGE2), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6) as inflammatory parameters. We also examined the effect of VBME on intracellular reactive oxygen species (ROS) production and the activity of nuclear factor-kappa B p65 (NF-κB p65). VBME significantly inhibited LPS-induced production of NO and PGE2 and LPS-mediated upregulation of iNOS and COX-2 expression in a dose-dependent manner; importantly, VBME was not cytotoxic. VBME also significantly reduced the generation of the pro-inflammatory cytokines TNF-α, IL-1β, and IL-6. In addition, VBME significantly dampened intracellular ROS production and suppressed NF-κB p65 translocation by blocking IκB-α phosphorylation and degradation in LPS-stimulated BV2 cells. Our findings indicate that VBME inhibits the production of inflammatory mediators in BV-2 microglial cells by suppressing NF-κB signaling. Thus, VBME may be useful in the treatment of neurodegenerative diseases due to its ability to inhibit inflammatory mediator production in activated BV-2 microglial cells. PMID:27169820
Vaccinium bracteatum Thunb. Exerts Anti-Inflammatory Activity by Inhibiting NF-κB Activation in BV-2 Microglial Cells.

PubMed

Kwon, Seung-Hwan; Ma, Shi-Xun; Ko, Yong-Hyun; Seo, Jee-Yeon; Lee, Bo-Ram; Lee, Taek Hwan; Kim, Sun Yeou; Lee, Seok-Yong; Jang, Choon-Gon

2016-09-01

This study was designed to evaluate the pharmacological effects of Vaccinium bracteatum Thunb. methanol extract (VBME) on microglial activation and to identify the underlying mechanisms of action of these effects. The anti-inflammatory properties of VBME were studied using lipopolysaccharide (LPS)-stimulated BV-2 microglial cells. We measured the production of nitric oxide (NO), inducible NO synthase (iNOS), cyclooxygenase (COX)-2, prostaglandin E₂ (PGE₂), tumor necrosis factor-alpha (TNF-α), interleukin-1 beta (IL-1β), and interleukin-6 (IL-6) as inflammatory parameters. We also examined the effect of VBME on intracellular reactive oxygen species (ROS) production and the activity of nuclear factor-kappa B p65 (NF-κB p65). VBME significantly inhibited LPS-induced production of NO and PGE2 and LPS-mediated upregulation of iNOS and COX-2 expression in a dose-dependent manner; importantly, VBME was not cytotoxic. VBME also significantly reduced the generation of the pro-inflammatory cytokines TNF-α, IL-1β, and IL-6. In addition, VBME significantly dampened intracellular ROS production and suppressed NF-κB p65 translocation by blocking IκB-α phosphorylation and degradation in LPS-stimulated BV2 cells. Our findings indicate that VBME inhibits the production of inflammatory mediators in BV-2 microglial cells by suppressing NF-κB signaling. Thus, VBME may be useful in the treatment of neurodegenerative diseases due to its ability to inhibit inflammatory mediator production in activated BV-2 microglial cells.
Acetylshikonin Inhibits Human Pancreatic PANC-1 Cancer Cell Proliferation by Suppressing the NF-κB Activity.

PubMed

Cho, Seok-Cheol; Choi, Bu Young

2015-09-01

Acetylshikonin, a natural naphthoquinone derivative compound, has been used for treatment of inflammation and cancer. In the present study, we have investigated whether acetylshikonin could regulate the NF-κB signaling pathway, thereby leading to suppression of tumorigenesis. We observed that acetylshikonin significantly reduced proliferation of several cancer cell lines, including human pancreatic PANC-1 cancer cells. In addition, acetylshikonin inhibited phorbol 12-myristate 13-acetate (PMA) or tumor necrosis-α (TNF-α)-induced NF-κB reporter activity. Proteome cytokine array and real-time RT-PCR results illustrated that acetylshikonin inhibition of PMA-induced production of cytokines was mediated at the transcriptional level and it was associated with suppression of NF-κB activity and matrix metalloprotenases. Finally, we observed that an exposure of acetylshikonin significantly inhibited the anchorage-independent growth of PANC-1 cells. Together, our results indicate that acetylshikonin could serve as a promising therapeutic agent for future treatment of pancreatic cancer.
Annexin A6 interacts with p65 and stimulates NF-κB activity and catabolic events in articular chondrocytes.

PubMed

Campbell, Kirk A; Minashima, Takeshi; Zhang, Ying; Hadley, Scott; Lee, You Jin; Giovinazzo, Joseph; Quirno, Martin; Kirsch, Thorsten

2013-12-01

ANXA6, the gene for annexin A6, is highly expressed in osteoarthritic (OA) articular chondrocytes but not in healthy articular chondrocytes. This study was undertaken to determine whether annexin A6 affects catabolic events in these cells. Articular chondrocytes were isolated from Anxa6-knockout mice, wild-type (WT) mice, and human articular cartilage in which ANXA6 was overexpressed. Cells were treated with interleukin-1β (IL-1β) or tumor necrosis factor α (TNFα), and expression of catabolic genes and activation of NF-κB were determined by real-time polymerase chain reaction and luciferase reporter assay. Anxa6(-/-) and WT mouse knee joints were injected with IL-1β or the medial collateral ligament was transected and partial resection of the medial meniscus was performed to determine the role of Anxa6 in IL-1β-mediated cartilage destruction and OA progression. The mechanism by which Anxa6 stimulates NF-κB activity was determined by coimmunoprecipitation and immunoblot analysis of nuclear and cytoplasmic fractions of IL-1β-treated Anxa6(-/-) and WT mouse chondrocytes for p65 and Anxa6. Loss of Anxa6 resulted in decreased NF-κB activation and catabolic marker messenger RNA (mRNA) levels in IL-1β- or TNFα-treated articular chondrocytes, whereas overexpression of ANXA6 resulted in increased NF-κB activity and catabolic marker mRNA levels. Annexin A6 interacted with p65, and loss of Anxa6 caused decreased nuclear translocation and retention of the active p50/p65 NF-κB complex. Cartilage destruction in Anxa6(-/-) mouse knee joints after IL-1β injection or partial medial meniscectomy was reduced as compared to that in WT mouse joints. Our data define a role of annexin A6 in the modulation of NF-κB activity and in the stimulation of catabolic events in articular chondrocytes. Copyright © 2013 by the American College of Rheumatology.
A new structural class of proteasome inhibitors that prevent NF-kappa B activation.

PubMed

Lum, R T; Kerwar, S S; Meyer, S M; Nelson, M G; Schow, S R; Shiffman, D; Wick, M M; Joly, A

1998-05-01

The multicatalytic proteinase or proteasome is a highly conserved cellular structure that is responsible for the ATP-dependent proteolysis of many proteins involved in important regulatory cellular processes. We have identified a novel class of inhibitors of the chymotrypsin-like proteolytic activity of the 20S proteasome that exhibit IC50 values ranging from 0.1 to 0.5 microgram/mL (0.1 to 1 microM). In cell proliferation assays, these compounds inhibit growth with an IC50 ranging from 5 to 10 micrograms/mL (10-20 microM). A representative member of this class of inhibitors was tested in other biological assays. CVT-634 (5-methoxy-1-indanone-3-acetyl-leu-D-leu-1-indanylamide) prevented lipopolysaccharide (LPS), tumor necrosis factor (TNF)-, and phorbol ester-induced activation of nuclear factor kappa B (NF-kappa B) in vitro by preventing signal-induced degradation of I kappa B-alpha. In these studies, the I kappa B-alpha that accumulated was hyperphosphorylated, indicating that CVT-634 did not inhibit I kappa B-alpha kinase, the enzyme responsible for signal-induced phosphorylation of I kappa B-alpha. In vivo studies indicated that CVT-634 prevented LPS-induced TNF synthesis in a murine macrophage cell line. In addition, in mice pretreated with CVT-634 at 25 and 50 mg/kg and subsequently treated with LPS, serum TNF levels were significantly lower (225 +/- 59 and 83 +/- 41 pg/mL, respectively) than in those mice that were treated only with LPS (865 +/- 282 pg/mL). These studies suggest that specific inhibition of the chymotrypsin-like activity of the proteasome is sufficient to prevent signal-induced NF-kappa B activation and that the proteasome is a novel target for the identification of agents that may be useful in the treatment of diseases whose etiology is dependent upon the activation of NF-kappa B.
IKK{epsilon} modulates RSV-induced NF-{kappa}B-dependent gene transcription

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bao Xiaoyong; Indukuri, Hemalatha; Liu Tianshuang

2010-12-20

Respiratory syncytial virus (RSV), a negative-strand RNA virus, is the most common cause of epidemic respiratory disease in infants and young children. RSV infection of airway epithelial cells induces the expression of immune/inflammatory genes through the activation of a subset of transcription factors, including Nuclear Factor-{kappa}B (NF-{kappa}B). In this study we have investigated the role of the non canonical I{kappa}B kinase (IKK){epsilon} in modulating RSV-induced NF-{kappa}B activation. Our results show that inhibition of IKK{epsilon} activation results in significant impairment of viral-induced NF-{kappa}B-dependent gene expression, through a reduction in NF-{kappa}B transcriptional activity, without changes in nuclear translocation or DNA-binding activity. Absencemore » of IKK{epsilon} results in a significant decrease of RSV-induced NF-{kappa}B phosphorylation on serine 536, a post-translational modification important for RSV-induced NF-{kappa}B-dependent gene expression, known to regulate NF-{kappa}B transcriptional activity without affecting nuclear translocation. This study identifies a novel mechanism by which IKK{epsilon} regulates viral-induced cellular signaling.« less
NleC, a type III secretion protease, compromises NF-κB activation by targeting p65/RelA.

PubMed

Yen, Hilo; Ooka, Tadasuke; Iguchi, Atsushi; Hayashi, Tetsuya; Sugimoto, Nakaba; Tobe, Toru

2010-12-16

The NF-κB signaling pathway is central to the innate and adaptive immune responses. Upon their detection of pathogen-associated molecular patterns, Toll-like receptors on the cell surface initiate signal transduction and activate the NF-κB pathway, leading to the production of a wide array of inflammatory cytokines, in attempt to eradicate the invaders. As a countermeasure, pathogens have evolved ways to subvert and manipulate this system to their advantage. Enteropathogenic and enterohemorrhagic Escherichia coli (EPEC and EHEC) are closely related bacteria responsible for major food-borne diseases worldwide. Via a needle-like protein complex called the type three secretion system (T3SS), these pathogens deliver virulence factors directly to host cells and modify cellular functions, including by suppressing the inflammatory response. Using gain- and loss-of-function screenings, we identified two bacterial effectors, NleC and NleE, that down-regulate the NF-κB signal upon being injected into a host cell via the T3SS. A recent report showed that NleE inhibits NF-κB activation, although an NleE-deficient pathogen was still immune-suppressive, indicating that other anti-inflammatory effectors are involved. In agreement, our present results showed that NleC was also required to inhibit inflammation. We found that NleC is a zinc protease that disrupts NF-κB activation by the direct cleavage of NF-κB's p65 subunit in the cytoplasm, thereby decreasing the available p65 and reducing the total nuclear entry of active p65. More importantly, we showed that a mutant EPEC/EHEC lacking both NleC and NleE (ΔnleC ΔnleE) caused greater inflammatory response than bacteria carrying ΔnleC or ΔnleE alone. This effect was similar to that of a T3SS-defective mutant. In conclusion, we found that NleC is an anti-inflammatory bacterial zinc protease, and that the cooperative function of NleE and NleC disrupts the NF-κB pathway and accounts for most of the immune suppression caused
Escherichia coli K1 inhibits proinflammatory cytokine induction in monocytes by preventing NF-kappaB activation.

PubMed

Selvaraj, Suresh K; Prasadarao, Nemani V

2005-08-01

Phagocytes are well-known effectors of the innate immune system to produce proinflammatory cytokines and chemokines such as tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-1beta, and IL-8 during infections. Here, we show that infection of monocytes with wild-type Escherichia coli K1, which causes meningitis in neonates, suppresses the production of cytokines and chemokines (TNF-alpha, regulated on activation, normal T expressed and secreted, macrophage-inflammatory protein-1beta, IL-1beta, and IL-8). In contrast, infection of monocytes with a mutant E. coli, which lacks outer membrane protein A (OmpA- E. coli) resulted in robust production of cytokines and chemokines. Wild-type E. coli K1 (OmpA+ E. coli) prevented the phosphorylation and its degradation of inhibitor of kappaB, thereby blocking the translocation of nuclear factor (NF)-kappaB to the nucleus. OmpA+ E. coli-infected cells, subsequently subjected to lipopolysaccharide challenge, were crippled severely in their ability to activate NF-kappaB to induce cytokine/chemokine production. Selective inhibitors of the extracellular signal-regulated kinase (ERK) 1/2 pathway and p38 mitogen-activated protein kinase (MAPK), but not Jun N-terminal kinase, significantly reduced the activation of NF-kappaB and the production of cytokines and chemokines induced by OmpA- E. coli, indicating a role for these kinases in the NF-kappaB/cytokine pathway. It is interesting that the phosphorylation of ERK 1/2 and p38 MAPK was notably reduced in monocytes infected with OmpA+ E. coli when compared with monocytes infected with OmpA- E. coli, suggesting that the modulation of upstream events common for NF-kappaB and MAPKs by the bacterium is possible. The ability of OmpA+ E. coli K1 to inhibit the macrophage response temporarily may enable bacterial survival and growth within the host for the onset of meningitis by E. coli K1.
Functional Analysis of Human NF1 in Drosophila

DTIC Science & Technology

2008-12-01

also have learning problem. Such learning phenotypes have been recapitulated in animal models, including in mouse and Drosophila mutants. This proposal...by examining the phenotypes of mutated human genes expressed in Drosophila NF1 null mutants. We also propose that Gsα/NF1 activated AC pathway...in both Drosophila and mouse NF1 models. Our previous work has shown that defective cAMP signaling leads to the learning phenotype in Drosophila Nf1
Molecular basis for the unique deubiquitinating activity of the NF-kappaB inhibitor A20.

PubMed

Lin, Su-Chang; Chung, Jee Y; Lamothe, Betty; Rajashankar, Kanagalaghatta; Lu, Miao; Lo, Yu-Chih; Lam, Amy Y; Darnay, Bryant G; Wu, Hao

2008-02-15

Nuclear factor kappaB (NF-kappaB) activation in tumor necrosis factor, interleukin-1, and Toll-like receptor pathways requires Lys63-linked nondegradative polyubiquitination. A20 is a specific feedback inhibitor of NF-kappaB activation in these pathways that possesses dual ubiquitin-editing functions. While the N-terminal domain of A20 is a deubiquitinating enzyme (DUB) for Lys63-linked polyubiquitinated signaling mediators such as TRAF6 and RIP, its C-terminal domain is a ubiquitin ligase (E3) for Lys48-linked degradative polyubiquitination of the same substrates. To elucidate the molecular basis for the DUB activity of A20, we determined its crystal structure and performed a series of biochemical and cell biological studies. The structure reveals the potential catalytic mechanism of A20, which may be significantly different from papain-like cysteine proteases. Ubiquitin can be docked onto a conserved A20 surface; this interaction exhibits charge complementarity and no steric clash. Surprisingly, A20 does not have specificity for Lys63-linked polyubiquitin chains. Instead, it effectively removes Lys63-linked polyubiquitin chains from TRAF6 without dissembling the chains themselves. Our studies suggest that A20 does not act as a general DUB but has the specificity for particular polyubiquitinated substrates to assure its fidelity in regulating NF-kappaB activation in the tumor necrosis factor, interleukin-1, and Toll-like receptor pathways.
Advanced Glycation End Products Enhance Macrophages Polarization into M1 Phenotype through Activating RAGE/NF-κB Pathway

PubMed Central

Jin, Xian; Yao, Tongqing; Zhou, Zhong'e; Zhu, Jian; Zhang, Song; Hu, Wei; Shen, Chengxing

2015-01-01

Atherosclerotic lesions are accelerated in patients with diabetes. M1 (classically activated in contrast to M2 alternatively activated) macrophages play key roles in the progression of atherosclerosis. Since advanced glycation end products (AGEs) are major pathogenic factors and active inflammation inducers in diabetes mellitus, this study assessed the effects of AGEs on macrophage polarization. The present study showed that AGEs significantly promoted macrophages to express IL-6 and TNF-α. M1 macrophage markers such as iNOS and surface markers including CD11c and CD86 were significantly upregulated while M2 macrophage markers such as Arg1 and CD206 remained unchanged after AGEs stimulation. AGEs significantly increased RAGE expression in macrophages and activated NF-κB pathway, and the aforementioned effects were partly abolished by administration of anti-RAGE antibody or NF-κB inhibitor PDTC. In conclusion, our results suggest that AGEs enhance macrophage differentiation into proinflammatory M1 phenotype at least partly via RAGE/NF-κB pathway activation. PMID:26114112
Mechanisms of NF-κB deregulation in lymphoid malignancies.

PubMed

Krappmann, Daniel; Vincendeau, Michelle

2016-08-01

Deregulations promoting constitutive activation of canonical and non-canonical NF-κB signaling are a common feature of many lymphoid malignancies. Due to their cellular origin and the pivotal role of NF-κB for the normal function of B lymphocytes, B-cell malignancies are particularly prone to genetic aberrations that affect the pathway. Key positive regulators of NF-κB signaling can act as oncogenes that are often prone to chromosomal translocation, amplifications or activating mutations. Negative regulators of NF-κB have tumor suppressor functions and are frequently inactivated either by genomic deletions or point mutations. Whereas some aberrations are found in a variety of different lymphoid malignancies, some oncogenic alterations are very restricted to distinct lymphoma subsets, reflecting the clonal and cellular origin of specific lymphoma entities. NF-κB activation in many lymphoma cells is also driven by the microenvironment or chronic signaling that does not rely on genetic alterations. A number of drugs that target the NF-κB pathway are in preclinical or clinical development, revealing that there will be new options for therapies in the future. Since each lymphoma entity utilizes distinct mechanisms to activate NF-κB, a major challenge is to elucidate the exact pathological processes in order to faithfully predict clinical responses to the different therapeutic approaches. Copyright © 2016 Elsevier Ltd. All rights reserved.
Lactoferrin and oral diseases: current status and perspective in periodontitis

PubMed Central

Berlutti, Francesca; Pilloni, Andrea; Pietropaoli, Miriam; Polimeni, Antonella; Valenti, Piera

2012-01-01

Summary Lactoferrin (Lf), an iron-binding glycoprotein able to chelate two ferric ions per molecule, is a component of human secretions synthesized by exocrine glands and neutrophils in infection/inflammation sites. Lactoferrin in saliva represents an important defence factor against bacterial injuries including those related to Streptococcus mutans and periodontopathic bacteria through its ability to decrease bacterial growth, biofilm development, iron overload, reactive oxygen formation and inflammatory processes. A growing body of research suggests that inflammatory periodontal disease involves a failure of resolution pathways to restore tissue homeostasis. There is an important distinction between anti-inflammation and resolution; anti-inflammation is pharmacologic intervention in inflammatory pathways, whereas resolution involves biologic pathways restoring inflammatory homeostasis. An appropriate regulation of pro-inflammatory cytokine synthesis might be useful in reducing periodontal tissue destruction. Recently, the multi-functional IL-6 is emerging as an important factor able to modulate bone, iron and inflammatory homeostasis. Here, we report an overview of Lf functions as well as for the first time Lf anti-inflammatory ability against periodontitis in in vitro model and observational clinical study. In in vitro model, represented by gingival fibroblasts infected with Prevotella intermedia, Lf exerted a potent anti-inflammatory activity. In the observational clinical trial performed through bovine Lf (bLf) topically administered to volunteers suffering from periodontitis, bLf decreased cytokines, including IL-6 in crevicular fluid, edema, bleeding, pocket depth, gingival and plaque index, thus improving clinical attachment levels. Even if other clinical trials are required, these results provide strong evidence for a instead of an therapeutic potential of this multifunctional natural protein. PMID:22545184

TNF-{alpha} promotes cell survival through stimulation of K{sup +} channel and NF{kappa}B activity in corneal epithelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang Ling; Reinach, Peter; Lu, Luo

2005-11-15

Tumor necrosis factor (TNF-{alpha}) in various cell types induces either cell death or mitogenesis through different signaling pathways. In the present study, we determined in human corneal epithelial cells how TNF-{alpha} also promotes cell survival. Human corneal epithelial (HCE) cells were cultured in DMEM/F-12 medium containing 10% FBS. TNF-{alpha} stimulation induced activation of a voltage-gated K{sup +} channel detected by measuring single channel activity using patch clamp techniques. The effect of TNF-{alpha} on downstream events included NF{kappa}B nuclear translocation and increases in DNA binding activities, but did not elicit ERK, JNK, or p38 limb signaling activation. TNF-{alpha} induced increases inmore » p21 expression resulting in partial cell cycle attenuation in the G{sub 1} phase. Cell cycle progression was also mapped by flow cytometer analysis. Blockade of TNF-{alpha}-induced K{sup +} channel activity effectively prevented NF{kappa}B nuclear translocation and binding to DNA, diminishing the cell-survival protective effect of TNF-{alpha}. In conclusion, TNF-{alpha} promotes survival of HCE cells through sequential stimulation of K{sup +} channel and NF{kappa}B activities. This response to TNF-{alpha} is dependent on stimulating K{sup +} channel activity because following suppression of K{sup +} channel activity TNF-{alpha} failed to activate NF{kappa}B nuclear translocation and binding to nuclear DNA.« less
Lymphotoxin activation by human T-cell leukemia virus type I-infected cell lines: role for NF-kappa B.

PubMed Central

Paul, N L; Lenardo, M J; Novak, K D; Sarr, T; Tang, W L; Ruddle, N H

1990-01-01

Human T-cell leukemia virus type I (HTLV-I)-infected T-cell lines constitutively produce high levels of biologically active lymphotoxin (LT; tumor necrosis factor-beta) protein and LT mRNA. To understand the regulation of LT transcription by HTLV-I, we analyzed the ability of a series of deletions of the LT promoter to drive the chloramphenicol acetyltransferase (CAT) reporter gene in HTLV-I-positive MT-2 cells. The smallest LT promoter fragment (-140 to +77) that was able to drive CAT activity contained a site that was similar to the immunoglobulin kappa-chain NF-kappa B-binding site. Since the HTLV-I tax gene activates the nuclear form of NF-kappa B, this finding suggested a possible means of HTLV-I activation of LT production. We found that the LT kappa B-like site specifically formed a complex with NF-kappa B-containing nuclear extract from MT-2, C81-66-45, and other activated T cells. Mutation of the LT kappa B site in the context of the LT promoter (-293 to +77) (mutant M1) reduced the ability of the promoter to drive the CAT gene in HTLV-I-infected and noninfected human T-cell lines. These data suggest a general role for NF-kappa B activation in the induction of LT gene transcription. Activation of LT in HTLV-I-infected cells may explain the pathology associated with HTLV-I infection, including the hypercalcemia that is prevalent in adult T-cell leukemia. Images PMID:1976820
Curcumin attenuates quinocetone induced apoptosis and inflammation via the opposite modulation of Nrf2/HO-1 and NF-kB pathway in human hepatocyte L02 cells.

PubMed

Dai, Chongshan; Li, Bin; Zhou, Yan; Li, Daowen; Zhang, Shen; Li, Hui; Xiao, Xilong; Tang, Shusheng

2016-09-01

The potential toxicity of quinocetone (QCT) has raised widely concern, but its mechanism is still unclear. This study aimed to investigate the protective effect of curcumin on QCT induced apoptosis and the underlying mechanism in human hepatocyte L02 cells. The results showed that QCT treatment significantly decreased the cell viability of L02 cell and increased the release of lactate dehydrogenase (LDH), which was attenuated by curcumin pre-treatment at 1.25, 2.5 and 5 μM. Compared to the QCT alone group, curcumin pre-treatment significantly attenuated QCT induced oxidative stress, mitochondrial dysfunction and apoptosis. In addition, curcumin pretreatment markedly attenuated QCT-induced increase of iNOS activity and NO production in a dose-dependent manner. Meanwhile, curcumin pretreatment markedly down-regulated the expression of nuclear factor -kB (NF-kB) and iNOS mRNAs, but up-regulated the expressions of Nrf2 and HO-1 mRNAs, compared to the QCT alone group. Zinc protoporphyrin IX, a HO-1 inhibitor, markedly partly abolished the cytoprotective effect of curcumin against QCT-induced caspase activation, NF-kB mRNA expression. These results indicate that curcumin could effectively inhibit QCT induced apoptosis and inflammatory response in L02 cells, which may involve the activation of Nrf2/HO-1 and inhibition of NF-kB pathway. Copyright © 2016 Elsevier Ltd. All rights reserved.
Suppressive effects of Lithospermum erythrorhizon extracts on lipopolysaccharide-induced activation of AP-1 and NF-kappaB via mitogen-activated protein kinase pathways in mouse macrophage cells.

PubMed

Han, Kyu Yeon; Kwon, Taek Hwan; Lee, Tae Hoon; Lee, Sung-Joon; Kim, Sung-Hoon; Kim, Jiyoung

2008-04-30

A variety of anti-inflammatory agents have been shown to exert chemopreventive activity via targeting of transcription factors such as NF-kappaB and AP-1. Lithospermum erythrorhizon (LE) has long been used in traditional oriental medicine. In this study, we demonstrated the inhibitory effects of LE extracts on lipopolysaccharide (LPS)-stimulated production of inflammatory cytokines. As an underlying mechanism of inhibition, LE extracts reduced LPS-induced transactivation of AP-1 as well as NF-kappaB in mouse macrophage cells. Electrophoretic mobility shift assays indicated that LE extracts inhibited the DNA binding activities of AP-1 and NF-kappaB. In addition, phosphorylation of IkappaB-alpha protein was suppressed by LE extracts. Moreover, LE extracts inhibited c-Jun N-terminal kinase and extracellular signal-regulated signaling pathways. Our results suggest that the anti-inflammatory activity of LE extracts may be mediated by the inhibition of signal transduction pathways that normally lead to the activation of AP-1and NF-kappaB. These inhibitory effects may be useful for chemoprevention of cancer or other chronic inflammatory diseases.
The E3 ubiquitin ligase mind bomb-2 (MIB2) protein controls B-cell CLL/lymphoma 10 (BCL10)-dependent NF-κB activation.

PubMed

Stempin, Cinthia C; Chi, Liying; Giraldo-Vela, Juan P; High, Anthony A; Häcker, Hans; Redecke, Vanessa

2011-10-28

B-cell CLL/lymphoma 10 (BCL10) is crucial for the activation of NF-κB in numerous immune receptor signaling pathways, including the T-cell receptor (TCR) and B-cell receptor signaling pathways. However, the molecular mechanisms that lead to signal transduction from BCL10 to downstream NF-κB effector kinases, such as TAK1 and components of the IKK complex, are not entirely understood. Here we used a proteomic approach and identified the E3 ligase MIB2 as a novel component of the activated BCL10 complex. In vitro translation and pulldown assays suggest direct interaction between BCL10 and MIB2. Overexpression experiments show that MIB2 controls BCL10-mediated activation of NF-κB by promoting autoubiquitination and ubiquitination of IKKγ/NEMO, as well as recruitment and activation of TAK1. Knockdown of MIB2 inhibited BCL10-dependent NF-κB activation. Together, our results identify MIB2 as a novel component of the activated BCL10 signaling complex and a missing link in the BCL10-dependent NF-κB signaling pathway.
Resveratrol promotes recovery of immune function of immunosuppressive mice by activating JNK/NF-κB pathway in splenic lymphocytes.

PubMed

Lai, Xin; Cao, Mei; Song, Xu; Jia, Renyong; Zou, Yuanfeng; Li, Lixia; Liang, Xiaoxia; He, Changliang; Yin, Lizi; Yue, Guizhou; Ye, Gang; Yin, Zhongqiong

2017-06-01

Resveratrol, a natural compound found in over 70 plants, is known to possess immunoregulatory effects and anti-inflammatory activity. It has been shown that resveratrol has regulatory effects on different signaling pathways in different diseases. However, few reports have evaluated the effects of resveratrol on reinforcing immunity recovery via activating nuclear factor-κB (NF-κB) pathway and Jun N-terminal kinases (JNK) pathway. The present study aimed to assess immune-enhancing activity and underlying mechanism of resveratrol in immunosuppressive mice. Previously, we reported that resveratrol could promote mouse spleen lymphocyte functions to recover the immune system effectively. In the present study, we show that resveratrol could upregulate the expressions of NF-κB, IκB kinase, JNK, and c-jun in splenic lymphocytes of immunosuppressive mice. Taken together, our results indicate that resveratrol could promote recovery of immunologic function in immunosuppressive mice by activating JNK/NF-κB pathway.
Geraniol attenuates 4NQO-induced tongue carcinogenesis through downregulating the activation of NF-κB in rats.

PubMed

Madankumar, Arumugam; Tamilarasi, Sasivarnam; Premkumar, Thandavamoorthy; Gopikrishnan, Mani; Nagabhishek, Natesh; Devaki, Thiruvengadam

2017-10-01

Geraniol, an acyclic monoterpene found in lemon grass and aromatic herb oil, has been shown to exert antitumor and antioxidant activities against various cancer types. The objective of this study was to investigate the potential chemoprotective role of geraniol against 4-nitroquinoline-1-oxide (4NQO)-induced oral carcinogenesis in male Wistar rats and furthermore to study anti-inflammatory mechanisms of action through possible NF-κB signaling. 4NQO was administered to rats at the dose of 50 ppm through drinking water to induce tongue cancer in 20 weeks. 4NQO provoked inflammation by upregulating the expressions of the p65 subunit nuclear factor kappa-β (NF-κB) in the nucleus, tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS). Additionally, staining for immature and mature mast cells in cancer niche by toluidine blue staining and alcian blue-safranin staining showed more accumulation. Co-treatment of geraniol 200 mg/kg b.w. showed a significant decrease in the level of p65 NF-κB in the nucleus, and this might be due to the inhibition of NF-κB activation/translocation into nucleus, which was further confirmed by decreased immature and mature mast cell density and the expression of inflammatory downstream mediators such as TNF-α, IL-1β, COX-2, and iNOS. Collectively, our results suggested that geraniol as a potential anti-inflammatory agent having the capability to obstruct 4NQO initiated NF-κB activation and modulated the expression of inflammatory mediators.
Pyropheophorbide-a methyl ester-mediated photosensitization activates transcription factor NF-kappaB through the interleukin-1 receptor-dependent signaling pathway.

PubMed

Matroule, J Y; Bonizzi, G; Morlière, P; Paillous, N; Santus, R; Bours, V; Piette, J

1999-01-29

Pyropheophorbide-a methyl ester (PPME) is a second generation of photosensitizers used in photodynamic therapy. We demonstrated that PPME photosensitization activated NF-kappaB transcription factor in colon cancer cells. Unexpectedly, this activation occurred in two separate waves, i.e. a rapid and transient one and a second slower but sustained phase. The former was due to photosensitization by PPME localized in the cytoplasmic membrane which triggered interleukin-1 receptor internalization and the transduction pathways controlled by the interleukin-1 type I receptor. Indeed, TRAF6 dominant negative mutant abolished NF-kappaB activation by PPME photosensitization, and TRAF2 dominant negative mutant was without any effect, and overexpression of IkappaB kinases increased gene transcription controlled by NF-kappaB. Oxidative stress was not likely involved in the activation. On the other hand, the slower and sustained wave could be the product of the release of ceramide through activation of the acidic sphingomyelinase. PPME localization within the lysosomal membrane could explain why ceramide acted as second messenger in NF-kappaB activation by PPME photosensitization. These data will allow a better understanding of the molecular basis of tumor eradication by photodynamic therapy, in particular the importance of the host cell response in the treatment.
Early diagnosis of mild cognitive impairment and Alzheimer's disease based on salivary lactoferrin.

PubMed

Carro, Eva; Bartolomé, Fernando; Bermejo-Pareja, Félix; Villarejo-Galende, Alberto; Molina, José Antonio; Ortiz, Pablo; Calero, Miguel; Rabano, Alberto; Cantero, José Luis; Orive, Gorka

2017-01-01

The Alzheimer's disease (AD) process is likely initiated many years before clinical onset. Biomarkers of preclinical disease are critical for the development of disease-modifying or even preventative therapies. Current biomarkers for early disease, including cerebrospinal fluid tau and amyloid β (Aβ) levels, structural and functional magnetic resonance imaging, and the use of brain amyloid imaging, are limited because they are very invasive or expensive. Noninvasive biomarkers may be a more accessible alternative, but none can currently detect preclinical AD with the required sensitivity and specificity. Here, we show a novel, straight-forward, and noninvasive approach for assessment of early stages of cognitive decline. Salivary samples from cases of amnestic mild cognitive impairment (aMCI) and AD, and neurology controls were analyzed. We have discovered and validated a new single saliva biomarker, lactoferrin, which in our cross-sectional investigation perfectly discriminates clinically diagnosed aMCI and AD patients from a cognitively healthy control group. The accuracy for AD diagnosis shown by salivary lactoferrin was greater than that obtained from core cerebrospinal fluid (CSF) biomarkers, including total tau and CSF Aβ 42 . Furthermore, salivary lactoferrin can be used for population screening and for identifying those underdiagnosed subjects with very early stages of mild cognitive impairment and AD. This biomarker may offer new insights in the early diagnostics for AD.
Time-dependent modulation of thioredoxin reductase activity might contribute to sulforaphane-mediated inhibition of NF-kappaB binding to DNA.

PubMed

Heiss, Elke; Gerhäuser, Clarissa

2005-01-01

The chemopreventive agent sulforaphane (SFN) exerts anti-inflammatory activity by thiol-dependent inhibition of nuclear factor kappaB (NF-kappaB) DNA binding. To further analyze the underlying mechanisms, we focused on the thioredoxin/thioredoxin reductase (TrxR) system as a key redox mechanism regulating NF-kappaB DNA binding. Using cultured Raw 264.7 mouse macrophages as a model, 1-chloro-2,4-dinitrobenzene (CDNB), a known inhibitor of TrxR, was identified as an inhibitor of lipopolysaccharide (LPS)-mediated nitric oxide (NO) production and of NF-kappaB DNA binding. CDNB and SFN acted synergistically with respect to inhibition of LPS-induced NO release, and we consequently identified SFN as a novel inhibitor of TrxR enzymatic activity in vitro. Short-term treatment of Raw macrophages with SFN or CDNB resulted in the inhibition of TrxR activity in vivo with half-maximal inhibitory concentration of 25.0 +/- 3.5 microM and 9.4 +/- 3.7 microM, respectively, whereas after a 24-h treatment with 25 microM SFN, TrxR activity was >1.5-fold elevated. In additional experiments, we could exclude that inhibition of trans-activating activity of NF-kappaB contributed to the reduced expression of pro-inflammatory proteins by SFN, based on transient transfection experiments with a (kappaB)(2)- chloramphenicol acetyltransferase construct and a lack of inhibition of protein kinase A activity. These findings further emphasize the importance of redox modulation or thiol reactivity for the regulation of NF-kappaB-dependent transcription by SFN. Antioxid. Redox Signal. 7, 1601-1611. Antioxid. Redox Signal. 7, 1601-1611.
CREB, NF-Y and MEIS1 conserved binding sites are essential to balance Myostatin promoter/enhancer activity during early myogenesis.

PubMed

Grade, Carla Vermeulen Carvalho; Mantovani, Carolina Stefano; Fontoura, Marina Alves; Yusuf, Faisal; Brand-Saberi, Beate; Alvares, Lúcia Elvira

2017-10-01

Myostatin (MSTN) is a strong inhibitor of skeletal muscle growth in human and other vertebrates. Its transcription is controlled by a proximal promoter/enhancer (Mstn P/E) containing a TATA box besides CREB, NF-Y, MEIS1 and FXR transcription factor binding sites (TFBSs), which are conserved throughout evolution. The aim of this work was to investigate the role of these TFBSs on Mstn P/E activity and evaluate the potential of their putative ligands as Mstn trans regulators. Mstn P/E mutant constructs were used to establish the role of conserved TFBSs using dual-luciferase assays. Expression analyses were performed by RT-PCR and in situ hybridization in C2C12 myoblasts and E10.5 mouse embryos, respectively. Our results revealed that CREB, NF-Y and MEIS1 sites are required to balance Mstn P/E activity, keeping Mstn transcription within basal levels during myoblast proliferation. Furthermore, our data showed that NF-Y site is essential, although not sufficient, to mediate Mstn P/E transcriptional activity. In turn, CREB and MEIS1 binding sites seem to depend on the presence of NF-Y site to induce Mstn P/E. FXR appears not to confer any effect on Mstn P/E activity, except in the absence of all other conserved TFBS. Accordingly, expression studies pointed to CREB, NF-Y and MEIS1 but not to FXR factors as possible regulators of Mstn transcription in the myogenic context. Altogether, our findings indicated that CREB, NF-Y and MEIS1 conserved sites are essential to control basal Mstn transcription during early myogenesis, possibly by interacting with these or other related factors.
Styrene induces an inflammatory response in human lung epithelial cells via oxidative stress and NF-{kappa}B activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Roeder-Stolinski, Carmen; Fischaeder, Gundula; Oostingh, Gertie Janneke

2008-09-01

Styrene is a volatile organic compound (VOC) that is widely used as a solvent in many industrial settings. Chronic exposure to styrene can result in irritation of the mucosa of the upper respiratory tract. Contact of styrene with epithelial cells stimulates the expression of a variety of inflammatory mediators, including the chemotactic cytokine monocyte chemoattractant protein-1 (MCP-1). To characterise the underlying mechanisms of the induction of inflammatory signals by styrene, we investigated the influence of this compound on the induction of oxidative stress and the activation of the nuclear factor-kappa B (NF-{kappa}B) signalling pathway in human lung epithelial cells (A549).more » The results demonstrate that styrene-induced MCP-1 expression, as well as the expression of the oxidative stress marker glutathione S-transferase (GST), is associated with a concentration dependent pattern of NF-{kappa}B activity. An inhibitor of NF-{kappa}B, IKK-NBD, and the anti-inflammatory antioxidant N-acetylcysteine (NAC) were both effective in suppressing styrene-induced MCP-1 secretion. In addition, NAC was capable of inhibiting the upregulation of GST expression. Our findings suggest that the activation of the NF-{kappa}B signalling pathway by styrene is mediated via a redox-sensitive mechanism.« less
Nuclear IL-33 is a transcriptional regulator of NF-{kappa}B p65 and induces endothelial cell activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Choi, Yeon-Sook; Park, Jeong Ae; Kim, Jihye

2012-05-04

Highlights: Black-Right-Pointing-Pointer IL-33 as nuclear factor regulated expression of ICAM-1 and VCAM-1. Black-Right-Pointing-Pointer Nuclear IL-33 increased the transcription of NF-{kappa}B p65 by binding to the p65 promoter. Black-Right-Pointing-Pointer Nuclear IL-33 controls NF-{kappa}B-dependent inflammatory responses. -- Abstract: Interleukin (IL)-33, an IL-1 family member, acts as an extracellular cytokine by binding its cognate receptor, ST2. IL-33 is also a chromatin-binding transcriptional regulator highly expressed in the nuclei of endothelial cells. However, the function of IL-33 as a nuclear factor is poorly defined. Here, we show that IL-33 is a novel transcriptional regulator of the p65 subunit of the NF-{kappa}B complex and ismore » involved in endothelial cell activation. Quantitative reverse transcriptase PCR and Western blot analyses indicated that IL-33 mediates the expression of intercellular adhesion molecule (ICAM)-1 and vascular cell adhesion molecule (VCAM)-1 in endothelial cells basally and in response to tumor necrosis factor-{alpha}-treatment. IL-33-induced ICAM-1/VCAM-1 expression was dependent on the regulatory effect of IL-33 on the nuclear factor (NF)-{kappa}B pathway; NF-{kappa}B p65 expression was enhanced by IL-33 overexpression and, conversely, reduced by IL-33 knockdown. Moreover, NF-{kappa}B p65 promoter activity and chromatin immunoprecipitation analysis revealed that IL-33 binds to the p65 promoter region in the nucleus. Our data provide the first evidence that IL-33 in the nucleus of endothelial cells participates in inflammatory reactions as a transcriptional regulator of NF-{kappa}B p65.« less
NLRP12 Suppresses Colon Inflammation and Tumorigenesis through the Negative Regulation of Non-canonical NF-κB Signaling and MAP Kinase Activation

PubMed Central

Allen, Irving C.; Wilson, Justin E.; Schneider, Monika; Lich, John D.; Roberts, Reid A.; Arthur, Janelle C.; Woodford, Rita-Marie T.; Davis, Beckley K.; Uronis, Joshua M.; Herfarth, Hans H.; Jobin, Christian; Rogers, Arlin B.; Ting, Jenny P.-Y.

2013-01-01

SUMMARY In vitro data suggest that a subgroup of NLR proteins, including NLRP12, inhibits the transcription factor NF-κB, although physiologic and disease-relevant evidence is largely missing. Dysregulated NF-κB activity is associated with colonic inflammation and cancer, and we found Nlrp12-/- mice were highly susceptible to colitis and colitis-associated colon cancer. Polyps isolated from Nlrp12-/- mice showed elevated non-canonical NF-κB activation and increased expression of target genes that were associated with cancer, including Cxcl13 and Cxcl12. NLRP12 negatively regulated ERK and AKT signaling pathways in affected tumor tissues. Both hematopoietic and nonhematopoietic-derived NLRP12 contributed to inflammation, but the latter dominantly contributed to tumorigenesis. The non-canonical NF-κB pathway was regulated upon degradation of TRAF3 and activation of NIK. NLRP12 interacted with both NIK and TRAF3, and Nlrp12-/- cells have constitutively elevated NIK, p100 processing to p52 and reduced TRAF3. Thus, NLRP12 is a checkpoint of noncanonical NF-κB, inflammation and tumorigenesis. PMID:22503542
OC-2-KB: A software pipeline to build an evidence-based obesity and cancer knowledge base.

PubMed

Lossio-Ventura, Juan Antonio; Hogan, William; Modave, François; Guo, Yi; He, Zhe; Hicks, Amanda; Bian, Jiang

2017-11-01

Obesity has been linked to several types of cancer. Access to adequate health information activates people's participation in managing their own health, which ultimately improves their health outcomes. Nevertheless, the existing online information about the relationship between obesity and cancer is heterogeneous and poorly organized. A formal knowledge representation can help better organize and deliver quality health information. Currently, there are several efforts in the biomedical domain to convert unstructured data to structured data and store them in Semantic Web knowledge bases (KB). In this demo paper, we present, OC-2-KB (Obesity and Cancer to Knowledge Base), a system that is tailored to guide the automatic KB construction for managing obesity and cancer knowledge from free-text scientific literature (i.e., PubMed abstracts) in a systematic way. OC-2-KB has two important modules which perform the acquisition of entities and the extraction then classification of relationships among these entities. We tested the OC-2-KB system on a data set with 23 manually annotated obesity and cancer PubMed abstracts and created a preliminary KB with 765 triples. We conducted a preliminary evaluation on this sample of triples and reported our evaluation results.
Dexamethasone potently enhances phorbol ester-induced IL-1beta gene expression and nuclear factor NF-kappaB activation.

PubMed

Wang, Y; Zhang, J J; Dai, W; Lei, K Y; Pike, J W

1997-07-15

The synthetic glucocorticoid dexamethasone, an immunosuppressive and anti-inflammatory agent, was investigated for its effect on PMA-mediated expression of the inflammatory cytokine IL-1beta in the human monocytic leukemic cell line THP-1. PMA alone induced the production of low levels of IL-1beta in THP-1 cells, whereas dexamethasone alone had no effect. However, dexamethasone potently enhanced PMA-mediated IL-1beta production. Using a selective and potent inhibitor of protein kinase C, we found that synergistic interaction between PMA and dexamethasone requires protein kinase C activation. PMA has been known to activate nuclear factor NF-kappaB in THP-1 cells. Using an oligonucleotide probe corresponding to an NF-kappaB DNA-binding motif of the IL-1beta gene promoter in gel electrophoresis mobility shift assays, we demonstrated that PMA-induced NF-kappaB activation was greatly potentiated by dexamethasone. Our results indicate that glucocorticoids can be positive regulators of inflammatory cytokine gene expression during monocytic cell differentiation.
KrasG12D-Induced IKK2/β/NF-κB Activation by IL-1α and p62 Feedforward Loops Is Required for Development of Pancreatic Ductal Adenocarcinoma

PubMed Central

Ling, Jianhua; Kang, Ya’an; Zhao, Ruiying; Xia, Qianghua; Lee, Dung-Fang; Chang, Zhe; Li, Jin; Peng, Bailu; Fleming, Jason B.; Wang, Huamin; Liu, Jinsong; Lemischka, Ihor R.; Hung, Mien-Chie; Chiao, Paul J.

2012-01-01

SUMMARY Constitutive Kras and NF-κB activation is identified as signature alterations in pancreatic ductal adenocarcinoma (PDAC). However, how NF-κB is activated in PDAC is not yet understood. Here, we report that pancreas-targeted IKK2/β inactivation inhibited NF-κB activation and PDAC development in KrasG12D and KrasG12D;Ink4a/ArfF/F mice, demonstrating a mechanistic link between IKK2/β and KrasG12D in PDAC inception. Our findings reveal that KrasG12D-activated AP-1 induces IL-1α, which, in turn, activates NF-κB and its target genes IL-1α and p62, to initiate IL-1α/p62 feedforward loops for inducing and sustaining NF-κB activity. Furthermore, IL-1α overexpression correlates with Kras mutation, NF-κB activity, and poor survival in PDAC patients. Therefore, our findings demonstrate the mechanism by which IKK2/β/NF-κB is activated by KrasG12D through dual feedforward loops of IL-1α/p62. PMID:22264792
Effects of mycobactin J and lactoferrin supplementation of drinking water on the in vivo multiplication of Mycobacterium paratuberculosis in gnotobiotic mice.

PubMed Central

Hamilton, H L; Czuprynski, C J

1992-01-01

In this study the effect of supplementation of drinking water with mycobactin J or lactoferrin on the multiplication of Mycobacterium paratuberculosis in gnotobiotic mice was investigated. The results indicated that neither mycobactin J nor lactoferrin, at the doses used, appeared to have any effect on fecal shedding or tissue burdens of M. paratuberculosis. PMID:1586898
LPS-induced NF-{kappa}B expression in THP-1Blue cells correlates with neopterin production and activity of indoleamine 2,3-dioxygenase

DOE Office of Scientific and Technical Information (OSTI.GOV)

Schroecksnadel, Sebastian; Jenny, Marcel; Division of Medical Biochemistry, Biocenter, Innsbruck Medical University, Innsbruck

2010-09-03

Research highlights: {yields} LPS induces NF-{kappa}B, neopterin formation and tryptophan degradation in THP-1 cells. {yields} Close dose- and time-dependent correlations exist between these biochemical events. {yields} Data provides some evidence for a parallel induction of them upon TLR stimulation. {yields} Results can be of considerable relevance also in vivo. -- Abstract: Neopterin production is induced in human monocyte-derived macrophages and dendritic cells upon stimulation with Th1-type cytokine interferon-{gamma} (IFN-{gamma}). In parallel, IFN-{gamma} induces the tryptophan-(trp)-degrading enzyme indoleamine 2,3-dioxygenase (IDO) and triggers the formation of reactive oxygen species (ROS). Translocation of the signal transduction element nuclear factor-{kappa}B (NF-{kappa}B) is induced bymore » ROS and accelerates the pro-inflammatory response by activation of other pro-inflammatory pathways. Therefore, a close relationship between NF-{kappa}B expression, the production of neopterin and the degradation of trp can be assumed, although this has not been demonstrated so far. In the present in vitro study we compared the influence of lipopolysaccharide (LPS) on NF-{kappa}B activation, neopterin formation and the degradation of trp in THP-1Blue cells, which represent the human myelomonocytic cell line THP-1 stably transfected with an NF-{kappa}B inducible reporter system. In cells stimulated with LPS, a significant induction of NF-{kappa}B was observed, and this was paralleled by an increase of kynureunine (kyn) and neopterin concentrations and a decline of trp. The increase of the kyn to trp quotient indicates accelerated IDO activity. Higher LPS concentrations and longer incubation of cells were associated with higher activities of all three biochemical pathways and significant correlations existed between NF-{kappa}B activation, neopterin release and trp degradation (all p < 0.001). We conclude that there is a parallel induction of NF-{kappa}B, neopterin
Human Milk Secretory Immunoglobulin A and Lactoferrin N-Glycans Are Altered in Women with Gestational Diabetes Mellitus123

PubMed Central

Smilowitz, Jennifer T.; Totten, Sarah M.; Huang, Jincui; Grapov, Dmitry; Durham, Holiday A.; Lammi-Keefe, Carol J.; Lebrilla, Carlito; German, J. Bruce

2013-01-01

Very little is known about the effects of gestational diabetes mellitus (GDM) on lactation and milk components. Recent reports suggested that hyperglycemia during pregnancy was associated with altered breast milk immune factors. Human milk oligosaccharides (HMOs) and N-glycans of milk immune-modulatory proteins are implicated in modulation of infant immunity. The objective of the current study was to evaluate the effect of GDM on HMO and protein-conjugated glycan profiles in breast milk. Milk was collected at 2 wk postpartum from women diagnosed with (n = 8) or without (n = 16) GDM at week 24–28 in pregnancy. Milk was analyzed for HMO abundances, protein concentrations, and N-glycan abundances of lactoferrin and secretory immunoglobulin A (sIgA). HMOs and N-glycans were analyzed by mass spectrometry and milk lactoferrin and sIgA concentrations were analyzed by the Bradford assay. The data were analyzed using multivariate modeling confirmed with univariate statistics to determine differences between milk of women with compared with women without GDM. There were no differences in HMOs between milk from women with vs. without GDM. Milk from women with GDM compared with those without GDM was 63.6% lower in sIgA protein (P < 0.05), 45% higher in lactoferrin total N-glycans (P < 0.0001), 36–72% higher in lactoferrin fucose and sialic acid N-glycans (P < 0.01), and 32–43% lower in sIgA total, mannose, fucose, and sialic acid N-glycans (P < 0.05). GDM did not alter breast milk free oligosaccharide abundances but decreased total protein and glycosylation of sIgA and increased glycosylation of lactoferrin in transitional milk. The results suggest that maternal glucose dysregulation during pregnancy has lasting consequences that may influence the innate immune protective functions of breast milk. PMID:24047700

42 CFR 483.132 - Evaluating the need for NF services and NF level of care (PASARR/NF).

Code of Federal Regulations, 2010 CFR

2010-10-01

... and 483.136, the State mental health or mental retardation authority must determine whether an NF... 42 Public Health 5 2010-10-01 2010-10-01 false Evaluating the need for NF services and NF level of care (PASARR/NF). 483.132 Section 483.132 Public Health CENTERS FOR MEDICARE & MEDICAID SERVICES...
Human T Cell Leukemia Virus Type 2 Tax-Mediated NF-κB Activation Involves a Mechanism Independent of Tax Conjugation to Ubiquitin and SUMO

PubMed Central

Journo, Chloé; Bonnet, Amandine; Favre-Bonvin, Arnaud; Turpin, Jocelyn; Vinera, Jennifer; Côté, Emilie; Chevalier, Sébastien Alain; Kfoury, Youmna; Bazarbachi, Ali

2013-01-01

Permanent activation of the NF-κB pathway by the human T cell leukemia virus type 1 (HTLV-1) Tax (Tax1) viral transactivator is a key event in the process of HTLV-1-induced T lymphocyte immortalization and leukemogenesis. Although encoding a Tax transactivator (Tax2) that activates the canonical NF-κB pathway, HTLV-2 does not cause leukemia. These distinct pathological outcomes might be related, at least in part, to distinct NF-κB activation mechanisms. Tax1 has been shown to be both ubiquitinated and SUMOylated, and these two modifications were originally proposed to be required for Tax1-mediated NF-κB activation. Tax1 ubiquitination allows recruitment of the IKK-γ/NEMO regulatory subunit of the IKK complex together with Tax1 into centrosome/Golgi-associated cytoplasmic structures, followed by activation of the IKK complex and RelA/p65 nuclear translocation. Herein, we compared the ubiquitination, SUMOylation, and acetylation patterns of Tax2 and Tax1. We show that, in contrast to Tax1, Tax2 conjugation to endogenous ubiquitin and SUMO is barely detectable while both proteins are acetylated. Importantly, Tax2 is neither polyubiquitinated on lysine residues nor ubiquitinated on its N-terminal residue. Consistent with these observations, Tax2 conjugation to ubiquitin and Tax2-mediated NF-κB activation is not affected by overexpression of the E2 conjugating enzyme Ubc13. We further demonstrate that a nonubiquitinable, non-SUMOylable, and nonacetylable Tax2 mutant retains a significant ability to activate transcription from a NF-κB-dependent promoter after partial activation of the IKK complex and induction of RelA/p65 nuclear translocation. Finally, we also show that Tax2 does not interact with TRAF6, a protein that was shown to positively regulate Tax1-mediated activation of the NF-κB pathway. PMID:23135727
Oral lactoferrin for the treatment of sepsis and necrotizing enterocolitis in neonates

USDA-ARS?s Scientific Manuscript database

Neonatal sepsis and necrotizing enterocolitis (NEC) cause significant neonatal mortality and morbidity in spite of appropriate antibiotic therapy. Enhancing host defense and modulating inflammation by using lactoferrin as an adjunct to antibiotics in the treatment of sepsis and/or NEC may improve cl...
Oral lactoferrin for the treatment of sepsis and necrotizing enterocolitis in neonates

USDA-ARS?s Scientific Manuscript database

Neonatal sepsis and necrotizing enterocolitis (NEC) cause significant neonatal mortality and morbidity in spite of appropriate antibiotic therapy. Enhancing host defence and modulating inflammation by using lactoferrin as an adjunct to antibiotics in the treatment of sepsis and/or NEC may improve cl...
Extracts of Porphyra tenera (Nori Seaweed) Activate the Immune Response in Mouse RAW264.7 Macrophages via NF-κB Signaling.

PubMed

Song, Ji-Hye; Kang, Hee-Bum; Park, Seung-Ho; Jeong, Ji-Hoon; Park, Jeongjin; You, Yanghee; Lee, Yoo-Hyun; Lee, Jeongmin; Kim, Eungpil; Choi, Kyung-Chul; Jun, Woojin

2017-12-01

Porphyra tenera, also known as nori, is a red algal species of seaweed. It is cultivated in Asia for culinary purposes. We report that P. tenera extract (PTE) enhances the immune response in mouse macrophages. We found that P. tenera extract regulates the NF-κB IκB kinase (IKK) signaling pathway, and we assessed the expression and translocation of p65, a subunit of NF-κB, in RAW264.7 mouse macrophage cells after treatment with PTE. We also investigated the effects of 10% ethanol PTE (PTE10) in RAW264.7 cells. The production of IL-10, IL-6, TNF-α, and IFN-γ was induced by PTE treatment of the macrophages, and PTE also enhanced p-IκB and p-AKT. PTE10 showed no cytotoxicity at 10-20 μg/mL in RAW264.7 cells. PTE10, in fact, increased cell viability at 24 h, stimulated macrophage cells, and induced the phosphorylation of Akt. Akt stimulates IKK activity through the phosphorylation of IKKα and enhances immune activity through the activation of NF-κB. In this study, NF-κB activation was induced by increasing p-NF-κB and p-IKK. A subunit of NF-κB, p65, was located in the nucleus and increased the expression of various cytokines. PTE thus enhanced the immune response through IκB-α immunostimulation signaling in RAW264.7 cells. PTE10 has potential therefore for development of future treatments requiring immune system stimulation.
A Critical Role for IL-17RB Signaling in HTLV-1 Tax-Induced NF-κB Activation and T-Cell Transformation

PubMed Central

Lavorgna, Alfonso; Matsuoka, Masao; Harhaj, Edward William

2014-01-01

Human T-cell leukemia virus type 1 (HTLV-1) infection is linked to the development of adult T-cell leukemia (ATL) and the neuroinflammatory disease HTLV-1 associated myelopathy/tropical spastic paraparesis (HAM/TSP). The HTLV-1 Tax protein functions as a potent viral oncogene that constitutively activates the NF-κB transcription factor to transform T cells; however, the underlying mechanisms remain obscure. Here, using next-generation RNA sequencing we identified the IL-25 receptor subunit IL-17RB as an aberrantly overexpressed gene in HTLV-1 immortalized T cells. Tax induced the expression of IL-17RB in an IκB kinase (IKK) and NF-κB-dependent manner. Remarkably, Tax activation of the canonical NF-κB pathway in T cells was critically dependent on IL-17RB expression. IL-17RB and IL-25 were required for HTLV-1-induced immortalization of primary T cells, and the constitutive NF-κB activation and survival of HTLV-1 transformed T cells. IL-9 was identified as an important downstream target gene of the IL-17RB pathway that drives the proliferation of HTLV-1 transformed cells. Furthermore, IL-17RB was overexpressed in leukemic cells from a subset of ATL patients and also regulated NF-κB activation in some, but not all, Tax-negative ATL cell lines. Together, our results support a model whereby Tax instigates an IL-17RB-NF-κB feed-forward autocrine loop that is obligatory for HTLV-1 leukemogenesis. PMID:25340344
Ezrin/NF-kB activation regulates epithelial- mesenchymal transition induced by EGF and promotes metastasis of colorectal cancer.

PubMed

Li, Yingru; Lin, Zhaoyu; Chen, Bin; Chen, Shuang; Jiang, Zhipeng; Zhou, Taicheng; Hou, Zehui; Wang, Youyuan

2017-08-01

There is growing evidence that epithelial mesenchymal-transition (EMT) plays significant roles in terms of tumor metastasis. There are a lot of cytokines inducing EMT of tumor cells, EGF is one of the important cytokines.Ezrin is a connexin between the cytoskeleton and the cell membrane, which is closely related to the morphological movement and metastasis of tumor cells.EGF can activate Ezrin and affects cell motility. In recent years, many studies have shown that NF-kB acts as an important transcription factor, involving in the process of EMT. However, does Ezrin participate in the regulation of EGF-induced EMT through the NF-kB pathway? This question needs us to discuss.In the present study, we found that EGF could induce colorectal cancer cells to develop EMT,enhance their ability to invade and migrate and promotes phosphorylation of Ezrin Tyr353.On the other hand, inhibition of Ezrin could reverse EGF-induced EMT and inhibit NF-kB P65 translocating into the nucleus. Finally, knockout of Ezrin inhibited EGF-induced lung metastasis of colorectal cancer xenografts and abnormal activation of Ezrin and NF-kB were related with colorectal cancer metastasis and poor prognosis. Our present results suggest that Ezrin/NF-kB pathway may provide experimental evidence for new targeted drugs for colorectal cancer metastasis. Copyright © 2017 Elsevier Masson SAS. All rights reserved.
Betulinic acid ameliorates experimental diabetic-induced renal inflammation and fibrosis via inhibiting the activation of NF-κB signaling pathway.

PubMed

Wang, Shaogui; Yang, Zhiying; Xiong, Fengxiao; Chen, Cheng; Chao, Xiaojuan; Huang, Junying; Huang, Heqing

2016-10-15

Diabetic nephropathy (DN) is the leading cause of end-stage renal failure and is characterized by excessive deposition of extracellular matrix (ECM) proteins such as fibronectin (FN), in the glomerular mesangium and tubulointerstitium. Betulinic acid (BA), a pentacyclic triterpene derived from the bark of the white birch tree, has been demonstrated to have many pharmacological activities. However, the effect of BA on DN has not been fully elucidated. To explore the possible anti-inflammatory effects of BA and their underlying mechanisms, we used streptozotocin-induced diabetic rat kidneys and high glucose-treated glomerular mesangial cells. Our study showed BA could inhibit the degradation of IκBα and the activity of NF-κB in diabetic rat kidneys and high glucose-induced mesangial cells, resulting in reduction of FN expression. In addition, BA suppressed the DNA binding activity and transcriptional activity of NF-κB in high glucose-induced glomerular mesangial cells (GMCs). Furthermore, BA enhanced the interaction between IκBα and β-arrestin2 in mesangial cells. Taken together, our data suggest BA inhibits NF-κB activation through stabilizing NF-κB inhibitory protein IκBα, thereby preventing diabetic renal fibrosis. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.
PSMB4 promotes multiple myeloma cell growth by activating NF-κB-miR-21 signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zheng, Peihao; Guo, Honggang; Li, Guangchao

2015-03-06

Proteasomal subunit PSMB4, was recently identified as potential cancer driver genes in several tumors. However, the regulatory mechanism of PSMB4 on carcinogenesis process remains unclear. In this study, we investigated the expression and roles of PSMB4 in multiple myeloma (MM). We found a significant up-regulation of PSMB4 in MM plasma and cell lines. Ectopic overexpression of PSMB4 promoted cell growth and colony forming ability of MM cells, whereas inhibition of PSMB4 led to a decrease of such events. Furthermore, our results demonstrated the up-regulation of miR-21 and a positive correlation between the levels of miR-21 and PSMB4 in MM. Re-expressionmore » of miR-21 markedly rescued PSMB4 knockdown-mediated suppression of cell proliferation and clone-formation. Additionally, while enforced expression of PSMB4 profoundly increased NF-κB activity and the level of miR-21, PSMB4 knockdown or NF-κB inhibition suppressed miR-21 expression in MM cells. Taken together, our results demonstrated that PSMB4 regulated MM cell growth in part by activating NF-κB-miR-21 signaling, which may represent promising targets for novel specific therapies. - Highlights: • First reported upregulation of PSMB4 in MM plasma and cell lines. • PSMB4 promoted MM cell growth and colony forming ability. • Further found miR-21 was up-regulated by PSMB4 in MM plasma and cell lines. • PSMB4-induced miR-21 expression was modulated by NF-κB. • PSMB4-NF-κB-miR-21 axis may be potential therapeutic targets of MM.« less
rsfMRI effects of KB220Z™ on Neural Pathways in Reward Circuitry of Abstinent Genotyped Heroin Addicts

PubMed Central

Blum, Kenneth; Liu, Yijun; Wang, Wei; Wang, Yarong; Zhang, Yi; Oscar-Berman, Marlene; Smolen, Andrew; Febo, Marcelo; Han, David; Simpatico, Thomas; Cronjé, Frans J; Demetrovics, Zsolt; Gold, Mark S.

2016-01-01

Recently Willuhn et al. reported that cocaine use and even non-substance related addictive behavior, increases, as dopaminergic function is reduced. Chronic cocaine exposure has been associated with decreases in D2/D3 receptors, also associated with lower activation to cues in occipital cortex and cerebellum in a recent PET study from Volkow’s group. Therefore, treatment strategies, like dopamine agonist therapy, that might conserve dopamine function may be an interesting approach to relapse prevention in psychoactive drug and behavioral addictions. To this aim, we evaluated the effect of KB220Z™ on reward circuitry of ten heroin addicts undergoing protracted abstinence, an average 16.9 months. In a randomized placebo-controlled crossover study of KB220Z™ five subjects completed a triple blinded–experiment in which the subject, the person administering the treatment and the person evaluating the response to treatment were blinded as to which treatment any particular subject was receiving. In addition, nine subjects total were genotyped utilizing the GARSRX™ test. We preliminarily report that KB220Z ™ induced an increase in BOLD activation in caudate-accumbens-dopaminergic pathways compared to placebo following one-hour acute administration. Furthermore, KB220Z™ also reduced resting state activity in the putamen of abstinent heroin addicts. In the second phase of this pilot study of all ten abstinent heroin-dependent subjects, three brain regions of interest (ROIs) we observed to be significantly activated from resting state by KB220Z compared to placebo (P < 0.05). Increased functional connectivity was observed in a putative network that included the dorsal anterior cingulate, medial frontal gyrus, nucleus accumbens, posterior cingulate, occipital cortical areas and cerebellum. These results and other qEEG study results suggest a putative anti-craving/anti-relapse role for KB220Z in addiction by direct or indirect dopaminergic interaction. Due to
In vitro anticancer property of a novel thalidomide analogue through inhibition of NF-kappaB activation in HL-60 cells.

PubMed

Li, Min; Sun, Wan; Yang, Ya-ping; Xu, Bo; Yi, Wen-yuan; Ma, Yan-xia; Li, Zhong-jun; Cui, Jing-rong

2009-01-01

To investigate the anticancer property and possible mechanism of action of a novel sugar-substituted thalidomide derivative (STA-35) on HL-60 cells in vitro. TNF-alpha-induced NF-kappaB activation was determined using a reporter gene assay. The MTT assay was used to measure cytotoxicity of the compound. The appearance of apoptotic Sub-G1 cells was detected by flow cytometry analysis. PARP cleavage and protein expression of NF-kappaB p65 and its inhibitor IkappaB were viewed by Western blotting. TA-35 (1-20 micromol/L) suppressed TNF-alpha-induced NF-kappaB activation in transfected cells (HEK293/pNiFty-SEAP) in a dose- (1-20 micromol/L) and time-dependent (0-48 h) manner. It was also shown that STA-35 exerted a dose-dependent inhibitory effect on HL-60 cell proliferation with an IC(50) value of 9.05 micromol/L. In addition, STA-35 induced apoptosis in HL-60 cells, as indicated by the appearance of a Sub-G1 peak in the cell cycle distribution, as well as poly ADP-ribose polymerase (PARP) cleavage. Subsequently, both NF-kappaB p65 and its inhibitor IkappaB gradually accumulated in cytoplasmic extracts in a dose- and time-dependent manner, indicating the blockage of NF-kappaB translocation induced by TNF-alpha from the cytoplasm to the nucleus. A novel sugar-substituted thalidomide derivative, STA-35, is potent toward HL-60 cells in vitro and induces apoptosis by the suppression of NF-kappaB activation.
Uptake of ingested bovine lactoferrin and its accumulation in adult mouse tissues.

PubMed

Fischer, Romy; Debbabi, Hajer; Blais, Anne; Dubarry, Michel; Rautureau, Michèle; Boyaka, Prosper N; Tome, Daniel

2007-10-01

Lactoferrin is a glycoprotein with antimicrobial and immunoregulatory properties, which is found in milk, other external secretions, and in the secondary granules of neutrophils. The present study examined the time course of uptake and the pattern of tissue accumulation of bovine lactoferrin (bLf) following intragastric intubation of a single dose to adult naïve mice or to mice daily fed bLf for 4 weeks. Following ingestion, bLf was transferred from the intestine into peripheral blood in a form with intact molecular weight (80 kDa) and localized within 10 to 20 min after oral administration in the liver, kidneys, gall bladder, spleen, and brain of both groups of mice. Immunoreactive bLf could also be detected in the luminal contents of the stomach, small intestine and colon 1 h after intragastric intubation. Interestingly, serum and tissue accumulation of bLf was approximately 50% lower in mice chronically fed this protein than in those given only the single oral dose. Furthermore, significant levels of bLf-specific IgA and IgG antibodies as well as bLf-containing IgA- and IgG immune complexes were detected in mice chronically fed bLf but not in those fed only once. Taken together, these results indicate that bLf resists major proteolytic degradation in the intestinal lumen and is readily absorbed in an antigenic form in blood and various mouse tissues. Chronic ingestion of lactoferrin reduces its uptake, probably through mechanisms such as immune exclusion, which minimize potential harmful reactions to food products.
Human papillomavirus genotypes in penile cancers from Japanese patients and HPV-induced NF-κB activation.

PubMed

Senba, Masachika; Mori, Naoki; Wada, Akihiro; Fujita, Shuichi; Yasunami, Michio; Irie, Sumiko; Hayashi, Tomayoshi; Igawa, Tsukasa; Kanetake, Hiroshi; Takahara, Osamu; Toriyama, Kan

2010-03-01

The causal relationship between chronic inflammation and cancer is widely accepted. Numerous investigations have identified nuclear factor-κB (NF-κB) as an important modulator in driving chronic inflammation to cancer. This study aimed to determine the prevalence of human papillomavirus (HPV) in penile cancer in Japanese patients and whether NF-κB is subsequently overexpressed in penile cancer. Thirty-four specimens of penile tissue (16 malignant and 18 benign cases) were examined to determine the association of HPV infection. An in situ hybridization (ISH) method was used to detect and localize HPV-DNA. A sensitive HPV polymerase chain reaction (PCR) procedure was used for the detection of HPV-DNA, and DNA sequencing was used to identify the HPV genotype. HPV-DNA was detected in 37.5 and 75% of cases of penile cancer, using ISH and PCR, respectively. Our efforts to detect HPV genotypes were unsuccessful as HPV-DNA could not be extracted from these materials. Using ISH, a prevalence of 68.2% of HPV infection was found in penile cancer in Kenyan patients in east Africa. In the present study, all 9 HPV-positive cases, (100%) were NF-κB-positive in the nucleus and/or cytoplasm. In contrast, of the 25 HPV-negative cases, 15 (60%) were NF-κB-positive in the nucleus and/or cytoplasm. Therefore, ISH is a method which is able to prove infection of a large quantity of HPV more effectively when compared with PCR. Thus, a large quantity of HPV infection leads to the activity of NF-κB. The most prevalent genotype was the HPV-22 found in 83.3% of the penile cancer cases. In addition, HPV-11 was found in 81.8% of the non-cancer cases. For cases with a high level of infection, the activity of NF-κB increased compared with those with a low level of HPV infection.
Enterovirus 71 2C Protein Inhibits NF-κB Activation by Binding to RelA(p65)

PubMed Central

Du, Haiwei; Yin, Peiqi; Yang, Xiaojie; Zhang, Leiliang; Jin, Qi; Zhu, Guofeng

2015-01-01

Viruses evolve multiple ways to interfere with NF-κB signaling, a key regulator of innate and adaptive immunity. Enterovirus 71 (EV71) is one of primary pathogens that cause hand-foot-mouth disease. Here, we identify RelA(p65) as a novel binding partner for EV71 2C protein from yeast two-hybrid screen. By interaction with IPT domain of p65, 2C reduces the formation of heterodimer p65/p50, the predominant form of NF-κB. We also show that picornavirus 2C family proteins inhibit NF-κB activation and associate with p65 and IKKβ. Our findings provide a novel mechanism how EV71 antagonizes innate immunity. PMID:26394554
Gastrin regulates ABCG2 to promote the migration, invasion and side populations in pancreatic cancer cells via activation of NF-κB signaling

DOE Office of Scientific and Technical Information (OSTI.GOV)

Wang, Juan; Xin, Beibei; Wang, Hui

Gastrin is absent in most normal adult pancreatic tissues but is highly expressed in pancreatic cancer tissues. Although Gastrin expression was reported to be associated with tumor proliferation in human pancreatic cancer, studies on the relationship between Gastrin and tumor metastasis in pancreatic cancer are rare. In this study, we performed an analysis to determine the effects of Gastrin on modulating the side populations, cell proportion and tumor cell metastatic potential and invasion activity and explored its mechanisms in pancreatic cancer. We indicated that Gastrin and ABCG2 were widely expressed in pancreatic cancer cell lines and overexpressed in cancer tissues.more » Gastrin induced ABCG2 expression, and this effect was mediated by NF-κB activation. Gastrin regulated the SP proportion of BxPC-3 cells via modulating ABCG2 expression. Through the regulation of the functions of NF-κB/ABCG2, Gastrin functionally promoted the migration and invasion in pancreatic cancer cell. The present study indicated that Gastrin induced ABCG2 expression by activating NF-κB and thereby modulated the SP proportion, tumor cell metastatic potential and invasion activity in pancreatic cancer. Gastrin could serve as an effective therapeutic target for the metastasis of pancreatic cancer. - Highlights: • Gastrin induces ABCG2 expression mediated by NF-κB activation. • Gastrin regulates NF-κB's function that binds to the ABCG2 promoter in BxPC-3 cells. • Gastrin promotes the SP proportion in BxPC-3 cells by modulating ABCG2 expression via activation of NF-κB molecule. • Gastrin induces an increase in migration and invasion potential in pancreatic cancer cell by regulating NF-κB/ABCG2 signaling.« less
The Pseudomonas aeruginosa exopolysaccharide Psl facilitates surface adherence and NF-kappaB activation in A549 cells.

PubMed

Byrd, Matthew S; Pang, Bing; Mishra, Meenu; Swords, W Edward; Wozniak, Daniel J

2010-06-29

In order for the opportunistic Gram-negative pathogen Pseudomonas aeruginosa to cause an airway infection, the pathogen interacts with epithelial cells and the overlying mucous layer. We examined the contribution of the biofilm polysaccharide Psl to epithelial cell adherence and the impact of Psl on proinflammatory signaling by flagellin. Psl has been implicated in the initial attachment of P. aeruginosa to biotic and abiotic surfaces, but its direct role in pathogenesis has not been evaluated (L. Ma, K. D. Jackson, R. M. Landry, M. R. Parsek, and D. J. Wozniak, J. Bacteriol. 188:8213-8221, 2006). Using an NF-kappaB luciferase reporter system in the human epithelial cell line A549, we show that both Psl and flagellin are necessary for full activation of NF-kappaB and production of the interleukin 8 (IL-8) chemokine. We demonstrate that Psl does not directly stimulate NF-kappaB activity, but indirectly as a result of increasing contact between bacterial cells and epithelial cells, it facilitates flagellin-mediated proinflammatory signaling. We confirm differential adherence of Psl and/or flagellin mutants by scanning electron microscopy and identify Psl-dependent membrane structures that may participate in adherence. Although we hypothesized that Psl would protect P. aeruginosa from recognition by the epithelial cell line A549, we instead observed a positive role for Psl in flagellin-mediated NF-kappaB activation, likely as a result of increasing contact between bacterial cells and epithelial cells.
The Pseudomonas aeruginosa Exopolysaccharide Psl Facilitates Surface Adherence and NF-κB Activation in A549 Cells

PubMed Central

Byrd, Matthew S.; Pang, Bing; Mishra, Meenu; Swords, W. Edward; Wozniak, Daniel J.

2010-01-01

In order for the opportunistic Gram-negative pathogen Pseudomonas aeruginosa to cause an airway infection, the pathogen interacts with epithelial cells and the overlying mucous layer. We examined the contribution of the biofilm polysaccharide Psl to epithelial cell adherence and the impact of Psl on proinflammatory signaling by flagellin. Psl has been implicated in the initial attachment of P. aeruginosa to biotic and abiotic surfaces, but its direct role in pathogenesis has not been evaluated (L. Ma, K. D. Jackson, R. M. Landry, M. R. Parsek, and D. J. Wozniak, J. Bacteriol. 188:8213–8221, 2006). Using an NF-κB luciferase reporter system in the human epithelial cell line A549, we show that both Psl and flagellin are necessary for full activation of NF-κB and production of the interleukin 8 (IL-8) chemokine. We demonstrate that Psl does not directly stimulate NF-κB activity, but indirectly as a result of increasing contact between bacterial cells and epithelial cells, it facilitates flagellin-mediated proinflammatory signaling. We confirm differential adherence of Psl and/or flagellin mutants by scanning electron microscopy and identify Psl-dependent membrane structures that may participate in adherence. Although we hypothesized that Psl would protect P. aeruginosa from recognition by the epithelial cell line A549, we instead observed a positive role for Psl in flagellin-mediated NF-κB activation, likely as a result of increasing contact between bacterial cells and epithelial cells. PMID:20802825
Resveratrol supplement inhibited the NF-κB inflammation pathway through activating AMPKα-SIRT1 pathway in mice with fatty liver.

PubMed

Tian, Yueli; Ma, Jingting; Wang, Wudong; Zhang, Lingjuan; Xu, Jia; Wang, Kai; Li, Dongfu

2016-11-01

Nonalcoholic fatty liver disease (NAFLD) is characterized by high levels of nonesterified fatty acids (NEFA), inflammation, and hepatic steatosis. Inflammation plays a crucial role in the development of fatty liver. Resveratrol (RSV) supplement could improve inflammatory response and hepatic steatosis, whereas the underlying mechanism was not well understood. In this study, mice fed with high-fat diet (HFD) exhibited severe hepatic injury and high blood concentrations of the inflammatory cytokines TNF-α, IL-6, and IL-1β. Hepatic NF-κB inflammatory pathway was over-induced in HFD mice. In vitro, NEFA treatment further increased NF-κB pathway activation in mice hepatocytes, which then promoted the synthesis of inflammatory cytokines. Interestingly, RSV treatment significantly inhibited overactivation of NF-κB pathway and improved hepatic steatosis. Furthermore, RSV further increased the AMP-activated protein kinaseα (AMPKα) phosphorylation and sirtuin1 (SIRT1) protein levels to inhibit overactivation of NF-κB pathway induced by HFD or high levels of NEFA. AMPKα or SIRT1 inhibition significantly decreased the improvement effect of RSV on the NF-κB pathway induced by high levels of NEFA. Taken together, these findings indicate that RSV supplement decreases the inflammatory level and improves hepatic steatosis through activating AMPKα-SIRT1 pathway. Therefore, these data suggested an important clinical application of RSV in preventing NAFLD in humans.
Constitutive activation of IKK2/NF-κB impairs osteogenesis and skeletal development.

PubMed

Swarnkar, Gaurav; Zhang, Kaihua; Mbalaviele, Gabriel; Long, Fanxin; Abu-Amer, Yousef

2014-01-01

Pathologic conditions impair bone homeostasis. The transcription factor NF-κB regulates bone homeostasis and is central to bone pathologies. Whereas contribution of NF-κB to heightened osteoclast activity is well-documented, the mechanisms underlying NF-κB impact on chondrocytes and osteoblasts are scarce. In this study, we examined the effect of constitutively active IKK2 (IKK2ca) on chondrogenic and osteogenic differentiation. We show that retroviral IKK2ca but not GFP, IKK2WT, or the inactive IKK2 forms IKK2KM and IKK2SSAA, strongly suppressed osteogenesis and chondrogenesis, in vitro. In order to explore the effect of constitutive NF-κB activation on bone formation in vivo, we activated this pathway in a conditional fashion. Specifically, we crossed the R26StopIKK2ca mice with mice carrying the Col2-cre in order to express IKK2ca in osteoblasts and chondrocytes. Both chondrocytes and osteoblasts derived from Col2Cre/IKK2ca expressed IKK2ca. Mice were born alive yet died shortly thereafter. Histologically, newborn Col2Cre+/RosaIKK2ca heterozygotes (Cre+IKK2ca_w/f (het)) and homozygotes (Cre+IKK2ca_f/f (KI)) showed smaller skeleton, deformed vertebrate and reduced or missing digit ossification. The width of neural arches, as well as ossification in vertebral bodies of Cre+IKK2ca_w/f and Cre+IKK2ca_f/f, was reduced or diminished. H&E staining of proximal tibia from new born pups revealed that Cre+IKK2ca_f/f displayed disorganized hypertrophic zones within the smaller epiphysis. Micro-CT analysis indicated that 4-wk old Cre+IKK2ca_w/f has abnormal trabecular bone in proximal tibia compared to WT littermates. Mechanistically, ex-vivo experiments showed that expression of differentiation markers in calvarial osteoblasts derived from newborn IKK2ca knock-in mice was diminished compared to WT-derived cells. In situ hybridization studies demonstrated that the hypertrophic chondrocyte marker type-X collagen, the pre-hypertrophic chondrocyte markers Indian hedgehog
[Expressions of HSP 70 and NF-kappaB in the peripheral blood lymphocyte of chronic gastritis patients of different syndrome patterns].

PubMed

Hu, Ling; Zheng, Xiao-Feng; Yan, Xue-Hui

2012-09-01

To study the expressions of heat shock protein 70 (HSP 70) and nuclear factor-kappa B (NF-kappaB) in the peripheral blood lymphocyte of chronic gastritis (CG) patients of Pi-Wei hygropyrexia syndrome (PWHS) and Pi-qi deficiency syndrome (PQDS), and to explore their correlation with Helicobacter pylori (Hp) infection. Recruited were totally 86 CG patients who visited at the clinics of gastroenterology, First Affiliated Hospital of Guangzhou University of Traditional Chinese Medicine, including 67 patients of PWHS (30 of predominant-dampness, 30 of equal dampness and heat, and 30 of predominant-heat) and 19 patients of PQDS. Another 12 volunteers from healthy employees and students of Guangzhou University of Traditional Chinese Medicine were recruited as the control group. Their peripheral blood was collected. The Hp infection was detected using ASSURE Hp rapid test. The expressions of HSP 70 and NF-kappaB in the peripheral blood lymphocyte were detected using flow cytometry. The Hp infection rate was 37. 31% in the GS patients of PWHS and 36. 84% in the GS patients of PQDS (P>0.05). Compared with the control group, the expression of HSP 70 decreased in the PWHS predominant-heat group, and the expression of NF-kappaB increased in the PWHS predominant-heat group and the PQDS group (P<0.05). The expression of NF-kappaB were higher in the positive Hp infection patients of PWHS and PQDS than in the control group (P<0.05). The expression of HSP 70 was higher in the positive Hp infection patients of PQDS than in the negative Hp infection patients of PQDS (P<0.05). Besides, the coefficient correlation was -0. 023 between HSP 70 and Hp infection, and 0. 027 between NF-KB and Hp infection (P>0.05). The increased expression of NF-KB in the peripheral blood lymphocyte of CG patients of PWHS and PQDS might reflect the pathogenic roles of "inner evil" in Chinese medicine theories. The increased expression of HSP 70 in CG patients of PQDS and decreased expression of HSP 70 in CG

Evaluation of NF-kappaB Pathway Inhibition for Space Radiation Biology Research

NASA Astrophysics Data System (ADS)

Koch, Kristina; Hellweg, Christine; Baumstark-Khan, Christa; Schmitz, Claudia; Lau, Patrick; Testard, Isabelle; Reitz, Guenther

Radiation is a potentially limiting factor for long term orbital and interplanetary missions. To improve risk estimation and to allow development of appropriate countermeasures, the study of the cellular radiation response is necessary. The anti-apoptotic factor nuclear factor κB (NF-κB) was identified as important modulating factor in the cellular response to heavy ions (Radiat. Res. 164: 527-530, 2005). This transcription factor could improve cellular survival after exposure to high radiation doses and influence the cancer risk of astronauts exposed to low doses of cosmic radiation. Therefore, the inhibition of selected NF-κB pathway compo-nents might help to identify possible pharmacological targets. It is supposed that the ATM kinase mediates the signal from damaged DNA in the nucleus to kinases in the cytoplasm. For liberation of NF-κB and its nuclear translocation, the inhibitor of NF-κB (IκB) has to be degraded in the proteasom. In this work, the efficacy and cytotoxicity of ATM, NF-κB and the proteasome inhibitors were analyzed using recombinant HEK-pNF-κB-d2EGFP/Neo cells. In the recommended concentration range, only the NF-κB inhibitor caffeic acid phenethyl ester (CAPE) displayed considerable cytotoxicity, while the others were not toxic. The inhibition of ATM by KU-55933 suppresses the X-ray and heavy ion (13 C, 35 MeV/u, LET 70 keV/m) induced activation of NF-κB dependent gene expression, indicating the central position of ATM in radiation induced NF-κB activation. CAPE and capsaicin partially inhibited NF-κB acti-vation by the cytokine tumor necrosis factor α. The proteasome inhibitor MG-132 completely abolished the activation and was therefore used for short-term incubation experiments with X-rays. MG-132 suppressed the X-ray induced NF-κB activation in HEK-pNF-κB-d2EGFP/Neo cells entirely. The results lead to the conclusion that ATM and the proteasomal degradation of IκB are essential prerequisites for radiation induced NF
Human lactoferrin stimulates skin keratinocyte function and wound re-epithelialization.

PubMed

Tang, L; Wu, J J; Ma, Q; Cui, T; Andreopoulos, F M; Gil, J; Valdes, J; Davis, S C; Li, J

2010-07-01

Human lactoferrin (hLF), a member of the transferrin family, is known for its antimicrobial and anti-inflammatory effects. Recent studies on various nonskin cell lines indicate that hLF may have a stimulatory effect on cell proliferation. To study the potential role of hLF in wound re-epithelialization. The effects of hLF on cell growth, migration, attachment and survival were assessed, with a rice-derived recombinant hLF (holo-rhLF), using proliferation analysis, scratch migration assay, calcein-AM/propidium iodide staining and terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL) method, respectively. The mechanisms of hLF on cell proliferation and migration were explored using specific pathway inhibitors. The involvement of lactoferrin receptor low-density lipoprotein receptor-related protein 1 (LRP1) was examined with RNA interference technique. An in vivo swine second-degree burn wound model was also used to assess wound re-epithelialization. Studies revealed that holo-rhLF significantly stimulated keratinocyte proliferation which could be blocked by mitogen-activated protein kinase (MAPK) kinase 1 inhibitor. Holo-rhLF also showed strong promoting effects on keratinocyte migration, which could be blocked by either inhibition of the MAPK, Src and Rho/ROCK pathways, or downregulation of the LRP1 receptor. With cells under starving or 12-O-tetradecanoylphorbol-13-acetate exposure, the addition of holo-rhLF was found greatly to increase cell viability and inhibit cell apoptosis. Additionally, holo-rhLF significantly increased the rate of wound re-epithelialization in swine second-degree burn wounds. Our studies demonstrate the direct effects of holo-rhLF on wound re-epithelialization including the enhancement of keratinocyte proliferation and migration as well as the protection of cells from apoptosis. The data strongly indicate its potential therapeutic applications in wound healing.
Blocking NF-κB: an inflammatory issue.

PubMed

Rahman, Arshad; Fazal, Fabeha

2011-11-01

The nuclear factor (NF)-κB is considered the master regulator of inflammatory responses. Studies in mouse models have established this transcription factor as an important mediator of many inflammatory disease states, including pulmonary diseases such as acute lung injury and acute respiratory distress syndrome. Endothelial cells provide the first barrier for leukocytes migrating to the inflamed sites and hence offer an attractive cellular context for targeting NF-κB for treatment of these diseases. However, recent studies showing that NF-κB also plays an important role in resolution phase of inflammation and in tissue repair and homeostasis have challenged the view of therapeutic inhibition of NF-κB. This article reviews the regulation of NF-κB in the context of endothelial cell signaling and provides a perspective on why "dampening" rather than "abolishing" NF-κB activation may be a safe and effective treatment strategy for inflammation-associated pulmonary and other inflammatory diseases.
Mesenchymal stem cells promote the sustained expression of CD69 on activated T lymphocytes: roles of canonical and non-canonical NF-κB signalling

PubMed Central

Saldanha-Araujo, Felipe; Haddad, Rodrigo; de Farias, Kelen C R Malmegrim; Souza, Alessandra de Paula Alves; Palma, Patrícia V; Araujo, Amélia G; Orellana, Maristela D; Voltarelli, Julio C; Covas, Dimas T; Zago, Marco A; Panepucci, Rodrigo A

2012-01-01

Abstract Mesenchymal stem cells (MSCs) are known to induce the conversion of activated T cells into regulatory T cells in vitro. The marker CD69 is a target of canonical nuclear factor kappa-B (NF-κB) signalling and is transiently expressed upon activation; however, stable CD69 expression defines cells with immunoregulatory properties. Given its enormous therapeutic potential, we explored the molecular mechanisms underlying the induction of regulatory cells by MSCs. Peripheral blood CD3+ T cells were activated and cultured in the presence or absence of MSCs. CD4+ cell mRNA expression was then characterized by microarray analysis. The drug BAY11-7082 (BAY) and a siRNA against v-rel reticuloendotheliosis viral oncogene homolog B (RELB) were used to explore the differential roles of canonical and non-canonical NF-κB signalling, respectively. Flow cytometry and real-time PCR were used for analyses. Genes with immunoregulatory functions, CD69 and non-canonical NF-κB subunits (RELB and NFKB2) were all expressed at higher levels in lymphocytes co-cultured with MSCs. The frequency of CD69+ cells among lymphocytes cultured alone progressively decreased after activation. In contrast, the frequency of CD69+ cells increased significantly following activation in lymphocytes co-cultured with MSCs. Inhibition of canonical NF-κB signalling by BAY immediately following activation blocked the induction of CD69; however, inhibition of canonical NF-κB signalling on the third day further induced the expression of CD69. Furthermore, late expression of CD69 was inhibited by RELB siRNA. These results indicate that the canonical NF-κB pathway controls the early expression of CD69 after activation; however, in an immunoregulatory context, late and sustained CD69 expression is promoted by the non-canonical pathway and is inhibited by canonical NF-κB signalling. PMID:21777379
The regulation of Jmjd3 upon the expression of NF-κB downstream inflammatory genes in LPS activated vascular endothelial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Yu, Shaoqing; Graduate School of Medicine, Nanchang University, Nanchang; Chen, Xia

Inflammatory mediators and adhesion molecules have been implicated in a variety of diseases including atherosclerosis. As both the mediator-releasing and targeted cells, vascular endothelial cells play key role in pathological processes. NF-κB signaling regulates a cluster of inflammatory factors in LPS-activated vascular endothelial cells but the underlying mechanisms remain largely unknown. Here, we investigated the epigenetic regulation of LPS upon the expression of inflammatory mediators and adhesion molecules. We found that LPS treatment promoted jmjd3 expression, enhanced Jmjd3 nuclear accumulation in human vascular endothelial cells. In addition, LPS enhanced the demethylation of H3K27me3, a specific substrate of Jmjd3. LPS treatmentmore » recruited Jmjd3 and NF-κB to the promoter region of target genes, suggesting Jmjd3 synergizes with NF-κB to activate the expression of target genes. We further found that Jmjd3 attenuated the methylation status in promoter region of target genes, culminating in target gene expression. Our findings unveil epigenetic regulations of LPS upon NF-κB pathway and identify Jmjd3 as a critical modulator of NF-κB pathway and potential therapeutic target for NF-κB related diseases including atherosclerosis.« less
NF-κB inhibition delays DNA damage–induced senescence and aging in mice

PubMed Central

Tilstra, Jeremy S.; Robinson, Andria R.; Wang, Jin; Gregg, Siobhán Q.; Clauson, Cheryl L.; Reay, Daniel P.; Nasto, Luigi A.; St Croix, Claudette M.; Usas, Arvydas; Vo, Nam; Huard, Johnny; Clemens, Paula R.; Stolz, Donna B.; Guttridge, Denis C.; Watkins, Simon C.; Garinis, George A.; Wang, Yinsheng; Niedernhofer, Laura J.; Robbins, Paul D.

2012-01-01

The accumulation of cellular damage, including DNA damage, is thought to contribute to aging-related degenerative changes, but how damage drives aging is unknown. XFE progeroid syndrome is a disease of accelerated aging caused by a defect in DNA repair. NF-κB, a transcription factor activated by cellular damage and stress, has increased activity with aging and aging-related chronic diseases. To determine whether NF-κB drives aging in response to the accumulation of spontaneous, endogenous DNA damage, we measured the activation of NF-κB in WT and progeroid model mice. As both WT and progeroid mice aged, NF-κB was activated stochastically in a variety of cell types. Genetic depletion of one allele of the p65 subunit of NF-κB or treatment with a pharmacological inhibitor of the NF-κB–activating kinase, IKK, delayed the age-related symptoms and pathologies of progeroid mice. Additionally, inhibition of NF-κB reduced oxidative DNA damage and stress and delayed cellular senescence. These results indicate that the mechanism by which DNA damage drives aging is due in part to NF-κB activation. IKK/NF-κB inhibitors are sufficient to attenuate this damage and could provide clinical benefit for degenerative changes associated with accelerated aging disorders and normal aging. PMID:22706308
KB-R7943 reduces 4-aminopyridine-induced epileptiform activity in adult rats after neuronal damage induced by neonatal monosodium glutamate treatment.

PubMed

Hernandez-Ojeda, Mariana; Ureña-Guerrero, Monica E; Gutierrez-Barajas, Paola E; Cardenas-Castillo, Jazmin A; Camins, Antoni; Beas-Zarate, Carlos

2017-05-09

Neonatal monosodium glutamate (MSG) treatment triggers excitotoxicity and induces a degenerative process that affects several brain regions in a way that could lead to epileptogenesis. Na + /Ca 2+ exchangers (NCX1-3) are implicated in Ca 2+ brain homeostasis; normally, they extrude Ca 2+ to control cell inflammation, but after damage and in epilepsy, they introduce Ca 2+ by acting in the reverse mode, amplifying the damage. Changes in NCX3 expression in the hippocampus have been reported immediately after neonatal MSG treatment. In this study, the expression level of NCX1-3 in the entorhinal cortex (EC) and hippocampus (Hp); and the effects of blockade of NCXs on the seizures induced by 4-Aminopyridine (4-AP) were analysed in adult rats after neonatal MSG treatment. KB-R7943 was applied as NCXs blocker, but is more selective to NCX3 in reverse mode. Neonatal MSG treatment was applied to newborn male rats at postnatal days (PD) 1, 3, 5, and 7 (4 g/kg of body weight, s.c.). Western blot analysis was performed on total protein extracts from the EC and Hp to estimate the expression level of NCX1-3 proteins in relative way to the expression of β-actin, as constitutive protein. Electrographic activity of the EC and Hp were acquired before and after intracerebroventricular (i.c.v.) infusion of 4-AP (3 nmol) and KB-R7943 (62.5 pmol), alone or in combination. All experiments were performed at PD60. Behavioural alterations were also recorder. Neonatal MSG treatment significantly increased the expression of NCX3 protein in both studied regions, and NCX1 protein only in the EC. The 4-AP-induced epileptiform activity was significantly higher in MSG-treated rats than in controls, and KB-R7943 co-administered with 4-AP reduced the epileptiform activity in more prominent way in MSG-treated rats than in controls. The long-term effects of neonatal MSG treatment include increases on functional expression of NCXs (mainly of NCX3) in the EC and Hp, which seems to contribute to
[Isolation and purification of recombinant human lactoferrin (rhLF) from transgenic rice and its antibacterial activities].

PubMed

Wu, Jinghuan; Yang, Lichen; Liu, Gaige; Gong, Zhaolong; Liu, Jikai; Hu, Yujie; Guo, Yunchang; Piao, Jianhua; Shen, Zhicheng; Yang, Xiaoguang

2013-05-01

To study the isolation and purification of recombinant human lactoferrin (rhLF) from transgenic rice, and to check its antibacterial activities. After isolated rhLF from transgenic rice via saturated ammonium sulfate precipitation, then purified it through CM Sepharose FF-exchange chromatography and molecular sieve chromatography Sephadex G25. The inhibition effects under different concentrations of rhLF (0.25, 0.5, 1.0, 2.0, 4.0, and 5.0 mg/ml) against Salmonella typhimurium, Staphyloccocus aureus, Bacillus cereus, Listeria monocytogenes were observed, using broth microdilution method. The rhLF was obtained at a higher purity (about 90%) through successful isolation and purification. After Coomassie blue staining, Westernblot and mass spectrometer analysis, it was identified as the purpose protein with the molecular weight of approximately 79 kDa. The antibacterial experiments showed that 5 mg/ml and 4 mg/ml rhLF could inhibite Salmonella typhimurium and Staphylococcus aureus persistently, 2 mg/ml and 1 mg/ml rhLF showed a significant inhibitory effects in the later period; while 0.5 mg/ml or lower concentration, showed no inhibitory effects. As to Bacillus cereus, only 5 mg/ml and 4 mg/ml rhLF exhibited certain inhibitory effects within 18 hours. Listeria monocytogenes was inhibited within 18 hours just at 5 mg/ml rhLF. The rhLF could be successfully separated and purified from transgenic rice, and the purified protein still has significant antibacterial activities.
Effect of Flash-Heat Treatment on Antimicrobial Activity of Breastmilk

PubMed Central

Wiedeman, Jean; Buehring, Gertrude; Peerson, Janet M.; Hayfron, Kweku; K'Aluoch, Okumu; Lonnerdal, Bo; Israel-Ballard, Kiersten; Coutsoudis, Anna; Abrams, Barbara

2011-01-01

Abstract Background and Objectives The World Health Organization recommends human immunodeficiency virus (HIV)-positive mothers in resource-poor regions heat-treat expressed breastmilk during periods of increased maternal-to-child transmission risk. Flash-heat, a “low tech” pasteurization method, inactivates HIV, but effects on milk protein bioactivity are unknown. The objectives were to measure flash-heat's effect on antimicrobial properties of lactoferrin, lysozyme, and whole milk and on the digestive resistance of lactoferrin and lysozyme. Methods Flash-heated and unheated breastmilk aliquots from HIV-positive mothers in South Africa were “spiked” with Staphylococcus aureus and Escherichia coli and then cultured for 0, 3, and 6 hours. Lysozyme and lactoferrin activities were determined by lysis of Micrococcus luteus cells and inhibition of enteropathogenic E. coli, respectively, measured spectrophotometrically. Percentages of proteins surviving in vitro digestion, lactoferrin and lysozyme activity, and bacteriostatic activity of whole milk in heated versus unheated samples were compared. Results There was no difference in rate of growth of E. coli or S. aureus in flash-heated versus unheated whole milk (p = 0.61 and p = 0.96, respectively). Mean (95% confidence interval) antibacterial activity of lactoferrin was diminished 11.1% (7.8%, 14.3%) and that of lysozyme by up to 56.6% (47.1%, 64.5%) by flash-heat. Digestion of lysozyme was unaffected (p = 0.12), but 25.4% less lactoferrin survived digestion (p < 0.0001). Conclusions In summary, flash-heat resulted in minimally decreased lactoferrin and moderately decreased lysozyme bioactivity, but bacteriostatic activity of whole milk against representative bacteria was unaffected. This suggests flash-heated breastmilk likely has a similar profile of resistance to bacterial contamination as that of unheated milk. Clinical significance of the decreased bioactivity should be tested in clinical
Effect of flash-heat treatment on antimicrobial activity of breastmilk.

PubMed

Chantry, Caroline J; Wiedeman, Jean; Buehring, Gertrude; Peerson, Janet M; Hayfron, Kweku; K'Aluoch, Okumu; Lonnerdal, Bo; Israel-Ballard, Kiersten; Coutsoudis, Anna; Abrams, Barbara

2011-06-01

The World Health Organization recommends human immunodeficiency virus (HIV)-positive mothers in resource-poor regions heat-treat expressed breastmilk during periods of increased maternal-to-child transmission risk. Flash-heat, a "low tech" pasteurization method, inactivates HIV, but effects on milk protein bioactivity are unknown. The objectives were to measure flash-heat's effect on antimicrobial properties of lactoferrin, lysozyme, and whole milk and on the digestive resistance of lactoferrin and lysozyme. Flash-heated and unheated breastmilk aliquots from HIV-positive mothers in South Africa were "spiked" with Staphylococcus aureus and Escherichia coli and then cultured for 0, 3, and 6 hours. Lysozyme and lactoferrin activities were determined by lysis of Micrococcus luteus cells and inhibition of enteropathogenic E. coli, respectively, measured spectrophotometrically. Percentages of proteins surviving in vitro digestion, lactoferrin and lysozyme activity, and bacteriostatic activity of whole milk in heated versus unheated samples were compared. There was no difference in rate of growth of E. coli or S. aureus in flash-heated versus unheated whole milk (p = 0.61 and p = 0.96, respectively). Mean (95% confidence interval) antibacterial activity of lactoferrin was diminished 11.1% (7.8%, 14.3%) and that of lysozyme by up to 56.6% (47.1%, 64.5%) by flash-heat. Digestion of lysozyme was unaffected (p = 0.12), but 25.4% less lactoferrin survived digestion (p < 0.0001). In summary, flash-heat resulted in minimally decreased lactoferrin and moderately decreased lysozyme bioactivity, but bacteriostatic activity of whole milk against representative bacteria was unaffected. This suggests flash-heated breastmilk likely has a similar profile of resistance to bacterial contamination as that of unheated milk. Clinical significance of the decreased bioactivity should be tested in clinical trials.
NF-κB gene signature predicts prostate cancer progression

PubMed Central

Jin, Renjie; Yi, Yajun; Yull, Fiona E.; Blackwell, Timothy S.; Clark, Peter E.; Koyama, Tatsuki; Smith, Joseph A.; Matusik, Robert J.

2014-01-01

In many prostate cancer (PCa) patients, the cancer will be recurrent and eventually progress to lethal metastatic disease after primary treatment, such as surgery or radiation therapy. Therefore, it would be beneficial to better predict which patients with early-stage PCa would progress or recur after primary definitive treatment. In addition, many studies indicate that activation of NF-κB signaling correlates with PCa progression; however, the precise underlying mechanism is not fully understood. Our studies show that activation of NF-κB signaling via deletion of one allele of its inhibitor, IκBα, did not induce prostatic tumorigenesis in our mouse model. However, activation of NF-κB signaling did increase the rate of tumor progression in the Hi-Myc mouse PCa model when compared to Hi-Myc alone. Using the non-malignant NF-κB activated androgen depleted mouse prostate, a NF-κB Activated Recurrence Predictor 21 (NARP21) gene signature was generated. The NARP21 signature successfully predicted disease-specific survival and distant metastases-free survival in patients with PCa. This transgenic mouse model derived gene signature provides a useful and unique molecular profile for human PCa prognosis, which could be used on a prostatic biopsy to predict indolent versus aggressive behavior of the cancer after surgery. PMID:24686169
The disintegrin, trimucrin, suppresses LPS-induced activation of phagocytes primarily through blockade of NF-κB and MAPK activation.

PubMed

Hung, Yu-Chun; Hsu, Chun-Chieh; Chung, Ching-Hu; Huang, Tur-Fu

2016-07-01

In addition to antiplatelet activity, disintegrin, a small-mass RGD-containing polypeptide, has been shown to exert anti-inflammatory effects but the mechanism involved remains unclear. In this study, we report that trimucrin, a disintegrin from the venom of Trimeresurus mucrosquamatus, inhibits lipopolysaccharide (LPS)-induced stimulation of THP-1 and RAW 264.7 cells. We also investigate the underlying mechanism. Trimucrin decreased the release of proinflammatory cytokines including tumor necrosis factor α (TNFα), interleukin-6 (IL-6), nitric oxide, and reactive oxygen species (ROS), and inhibited the adhesion and migration of LPS-activated phagocytes. Trimucrin significantly blocked the expression of nuclear factor kappaB (NF-κB)-related downstream inducible enzymes such as inducible nitric oxide synthase (iNOS) and COX-2. In addition, its anti-inflammatory effect was associated with the decreased mitogen-activated protein kinase (MAPK) phosphorylation. Furthermore, trimucrin concentration dependently inhibited LPS-induced phosphorylation of focal adhesion kinase (FAK), PI3K, and Akt. Trimucrin also reversed the DNA-binding activity of NF-κB by suppressing the LPS-induced nuclear translocation of p65 and the cytosolic IκB release. Flow cytometric analyses showed that trimucrin bound to cells in a concentration-dependent manner. The anti-αVβ3 mAb also specifically decreased the binding of fluorescein isothiocyanate (FITC)-conjugated trimucrin. Binding assays demonstrated that integrin αVβ3 was the binding site for trimucrin on THP-1 and RAW 264.7 cells. In conclusion, we showed that trimucrin decreases the inflammatory reaction through the attenuation of iNOS expression and nitric oxide (NO) production by blocking MAP kinase and the NF-κB activation in LPS-stimulated THP-1 and RAW 264.7 cells.
Rosiglitazone attenuates NF-{kappa}B-dependent ICAM-1 and TNF-{alpha} production caused by homocysteine via inhibiting ERK{sub 1/2}/p38MAPK activation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Bai, Yong-Ping; Liu, Yu-Hui; Chen, Jia

2007-08-17

Previous studies demonstrated an important interaction between nuclear factor-kappaB (NF-{kappa}B) activation and homocysteine (Hcy)-induced cytokines expression in endothelial cells and vascular smooth muscle cells. However, the underlying mechanism remains illusive. In this study, we investigated the effects of Hcy on NF-{kappa}B-mediated sICAM-1, TNF-{alpha} production and the possible involvement of ERK{sub 1/2}/p38MAPK pathway. The effects of rosiglitazone intervention were also examined. Our results show that Hcy increased the levels of sICAM-1 and TNF-{alpha} in cultured human umbilical vein endothelial cells (HUVECs) in a time- and concentration-dependent manner. This effect was significantly depressed by rosiglitazone and different inhibitors (PDTC, NF-{kappa}B inhibitor; PD98059,more » MEK inhibitor; SB203580, p38MAPK specific inhibitor; and staurosporine, PKC inhibitor). Next, we investigated the effect of Hcy on ERK{sub 1/2}/p38MAPK pathway and NF-{kappa}B activity in HUVECs. The results show that Hcy activated both ERK{sub 1/2}/p38MAPK pathway and NF-{kappa}B-DNA-binding activity. These effects were markedly inhibited by rosiglitazone as well as other inhibitors (SB203580, PD98059, and PDTC). Further, the pretreatment of staurosporine abrogated ERK{sub 1/2}/p38MAPK phosphorylation, suggesting that Hcy-induced ERK{sub 1/2}/p38MAPK activation is associated with PKC activity. Our results provide evidence that Hcy-induced NF-{kappa}B activation was mediated by activation of ERK{sub 1/2}/p38MAPK pathway involving PKC activity. Rosiglitazone reduces the NF-{kappa}B-mediated sICAM-1 and TNF-{alpha} production induced by Hcy via inhibition of ERK{sub 1/2}/p38MAPK pa0011thw.« less
Artesunate protects pancreatic beta cells against cytokine-induced damage via SIRT1 inhibiting NF-κB activation.

PubMed

Yu, L; Chen, J F; Shuai, X; Xu, Y; Ding, Y; Zhang, J; Yang, W; Liang, X; Su, D; Yan, C

2016-01-01

Artesunate (ART) has been known as the most effective and safe reagents to treat malaria for many years. In this study, we explored whether ART could protect pancreatic beta-cell against cytokine-induced damage. The production of nitrite (NO) was detected with the Griess Assay Kit. SIRT1 and inducible nitric oxide synthase (iNOS) expression were determined with Western blot. The transcriptional activity of NF-κB was evaluated by luciferase reporter assay. The expression of Sirt1 was silenced by RNA interference. Glucose-stimulated insulin secretion (GSIS) and potassium-stimulated insulin secretion (KSIS) assays were performed to measure the effect of ART on pancreatic beta-cells' function. The effect of ART on beta-cells apoptosis was evaluated by using Hochest/PI staining and TUNEL assay. ART enhanced GSIS (KSIS) and reduced apoptosis of pancreatic beta-cells induced by IL-1β. Further study showed that ART inhibited IL-1β-induced increase of NF-κB activity, iNOS expression, and NO production. Moreover, ART up-regulated SIRT1 expression in INS-1 cells and islets exposed to IL-1β. Inhibition of SIRT1 expression could partially abolished the inhibitory effect of ART on NF-κB activity in IL-1β-treated beta-cells. More importantly, the protective effect of ART on cytokine-induced damage was reversed by silencing SIRT1 expression. ART can elicit a protective effect on beta-cells exposed to IL-1β by stimulating SIRT1 expression, which resulted in the decrease of NF-κB activity, iNOS expression, and NO production. Hence, ART might be an effective drug for diabetes.
NF-kappaB Activity in Macrophages Determines Metastatic Potential of Breast Tumor Cells

DTIC Science & Technology

2011-08-01

Cheng DS, Chodosh LA, Blackwell TS, Yull FE: Activation of nuclear factor kappa B in mammary epithelium promotes milk loss during mammary... microbial products (15, 16). To date, the potential role of macrophages in the fetal lung innate immune response has not been closely examined. Studies...In this model, microbial products initially activate NF-kB in lung macrophages. The release of inflammatory mediators, particularly IL-1b and/or TNF-a
Growth characteristics of Lactobacillus brevis KB290 in the presence of bile.

PubMed

Kimoto-Nira, Hiromi; Suzuki, Shigenori; Suganuma, Hiroyuki; Moriya, Naoko; Suzuki, Chise

2015-10-01

Live Lactobacillus brevis KB290 have several probiotic activities, including immune stimulation and modulation of intestinal microbial balance. We investigated the adaptation of L. brevis KB290 to bile as a mechanism of intestinal survival. Strain KB290 was grown for 5 days at 37 °C in tryptone-yeast extract-glucose (TYG) broth supplemented with 0.5% sodium acetate (TYGA) containing 0.15%, 0.3%, or 0.5% bile. Growth was determined by absorbance at 620 nm or by dry weight. Growth was enhanced as the broth's bile concentration increased. Bile-enhanced growth was not observed in TYG broth or with xylose or fructose as the carbon source, although strain KB290 could assimilate these sugars. Compared with cells grown without bile, cells grown with bile had twice the cell yield (dry weight) and higher hydrophobicity, which may improve epithelial adhesion. Metabolite analysis revealed that bile induced more lactate production by glycolysis, thus enhancing growth efficiency. Scanning electron microscopy revealed that cells cultured without bile for 5 days in TYGA broth had a shortened rod shape and showed lysis and aggregation, unlike cells cultured for 1 day; cells grown with bile for 5 days had an intact rod shape and rarely appeared damaged. Cellular material leakage through autolysis was lower in the presence of bile than in its absence. Thus lysis of strain KB290 cells cultured for extended periods was suppressed in the presence of bile. This study provides new role of bile and sodium acetate for retaining an intact cell shape and enhancing cell yield, which are beneficial for intestinal survival. Copyright © 2015 Elsevier Ltd. All rights reserved.
CARMA3 is overexpressed in colon cancer and regulates NF-{kappa}B activity and cyclin D1 expression

DOE Office of Scientific and Technical Information (OSTI.GOV)

Miao, Zhifeng; Zhao, Tingting; Wang, Zhenning

2012-09-07

Highlights: Black-Right-Pointing-Pointer CARMA3 expression is elevated in colon cancers. Black-Right-Pointing-Pointer CARMA3 promotes proliferation and cell cycle progression in colon cancer cells. Black-Right-Pointing-Pointer CARMA3 upregulates cyclinD1 through NF-{kappa}B activation. -- Abstract: CARMA3 was recently reported to be overexpressed in cancers and associated with the malignant behavior of cancer cells. However, the expression of CARMA3 and its biological roles in colon cancer have not been reported. In the present study, we analyzed the expression pattern of CARMA3 in colon cancer tissues and found that CARMA3 was overexpressed in 30.8% of colon cancer specimens. There was a significant association between CARMA3 overexpression andmore » TNM stage (p = 0.0383), lymph node metastasis (p = 0.0091) and Ki67 proliferation index (p = 0.0035). Furthermore, knockdown of CARMA3 expression in HT29 and HCT116 cells with high endogenous expression decreased cell proliferation and cell cycle progression while overexpression of CARMA3 in LoVo cell line promoted cell proliferation and facilitated cell cycle transition. Further analysis showed that CARMA3 knockdown downregulated and its overexpression upregulated cyclin D1 expression and phospho-Rb levels. In addition, we found that CARMA3 depletion inhibited p-I{kappa}B levels and NF-{kappa}B activity and its overexpression increased p-I{kappa}B expression and NF-{kappa}B activity. NF-{kappa}B inhibitor BAY 11-7082 reversed the role of CARMA3 on cyclin D1 upregulation. In conclusion, our study found that CARMA3 is overexpressed in colon cancers and contributes to malignant cell growth by facilitating cell cycle progression through NF-{kappa}B mediated upregulation of cyclin D1.« less
Antiinflammatory effects of glucocorticoids in brain cells, independent of NF-kappa B.

PubMed

Bourke, E; Moynagh, P N

1999-08-15

Glucocorticoids are potent antiinflammatory drugs. They inhibit the expression of proinflammatory cytokines and adhesion molecules. It has recently been proposed that the underlying basis to such inhibition is the induction of the protein I kappa B, which inhibits the transcription factor NF-kappa B. The latter is a key activator of the genes encoding cytokines and adhesion molecules. The present study shows that the synthetic glucocorticoid, dexamethasone, inhibits the induction of the proinflammatory cytokine IL-8 and the adhesion molecules VCAM-1 and ICAM-1 in human 1321N1 astrocytoma and SK.N.SH neuroblastoma cells. However, dexamethasone failed to induce I kappa B or inhibit activation of NF-kappa B by IL-1 in the two cell types. EMSA confirmed the identity of the activated NF-kappa B by demonstrating that an oligonucleotide, containing the wild-type NF-kappa B-binding motif, inhibited formation of the NF-kappa B-DNA complexes whereas a mutated form of the NF-kappa B-binding motif was ineffective. In addition, supershift analysis showed that the protein subunits p50 and p65 were prevalent components in the activated NF-kappa B complexes. The lack of effect of dexamethasone on the capacity of IL-1 to activate NF-kappa B correlated with its inability to induce I kappa B and the ability of IL-1 to cause degradation of I kappa B, even in the presence of dexamethasone. The results presented in this paper strongly suggest that glucocorticoids may exert antiinflammatory effects in cells of neural origin by a mechanism(s) independent of NF-kappa B.
Differential IKK/NF-κB Activity is Mediated by TSC2 through mTORC1 in PTEN-null Prostate Cancer and Tuberous Sclerosis Complex Tumor Cells

PubMed Central

Gao, Yu; Gartenhaus, Ronald B.; Lapidus, Rena G.; Hussain, Arif; Zhang, Yanting; Wang, Xinghuan; Dan, Han C.

2015-01-01

The serine/threonine protein kinase Akt plays a critical role in regulating proliferation, growth and survival through phosphorylation of different downstream substrates. The mammalian target of rapamycin (mTOR) is a key target for Akt to promote tumorigenesis. It has been reported that Akt activates mTOR through phosphorylation and inhibition of the tuberous sclerosis complex (TSC) protein TSC2. Previously it was demonstrated that mTOR activates IKK/NF-κB signaling by promoting IKK activity downstream of Akt in conditions deficient of PTEN. In the current study, the mechanistic role of the tumor suppressor TSC2 was investigated in the regulation of IKK/NF-κB activity in PTEN-null prostate cancer and in TSC2 mutated tumor cells. The results demonstrate that TSC2 inhibits IKK/NF-κB activity downstream of Akt and upstream of mTORC1 in a PTEN deficient environment. However, TSC2 promotes IKK/NF-κB activity upstream of Akt and mTORC1 in TSC2 mutated tumor cells. These data indicate that TSC2 negatively or positively regulates IKK/NF-κB activity in a context-dependent manner depending on the genetic background. PMID:26374334
From the Cover: Activation of NF-κB-Autophagy Axis by 2-Hydroxyethyl Methacrylate Commits Dental Mesenchymal Cells to Apoptosis.

PubMed

Yu, Jing-Jing; Zhu, Ling-Xin; Zhang, Jie; Liu, Shan; Lv, Feng-Yuan; Cheng, Xue; Liu, Guo-Jing; Peng, Bin

2017-05-01

2-hydroxyethyl methacrylate (HEMA) is the major resin monomer that is released from incomplete polymerized dental restorative and adhesive biomaterials during dental therapy. Autophagy and apoptosis are biologically connected and the relationship between autophagy and apoptosis is complex under various circumstances. This study aimed to determine whether autophagy is activated by HEMA and further explore the function of autophagy during the HEMA-induced apoptosis of dental mesenchymal cells (DMCs). We exposed DMCs to different concentrations of HEMA. Cell viability showed a time- and concentration-dependent decrease when exposed to HEMA. We showed that HEMA exposure increased autophagic vacuoles and the expression of autophagic biomarkers (Beclin1, Atg5 and LC3). Pre-incubated with autophagy inhibitors (3-methyladenine and chloroquine) significantly prevented HEMA-induced apoptosis. Interestingly, HEMA initiated nuclear factor-κB (NF-κB) expression and nuclear translocation, whereas the NF-κB inhibitor (Bay 11-7082) markedly suppressed HEMA-induced autophagic activation and apoptosis. As is consistent with the in vitro results, HEMA treatment resulted in dental pulp tissue toxicity and activation of typical autophagic vacuoles in the tooth slice organ culture model ex vivo. In summary, we demonstrated that NF-κB signaling functioned upstream of HEMA-inducecd autophagy in DMCs and that the activation of NF-κB-autophagy axis was responsible for HEMA-induced apoptosis. Our findings provide novel insights into the mechanisms of resin monomer-mediated dental pulp damage during dental treatment, highlighting the activation of NF-κB-autophagy axis as an important mechanism of HEMA-mediated apoptosis. © The Author 2017. Published by Oxford University Press on behalf of the Society of Toxicology. All rights reserved. For Permissions, please e-mail: journals.permissions@oup.com.

[Effects of acupuncture on lactoferrin content in tears and tear secretion in patients suffering from dry eyes: a randomized controlled trial].

PubMed

Shi, Jing-lin; Miao, Wan-hong

2012-09-01

With the understanding of the immune inflammatory response in the pathogenesis of dry eyes, and the limitations of widely used artificial tears and numerous pharmaceuticals and methods to promote tear secretion, clinicians pay more attention to the therapies that can promote tear secretion actively. Acupuncture treatment for dry eye may meet this requirement. To observe the clinical efficacy of acupuncture treatment on dry eye and the effects on duration, and to examine the mechanisms of acupuncture in treating patients with dye eyes. The study was performed at Department of Ophthalmology, Shuguang Hospital, Shanghai University of Traditional Chinese Medicine from August 2010 to May 2011. Patients with the primary diagnosis of dry eye were enrolled. Sixty-five patients were randomly divided into treatment group and control group, and were given 3 weeks of acupuncture treatment or artificial tear therapy respectively. The enzyme-linked immunosorbent assay was used to detect the lactoferrin content of the tears before and after treatment. In order to evaluate the efficacy of the treatment methods, the Schirmer I test and break-up time were also measured. Compared with before treatment, the lactoferrin content in the tears of patients in the treatment group increased, break-up time was prolonged and the result of the Schirmer I test showed improvement after 3 weeks of treatment. The indexes mentioned above did not change in the control group after treatment. There were no significant differences in tear lactoferrin and Schirmer I test between one week after treatment and after 3-week treatment in the treatment group, but break-up time was significantly shortened. The result of Schirmer I test in the treatment group was significantly higher than that in the control group one week after treatment. Acupuncture can increase tear lactoferrin level, extend tear film break-up time and promote tear secretion in patients with dry eye in a time-limited trial. With the end of
NF-{kappa}B signaling is activated and confers resistance to apoptosis in three-dimensionally cultured EGFR-mutant lung adenocarcinoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Sakuma, Yuji, E-mail: ysakuma@gancen.asahi.yokohama.jp; Yamazaki, Yukiko; Nakamura, Yoshiyasu

2012-07-13

Highlights: Black-Right-Pointing-Pointer EGFR-mutant cells in 3D culture resist EGFR inhibition compared with suspended cells. Black-Right-Pointing-Pointer Degradation of I{kappa}B and activation of NF-{kappa}B are observed in 3D-cultured cells. Black-Right-Pointing-Pointer Inhibiting NF-{kappa}B enhances the efficacy of the EGFR inhibitor in 3D-cultured cells. -- Abstract: Epidermal growth factor receptor (EGFR)-mutant lung adenocarcinoma cells in suspension undergo apoptosis to a greater extent than adherent cells in a monolayer when EGFR autophosphorylation is inhibited by EGFR tyrosine kinase inhibitors (TKIs). This suggests that cell adhesion to a culture dish may activate an anti-apoptotic signaling pathway other than the EGFR pathway. Since the microenvironment of cellsmore » cultured in a monolayer are substantially different to that of cells existing in three-dimension (3D) in vivo, we assessed whether two EGFR-mutant lung adenocarcinoma cell lines, HCC827 and H1975, were more resistant to EGFR TKI-induced apoptosis when cultured in a 3D extracellular matrix (ECM) as compared with in suspension. The ECM-adherent EGFR-mutant cells in 3D were significantly less sensitive to treatment with WZ4002, an EGFR TKI, than the suspended cells. Further, a marked degradation of I{kappa}B{alpha}, the inhibitor of nuclear factor (NF)-{kappa}B, was observed only in the 3D-cultured cells, leading to an increase in the activation of NF-{kappa}B. Moreover, the inhibition of NF-{kappa}B with pharmacological inhibitors enhanced EGFR TKI-induced apoptosis in 3D-cultured EGFR-mutant cells. These results suggest that inhibition of NF-{kappa}B signaling would render ECM-adherent EGFR-mutant lung adenocarcinoma cells in vivo more susceptible to EGFR TKI-induced cell death.« less
NF-κB activation primes cells to a pro-inflammatory polarized response to a TLR7 agonist

PubMed Central

Lee, Jongdae; Hayashi, Masaaki; Lo, Jeng-Fan; Fearns, Colleen; Chu, Wen-Ming; Luo, Yunping; Xiang, Rong; Chuang, Tsung-Hsien

2009-01-01

Toll-like receptor 7 (TLR7) mediates anti-viral immunity by recognizing ssRNA viruses. Small molecular weight TLR7 agonists have been approved, or are being evaluated, for treatment of cancers or infectious diseases. Although TLR7 is predominantly expressed in a restricted set of immune cell types including plasmacytoid dendritic cells (pDCs), it is also expressed in non-native expressing cells (e.g., hepatocytes) under certain circumstances. To elucidate the molecular basis of TLR7 induction by pro-inflammatory stimulation and the subsequent cellular responses in these non-native TLR7-expressing cell types, we firstly cloned and characterized the 5′-promoter region of TLR7. The proximal region of this promoter drives the transcription of the TLR7 gene. Pro-inflammatory stimuli activated TLR7 transcription via a NF-κB binding motif in this region, and this activation could be blocked by mutation of the NF-κB binding site or addition of NF-κB inhibitors. Further studies showed that pretreatment of the Hep3B hepatocytes with TNF-α or IL-1 rendered them responsive to TLR7 activation by a TLR7 agonist. However, distinct from TLR7 activation in pDCs, which respond to stimulation with Th1 polarized cytokine production, TLR7 induction by pro-inflammatory signals in hepatocytes reconstitutes the NF-κB-dependent cascade but not the IRF7-dependent cascade, resulting in a pro-inflammatory polarized response rather than a Th1 polarized response. These results indicate that inflammatory stimulation is capable of priming cells to respond to TLR7 agonist with an immune response that differs from that in native TLR7-expressing cells. PMID:19426145
FGF-1-induced matrix metalloproteinase-9 expression in breast cancer cells is mediated by increased activities of NF-kappaB and activating protein-1.

PubMed

Lungu, Gina; Covaleda, Lina; Mendes, Odete; Martini-Stoica, Heidi; Stoica, George

2008-06-01

Matrix metalloproteinase-9 (MMP-9) plays a critical role in tumor invasion and metastasis. Here, we investigate the effect of fibroblast growth factor-1 (FGF-1) on the expression of MMP-9 in ENU1564, an ethyl-N-nitrosourea-induced rat mammary adenocarcinoma cell line. We observed that FGF-1 induces a dose-dependent increase in MMP-9 mRNA, protein, and activity in ENU1564 cells. To gain insight into the molecular mechanism of MMP-9 regulation by FGF-1, we investigated the role of components of PI3K-Akt and MEK1/2-ERK signaling pathways in our system since NF-kappaB and AP-1 transcription factor binding sites have been characterized in the upstream region of the MMP-9 gene. We demonstrated that FGF-1 increases Akt phosphorylation, triggers nuclear translocation of NF-kappaBp65, and enhances degradation of cytoplasmic IkappaBalpha. Pretreatment of cells with LY294002, a PI3K inhibitor, significantly inhibited MMP-9 protein expression in FGF-1-treated cells. Conversely, our data show that FGF-1 increases ERK phosphorylation in ENU1564 cells, increases c-jun and c-fos mRNA expression in a time-dependent manner, and triggers nuclear translocation of c-jun. Pretreatment of cells with PD98059, a MEK1/2 inhibitor significantly inhibited MMP-9 protein expression in FGF-1 treated cells. Finally, we observed increased DNA binding of NF-kappaB and AP-1 in FGF-1-treated cells and that mutation of either NF-kappaB or AP-1 response elements prevented MMP-9 promoter activation by FGF-1. Taken together, these results demonstrated that FGF-1-induced MMP-9 expression in ENU1564 cells is associated with increasing DNA binding activities of NF-kappaB and AP-1 and involve activation of a dual signaling pathway, PI3K-Akt and MEK1/2-ERK. (c) 2007 Wiley-Liss, Inc.
Thymol inhibits Staphylococcus aureus internalization into bovine mammary epithelial cells by inhibiting NF-κB activation.

PubMed

Wei, Zhengkai; Zhou, Ershun; Guo, Changming; Fu, Yunhe; Yu, Yuqiang; Li, Yimeng; Yao, Minjun; Zhang, Naisheng; Yang, Zhengtao

2014-01-01

Bovine mastitis is one of the most costly and prevalent diseases in the dairy industry and is characterised by inflammatory and infectious processes. Staphylococcus aureus (S. aureus), a Gram-positive organism, is a frequent cause of subclinical, chronic mastitis. Thymol, a monocyclic monoterpene compound isolated from Thymus vulgaris, has been reported to have antibacterial properties. However, the effect of thymol on S. aureus internalization into bovine mammary epithelial cells (bMEC) has not been investigated. In this study, we evaluated the effect of thymol on S. aureus internalization into bMEC, the expression of tracheal antimicrobial peptide (TAP) and β-defensin (BNBD5), and the inhibition of NF-κB activation in bMEC infected with S. aureus. Our results showed that thymol (16-64 μg/ml) could reduce the internalization of S. aureus into bMEC and down-regulate the mRNA expression of TAP and BNBD5 in bMEC infected with S. aureus. In addition, thymol was found to inhibit S. aureus-induced nitric oxide (NO) production in bMEC and suppress S. aureus-induced NF-κB activation in a dose-dependent manner. In conclusion, these results indicated that thymol inhibits S. aureus internalization into bMEC by inhibiting NF-κB activation. Copyright © 2014 Elsevier Ltd. All rights reserved.
Human T-Cell Leukemia Virus Type 1 Tax Induction of NF-κB Involves Activation of the IκB Kinase α (IKKα) and IKKβ Cellular Kinases

PubMed Central

Geleziunas, Romas; Ferrell, Sharon; Lin, Xin; Mu, Yajun; Cunningham, Emmett T.; Grant, Mark; Connelly, Margery A.; Hambor, John E.; Marcu, Kenneth B.; Greene, Warner C.

1998-01-01

Tax corresponds to a 40-kDa transforming protein from the pathogenic retrovirus human T-cell leukemia virus type 1 (HTLV-1) that activates nuclear expression of the NF-κB/Rel family of transcription factors by an unknown mechanism. Tax expression promotes N-terminal phosphorylation and degradation of IκBα, a principal cytoplasmic inhibitor of NF-κB. Our studies now demonstrate that HTLV-1 Tax activates the recently identified cellular kinases IκB kinase α (IKKα) and IKKβ, which normally phosphorylate IκBα on both of its N-terminal regulatory serines in response to tumor necrosis factor alpha (TNF-α) and interleukin-1 (IL-1) stimulation. In contrast, a mutant of Tax termed M22, which does not induce NF-κB, fails to activate either IKKα or IKKβ. Furthermore, endogenous IKK enzymatic activity was significantly elevated in HTLV-1-infected and Tax-expressing T-cell lines. Transfection of kinase-deficient mutants of IKKα and IKKβ into either human Jurkat T or 293 cells also inhibits NF-κB-dependent reporter gene expression induced by Tax. Similarly, a kinase-deficient mutant of NIK (NF-κB-inducing kinase), which represents an upstream kinase in the TNF-α and IL-1 signaling pathways leading to IKKα and IKKβ activation, blocks Tax induction of NF-κB. However, plasma membrane-proximal elements in these proinflammatory cytokine pathways are apparently not involved since dominant negative mutants of the TRAF2 and TRAF6 adaptors, which effectively block signaling through the cytoplasmic tails of the TNF-α and IL-1 receptors, respectively, do not inhibit Tax induction of NF-κB. Together, these studies demonstrate that HTLV-1 Tax exploits a distal part of the proinflammatory cytokine signaling cascade leading to induction of NF-κB. The pathological alteration of this cytokine pathway leading to NF-κB activation by Tax may play a central role in HTLV-1-mediated transformation of human T cells, clinically manifested as the adult T-cell leukemia. PMID
The cachectic mediator proteolysis inducing factor activates NF-kappaB and STAT3 in human Kupffer cells and monocytes.

PubMed

Watchorn, Tammy M; Dowidar, Nabil; Dejong, Cornelis H C; Waddell, Ian D; Garden, O James; Ross, James A

2005-10-01

A novel proteoglycan, proteolysis inducing factor (PIF), is capable of inducing muscle proteolysis during the process of cancer cachexia, and of inducing an acute phase response in human hepatocytes. We investigated whether PIF is able to activate pro-inflammatory pathways in human Kupffer cells, the resident macrophages of the liver, and in monocytes, resulting in the production of pro-inflammatory cytokines. Normal liver tissue was obtained from patients undergoing partial hepatectomy and Kupffer cells were isolated. Monocytes were isolated from peripheral blood. Following exposure to native PIF, pro-inflammatory cytokine production from Kupffer cells and monocytes was measured and the NF-kappaB and STAT3 transcriptional pathways were investigated using electrophoretic mobility shift assays. We demonstrate that PIF is able to activate the transcription factor NF-kappaB and NF-kappaB-inducible genes in human Kupffer cells, and in monocytes, resulting in the production of pro-inflammatory cytokines such as TNF-alpha, IL-8 and IL-6. PIF enhances the expression of the cell surface molecules LFA-1 and CD14 on macrophages. PIF also activates the transcription factor STAT3 in Kupffer cells. The pro-inflammatory effects of PIF, mediated via NF-kappaB and STAT3, are important in macrophage behaviour and may contribute to the inflammatory pro-cachectic process in the liver.
A20 Functional Domains Regulate Subcellular Localization and NF-Kappa B Activation

DTIC Science & Technology

2013-08-15

that the first function to be described for A20 was that of an anti -apoptotic protein (55). They based their choice of experiments and preliminary...mediated apoptosis (55). After positive selection of the resulting clones with neomycin and verification of A20 expression, they compared the...Karposi sarcoma herpesvirus (KSHV) mediated cell transformation (72). K13 can directly activate NF-κB by interacting with the IKK complex and is
NF-kappaB mediates FGF signal regulation of msx-1 expression.

PubMed

Bushdid, P B; Chen, C L; Brantley, D M; Yull, F; Raghow, R; Kerr, L D; Barnett, J V

2001-09-01

The nuclear factor-kappaB (NF-kappaB) family of transcription factors is involved in proliferation, differentiation, and apoptosis in a stage- and cell-dependent manner. Recent evidence has shown that NF-kappaB activity is necessary for both chicken and mouse limb development. We report here that the NF-kappaB family member c-rel and the homeodomain gene msx-1 have partially overlapping expression patterns in the developing chick limb. In addition, inhibition of NF-kappaB activity resulted in a decrease in msx-1 mRNA expression. Sequence analysis of the msx-1 promoter revealed three potential kappaB-binding sites similar to the interferon-gamma (IFN-gamma) kappaB-binding site. These sites bound to c-Rel, as shown by electrophoretic mobility shift assay (EMSA). Furthermore, inhibition of NF-kappaB activity significantly reduced transactivation of the msx-1 promoter in response to FGF-2/-4, known stimulators of msx-1 expression. These results suggest that NF-kappaB mediates the FGF-2/-4 signal regulation of msx-1 gene expression. Copyright 2001 Academic Press.
Cross Talk in HEK293 Cells Between Nrf2, HIF, and NF-κB Activities upon Challenges with Redox Therapeutics Characterized with Single-Cell Resolution.

PubMed

Johansson, Katarina; Cebula, Marcus; Rengby, Olle; Dreij, Kristian; Carlström, Karl E; Sigmundsson, Kristmundur; Piehl, Fredrik; Arnér, Elias S J

2017-02-20

Many transcription factors with importance in health and disease are redox regulated. However, how their activities may be intertwined in responses to redox-perturbing stimuli is poorly understood. To enable in-depth characterization of this aspect, we here developed a methodology for simultaneous determination of nuclear factor E2-related factor 2 (Nrf2), hypoxia-inducible factor (HIF), and nuclear factor kappa-light-chain-enhancer of activated B cell (NF-κB) activation at single-cell resolution, using a new tool named pTRAF (plasmid for transcription factor reporter activation based upon fluorescence). The pTRAF allowed determination of Nrf2, HIF, and NF-κB activities in a high-resolution and high-throughput manner, and we here assessed how redox therapeutics affected the activities of these transcription factors in human embryonic kidney cells (HEK293). Cross talk was detected between the three signaling pathways upon some types of redox therapeutics, also by using inducers typically considered specific for Nrf2, such as sulforaphane or auranofin, hypoxia for HIF activation, or tumor necrosis factor alpha (TNFα) for NF-κB stimulation. Doxorubicin, at low nontoxic doses, potentiated TNFα-induced activation of NF-κB and HIF, without effects in stand-alone treatment. Stochastic activation patterns in cell cultures were also considerable upon challenges with several redox stimuli. A novel strategy was here used to study simultaneous activation of Nrf2, HIF, and NF-κB in single cells. The method can also be adapted for studies of other transcription factors. The pTRAF provides new opportunities for in-depth studies of transcription factor activities. In this study, we found that upon challenges of cells with several redox-perturbing conditions, Nrf2, HIF, and NF-κB are uniquely responsive to separate stimuli, but can also display marked cross talk to each other within single cells. Antioxid. Redox Signal. 26, 229-246.
BRAF activated non-coding RNA (BANCR) promoting gastric cancer cells proliferation via regulation of NF-κB1

DOE Office of Scientific and Technical Information (OSTI.GOV)

Zhang, Zhi-Xin; Liu, Zhi-Qiang; Jiang, Biao

Background and objective: Long non-coding RNA, BANCR, has been demonstrated to contribute to the proliferation and migration of tumors. However, its molecular mechanism underlying gastric cancer is still unknown. In present study, we investigated whether BANCR was involved in the development of gastric cancer cells via regulation of NF-κB1. Methods: Human gastric cancer tissues were isolated as well as human gastric cell lines MGC803 and BGC823 were cultured to investigate the role of BANCR in gastric cancer. Results: BANCR expression was significantly up-regulated in gastric tumor tissues and gastric cell lines. Down-regulation of BANCR inhibited gastric cancer cell growth andmore » promoted cell apoptosis, and it also contributed to a significant decrease of NF-κB1 (P50/105) expression and 3′UTR of NF-κB1 activity. Overexpression of NF-κB1 reversed the effect of BANCR on cancer cell growth and apoptosis. MiroRNA-9 (miR-9) targeted NF-κB1, and miR-9 inhibitor also reversed the effects of BANCR on gastric cancer cell growth and apoptosis. Conclusion: BANCR was highly expressed both in gastric tumor tissues and in cancer cells. NF-κB1 and miR-9 were involved in the role of BANCR in gastric cancer cell growth and apoptosis. - Highlights: • BANCR up-regulated in gastric cancer (GC) tissues and cell lines MGC803 and BGC823. • Down-regulation of BANCR inhibited GC cell growth and promoted cell apoptosis. • Down-regulation of BANCR contributed to decreased 3′UTR of NF-κB1 and its expression. • Overexpressed NF-κB1 reversed the effect of BANCR on GC cell growth. • miR-9 inhibitor reversed the effect of BANCR on cancer GC cell growth.« less
The underlying mechanism of proinflammatory NF-κB activation by the mTORC2/Akt/IKKα pathway during skin aging

PubMed Central

Choi, Yeon Ja; Moon, Kyoung Mi; Chung, Ki Wung; Jeong, Ji Won; Park, Daeui; Kim, Dae Hyun; Yu, Byung Pal; Chung, Hae Young

2016-01-01

Mammalian target of rapamycin complex 2 (mTORC2), one of two different enzymatic complexes of mTOR, regulates a diverse set of substrates including Akt. mTOR pathway is one of well-known mediators of aging process, however, its role in skin aging has not been determined. Skin aging can be induced by physical age and ultraviolet (UV) irradiation which are intrinsic and extrinsic factors, respectively. Here, we report increased mTORC2 pathway in intrinsic and photo-induced skin aging, which is implicated in the activation of nuclear factor-κB (NF-κB). UVB-irradiated or aged mice skin revealed that mTORC2 activity and its component, rictor were significantly upregulated which in turn increased Akt activation and Akt-dependent IκB kinase α (IKKα) phosphorylation at Thr23 in vivo. We also confirmed that UVB induced the mTORC2/Akt/IKKα signaling pathway with HaCaT human normal keratinocytes. The increased mTORC2 signaling pathway during skin aging were associated to NF-κB activation. Suppression of mTORC2 activity by the treatment of a mTOR small inhibitor or knockdown of RICTOR partially rescued UVB-induced NF-κB activation through the downregulation of Akt/IKKα activity. Our data demonstrated the upregulation of mTORC2 pathway in intrinsic and photo-induced skin aging and its role in IKKα/NF-κB activation. These data not only expanded the functions of mTOR to skin aging but also revealed the therapeutic potential of inhibiting mTORC2 in ameliorating both intrinsic skin aging and photoaging. PMID:27486771
The Inositol Phosphatase SHIP-1 Inhibits NOD2-Induced NF-κB Activation by Disturbing the Interaction of XIAP with RIP2

PubMed Central

Condé, Claude; Rambout, Xavier; Lebrun, Marielle; Lecat, Aurore; Di Valentin, Emmanuel; Dequiedt, Franck; Piette, Jacques

2012-01-01

SHIP-1 is an inositol phosphatase predominantly expressed in hematopoietic cells. Over the ten past years, SHIP-1 has been described as an important regulator of immune functions. Here, we characterize a new inhibitory function for SHIP-1 in NOD2 signaling. NOD2 is a crucial cytoplasmic bacterial sensor that activates proinflammatory and antimicrobial responses upon bacterial invasion. We observed that SHIP-1 decreases NOD2-induced NF-κB activation in macrophages. This negative regulation relies on its interaction with XIAP. Indeed, we observed that XIAP is an essential mediator of the NOD2 signaling pathway that enables proper NF-κB activation in macrophages. Upon NOD2 activation, SHIP-1 C-terminal proline rich domain (PRD) interacts with XIAP, thereby disturbing the interaction between XIAP and RIP2 in order to decrease NF-κB signaling. PMID:22815893
NF-κB Directly Regulates Fas Transcription to Modulate Fas-mediated Apoptosis and Tumor Suppression*

PubMed Central

Liu, Feiyan; Bardhan, Kankana; Yang, Dafeng; Thangaraju, Muthusamy; Ganapathy, Vadivel; Waller, Jennifer L.; Liles, Georgia B.; Lee, Jeffrey R.; Liu, Kebin

2012-01-01

Fas is a member of the death receptor family. Stimulation of Fas leads to induction of apoptotic signals, such as caspase 8 activation, as well as “non-apoptotic” cellular responses, notably NF-κB activation. Convincing experimental data have identified NF-κB as a critical promoter of cancer development, creating a solid rationale for the development of antitumor therapy that suppresses NF-κB activity. On the other hand, compelling data have also shown that NF-κB activity enhances tumor cell sensitivity to apoptosis and senescence. Furthermore, although stimulation of Fas activates NF-κB, the function of NF-κB in the Fas-mediated apoptosis pathway remains largely undefined. In this study, we observed that deficiency of either Fas or FasL resulted in significantly increased incidence of 3-methylcholanthrene-induced spontaneous sarcoma development in mice. Furthermore, Fas-deficient mice also exhibited significantly greater incidence of azoxymethane and dextran sodium sulfate-induced colon carcinoma. In addition, human colorectal cancer patients with high Fas protein in their tumor cells had a longer time before recurrence occurred. Engagement of Fas with FasL triggered NF-κB activation. Interestingly, canonical NF-κB was found to directly bind to the FAS promoter. Blocking canonical NF-κB activation diminished Fas expression, whereas blocking alternate NF-κB increased Fas expression in human carcinoma cells. Moreover, although canonical NF-κB protected mouse embryo fibroblast (MEF) cells from TNFα-induced apoptosis, knocking out p65 diminished Fas expression in MEF cells, resulting in inhibition of FasL-induced caspase 8 activation and apoptosis. In contrast, knocking out p52 increased Fas expression in MEF cells. Our observations suggest that canonical NF-κB is a Fas transcription activator and alternate NF-κB is a Fas transcription repressor, and Fas functions as a suppressor of spontaneous sarcoma and colon carcinoma. PMID:22669972
Decoy receptor 3 suppresses TLR2-mediated B cell activation by targeting NF-κB.

PubMed

Huang, Zi-Ming; Kang, Jhi-Kai; Chen, Chih-Yu; Tseng, Tz-Hau; Chang, Chien-Wen; Chang, Yung-Chi; Tai, Shyh-Kuan; Hsieh, Shie-Liang; Leu, Chuen-Miin

2012-06-15

Decoy receptor 3 (DcR3) is a soluble protein in the TNFR superfamily. Its known ligands include Fas ligand, homologous to lymphotoxin, showing inducible expression, and competing with HSV glycoprotein D for herpes virus entry mediator, a receptor expressed by T lymphocytes, TNF-like molecule 1A, and heparan sulfate proteoglycans. DcR3 has been reported to modulate the functions of T cells, dendritic cells, and macrophages; however, its role in regulating B cell activation is largely unknown. In this study, we found that the DcR3.Fc fusion protein bound to human and mouse B cells and suppressed the activation of B cells. DcR3.Fc attenuated Staphylococcus aureus, IgM-, Pam(3)CSK(4)-, and LPS-mediated B cell proliferation but did not affect cytokine-induced B cell growth. In the presence of these mitogens, DcR3.Fc did not induce B cell apoptosis, suggesting that DcR3 may inhibit the signal(s) important for B cell activation. Because the combination of Fas.Fc, LT-βR.Fc (homologous to lymphotoxin, showing inducible expression, and competing with HSV glycoprotein D for herpes virus entry mediator, a receptor expressed by T lymphocytes receptor), and DR3.Fc (TNF-like molecule 1A receptor) did not suppress B cell proliferation and because the biological effect of DcR3.Fc on B cells was not blocked by heparin, we hypothesize that a novel ligand(s) of DcR3 mediates its inhibitory activity on B cells. Moreover, we found that TLR2-stimulated NF-κB p65 activation and NF-κB-driven luciferase activity were attenuated by DcR3.Fc. The TLR2-induced cytokine production by B cells was consistently reduced by DcR3. These results imply that DcR3 may regulate B cell activation by suppressing the activation of NF-κB.
Direct non transcriptional role of NF-Y in DNA replication.

PubMed

Benatti, Paolo; Belluti, Silvia; Miotto, Benoit; Neusiedler, Julia; Dolfini, Diletta; Drac, Marjorie; Basile, Valentina; Schwob, Etienne; Mantovani, Roberto; Blow, J Julian; Imbriano, Carol

2016-04-01

NF-Y is a heterotrimeric transcription factor, which plays a pioneer role in the transcriptional control of promoters containing the CCAAT-box, among which genes involved in cell cycle regulation, apoptosis and DNA damage response. The knock-down of the sequence-specific subunit NF-YA triggers defects in S-phase progression, which lead to apoptotic cell death. Here, we report that NF-Y has a critical function in DNA replication progression, independent from its transcriptional activity. NF-YA colocalizes with early DNA replication factories, its depletion affects the loading of replisome proteins to DNA, among which Cdc45, and delays the passage from early to middle-late S phase. Molecular combing experiments are consistent with a role for NF-Y in the control of fork progression. Finally, we unambiguously demonstrate a direct non-transcriptional role of NF-Y in the overall efficiency of DNA replication, specifically in the DNA elongation process, using a Xenopus cell-free system. Our findings broaden the activity of NF-Y on a DNA metabolism other than transcription, supporting the existence of specific TFs required for proper and efficient DNA replication. Copyright © 2015 The Authors. Published by Elsevier B.V. All rights reserved.
Nutritional composition analysis of meat from human lactoferrin transgenic bulls.

PubMed

Zhao, Jie; Xu, Jianxiang; Wang, Jianwu; Li, Ning

2013-01-01

Transgenic technology has many potential advantages in food production. However, the transgenic technology process may influence the composition of food products derived from genetically engineered (GE) animals, which may be adverse to human health. Therefore, it is very important to research the compositions of GE animal products. Here, we analyzed the compositions of meat from the offspring of human lactoferrin (hLF) transgenic cows, which can express human lactoferrin proteins in their mammary gland. Six hLF transgenic bulls and three wide-type (WT) bulls, 10 months of age, were slaughtered for meat composition analysis. To determine the comparative health of hLF bulls for meat analysis, hematological analyses, organ/body weight analyses and pathology analyses were conducted. Results of the meat analysis show that there were no significant differences in the hematological parameters, organ/body weight ratios of hLF and WT bulls (P>0.05), and histopathological examination of the main organs of hLF bulls revealed no abnormalities. Nutrient parameters of meat compositions of hLF and WT bulls did not show any significant differences (P>0.05). All of these results suggest that the hLF transgene did not have an impact on the meat nutrient compositions of hLF bulls.
The anti-papillomavirus activity of human and bovine lactoferricin.

PubMed

Mistry, Nitesh; Drobni, Peter; Näslund, Jonas; Sunkari, Vivekananda Gupta; Jenssen, Håvard; Evander, Magnus

2007-09-01

Human papillomavirus (HPV) cause common warts, laryngeal papilloma and genital condylomata and is necessary for the development of cervical cancer. We have previously found that lactoferrin has antiviral activity against HPV-16 and others have demonstrated that lactoferricin, an N-terminal fragment of lactoferrin, has inhibitory activities against several viruses. Two cell lines and two virus types, HPV-5 and HPV-16, were used to study if lactoferrin and lactoferricin could inhibit HPV pseudovirus (PsV) infection. We demonstrated that bovine lactoferrin (bLf) and human lactoferrin (hLf) were both potent inhibitors of HPV-5 and -16 PsV infections. Among the four lactoferricin derivatives we analyzed, a 15 amino acid peptide from bovine lactoferricin (bLfcin) 17-31 was the most potent inhibitor of both HPV-5 and HPV-16 PsV infection. Among the other derivatives, the human lactoferricin (hLfcin) 1-49 showed some antiviral activity against HPV PsV infection while bLfcin 17-42 inhibited only HPV-5 PsV infection in one of the cell lines. When we studied initial attachment of HPV-16, only bLfcin 17-42 and hLfcin 1-49 had an antiviral effect. This is the first time that lactoferricin was demonstrated to have an inhibitory effect on HPV infection and the antiviral activity differed depending on size, charge and structures of the lactoferricin.
LPS Increases 5-LO Expression on Monocytes via an Activation of Akt-Sp1/NF-κB Pathways.

PubMed

Lee, Seung Jin; Seo, Kyo Won; Kim, Chi Dae

2015-05-01

5-Lipoxygenase (5-LO) plays a pivotal role in the progression of atherosclerosis. Therefore, this study investigated the molecular mechanisms involved in 5-LO expression on monocytes induced by LPS. Stimulation of THP-1 monocytes with LPS (0~3 µg/ml) increased 5-LO promoter activity and 5-LO protein expression in a concentration-dependent manner. LPS-induced 5-LO expression was blocked by pharmacological inhibition of the Akt pathway, but not by inhibitors of MAPK pathways including the ERK, JNK, and p38 MAPK pathways. In line with these results, LPS increased the phosphorylation of Akt, suggesting a role for the Akt pathway in LPS-induced 5-LO expression. In a promoter activity assay conducted to identify transcription factors, both Sp1 and NF-κB were found to play central roles in 5-LO expression in LPS-treated monocytes. The LPS-enhanced activities of Sp1 and NF-κB were attenuated by an Akt inhibitor. Moreover, the LPS-enhanced phosphorylation of Akt was significantly attenuated in cells pretreated with an anti-TLR4 antibody. Taken together, 5-LO expression in LPS-stimulated monocytes is regulated at the transcriptional level via TLR4/Akt-mediated activations of Sp1 and NF-κB pathways in monocytes.
Contribution of Cage-Shaped Structure of Physalins to Their Mode of Action in Inhibition of NF-κB Activation.

PubMed

Ozawa, Masaaki; Morita, Masaki; Hirai, Go; Tamura, Satoru; Kawai, Masao; Tsuchiya, Ayako; Oonuma, Kana; Maruoka, Keiji; Sodeoka, Mikiko

2013-08-08

A library of oxygenated natural steroids, including physalins, withanolides, and perulactones, coupled with the synthetic cage-shaped right-side structure of type B physalins, was constructed. SAR studies for inhibition of NF-κB activation showed the importance of both the B-ring and the oxygenated right-side partial structure. The 5β,6β-epoxy derivatives of both physalins and withanolides showed similar profiles of inhibition of NF-κB activation and appeared to act on NF-κB signaling via inhibition of phosphorylation and degradation of IκBα. In contrast, type B physalins with C5-C6 olefin functionality inhibited nuclear translocation and DNA binding of RelA/p50 protein dimer, which lie downstream of IκBα degradation, although withanolides having the same AB-ring functionality did not. These results indicated that the right-side partial structure of these steroids influences their mode of action.

High glucose induces activation of NF-κB inflammatory signaling through IκBα sumoylation in rat mesangial cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Huang, Wei; Department of Endocrinology, The People’s Hospital of Xindu, Xindu, Sichuang, 610500; Xu, Ling

Highlights: •The expression of SUMO1, SUMO2/3 under high glucose was obviously enhanced. •High glucose induced degradation of IκBα and activation of NF-κB pathway. •Sumoylation of IκBα in high glucose were significantly decreased. •The proteasome inhibitor MG132 could partially revert the degradation of IκBα. -- Abstract: The posttranslational modification of proteins by small ubiquitin-like modifiers (SUMOs) has emerged as an important regulatory mechanism for the alteration of protein activity, stability, and cellular localization. The latest research demonstrates that sumoylation is extensively involved in the regulation of the nuclear factor κB (NF-κB) pathway, which plays a critical role in the regulation ofmore » inflammation and contributes to fibrosis in diabetic nephropathy (DN). However, the role of sumoylation in the regulation of NF-κB signaling in DN is still unclear. In the present study, we cultured rat glomerular mesangial cells (GMCs) stimulated by high glucose and divided GMCs into six groups: normal glucose group (5.6 mmol/L), high glucose groups (10, 20, and 30 mmol/L), mannitol group (i.e., osmotic control group), and MG132 intervention group (30 mmol/L glucose with MG132, a proteasome inhibitor). The expression of SUMO1, SUMO2/3, IκBα, NF-κBp65, and monocyte chemotactic protein 1 (MCP-1) was measured by Western blot, reverse-transcription polymerase chain reaction, and indirect immunofluorescence laser scanning confocal microscopy. The interaction between SUMO1, SUMO2/3, and IκBα was observed by co-immunoprecipitation. The results showed that the expression of SUMO1 and SUMO2/3 was dose- and time-dependently enhanced by high glucose (p < 0.05). However, the expression of IκBα sumoylation in high glucose was significantly decreased compared with the normal glucose group (p < 0.05). The expression of IκBα was dose- and time-dependently decreased, and NF-κBp65 and MCP-1 were increased under high glucose conditions
TRIM45 negatively regulates NF-{kappa}B-mediated transcription and suppresses cell proliferation

DOE Office of Scientific and Technical Information (OSTI.GOV)

Shibata, Mio; Sato, Tomonobu; Department of Pediatrics, Hokkaido University Graduate School of Medicine, Sapporo, Hokkaido 060-8638

2012-06-22

Highlights: Black-Right-Pointing-Pointer NF-{kappa}B plays an important role in cell survival and carcinogenesis. Black-Right-Pointing-Pointer TRIM45 negatively regulates TNF{alpha}-induced NF-{kappa}B-mediated transcription. Black-Right-Pointing-Pointer TRIM45 overexpression suppresses cell growth. Black-Right-Pointing-Pointer TRIM45 acts as a repressor for the NF-{kappa}B signal and regulates cell growth. -- Abstract: The NF-{kappa}B signaling pathway plays an important role in cell survival, immunity, inflammation, carcinogenesis, and organogenesis. Activation of NF-{kappa}B is regulated by several posttranslational modifications including phosphorylation, neddylation and ubiquitination. The NF-{kappa}B signaling pathway is activated by two distinct signaling mechanisms and is strictly modulated by the ubiquitin-proteasome system. It has been reported that overexpression of TRIM45, one ofmore » the TRIM family ubiquitin ligases, suppresses transcriptional activities of Elk-1 and AP-1, which are targets of the MAPK signaling pathway. In this study, we showed that TRIM45 also negatively regulates TNF{alpha}-induced NF-{kappa}B-mediated transcription by a luciferase reporter assay and that TRIM45 lacking a RING domain also has an activity to inhibit the NF-{kappa}B signal. Moreover, we found that TRIM45 overexpression suppresses cell growth. These findings suggest that TRIM45 acts as a repressor for the NF-{kappa}B signal and regulates cell growth.« less
Enhanced Optical Breakdown in KB Cells Labeled with Folate-Targeted Silver/Dendrimer Composite Nanodevices

PubMed Central

Tse, Christine; Zohdy, Marwa J.; Ye, Jing Yong; O'Donnell, Matthew; Lesniak, Wojciech; Balogh, Lajos

2010-01-01

Enhanced optical breakdown of KB cells (a human oral epidermoid cancer cell known to overexpress folate receptors) targeted with silver/dendrimer composite nanodevices (CNDs) is described. CNDs {(Ag0}25-PAMAM_E5.(NH2)42(NGly)74(NFA)2.7} were fabricated by reactive encapsulation, using a biocompatible template of dendrimer-folic acid (FA) conjugates. Preferential uptake of the folate-targeted CNDs (of various treatment concentrations and surface functionality) by KB cells was visualized with confocal microscopy and transmission electron microscopy (TEM). Intracellular laser-induced optical breakdown (LIOB) threshold and dynamics were detected and characterized by high-frequency ultrasonic monitoring of resulting transient bubble events. When irradiated with a near-infrared (NIR), femtosecond laser, the CND-targeted KB cells acted as well-confined activators of laser energy, enhancing nonlinear energy absorption, exhibiting a significant reduction in breakdown threshold, and thus selectively promoting intracellular LIOB. PMID:20883823
Coactivation of the PI3K/Akt and ERK signaling pathways in PCB153-induced NF-κB activation and caspase inhibition

DOE Office of Scientific and Technical Information (OSTI.GOV)

Liu, Changjiang; Key Lab of Birth Defects and Reproductive Health of National Health and Family Planning Commission, Chongqing Population and Family Planning Science and Technology Research Institute, Chongqing 400020; Yang, Jixin

2014-06-15

Polychlorinated biphenyls (PCBs) are a group of persistent and widely distributed environmental pollutants that have various deleterious effects, e.g., neurotoxicity, endocrine disruption and reproductive abnormalities. In order to verify the hypothesis that the PI3K/Akt and MAPK pathways play important roles in hepatotoxicity induced by PCBs, Sprague–Dawley (SD) rats were dosed with PCB153 intraperitoneally at 0, 4, 16 and 32 mg/kg for five consecutive days; BRL cells (rat liver cell line) were treated with PCB153 (0, 1, 5, and 10 μM) for 24 h. Results indicated that the PI3K/Akt and ERK pathways were activated in vivo and in vitro after exposuremore » to PCB153, and protein levels of phospho-Akt and phospho-ERK were significantly increased. Nuclear factor-κB (NF-κB) activation and caspase-3, -8 and -9 inhibition caused by PCB153 were also observed. Inhibiting the ERK pathway significantly attenuated PCB153-induced NF-κB activation, whereas inhibiting the PI3K/Akt pathway hardly influenced phospho-NF-κB level. However, inhibiting the PI3K/Akt pathway significantly elevated caspase-3, -8 and -9 activities, while the ERK pathway only synergistically regulated caspase-9. Proliferating cell nuclear antigen (PCNA), a reliable indicator of cell proliferation, was also induced. Moreover, PCB153 led to hepatocellular hypertrophy and elevated liver weight. Taken together, PCB153 leads to aberrant proliferation and apoptosis of hepatocytes through NF-κB activation and caspase inhibition, and coactivated PI3K/Akt and ERK pathways play critical roles in PCB153-induced hepatotoxicity. - Highlights: • PCB153 led to hepatotoxicity through NF-κB activation and caspase inhibition. • The PI3K/Akt and ERK pathways were coactivated in vivo and in vitro by PCB153. • The ERK pathway regulated levels of phospho-NF-κB and caspase-9. • The PI3K/Akt pathway regulated levels of caspase-3, -8 and -9.« less
Dihydroartemisinin inhibits catabolism in rat chondrocytes by activating autophagy via inhibition of the NF-κB pathway

PubMed Central

Jiang, Li-Bo; Meng, De-Hua; Lee, Soo-Min; Liu, Shu-Hao; Xu, Qin-Tong; Wang, Yang; Zhang, Jian

2016-01-01

Osteoarthritis is a disease with inflammatory and catabolic imbalance in cartilage. Dihydroartemisinin (DHA), a natural and safe anti-malarial agent, has been reported to inhibit inflammation, but its effects on chondrocytes have yet to be elucidated. We investigated the effects of DHA on catabolism in chondrocytes. Viability of SD rats chondrocytes was analyzed. Autophagy levels were determined via expression of autophagic markers LC3 and ATG5, GFP-LC3 analysis, acridine orange staining, and electron microscopy. ATG5 siRNA induced autophagic inhibition. Catabolic gene and chemokine expression was evaluated using qPCR. The NF-κB inhibitor SM7368 and p65 over-expression were used to analyze the role of NF-κB pathway in autophagic activation. A concentration of 1 μM DHA without cytotoxicity increased LC3-II and ATG5 levels as well as autophagosomal numbers in chondrocytes. DHA inhibited TNF-α-induced expression of MMP-3 and -9, ADAMTS5, CCL-2 and -5, and CXCL1, which was reversed by autophagic inhibition. TNF-α-stimulated nuclear translocation and degradation of the p65 and IκBα proteins, respectively, were attenuated in DHA-treated chondrocytes. NF-κB inhibition activated autophagy in TNF-α-treated chondrocytes, but p65 over-expression reduced the autophagic response to DHA. These results indicate that DHA might suppress the levels of catabolic and inflammatory factors in chondrocytes by promoting autophagy via NF-κB pathway inhibition. PMID:27941926
Mitogen- and Stress-Activated Kinase 1 (MSK1) Regulates Cigarette Smoke-Induced Histone Modifications on NF-κB-dependent Genes

PubMed Central

Sundar, Isaac K.; Chung, Sangwoon; Hwang, Jae-woong; Lapek, John D.; Bulger, Michael; Friedman, Alan E.; Yao, Hongwei; Davie, James R.; Rahman, Irfan

2012-01-01

Cigarette smoke (CS) causes sustained lung inflammation, which is an important event in the pathogenesis of chronic obstructive pulmonary disease (COPD). We have previously reported that IKKα (I kappaB kinase alpha) plays a key role in CS-induced pro-inflammatory gene transcription by chromatin modifications; however, the underlying role of downstream signaling kinase is not known. Mitogen- and stress-activated kinase 1 (MSK1) serves as a specific downstream NF-κB RelA/p65 kinase, mediating transcriptional activation of NF-κB-dependent pro-inflammatory genes. The role of MSK1 in nuclear signaling and chromatin modifications is not known, particularly in response to environmental stimuli. We hypothesized that MSK1 regulates chromatin modifications of pro-inflammatory gene promoters in response to CS. Here, we report that CS extract activates MSK1 in human lung epithelial (H292 and BEAS-2B) cell lines, human primary small airway epithelial cells (SAEC), and in mouse lung, resulting in phosphorylation of nuclear MSK1 (Thr581), phospho-acetylation of RelA/p65 at Ser276 and Lys310 respectively. This event was associated with phospho-acetylation of histone H3 (Ser10/Lys9) and acetylation of histone H4 (Lys12). MSK1 N- and C-terminal kinase-dead mutants, MSK1 siRNA-mediated knock-down in transiently transfected H292 cells, and MSK1 stable knock-down mouse embryonic fibroblasts significantly reduced CS extract-induced MSK1, NF-κB RelA/p65 activation, and posttranslational modifications of histones. CS extract/CS promotes the direct interaction of MSK1 with RelA/p65 and p300 in epithelial cells and in mouse lung. Furthermore, CS-mediated recruitment of MSK1 and its substrates to the promoters of NF-κB-dependent pro-inflammatory genes leads to transcriptional activation, as determined by chromatin immunoprecipitation. Thus, MSK1 is an important downstream kinase involved in CS-induced NF-κB activation and chromatin modifications, which have implications in pathogenesis
The transcription factor CCAAT-binding factor CBF/NF-Y regulates the proximal promoter activity in the human alpha 1(XI) collagen gene (COL11A1).

PubMed

Matsuo, Noritaka; Yu-Hua, Wang; Sumiyoshi, Hideaki; Sakata-Takatani, Keiko; Nagato, Hitoshi; Sakai, Kumiko; Sakurai, Mami; Yoshioka, Hidekatsu

2003-08-29

We have characterized the proximal promoter region of the human COL11A1 gene. Transient transfection assays indicate that the segment from -199 to +1 is necessary for the activation of basal transcription. Electrophoretic mobility shift assays (EMSAs) demonstrated that the ATTGG sequence, within the -147 to -121 fragment, is critical to bind nuclear proteins in the proximal COL11A1 promoter. We demonstrated that the CCAAT binding factor (CBF/NF-Y) bound to this region using an interference assay with consensus oligonucleotides and a supershift assay with specific antibodies in an EMSA. In a chromatin immunoprecipitation assay and EMSA using DNA-affinity-purified proteins, CBF/NF-Y proteins directly bound this region in vitro and in vivo. We also showed that four tandem copies of the CBF/NF-Y-binding fragment produced higher transcriptional activity than one or two copies, whereas the absence of a CBF/NF-Y-binding fragment suppressed the COL11A1 promoter activity. Furthermore, overexpression of a dominant-negative CBF-B/NF-YA subunit significantly inhibited promoter activity in both transient and stable cells. These results indicate that the CBF/NF-Y proteins regulate the transcription of COL11A1 by directly binding to the ATTGG sequence in the proximal promoter region.
R1: Platelets and Megakaryocytes contain functional NF-κB

PubMed Central

Spinelli, Sherry L.; Casey, Ann E.; Pollock, Stephen J.; Gertz, Jacqueline M.; McMillan, David H.; Narasipura, Srinivasa D.; Mody, Nipa A.; King, Michael R.; Maggirwar, Sanjay B.; Francis, Charles W.; Taubman, Mark B.; Blumberg, Neil; Phipps, Richard P.

2010-01-01

The Nuclear Factor (NF)-κB transcription factor family is well-known for their role in eliciting inflammation and promoting cell survival. We discovered that human megakaryocytes and platelets express the majority of NF-κB family members including the regulatory Inhibitor (I)-κB and Inhibitor Kappa Kinase (IKK) molecules. Objective Investigate the presence and role of NF-κB proteins in megakaryocytes and platelets. Methods and Results Anucleate platelets exposed to NF-κB inhibitors demonstrated impaired fundamental functions involved in repairing vascular injury and thrombus formation. Specifically, NF-κB inhibition diminished lamellapodia formation, decreased clot retraction times and reduced thrombus stability. Moreover, inhibition of I-κB-α phosphorylation (BAY-11-7082) reverts fully spread platelets back to a spheroid morphology. Addition of recombinant IKK-β or I-κB-α protein to BAY inhibitor-treated platelets partially restore platelet spreading in I-κB-α inhibited platelets, and addition of active IKK-β increased endogenous I-κB-α phosphorylation levels. Conclusions These novel findings support a crucial and non-classical role for the NF-κB family in modulating platelet function and reveal that platelets are sensitive to NF-κB inhibitors. As NF-κB inhibitors are being developed as anti-inflammatory and anti-cancer agents, they may have unintended effects on platelets. Based on these data, NF-κB is also identified as a new target to dampen unwanted platelet activation. PMID:20042710
Vimentin and PSF act in concert to regulate IbeA+ E. coli K1 induced activation and nuclear translocation of NF-κB in human brain endothelial cells.

PubMed

Chi, Feng; Bo, Tao; Wu, Chun-Hua; Jong, Ambrose; Huang, Sheng-He

2012-01-01

IbeA-induced NF-κB signaling through its primary receptor vimentin as well as its co-receptor PSF is required for meningitic E. coli K1 penetration and leukocyte transmigration across the blood-brain barrier (BBB), which are the hallmarks of bacterial meningitis. However, it is unknown how vimentin and PSF cooperatively contribute to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB, which are required for bacteria-mediated pathogenicities. IbeA-induced E. coli K1 invasion, polymorphonuclear leukocyte (PMN) transmigration and IKK/NF-κB activation are blocked by Caffeic acid phenethyl ester (CAPE), an inhibitor of NF-κB. IKKα/β phosphorylation is blocked by ERK inhibitors. Co-immunoprecipitation analysis shows that vimentin forms a complex with IκB, NF-κB and tubulins in the resting cells. A dissociation of this complex and a simultaneous association of PSF with NF-κB could be induced by IbeA in a time-dependent manner. The head domain of vimentin is required for the complex formation. Two cytoskeletal components, vimentin filaments and microtubules, contribute to the regulation of NF-κB. SiRNA-mediated knockdown studies demonstrate that IKKα/β phosphorylation is completely abolished in HBMECs lacking vimentin and PSF. Phosphorylation of ERK and nuclear translocation of NF-κB are entirely dependent on PSF. These findings suggest that vimentin and PSF cooperatively contribute to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB activation. PSF is essential for translocation of NF-κB and ERK to the nucleus. These findings reveal previously unappreciated facets of the IbeA-binding proteins. Cooperative contributions of vimentin and PSF to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB may represent a new paradigm in pathogen-induced signal transduction and lead to the development of novel strategies for the prevention and treatment of bacterial meningitis.
Vimentin and PSF Act in Concert to Regulate IbeA+ E. coli K1 Induced Activation and Nuclear Translocation of NF-κB in Human Brain Endothelial Cells

PubMed Central

Wu, Chun-Hua; Jong, Ambrose; Huang, Sheng-He

2012-01-01

Background IbeA-induced NF-κB signaling through its primary receptor vimentin as well as its co-receptor PSF is required for meningitic E. coli K1 penetration and leukocyte transmigration across the blood-brain barrier (BBB), which are the hallmarks of bacterial meningitis. However, it is unknown how vimentin and PSF cooperatively contribute to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB, which are required for bacteria-mediated pathogenicities. Methodology/Principal Findings IbeA-induced E. coli K1 invasion, polymorphonuclear leukocyte (PMN) transmigration and IKK/NF-κB activation are blocked by Caffeic acid phenethyl ester (CAPE), an inhibitor of NF-κB. IKKα/β phosphorylation is blocked by ERK inhibitors. Co-immunoprecipitation analysis shows that vimentin forms a complex with IκB, NF-κB and tubulins in the resting cells. A dissociation of this complex and a simultaneous association of PSF with NF-κB could be induced by IbeA in a time-dependent manner. The head domain of vimentin is required for the complex formation. Two cytoskeletal components, vimentin filaments and microtubules, contribute to the regulation of NF-κB. SiRNA-mediated knockdown studies demonstrate that IKKα/β phosphorylation is completely abolished in HBMECs lacking vimentin and PSF. Phosphorylation of ERK and nuclear translocation of NF-κB are entirely dependent on PSF. These findings suggest that vimentin and PSF cooperatively contribute to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB activation. PSF is essential for translocation of NF-κB and ERK to the nucleus. Conclusion/Significance These findings reveal previously unappreciated facets of the IbeA-binding proteins. Cooperative contributions of vimentin and PSF to IbeA-induced cytoplasmic activation and nuclear translocation of NF-κB may represent a new paradigm in pathogen-induced signal transduction and lead to the development of novel strategies for the prevention and
EWS-FLI1 inhibits TNF{alpha}-induced NF{kappa}B-dependent transcription in Ewing sarcoma cells

DOE Office of Scientific and Technical Information (OSTI.GOV)

Lagirand-Cantaloube, Julie, E-mail: julie.cantaloube@crbm.cnrs.fr; Laud, Karine, E-mail: karine.laud@curie.fr; Institut Curie, Genetique et biologie des cancers, Paris

2010-09-03

Research highlights: {yields} EWS-FLI1 interferes with TNF-induced activation of NF{kappa}B in Ewing sarcoma cells. {yields} EWS-FLI1 knockdown in Ewing sarcoma cells increases TNF-induced NF{kappa}B binding to DNA. {yields} EWS-FLI1 reduces TNF-stimulated NF{kappa}B-dependent transcriptional activation. {yields} Constitutive NF{kappa}B activity is not affected by EWS-FLI1. {yields} EWS-FLI1 physically interacts with NF{kappa}B p65 in vivo. -- Abstract: Ewing sarcoma is primarily caused by a t(11;22) chromosomal translocation encoding the EWS-FLI1 fusion protein. To exert its oncogenic function, EWS-FLI1 acts as an aberrant transcription factor, broadly altering the gene expression profile of tumor cells. Nuclear factor-kappaB (NF{kappa}B) is a tightly regulated transcription factor controllingmore » cell survival, proliferation and differentiation, as well as tumorigenesis. NF{kappa}B activity is very low in unstimulated Ewing sarcoma cells, but can be induced in response to tumor necrosis factor (TNF). We wondered whether NF{kappa}B activity could be modulated by EWS-FLI1 in Ewing sarcoma. Using a knockdown approach in Ewing sarcoma cells, we demonstrated that EWS-FLI1 has no influence on NF{kappa}B basal activity, but impairs TNF-induced NF{kappa}B-driven transcription, at least in part through inhibition of NF{kappa}B binding to DNA. We detected an in vivo physical interaction between the fusion protein and NF{kappa}B p65, which could mediate these effects. Our findings suggest that, besides directly controlling the activity of its primary target promoters, EWS-FLI1 can also indirectly influence gene expression in tumor cells by modulating the activity of key transcription factors such as NF{kappa}B.« less
The UniProtKB guide to the human proteome

PubMed Central

Breuza, Lionel; Poux, Sylvain; Estreicher, Anne; Famiglietti, Maria Livia; Magrane, Michele; Tognolli, Michael; Bridge, Alan; Baratin, Delphine; Redaschi, Nicole

2016-01-01

Advances in high-throughput and advanced technologies allow researchers to routinely perform whole genome and proteome analysis. For this purpose, they need high-quality resources providing comprehensive gene and protein sets for their organisms of interest. Using the example of the human proteome, we will describe the content of a complete proteome in the UniProt Knowledgebase (UniProtKB). We will show how manual expert curation of UniProtKB/Swiss-Prot is complemented by expert-driven automatic annotation to build a comprehensive, high-quality and traceable resource. We will also illustrate how the complexity of the human proteome is captured and structured in UniProtKB. Database URL: www.uniprot.org PMID:26896845
Indoxyl Sulfate Promotes Macrophage IL-1β Production by Activating Aryl Hydrocarbon Receptor/NF-κ/MAPK Cascades, but the NLRP3 inflammasome Was Not Activated

PubMed Central

Wakamatsu, Takuya; Yamamoto, Suguru; Ito, Toru; Sato, Yoko; Matsuo, Koji; Takahashi, Yoshimitsu; Kaneko, Yoshikatsu; Goto, Shin; Kazama, Junichiro James; Gejyo, Fumitake; Narita, Ichiei

2018-01-01

In chronic kidney disease (CKD) patients, accumulation of uremic toxins is associated with cardiovascular risk and mortality. One of the hallmarks of kidney disease-related cardiovascular disease is intravascular macrophage inflammation, but the mechanism of the reaction with these toxins is not completely understood. Macrophages differentiated from THP-1 cells were exposed to indoxyl sulfate (IS), a representative uremic toxin, and changes in inflammatory cytokine production and intracellular signaling molecules including interleukin (IL)-1, aryl hydrocarbon receptor (AhR), nuclear factor (NF)-κ, and mitogen-activated protein kinase (MAPK) cascades as well as the NLRP3 inflammasome were quantified by real-time PCR, Western blot analysis, and enzyme-linked immunosorbent assay. IS induced macrophage pro-IL-1β mRNA expression, although mature IL-1 was only slightly increased. IS increased AhR and the AhR-related mRNA expression; this change was suppressed by administration of proteasome inhibitor. IS promoted phosphorylation of NF-κB p65 and MAPK enzymes; the reaction and IL-1 expression were inhibited by BAY11-7082, an inhibitor of NF-κB. In contrast, IS decreased NLRP3 and did not change ASC, pro-caspase 1, or caspase-1 activation. IS-inducing inflammation in macrophages results from accelerating AhR-NF-κB/MAPK cascades, but the NLRP3 inflammasome was not activated. These reactions may restrict mature IL-1β production, which may explain sustained chronic inflammation in CKD patients. PMID:29543732
An RNA Interference Screen Identifies the Deubiquitinase STAMBPL1 as a Critical Regulator of Human T-Cell Leukemia Virus Type 1 Tax Nuclear Export and NF-κB Activation

PubMed Central

Lavorgna, Alfonso

2012-01-01

The human T-cell leukemia virus type 1 (HTLV-1) Tax oncoprotein actively shuttles between the nucleus, where it interacts with transcriptional and splicing regulatory proteins, and the cytoplasm, where it activates NF-κB. Posttranslational modifications of Tax such as ubiquitination regulate its subcellular localization and hence its function; however, the regulation of Tax trafficking and NF-κB activation by host factors is poorly understood. By screening a deubiquitinating (DUB) enzyme small interfering RNA (siRNA) library, we identified the metalloprotease STAM-binding protein-like 1 (STAMBPL1) as a positive regulator of Tax-mediated NF-κB activation. Overexpression of wild-type STAMBPL1, but not a catalytically inactive mutant, enhanced Tax-mediated NF-κB activation, whereas silencing of STAMBPL1 with siRNA impaired Tax activation of both the canonical and noncanonical NF-κB signaling pathways. STAMBPL1 regulated Tax-induced NF-κB signaling indirectly by controlling Tax nuclear/cytoplasmic transport and was required for DNA damage-induced Tax nuclear export. Together, these results reveal that the deubiquitinase STAMBPL1 is a key regulator of Tax trafficking and function. PMID:22258247
Double-edged swords as cancer therapeutics: novel, orally active, small molecules simultaneously inhibit p53-MDM2 interaction and the NF-κB pathway.

PubMed

Zhuang, Chunlin; Miao, Zhenyuan; Wu, Yuelin; Guo, Zizhao; Li, Jin; Yao, Jianzhong; Xing, Chengguo; Sheng, Chunquan; Zhang, Wannian

2014-02-13

Simultaneous inactivation of p53 and hyperactivation of nuclear factor-κB (NF-κB) is a common occurrence in human cancer. Currently, antitumor agents are being designed to selectively activate p53 or inhibit NF-κB. However, there is no concerted effort yet to deliberately design inhibitors that can simultaneously do both. This paper provided a proof-of-concept study that p53-MDM2 interaction and NF-κB pathway can be simultaneously targeted by a small-molecule inhibitor. A series of pyrrolo[3,4-c]pyrazole derivatives were rationally designed and synthesized as the first-in-class inhibitors of p53-MDM2 interaction and NF-κB pathway. Most of the compounds were identified to possess nanomolar p53-MDM2 inhibitory activity. Compounds 5q and 5s suppressed NF-κB activation through inhibition of IκBα phosphorylation and elevation of the cytoplasmic levels of p65 and phosphorylated IKKα/β. Biochemical assay for the kinases also supported the fact that pyrrolo[3,4-c]pyrazole compounds directly targeted the NF-κB pathway. In addition, four compounds (5j, 5q, 5s, and 5u) effectively inhibited tumor growth in the A549 xenograft model. Further pharmacokinetic study revealed that compound 5q exhibited excellent oral bioavailability (72.9%).
Pre-folding IkappaBalpha alters control of NF-kappaB signaling.

PubMed

Truhlar, Stephanie M E; Mathes, Erika; Cervantes, Carla F; Ghosh, Gourisankar; Komives, Elizabeth A

2008-06-27

Transcription complex components frequently show coupled folding and binding but the functional significance of this mode of molecular recognition is unclear. IkappaBalpha binds to and inhibits the transcriptional activity of NF-kappaB via its ankyrin repeat (AR) domain. The beta-hairpins in ARs 5-6 in IkappaBalpha are weakly-folded in the free protein, and their folding is coupled to NF-kappaB binding. Here, we show that introduction of two stabilizing mutations in IkappaBalpha AR 6 causes ARs 5-6 to fold cooperatively to a conformation similar to that in NF-kappaB-bound IkappaBalpha. Free IkappaBalpha is degraded by a proteasome-dependent but ubiquitin-independent mechanism, and this process is slower for the pre-folded mutants both in vitro and in cells. Interestingly, the pre-folded mutants bind NF-kappaB more weakly, as shown by both surface plasmon resonance and isothermal titration calorimetry in vitro and immunoprecipitation experiments from cells. One consequence of the weaker binding is that resting cells containing these mutants show incomplete inhibition of NF-kappaB activation; they have significant amounts of nuclear NF-kappaB. Additionally, the weaker binding combined with the slower rate of degradation of the free protein results in reduced levels of nuclear NF-kappaB upon stimulation. These data demonstrate clearly that the coupled folding and binding of IkappaBalpha is critical for its precise control of NF-kappaB transcriptional activity.
Lymphomagenic CARD11/BCL10/MALT1 signaling drives malignant B-cell proliferation via cooperative NF-κB and JNK activation.

PubMed

Knies, Nathalie; Alankus, Begüm; Weilemann, Andre; Tzankov, Alexandar; Brunner, Kristina; Ruff, Tanja; Kremer, Marcus; Keller, Ulrich B; Lenz, Georg; Ruland, Jürgen

2015-12-29

The aggressive activated B cell-like subtype of diffuse large B-cell lymphoma is characterized by aberrant B-cell receptor (BCR) signaling and constitutive nuclear factor kappa-B (NF-κB) activation, which is required for tumor cell survival. BCR-induced NF-κB activation requires caspase recruitment domain-containing protein 11 (CARD11), and CARD11 gain-of-function mutations are recurrently detected in human diffuse large B-cell lymphoma (DLBCL). To investigate the consequences of dysregulated CARD11 signaling in vivo, we generated mice that conditionally express the human DLBCL-derived CARD11(L225LI) mutant. Surprisingly, CARD11(L225LI) was sufficient to trigger aggressive B-cell lymphoproliferation, leading to early postnatal lethality. CARD11(L225LI) constitutively associated with B-cell CLL/lymphoma 10 (BCL10) and mucosa-associated lymphoid tissue lymphoma translocation gene 1 (MALT1) to simultaneously activate the NF-κB and c-Jun N-terminal kinase (JNK) signaling cascades. Genetic deficiencies of either BCL10 or MALT1 completely rescued the phenotype, and pharmacological inhibition of JNK was, similar to NF-κB blockage, toxic to autonomously proliferating CARD11(L225LI)-expressing B cells. Moreover, constitutive JNK activity was observed in primary human activated B cell-like (ABC)-DLBCL specimens, and human ABC-DLBCL cells were also sensitive to JNK inhibitors. Thus, our results demonstrate that enforced activation of CARD11/BCL10/MALT1 signaling is sufficient to drive transformed B-cell expansion in vivo and identify the JNK pathway as a therapeutic target for ABC-DLBCL.
NF-κB deregulation in splenic marginal zone lymphoma.

PubMed

Spina, Valeria; Rossi, Davide

2016-08-01

Splenic marginal zone lymphoma is a rare mature B-cell malignancy involving the spleen, bone marrow and blood. Over the past years, the rapid expansion of sequencing technologies allowing the genome-wide assessment of genomic, epigenetic and transcriptional changes has revolutionized our understanding of the biological basis of splenic marginal zone lymphoma by providing a comprehensive and unbiased view of the genes/pathways that are deregulated in this disease. NF-κB is a family of transcription factors that plays critical roles in development, survival, and activation of B lymphocytes. Consistent with the physiological involvement of NF-κB signalling in proliferation and commitment of mature B-cells to the marginal zone of the spleen, many oncogenic mutations involved in constitutive activation of the NF-κB pathway were recently identified in splenic marginal zone lymphoma. This review describes the progress in understanding the mechanism of NF-κB activation in splenic marginal zone lymphoma, including molecular, epigenetic and post-transcriptional modifications of NF-κB genes and of upstream pathways, and discusses how information gained from these efforts has provided new insights on potential targets of diagnostic, prognostic and therapeutic relevance for splenic marginal zone lymphoma. Copyright © 2016. Published by Elsevier Ltd.
The Nuclear Signaling of NF-κB – Current Knowledge, New Insights, and Future Perspectives

PubMed Central

Wan, Fengyi; Lenardo, Michael J.

2011-01-01

The nuclear factor-kappa B (NF-κB) transcription factor plays a critical role in diverse cellular processes associated with proliferation, cell death, development, as well as innate and adaptive immune responses. NF-κB is normally sequestered in the cytoplasm by a family of inhibitory proteins known as IκBs. The signal pathways leading to the liberation and nuclear accumulation of NF-κB, which can be activated by a wide variety of stimuli, have been extensively studied in the past two decades. After gaining access to the nucleus, NF-κB must be actively regulated to execute its fundamental function as a transcription factor. Recent studies have highlighted the importance of nuclear signaling in the regulation of NF-κB transcriptional activity. A non-Rel subunit of NF-κB, ribosomal protein S3 (RPS3), and numerous other nuclear regulators of NF-κB including Akirin, Nurr1, SIRT6, and others, have recently been identified, unveiling novel and exciting layers of regulatory specificity for NF-κB in the nucleus. Further insights into the nuclear events that govern NF-κB function will deepen our understanding of the elegant control of its transcriptional activity and better inform the potential rational design of therapeutics for NF-κB-associated diseases. PMID:19997086
Production of human lactoferrin and lysozyme in the milk of transgenic dairy animals: past, present, and future.

PubMed

Cooper, Caitlin A; Maga, Elizabeth A; Murray, James D

2015-08-01

Genetic engineering, which was first developed in the 1980s, allows for specific additions to animals' genomes that are not possible through conventional breeding. Using genetic engineering to improve agricultural animals was first suggested when the technology was in the early stages of development by Palmiter et al. (Nature 300:611-615, 1982). One of the first agricultural applications identified was generating transgenic dairy animals that could produce altered or novel proteins in their milk. Human milk contains high levels of antimicrobial proteins that are found in low concentrations in the milk of ruminants, including the antimicrobial proteins lactoferrin and lysozyme. Lactoferrin and lysozyme are both part of the innate immune system and are secreted in tears, mucus, and throughout the gastrointestinal (GI) tract. Due to their antimicrobial properties and abundance in human milk, multiple lines of transgenic dairy animals that produce either human lactoferrin or human lysozyme have been developed. The focus of this review is to catalogue the different lines of genetically engineered dairy animals that produce either recombinant lactoferrin or lysozyme that have been generated over the years as well as compare the wealth of research that has been done on the in vitro and in vivo effects of the milk they produce. While recent advances including the development of CRISPRs and TALENs have removed many of the technical barriers to predictable and efficient genetic engineering in agricultural species, there are still many political and regulatory hurdles before genetic engineering can be used in agriculture. It is important to consider the substantial amount of work that has been done thus far on well established lines of genetically engineered animals evaluating both the animals themselves and the products they yield to identify the most effective path forward for future research and acceptance of this technology.

Nuclear lamina defects cause ATM-dependent NF-κB activation and link accelerated aging to a systemic inflammatory response.

PubMed

Osorio, Fernando G; Bárcena, Clea; Soria-Valles, Clara; Ramsay, Andrew J; de Carlos, Félix; Cobo, Juan; Fueyo, Antonio; Freije, José M P; López-Otín, Carlos

2012-10-15

Alterations in the architecture and dynamics of the nuclear lamina have a causal role in normal and accelerated aging through both cell-autonomous and systemic mechanisms. However, the precise nature of the molecular cues involved in this process remains incompletely defined. Here we report that the accumulation of prelamin A isoforms at the nuclear lamina triggers an ATM- and NEMO-dependent signaling pathway that leads to NF-κB activation and secretion of high levels of proinflammatory cytokines in two different mouse models of accelerated aging (Zmpste24(-/-) and Lmna(G609G/G609G) mice). Causal involvement of NF-κB in accelerated aging was demonstrated by the fact that both genetic and pharmacological inhibition of NF-κB signaling prevents age-associated features in these animal models, significantly extending their longevity. Our findings provide in vivo proof of principle for the feasibility of pharmacological modulation of the NF-κB pathway to slow down the progression of physiological and pathological aging.
Inhibition of the NEMO/IKKβ association complex formation, a novel mechanism associated with the NF-κB activation suppression by Withania somnifera's key metabolite withaferin A.

PubMed

Grover, Abhinav; Shandilya, Ashutosh; Punetha, Ankita; Bisaria, Virendra S; Sundar, Durai

2010-12-02

Nuclear Factor kappa B (NF-κB) is a transcription factor involved in the regulation of cell signaling responses and is a key regulator of cellular processes involved in the immune response, differentiation, cell proliferation, and apoptosis. The constitutive activation of NF-κB contributes to multiple cellular outcomes and pathophysiological conditions such as rheumatoid arthritis, asthma, inflammatory bowel disease, AIDS and cancer. Thus there lies a huge therapeutic potential beneath inhibition of NF-κB signalling pathway for reducing these chronic ailments. Withania somnifera, a reputed herb in ayurvedic medicine, comprises a large number of steroidal lactones known as withanolides which show plethora of pharmacological activities like anti- inflammatory, antitumor, antibacterial, antioxidant, anticonvulsive, and immunosuppressive. Though a few studies have been reported depicting the effect of WA (withaferin A) on suppression of NF-κB activation, the mechanism behind this is still eluding the researchers. The study conducted here is an attempt to explore NF-κB signalling pathway modulating capability of Withania somnifera's major constituent WA and to elucidate its possible mode of action using molecular docking and molecular dynamics simulations studies. Formation of active IKK (IκB kinase) complex comprising NEMO (NF-κB Essential Modulator) and IKKβ subunits is one of the essential steps for NF-κB signalling pathway, non-assembly of which can lead to prevention of the above mentioned vulnerable disorders. As observed from our semi-flexible docking analysis, WA forms strong intermolecular interactions with the NEMO chains thus building steric as well as thermodynamic barriers to the incoming IKKβ subunits, which in turn pave way to naive complex formation capability of NEMO with IKKβ. Docking of WA into active NEMO/IKKβ complex using flexible docking in which key residues of the complex were kept flexible also suggest the disruption of the active
Inhibition of intestinal polyp growth by oral ingestion of bovine lactoferrin and immune cells in the large intestine.

PubMed

Iigo, Masaaki; Alexander, David B; Xu, Jiegou; Futakuchi, Mitsuru; Suzui, Masumi; Kozu, Takahiro; Akasu, Takayuki; Saito, Daizo; Kakizoe, Tadao; Yamauchi, Koji; Abe, Fumiaki; Takase, Mitsunori; Sekine, Kazunori; Tsuda, Hiroyuki

2014-10-01

Studies using animal models have demonstrated that ingestion of bovine lactoferrin (bLF) inhibits carcinogenesis in the colon and other organs of experimental animals. As a result of these studies, a blinded, randomized, controlled clinical trial was conducted in the National Cancer Center Hospital, Tokyo, Japan to determine whether ingestion of bLF had an effect on the growth of colorectal polyps in humans. Patients with colorectal polyps ≤5 mm diameter and likely to be adenomas ingested 0, 1.5, or 3.0 g bLF daily for 1 year. Ingestion of 3.0 g bLF suppressed the growth of colorectal polyps and increased the level of serum human lactoferrin in trial participants 63 years old or younger. The purpose of the present study was to investigate correlations between immune parameters and changes in polyp size. Trial participants with regressing polyps had increased NK cell activity, increased serum hLF levels (indicating increased neutrophil activity), and increased numbers of CD4+ cells in the polyps. These findings are consistent with a correlation between higher immune activity and suppression of colorectal polyps. In addition, participants with regressing polyps had lower numbers of PMNs and increased numbers of S100A8+ cells in the polyps, consistent with a correlation between lower inflammatory potential in the colon and suppression of colorectal polyps. Trial participants ingesting bLF had increased serum hLF levels, a possible increase in systemic NK cell activity, and increased numbers of CD4+ and CD161+ cells in the polyps. Taken together, our findings suggest that bLF suppressed colorectal polyps by enhancing immune responsiveness.
Ceftiofur impairs pro-inflammatory cytokine secretion through the inhibition of the activation of NF-{kappa}B and MAPK

DOE Office of Scientific and Technical Information (OSTI.GOV)

Ci Xinxin; Song Yu; Zeng Fanqin

2008-07-18

Ceftiofur is a new broad-spectrum, third-generation cephalosporin antibiotic for veterinary use. Immunopharmacological studies can provide new information on the immunomodulatory activities of some drugs, including their effect on cytokine productions. For this reason, we investigated the effect of ceftiofur on cytokine productions in vitro. We found that ceftiofur can downregulate tumor necrosis factor-{alpha} (TNF-{alpha}), interleukin-1{beta} (IL-1{beta}), and interleukin-6 (IL-6), but did not affect interleukin-10 (IL-10) production. We further investigated signal transduction mechanisms to determine how ceftiofur affects. RAW 264.7 cells were pretreated with 1, 5, or 10 mg/L of ceftiofur 1 h prior to treatment with 1 mg/L of LPS.more » Thirty minutes later, cells were harvested and mitogen activated protein kinases (MAPKs) activation was measured by Western blot. Alternatively, cells were fixed and nuclear factor-{kappa}B (NF-{kappa}B) activation was measured using immunocytochemical analysis. Signal transduction studies showed that ceftiofur significantly inhibited extracellular signal-regulated kinase (ERK), p38, and c-jun NH{sub 2}-terminal kinase (JNK) phosphorylation protein expression. Ceftiofur also inhibited p65-NF-{kappa}B translocation into the nucleus. Therefore, ceftiofur may inhibit LPS-induced production of inflammatory cytokines by blocking NF-{kappa}B and MAPKs signaling in RAW264.7 cells.« less
Activation of beta-major globin gene transcription is associated with recruitment of NF-E2 to the beta-globin LCR and gene promoter.

PubMed

Sawado, T; Igarashi, K; Groudine, M

2001-08-28

The mouse beta-globin gene locus control region (LCR), located upstream of the beta-globin gene cluster, is essential for the activated transcription of genes in the cluster. The LCR contains multiple binding sites for transactivators, including Maf-recognition elements (MAREs). However, little is known about the specific proteins that bind to these sites or the time at which they bind during erythroid differentiation. We have performed chromatin immunoprecipitation experiments to determine the recruitment of the erythroid-specific transactivator p45 NF-E2/MafK (p18 NF-E2) heterodimer and small Maf proteins to various regions in the globin gene locus before and after the induction of murine erythroleukemia (MEL) cell differentiation. We report that, before induction, the LCR is occupied by small Maf proteins, and, on erythroid maturation, the NF-E2 complex is recruited to the LCR and the active globin promoters, even though the promoters do not contain MAREs. This differentiation-coupled recruitment of NF-E2 complex correlates with a greater than 100-fold increase in beta-major globin transcription, but is not associated with a significant change in locus-wide histone H3 acetylation. These findings suggest that the beta-globin gene locus exists in a constitutively open chromatin conformation before terminal differentiation, and we speculate that recruitment of NF-E2 complex to the LCR and active promoters may be a rate-limiting step in the activation of beta-globin gene expression.
RANK-c attenuates aggressive properties of ER-negative breast cancer by inhibiting NF-κB activation and EGFR signaling.

PubMed

Sirinian, Chaido; Papanastasiou, Anastasios D; Schizas, Michail; Spella, Magda; Stathopoulos, Georgios T; Repanti, Maria; Zarkadis, Ioannis K; King, Tari A; Kalofonos, Haralabos P

2018-05-29

The RANK/RANKL axis emerges as a key regulator of breast cancer initiation, progression, and metastasis. RANK-c is a RANK receptor isoform produced through alternative splicing of the TNFRSF11A (RANK) gene and a dominant-negative regulator of RANK-induced nuclear factor-κB (NF-κB) activation. Here we report that RANK-c transcript is expressed in 3.2% of cases in The Cancer Genome Atlas breast cancer cohort evenly between ER-positive and ER-negative cases. Nevertheless, the ratio of RANK to RANK-c (RANK/RANK-c) is increased in ER-negative breast cancer cell lines compared to ER-positive breast cancer cell lines. In addition, forced expression of RANK-c in ER-negative breast cancer cell lines inhibited stimuli-induced NF-κB activation and attenuated migration, invasion, colony formation, and adhesion of cancer cells. Further, RANK-c expression in MDA-MB-231 cells inhibited lung metastasis and colonization in vivo. The RANK-c-mediated inhibition of cancer cell aggressiveness and nuclear factor-κB (NF-κB) activation in breast cancer cells seems to rely on a RANK-c/TNF receptor-associated factor-2 (TRAF2) protein interaction. This was further confirmed by a mutated RANK-c that is unable to interact with TRAF2 and abolishes the ability to attenuate NF-κB activation, migration, and invasion. Additional protein interaction characterization revealed epidermal growth factor receptor (EGFR) as a novel interacting partner for RANK-c in breast cancer cells with a negative effect on EGFR phosphorylation and EGF-dependent downstream signaling pathway activation. Our findings further elucidate the complex molecular biology of the RANKL/RANK system in breast cancer and provide preliminary data for RANK-c as a possible marker for disease progression and aggressiveness.
Bovine lactoferrin and lactoferricin interfere with intracellular trafficking of Herpes simplex virus-1.

PubMed

Marr, A K; Jenssen, H; Moniri, M Roshan; Hancock, R E W; Panté, N

2009-01-01

Although both lactoferrin (Lf), a component of the innate immune system of living organisms, and its N-terminal pepsin cleavage product lactoferricin (Lfcin) have anti-herpes activity, the precise mechanisms by which Lf and Lfcin bring about inhibition of herpes infections are not fully understood. In the present study, experiments were carried out to characterize the activity of bovine Lf and Lfcin (BLf and BLfcin) against the Herpes simplex virus-1 (HSV-1). HSV-1 cellular uptake and intracellular trafficking were studied by immunofluorescence microscopy. In comparison to the untreated infected control cells, both the BLf- and BLfcin-treated cells showed a significant reduction in HSV-1 cellular uptake. The few virus particles that were internalized appeared to have a delayed intracellular trafficking. Thus, in addition to their interference with the uptake of the virus into host cells, Lf and Lfcin also exert their antiviral effect intracellularly.
Tax-1 and Tax-2 similarities and differences: focus on post-translational modifications and NF-κB activation

PubMed Central

Shirinian, Margret; Kfoury, Youmna; Dassouki, Zeina; El-Hajj, Hiba; Bazarbachi, Ali

2013-01-01

Although human T cell leukemia virus type 1 and 2 (HTLV-1 and HTLV-2) share similar genetic organization, they have major differences in their pathogenesis and disease manifestation. HTLV-1 is capable of transforming T lymphocytes in infected patients resulting in adult T cell leukemia/lymphoma whereas HTLV-2 is not clearly associated with lymphoproliferative diseases. Numerous studies have provided accumulating evidence on the involvement of the viral transactivators Tax-1 versus Tax-2 in T cell transformation. Tax-1 is a potent transcriptional activator of both viral and cellular genes. Tax-1 post-translational modifications and specifically ubiquitylation and SUMOylation have been implicated in nuclear factor-kappaB (NF-κB) activation and may contribute to its transformation capacity. Although Tax-2 has similar protein structure compared to Tax-1, the two proteins display differences both in their protein–protein interaction and activation of signal transduction pathways. Recent studies on Tax-2 have suggested ubiquitylation and SUMOylation independent mechanisms of NF-κB activation. In this present review, structural and functional differences between Tax-1 and Tax-2 will be summarized. Specifically, we will address their subcellular localization, nuclear trafficking and their effect on cellular regulatory proteins. A special attention will be given to Tax-1/Tax-2 post-translational modification such as ubiquitylation, SUMOylation, phosphorylation, acetylation, NF-κB activation, and protein–protein interactions involved in oncogenecity both in vivo and in vitro. PMID:23966989
Magnesium isoglycyrrhizinate blocks fructose-induced hepatic NF-κB/NLRP3 inflammasome activation and lipid metabolism disorder.

PubMed

Zhao, Xiao-Juan; Yang, Yan-Zi; Zheng, Yan-Jing; Wang, Shan-Chun; Gu, Hong-Mei; Pan, Ying; Wang, Shui-Juan; Xu, Hong-Jiang; Kong, Ling-Dong

2017-08-15

Magnesium isoglycyrrhizinate as a hepatoprotective agent possesses immune modulation and anti-inflammation, and treats liver diseases. But its effects on immunological-inflammatory and metabolic profiles for metabolic syndrome with liver injury and underlying potential mechanisms are not fully understood. In this study, magnesium isoglycyrrhizinate alleviated liver inflammation and lipid accumulation in fructose-fed rats with metabolic syndrome. It also suppressed hepatic inflammatory signaling activation by reducing protein levels of phosphorylation of nuclear factor-kappa B p65 (p-NF-κB p65), inhibitor of nuclear factor kappa-B kinase α/β (p-IKKα/β) and inhibitor of NF-κB α (p-IκBα) as well as nucleotide-binding domain (NOD)-like receptor protein 3 (NLRP3), apoptosis-associated speck-like protein (ASC) and Caspase-1 in rats, being consistent with its reduction of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α) and IL-6 levels. Furthermore, magnesium isoglycyrrhizinate modulated lipid metabolism-related genes characterized by up-regulating peroxisome proliferator-activated receptor-α (PPAR-α) and carnitine palmitoyl transferase-1 (CPT-1), and down-regulating sensor for fatty acids to control-1 (SREBP-1) and stearoyl-CoA desaturase 1 (SCD-1) in the liver of fructose-fed rats, resulting in the reduction of triglyceride and total cholesterol levels. These effective actions were further confirmed in fructose-exposed BRL-3A and HepG2 cells. The molecular mechanisms underpinning these observations suggest that magnesium isoglycyrrhizinate may inhibit NF-κB/NLRP3 inflammasome activation to reduce immunological-inflammatory response, which in turn may prevent liver lipid metabolic disorder and accumulation under high fructose condition. Thus, blockade of NF-κB/NLRP3 inflammasome activation and lipid metabolism disorder by magnesium isoglycyrrhizinate may be the potential therapeutic approach for improving fructose-induced liver injury with
NF-kappaB: Two Sides of the Same Coin.

PubMed

Pires, Bruno R B; Silva, Rafael C M C; Ferreira, Gerson M; Abdelhay, Eliana

2018-01-09

Nuclear Factor-kappa B (NF-κB) is a transcription factor family that regulates a large number of genes that are involved in important physiological processes, including survival, inflammation, and immune responses. More recently, constitutive expression of NF-κB has been associated with several types of cancer. In addition, microorganisms, such as viruses and bacteria, cooperate in the activation of NF-κB in tumors, confirming the multifactorial role of this transcription factor as a cancer driver. Recent reports have shown that the NF-κB signaling pathway should receive attention for the development of therapies. In addition to the direct effects of NF-κB in cancer cells, it might also impact immune cells that can both promote or prevent tumor development. Currently, with the rise of cancer immunotherapy, the link among immune cells, inflammation, and cancer is a major focus, and NF-κB could be an important regulator for the success of these therapies. This review discusses the contrasting roles of NF-κB as a regulator of pro- and antitumor processes and its potential as a therapeutic target.
A Bio-Guided Fractionation to Assess the Inhibitory Activity of Calendula officinalis L. on the NF-κB Driven Transcription in Human Gastric Epithelial Cells

PubMed Central

Colombo, Elisa; Sangiovanni, Enrico; D'Ambrosio, Michele; Bosisio, Enrica; Ciocarlan, Alexandru; Fumagalli, Marco; Guerriero, Antonio; Harghel, Petru; Dell'Agli, Mario

2015-01-01

Calendula officinalis L. has been largely known for its topical anti-inflammatory properties; however, there are no experimental evidences about its antiphlogistic effect at the gastric level. To investigate whether marigold might exert an activity against gastric inflammation, a CH2Cl2 extract obtained from C. officinalis flowers was evaluated in vitro on the NF-κB pathway. The lipophilic extract demonstrated a significant inhibitory effect on the NF-κB driven transcription. The identification of active compounds was conducted by a bio-guided fractionation of the extract that afforded 16 fractions. Fraction J exhibited a concentration-dependent inhibitory activity on the NF-κB driven transcription and significantly contributed to the antiphlogistic effect showed by CH2Cl2 extract. The main components of fraction J were loliolide and the fucoside acetates of β-eudesmol and viridiflorol. HPLC analysis of fractions D and E led to the identification and isolation of triterpene esters that showed a concentration-dependent inhibition of the NF-κB driven transcription, with faradiol-3-myristate and the corresponding aglycone being the most active compounds. The present study provides some experimental evidences that Calendula officinalis L. may exert an anti-inflammatory activity on the gastric district by the inhibition of the NF-κB system, identifying the compounds responsible, at least in part, for the observed effect. PMID:26491463
Acidosis promotes invasiveness of breast cancer cells through ROS-AKT-NF-κB pathway

PubMed Central

Gupta, Subash C.; Singh, Ramesh; Pochampally, Radhika; Watabe, Kounosuke; Mo, Yin-Yuan

2014-01-01

It is well known that acidic microenvironment promotes tumorigenesis, however, the underlying mechanism remains largely unknown. In the present study, we show that acidosis promotes invasiveness of breast cancer cells through a series of signaling events. First, our study indicates that NF-κB is a key factor for acidosis-induced cell invasion. Acidosis activates NF-κB without affecting STAT3 activity; knockdown of NF-κB p65 abrogates the acidosis-induced invasion activity. Next, we show that the activation of NF-κB is mediated through phosphorylation and degradation of IκBα; and phosphorylation and nuclear translocation of p65. Upstream to NF-κB signaling, AKT is activated under acidic conditions. Moreover, acidosis induces generation of reactive oxygen species (ROS) which can be suppressed by ROS scavengers, reversing the acidosis-induced activation of AKT and NF-κB, and invasiveness. As a negative regulator of AKT, PTEN is oxidized and inactivated by the acidosis-induced ROS. Finally, inhibition of NADPH oxidase (NOX) suppresses acidosis-induced ROS production, suggesting involvement of NOX in acidosis-induced signaling cascade. Of considerable interest, acidosis-induced ROS production and activation of AKT and NF-κB can be only detected in cancer cells, but not in non-malignant cells. Together, these results demonstrate a cancer specific acidosis-induced signaling cascade in breast cancer cells, leading to cell invasion. PMID:25504433
Synergistic activation of NF-{kappa}B by nontypeable H. influenzae and S. pneumoniae is mediated by CK2, IKK{beta}-I{kappa}B{alpha}, and p38 MAPK

DOE Office of Scientific and Technical Information (OSTI.GOV)

Kweon, Soo-Mi; Wang, Beinan; Rixter, Davida

2006-12-15

In review of the past studies on NF-{kappa}B regulation, most of them have focused on investigating how NF-{kappa}B is activated by a single inducer at a time. Given the fact that, in mixed bacterial infections in vivo, multiple inflammation inducers, including both nontypeable Haemophilus influenzae (NTHi) and Streptococcus pneumoniae, are present simultaneously, a key issue that has yet to be addressed is whether NTHi and S. pneumoniae simultaneously activate NF-{kappa}B and the subsequent inflammatory response in a synergistic manner. Here, we show that NTHi and S. pneumoniae synergistically induce NF-{kappa}B-dependent inflammatory response via activation of multiple signaling pathways in vitromore » and in vivo. The classical IKK{beta}-I{kappa}B{alpha} and p38 MAPK pathways are involved in synergistic activation of NF-{kappa}B via two distinct mechanisms, p65 nuclear translocation-dependent and -independent mechanisms. Moreover, casein kinase 2 (CK2) is involved in synergistic induction of NF-{kappa}B via a mechanism dependent on phosphorylation of p65 at both Ser536 and Ser276 sites. These studies bring new insights into the molecular mechanisms underlying the NF-{kappa}B-dependent inflammatory response in polymicrobial infections and may lead to development of novel therapeutic strategies for modulating inflammation in mixed infections for patients with otitis media and chronic obstructive pulmonary diseases.« less
S100A8 and S100A9 Promotes Invasion and Migration through p38 Mitogen-Activated Protein Kinase-Dependent NF-κB Activation in Gastric Cancer Cells

PubMed Central

Kwon, Chae Hwa; Moon, Hyun Jung; Park, Hye Ji; Choi, Jin Hwa; Park, Do Youn

2013-01-01

S100A8 and S100A9 (S100A8/A9) are low-molecular weight members of the S100 family of calcium-binding proteins. Recent studies have reported S100A8/A9 promote tumorigenesis. We have previously reported that S100A8/A9 is mostly expressed in stromal cells and inflammatory cells between gastric tumor cells. However, the role of environmental S100A8/A9 in gastric cancer has not been defined. We observed in the present study the effect of S100A8/A9 on migration and invasion of gastric cancer cells. S100A8/A9 treatment increased migration and invasionat lower concentrations that did not affect cell proliferation and cell viability. S100A8/A9 caused activation of p38 mitogen-activated protein kinase (MAPK) and nuclear factor-κB (NF-κB). The phosphorylation of p38 MAPK was not affected by the NF-κB inhibitor Bay whereas activation of NF-κB was blocked by p38 MAPK inhibitor SB203580, indicating that S100A8/A9-induced NF-κB activation is mediated by phosphorylation of p38 MAPK. S100A8/A9-induced cell migration and invasion was inhibited by SB203580 and Bay, suggesting that activation of p38 MAPK and NF-κB is involved in the S100A8/A9 induced cell migration and invasion. S100A8/A9 caused an increase in matrix metalloproteinase 2 (MMP2) and MMP12 expression, which were inhibited by SB203580 and Bay. S100A8/A9-induced cell migration and invasion was inhibited by MMP2 siRNA and MMP12 siRNA, indicating that MMP2 and MMP12 is related to the S100A8/A9 induced cell migration and invasion. Taken together, these results suggest that S100A8/A9 promotes cell migration and invasion through p38 MAPK-dependent NF-κB activation leading to an increase of MMP2 and MMP12 in gastric cancer. PMID:23456298
IL-32γ promotes integrin αvβ6 expression through the activation of NF-κB in HSCs

PubMed Central

Liu, Hongcan; Pan, Xingfei; Cao, Hong; Shu, Xin; Sun, Haixia; Lu, Jianxi; Liang, Jiayin; Zhang, Ka; Zhu, Fengqin; Li, Gang; Zhang, Qi

2017-01-01

Hepatic stellate cell (HSC) activation is important in the pathogenesis of liver fibrosis. However, the molecular mechanism of HSC activation is not completely understood. In the present study, it was demonstrated that interleukin-32γ (IL-32γ) is capable of enhancing intefgrin αvβ6 expression by inducing integrin αvβ6 promoter activity in a dose-dependent manner in HSCs. Furthermore, it was determined that nuclear factor κB (NF-κB) activation is required for IL-32γ-induced integrin αvβ6 expression. Increased integrin αvβ6 expression is then able to activate HSCs. These results indicate that NF-κB activation is required for IL-32γ to induce integrin αvβ6 expression and consequently promote HSC activation. Therefore, IL-32γ activates HSCs and therefore may be associated with hepatic fibrogenesis. These results may enable the development of novel effective strategies to treat hepatic fibrosis. PMID:29042996
Human papillomavirus type 16 E6 and E 7 proteins alter NF-kB in cultured cervical epithelial cells and inhibition of NF-kB promotes cell growth and immortalization

DOE Office of Scientific and Technical Information (OSTI.GOV)

Vandermark, Erik R.; Deluca, Krysta A.; Gardner, Courtney R.

2012-03-30

The NF-kB family of transcription factors regulates important biological functions including cell growth, survival and the immune response. We found that Human Papillomavirus type 16 (HPV-16) E7 and E6/E7 proteins inhibited basal and TNF-alpha-inducible NF-kB activity in human epithelial cells cultured from the cervical transformation zone, the anatomic region where most cervical cancers develop. In contrast, HPV-16 E6 regulated NF-kB in a cell type- and cell growth-dependent manner. NF-kB influenced immortalization of cervical cells by HPV16. Inhibition of NF-kB by an IkB alpha repressor mutant increased colony formation and immortalization by HPV-16. In contrast, activation of NF-kB by constitutive expressionmore » of p65 inhibited proliferation and immortalization. Our results suggest that inhibition of NF-kB by HPV-16 E6/E7 contributes to immortalization of cells from the cervical transformation zone.« less
Dietary Apigenin Exerts Immune-Regulatory Activity in Vivo by Reducing NF-κB Activity, Halting Leukocyte Infiltration and Restoring Normal Metabolic Function

PubMed Central

Cardenas, Horacio; Arango, Daniel; Nicholas, Courtney; Duarte, Silvia; Nuovo, Gerard J.; He, Wei; Voss, Oliver H.; Gonzalez-Mejia, M. Elba; Guttridge, Denis C.; Grotewold, Erich; Doseff, Andrea I.

2016-01-01

The increasing prevalence of inflammatory diseases and the adverse effects associated with the long-term use of current anti-inflammatory therapies prompt the identification of alternative approaches to reestablish immune balance. Apigenin, an abundant dietary flavonoid, is emerging as a potential regulator of inflammation. Here, we show that apigenin has immune-regulatory activity in vivo. Apigenin conferred survival to mice treated with a lethal dose of Lipopolysaccharide (LPS) restoring normal cardiac function and heart mitochondrial Complex I activity. Despite the adverse effects associated with high levels of splenocyte apoptosis in septic models, apigenin had no effect on reducing cell death. However, we found that apigenin decreased LPS-induced apoptosis in lungs, infiltration of inflammatory cells and chemotactic factors’ accumulation, re-establishing normal lung architecture. Using NF-κB luciferase transgenic mice, we found that apigenin effectively modulated NF-κB activity in the lungs, suggesting the ability of dietary compounds to exert immune-regulatory activity in an organ-specific manner. Collectively, these findings provide novel insights into the underlying immune-regulatory mechanisms of dietary nutraceuticals in vivo. PMID:26938530
Dietary Apigenin Exerts Immune-Regulatory Activity in Vivo by Reducing NF-κB Activity, Halting Leukocyte Infiltration and Restoring Normal Metabolic Function.

PubMed

Cardenas, Horacio; Arango, Daniel; Nicholas, Courtney; Duarte, Silvia; Nuovo, Gerard J; He, Wei; Voss, Oliver H; Gonzalez-Mejia, M Elba; Guttridge, Denis C; Grotewold, Erich; Doseff, Andrea I

2016-03-01

The increasing prevalence of inflammatory diseases and the adverse effects associated with the long-term use of current anti-inflammatory therapies prompt the identification of alternative approaches to reestablish immune balance. Apigenin, an abundant dietary flavonoid, is emerging as a potential regulator of inflammation. Here, we show that apigenin has immune-regulatory activity in vivo. Apigenin conferred survival to mice treated with a lethal dose of Lipopolysaccharide (LPS) restoring normal cardiac function and heart mitochondrial Complex I activity. Despite the adverse effects associated with high levels of splenocyte apoptosis in septic models, apigenin had no effect on reducing cell death. However, we found that apigenin decreased LPS-induced apoptosis in lungs, infiltration of inflammatory cells and chemotactic factors' accumulation, re-establishing normal lung architecture. Using NF-κB luciferase transgenic mice, we found that apigenin effectively modulated NF-κB activity in the lungs, suggesting the ability of dietary compounds to exert immune-regulatory activity in an organ-specific manner. Collectively, these findings provide novel insights into the underlying immune-regulatory mechanisms of dietary nutraceuticals in vivo.
Human recombinant lactoferrin acts synergistically with antimicrobials commonly used in neonatal practice against coagulase-negative staphylococci and Candida albicans causing neonatal sepsis.

PubMed

Venkatesh, Mohan Pammi; Rong, Liang

2008-09-01

Neonatal sepsis causes significant mortality and morbidity. Coagulase-negative staphylococci (CoNS) and Candida frequently cause neonatal sepsis at >72 h of age. Lactoferrin, which is present in human milk, is a component of innate immunity and has broad-spectrum antimicrobial activity. The synergistic effects of lactoferrin with antibiotics against neonatal isolates have not been systematically evaluated. Here, eight clinical strains (seven neonatal) of CoNS and three strains (two neonatal) of Candida albicans were studied. MIC50 and MIC90 values of human recombinant lactoferrin (talactoferrin; TLF), vancomycin (VAN) and nafcillin (NAF) against CoNS, and of TLF, amphotericin B (AMB) and fluconazole (FLC) against C. albicans, were evaluated according to established guidelines. Antimicrobial combinations of TLF with NAF or VAN against CoNS, and TLF with AMB or FLC against C. albicans, were evaluated by a checkerboard method with serial twofold dilutions. Synergy was evaluated by the median effects principle, and combination indices and dose reduction indices were reported at 50, 75 and 90% inhibitory effect at several drug-dose ratios. It was found that TLF acted synergistically with NAF and VAN against CoNS, and with AMB and FLC against C. albicans, at multiple dose effects and drug-dose ratios with few exceptions. In synergistic combinations, drug reduction indices indicated a significant reduction in doses of antibiotics, which may be clinically relevant. Thus TLF acts synergistically with anti-staphylococcal and anti-Candida agents commonly used in neonatal practice and is a promising agent that needs to be evaluated in clinical studies.
Novel curcumin analogue UBS109 potently stimulates osteoblastogenesis and suppresses osteoclastogenesis: involvement in Smad activation and NF-κB inhibition.

PubMed

Yamaguchi, Masayoshi; Moore, Terry W; Sun, Aiming; Snyder, James P; Shoji, Mamoru

2012-08-01

Bone homeostasis is maintained through a balance between osteoblastic bone formation and osteoclastic bone resorption. Bone loss is induced due to decreased osteoblastic bone formation and increased osteoclastic bone resorption with various pathologic states. Osteoporosis with its accompanying decrease in bone mass is widely recognized as a major public health problem. Pharmacologic and functional food factors may play a role in the prevention of bone loss with aging. This study was undertaken to determine the effect of curcumin analogues (curcumin, EF31, ECMN909, and UBS109), which were newly synthesized, on osteoblastogenesis and osteoclastogenesis in vitro. Among these compounds, UBS109 had a unique stimulatory effect on osteoblastic differentiation and mineralization. UBS109 stimulated both basal and bone morphogenic protein-2 (BMP2)-increased Smad-luciferase activity, the Smad signaling of which is related to osteoblastogenesis. Such an effect was not seen with other compounds. Moreover, UBS109 potently suppressed tumor necrosis factor-α (TNF-α)-increased osteoblastic nuclear factor kappa B (NF-κB)-luciferase activity. In addition, EF31, ECMN909, and UBS109 had a suppressive effect on osteoclastogenesis as compared with that of curcumin. ECMN909 and UBS109 potently inhibited the receptor activator of NF-κB (RANK) ligand (RANKL)-increased preosteoclastic NF-κB-luciferase activity, in which NF-κB signaling plays a pivotal role in osteoclastogenesis. In the present study, curcumin analogue UBS109 was found to have a stimulating effect on osteoblastogenesis and a suppressive effect on osteoclastogenesis in vitro, suggesting an anabolic effect of the compound on bone mass.

Lactoferrin delivery systems: approaches for its more effective use.

PubMed

Onishi, Hiraku

2011-11-01

Recently, pharmacotherapy has advanced extensively, but there are still many refractory diseases which cannot be solved fully by existing therapeutic agents. Therefore, alternative medicine and health foods are now attracting much attention, for example, lactoferrin (LF): a multifunctional glycoprotein. As LF is non-toxic and low-cost, its application in healthcare and therapeutics is expected to be widespread. In this review, LF's general basic features are described. The interaction of LF with its receptors activates the immune system, including cytokine production and balance. In particular, the immune activation of orally administered LF is considered as a new strategy for the treatment of refractory diseases, such as inflammatory bowel disease, virus infection and tumor metastasis. Also mentioned are the problems associated with the use of LF. As LF is degraded rapidly in the body due to enzymatic hydrolysis, high amounts or frequent dosing is required; an appropriate delivery system may improve these problems and increase its efficiency. Chemical modifications, such as PEGylation, can enhance the stability of LF in the body, resulting in increased efficacy. Also, liposomes and enteric or microparticulate formulations can promote the function of LF in oral administration due to target site delivery and protection of LF from enzymatic hydrolysis. These delivery systems are expected to improve the utility of LF.
Pre-S2 Start Codon Mutation of Hepatitis B Virus Subgenotype B3 Effects on NF-κB Expression and Activation in Huh7 Cell Lines.

PubMed

Siburian, Marlinang Diarta; Suriapranata, Ivet Marita; Wanandi, Septelia Inawati

2018-03-19

A cross-sectional study on hepatitis B patients in Indonesia showed association of pre-S2 start codon mutation (M120 V) with cirrhosis and hepatocellular carcinoma (HCC), which was dissimilar from studies from other populations where pre-S2 deletion mutation was more prevalent. Different mutation patterns were attributed to different hepatitis B virus (HBV) subgenotypes in each population study. HBV surface proteins are reported to induce the activation of NF-κB, a transcriptional factor known to play an important role in the development of liver disease. This study aimed to see the effects of HBs variants in HBV subgenotype B3 on the expression and activation of NF-κB as one of the mechanisms in inducing advanced liver disease. HBV subgenotypes B3, each carrying wild-type (wt) HBs, M120 V, and pre-S2 deletion mutation were isolated from three HCC patients. HBs genes were amplified and cloned into pcDNA3.1 and were transfected using Lipofectamine into a Huh7 cell line. NF-κB activation was measured through IκB-α expression, which is regulated by NF-κB. RNA expressions for HBs, IκB-α, and NF-κB subunit (p50) were evaluated using real-time PCR. M120 V mutant had a significantly higher mRNA level compared with wt and pre-S2 deletion mutant; however, there were no significant differences in HBs protein expressions. The transcription level of p50 was higher in M120 V mutation compared with HBs wild-type and pre-S2 deletion mutant. NF-κB activation was higher in HBs wild-type compared with the two mutant variants. Pre-S2 mutations had no effect on the increment of NF-κB activation. However, M120 V mutation may utilize a different pathway in liver disease progression that involves high expression of NF-κB subunit, p50.
Therapeutic microRNAs targeting the NF-kappa B Signaling Circuits of Cancers

PubMed Central

Tong, Lingying; Yuan, Ye; Wu, Shiyong

2014-01-01

MicroRNAs (miRNAs) not only directly regulate NF-κB expression, but also up- or down-regulate NF-κB activity via upstream and downstream signaling pathways of NF-κB. In many cancer cells, miRNA expressions are altered accompanied with an elevation of NF-κB, which often plays a role in promoting cancer development and progression as well as hindering the effectiveness of chemo and radiation therapies. Thus NF-κB-targeting miRNAs have been identified and characterized as potential therapeutics for cancer treatment and sensitizers of chemo and radiotherapies. However, due to cross-targeting and instability of miRNAs, some limitations of using miRNA as cancer therapeutics still exist. In this review, the mechanisms for miRNA-mediated alteration of NF-κB expression and activation in different types of cancers will be discussed. The results of therapeutic use of NF-κB-targeting miRNA for cancer treatment will be examined. Some limitations, challenges and potential strategies in future development of miRNA as cancer therapeutics are also assessed. PMID:25220353
Free radical-triggered hepatic injury of experimental obstructive jaundice of rats involves overproduction of proinflammatory cytokines and enhanced activation of nuclear factor kappaB.

PubMed

Liu, T Z; Lee, K T; Chern, C L; Cheng, J T; Stern, A; Tsai, L Y

2001-10-01

Excessive production of hydroxyl radicals in blood and liver has previously been demonstrated by us in rats with obstructive jaundice induced by common bile duct ligation (CBDL). In this study, we demonstrate overproduction of superoxide radicals in circulating blood of CBDL rats by the lucigenin-amplified chemiluminescence technique. To pinpoint the molecular agents that mediate these processes, we measured circulating proinflammatory cytokines, such as tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta ( IL-1beta), and interleukin-6 (IL-6) in controls and CBDL rats. Concentrations of these cytokines in blood of CBDL rats were markedly elevated when compared to the controls (TNF-alpha: 36.7 +/- 5.0 vs 13.8 +/- 0.5 pg/mL; IL-6: 2,814 +/- 1,740 vs 0 pg/mL; IL-1beta: 11.9 +/- 2.6 vs 0 pg/mL). The overproduction of free radicals triggered by elevated cytokines in CBDL rats was correlated with the activation of NF-kappaB in hepatic tissue. Using the TdT-mediated dUTP nick-end label staining technique, we showed that hepatic tissue sections from CBDL rats had an increase in the apoptotic index (AI). Based on these findings, we propose that the severe hepatic injury in CBDL rats is mediated by a cycle that involves the activation of NF-kappaB by combined action of proinflammatory cytokines and reactive oxygen species (ROS). NF-KB, in turn, initiates the transcription of cytokine genes (eg, IL-6, IL-8, TNF-alpha), which triggers hepatic injury, at least in part, by a free radical-mediated apoptotic mechanism. Elevated ROS may be as a positive-feedback signal that triggers NF-KB reactivation; the severe hepatic injury of CBDL rats may result from perpetuation of this vicious cycle.
Hypoxia and P. gingivalis Synergistically Induce HIF-1 and NF-κB Activation in PDL Cells and Periodontal Diseases

PubMed Central

Gölz, L.; Memmert, S.; Rath-Deschner, B.; Jäger, A.; Appel, T.; Baumgarten, G.; Götz, W.; Frede, S.

2015-01-01

Periodontitis is characterized by deep periodontal pockets favoring the proliferation of anaerobic bacteria like Porphyromonas gingivalis (P. gingivalis), a periodontal pathogen frequently observed in patients suffering from periodontal inflammation. Therefore, the aim of the present study was to investigate the signaling pathways activated by lipopolysaccharide (LPS) of P. gingivalis (LPS-PG) and hypoxia in periodontal ligament (PDL) cells. The relevant transcription factors nuclear factor-kappa B (NF-κB) and hypoxia inducible factor-1 (HIF-1) were determined. In addition, we analyzed the expression of interleukin- (IL-) 1β, matrix metalloproteinase-1 (MMP-1), and vascular endothelial growth factor (VEGF) in PDL cells on mRNA and protein level. This was accomplished by immunohistochemistry of healthy and inflamed periodontal tissues. We detected time-dependent additive effects of LPS-PG and hypoxia on NF-κB and HIF-1α activation in PDL cells followed by an upregulation of IL-1β, MMP-1, and VEGF expression. Immunohistochemistry performed on tissue samples of gingivitis and periodontitis displayed an increase of NF-κB, HIF-1, and VEGF immunoreactivity in accordance with disease progression validating the importance of the in vitro results. To conclude, the present study underlines the significance of NF-κB and HIF-1α and their target genes VEGF, IL-1β, and MMP-1 in P. gingivalis and hypoxia induced periodontal inflammatory processes. PMID:25861162
Stabilisation of p53 enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-κB activation.

PubMed

Pan, D; Pan, L-Z; Hill, R; Marcato, P; Shmulevitz, M; Vassilev, L T; Lee, P W K

2011-09-27

Naturally oncolytic reovirus preferentially kills cancer cells, making it a promising cancer therapeutic. Mutations in tumour suppressor p53 are prevalent in cancers, yet the role of p53 in reovirus oncolysis is relatively unexplored. Human cancer cell lines were exposed to Nutlin-3a, reovirus or a combination of the two and cells were processed for reovirus titration, western blot, real-time PCR and apoptosis assay using Annexin V and 7-AAD staining. Confocal microscopy was used to determine translocation of the NF-κB p65 subunit. We show that despite similar reovirus replication in p53(+/+) and p53(-/-) cells, stabilisation of p53 by Nutlin-3a significantly enhanced reovirus-induced apoptosis and hence virus release and dissemination while having no direct effect on virus replication. Enhanced apoptosis by Nutlin-3a was not observed in p53(-/-) or p53 knockdown cells; however, increased expression of Bax and p21 are required. Moreover, elevated NF-κB activation in reovirus-infected cells following Nutlin-3a treatment was necessary for enhanced reovirus-induced apoptosis, as synergistic cytotoxicity was overcome by specific NF-κB inhibitors. Nutlin-3a treatment enhances reovirus-induced apoptosis and virus spread through p53-dependent NF-κB activation, and combination of reovirus and Nutlin-3a might represent an improved therapy against cancers harbouring wild-type p53.
Oleanane-triterpenoids from Panax stipuleanatus inhibit NF-κB

PubMed Central

Liang, Chun; Ding, Yan; Song, Seok Bean; Kim, Jeong Ah; Cuong, Nguyen Manh; Ma, Jin Yeul; Kim, Young Ho

2013-01-01

In continuation of our research to find biological components from Panax stipuleanatus, four oleanane-type triterpenes (12 to 15) were isolated successively. Fifteen oleanane-type saponins (1 to 15) were evaluated for nuclear factor (NF)-κB activity using a luciferase reporter gene assay in HepG2 cells. Compounds 6 to 11 inhibited NF-κB, with IC50 values between 3.1 to 18.9 μM. The effects on inducible nitric oxide synthase and cyclooxygenase-2 by compounds 8, 10, and 11 were also examined using reverse transcription-polymerase chain reaction. Three compounds (8, 10, and 11) inhibited NF-κB activity by reducing the concentration of inflammatory factors in HepG2 cells. PMID:23717159
Hypoxia and Inflammation in Cancer, Focus on HIF and NF-κB

PubMed Central

D’Ignazio, Laura; Batie, Michael; Rocha, Sonia

2017-01-01

Cancer is often characterised by the presence of hypoxia and inflammation. Paramount to the mechanisms controlling cellular responses under such stress stimuli, are the transcription factor families of Hypoxia Inducible Factor (HIF) and Nuclear Factor of κ-light-chain-enhancer of activated B cells (NF-κB). Although, a detailed understating of how these transcription factors respond to their cognate stimulus is well established, it is now appreciated that HIF and NF-κB undergo extensive crosstalk, in particular in pathological situations such as cancer. Here, we focus on the current knowledge on how HIF is activated by inflammation and how NF-κB is modulated by hypoxia. We summarise the evidence for the possible mechanism behind this activation and how HIF and NF-κB function impacts cancer, focusing on colorectal, breast and lung cancer. We discuss possible new points of therapeutic intervention aiming to harness the current understanding of the HIF-NF-κB crosstalk. PMID:28536364
The N-Terminus of Human Lactoferrin Displays Anti-biofilm Activity on Candida parapsilosis in Lumen Catheters.

PubMed

Fais, Roberta; Di Luca, Mariagrazia; Rizzato, Cosmeri; Morici, Paola; Bottai, Daria; Tavanti, Arianna; Lupetti, Antonella

2017-01-01

Candida parapsilosis is a major cause of hospital-acquired infection, often related to parenteral nutrition administered via catheters and hand colonization of health care workers, and its peculiar biofilm formation ability on plastic surfaces. The mortality rate of 30% points to the pressing need for new antifungal drugs. The present study aimed at analyzing the inhibitory activity of the N-terminal lactoferrin-derived peptide, further referred to as hLF 1-11, against biofilms produced by clinical isolates of C. parapsilosis characterized for their biofilm forming ability and fluconazole susceptibility. hLF 1-11 anti-biofilm activity was assessed in terms of reduction of biofilm biomass, metabolic activity, and observation of sessile cell morphology on polystyrene microtiter plates and using an in vitro model of catheter-associated C. parapsilosis biofilm production. Moreover, fluctuation in transcription levels of genes related to cell adhesion, hyphal development and extracellular matrix production upon peptide exposure were evaluated by quantitative real time RT-PCR. The results revealed that hLF 1-11 exhibits an inhibitory effect on biofilm formation by all the C. parapsilosis isolates tested, in a dose-dependent manner, regardless of their fluconazole susceptibility. In addition, hLF 1-11 induced a statistically significant dose-dependent reduction of preformed-biofilm cellular density and metabolic activity at high peptide concentrations only. Interestingly, when assessed in a catheter lumen, hLF 1-11 was able to induce a 2-log reduction of sessile cell viability at both the peptide concentrations used in RPMI diluted in NaPB. A more pronounced anti-biofilm effect was observed (3.5-log reduction) when a 10% glucose solution was used as experimental condition on both early and preformed C. parapsilosis biofilm. Quantitative real time RT-PCR experiments confirmed that hLF 1-11 down-regulates key biofilm related genes. The overall findings suggest hLF 1-11 as a
The N-Terminus of Human Lactoferrin Displays Anti-biofilm Activity on Candida parapsilosis in Lumen Catheters

PubMed Central

Fais, Roberta; Di Luca, Mariagrazia; Rizzato, Cosmeri; Morici, Paola; Bottai, Daria; Tavanti, Arianna; Lupetti, Antonella

2017-01-01

Candida parapsilosis is a major cause of hospital-acquired infection, often related to parenteral nutrition administered via catheters and hand colonization of health care workers, and its peculiar biofilm formation ability on plastic surfaces. The mortality rate of 30% points to the pressing need for new antifungal drugs. The present study aimed at analyzing the inhibitory activity of the N-terminal lactoferrin-derived peptide, further referred to as hLF 1-11, against biofilms produced by clinical isolates of C. parapsilosis characterized for their biofilm forming ability and fluconazole susceptibility. hLF 1-11 anti-biofilm activity was assessed in terms of reduction of biofilm biomass, metabolic activity, and observation of sessile cell morphology on polystyrene microtiter plates and using an in vitro model of catheter-associated C. parapsilosis biofilm production. Moreover, fluctuation in transcription levels of genes related to cell adhesion, hyphal development and extracellular matrix production upon peptide exposure were evaluated by quantitative real time RT-PCR. The results revealed that hLF 1-11 exhibits an inhibitory effect on biofilm formation by all the C. parapsilosis isolates tested, in a dose-dependent manner, regardless of their fluconazole susceptibility. In addition, hLF 1-11 induced a statistically significant dose-dependent reduction of preformed-biofilm cellular density and metabolic activity at high peptide concentrations only. Interestingly, when assessed in a catheter lumen, hLF 1-11 was able to induce a 2-log reduction of sessile cell viability at both the peptide concentrations used in RPMI diluted in NaPB. A more pronounced anti-biofilm effect was observed (3.5-log reduction) when a 10% glucose solution was used as experimental condition on both early and preformed C. parapsilosis biofilm. Quantitative real time RT-PCR experiments confirmed that hLF 1-11 down-regulates key biofilm related genes. The overall findings suggest hLF 1-11 as a
Hydroquinone stimulates inflammatory functions in microvascular endothelial cells via NF-κB nuclear activation.

PubMed

Hebeda, Cristina Bichels; Pinedo, Fernanda Júdice; Vinolo, Marco Aurélio Ramirez; Curi, Rui; Farsky, Sandra Helena Poliselli

2011-11-01

Hydroquinone impairs several leucocyte cell functions, which alter the immune response. Although endothelial cell functions are important for the development of immune responses, hydroquinone actions on endothelial cell have not been shown. Therefore, the effect of hydroquinone exposure (10 or 100 μM for 2 hr) on primary culture of microvascular endothelial cells (PMECs) obtained from the cremaster muscle of Wistar rats incubated in the presence or absence of lipopolysaccharide (LPS, 2 μg/mL) was investigated. Hydroquinone treatment induced the membrane expression of cell adhesion molecules (CAMs) from the immunoglobulin superfamilies ICAM-1 (intercellular), VCAM-1(vascular) and PECAM-1 (platelet endothelial) and induced the secretion of cytokines interleukin-1β (IL-1β) and tumour necrosis factor-α (TNF-α). The effects were dependent on transcriptional modifications because enhanced CAM mRNA expression as well as both cytokines and nuclear factor κB (NF-κB) nuclear activation was found. These effects may be due to the direct action of hydroquinone rather than its quinone metabolites, because endothelial cells do not present myeloperoxidase enzyme and hydroquinone incubation did not induce the expression of cytochrome P450 2E1 (CYP2E1) or prostaglandin H synthase 1. In addition, the incubation of endothelial cells with benzoquinone (10 μM, 2 hr) impaired PECAM-1 expression and did not modify NF-κB nuclear activation. Taken together, the data herein presented reveal that hydroquinone evokes pro-inflammatory properties in endothelial cells that are triggered by the enhancement of NF-κB nuclear translocation-dependent gene transcription. © 2011 The Authors. Basic & Clinical Pharmacology & Toxicology © 2011 Nordic Pharmacological Society.
Cardiac-specific overexpression of catalase prevents diabetes-induced pathological changes by inhibiting NF-κB signaling activation in the heart.

PubMed

Cong, Weitao; Ruan, Dandan; Xuan, Yuanhu; Niu, Chao; Tao, Youli; Wang, Yang; Zhan, Kungao; Cai, Lu; Jin, Litai; Tan, Yi

2015-12-01

Catalase is an antioxidant enzyme that specifically catabolizes hydrogen peroxide (H2O2). Overexpression of catalase via a heart-specific promoter (CAT-TG) was reported to reduce diabetes-induced accumulation of reactive oxygen species (ROS) and further prevent diabetes-induced pathological abnormalities, including cardiac structural derangement and left ventricular abnormity in mice. However, the mechanism by which catalase overexpression protects heart function remains unclear. This study found that activation of a ROS-dependent NF-κB signaling pathway was downregulated in hearts of diabetic mice overexpressing catalase. In addition, catalase overexpression inhibited the significant increase in nitration levels of key enzymes involved in energy metabolism, including α-oxoglutarate dehydrogenase E1 component (α-KGD) and ATP synthase α and β subunits (ATP-α and ATP-β). To assess the effects of the NF-κB pathway activation on heart function, Bay11-7082, an inhibitor of the NF-κB signaling pathway, was injected into diabetic mice, protecting mice against the development of cardiac damage and increased nitrative modifications of key enzymes involved in energy metabolism. In conclusion, these findings demonstrated that catalase protects mouse hearts against diabetic cardiomyopathy, partially by suppressing NF-κB-dependent inflammatory responses and associated protein nitration. Copyright © 2015 Elsevier Ltd. All rights reserved.
Effects of lactoferrin derived peptides on simulants of biological warfare agents.

PubMed

Sijbrandij, Tjitske; Ligtenberg, Antoon J; Nazmi, Kamran; Veerman, Enno C I; Bolscher, Jan G M; Bikker, Floris J

2017-01-01

Lactoferrin (LF) is an important immune protein in neutrophils and secretory fluids of mammals. Bovine LF (bLF) harbours two antimicrobial stretches, lactoferricin and lactoferampin, situated in close proximity in the N1 domain. To mimic these antimicrobial domain parts a chimeric peptide (LFchimera) has been constructed comprising parts of both stretches (LFcin17-30 and LFampin265-284). To investigate the potency of this construct to combat a set of Gram positive and Gram negative bacteria which are regarded as simulants for biological warfare agents, the effect on bacterial killing, membrane permeability and membrane polarity were determined in comparison to the constituent peptides and the native bLF. Furthermore we aimed to increase the antimicrobial potency of the bLF derived peptides by cationic amino acid substitutions. Overall, the bactericidal activity of the peptides could be related to membrane disturbing effects, i.e. membrane permeabilization and depolarization. Those effects were most prominent for the LFchimera. Arginine residues were found to be crucial for displaying antimicrobial activity, as lysine to arginine substitutions resulted in an increased antimicrobial activity, affecting mostly LFampin265-284 whereas arginine to lysine substitutions resulted in a decreased bactericidal activity, predominantly in case of LFcin17-30.
Pathogenetic Importance and Therapeutic Implications of NF-κB in Lymphoid Malignancies

PubMed Central

Lim, Kian-Huat; Yang, Yibin; Staudt, Louis M.

2014-01-01

Summary Derangement of the nuclear factor κB (NF-κB) pathway initiates and/or sustains many types of human cancer. B-cell malignancies are particularly affected by oncogenic mutations, translocations, and copy number alterations affecting key components the NF-κB pathway, most likely owing to the pervasive role of this pathway in normal B cells. These genetic aberrations cause tumors to be ‘addicted’ to NF-κB, which can be exploited therapeutically. Since each subtype of lymphoid cancer utilizes different mechanisms to activate NF-κB, several different therapeutic strategies are needed to address this pathogenetic heterogeneity. Fortunately, a number of drugs that block signaling cascades leading to NF-κB are in early phase clinical trials, several of which are already showing activity in lymphoid malignancies. PMID:22435566
The peroxisome proliferator-activated receptor β/δ (PPARβ/δ) agonist GW501516 prevents TNF-α-induced NF-κB activation in human HaCaT cells by reducing p65 acetylation through AMPK and SIRT1.

PubMed

Barroso, Emma; Eyre, Elena; Palomer, Xavier; Vázquez-Carrera, Manuel

2011-02-15

Nuclear factor (NF)-κB is a ubiquitously expressed transcription factor controlling the expression of numerous genes involved in inflammation. The aim of this study was to evaluate whether activation of the peroxisome proliferator-activated receptor (PPAR) β/δ prevented TNF-α-induced NF-κB activation in human HaCaT keratinocytes and, if so, to determine the mechanism involved. The PPARβ/δ agonist GW501516 inhibited the increase caused by TNF-α in the mRNA levels of the NF-κB target genes interleukin 8 (IL-8), TNF-α and thymic stromal lymphopoietin (TSLP). Likewise, GW501516 prevented the increase in NF-κB DNA-binding activity observed in cells exposed to TNF-α. The reduction in NF-κB activity following GW501516 treatment in cells stimulated with TNF-α did not involve either increased IκBα protein levels or a reduction in the translocation of the p65 subunit of NF-κB. In contrast, GW501516 treatment decreased TNF-α-induced p65 acetylation. Acetylation of p65 is mainly regulated by p300, a transcriptional co-activator that binds to and acetylates p65. Of note, AMP kinase (AMPK) activation phosphorylates p300 and reduces its binding to p65. GW501516 increased AMPK phosphorylation and the subsequent p300 phosphorylation, leading to a marked reduction in the association between p65 and this transcriptional co-activator. In addition, treatment with the PPARβ/δ agonist increased SIRT1 protein levels. Finally, the reduction in IL-8 mRNA levels following GW501516 treatment in TNF-α-stimulated cells was abolished in the presence of the PPARβ/δ antagonist GSK0660, the AMPK inhibitor compound C and the SIRT1 inhibitor sirtinol, indicating that the effects of GW501516 on NF-κB activity were dependent on PPARβ/δ, AMPK and SIRT1, respectively. Copyright Â© 2010 Elsevier Inc. All rights reserved.
MHY884, a newly synthesized tyrosinase inhibitor, suppresses UVB-induced activation of NF-κB signaling pathway through the downregulation of oxidative stress.

PubMed

Choi, Yeon Ja; Uehara, Yohei; Park, Ji Young; Kim, Seong Jin; Kim, So Ra; Lee, Hee Won; Moon, Hyung Ryong; Chung, Hae Young

2014-03-01

The skin is the primary target of prolonged and repeated ultraviolet (UVB) irradiation which induces cutaneous inflammation and pigmentation. Nuclear factor κB (NF-κB) is the major factor mediating UVB-induced inflammatory responses through the expression of various proinflammatory proteins such as inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2). We have previously reported that the synthetic novel compound 4-(5-chloro-2,3-dihydrobenzo[d]thiazol-2-yl)-2,6-dimethoxyphenol (MHY884) strongly suppressed tyrosinase activity and melanin synthesis in B16F10 melanoma cells. In the present study, we investigated the effect of MHY884 on the inhibition of UVB-induced NF-κB activation and its proinflammatory downstream proteins through the suppression of oxidative stress in an in vivo model of photoaging. Generation of reactive oxygen species (ROS) and peroxynitrite was measured in vitro and in B16F10 melanoma cells to verify the scavenging activity of MHY884. MHY884 suppressed oxidative stress both in vitro and in the melanoma cells in a dose-dependent manner. Next, melanin-possessing hairless mice were pre-treated with MHY884 and then irradiated with UVB repeatedly. Topical application of MHY884 attenuated UVB-induced oxidative stress, resulting in reduced NF-κB activity. Pre-treatment with MHY884 inhibited Akt and IκB kinase α/β signaling pathways, leading to decreased translocation and phosphorylation of p65, a subunit of NF-κB. This result correlated with the expression levels of iNOS and COX-2 in the skin of MHY884-treated mice. Thus, the novel tyrosinase inhibitor MHY884 suppressed NF-κB activation signaling pathway by scavenging UVB-induced oxidative stress. The discovery of MHY884, a novel tyrosinase inhibitor that targets NF-κB signaling, is significant, because this compound is a promising protective agent against UVB-induced skin damage. Copyright © 2014 Elsevier Ltd. All rights reserved.
NF-κB and androgen receptor variant expression correlate with human BPH progression.

PubMed

Austin, David C; Strand, Douglas W; Love, Harold L; Franco, Omar E; Jang, Alex; Grabowska, Magdalena M; Miller, Nicole L; Hameed, Omar; Clark, Peter E; Fowke, Jay H; Matusik, Robert J; Jin, Ren J; Hayward, Simon W

2016-04-01

Benign prostatic hyperplasia (BPH) is a common, chronic progressive disease. Inflammation is associated with prostatic enlargement and resistance to 5α-reductase inhibitor (5ARI) therapy. Activation of the nuclear factor-kappa B (NF-κB) pathway is linked to both inflammation and ligand-independent prostate cancer progression. NF-κB activation and androgen receptor variant (AR-V) expression were quantified in transition zone tissue samples from patients with a wide range of AUASS from incidental BPH in patients treated for low grade, localized peripheral zone prostate cancer to advanced disease requiring surgical intervention. To further investigate these pathways, human prostatic stromal and epithelial cell lines were transduced with constitutively active or kinase dead forms of IKK2 to regulate canonical NF-κB activity. The effects on AR full length (AR-FL) and androgen-independent AR-V expression as well as cellular growth and differentiation were assessed. Canonical NF-κB signaling was found to be upregulated in late versus early stage BPH, and to be strongly associated with non-insulin dependent diabetes mellitus. Elevated expression of AR-variant 7 (AR-V7), but not other AR variants, was found in advanced BPH samples. Expression of AR-V7 significantly correlated with the patient AUASS and TRUS volume. Forced activation of canonical NF-κB in human prostatic epithelial and stromal cells resulted in elevated expression of both AR-FL and AR-V7, with concomitant ligand-independent activation of AR reporters. Activation of NF-κB and over expression of AR-V7 in human prostatic epithelial cells maintained cell viability in the face of 5ARI treatment. Activation of NF-κB and AR-V7 in the prostate is associated with increased disease severity. AR-V7 expression is inducible in human prostate cells by forced activation of NF-κB resulting in resistance to 5ARI treatment, suggesting a potential mechanism by which patients may become resistant to 5ARI therapy. �
Cold induces reactive oxygen species production and activation of the NF-kappa B response in endothelial cells and inflammation in vivo.

PubMed

Awad, E M; Khan, S Y; Sokolikova, B; Brunner, P M; Olcaydu, D; Wojta, J; Breuss, J M; Uhrin, P

2013-09-01

Organs intended for transplantation are generally stored in the cold for better preservation of their function. However, following transplantation and reperfusion, the microvasculature of transplanted organs often proves to be activated. Extensive leukocyte adhesion and microthrombus formation contribute to failure of the transplanted organ. In this study we analyzed cold-induced changes to the activation status of cultured endothelial cells, possibly contributing to organ failure. We exposed human umbilical vein endothelial cells (HUVECs) to temperatures below 37 °C (mostly to 8 °C) for 30 min and upon rewarming to 37 °C kept incubating them for up to 24 h. We also in vivo locally exposed mice to cold. The exposure to low temperatures induced, in HUVECs, expression of the prothrombotic factors plasminogen activator inhibitor-1 (PAI-1) and tissue factor (TF) and of the inflammatory adhesion molecules, E-selectin, intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1). Furthermore, upon rewarming for 30 min, we detected activation of the inflammatory NF-κB pathway, as measured by transient NF-κB translocation to the nucleus and IκBα degradation. Using butylated hydroxytoluene (BHT), a scavenger of reactive oxygen species (ROS), we further demonstrated that cold-induced NF-κB activation depends on ROS production. Local exposure to cold also, in vivo, induced ROS production and ICAM-1 expression and resulted in leukocyte infiltration. Our results point to a causative link between ROS production and NF-κB activation, suppression of which had been shown to be beneficial during hypothermic storage and subsequent rewarming of organs for transplantation. © 2013 International Society on Thrombosis and Haemostasis.
Licochalcone A induces apoptosis in KB human oral cancer cells via a caspase-dependent FasL signaling pathway

PubMed Central

KIM, JAE-SUNG; PARK, MI-RA; LEE, SOOK-YOUNG; KIM, DO KYOUNG; MOON, SUNG-MIN; KIM, CHUN SUNG; CHO, SEUNG SIK; YOON, GOO; IM, HEE-JEONG; YOU, JAE-SEEK; OH, JI-SU; KIM, SU-GWAN

2014-01-01

Licochalcone A (Lico-A) is a natural phenol licorice compound with multiple bioactivities, including anti-inflammatory, anti-microbial, anti-fungal and osteogenesis-inducing properties. In the present study, we investigated the Lico-A-induced apoptotic effects and examined the associated apoptosis pathway in KB human oral cancer cells. Lico-A decreased the number of viable KB oral cancer cells. However, Lico-A did not have an effect on primary normal human oral keratinocytes. In addition, the IC50 value of Lico-A was determined to be ~50 μM following dose-dependent stimulation. KB oral cancer cells stimulated with Lico-A for 24 h showed chromatin condensation by DAPI staining, genomic DNA fragmentation by agarose gel electrophoresis and a gradually increased apoptotic cell population by FACS analysis. These data suggest that Lico-A induces apoptosis in KB oral cancer cells. Additionally, Lico-A-induced apoptosis in KB oral cancer cells was mediated by the expression of factor associated suicide ligand (FasL) and activated caspase-8 and −3 and poly(ADP-ribose) polymerase (PARP). Furthermore, in the KB oral cancer cells co-stimulation with a caspase inhibitor (Z-VAD-fmk) and Lico-A significantly abolished the apoptotic phenomena. Our findings demonstrated that Lico-A-induced apoptosis in KB oral cancer cells involves the extrinsic apoptotic signaling pathway, which involves a caspase-dependent FasL-mediated death receptor pathway. Our data suggest that Lico-A be developed as a chemotherapeutic agent for the management of oral cancer. PMID:24337492
NF-κB Regulates Caspase-4 Expression and Sensitizes Neuroblastoma Cells to Fas-Induced Apoptosis

PubMed Central

Yang, Hai-Jie; Wang, Mian; Wang, Lei; Cheng, Bin-Feng; Lin, Xiao-Yu; Feng, Zhi-Wei

2015-01-01

Found in neurons and neuroblastoma cells, Fas-induced apoptosis and accompanied activation of NF-κB signaling were thought to be associated with neurodegenerative diseases. However, the detailed functions of NF-κB activation in Fas killing and the effect of NF-κB activation on its downstream events remain unclear. Here, we demonstrated that agonistic Fas antibody induces cell death in a dose-dependent way and NF-κB signaling is activated as well, in neuroblastoma cells SH-EP1. Unexpectedly, NF-κB activation was shown to be pro-apoptotic, as suggested by the reduction of Fas-induced cell death with either a dominant negative form of IκBα (DN-IκBα) or an IκB kinase-specific inhibitor. To our interest, when analyzing downstream events of NF-κB signaling, we found that DN-IκBα only suppressed the expression of caspase-4, but not other caspases. Vice versa, enhancement of NF-κB activity by p65 (RelA) overexpression increased the expression of caspase-4 at both mRNA and protein levels. More directly, results from dual luciferase reporter assay demonstrated the regulation of caspase-4 promoter activity by NF-κB. When caspase-4 activity was blocked by its dominant negative (DN) form, Fas-induced cell death was substantially reduced. Consistently, the cleavage of PARP and caspase-3 induced by Fas was also reduced. In contrast, the cleavage of caspase-8 remained unaffected in caspase-4 DN cells, although caspase-8 inhibitor could rescue Fas-induced cell death. Collectively, these data suggest that caspase-4 activity is required for Fas-induced cell apoptosis and caspase-4 may act upstream of PARP and caspase-3 and downstream of caspase-8. Overall, we demonstrate that NF-κB can mediate Fas-induced apoptosis through caspase-4 protease, indicating that caspase-4 is a new mediator of NF-κB pro-apoptotic pathway in neuroblastoma cells. PMID:25695505

Some links on this page may take you to non-federal websites. Their policies may differ from this site.