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Sample records for factor-alpha induced expression

  1. Analysis of recombinant human tumour necrosis factor-alpha-induced CD4 expression on human eosinophils.

    PubMed Central

    Hossain, M; Okubo, Y; Horie, S; Sekiguchi, M

    1996-01-01

    We examined the hypothesis that one of the pro-inflammatory cytokines, tumour necrosis factor-alpha (TNF-alpha), could induce expression of the adhesion molecule CD4 on human eosinophils. We further examined the effector function of CD4 and the mechanisms regulating CD4 expression. Human eosinophils were cultured with various concentrations of recombinant human TNF-alpha (rhTNF-alpha) with or without various drugs for 24 hr. After culture, eosinophils were stained for CD4 using a monoclonal antibody and then analysed by flow cytometry. Eosinophil-derived neurotoxin (EDN) release as eosinophil degranulation was examined by cross-linking of CD4 on eosinophils. The rhTNF-alpha induced CD4 expression on human eosinophils in a dose- and time-dependent fashion; rhTNF-alpha-induced CD4 expression was significantly inhibited by 10(-6) M cycloheximide, 10(-8) M dexamethasone, or 10(-6) M herbimycin A. Recombinant human interferon-gamma inhibited rhTNF-alpha-induced CD4 expression in a dose-dependent manner. However, cross-linking of CD4 on eosinophils did not evoke EDN release, suggesting that newly expressed CD4 molecules on human eosinophils do not play any role in triggering degranulation. Our data indicate that TNF-alpha-induced CD4 expression on human eosinophils is dependent on protein synthesis and may be dependent on tyrosine kinase activity. PMID:8690465

  2. Grepafloxacin inhibits tumor necrosis factor-alpha-induced interleukin-8 expression in human airway epithelial cells.

    PubMed

    Hashimoto, S; Matsumoto, K; Gon, Y; Maruoka, S; Hayashi, S; Asai, Y; Machino, T; Horie, T

    2000-01-01

    We examined the effect of grepafloxacin (GPFX), a new fluoroquinolone antimicrobial agent, on interleukin-8 (IL-8) expression in tumor necrosis factor-alpha (TNF-alpha)-stimulated human airway epithelial cells (AEC). GPFX inhibited IL-8 protein production as well as mRNA expression in a concentration-dependent manner (2.5 - 25 micro g/ml), but the inhibition of IL-8 expression by corresponding concentrations of GPFX to serum and airway lining fluids was not complete. We discuss the modulatory effect of GPFX on IL-8 production in the context of its efficacy on controlling chronic airway inflammatory diseases.

  3. Transfection of influenza A virus nuclear export protein induces the expression of tumor necrosis factor alpha.

    PubMed

    Lara-Sampablo, Alejandra; Flores-Alonso, Juan Carlos; De Jesús-Ortega, Nereyda; Santos-López, Gerardo; Vallejo-Ruiz, Verónica; Rosas-Murrieta, Nora; Reyes-Carmona, Sandra; Herrera-Camacho, Irma; Reyes-Leyva, Julio

    2014-06-24

    Influenza A virus genomic segments eight codes for non-structural 1 (NS1) protein that is involved in evasion of innate antiviral response, and nuclear export protein (NEP) that participates in the export of viral ribonucleoprotein (RNP) complexes, transcription and replication. Tumor necrosis factor alpha (TNF-α) is highly expressed during influenza virus infections and is considered an anti-infective cytokine. NS1 and NEP proteins were overexpressed and their role on TNF-α expression was evaluated. Both TNF-α mRNA and protein increased in cells transfected with NEP but not with NS1. We further investigate if NS1 or NEP regulates the activity of TNF-α promoter. In the presence of NEP the activity of TNF-α promoter increased significantly compared with the control (83.5±2.9 vs. 30.9±2.8, respectively; p=0.001). This effect decreased 15-fold when the TNF-α promoter distal region was deleted, suggesting the involvement of mitogen-activated protein kinases (MAPK) and NF-kB response elements. This was corroborated by testing the effect produced on TNF-α promoter by the treatment with Raf/MEK/ERK (U0126), NF-kB (Bay-11-7082) and PI3K (Ly294-002) cell signaling inhibitors. Treatment with U0126 and Bay-117082 reduced the activity of TNF-α promoter mediated by NEP (41.5±3.2, 70% inhibition; and 80.6±7.4, 35% inhibition, respectively) compared to mock-treated control. The results suggest a new role for NEP protein that participates in the transcriptional regulation of human TNF-α expression.

  4. Pamidronate Down-regulates Tumor Necrosis Factor-alpha Induced Matrix Metalloproteinases Expression in Human Intervertebral Disc Cells

    PubMed Central

    Kang, Young-Mi; Hong, Seong-Hwan; Yang, Jae-Ho; Oh, Jin-Cheol; Park, Jin-Oh; Lee, Byung Ho; Lee, Sang-Yoon; Kim, Hak-Sun; Lee, Hwan-Mo

    2016-01-01

    Background N-containing bisphosphonates (BPs), such as pamidronate and risedronate, can inhibit osteoclastic function and reduce osteoclast number by inducing apoptotic cell death in osteoclasts. The aim of this study is to demonstrate the effect of pamidronate, second generation nitrogen-containing BPs and to elucidate matrix metallo-proteinases (MMPs) mRNA expression under serum starvation and/or tumor necrosis factor alpha (TNF-α) stimulation on metabolism of intervertebral disc (IVD) cells in vitro. Methods Firstly, to test the effect of pamidronate on IVD cells in vitro, various concentrations (10-12, 10-10, 10-8, and 10-6 M) of pamidronate were administered to IVD cells. Then DNA and proteoglycan synthesis were measured and messenger RNA (mRNA) expressions of type I collagen, type II collagen, and aggrecan were analyzed. Secondly, to elucidate the expression of MMPs mRNA in human IVD cells under the lower serum status, IVD cells were cultivated in full serum or 1% serum. Thirdly, to elucidate the expression of MMPs mRNA in IVD cells under the stimulation of 1% serum and TNF-α (10 ng/mL) In this study, IVD cells were cultivated in three dimensional alginate bead. Results Under the lower serum culture, IVD cells in alginate beads showed upregulation of MMP 2, 3, 9, 13 mRNA. The cells in lower serum and TNF-α also demonstrated upregulation of MMP-2, 3, 9, and 13 mRNA. The cells with various doses of pamidronate and lower serum and TNF-α were reveled partial down-regulation of MMPs. Conclusions Pamidronate, N-containing second generation BPs, was safe in metabolism of IVD in vitro maintaining chondrogenic phenotype and matrix synthesis, and down-regulated TNF-α induced MMPs expression. PMID:27622181

  5. The coffee diterpene kahweol inhibits tumor necrosis factor-{alpha}-induced expression of cell adhesion molecules in human endothelial cells

    SciTech Connect

    Kim, Hyung Gyun; Kim, Ji Young; Hwang, Yong Pil; Lee, Kyung Jin; Lee, Kwang Youl; Kim, Dong Hee; Kim, Dong Hyun; Jeong, Hye Gwang . E-mail: hgjeong@chosun.ac.kr

    2006-12-15

    Endothelial cells produce adhesion molecules after being stimulated with various inflammatory cytokines. These adhesion molecules play an important role in the development of atherogenesis. Recent studies have highlighted the chemoprotective and anti-inflammatory effects of kahweol, a coffee-specific diterpene. This study examined the effects of kahweol on the cytokine-induced monocyte/human endothelial cell interaction, which is a crucial early event in atherogenesis. Kahweol inhibited the adhesion of TNF{alpha}-induced monocytes to endothelial cells and suppressed the TNF{alpha}-induced protein and mRNA expression of the cell adhesion molecules, VCAM-1 and ICAM-1. Furthermore, kahweol inhibited the TNF{alpha}-induced JAK2-PI3K/Akt-NF-{kappa}B activation pathway in these cells. Overall, kahweol has anti-inflammatory and anti-atherosclerotic activities, which occurs partly by down-regulating the pathway that affects the expression and interaction of the cell adhesion molecules on endothelial cells.

  6. Ultraviolet radiation-induced tumor necrosis factor alpha, which is linked to the development of cutaneous SCC, modulates differential epidermal microRNAs expression

    PubMed Central

    Singh, Ashok; Willems, Estelle; Singh, Anupama; Hafeez, Bilal Bin; Ong, Irene M.; Mehta, Suresh L.; Verma, Ajit Kumar

    2016-01-01

    Chronic exposure to ultraviolet radiation (UVR) is linked to the development of cutaneous squamous cell carcinoma (SCC), a non-melanoma form of skin cancer that can metastasize. Tumor necrosis factor-alpha (TNFα), a pro-inflammatory cytokine, is linked to UVR-induced development of SCC. To find clues about the mechanisms by which TNFα may promote UVR-induced development of SCC, we investigated changes in the expression profiling of microRNAs (miRNA), a novel class of short noncoding RNAs, which affects translation and stability of mRNAs. In this experiment, TNFα knockout (TNFα KO) mice and their wild type (WT) littermates were exposed to acute UVR (2.0 kJ/m2) and the expression profiling of epidermal miRNA was determined 4hr post UVR exposure. TNFα deletion in untreated WT mice resulted in differential expression (log fold change>1) of seventeen miRNA. UVR exposure in WT mice induced differential expression of 22 miRNA. However, UVR exposure in TNFα KO mice altered only two miRNAs. Four miRNA, were differentially expressed between WT+UVR and TNFα KO+UVR groups. Differentially expressed selected miRNAs were further validated using real time PCR. Few of the differentially expressed miRNAs (miR-31-5p, miR-196a-5p, miR-127-3p, miR-206-3p, miR-411-5p, miR-709, and miR-322-5p) were also observed in UVR-induced SCC. Finally, bio-informatics analysis using DIANA, MIRANDA, Target Scan, and miRDB algorithms revealed a link with major UVR-induced pathways (MAPK, PI3K-Akt, transcriptional mis-regulation, Wnt, and TGF-beta). PMID:26918454

  7. Ultraviolet radiation-induced tumor necrosis factor alpha, which is linked to the development of cutaneous SCC, modulates differential epidermal microRNAs expression.

    PubMed

    Singh, Ashok; Willems, Estelle; Singh, Anupama; Hafeez, Bilal Bin; Ong, Irene M; Mehta, Suresh L; Verma, Ajit Kumar

    2016-04-05

    Chronic exposure to ultraviolet radiation (UVR) is linked to the development of cutaneous squamous cell carcinoma (SCC), a non-melanoma form of skin cancer that can metastasize. Tumor necrosis factor-alpha (TNFα), a pro-inflammatory cytokine, is linked to UVR-induced development of SCC. To find clues about the mechanisms by which TNFα may promote UVR-induced development of SCC, we investigated changes in the expression profiling of microRNAs (miRNA), a novel class of short noncoding RNAs, which affects translation and stability of mRNAs. In this experiment, TNFα knockout (TNFα KO) mice and their wild type (WT) littermates were exposed to acute UVR (2.0 kJ/m2) and the expression profiling of epidermal miRNA was determined 4hr post UVR exposure. TNFα deletion in untreated WT mice resulted in differential expression (log fold change>1) of seventeen miRNA. UVR exposure in WT mice induced differential expression of 22 miRNA. However, UVR exposure in TNFα KO mice altered only two miRNAs. Four miRNA, were differentially expressed between WT+UVR and TNFα KO+UVR groups. Differentially expressed selected miRNAs were further validated using real time PCR. Few of the differentially expressed miRNAs (miR-31-5p, miR-196a-5p, miR-127-3p, miR-206-3p, miR-411-5p, miR-709, and miR-322-5p) were also observed in UVR-induced SCC. Finally, bio-informatics analysis using DIANA, MIRANDA, Target Scan, and miRDB algorithms revealed a link with major UVR-induced pathways (MAPK, PI3K-Akt, transcriptional mis-regulation, Wnt, and TGF-beta).

  8. Apoptosis induced by tumor necrosis factor-alpha in rat hepatocyte cell lines expressing hepatitis B virus.

    PubMed Central

    Guilhot, S.; Miller, T.; Cornman, G.; Isom, H. C.

    1996-01-01

    Three well differentiated SV40-immortalized rat hepatocyte cell lines, CWSV1, CWSV2, and CWSV14, and Hepatitis B Virus (HBV)-producing cell lines derived from them were examined for sensitivity to tumor necrosis factor (TNF)-alpha. CWSV1, CWSV2, and CWSV14 cells were co-transfected with a DNA construct containing a dimer of the HBV genome and the neo gene and selected in G418 to generate stable cell lines. Characterization of these cell lines indicated that they contain integrated HBV DNA, contain low molecular weight HBV DNA compatible with the presence of HBV replication intermediates, express HBV transcripts, and produce HBV proteins. The viability of CWSV1, CWSV2, and CWSV2 cells was not significantly altered when they were treated with TNF-alpha at concentrations as high as 20,000 U/ml. The HBV-expressing CWSV1 cell line, SV1di36, and the HBV-expressing CWSV14 cell line, SV14di208, were also not killed when treated with TNF-alpha. However, the HBV-expressing CWSV2 cell line, SV2di366, was extensively killed when treated with TNF-alpha at concentrations ranging from 200 to 20,000 U/ml. Analysis of several different HBV-producing CWSV2 cell lines indicated that TNF-alpha killing depended upon the level of HBV expression. The TNF-alpha-induced cell killing in high HBV-producing CWSV2 cell lines was accompanied by the presence of an oligonucleosomal DNA ladder characteristic of apoptosis. Images Figure 2 Figure 3 Figure 4 Figure 6 Figure 9 Figure 10 Figure 11 PMID:8774135

  9. Tumour necrosis factor-alpha-induced protein 8 (TNFAIP8) expression associated with cell survival and death in cancer cell lines infected with canine distemper virus.

    PubMed

    Garcia, J A; Ferreira, H L; Vieira, F V; Gameiro, R; Andrade, A L; Eugênio, F R; Flores, E F; Cardoso, T C

    2015-09-16

    Oncolytic virotherapy is a novel strategy for treatment of cancer in humans and companion animals as well. Canine distemper virus (CDV), a paramyxovirus, has proven to be oncolytic through induction of apoptosis in canine-derived tumour cells, yet the mechanism behind this inhibitory action is poorly understood. In this study, three human mammary tumour cell lines and one canine-derived adenofibrosarcoma cell line were tested regarding to their susceptibility to CDV infection, cell proliferation, apoptosis, mitochondrial membrane potential and expression of tumour necrosis factor-alpha-induced protein 8 (TNFAIP8). CDV replication-induced cytopathic effect, decrease of cell proliferation rates, and >45% of infected cells were considered death and/or under late apoptosis/necrosis. TNFAIP8 and CDVM gene expression were positively correlated in all cell lines. In addition, mitochondrial membrane depolarization was associated with increase in virus titres (p < 0.005). Thus, these results strongly suggest that both human and canine mammary tumour cells are potential candidates for studies concerning CDV-induced cancer therapy.

  10. Haemophilus ducreyi lipooligosaccharides induce expression of the immunosuppressive enzyme indoleamine 2,3-dioxygenase via type I interferons and tumor necrosis factor alpha in human dendritic cells.

    PubMed

    Li, Wei; Katz, Barry P; Spinola, Stanley M

    2011-08-01

    Haemophilus ducreyi causes chancroid, a genital ulcer disease. In human inoculation experiments, most volunteers fail to clear the bacteria despite the infiltration of innate and adaptive immune cells to the infected sites. The immunosuppressive protein indoleamine 2,3-dioxygenase (IDO) is a rate-limiting enzyme in the L-tryptophan-kynurenine metabolic pathway. Tryptophan depletion and tryptophan metabolites contribute to pathogen persistence by inhibiting T cell proliferation, inducing T cell apoptosis, and promoting the expansion of FOXP3(+) regulatory T (Treg) cells. We previously found that FOXP3(+) Treg cells are enriched in experimental lesions and that H. ducreyi induced IDO transcription in dendritic cells (DC) derived from blood of infected volunteers who developed pustules. Here, we showed that enzymatically active IDO was induced in DC by H. ducreyi. Neutralizing antibodies against interferon alpha/beta receptor 2 chain (IFNAR2) and tumor necrosis factor alpha (TNF-α) inhibited IDO induction. Inhibitors of the mitogen-activated protein kinase (MAPK) p38 and nuclear factor-κB (NF-κB) also inhibited IDO expression. Neither bacterial contact with nor uptake by DC was required for IDO activation. H. ducreyi culture supernatant and H. ducreyi lipooligosaccharides (LOS) induced IDO expression, which required type I interferons, TNF-α, and the three MAPK (p38, c-Jun N-terminal kinase, and extracellular signal regulated kinase) and NF-κB pathways. In addition, LOS-induced IFN-β activated the JAK-STAT pathway. Blocking the LOS/Toll-like receptor 4 (TLR4) signaling pathway greatly reduced H. ducreyi-induced IDO production. These findings indicate that H. ducreyi-induced IDO expression in DC is largely mediated by LOS via type I interferon- and TNF-α-dependent mechanisms and the MAPK, NF-κB, and JAK-STAT pathways.

  11. Respiratory epithelial cell expression of human transforming growth factor-alpha induces lung fibrosis in transgenic mice.

    PubMed Central

    Korfhagen, T R; Swantz, R J; Wert, S E; McCarty, J M; Kerlakian, C B; Glasser, S W; Whitsett, J A

    1994-01-01

    Increased production of EGF or TGF-alpha by the respiratory epithelial cells has been associated with the pathogenesis of various forms of lung injury. Growth factors and cytokines are thought to act locally, via paracrine and autocrine mechanisms, to stimulate cell proliferation and matrix deposition by interstitial lung cells resulting in pulmonary fibrosis. To test whether TGF-alpha mediates pulmonary fibrotic responses, we have generated transgenic mice expressing human TGF-alpha under control of regulatory regions of the human surfactant protein C (SP-C) gene. Human TGF-alpha mRNA was expressed in pulmonary epithelial cells in the lungs of the transgenic mice. Adult mice bearing the SP-C-TGF-alpha transgene developed severe pulmonary fibrosis. Fibrotic lesions were observed in peribronchial, peribronchiolar, and perivascular regions, as well as subjacent to pleural surfaces. Lesions consisted of fibrous tissue that included groups of epithelial cells expressing endogenous SP-C mRNA, consistent with their identification as distal respiratory epithelial cells. Peripheral fibrotic regions consisted of thickened pleura associated with extensive collagen deposition. Alveolar architecture was disrupted in the transgenic mice with loss of alveoli in the lung parenchyma. Pulmonary epithelial cell expression of TGF-alpha in transgenic mice disrupts alveolar morphogenesis and produces fibrotic lesions mediated by paracrine signaling between respiratory epithelial and interstitial cells of the lung. Images PMID:8163670

  12. Changes in gene expression of tumor necrosis factor alpha and interleukin 6 in a canine model of caerulein-induced pancreatitis.

    PubMed

    Song, Ruhui; Yu, Dohyeon; Park, Jinho

    2016-07-01

    Acute pancreatitis is an inflammatory process that frequently involves peripancreatic tissues and remote organ systems. It has high morbidity and mortality rates in both human and veterinary patients. The severity of pancreatitis is generally determined by events that occur after acinar cell injury in the pancreas, resulting in elevated levels of various proinflammatory mediators, such as interleukin (IL) 1β and 6, as well as tumor necrosis factor alpha (TNF-α). When these mediators are excessively released into the systemic circulation, severe pancreatitis occurs with systemic complications. This pathophysiological process is similar to that of sepsis; thus, there are many striking clinical similarities between patients with septic shock and those with severe acute pancreatitis. We induced acute pancreatitis using caerulein in dogs and measured the change in the gene expression of proinflammatory cytokines. The levels of TNF-α and IL-6 mRNA peaked at 3 h, at twice the baseline levels, and the serum concentrations of amylase and lipase also increased. Histopathological examination revealed severe hyperemia of the pancreas and hyperemia in the duodenal villi and the hepatic sinusoid. Thus, pancreatitis can be considered an appropriate model to better understand the development of naturally occurring sepsis and to assist in the effective treatment and management of septic patients.

  13. Changes in gene expression of tumor necrosis factor alpha and interleukin 6 in a canine model of caerulein-induced pancreatitis

    PubMed Central

    Song, Ruhui; Yu, Dohyeon; Park, Jinho

    2016-01-01

    Acute pancreatitis is an inflammatory process that frequently involves peripancreatic tissues and remote organ systems. It has high morbidity and mortality rates in both human and veterinary patients. The severity of pancreatitis is generally determined by events that occur after acinar cell injury in the pancreas, resulting in elevated levels of various proinflammatory mediators, such as interleukin (IL) 1β and 6, as well as tumor necrosis factor alpha (TNF-α). When these mediators are excessively released into the systemic circulation, severe pancreatitis occurs with systemic complications. This pathophysiological process is similar to that of sepsis; thus, there are many striking clinical similarities between patients with septic shock and those with severe acute pancreatitis. We induced acute pancreatitis using caerulein in dogs and measured the change in the gene expression of proinflammatory cytokines. The levels of TNF-α and IL-6 mRNA peaked at 3 h, at twice the baseline levels, and the serum concentrations of amylase and lipase also increased. Histopathological examination revealed severe hyperemia of the pancreas and hyperemia in the duodenal villi and the hepatic sinusoid. Thus, pancreatitis can be considered an appropriate model to better understand the development of naturally occurring sepsis and to assist in the effective treatment and management of septic patients. PMID:27408338

  14. Different tumor necrosis factor-alpha-associated leptin expression in rats with dimethylnitrosamine and bile duct ligation-induced liver cirrhosis.

    PubMed

    Lin, Shih Yi; Chen, Wen Yin; Chiu, Yung Tsung; Lee, Wen Jane; Wu, Hurng Sheng; Sheu, Wayne Huey-Herng

    2005-04-01

    Although serum leptin concentrations are reported by several studies to increase in patients with liver cirrhosis, the mechanisms underpinning this increase remain unclear. Circulating tumor necrosis factor alpha (TNF-alpha) concentrations are also recognized to increase in liver cirrhosis. Furthermore, TNF-alpha administration to rodents results in increased expression and secretion of leptin from adipose tissue in a manner dependent on type 1 TNF-alpha receptor (TNF-RI). The present study was undertaken to examine adipose leptin expression and to explore potential relationships between leptin expression and TNF-alpha in subjects with liver cirrhosis. Liver cirrhosis was induced in male Sprague-Dawley rats by dimethylnitrosamine (DMN) administration or by common bile duct ligation (BDL). Ad libitum and pair-fed animals constituted controls. Serum leptin and TNF-alpha concentrations were determined by immunoassay. Gene expression was determined by the reverse transcription-polymerase chain reaction, and protein levels were measured by Western blotting. Serum leptin values after adjustment of body fat mass in DMN-treated rats were significantly higher than in pair-fed or ad libitum groups. Leptin mRNA and protein levels in epididymal fat in DMN rats increased by 1.8-fold and 2.3-fold, respectively, as compared with ad libitum controls, and by 4-fold and 6-fold, respectively, as compared with the pair-fed group. Epididymal TNF-alpha and membranous TNF-RI (mTNF-RI) concentrations were both 2.3 times higher in DMN rats than in ad libitum controls but did not differ between ad libitum and pair-fed groups. Adipose leptin protein levels correlated directly with TNF-alpha and mTNF-RI concentrations in combined DMN, ad libitum, and pair-fed rats (r=0.64 and r=0.49, respectively; P<.05). In BDL-treated rats, however, serum and adipose leptin concentrations were identical to those in ad libitum controls despite 2.1-fold and 2.4-fold increase in epididymal TNF-alpha and m

  15. Cellular localization of interleukin-8 and its inducer, tumor necrosis factor-alpha in psoriasis.

    PubMed Central

    Nickoloff, B. J.; Karabin, G. D.; Barker, J. N.; Griffiths, C. E.; Sarma, V.; Mitra, R. S.; Elder, J. T.; Kunkel, S. L.; Dixit, V. M.

    1991-01-01

    The importance of immunologic mechanisms in psoriasis has been deduced from the ability of immunosuppressive therapies to ameliorate this common and chronic skin disease. Certainly the histology of psoriatic lesions suggests a dialogue between the hyperplastic keratinocytes and infiltrating T lymphocytes and macrophages. To begin dissecting the cytokine network involved in the pathophysiology of psoriasis, the location, in both epidermal and dermal compartments, of tumor necrosis factor-alpha, interleukin-8, intercellular adhesion molecule-1, and transforming growth factor-alpha at the protein and/or mRNA levels were identified. Tumor necrosis factor-alpha was selected as a potentially key regulatory cytokine, first because it induces cultured keratinocyte interleukin-8, intercellular adhesion molecule-1, and transforming growth factor-alpha production, and second because intercellular adhesion molecule-1 expression by keratinocytes in psoriatic epidermis had been identified previously. Using immunohistochemical localization, tumor necrosis factor-alpha was identified in 12 psoriatic lesions as intense and diffuse expression by dermal dendrocytes (macrophages) in the papillary dermis (without significant staining of endothelial cells, mast cells, or dermal Langerhans cells), and focally by keratinocytes and intraepidermal Langerhans cells. Functional interaction between the dermal dendrocytes and keratinocytes was suggested by the presence of interleukin-8 expression of suprabasal keratinocytes immediately above the tumor necrosis factor-alpha-positive dermal dendrocytes. Interleukin-8 mRNA and transforming growth factor-alpha mRNA were detectable in the epidermal roof of psoriatic lesions, but neither was detectable at the protein or mRNA levels in any normal skin specimens. Treatment of cultured human keratinocytes with phorbol ester (which experimentally produces psoriasiform changes on mouse skin) or tumor necrosis factor-alpha also increased interleukin-8 and

  16. Expression of tumor necrosis factor-alpha-induced protein 8 in pancreas tissues and its correlation with epithelial growth factor receptor levels.

    PubMed

    Liu, Ke; Qin, Cheng-Kun; Wang, Zhi-Yi; Liu, Su-Xia; Cui, Xian-Ping; Zhang, Dong-Yuan

    2012-01-01

    Tumor necrosis factor (TNF)-alpha-induced protein 8 (TNFAIP8 or TIPE) is a recently identified protein considered to be associated with carcinogenesis. To investigate its expression pattern in pancreatic cancer patients and to analyse its correlation with clinicopathological significance and the expression levels of epithelial growth factor receptor (EGFR), immunohistochemistry was performed to detect the TNFAIP8 and EGFR proteins in pancreatic cancers, pancreatitis tissues, and healthy controls. The results showed stronger staining of TNFAIP8 protein in pancreatic cancer tissues compared with normal pancreas tissue. Furthermore, in 56 patients with pancreatic cancer, the expression levels of TNFAIP8 in patients with low tumor stage was higher than that with high tumor stage, and correlated with tumor staging and lymph node metastasis (P<0.05). Furthermore, TNFAIP8 expression positively correlated with EGFR levels (r=0.671135, P<0.05). These results indicate that TNFAIP8 may play important roles in the progression of pancreatic cancer.

  17. Increased tumor necrosis factor-alpha (TNF-alpha) gene expression in parainfluenza type 1 (Sendai) virus-induced bronchiolar fibrosis.

    PubMed Central

    Uhl, E. W.; Moldawer, L. L.; Busse, W. W.; Jack, T. J.; Castleman, W. L.

    1998-01-01

    Increased airway resistance and airway hyperresponsiveness induced in rats by infection with parainfluenza type I (Sendai) virus is associated with bronchiolar fibrosis. To determine whether increased tumor necrosis factor (TNF)-alpha gene expression is an important regulatory event in virus-induced bronchiolar fibrosis, pulmonary TNF-alpha mRNA and protein expression was assessed in rat strains that are susceptible (Brown Norway; BN) and resistant (Fischer 344; F344) to virus-induced bronchiolar fibrosis. Virus-inoculated BN rats had increased TNF-alpha pulmonary mRNA levels (P < 0.05) and increased numbers of bronchiolar macrophages and fibroblasts expressing TNF-alpha protein compared with virus-inoculated F344 rats (P < 0.05). Virus inoculation also induced elevated TNF-alpha mRNA and protein levels (P < 0.05) in cultured rat alveolar macrophages (NR8383 cells). A 55-kd soluble TNF receptor-immunoglobulin G fusion protein (sTNFR-IgG) was used to inhibit TNF-alpha bioactivity in virus-inoculated BN rats. Treated rats had fewer proliferating bronchiolar fibroblasts, as detected by bromodeoxyuridine incorporation, compared with virus-inoculated control rats (P < 0.05). There was also increased mortality in p55sTNFR-IgG-treated virus-inoculated rats associated with increased viral replication and decreased numbers of macrophages and lymphocytes in bronchoalveolar lavage fluid (P < 0.05). The results of this study indicate that 1) Sendai virus can directly up-regulate TNF-alpha mRNA and protein expression in macrophages, 2) TNF-alpha is an important mediator of virus-induced bronchiolar fibrosis, and 3) TNF-alpha has a critical role in the termination of Sendai viral replication in the lung. Images Figure 2 PMID:9466578

  18. Brucella abortus Invasion of Osteocytes Modulates Connexin 43 and Integrin Expression and Induces Osteoclastogenesis via Receptor Activator of NF-κB Ligand and Tumor Necrosis Factor Alpha Secretion

    PubMed Central

    Pesce Viglietti, Ayelén Ivana; Arriola Benitez, Paula Constanza; Gentilini, María Virginia; Velásquez, Lis Noelia; Fossati, Carlos Alberto; Giambartolomei, Guillermo Hernán

    2015-01-01

    Osteoarticular brucellosis is the most common localization of human active disease. Osteocytes are the most abundant cells of bone. They secrete factors that regulate the differentiation of both osteoblasts and osteoclasts during bone remodeling. The aim of this study is to determine if Brucella abortus infection modifies osteocyte function. Our results indicate that B. abortus infection induced matrix metalloproteinase 2 (MMP-2), receptor activator for NF-κB ligand (RANKL), proinflammatory cytokines, and keratinocyte chemoattractant (KC) secretion by osteocytes. In addition, supernatants from B. abortus-infected osteocytes induced bone marrow-derived monocytes (BMM) to undergo osteoclastogenesis. Using neutralizing antibodies against tumor necrosis factor alpha (TNF-α) or osteoprotegerin (OPG), RANKL's decoy receptor, we determined that TNF-α and RANKL are involved in osteoclastogenesis induced by supernatants from B. abortus-infected osteocytes. Connexin 43 (Cx43) and the integrins E11/gp38, integrin-α, integrin-β, and CD44 are involved in cell-cell interactions necessary for osteocyte survival. B. abortus infection inhibited the expression of Cx43 but did not modify the expression of integrins. Yet the expression of both Cx43 and integrins was inhibited by supernatants from B. abortus-infected macrophages. B. abortus infection was not capable of inducing osteocyte apoptosis. However, supernatants from B. abortus-infected macrophages induced osteocyte apoptosis in a dose-dependent manner. Taken together, our results indicate that B. abortus infection could alter osteocyte function, contributing to bone damage. PMID:26459511

  19. Upregulation of heat shock protein 70 and the differential protein expression induced by tumor necrosis factor-alpha enhances migration and inhibits apoptosis of hepatocellular carcinoma cell HepG2

    PubMed Central

    Huang, Bee-Piao; Lin, Chun-Shiang; Wang, Chau-Jong; Kao, Shao-Hsuan

    2017-01-01

    Tumor necrosis factor alpha (TNFα) plays diverse roles in liver damage and hepatocarcinogenesis with its multipotent bioactivity. However, the influence of TNFα on protein expression of hepatocellular carcinoma (HCC) is incompletely understood. Therefore, we aimed to investigate the differential protein expression of HCC in response to TNFα stimulus. We observed that HepG2 cell revealed a higher resistance to TNFα-induced apoptosis as compared to the non-tumorigenic hepatocyte THLE-2. By using a label-free quantitative proteomic analysis, we found that 520 proteins were differentially expressed in the HepG2 cells exposed to TNFα, including 211 up-regulated and 309 down-regulated proteins. We further confirmed several proteins with significant expression change (TNFα/control ratio>2.0 or <0.5) by immunoblotting using specific antibodies. We also analyzed the differential expressed proteins using Gene ontology and KEGG annotations, and the results implicated that TNFα might regulate ribosome, spliceosome, antigen processing and presentation, and energy metabolism in HepG2 cells. Moreover, we demonstrated that upregulation of heat shock protein 70 (HSP70) was involved in both the promoted migration and the inhibited apoptosis of HepG2 cells in response to TNFα. Collectively, these findings indicate that TNFα alters protein expression such as HSP70, which triggering specific molecular processes and signaling cascades that promote migration and inhibit apoptosis of HepG2 cells. PMID:28367089

  20. Sirtuin1 (SIRT1) Regulates Tumor Necrosis Factor-alpha (TNF-α-Induced) Aquaporin-2 (AQP2) Expression in Renal Medullary Collecting Duct Cells Through Inhibiting the NF-κB Pathway

    PubMed Central

    Lin, Qinqin; Geng, Yuanwen; Lin, Shuaishuai; Tian, Zhenjun

    2016-01-01

    Background Aquaporin-2 (AQP2) plays a major role in water reabsorption in the renal collecting duct, and is involved in a variety of renal disease. Recent studies have indicate that sirtuin1 (SIRT1) exerts renoprotective properties against kidney diseases. This study aimed to determine the potential role of SIRT1 in AQP2 expression induced by tumor necrosis factor-alpha (TNF-α) and to disclose the underlying mechanism in renal inner medullary collecting duct (IMCD) cells. Material/Methods Quantitative real-time PCR and Western blotting were respectively identified mRNA and protein expression. Immunofluorescence staining was used to detect the localization of AQP2. Small-interfering RNA (siRNA) was carried out for mechanism study. Results Results showed that AQP2 was clearly increased in the plasma membrane and decreased in the cytoplasm of IMCD cells treated with AVP. TNF-α treatment in IMCD cells significantly reduced SIRT1 and AQP2 expression, and increased acetylated NF-κBp65 protein level in time- and concentration-dependent manners. Moreover, SIRT1 overexpression or the activator SRT1720 augmented AQP2 expression and reduced the acetylation of NF-κBp65, which was reversed by SIRT1 siRNA or the inhibitors Ex527 and sirtinol in TNF-α-induced IMCD cells. Knockdown of NF-κBp65 or NF-κBp65 inhibition by pyrrolidine dithiocarbamate (PDTC) enhanced AQP2 expression in IMCD cells exposed to TNF-α. Importantly, knockdown of NF-κBp65 augmented the up-regulation of SIRT1 on AQP2 expression in IMCD cells induced by TNF-α. Conclusions These findings indicate that SIRT1 increases AQP2 expression in TNF-α-induced IMCD cells via the NF-κB-dependent signalling pathway, which might provide novel insight to understanding the renoprotective effects of SIRT1 in kidney diseases. PMID:27980322

  1. Inhibition of tumor necrosis factor-alpha-induced interleukin-6 expression by telmisartan through cross-talk of peroxisome proliferator-activated receptor-gamma with nuclear factor kappaB and CCAAT/enhancer-binding protein-beta.

    PubMed

    Tian, Qingping; Miyazaki, Ryohei; Ichiki, Toshihiro; Imayama, Ikuyo; Inanaga, Keita; Ohtsubo, Hideki; Yano, Kotaro; Takeda, Kotaro; Sunagawa, Kenji

    2009-05-01

    Telmisartan, an angiotensin II type 1 receptor antagonist, was reported to be a partial agonist of peroxisome proliferator-activated receptor-gamma. Although peroxisome proliferator-activated receptor-gamma activators have been shown to have an anti-inflammatory effect, such as inhibition of cytokine production, it has not been determined whether telmisartan has such effects. We examined whether telmisartan inhibits expression of interleukin-6 (IL-6), a proinflammatory cytokine, in vascular smooth muscle cells. Telmisartan, but not valsartan, attenuated IL-6 mRNA expression induced by tumor necrosis factor-alpha (TNF-alpha). Telmisartan decreased TNF-alpha-induced IL-6 mRNA and protein expression in a dose-dependent manner. Because suppression of IL-6 mRNA expression was prevented by pretreatment with GW9662, a specific peroxisome proliferator-activated receptor-gamma antagonist, peroxisome proliferator-activated receptor-gamma may be involved in the process. Telmisartan suppressed IL-6 gene promoter activity induced by TNF-alpha. Deletion analysis suggested that the DNA segment between -150 bp and -27 bp of the IL-6 gene promoter that contains nuclear factor kappaB and CCAAT/enhancer-binding protein-beta sites was responsible for telmisartan suppression. Telmisartan attenuated TNF-alpha-induced nuclear factor kappaB- and CCAAT/enhancer-binding protein-beta-dependent gene transcription and DNA binding. Telmisartan also attenuated serum IL-6 level in TNF-alpha-infused mice and IL-6 production from rat aorta stimulated with TNF-alpha ex vivo. These data suggest that telmisartan may attenuate inflammatory process induced by TNF-alpha in addition to the blockade of angiotensin II type 1 receptor. Because both TNF-alpha and angiotensin II play important roles in atherogenesis through enhancement of vascular inflammation, telmisartan may be beneficial for treatment of not only hypertension but also vascular inflammatory change.

  2. Tumor Necrosis Factor Alpha and Insulin-Like Growth Factor 1 Induced Modifications of the Gene Expression Kinetics of Differentiating Skeletal Muscle Cells

    PubMed Central

    Meyer, Swanhild U.; Krebs, Stefan; Thirion, Christian; Blum, Helmut; Krause, Sabine; Pfaffl, Michael W.

    2015-01-01

    Introduction TNF-α levels are increased during muscle wasting and chronic muscle degeneration and regeneration processes, which are characteristic for primary muscle disorders. Pathologically increased TNF-α levels have a negative effect on muscle cell differentiation efficiency, while IGF1 can have a positive effect; therefore, we intended to elucidate the impact of TNF-α and IGF1 on gene expression during the early stages of skeletal muscle cell differentiation. Methodology/Principal Findings This study presents gene expression data of the murine skeletal muscle cells PMI28 during myogenic differentiation or differentiation with TNF-α or IGF1 exposure at 0 h, 4 h, 12 h, 24 h, and 72 h after induction. Our study detected significant coregulation of gene sets involved in myoblast differentiation or in the response to TNF-α. Gene expression data revealed a time- and treatment-dependent regulation of signaling pathways, which are prominent in myogenic differentiation. We identified enrichment of pathways, which have not been specifically linked to myoblast differentiation such as doublecortin-like kinase pathway associations as well as enrichment of specific semaphorin isoforms. Moreover to the best of our knowledge, this is the first description of a specific inverse regulation of the following genes in myoblast differentiation and response to TNF-α: Aknad1, Cmbl, Sepp1, Ndst4, Tecrl, Unc13c, Spats2l, Lix1, Csdc2, Cpa1, Parm1, Serpinb2, Aspn, Fibin, Slc40a1, Nrk, and Mybpc1. We identified a gene subset (Nfkbia, Nfkb2, Mmp9, Mef2c, Gpx, and Pgam2), which is robustly regulated by TNF-α across independent myogenic differentiation studies. Conclusions This is the largest dataset revealing the impact of TNF-α or IGF1 treatment on gene expression kinetics of early in vitro skeletal myoblast differentiation. We identified novel mRNAs, which have not yet been associated with skeletal muscle differentiation or response to TNF-α. Results of this study may facilitate

  3. Expression of tumour necrosis factor alpha and accumulation of fibronectin in coronary artery restenotic lesions retrieved by atherectomy.

    PubMed Central

    Clausell, N.; de Lima, V. C.; Molossi, S.; Liu, P.; Turley, E.; Gotlieb, A. I.; Adelman, A. G.; Rabinovitch, M.

    1995-01-01

    BACKGROUND--The formation of coronary artery neointima experimentally induced in piglets after cardiac transplantation is related to an immune-inflammatory reaction associated with increased expression of T cells and inflammatory mediators (tumour necrosis factor alpha and interleukin 1 beta) and upregulation of fibronectin. In vivo blockade of tumour necrosis factor alpha in rabbits after cardiac transplantation results in reduced neointimal formation. The objective of this study was to investigate the hypothesis that coronary restenosis after atherectomy or percutaneous balloon angioplasty is associated with a similar inflammatory cascade initiated by mechanical injury. METHODS--Specimens taken at coronary atherectomy were analysed from 16 patients. Nine had had the procedure performed twice, firstly, to remove a primary lesion, and secondly, to remove a restenotic lesion. Seven had percutaneous balloon angioplasty after removal of restenotic tissue. Coronary atherectomy specimens were analysed by immunohistochemistry for the presence of T cells, macrophages, major histocompatibility complex II, interleukin 1 beta, tumour necrosis factor alpha, fibronectin, and the receptor for hyaluronan mediated motility. RESULTS--The groups were clinically and angiographically similar with equivalent lumens before and after atherectomy. Restenotic lesions had increased expression of tumour necrosis factor alpha and fibronectin compared with the primary lesions (P < 0.05 for both). There was also a trend towards a greater number of T cells and increased expression of interleukin 1 beta. CONCLUSIONS--Restenosis is associated with increased expression of tumour necrosis factor alpha and fibronectin, suggesting that an immune-inflammatory reaction probably contributes to neointimal formation and may represent a form of wound healing and repair secondary to mechanical injury. Images PMID:7626352

  4. Tumor necrosis factor-{alpha} enhances IL-15-induced natural killer cell differentiation

    SciTech Connect

    Lee, Jiwon; Lee, Suk Hyung; Shin, Nara; Jeong, Mira; Kim, Mi Sun; Kim, Mi Jeong; Yoon, Suk Ran; Chung, Jin Woong; Kim, Tae-Don; Choi, Inpyo

    2009-09-04

    The differentiation of natural killer (NK) cells is regulated by various factors including soluble growth factors and transcription factors. Here, we have demonstrated that tumor necrosis factor-{alpha} (TNF-{alpha}) is a positive regulator of NK cell differentiation. TNF-{alpha} augmented the IL-15-induced expression of NK1.1 and CD122 in mature NK cells, and TNF-{alpha} alone also induced NK cell maturation as well as IL-15. TNF-{alpha} also increased IFN-{gamma} production in NK cells in the presence of IL-15. Meanwhile, mRNA expression of several transcription factors, including T-bet and GATA-3, was increased by the addition of TNF-{alpha} and IL-15. In addition, TNF-{alpha} increased nuclear factor-kappa B (NF-{kappa}B) activity in NK cells and inhibition of NF-{kappa}B impeded TNF-{alpha}-enhanced NK cell maturation. Overall, these data suggest that TNF-{alpha} significantly increased IL-15-driven NK cell differentiation by increasing the expression of transcription factors that play crucial roles in NK cell maturation and inducing the NF-{kappa}B activity.

  5. Palmitate induces tumor necrosis factor-alpha expression in C2C12 skeletal muscle cells by a mechanism involving protein kinase C and nuclear factor-kappaB activation.

    PubMed

    Jové, Mireia; Planavila, Anna; Sánchez, Rosa M; Merlos, Manuel; Laguna, Juan Carlos; Vázquez-Carrera, Manuel

    2006-01-01

    The mechanisms responsible for increased expression of TNF-alpha in skeletal muscle cells in diabetic states are not well understood. We examined the effects of the saturated acid palmitate on TNF-alpha expression. Exposure of C2C12 skeletal muscle cells to 0.75 mm palmitate enhanced mRNA (25-fold induction, P < 0.001) and protein (2.5-fold induction) expression of the proinflammatory cytokine TNF-alpha. This induction was inversely correlated with a fall in GLUT4 mRNA levels (57% reduction, P < 0.001) and glucose uptake (34% reduction, P < 0.001). PD98059 and U0126, inhibitors of the ERK-MAPK cascade, partially prevented the palmitate-induced TNF-alpha expression. Palmitate increased nuclear factor (NF)-kappaB activation and incubation of the cells with the NF-kappaB inhibitors pyrrolidine dithiocarbamate and parthenolide partially prevented TNF-alpha expression. Incubation of palmitate-treated cells with calphostin C, a strong and specific inhibitor of protein kinase C (PKC), abolished palmitate-induced TNF-alpha expression, and restored GLUT4 mRNA levels. Palmitate treatment enhanced the expression of phospho-PKCtheta, suggesting that this PKC isoform was involved in the changes reported, and coincubation of palmitate-treated cells with the PKC inhibitor chelerythrine prevented the palmitate-induced reduction in the expression of IkappaBalpha and insulin-stimulated Akt activation. These findings suggest that enhanced TNF-alpha expression and GLUT4 down-regulation caused by palmitate are mediated through the PKC activation, confirming that this enzyme may be a target for either the prevention or the treatment of fatty acid-induced insulin resistance.

  6. Dehydrocostus lactone enhances tumor necrosis factor-alpha-induced apoptosis of human leukemia HL-60 cells.

    PubMed

    Oh, G S; Pae, H O; Chung, H T; Kwon, J W; Lee, J H; Kwon, T O; Kwon, S Y; Chon, B H; Yun, Young Gab

    2004-05-01

    Sesquiterpene lactones have raised considerable interest because of their ability to block the activation of nuclear transcription factor-kappaB (NF-kappaB). NF-kappaB plays an important role in the resistance of cancer cells to the induction of apoptosis by anticancer drugs and tumor necrosis factor-alpha (TNF-alpha). Pharmacological inhibition of NF-kappaB offers the promise of enhancing the efficacy of anticancer therapies. Here, we demonstrate that dehydrocostus lactone (DL), the major sesquiterpene lactone isolated from the roots of Saussurea lappa, inhibits NF-kappaB activation by preventing TNF-alpha-induced degradation and phosphorylation of its inhibitory protein I-kappaB alpha in human leukemia HL-60 cells and that DL renders HL-60 cells susceptible to TNF-alpha-induced apoptosis by enhancing caspase-8 and caspase-3 activities.

  7. Diesel exhaust particles induce the over expression of tumor necrosis factor-alpha (TNF-alpha) gene in alvelor machrophage and failed to induce apoptosis through activation of nuclear factor-kappaB (NF-kappaB)

    EPA Science Inventory

    Exposure to particulate matter (PM2.5-10), including diesel exhaust particles (DEP) has been reported to induce lung injury and exacerbation of asthma and chronic obstructive pulmonary disease. Alveolar macrophages play a major role in the lung's response to inhaled particles and...

  8. Interleukin-6 enhances whereas tumor necrosis factor alpha and interferons inhibit integrin expression and adhesion of human mast cells to extracellular matrix proteins.

    PubMed

    Schoeler, Dagmar; Grützkau, Andreas; Henz, Beate M; Küchler, Jens; Krüger-Krasagakis, Sabine

    2003-05-01

    Integrins are expressed on mast cells and constitute an essential prerequisite for the accumulation of the cells at sites of inflammation. In order to clarify a potential contribution of inflammatory cytokines to this process, we have studied the modulation of integrin expression and adhesion of immature human mast cells (HMC-1) to extracellular matrix proteins by interleukin-6, tumor necrosis factor alpha, interferon-alpha and interferon-gamma. Corticosteroids were used for comparison. On fluorescence-activated cell sorter analysis, preincubation of cells for 48 h with different concentrations of interleukin-6 induced a significant, up to 40%, increase of alpha v alpha 5, CD49b (alpha 2), CD49e (alpha 5), CD49f (alpha 6), and CD51 (alpha v). In contrast, different concentrations of tumor necrosis factor alpha, interferon-alpha, interferon-gamma, and dexamethasone (10-8-10-10 M) inhibited expression of adhesion receptors by up to 60%, reaching significance for some but not all integrins. On semiquantitative polymerase chain reaction analysis, interleukin-6, the other cytokines, and corticosteroids significantly modulated expression of alpha1, alpha v and alpha 5 integrin chains at mRNA level. Functional significance of these findings was proven in adhesion assays using fibronectin, laminin, and vitronectin, with interleukin-6 causing significant enhancement of adhesion in all cases, tumor necrosis factor alpha and dexamethasone inducing significant reduction of adhesion to fibronectin and laminin, and interferon-gamma significantly inhibiting adhesion to fibronectin only. Specificity of interleukin-6-induced changes was demonstrated using antibodies against alpha1 and alpha 5 integrins in unstimulated and interleukin-6-prestimulated cells. These data show that interleukin-6 stimulates mast cell adhesion to extracellular matrix and thus allows for the accumulation of the cells at tissue sites by enhancing integrin expression, whereas tumor necrosis factor alpha

  9. Glutathione regulation of redox-sensitive signals in tumor necrosis factor-alpha-induced vascular endothelial dysfunction.

    PubMed

    Tsou, Tsui-Chun; Yeh, Szu Ching; Tsai, Feng-Yuan; Chen, Jein-Wen; Chiang, Huai-Chih

    2007-06-01

    We investigated the regulatory role of glutathione in tumor necrosis factor-alpha (TNF-alpha)-induced vascular endothelial dysfunction as evaluated by using vascular endothelial adhesion molecule expression and monocyte-endothelial monolayer binding. Since TNF-alpha induces various biological effects on vascular cells, TNF-alpha dosage could be a determinant factor directing vascular cells into different biological fates. Based on the adhesion molecule expression patterns responding to different TNF-alpha concentrations, we adopted the lower TNF-alpha (0.2 ng/ml) to rule out the possible involvement of other TNF-alpha-induced biological effects. Inhibition of glutathione synthesis by l-buthionine-(S,R)-sulfoximine (BSO) resulted in down-regulations of the TNF-alpha-induced adhesion molecule expression and monocyte-endothelial monolayer binding. BSO attenuated the TNF-alpha-induced nuclear factor-kappaB (NF-kappaB) activation, however, with no detectable effect on AP-1 and its related mitogen-activated protein kinases (MAPKs). Deletion of an AP-1 binding site in intercellular adhesion molecule-1 (ICAM-1) promoter totally abolished its constitutive promoter activity and its responsiveness to TNF-alpha. Inhibition of ERK, JNK, or NF-kappaB attenuates TNF-alpha-induced ICAM-1 promoter activation and monocyte-endothelial monolayer binding. Our study indicates that TNF-alpha induces adhesion molecule expression and monocyte-endothelial monolayer binding mainly via activation of NF-kappaB in a glutathione-sensitive manner. We also demonstrated that intracellular glutathione does not modulate the activation of MAPKs and/or their downstream AP-1 induced by lower TNF-alpha. Although AP-1 activation by the lower TNF-alpha was not detected in our systems, we could not rule out the possible involvement of transiently activated MAPKs/AP-1 in the regulation of TNF-alpha-induced adhesion molecule expression.

  10. Phospholipase C-{delta}{sub 1} regulates interleukin-1{beta} and tumor necrosis factor-{alpha} mRNA expression

    SciTech Connect

    Chung, Eric; Jakinovich, Paul; Bae, Aekyung; Rebecchi, Mario

    2012-10-01

    Phospholipase C-{delta}{sub 1} (PLC{delta}{sub 1}) is a widely expressed highly active PLC isoform, modulated by Ca{sup 2+} that appears to operate downstream from receptor signaling and has been linked to regulation of cytokine production. Here we investigated whether PLC{delta}{sub 1} modulated expression of the pro-inflammatory cytokines interleukin-1{beta} (IL-1{beta}), tumor necrosis factor-{alpha} (TNF-{alpha}) and interleukin-6 (IL-6) in rat C6 glioma cells. Expression of PLC{delta}{sub 1} was specifically suppressed by small interfering RNA (siRNA) and the effects on cytokine mRNA expression, stimulated by the Toll-like receptor (TLR) agonist, lipopolysaccharide (LPS), were examined. Real-time polymerase chain reaction (RT-PCR) results showed that PLC{delta}{sub 1} knockdown enhanced expression IL-1{beta} and tumor necrosis factor-{alpha} (TNF-{alpha}) mRNA by at least 100 fold after 4 h of LPS stimulation compared to control siRNA treatment. PLC{delta}{sub 1} knock down caused persistently high Nf{kappa}b levels at 4 h of LPS stimulation compared to control siRNA-treated cells. PLC{delta}{sub 1} knockdown was also associated with elevated nuclear levels of c-Jun after 30 min of LPS stimulation, but did not affect LPS-stimulated p38 or p42/44 MAPK phosphorylation, normally associated with TLR activation of cytokine gene expression; rather, enhanced protein kinase C (PKC) phosphorylation of cellular proteins was observed in the absence of LPS stimulation. An inhibitor of PKC, bisindolylmaleimide II (BIM), reversed phosphorylation, prevented elevation of nuclear c-Jun levels, and inhibited LPS-induced increases of IL-1{beta} and TNF-{alpha} mRNA's induced by PLC{delta}{sub 1} knockdown. Our results show that loss of PLC{delta}{sub 1} enhances PKC/c-Jun signaling and up-modulates pro-inflammatory cytokine gene transcription in concert with the TLR-stimulated p38MAPK/Nf{kappa}b pathway. Our findings are consistent with the idea that PLC{delta}{sub 1} is a

  11. TERATOGENIC RESPONSES ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF)

    EPA Science Inventory

    TITLE:
    TERATOGENIC RESPONSES ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF). AUTHORS (ALL): Abbott, Barbara D.1; Best, Deborah S.1; Narotsky, Michael G.1. SPONSOR NAME: None INSTITUTIONS (ALL): 1. Repro Tox ...

  12. Cinnamaldehyde inhibits the tumor necrosis factor-alpha-induced expression of cell adhesion molecules in endothelial cells by suppressing NF-kappaB activation: effects upon IkappaB and Nrf2.

    PubMed

    Liao, Being-Chyuan; Hsieh, Chia-Wen; Liu, Yen-Chin; Tzeng, Tsai-Teng; Sun, Yung-Wei; Wung, Being-Sun

    2008-06-01

    The production of adhesion molecules and subsequent attachment of leukocytes to endothelial cells (ECs) are critical early events in atherogenesis. These adhesion molecules thus play an important role in the development of this disease. Recent studies have highlighted the chemoprotective and anti-inflammatory effects of cinnamaldehyde, a Cinnamomum cassia Presl-specific diterpene. In our current study, we have examined the effects of both cinnamaldehyde and extracts of C. cassia on cytokine-induced monocyte/human endothelial cell interactions. We find that these compounds inhibit the adhesion of TNFalpha-induced monocytes to endothelial cells and suppress the expression of the cell adhesion molecules, VCAM-1 and ICAM-1, at the transcriptional level. Moreover, in TNFalpha-treated ECs, the principal downstream signal of VCAM-1 and ICAM-1, NF-kappaB, was also found to be abolished in a time-dependent manner. Interestingly, cinnamaldehyde exerts its anti-inflammatory effects by blocking the degradation of the inhibitory protein IkappaB-alpha, but only in short term pretreatments, whereas it does so via the induction of Nrf2-related genes, including heme-oxygenase-1 (HO-1), over long term pretreatments. Treating ECs with zinc protoporphyrin, a HO-1 inhibitor, partially blocks the anti-inflammatory effects of cinnamaldehyde. Elevated HO-1 protein levels were associated with the inhibition of TNFalpha-induced ICAM-1 expression. In addition to HO-1, we also found that cinnamaldehyde can upregulate Nrf2 in nuclear extracts, and can increase ARE-luciferase activity and upregulate thioredoxin reductase-1, another Nrf2-related gene. Moreover, cinnamaldehyde exposure rapidly reduces the cellular GSH levels in ECs over short term treatments but increases these levels after 9 h exposure. Hence, our present findings indicate that cinnamaldehyde suppresses TNF-induced singling pathways via two distinct mechanisms that are activated by different pretreatment periods.

  13. Tumor necrosis factor-alpha is expressed by glomerular visceral epithelial cells in human membranous nephropathy.

    PubMed Central

    Neale, T. J.; Rüger, B. M.; Macaulay, H.; Dunbar, P. R.; Hasan, Q.; Bourke, A.; Murray-McIntosh, R. P.; Kitching, A. R.

    1995-01-01

    The role of tumor necrosis factor alpha (TNF-alpha) was examined in biopsy-proven glomerulonephritis by immunohistochemistry, in situ hybridization, immunogold electron microscopy, immunoassay in serum and urine, and urinary immunoblot. Striking glomerular capillary wall and visceral glomerular epithelial cell TNF-alpha protein staining was observed in all cases of membranous nephropathy and membranous lupus nephropathy. Staining was less frequently observed in crescentic glomerulonephritis and in isolated cases of other histological subtypes of glomerulonephritis, usually in association with glomerular macrophages. By immunogold electron microscopy TNF-alpha was localized in membranous nephropathy within the visceral glomerular epithelial cells, and also in the glomerular basement membrane, especially in relation to immune deposits. In situ hybridization localized TNF-alpha mRNA exclusively to glomerular epithelial cells in all biopsies with membranous morphology but not in other histological subtypes. Concentrations of TNF-alpha were significantly increased compared with normal controls in the urine of patients with membranous nephropathy and with crescentic glomerulonephritis. The expression of TNF-alpha by glomerular epithelial cells exclusively and universally in biopsies showing a membranous morphology strongly suggests this cytokine has a role in the pathogenesis of membranous nephropathy. Images Figure 1 Figure 2 Figure 3 Figure 5 PMID:7778683

  14. Macrophage-induced angiogenesis is mediated by tumour necrosis factor-alpha.

    PubMed

    Leibovich, S J; Polverini, P J; Shepard, H M; Wiseman, D M; Shively, V; Nuseir, N

    Macrophages are important in the induction of new blood vessel growth during wound repair, inflammation and tumour growth. We show here that tumour necrosis factor-alpha (TNF-alpha), a secretory product of activated macrophages that is believed to mediate tumour cytotoxicity, is a potent inducer of new blood vessel growth (angiogenesis). In vivo, TNF-alpha induces capillary blood vessel formation in the rat cornea and the developing chick chorioallantoic membrane at very low doses. In vitro, TNF-alpha stimulates chemotaxis of bovine adrenal capillary endothelial cells and induces cultures of these cells grown on type-1 collagen gels to form capillary-tube-like structures. The angiogenic activity produced by activated murine peritoneal macrophages is completely neutralized by a polyclonal antibody to TNF-alpha, suggesting immunological features are common to TNF-alpha and the protein responsible for macrophage-derived angiogenic activity. In inflammation and wound repair, TNF-alpha could augment repair by stimulating new blood vessel growth; in tumours, TNF-alpha might both stimulate tumour development by promoting vessel growth and participate in tumour destruction by direct cytotoxicity.

  15. Molecular Mechanisms of Hepatocellular Apoptosis Induced by Trovafloxacin-Tumor Necrosis Factor-alpha Interaction

    PubMed Central

    Roth, Robert A.

    2014-01-01

    Idiosyncratic drug-induced liver injury (IDILI) continues to be a significant human health problem. IDILI is characterized as occurring in a minority of individuals exposed to a drug, yet it accounts for as much as 17% of all cases of acute liver failure. Despite these concerns, the mechanisms underlying IDILI remain unknown. Trovafloxacin (TVX), which causes IDILI in humans, also causes hepatocellular death in vitro when combined with tumor necrosis factor-alpha (TNF) treatment. However, the molecular mechanisms involved in this toxicity are not fully characterized. The purpose of this study was to identify mechanisms by which TVX and TNF interact to cause hepatocellular death, with a focus on a human hepatocyte cell line. TVX and TNF interacted to cause cytotoxicity in HepG2 cells at drug concentrations similar to those in people undergoing TVX therapy. TVX/TNF treatment caused apoptosis and DNA damage in HepG2 cells that depended on caspase activation. Prolonged activation of JNK occurred in TVX/TNF-induced cytotoxicity, and treatment with the JNK selective inhibitor SP600125 attenuated cytotoxicity. TVX/TNF cotreatment also caused cytotoxicity in isolated primary murine hepatocytes that was dependent on caspase activation. These results increase understanding of molecular signaling pathways involved in hepatocellular death caused by a drug with idiosyncratic liability in the presence of TNF. PMID:24097668

  16. Regulation of the Drosophila hypoxia-inducible factor alpha Sima by CRM1-dependent nuclear export.

    PubMed

    Romero, Nuria M; Irisarri, Maximiliano; Roth, Peggy; Cauerhff, Ana; Samakovlis, Christos; Wappner, Pablo

    2008-05-01

    Hypoxia-inducible factor alpha (HIF-alpha) proteins are regulated by oxygen levels through several different mechanisms that include protein stability, transcriptional coactivator recruitment, and subcellular localization. It was previously reported that these transcription factors are mainly nuclear in hypoxia and cytoplasmic in normoxia, but so far the molecular basis of this regulation is unclear. We show here that the Drosophila melanogaster HIF-alpha protein Sima shuttles continuously between the nucleus and the cytoplasm. We identified the relevant nuclear localization signal and two functional nuclear export signals (NESs). These NESs are in the Sima basic helix-loop-helix (bHLH) domain and promote CRM1-dependent nuclear export. Site-directed mutagenesis of either NES provoked Sima nuclear retention and increased transcriptional activity, suggesting that nuclear export contributes to Sima regulation. The identified NESs are conserved and probably functional in the bHLH domains of several bHLH-PAS proteins. We propose that rapid nuclear export of Sima regulates the duration of cellular responses to hypoxia.

  17. Transforming growth factor alpha induces collagen degradation and cell migration in differentiating human epidermal raft cultures.

    PubMed Central

    Turksen, K; Choi, Y; Fuchs, E

    1991-01-01

    When cultured on plastic and treated with transforming growth factor alpha (TGF alpha), human keratinocytes exhibit an increase in proliferation at the colony periphery, apparently as a consequence of enhanced cell migration (Barrandon and Green, 1987). To investigate the effects of TGF alpha on a differentiating stratified squamous epithelium and to begin to examine the molecular basis mediating this influence, we cultured human epidermal cells on a gelled lattice of collagen and fibroblasts, floating on the air-liquid interface. Under these conditions, raft cultures differentiate and exhibit morphological and biochemical features of human skin in vivo (Asselineau et al., 1986; Kopan et al., 1987). When 3-wk-old raft cultures were treated with TGF alpha, basal cells showed a marked increase in cell proliferation. At elevated concentrations of TGF alpha, the organization of cells within the artificial tissue changed and islands of basal cells entered the collagen matrix. Biochemical analysis of the response revealed that type I collagenase and gelatinase were induced by keratinocytes within 12 h after TGF alpha treatment. In contrast, invasion of basal cells into the collagen matrix was not significant until 48-72 h post-treatment, suggesting that collagenase and gelatinase production may be a prerequisite to this phenomenon. These results have important implications for the possible role of TGF alpha in squamous cell carcinoma and tumor invasion. Images PMID:1663788

  18. Tumor necrosis factor alpha gene expression in human monocytic THP-1 cells exposed to beryllium.

    PubMed

    Galbraith, G M; Pandey, J P; Schmidt, M G; Arnaud, P; Goust, J M

    1996-01-01

    Chronic beryllium disease, which results from occupational exposure to particulate beryllium, is characterized by the development of lung granulomas and progressive pulmonary fibrosis. Increased production of proinflammatory cytokines (e.g., tumor necrosis factor alpha and interleukin-1 beta) by pulmonary alveolar macrophages occurs in many chronic fibrotic lung diseases and is thought to contribute to the disease process. The purpose of the present study was to investigate cytokine production by human monocytic cells exposed to beryllium in vitro. The results indicated that such cells respond to beryllium ions in the presence of fluoride by accumulation of messenger ribonucleic acid for both tumor necrosis factor alpha and interleukin-1 beta. These findings suggest that inhaled beryllium may directly stimulate the production of these cytokines by alveolar macrophages in vitro.

  19. Ribozyme modulation of lipopolysaccharide-induced tumor necrosis factor-alpha production by peritoneal cells in vitro and in vivo.

    PubMed

    Sioud, M

    1996-05-01

    We have utilized synthetic ribozymes to modulate the lipopolysaccharide (LPS)-induced production of tumor necrosis factor-alpha (TNF-alpha) by peritoneal cells. Two hammerhead ribozymes (mRz1 and mRz2) were prepared by transcription in vitro and their activities in vitro and in vivo were investigated. Both ribozymes cleaved their RNA target with an apparent turnover number (kcat) of 2 min(-1), and inhibited TNF-alpha gene expression in vitro by 50% and 70%, respectively. When mRz1 and mRz2, entrapped in liposomes, were delivered into mice by intraperitoneal injection, they inhibited LPS-induced TNF-alpha gene expression in vivo with mRz2 being the most effective. This enhanced activity could result from the facilitation of catalysis by cellular endogenous proteins, since they specifically bind to mRz2 as compared to mRz1. Furthermore, a significant mRz2 activity can be recovered from peritoneal cells 2 days post-administration in vivo. The anti-TNF-alpha ribozyme treatment in vivo resulted in a more significant reduction of LPS-induced IFN-gamma protein secretion compared to IL-10. In contrast to this pleiotropic effect, the anti-TNF-alpha ribozyme treatment did not affect the heterogenous expression of Fas ligand by peritoneal cells, indicating the specificity of the treatment. Taken together, the present data indicate that the biological effects of TNF-alpha can be modulated by ribozymes. In addition, the data suggest that ribozymes can be administered in a drug-like manner, and therefore indicate their potential in clinical applications.

  20. Sepsis-induced suppression of skeletal muscle translation initiation mediated by tumor necrosis factor alpha.

    PubMed

    Lang, Charles H; Frost, Robert A

    2007-01-01

    Inhibition of translational efficiency is responsible at least in part for the sepsis-induced decrease in protein synthesis observed in skeletal muscle. Moreover, infusion of the inflammatory cytokine tumor necrosis factor alpha (TNF-alpha) into naive rats produces a comparable decrement. Therefore, the purpose of the present study was to determine whether inhibition of TNF action under in vivo conditions could prevent the sepsis-induced decrease in translation initiation observed in the postabsorptive state. To address this aim, sepsis was produced by cecal ligation and puncture (CLP) and rats were studied in the fasted condition 20 to 24 hours thereafter. Both septic and time-matched nonseptic control rats were pretreated with TNF-binding protein (TNF(BP)) before CLP or sham surgery to neutralize endogenously produced TNF. Sepsis altered the distribution of eukaryotic initiation factor 4E (eIF4E) in the gastrocnemius by increasing the amount associated with 4E-BP1 (inactive complex) and decreasing the amount bound to eIF4G (active complex). This change in eIF4E availability was associated with a decreased phosphorylation of 4E-BP1. Furthermore, the phosphorylation of ribosomal protein S6 and mammalian target of rapamycin (mTOR) was also decreased in the gastrocnemius from septic rats. Pretreatment of septic rats with TNF(BP) largely ameliorated the altered distribution of eIF4E as well as the reduced phosphorylation of 4E-BP1, S6, and mTOR. In contrast, sepsis did not change either the total amount or the phosphorylation state of eIF2alpha or eIF2Bepsilon. Furthermore, no sepsis-induced change in eIFs was detected in the slow-twitch soleus muscle. The ability of TNF(BP) to prevent the sepsis-induced alterations in translation initiation was independent of change in plasma insulin and proportional to the insulinlike growth factor I content in blood and muscle but was associated with a reduction in plasma corticosterone. Hence, the decreased constitutive protein

  1. The tumor necrosis factor-alpha-induced protein 8 family in immune homeostasis and inflammatory cancer diseases.

    PubMed

    Luan, Y Y; Yao, Y M; Sheng, Z Y

    2013-01-01

    Within the immune system homeostasis is maintained by a myriad of mechanisms that include the regulation of immune cell activation and programmed cell death. The breakdown of immune homeostasis may lead to fatal inflammatory diseases. We set out to identify genes of tumor necrosis factor-alpha-induced protein 8 (TNFAIP8) family that has a functional role in the process of immune homeostasis. Tumor necrosis factor-alpha-induced protein 8 (TNFAIP8), which functions as an oncogenic molecule, is also associated with enhanced cell survival and inhibition of apoptosis. Tumor necrosis factor-alpha-induced protein 8-like 2 (TIPE2) governs immune homeostasis in both the innate and adaptive immune system and prevents hyper-responsiveness by negatively regulating signaling via T cell receptors and Toll-like receptors (TLRs). There also exist two highly homologous but uncharacterized proteins, TIPE1 and TIPE3. This review is an attempt to provide a summary of TNFAIP8 family associated with immune homeostasis and inflammatory cancer diseases.

  2. Cyclooxygenase 2, pS2, inducible nitric oxide synthase and transforming growth factor alpha in gastric adaptation to stress

    PubMed Central

    Nie, Shi-Nan; Sun, Hai-Chen; Wu, Xue-Hao; Qian, Xiao-Ming

    2004-01-01

    AIM: To determine the role of mucosal gene expression of cyclooxygenase 2 (COX-2), pS2 (belongs to trefoil peptides), inducible nitric oxide synthase (iNOS) and transforming growth factor alpha (TGFα ) in gastric adaptation to water immersion and restraint stress (WRS) in rats. METHODS: Wistar rats were exposed to single or repeated WRS for 4 h every other day for up to 6 d. Gastric mucosal blood flow (GMBF) was measured by laser Doppler flowmeter-3. The extent of gastric mucosal lesions were evaluated grossly and histologically and expressions of COX-2, pS2, iNOS and TGFα were determined by reverse transcriptase polymerase chain reaction (RT-PCR) and Western blot. RESULTS: The damage to the surface of gastric epithelium with focal areas of deep haemorrhagic necrosis was induced by repeated WRS.The adaptative cytoprotection against stress was developed with activation of cell proliferation in the neck regions of gastric glands. The ulcer index (UI) in groups II, III and IV was markedly reduced as compared with group I (I: 47.23 ± 1.20; IV: 10.39 ± 1.18,P < 0.01). GMBF significantly decreased after first exposure to WRS with an adaptive increasement of GMBF in experimental groups after repetitive challenges with WRS. After the 4th WRS, the value of GMBF almost restored to normal level (I: 321.87 ± 8.85; IV: 455.95 ± 11.81,P < 0.01). First WRS significantly decreased the expression of pS2 and significantly increased the expressions of COX-2, iNOS and TGFα . After repeated WRS, pS2 and TGFα expressions gradually increased (pS2: I: 0.37 ± 0.02; IV: 0.77 ± 0.01; TGFα : I: 0.86 ± 0.01; IV: 0.93 ± 0.03,P < 0.05) with a decrease in the expressions of COX-2 and iNOS (COX-2: I: 0.45 ± 0.02; IV: 0.22 ± 0.01; iNOS: I: 0.93 ± 0.01; IV: 0.56 ± 0.01, P < 0.01). Expressions of pS2, COX-2, iNOS and TGFα showed regular changes with a good relationship among them. CONCLUSION: Gastric adaptation to WRS injury involves enhanced cell proliferation,increased expression

  3. Involvement of Mst1 in tumor necrosis factor-{alpha}-induced apoptosis of endothelial cells

    SciTech Connect

    Ohtsubo, Hideki; Ichiki, Toshihiro Imayama, Ikuyo; Ono, Hiroki; Fukuyama, Kae; Hashiguchi, Yasuko; Sadoshima, Junichi; Sunagawa, Kenji

    2008-03-07

    Mammalian sterile 20-kinase 1 (Mst1), a member of the sterile-20 family protein kinase, plays an important role in the induction of apoptosis. However, little is know about the physiological activator of Mst1 and the role of Mst1 in endothelial cells (ECs). We examined whether Mst1 is involved in the tumor necrosis factor (TNF)-{alpha}-induced apoptosis of ECs. Western blot analysis revealed that TNF-{alpha} induced activation of caspase 3 and Mst1 in a time- and dose-dependent manner. TNF-{alpha}-induced Mst1 activation is almost completely prevented by pretreatment with Z-DEVD-FMK, a caspase 3 inhibitor. Nuclear staining with Hoechst 33258 and fluorescence-activated cell sorting of propidium iodide-stained cells showed that TNF-{alpha} induced apoptosis of EC. Diphenyleneiodonium, an inhibitor of NADPH oxidase, and N-acetylcysteine, a potent antioxidant, also inhibited TNF-{alpha}-induced activation of Mst1 and caspase 3, as well as apoptosis. Knockdown of Mst1 expression by short interfering RNA attenuated TNF-{alpha}-induced apoptosis but not cleavage of caspase 3. These results suggest that Mst1 plays an important role in the induction of TNF-{alpha}-induced apoptosis of EC. However, positive feedback mechanism between Mst1 and caspase 3, which was shown in the previous studies, was not observed. Inhibition of Mst1 function may be beneficial for maintaining the endothelial integrity and inhibition of atherogenesis.

  4. Evaluation of Cucurbita maxima extract against scopolamine-induced amnesia in rats: implication of tumour necrosis factor alpha.

    PubMed

    Jawaid, Talha; Shakya, Ashok K; Siddiqui, Hefazat Hussain; Kamal, Mehnaz

    2014-01-01

    Cucurbita maxima (CM) seed oil is commonly used in Indian folk medicine to treat various ailments. We have investigated the effect of CM seed oil on memory impairment induced by scopolamine in rats. Male adult Wistar rats were administered scopolamine 1 mg/kg body weight, i.p. or 1.25 mg/kg body weight, s.c. to induce memory impairment. The nootropic agent piracetam 100 mg/kg body weight, i.p. and CM seed oil 100 and 200 mg/kg body weight, p.o. were administered daily for five consecutive days. The memory function was evaluated in the Morris water maze (MWM) test, the social recognition test (SRT), the elevated plus maze (EPM) test, and the pole climbing test (PCT). Acetylcholinesterase (AChE) activity and oxidative stress parameters were estimated in the cortex, hippocampus, and cerebellum of the brains after completion of the behavioural studies. The effects of scopolamine on the levels of the tumour necrosis factor alpha (TNF-α) transcript were also investigated. Scopolamine caused memory impairment in all the behavioural paradigms along with a significant increase in the AChE activity and oxidative stress in the brain. Scopolamine also caused a significant increase in the expression of TNF-α in the hippocampus. CM seed oil exhibited antiamnesic activity as indicated by a significant reduction in the latency time in the MWM test and decreased social interaction during trial 2 in the SRT. Further, treatment with CM seed oil significantly decreased the AChE activity and malondialdehyde levels and increased the glutathione level in brain regions. CM seed oil also significantly decreased the expression of TNF-α in the hippocampus. The effect of CM seed oil on behavioural and biochemical parameters was comparable to that observed in rats treated with piracetam. These results indicate that CM seed oil may exert antiamnesic activity which may be attributed to the inhibition of AChE and inflammation as well as its antioxidant activity in the brain.

  5. Cardiotrophin-1 induces tumor necrosis factor alpha synthesis in human peripheral blood mononuclear cells.

    PubMed

    Fritzenwanger, Michael; Meusel, Katharina; Jung, Christian; Franz, Marcus; Wang, Zhenhua; Foerster, Martin; Figulla, Hans-R

    2009-01-01

    Chronic heart failure (CHF) is associated with elevated concentrations of tumor necrosis factor (TNF) alpha and cardiotrophin-1 (CT-1) and altered peripheral blood mononuclear cell (PBMC) function. Therefore, we tested whether CT-1 induces TNFalpha in PBMC of healthy volunteers. CT-1 induced in PBMC TNFalpha protein in the supernatant and TNFalpha mRNA in a concentration- and time-dependent manner determined by ELISA and real-time PCR, respectively. Maximal TNFalpha protein was achieved with 100 ng/mL CT-1 after 3-6 hours and maximal TNFalpha mRNA induction after 1 hour. ELISA data were confirmed using immunofluorescent flow cytometry. Inhibitor studies with actinomycin D and brefeldin A showed that both protein synthesis and intracellular transport are essential for CT-1 induced TNFalpha expression. CT-1 caused a dose dependent nuclear factor (NF) kappaB translocation. Parthenolide inhibited both NFkappaB translocation and TNFalpha protein expression indicating that NFkappaB seems to be necessary. We revealed a new mechanism for elevated serum TNFalpha concentrations and PBMC activation in CHF besides the hypothesis of PBMC activation by bacterial translocation from the gut.

  6. Tumor Necrosis Factor-Alpha and the ERK Pathway Drive Chemerin Expression in Response to Hypoxia in Cultured Human Coronary Artery Endothelial Cells

    PubMed Central

    Chua, Su-Kiat; Shyu, Kou-Gi; Lin, Yuh-Feng; Lo, Huey-Ming; Wang, Bao-Wei

    2016-01-01

    Background Chemerin, a novel adipokine, plays a role in the inflammation status of vascular endothelial cells. Hypoxia causes endothelial-cell proliferation, migration, and angiogenesis. This study was aimed at evaluating the protein and mRNA expression of chemerin after exposure of human coronary artery endothelial cells (HCAECs) to hypoxia. Methods and Results Cultured HCAECs underwent hypoxia for different time points. Chemerin protein levels increased after 4 h of hypoxia at 2.5% O2, with a peak of expression of tumor necrosis factor-alpha (TNF-alpha) at 1 h. Both hypoxia and exogenously added TNF-alpha during normoxia stimulated chemerin expression, whereas an ERK inhibitor (PD98059), ERK small interfering RNA (siRNA), or an anti-TNF-alpha antibody attenuated the chemerin upregulation induced by hypoxia. A gel shift assay indicated that hypoxia induced an increase in DNA-protein binding between the chemerin promoter and transcription factor SP1. A luciferase assay confirmed an increase in transcriptional activity of SP1 on the chemerin promoter during hypoxia. Hypoxia significantly increased the tube formation and migration of HCAECs, whereas PD98059, the anti-TNF-alpha antibody, and chemerin siRNA each attenuated these effects. Conclusion Hypoxia activates chemerin expression in cultured HCAECs. Hypoxia-induced chemerin expression is mediated by TNF-alpha and at least in part by the ERK pathway. Chemerin increases early processes of angiogenesis by HCAECs after hypoxic treatment. PMID:27792771

  7. Dynamic equilibrium unfolding pathway of human tumor necrosis factor-alpha induced by guanidine hydrochloride.

    PubMed

    Kim, Y R; Hahn, J S; Hong, H; Jeong, W; Song, N W; Shin, H C; Kim, D

    1999-01-11

    The dynamic equilibrium unfolding pathway of human tumor necrosis factor-alpha (TNF-alpha) during denaturation at different guanidine hydrochloride (GdnHCl) concentrations (0-4.2 M) was investigated by steady-state fluorescence spectroscopy, potassium iodide (KI) fluorescence quenching, far-UV circular dichroism (CD), picosecond time-resolved fluorescence lifetime, and anisotropy decay measurements. We utilized the intrinsic fluorescence of Trp-28 and Trp-114 to characterize the conformational changes involved in the equilibrium unfolding pathway. The detailed unfolding pathway under equilibrium conditions was discussed with respect to motional dynamics and partially folded structures. At 0-0.9 M [GdnHCl], the rotational correlation times of 22-25 ns were obtained from fluorescence anisotropy decay measurements and assigned to those of trimeric states by hydrodynamic calculation. In this range, the solvent accessibility of Trp residues increased with increasing [GdnHCl], suggesting the slight expansion of the trimeric structure. At 1.2-2.1 M [GdnHCl], the enhanced solvent accessibility and the rotational degree of freedom of Trp residues were observed, implying the loosening of the internal structure. In this [GdnHCl] region, TNF-alpha was thought to be in soluble aggregates having distinct conformational characteristics from a native (N) or fully unfolded state (U). At 4.2 M [GdnHCl], TNF-alpha unfolded to a U-state. From these results, the equilibrium unfolding pathway of TNF-alpha, trimeric and all beta-sheet protein, could not be viewed from the simple two state model (N-->U).

  8. Interleukin-1alpha and tumor necrosis factor-alpha expression during the early phases of orthodontic tooth movement in rats.

    PubMed

    Bletsa, Athanasia; Berggreen, Ellen; Brudvik, Pongsri

    2006-10-01

    Remodelling of the periodontium after application of mechanical forces constitutes the basis of clinical orthodontics and various immunoregulatory molecules are involved in this process. The aim of this study was to investigate the expression of the cytokines interleukin-1alpha (IL-1alpha) and tumor necrosis factor-alpha (TNF-alpha) in dental tissues during the early phases of orthodontic tooth movement. Eightteen male Wistar rats were used. All maxillary right first molars were moved orthodontically, with a force of 0.5 N, for 3 h, 1 d, and 3 d. The contralateral sides served as untreated controls. Parasagittal sections of the maxillary molars and the surrounding tissues were subjected to immunohistochemical staining for IL-1alpha or TNF-alpha, and were evaluated with light microscopy. IL-1alpha and TNF-alpha were expressed in the bone and periodontal ligament (PDL) along the roots of the orthodontically moved molars and in the gingiva. Increased expression of both cytokines was observed in the aforementioned areas after 1 and 3 d of tooth movement. The pulp tissue exhibited only minor changes in cytokine expression during tooth movement. The results suggest that mechanical stress results in almost immediate inflammatory reactions in various dental tissues.

  9. Erythropoietin protects myocardin-expressing cardiac stem cells against cytotoxicity of tumor necrosis factor-{alpha}

    SciTech Connect

    Madonna, Rosalinda; Shelat, Harnath; Xue, Qun; Willerson, James T.; De Caterina, Raffaele; Geng, Yong-Jian

    2009-10-15

    Cardiac stem cells are vulnerable to inflammation caused by infarction or ischemic injury. The growth factor, erythropoietin (Epo), ameliorates the inflammatory response of the myocardium to ischemic injury. This study was designed to assess the role of Epo in regulation of expression and activation of the cell death-associated intracellular signaling components in cardiac myoblasts stimulated with the proinflammatory cytokine tumor necrosis factor (TNF)-{alpha}. Cardiac myoblasts isolated from canine embryonic hearts characterized by expression of myocardin A, a promyogenic transcription factor for cardiovascular muscle development were pretreated with Epo and then exposed to TNF-{alpha}. Compared to untreated cells, the Epo-treated cardiac myoblasts exhibited better morphology and viability. Immunoblotting revealed lower levels of active caspase-3 and reductions in iNOS expression and NO production in Epo-treated cells. Furthermore, Epo pretreatment reduced nuclear translocation of NF-{kappa}B and inhibited phosphorylation of inhibitor of kappa B (I{kappa}B) in TNF-{alpha}-stimulated cardiac myoblasts. Thus, Epo protects cardiac myocyte progenitors or myoblasts against the cytotoxic effects of TNF-{alpha} by inhibiting NF-{kappa}B-mediated iNOS expression and NO production and by preventing caspase-3 activation.

  10. Nitric oxide mediates angiogenesis induced in vivo by platelet-activating factor and tumor necrosis factor-alpha.

    PubMed Central

    Montrucchio, G.; Lupia, E.; de Martino, A.; Battaglia, E.; Arese, M.; Tizzani, A.; Bussolino, F.; Camussi, G.

    1997-01-01

    We evaluated the role of an endogenous production of nitric oxide (NO) in the in vitro migration of endothelial cells and in the in vivo angiogenic response elicited by platelet-activating factor (PAF), tumor necrosis factor-alpha (TNF), and basic fibroblast growth factor (bFGF). The NO synthase inhibitor, N omega-nitro-L-arginine-methyl ester (L-NAME), but not its enantiomer D-NAME, prevented chemotaxis of endothelial cells induced in vitro by PAF and by TNF. The motogenic activity of TNF was also inhibited by WEB 2170, a specific PAF-receptor antagonist. In contrast, chemotaxis induced by bFGF was not prevented by L-NAME or by WEB 2170. Angiogenesis was studied in vivo in a murine model in which Matrigel was used as a vehicle for the delivery of mediators. In this model, the angiogenesis induced by PAF and TNF was inhibited by WEB 2170 and L-NAME but not by D-NAME. In contrast, angiogenesis induced by bFGF was not affected by L-NAME or by WEB 2170. TNF, but not bFGF, induced PAF synthesis within Matrigel. These results suggest that NO mediates the angiogenesis induced by PAF as well as that induced by TNF, which is dependent on the production of PAF. In contrast, the angiogenic effect of bFGF appears to be both PAF and NO independent. Images Figure 3 Figure 4 PMID:9250168

  11. Tumor necrosis factor-alpha expression in white-tailed deer (Odocoileus virginianus) infected with Epizootic haemorrhagic disease virus.

    PubMed

    Sharma, Prachi; Stallknech, David E; Quist, Charlotte F; Howerth, Elizabeth W

    2016-09-30

    Epizootic haemorrhagic disease (EHD) is the most important infectious disease of white‑tailed deer (WTD), however little is known about the role of inflammatory mediators in the pathogenesis. We characterized the expression of tumor necrosis factor‑alpha (TNF-α) ex vivo in tissues of WTD experimentally or naturally infected with EHD virus serotype 2 and in WTD peripheral blood mononuclear cells (PBMC) infected with EHD virus serotype 2 in vitro. Circulating levels of TNF-α were evaluated in serum from experimentally infected deer via cytotoxicity assay. The expression of TNF-α in tissues was evaluated via immunohistochemistry (IHC) in both experimentally and naturally infected deer. Semi‑quantitative reverse transcriptase polymerase chain reaction (RT-PCR) was used to assess the level of TNF-α mRNA in tissues from experimentally infected deer and WTD's PBMC. Circulating levels of TNF-α were not increased in infected animals and TNF-α was not detected in tissues of infected deer. Increased transcription of TNF-α was detected neither in infected WTD nor in the PBMC. Tumor necrosis factor-alpha may not play a significant role in the pathogenesis of EHD virus infection in WTD.

  12. Reduction of NO Synthase Expression and Tumor Necrosis Factor Alpha Production in Macrophages by Amphotericin B Lipid Carriers

    PubMed Central

    Larabi, Malika; Legrand, Philippe; Appel, Martine; Gil, Sophie; Lepoivre, Michel; Devissaguet, Jean-Philippe; Puisieux, Francis; Barratt, Gillian

    2001-01-01

    The present study compared the abilities of different lipid carriers of amphotericin B (AMB) to activate murine peritoneal macrophages, as assessed by their capacities to produce nitric oxide (NO) and tumor necrosis factor alpha (TNF-α). Although AMB alone did not induce NO production, synergy was observed with gamma interferon but not with lipopolysaccharide. This synergy could not be explained by the mobilization of the nuclear activation factor NF-κB by AMB. On the other hand, AMB induced TNF-α production without a costimulator and no synergy was observed. Anti-TNF-α antibodies did not influence NO production, and an inhibitor of NO synthase did not affect TNF-α production, indicating that the production of one of these effector molecules was independent of that of the other. The incorporation of AMB into lipid carriers reduced NO and TNF-α production with all formulations but more so with liposomes than with lipid complexes. NO production was correlated with the induction of NO synthase II, revealed by Western blotting. The extent of association of AMB with macrophages depended on the formulation, especially on the AMB/lipids ratio: the higher the ratio was, the greater the AMB association with macrophages. However, there was no clear correlation between AMB association with macrophages, whether internalized or bound to the membrane, and immunostimulating effects. These results may explain the reduced toxicities of lipid-based formulations of AMB. PMID:11158754

  13. Evolution of Ciona intestinalis Tumor necrosis factor alpha (CiTNFα): Polymorphism, tissues expression, and 3D modeling.

    PubMed

    Vizzini, Aiti; Giovanna, Parisi Maria; Cardinale, Laura; Testasecca, Lelia; Cammarata, Matteo

    2017-02-01

    Although the Tumor necrosis factor gene superfamily seems to be very conserved in vertebrates, phylogeny, tissue expression, genomic and gene organization, protein domains and polymorphism analyses showed that a strong change has happened mostly in invertebrates in which protochordates were a constraint during the immune-molecules history and evolution. RT PCR was used to investigate differential gene expression in different tissues. The expression shown was greater in the pharynx. Single-nucleotide polymorphism has been investigated in Ciona intestinalis Tumor necrosis factor alpha (CiTNFα) mRNA isolated from the pharynx of 30 ascidians collected from Licata, Sicily (Italy), by denaturing gradient gel electrophoresis (DGGE). For this analysis, CiTNFα nucleotide sequence was separated into two fragments, TNF-1 and -2, respectively, of 630 and 540 bp. We defined 23 individual DGGE patterns (named 1 to 10 for TNF-1 and 1 to 13 for TNF-2). Five patterns for TNF-1 accounted for <10% of the individuals, whereas the pattern 13 of TNF-2 accounted for >20% of the individuals. All the patterns were verified by direct sequencing. Single base-pair mutations were observed mainly within COOH-terminus, leading to 30 nucleotide sequence variants and 30 different coding sequences segregating in two main different clusters. Although most of the base mutations were silent, four propeptide variants were detected and six amino acid replacements occurred within COOH-terminus. Statistical tests for neutrality indicated negative selection pressure on signal and mature peptide domains, but possible positive selection pressure on COOH-terminus domain. Lastly we displayed the in silico 3D structure analysis including the CiTNFα variable region.

  14. Epithelial to mesenchymal transition in human skin wound healing is induced by tumor necrosis factor-alpha through bone morphogenic protein-2.

    PubMed

    Yan, Chunli; Grimm, Wesley A; Garner, Warren L; Qin, Lan; Travis, Taryn; Tan, Neiman; Han, Yuan-Ping

    2010-05-01

    Epithelial-mesenchymal transition (EMT), characterized by loss of epithelial adhesion and gain of mesenchymal features, is an important mechanism to empower epithelial cells into the motility that occurs during embryonic development and recurs in cancer and fibrosis. Whether and how EMT occurs in wound healing and fibrosis in human skin remains unknown. In this study we found that migrating epithelial cells in wound margins and deep epithelial ridges had gained mesenchymal features such as vimentin and FSP1 expression. In hypertrophic scars, EMT-related genes were elevated along with inflammatory cytokines, indicating a causal relationship. To reconstitute EMT in vitro, normal human skin and primary keratinocytes were exposed to cytokines such as tumor necrosis factor-alpha (TNF-alpha), resulting in expression of vimentin, FSP1, and matrix metalloproteinases. Moreover, TNF-alpha-induced EMT was impaired by antagonists against bone morphogen proteins (BMP) 2/4, suggesting that BMP mediates the TNF-alpha-induced EMT in human skin. Indeed, TNF-alpha could induce BMP-2 and its receptor (BMPR1A) in human skin and primary keratinocytes, and BMP2 could induce EMT features in skin explants and primary keratinocytes. In summary, we uncovered EMT features in both acute and fibrotic cutaneous wound healing of human skin. Moreover, we propose that the mesenchymal induction in wound healing is motivated by TNF-alpha, in part, through induction of BMP.

  15. Transforming growth factor alpha protection against drug-induced injury to the rat gastric mucosa in vivo.

    PubMed Central

    Romano, M; Polk, W H; Awad, J A; Arteaga, C L; Nanney, L B; Wargovich, M J; Kraus, E R; Boland, C R; Coffey, R J

    1992-01-01

    This study was designed to determine whether transforming growth factor alpha (TGF alpha) protects rat gastric mucosa against ethanol- and aspirin-induced injury. Systemic administration of TGF alpha dose-dependently decreased 100% ethanol-induced gastric mucosal injury; a dose of 50 micrograms/kg delivered intraperitoneally 15 min before ethanol decreased macroscopic mucosal injury by > 90%. At the microscopic level, TGF alpha prevented deep gastric necrotic lesions and reduced disruption of surface epithelium. Pretreatment with orogastric TGF alpha (200 micrograms/kg) only partially (40%) decreased macroscopic ethanol damage. Intraperitoneal administration of TGF alpha at a dose of 10 micrograms/kg, which does not significantly inhibit gastric acid secretion, decreased aspirin-induced macroscopic damage by > 80%. TGF alpha protection does not seem to be mediated by prostaglandin, glutathione, or ornithine decarboxylase-related events, as evidenced by lack of influence of the inhibition of their production. Pretreatment with the sulfhydryl blocking agent N-ethylmaleimide partially abolished (40%) the protective effect of TGF alpha. In addition, systemic administration of TGF alpha resulted in a two-fold increase in tyrosine phosphorylation of phospholipase C-gamma 1 and in a time- and dose-dependent increase in levels of immunoreactive insoluble gastric mucin; these events occurred in a time frame consistent with their participation in the protective effect of TGF alpha. Images PMID:1281834

  16. Infection of Human Fallopian Tube Epithelial Cells with Neisseria gonorrhoeae Protects Cells from Tumor Necrosis Factor Alpha-Induced Apoptosis

    PubMed Central

    Morales, Priscilla; Reyes, Paz; Vargas, Macarena; Rios, Miguel; Imarai, Mónica; Cardenas, Hugo; Croxatto, Horacio; Orihuela, Pedro; Vargas, Renato; Fuhrer, Juan; Heckels, John E.; Christodoulides, Myron; Velasquez, Luis

    2006-01-01

    Following infection with Neisseria gonorrhoeae, bacteria may ascend into the Fallopian tubes (FT) and induce salpingitis, a major cause of infertility. In the FT, interactions between mucosal epithelial cells and gonococci are pivotal events in the pathogen's infection cycle and the inflammatory response. In the current study, primary FT epithelial cells were infected in vitro with different multiplicities of infection (MOI) of Pil+ Opa+ gonococci. Bacteria showed a dose-dependent association with cells and induced the secretion of tumor necrosis factor alpha (TNF-α). A significant finding was that gonococcal infection (MOI = 1) induced apoptosis in approximately 30% of cells, whereas increasing numbers of bacteria (MOI = 10 to 100) did not induce apoptosis. Apoptosis was observed in only 11% of cells with associated bacteria, whereas >84% of cells with no adherent bacteria were apoptotic. TNF-α was a key contributor to apoptosis, since (i) culture supernatants from cells infected with gonococci (MOI = 1) induced apoptosis in naïve cultures, suggesting that a soluble factor was responsible; (ii) gonococcal infection-induced apoptosis was inhibited with anti-TNF-α antibodies; and (iii) the addition of exogenous TNF-α induced apoptosis, which was inhibited by the presence of increasing numbers of bacteria (MOI = 10 to 100). These data suggest that TNF-α-mediated apoptosis of FT epithelial cells is likely a primary host defense mechanism to prevent pathogen colonization. However, epithelial cell-associated gonococci have evolved a mechanism to protect the cells from undergoing TNF-α-mediated apoptosis, and this modulation of the host innate response may contribute to establishment of infection. Understanding the antiapoptotic mechanisms used by Neisseria gonorrhoeae will inform the pathogenesis of salpingitis and could suggest new intervention strategies for prevention and treatment of the disease. PMID:16714596

  17. Tumor necrosis factor alpha promotes the expression of immunosuppressive proteins and enhances the cell growth in a human bone marrow-derived stem cell culture

    SciTech Connect

    Miettinen, Johanna A.; Pietilae, Mika; Salonen, Riikka J.; Ohlmeier, Steffen; Ylitalo, Kari; Huikuri, Heikki V.; Lehenkari, Petri

    2011-04-01

    Mesenchymal stem cells (MSCs) are widely used in experimental treatments for various conditions that involve normal tissue regeneration via inflammatory repair. It is known that MSCs can secrete multiple soluble factors and suppress inflammation. Even though the effect of MSCs on inflammation has been extensively studied, the effect of inflammation on MSCs is poorly understood. One of the major cytokines released at the site of inflammation is tumor necrosis factor alpha (TNF-{alpha}) which is known to induce MSC invasion and proliferation. Therefore, we wanted to test the effects of TNF-{alpha} exposure on MSCs derived from human bone marrow. We found, as expected, that cell proliferation was significantly enhanced during TNF-{alpha} exposure. However, according to the cell surface marker analysis, the intensity of several antigens in the minimum criteria panel for MSCs proposed by International Society of Cellular Therapy (ISCT) was decreased dramatically, and in certain cases, the criteria for MSCs were not fulfilled. In addition, TNF-{alpha} exposure resulted in a significant but transient increase in human leukocyte antigen and CD54 expression. Additional proteomic analysis by two-dimensional difference gel electrophoresis and mass spectrometry revealed three proteins whose expression levels decreased and 8 proteins whose expression levels increased significantly during TNF-{alpha} exposure. The majority of these proteins could be linked to immunosuppressive and signalling pathways. These results strongly support reactive and immunosuppressive activation of MSCs during TNF-{alpha} exposure, which might influence MSC differentiation stage and capacity.

  18. Chrysin sensitizes tumor necrosis factor-alpha-induced apoptosis in human tumor cells via suppression of nuclear factor-kappaB.

    PubMed

    Li, Xin; Huang, Qing; Ong, Choon-Nam; Yang, Xing-Fen; Shen, Han-Ming

    2010-07-01

    Chrysin (5,7-dihydroxyflavone) is a natural flavonoid commonly found in many plants. The anti-cancer property of chrysin has been demonstrated although the molecular mechanisms remain to be further elucidated. In the present study, we found that, pretreatment with chrysin greatly sensitized various human cancer cells to tumor necrosis factor-alpha (TNFalpha)-induced apoptosis. In the search of the molecular mechanisms responsible for the sensitization effect of chrysin, we discovered that such sensitization is closely associated with the inhibitory effect of chrysin on TNFalpha-mediated nuclear transcription factor-kappaB (NF-kappaB) activation. Pretreatment with chrysin inhibited TNFalpha-induced degradation of Inhibitor of kappaB (IkappaB) protein and subsequent nuclear translocation of p65. As a result, chrysin suppressed the expression of NF-kappaB-targeted anti-apoptotic gene, c-FLIP-L. The role of c-FLIP-L was further confirmed by its ectopic expression, which significantly protected cell death induced by combined treatment with chrysin and TNFalpha. Data from this study thus reveal a novel function of chrysin and enhance the value of chrysin as an anti-cancer agent.

  19. Tumor necrosis factor alpha-induced angiogenesis depends on in situ platelet-activating factor biosynthesis

    PubMed Central

    1994-01-01

    Tumor necrosis factor (TNF) alpha, a potent inhibitor of endothelial cell growth in vitro, is angiogenic in vivo. Therefore, it was suggested that the angiogenic properties of this agent might be consequent to the production of secondary mediators. Since TNF-alpha stimulates the synthesis of platelet-activating factor (PAF) by monocytes and endothelial cells, we investigated the possible involvement of PAF in the angiogenic effect of TNF-alpha. Angiogenesis was studied in a murine model in which Matrigel was used as a vehicle for the delivery of mediators. In this model the angiogenesis induced by TNF-alpha was shown to be inhibited by WEB 2170, a specific PAF receptor antagonist. Moreover, in mice injected with TNF-alpha, PAF was detected within the Matrigel, 6 and 24 h after TNF-alpha injection. The synthesis of PAF within the Matrigel was concomitant with the early migration of endothelial cells and infiltration of monocytes. No infiltration of lymphocytes or polymorphonuclear leukocytes was observed. Synthetic PAF as well as PAF extracted and purified from mice challenged with TNF-alpha induced a rapid angiogenic response, inhibited by WEB 2170. These results suggest that the angiogenic effect of TNF-alpha is, at least in part, mediated by PAF synthesized from monocytes and/or endothelial cells infiltrating the Matrigel plug. PMID:7516414

  20. microRNA-142 is upregulated by tumor necrosis factor-alpha and triggers apoptosis in human gingival epithelial cells by repressing BACH2 expression

    PubMed Central

    Li, Song; Song, Zhongchen; Dong, Jiachen; Shu, Rong

    2017-01-01

    Tumor necrosis factor-alpha (TNF-α) has been shown to cause apoptosis of gingival epithelial cells (GECs) in periodontitis. However, the underlying molecular mechanism is still unclear. In this study, we showed that miR-142 expression was significantly elevated in human GECs after exposure to TNF-α. Such induction was in a time- and concentration-dependent manner. Serum miR-142 levels were positively correlated with serum TNF-α levels in patients with chronic periodontitis (r = 0.314, P = 0.0152). Depletion of miR-142 was found to attenuate TNF-α-induced apoptosis, as determined by TUNEL staining and caspase-3 activity assays. In contrast, overexpression of miR-142 significantly reduced viability and induced apoptosis in GECs. Basic leucine zipper transcription factor 2 (BACH2) was identified to be a functional target of miR-142. Overexpression of miR-142 caused a 3-fold reduction of BACH2 protein in primary GECs. Overexpression of BACH2 significantly reversed miR-142- or TNF-α-induced apoptosis of GECs. Similar to the findings with miR-142 mimic, depletion of BACH2 significantly promoted apoptosis in GECs, which was accompanied by decreased expression of Bcl-2 and Bcl-xL and increased expression of Bax and Bim. Overall, miR-142 mediates TNF-α-induced apoptosis in gingival epithelial cells by targeting BACH2 and may represent a potential therapeutic target for periodontitis. PMID:28123644

  1. Ginsenosides compound K and Rh(2) inhibit tumor necrosis factor-alpha-induced activation of the NF-kappaB and JNK pathways in human astroglial cells.

    PubMed

    Choi, Kyungsun; Kim, Myungsun; Ryu, Jeonghee; Choi, Chulhee

    2007-06-21

    Ginsenosides, the main component of Panax ginseng, have been known for the anti-inflammatory and anti-proliferative activities. In this study, we investigated the molecular mechanisms responsible for the anti-inflammatory effects of ginsenosides on activated astroglial cells. Among 13 different ginsenosides, intestinal bacterial metabolites Rh(2) and compound K (C-K) showed a significant inhibitory effect on tumor necrosis factor-alpha (TNF-alpha)-induced expression of intercellular adhesion molecule-1 in human astroglial cells. Pretreatment with C-K or Rh(2) suppressed TNF-alpha-induced phosphorylation of IkappaBalpha kinase and the subsequent phosphorylation and degradation of IkappaBalpha. Additionally, the same treatment inhibited TNF-alpha-induced phosphorylation of MKK4 and the subsequent activation of the JNK-AP-1 pathway. The inhibitory effect of ginsenosides on TNF-alpha-induced activation of the NF-kappaB and JNK pathways was not observed in human monocytic U937 cells. These results collectively indicate that ginsenoside metabolites C-K and Rh(2) exert anti-inflammatory effects by the inhibition of both NF-kappaB and JNK pathways in a cell-specific manner.

  2. Association of tumour necrosis factor alpha and its receptors with thymidine phosphorylase expression in invasive breast carcinoma.

    PubMed Central

    Leek, R. D.; Landers, R.; Fox, S. B.; Ng, F.; Harris, A. L.; Lewis, C. E.

    1998-01-01

    Angiogenesis is an essential requirement for tumour growth and metastasis and is regulated by a complex network of factors produced by both stromal cells and neoplastic cells within solid tumours. The cytokine tumour necrosis factor alpha (TNF-alpha) and the enzyme thymidine phosphorylase (TP) are two factors known to promote tumour angiogenesis. We have demonstrated recently that high numbers of tumour-associated macrophages (TAMs) are significantly associated with increased tumour angiogenesis and poor prognosis in invasive carcinoma of the breast. We have also shown that TAMs are a major source of TNF-alpha in invasive breast carcinomas, and that macrophage-like stromal cells as well as tumour cells synthesize TP in such tumours. However, little is known of the factors that regulate the production or activity of these factors in the tumour microenvironment. As TNF-alpha has been shown to up-regulate TP expression in tumour cells in vitro we performed an immunohistochemical study to investigate the possibility that TNF-alpha may be involved in the regulation of TP expression by malignant breast epithelial cells in vivo. To do this, we used a cocktail of non-neutralizing monoclonal anti-TNF-alpha antibodies to visualize both TNF-alpha-expressing macrophages and TNF-alpha bound to its receptors on tumour cells and endothelial cells in a series of 93 invasive carcinomas of the breast. A semiquantitative grading system was then used to compare these staining patterns with that for TP in the same biopsies. TNF-alpha immunoreactivity was also compared with various important tumour variables known to relate to outcome in this disease (microvessel density, node status, grade, stage, receptor status and macrophage infiltration), as well as relapse-free and overall survival data for these patients. Our data show significant positive correlations between TNF-alpha bound to its receptors on tumour cells and: (1) TP protein production by tumour cells, and (2) axillary lymph

  3. Amiodarone exposure during modest inflammation induces idiosyncrasy-like liver injury in rats: role of tumor necrosis factor-alpha.

    PubMed

    Lu, Jingtao; Jones, A Daniel; Harkema, Jack R; Roth, Robert A; Ganey, Patricia E

    2012-01-01

    Amiodarone [2-butyl-3-(3',5'-diiodo-4'α-diethylaminoethoxybenzoyl)-benzofuran] (AMD), a class III antiarrhythmic drug, is known to cause idiosyncratic hepatotoxic reactions in human patients. One hypothesis for the etiology of idiosyncratic adverse drug reactions is that a concurrent inflammatory stress results in decreased threshold for drug toxicity. To explore this hypothesis in an animal model, male Sprague-Dawley rats were treated with nonhepatotoxic doses of AMD or its vehicle and with saline vehicle or lipopolysaccharide (LPS) to induce low-level inflammation. Elevated alanine aminotransferase (ALT), aspartate aminotransferase, alkaline phosphatase, and gamma-glutamyltransferase activities as well as increased total bile acid concentrations in serum and midzonal hepatocellular necrosis were observed only in AMD/LPS-cotreated rats. The time interval between AMD and LPS administration was critical: AMD injected 16 h before LPS led to liver injury, whereas AMD injected 2-12 h before LPS failed to cause this response. The increase in ALT activity in AMD/LPS cotreatment showed a clear dose-response relationship with AMD as well as LPS. The metabolism and hepatic accumulation of AMD were not affected by LPS coexposure. Serum concentration of tumor necrosis factor-alpha (TNF) was significantly increased by LPS and was slightly prolonged by AMD. In Hepac1c7 cells, addition of TNF potentiated the cytotoxicity of both AMD and its primary metabolite, mono-N-desethylamiodarone. In vivo inhibition of TNF signaling by etanercept attenuated the AMD/LPS-induced liver injury in rats. In summary, AMD treatment during modest inflammation induced severe hepatotoxicity in rats, and TNF contributed to the induction of liver injury in this animal model of idiosyncratic AMD-induced liver injury.

  4. Tumor necrosis factor-alpha induced by hepatitis B virus core mediating the immune response for hepatitis B viral clearance in mice model.

    PubMed

    Tzeng, Horng-Tay; Tsai, Hwei-Fang; Chyuan, I-Tsu; Liao, Hsiu-Jung; Chen, Chun-Jen; Chen, Pei-Jer; Hsu, Ping-Ning

    2014-01-01

    Persistent hepatitis B viral (HBV) infection results in chronic hepatitis, liver cirrhosis, and hepatocellular carcinoma (HCC). An efficient control of virus infections requires the coordinated actions of both innate and adaptive immune responses. In order to define the role of innate immunity effectors against HBV, viral clearance was studied in a panel of immunodeficient mouse strains by the hydrodynamic injection approach. Our results demonstrate that HBV viral clearance is not changed in IFN-α/β receptor (IFNAR), RIG-I, MDA5, MYD88, NLRP3, ASC, and IL-1R knock-out mice, indicating that these innate immunity effectors are not required for HBV clearance. In contrast, HBV persists in the absence of tumor necrosis factor-alpha (TNF-α) or in mice treated with the soluble TNF receptor blocker, Etanercept. In these mice, there was an increase in PD-1-expressing CD8+ T-cells and an increase of serum HBV DNA, HBV core, and surface antigen expression as well as viral replication within the liver. Furthermore, the induction of TNF-α in clearing HBV is dependent on the HBV core, and TNF blockage eliminated HBV core-induced viral clearance effects. Finally, the intra-hepatic leukocytes (IHLs), but not the hepatocytes, are the cell source responsible for TNF-α production induced by HBcAg. These results provide evidences for TNF-α mediated innate immune mechanisms in HBV clearance and explain the mechanism of HBV reactivation during therapy with TNF blockage agents.

  5. Increased levels of inositol hexakisphosphate (InsP6) protect HEK293 cells from tumor necrosis factor (alpha)- and Fas-induced apoptosis.

    PubMed

    Verbsky, John; Majerus, Philip W

    2005-08-12

    The overexpression of inositol 1,3,4-trisphosphate 5/6-kinase has recently been shown to protect HEK293 cells from tumor necrosis factor alpha (TNF(alpha))-induced apoptosis. This overexpression leads to an increase in the levels of both inositol 1,3,4,5,6-pentakisphosphate (InsP5) and inositol 1,2,3,4,5,6-hexakisphosphate (InsP6). Cells that overexpress InsP5 2-kinase have increased levels of InsP6 and are also protected from TNFalpha-induced apoptosis; furthermore, cells that express an RNA interference construct to the 2-kinase are deficient in InsP6 and are sensitized to TNFalpha-induced apoptosis. Therefore the protective effect of 5/6-kinase on TNFalpha-mediated apoptosis is due to an increase of InsP6 or to a metabolite derived from InsP6. Furthermore, we find that the InsP6 also protects from Fas-mediated apoptosis. No effect was seen in the endocytic rate of transferrin receptor, caspase 8 activity, or TNF receptor number at the cell surface. Cells that overexpress 2-kinase do show an increase in the amount of receptor-interacting protein (RIP), while cells with reduced InsP6 levels show relatively less RIP, providing a possible mechanism for the effect on apoptosis.

  6. Lipopolysaccharide and Tumor Necrosis Factor Alpha Inhibit Interferon Signaling in Hepatocytes by Increasing Ubiquitin-Like Protease 18 (USP18) Expression

    PubMed Central

    MacParland, Sonya A.; Ma, Xue-Zhong; Chen, Limin; Khattar, Ramzi; Cherepanov, Vera; Selzner, Markus; Feld, Jordan J.; Selzner, Nazia

    2016-01-01

    ABSTRACT Inflammation may be maladaptive to the control of viral infection when it impairs interferon (IFN) responses, enhancing viral replication and spread. Dysregulated immunity as a result of inappropriate innate inflammatory responses is a hallmark of chronic viral infections such as, hepatitis B virus and hepatitis C virus (HCV). Previous studies from our laboratory have shown that expression of an IFN-stimulated gene (ISG), ubiquitin-like protease (USP)18 is upregulated in chronic HCV infection, leading to impaired hepatocyte responses to IFN-α. We examined the ability of inflammatory stimuli, including tumor necrosis factor alpha (TNF-α), lipopolysaccharide (LPS), interleukin-6 (IL-6) and IL-10 to upregulate hepatocyte USP18 expression and blunt the IFN-α response. Human hepatoma cells and primary murine hepatocytes were treated with TNF-α/LPS/IL-6/IL-10 and USP18, phosphorylated (p)-STAT1 and myxovirus (influenza virus) resistance 1 (Mx1) expression was determined. Treatment of Huh7.5 cells and primary murine hepatocytes with LPS and TNF-α, but not IL-6 or IL-10, led to upregulated USP18 expression and induced an IFN-α refractory state, which was reversed by USP18 knockdown. Liver inflammation was induced in vivo using a murine model of hepatic ischemia/reperfusion injury. Hepatic ischemia/reperfusion injury led to an induction of USP18 expression in liver tissue and promotion of lymphocytic choriomeningitis replication. These data demonstrate that certain inflammatory stimuli (TNF-α and LPS) but not others (IL-6 and IL-10) target USP18 expression and thus inhibit IFN signaling. These findings represent a new paradigm for how inflammation alters hepatic innate immune responses, with USP18 representing a potential target for intervention in various inflammatory states. IMPORTANCE Inflammation may prevent the control of viral infection when it impairs the innate immune response, enhancing viral replication and spread. Blunted immunity as a result of

  7. Tumor Necrosis Factor-alpha Induced Protein 3 Interacting Protein 1 Gene Polymorphisms and Pustular Psoriasis in Chinese Han Population

    PubMed Central

    Han, Jian-Wen; Wang, Yong; Alateng, Chulu; Li, Hong-Bin; Bai, Yun-Hua; Lyu, Xin-Xiang; Wu, Rina

    2016-01-01

    Background: Psoriasis is a common immune-mediated inflammatory dermatosis. Generalized pustular psoriasis (GPP) is the severe and rare type of psoriasis. The association between tumor necrosis factor-alpha induced protein 3 interacting protein 1 (TNIP1) gene and psoriasis was confirmed in people with multiple ethnicities. This study was to investigate the association between TNIP1 gene polymorphisms and pustular psoriasis in Chinese Han population. Methods: Seventy-three patients with GPP, 67 patients with palmoplantar pustulosis (PPP), and 476 healthy controls were collected from Chinese Han population. Six single nucleotide polymorphisms (SNPs) of the TNIP1 gene, namely rs3805435, rs3792798, rs3792797, rs869976, rs17728338, and rs999011 were genotyped by using polymerase chain reaction-ligase detection reaction. Statistical analyses were performed using the PLINK 1.07 package. Allele frequencies and genotyping frequencies for six SNPs were compared by using Chi-square test, odd ratio (OR) (including 95% confidence interval) were calculated. The haplotype analysis was conducted by Haploview software. Results: The frequencies of alleles of five SNPs were significantly different between the GPP group and the control group (P ≤ 7.22 × 10−3), especially in the GPP patients without psoriasis vulgaris (PsV). In the haplotype analysis, the most significantly different haplotype was H4: ACGAAC, with 13.1% frequency in the GPP group but only 3.4% in the control group (OR = 4.16, P = 4.459 × 10−7). However, no significant difference in the allele frequencies was found between the PPP group and control group for each of the six SNPs (P > 0.05). Conclusions: Polymorphisms in TNIP1 are associated with GPP in Chinese Han population. However, no association with PPP was found. These findings suggest that TNIP1 might be a susceptibility gene for GPP. PMID:27364786

  8. Suppressive effects of antimycotics on tumor necrosis factor-alpha-induced CCL27, CCL2, and CCL5 production in human keratinocytes.

    PubMed

    Kanda, Naoko; Watanabe, Shinichi

    2006-08-14

    Antimycotic agents are reported to improve cutaneous symptoms of atopic dermatitis or psoriasis vulgaris. Keratinocytes in these lesions excessively produce chemokines, CCL27, CCL2, or CCL5 which trigger inflammatory infiltrates. Tumor necrosis factor-alpha (TNF-alpha) induces production of these chemokines via activating nuclear factor-kappaB (NF-kappaB). We examined in vitro effects of antimycotics on TNF-alpha-induced CCL27, CCL2, and CCL5 production in human keratinocytes. Antimycotics ketoconazole and terbinafine hydrochloride suppressed TNF-alpha-induced CCL27, CCL2, and CCL5 secretion and mRNA expression in keratinocytes in parallel to the inhibition of NF-kappaB activity while fluconazole was ineffective. Anti-prostaglandin E2 (PGE2) antiserum or antisense oligonucleotides against PGE2 receptor EP2 or EP3 abrogated inhibitory effects of ketoconazole and terbinafine hydrochloride on TNF-alpha-induced NF-kappaB activity and CCL27, CCL2, and CCL5 production, indicating the involvement of endogenous PGE2 in the inhibitory effects. Prostaglandin H2, a precursor of PGE2 can be converted to thromboxane A2. Ketoconazole, terbinafine hydrochloride and thromboxane A2 synthase (EC 5.3.99.5) inhibitor, carboxyheptyl imidazole increased PGE2 release from keratinocytes and reduced that of thromboxane B2, a stable metabolite of thromboxane A2. Carboxyheptyl imidazole also suppressed TNF-alpha-induced NF-kappaB activity and CCL27, CCL2, and CCL5 production. These results suggest that ketoconazole and terbinafine hydrochloride may suppress TNF-alpha-induced NF-kappaB activity and CCL27, CCL2, and CCL5 production by increasing PGE2 release from keratinocytes. These antimycotics may suppress thromboxane A2 synthesis and redirect the conversion of PGH2 toward PGE2. These antimycotics may alleviate inflammatory infiltration in atopic dermatitis or psoriasis vulgaris by suppressing chemokine production.

  9. Regulation of semicarbazide-sensitive amine oxidase expression by tumor necrosis factor-alpha in adipocytes: functional consequences on glucose transport.

    PubMed

    Mercier, Nathalie; Moldes, Marthe; El Hadri, Khadija; Fève, Bruno

    2003-03-01

    Membrane-associated semicarbazide-sensitive amine oxidase (SSAO) is mainly present in the media of aorta and in adipose tissue. Recent works have reported that SSAO activation can stimulate glucose transport of fat cells and promote adipose conversion. In this study, the murine 3T3-L1 preadipose cell line was used to investigate SSAO regulation by tumor necrosis factor-alpha (TNF-alpha), a cytokine that is synthesized in fat cells and known to be involved in obesity-linked insulin resistance. SSAO mRNA and protein levels, and enzyme activity were decreased by TNF-alpha in a dose- and time-dependent manner, without any change of SSAO affinity for substrates or inhibitors. SSAO inhibition caused by TNF-alpha was spontaneously reversed along the time after TNF-alpha removal. The decrease in SSAO expression also occurred in white adipose tissue of C57BL/6 mice treated with mTNF-alpha. Overall, we demonstrated that reduction in SSAO expression induced by the cytokine had marked repercussions on amine-stimulated glucose transport, in a dose- and time-dependent manner. This effect was more pronounced than the inhibiting effect of TNF-alpha on insulin-stimulated glucose transport. Moreover, the peroxisome proliferator-activated receptor gamma agonists thiazolidinediones did not reverse either TNF-alpha effect on amine-sensitive glucose transport or the inhibition of SSAO activity, whereas they antagonized TNF-alpha effects on insulin-sensitive glucose transport. These results demonstrate that TNF-alpha can strongly down-regulate SSAO expression and activity, and through this mechanism can dramatically reduce amine-stimulated glucose transport. This suggests a potential role of this regulatory process in the pathogenesis of glucose homeostasis dysregulations observed during diseases accompanied by TNF-alpha overproduction, such as cachexia or obesity.

  10. Chronic sleep restriction elevates brain interleukin-1 beta and tumor necrosis factor-alpha and attenuates brain-derived neurotrophic factor expression.

    PubMed

    Zielinski, Mark R; Kim, Youngsoo; Karpova, Svetlana A; McCarley, Robert W; Strecker, Robert E; Gerashchenko, Dmitry

    2014-09-19

    Acute sleep loss increases pro-inflammatory and synaptic plasticity-related molecules in the brain, including interleukin-1 beta (IL-1β), tumor necrosis factor-alpha (TNF-α), and brain-derived neurotrophic factor (BDNF). These molecules enhance non-rapid eye movement sleep slow wave activity (SWA), also known as electroencephalogram delta power, and modulate neurocognitive performance. Evidence suggests that chronic sleep restriction (CSR), a condition prevalent in today's society, does not elicit the enhanced SWA that is seen after acute sleep loss, although it cumulatively impairs neurocognitive functioning. Rats were continuously sleep deprived for 18h per day and allowed 6h of ad libitum sleep opportunity for 1 (SR1), 3 (SR3), or 5 (SR5) successive days (i.e., CSR). IL-1β, TNF-α, and BDNF mRNA levels were determined in the somatosensory cortex, frontal cortex, hippocampus, and basal forebrain. Largely, brain IL-1β and TNF-α expression were significantly enhanced throughout CSR. In contrast, BDNF mRNA levels were similar to baseline values in the cortex after 1 day of SR and significantly lower than baseline values in the hippocampus after 5 days of SR. In the basal forebrain, BDNF expression remained elevated throughout the 5 days of CSR, although IL-1β expression was significantly reduced. The chronic elevations of IL-1β and TNF-α and inhibition of BDNF might contribute to the reported lack of SWA responses reported after CSR. Further, the CSR-induced enhancements in brain inflammatory molecules and attenuations in hippocampal BDNF might contribute to neurocognitive and vigilance detriments that occur from CSR.

  11. Synergistic effect of interferon-gamma and tumor necrosis factor-alpha on coxsackievirus and adenovirus receptor expression: an explanation of cell sloughing during testicular inflammation in mice.

    PubMed

    Gao, Ying; Lui, Wing-Yee

    2014-03-01

    Coxsackievirus and adenovirus receptor (CAR) is a junction molecule that expresses on Sertoli and germ cells. It mediates Sertoli-germ cell adhesion and facilitates migration of preleptotene/leptotene spermatocytes across the blood-testis barrier, suggesting that CAR-based cell adhesion and migration are crucial for spermatogenesis. Interferon-gamma (IFNG) and tumor necrosis factor alpha (TNF) are two major cytokines that are elevated during testicular inflammation and cause reduced fertility. We investigated the mechanism by which IFNG and TNF exert their disruptive effects on testicular cell adhesion. We have demonstrated that combined treatment with IFNG and TNF (IFNG+TNF) exerts a synergistic effect by downregulating CAR mRNA and protein levels. Immunofluorescence staining revealed that IFNG+TNF treatment effectively removes CAR from the site of cell-cell contact. Using inhibitor and co-immunoprecipitation, we confirmed that IFNG+TNF mediates CAR protein degradation via ubiquitin-proteasome and NFKB pathways. Blockage of ubiquitin-proteasome pathway significantly inhibits CAR degradation, as indicated by the reappearance of CAR at the site of cell-cell contact. Additionally, IFNG+TNF reduces CAR mRNA via transcriptional regulation. Mutational studies have shown that IFNG+TNF-induced CAR repression is achieved by suppression of the basal transcription. Electrophoretic mobility shift assay and chromatin immunoprecipitation assays further confirmed that IFNG+TNF treament not only inhibits binding of the basal transcription factors but also promotes binding of NFKB subunits and Sp1 (negative regulators) to the CAR promoter region. Taken together, IFNG+TNF treatment significantly downregulates CAR expression, which provides an explanation of how cell sloughing in the epithelium mediates, by loss of CAR-based cell adhesion, during testicular inflammation.

  12. Tumor necrosis factor alpha induces spermidine/spermine N1-acetyltransferase through nuclear factor kappaB in non-small cell lung cancer cells.

    PubMed

    Babbar, Naveen; Hacker, Amy; Huang, Yi; Casero, Robert A

    2006-08-25

    Tumor necrosis factor alpha (TNFalpha) is a potent pleiotropic cytokine produced by many cells in response to inflammatory stress. The molecular mechanisms responsible for the multiple biological activities of TNFalpha are due to its ability to activate multiple signal transduction pathways, including nuclear factor kappaB (NFkappaB), which plays critical roles in cell proliferation and survival. TNFalpha displays both apoptotic and antiapoptotic properties, depending on the nature of the stimulus and the activation status of certain signaling pathways. Here we show that TNFalpha can lead to the induction of NFkappaB signaling with a concomitant increase in spermidine/spermine N(1)-acetyltransferase (SSAT) expression in A549 and H157 non-small cell lung cancer cells. Induction of SSAT, a stress-inducible gene that encodes a rate-limiting polyamine catabolic enzyme, leads to lower intracellular polyamine contents and has been associated with decreased cell growth and increased apoptosis. Stable overexpression of a mutant, dominant negative IkappaBalpha protein led to the suppression of SSAT induction by TNFalpha in these cells, thereby substantiating a role of NFkappaB in the induction of SSAT by TNFalpha. SSAT promoter deletion constructs led to the identification of three potential NFkappaB response elements in the SSAT gene. Electromobility shift assays, chromatin immunoprecipitation experiments and mutational studies confirmed that two of the three NFkappaB response elements play an important role in the regulation of SSAT in response to TNFalpha. The results of these studies indicate that a common mediator of inflammation can lead to the induction of SSAT expression by activating the NFkappaB signaling pathway in non-small cell lung cancer cells.

  13. Phenotypic alterations in human saphenous vein culture induced by tumor necrosis factor-alpha and lipoproteins: a preliminary development of an initial atherosclerotic plaque model

    PubMed Central

    2013-01-01

    Background Atherosclerosis is a chronic progressive inflammatory disease of blood vessels particularly the arteries. The development of atherosclerotic plaques or atherogenesis is a complex process that is influenced by cardiovascular risk factors such as vascular inflammation and dyslipidemia. This study demonstrates the ability of tumor necrosis factor-alpha (TNF-α) and low density lipoproteins (LDL) to induce atherosclerotic plaque in human saphenous vein (HSV) organ culture. Methods Normal HSV segments, from male patients who had coronary bypass graft, were cultured in DMEM containing 5% heat inactivated fetal bovine serum. TNF-α (5 ng/ml) was applied in combination with native LDL (nLDL) or oxidized LDL (oxLDL) at the dose of 50 μg/ml for 14 days. The phenotypic changes of the organ cultures characteristic of initial atherosclerotic plaques were evaluated. The effect of anti-atherogenic agent, 17-β estradiol (E2), was also determined. Results Histologic, histomorphometric, and immunohistochemical examinations revealed that HSV rings stimulated with TNF-α + nLDL or TNF-α + oxLDL can exhibit the essential morphological features of atherogenesis, including fibrous cap formation, cholesterol clefts, evident thickening of the intimal layer, increased proliferation of smooth muscle cells (SMC) and migration to the subendothelial layer, significant SMC foam cell formation, and increased expression of adhesion molecules in the vascular wall. Addition of E2 (50 nM) to the culture significantly modulated the critical changes. Consistently, mRNA profiling of the HSV model revealed that 50 of 84 genes of atherosclerosis were up-regulated. Conclusions Phenotypic changes characteristic of the initial development of atherosclerotic plaques can be induced in HSV organ culture. PMID:24010774

  14. Transcription Factor AP-2α Is Preferentially Cleaved by Caspase 6 and Degraded by Proteasome during Tumor Necrosis Factor Alpha-Induced Apoptosis in Breast Cancer Cells

    PubMed Central

    Nyormoi, Okot; Wang, Zhi; Doan, Dao; Ruiz, Maribelis; McConkey, David; Bar-Eli, Menashe

    2001-01-01

    Several reports have linked activating protein 2α (AP-2α) to apoptosis, leading us to hypothesize that AP-2α is a substrate for caspases. We tested this hypothesis by examining the effects of tumor necrosis factor alpha (TNF-α) on the expression of AP-2 in breast cancer cells. Here, we provide evidence that TNF-α downregulates AP-2α and AP-2γ expression posttranscriptionally during TNF-α-induced apoptosis. Both a general caspase antagonist (zVADfmk) and a caspase 6-preferred antagonist (zVEIDfmk) inhibited TNF-α-induced apoptosis and AP-2α downregulation. In vivo tests showed that AP-2α was cleaved by caspases ahead of the DNA fragmentation phase of apoptosis. Recombinant caspase 6 cleaved AP-2α preferentially, although caspases 1 and 3 also cleaved it, albeit at 50-fold or higher concentrations. Activated caspase 6 was detected in TNF-α-treated cells, thus confirming its involvement in AP-2α cleavage. All three caspases cleaved AP-2α at asp19 of the sequence asp-arg-his-asp (DRHD19). Mutating D19 to A19 abrogated AP-2α cleavage by all three caspases. TNF-α-induced cleavage of AP-2α in vivo led to AP-2α degradation and loss of DNA-binding activity, both of which were prevented by pretreatment with zVEIDfmk. AP-2α degradation but not cleavage was inhibited in vivo by PS-431 (a proteasome antagonist), suggesting that AP-2α is degraded subsequent to cleavage by caspase 6 or caspase 6-like enzymes. Cells transfected with green fluorescent protein-tagged mutant AP-2α are resistant to TNF-α-induced apoptosis, further demonstrating the link between caspase-mediated cleavage of AP-2α and apoptosis. This is the first report to demonstrate that degradation of AP-2α is a critical event in TNF-α-induced apoptosis. Since the DRHD sequence in vertebrate AP-2 is widely conserved, its cleavage by caspases may represent an important mechanism for regulating cell survival, proliferation, differentiation, and apoptosis. PMID:11438643

  15. Tumor necrosis factor alpha leads to increased cell surface expression of CXCR4 in SK-N-MC cells.

    PubMed

    Rostasy, Kevin; Gorgun, Gullue; Kleyner, Yelena; Garcia, Anthony; Kramer, Michael; Melanson, Suzanne M; Mathys, Jean Marie; Yiannoutsos, Constantin; Skolnik, Paul R; Navia, Bradford A

    2005-07-01

    Both host and viral factors play an important role in the pathogenesis of human immunodeficiency virus (HIV)-associated bran injury. In this study, the authors examined the interactions between tumor necrosis factor (TNF)-alpha, CXCR4, the alpha chemokine receptor, and three HIV isolates, including the T-tropic viruses, HIV-1(MN) and HIV-1(IIIB), and the dual tropic virus, HIV-1(89.6). The authors show by flow cytometry that treatment of differentiated SK-N-MC cells with TNF-alpha induces a significant increase in the cell surface expression of CXCR4 in a time- and dose-dependent manner. The effect is partly regulated at the level of transcription. To assess the biological significance of this finding, we show that TNF-alpha potentiates the ability of the above mentioned HIV isolates to induce neuronal apoptosis and that the effect is significantly reduced by pretreating cells with monoclonal antibodies to either CXCR4 and TNF-alpha. Together these results suggest that TNF-alpha may render neuronal cells vulnerable to the apoptotic effects of HIV by increasing the cell surface expression of CXCR4 and thus identify another mechanism by which TNF-alpha contributes to the pathogenesis of HIV-associated brain injury.

  16. Evaluation of a novel subtilisin inhibitor gene and mutant derivatives for the expression and secretion of mouse tumor necrosis factor alpha by Streptomyces lividans.

    PubMed Central

    Lammertyn, E; Van Mellaert, L; Schacht, S; Dillen, C; Sablon, E; Van Broekhoven, A; Anné, J

    1997-01-01

    In order to evaluate the expression and secretion signals of the highly secreted subtilisin inhibitor of Streptomyces venezuelae CBS762.70 (VSI) for the production of heterologous proteins by Streptomyces lividans, mouse tumor necrosis factor alpha (mTNF) was chosen as a model protein. The mTNF cDNA was fused to the vsi signal sequence. The analysis of secretion by sodium dodecyl sulfate-polyacrylamide gel electrophoresis and biological activity measurements revealed an efficient translocation of mTNF. Up to 300 mg of secreted biologically active mTNF per liter could be obtained in shaken-flask cultures. By analyzing the effects of mutations in the N region of the VSI signal peptide on secretion, we found that decreasing the +3 charge of the wild-type protein to +2 resulted in a 3- to 10-fold increase in secretion. PMID:9143114

  17. Experimental study on inhibitory effect of niacinamide on tumor necrosis factor-alpha-induced matrix degradation of annulus fibrous tissue in vitro.

    PubMed

    Xu, Runbing; Shao, Zengwu; Xiong, Liming

    2008-10-01

    The inhibitory effect of niacinamide on tumor necrosis factor-alpha (TNF-alpha) induced annulus fibrous (AF) degradation was assessed, and the mechanism of the inhibition was investigated. Chiba's intervertebral disc (IVD) culture model was established. Forty-eight IVDs from 12 adult Japanese white rabbits were randomly divided into 4 groups (12 IVDs in each group), and various concentrations of niacinamide and TNF-alpha were added to the medium for intervention: negative control group, niacinamide control group (0.5 mg/mL niacinamide), degeneration group (10 ng/mL TNF-alpha), and treatment group (0.5 mg/mL niacinamide and 10 ng/mL TNF-alpha). After one week's culture, AFs were collected for glycosaminoglycan (GS) content measurement, safranin O-fast green staining, and immunohistochemical staining for type I, II collagen and cysteine containing aspartate specific protease-3 (Caspase-3). It was found that the GS content in treatment group was increased by about 48% as compared with degeneration group (t=16.93, P<0.001), and close to that in niacinamide control group (t=0.71, P=0.667). Safranine O-fast green staining exhibited higher staining density and better histological structure of AF in the treatment group as compared with the degeneration group. Immunohistochemical staining for both Type I and II collagen demonstrated that lamellar structure and continuity of collagen in treatment group were better reserved than in degeneration group. Positive staining rate of Caspase-3 in AFs of negative control group, niacinamide control group, degeneration group and treatment group was 3.4%, 4.3%, 17.9% and 10.3% respectively. The positive rate in treatment group was significantly lower than in degeneration group (P<0.01). It was concluded that niacinamide could effectively alleviate TNF-alpha induced destruction and synthesis inhibition of matrix ingredients in AFs. The inhibition may be related with reduction of expression of Caspase-3. Thus, niacinamide is of potential

  18. Garlic (Allium sativum) stimulates lipopolysaccharide-induced tumor necrosis factor-alpha production from J774A.1 murine macrophages.

    PubMed

    Sung, Jessica; Harfouche, Youssef; De La Cruz, Melissa; Zamora, Martha P; Liu, Yan; Rego, James A; Buckley, Nancy E

    2015-02-01

    Garlic (Allium sativum) is known to have many beneficial attributes such as antimicrobial, antiatherosclerotic, antitumorigenetic, and immunomodulatory properties. In the present study, we investigated the effects of an aqueous garlic extract on macrophage cytokine production by challenging the macrophage J774A.1 cell line with the garlic extract in the absence or presence of lipopolysaccharide (LPS) under different conditions. The effect of allicin, the major component of crushed garlic, was also investigated. Using enzyme-linked immunosorbent assay and reverse transcriptase-quantitative polymerase chain reaction, it was found that garlic and synthetic allicin greatly stimulated tumor necrosis factor-alpha (TNF-α) production in macrophages treated with LPS. The TNF-α secretion levels peaked earlier and were sustained for a longer time in cells treated with garlic and LPS compared with cells treated with LPS alone. Garlic acted in a time-dependent manner. We suggest that garlic, at least partially via its allicin component, acts downstream from LPS to stimulate macrophage TNF-α secretion.

  19. Tumor necrosis factor-alpha increases reactive oxygen species by inducing spermine oxidase in human lung epithelial cells: a potential mechanism for inflammation-induced carcinogenesis.

    PubMed

    Babbar, Naveen; Casero, Robert A

    2006-12-01

    Inflammation has been implicated in the development of many human epithelial cancers, including those of the stomach, lung, colon, and prostate. Tumor necrosis factor-alpha (TNF-alpha) is a potent pleiotropic, proinflammatory cytokine produced by many cells in response to injury and inflammation. Here, we show that TNF-alpha exposure results in increased production of reactive oxygen species (ROS), with a concomitant increase in the production of 8-oxo-deoxyguanosine, a marker for oxidative DNA damage, in human lung bronchial epithelial cells. The source of the ROS in TNF-alpha-treated cells was determined by both pharmacologic and small interfering RNA (siRNA) strategies to be spermine oxidase (SMO/PAOh1). SMO/PAOh1 oxidizes spermine into spermidine, 3-aminopropanal, and H(2)O(2). Inhibition of TNF-alpha-induced SMO/PAOh1 activity with MDL 72,527 or with a targeted siRNA prevented ROS production and oxidative DNA damage. Further, similar induction in SMO/PAOh1 is observed with treatment of another inflammatory cytokine, interleukin-6. The data are consistent with a model that directly links inflammation and DNA damage through the production of H(2)O(2) by SMO/PAOh1. Further, these results suggest a common mechanism by which inflammation from multiple sources can lead to the mutagenic changes necessary for the development and progression of epithelial cancers.

  20. Role of hypoxia-inducible factor-{alpha} in hepatitis-B-virus X protein-mediated MDR1 activation

    SciTech Connect

    Han, Hyo-Kyung; Han, Chang Yeob; Cheon, Eun-Pa; Lee, Jaewon; Kang, Keon Wook . E-mail: kwkang@chosun.ac.kr

    2007-06-01

    The transition from chemotherapy-responsive cancer cells to chemotherapy-resistant cancer cells is mainly accompanied by the increased expression of multi-drug resistance 1 (MDR1). We found that hepatitis-B-virus X protein (HBx) increases the transcriptional activity and protein level of MDR1 in a hepatoma cell line, H4IIE. In addition, HBx overexpression made H4IIE cells more resistant to verapamil-uptake. HBx stabilized hypoxia-inducible factor-1{alpha} (HIF-1{alpha}) and induced the nuclear translocation of C/EBP{beta}. Reporter gene analyses showed that HBx increased the reporter activity in the cells transfected with the reporter containing MDR1 gene promoter. Moreover, the luciferase reporter gene activity was significantly inhibited by HIF-1{alpha} siRNA but not by overexpression of C/EBP dominant negative mutant. These results imply that HBx increases the MDR1 transporter activity through the transcriptional activation of the MDR1 gene with HIF-1{alpha} activation, and suggest HIF-1{alpha} for the therapeutic target of HBV-mediated chemoresistance.

  1. Expression of anti-tumor necrosis factor alpha (TNFα) single-chain variable fragment (scFv) in Spirodela punctata plants transformed with Agrobacterium tumefaciens.

    PubMed

    Balaji, Parthasarathy; Satheeshkumar, P K; Venkataraman, Krishnan; Vijayalakshmi, M A

    2016-05-01

    Therapeutic antibodies against tumor necrosis factor alpha (TNFα) have been considered effective for some of the autoimmune diseases such as rheumatoid arthritis, Crohn's diseases, and so on. But associated limitations of the current therapeutics in terms of cost, availability, and immunogenicity have necessitated the need for alternative candidates. Single-chain variable fragment (scFv) can negate the limitations tagged with the anti-TNFα therapeutics to a greater extent. In the present study, Spirodela punctata plants were transformed with anti-TNFα through in planta transformation using Agrobacterium tumefaciens strain, EHA105. Instead of cefotaxime, garlic extract (1 mg/mL) was used to remove the agrobacterial cells after cocultivation. To the best of our knowledge, this report shows for the first time the application of plant extracts in transgenic plant development. 95% of the plants survived screening under hygromycin. ScFv cDNA integration in the plant genomic DNA was confirmed at the molecular level by PCR. The transgenic protein expression was followed up to 10 months. Expression of scFv was confirmed by immunodot blot. Protein expression levels of up to 6.3% of total soluble protein were observed. β-Glucuronidase and green fluorescent protein expressions were also detected in the antibiotic resistant plants. The paper shows the generation of transgenic Spirodela punctuata plants through in planta transformation.

  2. Effects of doxepin on brain-derived neurotrophic factor, tumor necrosis factor alpha, mitogen-activated protein kinase 14, and AKT1 genes expression in rat hippocampus

    PubMed Central

    Eidelkhani, Nastaran; Radahmadi, Maryam; Kazemi, Mohammad; Rafiee, Laleh; Alaei, Hojjatallah; Reisi, Parham

    2015-01-01

    Background: It has been suggested that doxepin in addition to enhancement of noradrenaline and serotonin levels may have neuroprotective effects. Therefore, this study investigated the effect of doxepin on gene expression of brain-derived neurotrophic factor (BDNF), tumor necrosis factor alpha (TNF-α), mitogen-activated protein kinase 14 (MAPK14), and serine-threonine protein kinase AKT1 in rat hippocampus. Materials and Methods: Male rats were divided randomly into three groups: Control, doxepin 1 mg/kg, and doxepin 5 mg/kg. Rats received an i.p injection of doxepin for 21 days. Then the hippocampi were dissected for the measurement of the expression of BDNF, TNF-α, MAPK14, and AKT1 genes. Results: Our results showed no significant effects of doxepin on gene expression of BDNF, TNF-α, MAPK14, and AKT1 genes in the hippocampus. Conclusions: These results did not show significant effects of doxepin on the genes that affect the neuronal survival in intact animals. However, more studies need to be done, especially in models associated with neuronal damage. PMID:26601091

  3. Human eosinophils can express the cytokines tumor necrosis factor-alpha and macrophage inflammatory protein-1 alpha.

    PubMed Central

    Costa, J J; Matossian, K; Resnick, M B; Beil, W J; Wong, D T; Gordon, J R; Dvorak, A M; Weller, P F; Galli, S J

    1993-01-01

    By in situ hybridization, 44-100% of the blood eosinophils from five patients with hypereosinophilia and four normal subjects exhibited intense hybridization signals for TNF-alpha mRNA. TNF-alpha protein was detectable by immunohistochemistry in blood eosinophils of hypereosinophilic subjects, and purified blood eosinophils from three atopic donors exhibited cycloheximide-inhibitable spontaneous release of TNF-alpha in vitro. Many blood eosinophils (39-91%) from hypereosinophilic donors exhibited intense labeling for macrophage inflammatory protein-1 alpha (MIP-1 alpha) mRNA, whereas eosinophils of normal donors demonstrated only weak or undetectable hybridization signals for MIP-1 alpha mRNA. Most tissue eosinophils infiltrating nasal polyps were strongly positive for both TNF-alpha and MIP-1 alpha mRNA. By Northern blot analysis, highly enriched blood eosinophils from a patient with the idiopathic hypereosinophilic syndrome exhibited differential expression of TNF-alpha and MIP-1 alpha mRNA. These findings indicate that human eosinophils represent a potential source of TNF-alpha and MIP-1 alpha, that levels of expression of mRNA for both cytokines are high in the blood eosinophils of hypereosinophilic donors and in eosinophils infiltrating nasal polyps, that the eosinophils of normal subjects express higher levels of TNF-alpha than MIP-1 alpha mRNA, and that eosinophils purified from the blood of atopic donors can release TNF-alpha in vitro. Images PMID:8514874

  4. Functional NF-IL6/CCAAT enhancer-binding protein is required for tumor necrosis factor alpha-inducible expression of the granulocyte colony- stimulating factor (CSF), but not the granulocyte/macrophage CSF or interleukin 6 gene in human fibroblasts [retracted by Adler G. In: J Exp Med 1997 Jul 7;186(1):171

    PubMed Central

    1995-01-01

    Tumor necrosis factor (TNF) alpha participates in the regulation of the acute-phase, immune, and inflammatory responses. Target genes known to be transcriptionally activated by TNF-alpha include the granulocyte (G)- colony-stimulating factor (CSF) gene, the granulocyte/macrophage (GM)- CSF gene, as well as the interleukin (IL) 6 gene. Functional nuclear factor (NF)-IL6 recognition sites have been identified in regulatory regions of these genes by transient transfection studies using deleted promoter constructs. In addition, NF-IL6 is known to form heterodimeric complexes with the NF-kappa B transcription factor, which is also engaged in the transcriptional regulation of these genes. The indispensable importance of NF-IL6 for regulating gene expression of proinflammatory cytokine genes in response to inflammatory stimuli in vivo remains, however, unclear. We here report, by using both antisense (AS) oligodesoxyribonucleotide (ODN) and ribozyme (RZ)-mediated specific elimination of NF-IL6 transcripts in human fibroblasts, that TNF-alpha-induced synthesis of G-CSF, but not of GM-CSF or IL-6, is abolished in the absence of functional NF-IL6 in vivo. Both AS ODN and RZ targeting of the NF-IL6 transcript eliminate NF-IL6 protein, as shown in Western blot analysis and electrophoretic mobility shift assays. Similarly, fibroblasts exposed to either the AS NF-IL6 ODN or the NF-IL6 RZ, but not to the sense or nonsense ODN or a mutated ribozyme, also failed to respond with functional activation of NF-IL6 as assayed in transient transfection studies using heterologous promoter constructs harboring the NF-IL6 recognition site. In contrast, protein synthesis, DNA-binding activity, and transcriptional activation capacity of the NF-kappa B transcription factor is not impaired upon exposure to either ODN or RZ. Fibroblasts that had been cultured in the presence of the AS NF-IL6 ODN or the NF-IL6RZ failed to synthesize G- CSF protein in response to TNF-alpha, while TNF-alpha-inducible

  5. Molecular cloning and expression analysis of sea bass (Dicentrarchus labrax L.) tumor necrosis factor-alpha (TNF-alpha).

    PubMed

    Nascimento, Diana S; Pereira, Pedro J B; Reis, Marta I R; do Vale, Ana; Zou, Jun; Silva, Manuel T; Secombes, Christopher J; dos Santos, Nuno M S

    2007-09-01

    In the search for pro-inflammatory genes in sea bass a TNF-alpha gene was cloned and sequenced. The sea bass TNF-alpha (sbTNF-alpha) putative protein conserves the TNF-alpha family signature, as well as the two cysteines usually involved in the formation of a disulfide bond. The mouse TNF-alpha Thr-Leu cleavage sequence and a potential transmembrane domain were also found, suggesting that sbTNF-alpha exists as two forms: a approximately 28 kDa membrane-bound form and a approximately 18.4 kDa soluble protein. The single copy sbTNF-alpha gene contains a four exon-three intron structure similar to other known TNF-alpha genes. Homology modeling of sbTNF-alpha is compatible with the trimeric quaternary architecture of its mammalian counterparts. SbTNF-alpha is constitutively expressed in several unstimulated tissues, and was not up-regulated in the spleen and head-kidney, in response to UV-killed Photobacterium damselae subsp. piscicida. However, an increase of sbTNF-alpha expression was detected in the head-kidney during an experimental infection using the same pathogen.

  6. Trovafloxacin-induced Replication Stress Sensitizes HepG2 Cells to Tumor Necrosis Factor-alpha-induced Cytotoxicity Mediated by Extracellular Signal-regulated Kinase and Ataxia Telangiectasia and Rad3-related

    PubMed Central

    Beggs, Kevin M.; Maiuri, Ashley R.; Fullerton, Aaron M.; Poulsen, Kyle L.; Breier, Anna B.; Ganey, Patricia E.; Roth, Robert A.

    2015-01-01

    Use of the fluoroquinolone antibiotic trovafloxacin (TVX) was restricted due to idiosyncratic, drug-induced liver injury (IDILI). Previous studies demonstrated that tumor necrosis factor-alpha (TNF) and TVX interact to cause death of hepatocytes in vitro that was associated with prolonged activation of c-Jun N-terminal kinase (JNK), activation of caspases 9 and 3, and DNA damage. The purpose of this study was to explore further the mechanism by which TVX interacts with TNF to cause cytotoxicity. Treatment with TVX caused cell cycle arrest, enhanced expression of p21 and impaired proliferation, but cell death only occurred after cotreatment with TVX and TNF. Cell death involved activation of extracellular signal-related kinase (ERK), which in turn activated caspase 3 and ataxia telangiectasia and Rad3-related (ATR), both of which contributed to cytotoxicity. Cotreatment of HepG2 cells with TVX and TNF caused double-strand breaks in DNA, and ERK contributed to this effect. Inhibition of caspase activity abolished the DNA strand breaks. The data suggest a complex interaction of TVX and TNF in which TVX causes replication stress, and the downstream effects are exacerbated by TNF, leading to hepatocellular death. These results raise the possibility that IDILI from TVX results from MAPK and ATR activation in hepatocytes initiated by interaction of cytokine signaling with drug-induced replication stress. PMID:25748550

  7. The herbal medicine shoseiryu-to inhibits allergen-induced synthesis of tumour necrosis factor alpha by peripheral blood mononuclear cells in patients with perennial allergic rhinitis.

    PubMed

    Tanaka, A; Ohashi, Y; Kakinoki, Y; Washio, Y; Yamada, K; Nakai, Y; Nakano, T; Nakai, Y; Ohmoto, Y

    1998-01-01

    The herbal medicine shoseiryu-to is an effective agent in the treatment of allergic rhinitis. However, the mechanism by which it exerts its action in improving patient symptoms remains unclear. It might affect the allergen-induced TH1 and/or TH2 responses. This study investigated whether the herbal medicine could affect cytokine synthesis by peripheral blood mononuclear cells (PBMCs) in response to the major Dermatophagoides farinae (D. farinae) allergen, Der f 1. PBMCs were obtained from 15 patients with perennial allergic rhinitis due to D. farinae, and were stimulated for 96 h with 10 micrograms/ml Der f 1 in the presence or absence of 45 mg/ml shoseiryu-to. The culture supernatants were harvested to determine the synthesis of IgE, interleukin 5 (IL-5), IL-6, IL-10, interferon-gamma (IFN-gamma) and tumour necrosis factor alpha (TNF-alpha). The agent did not affect the allergen-induced synthesis of IL-5, IL-6 and IFN-gamma, but somewhat decreased the synthesis of IgE and IL-10. This study highlighted an interesting pharmacological action of shoseiryu-to to substantially inhibit the allergen-induced synthesis of TNF-alpha. Our study suggests that the shoseiryu-to may alleviate nasal symptoms in allergic rhinitis through control of the allergen-induced inflammatory process.

  8. Sera of patients with high titers of immunoglobulin G against Toxoplasma gondii induce secretion of tumor necrosis factor alpha by human monocytes.

    PubMed Central

    Pelloux, H; Chumpitazi, B F; Santoro, F; Polack, B; Vuillez, J P; Ambroise-Thomas, P

    1992-01-01

    Toxoplasma gondii alone does not induce tumor necrosis factor alpha (TNF-alpha) secretion by human monocytes and macrophages. Nevertheless, sera from infected patients with high titers of specific immunoglobulin G antibodies against T. gondii induce TNF-alpha secretion, which is significantly higher than the corresponding induction by negative sera (P less than 0.05). After incubation with the positive serum, parasites also induce secretion of this cytokine, but TNF-alpha levels are lower (11.4 to 71.8%) than those obtained with positive serum alone. Therefore, this secretion seems to be elicited in part by antibody-T. gondii complexes and/or another unidentified factor(s), probably different from lipopolysaccharide, interleukin-1, TNF-alpha, and gamma interferon. In this study, monocytes secreted more TNF-alpha into the culture fluid than macrophages did (P less than 0.05), and no correlation was observed between secretion of this cytokine by the monocytes and the intracellular multiplication of the parasites, evaluated by [3H]uracil incorporation. Sera from patients with other infections diseases did not induce secretion of TNF-alpha; however, serum free of antibodies to T. gondii, obtained from patients with leishmaniosis, also stimulated secretion of the cytokine. PMID:1612737

  9. Omega-3 fatty acids decreases oxidative stress, tumor necrosis factor-alpha, and interleukin-1 beta in hyperthyroidism-induced hepatic dysfunction rat model.

    PubMed

    Gomaa, Asmaa M S; Abd El-Aziz, Ebtihal A

    2016-12-01

    Hyperthyroidism is associated with abnormalities of the liver. Omega-3 polyunsaturated fatty acids, especially their long-chain forms: eicosapentaenoic acid (EPA) and docosahexaenoic acid (DHA) have beneficial health effects. The objectives of the present study were to assess hyperthyroidism-induced hepatic dysfunction in adult male rats and to evaluate the ameliorative effects of omega-3 on hyperthyroidism-induced hepatic dysfunction and the underlying mechanisms. Twenty four adult male rats were randomly divided into three equal groups; control group which received water for 6 weeks, hyperthyroid group which received L-thyroxine orally for 6 weeks and hyperthyroid omega-3 treated group which received L-thyroxine for 2 weeks and then co-treated with L-thyroxine and omega-3 oral compound containing 18% of EPA and 12% of DHA for 4 weeks. Hyperthyroid omega-3 treated group showed significantly increased final body weight and body weight gain, decreased liver weight to body weight ratio, decreased serum triiodo-l-thyronine level, increased serum thyroid stimulating hormone level, decreased serum levels of alanine transaminase, aspartate transaminase and tumor necrosis factor-alpha, increased hepatic levels of total antioxidant capacity and decreased hepatic levels of total peroxide and interleukin-1 beta when compared with the hyperthyroid group. Furthermore, histopathological studies revealed also marked improvement. We concluded that omega-3 had encouraging therapeutic effects against hyperthyroidism-induced hepatic dysfunction attributable to more than one mechanism: antioxidant, anti-inflammatory and anti-fibrotic effects.

  10. Dexamethasone inhibits induction of liver tumor necrosis factor-alpha mRNA and liver growth induced by lead nitrate and ethylene dibromide.

    PubMed Central

    Ledda-Columbano, G. M.; Columbano, A.; Cannas, A.; Simbula, G.; Okita, K.; Kayano, K.; Kubo, Y.; Katyal, S. L.; Shinozuka, H.

    1994-01-01

    We have recently demonstrated that a single injection of the mitogen lead nitrate to rats induced a rapid increase of tumor necrosis factor-alpha (TNF-alpha) mRNA in the liver and suggested that this cytokine may be involved in triggering hepatocyte proliferation in this model of direct hyperplasia. In this study, we examined whether a similar induction of liver TNF-alpha mRNA could be observed preceding the onset of hepatocyte proliferation induced by ethylene dibromide, another hepatocyte mitogen. In addition, we used dexamethasone, a well known inhibitor of TNF-alpha production, to determine whether its administration could suppress hepatocyte proliferation induced by lead nitrate and ethylene dibromide. A single intragastric administration of ethylene dibromide (100 mg/kg) to male Wistar rats enhanced liver TNF-alpha mRNA after 4 and 7 hours, which then returned to control levels by 24 hours. TNF-alpha mRNA was detectable only in a nonparenchymal cell fraction of the liver. Pretreatment of rats with a single dose of dexamethasone (2 mg/kg) 60 minutes before lead nitrate (100 mumol/kg) or ethylene dibromide completely abolished the increased levels of liver TNF-alpha mRNA induced by these agents. Inhibition by dexamethasone of TNF-alpha mRNA was associated with an inhibition of liver cell proliferation induced by these mitogens, as measured by [3H]thymidine incorporation into hepatic DNA, mitotic index, and DNA content. These results further support the hypothesis that TNF-alpha may be involved in triggering hepatocyte proliferation induced by primary mitogens. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 PMID:7943184

  11. Lactate induces tumour necrosis factor-alpha, interleukin-6 and interleukin-1beta release in microglial- and astroglial-enriched primary cultures.

    PubMed

    Andersson, Anna K; Rönnbäck, Lars; Hansson, Elisabeth

    2005-06-01

    Hyperammonaemia has deleterious effects on the CNS in patients with liver dysfunction. Cellular mechanisms underlying the effects of hyperammonaemia are largely unknown, although astrocytes have been the main target of interest. This study investigated how treatment with NH4Cl and lactate, which increase in the brain as a consequence of hyperammonaemia, affects cells in primary rat cultures enriched in either astrocytes or microglia. Morphological changes were studied over time using light microscopy. Release of the proinflammatory cytokines tumour necrosis factor-alpha (TNF-alpha), interleukin (IL)-6 and IL-1beta was measured using ELISA. NH4Cl was found to induce vacuole formation in both culture systems. Lactate treatment altered astrocytic appearance, resulting in increased space between individual cells. Microglia adopted a round morphology with either NH4Cl or lactate treatment. Lactate, but not NH4Cl, induced release of TNF-alpha and IL-6 in both astroglial- and microglial-enriched cultures, while IL-1beta was released only in microglial cultures. Cytokine release was higher in the microglial- than in the astroglial-enriched cultures. Additionally, the astroglial-enriched cultures containing approximately 10% microglial cells released more cytokines than cultures containing about 5% microglial cells. Taken together, our data suggest that most TNF-alpha, IL-6 and IL-1beta release comes from microglia. Thus, microglia could play an important role in the pathological process of hyperammonaemia.

  12. Modulator effects of interleukin-1beta and tumor necrosis factor-alpha on AMPA-induced excitotoxicity in mouse organotypic hippocampal slice cultures.

    PubMed

    Bernardino, Liliana; Xapelli, Sara; Silva, Ana P; Jakobsen, Birthe; Poulsen, Frantz R; Oliveira, Catarina R; Vezzani, Annamaria; Malva, João O; Zimmer, Jens

    2005-07-20

    The inflammatory cytokines interleukin-1beta and tumor necrosis factor-alpha (TNF-alpha) have been identified as mediators of several forms of neurodegeneration in the brain. However, they can produce either deleterious or beneficial effects on neuronal function. We investigated the effects of these cytokines on neuronal death caused by exposure of mouse organotypic hippocampal slice cultures to toxic concentrations of AMPA. Either potentiation of excitotoxicity or neuroprotection was observed, depending on the concentration of the cytokines and the timing of exposure. A relatively high concentration of mouse recombinant TNF-alpha (10 ng/ml) enhanced excitotoxicity when the cultures were simultaneously exposed to AMPA and to this cytokine. Decreasing the concentration of TNF-alpha to 1 ng/ml resulted in neuroprotection against AMPA-induced neuronal death independently on the application protocol. By using TNF-alpha receptor (TNFR) knock-out mice, we demonstrated that the potentiation of AMPA-induced toxicity by TNF-alpha involves TNF receptor-1, whereas the neuroprotective effect is mediated by TNF receptor-2. AMPA exposure was associated with activation and proliferation of microglia as assessed by macrophage antigen-1 and bromodeoxyuridine immunohistochemistry, suggesting a functional recruitment of cytokine-producing cells at sites of neurodegeneration. Together, these findings are relevant for understanding the role of proinflammatory cytokines and microglia activation in acute and chronic excitotoxic conditions.

  13. ROLES OF EPIDERMAL GROWTH FACTOR (EGF) AND TRANSFORMING GROWTH FACTOR-ALPHA (TGF-A) IN MEDIATION OF DIOXIN (TCDD)-INDUCED DELAYS IN DEVELOPMENT OF THE MOUSE MAMMARY GLAND

    EPA Science Inventory

    Roles of Epidermal Growth Factor (EGF) and Transforming Growth Factor-alpha (TGF-a) in Mediation of Dioxin (TCDD)-Induced Delays in Development of the Mouse Mammary Gland.
    Suzanne E. Fenton, Barbara Abbott, Lamont Bryant, and Angela Buckalew. U.S. EPA, NHEERL, Reproductive Tox...

  14. Tumor necrosis factor-alpha inversely regulates prostaglandin D2 and prostaglandin E2 production in murine macrophages. Synergistic action of cyclic AMP on cyclooxygenase-2 expression and prostaglandin E2 synthesis.

    PubMed

    Fournier, T; Fadok, V; Henson, P M

    1997-12-05

    Increased synthesis of insulin-like growth factor-1 is induced in murine macrophages by prostaglandin E2 (PGE2) and tumor necrosis factor-alpha (TNFalpha). Accordingly, we have investigated mechanisms regulating synthesis of PGE2 that might contribute to autocrine/paracrine effects on insulin-like growth factor-1 production. In response to zymosan, TNFalpha specifically induced a 5-fold increase in PGE2 synthesis, at the same time decreasing PGD2 production in a reciprocal fashion. Activators of cyclic AMP-dependent protein kinase (PKA), such as PGE2 itself or dibutyryl cyclic AMP, did not modify PGE2 production by themselves but potentiated the TNFalpha-induced increase in PGE2; this effect required both RNA and protein synthesis. No significant change in arachidonate release or production of other eicosanoids was observed. The inducible form of cyclooxygenase-2 (COX2) but not of the constitutive form COX1 was implicated in the generation of both PGE2 and PGD2 in these cells by use of specific inhibitors and effects of dexamethasone. Neither COX1 nor COX2 protein levels were affected by TNFalpha or PKA activators used alone, whereas in association, marked up-regulation of COX2 mRNA and protein was observed. Incubations of cells carried out with PGH2 demonstrated that PGE2 synthase activity was increased after a TNFalpha pretreatment. Taken together, our results suggest that TNFalpha induced a switch from the PGD2 to PGE2 synthesis pathway by regulating PGE2 synthase expression and/or activity and that activators of PKA markedly potentiated the TNFalpha-induced increase in PGE2 through up-regulation of COX2 gene expression.

  15. Interleukin-1beta, -6, -12 and tumor necrosis factor-alpha expression in brains of dogs with canine distemper virus infection.

    PubMed

    Gröne, A; Alldinger, S; Baumgärtner, W

    2000-10-02

    Canine distemper virus infection in dogs is commonly associated with demyelinating central nervous system lesions. Investigations on viral protein expression by studying mRNA and protein distribution together with the characterization of CD4 and CD8 inflammatory cells and MHC class II up-regulation revealed a biphasic disease process. To further investigate the cellular interactions in the different plaque types the cerebella of 14 dogs with confirmed distemper infection were investigated for expression of interleukin (IL)-1beta, -6, -12 and tumor necrosis factor-alpha (TNF) by immunohistochemistry using rabbit polyclonal anti-cytokine antibodies. T-cells and astrocytes were identified with rabbit anti CD3- and GFAP-monoclonal and polyclonal antibodies, respectively; and microglia/macrophages were characterized by their ability to bind lectin from Bandeiraea simplicifolia (BS-1). To further name the cytokine expressing cells immunoenzymatic double staining using DAB and New Fuchsin was performed. White matter lesions were classified according to histopathological criteria into acute, subacute and chronic. Canine distemper virus nucleoprotein antigen was demonstrated in nearly all plaques, except in older plaques where virus was not present within the plaque but adjacent to the lesion. IL-1 expression was observed to varying degrees in all types of lesions. Most often IL-1 was present in CD3 and BS-1 positive cells in the brain parenchyma in earlier plaques and comprising perivascular cuffs found in chronic plaques. IL-6 expression was present in all lesions, and followed a similar distribution pattern as IL-1. IL-12 displayed very often a granular extracellular pattern of immunoreactivity, especially in the brain parenchyma, and was found only in individual perivascular cells. TNF staining, predominantly found in astrocytes, was present in lesions of various types; however, staining appeared to be stronger in acute lesions and decreased in chronic plaques. In the

  16. Epigallocatechin-3-gallate protects against tumor necrosis factor alpha induced inhibition of osteogenesis of mesenchymal stem cells.

    PubMed

    Liu, Wei; Fan, Jian-Bo; Xu, Da-Wei; Zhang, Jie; Cui, Zhi-Ming

    2016-03-01

    Anabolic bone accruement through osteogenic differentiation is important for the maintenance of physiological bone mass and often disrupted in various inflammatory diseases. Epigallocatechin-3-gallate, as an antioxidant and anti-inflammatory agent, has been suggested for potential therapeutic use in this context, possibly by the inhibition of bone resorption as well as the enhancement of bone formation through directly activating osteoblast differentiation. However, the reported effects of epigallocatechin-3-gallate modulating osteoblast differentiation are mixed, and the underlying molecular mechanism is still elusive. Moreover, there is limited information regarding the effects of epigallocatechin-3-gallate on osteogenic potential of mesenchymal stem cell in inflammation. Here, we examined the in vitro osteogenic differentiation of human mesenchymal stem cells. We found that the cell viability and osteoblast differentiation of human bone marrow-derived mesenchymal stem cells are significantly inhibited by inflammatory cytokine TNFα treatment. Epigallocatechin-3-gallate is able to enhance the cell viability and osteoblast differentiation of mesenchymal stem cells and is capable of reversing the TNFα-induced inhibition. Notably, only low doses of epigallocatechin-3-gallate have such benefits, which potentially act through the inhibition of NF-κB signaling that is stimulated by TNFα. These data altogether clarify the controversy on epigallocatechin-3-gallate promoting osteoblast differentiation and further provide molecular basis for the putative clinical use of epigallocatechin-3-gallate in stem cell-based bone regeneration for inflammatory bone loss diseases, such as rheumatoid arthritis and prosthetic osteolysis.

  17. Ataxia-telangiectasia mutated activation mediates tumor necrosis factor-alpha induced MMP-13 up-regulation and metastasis in lung cancer cells

    PubMed Central

    Yan, Hong Qiong; Zhang, Di; Shi, Yuan Yuan; You, Xiang; Shi, Lei; Li, Qing; Gao, Feng Guang

    2016-01-01

    Despite that ataxia-telangiectasia mutated (ATM) is involved in IL-6 promoted lung cancer chemotherapeutic resistance and metastasis, the exact role of ATM in tumor necrosis factor-alpha (TNF-α) increasing tumor migration is still elusive. In the present study, we demonstrated that TNF-α promoted lung cancer cell migration by up-regulation of matrix metalloproteinase-13 (MMP-13). Notably, by gene silencing or kinase inhibition, we proposed for the first time that ATM is a key up-stream regulator of TNF-α activated ERK/p38-NF-κB pathway. The existence of TNF-α secreted in autocrine or paracrine manner by components of tumor microenvironment highlights the significance of TNF-α in inflammation-associated tumor metastasis. Importantly, in vivo lung cancer metastasis test showed that ATM depletion actually reduce the number of metastatic nodules and cancer nests in lung tissues, verifying the critical role of ATM in metastasis. In conclusion, our findings demonstrate that ATM, which could be activated by lung cancer-associated TNF-α, up-regulate MMP-13 expression and thereby augment tumor metastasis. Therefore, ATM might be a promising target for prevention of inflammation-associated lung cancer metastasis. PMID:27556690

  18. Tumor necrosis factor-alpha-induced inhibition of phosphatidylcholine synthesis by human type II pneumocytes is partially mediated by prostaglandins.

    PubMed Central

    Arias-Díaz, J; Vara, E; García, C; Balibrea, J L

    1994-01-01

    TNF alpha seems to play an important role in the pathogenesis of adult respiratory distress syndrome. We studied the effect of TNF alpha on phospholipid synthesis by isolated type II pneumocytes and attempted to characterize the role of arachidonate metabolites and the influence of pentoxifylline on such an effect. Lung tissue obtained from both multiple organ donors (n = 14) and lung cancer patients (n = 11) was used for cell isolation. Surfactant synthesis was measured by the incorporation of D-[U-14C]glucose into phosphatidylcholine (PC). The basal PC synthesis was higher in the donor group than in the malignant group (3.44 +/- 0.19 vs 2.15 +/- 0.15 pmol/microgram protein x 120 min, P < 0.01), and, in the presence of 100 ng/ml of TNF alpha, the incorporation of labeled glucose into PC was reduced significantly in both donor (1.13 +/- 0.11 vs 3.44 +/- 0.19 pmol/microgram protein x 120 min, P < 0.01) and cancer (0.99 +/- 0.11 vs 2.15 +/- 0.15 pmol/microgram protein x 120 min, P < 0.01) groups. Indomethacin was able to completely block the cytokine-induced decrease in PC synthesis by pneumocytes from the malignant group and to attenuate the inhibitory effect of TNF alpha in those from donors, nordihydroguaiaretic acid having a similar effect. The TNF alpha effect can be blocked by pentoxifylline (100 micrograms/ml), a substance which can even succeed in reverting the basal secretory inhibition of cancer patients' pneumocytes to levels similar to those of the donor group. TNF alpha may contribute to the pathophysiology of adult respiratory distress syndrome by inhibiting the synthesis of surfactant. TNF alpha might be produced in lung tumors, resulting in chronic paracrine or systemic exposure of pneumocytes to low concentrations of the cytokine. The TNF alpha effect was not prevented completely by the blockage of the arachidonic acid metabolism, hence other mediators should also be implicated. PMID:8040266

  19. Tumor Necrosis Factor Alpha Blocker-Induced Erythrodermic Sarcoidosis in with Juvenile Rheumatoid Arthritis: A Case Report and Review of the Literature

    PubMed Central

    Park, Su-Kyung; Hwang, Pyung-Han; Yun, Seok-Kweon; Kim, Han-Uk

    2017-01-01

    The development of cutaneous sarcoidosis as a paradoxical adverse event of tumor necrosis factor alpha (TNF-α) blockers has been reported in the literature; however, an erythrodermic form of cutaneous sarcoidosis during anti-TNF-α therapy has not yet been reported. Herein, we report the first case of an erythrodermic form of cutaneous sarcoidosis during anti-TNF-α therapy and review previous studies of cutaneous sarcoidosis. A 6-year-old Korean girl who had been suffering from juvenile rheumatoid arthritis presented with generalized erythematous skin eruption involving more than about 90% of her body surface area. After 14 months of etanercept treatment, the new erythematous skin eruption had developed and progressed into generalized erythroderma. Exclusion of suspected co-medication had been performed based on medication history. She had no other systemic symptoms, and ophthalmologic and neurologic examinations were normal. Histopathologic findings of the skin lesion revealed diffuse non-caseating granulomatous infiltrates composed of epithelioid histiocytes with sparse lymphocytes involving the entire dermis. Periodic-acid-Schiff and acid-fast stains were negative, and acid-fast bacilli was not detected by polymerase chain reaction of the skin biopsy. Based on clinicopathologic findings, she was diagnosed with etanercept-induced sarcoidal granuloma. After discontinuation of the suspected agent, the lesions spontaneously disappeared. PMID:28223750

  20. N-acetylcysteine supplementation controls total antioxidant capacity, creatine kinase, lactate, and tumor necrotic factor-alpha against oxidative stress induced by graded exercise in sedentary men.

    PubMed

    Leelarungrayub, Donrawee; Khansuwan, Raphiphat; Pothongsunun, Prapas; Klaphajone, Jakkrit

    2011-01-01

    Aim of this study was to evaluate the effects of short-term (7 days) N-acetylcysteine (NAC) at 1,200 mg daily supplementation on muscle fatigue, maximal oxygen uptake (VO(2max)), total antioxidant capacity (TAC), lactate, creatine kinase (CK), and tumor necrotic factor-alpha (TNF-α). Twenty-nine sedentary men (13 controls; 16 in the supplement group) from a randomized control were included. At before and after supplementation, fatigue index (FI) was evaluated in the quadriceps muscle, and performed a graded exercise treadmill test to induce oxidative stress, and as a measure of VO(2max). Blood samples were taken before exercise and 20 minutes after it at before and after supplementation, to determine TAC, CK, lactate, and TNF-α levels. Results showed that FI and VO(2max) increased significantly in the supplement group. After exercise decreased the levels of TAC and increased lactate, CK, and TNF-α of both groups at before supplementation. After supplementation, lactate, CK, and TNF-α levels significantly increased and TAC decreased after exercise in the control group. Whereas the TAC and lactate levels did not change significantly, but CK and TNF-α increased significantly in the supplement group. Therefore, this results showed that NAC improved the muscle fatigue, VO(2max), maintained TAC, controlled lactate production, but had no influence on CK and TNF-α.

  1. Tumor Necrosis Factor Alpha Blocker-Induced Erythrodermic Sarcoidosis in with Juvenile Rheumatoid Arthritis: A Case Report and Review of the Literature.

    PubMed

    Park, Su-Kyung; Hwang, Pyung-Han; Yun, Seok-Kweon; Kim, Han-Uk; Park, Jin

    2017-02-01

    The development of cutaneous sarcoidosis as a paradoxical adverse event of tumor necrosis factor alpha (TNF-α) blockers has been reported in the literature; however, an erythrodermic form of cutaneous sarcoidosis during anti-TNF-α therapy has not yet been reported. Herein, we report the first case of an erythrodermic form of cutaneous sarcoidosis during anti-TNF-α therapy and review previous studies of cutaneous sarcoidosis. A 6-year-old Korean girl who had been suffering from juvenile rheumatoid arthritis presented with generalized erythematous skin eruption involving more than about 90% of her body surface area. After 14 months of etanercept treatment, the new erythematous skin eruption had developed and progressed into generalized erythroderma. Exclusion of suspected co-medication had been performed based on medication history. She had no other systemic symptoms, and ophthalmologic and neurologic examinations were normal. Histopathologic findings of the skin lesion revealed diffuse non-caseating granulomatous infiltrates composed of epithelioid histiocytes with sparse lymphocytes involving the entire dermis. Periodic-acid-Schiff and acid-fast stains were negative, and acid-fast bacilli was not detected by polymerase chain reaction of the skin biopsy. Based on clinicopathologic findings, she was diagnosed with etanercept-induced sarcoidal granuloma. After discontinuation of the suspected agent, the lesions spontaneously disappeared.

  2. Systemic administration of an anti-tumor necrosis factor-alpha monoclonal antibody protects against endotoxin-induced uveitis in rats

    PubMed Central

    Ge, Qingman; Wang, Shaocheng; Zheng, Yuezhong

    2016-01-01

    Objective: This study was to evaluate the effect of systemic injection of an anti-tumor necrosis factor alpha (TNF-α) monoclonal antibody (mAb) on endotoxin-induced uveitis (EIU). Materials and Methods: Fifty-six male Wistar rats (6–8 weeks old) were randomly divided into three groups: EIU, anti-TNF-α mAb + EIU, and control. EIU was induced by injecting Escherichia coli O55:B5 lipopolysaccharide (LPS) into the hind footpad of the rats (150 μg/rat). The anti-TNF-α mAb (1 μg/kg) was administrated 30 min before LPS injection through one-time intravenous injection. The onset time and peak time of EIU were recorded. The serum and aqueous humor (AH) TNF-α, interleukin (IL)-6, and IL-10 levels were measured by ELISA at 4, 24, and 72 h post-LPS injection. Clinical manifestations of EIU and eye histopathology were scored. Results: Compared with the EIU rats, anti-TNF-α mAb + EIU rats showed significantly delayed onset of uveitis (t = 7.41, P < 0.001), lower clinical scores and histopathological grades (t = 3.18/2.22, P < 0.001), reduced levels of TNF-α (F = 15.06/59.43, P < 0.001) and IL-6 (F = 99.63/14.92, P < 0.001), and increased levels of IL-10 (F = 24.94/8.99, P < 0.001) in the serum and AH. AH TNF-α, serum IL-6, and AH IL-6 levels are positively correlated, whereas serum IL-10 levels were negatively correlated with EIU activity. Conclusion: Antagonizing TNF-α by system injection of the anti-TNF-α mAb protects against EIU in rats. Blocking TNF-α signaling could be a useful strategy for managing uveitis. PMID:28112125

  3. Local expression of tumor necrosis factor alpha and interleukin-2 correlates with protection against corneal scarring after ocular challenge of vaccinated mice with herpes simplex virus type 1.

    PubMed Central

    Ghiasi, H; Wechsler, S L; Kaiwar, R; Nesburn, A B; Hofman, F M

    1995-01-01

    To correlate specific local immune responses with protection from corneal scarring, we examined immune cell infiltrates in the cornea after ocular challenge of vaccinated mice with herpes simplex virus type 1 (HSV-1). This is the first report to examine corneal infiltrates following ocular challenge of a vaccinated mouse rather than following infection of a naive mouse. Mice were vaccinated systemically with vaccines that following ocular challenge with HSV-1 resulted in (i) complete protection against corneal disease (KOS, an avirulent strain of HSV-1); (ii) partial protection, resulting in moderate corneal disease (baculovirus-expressed HSV-1 glycoprotein E [gE]); and (iii) no protection, resulting in severe corneal disease (mock vaccine). Infiltration into the cornea of CD4+ T cells, CD8+ T cells, macrophages, and cells containing various lymphokines was monitored on days 0, 1, 3, 7, and 10 postchallenge by immunocytochemistry of corneal sections. Prior to ocular challenge, no eye disease or corneal infiltrates were detected in any mice. KOS-vaccinated mice developed high HSV-1 neutralizing antibody titers (> 1:640) in serum. After ocular challenge, they were completely protected against death, developed no corneal disease, and had no detectable virus in their tear films at any time examined. In response to the ocular challenge, these mice developed high local levels of infiltrating CD4+ T cells and cells containing interleukin-2 (IL-2), IL-4, IL-6, or tumor necrosis factor alpha (TNF-alpha). In contrast, only low levels of infiltrating CD8+ T cells were found, and gamma interferon (IFN-gamma)-containing cells were not present until day 10. gE-vaccinated mice developed neutralizing antibody titers in serum almost as high as those of the KOS-vaccinated mice (> 1:320). After ocular challenge, they were also completely protected against death. However, the gE-vaccinated mice developed low levels of corneal disease and virus was detected in one-third of their eyes

  4. Tumor Necrosis Factor Alpha-Induced Recruitment of Inflammatory Mononuclear Cells Leads to Inflammation and Altered Brain Development in Murine Cytomegalovirus-Infected Newborn Mice.

    PubMed

    Seleme, Maria C; Kosmac, Kate; Jonjic, Stipan; Britt, William J

    2017-04-15

    Congenital human cytomegalovirus (HCMV) infection is a significant cause of abnormal neurodevelopment and long-term neurological sequelae in infants and children. Resident cell populations of the developing brain have been suggested to be more susceptible to virus-induced cytopathology, a pathway thought to contribute to the clinical outcomes following intrauterine HCMV infection. However, recent findings in a newborn mouse model of the infection in the developing brain have indicated that elevated levels of proinflammatory mediators leading to mononuclear cell activation and recruitment could underlie the abnormal neurodevelopment. In this study, we demonstrate that treatment with tumor necrosis factor alpha (TNF-α)-neutralizing antibodies decreased the frequency of CD45(+) Ly6C(hi) CD11b(+) CCR2(+) activated myeloid mononuclear cells (MMCs) and the levels of proinflammatory cytokines in the blood and the brains of murine CMV-infected mice. This treatment also normalized neurodevelopment in infected mice without significantly impacting the level of virus replication. These results indicate that TNF-α is a major component of the inflammatory response associated with altered neurodevelopment that follows murine CMV infection of the developing brain and that a subset of peripheral blood myeloid mononuclear cells represent a key effector cell population in this model of virus-induced inflammatory disease of the developing brain.IMPORTANCE Congenital human cytomegalovirus (HCMV) infection is the most common viral infection of the developing human fetus and can result in neurodevelopmental sequelae. Mechanisms of disease leading to neurodevelopmental deficits in infected infants remain undefined, but postulated pathways include loss of neuronal progenitor cells, damage to the developing vascular system of the brain, and altered cellular positioning. Direct virus-mediated cytopathic effects cannot explain the phenotypes of brain damage in most infected infants. Using a

  5. Protein kinase C-{beta}, fibronectin, {alpha}{sub 5}{beta}{sub 1}-integrin and tumor necrosis factor-{alpha} are required for phorbol diester-induced apoptosis in human myeloid leukemia cells in human myeloid leukemia cells.

    SciTech Connect

    Laouar, A.; Glesne, D.; Huberman, E.

    2001-12-01

    The human myeloid HL-60 cell line and its cell variant HL-525 were used to study signaling events leading to apoptosis induction by phorbol 12-myristate 13-acetate (PMA), an activator of protein kinase C (PKC) enzymes. Unlike parental cells, HL-525 cells are PKC-{beta} deficient and resistant to PMA-induced apoptosis. These cells regain susceptibility to apoptosis induction after transfection with a PKC-{beta} expression vector. By using this vector and specific neutralizing monoclonal antibodies (mAbs), it was established that PMA-induced apoptosis also called for an interaction between cell-surface {alpha}{sub 5}{beta}{sub 1}-integrin and its deposited ligand fibronectin (FN), which is downstream of PKC-{beta} activation. Experiments with mAbs, the PKC-{beta} vector, and exogenous FN revealed that the next step entailed an interaction between secreted tumor necrosis factor-{alpha} and its type I receptor. By using a sphingomyelinase inhibitor, it was concluded that the subsequent step involved ceramide production. Moreover, a permeable ceramide was effective in inducing apoptosis in both HL-60 and HL-525 cells, and this induction was caspase-1 and/or -4 dependent because an inhibitor of these caspases abrogated the induced apoptosis. Based on these and related differentiation studies, we conclude that the above signaling events, the early ones in particular, are shared with PMA-induced macrophage differentiation in the HL-60 cells. It is likely that once these cells acquire their macrophage phenotype and perform their tasks, they become superfluous and are eliminated from the body by a self-triggered apoptotic process that involves our proposed signaling scheme.

  6. Ketamine inhibits tumor necrosis factor-{alpha} and interleukin-6 gene expressions in lipopolysaccharide-stimulated macrophages through suppression of toll-like receptor 4-mediated c-Jun N-terminal kinase phosphorylation and activator protein-1 activation

    SciTech Connect

    Wu, G.-J.; Chen, T.-L.; Ueng, Y.-F.; Chen, R.-M.

    2008-04-01

    Our previous study showed that ketamine, an intravenous anesthetic agent, has anti-inflammatory effects. In this study, we further evaluated the effects of ketamine on the regulation of tumor necrosis factor-{alpha} (TNF-{alpha}) and interlukin-6 (IL-6) gene expressions and its possible signal-transducing mechanisms in lipopolysaccharide (LPS)-activated macrophages. Exposure of macrophages to 1, 10, and 100 {mu}M ketamine, 100 ng/ml LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. A concentration of 1000 {mu}M of ketamine alone or in combined treatment with LPS caused significant cell death. Administration of LPS increased cellular TNF-{alpha} and IL-6 protein levels in concentration- and time-dependent manners. Meanwhile, treatment with ketamine concentration- and time-dependently alleviated the enhanced effects. LPS induced TNF-{alpha} and IL-6 mRNA syntheses. Administration of ketamine at a therapeutic concentration (100 {mu}M) significantly inhibited LPS-induced TNF-{alpha} and IL-6 mRNA expressions. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA into macrophages decreased cellular TLR4 levels. Co-treatment of macrophages with ketamine and TLR4 siRNA decreased the LPS-induced TNF-{alpha} and IL-6 productions more than alone administration of TLR4 siRNA. LPS stimulated phosphorylation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos from the cytoplasm to nuclei. However, administration of ketamine significantly decreased LPS-induced activation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos. LPS increased the binding of nuclear extracts to activator protein-1 consensus DNA oligonucleotides. Administration of ketamine significantly ameliorated LPS-induced DNA binding activity of activator protein-1. Therefore, a clinically relevant concentration of ketamine can inhibit TNF-{alpha} and IL-6 gene expressions in LPS-activated macrophages. The suppressive mechanisms

  7. Ketamine inhibits tumor necrosis factor-alpha and interleukin-6 gene expressions in lipopolysaccharide-stimulated macrophages through suppression of toll-like receptor 4-mediated c-Jun N-terminal kinase phosphorylation and activator protein-1 activation.

    PubMed

    Wu, Gone-Jhe; Chen, Ta-Liang; Ueng, Yune-Fang; Chen, Ruei-Ming

    2008-04-01

    Our previous study showed that ketamine, an intravenous anesthetic agent, has anti-inflammatory effects. In this study, we further evaluated the effects of ketamine on the regulation of tumor necrosis factor-alpha (TNF-alpha) and interlukin-6 (IL-6) gene expressions and its possible signal-transducing mechanisms in lipopolysaccharide (LPS)-activated macrophages. Exposure of macrophages to 1, 10, and 100 microM ketamine, 100 ng/ml LPS, or a combination of ketamine and LPS for 1, 6, and 24 h was not cytotoxic to macrophages. A concentration of 1000 microM of ketamine alone or in combined treatment with LPS caused significant cell death. Administration of LPS increased cellular TNF-alpha and IL-6 protein levels in concentration- and time-dependent manners. Meanwhile, treatment with ketamine concentration- and time-dependently alleviated the enhanced effects. LPS induced TNF-alpha and IL-6 mRNA syntheses. Administration of ketamine at a therapeutic concentration (100 microM) significantly inhibited LPS-induced TNF-alpha and IL-6 mRNA expressions. Application of toll-like receptor 4 (TLR4) small interfering (si)RNA into macrophages decreased cellular TLR4 levels. Co-treatment of macrophages with ketamine and TLR4 siRNA decreased the LPS-induced TNF-alpha and IL-6 productions more than alone administration of TLR4 siRNA. LPS stimulated phosphorylation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos from the cytoplasm to nuclei. However, administration of ketamine significantly decreased LPS-induced activation of c-Jun N-terminal kinase and translocation of c-Jun and c-Fos. LPS increased the binding of nuclear extracts to activator protein-1 consensus DNA oligonucleotides. Administration of ketamine significantly ameliorated LPS-induced DNA binding activity of activator protein-1. Therefore, a clinically relevant concentration of ketamine can inhibit TNF-alpha and IL-6 gene expressions in LPS-activated macrophages. The suppressive mechanisms occur through

  8. Effects of carbocisteine on sialyl-Lewis x expression in an airway carcinoma cell line stimulated with tumor necrosis factor-alpha.

    PubMed

    Ishibashi, Yuji; Imai, Shigeru; Inouye, Yoshio; Okano, Teruo; Taniguchi, Akiyoshi

    2006-01-20

    Carbocisteine is a mucoregulatory drug normalizing sialic acid and fucose contents in mucins through the regulation of glycosyltransferase activities. Tumor necrosis factor (TNF)-alpha-induced overexpression of sialyl-Lewis x epitopes, containing sialic acid and fucose, in mucins were previously reported to be regulated by glycosyltransferase mRNAs expression through phosphatidyl inositol-specific phospholipase C (PI-PLC) signaling pathways [Ishibashi, Y., Inouye, Y., Okano, T., Taniguchi, A., 2005. Regulation of sialyl-Lewis x epitope expression by TNF-alpha and EGF in an airway carcinoma cell line. Glycoconj. J. 22, 53-62]. To investigate the mechanism behind the mucoregulatory action of carbocisteine, the present study evaluated the effects of carbocisteine on TNF-alpha-induced overexpression of sialyl-Lewis x epitopes in NCI-H292 cells. 100 mug/ml of carbocisteine was able to inhibit the TNF-alpha-induced expression of hST3GallV mRNA, FUT3 mRNA, C2/4GnT mRNA and sialyl-Lewis x epitopes as well as the TNF-alpha-induced activity of PI-PLC in NCI-H292 cells. These findings suggest that carbocisteine may normalize the sialyl-Lewis x epitopes expression in mucins through the inhibition of cellular PI-PLC activity in vivo.

  9. The role of glucagon-like peptide-2 on apoptosis, cell proliferation, and oxidant-antioxidant system at a mouse model of intestinal injury induced by tumor necrosis factor-alpha/actinomycin D.

    PubMed

    Arda-Pirincci, Pelin; Bolkent, Sehnaz

    2011-04-01

    Tumor necrosis factor-alpha (TNF-α) is a multifunctional cytokine, which has the ability to produce cytotoxicity via induction of cell death and cell cycle arrest. Blocking the synthesis of protective proteins through a transcriptional inhibitor such as actinomycin D (Act D) sensitizes many cell types to TNF-α toxicity. Teduglutide, h[Gly(2)]GLP-2, is a protease-resistant synthetic analog of glucagon-like peptide-2 (GLP-2) which is an intestinotrophic peptide. In this study, we evaluated this potential of GLP-2 on apoptosis, cell proliferation, and oxidant-antioxidant system on a mouse model of intestinal injury induced by TNF-α/Act D. The intestinal injury was induced by intraperitoneal administration of 15 μg/kg TNF-α and 800 μg/kg Act D per mouse. Animals were injected subcutaneously 200 μg/kg h[Gly(2)]GLP-2 every 12 h for 10 consecutive days prior to the administration of TNF-α and Act D. The model of intestinal injury induced by TNF-α/Act D, which is the new animal model for the intestinal disorders, was characterized by the degeneration of intestinal mucosa, an increase in apoptotic index, expression of active caspase-3, lipid peroxidation and glutathione (GSH) levels, glutathione peroxidase (GPx) and superoxide dismutase (SOD) activities; a decrease in cell proliferation and catalase (CAT) activity. h[Gly(2)]GLP-2 pretreatment prevented the TNF-α/Act D-induced oxidative injury by a significant reduction in the intestinal injury, apoptotic index, expression of active caspase-3, lipid peroxidation and GSH levels, GPx and SOD activities; a markedly increase in cell proliferation, and CAT activity. These results demonstrate that GLP-2 has a protective, antiapoptotic, proliferative, and antioxidant effects against to TNF-α/Act D-induced intestinal injury. It is suggested that GLP-2 may potentially be useful as a therapeutic agent in TNF-α-mediated intestinal disorders.

  10. Interferon-gamma and tumor necrosis factor-alpha regulate amyloid-beta plaque deposition and beta-secretase expression in Swedish mutant APP transgenic mice.

    PubMed

    Yamamoto, Masaru; Kiyota, Tomomi; Horiba, Masahide; Buescher, James L; Walsh, Shannon M; Gendelman, Howard E; Ikezu, Tsuneya

    2007-02-01

    Reactive astrocytes and microglia in Alzheimer's disease surround amyloid plaques and secrete proinflammatory cytokines that affect neuronal function. Relationship between cytokine signaling and amyloid-beta peptide (Abeta) accumulation is poorly understood. Thus, we generated a novel Swedish beta-amyloid precursor protein mutant (APP) transgenic mouse in which the interferon (IFN)-gamma receptor type I was knocked out (APP/GRKO). IFN-gamma signaling loss in the APP/GRKO mice reduced gliosis and amyloid plaques at 14 months of age. Aggregated Abeta induced IFN-gamma production from co-culture of astrocytes and microglia, and IFN-gamma elicited tumor necrosis factor (TNF)-alpha secretion in wild type (WT) but not GRKO microglia co-cultured with astrocytes. Both IFN-gamma and TNF-alpha enhanced Abeta production from APP-expressing astrocytes and cortical neurons. TNF-alpha directly stimulated beta-site APP-cleaving enzyme (BACE1) expression and enhanced beta-processing of APP in astrocytes. The numbers of reactive astrocytes expressing BACE1 were increased in APP compared with APP/GRKO mice in both cortex and hippocampus. IFN-gamma and TNF-alpha activation of WT microglia suppressed Abeta degradation, whereas GRKO microglia had no changes. These results support the idea that glial IFN-gamma and TNF-alpha enhance Abeta deposition through BACE1 expression and suppression of Abeta clearance. Taken together, these observations suggest that proinflammatory cytokines are directly linked to Alzheimer's disease pathogenesis.

  11. A role for protein phosphatase 2A in regulating p38 mitogen activated protein kinase activation and tumor necrosis factor-alpha expression during influenza virus infection.

    PubMed

    Law, Anna H Y; Tam, Alex H M; Lee, Davy C W; Lau, Allan S Y

    2013-04-02

    Influenza viruses of avian origin continue to pose pandemic threats to human health. Some of the H5N1 and H9N2 virus subtypes induce markedly elevated cytokine levels when compared with the seasonal H1N1 virus. We previously showed that H5N1/97 hyperinduces tumor necrosis factor (TNF)-alpha through p38 mitogen activated protein kinase (MAPK). However, the detailed mechanisms of p38MAPK activation and TNF-alpha hyperinduction following influenza virus infections are not known. Negative feedback regulations of cytokine expression play important roles in avoiding overwhelming production of proinflammatory cytokines. Here we hypothesize that protein phosphatases are involved in the regulation of cytokine expressions during influenza virus infection. We investigated the roles of protein phosphatases including MAPK phosphatase-1 (MKP-1) and protein phosphatase type 2A (PP2A) in modulating p38MAPK activation and downstream TNF-alpha expressions in primary human monocyte-derived macrophages (PBMac) infected with H9N2/G1 or H1N1 influenza virus. We demonstrate that H9N2/G1 virus activated p38MAPK and hyperinduced TNF-alpha production in PBMac when compared with H1N1 virus. H9N2/G1 induced PP2A activity in PBMac and, with the treatment of a PP2A inhibitor, p38MAPK phosphorylation and TNF-alpha production were further increased in the virus-infected macrophages. However, H9N2/G1 did not induce the expression of PP2A indicating that the activation of PP2A is not mediated by p38MAPK in virus-infected PBMac. On the other hand, PP2A may not be the targets of H9N2/G1 in the upstream of p38MAPK signaling pathways since H1N1 also induced PP2A activation in primary macrophages. Our results may provide new insights into the control of cytokine dysregulation.

  12. Expression of a tumor necrosis factor alpha transgene in murine pancreatic beta cells results in severe and permanent insulitis without evolution towards diabetes

    PubMed Central

    1992-01-01

    Mice bearing a tumor necrosis factor (TNF) alpha transgene controlled by an insulin promoter developed an increasingly severe lymphocytic insulitis, apparently resulting from the induction of endothelial changes with features similar to those observed in other places of intense lymphocytic traffic. This was accompanied by dissociation of the endocrine tissue (without marked decrease in its total mass), islet fibrosis, and the development of intraislet ductules containing, by places, beta cells in their walls, suggesting a regenerative capacity. Islet disorganization and fibrosis did not result from lymphocytic infiltration, since they were also observed in SCID mice bearing the transgene. Diabetes never developed, even though a number of potentially inducing conditions were used, including the prolonged perfusion of interferon gamma and the permanent expression of a nontolerogenic viral protein on beta cells (obtained by using mice bearing two transgenes). It is concluded that (a) a slow process of TNF release in pancreatic islets induces insulitis, and may be instrumental in the insulitis resulting from local cell-mediated immune reactions, but (b) that insulitis per se is not diabetogenic, lymphocyte stimulation by cells other than beta cells being necessary to trigger extensive beta cell damage. This provides an explanation for the discrepancy between the occurrence of insulitis and that of clinical disease in autoimmune diabetes. PMID:1460428

  13. Alpha-Tocopherol Alters Transcription Activities that Modulates Tumor Necrosis Factor Alpha (TNF-α) Induced Inflammatory Response in Bovine Cells.

    PubMed

    Li, Cong-Jun; Li, Robert W; Kahl, Stanislaw; Elsasser, Theodore H

    2012-01-01

    To further investigate the potential role of α-tocopherol in maintaining immuno-homeostasis in bovine cells (Madin-Darby bovine kidney epithelial cell line), we undertook in vitro experiments using recombinant TNF-α as an immuno-stimulant to simulate inflammation response in cells with or without α-tocopherol pre-treatment. Using microarray global-profiling and IPA (Ingenuity Pathways Analysis, Ingenuity(®) Systems, http://www.ingenuity.com) data analysis on TNF-α-induced gene perturbation in those cells, we focused on determining whether α-tocopherol treatment of normal bovine cells in a standard cell culture condition can modify cell's immune response induced by TNF-α challenge. When three datasets were filtered and compared using IPA, there were a total of 1750 genes in all three datasets for comparison, 97 genes were common in all three sets; 615 genes were common in at least two datasets; there were 261 genes unique in TNF-α challenge, 399 genes were unique in α-tocopherol treatment, and 378 genes were unique in the α-tocopherol plus TNF-α treatment. TNF-α challenge induced significant change in gene expression. Many of those genes induced by TNF-α are related to the cells immune and inflammatory responses. The results of IPA data analysis showed that α-tocopherol-pretreatment of cells modulated cell's response to TNF-α challenge. In most of the canonical pathways, α-tocopherol pretreatment showed the antagonistic effect against the TNF-α-induced pro-inflammatory responses. We concluded that α-tocopherol pre-treatment has a significant antagonistic effect that modulates the cell's response to the TNF-α challenge by altering the gene expression activities of some important signaling molecules.

  14. Adjusting effects of baicalin for nuclear factor-kappaB and tumor necrosis factor-alpha on rats with caerulein-induced acute pancreatitis.

    PubMed

    Xue, Dongbo; Zhang, Weihui; Zhang, Yingmei; Wang, Haiyang; Zheng, Biao; Shi, Xingye

    2006-01-01

    Forty Wistar rats were divided into 5 groups, including the control group, the acute pancreatitis group (AP group, induced by intraperitoneal injections of caerulein), and the AP group treated with baicalin, the AP group treated with LPS, and the AP group treated with LPS and baicalin. Pathological damage of pancreatic tissue was scored with hematoxylin and eosin (HE) staining. The mRNA expression of TNF-alpha was measured with semiquantitative RT-PCR, and activation of NF-kappaB was detected with flow cytometry assay. It was shown in the results that the expression of TNF-alpha mRNA, activation of NF-kappaB, and pathological score of AP group were all obviously higher than those of control group (P < .01). In AP group treated with LPS, further rise of these values were observed (P < .01). In the AP group treated with baicalin, activation of NF-kappaB decreased (P < .05), and expression of TNF-alpha mRNA also obviously decreased (P < .01), while pancreatic pathological damage was alleviated at the same time (P < .01); similar results were observed in AP group treated with LPS and baicalin (P < .01), which indicated that baicalin might be applied to inhibit NF-kappaB activating and TNF-alpha expressing so as to treat AP.

  15. Tumor necrosis factor-alpha upregulates 11beta-hydroxysteroid dehydrogenase type 1 expression by CCAAT/enhancer binding protein-beta in HepG2 cells.

    PubMed

    Ignatova, Irena D; Kostadinova, Radina M; Goldring, Christopher E; Nawrocki, Andrea R; Frey, Felix J; Frey, Brigitte M

    2009-02-01

    The enzyme 11beta-hydroxysteroid dehydrogenase type 1 (11beta-HSD1) catalyzes the conversion of inactive to active glucocorticoids. 11beta-HSD1 plays a crucial role in the pathogenesis of obesity and controls glucocorticoid actions in inflammation. Several studies have demonstrated that TNF-alpha increases 11beta-HSD1 mRNA and activity in various cell models. Here, we demonstrate that mRNA and activity of 11beta-HSD1 is increased in liver tissue from transgenic mice overexpressing TNF-alpha, indicating that this effect also occurs in vivo. To dissect the molecular mechanism of this increase, we investigated basal and TNF-alpha-induced transcription of the 11beta-HSD1 gene (HSD11B1) in HepG2 cells. We found that TNF-alpha acts via p38 MAPK pathway. Transient transfections with variable lengths of human HSD11B1 promoter revealed highest activity with or without TNF-alpha in the proximal promoter region (-180 to +74). Cotransfection with human CCAAT/enhancer binding protein-alpha (C/EBPalpha) and C/EBPbeta-LAP expression vectors activated the HSD11B1 promoter with the strongest effect within the same region. Gel shift and RNA interference assays revealed the involvement of mainly C/EBPalpha, but also C/EBPbeta, in basal and only of C/EBPbeta in the TNF-alpha-induced HSD11B1 expression. Chromatin immunoprecipitation assay confirmed in vivo the increased abundance of C/EBPbeta on the proximal HSD11B1 promoter upon TNF-alpha treatment. In conclusion, C/EBPalpha and C/EBPbeta control basal transcription, and TNF-alpha upregulates 11beta-HSD1, most likely by p38 MAPK-mediated increased binding of C/EBPbeta to the human HSD11B1 promoter. To our knowledge, this is the first study showing involvement of p38 MAPK in the TNF-alpha-mediated 11beta-HSD1 regulation, and that TNF-alpha stimulates enzyme activity in vivo.

  16. Microglia-derived proinflammatory cytokines tumor necrosis factor-alpha and interleukin-1beta induce Purkinje neuronal apoptosis via their receptors in hypoxic neonatal rat brain.

    PubMed

    Kaur, Charanjit; Sivakumar, Viswanathan; Zou, Zhirong; Ling, Eng-Ang

    2014-01-01

    The developing cerebellum is extremely vulnerable to hypoxia which can damage the Purkinje neurons. We hypothesized that this might be mediated by tumor necrosis factor-α (TNF-α) and interleukin-1β (IL-1β) derived from activated microglia as in other brain areas. One-day-old rats were subjected to hypoxia following, which the expression changes of various proteins in the cerebellum including hypoxia inducible factor-1α, TNF-α, IL-1β, TNF-R1 and IL-1R1 were analyzed. Following hypoxic exposure, TNF-α and IL-1β immunoexpression in microglia was enhanced coupled by that of TNF-R1 and IL-1R1 in the Purkinje neurons. Along with this, hypoxic microglia in vitro showed enhanced release of TNF-α and IL-1β whose receptor expression was concomitantly increased in the Purkinje neurons. In addition, nitric oxide (NO) level was significantly increased in the cerebellum and cultured microglia subjected to hypoxic exposure. Moreover, cultured Purkinje neurons treated with conditioned medium derived from hypoxic microglia underwent apoptosis but the incidence was significantly reduced when the cells were treated with the same medium that was neutralized with TNF-α/IL-1β antibody. We conclude that hypoxic microglia in the neonatal cerebellum produce increased amounts of NO, TNF-α and IL-1β which when acting via their respective receptors could induce Purkinje neuron death.

  17. Effects of the −791(C→T) mutation in the promoter for tumor necrosis factor alpha on gene expression and resistance of Large White pigs to enterotoxigenic Escherichia coli F18

    PubMed Central

    Liu, Ying; Dai, Chaohui; Sun, Li; Zhu, Guoqiang; Wu, Shenglong; Bao, Wenbin

    2016-01-01

    Tumor necrosis factor alpha (TNF-α) plays an important role in the immune system. In this study, TNF-α expression was analyzed in 11 tissues of 8 piglets resistant to enterotoxigenic Escherichia coli (ETEC) F18 and 8 ETEC F18-susceptible piglets from the Large White breed. The expression levels of TNF-α were high in immune organs (spleen, lung, thymus, and lymph nodes). The levels were higher in ETEC F18-resistant piglets than in ETEC F18-susceptible piglets, with significant differences in spleen, kidney, thymus, lymph node, and duodenum (P < 0.05). The mutation TNF-α −791(C→T) and 3 genotypes (CC, CT, and TT) were identified. The TNF-α expression levels in the spleen, kidney, lymph nodes, and duodenum were significantly higher in the TT pigs than in the CC pigs (P < 0.05). Thus, TNF-α −791(C→T) has significant effects on mRNA expression and may regulate ETEC F18 resistance of weaning piglets. Therefore, the −791(C→T) mutation of the TNF-α gene could be considered an important potential genetic marker of ETEC F18 resistance. PMID:27408333

  18. Constitutive stabilization of hypoxia-inducible factor alpha selectively promotes the self-renewal of mesenchymal progenitors and maintains mesenchymal stromal cells in an undifferentiated state.

    PubMed

    Park, In-Ho; Kim, Kwang-Ho; Choi, Hyun-Kyung; Shim, Jae-Seung; Whang, Soo-Young; Hahn, Sang June; Kwon, Oh-Joo; Oh, Il-Hoan

    2013-09-27

    With the increasing use of culture-expanded mesenchymal stromal cells (MSCs) for cell therapies, factors that regulate the cellular characteristics of MSCs have been of major interest. Oxygen concentration has been shown to influence the functions of MSCs, as well as other normal and malignant stem cells. However, the underlying mechanisms of hypoxic responses and the precise role of hypoxia-inducible factor-1α (Hif-1α), the master regulatory protein of hypoxia, in MSCs remain unclear, due to the limited span of Hif-1α stabilization and the complex network of hypoxic responses. In this study, to further define the significance of Hif-1α in MSC function during their self-renewal and terminal differentiation, we established adult bone marrow (BM)-derived MSCs that are able to sustain high level expression of ubiquitin-resistant Hif-1α during such long-term biological processes. Using this model, we show that the stabilization of Hif-1α proteins exerts a selective influence on colony-forming mesenchymal progenitors promoting their self-renewal and proliferation, without affecting the proliferation of the MSC mass population. Moreover, Hif-1α stabilization in MSCs led to the induction of pluripotent genes (oct-4 and klf-4) and the inhibition of their terminal differentiation into osteogenic and adipogenic lineages. These results provide insights into the previously unrecognized roles of Hif-1α proteins in maintaining the primitive state of primary MSCs and on the cellular heterogeneities in hypoxic responses among MSC populations.

  19. Anti-inflammatory effects of a new tumour necrosis factor-alpha (TNF-α) inhibitor (CNI-1493) in collagen-induced arthritis (CIA) in rats

    PubMed Central

    ÅKerlund, K; Erlandsson Harris, H; Tracey, K J; Wang, H; Fehniger, T; Klareskog, L; Andersson, J; Andersson, U

    1999-01-01

    A recently developed compound, a multivalent guanylhydrazone (CNI-1493) that inhibits TNF-α production by suppressing TNF-α translational efficiency, was administered in an experimental model of collagen type II-induced arthritis in DA rats. CNI-1493 was injected daily intraperitoneally either before the onset of arthritis or after the establishment of clinical disease. Prophylactic treatment with CNI-1493 significantly prevented or delayed the onset and suppressed the severity of arthritis in a dose-dependent manner. Therapeutic intervention with CNI-1493 in established joint disease also resulted in a significant reduction of clinical signs of arthritis in treated animals. No severe side-effects were noted when animals were treated with daily CNI-1493 doses up to 5 mg/kg. An immunohistochemical study was performed which demonstrated that CNI-1493 led to a reduced expression of TNF-α at the site of disease activity. Thus, CNI-1493 with documented inhibitory effects on TNF-α synthesis, has proven successful in ameliorating the course of arthritis in CIA. We believe that the use of a compound such as CNI-1493 with a defined mode of action provides a useful tool for dissecting and understanding important pathogenic mechanisms operating in the development of chronic arthritis. PMID:9933418

  20. Tumor necrosis factor-alpha expression in peripheral blood mononuclear cells correlates with early childhood social interaction in autism spectrum disorder.

    PubMed

    Makinodan, Manabu; Iwata, Keiko; Ikawa, Daisuke; Yamashita, Yasunori; Yamamuro, Kazuhiko; Toritsuka, Michihiro; Kimoto, Sohei; Okumura, Kazuki; Yamauchi, Takahira; Yoshino, Hiroki; Tsujii, Masatsugu; Sugiyama, Toshiro; Tsuchiya, Kenji; Mori, Norio; Matsuzaki, Hideo; Kishimoto, Toshifumi

    2017-03-01

    Autism spectrum disorder is a neurodevelopmental disorder characterized by impaired social interaction, poor communication skills, and repetitive/restrictive behaviors. Elevated blood levels of pro-inflammatory cytokines have been reported in subjects with autism spectrum disorder. On the other hand, early childhood adverse experience also increases blood levels of these cytokines. Since social experience of children with autism spectrum disorder is generally unlike to typically developing children, we hypothesized that social interaction during childhood contribute to pro-inflammatory cytokine expression in subjects with autism spectrum disorder. We compared revised Autism Diagnostic Interview scores and expression levels of pro-inflammatory cytokines in peripheral blood mononuclear cells of subjects with autism spectrum disorder (n = 30). The score of domain A on the revised Autism Diagnostic Interview, indicating social interaction impairment in early childhood, was negatively correlated with tumor necrosis factor-α mRNA expression level in peripheral blood mononuclear cells but not interleukin-1β or -6. Consistently, tumor necrosis factor-α mRNA expression was markedly low in subjects with autism spectrum disorder compared to typically developing children who presumably experienced the regular levels of social interaction. These findings suggest that the low blood levels of tumor necrosis factor-α mRNA in subjects with autism spectrum disorder might be due to impaired social interaction in early childhood.

  1. TERATOGENIC EFFECTS OF RETINOIC ACID ARE MODULATED IN MICE LACKING EXPRESSION OF EPIDERMAL GROWTH FACTOR AND TRANSFORMING GROWTH FACTOR-ALPHA

    EPA Science Inventory

    Background: EGF and TGF regulate cell proliferation and differentiation in the embryo. The induction of cleft palate (CP) by all trans retinoic acid (RA) was associated with altered expression of TGF, EGF receptor and binding of EGF. The present study uses knockout (KO) mice to e...

  2. Effects of tumor necrosis factor-alpha central administration on hippocampal damage in rat induced by amyloid beta-peptide (25-35).

    PubMed

    Stepanichev, Mikhail Yu; Zdobnova, Irina M; Yakovlev, Alexander A; Onufriev, Mikhail V; Lazareva, Natalia A; Zarubenko, Irina I; Gulyaeva, Natalia V

    2003-01-01

    Male Wistar rats received unilateral intrahippocampal injections of 3 nmol (3.18 microg) aggregated Abeta(25-35), intracerebroventricular bilateral injections of 0.5 microg human recombinant TNFalpha or both (Abeta(25-35) + TNFalpha-treated animals). Seven days after the surgery brain sections were stained with cresyl violet (Nissl), for fragmented DNA (TUNEL), glial fibrillar acidic protein (GFAP) and isolectin B4-reactive microglia. In addition, caspase-3 activity in brain regions was measured fluorometrically. The morphology of the hippocampus after the injection of Abeta(25-35) or both Abeta(25-35) and TNFalpha (but not TNFalpha alone) showed cell loss in the CA1 pyramidal cell layer. The extension of neuronal degeneration measured in the CA1 field was significantly larger in Abeta(25-35)-treated groups compared to the contralateral hemisphere of both vehicle-treated controls and animals injected with TNFalpha alone. TNFalpha augmented the Abeta(25-35)-induced damage, significantly increasing the extension of degenerating area. Administration of Abeta(25-35) caused reactive gliosis in the ipsilateral hemisphere as demonstrated by upregulation of GFAP expression and the presence of hypertrophic astrocytes in the hippocampus. This effect was much more prominent in the hippocampi of rats treated with Abeta(25-35) + TNFalpha but absent after administration of TNFalpha alone. In both Abeta(25-35)-treated groups, the damaged area of the hippocampal CA1 field and lateral band of dentate gyrus displayed many darkly stained round isolectin B4-positive phagocyte-like microglial cells. Sparse TUNEL-positive nuclei were found in the hippocampi of rats treated with Abeta(25-35) alone or together with TNFalpha, but not in the control brain sections or in brain sections of TNFalpha-injected animals. The activity of caspase-3 increased significantly in the ipsilateral hippocampus after the injection of Abeta(25-35). Surprisingly, administration of TNFalpha into the cerebral

  3. Neonatal exposure to estradiol-17β modulates tumour necrosis factor alpha and cyclooxygenase-2 expression in brain and also in ovaries of adult female rats.

    PubMed

    Shridharan, Radhika Nagamangalam; Krishnagiri, Harshini; Govindaraj, Vijayakumar; Sarangi, SitiKantha; Rao, Addicam Jagannadha

    2016-02-01

    The sexually dimorphic organization in perinatal rat brain is influenced by steroid hormones. Exposure to high levels of estrogen or endocrine-disrupting compounds during perinatal period may perturb this process, resulting in compromised reproductive physiology and behavior as observed in adult In our recent observation neonatal exposure of the female rats to estradiol-17β resulted in down-regulation of TNF-α, up-regulation of COX-2 and increase in SDN-POA size in pre-optic area in the adulthood. It is known that the control of reproductive performance in female involves a complex interplay of the hypothalamus, pituitary, and ovary. The present study was undertaken to understand the possible molecular mechanism involved in changes observed in the ovarian morphology and expression of selected genes in the ovary. Administration of estradiol-17β (100 μg) on day 2 and 3 after birth revealed up-regulation of ER-α, ER-β, COX-2 and down-regulation of TNF-α expression. Also the decrease in the ovarian weight, altered ovarian morphology and changes in the 2D protein profiles were also seen. This is apparently the first report documenting that neonatal estradiol exposure modulates TNF-α and COX-2 expression in the ovary as seen during adult stage. Our results permit us to suggest that cues originating from the modified brain structure due to neonatal exposure of estradiol-17β remodel the ovary at the molecular level in such a way that there is a disharmony in the reproductive function during adulthood and these changes are perennial and can lead to infertility and changes of reproductive behavior.

  4. Up-regulated expression of transforming growth factor-alpha in the bronchiolar-alveolar duct regions of asbestos-exposed rats.

    PubMed Central

    Liu, J. Y.; Morris, G. F.; Lei, W. H.; Corti, M.; Brody, A. R.

    1996-01-01

    It has become apparent that the numerous growth factors and cytokines are produced during the development of fibroproliferative lung disease. Investigators must sort out which combinations of these factors are playing mechanistic roles in the disease process. Here we demonstrate that transforming growth factor (TGF)-alpha, a potent epithelial and mesenchymal cell mitogen, is upregulated specifically at the sites of asbestos fiber deposition in the lungs of rats exposed for 5 hours. Unexposed animals and those exposed to high concentrations of iron spheres exhibited no increase in TGF-alpha expression at any time during the experiment. Inhaled asbestos fibers deposit initially at the bronchiolar-alveolar duct regions and alveolar macrophages accumulate at these sites within hours. Non-isotopic in situ hybridization and immunohistochemistry were used to show that the mRNA that codes for TGF-alpha along with the peptide were clearly up-regulated at the bronchiolar-alveolar duct regions by 24 hours after the single asbestos exposure. The numbers of labeled cells demonstrated that expression of the mRNA and protein remained significantly above background for at least 2 weeks after exposure along with increased cell proliferation assessed by staining for proliferating cell nuclear antigen. This, to our knowledge, is the first demonstration of TGF-alpha expression at sites of lung injury in developing fibroproliferative disease. This finding supports the hypothesis that the growth factor is involved in the dramatic epithelial and mesenchymal proliferation we documented previously, although additional experiments will be essential to establish the precise role of TGF-alpha. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:8686744

  5. Up-regulated expression of transforming growth factor-alpha in the bronchiolar-alveolar duct regions of asbestos-exposed rats.

    PubMed

    Liu, J Y; Morris, G F; Lei, W H; Corti, M; Brody, A R

    1996-07-01

    It has become apparent that the numerous growth factors and cytokines are produced during the development of fibroproliferative lung disease. Investigators must sort out which combinations of these factors are playing mechanistic roles in the disease process. Here we demonstrate that transforming growth factor (TGF)-alpha, a potent epithelial and mesenchymal cell mitogen, is upregulated specifically at the sites of asbestos fiber deposition in the lungs of rats exposed for 5 hours. Unexposed animals and those exposed to high concentrations of iron spheres exhibited no increase in TGF-alpha expression at any time during the experiment. Inhaled asbestos fibers deposit initially at the bronchiolar-alveolar duct regions and alveolar macrophages accumulate at these sites within hours. Non-isotopic in situ hybridization and immunohistochemistry were used to show that the mRNA that codes for TGF-alpha along with the peptide were clearly up-regulated at the bronchiolar-alveolar duct regions by 24 hours after the single asbestos exposure. The numbers of labeled cells demonstrated that expression of the mRNA and protein remained significantly above background for at least 2 weeks after exposure along with increased cell proliferation assessed by staining for proliferating cell nuclear antigen. This, to our knowledge, is the first demonstration of TGF-alpha expression at sites of lung injury in developing fibroproliferative disease. This finding supports the hypothesis that the growth factor is involved in the dramatic epithelial and mesenchymal proliferation we documented previously, although additional experiments will be essential to establish the precise role of TGF-alpha.

  6. The Neutrophil Response Induced by an Agonist for Free Fatty Acid Receptor 2 (GPR43) Is Primed by Tumor Necrosis Factor Alpha and by Receptor Uncoupling from the Cytoskeleton but Attenuated by Tissue Recruitment

    PubMed Central

    Björkman, Lena; Mårtensson, Jonas; Winther, Malene; Gabl, Michael; Holdfeldt, André; Uhrbom, Martin; Bylund, Johan; Højgaard Hansen, Anders; Pandey, Sunil K.; Ulven, Trond; Forsman, Huamei

    2016-01-01

    Ligands with improved potency and selectivity for free fatty acid receptor 2 (FFA2R) have become available, and we here characterize the neutrophil responses induced by one such agonist (Cmp1) and one antagonist (CATPB). Cmp1 triggered an increase in the cytosolic concentration of Ca2+, and the neutrophils were then desensitized to Cmp1 and to acetate, a naturally occurring FFA2R agonist. The antagonist CATPB selectively inhibited responses induced by Cmp1 or acetate. The activated FFA2R induced superoxide anion secretion at a low level in naive blood neutrophils. This response was largely increased by tumor necrosis factor alpha (TNF-α) in a process associated with a recruitment of easily mobilizable granules, but neutrophils recruited to an aseptic inflammation in vivo were nonresponding. Superoxide production induced by Cmp1 was increased in latrunculin A-treated neutrophils, but no reactivation of desensitized FFA2R was induced by this drug, suggesting that the cytoskeleton is not directly involved in terminating the response. The functional and regulatory differences between the receptors that recognize short-chain fatty acids and formylated peptides, respectively, imply different roles of these receptors in the orchestration of inflammation and confirm the usefulness of a selective FFA2R agonist and antagonist as tools for the exploration of the precise role of the FFA2R. PMID:27503855

  7. Murine transcription factor alpha A-crystallin binding protein I. Complete sequence, gene structure, expression, and functional inhibition via antisense RNA.

    PubMed

    Brady, J P; Kantorow, M; Sax, C M; Donovan, D M; Piatigorsky, J

    1995-01-20

    alpha A-crystallin binding protein I (alpha A-CRYBP1) is a ubiquitously expressed DNA binding protein that was previously identified by its ability to interact with a functionally important sequence in the mouse alpha A-crystallin gene promoter. Here, we have cloned a single copy gene with 10 exons spanning greater than 70 kb of genomic DNA that encodes alpha A-CRYBP1. The mouse alpha A-CRYBP1 gene specifies a 2,688-amino acid protein with 72% amino acid identity to its human homologue, PRDII-BF1. Both the human and the mouse proteins contain two sets of consensus C2H2 zinc fingers at each end as well a central nonconsensus zinc finger. The alpha A-CRYBP1 gene produces a 9.5-kb transcript in 11 different tissues as well as a testis-specific, 7.7-kb transcript. alpha A-CRYBP1 cDNA clones were isolated from adult mouse brain and testis as well as from cell lines derived from mouse lens (alpha TN4-1) and muscle (C2C12). A single clone isolated from the muscle C2C12 library contains an additional exon near the 5'-end that would prevent production of a functional protein if the normal translation start site were utilized; however, there is another potential initiation codon located downstream that is in frame with the rest of the coding region. In addition, we identified multiple cDNAs from the testis in which the final intron is still present. Finally, we used an antisense expression construct derived from an alpha A-CRYBP1 cDNA clone to provide the first functional evidence that alpha A-CRYBP1 regulates gene expression. When introduced into the alpha TN4-1 mouse lens cell line, the antisense construct significantly inhibited expression from a heterologous promoter that utilized the alpha A-CRYBP1 binding site as an enhancer.

  8. Decreased expression of tumor necrosis factor-alpha-stimulated gene 6 in cumulus cells of the cyclooxygenase-2 and EP2 null mice.

    PubMed

    Ochsner, Scott A; Russell, Darryl L; Day, Anthony J; Breyer, Richard M; Richards, Joanne S

    2003-03-01

    Ovulation, the release of fertilizable oocytes from mature follicles, involves tissue remodeling and increased prostaglandin (PG) signaling. Cyclooxygenase (COX)-2 is the rate-limiting enzyme during PG synthesis. Female mice null for either COX-2 or the PGE(2) receptor EP2 are infertile, show decreased ovulation, and exhibit abnormal cumulus expansion. Cumulus expansion is the production of a complex extracellular matrix surrounding the cumulus-oocyte complex (COC). Matrix components consist of hyaluronan, proteoglycans, and proteins with hyaluronan binding domains. One such hyaluronan binding protein is TNFalpha-stimulated gene 6 (TSG-6). By various methods, we show induction of TSG-6 and hyaluronan synthase-2 mRNA in ovaries of mice treated with pregnant mare serum gonadotropin and human chorionic gonadotropin. By in situ hybridization, we show that both genes are expressed in periantral mural granulosa cells and cumulus cells of the mouse ovary. Notably, RT-PCR and in situ hybridization show that TSG-6 mRNA but not hyaluronan synthase-2 mRNA expression is selectively reduced in cumulus cells of COX-2 and EP2 null mice. Western analysis further confirms that TSG-6 protein is reduced in isolated COCs but remains covalently associated with inter alpha-trypsin inhibitor in COX-2 null mice. These observations identify TSG-6 as a target of PG action and show that its production in ovulatory follicles is associated with proper formation of the cumulus-derived extracellular matrix.

  9. Expression of a tumor necrosis factor-alpha transgene in murine lung causes lymphocytic and fibrosing alveolitis. A mouse model of progressive pulmonary fibrosis.

    PubMed Central

    Miyazaki, Y; Araki, K; Vesin, C; Garcia, I; Kapanci, Y; Whitsett, J A; Piguet, P F; Vassalli, P

    1995-01-01

    The murine TNF-alpha gene was expressed under the control of the human surfactant protein SP-C promoter in transgenic mice. A number of the SP-C TNF-alpha mice died at birth or after a few weeks with very severe lung lesions. Surviving mice transmitted a pulmonary disease to their offspring, the severity and evolution of which was related to the level of TNF-alpha mRNA in the lung; TNF-alpha RNA was detected in alveolar epithelium, presumably in type II epithelial cells. In a longitudinal study of two independent mouse lines, pulmonary pathology, at 1-2 mo of age, consisted of a leukocytic alveolitis with a predominance of T lymphocytes. Leukocyte infiltration was associated with endothelial changes and increased levels of mRNA for the endothelial adhesion molecule VCAM-1. In the following months, alveolar spaces enlarged in association with thickening of the alveolar walls due to an accumulation of desmin-containing fibroblasts, collagen fibers, and lymphocytes. Alveolar surfaces were lined by regenerating type II epithelial cells, and alveolar spaces contained desquamating epithelial cells in places. Platelet trapping in the damaged alveolar capillaries was observed. Pulmonary pathology in the SP-C TNF-alpha mice bears a striking resemblance to human idiopathic pulmonary fibrosis, in which increased expression of TNF-alpha in type II epithelial cells has also been noted. These mice provide a valuable animal model for understanding the pathogenesis of pulmonary fibrosis and exploring possible therapeutic approaches. Images PMID:7542280

  10. Inhibitory activity of the white wine compounds, tyrosol and caffeic acid, on lipopolysaccharide-induced tumor necrosis factor-alpha release in human peripheral blood mononuclear cells.

    PubMed

    Giovannini, L; Migliori, M; Filippi, C; Origlia, N; Panichi, V; Falchi, M; Bertelli, A A E; Bertelli, A

    2002-01-01

    The objective of this study was to assess whether tyrosol and caffeic acid are able to inhibit lipopolysaccharide (LPS)-induced tumor necrosis factor (TNF)-alpha release. TNF is one of the most important cytokines involved in inflammatory reactions. The results show that both tyrosol and caffeic acid are able to inhibit LPS-induced TNF-alpha release from human monocytes, even at low doses. Their mechanisms of action are discussed and we conclude that high doses of the two compounds are not required to achieve effective inhibition of inflammatory reactions due to TNF-alpha release.

  11. Human tumor necrosis factor-alpha gene 3' untranslated region confers inducible toxin responsiveness to homologous promoter in monocytic THP-1 cells.

    PubMed

    Seiler-Tuyns, A; Dufour, N; Spertini, F

    1999-07-30

    To better define the role of 3' untranslated region (3'UTR) on transcriptional regulation of the human tumor necrosis factor (TNF)-alpha gene, monocytic human THP-1 cells were transfected with two TNF-alpha promoter constructs spanning base pairs -1897/-1 and -1214/-1, respectively, and linked to the rabbit beta-globin gene. Quantitative globin gene expression of chimerae was measured by reverse transcription-polymerase chain reaction. A construct linking the chicken beta-actin promoter and a deleted portion of the beta-globin gene was cotransfected and used as internal standard. Unexpectedly, when THP-1 cells were stimulated with lipopolysaccharide or toxic shock syndrome toxin-1, gene regulation was hardly detected. In contrast, endogenous TNF-alpha gene regulation measured by the same reverse transcription-polymerase chain reaction procedure was vigorous. Remarkably, ligation of 3'UTR to chimeric constructs led to a drastic drop in the basal level of chimeric gene expression, resulting in a 15- to 40-fold induction of the reporter gene. Consistently, when the TNF-alpha promoter was replaced by the cytomegalovirus early immediate promoter, gene expression was also uniformly reduced but was no longer up-regulated upon stimulation with lipopolysaccharide and toxic shock syndrome toxin-1. These data provide the first line of evidence that, in addition to its role in TNF-alpha transcript stability and translation, human TNF-alpha 3'UTR also participates in modulating gene expression at the transcriptional level.

  12. Blast-Induced Moderate Neurotrauma (BINT) Elicits Early Complement Activation and Tumor Necrosis Factor Alpha (TNFalpha) Release in a Rat Brain

    DTIC Science & Technology

    2012-04-25

    Morganti-Kossmann MC, Jones JL, Barnum SR. Elevated levels of the complement components C3 and factor B in ventricular cerebrospinal fluid of patients with...Ingersoll SA, Martin CB, Barnum SR, Martin BK. CNS-specific expression of C3a and C5a exacerbate demyelination severity in the cuprizone model. Mol Immunol

  13. The tumor necrosis factor alpha-induced protein 3 (TNFAIP3, A20) imposes a brake on antitumor activity of CD8 T cells

    PubMed Central

    Giordano, Marilyn; Roncagalli, Romain; Bourdely, Pierre; Chasson, Lionel; Buferne, Michel; Yamasaki, Sho; Beyaert, Rudi; van Loo, Geert; Auphan-Anezin, Nathalie; Schmitt-Verhulst, Anne-Marie; Verdeil, Grégory

    2014-01-01

    The transcription factor NF-κB is central to inflammatory signaling and activation of innate and adaptive immune responses. Activation of the NF-κB pathway is tightly controlled by several negative feedback mechanisms, including A20, an ubiquitin-modifying enzyme encoded by the tnfaip3 gene. Mice with selective deletion of A20 in myeloid, dendritic, or B cells recapitulate some human inflammatory pathology. As we observed high expression of A20 transcripts in dysfunctional CD8 T cells in an autochthonous melanoma, we analyzed the role of A20 in regulation of CD8 T-cell functions, using mice in which A20 was selectively deleted in mature conventional T cells. These mice developed lymphadenopathy and some organ infiltration by T cells but no splenomegaly and no detectable pathology. A20-deleted CD8 T cells had increased sensitivity to antigen stimulation with production of large amounts of IL-2 and IFNγ, correlated with sustained nuclear expression of NF-κB components reticuloendotheliosis oncogene c-Rel and p65. Overexpression of A20 by retroviral transduction of CD8 T cells dampened their intratumor accumulation and antitumor activity. In contrast, relief from the A20 brake in NF-κB activation in adoptively transferred antitumor CD8 T cells led to improved control of melanoma growth. Tumor-infiltrating A20-deleted CD8 T cells had enhanced production of IFNγ and TNFα and reduced expression of the inhibitory receptor programmed cell death 1. As manipulation of A20 expression in CD8 T cells did not result in pathologic manifestations in the mice, we propose it as a candidate to be targeted to increase antitumor efficiency of adoptive T-cell immunotherapy. PMID:25024217

  14. Role of the adhesion molecule lymphocyte function associated antigen 1 in toxic shock syndrome toxin 1-induced tumor necrosis factor alpha and interleukin-1 beta secretion by human monocytes.

    PubMed Central

    See, R H; Chow, A W

    1992-01-01

    We previously demonstrated that the induction by staphylococcal toxic shock syndrome toxin 1 (TSST-1) of tumor necrosis factor alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) secretion by human monocytes requires direct T cell-monocyte contact. In the present study, a role for the adhesion molecule lymphocyte function associated antigen 1 (LFA-1) in TSST-1-induced cytokine secretion by human monocytes among 12 normal healthy donors was investigated. Monoclonal antibodies to the alpha chain (anti-CD11a) and to the beta chain (anti-CD18) of LFA-1 significantly inhibited TSST-1-induced TNF-alpha and IL-1 beta secretion (P < 0.025; Wilcoxon signed-rank test, two tailed), while a control monoclonal antibody directed against the monocyte CD14 antigen had no effect. These results suggest that LFA-1 may play an important role in the secretion of TNF-alpha and IL-1 beta by TSST-1-stimulated human monocytes, likely by promoting cell-cell adhesion between monocytes and lymphocytes. PMID:1399006

  15. Shiga Toxin 1 Induces on Lipopolysaccharide-Treated Astrocytes the Release of Tumor Necrosis Factor-alpha that Alter Brain-Like Endothelium Integrity

    PubMed Central

    Landoni, Verónica I.; Schierloh, Pablo; de Campos Nebel, Marcelo; Fernández, Gabriela C.; Calatayud, Cecilia; Lapponi, María J.; Isturiz, Martín A.

    2012-01-01

    The hemolytic uremic syndrome (HUS) is characterized by hemolytic anemia, thrombocytopenia and renal dysfunction. The typical form of HUS is generally associated with infections by Gram-negative Shiga toxin (Stx)-producing Escherichia coli (STEC). Endothelial dysfunction induced by Stx is central, but bacterial lipopolysaccharide (LPS) and neutrophils (PMN) contribute to the pathophysiology. Although renal failure is characteristic of this syndrome, neurological complications occur in severe cases and is usually associated with death. Impaired blood-brain barrier (BBB) is associated with damage to cerebral endothelial cells (ECs) that comprise the BBB. Astrocytes (ASTs) are inflammatory cells in the brain and determine the BBB function. ASTs are in close proximity to ECs, hence the study of the effects of Stx1 and LPS on ASTs, and the influence of their response on ECs is essential. We have previously demonstrated that Stx1 and LPS induced activation of rat ASTs and the release of inflammatory factors such as TNF-α, nitric oxide and chemokines. Here, we demonstrate that rat ASTs-derived factors alter permeability of ECs with brain properties (HUVECd); suggesting that functional properties of BBB could also be affected. Additionally, these factors activate HUVECd and render them into a proagregant state promoting PMN and platelets adhesion. Moreover, these effects were dependent on ASTs secreted-TNF-α. Stx1 and LPS-induced ASTs response could influence brain ECs integrity and BBB function once Stx and factors associated to the STEC infection reach the brain parenchyma and therefore contribute to the development of the neuropathology observed in HUS. PMID:22479186

  16. Reversion of lethality and growth defects in Fatiga oxygen-sensor mutant flies by loss of hypoxia-inducible factor-alpha/Sima.

    PubMed

    Centanin, Lázaro; Ratcliffe, Peter J; Wappner, Pablo

    2005-11-01

    Hypoxia-Inducible Factor (HIF) prolyl hydroxylase domains (PHDs) have been proposed to act as sensors that have an important role in oxygen homeostasis. In the presence of oxygen, they hydroxylate two specific prolyl residues in HIF-alpha polypeptides, thereby promoting their proteasomal degradation. So far, however, the developmental consequences of the inactivation of PHDs in higher metazoans have not been reported. Here, we describe novel loss-of-function mutants of fatiga, the gene encoding the Drosophila PHD oxygen sensor, which manifest growth defects and lethality. We also report a null mutation in dHIF-alpha/sima, which is unable to adapt to hypoxia but is fully viable in normoxic conditions. Strikingly, loss-of-function mutations of sima rescued the developmental defects observed in fatiga mutants and enabled survival to adulthood. These results indicate that the main functions of Fatiga in development, including control of cell size, involve the regulation of dHIF/Sima.

  17. Inhibition of tumor necrosis factor-alpha and cyclooxigenase-2 by Isatin: a molecular mechanism of protection against TNBS-induced colitis in rats.

    PubMed

    Socca, Eduardo Augusto Rabelo; Luiz-Ferreira, Anderson; de Faria, Felipe Meira; de Almeida, Ana Cristina; Dunder, Ricardo José; Manzo, Luis Paulo; Brito, Alba Regina Monteiro Souza

    2014-02-25

    Isatin, an indole alkaloid has been shown to have anti-microbial, anti-tumor and anti-inflammatory effects. Due to its findings, we evaluated whether this alkaloid would have any effect on TNBS-induced colitis. Animals (male Unib:WH rats, aged 8 weeks old) were induced colitis through a rectal administration of 2,4,6-trinitrobenzene sulphonic acid using a catheter inserted 8 cm into the rectum of the animals. The rats were divided into two major groups: non-colitic and colitic. The colitic group was sub-divided into 6 groups (10 animals per group): colitic non-treated, Isatin 3; 6; 12.5; 18.75 and 25 mg/kg. Our main results showed that the oral treatment with Isatin 6 and 25 mg/kg were capable of avoiding the increase in TNF-α, COX-2 and PGE₂ levels when compared to the colitic non-treated group. Interestingly, the same doses (6 and 25 mg/kg) were also capable of preventing the decrease in IL-10 levels comparing with the colitic non-treated group. The levels of MPO, (an indirect indicator of neutrophil presence), were also maintained lower than those of the colitic non-treated group. Isatin also prevented the decrease of SOD activity and increase of GSH-Px and GSH-Rd activity as well as the depletion of GSH levels. In conclusion, both pre-treatments (6 and 25 mg/kg) were capable of protecting the gut mucosa against the injury caused by TNBS, through the combination of antioxidant and anti-inflammatory properties, which, together, showed a protective activity of the indole alkaloid Isatin.

  18. Cytosine-Phosphorothionate-Guanine Oligodeoxynucleotides Exacerbates Hemophagocytosis by Inducing Tumor Necrosis Factor-Alpha Production in Mice after Bone Marrow Transplantation.

    PubMed

    Liu, Jiajia; Guo, Yong-Mei; Onai, Nobuyuki; Ohyagi, Hideaki; Hirokawa, Makoto; Takahashi, Naoto; Tagawa, Hiroyuki; Ubukawa, Kumi; Kobayashi, Isuzu; Tezuka, Hiroyuki; Minamiya, Yoshihiro; Ohteki, Toshiaki; Sawada, Kenichi

    2016-04-01

    Hemophagocytic syndrome (HPS) is frequently associated with hematopoietic stem cell transplantation and is treated with some benefit derived from TNF-α inhibitors. However, the mechanisms of how HPS occurs and how a TNF-α inhibitor exerts some benefit to HPS management have remained unclear. We evaluated the effect of toll-like receptor (TLR) ligands, especially focusing on cytosine-phosphorothionate-guanine oligodeoxynucleotide (CpG), a TLR9 ligand, on HPS in mice that underwent transplantation with syngeneic or allogeneic bone marrow (BM) cells (Syn-BMT, Allo-BMT), or with allogeneic BM cells plus splenocytes to promote graft-versus-host disease (GVHD mice). Hemophagocytosis was a common feature early after all BMT, but it subsided in Syn-BMT and Allo-BMT mice. In GVHD mice, however, hemophagocytosis persisted and was accompanied by upregulated production of IFN-γ but not TNF-α, and it was suppressed by blockade of IFN-γ but not TNF-α. A single injection of the TLR9 ligand CpG promoted HPS in all BMT mice and was lethal in GVHD mice, accompanied by greatly upregulated production of TNF-α, IL-6, and IFN-γ. Blocking of TNF-α, but not IL-6 or IFN-γ, suppressed CpG-induced HPS in all BMT mice and rescued GVHD mice from CpG-induced mortality. Thus, TLR9 signaling mediates TNF-α-driven HPS in BMT mice and is effectively treated through TNF-α inhibition.

  19. Antioxidant alpha-lipoic acid inhibits osteoclast differentiation by reducing nuclear factor-kappaB DNA binding and prevents in vivo bone resorption induced by receptor activator of nuclear factor-kappaB ligand and tumor necrosis factor-alpha.

    PubMed

    Kim, Hyon Jong; Chang, Eun-Ju; Kim, Hyun-Man; Lee, Seung Bok; Kim, Hyun-Duck; Su Kim, Ghi; Kim, Hong-Hee

    2006-05-01

    The relationship between oxidative stress and bone mineral density or osteoporosis has recently been reported. As bone loss occurring in osteoporosis and inflammatory diseases is primarily due to increases in osteoclast number, reactive oxygen species (ROS) may be relevant to osteoclast differentiation, which requires receptor activator of nuclear factor-kappaB ligand (RANKL). Tumor necrosis factor-alpha (TNF-alpha) frequently present in inflammatory conditions has a profound synergy with RANKL in osteoclastogenesis. In this study, we investigated the effects of alpha-lipoic acid (alpha-LA), a strong antioxidant clinically used for some time, on osteoclast differentiation and bone resorption. At concentrations showing no growth inhibition, alpha-LA potently suppressed osteoclastogenesis from bone marrow-derived precursor cells driven either by a high-dose RANKL alone or by a low-dose RANKL plus TNF-alpha (RANKL/TNF-alpha). alpha-LA abolished ROS elevation by RANKL or RANKL/TNF-alpha and inhibited NF-kappaB activation in osteoclast precursor cells. Specifically, alpha-LA reduced DNA binding of NF-kappaB but did not inhibit IKK activation. Furthermore, alpha-LA greatly suppressed in vivo bone loss induced by RANKL or TNF-alpha in a calvarial remodeling model. Therefore, our data provide evidence that ROS plays an important role in osteoclast differentiation through NF-kappaB regulation and the antioxidant alpha-lipoic acid has a therapeutic potential for bone erosive diseases.

  20. The major acute-phase protein, serum amyloid P component, in mice is not involved in endogenous resistance against tumor necrosis factor alpha-induced lethal hepatitis, shock, and skin necrosis.

    PubMed

    Van Molle, W; Hochepied, T; Brouckaert, P; Libert, C

    2000-09-01

    The proinflammatory cytokine tumor necrosis factor alpha (TNF-alpha) induces lethal hepatitis when injected into D-(+)-galactosamine-sensitized mice on the one hand or systemic inflammatory response syndrome (SIRS) in normal mice on the other hand. We studied whether serum amyloid P component (SAP), the major acute-phase protein in mice, plays a protective role in both lethal models. For this purpose, we used SAP(0/0) mice generated by gene targeting. We studied the lethal response of SAP(0/0) or SAP(+/+) mice to both lethal triggers but found no differences in the sensitivity of both types of mice. We also investigated whether SAP is involved in establishing two types of endogenous protection: one using a single injection of interleukin-1beta (IL-1beta) for desensitization and clearly involving a liver protein, the other by tolerizing mice for 5 days using small doses of human TNF-alpha. Although after IL-1beta or after tolerization the SAP levels in the serum had risen fourfold in the control mice and not in the SAP(0/0) mice, the same extents of desensitization and tolerization were achieved. Finally, we observed that the induction of hemorrhagic necrosis in the skin of mice by two consecutive local injections with TNF-alpha was not altered in SAP(0/0) mice. We conclude that the presence or absence of SAP has no influence on the sensitivity of mice to TNF-alpha-induced hepatitis, SIRS, and hemorrhagic necrosis or on the endogenous protective mechanisms of desensitization or tolerization.

  1. Toll-like receptor 2- and 6-mediated stimulation by macrophage-activating lipopeptide 2 induces lipopolysaccharide (LPS) cross tolerance in mice, which results in protection from tumor necrosis factor alpha but in only partial protection from lethal LPS doses.

    PubMed

    Deiters, Ursula; Gumenscheimer, Marina; Galanos, Chris; Mühlradt, Peter F

    2003-08-01

    Patients or experimental animals previously exposed to lipopolysaccharide (LPS) become tolerant to further LPS challenge. We investigated the potential of the macrophage-activating lipopeptide 2 (MALP-2) to induce in vivo cross tolerance to tumor necrosis factor alpha (TNF-alpha) and LPS. MALP-2-induced tolerance could be of practical interest, as MALP-2 proved much less pyrogenic in rabbits than LPS. Whereas LPS signals via Toll-like receptor 4 (TLR4), MALP-2 uses TLR2 and TLR6. LPS-mediated cytokine release was studied in mice pretreated with intraperitoneal injections of MALP-2. No biologically active TNF-alpha could be detected in the serum of MALP-2-treated animals when challenged with LPS 24 or 72 h later, whereas suppression of LPS-dependent interleukin (IL)-6 lasted for only 24 h. Protection from lethal TNF-alpha shock was studied in galactosamine-treated mice. Dose dependently, MALP-2 prevented death from lethal TNF-alpha doses in TLR4(-/-) but not in TLR2(-/-) mice, with protection lasting from 5 to 24 h. To assay protection from LPS, mice were pretreated with MALP-2 doses of up to 10 micro g. Five and 24 h later, the animals were simultaneously sensitized and challenged by intravenous coinjection of galactosamine and a lethal dose of 50 ng of LPS. There was only limited protection (four of seven mice survived) when mice were challenged 5 h after MALP-2 pretreatment, and no protection when mice were challenged at later times. The high effectiveness of MALP-2 in suppressing TNF-alpha, the known ways of biological inactivation, and low pyrogenicity make MALP-2 a potential candidate for clinical use.

  2. Roles of tumor necrosis factor alpha, granulocyte-macrophage colony-stimulating factor, platelet-activating factor, and arachidonic acid metabolites in interleukin-1-induced resistance to infection in neutropenic mice.

    PubMed Central

    Vogels, M T; Hermsen, C C; Huys, H L; Eling, W M; van der Meer, J W

    1994-01-01

    Treatment with a single low dose (80 to 800 ng) of interleukin-1 (IL-1) 24 h before a lethal bacterial challenge in granulocytopenic and in normal mice enhances nonspecific resistance. The mechanism behind this protection has only partially been elucidated. Since IL-1 induces production of tumor necrosis factor alpha (TNF-alpha), granulocyte-macrophage colony-stimulating factor (GM-CSF), platelet-activating factor (PAF), and arachidonic acid metabolites, we investigated the potential role of these substances in IL-1-induced protection. Low doses of murine TNF-alpha but not of human TNF-alpha enhanced survival, suggesting an effect via the type II TNF receptor rather than the type I TNF receptor, which has little species specificity. In line with this TNF-alpha-induced protection from infection, pretreatment with a low dose of a rat anti-murine TNF-alpha monoclonal antibody tended to inhibit IL-1-induced protection, suggesting a role of TNF-alpha as a mediator of IL-1-induced enhanced resistance to infection. Pretreatment with higher doses of anti-TNF-alpha, however, showed a dose-related protective effect per se, which could be further enhanced by a suboptimal dose of IL-1. A combination of optimal doses of anti-TNF-alpha and IL-1 produced an increase in survival similar to that produced by separate pretreatments. This lack of further enhancement of survival by combined optimal pretreatments suggests a similar mechanism of protection, most likely attenuation of deleterious effects of overproduced proinflammatory cytokines like TNF-alpha during lethal infection. Pretreatment with different doses of GM-CSF before a lethal Pseudomonas aeruginosa challenge in neutropenic mice did not enhance survival. Different doses of WEB 2170, a selective PAF receptor antagonist, of MK-886, a selective inhibitor of leukotriene biosynthesis, or of several cyclooxygenase inhibitors did not reduce the protective effect of IL-1 pretreatment. We conclude that IL-1-induced nonspecific

  3. Growth and major histocompatibility antigen expression regulation by IL-4, interferon-gamma (IFN-gamma) and tumour necrosis factor-alpha (TNF-alpha) on human renal cell carcinoma.

    PubMed Central

    Hillman, G G; Puri, R K; Kukuruga, M A; Pontes, J E; Haas, G P

    1994-01-01

    We have recently shown that human renal cell carcinoma (RCC) tumour lines express high-affinity IL-4 receptors. Binding of IL-4 to RCC cells induced a growth inhibition in the range of 20-68%. To enhance the growth inhibitory effect of IL-4, we have tested the effects of two additional cytokines capable of directly affecting tumour cell growth. IFN-gamma caused a significant inhibition of RCC tumour cell growth (up to 70%) in a dose-dependent manner, whereas the effect of TNF-alpha was more limited (0-20% inhibition). The addition of IL-4 to IFN-gamma on RCC cells sensitive to IL-4 induced a greater inhibition of cell growth than that seen with each cytokine alone. IL-4 and IFN-gamma rendered RCC cells more responsive to the inhibitory effect mediated by TNF-alpha. The combination of TNF-alpha with IL-4 and IFN-gamma induced an optimal growth inhibition (up to 90-98%) of RCC cells. In addition to a direct anti-proliferative effect, we have demonstrated that these cytokines can also enhance the expression of MHC antigens on the surface of RCC tumour cell lines which may render the cells more immunogenic. All RCC lines tested expressed class I antigens, but not class II antigens. IFN-gamma induced class II expression and up-regulated the expression of class I antigens on RCC cells. Class II antigen expression was detectable following 48 h incubation, and greater after 72 h with IFN-gamma. IL-4 minimally affected class I expression, whereas TNF-alpha up-regulated class I antigen expression. IL-4 or TNF-alpha did not induce class II expression. The combination of the three cytokines slightly augmented the up-regulation of class I and class II antigens observed with IFN-gamma alone. These observations confirm the direct interaction of IL-4, IFN-gamma and TNF-alpha with RCC tumour cells, both at the level of growth regulation and MHC antigen expression, and suggest a therapeutic potential of the combination of the three cytokines for renal cell carcinoma. PMID:8004818

  4. Cytokines in chronic inflammatory arthritis. V. Mutual antagonism between interferon-gamma and tumor necrosis factor-alpha on HLA-DR expression, proliferation, collagenase production, and granulocyte macrophage colony-stimulating factor production by rheumatoid arthritis synoviocytes.

    PubMed Central

    Alvaro-Gracia, J M; Zvaifler, N J; Firestein, G S

    1990-01-01

    The effects of a broad array of cytokines, individually and in combination, were determined on separate functions (proliferation, collagenase production, and granulocyte macrophage colony-stimulating factor [GM-CSF] production) and phenotype (expression of class II MHC antigens) of cultured fibroblast-like RA synoviocytes. The following recombinant cytokines were used: IL-1 beta, IL-2, IL-3, IL-4, IFN-gamma, tumor necrosis factor (TNF)-alpha, GM-CSF, and macrophage colony-stimulating factor (M-CSF). Only IFN-gamma induced HLA-DR (but not HLA-DQ) expression. TNF-alpha inhibited IFN-gamma-mediated HLA-DR expression (46.7 +/- 4.1% inhibition) and HLA-DR mRNA accumulation. This inhibitory effect was also observed in osteoarthritis synoviocytes. Only TNF-alpha and IL-1 increased synoviocyte proliferation (stimulation index 3.60 +/- 1.03 and 2.31 +/- 0.46, respectively). IFN-gamma (but none of the other cytokines) inhibited TNF-alpha-induced proliferation (70 +/- 14% inhibition) without affecting the activity of IL-1. Only IL-1 beta and TNF-alpha induced collagenase production (from less than 0.10 U/ml to 1.10 +/- 0.15 and 0.72 +/- 0.24, respectively). IFN-gamma decreased TNF-alpha-mediated collagenase production (69 +/- 19% inhibition) and GM-CSF production but had no effect on the action of IL-1. These data demonstrate mutual antagonism between IFN-gamma and TNF-alpha on fibroblast-like synoviocytes and suggest a novel homeostatic control mechanism that might be defective in RA where very little IFN-gamma is produced. Images PMID:2174906

  5. Mesenchymal stromal cells expressing ErbB-2/neu elicit protective antibreast tumor immunity in vivo, which is paradoxically suppressed by IFN-gamma and tumor necrosis factor-alpha priming.

    PubMed

    Romieu-Mourez, Raphaëlle; François, Moïra; Abate, Amanda; Boivin, Marie-Noëlle; Birman, Elena; Bailey, Dana; Bramson, Jonathan L; Forner, Kathy; Young, Yoon-Kow; Medin, Jeffrey A; Galipeau, Jacques

    2010-10-15

    It is unknown whether mesenchymal stromal cells (MSC) can regulate immune responses targeting tumor autoantigens of low immunogenicity. We tested here whether immunization with MSC could break immune tolerance towards the ErbB-2/HER-2/neu tumor antigen and the effects of priming with IFN-γ and tumor necrosis factor-α (TNF-α) on this process. BALB/c- and C57BL/6-derived MSC were lentivirally transduced to express a kinase-inactive rat neu mutant (MSC/Neu). Immunization of BALB/c mice with nontreated or IFN-γ-primed allogeneic or syngeneic MSC/Neu induced similar levels of anti-neu antibody titers; however, only syngeneic MSC/Neu induced protective neu-specific CD8(+) T cell responses. Compared to immunization with nontreated or IFN-γ-primed syngeneic MSC/Neu, the number of circulating neu-specific CD8(+) T cells and titers of anti-neu antibodies were observed to be decreased after immunizations with IFN-γ- plus TNF-α-primed MSC/Neu. In addition, syngeneic MSC/Neu seemed more efficient than IFN-γ-primed MSC/Neu at inducing a protective therapeutic antitumor immune response resulting in the regression of transplanted neu-expressing mammary tumor cells. In vitro antigen-presenting cell assays performed with paraformaldehyde-fixed or live MSC showed that priming with IFN-γ plus TNF-α, compared to priming with IFN-γ alone, increased antigen presentation as well as the production of immunosuppressive factors. These data suggest that whereas MSC could effectively serve as antigen-presenting cells to induce immune responses aimed at tumor autoantigens, these functions are critically regulated by IFN-γ and TNF-α.

  6. The interferon-gamma-induced GTPase, mGBP-2, inhibits tumor necrosis factor alpha (TNF-alpha) induction of matrix metalloproteinase-9 (MMP-9) by inhibiting NF-kappaB and Rac protein.

    PubMed

    Balasubramanian, Sujata; Fan, Meiyun; Messmer-Blust, Angela F; Yang, Chuan H; Trendel, Jill A; Jeyaratnam, Jonathan A; Pfeffer, Lawrence M; Vestal, Deborah J

    2011-06-03

    Matrix metalloproteinase-9 (MMP-9) is important in numerous normal and pathological processes, including the angiogenic switch during tumor development and tumor metastasis. Whereas TNF-α and other cytokines up-regulate MMP-9 expression, interferons (IFNs) inhibit MMP-9 expression. We found that IFN-γ treatment or forced expression of the IFN-induced GTPase, mGBP-2, inhibit TNF-α-induced MMP-9 expression in NIH 3T3 fibroblasts, by inhibiting MMP-9 transcription. The NF-κB transcription factor is required for full induction of MMP-9 by TNF-α. Both IFN-γ and mGBP-2 inhibit the transcription of a NF-κB-dependent reporter construct, suggesting that mGBP-2 inhibits MMP-9 induction via inhibition of NF-κB-mediated transcription. Interestingly, mGBP-2 does not inhibit TNF-α-induced degradation of IκBα or p65/RelA translocation into the nucleus. However, mGBP-2 inhibits p65 binding to a κB oligonucleotide probe in gel shift assays and to the MMP-9 promoter in chromatin immunoprecipitation assays. In addition, TNF-α activation of NF-κB in NIH 3T3 cells is dependent on Rac activation, as evidenced by the inhibition of TNF-α induction of NF-κB-mediated transcription by a dominant inhibitory form of Rac1. A role for Rac in the inhibitory action of mGBP-2 on NF-κB is further shown by the findings that mGBP-2 inhibits TNF-α activation of endogenous Rac and constitutively activate Rac can restore NF-κB transcription in the presence of mGBP-2. This is a novel mechanism by which IFNs can inhibit the cytokine induction of MMP-9 expression.

  7. Hypoxia augments lipopolysaccharide-induced cytokine expression in periodontal ligament cells.

    PubMed

    Jian, Congxiang; Li, Chenjun; Ren, Yu; He, Yong; Li, Yunming; Feng, Xiaodan; Zhang, Gang; Tan, Yinghui

    2014-10-01

    Periodontitis is a chronic inflammatory disease characterized by the destruction of tooth supporting tissues. Hypoxia, the mainly changes of the plateau environment, can induce severe periodontitis by animal experiments. There is, however, very little information on hypoxia and lipopolysaccharide (LPS) induced cytokine expression in periodontal ligament (PDL) cells. In this article, we characterized hypoxia or P. gingivalis lipopolysaccharide (Pg LPS) induced tumor necrosis factor alpha (TNF-α), interleukin (IL)-1β, and IL-6 expression by human periodontal ligament (hPDL) cells. We found that hypoxia augmented Pg LPS induced TNF-α, IL-1β, and IL-6 expression in hPDL cells. We also demonstrated that nuclear factor kappa B pathway was involved in hypoxia augmenting Pg LPS induced cytokine expression in hPDL cells. Thus, our results suggest that the hypoxic environment may enhance the immune function of hPDL cells that is induced by Pg LPS.

  8. Tumor necrosis factor-alpha and interferon-gamma, but not HTLV-I tax, are likely factors in the epidermotropism of cutaneous T-cell lymphoma via induction of interferon-inducible protein-10.

    PubMed

    Daliani, D; Ulmer, R A; Jackow, C; Pugh, W; Gansbacher, B; Cabanillas, F; Duvic, M; Sarris, A H

    1998-04-01

    We have previously shown that Interferon-Inducible Protein-10 (IP-10), a cytokine chemotactic for CD4-positive lymphocytes, is overexpressed by lesional epidermal keratinocytes and probably accounts for the epidermotropism of cutaneous T-cell lymphoma (CTCL). The tax gene of human T-lymphotropic virus-I (HTLV-I) immortalizes CD4-positive lymphocytes, induces IFN-gamma, and has been detected in patients with classical CTCL who are seronegative for HTLV-I. TNF-alpha is synergistic with IFN-gamma for the induction of IP-10. We therefore decided to define the presence of tax, IFN-gamma, TNF-alpha, and IP-10 in lesions of 19 adults with classical CTCL who were seronegative for HTLV-I. Lesional mRNAs for actin, TNF-alpha, IFN-gamma, and tax were detected by reverse-transcriptase polymerase chain reaction (RT-PCR) amplification. In addition IP-10, TNF-alpha, and IFN-gamma were detected and localized with immunocytochemistry of frozen sections. In agreement with previous observations IP-10 was overexpressed in lesional keratinocytes of all 19 patients. By RT-PCR, mRNA for IFN-gamma was detected in lesions of 8, and for TNF-alpha in lesions of 13 patients. By immunocytochemistry, TNF-alpha was expressed by lesional keratinocytes in 10 of 13 tested patients, whereas IFN-gamma was focally expressed by lesional lymphocytes and faintly by lesional keratinocytes in 9 of 13 tested patients. tax mRNA was not detected in lesions of any patient, but was easily detectable in cutaneous lesions or peripheral blood of control patients who were seropositive for HTLV-I. We conclude that TNF-alpha and IFN-gamma may cause epidermotropism by inducing IP-10. However, the tax gene of HTLV-I does not appear to be involved in the pathogenesis of classical CTCL.

  9. Effect of tetrandrine on calcium-dependent tumour necrosis factor-alpha production in glia-neurone mixed cultures.

    PubMed

    Wang, Bin; Yang, Li; Yan, Hong-Li; Wang, Meng; Xiao, Ji-Gao

    2005-10-01

    Tumour necrosis factor-alpha is believed to have a deleterious role in the pathophysiology of brain injury. Tetrandrine has protective effect on neuronal cells, however, the mechanisms involved in its action have not been clearly established. The aim of this study was to investigate the role of tetrandrine on calcium-dependent tumour necrosis factor-alpha production in glia-neurone mixed cultures. Glia-neurone mixed cultures were treated by addition of Ca2+ regulating agents for a period of 6 hr. Tetrandrine or/and TMB-8 were added 30 min. before the stimulation. The supernatant tumour necrosis factor-alpha levels were quantified by enzyme-linked immunosorbent assay. Exposure of lipopolysaccharide 10 and 100 ng/ml caused significant increase in tumour necrosis factor-alpha production respectively, with no alteration in cultures treated with 1 ng/ml lipopolysaccharide. Glia-neurone mixed cultures exhibited a marked elevation in tumour necrosis factor-alpha production after exposure to CaCl2, KCl, thapsigargin, BHQ and norepinephrine in the presence of lipopolysaccharide at 1 ng/ml respectively. Tetrandrine 0.3, 1, and 3 microM concentration-dependently reduced tumour necrosis factor-alpha production evoked by CaCl2 or KCl. Tetrandrine preincubation had no significant effect on the response to Ca2+-ATPase inhibitor thapsigargin or BHQ. Norepinephrine-induced tumour necrosis factor-alpha production was significantly reduced by tetrandrine and almost abolished by combination of tetrandrine and intracellular Ca2+ release inhibitor TMB-8. These results suggested that tetrandrine at a concentration of 0.3, 1, or 3 microM inhibited tumour necrosis factor-alpha production induced by Ca2+ entry in glia-neurone mixed cultures.

  10. Tumor necrosis factor-alpha inhibits pre-osteoblast differentiation through its type-1 receptor.

    PubMed

    Abbas, Sabiha; Zhang, Yan-Hong; Clohisy, John C; Abu-Amer, Yousef

    2003-04-01

    Tumor necrosis factor-alpha (TNF) is a pro-inflammatory cytokine with a profound role in many skeletal diseases. The cytokine has been described as a mediator of bone loss in osteolysis and other inflammatory bone diseases. In addition to its known bone resorptive action, TNF reduces bone formation by inhibiting osteoblast differentiation. Using primary and transformed osteoblastic cells, we first document that TNF inhibits expression of alkaline phosphatase and matrix deposition, both considered markers of osteoblast differentiation. The effects are dose- and time-dependent. Core-binding factor A1 (cbfa1) is a transcription factor critical for osteoblast differentiation, and we show here that it is activated by the osteoblast differentiation agent, beta-glycerophosphate. Therefore, we investigated whether the inhibitory effects of TNF were associated with altered activity of this transcription factor. Using retardation assays, we show that TNF significantly inhibits cbfal activation by beta-glycerophosphate, manifested by reduced DNA-binding activity. Next, we turned to determine the signaling pathway by which TNF inhibits osteoblast differentiation. Utilizing animals lacking individual TNF receptors, we document that TNFr1 is required for transmitting the cytokine's inhibitory effect. In the absence of this receptor, TNF failed to impact all osteoblast differentiation markers tested. In summary, TNF blocks expression of osteoblast differentiation markers and inhibits beta-glycerophosphate-induced activation of the osteoblast differentiation factor cbfa1. Importantly, these effects are mediated via a mechanism requiring the TNF type-1 receptor.

  11. Targeted Cancer Therapy with Tumor Necrosis Factor-Alpha

    PubMed Central

    Cai, Weibo; Kerner, Zachary J.; Hong, Hao; Sun, Jiangtao

    2013-01-01

    Tumor necrosis factor-alpha (TNF-α), a member of the TNF superfamily, was the first cytokine to be evaluated for cancer biotherapy. However, the clinical use of TNF-α is severely limited by its toxicity. Currently, TNF-α is administered only through locoregional drug delivery systems such as isolated limb perfusion and isolated hepatic perfusion. To reduce the systemic toxicity of TNF-α, various strategies have been explored over the last several decades. This review summarizes current state-of-the-art targeted cancer therapy using TNF-α. Passive targeting, cell-based therapy, gene therapy with inducible or tissue-specific promoters, targeted polymer-DNA complexes, tumor pre-targeting, antibody-TNF-α conjugate, scFv/TNF-α fusion proteins, and peptide/TNF-α fusion proteins have all been investigated to combat cancer. Many of these agents are already in advanced clinical trials. Molecular imaging, which can significantly speed up the drug development process, and nanomedicine, which can integrate both imaging and therapeutic components, has the potential to revolutionize future cancer patient management. Cooperative efforts from scientists within multiple disciplines, as well as close partnerships among many organizations/entities, are needed to quickly translate novel TNF-α-based therapeutics into clinical investigation. PMID:24115841

  12. Autophagosomal IkappaB alpha degradation plays a role in the long term control of tumor necrosis factor-alpha-induced nuclear factor-kappaB (NF-kappaB) activity.

    PubMed

    Colleran, Amy; Ryan, Aideen; O'Gorman, Angela; Mureau, Coralie; Liptrot, Catherine; Dockery, Peter; Fearnhead, Howard; Egan, Laurence J

    2011-07-01

    Transcription factor NF-κB is persistently activated in many chronic inflammatory diseases and cancers. The short term regulation of NF-κB is well understood, but little is known about the mechanisms of its long term activation. We studied the effect of a single application of TNF-α on NF-κB activity for up to 48 h in intestinal epithelial cells. Results show that NF-κB remained persistently activated up to 48 h after TNF-α and that the long term activation of NF-κB was accompanied by a biphasic degradation of IκBα. The first phase of IκBα degradation was proteasome-dependent, but the second was not. Further investigation showed that TNF-α stimulated formation of autophagosomes in intestinal epithelial cells and that IκBα co-localized with autophagosomal vesicles. Pharmacological or genetic blockade of autophagosome formation or the inhibition of lysosomal proteases decreased TNF-α-induced degradation of IκBα and lowered NF-κB target gene expression. Together, these findings indicate a role of autophagy in the control of long term NF-κB activity. Because abnormalities in autophagy have been linked to ineffective innate immunity, we propose that alterations in NF-κB may mediate this effect.

  13. Angiotensin-converting enzyme inhibitors improve hepatic steatosis by modulating expression of tumour necrosis factor-alpha, interleukin-6 and adiponectin receptor-2 in rats with type 2 diabetes.

    PubMed

    Zhang, Xia; Li, Zhong-Zhuan; Liu, Dong-Fang; Xu, Xin; Mei, Zhe-Chuan; Shen, Wei

    2009-07-01

    1. Angiotensin-converting enzyme inhibitors (ACEI) are hypotensive drugs that have been shown to prevent Type 2 diabetes mellitus (T2DM) in high-risk individuals. However, in T2DM, the effects of ACEI on hepatic steatosis are not known. The aim of the present study was to examine the effects of ACEI on changes in liver histology and hepatic mRNA expression of adipokines in rats with T2DM. 2. Thirty-six rats were divided into a normal control group, a T2DM group and a fosinopril-treated group. After six weeks of treatment with 5 mg/kg per day fosinopril, an ACEI, changes in liver histology, serum fasting glucose (FG), insulin, triglyceride (TG), total cholesterol (TC), alanine aminotransferase (ALT), aspartate aminotransferase (AST), tumour necrosis factor (TNF)-alpha, interleukin (IL)-6, adiponectin were evaluated, as was hepatic TNF-alpha, IL-6 and adiponectin receptor-2 (adipoR2) mRNA expression. 3. The degree of hepatic steatosis and inflammation, serum FG, insulin, TG, TC, ALT, TNF-alpha and IL-6 concentrations and hepatic TNF-alpha and IL-6 mRNA expression were significantly higher in rats with T2DM than in normal controls. Serum adiponectin concentrations and hepatic adipoR2 mRNA expression in rats with T2DM were significantly lower than in normal controls. Fosinopril significantly reduced the degree of hepatic steatosis, serum FG, insulin, ALT, TNF-alpha and IL-6 concentrations and hepatic TNF-alpha and IL-6 mRNA expression. Fosinopril significantly increased serum adiponectin concentrations and hepatic adipoR2 mRNA expression. 4. In conclusion, the ACEI improved insulin sensitivity and hepatic steatosis in rats with T2DM by increasing circulating adiponectin and hepatic adipoR2 levels, in addition to reducing pro-inflammatory cytokine levels in the circulation and liver.

  14. Role of tissue factor in the antitumor effect of recombinant human tumor necrosis factor-alpha in mice.

    PubMed

    Nishigaki, F; Miyayasu, K; Tsujimoto, S; Manda, T; Shimomura, K

    1994-01-01

    Recombinant tumor necrosis factor-alpha (rTNF-alpha) inhibited tumor growth of Meth A fibrosarcoma (Meth A) solid tumor in mice, and the antitumor effect of rTNF-alpha was significantly decreased by pretreatment with small doses or rTNF-alpha in mice. In in vitro experiments, incubation of human umbilical vein endothelial cells with rTNF-alpha enhanced procoagulant activity (PCA), which was drastically augmented after an addition of the conditioned medium of Meth A tumor cells. Furthermore, rTNF-alpha-induced PCA was decreased by pretreatment with rTNF-alpha in endothelial cells. This PCA was completely blocked after the addition of anti-human tissue factor (TF) murine monoclonal antibody. These results imply that in vivo antitumor effects of rTNF-alpha are mediated by expression of TF in endothelial cells, which is augmented by tumor released factor(s).

  15. Reduced tumor necrosis factor-alpha and transforming growth factor-beta1 expression in the lungs of inbred mice that fail to develop fibroproliferative lesions consequent to asbestos exposure.

    PubMed

    Brass, D M; Hoyle, G W; Poovey, H G; Liu, J Y; Brody, A R

    1999-03-01

    Tumor necrosis factor (TNF)-alpha and transforming growth factor (TGF)-beta mRNA and protein expression and the degree of fibroproliferative response to inhaled asbestos fibers are clearly reduced in the 129 inbred mouse strain as compared with typical fibrogenesis observed in the C57BL/6 inbred strain. The C57BL/6 mice showed prominent lesions at bronchiolar-alveolar duct (BAD) junctions where asbestos fibers deposit and responding macrophages accumulate. The 129 mice, however, were generally indistinguishable from controls even though the numbers of asbestos fibers deposited in the lungs of all exposed animals were the same. Quantitative morphometry of H&E-stained lung sections comparing the C57BL/6 and 129 mice showed significantly less mean cross-sectional area of the BAD junctions in the 129 animals, apparent at both 48 hours and 4 weeks after exposure. In addition, fewer macrophages had accumulated at these sites in the 129 mice. Nuclear bromodeoxyuridine immunostaining demonstrated that the number of proliferating cells at first alveolar duct bifurcations and in adjacent terminal bronchioles was significantly reduced in the 129 strain compared with C57BL/6 mice at 48 hours after exposure (P < 0.01). TNF-alpha and TGF-beta1 gene expression, as measured by in situ hybridization, was reduced in the 129 mice at 48 hours after exposure, and expression of TNF-alpha and TGF-beta1 protein, as measured by immunohistochemistry, was similarly reduced or absent in the 129 animals. We postulate that the protection afforded the 129 mice is related to reduction of growth factor expression by the bronchiolar-alveolar epithelium and lung macrophages.

  16. Expression of messenger ribonucleic acid and presence of immunoreactive proteins for epidermal growth factor (EGF), transforming growth factor alpha (TGF alpha) and EGF/TGF alpha receptors and 125I-EGF binding sites in human fallopian tube.

    PubMed

    Chegini, N; Zhao, Y; McLean, F W

    1994-05-01

    Reverse transcription polymerase chain reaction (RT-PCR) revealed that the Fallopian tubes express epidermal growth factor (EGF), transforming growth factor (TGF alpha), and EGF receptor (EGF-R) mRNA. The RT-PCR product was verified by restriction enzyme digestion analysis. Immunohistochemically, EGF, TGF alpha, and EGF-R were localized in Fallopian tubes by use of specific antibodies to human EGF, mature fragments of human TGF alpha, and monoclonal antibodies to the extracellular binding domain of EGF-R. The tubal epithelial cells were the primary site of immunoreactive EGF, TGF alpha, and EGF-R, which were present to a lesser extent in the stromal cells, smooth muscle cell layers, fibroblasts of serosal tissue, and arterial endothelial and smooth muscle cells. Using antibodies generated against the amino and carboxy termini of TGF alpha precursor produced a similar cellular distribution to that observed for mature TGF alpha. The intensity of immunoreactive TGF alpha with these antibodies was similar to that seen with EGF. The ciliated and nonciliated epithelial cells in the ampullary and isthmus regions immunostained with similar intensity for EGF, TGF alpha, and EGF-R. The immunostaining for EGF, TGF alpha, and EGF-R was cycle-dependent, was considerably higher during late proliferative and early-to-mid-secretory phases than during early proliferative and late secretory phases of the menstrual cycle, and was reduced during the postmenopausal period. Specimens obtained 5-12 yr after tubal ligation immunostained for EGF, TGF alpha, and EGF-R similarly to sections from unligated tubes taken during the same phase of the cycle. Quantitative autoradiography of 125I-EGF binding generated a pattern similar to that of immunostaining for EGF-R binding. Net grain density/100 microns 2 calculated for different cell types indicated that the epithelial cells had a significantly higher grain density than did other tubal cell types (p < 0.05) without the cycle dependency seen

  17. Expression of inducible nitric oxide in human lung epithelial cells.

    PubMed

    Robbins, R A; Barnes, P J; Springall, D R; Warren, J B; Kwon, O J; Buttery, L D; Wilson, A J; Geller, D A; Polak, J M

    1994-08-30

    Nitric oxide (NO) is increased in the exhaled air of subjects with several airway disorders. To determine if cytokines could stimulate epithelial cells accounting for the increased NO, the capacity of the proinflammatory cytokines (cytomix: tumor necrosis factor-alpha, interleukin-1 beta, and interferon-gamma) to increase inducible nitric oxide synthase (iNOS) was investigated in A549 and primary cultures of human bronchial epithelial cells. Cytomix induced a time-dependent increase in nitrite levels in culture supernatant fluids (p < 0.05). Increased numbers of cells stained for iNOS and increased iNOS mRNA was detected in the cytokine-stimulated cells compared to control (p < 0.05). Dexamethasone diminished the cytokine-induced increase in nitrite, iNOS by immunocytochemistry, and iNOS mRNA. These data demonstrate that cytokines, such as those released by mononuclear cells, can induce lung epithelial iNOS expression and NO release, and that this is attenuated by dexamethasone.

  18. Enhanced effects of combined bu-zhong-yi-qi-tang (TJ-41) and interleukin-18 on the production of tumour necrosis factor-alpha and interferon-gamma in human peripheral blood mononuclear cells.

    PubMed

    Tamura, R; Takahashi, H K; Xue, D; Kubo, S; Saito, S; Nishibori, M; Iwagaki, H; Tanaka, N

    2004-01-01

    Co-stimulatory molecules play important roles in immune responses. We investigated the effect of Bu-Zhong-Yi-Qi-Tang (TJ-41) on the expression of intercellular adhesion molecule-1 (ICAM-1), B7.1 and B7.2 by peripheral blood mononuclear cells stimulated by interleukin-18 (IL-18) using fluorescence-activated cell sorter analysis. TJ-41 increased IL-18-induced ICAM-1 and B7.2 expression, resulting in enhanced production of tumour necrosis factor-alpha and interferon-gamma. These results suggest that TJ-41 enhances IL-18-induced cell-mediated immunity and may enhance host defence mechanisms against pathogens.

  19. Localization of tumour necrosis factor-alpha (TNF-alpha) and its receptors in normal and psoriatic skin: epidermal cells express the 55-kD but not the 75-kD TNF receptor.

    PubMed Central

    Kristensen, M; Chu, C Q; Eedy, D J; Feldmann, M; Brennan, F M; Breathnach, S M

    1993-01-01

    The distribution of TNF-alpha, p55 TNF receptor (TNF-R) and p75 TNF-R in normal skin and uninvolved and lesional skin from psoriasis patients has been investigated, using specific mono- and polyclonal antibodies. In normal skin, and uninvolved and lesional skin from psoriasis patients, p55 TNF-R is associated with epidermal keratinocytes and a network of upper dermal dendritic cells. This suggests that the actions of TNF-alpha on epidermal cells in vivo are mediated by binding to the p55 TNF-R. In lesional psoriasis skin, there was staining of the parakeratotic stratum corneum and increased expression of p55 TNF-R in association with upper dermal blood vessels. Staining for p75 TNF-R in normal skin was restricted to eccrine sweat ducts and dermal dendritic cells, and was absent from the epidermis. In lesional psoriasis skin, there was staining for p75 TNF-R in association with upper dermal blood vessels and perivascular infiltrating cells. TNF-alpha in normal skin was predominantly localized to the basal cell layers of the epidermis, and was seen in association with eccrine ducts and sebaceous glands. In lesional psoriasis skin, and to a lesser extent in uninvolved psoriasis skin, TNF-alpha was distributed throughout the epidermis, and was also specifically localized to upper dermal blood vessels. Up-regulation of TNF-alpha, p55 TNF-R and p75 TNF-R on dermal blood vessels in psoriasis may play an important role in the pathogenesis of this condition by promoting cutaneous recruitment of inflammatory cells. Images Fig. 4 Fig. 5 Fig. 1 Fig. 2 Fig. 3 Fig. 6 Fig. 7 Fig. 8 PMID:8222328

  20. Phenol induced acute cutaneous inflammation (AI) in mice: Diminished response in mast cell-deficient (W/W sup v ) mice and evidence of a role for tumor necrosis factor-alpha (TNF)

    SciTech Connect

    Wershil, B.K.; Wang, Z.S.; Gordon, J.R.; Galli, S.J. Harvard Medical School, Boston, MA )

    1991-03-11

    AI can be induced by a variety of chemical agents. The authors examined AI in mast cell-deficient (WBB6F{sub 1}-W/W{sup v}) and congenic normal (WBB6F{sub 1}-+/+) mice; AI was induced by the epicutaneous application to the ear of phenol (2 mg), benzalkonium chloride (BC; 1 mg) and ethyl phenylpropiolate (EPP, 2 or 5 mg). Phenol induced significantly greater swelling in +/+ than in W/W{sup v} mice. No difference in swelling was seen in +/+ versus W/W{sup v} mice with BC or EEP. Phenol application induced significantly greater neutrophil infiltration in +/+ than in W/W{sup v} mice. Mast cells represent a rich source of TNF and TNF has been shown to participate in the neutrophil accumulation seen in mast cell-dependent, IgE-mediated cutaneous late phase reactions. The authors injected +/+ mice i.d. with 20 {mu}l of 1:100 dilution of a polyclonal rabbit anti-mouse TNF antiserum or 20 {mu}l of medium and then applied 2 mg phenol at the same sites. At 24 hrs, significantly less neutrophil accumulation was seen in the ear treated with anti-TNF antibodies than in the control ear. The authors conclude that mast cells may participate in phenol-induced AI, and that TNF contributes to this response.

  1. Interferon-gamma and transforming growth factor-beta modulate the activation of mitogen-activated protein kinases and tumor necrosis factor-alpha production induced by Fc gamma-receptor stimulation in murine macrophages.

    PubMed

    Rose, D M; Winston, B W; Chan, E D; Riches, D W; Henson, P M

    1997-09-08

    Engagement of receptors for the Fc region of IgG (Fc gamma R) can activate a variety of biological responses in macrophages, and these responses can be modulated either positively or negatively by co-stimulation with a variety of agents including cytokines such as interferon-gamma (IFN-gamma) and transforming growth factor-beta (TGF-beta). We have previously demonstrated that Fc gamma R crosslinking activates the mitogen-activated protein kinase (MAPK) family members p42MAPK, p38, and JNK. Herein, we examined the modulatory effect of IFN-gamma, TGF-beta, and platelet-activating factor (PAF) on Fc gamma R-induced MAPK activation in murine macrophages. Fc gamma R-induced activation of p42MAPK and JNK was augmented nearly two-fold by pretreatment with IFN-gamma. Conversely, TGF-beta pretreatment suppressed Fc gamma R-induced activation of p42MAPK, JNK, and p38. These modulatory effects of IFN-gamma and TGF-beta on MAPK activation correlated with changes in Fc gamma R-stimulated TNF-alpha production by these two cytokines.

  2. Key roles for GRB2-associated-binding protein 1, phosphatidylinositol-3-kinase, cyclooxygenase 2, prostaglandin E2 and transforming growth factor alpha in linoleic acid-induced upregulation of lung and breast cancer cell growth

    PubMed Central

    Johnson, E.D.; Beck, K.L.; Pardini, R.S.

    2014-01-01

    SUMMARY The omega-6 polyunsaturated fatty acid linoleic acid (LA; C18:2 n-6) is prevalent in Western diets and has been shown to enhance tumorigenesis of several cancer models. However, the modes by which LA affects carcinogenesis have not been fully elucidated. In this study, a mechanism for LA-induced upregulation of cancer cell growth is defined. Cellular proliferation was enhanced with LA treatment in BT-474 human breast ductal carcinoma and A549 human lung adenocarcinoma cell lines. Enrichment of LA increased COX activity and led to increases in PGE2 production, followed by increases in MMP and TGF-α levels, which are all key elements involved in the enhancement of cancer cell growth. Further investigation revealed that LA supplementation in both BT-474 breast and A549 lung cancer cell lines greatly increased the association between the scaffolding protein Gab1 and EGFR, while at the same time dramatically decreasing Gab1 protein levels. These changes are concomitant with increases in activated Akt (pAkt), a downstream signaling component in the PI3K signaling pathway. Moreover, inhibitors of EGFR, PI3K and Gab1-specific siRNAs were capable of reversing LA-induced upregulation of pAkt, as well as observed increases in cell proliferation for these models. These data establish Gab1 as major target in LA-induced enhancement of tumorigenesis. PMID:24374147

  3. Role for Tumor Necrosis Factor Alpha in Murine Cytomegalovirus Transcriptional Reactivation in Latently Infected Lungs

    PubMed Central

    Simon, Christian O.; Seckert, Christof K.; Dreis, Doris; Reddehase, Matthias J.; Grzimek, Natascha K. A.

    2005-01-01

    Interstitial pneumonia is a major clinical manifestation of primary or recurrent cytomegalovirus (CMV) infection in immunocompromised recipients of a bone marrow transplant. In a murine model, lungs were identified as a prominent site of CMV latency and recurrence. Pulmonary latency of murine CMV is characterized by high viral genome burden and a low incidence of variegated immediate-early (IE) gene expression, reflecting a sporadic activity of the major IE promoters (MIEPs) and enhancer. The enhancer-flanking promoters MIEP1/3 and MIEP2 are switched on and off during latency in a ratio of ∼2:1. MIEP1/3 latency-associated activity generates the IE1 transcript of the ie1/3 transcription unit but not the alternative splicing product IE3 that encodes the essential transactivator of early gene expression. Splicing thus appeared to be an important checkpoint for maintenance of latency. In accordance with previous work of others, we show here that signaling by the proinflammatory cytokine tumor necrosis factor alpha (TNF-α) activates IE1/3 transcription in vivo. As an addition to current knowledge, Poisson distribution analysis revealed an increased incidence of IE1/3 transcriptional events as well as a higher amount of transcripts per event. Notably, TNF-α promoted the splicing to IE3 transcripts, but transcription did not proceed to the M55/gB early gene. Moreover, the activated transcriptional state induced by TNF-α did not predispose latently infected mice to a higher incidence of virus recurrence after hematoablative treatment. In conclusion, TNF-α is an important inductor of IE gene transcriptional reactivation, whereas early genes downstream in the viral replicative cycle appear to be the rate-limiting checkpoint(s) for virus recurrence. PMID:15596827

  4. Effects of tumor necrosis factor alpha on host immune response in chronic persistent tuberculosis: possible role for limiting pathology.

    PubMed

    Mohan, V P; Scanga, C A; Yu, K; Scott, H M; Tanaka, K E; Tsang, E; Tsai, M M; Flynn, J L; Chan, J

    2001-03-01

    Reactivation of latent tuberculosis contributes significantly to the incidence of disease caused by Mycobacterium tuberculosis. The mechanisms involved in the containment of latent tuberculosis are poorly understood. Using the low-dose model of persistent murine tuberculosis in conjunction with MP6-XT22, a monoclonal antibody that functionally neutralizes tumor necrosis factor alpha (TNF-alpha), we examined the effects of TNF-alpha on the immunological response of the host in both persistent and reactivated tuberculous infections. The results confirm an essential role for TNF-alpha in the containment of persistent tuberculosis. TNF-alpha neutralization resulted in fatal reactivation of persistent tuberculosis characterized by a moderately increased tissue bacillary burden and severe pulmonic histopathological deterioration that was associated with changes indicative of squamous metaplasia and fluid accumulation in the alveolar space. Analysis of pulmonic gene and protein expression of mice in the low-dose model revealed that nitric oxide synthase was attenuated during MP6-XT22-induced reactivation, but was not totally suppressed. Interleukin-12p40 and gamma interferon gene expression in TNF-alpha-neutralized mice was similar to that in control mice. In contrast, interleukin-10 expression was augmented in the TNF-alpha-neutralized mice. In summary, results of this study suggest that TNF-alpha plays an essential role in preventing reactivation of persistent tuberculosis, modulates the pulmonic expression of specific immunologic factors, and limits the pathological response of the host.

  5. Transforming growth factor alpha promotes osteosarcoma metastasis by ICAM-1 and PI3K/Akt signaling pathway.

    PubMed

    Hou, Chun-Han; Lin, Feng-Ling; Tong, Kai-Biao; Hou, Sheng-Mon; Liu, Ju-Fang

    2014-06-15

    Osteosarcoma is the most common primary malignancy of bone and is characterized by a high malignant and metastatic potential. Transforming growth factor alpha (TGF-α) is classified as the EGF (epidermal growth factor)-like family, which is involved in cancer cellular activities such as proliferation, motility, migration, adhesion and invasion abilities. However, the effect of TGF-α on human osteosarcoma is largely unknown. We found that TGF-α increased the cell migration and expression of intercellular adhesion molecule-1 (ICAM-1) in human osteosarcoma cells. Transfection of cells with ICAM-1 siRNA reduced TGF-α-mediated cell migration. We also found that the phosphatidylinositol 3'-kinase (PI3K)/Akt/NF-κB pathway was activated after TGF-α treatment, and TGF-α-induced expression of ICAM-1 and cell migration was inhibited by the specific inhibitors and siRNAs of PI3K, Akt, and NF-κB cascades. In addition, knockdown of TGF-α expression markedly decreased cell metastasis in vitro and in vivo. Our results indicate that TGF-α/EGFR interaction elicits PI3K and Akt activation, which in turn activates NF-κB, resulting in the expression of ICAM-1 and contributing the migration of human osteosarcoma cells.

  6. Tumor necrosis factor-alpha regulation of the Id gene family in astrocytes and microglia during CNS inflammatory injury.

    PubMed

    Tzeng, S F; Kahn, M; Liva, S; De Vellis, J

    1999-04-01

    The inhibitors of DNA binding (Id) gene family is highly expressed during embryogenesis and throughout adulthood in the rat central nervous system (CNS). In vitro studies suggest that the Id gene family is involved in the regulation of cell proliferation and differentiation. Recently, Id gene expression was shown to be expressed in immature and mature astrocytes during development and upregulated in reactive astrocytes after spinal cord injury. These results suggest that the Id gene family may play an important role in regulating astrocyte development and reactivity; however, the factors regulating Id expression in astrocytes remain undefined. Tumor necrosis factor-alpha (TNF alpha), a proinflammatory cytokine, is thought to play a crucial role in astrocyte/microglia activation after injury to the CNS. To determine if TNF alpha plays a role in Id gene expression, we exogenously administered TNF alpha into developing postnatal rats. We report that TNF alpha injections resulted in a rapid and transient increase in both cell number and mRNA expression for Id2 and Id3 when compared to levels observed in noninjected or control-injected animals. Id1 mRNA levels were also upregulated after TNF alpha treatment, but to a lesser degree. Significant increases in TNF alpha-induced Id2 and Id3 mRNA were observed in the ventricular/subventricular zone, cingulum and corpus callosum. TNF alpha also increased Id2 mRNA expression in the caudate putamen and hippocampus at the injection site. Id2 and Id3 mRNA+ cells were identified as GFAP+ and S100 alpha + astrocytes as well as ED1+ microglia. This is the first report to show TNF-alpha-induced modulation of the Id gene family and suggests that Id may be involved in the formation of reactive astrocytes and activated microglia in the rodent brain. These results suggest a putative role for the Id family in the molecular mechanisms regulating cellular responsiveness to TNF alpha and CNS inflammation.

  7. Raised serum levels of cachectin/tumor necrosis factor alpha in renal allograft rejection

    PubMed Central

    1987-01-01

    A sensitive radioimmunoassay was used for monitoring serum levels of endogenous cachectin/tumor necrosis factor alpha (TNF) in 10 renal transplant recipients. Acute allograft rejections were associated with marked elevations of circulating TNF. The peak levels of TNF (median 140 pg/ml) were in the same concentration range as previously reported in parasitic infections. The results show that the release of TNF into circulation is an early event in renal allograft rejection and that raised levels of TNF in man can also be induced by noninfectious stimuli. PMID:3309124

  8. Virus-cell interactions regulating induction of tumor necrosis factor alpha production in macrophages infected with herpes simplex virus.

    PubMed

    Paludan, S R; Mogensen, S C

    2001-11-01

    Macrophages respond to virus infections by rapidly secreting proinflammatory cytokines, which play an important role in the first line of defense. Tumor necrosis factor alpha (TNF-alpha) is one of the major macrophage-produced cytokines. In this study we have investigated the virus-cell interactions responsible for induction of TNF-alpha expression in herpes simplex virus (HSV)-infected macrophages. Both HSV type 1 (HSV-1) and HSV-2 induced TNF-alpha expression in macrophages activated with gamma interferon (IFN-gamma). This induction was to some extent sensitive to UV treatment of the virus. Virus particles unable to enter the cells displayed reduced capacity to stimulate TNF-alpha expression but retained a significant portion which was abolished by HSV-specific antibodies. Recombinant HSV-1 glycoprotein D was able to trigger TNF-alpha secretion in concert with IFN-gamma. Sugar moieties of HSV glycoproteins have been reported to be involved in induction of IFN-alpha but did not contribute to TNF-alpha expression in macrophages. Moreover, the entry-dependent portion of the TNF-alpha induction was investigated with HSV-1 mutants and found to be independent of the tegument proteins VP16 and UL13 and partly dependent on nuclear translocation of the viral DNA. Finally, we found that macrophages expressing an inactive mutant of the double-stranded RNA (dsRNA)-activated protein kinase (PKR) produced less TNF-alpha in response to infectious HSV infection than the empty-vector control cell line but displayed the same responsiveness to UV-inactivated virus. These results indicate that HSV induces TNF-alpha expression in macrophages through mechanisms involving (i) viral glycoproteins, (ii) early postentry events occurring prior to nuclear translocation of viral DNA, and (iii) viral dsRNA-PKR.

  9. Tumor necrosis factor-alpha induced enhancement of cryosurgery

    NASA Astrophysics Data System (ADS)

    Goel, Raghav; Paciotti, Guilio F.; Bischof, John C.

    2008-02-01

    Local recurrence of cancer after cryosurgery is related to the inability to monitor and predict destruction of cancer (temperatures > -40°C) within an iceball. We previously reported that a cytokine adjuvant TNF-α could be used to achieve complete cancer destruction at the periphery of an iceball (0 to -40°C). This study is a further development of that work in which cryosurgery was performed using cryoprobes operating at temperatures > -40°C. LNCaP Pro 5 tumor grown in a dorsal skin fold chamber (DSFC) was frozen at -6°C after TNF-α incubation for 4 or 24 hours. Tumors grown in the hind limb were frozen with a probe tip temperature of -40°C, 4 or 24 hours after systemic injection with TNF-α. Both cryosurgery alone or TNF-α treatment alone caused only a minimal damage to the tumor tissue at the conditions used in the study. The combination of TNF-α and cryosurgery produced a significant damage to the tumor tissue in both the DSFC and the hind limb model system. This augmentation in cryoinjury was found to be time-dependent with 4-hour time period between the two treatments being more effective than 24-hour. These results suggests the possibility of cryotreatment at temperatures > -40°C with the administration of TNF-α.

  10. Immediate anti-tumor necrosis factor-alpha (etanercept) therapy enhances axonal regeneration after sciatic nerve crush.

    PubMed

    Kato, Kinshi; Liu, Huaqing; Kikuchi, Shin-ichi; Myers, Robert R; Shubayev, Veronica I

    2010-02-01

    Peripheral nerve regeneration begins immediately after injury. Understanding the mechanisms by which early modulators of axonal degeneration regulate neurite outgrowth may affect the development of new strategies to promote nerve repair. Tumor necrosis factor-alpha (TNF-alpha) plays a crucial role in the initiation of degenerative cascades after peripheral nerve injury. Here we demonstrate using real-time Taqman quantitative RT-PCR that, during the time course (days 1-60) of sciatic nerve crush, TNF-alpha mRNA expression is induced at 1 day and returned to baseline at 5 days after injury in nerve and the corresponding dorsal root ganglia (DRG). Immediate therapy with the TNF-alpha antagonist etanercept (fusion protein of TNFRII and human IgG), administered systemically (i.p.) and locally (epineurially) after nerve crush injury, enhanced the rate of axonal regeneration, as determined by nerve pinch test and increased number of characteristic clusters of regenerating nerve fibers distal to nerve crush segments. These fibers were immunoreactive for growth associated protein-43 (GAP-43) and etanercept, detected by anti-human IgG immunofluorescence. Increased GAP-43 expression was found in the injured nerve and in the corresponding DRG and ventral spinal cord after systemic etanercept compared with vehicle treatments. This study established that immediate therapy with TNF-alpha antagonist supports axonal regeneration after peripheral nerve injury.

  11. Effects of leptin and tumor necrosis factor-alpha on degranulation and superoxide production of polymorphonuclear neutrophils from Holstein cows.

    PubMed

    Ahmed, Mohamed; Kimura, Kazuhiro; Soliman, Mohamed; Yamaji, Daisuke; Okamatsu-Ogura, Yuko; Makondo, Kennedy; Inanami, Osamu; Saito, Masayuki

    2007-02-01

    Leptin, a pleiotropic hormone regulating food intake and energy expenditure, has been shown to directly modulate human polymorphonuclear neutrophil (PMN) functions or indirectly through the action of tumor necrosis factor-alpha (TNF-alpha). Bovine PMN have considerable different characteristics from human PMN. For example, it does not respond to N-formyl-Methionyl-Leucyl-phenylalanine, a well known human PMN activator. In the present study, we tested the effects of leptin and TNF-alpha on superoxide production and degranulation of bovine peripheral PMN, in which both long isoform of leptin receptor (Ob-Rb) and TNF receptor 1 were expressed. Human leptin, human TNF-alpha, phorbol myristate acetate (PMA) and opsonized zymosan particles (OZP) did not stimulate degranulation responses, while zymosan-activated serum (ZAS) did. Neither leptin nor TNF-alpha enhanced the ZAS-induced degranulation responses. TNF-alpha, PMA, OZP and ZAS increased superoxide production in different magnitudes, whereas leptin did not. TNF-alpha, but not leptin, enhanced OZP- and ZAS-induced superoxide production, possibly, in part due to facilitating translocation of p47(phox), a component of NADPH oxidase. These results indicate that, unlike in human PMN, leptin does not have any direct effect on degranulation and superoxide production in bovine PMN, although TNF-alpha influences superoxide production.

  12. The role of tumour necrosis factor alpha and soluble tumour necrosis factor alpha receptors in the symptomatology of schizophrenia.

    PubMed

    Turhan, Levent; Batmaz, Sedat; Kocbiyik, Sibel; Soygur, Arif Haldun

    2016-07-01

    Background Immunological mechanisms may be responsible for the development and maintenance of schizophrenia symptoms. Aim The aim of this study is to measure tumour necrosis factor-alpha (TNF-α), soluble tumour necrosis factor-alpha receptor I (sTNF-αRI), and soluble tumour necrosis factor-alpha receptor II (sTNF-αRII) levels in patients with schizophrenia and healthy individuals, and to determine their relationship with the symptoms of schizophrenia. Methods Serum TNF-α, sTNF-αRI and sTNF-αRII levels were measured. The Positive and Negative Syndrome Scale (PANSS) was administered for patients with schizophrenia (n = 35), and the results were compared with healthy controls (n = 30). Hierarchical regression analyses were undertaken to predict the levels of TNF-α, sTNF-αRI and sTNF-αRII. Results No significant difference was observed in TNF-α levels, but sTNF-αRI and sTNF-αRII levels were lower in patients with schizophrenia. Serum sTNF-αRI and sTNF-αRII levels were found to be negatively correlated with the negative subscale score of the PANSS, and sTNF-αRI levels were also negatively correlated with the total score of the PANSS. Smoking, gender, body mass index were not correlated with TNF-α and sTNF-α receptor levels. Conclusions These results suggest that there may be a change in anti-inflammatory response in patients with schizophrenia due to sTNF-αRI and sTNF-αRII levels. The study also supports low levels of TNF activity in schizophrenia patients with negative symptoms.

  13. The reduction of tumor necrosis factor-alpha release and tissue damage by pentobarbital in the experimental endotoxemia model.

    PubMed

    Yang, Fwu Lin; Li, Chi Han; Hsu, Bang Gee; Tsai, Nu-Man; Lin, Shinn Zong; Harn, Horng Jyh; Chen, Hsing I; Liao, Kuang Wen; Lee, Ru Ping

    2007-09-01

    Sepsis is the leading cause of death for intensive care patients. Lipopolysaccharide (LPS) administration to animals under anesthesia is a strategy for the study of uncontrolled release of proinflammatory cytokines. Anesthetics have been indicated that they can specially affect immune responses, such as the inflammatory response. Pentobarbital is an anesthetic used mainly in animal studies. Thus, the effect of pentobarbital on tumor necrosis factor-alpha (TNF-alpha) release was determined. The results revealed that pentobarbital suppressed the expression of TNF-alpha mRNA and its proteins, which may result from the decrease in the activities of nuclear factor-kappaB and activator protein 1 and the reduction of the expression of p38 mitogen-activated protein kinase by pentobarbital. After the inhibitory activity of the pentobarbital for TNF-alpha release was proven in vivo, the cytotoxic effects of LPS were examined in vivo with or without pentobarbital treatments. In vivo results indicated that plasma levels of alanine aminotransferase, aspartate aminotransferase, lactic dehydrogenase, creatine kinase, serum urea nitrogen, and amylase decreased dramatically in the anesthetic group with pentobarbital administration. Finally, the effect of pentobarbital on TNF-alpha-related cell death was monitored in vitro, and the results indicated the pentobarbital could directly enhance the viabilities of cells under the treatment of TNF-alpha and protected cells from apoptosis induced by deferoxamine mesylate-induced hypoxia. These results suggest that pentobarbital significantly influences the LPS-induced inflammatory response and protects cells from death directly and indirectly induced by TNF-alpha. The information provides a perspective to re-evaluate the results of the experiments in which animals were anesthetized with pentobarbital. The anti-inflammatory effects of the drugs may have been caused by the synergistic effect of pentobarbital.

  14. CT-2576, an inhibitor of phospholipid signaling, suppresses constitutive and induced expression of human immunodeficiency virus.

    PubMed Central

    Leung, D W; Peterson, P K; Weeks, R; Gekker, G; Chao, C C; Kaplan, A H; Balantac, N; Tompkins, C; Underiner, G E; Bursten, S

    1995-01-01

    Viruses such as human immunodeficiency virus (HIV) require cellular activation for expression. Cellular activation in lymphoid cells is associated with augmented accumulation of certain phosphatidic acid (PA) species derived from the hydrolysis of glycan phosphatidylinositol (GPI). This suggests that activation of a phospholipid pathway may play a role in initiation of viral replication. To test this hypothesis, we examined the effect of tat gene expression on the production of cellular PA species, as the Tat protein is essential for HIV expression and has been implicated in activating the expression of multiple host cellular genes. Expression of tat increased the expression of PA. We then tested whether synthetic inhibitors of PA metabolism would inhibit activation of the HIV long terminal repeat by Tat and tumor necrosis factor alpha (TNF-alpha). CT-2576 suppressed both PA generation induced by Tat and HIV long terminal repeat-directed gene expression in response to Tat or TNF-alpha at a posttranscriptional step. CT-2576 also inhibited constitutive as well as TNF-alpha- and interleukin 6-induced expression of HIV p24 antigen in chronically infected U1 cells and in peripheral blood lymphocytes acutely infected with a clinical isolate of HIV. Pharmacological inhibition of synthesis of selected PA species may therefore provide a therapeutic approach to suppression of HIV replication. Images Fig. 1 Fig. 3 Fig. 5 PMID:7761405

  15. Altered pattern of tumor necrosis factor-alpha production in peripheral blood monocytes from Crohn's disease

    PubMed Central

    Loganes, Claudia; Pin, Alessia; Naviglio, Samuele; Girardelli, Martina; Bianco, Anna Monica; Martelossi, Stefano; Tommasini, Alberto; Piscianz, Elisa

    2016-01-01

    AIM To evaluate the inflammatory state in Crohn’s disease (CD) patients and correlate it with genetic background and microbial spreading. METHODS By means of flow cytometry, production of tumor necrosis factor-alpha (TNF-α) was measured in peripheral blood monocytes from patients suffering from CD, ulcerative colitis (UC) and in healthy subjects after stimulation of the NOD2 and TLR pathways. CD patients were genotyped for the three most common NOD2 variants (R702W, G908R and L1007Pfs*2) and basal production of TNF-α was correlated to NOD2 genotype. Also, production of TNF-α was correlated to plasmatic levels of LPS Binding Protein (LBP), soluble (s) CD14 and to the activity state of the disease. RESULTS The patients with CD were characterized by a significantly higher monocyte basal expression of TNF-α compared with healthy subjects and UC patients, and after stimulation with Pam3CSK4 (ligand of TLR2/1) and MDP-L18 (ligand of NOD2) this difference was maintained, while other microbial stimuli (LPS, ligand of TLR4 and PolyI:C, ligand of TLR3) induced massive activation in CD monocytes as well as in UC and in healthy control cells. There was no significant difference in the production of TNF-α between patients who carried CD-associated heterozygous or homozygous variants in NOD2 and patients with wild type NOD2 genotype. Although serum LBP levels have been shown to correlate positively with the state of activity of the disease, TNF-α production did not show a clear correlation with either LBP or sCD14 levels in plasma. Moreover, no clear correlation was seen between TNF-α production and activity indices in either CD or UC. CONCLUSION Peripheral monocytes from CD express higher basal and stimulated TNF-α than controls, regardless of NOD2 genotype and without a clear correlation with disease activity. PMID:27895399

  16. The development of novel inhibitors of tumor necrosis factor-alpha production based on substituted [5,5]-bicyclic pyrozolones

    SciTech Connect

    Laufersweiler, Matthew; Brugel, Todd; Clark, Michael; Golebiowski, Adam; Bookland, Roger; Laughlin, Steven; Sabat, Mark; Townes, Jennifer; VanRens, John; De, Biswanath; Hsieh, Lily; Heitmeyer, Sandra; Juergens, Karen; Brown, Kimberly; Mekel, Marlene; Walter, Richard; Janusz, Michael

    2010-11-16

    Novel substituted [5,5]-bicyclic pyrzazolones are presented as inhibitors of tumor necrosis factor-{alpha} (TNF-{alpha}) production. Many of these compounds show low nanomolar activity against lipopolysaccaride (LPS)-induced TNF-{alpha} production in THP-1 cells. This class of molecules was co-crystallized with mutated p38, and several analogs showed good oral bioavailability in the rat. Oral activity of these compounds in the rat iodoacetate model for osteoarthritis is discussed.

  17. Retinoic acid inhibits inducible nitric oxide synthase expression in 3T3-L1 adipocytes.

    PubMed

    Yang, Jeong-Yeh; Koo, Bon-Sun; Kang, Mi-Kyung; Rho, Hye-Won; Sohn, Hee-Sook; Jhee, Eun-Chung; Park, Jin-Woo

    2002-11-30

    The present study was undertaken to explore whether retinoids, which are known to have immunomodulatory actions, could attenuate tumor necrosis factor-alpha (TNF)-stimulated inducible nitric oxide synthase (iNOS) expression in 3T3-L1 adipocytes. Adipocytes incubated with TNF induced dose- and time-dependent accumulation of nitrite in the culture medium through the iNOS induction as confirmed by Western blotting. Treatment of cells with TNF in the presence of all-trans-retinoic acid (RA) significantly decreased their ability to produce nitrite and iNOS induction. Both 13-cis- and all- trans-RA-induced suppression was dose-dependent, and all-trans-RA was somewhat potent than 13-cis-RA. The inhibitory effect of RA on TNF-induced iNOS induction was reversible, completely recovered after 2 days, and was exerted through the inhibition of NF-kappaB activation. TNF also suppressed the lipoprotein lipase (LPL) activity of 3T3-L1 adipocytes. RA could not reverse the TNF- induced LPL suppression at RA levels causing near complete inhibition of the TNF-induced NO production. These results indicate that RAs attenuate iNOS expression reversibly in TNF-stimulated 3T3-L1 adipocytes, and that the TNF-induced LPL suppression is not the result of NO overproduction.

  18. Tumor necrosis factor alpha-mediated inhibition of melanogenesis is dependent on nuclear factor kappa B activation.

    PubMed

    Englaro, W; Bahadoran, P; Bertolotto, C; Buscà, R; Dérijard, B; Livolsi, A; Peyron, J F; Ortonne, J P; Ballotti, R

    1999-02-25

    Melanogenesis is a physiological process resulting in the synthesis of melanin pigments which play a crucial protective role against skin photocarcinogenesis. In vivo, solar ultraviolet light triggers the secretion of numerous keratinocyte-derived factors that are implicated in the regulation of melanogenesis. Among these, tumor necrosis factor alpha (TNFalpha), a cytokine implicated in the pro-inflammatory response, down-regulates pigment synthesis in vitro. In this report, we aimed to determine the molecular mechanisms by which this cytokine inhibits melanogenesis in B16 melanoma cells. First, we show that TNFalpha inhibits the activity and protein expression of tyrosinase which is the key enzyme of melanogenesis. Further, we demonstrate that this effect is subsequent to a down-regulation of the tyrosinase promoter activity in both basal and cAMP-induced melanogenesis. Finally, we present evidence indicating that the inhibitory effect of TNFalpha on melanogenesis is dependent on nuclear factor kappa B (NFkappaB) activation. Indeed, overexpression of this transcription factor in B16 cells is sufficient to inhibit tyrosinase promoter activity. Furthermore, a mutant of inhibitory kappa B (IkappaB), that prevents NFkappaB activation, is able to revert the effect of TNFalpha on the tyrosinase promoter activity. Taken together, our results clarify the mechanisms by which TNFalpha inhibits pigmentation and point out the key role of NFkappaB in the regulation of melanogenesis.

  19. Tumor necrosis factor alpha converting enzyme: an encouraging target for various inflammatory disorders.

    PubMed

    Bahia, Malkeet S; Silakari, Om

    2010-05-01

    Tumor necrosis factor alpha is one of the most common pro-inflammatory cytokines responsible for various inflammatory disorders. It plays an important role in the origin and progression of rheumatoid arthritis and also in other autoimmune disease conditions. Some anti-tumor necrosis factor alpha antibodies like Enbrel, Humira and Remicade have been successfully used in these disease conditions as antagonists of tumor necrosis factor alpha. Inhibition of generation of active form of tumor necrosis factor alpha is a promising therapy for various inflammatory disorders. Therefore, the inhibition of an enzyme (tumor necrosis factor alpha converting enzyme), which is responsible for processing inactive form of tumor necrosis factor alpha into its active soluble form, is an encouraging target. Many tumor necrosis factor alpha converting enzyme inhibitors have been the candidates of clinical trials but none of them have reached in to the market because of their broad spectrum inhibitory activity for other matrix metalloproteases. Selectivity of tumor necrosis factor alpha converting enzyme inhibition over matrix metalloproteases is of utmost importance. If selectivity is achieved successfully, side-effects can be over-ruled and this approach may become a novel therapy for treatment of rheumatoid arthritis and other inflammatory disorders. This cytokine not only plays a pivotal role in inflammatory conditions but also in some cancerous conditions. Thus, successful targeting of tumor necrosis factor alpha converting enzyme may result in multifunctional therapy.

  20. Effects of tumor necrosis factor-alpha (TNF alpha) in epidermal keratinocytes revealed using global transcriptional profiling.

    PubMed

    Banno, Tomohiro; Gazel, Alix; Blumenberg, Miroslav

    2004-07-30

    Identification of tumor necrosis factor-alpha (TNF alpha) as the key agent in inflammatory disorders, e.g. rheumatoid arthritis, Crohn's disease, and psoriasis, led to TNF alpha-targeting therapies, which, although avoiding many of the side-effects of previous drugs, nonetheless causes other side-effects, including secondary infections and cancer. By controlling gene expression, TNF alpha orchestrates the cutaneous responses to environmental damage and inflammation. To define TNF alpha action in epidermis, we compared the transcriptional profiles of normal human keratinocytes untreated and treated with TNF alpha for 1, 4, 24, and 48 h by using oligonucleotide microarrays. We found that TNF alpha regulates not only immune and inflammatory responses but also tissue remodeling, cell motility, cell cycle, and apoptosis. Specifically, TNF alpha regulates innate immunity and inflammation by inducing a characteristic large set of chemokines, including newly identified TNF alpha targets, that attract neutrophils, macrophages, and skin-specific memory T-cells. This implicates TNF alpha in the pathogenesis of psoriasis, fixed drug eruption, atopic and allergic contact dermatitis. TNF alpha promotes tissue repair by inducing basement membrane components and collagen-degrading proteases. Unexpectedly, TNF alpha induces actin cytoskeleton regulators and integrins, enhancing keratinocyte motility and attachment, effects not previously associated with TNF alpha. Also unanticipated was the influence of TNF alpha upon keratinocyte cell fate by regulating cell-cycle and apoptosis-associated genes. Therefore, TNF alpha initiates not only the initiation of inflammation and responses to injury, but also the subsequent epidermal repair. The results provide new insights into the harmful and beneficial TNF alpha effects and define the mechanisms and genes that achieve these outcomes, both of which are important for TNF alpha-targeted therapies.

  1. Cellular and molecular evidence for a role of tumor necrosis factor alpha in the ovulatory mechanism of trout

    PubMed Central

    2010-01-01

    Background The relevance of immune-endocrine interactions to the regulation of ovarian function in teleosts is virtually unexplored. As part of the innate immune response during infection, a number of cytokines such as tumor necrosis factor alpha (TNF alpha) and other immune factors, are produced and act on the reproductive system. However, TNF alpha is also an important physiological player in the ovulatory process in mammals. In the present study, we have examined for the first time the effects of TNF alpha in vitro in preovulatory ovarian follicles of a teleost fish, the brown trout (Salmo trutta). Methods To determine the in vivo regulation of TNF alpha expression in the ovary, preovulatory brook trout (Salvelinus fontinalis) were injected intraperitoneally with either saline or bacterial lipopolysaccharide (LPS). In control and recombinant trout TNF alpha (rtTNF alpha)-treated brown trout granulosa cells, we examined the percentage of apoptosis by flow cytometry analysis and cell viability by propidium iodide (PI) staining. Furthermore, we determined the in vitro effects of rtTNF alpha on follicle contraction and testosterone production in preovulatory brown trout ovarian follicles. In addition, we analyzed the gene expression profiles of control and rtTNF alpha-treated ovarian tissue by microarray and real-time PCR (qPCR) analyses. Results LPS administration in vivo causes a significant induction of the ovarian expression of TNF alpha. Treatment with rtTNF alpha induces granulosa cell apoptosis, decreases granulosa cell viability and stimulates the expression of genes known to be involved in the normal ovulatory process in trout. In addition, rtTNF alpha causes a significant increase in follicle contraction and testosterone production. Also, using a salmonid-specific microarray platform (SFA2.0 immunochip) we observed that rtTNF alpha induces the expression of genes known to be involved in inflammation, proteolysis and tissue remodeling. Furthermore, the

  2. Analysis of Subcellular RNA Fractions Revealed a Transcription-Independent Effect of Tumor Necrosis Factor Alpha on Splicing, Mediated by Spt5

    PubMed Central

    Diamant, Gil; Eisenbaum, Tal; Leshkowitz, Dena

    2016-01-01

    The proinflammatory cytokine tumor necrosis factor alpha (TNF-α) modulates the expression of many genes, primarily through activation of NF-κB. Here, we examined the global effects of the elongation factor Spt5 on nascent and mature mRNAs of TNF-α-induced cells using chromatin and cytosolic subcellular fractions. We identified several classes of TNF-α-induced genes controlled at the level of transcription, splicing, and chromatin retention. Spt5 was found to facilitate splicing and chromatin release in genes displaying high induction rates. Further analysis revealed striking effects of TNF-α on the splicing of 25% of expressed genes; the vast majority were not transcriptionally induced. Splicing enhancement of noninduced genes by TNF-α was transient and independent of NF-κB. Investigating the underlying basis, we found that Spt5 is required for the splicing facilitation of the noninduced genes. In line with this, Spt5 interacts with Sm core protein splicing factors. Furthermore, following TNF-α treatment, levels of RNA polymerase II (Pol II) but not Spt5 are reduced from the splicing-induced genes, suggesting that these genes become enriched with a Pol II-Spt5 form. Our findings revealed the Pol II-Spt5 complex as a highly competent coordinator of cotranscriptional splicing. PMID:26903558

  3. Analysis of Subcellular RNA Fractions Revealed a Transcription-Independent Effect of Tumor Necrosis Factor Alpha on Splicing, Mediated by Spt5.

    PubMed

    Diamant, Gil; Eisenbaum, Tal; Leshkowitz, Dena; Dikstein, Rivka

    2016-05-01

    The proinflammatory cytokine tumor necrosis factor alpha (TNF-α) modulates the expression of many genes, primarily through activation of NF-κB. Here, we examined the global effects of the elongation factor Spt5 on nascent and mature mRNAs of TNF-α-induced cells using chromatin and cytosolic subcellular fractions. We identified several classes of TNF-α-induced genes controlled at the level of transcription, splicing, and chromatin retention. Spt5 was found to facilitate splicing and chromatin release in genes displaying high induction rates. Further analysis revealed striking effects of TNF-α on the splicing of 25% of expressed genes; the vast majority were not transcriptionally induced. Splicing enhancement of noninduced genes by TNF-α was transient and independent of NF-κB. Investigating the underlying basis, we found that Spt5 is required for the splicing facilitation of the noninduced genes. In line with this, Spt5 interacts with Sm core protein splicing factors. Furthermore, following TNF-α treatment, levels of RNA polymerase II (Pol II) but not Spt5 are reduced from the splicing-induced genes, suggesting that these genes become enriched with a Pol II-Spt5 form. Our findings revealed the Pol II-Spt5 complex as a highly competent coordinator of cotranscriptional splicing.

  4. Role of Eosinophils and Tumor Necrosis Factor Alpha in Interleukin-25-Mediated Protection from Amebic Colitis

    PubMed Central

    Noor, Zannatun; Watanabe, Koji; Abhyankar, Mayuresh M.; Burgess, Stacey L.; Buonomo, Erica L.

    2017-01-01

    ABSTRACT The parasite Entamoeba histolytica is a cause of diarrhea in infants in low-income countries. Previously, it was shown that tumor necrosis factor alpha (TNF-α) production was associated with increased risk of E. histolytica diarrhea in children. Interleukin-25 (IL-25) is a cytokine that is produced by intestinal epithelial cells that has a role in maintenance of gut barrier function and inhibition of TNF-α production. IL-25 expression was decreased in humans and in the mouse model of amebic colitis. Repletion of IL-25 blocked E. histolytica infection and barrier disruption in mice, increased gut eosinophils, and suppressed colonic TNF-α. Depletion of eosinophils with anti-Siglec-F antibody prevented IL-25-mediated protection. In contrast, depletion of TNF-α resulted in resistance to amebic infection. We concluded that IL-25 provides protection from amebiasis, which is dependent upon intestinal eosinophils and suppression of TNF-α. PMID:28246365

  5. Stable Escherichia coli-Clostridium acetobutylicum shuttle vector for secretion of murine tumor necrosis factor alpha.

    PubMed

    Theys, J; Nuyts, S; Landuyt, W; Van Mellaert, L; Dillen, C; Böhringer, M; Dürre, P; Lambin, P; Anné, J

    1999-10-01

    Recombinant plasmids were constructed to secrete mouse tumor necrosis factor alpha (mTNF-alpha) from Clostridium acetobutylicum. The shuttle plasmids contained the clostridial endo-beta1, 4-glucanase (eglA) promoter and signal sequence that was fused in frame to the mTNF-alpha cDNA. The construction was first tested in Escherichia coli and then introduced in C. acetobutylicum DSM792 by electroporation. Controls confirmed the presence and stability of the recombinant plasmids in this organism. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis and an in vitro cytotoxic assay were used to monitor expression and secretion of mTNF-alpha during growth. Significant levels of biologically active mTNF-alpha were measured in both lysates and supernatants. The present report deals with investigations on the elaboration of a gene transfer system for cancer treatment using anaerobic bacteria.

  6. Etanercept attenuates TNBS-induced experimental colitis: role of TNF-α expression.

    PubMed

    Paiotti, Ana Paula Ribeiro; Miszputen, Sender Jankiel; Oshima, Celina Tizuko Fujiyama; Artigiani Neto, Ricardo; Ribeiro, Daniel Araki; Franco, Marcello

    2011-10-01

    Crohn's disease (CD) is associated with gut barrier dysfunction. Tumour necrosis factor-α (TNF-α) plays an important role into the pathogenesis of several inflammatory diseases because its expression is increased in inflamed mucosa of CD patients. Anti-TNF therapy improves significantly mucosal inflammation. Thus, this study aimed to evaluate the effect of Etanercept (ETC), a tumour necrosis factor alpha (TNF-α) antagonist on the 2,4,6-trinitrobenzene sulfonic acid (TNBS)-induced experimental colitis. A total of 18 Wistar rats were randomized into four groups, as follows: (1) Sham: sham induced-colitis; (2) TNBS: non-treated induced-colitis; (3) ETC control; (4) ETC-treated induced-colitis. Rats from group 4 presented significant improvement either of macroscopic or of histopathological damage in the distal colon. The gene expression of TNF-α mRNA, decreased significantly in this group compared to the TNBS non-treated group. The treatment with etanercept attenuated the colonic damages and reduced the inflammation caused by TNBS. Taken together, our results suggest that ETC attenuates intestinal colitis induced by TNBS in Wistar rats by TNF-α downregulation.

  7. Collagenase-3 (matrix metalloproteinase-13) expression is induced in oral mucosal epithelium during chronic inflammation.

    PubMed Central

    Uitto, V. J.; Airola, K.; Vaalamo, M.; Johansson, N.; Putnins, E. E.; Firth, J. D.; Salonen, J.; López-Otín, C.; Saarialho-Kere, U.; Kähäri, V. M.

    1998-01-01

    Increased proliferation of mucosal epithelium during inflammation is associated with degradation of subepithelial connective tissue matrix and local invasion of the epithelial cells. Here we have studied, whether collagenase-3 (MMP-13), a collagenolytic matrix metalloproteinase with an exceptionally wide substrate specificity, is expressed in the epithelium of chronically inflamed mucosa. Examination of human gingival tissue sections from subjects with chronic adult periodontitis with in situ hybridization revealed marked expression of MMP-13 in basal cells of some epithelial rete ridges expanding into connective tissue. Immunohistochemical staining demonstrated that these cells also expressed strongly laminin-5, suggesting that they are actively migrating cells. A strong signal for MMP-13 mRNA was occasionally also noted in the suprabasal epithelial cells facing the gingival pocket, whereas no collagenase-1 (MMP-1) mRNA was detected in any areas of the epithelium. MMP-13 expression was also detected in fibroblast-like cells associated with collagen fibers of the inflamed subepithelial connective tissue. In organ culture of human oral mucosa, MMP-13 mRNA expression was observed in epithelial cells growing into connective tissue of the specimens. Regulation of MMP-13 expression was examined in cultured normal nonkeratinizing epithelial cells isolated from porcine periodontal ligament. In these cells, MMP-13 expression at the mRNA and protein level was potently enhanced (up to sixfold) by tumor necrosis factor-alpha, transforming growth factor-beta(1), and transforming growth factor-alpha and by keratinocyte growth factor in the presence of heparin. In addition, plating periodontal ligament epithelial cells on type I collagen stimulated MMP-13 expression (sevenfold) as compared with cells grown on tissue culture plastic. The results of this study show, that expression of MMP-13 is specifically induced in undifferentiated epithelial cells during chronic inflammation

  8. Tumor necrosis factor-alpha antagonism by the murine tumor necrosis factor-alpha receptor 2-Fc fusion protein exacerbates histoplasmosis in mice.

    PubMed

    Deepe, George S

    2007-06-01

    Treatment of some inflammatory conditions with tumor necrosis factor-alpha (TNF-alpha) antagonists is efficacious, but such treatments are associated with infections with intracellular pathogens, including Histoplasma capsulatum. We explored protective immunity to H. capsulatum in mice given a fusion protein consisting of TNF-alpha receptor 2 (TNFR2) bound to the Fc portion of mouse IgG1. Intraperitoneal administration of this inhibitor exacerbated primary or secondary pulmonary infection at dosages ranging from 1 to 5 mg/kg. All mice with primary infection given the inhibitor succumbed to infection within 10-21 days of treatment. In secondary histoplasmosis, mice receiving 1, but not 5, mg/kg survived treatment. Fungal burden was increased even if treatment with the inhibitor was initiated after the onset of infection. The inflammatory response of the lungs of mice given the inhibitor did not differ from that of mice given control vehicle. Susceptibility was not associated with major alterations in cytokines known to protect or exacerbate infection. However, expression of nitric oxide synthase 2 (NOS2) was depressed early in primary infection. These results demonstrate that antagonism of endogenous TNF-alpha by this fusion protein modulates susceptibility. Impaired immunity is not a result of altered cytokine responses or changes in the inflammation and may not be demonstrable in other murine strains.

  9. C1q tumor necrosis factor alpha-related protein isoform 5 is increased in mitochondrial DNA-depleted myocytes and activates AMP-activated protein kinase.

    PubMed

    Park, Seung-Yoon; Choi, Jung Hyun; Ryu, Hyun Su; Pak, Youngmi Kim; Park, Kyong Soo; Lee, Hong Kyu; Lee, Wan

    2009-10-09

    Depletion of mtDNA in myocytes causes insulin resistance and alters nuclear gene expression that may be involved in rescuing processes against cellular stress. Here we show that the expression of C1q tumor necrosis factor alpha-related protein isoform 5 (C1QTNF5) is drastically increased following depletion of mtDNA in myocytes. C1QTNF5 is homologous to adiponectin in respect to domain structure, and its expression and secretion from myocytes correlated negatively with the cellular mtDNA content. Similar to adiponectin, C1QTNF5 induced the phosphorylation of AMP-activated protein kinase (AMPK), leading to increased cell surface recruitment of GLUT4 and increased glucose uptake. Treatment of cells with purified recombinant C1QTNF5 increased the phosphorylation of acetyl-CoA carboxylase and stimulated fatty acid oxidation. C1QTNF5-mediated phosphorylation of AMPK or acetyl-CoA carboxylase was unaffected by depletion of adiponectin receptors such as AdipoR1 or AdipoR2, which indicated that adiponectin receptors do not participate in C1QTNF5-induced activation of AMPK. Serum C1QTNF5 levels were significantly higher in obese/diabetic animals (OLETF rats, ob/ob mice, and db/db mice). These results highlight C1QTNF5 as a putative biomarker for mitochondrial dysfunction and a potent activator of AMPK.

  10. Dexmedetomidine inhibits tumor necrosis factor-alpha and interleukin 6 in lipopolysaccharide-stimulated astrocytes by suppression of c-Jun N-terminal kinases.

    PubMed

    Zhang, Xiaobao; Wang, Jun; Qian, Wenyi; Zhao, Jingjing; Sun, Li; Qian, Yanning; Xiao, Hang

    2014-06-01

    Astrocytes play an important role in immune regulation in the central nervous system (CNS). Dexmedetomidine (DEX) has been reported to exert anti-inflammatory effects on astrocytes stimulated by lipopolysaccharide (LPS) both in vitro and in vivo studies. However, the underlying molecular mechanisms remain poorly understood. This study was designed to evaluate the effects of DEX on tumor necrosis factor-alpha (TNF-α) and interleukin 6 (IL-6) gene expressions in LPS-challenged astrocytes. Moreover, c-Jun N-terminal kinases (JNKs) and p38 mitogen-activated protein kinase (MAPK) pathways in LPS-challenged astrocytes were also investigated. In the present study, astrocytes were stimulated with LPS in the absence and presence of various concentrations of DEX. With real-time PCR assay, we found that LPS significantly increased expressions of TNF-α and IL-6 in mRNA level; however, these effects could be attenuated by DEX. Furthermore, JNK pathway might be involved in LPS-induced astrocyte activation because JNK phosphorylation was significantly increased, and the inhibition of this pathway mediated by DEX as well as SP600125 (JNK inhibitor) decreased TNF-α and IL-6 expressions. Moreover, p38 MAPK was also activated by LPS; however, this pathway seemed to have not participated in DEX-mediated LPS-induced inflammation. These results, taken together, suggest that JNK rather than p38 MAPK signal pathway, provides the potential target for the therapeutic effects of DEX for neuronal inflammatory reactions.

  11. Tumor necrosis factor alpha and lymphotoxin production in Hodgkin's disease.

    PubMed

    Kretschmer, C; Jones, D B; Morrison, K; Schlüter, C; Feist, W; Ulmer, A J; Arnoldi, J; Matthes, J; Diamantstein, T; Flad, H D

    1990-08-01

    It is likely that the characteristic histologic features of Hodgkin's disease reflect cytokine production by the tumor cell population. Tumor necrosis factor alpha (TNF-alpha) and lymphotoxin (tumor necrosis factor beta [TNF-beta]) are important inflammatory mediators with wide-ranging effects within the lymphoreticular system. The aim of the present study was to investigate TNF-alpha and lymphotoxin production in the Hodgkin's disease-derived cell lines L428 and L540. At the product level, both cytokines could be demonstrated by immunostaining with specific monoclonal antibodies. TNF-alpha could be demonstrated by means of an enzyme-linked immunosorbent assay in culture supernatants from both cell lines as well as in cell lysates of L428 and L540 cells. Cytotoxic activity could be achieved only in L428 supernatants. This cytotoxic activity could not be blocked by the addition of a polyclonal antibody against TNF-alpha, but was partially inhibited with the monoclonal antibody against lymphotoxin. Synthesis of TNF-alpha and lymphotoxin in both L428 and L540 was confirmed by demonstrating the intracellular-specific messenger RNA (mRNA) using specific cDNA clones in Northern blot analysis. In situ hybridization studies with the TNF-alpha cDNA probe gave positive hybridization signals in L428 and in L540. These results demonstrate the transcription, translation, and export of TNF-alpha and lymphotoxin in cultured Hodgkin's disease-derived cell lines. In addition, results of preliminary experiments are presented in which we demonstrate Reed-Sternberg cells positive for TNF-alpha protein and mRNA in different Hodgkin's disease tissue biopsies, indicating that, at least for TNF-alpha, our cell line data are relevant to the neoplastic population present in Hodgkin's disease tissue.

  12. Macrophage-elicited osteoclastogenesis in response to bacterial stimulation requires Toll-like receptor 2-dependent tumor necrosis factor-alpha production.

    PubMed

    Ukai, Takashi; Yumoto, Hiromichi; Gibson, Frank C; Genco, Caroline Attardo

    2008-02-01

    The receptor activator of NF-kappaB ligand (RANKL) and the proinflammatory cytokines are believed to play important roles in osteoclastogenesis. We recently reported that the innate immune recognition receptor, Toll-like receptor 2 (TLR2), is crucial for inflammatory bone loss in response to infection by Porphyromonas gingivalis, the primary organism associated with chronic inflammatory periodontal disease. However, the contribution of macrophage-expressed TLRs to osteoclastogenesis has not been defined. In this study, we defined a requirement for TLR2 in tumor necrosis factor-alpha (TNF-alpha)-elicited osteoclastogenesis in response to exposure to P. gingivalis. Culture supernatant (CS) fluids from P. gingivalis-stimulated macrophages induced bone marrow macrophage-derived osteoclastogenesis. This activity was dependent on TNF-alpha and occurred independently of RANKL, interleukin-1beta (IL-1beta), and IL-6. CS fluids from P. gingivalis-stimulated TLR2(-/-) macrophages failed to express TNF-alpha, and these fluids induced significantly less osteoclast formation compared with that of the wild-type or the TLR4(-/-) macrophages. In addition, P. gingivalis exposure induced up-regulation of TLR2 expression on the cell surface of macrophages, which was demonstrated to functionally react to reexposure to P. gingivalis, as measured by a further increase in TNF-alpha production. These results demonstrate that macrophage-dependent TLR2 signaling is crucial for TNF-alpha-dependent/RANKL-independent osteoclastogenesis in response to P. gingivalis infection. Furthermore, the ability of P. gingivalis to induce the cell surface expression of TLR2 may contribute to the chronic inflammatory state induced by this pathogen.

  13. Hyper-inducible expression system for streptomycetes.

    PubMed

    Herai, Sachio; Hashimoto, Yoshiteru; Higashibata, Hiroki; Maseda, Hideaki; Ikeda, Haruo; Omura, Satoshi; Kobayashi, Michihiko

    2004-09-28

    Streptomycetes produce useful enzymes and a wide variety of secondary metabolites with potent biological activities (e.g., antibiotics, immunosuppressors, pesticides, etc.). Despite their importance in the pharmaceutical and agrochemical fields, there have been no reports for practical expression systems in streptomycetes. Here, we developed a "P(nitA)-NitR" system for regulatory gene expression in streptomycetes based on the expression mechanism of Rhodococcus rhodochrous J1 nitrilase, which is highly induced by an inexpensive and safe inducer, epsilon-caprolactam. Heterologous protein expression experiments demonstrated that the system allowed suppressed basal expression and hyper-inducible expression, yielding target protein levels of as high as approximately 40% of all soluble protein. Furthermore, the system functioned in important streptomycete strains. Thus, the P(nitA)-NitR system should be a powerful tool for improving the productivity of various useful products in streptomycetes.

  14. Induction of bone-type alkaline phosphatase in human vascular smooth muscle cells: roles of tumor necrosis factor-alpha and oncostatin M derived from macrophages.

    PubMed

    Shioi, Atsushi; Katagi, Miwako; Okuno, Yasuhisa; Mori, Katsuhito; Jono, Shuichi; Koyama, Hidenori; Nishizawa, Yoshiki

    2002-07-12

    Inflammatory cells such as macrophages and T lymphocytes play an important role in vascular calcification associated with atherosclerosis and cardiac valvular disease. In particular, macrophages activated with cytokines derived from T lymphocytes such as interferon-gamma (IFN-gamma) may contribute to the development of vascular calcification. Moreover, we have shown the stimulatory effect of 1alpha,25-dihydroxyvitamin D3 (1,25(OH)2D3) on in vitro calcification through increasing the expression of alkaline phosphatase (ALP), an ectoenzyme indispensable for bone mineralization, in vascular smooth muscle cells. Therefore, we hypothesized that macrophages may induce calcifying phenotype, especially the expression of ALP in human vascular smooth muscle cells (HVSMCs) in the presence of IFN-gamma and 1,25(OH)2D3. To test this hypothesis, we used cocultures of HVSMCs with human monocytic cell line (THP-1) or peripheral blood monocytes (PBMCs) in the presence of IFN-gamma and 1,25(OH)2D3. THP-1 cells or PBMCs induced ALP activity and its gene expression in HVSMCs and the cells with high expression of ALP calcified their extracellular matrix by the addition of beta-glycerophosphate. Thermostability and immunoassay showed that ALP induced in HVSMCs was bone-specific enzyme. We further identified tumor necrosis factor-alpha (TNF-alpha) and oncostatin M (OSM) as major factors inducing ALP in HVSMCs in the culture supernatants of THP-1 cells. TNF-alpha and OSM, only when applied together, increased ALP activities and in vitro calcification in HVSMCs in the presence of IFN-gamma and 1,25(OH)2D3. These results suggest that macrophages may contribute to the development of vascular calcification through producing various inflammatory mediators, especially TNF-alpha and OSM.

  15. Human tumor necrosis factor alpha gene regulation by virus and lipopolysaccharide.

    PubMed

    Goldfeld, A E; Doyle, C; Maniatis, T

    1990-12-01

    We have identified a region of the human tumor necrosis factor alpha (TNF-alpha) gene promoter that is necessary for maximal constitutive, virus-induced, and lipopolysaccharide (LPS)-induced transcription. This region contains three sites that match an NF-kappa B binding-site consensus sequence. We show that these three sites specifically bind NF-kappa B in vitro, yet each of these sites can be deleted from the TNF-alpha promoter with little effect on the induction of the gene by virus or LPS. Moreover, when multimers of these three sites are placed upstream from a truncated TNF-alpha promoter, or a heterologous promoter, an increase in the basal level of transcription is observed that is influenced by sequence context and cell type. However, these multimers are not sufficient for virus or LPS induction of either promoter. Thus, unlike other virus- and LPS-inducible promoters that contain NF-kappa B binding sites, these sites from the TNF-alpha promoter are neither required nor sufficient for virus or LPS induction. Comparison of the sequence requirements of virus induction of the human TNF-alpha gene in mouse L929 and P388D1 cells reveals significant differences, indicating that the sequence requirements for virus induction of the gene are cell type-specific. However, the sequences required for virus and LPS induction of the gene in a single cell type, P388D1, overlap.

  16. Tumor necrosis factor-alpha regulates transforming growth factor-beta-dependent epithelial-mesenchymal transition by promoting hyaluronan-CD44-moesin interaction.

    PubMed

    Takahashi, Eri; Nagano, Osamu; Ishimoto, Takatsugu; Yae, Toshifumi; Suzuki, Yoshimi; Shinoda, Takeshi; Nakamura, Satoshi; Niwa, Shinichiro; Ikeda, Shun; Koga, Hisashi; Tanihara, Hidenobu; Saya, Hideyuki

    2010-02-05

    Aberrant epithelial-mesenchymal transition (EMT) is involved in development of fibrotic disorders and cancer invasion. Alterations of cell-extracellular matrix interaction also contribute to those pathological conditions. However, the functional interplay between EMT and cell-extracellular matrix interactions remains poorly understood. We now show that the inflammatory mediator tumor necrosis factor-alpha (TNF-alpha) induces the formation of fibrotic foci by cultured retinal pigment epithelial cells through activation of transforming growth factor-beta (TGF-beta) signaling in a manner dependent on hyaluronan-CD44-moesin interaction. TNF-alpha promoted CD44 expression and moesin phosphorylation by protein kinase C, leading to the pericellular interaction of hyaluronan and CD44. Formation of the hyaluronan-CD44-moesin complex resulted in both cell-cell dissociation and increased cellular motility through actin remodeling. Furthermore, this complex was found to be associated with TGF-beta receptor II and clathrin at actin microdomains, leading to activation of TGF-beta signaling. We established an in vivo model of TNF-alpha-induced fibrosis in the mouse eye, and such ocular fibrosis was attenuated in CD44-null mice. The production of hyaluronan and its interaction with CD44, thus, play an essential role in TNF-alpha-induced EMT and are potential therapeutic targets in fibrotic disorders.

  17. Loss of ADAM17-Mediated Tumor Necrosis Factor Alpha Signaling in Intestinal Cells Attenuates Mucosal Atrophy in a Mouse Model of Parenteral Nutrition

    PubMed Central

    Feng, Yongjia; Tsai, Yu-Hwai; Xiao, Weidong; Ralls, Matthew W.; Stoeck, Alex; Wilson, Carole L.; Raines, Elaine W.

    2015-01-01

    Total parenteral nutrition (TPN) is commonly used clinically to sustain patients; however, TPN is associated with profound mucosal atrophy, which may adversely affect clinical outcomes. Using a mouse TPN model, removing enteral nutrition leads to decreased crypt proliferation, increased intestinal epithelial cell (IEC) apoptosis and increased mucosal tumor necrosis factor alpha (TNF-α) expression that ultimately produces mucosal atrophy. Upregulation of TNF-α signaling plays a central role in mediating TPN-induced mucosal atrophy without intact epidermal growth factor receptor (EGFR) signaling. Currently, the mechanism and the tissue-specific contributions of TNF-α signaling to TPN-induced mucosal atrophy remain unclear. ADAM17 is an ectodomain sheddase that can modulate the signaling activity of several cytokine/growth factor receptor families, including the TNF-α/TNF receptor and ErbB ligand/EGFR pathways. Using TPN-treated IEC-specific ADAM17-deficient mice, the present study demonstrates that a loss of soluble TNF-α signaling from IECs attenuates TPN-induced mucosal atrophy. Importantly, this response remains dependent on the maintenance of functional EGFR signaling in IECs. TNF-α blockade in wild-type mice receiving TPN confirmed that soluble TNF-α signaling is responsible for downregulation of EGFR signaling in IECs. These results demonstrate that ADAM17-mediated TNF-α signaling from IECs has a significant role in the development of the proinflammatory state and mucosal atrophy observed in TPN-treated mice. PMID:26283731

  18. Divergent mechanisms utilized by SOCS3 to mediate interleukin-10 inhibition of tumor necrosis factor alpha and nitric oxide production by macrophages.

    PubMed

    Qasimi, Pooran; Ming-Lum, Andrew; Ghanipour, Ali; Ong, Christopher J; Cox, Michael E; Ihle, James; Cacalano, Nicolas; Yoshimura, Akihiko; Mui, Alice L-F

    2006-03-10

    The cytokine interleukin-10 (IL-10) potently inhibits macrophage function through activation of the transcription factor STAT3. The expression of SOCS3 (suppressor of cytokine signaling-3) has been shown to be induced by IL-10 in a STAT3-dependent manner. However, the relevance of SOCS3 expression to the anti-inflammatory effect of IL-10 on macrophages has been controversial. Through kinetic analysis of the requirement for SOCS3 in IL-10 inhibition of lipopolysaccharide (LPS)-stimulated tumor necrosis factor-alpha (TNFalpha) transcription and translation, SOCS3 was found to be necessary for TNFalpha expression during the early phase, but not the late phase of IL-10 action. SOCS3 was essential for IL-10 inhibition of LPS-stimulated production of iNOS (inducible nitric-oxide synthase) protein and nitric oxide (NO). To determine the domains of SOCS3 protein important in mediating these effects, SOCS3-/- macrophages were reconstituted with SOCS3 mutated for the SH2, KIR, SOCS box domains, and tyrosines 204 (Tyr204) and 221 (Tyr221). The SH2 domain, SOCS box, and both Tyr204 and Tyr221 were required for IL-10 inhibition of TNFalpha mRNA and protein expression, but interestingly the KIR domain was necessary only for IL-10 inhibition of TNFalpha protein expression. In contrast, Tyr204 and Tyr221 were the only structural features of SOCS3 that were necessary in mediating IL-10 inhibition of iNOS protein expression and NO production. These data define SOCS3 as an important mediator of IL-10 inhibition of macrophage activation and that SOCS3 interferes with distinct LPS-stimulated signal transduction events through differing mechanisms.

  19. Molecular design of conjugated tumor necrosis factor-alpha: synthesis and characteristics of polyvinyl pyrrolidone modified tumor necrosis factor-alpha.

    PubMed

    Kamada, H; Tsutsumi, Y; Tsunoda, S; Kihira, T; Kaneda, Y; Yamamoto, Y; Nakagawa, S; Horisawa, Y; Mayumi, T

    1999-04-13

    We conjugated tumor necrosis factor-alpha (TNF-alpha) with the synthetic polymeric modifier polyvinyl pyrrolidone (PVP) to facilitate its clinical use for anti-tumor therapy. TNF-alpha was chemically conjugated with the terminal carboxyl-bearing PVP at one end of its main chain, which was radically polymerized via the formation of an amide bond between the lysine amino groups of TNF-alpha and carboxyl group of PVP. In vitro specific bioactivity of PVP-conjugated TNF-alpha (PVP-TNF-alpha) relative to that of native TNF-alpha gradually decreased with increases in the degree of PVP attachment. In contrast, PVP-TNF-alpha in which 40% of TNF-alpha lysine residues were coupled with PVP (MPVP-TNF-alpha) exhibited the highest anti-tumor activity among the conjugated derivatives examined. MPVP-TNF-alpha had more than 200-fold higher anti-tumor efficacy than native TNF-alpha, and the anti-tumor activity of MPVP- TNF-alpha was more than 5-fold stronger than that MPEG- TNF-alpha which had the highest anti-tumor activity among PEG-conjugated TNF-alphas examined. Additionally, a high dose of native TNF-alpha induced toxic side-effects such as body weight reduction, piloerection and tissue inflammation, while no side effects were observed following i.v. administration of MPVP-TNF-alpha. The plasma half-life of MPVP-TNF-alpha (360 min) was about 80 and 3-fold longer than those of native TNF-alpha (4.6 min) and MPEG-TNF-alpha (122 min), respectively. These results suggested that PVP is a useful polymeric modifier for increasing the anti-tumor activity of PVP.

  20. Omentin inhibits TNF-{alpha}-induced expression of adhesion molecules in endothelial cells via ERK/NF-{kappa}B pathway

    SciTech Connect

    Zhong, Xia; Li, Xiaonan; Liu, Fuli; Tan, Hui; Shang, Deya

    2012-08-24

    Highlights: Black-Right-Pointing-Pointer Omentin inhibited TNF-{alpha}-induced adhesion of THP-1 cells to HUVECs. Black-Right-Pointing-Pointer Omentin reduces expression of ICAM-1 and VCAM-1 induced by TNF-{alpha} in HUVECs. Black-Right-Pointing-Pointer Omentin inhibits TNF-{alpha}-induced ERK and NF-{kappa}B activation in HUVECs. Black-Right-Pointing-Pointer Omentin supreeses TNF-{alpha}-induced expression of ICAM-1 and VCAM-1 via ERK/NF-{kappa}B pathway. -- Abstract: In the present study, we investigated whether omentin affected the expression of intracellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1 (VCAM-1) in tumor necrosis factor-{alpha} (TNF-{alpha}) induced human umbilical vein endothelial cells (HUVECs). Our data showed that omentin decreased TNF-{alpha}-induced expression of ICAM-1 and VCAM-1 in HUVECs. In addition, omentin inhibited TNF-{alpha}-induced adhesion of THP-1 cells to HUVECs. Further, we found that omentin inhibited TNF-{alpha}-activated signal pathway of nuclear factor-{kappa}B (NF-{kappa}B) by preventing NF-{kappa}B inhibitory protein (I{kappa}B{alpha}) degradation and NF-{kappa}B/DNA binding activity. Omentin pretreatment significantly inhibited TNF-{alpha}-induced ERK activity and ERK phosphorylation in HUVECs. Pretreatment with PD98059 suppressed TNF-{alpha}-induced NF-{kappa}B activity. Omentin, NF-kB inhibitor (BAY11-7082) and ERK inhibitor (PD98059) reduced the up-regulation of ICAM-1 and VCAM-1 induced by TNF-{alpha}. These results suggest that omentin may inhibit TNF-{alpha}-induced expression of adhesion molecules in endothelial cells via blocking ERK/NF-{kappa}B pathway.

  1. Immunological effects of a tumor necrosis factor alpha-armed oncolytic adenovirus.

    PubMed

    Hirvinen, Mari; Rajecki, Maria; Kapanen, Mika; Parviainen, Suvi; Rouvinen-Lagerström, Noora; Diaconu, Iulia; Nokisalmi, Petri; Tenhunen, Mikko; Hemminki, Akseli; Cerullo, Vincenzo

    2015-03-01

    For long it has been recognized that tumor necrosis factor alpha (TNFa) has anticancer characteristics, and its use as a cancer therapeutic was proposed already in the 1980s. However, its systemic toxicity has limited its usability. Oncolytic viruses, selectively cancer-killing viruses, have shown great potency, and one of their most useful aspects is their ability to produce high amounts of transgene products locally, resulting in high local versus systemic concentrations. Therefore, the overall magnitude of tumor cell killing results from the combination of oncolysis, transgene-mediated direct effect such as TNFa-mediated apoptosis, and, perhaps most significantly, from activation of the host immune system against the tumor. We generated a novel chimeric oncolytic adenovirus expressing human TNFa, Ad5/3-D24-hTNFa, whose efficacy and immunogenicity were tested in vitro and in vivo. The hTNFa-expressing adenovirus showed increased cancer-eradicating potency, which was shown to be because of elevated apoptosis and necrosis rates and induction of various immune responses. Interestingly, we saw increase in immunogenic cell death markers in Ad5/3-d24-hTNFa-treated cells. Moreover, tumors treated with Ad5/3-D24-hTNFa displayed enhanced presence of OVA-specific cytotoxic T cells. We thus can conclude that tumor eradication and antitumor immune responses mediated by Ad5/3-d24-hTNFa offer a new potential drug candidate for cancer therapy.

  2. Mitochondria mediate tumor necrosis factor-alpha/NF-kappaB signaling in skeletal muscle myotubes

    NASA Technical Reports Server (NTRS)

    Li, Y. P.; Atkins, C. M.; Sweatt, J. D.; Reid, M. B.; Hamilton, S. L. (Principal Investigator)

    1999-01-01

    Tumor necrosis factor-alpha (TNF-alpha) is implicated in muscle atrophy and weakness associated with a variety of chronic diseases. Recently, we reported that TNF-alpha directly induces muscle protein degradation in differentiated skeletal muscle myotubes, where it rapidly activates nuclear factor kappaB (NF-kappaB). We also have found that protein loss induced by TNF-alpha is NF-kappaB dependent. In the present study, we analyzed the signaling pathway by which TNF-alpha activates NF-kappaB in myotubes differentiated from C2C12 and rat primary myoblasts. We found that activation of NF-kappaB by TNF-alpha was blocked by rotenone or amytal, inhibitors of complex I of the mitochondrial respiratory chain. On the other hand, antimycin A, an inhibitor of complex III, enhanced TNF-alpha activation of NK-kappaB. These results suggest a key role of mitochondria-derived reactive oxygen species (ROS) in mediating NF-kappaB activation in muscle. In addition, we found that TNF-alpha stimulated protein kinase C (PKC) activity. However, other signal transduction mediators including ceramide, Ca2+, phospholipase A2 (PLA2), and nitric oxide (NO) do not appear to be involved in the activation of NF-kappaB.

  3. Induction of sensitivity to the cytotoxic action of tumor necrosis factor alpha by adenovirus E1A is independent of transformation and transcriptional activation.

    PubMed Central

    Ames, R S; Holskin, B; Mitcho, M; Shalloway, D; Chen, M J

    1990-01-01

    We have previously shown that expression of the adenovirus E1A 12S or 13S products in NIH 3T3 fibroblasts induces susceptibility to the cytotoxic actions of tumor necrosis factor alpha (TNF alpha). A large number of studies have mapped the multiple biological functions of the 12S and 13S products to three highly conserved regions (CR) within the E1A sequence. Here we used plasmids coding for E1A deletion and point mutants in these regions to generate target cell lines for TNF alpha cytotoxicity assays to determine which regions and functions are necessary for the induction of TNF alpha sensitivity. Expression of CR1 was required for the induction of TNF alpha sensitivity. This finding did not reflect a requirement for transforming or transcriptional repression activity, since some mutants that were defective in both of these properties were able to induce TNF alpha sensitivity. CR2 transformation-defective point mutants, but not a CR2/3 region deletion mutant, were also able to induce sensitivity. In addition, NIH 3T3 cells expressing the retroviral transcription activators tat from human immunodeficiency virus type 1 and tax from human T-lymphotropic virus type I were not sensitive to TNF alpha. However, the possibility that E1A-mediated transcriptional activation can augment the induction of TNF alpha sensitivity is not excluded. Comparison of data from previous biological studies with the TNF alpha cytotoxicity assays presented here suggested that the mechanism by which E1A induces sensitivity to TNF alpha in NIH 3T3 cells is independent of many of the known E1A biological functions, including transformation in cooperation with ras, immortalization, induction of DNA synthesis in quiescent cells, and transcriptional repression. A novel E1A-mediated effect may be involved, although our data do not exclude the possibility that sensitization to TNF alpha is mediated through E1A binding to cellular proteins. Images PMID:2143540

  4. Differential cytokine expression in skin graft healing in inducible nitric oxide synthase knockout mice.

    PubMed

    Most, D; Efron, D T; Shi, H P; Tantry, U S; Barbul, A

    2001-10-01

    Inducible nitric oxide synthase (iNOS) and its product, nitric oxide, have been shown to play important roles in wound biology. The present study was performed to investigate the role of iNOS in modulating the cytokine cascade during the complex process of skin graft wound healing.Fifteen iNOS-knockout mice and 15 wild-type C57BL/6J mice were subjected to autogenous 1-cm2 intrascapular full-thickness skin grafts. Three animals in each group were killed on postoperative days 3, 5, 7, 10, and 14. Specimens were then analyzed using nonisotopic in situ hybridization versus mRNA of tumor growth factor-beta1, vascular endothelial growth factor, iNOS, endothelial nitric oxide synthase (eNOS), tumor necrosis factor-alpha, and basic fibroblast growth factor, as well as positive and negative control probes. Positive cells in both grafts and wound beds were counted using a Leica microgrid. Scar thickness was measured with a Leica micrometer. Data were analyzed using the unpaired Student's t test. Expression of iNOS was 2- to 4-fold higher in knockout mice than in wild-type mice on postoperative days 5, 7, and 14. Expression of eNOS was 2- to 2.5-fold higher in knockout mice than in wild-type mice on postoperative days 5 and 7. Tumor necrosis factor-alpha expression was 2- to 7-fold higher in knockout mice than in wild-type mice on all postoperative days. In contrast, expression levels of angiogenic/fibrogenic cytokines (vascular endothelial growth factor, basis fibroblast growth factor, and tumor growth factor-beta1) were 2.5- to 4-fold higher in wild-type mice than in knockout mice. Scars were 1.5- to 2.5-fold thicker in knockout mice than in wild-type mice at all time points. All of the above results represent statistically significant differences (p < 0.05). Significantly different patterns of cytokine expression were seen in knockout and wild-type mice. Although the scar layer was thicker in knockout mice, it showed much greater infiltration with inflammatory cells. These

  5. Prior peritoneal lavage with hot 0.9 % saline induces HSP70 expression and protects against cerulein-induced acute pancreatitis in rats.

    PubMed

    Meng, Ke; Liu, Qingsen; Dou, Yan; Huang, Qiyang

    2013-02-01

    Recent studies have indicated that pre-induction of heat shock protein 70 (HSP70) expression in the pancreas protects against secretagogue-induced pancreatitis. In those studies, the HSP70 was mostly induced by unfeasible conditions. The aim of this current study was to investigate the effect of peritoneal lavage with hot 0.9 % saline (42 °C) on the pancreatic expression of HSP70 and its protective effect on cerulein-induced acute pancreatitis in rats. Male Wistar rats were peritoneally lavaged with 0.9 % saline at 42 °C for 30 min. HSP70 expression was evaluated by western blotting analysis. Prior peritoneal lavages with hot and warm saline were performed. Acute pancreatitis was induced by administration of intraperitoneal injection of cerulein (20 μg/kg) four times, and its severity was assessed by measuring serum amylase, tumor necrosis factor-alpha (TNF-α), interleukin-6 (IL-6), and trypsinogen activation peptide (TAP) levels. Pancreatic sections were stained with hematoxylin and eosin for histological evaluation. Peritoneal lavage with hot 0.9 % saline increased intrapancreatic HSP70 expression and ameliorated the cerulein-induced pancreatitis in rats, judged by the significantly reduced serum amylase, TNF-α, and IL-6 concentrations; histopathological scores, and serum TAP levels. Peritoneal lavage with hot 0.9 % saline can induce HSP70 expression and prevent cerulein-induced acute pancreatitis in rats. The results suggest that HSP70 protects against cerulein-induced pancreatitis by preventing proinflammatory cytokine synthesis and trypsinogen activation during acute pancreatitis.

  6. p53-dependent NDRG1 expression induces inhibition of intestinal epithelial cell proliferation but not apoptosis after polyamine depletion.

    PubMed

    Zhang, Ai-Hong; Rao, Jaladanki N; Zou, Tongtong; Liu, Lan; Marasa, Bernard S; Xiao, Lan; Chen, Jie; Turner, Douglas J; Wang, Jian-Ying

    2007-07-01

    Normal intestinal mucosal growth requires polyamines that regulate expression of various genes involved in cell proliferation, growth arrest, and apoptosis. Our previous studies have shown that polyamine depletion stabilizes p53, resulting in inhibition of intestinal epithelial cell (IEC) proliferation, but the exact downstream targets of induced p53 are still unclear. The NDRG1 (N-myc downregulated gene-1) gene encodes a growth-related protein, and its transcription can be induced in response to stress. The current study tests the hypothesis that induced p53 inhibits IEC proliferation by upregulating NDRG1 expression following polyamine depletion. Depletion of cellular polyamines by inhibiting ornithine decarboxylase (ODC) with alpha-difluoromethylornithine not only induced p53 but also increased NDRG1 transcription as indicated by induction of the NDRG1 promoter activity and increased levels of NDRG1 mRNA and protein, all of which were prevented by using specific p53 siRNA and in cells with a targeted deletion of p53. In contrast, increased levels of cellular polyamines by ectopic expression of the ODC gene decreased p53 and repressed expression of NDRG1. Consistently, polyamine depletion-induced activation of the NDRG1-promoter was decreased when p53-binding sites within the NDRG1 proximal promoter region were deleted. Ectopic expression of the wild-type NDRG1 gene inhibited DNA synthesis and decreased final cell numbers regardless of the presence or absence of endogenous p53, whereas silencing NDRG1 promoted cell growth. However, overexpression of NDRG1 failed to directly induce cell death and to alter susceptibility to apoptosis induced by tumor necrosis factor-alpha/cycloheximide. These results indicate that NDRG1 is one of the direct mediators of induced p53 following polyamine depletion and that p53-dependent NDRG1 expression plays a critical role in the negative control of IEC proliferation.

  7. Tumor necrosis factor-alpha inhibitors suppress CCL2 chemokine in monocytes via epigenetic modification.

    PubMed

    Lin, Yi-Ching; Lin, Yu-Chih; Huang, Ming-Yii; Kuo, Po-Lin; Wu, Cheng-Chin; Lee, Min-Sheng; Hsieh, Chong-Chao; Kuo, Hsuan-Fu; Kuo, Chang-Hung; Tsai, Wen-Chan; Hung, Chih-Hsing

    2017-03-01

    The treatment of rheumatoid arthritis (RA) with tumor necrosis factor-alpha (TNF-α) inhibitors could lead to adverse effects. Therefore, the identification of downstream therapeutic targets is important. Monocyte chemoattractant protein-1 (MCP-1, also called CCL2) is related to RA disease activity, and epigenetic modifications are hypothesized to regulate gene expression in RA pathogenesis. We studied the effects of two TNF-α inhibitors, etanercept and adalimumab, on CCL2 expression and the potentially associated intracellular mechanisms, including epigenetic regulation. Etanercept and adalimumab decreased CCL2 production in THP-1 cells and human primary monocytes, as detected using enzyme-linked immunosorbent assays, and these changes in the CCL2 levels were independent of the TNF-α levels. Etanercept and adalimumab suppressed mitogen-activated protein kinase (MAPK) phospho-p38, phospho-JNK, phospho-ERK and nuclear factor-κB (NF-κB) phospho-p65, as demonstrated using western blot analyses. The investigation of epigenetic modifications using chromatin immunoprecipitation revealed that etanercept and adalimumab down-regulated acetylation of histone (H)3 and H4 in the CCL2 promoter region by decreasing the recruitment of the NF-κB associated acetyltransferases p300, CBP and PCAF. Etanercept and adalimumab also down-regulated trimethylation of H3K4, H3K27, H3K36 and H3K79 in the CCL2 promoter region by decreasing the expression of the related methyltransferases WDR5 and Smyd2. We demonstrated that TNF-α inhibitors exert immunomodulatory effects on CCL2 expression in human monocytes via MAPKs, NF-κB and epigenetic modifications. These findings broaden the mechanistic knowledge related to TNF-α inhibitors and provide novel therapeutic targets for RA.

  8. The 19-kilodalton adenovirus E1B transforming protein inhibits programmed cell death and prevents cytolysis by tumor necrosis factor alpha.

    PubMed Central

    White, E; Sabbatini, P; Debbas, M; Wold, W S; Kusher, D I; Gooding, L R

    1992-01-01

    The adenovirus E1A and E1B proteins are required for transformation of primary rodent cells. When expressed in the absence of the 19,000-dalton (19K) E1B protein, however, the E1A proteins are acutely cytotoxic and induce host cell chromosomal DNA fragmentation and cytolysis, analogous to cells undergoing programmed cell death (apoptosis). E1A alone can efficiently initiate the formation of foci which subsequently undergo abortive transformation whereby stimulation of cell growth is counteracted by continual cell death. Cell lines with an immortalized growth potential eventually arise with low frequency. Coexpression of the E1B 19K protein with E1A is sufficient to overcome abortive transformation to produce high-frequency transformation. Like E1A, the tumoricidal cytokine tumor necrosis factor alpha (TNF-alpha) evokes a programmed cell death response in many tumor cell lines by inducing DNA fragmentation and cytolysis. Expression of the E1B 19K protein by viral infection, by transient expression, or in transformed cells completely and specifically blocks this TNF-alpha-induced DNA fragmentation and cell death. Cosegregation of 19K protein transforming activity with protection from TNF-alpha-mediated cytolysis demonstrates that both activities are likely the consequence of the same function of the protein. Therefore, we propose that by suppressing an intrinsic cell death mechanism activated by TNF-alpha or E1A, the E1B 19K protein enhances the transforming activity of E1A and enables adenovirus to evade TNF-alpha-dependent immune surveillance. Images PMID:1317006

  9. Role of cytochrome C in apoptosis: increased sensitivity to tumor necrosis factor alpha is associated with respiratory defects but not with lack of cytochrome C release.

    PubMed

    Vempati, Uma D; Diaz, Francisca; Barrientos, Antoni; Narisawa, Sonoko; Mian, Abdul M; Millán, José Luis; Boise, Lawrence H; Moraes, Carlos T

    2007-03-01

    Although the role of cytochrome c in apoptosis is well established, details of its participation in signaling pathways in vivo are not completely understood. The knockout for the somatic isoform of cytochrome c caused embryonic lethality in mice, but derived embryonic fibroblasts were shown to be resistant to apoptosis induced by agents known to trigger the intrinsic apoptotic pathway. In contrast, these cells were reported to be hypersensitive to tumor necrosis factor alpha (TNF-alpha)-induced apoptosis, which signals through the extrinsic pathway. Surprisingly, we found that this cell line (CRL 2613) respired at close to normal levels because of an aberrant activation of a testis isoform of cytochrome c, which, albeit expressed at low levels, was able to replace the somatic isoform for respiration and apoptosis. To produce a bona fide cytochrome c knockout, we developed a mouse knockout for both the testis and somatic isoforms of cytochrome c. The mouse was made viable by the introduction of a ubiquitously expressed cytochrome c transgene flanked by loxP sites. Lung fibroblasts in which the transgene was deleted showed no cytochrome c expression, no respiration, and resistance to agents that activate the intrinsic and to a lesser but significant extent also the extrinsic pathways. Comparison of these cells with lines with a defective oxidative phosphorylation system showed that cells with defective respiration have increased sensitivity to TNF-alpha-induced apoptosis, but this process was still amplified by cytochrome c. These studies underscore the importance of oxidative phosphorylation and apoptosome function to both the intrinsic and extrinsic apoptotic pathways.

  10. Stress downregulates lipopolysaccharide-induced expression of proinflammatory cytokines in the spleen, pituitary, and brain of mice.

    PubMed

    Goujon, E; Parnet, P; Laye, S; Combe, C; Kelley, K W; Dantzer, R

    1995-12-01

    Mice injected with LPS (10 mu g/mouse, sc) or saline were submitted to a 15-min restraint stress and sacrificed 1 or 2 h later to assess the effect of stress on the induction of interleukin-1beta (IL-1beta) and other proinflammatory cytokines (IL-1alpha, IL-1ra, IL-6, and tumor necrosis factor-alpha) in the spleen, pituitary, hypothalamus, hippocampus, and striatum. LPS-induced cytokine gene expression, as determined by comparative RT-PCR, was lower in stressed than in nonstressed mice. LPS increased plasma and tissue levels of IL-1beta, as determined by ELISA, but this effect was less marked in stressed than in nonstressed mice. These results are discussed in relation to the modulatory effects of glucocorticoids on cytokine production.

  11. PPARγ regulates the mitochondrial dysfunction in human neural stem cells with tumor necrosis factor alpha.

    PubMed

    Chiang, M-C; Cheng, Y-C; Lin, K-H; Yen, C-H

    2013-01-15

    Peroxisome proliferator-activated receptor gamma (PPARγ) belongs to a family of ligand-activated transcription factors, and its ligands are known to control many physiological and pathological conditions. The hypothesis of our study was that the PPARγ agonist (rosiglitazone) could mediate tumor necrosis factor alpha (TNFα) related to the regulation of human neural stem cells (hNSCs), by which TNFα possibly fulfills important roles in neuronal impairment. The results show that PPARγ mediates the cell viability of hNSCs via the downregulation of the activity of caspase 3, indicating that this rescue effect of PPARγ could improve the reduced levels of two mitochondrial regulators, adenosine monophosphate-activated protein kinase (AMPK) and Sirtuin 1 (SIRT1) in the hNSCs with TNFα. The stimulation of mitochondrial function by PPARγ was associated with activation of the PPAR coactivator1 alpha (PGC1α) pathway by up-regulation of oxidative defense and mitochondrial systems. The above protective effects appeared to be exerted by a direct activation of the rosiglitazone, because it protected hNSCs from TNFα-evoked oxidative stress and mitochondrial deficiency. Here we show that the rosiglitazone protects hNSCs against Aβ-induced apoptosis and promotes cell survival. These findings extend our understanding of the central role of PPARγ in TNFα-related neuronal impairment, which probably increases risks of neurodegenerative diseases. The anti-inflammatory effects of PPARγ in the hNSCs with TNFα, and the involved mechanisms were also characterized.

  12. Involvement of tumor necrosis factor-alpha in the pathogenesis of autoimmune orchitis in rats.

    PubMed

    Suescun, María O; Rival, Claudia; Theas, María S; Calandra, Ricardo S; Lustig, Livia

    2003-06-01

    We studied the testicular macrophages of rats with experimental autoimmune orchitis (EAO) and analyzed whether the tumor necrosis factor-alpha (TNFalpha) is involved in germ cell apoptosis and in Leydig cell steroidogenesis. The EAO was induced in adult male Sprague-Dawley rats by active immunization with testicular homogenate and adjuvants. In the experimental group, a severe orchitis was observed 80 days after the first immunization. ED1- and ED2-positive macrophages were quantified by immunohistochemistry. The TNFalpha concentration of conditioned media from testicular macrophages (TMCM) was determined by ELISA. The number of apoptotic TNF receptor 1 (TNFR1)-positive germ cells was identified by combining in situ end labeling of apoptotic DNA and immunohistochemical techniques. The effect of TNFalpha on Leydig cell testosterone production was determined by RIA. In rats with EAO, we observed a significant increase in the number of TNFalpha-positive testicular macrophages, the TNFalpha concentration in TMCM, and the number of TNFR1-positive germ cells. Sixty percent of TNFR1-positive germ cells were apoptotic. These results suggest that TNFalpha could be involved in the pathogenesis of EAO. Acting together with other local factors such as Fas-FasL, TNFalpha could trigger germ cell apoptosis. We also demonstrated that TNFalpha inhibited in vitro testosterone production in basal and hCG-stimulated Leydig cells from rats with orchitis.

  13. Interleukin-13 inhibits inducible nitric oxide synthase expression in human mesangial cells.

    PubMed Central

    Saura, M; Martínez-Dalmau, R; Minty, A; Pérez-Sala, D; Lamas, S

    1996-01-01

    The synthesis of nitric oxide in inflammatory situations requires the expression of an inducible isoform of nitric oxide synthase (iNOS). Human mesangial cells (HMC) express an iNOS enzyme after exposure to multiple co-stimuli. In this study we have observed that while tumour necrosis factor-alpha, interleukin (IL)-1 beta, interferon-gamma and bacterial lipopolysaccharide (LPS) were unable to significantly induce NO synthesis when used alone, they induced an evident stimulation of NO synthesis when used in various combinations. A mixture of the three cytokines (CM) and LPS resulted in a 10-15-fold stimulation of NO synthesis over control values which started to be significant after 16 h. The addition of IL-13, a cytokine with anti-inflammatory properties, inhibited CM/LPS-induced NO synthesis in a concentration-dependent manner. A marked inhibitory effect (60-65%) could be observed when HMC were treated with IL-13 (10 ng/ml) 24 h before, at the same time as, or even 4 h after the addition of CM/LPS. This inhibitory effect was still significant (25%) when IL-13 was added 16 h after CM/LPS. Northern analysis showed that IL-13-mediated iNOS inhibition was closely correlated with the suppression of iNOS mRNA expression. These results identify IL-13 as a powerful regulatory tool for the inhibition of NO synthesis in human cells, a property which may be pathophysiologically relevant in NO-related inflammatory processes. PMID:8573104

  14. Cryptotanshinone inhibits TNF-α-induced LOX-1 expression by suppressing reactive oxygen species (ROS) formation in endothelial cells.

    PubMed

    Ran, Xiaoli; Zhao, Wenwen; Li, Wenping; Shi, Jingshan; Chen, Xiuping

    2016-07-01

    Cryptotanshinone (CPT) is a natural compound isolated from traditional Chinese medicine Salvia miltiorrhiza Bunge. In the present study, the regulatory effect and potential mechanisms of CPT on tumor necrosis factor alpha (TNF-α) induced lectin-like receptor for oxidized low density lipoprotein (LOX-1) were investigated. Human umbilical vein endothelial cells (HUVECs) were cultured and the effect of TNF-α on LOX-1 expression at mRNA and protein levels was determined by Real-time PCR and Western blotting respectively. The formation of intracellular ROS was determined with fluorescence probe CM-DCFH2-DA. The endothelial ox-LDL uptake was evaluated with DiI-ox-LDL. The effect of CPT on LOX-1 expression was also evaluated with SD rats. TNF-α induced LOX-1 expression in a dose- and time-dependent manner in endothelial cells. TNF-α induced ROS formation, phosphorylation of NF-κB p65 and ERK, and LOX-1 expression, which were suppressed by rotenone, DPI, NAC, and CPT. NF-κB inhibitor BAY11-7082 and ERK inhibitor PD98059 inhibited TNF-α-induced LOX-1 expression. CPT and NAC suppressed TNF-α-induced LOX-1 expression and phosphorylation of NF-κB p65 and ERK in rat aorta. These data suggested that TNF-α induced LOX-1 expression via ROS activated NF-κB/ERK pathway, which could be inhibited by CPT. This study provides new insights for the anti-atherosclerotic effect of CPT.

  15. Cryptotanshinone inhibits TNF-α-induced LOX-1 expression by suppressing reactive oxygen species (ROS) formation in endothelial cells

    PubMed Central

    Ran, Xiaoli; Zhao, Wenwen; Li, Wenping

    2016-01-01

    Cryptotanshinone (CPT) is a natural compound isolated from traditional Chinese medicine Salvia miltiorrhiza Bunge. In the present study, the regulatory effect and potential mechanisms of CPT on tumor necrosis factor alpha (TNF-α) induced lectin-like receptor for oxidized low density lipoprotein (LOX-1) were investigated. Human umbilical vein endothelial cells (HUVECs) were cultured and the effect of TNF-α on LOX-1 expression at mRNA and protein levels was determined by Real-time PCR and Western blotting respectively. The formation of intracellular ROS was determined with fluorescence probe CM-DCFH2-DA. The endothelial ox-LDL uptake was evaluated with DiI-ox-LDL. The effect of CPT on LOX-1 expression was also evaluated with SD rats. TNF-α induced LOX-1 expression in a dose- and time-dependent manner in endothelial cells. TNF-α induced ROS formation, phosphorylation of NF-κB p65 and ERK, and LOX-1 expression, which were suppressed by rotenone, DPI, NAC, and CPT. NF-κB inhibitor BAY11-7082 and ERK inhibitor PD98059 inhibited TNF-α-induced LOX-1 expression. CPT and NAC suppressed TNF-α-induced LOX-1 expression and phosphorylation of NF-κB p65 and ERK in rat aorta. These data suggested that TNF-α induced LOX-1 expression via ROS activated NF-κB/ERK pathway, which could be inhibited by CPT. This study provides new insights for the anti-atherosclerotic effect of CPT. PMID:27382351

  16. Dissociation of lipopolysaccharide (LPS)-inducible gene expression in murine macrophages pretreated with smooth LPS versus monophosphoryl lipid A.

    PubMed Central

    Henricson, B E; Manthey, C L; Perera, P Y; Hamilton, T A; Vogel, S N

    1993-01-01

    Lipopolysaccharide (LPS) and the nontoxic derivative of lipid A, monophosphoryl lipid A (MPL), were employed to assess the relationship between expression of LPS-inducible inflammatory genes and the induction of tolerance to LPS in murine macrophages. Both LPS and MPL induced expression (as assessed by increased steady-state mRNA levels) of a panel of seven "early" inflammatory genes including the tumor necrosis factor alpha (TNF-alpha), interleukin-1 beta, type 2 TNF receptor (TNFR-2), IP-10, D3, D8, and D2 genes (the last four represent LPS-inducible early genes whose functions remain unknown). In addition, LPS and MPL were both capable of inducing tolerance to LPS. The two stimuli differed in the relative concentration required to induce various outcome measures, with LPS being 100- to 1,000-fold more potent on a mass concentration basis. Characterization of the tolerant state identified three distinct categories of responsiveness. Two genes (IP-10 and D8) exhibited strong desensitization in macrophages pretreated with tolerance-inducing concentrations of either LPS or MPL. In macrophages rendered tolerant by pretreatment with LPS or MPL, a second group of inducible mRNAs (TNF-alpha, interleukin-1 beta, and D3) showed moderate suppression of response to secondary stimulation by LPS. The third category of inducible genes (TNFR-2 and D2) showed increased expression in macrophages pretreated with tolerance-inducing concentrations of either LPS or MPL. All of the LPS-inducible genes examined exhibited modest superinduction with less than tolerance-inducing concentrations of either stimulus, suggesting a priming effect of these adjuvants at low concentration. The differential behavior of the members of this panel of endotoxin-responsive genes thus offers insight into molecular events associated with acquisition of transient tolerance to LPS. PMID:8388859

  17. Dynamic distributions of tumor necrosis factor-alpha and its receptors in the red nucleus of rats with spared nerve injury.

    PubMed

    Wang, Jing; Ding, Cui-Ping; Yu, Jing; Zeng, Xiao-Yan; Han, Shui-Ping; Wang, Jun-Yang

    2016-08-01

    Previous studies have demonstrated that tumor necrosis factor-alpha (TNF-α) in the red nucleus (RN) plays a facilitated role in the development of neuropathic pain, and its effect is transmitted through TNF-α receptor (TNFR) subtypes 1 and 2. Here, the dynamic distributions of TNF-α and TNFRs in the RN of rats with spared nerve injury (SNI) were investigated. Western blot analysis and immunofluorescence staining indicated that TNF-α was hardly expressed in the RN of normal rats but significantly increased at 1 week and peaked at 2 weeks after SNI. Neurons and oligodendrocytes showed TNF-α expression at both 1 week and 2 weeks after SNI, while astrocytes and microglia produced TNF-α later than neurons and oligodendrocytes starting at 2 weeks after SNI. TNFR1 was constitutively expressed in the RN of normal rats and significantly enhanced at 2 weeks but not 1 week after SNI; it was mainly localized in neurons, oligodendrocytes and microglia. Astrocytes were not immunopositive for TNFR1 under normal conditions and at 1 week after injury, but small amounts of astrocytes showed TNFR1 expression at 2 weeks after SNI. A low level of TNFR2 was expressed in the RN of normal rats, but it was significantly increased at 1 week and 2 weeks after SNI and localized in neurons and all three types of glia. These findings suggest that neurons and three types of glia in the RN all contribute to TNF-α production and participate in the initiation and/or maintenance of neuropathic pain induced by SNI. TNF-α exerts its effects in different types of cells maybe through different receptors, TNFR1 and/or TNFR2, in the different stages of neuropathic pain.

  18. Human macrophage inflammatory protein-3alpha/CCL20/LARC/Exodus/SCYA20 is transcriptionally upregulated by tumor necrosis factor-alpha via a non-standard NF-kappaB site.

    PubMed

    Harant, H; Eldershaw, S A; Lindley, I J

    2001-12-14

    The 5'-flanking sequences of the human macrophage inflammatory protein-3alpha/CCL20 gene were cloned and transfected into G-361 human melanoma cells in a luciferase reporter construct. Tumor necrosis factor-alpha (TNF-alpha) treatment stimulated luciferase expression, and promoter truncations demonstrated that TNF-alpha inducibility is conferred by a region between nt -111 and -77, which contains a non-standard nuclear factor-kappaB (NF-kappaB) binding site. The requirement for NF-kappaB was demonstrated as follows: (i) mutations in this NF-kappaB site abrogated TNF-alpha responsiveness; (ii) TNF-alpha activated a construct containing two copies of the CCL20 NF-kappaB binding site; (iii) overexpression of NF-kappaB p65 activated the CCL20 promoter; (iv) NF-kappaB from nuclear extracts of TNF-alpha-stimulated cells bound specifically to this NF-kappaB site.

  19. Specific vaginal lactobacilli suppress the inflammation induced by lipopolysaccharide stimulation through downregulation of toll-like receptor 4 expression in human embryonic intestinal epithelial cells

    PubMed Central

    TOBITA, Keisuke; WATANABE, Itsuki; SAITO, Masanori

    2016-01-01

    Vaginal lactobacilli (VLB) spread from the mother to the infant during vaginal delivery. However, the effects of VLB on infant intestinal function remain unclear. We investigated the probiotic function and immune effects of VLB on the human embryonic intestinal epithelial cell line INT-407. VLB survived artificial gastric juice and adhered to INT-407 cells. Exposure of INT-407 cells to VLB attenuated both the lipopolysaccharide (LPS)-induced stimulation of interleukin-8 and tumor necrosis factor alpha production and the LPS-stimulated upregulation of TLR4 expression. These results suggest that specific VLB suppresses the inflammation induced by LPS stimulation through downregulation of TLR4 expression in human embryonic intestinal epithelial cells. PMID:28243550

  20. Ulstrastructural Localization of Tumour Necrosis Factor-Alpha

    DTIC Science & Technology

    1994-01-01

    secretory granules of all eosinophils , neutrophils and monocytes in the bone marrow, spleen, lung and the proximal intestine. TNF was also observed in the...TNF in the secretory granules of all 0 others, that endotoxin is not a requirement for septic neutrophils, eosinophils and monocytes examined. TNF shock...endotoxin-induced TNF appearance of TNF in the secretory granules of all might have been secreted by the smooth muscle cells with eosinophils

  1. Role of tumor necrosis factor-alpha in zebrafish retinal neurogenesis and myelination

    PubMed Central

    Lei, Xu-Dan; Sun, Yan; Cai, Shi-Jiao; Fang, Yang-Wu; Cui, Jian-Lin; Li, Yu-Hao

    2016-01-01

    AIM To investigate the role of tumor necrosis factor-alpha (TNF-α) in zebrafish retinal development and myelination. METHODS Morpholino oligonucleotides (MO), which are complementary to the translation start site of the wild-type embryonic zebrafish TNF-α mRNA sequence, were synthesized and injected into one- to four-cell embryos. The translation blocking specificity was verified by Western blotting using an anti-TNF-α antibody, whole-mount in situ hybridization using a hepatocyte-specific mRNA probe ceruloplasmin (cp), and co-injection of TNF-α MO and TNF-α mRNA. An atonal homolog 7 (atoh7) mRNA probe was used to detect neurogenesis onset. The retinal neurodifferentiation was analyzed by immunohistochemistry using antibodies Zn12, Zpr1, and Zpr3 to label ganglion cells, cones, and rods, respectively. Myelin basic protein (mbp) was used as a marker to track and observe the myelination using whole-mount in situ hybridization. RESULTS Targeted knockdown of TNF-α resulted in specific suppression of TNF-α expression and a severely underdeveloped liver. The co-injection of TNF-α MO and mRNA rescued the liver development. Retinal neurogenesis in TNF-α morphants was initiated on time. The retina was fully laminated, while ganglion cells, cones, and rods were well differentiated at 72 hours post-fertilization (hpf). mbp was expressed in Schwann cells in the lateral line nerves and cranial nerves from 3 days post-fertilization (dpf) as well as in oligodendrocytes linearly along the hindbrain bundles and the spinal cord from 4 dpf, which closely resembled its endogenous profile. CONCLUSION TNF-α is not an essential regulator for retinal neurogenesis and optic myelination. PMID:27366683

  2. Effect of Tumour Necrosis Factor-Alpha on Estrogen Metabolic Pathways in Breast Cancer Cells

    PubMed Central

    Kamel, Marwa; Shouman, Samia; El-Merzebany, Mahmoud; Kilic, Gokhan; Veenstra, Timothy; Saeed, Muhammad; Wagih, Mohamed; Diaz-Arrastia, Concepcion; Patel, Deepa; Salama, Salama

    2012-01-01

    Tumor necrosis factor-alpha (TNF-α) is a proinflammatory cytokine that has been linked to breast cancer development. Estrogen metabolic pathway is also involved in breast carcinogenesis and DNA adducts formation. In this study we investigated the effect of TNF-α on the estrogen metabolic pathway in MCF-7, a breast cancer cell line. Capillary liquid chromatography/mass spectrometry (LC/MS) and High performance liquid chromatography (HPLC) were used for analysis of estrogen metabolites and estrogen-DNA adducts levels respectively. Reporter gene assay, Real time reverse transcription polymerase chain reaction (real time RT-PCR) and Western blot were used to assess the expression of estrogen metabolizing genes and enzymes. TNF-α significantly increased the total EM and decreased the estrone (E1) / 17-β estradiol (E2) ratio. Moreover, it altered the expression of genes and enzymes involved in E2 activation and deactivation pathways e.g. Cytochrome P-450 1A1 (CYP1A1), Cytochrome P-450 1B1 (CYP1B1), Catechol-O-methyl transferase (COMT) and Nicotinamide adenine dinucleotide phosphate-quinone oxidoreductase 1 (NQO1). In addition, there were increased levels of some catechol estrogens e.g. 4-hydroxy-estrone (4-OHE1) and 2-hydroxyestradiol (2-OHE2) with decreased levels of methylated catechols e.g. 2-methoxy estradiol (2-MeOE2). DNA adducts especially 4-OHE1-[2]-1-N3 Adenine was significantly increased. TNF-α directs the estrogen metabolism into more hormonally active and carcinogenic products in MCF-7. This may implicate a new possible explanation for inflammation associated breast cancer. PMID:22866165

  3. Characterization of the rat transforming growth factor alpha gene and identification of promoter sequences.

    PubMed Central

    Blasband, A J; Rogers, K T; Chen, X R; Azizkhan, J C; Lee, D C

    1990-01-01

    We have determined the complete nucleotide sequence of rat transforming growth factor alpha (TGF alpha) mRNA and characterized the six exons that encode this transcript. These six exons span approximately 85 kilobases of genomic DNA, with exons 1 to 3 separated by particularly large introns. What had previously been thought to represent a species-specific difference in the size of the TGF alpha precursor (proTGF alpha) is now shown to be due to microheterogeneity in the splicing of exons 2 and 3. This results from a tandem duplication of the acceptor CAG and gives rise to two alternate forms (159 and 160 amino acids) of the integral membrane precursor. Exon 6, which encodes the 3' untranslated region of TGF alpha mRNA, also encodes, on the opposite strand, a small (approximately 200-nucleotide) transcript whose sequence predicts an open reading frame of 51 amino acids. Expression of this latter transcript does not appear to be coregulated with that of TGF alpha mRNA. Primer extension and S1 nuclease analyses of authentic TGF alpha transcripts revealed two major and multiple minor 5' ends which span more than 200 base pairs of DNA in a G + C-rich region that lacks canonical CCAAT or TATA sequences. The 5' ends of six independently derived cDNAs localized to five different sites in this same region. Restriction fragments that overlap these transcription start sites and extend approximately 300 base pairs in the 5' direction faithfully promote transcription in vitro with HeLa cell nuclear extracts. In addition, they direct the expression of the bacterial chloramphenicol acetyltransferase gene in transient-transfection assays. Images PMID:2325647

  4. Pseudomonas aeruginosa and tumor necrosis factor-alpha attenuate Clara cell secretory protein promoter function.

    PubMed

    Harrod, Kevin S; Jaramillo, Richard J

    2002-02-01

    The Clara cell secretory protein (CCSP, also CC-10/uterglobin) is a 16-kD homodimeric protein abundantly expressed in the airways of mammals. Although the molecular function is unknown, gene-targeting studies indicate CCSP as a regulator of lung inflammation following acute respiratory infection or injury. CCSP is decreased in the lungs of mice following acute Pseudomonas aeruginosa (P.a.) infection. In the present study, the role of decreased promoter function in the regulation of CCSP by P.a. was assessed using an in vitro co-culture system and in vivo studies of transgenic mice. CCSP promoter activity in lung epithelial cells was markedly decreased by P.a. or tumor necrosis factor-alpha (TNF-alpha) in a dose-dependent manner. Regulation of CCSP promoter function by either P.a. or TNF-alpha was localized to the proximal 166 bp flanking region of the CCSP promoter activity. Decreased regulation of the CCSP promoter by P.a. or TNF-alpha was specific to CCSP, as human surfactant protein D (SP-D) promoter activity was unaffected or increased by P.a. or TNF-alpha, respectively. A neutralizing antibody against human TNF-alpha was able to reverse both the TNF-alpha- mediated as well as P.a.-mediated decrease in CCSP promoter function in lung epithelial cells. TNF-alpha secretion by lung epithelial cells coincided with the decrease in CCSP promoter function following P.a. administration. Using a transgenic mouse model, P.a. administration to the lung markedly attenuated CCSP promoter-conferred gene expression in vivo. The attenuation of CCSP promoter activity in lung epithelial cells by P.a. involves, in part, autocrine/paracrine secretion of TNF-alpha, which in turn regulates CCSP transcription through cis-active elements in the proximal promoter region.

  5. The role of transforming growth factor alpha in rat craniofacial development and chondrogenesis.

    PubMed

    Huang, L; Solursh, M; Sandra, A

    1996-08-01

    To explore the possible role of transforming growth factor alpha (TGF-alpha) in craniofacial development, its expression in the craniofacial region of rat embryos from embryonic day (d) 9 to d 20 was examined by in situ hybridisation and immunostaining. The TGF-alpha transcripts were first detected in the neural fold of embryonic d 9 and 10 embryos. In the craniofacial region, the TGF-alpha transcripts were not detected until embryonic d 16 in mesenchyme surrounding the olfactory bulb, within the olfactory bulb, the nasal capsule, vomeronasal organ, and vibrissal follicle. In addition, TGF-alpha message was detected in mesenchyme in the vicinity of Meckel's cartilage, and in the dental epithelium and lamina. This expression pattern of TGF-alpha transcripts persisted until embryonic d 17 but disappeared by d 18. The presence of TGF-alpha protein largely coincided with TGF-alpha message although, unlike the message, it persisted throughout later embryogenesis in the craniofacial region. The possible function of TGF-alpha in chondrogenesis was explored by employing the micromass culture technique. Cartilage nodule formation in mesenchymal cells cultured from rat mandibles in the presence of TGF-alpha was significantly inhibited. This inhibitory effect of TGF-alpha on chondrogenesis was reversed by addition of antibody against the EGF receptor, which crossreacts with the TGF-alpha receptor. The inhibitory effect of TGF-alpha on chondrogenesis in vitro was further confirmed by micromass culture using mesenchymal cells from rat embryonic limb bud. Taken together, these results demonstrate the involvement of TGF-alpha in chondrogenesis during embryonic development, possibly by way of a specific inhibition of cartilage formation from mesenchymal precursor cells.

  6. The role of transforming growth factor alpha in rat craniofacial development and chondrogenesis.

    PubMed Central

    Huang, L; Solursh, M; Sandra, A

    1996-01-01

    To explore the possible role of transforming growth factor alpha (TGF-alpha) in craniofacial development, its expression in the craniofacial region of rat embryos from embryonic day (d) 9 to d 20 was examined by in situ hybridisation and immunostaining. The TGF-alpha transcripts were first detected in the neural fold of embryonic d 9 and 10 embryos. In the craniofacial region, the TGF-alpha transcripts were not detected until embryonic d 16 in mesenchyme surrounding the olfactory bulb, within the olfactory bulb, the nasal capsule, vomeronasal organ, and vibrissal follicle. In addition, TGF-alpha message was detected in mesenchyme in the vicinity of Meckel's cartilage, and in the dental epithelium and lamina. This expression pattern of TGF-alpha transcripts persisted until embryonic d 17 but disappeared by d 18. The presence of TGF-alpha protein largely coincided with TGF-alpha message although, unlike the message, it persisted throughout later embryogenesis in the craniofacial region. The possible function of TGF-alpha in chondrogenesis was explored by employing the micromass culture technique. Cartilage nodule formation in mesenchymal cells cultured from rat mandibles in the presence of TGF-alpha was significantly inhibited. This inhibitory effect of TGF-alpha on chondrogenesis was reversed by addition of antibody against the EGF receptor, which crossreacts with the TGF-alpha receptor. The inhibitory effect of TGF-alpha on chondrogenesis in vitro was further confirmed by micromass culture using mesenchymal cells from rat embryonic limb bud. Taken together, these results demonstrate the involvement of TGF-alpha in chondrogenesis during embryonic development, possibly by way of a specific inhibition of cartilage formation from mesenchymal precursor cells. Images Fig. 1 Fig. 2 Fig. 3 Fig. 4 Fig. 5 Fig. 6 Fig. 7 Fig. 9 PMID:8771398

  7. Mitochondrial Carrier Homolog 2 Is a Target of tBID in Cells Signaled To Die by Tumor Necrosis Factor Alpha

    PubMed Central

    Grinberg, Michal; Schwarz, Michal; Zaltsman, Yehudit; Eini, Tzipi; Niv, Hagit; Pietrokovski, Shmuel; Gross, Atan

    2005-01-01

    BID, a proapoptotic BCL-2 family member, plays an essential role in the tumor necrosis factor alpha (TNF-α)/Fas death receptor pathway in vivo. Activation of the TNF-R1 receptor results in the cleavage of BID into truncated BID (tBID), which translocates to the mitochondria and induces the activation of BAX or BAK. In TNF-α-activated FL5.12 cells, tBID becomes part of a 45-kDa cross-linkable mitochondrial complex. Here we describe the biochemical purification of this complex and the identification of mitochondrial carrier homolog 2 (Mtch2) as part of this complex. Mtch2 is a conserved protein that is similar to members of the mitochondrial carrier protein family. Our studies with mouse liver mitochondria indicate that Mtch2 is an integral membrane protein exposed on the surface of mitochondria. Using blue-native gel electrophoresis we revealed that in viable FL5.12 cells Mtch2 resides in a protein complex of ca. 185 kDa and that the addition of TNF-α to these cells leads to the recruitment of tBID and BAX to this complex. Importantly, this recruitment was partially inhibited in FL5.12 cells stably expressing BCL-XL. These results implicate Mtch2 as a mitochondrial target of tBID and raise the possibility that the Mtch2-resident complex participates in the mitochondrial apoptotic program. PMID:15899861

  8. Tumor Necrosis Factor Alpha Inhibits L-Type Ca(2+) Channels in Sensitized Guinea Pig Airway Smooth Muscle through ERK 1/2 Pathway.

    PubMed

    Reyes-García, Jorge; Flores-Soto, Edgar; Solís-Chagoyán, Héctor; Sommer, Bettina; Díaz-Hernández, Verónica; García-Hernández, Luz María; Montaño, Luis M

    2016-01-01

    Tumor necrosis factor alpha (TNF-α) is a potent proinflammatory cytokine that plays a significant role in the pathogenesis of asthma by inducing hyperresponsiveness and airway remodeling. TNF-α diminishes the L-type voltage dependent Ca(2+) channel (L-VDCC) current in cardiac myocytes, an observation that seems paradoxical. In guinea pig sensitized tracheas KCl responses were lower than in control tissues. Serum from sensitized animals (Ser-S) induced the same phenomenon. In tracheal myocytes from nonsensitized (NS) and sensitized (S) guinea pigs, an L-VDCC current (ICa) was observed and diminished by Ser-S. The same decrease was detected in NS myocytes incubated with TNF-α, pointing out that this cytokine might be present in Ser-S. We observed that a small-molecule inhibitor of TNF-α (SMI-TNF) and a TNF-α receptor 1 (TNFR1) antagonist (WP9QY) reversed ICa decrease induced by Ser-S in NS myocytes, confirming the former hypothesis. U0126 (a blocker of ERK 1/2 kinase) also reverted the decrease in ICa. Neither cycloheximide (a protein synthesis inhibitor) nor actinomycin D (a transcription inhibitor) showed any effect on the TNF-α-induced ICa reduction. We found that CaV1.2 and CaV1.3 mRNA and proteins were expressed in tracheal myocytes and that sensitization did not modify them. In cardiac myocytes, ERK 1/2 phosphorylates two sites of the L-VDCC, augmenting or decreasing ICa; we postulate that, in guinea pig tracheal smooth muscle, TNF-α diminishes ICa probably by phosphorylating the L-VDCC site that reduces its activity through the ERK1/2 MAP kinase pathway.

  9. Tumor Necrosis Factor Alpha Inhibits L-Type Ca2+ Channels in Sensitized Guinea Pig Airway Smooth Muscle through ERK 1/2 Pathway

    PubMed Central

    Reyes-García, Jorge; Flores-Soto, Edgar; Solís-Chagoyán, Héctor; Sommer, Bettina; Díaz-Hernández, Verónica; García-Hernández, Luz María

    2016-01-01

    Tumor necrosis factor alpha (TNF-α) is a potent proinflammatory cytokine that plays a significant role in the pathogenesis of asthma by inducing hyperresponsiveness and airway remodeling. TNF-α diminishes the L-type voltage dependent Ca2+ channel (L-VDCC) current in cardiac myocytes, an observation that seems paradoxical. In guinea pig sensitized tracheas KCl responses were lower than in control tissues. Serum from sensitized animals (Ser-S) induced the same phenomenon. In tracheal myocytes from nonsensitized (NS) and sensitized (S) guinea pigs, an L-VDCC current (ICa) was observed and diminished by Ser-S. The same decrease was detected in NS myocytes incubated with TNF-α, pointing out that this cytokine might be present in Ser-S. We observed that a small-molecule inhibitor of TNF-α (SMI-TNF) and a TNF-α receptor 1 (TNFR1) antagonist (WP9QY) reversed ICa decrease induced by Ser-S in NS myocytes, confirming the former hypothesis. U0126 (a blocker of ERK 1/2 kinase) also reverted the decrease in ICa. Neither cycloheximide (a protein synthesis inhibitor) nor actinomycin D (a transcription inhibitor) showed any effect on the TNF-α-induced ICa reduction. We found that CaV1.2 and CaV1.3 mRNA and proteins were expressed in tracheal myocytes and that sensitization did not modify them. In cardiac myocytes, ERK 1/2 phosphorylates two sites of the L-VDCC, augmenting or decreasing ICa; we postulate that, in guinea pig tracheal smooth muscle, TNF-α diminishes ICa probably by phosphorylating the L-VDCC site that reduces its activity through the ERK1/2 MAP kinase pathway. PMID:27445440

  10. Involvement of interleukin-8, vascular endothelial growth factor, and basic fibroblast growth factor in tumor necrosis factor alpha-dependent angiogenesis.

    PubMed Central

    Yoshida, S; Ono, M; Shono, T; Izumi, H; Ishibashi, T; Suzuki, H; Kuwano, M

    1997-01-01

    Tumor necrosis factor alpha (TNF-alpha) is a macrophage/monocyte-derived polypeptide which modulates the expression of various genes in vascular endothelial cells and induces angiogenesis. However, the underlying mechanism by which TNF-alpha mediates angiogenesis is not completely understood. In this study, we assessed whether TNF-alpha-induced angiogenesis is mediated through TNF-alpha itself or indirectly through other TNF-alpha-induced angiogenesis-promoting factors. Cellular mRNA levels of interleukin-8 (IL-8), vascular endothelial growth factor (VEGF), basic fibroblast growth factor (bFGF), and their receptors were increased after the treatment of human microvascular endothelial cells with TNF-alpha (100 U/ml). TNF-alpha-dependent tubular morphogenesis in vascular endothelial cells was inhibited by the administration of anti-IL-8, anti-VEGF, and anti-bFGF antibodies, and coadministration of all three antibodies almost completely abrogated tubular formation. Moreover, treatment with Sp1, NF-kappaB, and c-Jun antisense oligonucleotides inhibited TNF-alpha-dependent tubular morphogenesis by microvascular endothelial cells. Administration of a NF-kappaB antisense oligonucleotide almost completely inhibited TNF-alpha-dependent IL-8 production and partially abrogated TNF-alpha-dependent VEGF production, and an Sp1 antisense sequence partially inhibited TNF-alpha-dependent production of VEGF. A c-Jun antisense oligonucleotide significantly inhibited TNF-alpha-dependent bFGF production but did not affect the production of IL-8 and VEGF. Administration of an anti-IL-8 or anti-VEGF antibody also blocked TNF-alpha-induced neovascularization in the rabbit cornea in vivo. Thus, angiogenesis by TNF-alpha appears to be modulated through various angiogenic factors, both in vitro and in vivo, and this pathway is controlled through paracrine and/or autocrine mechanisms. PMID:9199336

  11. Interleukin-2 enhances the production of tumor necrosis factor-alpha in activated B-type chronic lymphocytic leukemia (B-CLL) cells.

    PubMed

    Larsson, L G; Carlsson, M; Schena, M; Lantz, M; Caligaris-Cappio, F; Nilsson, K

    1993-02-01

    Tumor necrosis factor-alpha (TNF-alpha) has recently been implicated as a regulator growth and differentiation of normal and malignant B cells. We utilized a selected clone (I-83) of primary resting B-type chronic lymphocytic leukemia (B-CLL) cells, inducible to activation, growth and differentiation in vitro, as a model system to study the possible role of TNF-alpha as an autocrine growth factor for such cells. Our results show that unstimulated I-83 B-CLL cells produced a low level of TNF-alpha mRNA, as shown by Northern blot analysis, and cytoplasmic TNF-alpha, determined in individual cells by immunocytochemistry. Secreted TNF-alpha could, however, not be detected in the medium by ELISA. TNF-alpha synthesis and secretion was, however, induced to high levels by stimulation of the B-CLL cells with interleukin-2 (IL-2) after activation by 12-O-tetradecanoylphorbol-13-acetate (TPA) or Staphylococcus aureus Cowan strain I (SAC) and B-cell stimulatory factor-MP6 (thioredoxin). A moderate increase in TNF-alpha secretion was also induced by TPA or IL-2 alone. IL-4 did not have any major effects on the production of TNF-alpha in activated cells, but inhibited the IL-2-induced production of TNF-alpha in SAC-activated cells. The cell surface expression of TNF-alpha receptors (TNF-R), as determined by binding assay using 125I-labelled recombinant TNF-alpha (rTNF-alpha), was also induced after SAC or TPA activation, but shed receptors (TNF-binding proteins) were only observed after TPA activation. Exogenously added rTNF-alpha in combination with TPA or SAC induced a high level of DNA synthesis in I-83 B-CLL cells. The increased endogenous production and secretion of TNF-alpha during induced growth stimulation, the induced expression of TNF-R, and the mitogenic effect of TNF-alpha on activated B-CLL cells raise the question whether TNF-alpha may function as an autocrine co-stimulator of B-CLL cell growth as recently suggested. anti-TNF-alpha and anti-TNF-R antibodies

  12. Inducible expression of vascular cell adhesion molecule-1 by vascular smooth muscle cells in vitro and within rabbit atheroma.

    PubMed Central

    Li, H.; Cybulsky, M. I.; Gimbrone, M. A.; Libby, P.

    1993-01-01

    Vascular cell adhesion molecule-1 (VCAM-1), a mononuclear leukocyte adhesion molecule, is expressed in cultured vascular endothelial cells activated by cytokines and is induced in rabbit aortic endothelium in vivo within 1 week after initiation of an atherogenic diet. We now demonstrate that vascular smooth muscle cells can also express VCAM-1 in rabbit atherosclerotic lesions in vivo and in response to cytokines in vitro. Immunohistochemical staining of aortas from rabbits fed a 0.3% cholesterol-containing diet revealed that a portion of smooth muscle cells within intimal foam cell-rich lesions expressed VCAM-1. The intimal VCAM-1-expressing cells localized predominantly in regions above the internal elastic lamina. These VCAM-1-positive cells had the typical spindle shape of smooth muscle cells but had reduced alpha-actin expression in comparison to normal medial smooth muscle cells, and did not bear markers for endothelium, macrophages, and T cells. In culture, rabbit aortic smooth muscle cells expressed VCAM-1 mRNA and protein in a time- and concentration-dependent fashion when exposed to interferon-gamma or Gram-negative bacterial lipopolysaccharide. Cultured human vascular smooth muscle cells also expressed VCAM-1 mRNA and protein in response to lipopolysaccharide, interferon-gamma, and interleukin-4. The monokines interleukin-1 alpha and tumor necrosis factor-alpha did not induce VCAM-1 expression in either rabbit or human vascular smooth muscle cells. Inducible VCAM-1 expression by vascular smooth muscle cells in vivo during hypercholesterolemia and in vitro in response to certain cytokines suggests a broader range of VCAM-1 functions in vascular biology than heretofore appreciated. Images Figure 1 Figure 2 Figure 3 Figure 4 PMID:7504883

  13. Astrocyte-targeted expression of interleukin-6 protects the central nervous system during neuroglial degeneration induced by 6-aminonicotinamide.

    PubMed

    Penkowa, Milena; Camats, Jordi; Hadberg, Hanne; Quintana, Albert; Rojas, Santiago; Giralt, Mercedes; Molinero, Amalia; Campbell, Iain L; Hidalgo, Juan

    2003-08-15

    6-aminonicotinamide (6-AN) is a niacin antagonist, which leads to degeneration of gray matter astrocytes mainly in the brainstem. We have examined the role of interleukin-6 (IL-6) in this degenerative process by using transgenic mice with astrocyte-targeted IL-6 expression (GFAP-IL6 mice). This study demonstrates that transgenic IL-6 expression significantly increases the 6-AN-induced inflammatory response of reactive astrocytes, microglia/macrophages, and lymphocytes in the brainstem. Also, IL-6 induced significant increases in proinflammatory cytokines IL-1, IL-12, and tumor necrosis factor-alpha as well as growth factors basic fibroblast growth factor (bFGF), transforming growth factor-beta, neurotrophin-3, angiopoietin, vascular endothelial growth factor, and the receptor for bFGF. In accordance, angiogenesis was increased in GFAP-IL6 mice relative to controls after 6-AN. Moreover, oxidative stress and apoptotic cell death were significantly reduced by transgenic IL-6 expression. IL-6 is also a major inducer in the CNS of metallothionein I and II (MT-I+II), which were significantly increased in the GFAP-IL6 mice. MT-I+II are antioxidants and neuroregenerative factors in the CNS, so increased MT-I+II levels in GFAP-IL6 mice could contribute to the reduction of oxidative stress and cell death in these mice.

  14. Borage oil reduction of rheumatoid arthritis activity may be mediated by increased cAMP that suppresses tumor necrosis factor-alpha.

    PubMed

    Kast, R E

    2001-11-01

    Recent double blind studies have shown some benefit of borage oil in treatment of rheumatoid arthritis. Tumor necrosis factor-alpha has been shown to be a central mediator of inflammatory and joint destructive processes in rheumatoid arthritis. In this paper, evidence from published research is reviewed that indicates gamma linolenic acid component of borage oil increases prostaglandin E levels that increase cAMP levels that in turn suppress tumor necrosis factor-alpha synthesis. If this biochemical path of borage oil is correct then (1) concomitant non-steroidal anti-inflammatory drug use would tend to undermine borage oil effects, and (2) borage oil would be contraindicated in pregnancy given the teratogenic and labor inducing effects of prostaglandin E agonists.

  15. Hemoglobin stimulates mononuclear leukocytes to release interleukin-8 and tumor necrosis factor alpha.

    PubMed

    McFaul, S J; Bowman, P D; Villa, V M; Gutierrez-Ibanez, M J; Johnson, M; Smith, D

    1994-11-01

    Incubation of human mononuclear leukocytes (MNL) with human stroma-free hemolysate (SFH), purified adult hemoglobin Ao (HbAo), and oxidized HbAo (METHb) caused MNL to release compounds into the supernate that mediated neutrophil (polymorphonuclear leukocytes, PMN) chemotaxis and PMN adherence to human umbilical vein endothelial cells (HUVEC). Chemotaxis and PMN adherence to HUVEC were reduced significantly when supernates were preincubated with neutralizing antibodies to interleukin-8 (IL-8) and tumor necrosis factor alpha (TNF-alpha), respectively, suggesting that IL-8 and TNF-alpha played significant roles in mediating these activities. Greatest chemotactic activity was observed in supernates of MNL treated with HbAo; while greatest PMN/endothelial cell (EC) adherence activity was observed in supernates of MNL treated with METHb. Furthermore, PMN/EC adherence activity was a function of METHb content in each hemoglobin solution. PMN chemotaxis, PMN adherence to HUVEC, and cytokine release increased as a function of increasing incubation time. Chemotactic activity was detected in HbAo-treated and METHb-treated MNL supernates after incubation for 6 hours and was maximal by 10 hours. IL-8 was detected in both HbAo and METHb-MNL supernates by 4 hours. PMN/EC adherence activity was detected in HbAo-MNL supernates at 10 hours and in METHb-MNL supernates at 4 hours. TNF-alpha was detected in METHb and HbAo-MNL supernates at 4 and 12 hours, respectively. These results suggest that hemoglobin solutions stimulate MNL to release IL-8 and TNF-alpha in quantities sufficient to induce PMN chemotaxis and PMN adherence to HUVEC. This is a US government work. There are no restrictions on its use.

  16. Isolated rat stomach ECL cells generate prostaglandin E(2) in response to interleukin-1 beta, tumor necrosis factor-alpha and bradykinin.

    PubMed

    Lindström, E; Lerner, U H; Håkanson, R

    2001-03-30

    The ECL cells control parietal cells by releasing histamine in their immediate vicinity. Gastrin and pituitary adenylate cyclase-activating peptide (PACAP) stimulate histamine secretion from isolated ECL cells, while somatostatin and galanin inhibit stimulated secretion. Prostaglandin E2 and related prostaglandins likewise suppress ECL-cell histamine secretion. Conceivably, that is how they inhibit acid secretion. In the present study, we examined if prostaglandin E2 can be generated by isolated ECL cells. Rat stomach ECL cells were purified (>90% purity) by counterflow elutriation and gradient centrifugation and cultured for 48 h. ECL cell stimulants (gastrin and PACAP) and inflammatory agents (interleukin-1 beta, tumor necrosis factor-alpha and bradykinin) were tested for their ability to induce prostaglandin E2 accumulation (24-h incubation), measured by radioimmunoassay. Gastrin and PACAP did not affect prostaglandin E2 accumulation but interleukin-1 beta (300 pg/ml), tumor necrosis factor-alpha (10 ng/ml) and bradykinin (1 microM) induced a 2- to 3-fold increase in the amount of prostaglandin E2 accumulated. While the combination of interleukin-1 beta and bradykinin induced a 9-fold increase, the combination interleukin-1 beta+tumor necrosis factor-alpha and bradykinin + tumor necrosis factor-alpha induced additive effects only. The combination of interleukin-1 beta + tumor necrosis factor-alpha + bradykinin did not induce a greater effect than interleukin-1 beta + bradykinin. The effect of interleukin-1 beta + bradykinin was abolished by adding 10 nM hydrocortisone (suppressing phospholipase A2 and cyclooxygenase) or 1 microM indomethacin (inhibiting cyclooxygenase). Incubating ECL cells in the presence of interleukin-1 beta+bradykinin for 24 h reduced their ability to secrete histamine in response to gastrin. The inhibitory effect was reversed by 1 microM indomethacin. Also, increasing the concentrations of hydrocortisone in the medium resulted in an

  17. Natural soluble interleukin-15Ralpha is generated by cleavage that involves the tumor necrosis factor-alpha-converting enzyme (TACE/ADAM17).

    PubMed

    Budagian, Vadim; Bulanova, Elena; Orinska, Zane; Ludwig, Andreas; Rose-John, Stefan; Saftig, Paul; Borden, Ernest C; Bulfone-Paus, Silvia

    2004-09-24

    This study shows that the high affinity alpha-chain of the interleukin (IL)-15 receptor exists not only in membrane-anchored but also in soluble form. Soluble IL-15Ralpha (sIL-15Ralpha) can be detected in mouse sera and cell-conditioned media by enzyme-linked immunosorbent assay and by immunoprecipitation and Western blotting. This protein has a molecular mass of about 30 kDa because of the presence of a single N-glycosylation site, which is reduced to 26 kDa after N-glycosidase treatment. Transmembrane IL-15Ralpha is constitutively converted into its soluble form by proteolytic cleavage that involves tumor necrosis factor-alpha-converting enzyme (TACE), and this process is further enhanced by phorbol 12-myristate 13-acetate (PMA) stimulation. The hydroxamate GW280264X, which is capable of blocking TACE and the closely related disintegrin-like metalloproteinase 10 (ADAM10), effectively inhibited both spontaneous and PMA-inducible cleavage of IL-15Ralpha, whereas GI254023X, which preferentially blocks ADAM10, was ineffective. Overexpression of TACE but not ADAM10 in COS-7 cells enhanced the constitutive and PMA-inducible cleavage of IL-15Ralpha. Moreover, murine fibroblasts deficient in TACE but not ADAM10 expression exhibited a significant reduction in the spontaneous and inducible IL-15Ralpha shedding, whereas a reconstitution of TACE in these cells restored the release of sIL-15Ralpha, thereby suggesting that TACE-mediated proteolysis may represent a major mechanism for sIL-15Ralpha generation in mice. The existence of natural sIL-15Ralpha offers novel insights into the complex biology of IL-15 and envisages a new level for therapeutic intervention.

  18. Substance P Induces HO-1 Expression in RAW 264.7 Cells Promoting Switch towards M2-Like Macrophages

    PubMed Central

    Montana, Giovanna

    2016-01-01

    Substance P (SP) is a neuropeptide that mediates many physiological as well as inflammatory responses. Recently, SP has been implicated in the resolution of inflammation through induction of M2 macrophages phenotype. The shift between M1-like and M2-like, allowing the resolution of inflammatory processes, also takes place by means of hemeoxygenase-1 (HO-1). HO-1 is induced in response to oxidative stress and inflammatory stimuli and modulates the immune response through macrophages polarisation. SP induces HO-1 expression in human periodontal ligament (PDL), the latter potentially plays a role in cytoprotection. We demonstrated that SP promotes M2-like phenotype from resting as well as from M1 macrophages. Indeed, SP triggers the production of interleukine-10 (IL-10), interleukine-4 (IL-4) and arginase-1 (Arg1) without nitric oxide (NO) generation. In addition, SP increases HO-1 expression in a dose- and time-dependent manner. Here we report that SP, without affecting cell viability, significantly reduces the production of pro-inflammatory cytokines and enzymes, such as tumor necrosis factor-alpha (TNF-α), interleukine-6 (IL-6), inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2), and ameliorates migration and phagocytic properties in LPS-stimulated RAW 264.7 cells. M2-like conversion required retention of NF-κB p65 into the cytoplasm and HO-1 induced expression. Silencing of the HO-1 mRNA expression reversed the induction of pro-inflammatory cytokines in RAW 264.7 stimulated by LPS and down-regulated anti-inflammatory hallmarks of M2 phenotype. In conclusion, our data show that SP treatment might be associated with anti-inflammatory effects in LPS-stimulated RAW 264.7 cells by suppressing NF-κB activation and inducing HO-1 expression. PMID:27907187

  19. The aporphine alkaloid boldine induces adiponectin expression and regulation in 3T3-L1 cells.

    PubMed

    Yu, Bangning; Cook, Carla; Santanam, Nalini

    2009-10-01

    Adiponectin is an adipokine secreted by differentiated adipocytes. Clinical studies suggest a negative correlation between oxidative stress and adiponectin levels in patients with metabolic syndrome or cardiovascular disease. Natural compounds that can prevent oxidative stress mediated inhibition of adiponectin may be potentially therapeutic. Boldine, an aporphine alkaloid abundant in the medicinal plant Peumus boldus, is a powerful antioxidant. The current study demonstrates the effects of boldine on the expression of adiponectin and its regulators, CCAAT/enhancer binding protein-alpha (C/EBPalpha) and peroxisome proliferator-activated receptor (PPAR)-gamma, in 3T3-L1 cells. Differentiated 3T3-L1 adipocytes were exposed to either hydrogen peroxide (H(2)O(2)) (100 microM) or tumor necrosis factor-alpha (TNFalpha) (1 ng/mL) for 24 hours in the presence or absence of increasing concentrations of boldine (5-100 microM). Quantitative polymerase chain reaction showed that both the oxidants decreased the mRNA levels of adiponectin, PPARgamma, and C/EBPalpha to half of the control levels. Boldine, at all concentrations, counteracted the inhibitory effect of H(2)O(2) or TNFalpha and increased the expression of adiponectin and its regulators. The effect of boldine on adiponectin expression was biphasic, with the lower concentrations (5-25 microM) having a larger inductive effect compared to higher concentrations (50-100 microM). Boldine treatment alone in the absence of H(2)O(2) or TNFalpha was also able to induce adiponectin at the inductive phase of adipogenesis. Peroxisome proliferator response element-luciferase promoter transactivity analysis showed that boldine interacts with the PPAR response element and could potentially modulate PPAR responsive genes. Our results indicate that boldine is able to modulate the expression of adiponectin and its regulators in 3T3-L1 cells and has the potential to be beneficial in obesity-related cardiovascular disease.

  20. Inducible gene expression systems and plant biotechnology.

    PubMed

    Corrado, Giandomenico; Karali, Marianthi

    2009-01-01

    Plant biotechnology relies heavily on the genetic manipulation of crops. Almost invariantly, the gene of interest is expressed in a constitutive fashion, although this may not be strictly necessary for several applications. Currently, there are several regulatable expression systems for the temporal, spatial and quantitative control of transgene activity. These molecular switches are based on components derived from different organisms, which range from viruses to higher eukaryotes. Many inducible systems have been designed for fundamental and applied research and since their initial development, they have become increasingly popular in plant molecular biology. This review covers a broad number of inducible expression systems examining their properties and relevance for plant biotechnology in its various guises, from molecular breeding to pharmaceutical and industrial applications. For each system, we examine some advantages and limitations, also in relation to the strategy on which they rely. Besides being necessary to control useful genes that may negatively affect crop yield and quality, we discuss that inducible systems can be also used to increase public acceptance of GMOs, reducing some of the most common concerns. Finally, we suggest some directions and future developments for their further diffusion in agriculture and biotechnology.

  1. Expression of IP-10, a lipopolysaccharide- and interferon-gamma-inducible protein, in murine mesangial cells in culture.

    PubMed Central

    Gómez-Chiarri, M.; Hamilton, T. A.; Egido, J.; Emancipator, S. N.

    1993-01-01

    IP-10 is an early gene induced in multiple cell types by a variety of proinflammatory agents, notably interferons (IFNs) and lipopolysaccharide (LPS). To determine whether this protein might play a role in amplifying immune-mediated glomerular injury, we cultured mouse mesangial cells with several stimuli for various times. Increasing amounts of IFN-gamma (to 100 units/ml) elicited increasing levels of IP-10 messenger RNA (mRNA), sustained to 24 hours, but had no effect on tumor necrosis factor-alpha (TNF-alpha) mRNA. LPS induced transient IP-10 mRNA expression that peaked at 8 hours; TNF-alpha mRNA was also increased. TNF-alpha at doses up to 10 ng/ml and soluble immune complexes up to 150 micrograms/ml antibody evoked 3- to 5-fold increases in IP-10 mRNA expression, much less than the 30- to 70-fold increases seen with IFN-gamma and LPS. We conclude that IFN-gamma, LPS, and other agonists can amplify glomerular immune injury, perhaps via elevated expression of IP-10. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 PMID:8434640

  2. Evaluation of a nanotechnology-based approach to induce gene-expression in human THP-1 macrophages under inflammatory conditions.

    PubMed

    Bernal, Laura; Alvarado-Vázquez, Abigail; Ferreira, David Wilson; Paige, Candler A; Ulecia-Morón, Cristina; Hill, Bailey; Caesar, Marina; Romero-Sandoval, E Alfonso

    2017-02-01

    Macrophages orchestrate the initiation and resolution of inflammation by producing pro- and anti-inflammatory products. An imbalance in these mediators may originate from a deficient or excessive immune response. Therefore, macrophages are valid therapeutic targets to restore homeostasis under inflammatory conditions. We hypothesize that a specific mannosylated nanoparticle effectively induces gene expression in human macrophages under inflammatory conditions without undesirable immunogenic responses. THP-1 macrophages were challenged with lipopolysaccharide (LPS, 5μg/mL). Polyethylenimine (PEI) nanoparticles grafted with a mannose receptor ligand (Man-PEI) were used as a gene delivery method. Nanoparticle toxicity, Man-PEI cellular uptake rate and gene induction efficiency (GFP, CD14 or CD68) were studied. Potential immunogenic responses were evaluated by measuring the production of tumor necrosis factor-alpha (TNF-α), Interleukin (IL)-6 and IL-10. Man-PEI did not produce cytotoxicity, and it was effectively up-taken by THP-1 macrophages (69%). This approach produced a significant expression of GFP (mRNA and protein), CD14 and CD68 (mRNA), and transiently and mildly reduced IL-6 and IL-10 levels in LPS-challenged macrophages. Our results indicate that Man-PEI is suitable for inducing an efficient gene overexpression in human macrophages under inflammatory conditions with limited immunogenic responses. Our promising results set the foundation to test this technology to induce functional anti-inflammatory genes.

  3. Hyperbaric oxygen activates discoidin domain receptor 2 via tumour necrosis factor-alpha and the p38 MAPK pathway to increase vascular smooth muscle cell migration through matrix metalloproteinase 2.

    PubMed

    Shyu, Kou-Gi; Wang, Bao-Wei; Chang, Hang

    2009-04-01

    DDR2 (discoidin domain receptor 2) regulates collagen turnover mediated by SMCs (smooth muscle cells) in atherosclerosis. HBO (hyperbaric oxygen) has been used in medical practice; however, the molecular mechanism of the beneficial effects of HBO is poorly understood. Furthermore, the effect of HBO on DDR2 has not been reported previously. In the present study, we investigated the cellular and molecular mechanisms of DDR2 regulation by HBO in VSMCs (vascular SMCs). Cells were exposed to 2.5 ATA (atmosphere absolute) of oxygen in a hyperbaric chamber. DDR2 protein (3.63-fold) and mRNA (2.34-fold) expression were significantly increased after exposure to 2.5 ATA HBO for 1 h. Addition of SB203580 and p38 MAPK (mitogen-activated protein kinase) siRNA (small interfering RNA) 30 min before HBO inhibited the induction of DDR2 protein. HBO also significantly increased DNA-protein binding activity of Myc/Max. Addition of SB203580 and an anti-TNF-alpha (tumour necrosis factor-alpha) monoclonal antibody 30 min before HBO abolished the DNA-protein binding activity induced by HBO. HBO significantly increased the secretion of TNF-alpha from cultured VSMCs. Exogenous addition of TNF-alpha significantly increased DDR2 protein expression, whereas anti-TNF-alpha and anti-(TNF-alpha receptor) antibodies blocked the induction of DDR2 protein expression. HBO significantly increased VSMC migration and proliferation, whereas DDR2 siRNA inhibited the migration induced by HBO. HBO increased activated MMP2 (matrix metalloproteinase 2) protein expression, and DDR2 siRNA abolished the induction of activated MMP2 expression induced by HBO. In conclusion, HBO activates DDR2 expression in cultured rat VSMCs. HBO-induced DDR2 is mediated by TNF-alpha and at least in part through the p38 MAPK and Myc pathways.

  4. Human T-cell lymphotropic virus type 1-infected T lymphocytes impair catabolism and uptake of glutamate by astrocytes via Tax-1 and tumor necrosis factor alpha.

    PubMed

    Szymocha, R; Akaoka, H; Dutuit, M; Malcus, C; Didier-Bazes, M; Belin, M F; Giraudon, P

    2000-07-01

    Human T-cell lymphotropic virus type 1 (HTLV-1) is the causative agent of a chronic progressive myelopathy called tropical spastic paraparesis/HTLV-1-associated myelopathy (TSP/HAM). In this disease, lesions of the central nervous system (CNS) are associated with perivascular infiltration by lymphocytes. We and others have hypothesized that these T lymphocytes infiltrating the CNS may play a prominent role in TSP/HAM. Here, we show that transient contact of human or rat astrocytes with T lymphocytes chronically infected by HTLV-1 impairs some of the major functions of brain astrocytes. Uptake of extracellular glutamate by astrocytes was significantly decreased after transient contact with infected T cells, while the expression of the glial transporters GLAST and GLT-1 was decreased. In two-compartment cultures avoiding direct cell-to-cell contact, similar results were obtained, suggesting possible involvement of soluble factors, such as cytokines and the viral protein Tax-1. Recombinant Tax-1 and tumor necrosis factor alpha (TNF-alpha) decreased glutamate uptake by astrocytes. Tax-1 probably acts by inducing TNF-alpha, as the effect of Tax-1 was abolished by anti-TNF-alpha antibody. The expression of glutamate-catabolizing enzymes in astrocytes was increased for glutamine synthetase and decreased for glutamate dehydrogenase, the magnitudes of these effects being correlated with the level of Tax-1 transcripts. In conclusion, Tax-1 and cytokines produced by HTLV-1-infected T cells impair the ability of astrocytes to manage the steady-state level of glutamate, which in turn may affect neuronal and oligodendrocytic functions and survival.

  5. Intracellular reactive oxygen species as apparent modulators of heat-shock protein 27 (hsp27) structural organization and phosphorylation in basal and tumour necrosis factor alpha-treated T47D human carcinoma cells.

    PubMed Central

    Mehlen, P; Kretz-Remy, C; Briolay, J; Fostan, P; Mirault, M E; Arrigo, A P

    1995-01-01

    The small stress protein heat-shock protein 27 (hsp27) is an oligomeric phosphoprotein, constitutively expressed in most human cells, which enhances cellular resistance to tumour necrosis factor alpha (TNF alpha). This phenomenon correlates with dramatic changes in hsp27 cellular location, structural organization and phosphorylation. To gain a better understanding of the molecular mechanisms regulating these properties of hsp27, we investigated whether they were a consequence of the intracellular production of reactive oxygen species (ROS) generated by TNF alpha. Here, we report that, in T47D carcinoma cell lines, the rapid burst of intracellular ROS production and changes in hsp27 locale, structural organization and phosphoisoform composition induced by TNF alpha were abolished by the overexpression of the antioxidant enzyme seleno-glutathione peroxidase (GSHPx). These effects were greatly diminished when GSHPx-expressing cells were grown in the absence of selenium, a cofactor that is essential for seleno-GSHPx activity, indicating that they are directly linked to the increased GSHPx activity. Moreover, in growing T47D cells, GSHPx expression induced intracellular redistribution of hsp27 and decreased the phosphorylation of this protein without altering its pattern of oligomerization. In contrast, the heat-mediated phosphorylation of hsp27 was not altered by decreased intracellular ROS levels. Hence, in growing and TNF-treated cells, several hsp27 properties appear to be modulated by fluctuations in intracellular ROS levels. Images Figure 1 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7 PMID:8526844

  6. Transforming growth factor-beta 1 induces alpha-smooth muscle actin expression in granulation tissue myofibroblasts and in quiescent and growing cultured fibroblasts

    PubMed Central

    1993-01-01

    Granulation tissue fibroblasts (myofibroblasts) develop several ultrastructural and biochemical features of smooth muscle (SM) cells, including the presence of microfilament bundles and the expression of alpha-SM actin, the actin isoform typical of vascular SM cells. Myofibroblasts have been proposed to play a role in wound contraction and in retractile phenomena observed during fibrotic diseases. We show here that the subcutaneous administration of transforming growth factor- beta 1 (TGF beta 1) to rats results in the formation of a granulation tissue in which alpha-SM actin expressing myofibroblasts are particularly abundant. Other cytokines and growth factors, such as platelet-derived growth factor and tumor necrosis factor-alpha, despite their profibrotic activity, do not induce alpha-SM actin in myofibroblasts. In situ hybridization with an alpha-SM actin probe shows a high level of alpha-SM actin mRNA expression in myofibroblasts of TGF beta 1-induced granulation tissue. Moreover, TGF beta 1 induces alpha-SM actin protein and mRNA expression in growing and quiescent cultured fibroblasts and preincubation of culture medium containing whole blood serum with neutralizing antibodies to TGF beta 1 results in a decrease of alpha-SM actin expression by fibroblasts in replicative and non-replicative conditions. These results suggest that TGF beta 1 plays an important role in myofibroblast differentiation during wound healing and fibrocontractive diseases by regulating the expression of alpha-SM actin in these cells. PMID:8314838

  7. Hyperbaric oxygen treatment induces antioxidant gene expression.

    PubMed

    Godman, Cassandra A; Joshi, Rashmi; Giardina, Charles; Perdrizet, George; Hightower, Lawrence E

    2010-06-01

    Although the underlying molecular causes of aging are not entirely clear, hormetic agents like exercise, heat, and calorie restriction may generate a mild pro-oxidant stress that induces cell protective responses to promote healthy aging. As an individual ages, many cellular and physiological processes decline, including wound healing and reparative angiogenesis. This is particularly critical in patients with chronic non-healing wounds who tend to be older. We are interested in the potential beneficial effects of hyperbaric oxygen as a mild hormetic stress on human microvascular endothelial cells. We analyzed global gene expression changes in human endothelial cells following a hyperbaric exposure comparable to a clinical treatment. Our analysis revealed an upregulation of antioxidant, cytoprotective, and immediate early genes. This increase coincided with an increased resistance to a lethal oxidative stress. Our data indicate that hyperbaric oxygen can induce protection against oxidative insults in endothelial cells and may provide an easily administered hormetic treatment to help promote healthy aging.

  8. Endothelin-1 inhibits TNF alpha-induced iNOS expression in 3T3-F442A adipocytes.

    PubMed

    Mérial-Kieny, Christelle; Lonchampt, Michel; Cogé, Francis; Verwaerde, Patrick; Galizzi, Jean-Pierre; Boutin, Jean A; Lafontan, Max; Levens, Nigel; Galitzky, Jean; Félétou, Michel

    2003-07-01

    1. Endothelin-1 (ET-1) and tumor necrosis factor alpha (TNFalpha) by their action on adipocytes have been independently linked to the pathogenesis of insulino-resistance. In isolated adipocytes, TNFalpha induces the expression of the inducible nitric oxide synthase (iNOS). The purpose of the present work was, in the 3T3-F442A adipocyte cell line, to characterise TNFalpha-induced iNOS expression and to determine whether or not ET-1 could influence TNFalpha-induced iNOS expression and NO production. 2. In differentiated 3T3-F442A, treatment with TNFalpha (20 ng ml(-1)) induced the expression of a functional iNOS as demonstrated by nitrite assay, Western blot, reverse transcription-polymerase chain reaction and Northern blot analysis. TNFalpha-induced iNOS expression requires nuclear factor kappaB activation, but does not necessitate the activation of the PI-3 kinase/Akt and P38-MAP kinase pathways. 3. ET-1, but not ET-3, inhibited the TNFalpha-induced expression of iNOS protein and mRNA as well as nitrite production. The effects of ET-1 were blocked by a specific ETA (BQ123, pA(2) 7.4) but not by a specific ETB receptor antagonist (BQ788). 3T3-F442A adipocytes express the mRNAs for prepro-ET-1 and the ET-A receptor subtype, but not for the ET-B subtype. 4. The inhibitory effect of ET-1 was not affected by bisindolylmaleimide, SB 203580 or indomethacin, inhibitors of protein kinase C, p38-MAP kinase and cyclooxygenase, respectively, and was not associated with cAMP production. However, the effect of ET-1 was partially reversed by wortmannin, suggesting the involvement of PI3 kinase in the transduction signal of ET-1. 5. Differentiated 3T3-F442A adipocytes did not release ET-1 with or without exposure to TNFalpha, although the mRNA for preproET-1 was detected in both pre- and differentiated adipocytes. 6. Thus, these results confirm that adipocytes are a target for circulating ET-1 and demonstrate that the activation of the ETA receptor subtype can prevent TNFalpha-induced

  9. Oral N-acetylcysteine reduces bleomycin-induced lung damage and mucin Muc5ac expression in rats.

    PubMed

    Mata, M; Ruíz, A; Cerdá, M; Martinez-Losa, M; Cortijo, J; Santangelo, F; Serrano-Mollar, A; Llombart-Bosch, A; Morcillo, E J

    2003-12-01

    Oxidative stress is involved in the pathogenesis of pulmonary fibrosis, therefore antioxidants may be of therapeutic value. Clinical work indicates that N-acetylcysteine (NAC) may be beneficial in this disease. The activity of this antioxidant was examined on bleomycin-induced lung damage, mucus secretory cells hyperplasia and mucin Muc5ac gene expression in rats. NAC (3 mmol x kg(-1) x day(-1)) or saline was given orally to Sprague-Dawley rats for 1 week prior to a single intratracheal instillation of bleomycin (2.5 U x kg(-1)) and for 14 days postinstillation. NAC decreased collagen deposition in bleomycin-exposed rats (hydroxyproline content was 4,257+/-323 and 3,200+/-192 microg x lung(-1) in vehicle- and NAC-treated rats, respectively) and lessened the fibrotic area assessed by morphometric analysis. The bleomycin-induced increases in lung tumour necrosis factor-alpha and myeloperoxidase activity were reduced by NAC treatment. The numbers of mucus secretory cells in airway epithelium, and the Muc5ac messenger ribonucleic acid and protein expression, were markedly augmented in rats exposed to bleomycin. These changes were significantly reduced in NAC-treated rats. These results indicate that bleomycin increases the number of airway secretory cells and their mucin production, and that oral N-acetylcysteine improved pulmonary lesions and reduced the mucus hypersecretion in the bleomycin rat model.

  10. TNF-{alpha} promotes human retinal pigment epithelial (RPE) cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression through activation of Akt/mTORC1 signaling

    SciTech Connect

    Wang, Cheng-hu; Cao, Guo-Fan; Jiang, Qin; Yao, Jin

    2012-08-17

    Highlights: Black-Right-Pointing-Pointer TNF-{alpha} induces MMP-9 expression and secretion to promote RPE cell migration. Black-Right-Pointing-Pointer MAPK activation is not critical for TNF-{alpha}-induced MMP-9 expression. Black-Right-Pointing-Pointer Akt and mTORC1 signaling mediate TNF-{alpha}-induced MMP-9 expression. Black-Right-Pointing-Pointer SIN1 knockdown showed no significant effect on MMP-9 expression by TNF-{alpha}. -- Abstract: Tumor necrosis factor-alpha (TNF-{alpha}) promotes in vitro retinal pigment epithelial (RPE) cell migration to initiate proliferative vitreoretinopathy (PVR). Here we report that TNF-{alpha} promotes human RPE cell migration by inducing matrix metallopeptidase 9 (MMP-9) expression. Inhibition of MMP-9 by its inhibitor or its neutralizing antibody inhibited TNF-{alpha}-induced in vitro RPE cell migration. Reversely, exogenously-added active MMP-9 promoted RPE cell migration. Suppression Akt/mTOR complex 1(mTORC1) activation by LY 294002 and rapamycin inhibited TNF-{alpha}-mediated MMP-9 expression. To introduce a constitutively active Akt (CA-Akt) in cultured RPE cells increased MMP-9 expression, and to block mTORC1 activation by rapamycin inhibited its effect. RNA interference (RNAi)-mediated silencing of SIN1, a key component of mTOR complex 2 (mTORC2), had no effect on MMP-9 expression or secretion. In conclusion, this study suggest that TNF-{alpha} promotes RPE cell migration by inducing MMP-9 expression through activation of Akt/ mTORC1, but not mTORC2 signaling.

  11. Tumour necrosis factor-alpha impairs neuronal differentiation but not proliferation of hippocampal neural precursor cells: Role of Hes1.

    PubMed

    Keohane, Aoife; Ryan, Sinead; Maloney, Eimer; Sullivan, Aideen M; Nolan, Yvonne M

    2010-01-01

    Tumour necrosis factor-alpha (TNFalpha) is a pro-inflammatory cytokine, which influences neuronal survival and function yet there is limited information available on its effects on hippocampal neural precursor cells (NPCs). We show that TNFalpha treatment during proliferation had no effect on the percentage of proliferating cells prepared from embryonic rat hippocampal neurosphere cultures, nor did it affect cell fate towards either an astrocytic or neuronal lineage when cells were then allowed to differentiate. However, when cells were differentiated in the presence of TNFalpha, significantly reduced percentages of newly born and post-mitotic neurons, significantly increased percentages of astrocytes and increased expression of TNFalpha receptors, TNF-R1 and TNF-R2, as well as expression of the anti-neurogenic Hes1 gene, were observed. These data indicate that exposure of hippocampal NPCs to TNFalpha when they are undergoing differentiation but not proliferation has a detrimental effect on their neuronal lineage fate, which may be mediated through increased expression of Hes1.

  12. Gravity-Induced Gene Expression in Plants.

    NASA Astrophysics Data System (ADS)

    Sederoff, Heike; Heber, Steffen; Howard, Brian; Myburg-Nichols, Henrietta; Hammond, Rebecca; Salinas-Mondragon, Raul; Brown, Christopher S.

    Plants sense changes in their orientation towards the vector of gravity and respond with directional growth. Several metabolites in the signal transduction cascade have been identified. However, very little is known about the interaction between these sensing and signal transduction events and even less is known about their role in the differential growth response. Gravity induced changes in transcript abundance have been identified in Arabidopsis whole seedlings and root apices (Moseyko et al. 2002; Kimbrough et al. 2004). Gravity induced transcript abundance changes can be observed within less than 1 min after stimulation (Salinas-Mondragon et al. 2005). Gene expression however requires not only transcription but also translation of the mRNA. Translation can only occur when mRNA is associated with ribosomes, even though not all mRNA associated with ribosomes is actively translated. To approximate translational capacity we quantified whole genome transcript abundances in corn stem pulvini during the first hour after gravity stimulation in total and poly-ribosomal fractions. As in Arabidopsis root apices, transcript abundances of several clusters of genes responded to gravity stimulation. The vast majority of these transcripts were also found to associate with polyribosomes in the same temporal and quantitative pattern. These genes are transcriptionally regulated by gravity stimulation, but do not exhibit translational regulation. However, a small group of genes showed increased transcriptional regulation after gravity stimulation, but no association with polysomes. These transcripts likely are translationally repressed. The mechanism of translational repression for these transcripts is unknown. Based on the hypothesis that the genes essential for gravitropic responses should be expressed in most or all species, we compared the temporal gravity induced expression pattern of all orthologs identified between maize and Arabidopsis. A small group of genes showed high

  13. Identification of a novel cyclosporin-sensitive element in the human tumor necrosis factor alpha gene promoter

    PubMed Central

    1993-01-01

    Tumor necrosis factor alpha (TNF-alpha), a cytokine with pleiotropic biological effects, is produced by a variety of cell types in response to induction by diverse stimuli. In this paper, TNF-alpha mRNA is shown to be highly induced in a murine T cell clone by stimulation with T cell receptor (TCR) ligands or by calcium ionophores alone. Induction is rapid, does not require de novo protein synthesis, and is completely blocked by the immunosuppressant cyclosporin A (CsA). We have identified a human TNF-alpha promoter element, kappa 3, which plays a key role in the calcium-mediated inducibility and CsA sensitivity of the gene. In electrophoretic mobility shift assays, an oligonucleotide containing kappa 3 forms two DNA protein complexes with proteins that are present in extracts from unstimulated T cells. These complexes appear in nuclear extracts only after T cell stimulation. Induction of the inducible nuclear complexes is rapid, independent of protein synthesis, and blocked by CsA, and thus, exactly parallels the induction of TNF-alpha mRNA by TCR ligands or by calcium ionophore. Our studies indicate that the kappa 3 binding factor resembles the preexisting component of nuclear factor of activated T cells. Thus, the TNF-alpha gene is an immediate early gene in activated T cells and provides a new model system in which to study CsA-sensitive gene induction in activated T cells. PMID:8376940

  14. The increased gastroprotective effect of pioglitazone in cholestatic rats: role of nitric oxide and tumour necrosis factor alpha.

    PubMed

    Moezi, Leila; Janahmadi, Zeinab; Amirghofran, Zahra; Nekooeian, Ali Akbar; Dehpour, Ahmad R

    2014-02-01

    The prevalence of gastric ulcers is high in cholestatic patients, but the exact mechanism of this increased frequency remains uncertain. It has been shown that pioglitazone accelerates the healing of pre-existing gastric ulcers. The present study was designed to investigate the effect of pioglitazone, on the gastric mucosal lesions in cholestatic rats. Cholestasis was induced by surgical ligation of common bile duct and sham-operated rats served as control. Different groups of sham and cholestatic animals received solvent or pioglitazone (5, 15, 30 mg/kg) for 7 days. On the day eight rats were killed after oral ethanol administration and the area of gastric lesions was measured. The serums of rats were also collected to determine serum levels of tumour necrosis factor alpha (TNF-α), IL-1β and bilirubin. The ethanol-induced gastric mucosal damage was significantly more severe in cholestatic rats than sham-operated ones. Pretreatment with pioglitazone dose-dependently attenuated gastric lesions induced by ethanol in both sham and cholestatic rats, but this effect was more prominent in cholestatic ones. The effect of pioglitazone was associated with a significant fall in serum levels of TNF-α in cholestatic rats. L-NAME, a non-selective nitric oxide synthase (NOS) inhibitor, and decreased pioglitazone-induced gastroprotective effect in cholestatic rats, while aminoguanidine, a selective inducible NOS inhibitor, potentiated pioglitazone-induced gastroprotective effect in the cholestatic rats. Chronic treatment with pioglitazone exerts an enhanced gastroprotective effect on the stomach ulcers of cholestatic rats compared to sham rats probably due to constitutive NOS induction and/or inducible NOS inhibition and attenuating release of TNF-α.

  15. Expression of polyalanine stretches induces mitochondrial dysfunction.

    PubMed

    Toriumi, Kazuya; Oma, Yoko; Kino, Yoshihiro; Futai, Eugene; Sasagawa, Noboru; Ishiura, Shoichi

    2008-05-15

    In recent years, several novel types of disorders have been characterized, including what have been termed polyalanine diseases, in which patients have expanded triplet repeats in specific genes, resulting in the translation of aberrantly elongated polyalanine stretches. In this study, we showed that yellow fluorescent protein (YFP)-fused elongated polyalanine stretches localized exclusively to the cytoplasm and formed aggregates. Additionally, the polyalanine stretches themselves were toxic. We sought to identify proteins that bound directly to the polyalanine stretches, as factors that might be involved in triggering cell death. Many mitochondrial proteins were identified as polyalanine-binding proteins. We showed that one of the identified proteins, succinate dehydrogenase subunit A, was decreased in the mitochondria of cells expressing polyalanine stretches; as a result, succinate oxidative activity was decreased. Furthermore, the polyalanine stretches also associated directly with mitochondria. This suggests that polya-lanine stretches might directly induce cell death. Additionally, the mitochondrial membrane potential was reduced in cells expressing polyalanine stretches. We propose a novel mechanism by which polyalanine stretches may cause cytotoxicity through mitochondrial dysfunction. This may be a common mechanism underlying the pathogenesis of all polyalanine diseases.

  16. Constitutive expression of MC1R in HaCaT keratinocytes inhibits basal and UVB-induced TNF-alpha production.

    PubMed

    Garcin, Geneviève; Le Gallic, Lionel; Stoebner, Pierre-Emmanuel; Guezennec, Anne; Guesnet, Joelle; Lavabre-Bertrand, Thierry; Martinez, Jean; Meunier, Laurent

    2009-01-01

    Alpha-melanocyte stimulating hormone (alpha-MSH) binds to melanocortin-1 receptor (MC1R) on melanocytes to stimulate pigmentation and modulate various cutaneous inflammatory responses. MC1R expression is not restricted to melanocytic cells and may be induced in keratinocytes after UVB exposure. We hypothesized that MC1R signaling in keratinocytes, wherein basal conditions are barely expressed, may modulate mediators of inflammation, such as nuclear factor-kappa B (NF-kappaB) and tumor necrosis factor-alpha (TNF-alpha). Therefore, we generated HaCaT cells that stably express human MC1R or the Arg151Cys (R151C) nonfunctional variant. We demonstrate that: (1) the constitutive activity of MC1R results in elevated intracellular cAMP level, reduced NF-kappaB activity and decreased TNF-alpha transcription; (2) binding of alpha-MSH to MC1R and the subsequent increase in cAMP production do not inhibit TNFalpha-mediated NF-kappaB activation; (3) MC1R signaling is sufficient to strongly inhibit UVB-induced TNF-alpha expression and this inhibitory effect is further enhanced by alpha-MSH stimulation. Our findings suggest that the constitutive activity of the G-protein-coupled MC1R in keratinocytes may contribute to the modulation of inflammatory events and immune response induced by UV light.

  17. Asiatic acid alleviates hemodynamic and metabolic alterations via restoring eNOS/iNOS expression, oxidative stress, and inflammation in diet-induced metabolic syndrome rats.

    PubMed

    Pakdeechote, Poungrat; Bunbupha, Sarawoot; Kukongviriyapan, Upa; Prachaney, Parichat; Khrisanapant, Wilaiwan; Kukongviriyapan, Veerapol

    2014-01-16

    Asiatic acid is a triterpenoid isolated from Centella asiatica. The present study aimed to investigate whether asiatic acid could lessen the metabolic, cardiovascular complications in rats with metabolic syndrome (MS) induced by a high-carbohydrate, high-fat (HCHF) diet. Male Sprague-Dawley rats were fed with HCHF diet with 15% fructose in drinking water for 12 weeks to induce MS. MS rats were treated with asiatic acid (10 or 20 mg/kg/day) or vehicle for a further three weeks. MS rats had an impairment of oral glucose tolerance, increases in fasting blood glucose, serum insulin, total cholesterol, triglycerides, mean arterial blood pressure, heart rate, and hindlimb vascular resistance; these were related to the augmentation of vascular superoxide anion production, plasma malondialdehyde and tumor necrosis factor-alpha (TNF-α) levels (p<0.05). Plasma nitrate and nitrite (NOx) were markedly high with upregulation of inducible nitric oxide synthase (iNOS) expression, but dowregulation of endothelial nitric oxide synthase (eNOS) expression (p<0.05). Asiatic acid significantly improved insulin sensitivity, lipid profiles, hemodynamic parameters, oxidative stress markers, plasma TNF-α, NOx, and recovered abnormality of eNOS/iNOS expressions in MS rats (p<0.05). In conclusion, asiatic acid improved metabolic, hemodynamic abnormalities in MS rats that could be associated with its antioxidant, anti-inflammatory effects and recovering regulation of eNOS/iNOS expression.

  18. High Serum Interleukin-10 and Tumor Necrosis Factor Alpha Levels in Chronic Paracoccidioidomycosis

    PubMed Central

    Fornari, M. C.; Bava, A. J.; Guereño, M. T.; Berardi, V. E.; Silaf, M. R.; Negroni, R.; Diez, R. A.

    2001-01-01

    In patients with chronic paracoccidioidomycosis (n = 10), levels of tumor necrosis factor alpha, interleukin-10, and interleukin-2 in serum, measured by enzyme-linked immunosorbent assay (in picograms per milliliter, as mean ± standard error of the mean), were higher than in normal controls (n = 8): 186 ± 40 versus 40 ± 7 (P < 0.05), 203 ± 95 versus 20 ± 8 (P = 0.001), and 96.3 ± 78.57 versus 1.19 ± 1.19 (P = 0.045), respectively. Gamma interferon and interleukin-4 levels were similar in patients and controls. PMID:11527826

  19. Interleukin 6 and tumour necrosis factor alpha serum levels in monoclonal gammopathy of undetermined significance.

    PubMed

    Bladé, Joan; Filella, Xabier; Montoto, Silvia; Bosch, Francesc; Rosiñol, Laura; Coca, Francesca; Giné, Eva; Nadal, Elisabeth; Aymerich, Marta; Rozman, María; Montserrat, Emili

    2002-05-01

    The objectives of the present study were to compare the interleukin 6 (IL-6) and tumour necrosis factor-alpha (TNF-alpha) serum levels of individuals with monoclonal gammopathy of undetermined significance (MGUS) with those of healthy controls, and to ascertain the predictor value of these cytokines in the evolution from MGUS to multiple myeloma. After a median follow-up of 7 years from the initial cytokine measurements, nine patients with MGUS have evolved to a malignant condition. The actuarial probability of malignant transformation in patients with increased IL-6 and TNF-alpha was not significantly higher than in those with normal values.

  20. Effects of Tumor Necrosis Factor Alpha on Sin Nombre Virus Infection In Vitro

    PubMed Central

    Khaiboullina, Svetlana F.; Netski, Dale M.; Krumpe, Peter; St. Jeor, Stephen C.

    2000-01-01

    Previous data indicate that immune mechanisms may be involved in developing capillary leakage during Sin Nombre virus (SNV) infection. Therefore, we investigated production of tumor necrosis factor alpha (TNF-α) by human alveolar macrophages and human umbilical vein endothelial cells (HUVEC) after infection with SNV. In addition, we examined the effect of TNF-α on HUVEC monolayer leakage. Our results reveal that although TNF-α decreases accumulation of viral nucleoproteins, TNF-α levels do not change in SNV-infected cells. In addition, supernatants from SNV-infected human alveolar macrophages did not cause a significant increase in endothelial monolayer permeability. PMID:11090198

  1. Effects of interferon-gamma and tumor necrosis factor-alpha on macrophage enzyme levels

    NASA Technical Reports Server (NTRS)

    Pierangeli, Silvia S.; Sonnenfeld, Gerald

    1989-01-01

    Murine peritoneal macrophages were treated with interferon-gamma (IFN-gamma) or tumor necrosis factor-alpha (TNF). Measurements of changes in acid phosphatase and beta-glucuronidase levels were made as an indication of activation by cytokine treatment. IFN-gamma or TNF-gamma treatment resulted in a significant increase in the activities of both enzymes measured in the cell lysates. This increase was observable after 6 h of incubation, but reached its maximum level after 24 h of incubation. The effect of the treatment of the cell with both cytokines together was additive. No synergistic effect of addition of both cytokines on the enzyme levels was observed.

  2. Implication of TNF-alpha convertase (TACE/ADAM17) in inducible nitric oxide synthase expression and inflammation in an experimental model of colitis.

    PubMed

    Colón, A L; Menchén, L A; Hurtado, O; De Cristóbal, J; Lizasoain, I; Leza, J C; Lorenzo, P; Moro, M A

    2001-12-21

    Tumour necrosis factor-alpha (TNF-alpha) is a pro-inflammatory cytokine which is shed in its soluble form by a disintegrin and metalloproteinase (ADAM) called TNF-alpha convertase (TACE; ADAM17). TNF-alpha plays a role in inflammatory bowel disease (IBD) and is involved in the expression of inducible nitric oxide synthase (iNOS) which has also been implicated in IBD. The study was designed to investigate whether colitis induced by trinitrobenzene sulphonic acid (TNBS) in rats produces an increase in TACE activity and/or expression and whether its pharmacological inhibition reduces TNF-alpha levels, iNOS expression and colonic damage in this model. TNBS (30 mg in 0.4 ml of 50% ethanol) was instilled into the colon of female Wistar rats. Saline or TACE inhibitor BB1101 (10 mg/kg/day) was administered intraperitoneally 5 days after TNBS instillation. On day 10, colons were removed and assessed for pathological score, myeloperoxidase (MPO), NO synthase (NOS), TACE enzymatic activity and protein levels, colonic TNF-alpha and NOx- levels. Instillation of TNBS caused an increase in TACE activity and expression and the release of TNF-alpha. TNBS also resulted in iNOS expression and colonic damage. BB1101 blocked TNBS-induced increase in TACE activity, TNF-alpha release and iNOS expression. Concomitantly, BB1101 ameliorated TNBS-induced colonic damage and inflammation. TNBS causes TNF-alpha release by an increase in TACE activity and expression and this results in the expression of iNOS and subsequent inflammation, suggesting that TACE inhibition may prove useful as a therapeutic means in IBD.

  3. Differential induction of tumor necrosis factor alpha in murine and human leukocytes by Mycoplasma arthritidis-derived superantigen.

    PubMed Central

    Rink, L; Nicklas, W; Alvarez-Ossorio, L; Koester, M; Kirchner, H

    1994-01-01

    Mycoplasma arthritidis-derived superantigen (MAS) is exclusively produced by M. arthritidis, which is the only known mycoplasma to produce a superantigen. As a superantigen, MAS shows properties similar to those of the staphylococcal enterotoxins and related substances, such as binding to major histocompatibility complex (MHC) class II and V beta-specific stimulation of T cells. In this series of experiments, we demonstrate some differences between MAS and other superantigens. MAS induced the production of tumor necrosis factor alpha (TNF-alpha) mRNA in human as well as in murine leukocytes. However, only in murine leukocytes was the mRNA adequately translated into the protein. In human peripheral blood mononuclear cells, we found only small amounts of TNF, whereas in murine spleen cells we detected levels more than three times higher. The proliferative response to MAS has been shown to be restricted to I-E alpha in the murine MHC. Furthermore, TNF was induced in I-E alpha+ bone marrow-derived macrophages by MAS. In these cells, MAS rapidly induced very high levels of TNF and the amounts of mRNA detected correlated to the amount of protein produced. In comparison with other superantigens, including the staphylococcal enterotoxins, toxic shock syndrome toxin 1, and exfoliative toxin A, the failure of MAS to induce TNF-alpha in human peripheral blood mononuclear cells is specific for MAS and not common to all superantigens. The direct activation of bone marrow-derived macrophages also seems to be specific for MAS. These data suggest that the induction of TNF-alpha by MAS is dependent on the strength of binding to the MHC class II molecule. Images PMID:8300207

  4. Sleep deprivation attenuates endotoxin-induced cytokine gene expression independent of day length and circulating cortisol in male Siberian hamsters (Phodopus sungorus).

    PubMed

    Ashley, Noah T; Walton, James C; Haim, Achikam; Zhang, Ning; Prince, Laura A; Fruchey, Allison M; Lieberman, Rebecca A; Weil, Zachary M; Magalang, Ulysses J; Nelson, Randy J

    2013-07-15

    Sleep is restorative, whereas reduced sleep leads to negative health outcomes, such as increased susceptibility to disease. Sleep deprivation tends to attenuate inflammatory responses triggered by infection or exposure to endotoxin, such as bacterial lipopolysaccharide (LPS). Previous studies have demonstrated that Siberian hamsters (Phodopus sungorus), photoperiodic rodents, attenuate LPS-induced fever, sickness behavior and upstream pro-inflammatory gene expression when adapted to short day lengths. Here, we tested whether manipulation of photoperiod alters the suppressive effects of sleep deprivation upon cytokine gene expression after LPS challenge. Male Siberian hamsters were adapted to long (16 h:8 h light:dark) or short (8 h:16 h light:dark) photoperiods for >10 weeks, and were deprived of sleep for 24 h using the multiple platform method or remained in their home cage. Hamsters received an intraperitoneal injection of LPS or saline (control) 18 h after starting the protocol, and were killed 6 h later. LPS increased liver and hypothalamic interleukin-1 (IL-1) and tumor necrosis factor-alpha (TNF) gene expression compared with vehicle. Among LPS-challenged hamsters, sleep deprivation reduced IL-1 mRNA levels in liver and hypothalamus, but not TNF. IL-1 attenuation was independent of circulating baseline cortisol, which did not increase after sleep deprivation. Conversely, photoperiod altered baseline cortisol, but not pro-inflammatory gene expression in sleep-deprived hamsters. These results suggest that neither photoperiod nor glucocorticoids influence the suppressive effect of sleep deprivation upon LPS-induced inflammation.

  5. Cardioinductive network guiding stem cell differentiation revealed by proteomic cartography of tumor necrosis factor alpha-primed endodermal secretome.

    PubMed

    Arrell, D Kent; Niederländer, Nicolas J; Faustino, Randolph S; Behfar, Atta; Terzic, Andre

    2008-02-01

    In the developing embryo, instructive guidance from the ventral endoderm secures cardiac program induction within the anterolateral mesoderm. Endoderm-guided cardiogenesis, however, has yet to be resolved at the proteome level. Here, through cardiopoietic priming of the endoderm with the reprogramming cytokine tumor necrosis factor alpha (TNFalpha), candidate effectors of embryonic stem cell cardiac differentiation were delineated by comparative proteomics. Differential two-dimensional gel electrophoretic mapping revealed that more than 75% of protein species increased >1.5-fold in the TNFalpha-primed versus unprimed endodermal secretome. Protein spot identification by linear ion trap quadrupole (LTQ) tandem mass spectrometry (MS/MS) and validation by shotgun LTQ-Fourier transform MS/MS following multidimensional chromatography mapped 99 unique proteins from 153 spot assignments. A definitive set of 48 secretome proteins was deduced by iterative bioinformatic screening using algorithms for detection of canonical and noncanonical indices of secretion. Protein-protein interaction analysis, in conjunction with respective expression level changes, revealed a nonstochastic TNFalpha-centric secretome network with a scale-free hierarchical architecture. Cardiovascular development was the primary developmental function of the resolved TNFalpha-anchored network. Functional cooperativity of the derived cardioinductive network was validated through direct application of the TNFalpha-primed secretome on embryonic stem cells, potentiating cardiac commitment and sarcomerogenesis. Conversely, inhibition of primary network hubs negated the procardiogenic effects of TNFalpha priming. Thus, proteomic cartography establishes a systems biology framework for the endodermal secretome network guiding stem cell cardiopoiesis.

  6. S. aureus blocks efferocytosis of neutrophils by macrophages through the activity of its virulence factor alpha toxin

    PubMed Central

    Cohen, Taylor S.; Jones-Nelson, Omari; Hotz, Meghan; Cheng, Lily; Miller, Lloyd S.; Suzich, JoAnn; Stover, C. Kendall; Sellman, Bret R.

    2016-01-01

    Bacterial pneumonia, such as those caused by Staphylococcus aureus, is associated with an influx of inflammatory neutrophils into the lung tissue and airways. Regulation and clearance of recruited neutrophils is essential for preventing tissue damage by “friendly fire”, a responsibility of macrophages in a process called efferocytosis. We hypothesized that S. aureus impairs efferocytosis by alveolar macrophages (AMs) through the activity of the secreted virulence factor alpha toxin (AT), which has been implicated in altering the antimicrobial function of AMs. Infection of mice lacking AMs resulted in significantly increased numbers of neutrophils in the lung, while clearance of neutrophils delivered intranasally into uninfected mice was reduced in AM depleted animals. In vitro, sublytic levels of AT impaired uptake of apoptotic neutrophils by purified AMs. In vivo, the presence of AT reduced uptake of neutrophils by AMs. Differential uptake of neutrophils was not due to changes in either the CD47/CD172 axis or CD36 levels. AT significantly reduced lung expression of CCN1 and altered AM surface localization of DD1α, two proteins known to influence efferocytosis. We conclude that AT may contribute to tissue damage during S. aureus pneumonia by inhibiting the ability of AM to clear neutrophils at the site of infection. PMID:27739519

  7. Electrical remodeling of cardiac myocytes from mice with heart failure due to the overexpression of tumor necrosis factor-alpha.

    PubMed

    Petkova-Kirova, Polina S; Gursoy, Erdal; Mehdi, Haider; McTiernan, Charles F; London, Barry; Salama, Guy

    2006-05-01

    Mice that overexpress the inflammatory cytokine tumor necrosis factor-alpha in the heart (TNF mice) develop heart failure characterized by atrial and ventricular dilatation, decreased ejection fraction, atrial and ventricular arrhythmias, and increased mortality (males > females). Abnormalities in Ca2+ handling, prolonged action potential duration (APD), calcium alternans, and reentrant atrial and ventricular arrhythmias were previously observed with the use of optical mapping of perfused hearts from TNF mice. We therefore tested whether altered voltage-gated outward K+ and/or inward Ca2+ currents contribute to the altered action potential characteristics and the increased vulnerability to arrhythmias. Whole cell voltage-clamp recordings of K+ currents from left ventricular myocytes of TNF mice revealed an approximately 50% decrease in the rapidly activating, rapidly inactivating transient outward K+ current Ito and in the rapidly activating, slowly inactivating delayed rectifier current IK,slow1, an approximately 25% decrease in the rapidly activating, slowly inactivating delayed rectifier current IK,slow2, and no significant change in the steady-state current Iss compared with controls. Peak amplitudes and inactivation kinetics of the L-type Ca2+ current ICa,L were not altered. Western blot analyses revealed a reduction in the proteins underlying Kv4.2, Kv4.3, and Kv1.5. Thus decreased K+ channel expression is largely responsible for the prolonged APD in the TNF mice and may, along with abnormalities in Ca2+ handling, contribute to arrhythmias.

  8. Increased intestinal protein synthesis during sepsis and following the administration of tumour necrosis factor alpha or interleukin-1 alpha.

    PubMed Central

    von Allmen, D; Hasselgren, P O; Higashiguchi, T; Frederick, J; Zamir, O; Fischer, J E

    1992-01-01

    The influence of sepsis on intestinal protein synthesis was studied in rats. Sepsis was induced by caecal ligation and puncture (CLP); control rats were sham-operated. Protein synthesis was measured in vivo in the jejunum and ileum following a flooding dose of [14C]leucine. At 8 h after CLP the protein synthesis rate was increased by approx. 15% in jejunal mucosa, and at 16 h after CLP, the protein synthesis rate was increased by 50-60% in the mucosa and seromuscular layer of both jejunum and ileum. In a second series of experiments, rats were treated with recombinant tumour necrosis factor alpha (rTNF alpha) or recombinant interleukin-1 alpha (rIL-1 alpha) administered at a total dose of 300 micrograms/kg body weight over 16 h. Control rats received corresponding volumes of solvent. Treatment with rTNF alpha resulted in an approx. 25% increase in mucosal protein synthesis in jejunum. Following treatment with rIL-1 alpha, protein synthesis increased by 25% in jejunal mucosa and almost doubled in ileal mucosa. The results suggest that sepsis stimulates intestinal protein synthesis and that this response may, at least in part, be mediated by TNF and/or IL-1. PMID:1530589

  9. Molecular design of hybrid tumour necrosis factor alpha with polyethylene glycol increases its anti-tumour potency.

    PubMed Central

    Tsutsumi, Y.; Kihira, T.; Tsunoda, S.; Kanamori, T.; Nakagawa, S.; Mayumi, T.

    1995-01-01

    This study was conducted to increase the anti-tumour potency and reduce the toxic side-effects of tumour necrosis factor alpha (TNF-alpha). Natural human TNF-alpha was chemically conjugated with monomethoxy polyethylene glycol (PEG) using succinimidyl coupling of lysine amino groups of TNF-alpha. The number-average molecular weight of PEG-modified TNF-alpha (PEG-TNF-alpha) increased with an increase in the reaction time and the initial molar ratio of PEG relative to TNF-alpha. The resulting modified TNF-alpha was separated into fractions of various molecular weights. The specific activity of separated PEG-TNF-alpha s relative to that of native TNF-alpha gradually decreased with an increase in the degree of PEG modification, but the plasma half-life was drastically increased with the increase in molecular weight of modified TNF-alpha. PEG-TNF-alpha s, in which 29% and 56% of lysine residues were coupled to PEG, had anti-tumour activity approximately 4 and 100 times greater than unmodified TNF-alpha in the murine Meth-A fibrosarcoma model. Extensive PEG modification did not increase its in vivo activity. A high dose of unmodified TNF-alpha induced toxic side-effects, but these were not observed with the modified TNF-alpha s. Optimal PEG modification of TNF-alpha markedly increased its bioavailability and may facilitate its potential anti-tumour therapeutic use. PMID:7734321

  10. Expression changes of inflammatory factors in the rat lung of decompression sickness induced by fast buoyancy ascent escape.

    PubMed

    Wang, Hai-Tao; Fang, Yi-Qun; You, Pu; Bao, Xiao-Chen; Yuan, Heng-Rong; Ma, Jun; Wang, Fang-Fang; Li, Kai-Cheng

    2015-01-01

    Fast buoyancy ascent escape is one of the major naval submarine escape maneuvers. Decompression sickness (DCS) is the major bottleneck to increase the depth of fast buoyancy ascent escape. Rapid decompression induces the release of inflammatory mediators and results in tissue inflammation cascades and a protective anti-inflammatory response. In our previous study, we found that DCS caused by simulated fast buoyancy ascent escape could induce acute lung injury (ALI) and the expression changes of the proinflammatory cytokines: tumor necrosis factor alpha (TNF-α), interleukin (IL)-1β and IL-6 in rat lung tissue. In order to study the expression change characteristics of TNF-α, IL-1β, IL-6, IL-10 and IL-13 in the rat lung of DCS caused by simulated fast buoyancy ascent escape, we detected the rat lung mRNA and protein levels of TNF-α, IL-1β, IL-6, IL-10 and IL-13 at 0.5 hour after DCS caused by simulated fast buoyancy ascent escape (fast escape group), compared with the normal control group (control group) and diving DCS (decompression group). We observed that DCS caused by simulated fast buoyancy ascent escape could increase the mRNA levels of TNF-α, IL-1β, IL-6, IL-10, and the protein levels of TNF-α, IL-10 in rat lung tissue. At the same time, we found that the protein level of IL-13 was also downregulated in rat lung tissue. TNF-α, IL-10 and IL-13 may be involved in the process of the rat lung injury of DCS caused by simulated fast buoyancy ascent escape. In conclusion, the expression changes of inflammatory factors in the rat lung of DCS caused by simulated fast buoyancy ascent escape were probably different from that in the rat lung of diving DCS, which indicated that the pathological mechanism of DCS caused by simulated fast buoyancy ascent escape might be different from that of diving DCS.

  11. Transgenic cyclooxygenase-2 expression and high salt enhanced susceptibility to chemical-induced gastric cancer development in mice.

    PubMed

    Leung, Wai K; Wu, Kai-chun; Wong, Christine Y P; Cheng, Alfred S L; Ching, Arthur K K; Chan, Anthony W H; Chong, Wilson W S; Go, Minnie Y Y; Yu, Jun; To, Ka-Fai; Wang, Xin; Chui, Y L; Fan, D M; Sung, Joseph J Y

    2008-08-01

    Cyclooxoygenase (COX)-2 overexpression is involved in gastric carcinogenesis. While high-salt intake is a known risk factor for gastric cancer development, we determined the effects of high salt on gastric chemical carcinogenesis in COX-2 transgenic (TG) mice. COX-2 TG mice were developed in C57/BL6 strain using the full-length human cox-2 complementary DNA construct. Six-week-old COX-2 TG and wild-type (WT) littermates were randomly allocated to receive alternate week of N-methyl-N-nitrosourea (MNU, 240 p.p.m.) in drinking water or control for 10 weeks. Two groups of mice were further treated with 10% NaCl during the initial 10 weeks. All mice were killed at the end of week 50. Both forced COX-2 overexpression and high-salt intake significantly increased the frequency of gastric cancer development in mice as compared with WT littermates treated with MNU alone. However, no additive effect was observed on the combination of high salt and COX-2 expression. We further showed that MNU and high-salt treatment increased chronic inflammatory infiltrates and induced prostaglandin E(2) (PGE(2)) production in the non-cancerous stomach. Whereas high-salt treatment markedly increased the expression of inflammatory cytokines (tumor necrosis factor-alpha, interferon-gamma, interleukin (IL)-1 beta and IL-6) in the gastric mucosa, COX-2 overexpression significantly altered the cell kinetics in the MNU-induced gastric cancer model. In conclusion, both high salt and COX-2 overexpression promote chemical-induced gastric carcinogenesis, possibly related to chronic inflammation, induction of PGE(2), disruption of cell kinetics and induction of inflammatory cytokines.

  12. Eugenol reduces acute pain in mice by modulating the glutamatergic and tumor necrosis factor alpha (TNF-α) pathways.

    PubMed

    Dal Bó, Wladmir; Luiz, Ana Paula; Martins, Daniel F; Mazzardo-Martins, Leidiane; Santos, Adair R S

    2013-10-01

    Eugenol is utilized together with zinc oxide in odontological clinical for the cementation of temporary prostheses and the temporary restoration of teeth and cavities. This work explored the antinociceptive effects of the eugenol in different models of acute pain in mice and investigated its possible modulation of the inhibitory (opioid) and excitatory (glutamatergic and pro-inflammatory cytokines) pathways of nociceptive signaling. The administration of eugenol (3-300 mg/kg, p.o., 60 min or i.p., 30 min) inhibited 82 ± 10% and 90 ± 6% of the acetic acid-induced nociception, with ID₅₀ values of 51.3 and 50.2 mg/kg, respectively. In the glutamate test, eugenol (0.3-100 mg/kg, i.p.) reduced the response behavior by 62 ± 5% with an ID₅₀ of 5.6 mg/kg. In addition, the antinociceptive effect of eugenol (10 mg/kg, i.p.) in the glutamate test was prevented by the i.p. treatment for mice with naloxone. The pretreatment of mice with eugenol (10 mg/kg, i.p.) was able to inhibit the nociception induced by the intrathecal (i.t.) injection of glutamate (37 ± 9%), kainic (acid kainite) (41 ± 12%), α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) (55 ± 5%), and substance P (SP) (39 ± 8%). Furthermore, eugenol (10 mg/kg, i.p.) also inhibited biting induced by tumor necrosis factor alpha (TNF-α, 65 ± 8%). These results extend our current knowledge of eugenol and confirm that it promotes significant antinociception against different mouse models of acute pain. The mechanism of action appears to involve the modulation of the opioid system and glutamatergic receptors (i.e., kainate and AMPA), and the inhibition of TNF-α. Thus, eugenol could represent an important compound in the treatment for acute pain.

  13. Effect of Egg White Combined with Chalcanthite on Lipopolysaccharide induced Inflammatory Cytokine Expression in RAW 264.7 cells

    PubMed Central

    Choi, Eun-A; Yoon, Jeung-Won; Choi, Hak-Joo; Kim, Dong-Hee; Yoo, Hwa-Seung

    2012-01-01

    Aim: Historically, mineral compound herbal medicines have long been used in treatments of immune-related diseases in Korea, China and other Asian countries. In this study, we investigated the anti-inflammatory effect of egg white combined with chalcanthite (IS4) on lipopolysaccharide (LPS)-stimulated RAW 264.7 cells. Methods: RAW 264.7 cells cultured with LPS and various concentrations of IS4 were analyzed to determine the production of pro-inflammatory cytokines and mediators by using enzyme-linked immune sorbent assays (ELISAs). Results: IS4 concentration inhibited the production of interleukin-1beta (IL-1 β), interleukin-6 (IL-6), and granulocyte -macrophage colony-stimulating factor (GM-CSF) induced by LPS. IS4 at high concentrations (25 and 50㎍/ml) inhibited, in concentration-dependent manner, the expression of tumor necrosis factor-alpha (TNF– α) stimulated by LPS. Conclusion: IS4 has shown an anti-inflammatory effect in RAW 264.7 cells. PMID:25780629

  14. Brain cytokine and chemokine mRNA expression in mice induced by intranasal instillation with ultrafine carbon black.

    PubMed

    Tin-Tin-Win-Shwe; Yamamoto, Shoji; Ahmed, Sohel; Kakeyama, Masaki; Kobayashi, Takahiro; Fujimaki, Hidekazu

    2006-05-25

    Ambient air ultrafine particles (UFPs) have gained enormous attention to many researchers with recent evidence showing them to have more hazardous effects on human health than larger ambient particles. Studies focusing the possibility of effects on brain are quite limited. To examine the effect of ultrafine carbon black (ufCB) on mice brain, we instilled 125 microg of 14 nm or 95 nm CB into the nostrils of 8-week-old male BALB/c mice, once a week for 4 weeks. Four hours after the last instillation, we collected olfactory bulb and hippocampus and detected the expression of cytokine and chemokine mRNA by quantitative real-time PCR method. In this study, we found the induction of proinflammatory cytokines (interleukin-1 beta and tumor necrosis factor-alpha and chemokines (monocyte chemoattractant protein-1/CCL2, macrophage inflammatory protein-1 alpha/CCL3), and monokine induced interferon-gamma/CXC chemokine ligand (CXCL9) mRNA in brain olfactory bulb, not in the hippocampus of mice instilled with 14 nm ufCB intranasally. We suggest that the intranasal instillation of ufCB may influence the brain immune function depending on their size. To our knowledge, this is the first study to demonstrate region-specific brain cytokine and chemokine mRNA-induction in mice triggered by intranasal instillation of specific-sized ufCB, in a physiologically relevant condition.

  15. Role of very-late antigen-4 (VLA-4) in myelin basic protein-primed T cell contact-induced expression of proinflammatory cytokines in microglial cells.

    PubMed

    Dasgupta, Subhajit; Jana, Malabendu; Liu, Xiaojuan; Pahan, Kalipada

    2003-06-20

    The presence of neuroantigen-primed T cells recognizing self-myelin antigens within the CNS is necessary for the development of demyelinating autoimmune disease like multiple sclerosis. This study was undertaken to investigate the role of myelin basic protein (MBP)-primed T cells in the expression of proinflammatory cytokines in microglial cells. MBP-primed T cells alone induced specifically the microglial expression of interleukin (IL)-1beta, IL-1alpha tumor necrosis factor alpha, and IL-6, proinflammatory cytokines that are primarily involved in the pathogenesis of MS. This induction was primarily dependent on the contact between MBP-primed T cells and microglia. The activation of microglial NF-kappaB and CCAAT/enhancer-binding protein beta (C/EBPbeta) by MBP-primed T cell contact and inhibition of contact-mediated microglial expression of proinflammatory cytokines by dominant-negative mutants of p65 and C/EBPbeta suggest that MBP-primed T cells induce microglial expression of cytokines through the activation of NF-kappaB and C/EBPbeta. In addition, we show that MBP-primed T cells express very late antigen-4 (VLA-4), and functional blocking antibodies to alpha4 chain of VLA-4 (CD49d) inhibited the ability of MBP-primed T cells to induce microglial proinflammatory cytokines. Interestingly, the blocking of VLA-4 impaired the ability of MBP-primed T cells to induce microglial activation of only C/EBPbeta but not that of NF-kappaB. This study illustrates a novel role of VLA-4 in regulating neuroantigen-primed T cell-induced activation of microglia through C/EBPbeta

  16. Preliminary characterisation of tumor necrosis factor alpha and interleukin-10 responses to Chlamydia pecorum infection in the koala (Phascolarctos cinereus).

    PubMed

    Mathew, Marina; Beagley, Kenneth W; Timms, Peter; Polkinghorne, Adam

    2013-01-01

    Debilitating infectious diseases caused by Chlamydia are major contributors to the decline of Australia's iconic native marsupial species, the koala (Phascolarctos cinereus). An understanding of koala chlamydial disease pathogenesis and the development of effective strategies to control infections continue to be hindered by an almost complete lack of species-specific immunological reagents. The cell-mediated immune response has been shown to play an influential role in the response to chlamydial infection in other hosts. The objective of this study, hence, was to provide preliminary data on the role of two key cytokines, pro-inflammatory tumour necrosis factor alpha (TNFα) and anti-inflammatory interleukin 10 (IL10), in the koala Chlamydia pecorum response. Utilising sequence homology between the cytokine sequences obtained from several recently sequenced marsupial genomes, this report describes the first mRNA sequences of any koala cytokine and the development of koala specific TNFα and IL10 real-time PCR assays to measure the expression of these genes from koala samples. In preliminary studies comparing wild koalas with overt chlamydial disease, previous evidence of C. pecorum infection or no signs of C. pecorum infection, we revealed strong but variable expression of TNFα and IL10 in wild koalas with current signs of chlamydiosis. The description of these assays and the preliminary data on the cell-mediated immune response of koalas to chlamydial infection paves the way for future studies characterising the koala immune response to a range of its pathogens while providing reagents to assist with measuring the efficacy of ongoing attempts to develop a koala chlamydial vaccine.

  17. Role of Tumor Necrosis Factor Alpha in Disease Using a Mouse Model of Shiga Toxin-Mediated Renal Damage ▿

    PubMed Central

    Lentz, Erin K.; Cherla, Rama P.; Jaspers, Valery; Weeks, Bradley R.; Tesh, Vernon L.

    2010-01-01

    Mice have been extensively employed as an animal model of renal damage caused by Shiga toxins. In this study, we examined the role of the proinflammatory cytokine tumor necrosis factor alpha (TNF-α) in the development of toxin-mediated renal disease in mice. Mice pretreated with TNF-α and challenged with Shiga toxin type 1 (Stx1) showed increased survival compared to that of mice treated with Stx1 alone. Conversely, mice treated with Stx1 before TNF-α administration succumbed more quickly than mice given Stx1 alone. Increased lethality in mice treated with Stx1 followed by TNF-α was associated with evidence of glomerular damage and the loss of renal function. No differences in renal histopathology were noted between animals treated with Stx1 alone and the TNF-α pretreatment group, although we noted a sparing of renal function when TNF-α was administered before toxin. Compared to that of treatment with Stx1 alone, treatment with TNF-α after toxin altered the renal cytokine profile so that the expression of proinflammatory cytokines TNF-α and interleukin-1β (IL-1β) increased, and the expression of the anti-inflammatory cytokine IL-10 decreased. Increased lethality in mice treated with Stx1 followed by TNF-α was associated with higher numbers of dUTP-biotin nick end labeling-positive renal tubule cells, suggesting that increased lethality involved enhanced apoptosis. These data suggest that the early administration of TNF-α is a candidate interventional strategy blocking disease progression, while TNF-α production after intoxication exacerbates disease. PMID:20605983

  18. Use of anti tumor necrosis factor-alpha monoclonal antibody for ulcerative jejunoileitis

    PubMed Central

    Seven, Gulseren; Assaad, Adel; Biehl, Thomas; Kozarek, Richard A

    2012-01-01

    Ulcerative jejunoileitis is an uncommon clinical syndrome consisting of abdominal pain, weight loss associated with diarrhea, and multiple inflammatory ulcerations and strictures of the small bowel. Ulcerative jejunoileitis can complicate established celiac disease or develop in patients de novo. Increased levels of tumor necrosis factor-alpha (TNF-α) in the small intestine of patients with untreated celiac disease are associated with a role in the immune pathogenesis of this disorder. No specific therapy has been shown to change the course of ulcerative jejunoileitis. We report a case of severe ulcerative jejunoileitis previously unresponsive to traditional therapies, including high dose corticosteroids and cyclosporine. The patient had a dramatic resolution of symptoms and a complete normalization of endoscopic findings after anti-TNF-α monoclonal antibody, infliximab (Remicade®). PMID:23049226

  19. Transforming growth factor alpha and epidermal growth factor levels in normal human gastrointestinal mucosa.

    PubMed Central

    Cartlidge, S. A.; Elder, J. B.

    1989-01-01

    Acid soluble proteins from 23 samples of normal human gastrointestinal mucosa derived from four normal adult organ donors were extracted and subjected to specific radiommunoassays for transforming growth factor alpha (TGF alpha) and urogastrone epidermal growth factor (URO-EGF). All tissues were found to contain immunoreactive TGF alpha and levels ranged from 57 to 4,776 pg-1 wet weight of tissue. Although levels varied between tissue donors, the distribution of TGF alpha throughout the gastrointestinal tract appeared similar in all cases. URO-EGF levels were much lower (0-216 pg g-1 wet weight). TGF alpha levels in extracts of gastrointestinal mucosa from a 7-year-old female donor were higher and the observed distribution was markedly different from adult levels. URO-EGF was not detected in mucosal or submucosal tissue extracts from this patient. Further studies in juveniles are indicated. PMID:2803941

  20. Effect of transforming growth factor-alpha on inositol phospholipid metabolism in human epidermoid carcinoma cells

    SciTech Connect

    Kato, M.; Takenawa, T.; Twardzik, D.R.

    1988-08-01

    Transforming growth factor-alpha (TGF-alpha) stimulates (in a dose-dependent manner) the incorporation of (/sup 32/P)Pi into phosphatidylinositol (PI), phosphatidylinositol 4-phosphate (PIP), phosphatidylinositol 4,5-bisphosphate (PIP2), and phosphatidic acid (PA) in the human epidermoid carcinoma cell line (A431). The effect of TGF-alpha on the incorporation was found to be similar to that of EGF. On the other hand, a striking difference in the activation of diacylglycerol (DG) kinase activity was seen between TGF-alpha and EGF. At least 100 times more TGF-alpha was required to achieve maximal stimulation of DG kinase activity relative to EGF. These results suggest that the activation of DG kinase by TGF-alpha may involve a mechanism independent from or subsequent to activation of the EGF receptor.

  1. N-Formyl-Methionyl-Leucyl-Phenylalanine Inhibits both Gamma Interferon- and Interleukin-10-Induced Expression of FcγRI on Human Monocytes

    PubMed Central

    Beigier-Bompadre, Macarena; Barrionuevo, Paula; Alves-Rosa, Fernanda; Rubel, Carolina J.; Palermo, Marina S.; Isturiz, Martín A.

    2001-01-01

    Three different classes of receptors for the Fc portion of immunoglobulin G (FcγRs), FcγRI, FcγRII, and FcγRIII, have been identified on human leukocytes. One of them, FcγRI, is a high-affinity receptor capable of induction of functions that include phagocytosis, respiratory burst, antibody-dependent cell-mediated cytotoxicity (ADCC), and secretion of cytokines. This receptor is expressed on mononuclear phagocytes, and this expression is regulated by cytokines and hormones such as gamma interferon (IFN-γ), IFN-β, interleukin-10 (IL-10), and glucocorticoids. We have recently demonstrated that the chemotactic peptide N-formyl-methionyl-leucyl-phenylalanine (FMLP) is capable of inducing a time-dependent downregulation of both FcγRIIIB and FcγRII in human neutrophils, altering FcγR-dependent functions. Considering the biological relevance of the regulation of FcγRI, we investigated the effect of FMLP on the overexpression of FcγRI induced by both IFN-γ and IL-10 on human monocytes. We demonstrate that FMLP significantly abrogated IFN-γ- and IL-10-induced FcγRI expression, although its basal level of expression was not altered. However, other IFN-γ-mediated effects such as the overexpression of the major histocompatibility complex class II antigens and the enhancement of lipopolysaccharide-induced secretion of tumor necrosis factor alpha were not affected by FMLP treatment. The formyl peptide completely inhibited the IFN-γ- and IL-10-induced enhancement of ADCC and phagocytosis carried out by adherent cells. The inhibitory effect of FMLP on FcγRI upregulation could exert an important regulatory effect during the evolution of bacterial infections. PMID:11238229

  2. Modulation of gene-expression profiles associated with sodium arsenite-induced cardiotoxicity by p-coumaric acid, a common dietary polyphenol.

    PubMed

    Prasanna, Nagalakshmi; Rasool, Mahaboobkhan

    2014-04-01

    In the present study, the purpose was to investigate the effect of p-coumaric acid on the mRNA-expression levels of inflammatory cytokines, transcription factor, MAP kinases, and apoptotic proteins by real time reverse transcription polymerase chain reaction in the cardiac tissue of sodium arsenite exposed rats. Sodium arsenite administration (5 mg/kg/b.wt, once daily for 30 days) upregulated the mRNA-expression levels of inflammatory cytokines (interleukin-1 beta, interleukin-6, tumor necrosis factor-alpha, and tumor growth factor-beta), transcription factor (NF-Kb-Rel A), protein kinases (Janus kinase and p38), caspase 3, and proapoptotic protein Bax in the cardiac tissue of rats, but the antiapoptotic protein Bcl-2 mRNA expression was found be downregulated. However, p-coumaric acid (75, 100 mg/kg/b. wt. oral) pretreatment daily before the sodium arsenite exposure protected the changes in the above mRNA-expression profiles observed in the cardiac tissues. In conclusion, this study confirmed that p-coumaric acid could be a promising dietary agent for protecting against the sodium arsenite-induced cardiotoxicity.

  3. Hypoxia-induced paracrine regulation of vascular endothelial growth factor receptor expression.

    PubMed Central

    Brogi, E; Schatteman, G; Wu, T; Kim, E A; Varticovski, L; Keyt, B; Isner, J M

    1996-01-01

    Vascular endothelial growth factor (VEGF)/vascular permeability factor (VPF), an endothelial cell (EC)-specific mitogen, stimulates angiogenesis in vivo, particularly in ischemic regions. VEGF/VPF expression by cells of hypoxic tissues coincides with expression of its two receptors, KDR and flt-1, by ECs in the same tissues. We investigated whether hypoxia or hypoxia-dependent conditions operate in coordinating this phenomenon. Human umbilical vein and microvascular ECs were exposed to direct hypoxia or to medium conditioned (CM) by myoblasts maintained in hypoxia for 4 d. Control ECs were maintained in normoxia or normoxia-CM. Binding of 125I-VEGF to ECs was then evaluated. Hypoxic treatment of ECs had no effect on 125I-VEGF binding. However, treatment of ECs with hypoxia-CM produced a threefold increase in 125I-VEGF binding, with peak at 24 h (P < 0.001, ANOVA). Scatchard analysis disclosed that increased binding was due to a 13-fold increase in KDR receptors/cell, with no change in KDR affinity (Kd = 260 +/- 51 pM, normoxia-CM versus Kd = 281 +/- 94 pM, hypoxia-CM) and no change in EC number (35.6 +/- 5.9 x 10(3) ECs/cm2, normoxia-CM versus 33.5 +/- 5.5 x 10(3) ECs/cm2, hypoxia-CM). Similar results were obtained using CM from hypoxic smooth muscle cells. KDR upregulation was not prevented by addition to the hypoxia-CM of neutralizing antibodies against VEGF, tumor necrosis factor-alpha, transforming growth factor beta 1 or basic fibroblast growth factor. Similarly, addition of VEGF or lactic acid to the normoxia-CM had no effect on VEGF binding. We conclude that mechanism(s) initiated by hypoxia can induce KDR receptor upregulation in ECs. Hypoxic cells, normal or neoplastic, not only can produce VEGF/VPF, but can also modulate its effects via paracrine induction of VEGF/VPF receptors in ECs. PMID:8567969

  4. Troglitazone inhibits the expression of inducible nitric oxide synthase in adipocytes in vitro and in vivo study in 3T3-L1 cells and Otsuka Long-Evans Tokushima Fatty rats.

    PubMed

    Dobashi, K; Asayama, K; Nakane, T; Kodera, K; Hayashibe, H; Nakazawa, S

    2000-09-15

    The aim of this study was to determine the mechanism of troglitazone action on nitric oxide (NO) production via inducible NO synthase (iNOS) in adipocytes in vitro and in vivo. The treatment of 3T3-L1 adipocytes with the combination of lipopolysaccharide (LPS), tumor necrosis factor-alpha and interferon-gamma synergistically induced de novo iNOS expression leading to enhanced NO production. The NO production was inhibited by co-treatment with aminoguanidine or N-nitro-L-arginine methylester hydrochloride. Troglitazone inhibited the NO production in a dose dependent manner by the suppression of iNOS expression. In the 24 week-old Otsuka Long-Evans Tokushima Fatty (OLETF) rats, the mean weight and the blood glucose were 21% and 30%, respectively, higher than in their lean counterparts. The serum nitrite concentration was increased after injection of LPS (4 mg/kg, i.p.), more markedly in OLETF rats than in the lean rats. The epididymal fats from LPS-injected groups, but not the ones from the non-injected groups, expressed mRNA and protein of iNOS. Troglitazone pre-treatment blocked the LPS-induced expression of iNOS in adipose tissue and the increase in serum nitrite concentration. These results suggest that troglitazone inhibits the cytokine-induced NO production in adipocytes by blocking iNOS expression both in vitro and in vivo.

  5. Functional analysis of a human tumor necrosis factor alpha (TNF-alpha) promoter polymorphism related to joint damage in rheumatoid arthritis.

    PubMed Central

    Kaijzel, E. L.; van Krugten, M. V.; Brinkman, B. M.; Huizinga, T. W.; van der Straaten, T.; Hazes, J. M.; Ziegler-Heitbrock, H. W.; Nedospasov, S. A.; Breedveld, F. C.; Verweij, C. L.

    1998-01-01

    BACKGROUND: Functional heterogeneity in the tumor necrosis factor alpha (TNF-alpha) gene may be responsible for the TNF-alpha response in infectious and autoimmune diseases. Recently, the TNF-238 promoter polymorphism was observed as being associated with a more destructive disease in rheumatoid arthritis (RA). To determine the relation between TNF-238 and disease progression, the extent of joint destruction in a cohort of 101 RA patients followed for 12 years was analyzed. Furthermore, we have attempted to link this polymorphism to TNF-alpha gene transcription in monocytes and lymphocytes in vitro. PATIENTS, MATERIALS, AND METHODS: The extent of joint destruction determined on X-rays of hands and feet assessed after 0, 3, 6, and 12 years was compared with TNF-238 genotypes. Functional consequences of TNF-alpha gene polymorphisms using reporter gene constructs were analyzed in cells of the monocyte and lymphocyte lineage by means of transient transfection systems. RESULTS: The rate of joint damage in -238GA patients was lower than that in the -238GG patients, independent of HLA-DR4. Damage after 12 years was 76 +/- 30 for the -238GA versus 126 +/- 13 for the -238GG patients as determined by the van der Heijde's modification of Sharp's method. Furthermore, TNF-238A was found to be in linkage disequilibrium with an additional polymorphism at position -376. Functional assays revealed no significant differences in the level of inducible reporter gene expression between the TNF-238/-376 promoter constructs in the cell types tested. CONCLUSION: In a prospective study, we show that the TNF-238GG genotype contributes to progression of joint destruction in RA, independent of the presence of HLA-DR4. However, in vitro transfection assays indicate that TNF-238A by itself or in combination with TNF-376A is not likely to be of direct functional relevance for transcriptional activation. Therefore, these polymorphisms may serve as markers for additional polymorphisms in the TNF

  6. Vocalization Induced CFos Expression in Marmoset Cortex

    PubMed Central

    Miller, Cory T.; DiMauro, Audrey; Pistorio, Ashley; Hendry, Stewart; Wang, Xiaoqin

    2010-01-01

    All non-human primates communicate with conspecifics using vocalizations, a system involving both the production and perception of species-specific vocal signals. Much of the work on the neural basis of primate vocal communication in cortex has focused on the sensory processing of vocalizations, while relatively little data are available for vocal production. Earlier physiological studies in squirrel monkeys had shed doubts on the involvement of primate cortex in vocal behaviors. The aim of the present study was to identify areas of common marmoset (Callithrix jacchus) cortex that are potentially involved in vocal communication. In this study, we quantified cFos expression in three areas of marmoset cortex – frontal, temporal (auditory), and medial temporal – under various vocal conditions. Specifically, we examined cFos expression in these cortical areas during the sensory, motor (vocal production), and sensory–motor components of vocal communication. Our results showed an increase in cFos expression in ventrolateral prefrontal cortex as well as the medial and lateral belt areas of auditory cortex in the vocal perception condition. In contrast, subjects in the vocal production condition resulted in increased cFos expression only in dorsal premotor cortex. During the sensory–motor condition (antiphonal calling), subjects exhibited cFos expression in each of the above areas, as well as increased expression in perirhinal cortex. Overall, these results suggest that various cortical areas outside primary auditory cortex are involved in primate vocal communication. These findings pave the way for further physiological studies of the neural basis of primate vocal communication. PMID:21179582

  7. The clinical significance of tumor necrosis factor-alpha plasma level in patients having chronic lymphocytic leukemia.

    PubMed

    Ferrajoli, Alessandra; Keating, Michael J; Manshouri, Taghi; Giles, Francis J; Dey, Amanda; Estrov, Zeev; Koller, Charles A; Kurzrock, Razelle; Thomas, Deborah A; Faderl, Stefan; Lerner, Susan; O'Brien, Susan; Albitar, Maher

    2002-08-15

    Tumor necrosis factor-alpha (TNF-alpha), a cytokine possessing pleiotropic biological activities, is produced by leukemic lymphocytes in patients with chronic lymphocytic leukemia (CLL) and acts as an autocrine and paracrine growth factor in this disease. In this study, TNF-alpha levels were determined in 150 patients with CLL and correlated with disease characteristics, prognostic factors, and survival. The mean TNF-alpha plasma concentration in the patients with CLL was significantly higher than in the healthy control population (16.4 versus 8.7 pg/mL; P <.0001). Patients having an elevated TNF-alpha level had more advanced Rai and Binet stage disease, higher serum beta(2)-microglobulin (beta(2)M) levels, a greater percentage of cells expressing CD38, and lower hemoglobin and platelet levels. Patients having chromosomal abnormalities such as 11q deletion, trisomy 12, and chromosome 17 aberrations had a higher mean TNF-alpha level (27.5 pg/mL) than patients having a diploid karyotype or other miscellaneous cytogenetic abnormalities (14.2 pg/mL; P <.001). The TNF-alpha level was a predictor of survival when the Cox proportional hazards model was used with TNF-alpha entered as a continuous variable (P =.0001). Also, patients having a TNF-alpha level above the mean value of 14 pg/mL had significantly shorter survival duration (P =.00001). The TNF-alpha level remained predictive of survival in Cox multivariate analysis independent of Rai staging and beta(2)M, hemoglobin, prior therapy, white cell count, and platelet level (P =.005). We conclude that the TNF-alpha level serves as a prognostic factor in patients with CLL and that inhibition of TNF-alpha in these patients could have therapeutic importance.

  8. Brucella suis prevents human dendritic cell maturation and antigen presentation through regulation of tumor necrosis factor alpha secretion.

    PubMed

    Billard, Elisabeth; Dornand, Jacques; Gross, Antoine

    2007-10-01

    Brucella is a facultative intracellular pathogen and the etiological agent of brucellosis. In some cases, human brucellosis results in a persistent infection that may reactivate years after the initial exposure. The mechanisms by which the parasite evades clearance by the immune response to chronically infect its host are unknown. We recently demonstrated that dendritic cells (DCs), which are critical components of adaptive immunity, are highly susceptible to Brucella infection and are a preferential niche for the development of the bacteria. Here, we report that in contrast to several intracellular bacteria, Brucella prevented the infected DCs from engaging in their maturation process and impaired their capacities to present antigen to naïve T cells and to secrete interleukin-12. Moreover, Brucella-infected DCs failed to release tumor necrosis factor alpha (TNF-alpha), a defect involving the bacterial protein Omp25. Exogenous TNF-alpha addition to Brucella-infected DCs restored cell maturation and allowed them to present antigens. Two avirulent mutants of B. suis, B. suis bvrR and B. suis omp25 mutants, which do not express the Omp25 protein, triggered TNF-alpha production upon DC invasion. Cells infected with these mutants subsequently matured and acquired the ability to present antigens, two properties which were dramatically impaired by addition of anti-TNF-alpha antibodies. In light of these data, we propose a model in which virulent Brucella alters the maturation and functions of DCs through Omp25-dependent control of TNF-alpha production. This model defines a specific evasion strategy of the bacteria by which they can escape the immune response to chronically infect their host.

  9. Transforming growth factor alpha: mutation of aspartic acid 47 and leucine 48 results in different biological activities.

    PubMed Central

    Lazar, E; Watanabe, S; Dalton, S; Sporn, M B

    1988-01-01

    To study the relationship between the primary structure of transforming growth factor alpha (TGF-alpha) and some of its functional properties (competition with epidermal growth factor (EGF) for binding to the EGF receptor and induction of anchorage-independent growth), we introduced single amino acid mutations into the sequence for the fully processed, 50-amino-acid human TGF-alpha. The wild-type and mutant proteins were expressed in a vector by using a yeast alpha mating pheromone promoter. Mutations of two amino acids that are conserved in the family of the EGF-like peptides and are located in the carboxy-terminal part of TGF-alpha resulted in different biological effects. When aspartic acid 47 was mutated to alanine or asparagine, biological activity was retained; in contrast, substitutions of this residue with serine or glutamic acid generated mutants with reduced binding and colony-forming capacities. When leucine 48 was mutated to alanine, a complete loss of binding and colony-forming abilities resulted; mutation of leucine 48 to isoleucine or methionine resulted in very low activities. Our data suggest that these two adjacent conserved amino acids in positions 47 and 48 play different roles in defining the structure and/or biological activity of TGF-alpha and that the carboxy terminus of TGF-alpha is involved in interactions with cellular TGF-alpha receptors. The side chain of leucine 48 appears to be crucial either indirectly in determining the biologically active conformation of TGF-alpha or directly in the molecular recognition of TGF-alpha by its receptor. PMID:3285178

  10. Does inhibition of tumor necrosis factor alpha affect chlamydial genital tract infection in mice and guinea pigs?

    PubMed

    Darville, T; Andrews, C W; Rank, R G

    2000-09-01

    The role of tumor necrosis factor alpha (TNF-alpha) in host defense against chlamydial infection remains unclear. In order to further evaluate the relevance of TNF-alpha to host resistance in chlamydial genital tract infection, we examined the effect of local inhibition of the TNF-alpha response in normal C57 mice and in interferon gamma gene-deficient C57 mice infected intravaginally with the mouse pneumonitis agent of Chlamydia trachomatis. Since the guinea pig model of female genital tract infection more closely approximates the human in terms of ascending infection and development of pathology, we also examined the effect of local inhibition of the TNF-alpha response in guinea pigs infected intravaginally with the guinea pig strain of Chlamydia psittaci. We successfully blocked the early TNF-alpha response in the respective animal models. This blockade had no effect on the numbers of organisms isolated from the genital tract during the time of TNF-alpha inhibition in mice or guinea pigs. Analysis of interleukin-1beta, macrophage inflammatory protein-2, and granulocyte macrophage-colony stimulating factor in the mouse model revealed that blockade of the TNF-alpha response did not alter the release of these proinflammatory proteins. Yet, in TNF-alpha-depleted mice, increased numbers of neutrophils were detected in the genital tract, and, in TNF-alpha-depleted guinea pigs, increased numbers of neutrophils as well as infiltrating lymphocytes were seen in the endocervix. Blockade of TNF-alpha does not affect the level of infection in mice or guinea pigs, but it may decrease TNF-alpha-induced apoptosis of infiltrating inflammatory cells.

  11. Contrast-enhanced magnetic resonance imaging of tumor-bearing mice treated with human recombinant tumor necrosis factor alpha.

    PubMed

    Aicher, K P; Dupon, J W; White, D L; Aukerman, S L; Moseley, M E; Juster, R; Rosenau, W; Winkelhake, J L; Brasch, R C

    1990-11-15

    Pharmacological effects of recombinant human tumor necrosis factor alpha (TNF) were studied in a mouse fibrosarcoma model using magnetic resonance imaging enhanced with a macromolecular contrast agent, albumin(gadolinium-diethylenetriamine pentaacetic acid)35. TNF was administered i.v. in a dose of 150 micrograms/kg, 60 to 80 min prior to imaging. Contrast-enhanced and nonenhanced magnetic resonance images of TNF-treated (n = 10) and untreated (n = 8) Meth A fibrosarcomas were obtained at 2.0 Tesla using T1-weighted spin-echo pulse sequences. Serial images spanning an interval of 60 to 120 min after TNF administration showed that the TNF-treated tumors enhanced significantly more overall than did untreated tumors (43% versus 31%). The most marked differential tumor enhancement was observed in the tumor rim (59% versus 40%). Nontumorous tissue, including muscle and brain, revealed no significant enhancement differences between TNF-treated animals and controls. The observed tumor enhancement corresponded strongly with Evans blue staining; the TNF-treated tumors stained deep blue, while untreated tumors and normal tissues observed did not stain. The different enhancement and Evans blue staining patterns between TNF-treated tumors and untreated tumors are attributed to TNF-induced changes in tumor capillary integrity. The data indicate that TNF effects on tumors include an increased capillary permeability for macromolecules at early times after administration. The ability to detect changes in capillary permeability in vivo using contrast-enhanced magnetic resonance imaging may prove to be clinically useful to monitor tumor response to TNF.

  12. Asbestos fibres and man made mineral fibres: induction and release of tumour necrosis factor-alpha from rat alveolar macrophages.

    PubMed Central

    Ljungman, A G; Lindahl, M; Tagesson, C

    1994-01-01

    OBJECTIVES--Mounting evidence suggests that asbestos fibres can stimulate alveolar macrophages to generate the potent inflammatory and fibrogenic mediator, tumour necrosis factor-alpha (TNF-alpha), and that this may play an important part in the onset and development of airway inflammation and lung fibrosis due to asbestos fibre inhalation. Little is known, however, about the ability of other mineral fibres to initiate formation and release of TNF-alpha by alveolar macrophages. Therefore the effects of different fibres (crocidolite, chrysotile A, chrysotile B, two man made mineral fibres (MMVF 21 and MMVF 22), a ceramic fibre (RCF 1), and a silicon carbide whisker fibre (SiCwh)) on formation and release of TNF-alpha by rat alveolar macrophages were examined. METHODS--Cells were isolated and incubated at 37 degrees C with the different fibres, or with culture medium alone (controls), and the amounts of TNF-alpha messenger RNA (mRNA) in the cells and TNF-alpha bioactivity released into the culture medium were measured at different time points. RESULTS--Significantly (P < 0.05 v control) increased amounts of TNF-alpha mRNA were found in cells exposed to crocidolite, chrysotile A, chrysotile B, MMVF 21, RCF 1, or SiCwh for 90 minutes, and significantly (P < 0.05 v control) increased activities of TNF-alpha were found in the medium of macrophages exposed to crocidolite, chrysotile A, chrysotile B, or MMVF 21 for four hours. CONCLUSION--These observations suggest that not only natural mineral fibres but also certain man made mineral fibres are able to induce the formation and release of TNF-alpha by alveolar macrophages in vitro. Images Figure 1 Figure 3 PMID:7849857

  13. Pulmonary microRNA expression profiling in an immature piglet model of cardiopulmonary bypass-induced acute lung injury.

    PubMed

    Li, Wenlei; Ma, Kai; Zhang, Sen; Zhang, Hao; Liu, Jinping; Wang, Xu; Li, Shoujun

    2015-04-01

    After surgery performed under cardiopulmonary bypass (CPB), severe lung injury often occurs in infants. MicroRNAs (miRNAs) are potentially involved in diverse pathophysiological processes via regulation of gene expression. The objective of this study was to investigate differentially expressed miRNAs and their potential target genes in immature piglet lungs in response to CPB. Fourteen piglets aged 18.6 ± 0.5 days were equally divided into two groups that underwent sham sternotomy or CPB. The duration of aortic cross-clamping was 2 h, followed by 2 h reperfusion. Lung injury was evaluated by lung function indices, levels of cytokines, and histological changes. We applied miRNA microarray and quantitative real-time polymerase chain reaction (qRT-PCR) analysis to determine miRNA expression. Meanwhile, qRT-PCR and enzyme-linked immunosorbent assay were used for validation of predicted mRNA targets. The deterioration of lung function and histopathological changes revealed the piglets' lungs were greatly impaired due to CPB. The levels of tumor necrosis factor alpha, interleukin 6, and interleukin 10 increased in the lung tissue after CPB. Using miRNA microarray, statistically significant differences were found in the levels of 16 miRNAs in the CPB group. Up-regulation of miR-21 was verified by PCR. We also observed down-regulation in the levels of miR-127, miR-145, and miR-204, which were correlated with increases in the expression of the products of their potential target genes PIK3CG, PTGS2, ACE, and IL6R in the CPB group, suggesting a potential role for miRNA in the regulation of inflammatory response. Our results show that CPB induces severe lung injury and dynamic changes in miRNA expression in piglet lungs. Moreover, the changes in miRNA levels and target gene expression may provide a basis for understanding the pathogenesis of CPB-induced injury to immature lungs.

  14. Costimulator B7-1 confers antigen-presenting-cell function to parenchymal tissue and in conjunction with tumor necrosis factor alpha leads to autoimmunity in transgenic mice.

    PubMed Central

    Guerder, S; Picarella, D E; Linsley, P S; Flavell, R A

    1994-01-01

    Tolerance to peripheral antigens is thought to result from the inability of parenchymal tissue to stimulate T cells--an inability that is believed to relate to the lack of expression of the costimulatory signal(s) required for T-cell activation. To test this model, we generated transgenic mice expressing costimulatory molecule B7-1 on the B cells of the pancreas. We find that islets from these transgenic mice are immunogenic for naive T cells in vitro and in vivo. Nonetheless, mice expressing the costimulator B7-1 specifically on beta cells do not develop diabetes, suggesting that expression of the B7-1 costimulator is not sufficient to abrogate the tolerance to peripheral antigens. We have reported that tumor necrosis factor alpha subunit (TNF-alpha) expressed by beta cells leads to a local inflammation but no islet destruction. Strikingly, however, the combination of a local inflammation due to the expression of the cytokine TNF-alpha and the expression of B7-1 results in tissue destruction and diabetes. Images PMID:7515187

  15. Tristetraprolin Down-Regulation Contributes to Persistent TNF-Alpha Expression Induced by Cigarette Smoke Extract through a Post-Transcriptional Mechanism

    PubMed Central

    Cheng, Ming-Liang; Zhang, Quan; Mu, Mao; Li, Hong; Luo, Yuan; Liang, Yue-Dong; Luo, Xin-Hua; Gao, Chang-Qing; Jackson, Patricia L.; Wells, J. Michael; Zhou, Yong; Hu, Meng; Cai, Guoqiang; Thannickal, Victor J.; Steele, Chad; Blalock, J. Edwin; Han, Xiaosi; Chen, Ching-Yi; Ding, Qiang

    2016-01-01

    Rationale Tumor necrosis factor-alpha (TNF-α) is a potent pro-inflammatory mediator and its expression is up-regulated in chronic obstructive pulmonary disease (COPD). Tristetraprolin (TTP) is implicated in regulation of TNF-α expression; however, whether TTP is involved in cigarette smoke-induced TNF-α expression has not been determined. Methods TTP expression was examined by western blot analysis in murine alveolar macrophages and alveolar epithelial cells challenged without or with cigarette smoke extract (CSE). TNF-α mRNA stability, and the decay of TNF-α mRNA, were determined by real-time quantitative RT-PCR. TNF-α protein levels were examined at the same time in these cells. To identify the molecular mechanism involved, a construct expressing the human beta-globin reporter mRNA containing the TNF-α 3’-untranslated region was generated to characterize the TTP targeted site within TNF-α mRNA. Results CSE induced TTP down-regulation in alveolar macrophages and alveolar epithelial cells. Reduced TTP expression resulted in significantly increased TNF-α mRNA stability. Importantly, increased TNF-α mRNA stability due to impaired TTP function resulted in significantly increased TNF-α levels in these cells. Forced TTP expression abrogated the increased TNF-α mRNA stability and expression induced by CSE. By using the globin reporter construct containing TNF-α mRNA 3’-untranslated region, the data indicate that TTP directly targets the adenine- and uridine-rich region (ARE) of TNF-α mRNA and negatively regulates TNF-α expression at the post-transcriptional level. Conclusion The data demonstrate that cigarette smoke exposure reduces TTP expression and impairs TTP function, resulting in significantly increased TNF-α mRNA stability and excessive TNF-α expression in alveolar macrophages and epithelial cells. The data suggest that TTP is a novel post-transcriptional regulator and limits excessive TNF-α expression and inflammatory response induced by

  16. An IPTG Inducible Conditional Expression System for Mycobacteria

    PubMed Central

    Ravishankar, Sudha; Ambady, Anisha; Ramu, Haripriya; Mudugal, Naina Vinay; Tunduguru, Ragadeepthi; Anbarasu, Anand; Sharma, Umender K.; Sambandamurthy, Vasan K.; Ramaiah, Sudha

    2015-01-01

    Conditional expression strains serve as a valuable tool to study the essentiality and to establish the vulnerability of a target under investigation in a drug discovery program. While essentiality implies an absolute requirement of a target function, vulnerability provides valuable information on the extent to which a target function needs to be depleted to achieve bacterial growth inhibition followed by cell death. The critical feature of an ideal conditional expression system is its ability to tightly regulate gene expression to achieve the full spectrum spanning from a high level of expression in order to support growth and near zero level of expression to mimic conditions of gene knockout. A number of bacterial conditional expression systems have been reported for use in mycobacteria. The utility of an isopropylthiogalactoside (IPTG) inducible system in mycobacteria has been reported for protein overexpression and anti-sense gene expression from a replicating multi-copy plasmid. Herein, we report the development of a versatile set of non-replicating IPTG inducible vectors for mycobacteria which can be used for generation of conditional expression strains through homologous recombination. The role of a single lac operator versus a double lac operator to regulate gene expression was evaluated by monitoring the expression levels of β-galactosidase in Mycobacterium smegmatis. These studies indicated a significant level of leaky expression from the vector with a single lac operator but none from the vector with double lac operator. The significance of the double lac operator vector for target validation was established by monitoring the growth kinetics of an inhA, a rpoB and a ftsZ conditional expression strain grown in the presence of different concentrations of IPTG. The utility of this inducible system in identifying target specific inhibitors was established by screening a focussed library of small molecules using an inhA and a rpoB conditional expression

  17. Tumor Necrosis Factor-alpha Stimulates the Overproduction of Intestinal Apolipoprotein B48-containing Very Low Density Lipoproproteins

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Tumor necrosis factor-alpha(a)(TNFa), a proinflammatory cytokine, is involved in obesity-associated pathologies including type 2 diabetes and atherosclerosis. TNFa enhanced postprandial apoB48-VLDL1 overproduction by about 89% compared with the control after 90 min olive oil loading; TNFa did not si...

  18. Early activation of splenic macrophages by tumor necrosis factor alpha is important in determining the outcome of experimental histoplasmosis in mice.

    PubMed Central

    Wu-Hsieh, B A; Lee, G S; Franco, M; Hofman, F M

    1992-01-01

    Experimental infection of animals with Histoplasma capsulatum caused a massive macrophage infiltration into the spleen and induced the production of tumor necrosis factor alpha (TNF-alpha) locally. The cytokine was also produced in vitro by peritoneal exudate macrophages exposed to a large inoculum of yeast cells. Depletion of the cytokine by injection of polyclonal sheep anti-TNF-alpha antibody was detrimental to sublethally infected mice. Fungous burdens in the spleens of TNF-alpha-depleted mice were higher than they were in the infected control mice at days 2, 7, and 9 after infection, and the antibody-treated animals succumbed to the infection. Histopathological study of spleen sections revealed that splenic macrophages were not able to control proliferation of intracellular yeasts as a result of TNF-alpha depletion. It seems that TNF-alpha plays a role in early activation of splenic macrophages which is important in controlling the outcome of an infection. Images PMID:1398934

  19. Differential expression of uterine NO in pregnant and nonpregnant rats with intrauterine bacterial infection.

    PubMed

    Fang, L; Nowicki, B; Yallampalli, C

    2001-05-01

    Previous studies have demonstrated that nitric oxide (NO) is involved in the uterine host defense against bacterial infection. In nonpregnant rats, NO production in the uterus was shown to be lower, and inducible NO synthase (NOS) expression was undetectable. However, studies in pregnant rats show abundant expression of inducible NOS with significant elevation in NO production in the uterus. We have recently reported that intrauterine Escherichia coli infection caused a localized increase in uterine NO production and inducible NOS expression in the nonpregnant rat. In our present study, we examined whether the uterine NO production, NOS expression, and uterine tumor necrosis factor-alpha protein are increased in pregnant rats with intrauterine pathogenic Escherichia coli infection. Unlike the nonpregnant state, the NO production in the infected uterine horn of pregnant rats was not significantly elevated after bacterial inoculation compared with the contralateral uterine horn. The expression of uterine NOS (types II and III) also did not show significant upregulation in the infected horn. This is in contrast to that in nonpregnant animals, in which type II NOS was induced in the uterus on infection. Moreover, intrauterine infection induced an elevated expression of tumor necrosis factor-alpha protein in the infected horn both of nonpregnant and of pregnant rats. These data suggest that the sequential stimulation of NOS expression, especially the inducible isoform, and generation of uterine NO are lacking during pregnancy despite an elevated tumor necrosis factor-alpha after infection. In summary, NO synthesis response may be maximal at pregnancy, and infection may not further induce the NO system. Present studies, together with our previous report that intrauterine infection-induced lethality in pregnancy rats was amplified with the inhibition of NO, suggest that pregnancy is a state predisposed for increased complications associated with intrauterine infection and

  20. Detection of tumour necrosis factor-alpha in dogs with lymphoma(*).

    PubMed

    Hofer, J; DeFrancesco, T C; Williams, L E

    2011-12-01

    Tumour necrosis factor-alpha (TNF-α) production by malignant lymphoblasts has been identified in vitro and in vivo in mice and humans, respectively. The goals of this study were (1) to evaluate a novel single-sample TNF-α assay and (2) to determine whether TNF-α is increased in dogs with lymphoma prior to and following treatment. Canine TNF-α was analysed concurrently using the novel Siemens Immulite® single-sample automated ELISA and the previously validated Quantikine® standard ELISA. Serum from dogs with lymphoma and from breed-, age- and gender-matched control dogs was evaluated at two time points. Three of 25 (12%) dogs with lymphoma had detectable TNF-α at diagnosis, whereas none had detectable TNF-α following complete or partial remission. TNF-α was not detectable in control dogs. Despite 91% homology between human and canine TNF-α, the Immulite® automated ELISA failed to detect canine TNF-α. Serum TNF-α appears to have limited value as a tumour marker in dogs with lymphoma.

  1. Interleukin 10 and Tumor Necrosis Factor-Alpha in Pregnancy: Aspects of Interest in Clinical Obstetrics

    PubMed Central

    Brogin Moreli, Jusciele; Cirino Ruocco, Ana Maria; Vernini, Joice Monaliza; Rudge, Marilza Vieira Cunha; Calderon, Iracema Mattos Paranhos

    2012-01-01

    The purpose of this study was to review the literature regarding the action of the cytokines interleukin 10 (IL-10) and tumor necrosis factor-alpha (TNF-α) in pregnancy and to emphasize the factors that are of interest to clinical obstetrics. The literature highlights several actions of IL-10 and TNF-α during pregnancy. The actions of these cytokines seem to be antagonistic and dependent on the balance between them, which is orchestrated by the specific immunosuppressive action of IL-10. TNF-α has a characteristic inflammatory action, and it is an additional diabetogenic factor in pregnancy. The loss of the control of the production of these cytokines, with increase of TNF-α, is related to the risk for developing obstetric complications, particularly recurrent fetal loss, gestational diabetes mellitus, hypertensive syndromes, and fetal growth restriction. However, study results are controversial and are not clearly defined. These issues are attributed to the heterogeneity of the studies, particularly regarding their sample sizes and sources, the evaluation methods, and the multiplicity of factors and conditions that influence cytokine production. These questions are fundamental and should be addressed in future investigations to obtain more consistent results that can be applied to obstetric practice. PMID:22462002

  2. Interleukin-6 and tumor necrosis factor-alpha values in elk neonates

    USGS Publications Warehouse

    Barber-Meyer, S. M.; Johnson, C.R.; Murtaugh, M.P.; Mech, L.D.; White, P.J.

    2007-01-01

    Serological indicators of general condition would be helpful for monitoring or assessing ungulate wildlife. Toward that end, we report the 1st reference values for 2 cytokines, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-??), in neonatal elk (Cervus elaphus). We obtained blood samples from 140 calves ??? 6 days old in Yellowstone National Park during summer 2003-2005. TL-6 values ranged from 0 to 1.21 pg/ml with a median of 0.03 pg/ml. TNF-?? values ranged from 0 to 225.43 pg/ml with a median of 1.85 pg/ml. IL-6 and TNF-?? concentrations were not significant predictors of elk calf survival through 21 days. Development of ungulate-based IL-6 and TNF-?? assays that provide greater sensitivity than cross-reacting human-based assays could be helpful in monitoring ungulate condition and health status comparisons among herds. Such information could provide indirect assessments of range quality or environmental influences among herds. ?? 2007 American Society of Mammalogists.

  3. Increased tumour necrosis factor alpha production by neutrophils in patients with hepatitis B.

    PubMed Central

    Fan, X; Zhang, Z

    1994-01-01

    AIMS--To investigate the role of serum and neutrophil tumour necrosis factor alpha (TNF alpha) in patients with viral hepatitis. METHODS--The activities of serum and neutrophil TNF alpha were measured using a bioassay of in vitro cytotoxicity against L929 cells in 57 patients with viral hepatitis and 20 healthy blood donors. RESULTS--Both serum and neutrophil TNF alpha in patients with chronic active hepatitis (CAH) and subacute fulminant hepatitis (SAFH) increased compared with those in normal controls (p < 0.01). No such differences were seen in patients with acute hepatitis. Serum and neutrophil TNF alpha were obviously reduced in patients with CAH and SAFH during convalescence compared with the active period (p < 0.05; p < 0.01). Furthermore, serum TNF alpha was significantly increased in patients with SAFH and complications compared with those without (p < 0.01), and in patients with SAFH who died compared with those who survived (p < 0.01). Neutrophil TNF alpha was significantly higher in patients with SAFH and secondary bacterial infections (p < 0.05). CONCLUSIONS--Production of serum and neutrophil TNF alpha is increased in patients with CAH and SAFH, suggesting that neutrophil TNF alpha causes liver injury in these patients. PMID:8089217

  4. Plasma Levels of Tumor Necrosis Factor-Alpha and Interleukin-6 in Obsessive Compulsive Disorder

    PubMed Central

    Konuk, N.; Tekın, I. O.; Ozturk, U.; Atik, L.; Atasoy, N.; Bektas, S.; Erdogan, A.

    2007-01-01

    Aim. Recent research implicated place of an immune mechanism in the pathophysiology of obsessive-compulsive disorder (OCD). Despite increasing evidence involvement of cytokine release in OCD, results of the studies are inconsistent. The aim of this study was to evaluate the plasma levels of the cytokines; tumor necrosis factor-alpha (TNF-α) and interleukin-6 (IL-6) in OCD patients. Methods. Plasma concentrations of TNF-α and IL-6 were measured in 31 drug-free outpatients with OCD, and 31-year age and sex-matched healthy controls. TNF-α and IL-6 concentrations in blood were determined by enzyme-linked immunosorbent assay (ELISA). Results. Both TNF-α and IL-6 levels showed statistically significant increases in OCD patients compared to controls (P < .000, P < .001, resp.). In addition, the age of onset was negatively correlated with TNF-α level (r = −.402, P = .025) and duration of illness was weakly correlated with IL-6 levels (r : .357; P : .048) in patients group. Conclusion. OCD patients showed increases in TNF-α and IL-6 levels compared to the healthy controls. This study provides evidence for alterations in the proinflamatory cytokines which suggest the involvement of the immune system in the pathophysiology of OCD. PMID:17497035

  5. Anti-tumor necrosis factor alpha for retinal diseases: current knowledge and future concepts.

    PubMed

    Mirshahi, Alireza; Hoehn, René; Lorenz, Katrin; Kramann, Christina; Baatz, Holger

    2012-01-01

    Tumor necrosis factor alpha (TNF-α) is a pro-inflammatory cytokine produced by macrophages and T-cells. It plays an important role both in inflammation and apoptosis. In the eye, TNF-α appears to have a role in the pathogenesis of inflammatory, edematous, neovascular and neurodegenerative disorders. Several TNF-blocking drugs have been developed and approved, and are in clinical use for inflammatory diseases such as rheumatoid arthritis, psoriasis and ankylosing spondylitis. TNF-α blockers are widely used in ophthalmology as an off-label alternative to "traditional" immunosuppressive and immune-modulatory treatments in noninfectious uveitis. Preliminary studies suggest a positive effect of intravenously administered TNF-α blockers, mainly infliximab, for treating refractory diabetic macular edema and neovascular age-related macular degeneration. Unfortunately, much of the current data raises considerable safety concerns for intravitreal use of TNF-α inhibitors, in particular, intraocular inflammatory responses have been reported after intravitreal injection of infliximab. Results of dose-finding studies and humanized antibody or antibody fragments (e.g. adalimumab) are anticipated in the coming years; these will shed light on potential benefits and risks of local and systemic TNF-α blockers used for treatment of diseases of the retina and choroid.

  6. Diagnostic accuracy of tumor necrosis factor-alpha assay for tuberculous pleurisy

    PubMed Central

    Li, Min; Luo, Zhuang; Zhu, Wenye; Khan, Rana Sami Ullah; Ummair, Saeed Ummai; Shi, Shaoqing

    2016-01-01

    Abstract Background: The diagnosis of tuberculous pleurisy is difficult and traditional methods are not always helpful. Many studies have focused on the tumor necrosis factor-alpha (TNF-α) assay in pleural effusion for the diagnosis of tuberculous pleurisy, but the results remain controversial. This meta-analysis was conducted to determine the overall diagnostic accuracy of TNF-α. Methods: Relevant studies were searched from PubMed, Embase, Web of Science, Chinese National Knowledge Infrastructure (CNKI), Wangfang, and Weipu. We pooled the published results and computed the accuracy measures, including sensitivity, specificity, positive likelihood ratio (PLR), negative likelihood ratio (NLR), and diagnostic odds ratio (DOR). Receiver operating characteristic curves (SROC) and the area under the curve (AUC) were used to summarize the overall test performance. Results: Twelve studies with 1022 patients met the inclusion criteria. The pooled sensitivity and specificity were 0.85 (95%CI, 0.81–0.89) and 0.80 (95% CI, 0.77–0.83) respectively. The area under the SROC curve was 0.89. Conclusions: The results of meta-analysis suggested that the TNF-α assay plays a vital role in the diagnosis of tuberculous pleurisy, whereas other test results or clinical findings should be interpreted together with the TNF-α assay to improve the overall diagnostic accuracy. PMID:27902616

  7. Nickel hydroxy carbonate increases tumour necrosis factor alpha and interleukin 6 secretion by alveolar macrophages.

    PubMed

    Arsalane, K; Gosset, P; Hildebrand, H F; Voisin, C; Tonnel, A B; Wallaert, B

    1994-01-01

    The aim of the current study was to assess the in vitro effects of nickel hydroxy carbonate (NiHC) at noncytotoxic concentrations on the production of cytokines such as tumour necrosis factor alpha (TNF-alpha) and interleukin 6 (IL-6) in alveolar macrophages (AMs). The effect of NiHC was evaluated in both unstimulated AMs and cells activated by lipopolysaccharide (LPS). Cytotoxicity was related to lactate dehydrogenase release and ATP cell content. The results confirm that NiHC at concentrations of 0.125, 1.25 and 3.125 micrograms NiHC 10(-6) cells was not cytotoxic. The NiHC exposure of unstimulated AMs significantly increased the release of TNF-alpha at all concentrations and that of IL-6 at 1.25 micrograms NiHC 10(-6) cells. LPS addition significantly increased the secretion of both cytokines. However, NiHC did not cause a significant increase in the release of TNF-alpha and IL-6 in LPS-stimulated cells. In conclusion, the ability of NiHC to activate AMs and to release increased amounts of pro-inflammatory mediators may be responsible, at least partly, for inflammation and pneumotoxicity associated with nickel exposure.

  8. Structure-function analysis of synthetic and recombinant derivatives of transforming growth factor alpha.

    PubMed Central

    Defeo-Jones, D; Tai, J Y; Wegrzyn, R J; Vuocolo, G A; Baker, A E; Payne, L S; Garsky, V M; Oliff, A; Riemen, M W

    1988-01-01

    Transforming growth factor alpha (TGF-alpha) is a 50-amino-acid peptide that stimulates cell proliferation via binding to cell surface receptors. To identify the structural features of TGF-alpha that govern receptor-ligand interactions, we prepared synthetic peptide fragments and recombinant mutant proteins of TGF-alpha. These TGF-alpha derivatives were tested in receptor binding and mitogenesis assays. Synthetic peptides representing the N terminus, the C terminus, or the individual disulfide constrained rings of TGF-alpha did not exhibit receptor-binding or mitogenic activity. Replacement of the cysteines with alanines at positions 8 and 21, 16 and 32, and 34 and 43 or at positions 8 and 21 and 34 and 43 yielded inactive mutant proteins. However, mutant proteins containing substitutions or deletions in the N-terminal region retained significant biologic activity. Conservative amino acid changes at residue 29 or 38 or both and a nonconservative amino acid change at residue 12 had little effect on binding or mitogenesis. However, nonconservative amino acid changes at residues 15, 38, and 47 produced dramatic decreases in receptor binding (23- to 71-fold) and mitogenic activity (38- to 125-fold). These studies indicate that at least three distinct regions of TGF-alpha contribute to biologic activity. PMID:2850475

  9. Herbicide safener-inducible gene expression in Arabidopsis thaliana.

    PubMed

    De Veylder, L; Van Montagu, M; Inzé, D

    1997-05-01

    The potential use of a new chemical-inducible gene expression system in Arabidopsis thaliana has been examined. The system is based on the maize In2-2 promoter which is activated by benzenesulfonamide herbicide safeners. Plants transformed with the beta-glucuronidase (gus) reporter gene under the control of the In2-2 promoter were grown in the presence of different safeners and the induced GUS activity pattern was studied histochemically. In the absence of safeners, the In2-2 promoter was not active. Application of different safeners induced distinct gus expression patterns, including expression in the root, hydathodes, and the shoot apical meristem. Plants maintained continuously on inducing concentrations of the safeners were retarded in growth. The growth inhibition effects of the Sa5 safener could be overcome in a sulfonylurea-resistant background. In2-2 promoter activity could also be induced by the sulfonylurea herbicide chlorsulfuron. In the sulfonylurea-resistant background, which derives from herbicide-resistant acetolactate synthase activity, induction of the In2-2 promoter by chlorsulfuron was lower. Furthermore, branched-chain amino acids, known to inhibit acetolactate synthase activity, also induced In2-2 promoter activity. Our data suggest a strong correlation between In2-2 expression and inhibition of the acetolactate synthase activity.

  10. Stretch-induced VEGF expression in the heart.

    PubMed Central

    Li, J; Hampton, T; Morgan, J P; Simons, M

    1997-01-01

    Vascular endothelial growth factor (VEGF) is an endothelial cell mitogen involved in vascular development and angiogenesis. Recently we have observed increased VEGF expression in the normal myocardium after myocardial infarction in a rat heart. This study was designed to explore the mechanism responsible for this increase in VEGF expression. Induction of myocardial stretch in an isolated perfused Langendorff preparation by inflation of an intraventricular balloon to an end-diastolic load of 35 mmHg for 30 min resulted in a nearly sixfold increase in VEGF message level not only in the chamber subjected to stretch (left ventricle) but also in the unstretched right ventricle, thus raising the possibility of a soluble factor mediating stretch- induced induction of VEGF expression. This was further confirmed by demonstrating that coronary venous effluent collected from the stretched heart and used to perfuse isolated hearts in which no balloon was present was able to induce VEGF expression in these normal hearts. Inhibition of TGF-beta activity using a neutralizing antibody, but not antagonists/inhibitors of endothelin and angiotensin II, eliminated stretch-induced increase in VEGF expression. Staurosporine, a protein kinase C inhibitor, also blocked stretch-induced increase of VEGF expression. Measurement of TGF-beta concentration in the perfusate demonstrated increased amounts of the cytokine after myocardial stretch, and addition of TGF-beta protein to the perfusion buffer resulted in increased VEGF expression in control hearts. These results suggest that stretch-induced increase of VEGF expression in the heart is mediated at least in part by TGF-beta. PMID:9202052

  11. Regulation of hepcidin expression by inflammation-induced activin B

    PubMed Central

    Kanamori, Yohei; Sugiyama, Makoto; Hashimoto, Osamu; Murakami, Masaru; Matsui, Tohru; Funaba, Masayuki

    2016-01-01

    Activin B is induced in response to inflammation in the liver and enhances hepcidin expression, but the source of activin B and the molecular mechanism underlying hepcidin induction are not clear yet. Lipopolysaccharide (LPS)-induced inflammation induced inhibin βB but not inhibin α or inhibin βA expression in the liver, implicating activin B induction. Immunoreactive inhibin βB was detected in endothelial cells and Kupffer cells in LPS-treated liver. Activin B, but not activin A or activin AB, directly increased hepcidin expression. Activin B induced phosphorylation and activation of Smad1/5/8, the BMP-regulated (BR)-Smads. The stimulation of hepcidin transcription by activin B was mediated by ALK2 and ActRIIA, receptors for the TGF-β family. Unexpectedly, activin B-induced hepcidin expression and BR-Smad phosphorylation were resistant to the effects of LDN-193189, an ALK2/3/6 inhibitor. ALK2 and ActRIIA complex formation in response to activin B may prevent the approach of LDN-193189 to ALK2 to inhibit its activity. Activin B also induced phosphorylation of Smad2/3, the TGF-β/activin-regulated (AR)-Smad, and increased expression of connective tissue growth factor, a gene related to liver fibrogenesis, through ALK4 and ActRIIA/B. Activin B-induced activation of the BR-Smad pathway was also detected in non-liver-derived cells. The present study reveals the broad signaling of activin B, which is induced in non-parenchymal cells in response to hepatic inflammation, in hepatocytes. PMID:27922109

  12. Radiocurability by Targeting Tumor Necrosis Factor-{alpha} Using a Bispecific Antibody in Carcinoembryonic Antigen Transgenic Mice

    SciTech Connect

    Larbouret, Christel; Robert, Bruno; Linard, Christine; Teulon, Isabelle; Gourgou, Sophie M.Sc.; Bibeau, Frederic; Martineau, Pierre; Santoro, Lore; Pouget, Jean-Pierre; Pelegrin, Andre; Azria, David

    2007-11-15

    Purpose: Tumor necrosis factor-{alpha} (TNF-{alpha}) enhances radiotherapy (RT) killing of tumor cells in vitro and in vivo. To overcome systemic side effects, we used a bispecific antibody (BsAb) directed against carcinoembryonic antigen (CEA) and TNF-{alpha} to target this cytokine in a CEA-expressing colon carcinoma. We report the evaluation of this strategy in immunocompetent CEA-transgenic mice. Methods and Materials: The murine CEA-transfected colon carcinoma MC-38 was used for all experiments. In vitro, clonogenic assays were performed after RT alone, TNF-{alpha} alone, and RT plus TNF-{alpha}. In vivo, the mice were randomly assigned to treatment groups: control, TNF-{alpha}, BsAb, BsAb plus TNF-{alpha}, RT, RT plus TNF-{alpha}, and RT plus BsAb plus TNF-{alpha}. Measurements of endogenous TNF-{alpha} mRNA levels and evaluation of necrosis (histologic evaluation) were assessed per treatment group. Results: In vitro, combined RT plus TNF-{alpha} resulted in a significant decrease in the survival fraction at 2 Gy compared with RT alone (p < 0.00001). In vivo, we observed a complete response in 5 (50%) of 10, 2 (20%) of 10, 2 (18.2%) of 11, and 0 (0%) of 12 treated mice in the RT plus BsAb plus TNF-{alpha}, RT plus TNF-{alpha}, RT alone, and control groups, respectively. This difference was statistically significant when TNF-{alpha} was targeted with the BsAb (p = 0.03). The addition of exogenous TNF-{alpha} to RT significantly increased the endogenous TNF-{alpha} mRNA level, particularly when TNF-{alpha} was targeted with BsAb (p < 0.01). The percentages of necrotic area were significantly augmented in the RT plus BsAb plus TNF-{alpha} group. Conclusion: These results suggest that targeting TNF-{alpha} with the BsAb provokes RT curability in a CEA-expressing digestive tumor syngenic model and could be considered as a solid rationale for clinical trials.

  13. Assessment of Anti-TNF-α Activities in Keratinocytes Expressing Inducible TNF- α: A Novel Tool for Anti-TNF-α Drug Screening

    PubMed Central

    Udommethaporn, Sutthirat; Tencomnao, Tewin; McGowan, Eileen M.

    2016-01-01

    Tumor necrosis factor alpha (TNF-α) is a pro-inflammatory cytokine important in normal and pathological biological processes. Newly synthesized pro-TNF-α is expressed on the plasma membrane and cleaved to release soluble TNF-α protein: both are biologically active. Secreted TNF-α signals through TNF receptors and the membrane-bound TNF-α acts by cell contact-dependent signaling. Anti-TNF-α antibodies have been used effectively for treatment of chronic inflammation, however with adverse side effects. Thus, there is a need for new anti-TNF-α small molecule compounds. Anti-TNF-α activity assays involve treatment of keratinocytes with exogenous TNF-α before or after anti-TNF-α incubation. However, this model fails to address the dual signaling of TNF-α. Here we describe a Doxycycline (Dox)-inducible TNF-α (HaCaT-TNF-α) expression system in keratinocytes. Using this in-vitro model, we show cell inhibition and induced expression of pro-inflammatory cytokines and markers, including IL-1β, IL-6, IL-8, NF-κB1, and KRT-16, similar to cells treated with exogenous TNF-α. Sufficient secreted TNF-α produced also activated IL-1β and IL-8 expression in wt HaCaT cells. Importantly, stimulated expression of IL-1β and IL-8 in HaCaT-TNF-α were blocked by Quercetin, a flavanol shown to possess anti-TNF-α activities. This novel in vitro cell model provides an efficient tool to investigate the dual signaling of TNF-α. Importantly, this model provides an effective, fast, and simple screening for compounds with anti-TNF-α activities for chronic inflammatory disease therapies. PMID:27415000

  14. Leptin deficiency-induced obesity exacerbates ultraviolet B radiation-induced cyclooxygenase-2 expression and cell survival signals in ultraviolet B-irradiated mouse skin

    SciTech Connect

    Sharma, Som D.; Katiyar, Santosh K.

    2010-05-01

    Obesity has been implicated in several inflammatory diseases and in different types of cancer. Chronic inflammation induced by exposure to ultraviolet (UV) radiation has been implicated in various skin diseases, including melanoma and nonmelanoma skin cancers. As the relationship between obesity and susceptibility to UV radiation-caused inflammation is not clearly understood, we assessed the role of obesity on UVB-induced inflammation, and mediators of this inflammatory response, using the genetically obese (leptin-deficient) mouse model. Leptin-deficient obese (ob/ob) mice and wild-type counterparts (C57/BL6 mice) were exposed to UVB radiation (120 mJ/cm{sup 2}) on alternate days for 1 month. The mice were then euthanized and skin samples collected for analysis of biomarkers of inflammatory responses using immunohistochemistry, western blotting, ELISA and real-time PCR. Here, we report that the levels of inflammatory responses were higher in the UVB-exposed skin of the ob/ob obese mice than those in the UVB-exposed skin of the wild-type non-obese mice. The levels of UVB-induced cyclooxygenase-2 expression, prostaglandin-E{sub 2} production, proinflammatory cytokines (i.e., tumor necrosis factor-alpha, interleukin-1beta, interleukin-6), and proliferating cell nuclear antigen and cell survival signals (phosphatidylinositol-3-kinase and p-Akt-Ser{sup 473}) were higher in the skin of the ob/ob obese mice than the those in skin of their wild-type non-obese counterparts. Compared with the wild-type non-obese mice, the leptin-deficient obese mice also exhibited greater activation of NF-kappaB/p65 and fewer apoptotic cells in the UVB-irradiated skin. Our study suggests for the first time that obesity in mice is associated with greater susceptibility to UVB-induced inflammatory responses and, therefore, obesity may increase susceptibility to UVB-induced inflammation-associated skin diseases, including the risk of skin cancer.

  15. Tumor necrosis factor alpha promotes the proliferation of human nucleus pulposus cells via nuclear factor-κB, c-Jun N-terminal kinase, and p38 mitogen-activated protein kinase.

    PubMed

    Wang, Xiao-Hu; Hong, Xin; Zhu, Lei; Wang, Yun-Tao; Bao, Jun-Ping; Liu, Lei; Wang, Feng; Wu, Xiao-Tao

    2015-04-01

    Although tumor necrosis factor alpha (TNF-α) is known to play a critical role in intervertebral disc (IVD) degeneration, the effect of TNF-α on nucleus pulposus (NP) cells has not yet been elucidated. The aim of this study was to explore the effect of TNF-α on proliferation of human NP cells. NP cells were treated with different concentrations of TNF-α. Cell proliferation was determined by cell counting kit-8 (CCK-8) analysis and Ki67 immunofluorescence staining, and expression of cyclin B1 was studied by quantitative real-time RT-PCR. Cell cycle was measured by flow cytometry and cell apoptosis was analyzed using an Annexin V-fluorescein isothiocyanate (FITC) & propidium iodide (PI) apoptosis detection kit. To identify the mechanism by which TNF-α induced proliferation of NP cells, selective inhibitors of major signaling pathways were used and Western blotting was carried out. Treatment with TNF-α increased cell viability (as determined by CCK-8 analysis) and expression of cyclin B1 and the number of Ki67-positive and S-phase NP cells, indicating enhancement of proliferation. Consistent with this, NP cell apoptosis was suppressed by TNF-α treatment. Moreover, inhibition of NF-κB, c-Jun N-terminal kinase (JNK), and p38 mitogen-activated protein kinase (MAPK) blocked TNF-α-stimulated proliferation of NP cells. In conclusion, the current findings suggest that the effect of TNF-α on IVD degeneration involves promotion of the proliferation of human NP cells via the NF-κB, JNK, and p38 MAPK pathways.

  16. Production of tumor necrosis factor-alpha and interferon-gamma from human peripheral blood lymphocytes by MGN-3, a modified arabinoxylan from rice bran, and its synergy with interleukin-2 in vitro.

    PubMed

    Ghoneum, M; Jewett, A

    2000-01-01

    Recently, we presented evidence for the role of MGN-3, an enzymatically modified arabinoxylan extracted from rice bran, in potent activation of human natural killer (NK) cell function in vivo and in vitro. In the current study, we examined the mechanism by which MGN-3 elevated NK cytotoxic activity. We did this by testing the action of MGN-3 on the levels of both tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) secretions and MGN-3 function on the expression of key cell surface receptors. Peripheral blood lymphocytes were treated with MGN-3 at concentrations of 0.1 mg/ml and 1 mg/ml, and supernatants were subjected to enzyme-linked immunosorbent assay. Results showed that MGN-3 is a potent TNF-alpha inducer. The effect was dose-dependent. MGN-3 concentration at 0.1 and 1 mg/ml increased TNF-alpha production by 22.8- and 47. 1-fold, respectively. MGN-3 also increased production of IFN-gamma but at lower levels as compared to TNF-alpha With respect to key cell surface receptors, MGN-3 increases the expression of CD69, an early activation antigen at 16 hours after treatment. Furthermore, the interleukin-2 receptor CD25 and the adhesion molecule ICAM-1 (CD54) were upregulated after treatment with MGN-3. Treating highly purified NK cells with MGN-3 also resulted in increased levels of TNF-alpha and IFN-gamma secretion in conjunction with augmentation of NK cell cytotoxic function. Furthermore, addition of MGN-3 to interleukin-2-activated NK cells resulted in a synergistic induction of TNF-alpha and IFN-gamma secretion. Overall, our data suggest that MGN-3, a novel biological response modifier, can be used as a safe alternative or as an adjuvant to the existing immunotherapeutic modalities.

  17. Dendritic Cell Activation and Cytokine Production Induced by Group B Neisseria meningitidis: Interleukin-12 Production Depends on Lipopolysaccharide Expression in Intact Bacteria

    PubMed Central

    Dixon, Garth L. J.; Newton, Phillippa J.; Chain, Benjamin M.; Katz, David; Andersen, Svein Rune; Wong, Simon; van der Ley, Peter; Klein, Nigel; Callard, Robin E.

    2001-01-01

    Interactions between dendritic cells (DCs) and microbial pathogens are fundamental to the generation of innate and adaptive immune responses. Upon stimulation with bacteria or bacterial components such as lipopolysaccharide (LPS), immature DCs undergo a maturation process that involves expression of costimulatory molecules, HLA molecules, and cytokines and chemokines, thus providing critical signals for lymphocyte development and differentiation. In this study, we investigated the response of in vitro-generated human DCs to a serogroup B strain of Neisseria meningitidis compared to an isogenic mutant lpxA strain totally deficient in LPS and purified LPS from the same strain. We show that the parent strain, lpxA mutant, and meningococcal LPS all induce DC maturation as measured by increased surface expression of costimulatory molecules and HLA class I and II molecules. Both the parent and lpxA strains induced production of tumor necrosis factor alpha (TNF-α), interleukin-1α (IL-1α), and IL-6 in DCs, although the parent was the more potent stimulus. In contrast, high-level IL-12 production was only seen with the parent strain. Compared to intact bacteria, purified LPS was a very poor inducer of IL-1α, IL-6, and TNF-α production and induced no detectable IL-12. Addition of exogenous LPS to the lpxA strain only partially restored cytokine production and did not restore IL-12 production. These data show that non-LPS components of N. meningitidis induce DC maturation, but that LPS in the context of the intact bacterium is required for high-level cytokine production, especially that of IL-12. These findings may be useful in assessing components of N. meningitidis as potential vaccine candidates. PMID:11401973

  18. Dendritic cell activation and cytokine production induced by group B Neisseria meningitidis: interleukin-12 production depends on lipopolysaccharide expression in intact bacteria.

    PubMed

    Dixon, G L; Newton, P J; Chain, B M; Katz, D; Andersen, S R; Wong, S; van der Ley, P; Klein, N; Callard, R E

    2001-07-01

    Interactions between dendritic cells (DCs) and microbial pathogens are fundamental to the generation of innate and adaptive immune responses. Upon stimulation with bacteria or bacterial components such as lipopolysaccharide (LPS), immature DCs undergo a maturation process that involves expression of costimulatory molecules, HLA molecules, and cytokines and chemokines, thus providing critical signals for lymphocyte development and differentiation. In this study, we investigated the response of in vitro-generated human DCs to a serogroup B strain of Neisseria meningitidis compared to an isogenic mutant lpxA strain totally deficient in LPS and purified LPS from the same strain. We show that the parent strain, lpxA mutant, and meningococcal LPS all induce DC maturation as measured by increased surface expression of costimulatory molecules and HLA class I and II molecules. Both the parent and lpxA strains induced production of tumor necrosis factor alpha (TNF-alpha), interleukin-1alpha (IL-1alpha), and IL-6 in DCs, although the parent was the more potent stimulus. In contrast, high-level IL-12 production was only seen with the parent strain. Compared to intact bacteria, purified LPS was a very poor inducer of IL-1alpha, IL-6, and TNF-alpha production and induced no detectable IL-12. Addition of exogenous LPS to the lpxA strain only partially restored cytokine production and did not restore IL-12 production. These data show that non-LPS components of N. meningitidis induce DC maturation, but that LPS in the context of the intact bacterium is required for high-level cytokine production, especially that of IL-12. These findings may be useful in assessing components of N. meningitidis as potential vaccine candidates.

  19. Lotus hairy roots expressing inducible arginine decarboxylase activity.

    PubMed

    Chiesa, María A; Ruiz, Oscar A; Sánchez, Diego H

    2004-05-01

    Biotechnological uses of plant cell-tissue culture usually rely on constitutive transgene expression. However, such expression of transgenes may not always be desirable. In those cases, the use of an inducible promoter could be an alternative approach. To test this hypothesis, we developed two binary vectors harboring a stress-inducible promoter from Arabidopsis thaliana, driving the beta-glucuronidase reporter gene and the oat arginine decarboxylase. Transgenic hairy roots of Lotus corniculatus were obtained with osmotic- and cold-inducible beta-glucuronidase and arginine decarboxylase activities. The increase in the activity of the latter was accompanied by a significant rise in total free polyamines level. Through an organogenesis process, we obtained L. corniculatus transgenic plants avoiding deleterious phenotypes frequently associated with the constitutive over-expression of arginine decarboxylation and putrescine accumulation.

  20. Short-Lived Effector CD8 T Cells Induced by Genetically Attenuated Malaria Parasite Vaccination Express CD11c

    PubMed Central

    Cooney, Laura A.; Gupta, Megha; Thomas, Sunil; Mikolajczak, Sebastian; Choi, Kimberly Y.; Gibson, Claire; Jang, Ihn K.; Danziger, Sam; Aitchison, John; Gardner, Malcolm J.; Kappe, Stefan H. I.

    2013-01-01

    Vaccination with a single dose of genetically attenuated malaria parasites can induce sterile protection against sporozoite challenge in the rodent Plasmodium yoelii model. Protection is dependent on CD8+ T cells, involves perforin and gamma interferon (IFN-γ), and is correlated with the expansion of effector memory CD8+ T cells in the liver. Here, we have further characterized vaccine-induced changes in the CD8+ T cell phenotype and demonstrated significant upregulation of CD11c on CD3+ CD8b+ T cells in the liver, spleen, and peripheral blood. CD11c+ CD8+ T cells are predominantly CD11ahi CD44hi CD62L−, indicative of antigen-experienced effector cells. Following in vitro restimulation with malaria-infected hepatocytes, CD11c+ CD8+ T cells expressed inflammatory cytokines and cytotoxicity markers, including IFN-γ, tumor necrosis factor alpha (TNF-α), interleukin-2 (IL-2), perforin, and CD107a. CD11c− CD8+ T cells, on the other hand, expressed negligible amounts of all inflammatory cytokines and cytotoxicity markers tested, indicating that CD11c marks multifunctional effector CD8+ T cells. Coculture of CD11c+, but not CD11c−, CD8+ T cells with sporozoite-infected primary hepatocytes significantly inhibited liver-stage parasite development. Tetramer staining for the immunodominant circumsporozoite protein (CSP)-specific CD8+ T cell epitope demonstrated that approximately two-thirds of CSP-specific cells expressed CD11c at the peak of the CD11c+ CD8+ T cell response, but CD11c expression was lost as the CD8+ T cells entered the memory phase. Further analyses showed that CD11c+ CD8+ T cells are primarily KLRG1+ CD127− terminal effectors, whereas all KLRG1− CD127+ memory precursor effector cells are CD11c− CD8+ T cells. Together, these results suggest that CD11c marks a subset of highly inflammatory, short-lived, antigen-specific effector cells, which may play an important role in eliminating infected hepatocytes. PMID:23980113

  1. Flavonoids induce the expression of acetylcholinesterase in cultured osteoblasts.

    PubMed

    Xu, Miranda L; Bi, Cathy W C; Kong, Ava Y Y; Dong, Tina T X; Wong, Yung H; Tsim, Karl W K

    2016-11-25

    Flavonoids, a group of natural compounds mainly derived from plants, are known to possess osteogenic effects in bone cells. Here, we aimed to test if flavonoid could induce a cholinergic enzyme, acetylcholinesterase (AChE), as well as bone differentiation. In cultured rat osteoblasts, twenty flavonoids, deriving from Chinese herbs and having known induction of alkaline phosphatase (ALP(1)) expression, were tested for its induction activity on AChE expression. Eleven flavonoids showed the induction, and five of them had robust activation of AChE expression, including baicalin, calycosin, genistin, hyperin and pratensein: the induction of AChE included the levels of mRNA, protein and enzymatic activity. Moreover, the flavonoid-induced AChE expression in cultured osteoblast was in proline-rich membrane anchor (PRiMA)-linked tetrameric globular form (G4) only. In parallel, the expression of PRiMA was also induced by the application of flavonoids. The flavonoid-induced AChE in the cultures was not affected by estrogen receptor blocker, ICI 182,780. Taken together, the induction of PRiMA-linked AChE in osteoblast should be independent to classical estrogen signaling pathway.

  2. Tumor necrosis factor-alpha release during systemic reaction in cold urticaria.

    PubMed

    Tillie-Leblond, I; Gosset, P; Janin, A; Dalenne, R; Joseph, M; Wallaert, B; Tonnel, A B

    1994-02-01

    Primary cold urticaria (PCU) characterized by the association of urticaria, angioedema, and sometimes a shock-like reaction after cold exposure, is usually considered to be linked with histamine and prostaglandin D2 release by mast cells. To determine the involvement of cytokines, we studied the release of tumor necrosis factor-alpha (TNF-alpha) in the blood of the efferent vein after immersion of the hand in chilled water. Five patients with PCU were compared with a control population (three patients with nonphysical urticaria and three healthy subjects). Among patients with PCU who underwent the cold immersion test, two exhibited a shock-like reaction with a large urticarial plaque (patients 1 and 2), one had only a mild cutaneous reaction, and two had no reaction. Patient 1 was reevaluated after 6 months of treatment with H1 and H2 antihistamines: he did not respond to this challenge. All controls were strictly negative. Histamine was released within the first minute after the challenge in the three patients with PCU, but at a higher level for the two patients who had a systemic reaction. TNF-alpha was undetectable in the blood of the patient with only a mild cutaneous reaction, whereas TNF-alpha release was observed for the two patients with a systemic reaction, 2 and 6 minutes after the end of the cold immersion test. The two other patients and the control subjects released neither histamine nor TNF-alpha. In parallel, pathologic and immunohistochemical (with a rabbit anti-TNF-alpha antibody) studies were performed on skin biopsy specimens collected 10 minutes after ice-cube test.(ABSTRACT TRUNCATED AT 250 WORDS)

  3. Tumour necrosis factor-alpha (TNF-alpha) in patients who have asbestosis and develop cancer.

    PubMed Central

    Partanen, R; Koskinen, H; Hemminki, K

    1995-01-01

    OBJECTIVES--Concentrations of tumour necrosis factor-alpha (TNF-alpha) were assayed by radioimmunoassay in serum samples collected between 1981 and 1987 from 111 patients with asbestosis who were at a high risk of cancer. Follow up of these patients until 1993 showed that 38 had developed cancer (27 lung, three mesotheliomas, and eight diverse malignancies). RESULTS--The mean serum concentrations of TNF-alpha given in fmol/100 microliters serum in all the cases with cancer (14.1) and the cases with lung cancer (13.6) were significantly higher (P < 0.05) than the mean concentrations in the exposed controls (10.5). A positive increase was considered to be any value that was > 2 SDs above the mean of the exposed controls. 22% (six of 27) of the cases with lung cancer were positive compared with 4% (three of 73) of the exposed controls, a significant difference (P < 0.001). The serum concentrations of TNF-alpha correlated moderately with cancer (r = 0.3), lung cancer (r = 0.3), and Neu oncoproteins and epidermal growth factor receptor (EGFR) (r = 0.3, 0.5 respectively). Also, there was a significant correlation between development of cancer and severity or progression of asbestosis. There was no correlation between the concentrations of TNF-alpha and severity or progression of asbestosis. CONCLUSIONS--These results showed high concentrations of TNF-alpha in the patients who had cancer. TNF-alpha may offer an auxiliary method in early diagnosis of cancers related to asbestosis. PMID:7795753

  4. Polymer-functionalized silica nanosphere labels for ultrasensitive detection of tumor necrosis factor-alpha.

    PubMed

    Yuan, Liang; Hua, Xin; Wu, Yafeng; Pan, Xiaohu; Liu, Songqin

    2011-09-01

    A signal amplification strategy for sensitive detection of tumor necrosis factor-alpha (TNF-α) using quantum dots (QDs)-polymer-functionalized silica nanosphere as the label was proposed. In this approach, silica nanospheres with good monodispersity and uniform structure were employed as carriers for surface-initiated atom transfer radical polymerization of glycidyl methacrylate, which is readily available functional monomer that possessing easily transformable epoxy groups for subsequent CdTe QDs binding through ring-open reaction. Then, human anti rabbit TNF-α antibody (anti-TNF-α, Ab2, served as a model protein) was bonded to CdTe QDs-modified silica nanospheres coated with polymer to obtain QDs-polymer-functionalized silica nanosphere labels (Si/PGMA/QD/Ab2). The Si/PGMA/QD/Ab2 labels were attached onto a gold electrode surface through a subsequent "sandwich" immunoreaction. This reaction was confirmed by scanning electron microscopy (SEM) and fluorescence microscopic images. Enhanced sensitivity could be achieved by an increase of CdTe QD loading per immunoassay event, because of a large number of surface functional epoxy groups offered by the PGMA. As a result, the electrochemiluminescence (ECL) and square-wave voltammetry (SWV) measurements showed 10.0- and 5.5-fold increases in detection signals, respectively, in comparison with the unamplified method. The detection limits of 7.0 pg mL(-1) and 3.0 pg mL(-1) for TNF-α antibodies by ECL and SWV measurements, respectively, were achieved. The proposed strategy successfully demonstrated a simple, reproducible, specific, and potent method that can be expanded to detect other proteins and DNA.

  5. Orofacial clefts, parental cigarette smoking, and transforming growth factor-alpha gene variants

    SciTech Connect

    Shaw, G.M.; Wasserman, C.R.; O`Malley, C.D.

    1996-03-01

    Results of studies determine whether women who smoke during early pregnancy are at increased risk of delivering infants with orofacial clefts have been mixed, and recently a gene-environment interaction between maternal smoking, transforming growth factor-alpha (TGFa), and clefting has been reported. Using a large population-based case-control study, we investigated whether parental periconceptional cigarette smoking was associated with an increased risk for having offspring with orofacial clefts. We also investigated the influence of genetic variation of the TGFa locus on the relation between smoking and clefting. Parental smoking information was obtained from telephone interviews with mothers of 731 (84.7% of eligible) orofacial cleft case infants and with mothers of 734 (78.2%) nonmalformed control infants. DNA was obtained from newborn screening blood spots and genotyped for the allelic variants of TGFa. We found that risks associated with maternal smoking were most elevated for isolated cleft lip with or without cleft palate, (odds ratio 2.1 [95% confidence interval 1.3-3.6]) and for isolated cleft palate (odds ratio 2.2 [1.1-4.5]) when mothers smoked {ge} 20 cigarrettes/d. These risks for white infants ranged from 3-fold to 11-fold across phenotypic groups. Paternal smoking was not associated with clefting among the offspring of nonsmoking mothers, and passive smoke exposures were associated with at most slightly increased risks. This study offers evidence that the risk for orofacial clefting in infants may be influenced by maternal smoke exposures alone as well as in combination (gene-environment interaction) with the presence of the uncommon TGFa allele. 56 refs., 5 tabs.

  6. A novel cold-inducible expression system for Bacillus subtilis.

    PubMed

    Thuy Le, Ai Thi; Schumann, Wolfgang

    2007-06-01

    Production of recombinant proteins at low temperatures is one strategy to prevent formation of protein aggregates and the use of an expensive inducer such as IPTG. We report on the construction of two expression vectors both containing the cold-inducible des promoter of Bacillus subtilis, where one allows intra- and the other extracellular synthesis of recombinant proteins. Production of recombinant proteins started within the first 30min after temperature downshock to 25 degrees C and continued for about 5h.

  7. Thiostrepton-induced gene expression in Streptomyces lividans.

    PubMed Central

    Murakami, T; Holt, T G; Thompson, C J

    1989-01-01

    Thiostrepton induced the expression of four proteins (17, 19, 30, and 56 kilodaltons) of unknown function in Streptomyces lividans. The chromosomal gene which encoded the 19-kilodalton protein (tipA) was cloned and sequenced. Transcription of the tipA promoter was induced at least 200-fold by thiostrepton. The tipA 200-fold by thiostrepton. The tipA transcriptional start site (located by S1 mapping and primer extension experiments) was preceded by a 45-base-pair imperfect inverted-repeat sequence which included the -10 and -35 regions of the promoter. Under noninducing conditions in vivo, this might form a cruciform structure which is not recognized by RNA polymerase. A 143-base-pair fragment including this region was cloned into a promoter probe vector, pIJ486. In this plasmid, pAK114, the thiostrepton-inducible tipA promoter controlled the expression of a kanamycin resistance gene encoding an aminoglycoside phosphotransferase. As little as 1 ng of thiostrepton spotted on a lawn of S. lividans(pAK114) induced kanamycin-resistant growth. Other thiostreptonlike antibiotics also induced tipA, but structurally unrelated antibiotics which inhibit translation had no effect. In S. lividans, the promoter could be induced by thiostrepton during either growth or stationary phase. The tipA promoter should be a valuable tool for expression studies in streptomycetes. Images PMID:2537819

  8. Interleukin 1 induces expression of the human immunodeficiency virus alone and in synergy with interleukin 6 in chronically infected U1 cells: inhibition of inductive effects by the interleukin 1 receptor antagonist.

    PubMed Central

    Poli, G; Kinter, A L; Fauci, A S

    1994-01-01

    In the present study we have observed that interleukin (IL) 1 alpha or IL-1 beta directly induced expression of human immunodeficiency virus (HIV) in the latently infected human promonocytic cell line U1. In addition, IL-1 synergized with IL-6, but not with tumor necrosis factor, in the upregulation of virus expression in U1 cells as measured by accumulation of steady-state mRNAs and production of reverse transcriptase activity. The HIV inductive effect of IL-1 was blocked by transforming growth factor beta, anti-IL-1 antibodies, or monoclonal antibodies directed to the type 1, but not to the type 2, cell surface receptor for IL-1; the latter actually caused enhancement of the IL-1-mediated effect. Unlike tumor necrosis factor alpha, IL-1 either alone or in combination with IL-6 did not induce activation of the transcription activating factor NF-kappa B above the constitutive levels of unstimulated U1 cells. Finally, the IL-1 receptor antagonist effectively blocked IL-1-mediated direct and synergistic inductive effects on virus production. Thus, IL-1 may be an important mediator of HIV expression, and blocking of IL-1 expression and/or its effects may have a potential therapeutic role in the inhibition of HIV expression in infected individuals. Images Fig. 2 Fig. 3 PMID:7506410

  9. Running exercise increases tumor necrosis factor-alpha secreting from mesenteric fat in insulin-resistant rats.

    PubMed

    Nara, M; Kanda, T; Tsukui, S; Inukai, T; Shimomura, Y; Inoue, S; Kobayashi, I

    1999-01-01

    Tumor necrosis factor-alpha (TNF-alpha) is an important mediator of insulin resistance in obese subjects, through its overexpression in fat tissue. However, how exercise can modify the expression of TNF-alpha is controversial. We examined TNF-alpha in adipose tissue using an animal model of insulin resistance that was produced by feeding rats a diet high in sucrose. The rats were allocated to one of three groups: those receiving a starch-based diet (control group): those fed a high-sucrose diet (sucrose-fed group): and those fed a high-sucrose diet and given wheel exercise (exercised group). The animals were allowed to eat and drink ad lib for 4 or 12 weeks (4 wk: control n=7, sucrose-fed n=7, exercised n=10; 12 wk: control n=5, sucrose-fed n=5, exercised n=9). The voluntary wheel exercise was initiated with the feeding of the high-sucrose diet. The rats in the exercise groups ran 15 +/- 3 km/week. We showed that 12-week voluntary running exercise significantly (P<0.05) increased both TNF-alpha protein (5-fold) and mRNA (1.4 fold) in the mesenteric fat of insulin-resistant rats compared to non-exercised sucrose-fed mice. Accordingly, in exercised group, plasma glucose (124 +/- 9 mEq/L vs 141 +/- 11 mEq/L). and free fatty acid (0.98 +/- 0.07 mEq/L vs 1.4 +/- 0.05 mEq/L) concentrating in portal vein blood were reduced compared to sucrose-fed group. The amounts of fatty tissue both in mesenteric and subcutaneous tissues were significantly (P<0.05) decreased through running exercise. We consider that up-regulation of TNF-alpha in mesenteric fat may be a compensatory mechanism for the reduction of fatty acid in adipose tissues and this change could control metabolic homeostasis during exercise to modulate a hyperinsulinemic state.

  10. TLR4 mediates LPS-induced VEGF expression in odontoblasts.

    PubMed

    Botero, Tatiana M; Shelburne, Charles E; Holland, G Rex; Hanks, Carl T; Nör, Jacques E

    2006-10-01

    Lipopolysaccharide (LPS) from gram-negative bacteria cell walls such as Prevotella intermedia and Escherichia coli induce vascular endothelial growth factor (VEGF) expression in odontoblasts, but not in undifferentiated dental pulp cells. CD14 and TLR4 are responsible for LPS signaling in macrophages, but their expression levels and function in dental pulp cells are unknown. We showed here that murine odontoblast-like cells (MDPC-23) express CD14 and TLR4 by immunohistochemistry and flow cytometry. In contrast, undifferentiated dental pulp cells (OD-21) presented low or no expression of these two receptors. MDPC-23 cells showed CD14 and TLR4 up-regulation upon exposure to LPS, as determined by real time PCR. Dominant negative murine TLR4 (DN-mTLR4) transfected MDPC-23 cells did not show upregulated VEGF expression in response to LPS stimulation. These results demonstrate that odontoblast-like cells express CD14 and TLR4, and that LPS-induced VEGF expression is mediated, at least in part, by TLR4 signaling.

  11. Phase I trial of ISIS 104838, a 2'-methoxyethyl modified antisense oligonucleotide targeting tumor necrosis factor-alpha.

    PubMed

    Sewell, K Lea; Geary, Richard S; Baker, Brenda F; Glover, Josephine M; Mant, Timothy G K; Yu, Rosie Z; Tami, Joseph A; Dorr, F Andrew

    2002-12-01

    ISIS 104838 is a 20-mer phosphorothioate antisense oligonucleotide (ASO) that binds tumor necrosis factor-alpha (TNF-alpha) mRNA. It carries a 2'-methoxyethyl modification on the five 3' and 5' nucleotide sugars, with 10 central unmodified deoxynucleotides. ISIS 104838 was identified from a 264 ASO screen in phorbol myristate acetate-activated keratinocytes, and the dose response was assessed in lipopolysaccharide (LPS)-activated monocytes. Healthy males received multiple intravenous (i.v.) ISIS 104838 infusions in a placebo-controlled dose escalation trial (0.1-6 mg/kg). Additional volunteers received single or multiple subcutaneous (s.c.) injections. ISIS 104838 suppressed TNF-alpha protein by 85% in stimulated keratinocytes. The IC50 for TNF-alpha mRNA inhibition in stimulated monocytes was <1 microM. For i.v., C(max) occurred at the end of infusion. The effective plasma half-life was 15 to 45 min at 0.1 to 0.5 mg/kg and 1 to 1.8 h for higher doses. The apparent terminal plasma elimination half-life approximated 25 days. Obese subjects had higher plasma levels following equivalent mg/kg doses. For s.c. injections, C(max) occurred at 2 to 4 h and was lower than with equivalent i.v. dosing. Plasma bioavailability compared with i.v. was 82% following a 200 mg/ml s.c. injection. Transient activated partial thromboplastin time prolongation occurred after i.v. infusions and minimally after s.c. injections. Two subjects experienced rash, one a reversible platelet decrease, and mild injection site tenderness was noted. TNF-alpha production by peripheral blood leukocytes, induced ex vivo by LPS, was decreased by ISIS 104838 (p < 0.01). ISIS 104838, a second-generation antisense oligonucleotide, was generally well tolerated intravenously and subcutaneously. The pharmacokinetics support an infrequent dosing interval. Inhibition of TNF-alpha production ex vivo was demonstrated.

  12. Zinc pyrithione induces apoptosis and increases expression of Bim.

    PubMed

    Mann, J J; Fraker, P J

    2005-03-01

    We demonstrate herein that zinc pyrithione can induce apoptosis at nanomolar concentrations. Zinc pyrithione was a potent inducer of cell death causing greater than 40-60% apoptosis among murine thymocytes, murine splenic lymphocytes and human Ramos B and human Jurkat T cells. Conversely, the addition of a zinc chelator protected thymocytes against zinc pyrithione induced apoptosis indicating these responses were specific for zinc. Zinc-induced apoptosis was dependent on transcription and translation which suggested possible regulation by a proapoptotic protein. Indeed, zinc induced a 1.9 and 3.4 fold increase respectively in expression of the BimEL and BimL isoforms and also stimulated production of the most potent isoform, BimS. This increase in Bim isoform expression was dependent on transcription being blocked by treatment with actinomycin D. Overexpression of Bcl-2 or Bcl-xL provided substantial protection of Ramos B and Jurkat T cells against zinc-induced apoptosis. Zinc also activated the caspase cascade demonstrated by cleavage of caspase 9. Addition of specific inhibitors for caspase 9 and caspase 3 also blocked zinc-induced apoptosis. The data herein adds to the growing evidence that free or unbound zinc could be harmful to cells of the immune system.

  13. Effect of Selenium Against Lead-Induced Damage on the Gene Expression of Heat Shock Proteins and Inflammatory Cytokines in Peripheral Blood Lymphocytes of Chickens.

    PubMed

    Sun, G X; Chen, Y; Liu, C P; Li, S; Fu, J

    2016-08-01

    The possible beneficial role of selenium (Se) in heat shock proteins (HSPs) and inflammation damage induced by lead (Pb) in chickens is unclear. Therefore, the aim of this study was to investigate the effect of Se against Pb on the messenger RNA (mRNA) expression levels of HSPs (HSP 27, 40, 60, 70, and 90); heme oxygenase-1 (HO-1); and the inflammatory cytokines nuclear factor kappa B (NF-κB), tumor necrosis factor alpha (TNF-α), cyclooxygenase-2 (COX-2), and inducible nitric oxide synthase (iNOS) in the peripheral blood lymphocytes of chickens. A total of 360 1-day-old broiler chickens were randomly allocated into four groups (n = 90/group). The control group was fed a basic diet containing 0.2 mg/kg Se and 0.5 mg/kg Pb; the Se supplementation group (+Se group) was fed a Se-adequate (sodium selenite) diet containing 1 mg/kg Se and 0.5 mg/kg Pb; the Pb-supplemented group (+Pb group) was fed a Pb acetate diet containing 0.2 mg/kg Se and 350 mg/kg Pb; and the Se and Pb compound group (Se + Pb group) was fed a diet containing 1 mg/kg Se and 350 mg/kg Pb. The blood was collected and examined for the mRNA levels of HSP and inflammatory cytokine genes at 30 and 60 days old. The results showed that Pb poisoning induced the mRNA expression of HSPs and inflammatory cytokines in the peripheral blood lymphocytes of chickens. In addition, Se alleviated the Pb-induced increase in HSP and inflammatory cytokine mRNA levels in chicken peripheral blood lymphocytes. In conclusion, Se can antagonize the toxic effects of Pb on chickens and protect the chickens' peripheral blood lymphocytes in normal physiological function.

  14. Three polymorphisms of tumor necrosis factor-alpha and hepatitis B virus related hepatocellular carcinoma

    PubMed Central

    Xiao, Qi; Fu, BiQi; Chen, Ping; Liu, Zhong Zhong; Wang, Wei; Ye, QiFa

    2016-01-01

    Abstract Background: To assess the association between tumor necrosis factor-alpha (TNF-α) G308A, G238A and C863T polymorphisms and hepatitis B virus related hepatocellular carcinoma (HBV-HCC) susceptibility. Methods: We interrogated the databases of Pubmed, Sciencedirect and Viley online library up to March 8, 2016. Odds ratios (ORs) and corresponding 95% confidence intervals (95%CIs) were calculated in a fixed-effects model or a random-effects model when appropriate. Results: In total, 12 case–control studies which containing 1580 HBV-HCC cases, 2033 HBV carrier controls, 395 HBV spontaneously recovered (SR) controls and 1116 healthy controls were included. Compared with GG genotype, the genotypes GA/AA of G308A were associated with a significantly increased HBV-HCC risk when the controls were all healthy individuals (AA vs. GG, OR 2.483, 95%CI 1.243 to 4.959; GA vs. GG, OR 1.383, 95%CI 1.028 to 1.860; GA/AA vs. GG, OR 1.381, 95%CI 1.048 to 1.820). Meanwhile, only the AA vs. GG model of G238A and HBV-HCC showed a statistic significance when the controls were healthy individuals (OR 4.776, 95%CI 1.280 to 17.819). CT genotype of TNF-α C863T could increase HBV-HCC risk whenever the controls were healthy individuals, HBV carriers or HBV recovers. Conclusion: This meta-analysis shows that AA genotype in TNF-α G308A and TNF-α G238A and CT genotype in TNF-α C863T may increase HBV-HCC risk. Therefore, HBV infection seemed to be a more important factor for tumorigenesis of HCC than genetic predisposition in G308A of TNF-α, and interaction between TNF-α C863T polymorphisms and HBV infection might be associated with increased HCC risk. PMID:27977601

  15. Tumor necrosis factor-alpha/interleukin-10 balance in normal and cystic fibrosis children.

    PubMed Central

    Shmarina, G V; Pukhalsky, A L; Kokarovtseva, S N; Pukhalskaya, D A; Shabalova, L A; Kapranov, N I; Kashirskaja, N J

    2001-01-01

    BACKGROUND: The balance between tumor necrosis factor-alpha (TNF-alpha) and interleukin-10 (IL-10) is important for immune homeostasis maintenance. Exuberant production of TNF-alpha contributes to overwhelming inflammatory response and tissue damage. But, commonly, increase in TNF-alpha is counterbalanced by simultaneous synthesis of an anti-inflammatory cytokine IL-10, which suppresses production of many activating and regulatory mediators. AIMS: In the present study, the relationships between TNF-alpha and IL-10 in the plasma of healthy school-children and cystic fibrosis (CF) patients have been investigated. METHODS: Blood samples were obtained from 12 CF patients with chronic pulmonary disease and 18 healthy schoolchildren vaccinated with live attenuated rubella vaccine. IL-10 and TNF-alpha were determined in the plasma samples using commercially available enzyme-linked immunosorbent assay kits. RESULTS: Before vaccination, most healthy children (13 of 18) demonstrated superiority of pro-inflammatory TNF-alpha over anti-inflammatory IL-10 (TNF-alpha/IL-10 > 1). In these subjects, a significant positive linear association between the cytokine values has been found. Vaccine challenge resulted in a marked reduction of TNF-alpha/IL-10 ratios. In addition, a disappearance of correlation between the cytokine values was observed. Such disturbance was related to exuberant elevation of the IL-10 levels after inoculation. On the contrary, in CF individuals, plasma cytokine values remained in strong linear association independently of TNF-alpha or IL-10 predominance. No spikes in the plasma levels of IL-10 in CF patients during a 6-month observation period have been revealed. CONCLUSIONS: There were no fundamental differences between CF and healthy children in the regulation of TNF-alpha and IL-10 secretion. Thus, immune quiescence seemed to be associated with the predominance of TNF-alpha, whereas immune disturbance was characterized by IL-10 superiority. The only

  16. Dexmedetomidine Attenuates Lipopolysaccharide Induced MCP-1 Expression in Primary Astrocyte

    PubMed Central

    Liu, Huan; Faez Abdelgawad, Amro

    2017-01-01

    Background. Neuroinflammation which presents as a possible mechanism of delirium is associated with MCP-1, an important proinflammatory factor which is expressed on astrocytes. It is known that dexmedetomidine (DEX) possesses potent anti-inflammatory properties. This study aimed to investigate the potential effects of DEX on the production of MCP-1 in lipopolysaccharide-stimulated astrocytes. Materials and Methods. Astrocytes were treated with LPS (10 ng/ml, 50 ng/ml, 100 ng/ml, and 1000 ng/ml), DEX (500 ng/mL), LPS (100 ng/ml), and DEX (10, 100, and 500 ng/mL) for a duration of three hours; expression levels of MCP-1 were measured by real-time PCR. The double immunofluorescence staining protocol was utilized to determine the expression of α2-adrenoceptors (α2AR) and glial fibrillary acidic protein (GFAP) on astrocytes. Results. Expressions of MCP-1 mRNA in astrocytes were induced dose-dependently by LPS. Administration of DEX significantly inhibited the expression of MCP-1 mRNA (P < 0.001). Double immunofluorescence assay showed that α2AR colocalize with GFAP, which indicates the expression of α2-adrenoceptors in astrocytes. Conclusions. DEX is a potent suppressor of MCP-1 in astrocytes induced with lipopolysaccharide through α2A-adrenergic receptors, which potentially explains its beneficial effects in the treatment of delirium by attenuating neuroinflammation. PMID:28286770

  17. Dexmedetomidine Attenuates Lipopolysaccharide Induced MCP-1 Expression in Primary Astrocyte.

    PubMed

    Liu, Huan; Davis, Jacques R J; Wu, Zhi-Lin; Faez Abdelgawad, Amro

    2017-01-01

    Background. Neuroinflammation which presents as a possible mechanism of delirium is associated with MCP-1, an important proinflammatory factor which is expressed on astrocytes. It is known that dexmedetomidine (DEX) possesses potent anti-inflammatory properties. This study aimed to investigate the potential effects of DEX on the production of MCP-1 in lipopolysaccharide-stimulated astrocytes. Materials and Methods. Astrocytes were treated with LPS (10 ng/ml, 50 ng/ml, 100 ng/ml, and 1000 ng/ml), DEX (500 ng/mL), LPS (100 ng/ml), and DEX (10, 100, and 500 ng/mL) for a duration of three hours; expression levels of MCP-1 were measured by real-time PCR. The double immunofluorescence staining protocol was utilized to determine the expression of α2-adrenoceptors (α2AR) and glial fibrillary acidic protein (GFAP) on astrocytes. Results. Expressions of MCP-1 mRNA in astrocytes were induced dose-dependently by LPS. Administration of DEX significantly inhibited the expression of MCP-1 mRNA (P < 0.001). Double immunofluorescence assay showed that α2AR colocalize with GFAP, which indicates the expression of α2-adrenoceptors in astrocytes. Conclusions. DEX is a potent suppressor of MCP-1 in astrocytes induced with lipopolysaccharide through α2A-adrenergic receptors, which potentially explains its beneficial effects in the treatment of delirium by attenuating neuroinflammation.

  18. The stem of sinomenium acutum inhibits mast cell-mediated anaphylactic reactions and tumor necrosis factor-alpha production from rat peritoneal mast cells.

    PubMed

    Kim, H M; Moon, P D; Chae, H J; Kim, H R; Chung, J G; Kim, J J; Lee, E J

    2000-05-01

    The aqueous extract of Sinomenium acutum stem (SSAE) (0.1-1000 mg/kg) dose-dependently inhibited systemic anaphylactic reaction induced by compound 48/80 in mice. In particular, SSAE reduced compound 48/80-induced anaphylactic reaction with 50% at the dose of 1000 mg/kg. SSAE (100-1000 mg/kg) also significantly inhibited local anaphylactic reaction activated by anti-dinitrophenyl (DNP) IgE. When mice were pretreated with SSAE at a concentration ranging from 0.1 to 1000 mg/kg, the plasma histamine levels were reduced in a dose-dependent manner. SSAE (1-1000 microg/ml) dose-dependently inhibited histamine release from the rat peritoneal mast cells (RPMCs) activated by compound 48/80 or anti-DNP IgE. In addition, SSAE (0.1 microg/ml) had a significant inhibitory effect on anti-DNP IgE-induced tumor necrosis factor-alpha (TNF-alpha) production. These results indicate that SSAE inhibits mast cell-mediated anaphylactic reactions and TNF-alpha production from mast cells.

  19. Recombinant human tumor necrosis factor alpha does not potentiate cell killing after photodynamic therapy with a silicon phthalocyanine in A431 human epidermoid carcinoma cells.

    PubMed

    Azizuddin, K; Kalka, K; Chiu, S M; Ahmad, N; Mukhtar, H; Separovic, D

    2001-02-01

    Photodynamic therapy (PDT) is a novel cancer treatment utilizing a photosensitizer, visible light and oxygen. PDT with the silicon phthalocyanine Pc 4, a new photosensitizer, is highly effective in cancer cell destruction and tumor ablation. The mechanisms underlying cancer cell killing by PDT are not fully understood. Tumor necrosis factor alpha (TNF) is a multifunctional cytokine that has been implicated in photocytotoxicity. We asked whether recombinant human TNF (rhTNF) affects Pc 4-PDT cytotoxicity in A431 human epidermoid carcinoma cells. Co-treatment of A431 cells with various doses of Pc 4-PDT and a sub-lethal rhTNF dose led to a sub-additive reduction in cell survival. In addition, in the presence of Pc 4-PDT or rhTNF, caspase-3 activity and apoptosis were induced. The combined treatment, however, did not potentiate either caspase-3 activity or apoptosis. Similar to previous findings we observed that Pc 4-PDT initiated a time-dependent extracellular TNF accumulation. The data suggest that: a) PDT and rhTNF induce cancer cell killing through different mechanisms; and b) Pc 4-PDT-induced TNF production is a stress response that may not directly affect photocytotoxicity.

  20. Nickel-induced heritable alterations in retroviral transforming gene expression.

    PubMed Central

    Biggart, N W; Gallick, G E; Murphy, E C

    1987-01-01

    Determination of the mutagenic effects of carcinogenic nickel compounds has been difficult because, like many metals, nickel is poorly or nonmutagenic in procaryotic mutagenicity assays. We attempted to characterize nickel-induced genetic lesions by assessing the effect of nickel chloride on the conditionally defective expression of the v-mos transforming gene in normal rat kidney cells infected with the Murine sarcoma virus mutant ts110 (MuSVts110) retrovirus. MuSVts110 contains an out-of-frame gag gene-mos gene junction that prevents the expression of the v-mos gene at the nonpermissive temperature (39 degrees C). In MuSVts110-infected cells (6m2 cells) grown at 33 degrees C, however, this defect can be suppressed by a splicing event that restores the mos reading frame, allowing the expression of a gag-mos fusion protein which induces the transformed phenotype. The capacity to splice the viral transcript at 33 degrees C, but not at 39 degrees C, is an intrinsic property of the viral RNA. This property allowed us to target the MuSVts110 genome using a positive selection scheme whereby nickel was used to induce genetic changes which resulted in expression of the transformed phenotype at 39 degrees C. We treated 6m2 cells with NiCl2 and isolated foci consisting of cells which had reverted to the transformed phenotype at 39 degrees C. We found that brief nickel treatment increased the reversion frequency of 6m2 cells grown at 39 degrees C sevenfold over the spontaneous reversion frequency. The nickel-induced revertants displayed the following heritable characteristics: They stably maintained the transformed phenotype at 39 degrees C; unlike the MuSVts110 RNA in 6m2 cells, the nickel-induced revertant viral RNA could be spliced efficiently at 39 degrees C; as a consequence of the enhanced accumulation of spliced viral RNA, the nickel-induced revertants produced substantial amounts of the transforming v-mos protein P85gag-mos at 39 degrees C; the nickel-induced

  1. Ectoine from halophilic microorganisms induces the expression of hsp70 and hsp70B' in human keratinocytes modulating the proinflammatory response.

    PubMed

    Buommino, Elisabetta; Schiraldi, Chiara; Baroni, Adone; Paoletti, Iole; Lamberti, Monica; De Rosa, Mario; Tufano, Maria Antonietta

    2005-01-01

    The heat shock proteins (Hsps) have an important role in the cytoprotection and repair of cells and tissues. One potential mechanism of protection is the ability of Hsp to inhibit genetic expression of proinflammatory cytokines, the transcription of which is dependent on nuclear factor-kappa B (NF-kappaB) activation. In this study, we evaluated the ability of ectoine, a novel natural biomolecule produced by halophilic microorganisms, to activate the hsp70 and hsp70B'. By reverse transcriptase-polymerase chain reaction and Western blot analysis, we demonstrated increased hsp70B' gene expression in human keratinocytes treated with ectoine and heat stressed. In contrast, in the absence of heat shock, ectoine was unable to induce hsp70B' but had the ability to induce another member of the Hsp family, the hsp70. The latter is not only elevated in response to stress but is also present at basal level in unstressed cells. In addition, ectoine had no effect on proinflammatory cytokines interleukin (IL)-1alpha, IL-6, IL-8, and tumor necrosis factor-alpha and on NF-kappaB and IkappaB-alpha pathway, whereas it downregulated the expression of cited proinflammatory cytokines, in lipopolysaccharides-treated keratinocytes. These results highlighted the ability of ectoine to protect cells from stress conditions and to prevent cell damage by maintaining an elevated level of the Hsp70. Overall, these data might suggest the use of this compatible solute in cosmetic and even pharmaceutical preparations aiming to activate a cytoprotective heat shock response in human cells.

  2. Salmonella enterica induces joint inflammation and expression of interleukin-17 in draining lymph nodes early after onset of enterocolitis in mice.

    PubMed

    Noto Llana, Mariángeles; Sarnacki, Sebastián Hernán; Vázquez, María Victoria; Gartner, Alejandra Sonia; Giacomodonato, Mónica Nancy; Cerquetti, María Cristina

    2012-06-01

    In developing countries, one-third of reactive arthritis (ReA) cases are associated with Salmonella enterocolitis; nevertheless, there is no animal model for studying this pathology. Here we induced a self-limiting Salmonella enterica serovar Enteritidis enterocolitis in mice to analyze the onset of ReA. BALB/c mice received orally 20 μg of streptomycin 24 h before intragastric inoculation of a low dose (3 × 10(3) to 4 × 10(3) CFU) of S. Enteritidis. In response to Salmonella infection, a 30-fold increase in the expression of interleukin-17 (IL-17), measured by quantitative PCR, was observed in mesenteric lymph nodes 5 days postinfection. At this time synovitis was already evident, and concomitantly, a significant increase in joint tumor necrosis factor alpha (TNF-α) was detected by enzyme-linked immunosorbent assay (ELISA). The early development of joint lesions was accompanied by an increased expression of IL-17 in inguinal and popliteal lymph nodes. Infection with 10(7) CFU of an isogenic ΔinvG mutant bearing a defective type III secretion system of Salmonella encoded in the pathogenicity island 1 apparatus (TTSS-1) induced enterocolitis histologically similar to that triggered by the wild-type strain. Interestingly, despite the higher infective dose used, the mutant did not trigger intestinal IL-17. Moreover, no synovitis was observed in mice suffering ΔinvG enterocolitis. Neutralization of IL-17 in mice infected with S. Enteritidis prevented both synovitis and the increment of TNF-α in the joints, suggesting that IL-17 participates in the generation of Salmonella-induced ReA through the induction of TNF-α in the joints.

  3. XPB Induces C1D Expression to Counteract UV-Induced Apoptosis

    PubMed Central

    Li, Guang; Liu, Juhong; Abu-Asab, Mones; Masabumi, Shibuya; Maru, Yoshiro

    2010-01-01

    Although C1D has been shown to be involved in DNA double-strand breaks repair, how C1D expression was induced and the mechanism(s) by which C1D facilitates DNA repair in mammalian cells remain poorly understood. We and others have previously shown that expression of XPB protein efficiently compensated the UV-irradiation sensitive phenotype of 27-1 cells which lacks functional XPB. To further explore XPB-regulated genes that could be involved in UV-induced DNA repair, Differential Display analysis of mRNA level from CHO-9, 27-1 and 27-1 complemented with wild-type XPB were performed and C1D gene was identified as one of the major genes whose expression was significantly up-regulated by restoring XPB function. We found that XPB is essential to induce C1D transcription after UV-irradiation. The increase of C1D expression effectively compensates the UV-induced proteolysis of C1D and thus maintains cellular C1D level to cope with DNA damage inflicted by UV-irradiation. We further showed that although insufficient to rescue 27-1 cells from UV-induced apoptosis by itself, C1D facilitates XPB DNA repair through direct interaction with XPB. Our findings provided direct evidence that C1D is associated with DNA repair complex and may promote repair of UV-induced DNA damage. PMID:20530579

  4. Microarray studies of psychostimulant-induced changes in gene expression.

    PubMed

    Yuferov, Vadim; Nielsen, David; Butelman, Eduardo; Kreek, Mary Jeanne

    2005-03-01

    Alterations in the expression of multiple genes in many brain regions are likely to contribute to psychostimulant-induced behaviours. Microarray technology provides a powerful tool for the simultaneous interrogation of gene expression levels of a large number of genes. Several recent experimental studies, reviewed here, demonstrate the power, limitations and progress of microarray technology in the field of psychostimulant addiction. These studies vary in the paradigms of cocaine or amphetamine administration, drug doses, route and also mode of administration, duration of treatment, animal species, brain regions studied and time of tissue collection after final drug administration. The studies also utilize different microarray platforms and statistical techniques for analysis of differentially expressed genes. These variables influence substantially the results of these studies. It is clear that current microarray techniques cannot detect small changes reliably in gene expression of genes with low expression levels, including functionally significant changes in components of major neurotransmission systems such as glutamate, dopamine, opioid and GABA receptors, especially those that may occur after chronic drug administration or drug withdrawal. However, the microarray studies reviewed here showed cocaine- or amphetamine-induced alterations in the expression of numerous genes involved in the modulation of neuronal growth, cytoskeletal structures, synaptogenesis, signal transduction, apoptosis and cell metabolism. Application of laser capture microdissection and single-cell cDNA amplification may greatly enhance microarray studies of gene expression profiling. The combination of rapidly evolving microarray technology with established methods of neuroscience, molecular biology and genetics, as well as appropriate behavioural models of drug reinforcement, may provide a productive approach for delineating the neurobiological underpinnings of drug responses that lead to

  5. [Expression of tobacco SKP1 gene induced by oligochitosan].

    PubMed

    Zhang, Fu-Yun; Feng, Bin; Du, Yu-Guang; Bai, Xue-Fang; Zhang, Yu-Kui

    2005-04-01

    Oligochitosan is an effective inductor for plant resistance. To understand the induced resistance mechanism, mRNA differential display was used to isolate genes from Nicotiana tabacum var. Samsun NN and four enhanced-expression gene fragments were obtained and were reamplified. The reamplified products of appropriate size were isolated and purified before they were subcloned into PMD18-T vector. The results of plasmids digestion by EcoRI and HindIII and analysis of reverse Northern blot indicated that the expression of the four genes was enhanced by oligochitosan induction. Sequence and homology analysis show that they share 82% identity in nucleotide sequence with Nicotiana benthamiana SKP1 gene. Because the SKP1 protein as the core component of SCF (ubiquitin ligase enzyme E3) is relevant to plant resistance to virus, so these results suggested that oligochitosan can induce plant resistance and its mechanism may be relevant to ubiquitination.

  6. Inducible expression of endomorphins in murine dendritic cells.

    PubMed

    Yang, Xiaohuai; Xia, Hui; Chen, Yong; Liu, Xiaofen; Zhou, Cheng; Gao, Qin; Li, Zhenghong

    2012-12-15

    Bone marrow precursor cells were extracted from C57BL/6J mice aged 7-8 weeks, and dendritic cells were purified using anti-CD11c (a specific marker for dendritic cells) antibody-coated magnetic beads. Immunofluorescence staining revealed that the expression levels of endomorphin-1 and endomorphin-2 were upregulated in dendritic cells activated by lipopolysaccharide. An enzyme immunoassay showed that lipopolysaccharide and other Toll-like receptor ligands promoted the secretion of endomorphin-1 and endomorphin-2 from activated dendritic cells. [(3)H]-thymidine incorporation demonstrated that endomorphin-1 and endomorphin-2 both inhibited the proliferation of T lymphocyte induced by activated dendritic cells. Furthermore, this immunosuppressive effect was blocked by CTOP, a specific antagonist of µ-opioid receptors. Our experimental findings indicate that activated dendritic cells can induce the expression and secretion of endomorphins, and that endomorphins suppress T lymphocyte proliferation through activation of µ-opioid receptors.

  7. Tyrosine hydroxylase, interleukin-1beta and tumor necrosis factor-alpha are overexpressed in peripheral blood mononuclear cells from schizophrenia patients as determined by semi-quantitative analysis.

    PubMed

    Liu, Liang; Jia, Fujun; Yuan, Guozhen; Chen, Zaohuo; Yao, Jianjun; Li, Hengfen; Fang, Chunxia

    2010-03-30

    The aim of this study is to profile the peripheral biomarkers (tyrosine hydroxylase, TH; interleukin-1beta, IL-1beta; and tumor necrosis factor-alpha, TNF-alpha) for schizophrenia and explore their relations with clinical symptoms. Thirty-nine patients with schizophrenia were evaluated using the Positive and Negative Syndrome Scale (PANSS), and 25 siblings and 30 normal healthy subjects were used as controls. The mRNA expression levels of TH, IL-1beta and TNF-alpha in peripheral blood mononuclear cells, as determined with semi-quantitative reverse transcription-polymerase chain reaction, were all significantly increased in both patients (3-fold) and siblings (2-fold) as compared with normal control. Both IL-1beta and TNF-alpha were significantly correlated with scores on the general psychopathology subscale of the PANSS. A significant positive correlation between IL-1beta and TH expression was found in both sibling and normal controls, but not in patients, while a positive correlation between IL-1beta and TNF-alpha was significant in all the groups. These results suggest that TH, IL-1beta and TNF-alpha are overexpressed in the peripheral blood mononuclear cells of schizophrenia patients, perhaps due to the hereditary factors. IL-1beta and TNF-alpha may influence the symptoms of schizophrenia in the cognition dysfunction and anxiety/depression domains of the PANSS.

  8. Interleukin-1beta but not tumor necrosis factor-alpha potentiates neuronal damage by quinolinic acid: protection by an adenosine A2A receptor antagonist.

    PubMed

    Stone, Trevor W; Behan, Wilhelmina M H

    2007-04-01

    Quinolinic acid is an agonist at glutamate receptors sensitive to N-methyl-D-aspartate (NMDA). It has been implicated in neural dysfunction associated with infections, trauma, and ischemia, although its neurotoxic potency is relatively low. This study was designed to examine the effects of a combination of quinolinic acid and the proinflammatory cytokines interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha). Compounds were administered to the hippocampus of anesthetized male rats, animals being allowed to recover for 7 days before histological analysis of the hippocampus for neuronal damage estimated by counting of intact, healthy neurons. A low dose of quinolinic acid or IL-1beta produced no damage by itself, but the two together induced a significant loss of pyramidal neurons in the hippocampus. Higher doses produced almost total loss of pyramidal cells. Intrahippocampal TNF-alpha produced no effect alone but significantly reduced the neuronal loss produced by quinolinic acid. The adenosine A(2A) receptor antagonist ZM241385 reduced neuronal loss produced by the combinations of quinolinic acid and IL-1beta. The results suggest that simultaneous quinolinic acid and IL-1beta, both being induced by cerebral infection or injury, are synergistic in the production of neuronal damage and could together contribute substantially to traumatic, infective, or ischemic cerebral damage. Antagonism of adenosine A(2A) receptors protects neurons against the combination of quinolinic acid and IL-1beta.

  9. Monocytes can be induced to express lymphatic phenotypes.

    PubMed

    Changming, W; Xin, L; Hua, T; Shikun, W; Qiong, X; Zhigeng, Z; Xueying, W

    2011-06-01

    Although it has been recently shown that monocytes can transdifferentiate into blood vascular endothelial cells which are involved in angiogenesis, little attention has been paid to their potential to transdifferentiate into lymphatic endothelial cells. Therefore, we examined this question in our study. We first stimulated monocytes with either fibronectin (FN), VEGF-C, TNF-alpha, LPS, or IL-3 for 24h. Then we examined the expression of several markers of lymphatic endothelium and found that the monocytes expressed specific lymphatic endothelial markers, LYVE-1, Podoplanin, and Prox-1, but not common endothelial markers vWF or eNOS. Next, monocytes were incubated in endothelial growth medium with FN and VEGF-C for 6d. These monocytes were also found to express LYVE-1, Podoplanin and Prox-1, but not vWF or eNOS. Our results indicate that monocytes in vitro can be easily induced to present lymphatic phenotypes in an inflammatory environment.

  10. Expression of hyaluronidase by tumor cells induces angiogenesis in vivo.

    PubMed Central

    Liu, D; Pearlman, E; Diaconu, E; Guo, K; Mori, H; Haqqi, T; Markowitz, S; Willson, J; Sy, M S

    1996-01-01

    Hyaluronic acid is a proteoglycan present in the extracellular matrix and is important for the maintenance of tissue architecture. Depolymerization of hyaluronic acid may facilitate tumor invasion. In addition, oligosaccharides of hyaluronic acid have been reported to induce angiogenesis. We report here that a hyaluronidase similar to the one on human sperm is expressed by metastatic human melanoma, colon carcinoma, and glioblastoma cell lines and by tumor biopsies from patients with colorectal carcinomas, but not by tissues from normal colon. Moreover, angiogenesis is induced by hyaluronidase+ tumor cells but not hyaluronidase- tumor cells and can be blocked by an inhibitor of hyaluronidase. Tumor cells thus use hyaluronidase as one of the "molecular saboteurs" to depolymerize hyaluronic acid to facilitate invasion. As a consequence, breakdown products of hyaluronic acid can further promote tumor establishment by inducing angiogenesis. Hyaluronidase on tumor cells may provide a target for anti-neoplastic drugs. Images Fig. 1 Fig. 2 Fig. 3 PMID:8755562

  11. Expression of Hyaluronidase by Tumor Cells Induces Angiogenesis in vivo

    NASA Astrophysics Data System (ADS)

    Liu, Dacai; Pearlman, Eric; Diaconu, Eugenia; Guo, Kun; Mori, Hiroshi; Haqqi, Tariq; Markowitz, Sanford; Willson, James; Sy, Man-Sun

    1996-07-01

    Hyaluronic acid is a proteoglycan present in the extracellular matrix and is important for the maintenance of tissue architecture. Depolymerization of hyaluronic acid may facilitate tumor invasion. In addition, oligosaccharides of hyaluronic acid have been reported to induce angiogenesis. We report here that a hyaluronidase similar to the one on human sperm is expressed by metastatic human melanoma, colon carcinoma, and glioblastoma cell lines and by tumor biopsies from patients with colorectal carcinomas, but not by tissues from normal colon. Moreover, angiogenesis is induced by hyaluronidase+ tumor cells but not hyaluronidase- tumor cells and can be blocked by an inhibitor of hyaluronidase. Tumor cells thus use hyaluronidase as one of the ``molecular saboteurs'' to depolymerize hyaluronic acid to facilitate invasion. As a consequence, breakdown products of hyaluronic acid can further promote tumor establishment by inducing angiogenesis. Hyaluronidase on tumor cells may provide a target for anti-neoplastic drugs.

  12. Early growth response protein 1 (EGR1) regulates pro-inflammatory gene expression in response to palmitate and TNF alpha in human placenta cells and is induced in obese placenta

    Technology Transfer Automated Retrieval System (TEKTRAN)

    Maternal obesity has been hypothesized to induce a pro-inflammatory response in the placenta. However, the specific factors contributing to this pro-infalmmatory response are yet to be determined. Our objective was to examine the effects of palmitic acid (PA), tumor necrosis factor alpha (TNF alph...

  13. Calcium oxalate calculi-induced clusterin expression in kidney.

    PubMed

    Li, Jin-Yi; Liu, Junjiang; Jiang, Junyi; Pumill, Chris; Elaiho, Cordelia; Zhang, Yunxia; Li, Shoubin; Zhou, Tie

    2015-10-01

    The aim of the study was to investigate clusterin expression in the kidney and evaluate the urine clusterin level in the kidney stone formers. (1) In vitro, we treated the Madin-Darby canine kidney (MDCK) cell line with different concentrations of calcium oxalate (CaOx), and then the clusterin protein expression in the cells was evaluated by Western blotting. (2) Kidney stone patients who received percutaneous nephrolithotomy were enrolled in our study. Urine samples were collected before surgery, the kidney punctured to obtain kidney tissue guided by ultrasound intraoperatively. Clusterin expression in the human kidney tissue was evaluated by immunochemistry. The urine clusterin level was determined by enzyme-linked immunosorbent assay. Non-kidney disease subjects were chosen as controls. In vitro, the clusterin expression was up-regulated in the MDCK cells induced by CaOx. The study included 49 patients and 41 non-kidney disease subjects. All calculi were composed of calcium oxalate monohydrate or calcium oxalate dihydrate and a few also contained protein or uric acid. Mean ± SD urine clusterin level was 17.47 ± 18.61 μg/ml in patients, and 3.31 ± 5.42 μg/ml in non-kidney disease subjects, respectively (p < 0.001). Immunohistochemistry revealed the clusterin was located in the cytoplasm of the renal distal and collecting tubular epithelial cells. Also the tissue clusterin expression increased significantly in the kidney stone formers compared to the control groups (p = 0.001). CaOx could induce clusterin expression in renal tubular cells, and increase clusterin levels in the kidney and urine from the kidney stone formers.

  14. Molecular characterisation of tumour necrosis factor alpha and its potential connection with lipoprotein lipase and peroxisome proliferator-activated receptors in blunt snout bream (Megalobrama amblycephala).

    PubMed

    Zhou, Man; Mi, Hai-Feng; Liu, Wen-Bin; Wu, Ye-Yang; Wang, Kai-Zhou; Jiang, Guang-Zhen

    2017-02-13

    Tumour necrosis factor alpha (TNF-α) is one kind of cytokines which is related to inflammation and lipid metabolism. TNF-α cDNA was cloned from the liver of blunt snout bream (Megalobrama amblycephala) through real-time polymerase chain reaction (PCR) and rapid amplification of cDNA ends (RACE) methods. The full-length cDNA of TNF-α covered 1467 bp, with an open reading frame (ORF) of 723 bp, which encodes 240 amino acids. It possessed the TNF family signature IIIPDDGIYFVYSQ. After the lipopolysaccharide (LPS) challenge test, a graded tissue-specific expression pattern of TNF-α was observed and there was high expression abundance in the kidney, brain and liver. After 8 weeks feeding trial, liver samples, two groups fed with 6% and 11% lipid levels, were collected. The results showed that, for fish fed with high-fat diet, the triglyceride of serum and lipid content of liver were elevated. Furthermore, TNF-α and peroxisome proliferator-activated receptors (PPARα, β) mRNA expression of fish fed 11% lipid diet were significantly up-regulated (p < 0.05). Lipoprotein lipase (LPL) and PPARγ mRNA expression of fish fed 11% lipid lever diet were significantly decreased compared to those of fish fed 6% (p < 0.05). The differences between the various expression of related genes in the high and low fat groups demonstrated that TNF-α played a key role in lipid metabolism, which may have an influence on fat metabolism through reducing fat synthesis and strengthening the β-oxidation of fatty acid. These discrepancies warrant further research.

  15. Tumor necrosis factor-{alpha} enhanced fusions between oral squamous cell carcinoma cells and endothelial cells via VCAM-1/VLA-4 pathway

    SciTech Connect

    Song, Kai; Zhu, Fei; Zhang, Han-zhong; Shang, Zheng-jun

    2012-08-15

    Fusion between cancer cells and host cells, including endothelial cells, may strongly modulate the biological behavior of tumors. However, no one is sure about the driving factors and underlying mechanism involved in such fusion. We hypothesized in this study that inflammation, one of the main characteristics in tumor microenvironment, serves as a prominent catalyst for fusion events. Our results showed that oral cancer cells can fuse spontaneously with endothelial cells in co-culture and inflammatory cytokine tumor necrosis factor-{alpha} (TNF-{alpha}) increased fusion of human umbilical vein endothelium cells and oral cancer cells by up to 3-fold in vitro. Additionally, human oral squamous cell carcinoma cell lines and 35 out of 50 (70%) oral squamous carcinoma specimens express VLA-4, an integrin, previously implicated in fusions between human peripheral blood CD34-positive cells and murine cardiomyocytes. Expression of VCAM-1, a ligand for VLA-4, was evident on vascular endothelium of oral squamous cell carcinoma. Moreover, immunocytochemistry and flow cytometry analysis revealed that expression of VCAM-1 increased obviously in TNF-{alpha}-stimulated endothelial cells. Anti-VLA-4 or anti-VCAM-1 treatment can decrease significantly cancer-endothelial adhesion and block such fusion. Collectively, our results suggested that TNF-{alpha} could enhance cancer-endothelial cell adhesion and fusion through VCAM-1/VLA-4 pathway. This study provides insights into regulatory mechanism of cancer-endothelial cell fusion, and has important implications for the development of novel therapeutic strategies for prevention of metastasis. -- Highlights: Black-Right-Pointing-Pointer Spontaneous oral cancer-endothelial cell fusion. Black-Right-Pointing-Pointer TNF-{alpha} enhanced cell fusions. Black-Right-Pointing-Pointer VCAM-1/VLA-4 expressed in oral cancer. Black-Right-Pointing-Pointer TNF-{alpha} increased expression of VCAM-1 on endothelial cells. Black

  16. Gemcitabine-induced CXCL8 expression counteracts its actions by inducing tumor neovascularization

    SciTech Connect

    Song, Yao; Baba, Tomohisa; Li, Ying-Yi; Furukawa, Kaoru; Tanabe, Yamato; Matsugo, Seiichi; Sasaki, Soichiro; Mukaida, Naofumi

    2015-03-06

    Patients with pancreatic ductal adenocarcinoma (PDAC) are frequently complicated with metastatic disease or locally advanced tumors, and consequently need chemotherapy. Gemcitabine is commonly used for PDAC treatment, but with limited efficacy. The capacity of gemcitabine to generate reactive oxygen species (ROS) in human pancreatic cancer cells, prompted us to examine its effects on the expression of pro-inflammatory cytokines and chemokines. We observed that gemcitabine enhanced selectively the expression of CXCL8 in human pancreatic cancer cells through ROS generation and NF-κB activation. In vitro blocking of CXCL8 failed to modulate gemcitabine-mediated inhibition of cell proliferation in human pancreatic cancer cells. Gemcitabine also enhanced CXCL8 expression in pancreatic cancer cells in xenografted tumor tissues. Moreover, anti-CXCL8 antibody treatment in vivo attenuated tumor formation as well as intra-tumoral vascularity in nude mice, which were transplanted with Miapaca-2 cells and treated with gemcitabine. Thus, gemcitabine-induced CXCL8 may counteract the drug through inducing neovascularization. - Highlights: • Gemcitabine induced CXCL8 expression in human pancreatic cancer cells. • CXCL8 expression required ROS generation and NF-κB activation. • CXCL8 did not affect in vitro proliferation of human pancreatic cancer cells. • CXCL8 in vivo counteracted gemcitabine by inducing neovascularization.

  17. Chronic DPP-IV inhibition with PKF-275-055 attenuates inflammation and improves gene expressions responsible for insulin secretion in streptozotocin induced diabetic rats.

    PubMed

    Akarte, Atul Sureshrao; Srinivasan, B P; Gandhi, Sonia; Sole, Sushant

    2012-09-29

    Inhibitors of dipeptidyl peptidase-4 (DPP-IV), a key regulator of the actions of incretin hormones, exert antihyperglycemic effects in type 2 diabetic patients. A major question concerns the potential ability of long term DPP-IV inhibition to have beneficial disease-modifying effects, specifically to attenuate loss of pancreatic β-cell mass due to oxidative stress induced inflammation. Here, we investigated the effects of a potent and selective DPP-4 inhibitor, an analog of vildagliptin (PKF-275-055), on glycemic control, pancreatic β-cell mass, genes and proteins expressions, tumor necrosis factor-alpha, and nitric oxide in an n2-STZ diabetic model of rat with defects in insulin sensitivity and secretion. To induce NIDDM, streptozotocin (STZ) 90 mg/kg was administered i.p. to a group of 2 days old pups. Diabetic rats were administered orally with vildagliptin analog PKF-275-055. Saline treated animals served as diabetic control. Significant and dose-dependent correction of postprandial hyperglycemia was observed in diabetic rats following 8 weeks of chronic therapy. Treatment with PKF-275-055 showed increased the number of insulin-positive β-cells in islets and improved the expressions of genes and proteins are responsible for insulin secretions. In addition, treatment of rats with PKF-275-055 significantly increased insulin content, glycogen content and total proteins content; and decreased the inflammatory markers i.e. nitric oxide and TNF-alpha. The present studies indicate that PKF-275-055 is a novel selective DPP-IV inhibitor having potential to reduce inflammation that might be a potential agent for type 2 diabetes.

  18. Low doses of neutrons induce changes in gene expression

    SciTech Connect

    Woloschak, G.E.; Chang-Liu, C.M. ); Panozzo, J.; Libertin, C.R. )

    1993-01-01

    Studies were designed to identify genes induced following low-dose neutron but not following [gamma]-ray exposure in fibroblasts. Our past work had shown differences in the expression of [beta]-protein kinase C and c-fos genes, both being induced following [gamma]-ray but not neutron exposure. We have identified two genes that are induced following neutron, but not [gamma]-ray, exposure: Rp-8 (a gene induced by apoptosis) and the long terminal repeat (LTR) of the human immunodeficiency (HIV). Rp-8 mRNA induction was demonstrated in Syrian hamster embryo fibroblasts and was found to be induced in cells exposed to neutrons administered at low (0.5 cGy/min) and at high dose rate (12 cGy/min). The induction of transcription from the LTR of HIV was demonstrated in HeLa cells bearing a transfected construct of the chloramphenicol acetyl transferase (CAT) gene driven by the HIV-LTR promoter. Measures of CAT activity and CAT transcripts following irradiation demonstrated an unresponsiveness to [gamma] rays over a broad range of doses. Twofold induction of the HIV-LTR was detected following neutron exposure (48 cGy) administered at low (0.5 cGy/min) but not high (12 cGy/min) dose rates. Ultraviolet-mediated HIV-LTR induction was inhibited by low-dose-rate neutron exposure.

  19. Low doses of neutrons induce changes in gene expression

    SciTech Connect

    Woloschak, G.E.; Chang-Liu, C.M.; Panozzo, J.; Libertin, C.R.

    1993-06-01

    Studies were designed to identify genes induced following low-dose neutron but not following {gamma}-ray exposure in fibroblasts. Our past work had shown differences in the expression of {beta}-protein kinase C and c-fos genes, both being induced following {gamma}-ray but not neutron exposure. We have identified two genes that are induced following neutron, but not {gamma}-ray, exposure: Rp-8 (a gene induced by apoptosis) and the long terminal repeat (LTR) of the human immunodeficiency (HIV). Rp-8 mRNA induction was demonstrated in Syrian hamster embryo fibroblasts and was found to be induced in cells exposed to neutrons administered at low (0.5 cGy/min) and at high dose rate (12 cGy/min). The induction of transcription from the LTR of HIV was demonstrated in HeLa cells bearing a transfected construct of the chloramphenicol acetyl transferase (CAT) gene driven by the HIV-LTR promoter. Measures of CAT activity and CAT transcripts following irradiation demonstrated an unresponsiveness to {gamma} rays over a broad range of doses. Twofold induction of the HIV-LTR was detected following neutron exposure (48 cGy) administered at low (0.5 cGy/min) but not high (12 cGy/min) dose rates. Ultraviolet-mediated HIV-LTR induction was inhibited by low-dose-rate neutron exposure.

  20. Regulation of LPS-induced mRNA expression of pro-inflammatory cytokines via alteration of NF-κB activity in mouse peritoneal macrophages exposed to fluoride.

    PubMed

    Tian, Yuhu; Huo, Meijun; Li, Guangsheng; Li, Yanyan; Wang, Jundong

    2016-10-01

    F toxicity to immune system, especially to macrophage, has been studied a lot recently. Nuclear factor-kappa B (NF-κB), as a transcription factor, plays a central role in immune and inflammatory responses via the regulation of downstream gene expression. Recent studies indicated that fluoride effect on inflammatory cytokine secretion, however, the molecular mechanism was less understood. In our study, peritoneal macrophages (PMs) were divided several groups and were administrated sodium fluoride (NaF, 50, 100, 200, 400, 800 μM) and/or lipopolysaccharide (LPS, 30 ng/mg). The mRNA expression of p65, inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF-α) and interleukin-1 beta (IL-1β) in macrophages exposed to fluoride was determined by quantitative real-time RT-PCR respectively. The translocation of NF-κB from cytoplasm to nucleus, which in a way reflects NF-κB activity, was demonstrated by Immunofluorescence and ELISA. Our results showed that fluoride had a dose-dependent effect on NF-κB activity, which coincided with LPS-induced mRNA expression of its downstream genes, iNOS and IL-1β. Fluoride alone causes no effect on gene expression. However, the mRNA expression of TNF-α showed non-NF-κB-dependent manner. Therefore, we come to the conclusion that fluoride can regulate LPS-induced mRNA expression of iNOS and IL-1β via NF-κB pathway in mouse peritoneal macrophages.

  1. EGR-1 regulates Ho-1 expression induced by cigarette smoke

    SciTech Connect

    Chen, Huaqun; Wang, Lijuan; Gong, Tao; Yu, Yang; Zhu, Chunhua; Li, Fen; Wang, Li; Li, Chaojun

    2010-05-28

    As an anti-oxidant molecule, heme oxygenase-1 (HO-1) has been implicated in the protection of lung injury by cigarette smoke (CS). The mechanisms regulating its expression have not been defined. In this report, the role of early growth response 1 (EGR-1) in the regulation of Ho-1 expression was investigated. In C57BL/6 mice with CS exposure, HO-1 was greatly increased in bronchial epithelial cells and alveolar inflammatory cells. In primary cultured mouse lung fibroblasts and RAW264.7 cells exposed to cigarette smoke water extract (CSE), an increase in HO-1 protein level was detected. In addition, CSE induced HO-1 expression was decreased in Egr-1 deficient mouse embryo fibroblasts (Egr-1{sup -/-} MEFs). Nuclear localization of EGR-1 was examined in mouse lung fibroblasts after exposure to CSE. Luciferase reporter activity assays showed that the enhancer region of the Ho-1 gene containing a proposed EGR-1 binding site was responsible for the induction of HO-1. A higher increase of alveolar mean linear intercept (Lm) was observed in lung tissues, and a larger increase in the number of total cells and monocytes/macrophages from bronchial alveolar lavage fluid was found in CS-exposed mice by loss of function of EGR-1 treatment. In summary, the present data demonstrate that EGR-1 plays a critical role in HO-1 production induced by CS.

  2. Lipopolysaccharide induces autotaxin expression in human monocytic THP-1 cells

    SciTech Connect

    Li Song; Zhang Junjie

    2009-01-09

    Autotaxin (ATX) is a secreted enzyme with lysophospholipase D (lysoPLD) activity, which converts lysophosphatidylcholine (LPC) into lysophosphatidic acid (LPA), a bioactive phospholipid involved in numerous biological activities, including cell proliferation, differentiation, and migration. In the present study, we found that bacterial lipopolysaccharide (LPS), a well-known initiator of the inflammatory response, induced ATX expression in monocytic THP-1 cells. The activation of PKR, JNK, and p38 MAPK was required for the ATX induction. The LPS-induced ATX in THP-1 cells was characterized as the {beta} isoform. In the presence of LPC, ATX could promote the migrations of THP-1 and Jurkat cells, which was inhibited by pertussis toxin (PTX), an inhibitor of Gi-mediated LPA receptor signaling. In summary, LPS induces ATX expression in THP-1 cells via a PKR, JNK and p38 MAPK-mediated mechanism, and the ATX induction is likely to enhance immune cell migration in proinflammatory response by regulating LPA levels in the microenvironment.

  3. The immune adaptor molecule SARM modulates tumor necrosis factor alpha production and microglia activation in the brainstem and restricts West Nile Virus pathogenesis.

    PubMed

    Szretter, Kristy J; Samuel, Melanie A; Gilfillan, Susan; Fuchs, Anja; Colonna, Marco; Diamond, Michael S

    2009-09-01

    Sterile alpha and HEAT/Armadillo motif (SARM) is a highly conserved Toll/interleukin-1 receptor (TIR)-containing adaptor protein that is believed to negatively regulate signaling of the pathogen recognition receptors Toll-like receptor 3 (TLR3) and TLR4. To test its physiological function in the context of a microbial infection, we generated SARM(-/-) mice and evaluated the impact of this deficiency on the pathogenesis of West Nile virus (WNV), a neurotropic flavivirus that requires TLR signaling to restrict infection. Although SARM was preferentially expressed in cells of the central nervous system (CNS), studies with primary macrophages, neurons, or astrocytes showed no difference in viral growth kinetics. In contrast, viral replication was increased specifically in the brainstem of SARM(-/-) mice, and this was associated with enhanced mortality after inoculation with a virulent WNV strain. A deficiency of SARM was also linked to reduced levels of tumor necrosis factor alpha (TNF-alpha), decreased microglia activation, and increased neuronal death in the brainstem after WNV infection. Thus, SARM appears to be unique among the TIR adaptor molecules, since it functions to restrict viral infection and neuronal injury in a brain region-specific manner, possibly by modulating the activation of resident CNS inflammatory cells.

  4. 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.

  5. Tumor necrosis factor alpha -238 G/A and -308 G/A polymorphisms and soluble TNF-α levels in chronic kidney disease: correlation with clinical variables

    PubMed Central

    Vázquez-Huerta, Diana I; Alvarez-Rodríguez, Bertha A; Topete-Reyes, Jorge F; Muñoz-Valle, José F; Parra-Michel, Renato; Fuentes-Ramírez, Francisco; Salazar-López, María A; Valle, Yeminia; Reyes-Castillo, Zyanya; Cruz-González, A; Brennan-Bourdon, Lorena M; Torres-Carrillo, Norma

    2014-01-01

    Chronic kidney disease (CKD) is characterized by accumulation of proinflammatory cytokines, mainly tumor necrosis factor alpha (TNF-α). Single nucleotide polymorphisms (SNPs) of TNFA gene, including -238 G/A and -308 G/A, have been associated with alteration in the soluble TNF-α (sTNF-α) expression. The aim was to investigate the association of -238 y -308 TNFA gene SNPs with sTNF-α levels in CKD patients. We included 150 CKD patients and 192 control subjects (CS). Both SNPs were genotyped with polymerase chain reaction-restriction fragment length polymorphism technique and sTNF-α levels were measured by enzyme-linked immunosorbent assay. The genotypic distribution of -238 and -308 SNPs was not significantly different between CKD patients and CS (p > 0.001). However, the sTNF-α levels were higher in CKD, compared to CS (p < 0.001). Also, sTNF-α correlated with creatinine (r = 0.279, p = 0.004), urea (r = 0.325, p = 0.001), phosphorus (r = 0.479, p = 0.001), glomerular filtration rate (r = -0.236, p = 0.019) and monocyte count (r = 0.276, p = 0.010). In conclusion, elevated sTNF-α levels are associated with CKD. However, the -238 and -308 TNFA gene SNPs were not associated with susceptibility to CKD and sTNF-α levels in a Mexican population. PMID:25232395

  6. Role of ascorbate in the activation of NF-kappaB by tumour necrosis factor-alpha in T-cells.

    PubMed

    Muñoz, E; Blázquez, M V; Ortiz, C; Gomez-Díaz, C; Navas, P

    1997-07-01

    The first product of ascorbate oxidation, the ascorbate free radical (AFR), acts in biological systems mainly as an oxidant, and through its role in the plasma membrane redox system exerts different effects on the cell. We have investigated the role of ascorbate, AFR and dehydroascorbate (DHA) in the activation of the NF-kappaB transcription factor in Jurkat T-cells stimulated by tumour necrosis factor-alpha (TNF-alpha). Here we show, by electrophoretic mobility shift assays, that ascorbate increases the binding of NF-kappaB to DNA in TNF-alpha-stimulated Jurkat cells. The ability of ascorbate to enhance cytoplasmic inhibitory IkBalpha protein degradation correlates completely with its capacity to induce NF-kappaB binding to DNA and to potentiate NF-kappaB-mediated transactivation of the HIV-1 long terminal repeat promoter in TNF-alpha-stimulated Jurkat cells but not in cells stimulated with PMA plus ionomycin. AFR behaves like ascorbate, while DHA and ascorbate phosphate do not affect TNF-alpha-mediated NF-kappaB activation. These results provide new evidence for a possible relationship between the activation of the electron-transport system at the plasma membrane by ascorbate or its free radical and redox-dependent gene transcription in T-cells.

  7. Effects of anti-tumor necrosis factor-alpha and anti-intercellular adhesion molecule-1 antibodies on ischemia/reperfusion lung injury.

    PubMed

    Chiang, Chi-Huei

    2006-10-31

    Inhibition of neutrophil activation and adherence to endothelium by antibodies to tumor necrosis factor-alpha (TNF-alpha) and intercellular adhesion molecules (ICAM-1), respectively, might attenuate ischemia-reperfusion injury (I/R). I/R was conducted in an isolated rat lung model. Anti-TNF-alpha antibody and/or anti-ICAM-1 antibody were added before ischemia or after reperfusion. Hemodynamic changes, lung weight gain (LWG), capillary filtration coefficients (Kfc), and pathologic changes were assessed to evaluate the severity of I/R. The LWG, Kfc, pathological changes and lung injury score of treatment groups with anti-TNF-alpha antibody treatment, either pre-ischemia or during reperfusion, were less than those observed in control groups. Similar findings were found in group treated with anti-ICAM-1 antibody or combination therapy during reperfusion. In contrast, pre-I/R treatment with anti-ICAM-1 antibody induced severe lung edema and failure to complete the experimental procedure. No additional therapeutic effect was found in combination therapy. We conclude that TNF-alpha and ICAM-1 play important roles in I/R. Anti-TNF-alpha antibody has therapeutic and preventive effects on I/R. However, combined therapy with anti-TNF-alpha antibody and anti-ICAM-1 antibody may have no additive effect and need further investigation.

  8. In vitro protective effects of two extracts from bergamot peels on human endothelial cells exposed to tumor necrosis factor-alpha (TNF-alpha).

    PubMed

    Trombetta, Domenico; Cimino, Francesco; Cristani, Mariateresa; Mandalari, Giuseppina; Saija, Antonella; Ginestra, Giovanna; Speciale, Antonio; Chirafisi, Joselita; Bisignano, Giuseppe; Waldron, Keith; Narbad, Arjan; Faulds, Craig B

    2010-07-28

    Bergamot ( Citrus bergamia Risso) is a less commercialized Citrus fruit, mainly used for its essential oil extracted from the peel. Bergamot peel (BP) represents about 60% of the processed fruits and is regarded as primary waste. However, it contains good amounts of useful compounds, such as pectins and flavonoids. Many of the bioactivities of Citrus flavonoids appear to impact vascular endothelial cells. Herein, we report the protective effect of two flavonoid-rich extracts from BP (endowed with radical-scavenging properties and lacking genotoxic activity) against alterations in cell modifications induced by the pleiotropic inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) on human umbilical vein endothelial cells (HUVECs), as demonstrated by monitoring intracellular levels of malondialdehyde/4-hydroxynonenal, reduced and oxidized glutathione and superoxide dismutase activity, and the activation status of nuclear factor-kappaB (NF-kappaB). Thus, BP appears to be a potential source of natural antioxidant/anti-inflammatory phytocomplexes to be employed as ingredients of nutraceutical products or functional foods.

  9. Anti-inflammatory drugs and tumor necrosis factor-alpha production from monocytes: role of transcription factor NF-kappa B and implication for rheumatoid arthritis therapy.

    PubMed

    Lavagno, Luisa; Gunella, Gabriele; Bardelli, Claudio; Spina, Simona; Fresu, Luigia Grazia; Viano, Ilario; Brunelleschi, Sandra

    2004-10-06

    Inhibition of tumor necrosis factor-alpha (TNF-alpha) represents a relevant target in rheumatoid arthritis therapy. Besides inhibiting cyclooxygenase, anti-inflammatory drugs can affect the activation of transcription factors. We investigated the ability of dexamethasone, indomethacin, and rofecoxib to modulate nuclear factor-kappaB (NF-kappaB) activation and TNF-alpha release from human monocytes challenged with lipopolysaccharide (LPS) or phorbol 12-myristate 13-acetate (PMA). Both stimuli induced NF-kappaB nuclear translocation and TNF-alpha secretion. Dexamethasone potently inhibited TNF-alpha release, indomethacin inhibited only PMA-evoked release, while rofecoxib had no effect. In the electrophoretic mobility shift assay, dexamethasone and rofecoxib dose-dependently inhibited the DNA binding activity of NF-kappaB in stimulated monocytes, whereas indomethacin failed to inhibit the LPS-evoked one. These results were further confirmed by evaluating the drugs' ability to reduce nuclear NF-kappaB subunits, as well as the amount of phosphorylated IkappaBalpha in cytosolic fractions. In conclusion, these results indicate that anti-inflammatory drugs differ largely in their ability to inhibit NF-kappaB activity and/or TNF-alpha release from human monocytes. These effects can be relevant to rheumatoid arthritis therapy.

  10. Interaction of the CD43 Sialomucin with the Mycobacterium tuberculosis Cpn60.2 Chaperonin Leads to Tumor Necrosis Factor Alpha Production.

    PubMed

    Torres-Huerta, Alvaro; Villaseñor, Tomás; Flores-Alcantar, Angel; Parada, Cristina; Alemán-Navarro, Estefanía; Espitia, Clara; Pedraza-Alva, Gustavo; Rosenstein, Yvonne

    2017-03-01

    Mycobacterium tuberculosis is the causal agent of tuberculosis. Tumor necrosis factor alpha (TNF-α), transforming growth factor β (TGF-β), and gamma interferon (IFN-γ) secreted by activated macrophages and lymphocytes are considered essential to contain Mycobacterium tuberculosis infection. The CD43 sialomucin has been reported to act as a receptor for bacilli through its interaction with the chaperonin Cpn60.2, facilitating mycobacterium-macrophage contact. We report here that Cpn60.2 induces both human THP-1 cells and mouse-derived bone marrow-derived macrophages (BMMs) to produce TNF-α and that this production is CD43 dependent. In addition, we present evidence that the signaling pathway leading to TNF-α production upon interaction with Cpn60.2 requires active Src family kinases, phospholipase C-γ (PLC-γ), phosphatidylinositol 3-kinase (PI3K), p38, and Jun N-terminal protein kinase (JNK), both in BMMs and in THP-1 cells. Our data highlight the role of CD43 and Cpn60.2 in TNF-α production and underscore an important role for CD43 in the host-mycobacterium interaction.

  11. EHMT1 and EHMT2 inhibition induces fetal hemoglobin expression

    PubMed Central

    Renneville, Aline; Van Galen, Peter; Canver, Matthew C.; McConkey, Marie; Krill-Burger, John M.; Dorfman, David M.; Holson, Edward B.; Bernstein, Bradley E.; Orkin, Stuart H.; Bauer, Daniel E.

    2015-01-01

    Fetal hemoglobin (HbF, α2γ2) induction is a well-validated strategy for sickle cell disease (SCD) treatment. Using a small-molecule screen, we found that UNC0638, a selective inhibitor of EHMT1 and EHMT2 histone methyltransferases, induces γ-globin expression. EHMT1/2 catalyze mono- and dimethylation of lysine 9 on histone 3 (H3K9), raising the possibility that H3K9Me2, a repressive chromatin mark, plays a role in silencing γ-globin expression. In primary human adult erythroid cells, UNC0638 and EHMT1 or EHMT2 short hairpin RNA–mediated knockdown significantly increased γ-globin expression, HbF synthesis, and the percentage of cells expressing HbF. At effective concentrations, UNC0638 did not alter cell morphology, proliferation, or erythroid differentiation of primary human CD34+ hematopoietic stem and progenitor cells in culture ex vivo. In murine erythroleukemia cells, UNC0638 and Ehmt2 CRISPR/Cas9-mediated knockout both led to a marked increase in expression of embryonic β-globin genes Hbb-εy and Hbb-βh1. In primary human adult erythroblasts, chromatin immunoprecipitation followed by sequencing analysis revealed that UNC0638 treatment leads to genome-wide depletion in H3K9Me2 and a concomitant increase in the activating mark H3K9Ac, which was especially pronounced at the γ-globin gene region. In RNA-sequencing analysis of erythroblasts, γ-globin genes were among the most significantly upregulated genes by UNC0638. Further increase in γ-globin expression in primary human adult erythroid cells was achieved by combining EHMT1/2 inhibition with the histone deacetylase inhibitor entinostat or hypomethylating agent decitabine. Our data provide genetic and pharmacologic evidence that EHMT1 and EHMT2 are epigenetic regulators involved in γ-globin repression and represent a novel therapeutic target for SCD. PMID:26320100

  12. EHMT1 and EHMT2 inhibition induces fetal hemoglobin expression.

    PubMed

    Renneville, Aline; Van Galen, Peter; Canver, Matthew C; McConkey, Marie; Krill-Burger, John M; Dorfman, David M; Holson, Edward B; Bernstein, Bradley E; Orkin, Stuart H; Bauer, Daniel E; Ebert, Benjamin L

    2015-10-15

    Fetal hemoglobin (HbF, α2γ2) induction is a well-validated strategy for sickle cell disease (SCD) treatment. Using a small-molecule screen, we found that UNC0638, a selective inhibitor of EHMT1 and EHMT2 histone methyltransferases, induces γ-globin expression. EHMT1/2 catalyze mono- and dimethylation of lysine 9 on histone 3 (H3K9), raising the possibility that H3K9Me2, a repressive chromatin mark, plays a role in silencing γ-globin expression. In primary human adult erythroid cells, UNC0638 and EHMT1 or EHMT2 short hairpin RNA-mediated knockdown significantly increased γ-globin expression, HbF synthesis, and the percentage of cells expressing HbF. At effective concentrations, UNC0638 did not alter cell morphology, proliferation, or erythroid differentiation of primary human CD34(+) hematopoietic stem and progenitor cells in culture ex vivo. In murine erythroleukemia cells, UNC0638 and Ehmt2 CRISPR/Cas9-mediated knockout both led to a marked increase in expression of embryonic β-globin genes Hbb-εy and Hbb-βh1. In primary human adult erythroblasts, chromatin immunoprecipitation followed by sequencing analysis revealed that UNC0638 treatment leads to genome-wide depletion in H3K9Me2 and a concomitant increase in the activating mark H3K9Ac, which was especially pronounced at the γ-globin gene region. In RNA-sequencing analysis of erythroblasts, γ-globin genes were among the most significantly upregulated genes by UNC0638. Further increase in γ-globin expression in primary human adult erythroid cells was achieved by combining EHMT1/2 inhibition with the histone deacetylase inhibitor entinostat or hypomethylating agent decitabine. Our data provide genetic and pharmacologic evidence that EHMT1 and EHMT2 are epigenetic regulators involved in γ-globin repression and represent a novel therapeutic target for SCD.

  13. Brain CB₁ receptor expression following lipopolysaccharide-induced inflammation.

    PubMed

    Hu, H; Ho, W; Mackie, K; Pittman, Q J; Sharkey, K A

    2012-12-27

    Cannabinoid 1 receptors (CB(1)) are highly expressed on presynaptic terminals in the brain where they are importantly involved in the control of neurotransmitter release. Alteration of CB(1) expression is associated with a variety of neurological and psychiatric disorders. There is now compelling evidence that peripheral inflammatory disorders are associated with depression and cognitive impairments. These can be modeled in rodents with peripheral administration of lipopolysaccharide (LPS), but central effects of this treatment remain to be fully elucidated. As a reduction in endocannabinoid tone is thought to contribute to depression, we asked whether the expression of CB(1) in the CNS is altered following LPS treatment. CD1 mice received LPS (0.1-1mg/kg, ip) and 6h later activated microglial cells were observed only in circumventricular organs and only at the higher dose. At 24h, activated microglial cells were identified in other brain regions, including the hippocampus, a structure implicated in some mood disorders. Immunohistochemistry and real-time polymerase chain reaction (PCR) were utilized to evaluate the change of CB(1) expression 24h after inflammation. LPS induced an increase of CB(1) mRNA in the hippocampus and brainstem. Subsequent immunohistochemical analysis revealed reduced CB(1) in the hippocampus, especially in CA3 pyramidal layer. Analysis of co-localization with markers of excitatory and inhibitory terminals indicated that the decrease in CB(1) expression was restricted to glutamatergic terminals. Despite widespread microglial activation, these results suggest that peripheral LPS treatment leads to limited changes in CB(1) expression in the brain.

  14. A chloride-inducible gene expression cassette and its use in induced lysis of Lactococcus lactis.

    PubMed Central

    Sanders, J W; Venema, G; Kok, J

    1997-01-01

    A chloride-inducible promoter previously isolated from the chromosome of Lactococcus lactis (J. W. Sanders, G. Venema, J. Kok, and K. Leenhouts, Mol. Gen. Genet., in press) was exploited for the inducible expression of homologous and heterologous genes. An expression cassette consisting of the positive-regulator gene gadR, the chloride-inducible promoter Pgad, and the translation initiation signals of gadC was amplified by PCR. The cassette was cloned upstream of Escherichia coli lacZ, the holin-lysin cassette (lytPR) of the lactococcal bacteriophage r1t, and the autolysin gene of L. lactis, acmA. Basal activity of Pgad resulted in a low level of expression of all three proteins. Growth in the presence of 0.5 M NaCl of a strain containing the gadC::lacZ fusion resulted in a 1,500-fold increase of beta-galactosidase activity. The background activity levels of LytPR and AcmA had no deleterious effects on cell growth, but induction of lysin expression by addition of 0.5 M NaCl resulted in inhibition of growth. Lysis was monitored by following the release of the cytoplasmic marker enzyme PepX. Released PepX activity was maximal at 1 day after induction of lytPR expression with 0.1 M NaCl. Induction of acmA expression resulted in slower release of PepX from the cells. The presence of the inducing agent NaCl resulted in the stabilization of osmotically fragile cells. PMID:9406408

  15. Inducible nitric oxide synthase is expressed in synovial fluid granulocytes.

    PubMed

    Cedergren, J; Forslund, T; Sundqvist, T; Skogh, T

    2002-10-01

    The objective of the study was to evaluate the NO-producing potential of synovial fluid (SF) cells. SF from 15 patients with arthritis was compared with blood from the same individuals and with blood from 10 healthy controls. Cellular expression of inducible nitric oxide synthase (iNOS) was analysed by flow cytometry. High-performance liquid chromatography was used to measure l-arginine and l-citrulline. Nitrite and nitrate were measured colourimetrically utilizing the Griess' reaction. Compared to whole blood granulocytes in patients with chronic arthritis, a prominent iNOS expression was observed in SF granulocytes (P < 0.001). A slight, but statistically significant, increase in iNOS expression was also recorded in lymphocytes and monocytes from SF. l-arginine was elevated in SF compared to serum (257 +/- 78 versus 176 +/- 65 micro mol/l, P = 0.008), whereas a slight increase in l-citrulline (33 +/- 11 versus 26 +/- 9 micro mol/l), did not reach statistical significance. Great variations but no significant differences were observed comparing serum and SF levels of nitrite and nitrate, respectively, although the sum of nitrite and nitrate tended to be elevated in SF (19.2 +/- 20.7 versus 8.6 +/- 6.5 micro mol/l, P = 0.054). Synovial fluid leucocytes, in particular granulocytes, express iNOS and may thus contribute to intra-articular NO production in arthritis.

  16. Thyroidectomy induces neurofilament expression in adenohypophyses of rats.

    PubMed

    Salinas, E; Quintanar, J L

    2001-08-01

    We studied the effect of thyroidectomy on neurofilament expression in adenohypophyses of rats. The question of whether thyroxine (T4) administration can reduce this effect was also investigated. Rats were divided into: 1. Euthyroid controls, 2. Thyroidectomized 20 d (Tx 20 d), 3. Thyroidectomized 20 d with replacement of T4 (Tx 20 d + T4 20 d), 4. Thyroidectomized 40 d (Tx 40 d), 5. Thyroidectomized 40 d with replacement of T4 20 d after surgery (Tx 40 d + T4 20 d). Adenohypophyses were studied by immunohistochemistry and Western blot analysis using antibodies against neurofilament 200 kDa (NF-H) and thyroid-stimulating hormone (TSH). The number of thyrotrophs with immunoreactivity for NF-H was increased in Tx 20 d and Tx 40 d rats, whereas T4 administration protected the effect of thyroidectomy. In the thyroidectomized animals, thyrotrophs showed eccentric nuclei and the cytoplasm was full of NF-H immunoreactivity, whereas in T4 treated rats, the thyrotrophs were similar to control. Western blot analysis showed that NF-H expression increased in rats thyroidectomized for 20 and 40 d. T4 given immediately or 20 d after thyroidectomy caused no changes in NF-H expression. We conclude that thyroidectomy induces NF-H expression in adenohypophyses of rats and administration of T4 decreases this effect.

  17. Strong Magnetic Field Induced Changes of Gene Expression in Arabidopsis

    NASA Astrophysics Data System (ADS)

    Paul, A.-L.; Ferl, R. J.; Klingenberg, B.; Brooks, J. S.; Morgan, A. N.; Yowtak, J.; Meisel, M. W.

    2005-07-01

    We review our studies of the biological impact of magnetic field strengths of up to 30 T on transgenic arabidopsis plants engineered with a stress response gene consisting of the alcohol dehydrogenase (Adh) gene promoter driving the β-glucuronidase (GUS) gene reporter. Field strengths in excess of 15 T induce expression of the Adh/GUS transgene in the roots and leaves. Microarray analyses indicate that such field strengths have a far reaching effect on the genome. Wide spread induction of stress-related genes and transcription factors, and a depression of genes associated with cell wall metabolism are prominent examples.

  18. Lactobacillus fermentum ZYL0401 Attenuates Lipopolysaccharide-Induced Hepatic TNF-α Expression and Liver Injury via an IL-10- and PGE2-EP4-Dependent Mechanism

    PubMed Central

    Lv, Longxian; Yang, Jianzhuan; Lu, Haifeng; Li, Lanjuan

    2015-01-01

    Lipopolysaccharide (LPS) has essential role in the pathogenesis of D-galactosamine-sensitized animal models and alcoholic liver diseases of humans, by stimulating release of pro-inflammatory mediators that cause hepatic damage and intestinal barrier impairment. Oral pretreatment of probiotics has been shown to attenuate LPS-induced hepatic injury, but it is unclear whether the effect is direct or due to improvement in the intestinal barrier. The present study tested the hypothesis that pretreatment with probiotics enables the liver to withstand directly LPS-induced hepatic injury and inflammation. In a mouse model of LPS-induced hepatic injury, the levels of hepatic tumor necrosis factor-alpha (TNF-α) and serum alanine aminotransferase (ALT) of mice with depleted intestinal commensal bacteria were not significantly different from that of the control models. Pre-feeding mice for 10 days with Lactobacillus fermentum ZYL0401 (LF41), significantly alleviated LPS-induced hepatic TNF-α expression and liver damage. After LF41 pretreatment, mice had dramatically more L.fermentum-specific DNA in the ileum, significantly higher levels of ileal cyclooxygenase (COX)-2 and interleukin 10 (IL-10) and hepatic prostaglandin E2 (PGE2). However, hepatic COX-1, COX-2, and IL-10 protein levels were not changed after the pretreatment. There were also higher hepatic IL-10 protein levels after LPS challenge in LF41-pretreaed mice than in the control mice. Attenuation of hepatic TNF-α was mediated via the PGE2/E prostanoid 4 (EP4) pathway, and serum ALT levels were attenuated in an IL-10-dependent manner. A COX-2 blockade abolished the increase in hepatic PGE2 and IL-10 associated with LF41. In LF41-pretreated mice, a blockade of IL-10 caused COX-2-dependent promotion of hepatic PGE2, without affecting hepatic COX-2levels. In LF41-pretreated mice, COX2 prevented enhancing TNF-α expression in both hepatic mononuclear cells and the ileum, and averted TNF-α-mediated increase in

  19. Lactobacillus fermentum ZYL0401 Attenuates Lipopolysaccharide-Induced Hepatic TNF-α Expression and Liver Injury via an IL-10- and PGE2-EP4-Dependent Mechanism.

    PubMed

    Jin, Pengfeng; Chen, Yunbo; Lv, Longxian; Yang, Jianzhuan; Lu, Haifeng; Li, Lanjuan

    2015-01-01

    Lipopolysaccharide (LPS) has essential role in the pathogenesis of D-galactosamine-sensitized animal models and alcoholic liver diseases of humans, by stimulating release of pro-inflammatory mediators that cause hepatic damage and intestinal barrier impairment. Oral pretreatment of probiotics has been shown to attenuate LPS-induced hepatic injury, but it is unclear whether the effect is direct or due to improvement in the intestinal barrier. The present study tested the hypothesis that pretreatment with probiotics enables the liver to withstand directly LPS-induced hepatic injury and inflammation. In a mouse model of LPS-induced hepatic injury, the levels of hepatic tumor necrosis factor-alpha (TNF-α) and serum alanine aminotransferase (ALT) of mice with depleted intestinal commensal bacteria were not significantly different from that of the control models. Pre-feeding mice for 10 days with Lactobacillus fermentum ZYL0401 (LF41), significantly alleviated LPS-induced hepatic TNF-α expression and liver damage. After LF41 pretreatment, mice had dramatically more L.fermentum-specific DNA in the ileum, significantly higher levels of ileal cyclooxygenase (COX)-2 and interleukin 10 (IL-10) and hepatic prostaglandin E2 (PGE2). However, hepatic COX-1, COX-2, and IL-10 protein levels were not changed after the pretreatment. There were also higher hepatic IL-10 protein levels after LPS challenge in LF41-pretreated mice than in the control mice. Attenuation of hepatic TNF-α was mediated via the PGE2/E prostanoid 4 (EP4) pathway, and serum ALT levels were attenuated in an IL-10-dependent manner. A COX-2 blockade abolished the increase in hepatic PGE2 and IL-10 associated with LF41. In LF41-pretreated mice, a blockade of IL-10 caused COX-2-dependent promotion of hepatic PGE2, without affecting hepatic COX-2 levels. In LF41-pretreated mice, COX2 prevented enhancing TNF-α expression in both hepatic mononuclear cells and the ileum, and averted TNF-α-mediated increase in

  20. Benzo(a)pyrene induces oxidative stress, pro-inflammatory cytokines, expression of nuclear factor-kappa B and deregulation of wnt/beta-catenin signaling in colons of BALB/c mice.

    PubMed

    Ajayi, Babajide O; Adedara, Isaac A; Farombi, Ebenezer O

    2016-09-01

    The incidence of colonic toxicity has been epidemiologically linked to the consumption of foods contaminated with benzo(a)pyrene (B[a]P). The present study investigated the effects of B[a]P on biomarkers of oxidative stress, inflammation and wnt-signaling in colon of BALB/c mice following exposure to 62.5, 125 and 250 mg/kg of B[a]P for 7 days by oral gavage. Exposure to B[a]P significantly decreased the colonic antioxidant enzymes activities and glutathione level with concomitant significant increase in myeloperoxidase activity, nitric oxide and lipid peroxidation levels. Colon histopathology results showed treatment-related lesions characterized by atrophy, mucosal ulceration and gland erosion in the B[a]P-treated mice. Immunohistochemistry analysis showed that B[a]P treatment increased the protein expression of nuclear factor kappa B, pro-inflammatory cytokines namely tumor necrosis factor alpha and interleukin-1β, as well as cyclooxygenase-2 and inducible nitric oxide synthase in the mice colon. Altered canonical wnt-signaling was confirmed by strong diaminobenzidine staining for p38 mitogen activated protein kinase, β-catenin expression and absence of adenomatous polyposis coli following B[a]P administration. The present data highlight that exposure to B[a]P induces colon injury via induction of oxidative and nitrosative stress, inflammatory biomarkers and dsyregulation wnt/β-catenin signaling, thus confirming the role of B[a]P in the pathogenesis of colonic toxicity.

  1. Down-regulation of HSP60 expression by RNAi increases lipopolysaccharide- and cerulein-induced damages on isolated rat pancreatic tissues

    PubMed Central

    Lu, Shuai; Li, Kun; Feng, Jia-Yan; Li, Yan-Na; Gao, Zhi-Rong; Chen, Chang-Jie

    2010-01-01

    The objective of this study was to investigate the function of heat shock protein 60 (HSP60) on pancreatic tissues by applying HSP60 small interfering RNA (siRNA) to reduce HSP60 expression. Rat pancreas was isolated and pancreatic tissue snips were prepared, cultured, and stimulated with low and high concentrations of cerulein (10−11 and 10−5 mol/L) or lipopolysaccharide (LPS, 10 and 20 μg/mL). Before the stimulation and 1 and 4 h after the stimulation, the viability and the level of trypsinogen activation peptide (TAP) in the tissue fragments were determined and the levels of tumor necrosis factor-alpha (TNF-α) and interleukin 6 (IL-6) in the culture supernatants were measured. Real-time PCR and Western blotting were used to evaluate the HSP60 mRNA and protein expression. After the administration of siRNA to inhibit HSP60 expression in the isolated tissues, these injury parameters were measured and compared. The pancreatic tissues in the control (mock-interfering) group showed a decreased viability to varying degrees after being stimulated with cerulein or LPS, and the levels of TAP, TNF-α, and IL-6 increased significantly (p < 0.05) in the tissues and/or in the culture supernatant. The expressions of HSP60 mRNA and protein were raised moderately after stimulating 1 h with low concentrations of cerulein or LPS, but decreased with high concentrations of the toxicants. In particular, the expression of HSP60 protein was reduced significantly (p < 0.05) when the tissues were stimulated by the two toxicants for 4 h. In contrast, the tissue fragments in which HSP60 siRNA was applied showed much lower tissue viability (p < 0.01) and higher levels of TNF-a, IL-6, and TAP (p < 0.01) in the tissues or culture supernatant after stimulating with the toxicants at the same dose and for the same time duration as compared with those of the control groups (p < 0.05). The results indicated that both cerulein and LPS can induce injuries on isolated

  2. Regulation of cytokine production by soluble CD23: costimulation of interferon gamma secretion and triggering of tumor necrosis factor alpha release

    PubMed Central

    1994-01-01

    Soluble CD23 (sCD23) has multiple IgE-independent biological activities. In the present study, we examined the regulatory effect of sCD23 on cytokine production by human peripheral blood mononuclear cells (PBMC). We show that sCD23 enhances by about 80-fold the interleukin 2 (IL-2)-induced interferon gamma (IFN-gamma) production and by about 10-fold the response to IL-12. This potentiating activity is time and dose dependent and is not associated with a significant effect on DNA synthesis. The sCD23 costimulatory activity for IFN-gamma synthesis is drastically reduced in monocyte-depleted PBMC, suggesting that monocytes may be the target for sCD23. This hypothesis was supported by the following observations. First, sCD23 alone is a potent inducer of tumor necrosis factor alpha (TNF-alpha) production by PBMC and this effect disappears after monocyte depletion. The triggering of TNF-alpha release is specifically inhibited by neutralizing anti-CD23 monoclonal antibody (mAb). In addition, IL-2 and IL-12 synergize with sCD23 to induce TNF-alpha production. Second, sCD23 triggers the release of other inflammatory mediators such as IL-1 alpha, IL-1 beta, and IL-6. Finally, TNF-alpha production in response to IL-2 and sCD23 precedes IFN-gamma and IFN-gamma secretion is significantly inhibited by anti-TNF-alpha mAb, indicating that the sCD23 costimulatory signal for IFN-gamma production may be partially mediated by TNF-alpha release. It is proposed that sCD23 is a proinflammatory cytokine that, in addition, may play an important role in the control of the immune response via the enhancement of IFN-gamma production. PMID:8064221

  3. Effect of Interleukin-1beta and Tumor Necrosis Factor-alpha on Gene Expression in Human Endothelial Cells

    DTIC Science & Technology

    2003-06-01

    bela 3 (phosphatidylinositoJ.specific) AA846573 CD47 antigen (Rh-related antigen, integrin-associated signal transducer) AA455448 proteasome ( prosome ...macropain) activator subunit 1 (PA28 alpha) T47815 proteasome ( prosome , macropain) activator subunH 2 (PA28 beta) H65395 stimulated trans-acting...class I, C AA464246 natural kiter cell transcript 4 AA458965 diubiqu~in N33920 proteasome ( prosome , macropain) subunit, alpha type, 6 AA047338 matrix

  4. Immunomodulatory Factors Galectin-9 and Interferon-Gamma Synergize to Induce Expression of Rate-Limiting Enzymes of the Kynurenine Pathway in the Mouse Hippocampus

    PubMed Central

    Brooks, Alexandra K.; Lawson, Marcus A.; Rytych, Jennifer L.; Yu, Kevin C.; Janda, Tiffany M.; Steelman, Andrew J.; McCusker, Robert H.

    2016-01-01

    Elevated levels of circulating pro-inflammatory cytokines are associated with symptomology of several psychiatric disorders, notably major depressive disorder. Symptomology has been linked to inflammation/cytokine-dependent induction of the Kynurenine Pathway. Galectins, like pro-inflammatory cytokines, play a role in neuroinflammation and the pathogenesis of several neurological disorders but without a clearly defined mechanism of action. Their involvement in the Kynurenine Pathway has not been investigated. Thus, we searched for a link between galectins and the Kynurenine Pathway using in vivo and ex vivo models. Mice were administered LPS and pI:C to determine if galectins (Gal’s) were upregulated in the brain following in vivo inflammatory challenges. We then used organotypic hippocampal slice cultures (OHSCs) to determine if Gal’s, alone or with inflammatory mediators [interferon-gamma (IFNγ), tumor necrosis factor-alpha (TNFα), interleukin-1beta (IL-1β), polyinosine-polycytidylic acid (pI:C), and dexamethasone (Dex; synthetic glucocorticoid)], would increase expression of indoleamine/tryptophan-2,3-dioxygenases (DO’s: Ido1, Ido2, and Tdo2; Kynurenine Pathway rate-limiting enzymes). In vivo, hippocampal expression of cytokines (IL-1β, TNFα, and IFNγ), Gal-3, and Gal-9 along with Ido1 and Ido2 were increased by LPS and pI:C (bacterial and viral mimetics). Of the cytokines induced in vivo, only IFNγ increased expression of two Ido1 transcripts (Ido1-FL and Ido1-v1) by OHSCs. Although ineffective alone, Gal-9 accentuated IFNγ-induced expression of only Ido1-FL. Similarly, IFNγ induced expression of several Ido2 transcripts (Ido2-v1, Ido2-v3, Ido2-v4, Ido2-v5, and Ido2-v6). Gal-9 accentuated IFNγ-induced expression of only Ido2-v1. Surprisingly, Gal-9 alone, slightly but significantly, induced expression of Tdo2 (Tdo2-v1 and Tdo2-v2, but not Tdo2-FL). These effects were specific to Gal-9 as Gal-1 and Gal-3 did not alter DO expression. These results

  5. Bitumen fume-induced gene expression profile in rat lung

    SciTech Connect

    Gate, Laurent . E-mail: laurent.gate@inrs.fr; Langlais, Cristina; Micillino, Jean-Claude; Nunge, Herve; Bottin, Marie-Claire; Wrobel, Richard; Binet, Stephane

    2006-08-15

    Exposure to bitumen fumes during paving and roofing activities may represent an occupational health risk. To date, most of the studies performed on the biological effect of asphalt fumes have been done with regard to their content in carcinogenic polycyclic aromatic hydrocarbons (PAH). In order to gain an additional insight into the mechanisms of action of bitumen fumes, we studied their pulmonary effects in rodents following inhalation using the microarray technology. Fisher 344 rats were exposed for 5 days, 6 h/day to bitumen fumes generated at road paving temperature (170 {sup o}C) using a nose-only exposition device. With the intention of studying the early transcriptional events induced by asphalt fumes, lung tissues were collected immediately following exposure and gene expression profiles in control and exposed rats were determined by using oligonucleotide microarrays. Data analysis revealed that genes involved in lung inflammatory response as well as genes associated with PAH metabolization and detoxification were highly expressed in bitumen-exposed animals. In addition, the expression of genes related to elastase activity and its inhibition which are associated with emphysema was also modulated. More interestingly genes coding for monoamine oxidases A and B involved in the metabolism of neurotransmitters and xenobiotics were downregulated in exposed rats. Altogether, these data give additional information concerning the bitumen fumes biological effects and would allow to better review the health effects of occupational asphalt fumes exposure.

  6. Bitumen fume-induced gene expression profile in rat lung.

    PubMed

    Gate, Laurent; Langlais, Cristina; Micillino, Jean-Claude; Nunge, Hervé; Bottin, Marie-Claire; Wrobel, Richard; Binet, Stéphane

    2006-08-15

    Exposure to bitumen fumes during paving and roofing activities may represent an occupational health risk. To date, most of the studies performed on the biological effect of asphalt fumes have been done with regard to their content in carcinogenic polycyclic aromatic hydrocarbons (PAH). In order to gain an additional insight into the mechanisms of action of bitumen fumes, we studied their pulmonary effects in rodents following inhalation using the microarray technology. Fisher 344 rats were exposed for 5 days, 6 h/day to bitumen fumes generated at road paving temperature (170 degrees C) using a nose-only exposition device. With the intention of studying the early transcriptional events induced by asphalt fumes, lung tissues were collected immediately following exposure and gene expression profiles in control and exposed rats were determined by using oligonucleotide microarrays. Data analysis revealed that genes involved in lung inflammatory response as well as genes associated with PAH metabolization and detoxification were highly expressed in bitumen-exposed animals. In addition, the expression of genes related to elastase activity and its inhibition which are associated with emphysema was also modulated. More interestingly genes coding for monoamine oxidases A and B involved in the metabolism of neurotransmitters and xenobiotics were downregulated in exposed rats. Altogether, these data give additional information concerning the bitumen fumes biological effects and would allow to better review the health effects of occupational asphalt fumes exposure.

  7. MicroRNA (miRNA) expression is regulated by butyrate induced epigenetic modulation of gene expression in bovine cells

    Technology Transfer Automated Retrieval System (TEKTRAN)

    We present evidence that butyrate induced histone acetylation regulates miRNA expression. MicroRNA expression microarray profiling revealed that 35 miRNA transcripts are significantly (p <0.05) differentially expressed after cells were treated with 10 mM butyrate. Among them, 11 transcripts are dif...

  8. Verproside inhibits TNF-α-induced MUC5AC expression through suppression of the TNF-α/NF-κB pathway in human airway epithelial cells.

    PubMed

    Lee, Su Ui; Sung, Min Hee; Ryu, Hyung Won; Lee, Jinhyuk; Kim, Hui-Seong; In, Hyun Ju; Ahn, Kyung-Seop; Lee, Hyun-Jun; Lee, Hyeong-Kyu; Shin, Dae-Hee; Lee, Yongnam; Hong, Sung-Tae; Oh, Sei-Ryang

    2016-01-01

    Airway mucus secretion is an essential innate immune response for host protection. However, overproduction and hypersecretion of mucus, mainly composed of MUC5AC, are significant risk factors in asthma and chronic obstructive pulmonary disease (COPD) patients. Previously, we reported that verproside, a catalpol derivative iridoid glycoside isolated from Pseudolysimachion rotundum var. subintegrum, is a potent anti-asthmatic candidate drug in vivo. However, the molecular mechanisms underlying the pharmacological actions of verproside remain unknown. Here, we found that verproside significantly reduces the expression levels of tumor necrosis factor alpha (TNF-α)-induced MUC5AC mRNA and protein by inhibiting both nuclear factor kappa B (NF-κB) transcriptional activity and the phosphorylation of its upstream effectors such as IκB kinase (IKK)β, IκBα, and TGF-β-activated kinase 1 (TAK1) in NCI-H292 cells. Moreover, verproside attenuated TNF-α-induced MUC5AC transcription more effectively when combined with an IKK (BAY11-7082) or a TAK1 (5z-7-oxozeaenol) inhibitor than when administered alone. Importantly, we demonstrated that verproside negatively modulates the formation of the TNF-α-receptor (TNFR) 1 signaling complex [TNF-RSC; TNFR1-recruited TNFR1-associated death domain protein (TRADD), TNFR-associated factor 2 (TRAF2), receptor-interacting protein kinase 1 (RIP1), and TAK1], the most upstream signaling factor of NF-κB signaling. In silico molecular docking studies show that verproside binds between TRADD and TRAF2 subunits. Altogether, these results suggest that verproside could be a good therapeutic candidate for treatment of inflammatory airway diseases such as asthma and COPD by blocking the TNF-α/NF-κB signaling pathway.

  9. UV radiation induces CXCL5 expression in human skin.

    PubMed

    Reichert, Olga; Kolbe, Ludger; Terstegen, Lara; Staeb, Franz; Wenck, Horst; Schmelz, Martin; Genth, Harald; Kaever, Volkhard; Roggenkamp, Dennis; Neufang, Gitta

    2015-04-01

    CXCL5 has recently been identified as a mediator of UVB-induced pain in rodents. To compare and to extend previous knowledge of cutaneous CXCL5 regulation, we performed a comprehensive study on the effects of UV radiation on CXCL5 regulation in human skin. Our results show a dose-dependent increase in CXCL5 protein in human skin after UV radiation. CXCL5 can be released by different cell types in the skin. We presumed that, in addition to immune cells, non-immune skin cells also contribute to UV-induced increase in CXCL5 protein. Analysis of monocultured dermal fibroblasts and keratinocytes revealed that only fibroblasts but not keratinocytes displayed up regulated CXCL5 levels after UV stimulation. Whereas UV treatment of human skin equivalents, induced epidermal CXCL5 mRNA and protein expression. Up regulation of epidermal CXCL5 was independent of keratinocyte differentiation and keratinocyte-keratinocyte interactions in epidermal layers. Our findings provide first evidence on the release of CXCL5 in UV-radiated human skin and the essential role of fibroblast-keratinocyte interaction in the regulation of epidermal CXCL5.

  10. Chemically inducible expression of the PHB biosynthetic pathway in Arabidopsis.

    PubMed

    Kourtz, Lauralynn; Dillon, Kevin; Daughtry, Sean; Peoples, Oliver P; Snell, Kristi D

    2007-12-01

    Arabidopsis plants were transformed with a multi-gene construct for expression of the polyhydroxybutyrate (PHB) biosynthetic pathway containing a gene switch that can be activated by commercially available non-steroidal ecdysone analogs approved for use on some crops as pesticides. T(1) progeny of transgenic Arabidopsis plants were isolated and screened for PHB production in the presence of ecdysone analogs. T(2) progeny derived from selected T(1) lines were subjected to further analysis by comparing PHB production levels prior to treatment with inducing agent and 21 days after initiation of induction. Significant PHB production was delayed in many of the engineered plants until after induction. PHB levels of up to 14.3% PHB per unit dry weight were observed in young leaves harvested from engineered T(2) plants after applications of the commercial ecdysone analog Mimic. PHB in older leaves reached levels of up to 7% PHB per unit dry weight. This study represents a first step towards engineering a chemically inducible gene switch for PHB production in plants using inducing agents that are approved for field use.

  11. Transforming growth factor alpha and epidermal growth factor levels in bladder cancer and their relationship to epidermal growth factor receptor.

    PubMed Central

    Mellon, J. K.; Cook, S.; Chambers, P.; Neal, D. E.

    1996-01-01

    We have examined levels of epidermal growth factor (EGF) and transforming growth factor alpha (TGF-alpha) in neoplastic and non-neoplastic bladder tissue using a standard radioimmunoassay technique. Tumour samples had much higher TGF-alpha levels compared with EGF and TGF-alpha levels in malignant tissue were significantly higher than in benign bladder samples. There was, in addition, a difference in mean EGF levels from 'normal' bladder samples from non-tumour bearing areas of bladder in patients with bladder cancer compared with 'normal' bladder tissue obtained at the time of organ retrieval surgery. Levels of EGF and TGF-alpha did not correlate with levels of EGF receptor (EGFR) as determined by a radioligand binding method but levels of TGF-alpha > 10 ng gm-1 of tumour tissue did correlate with EGFR positivity defined using immunohistochemistry. These data suggest that TGF-alpha is the likely ligand for EGFR in bladder tumours. PMID:8605103

  12. Expression of Inflammatory and Cell Death Program Genes and Comet DNA Damage Assay Induced by Escherichia coli in Layer Hens

    PubMed Central

    Mehaisen, Gamal M. K.; Eshak, Mariam G.; El Sabry, M. I.; Abass, Ahmed O.

    2016-01-01

    Modern methods of industrial poultry and egg production systems involve stressful practices that stimulate Escherichia coli (E. coli) activity causing endotoxic shock. This investigation was conducted to evaluate the expression of pro-inflammatory cytokines and cell death program genes and DNA damage induced by E. coli in the brain and liver tissues of laying hens. A total of two hundred and ten H&N brown layer hens with 20 week age, were used in this research. First, preliminary experiments were designed (60 hens in total) to establish the optimal exposure dose of E. coli and to determine the nearest time of notable response to be used in the remainder studies of this research. At 35-wk of age, 150 hens were randomly assigned into 2 groups with 3 replicates of 25 birds each; the first group was injected in the brachial wing vein with 107 E. coli colony/hen, while the second group was injected with saline and served as a control. The body temperature and plasma corticosterone concentration were measured 3 hr after injection. Specimens of liver and brain were obtained from each group and the gene expression of p38 mitogen-activated protein kinase, interlukin-1β (IL-1β), tumor necrosis factor alpha (TNF-α), Bax, and caspase-3 genes were measured by quantitative real-time PCR. DNA damage in the brain and liver tissues were also measured by comet assay. Hens treated with E. coli showed significant (P<0.05) increase of body temperature and plasma corticosterone (42.6°C and 14.5 ng/ml, respectively) compared to the control group (41.1°C and 5.5 ng/ml, respectively). Additional remarkable over-inflammation gene expression of p38, IL-1β and TNF-α.genes were also detected in the brain (2.2-fold, 2.0-fold and 3.3-fold, respectively) and the liver (2.1-fold, 1.9-fold and 3.0-fold, respectively) tissues of the infected chickens. It is also important to note that hens injected with E. coli showed an increase in DNA damage in the brain and liver cells (P<0.05). These

  13. Major Outer Membrane Protein Omp25 of Brucella suis Is Involved in Inhibition of Tumor Necrosis Factor Alpha Production during Infection of Human Macrophages

    PubMed Central

    Jubier-Maurin, Véronique; Boigegrain, Rose-Anne; Cloeckaert, Axel; Gross, Antoine; Alvarez-Martinez, Maria-Teresa; Terraza, Annie; Liautard, Janny; Köhler, Stephan; Rouot, Bruno; Dornand, Jacques; Liautard, Jean Pierre

    2001-01-01

    Brucella spp. can establish themselves and cause disease in humans and animals. The mechanisms by which Brucella spp. evade the antibacterial defenses of their host, however, remain largely unknown. We have previously reported that live brucellae failed to induce tumor necrosis factor alpha (TNF-α) production upon human macrophage infection. This inhibition is associated with a nonidentified protein that is released into culture medium. Outer membrane proteins (OMPs) of gram-negative bacteria have been shown to modulate macrophage functions, including cytokine production. Thus, we have analyzed the effects of two major OMPs (Omp25 and Omp31) of Brucella suis 1330 (wild-type [WT] B. suis) on TNF-α production. For this purpose, omp25 and omp31 null mutants of B. suis (Δomp25 B. suis and Δomp31 B. suis, respectively) were constructed and analyzed for the ability to activate human macrophages to secrete TNF-α. We showed that, in contrast to WT B. suis or Δomp31 B. suis, Δomp25 B. suis induced TNF-α production when phagocytosed by human macrophages. The complementation of Δomp25 B. suis with WT omp25 (Δomp25-omp25 B. suis mutant) significantly reversed this effect: Δomp25-omp25 B. suis-infected macrophages secreted significantly less TNF-α than did macrophages infected with the Δomp25 B. suis mutant. Furthermore, pretreatment of WT B. suis with an anti-Omp25 monoclonal antibody directed against an epitope exposed at the surface of the bacteria resulted in substancial TNF-α production during macrophage infection. These observations demonstrated that Omp25 of B. suis is involved in the negative regulation of TNF-α production upon infection of human macrophages. PMID:11447156

  14. Tumor necrosis factor alpha activates transcription of the NADPH oxidase organizer 1 (NOXO1) gene and upregulates superoxide production in colon epithelial cells.

    PubMed

    Kuwano, Yuki; Tominaga, Kumiko; Kawahara, Tsukasa; Sasaki, Hidekazu; Takeo, Keiko; Nishida, Kensei; Masuda, Kiyoshi; Kawai, Tomoko; Teshima-Kondo, Shigetada; Rokutan, Kazuhito

    2008-12-15

    NADPH oxidase 1 (Nox1) is a multicomponent enzyme consisting of p22(phox), Nox organizer 1 (NOXO1), Nox1 activator 1, and Rac1. Interleukin-1beta, flagellin, interferon-gamma, and tumor necrosis factor alpha (TNF-alpha) similarly induced Nox1 in a colon cancer cell line (T84), whereas only TNF-alpha fully induced NOXO1 and upregulated superoxide-producing activity by ninefold. This upregulation was canceled by knockdown of NOXO1 with small interfering RNAs. TNF-alpha rapidly phosphorylated p38 mitogen-activated protein kinase and c-Jun N-terminal kinase 1/2, followed by phosphorylation of c-Jun and c-Fos and appearance of an AP-1 binding activity within 30 min. We cloned the 5' flank of the human NOXO1 gene (-3888 to +263 bp), and found that the region between -585 and -452 bp, which contains consensus elements of YY-1, AP-1, and Ets, and the GC-rich region encoding three putative binding sites for SP-1, was crucial for TNF-alpha-dependent promoter activity. Serial mutation analysis of the elements identified an AP-1 binding site (from -561 to -551 bp, agtAAGtcatg) as a crucial element for TNF-alpha-stimulated transcription of the human NOXO1 gene, which was also confirmed by the AP-1 decoy experiments. Thus, TNF-alpha acts as a potent activator of Nox1-based oxidase in colon epithelial cells, suggesting a potential role of this oxidase in inflammation of the colon.

  15. Tumor necrosis factor-alpha during neonatal brain development affects anxiety- and depression-related behaviors in adult male and female mice.

    PubMed

    Babri, Shirin; Doosti, Mohammad-Hossein; Salari, Ali-Akbar

    2014-03-15

    A nascent literature suggests that neonatal infection is a risk factor for the development of brain, behavior and hypothalamic-pituitary-adrenal axis which can affect anxiety- and depression-related behaviors in later life. It has been documented that neonatal infection raises the concentrations of tumor necrosis factor-alpha (TNF-α) in neonate rodents and such infections may result in neonatal brain injury, at least in part, through pro-inflammatory cytokines. In addition, previous studies have shown that TNF-α is involved in cellular differentiation, neurogenesis and programmed cell death during the development of the central nervous system. We investigated for the first time whether neonatal exposure to TNF-α can affect body weight, stress-induced corticosterone (COR), anxiety- and depression-related behaviors in adult mice. In the present study, neonatal mice were treated to recombinant mouse TNF-α (0.2, 0.4, 0.7 and 1 μg/kg) or saline on postnatal days 3 and 5, then adult male and female mice were exposed to different behavioral tests. The results indicated that neonatal TNF-α treatment reduced body weight in neonatal period in both sexes. In addition, this study presents findings indicating that high doses of TNF- increase stress-induced COR levels, anxiety- and depression-related behaviors in adult males, but increase levels of anxiety without significantly influencing depression in adult female mice [corrected]. Our findings suggest that TNF-α exposure during neonatal period can alter brain and behavior development in a dose and sex-dependent manner in mice.

  16. Infection of peripheral blood mononuclear cells by herpes simplex and Epstein-Barr viruses. Differential induction of interleukin 6 and tumor necrosis factor-alpha.

    PubMed Central

    Gosselin, J; Flamand, L; D'Addario, M; Hiscott, J; Menezes, J

    1992-01-01

    Infection by herpesviruses can result in profound immunosuppressive or immunomodulatory effects. However, no significant information is available on the effect of such infections on the production of immunoregulatory cytokines. We studied the kinetics of production of two monocyte-derived cytokines, interleukin 6 (IL-6) and tumor necrosis factor-alpha (TNF alpha), induced by Epstein-Barr virus (EBV) and herpes simplex virus type 1 (HSV-1) in peripheral blood mononuclear cell cultures and in fractionated cell populations. We observed that, when compared to HSV-1, EBV is a stronger inducer of IL-6. In EBV-infected cultures, IL-6 protein was detected at day 1 postinfection and gradually increased with time. In contrast, lower amounts of IL-6 were detected 5 d postinfection in HSV-1-infected cultures. HSV-1-infected cultures secreted significant amounts of TNF alpha protein after 5 d of culture and reached a maximal level of production at day 7, whereas EBV inhibited TNF alpha production. In fractionated cell populations, monocytic cells were found to be the main source of IL-6 synthesis after EBV or HSV-1 infection. However, TNF alpha synthesis in HSV-1-infected cultures was from both B and monocytic cells. By using the polymerase chain reaction technique we show that, after infection by these two herpesviruses, differences in cytokine gene products are also observed at the transcriptional level. These observations demonstrate that EBV and HSV-1 exert differential effects on IL-6 and TNF alpha gene transcription and on the resulting protein secretion in human mononuclear blood cells. Images PMID:1318324

  17. Inducible Expression of CXCL1 within the Central Nervous System Amplifies Viral-Induced Demyelination

    PubMed Central

    Marro, Brett S.; Grist, Jonathan J.

    2016-01-01

    The functional role of the ELR+ chemokine CXCL1 in host defense and disease following infection of the CNS with the neurotropic JHM strain of mouse hepatitis virus (JHMV) was examined. Mice in which expression of CXCL1 is under the control of a tetracycline-inducible promoter active within glial fibrillary acidic protein–positive cells were generated and this allowed for selectively increasing CNS expression of CXCL1 in response to JHMV infection and evaluating the effects on neuroinflammation, control of viral replication, and demyelination. Inducible expression of CNS-derived CXCL1 resulted in increased levels of CXCL1 protein within the serum, brain, and spinal cord that correlated with increased frequency of Ly6G+CD11b+ neutrophils present within the CNS. Elevated levels of CXCL1 did not influence the generation of virus-specific T cells, and there was no difference in control of JHMV replication compared with control mice, indicating that T cell infiltration into the CNS is CXCL1-independent. Sustained CXCL1 expression within the CNS resulted in increased mortality that correlated with elevated neutrophil infiltration, diminished numbers of mature oligodendrocytes, and an increase in the severity of demyelination. Neutrophil ablation in CXCL1-transgenic mice reduced the severity of demyelination in mice, arguing for a role for these cells in white matter damage. Collectively, these findings illustrate that sustained CXCL1 expression amplifies the severity of white matter damage and that neutrophils can contribute to this process in a model of viral-induced neurologic disease. PMID:26773148

  18. Personalized medicine: theranostics (therapeutics diagnostics) essential for rational use of tumor necrosis factor-alpha antagonists.

    PubMed

    Bendtzen, Klaus

    2013-04-01

    With the discovery of the central pathogenic role of tumor necrosis factor (TNF)-alpha in many immunoinflammatory diseases, specific inhibition of this pleiotropic cytokine has revolutionized the treatment of patients with several non-infectious inflammatory disorders. As a result, genetically engineered anti-TNF-alpha antibody constructs now constitute one of the heaviest medicinal expenditures in many countries. All currently used TNF antagonists may dramatically lower disease activity and, in some patients, induce remission. Unfortunately, however, not all patients respond favorably, and safety can be severely impaired by immunogenicity, i.e., the ability of a drug to induce anti-drug antibodies (ADA). Assessment of ADA is therefore an important component of the evaluation of drug safety in both pre-clinical and clinical studies and in the process of developing less immunogenic and safer biopharmaceuticals. Therapeutics diagnostics, also called theranostics, i.e., monitoring functional drug levels and neutralizing ADA in the circulation, is central to more effective use of biopharmaceuticals. Hence, testing-based strategies rather than empirical dose-escalation may provide more cost-effective use of TNF antagonists as this allows therapies tailored according to individual requirements rather than the current universal approach to diagnosis. The objective of the present review is to discuss the reasons for recommending theranostics to implement an individualized use of TNF antagonists and to highlight some of the methodological obstacles that have obscured cost-effective ways of using these therapies.

  19. Oxalomalate affects the inducible nitric oxide synthase expression and activity.

    PubMed

    Irace, Carlo; Esposito, Giuseppe; Maffettone, Carmen; Rossi, Antonietta; Festa, Michela; Iuvone, Teresa; Santamaria, Rita; Sautebin, Lidia; Carnuccio, Rosa; Colonna, Alfredo

    2007-03-13

    Inducible nitric oxide synthase (iNOS) is an homodimeric enzyme which produces large amounts of nitric oxide (NO) in response to inflammatory stimuli. Several factors affect the synthesis and catalytic activity of iNOS. Particularly, dimerization of NOS monomers is promoted by heme, whereas an intracellular depletion of heme and/or L-arginine considerably decreases NOS resistance to proteolysis. In this study, we found that oxalomalate (OMA, oxalomalic acid, alpha-hydroxy-beta-oxalosuccinic acid), an inhibitor of both aconitase and NADP-dependent isocitrate dehydrogenase, inhibited nitrite production and iNOS protein expression in lipopolysaccharide (LPS)-activated J774 macrophages, without affecting iNOS mRNA content. Furthermore, injection of OMA precursors to LPS-stimulated rats also decreased nitrite production and iNOS expression in isolated peritoneal macrophages. Interestingly, alpha-ketoglutarate or succinyl-CoA administration reversed OMA effect on NO production, thus correlating NO biosynthesis with the anabolic capacity of Krebs cycle. When protein synthesis was blocked by cycloheximide in LPS-activated J774 cells treated with OMA, iNOS protein levels, evaluated by Western blot analysis and (35)S-metabolic labelling, were decreased, suggesting that OMA reduces iNOS biosynthesis and induces an increase in the degradation rate of iNOS protein. Moreover, we showed that OMA inhibits the activity of the iNOS from lung of LPS-treated rats by enzymatic assay. Our results, demonstrating that OMA acts regulating synthesis, catalytic activity and degradation of iNOS, suggest that this compound might have a potential role in reducing the NO overproduction occurring in some pathological conditions.

  20. Development of a mouse-feline chimeric antibody against feline tumor necrosis factor-alpha

    PubMed Central

    DOKI, Tomoyoshi; TAKANO, Tomomi; HOHDATSU, Tsutomu

    2016-01-01

    Feline infectious peritonitis (FIP) is a fatal inflammatory disease caused by FIP virus infection. Feline tumor necrosis factor (fTNF)-alpha is closely involved in the aggravation of FIP pathology. We previously described the preparation of neutralizing mouse anti-fTNF-alpha monoclonal antibody (mAb 2–4) and clarified its role in the clinical condition of cats with FIP using in vitro systems. However, administration of mouse mAb 2–4 to cat may lead to a production of feline anti-mouse antibodies. In the present study, we prepared a mouse-feline chimeric mAb (chimeric mAb 2–4) by fusing the variable region of mouse mAb 2–4 to the constant region of feline antibody. The chimeric mAb 2–4 was confirmed to have fTNF-alpha neutralization activity. Purified mouse mAb 2–4 and chimeric mAb 2–4 were repeatedly administered to cats, and the changes in the ability to induce feline anti-mouse antibody response were investigated. In the serum of cats treated with mouse mAb 2–4, feline anti-mouse antibody production was induced, and the fTNF-alpha neutralization effect of mouse mAb 2–4 was reduced. In contrast, in cats treated with chimeric mAb 2–4, the feline anti-mouse antibody response was decreased compared to that of mouse mAb 2–4-treated cats. PMID:27264736

  1. Chemical memory reactions induced bursting dynamics in gene expression.

    PubMed

    Tian, Tianhai

    2013-01-01

    Memory is a ubiquitous phenomenon in biological systems in which the present system state is not entirely determined by the current conditions but also depends on the time evolutionary path of the system. Specifically, many memorial phenomena are characterized by chemical memory reactions that may fire under particular system conditions. These conditional chemical reactions contradict to the extant stochastic approaches for modeling chemical kinetics and have increasingly posed significant challenges to mathematical modeling and computer simulation. To tackle the challenge, I proposed a novel theory consisting of the memory chemical master equations and memory stochastic simulation algorithm. A stochastic model for single-gene expression was proposed to illustrate the key function of memory reactions in inducing bursting dynamics of gene expression that has been observed in experiments recently. The importance of memory reactions has been further validated by the stochastic model of the p53-MDM2 core module. Simulations showed that memory reactions is a major mechanism for realizing both sustained oscillations of p53 protein numbers in single cells and damped oscillations over a population of cells. These successful applications of the memory modeling framework suggested that this innovative theory is an effective and powerful tool to study memory process and conditional chemical reactions in a wide range of complex biological systems.

  2. Light-induced currents in Xenopus oocytes expressing bovine rhodopsin.

    PubMed Central

    Knox, B E; Khorana, H G; Nasi, E

    1993-01-01

    1. We have investigated the functioning of bovine rod opsin, which is efficiently synthesized from RNA made by in vitro transcription, following injection into Xenopus oocytes. We found that oocytes expressing the gene for opsin exhibit light-dependent ionic currents only after pigment generation by incubation with 11-cis-retinal. These currents are similar to the endogenous muscarinic acetylcholine (ACh) response of oocytes, but their amplitude is substantially smaller. 2. In order to optimize the conditions for obtaining light-induced currents in RNA-injected oocytes, the native ACh response was examined under several conditions. It was found that elevated external calcium markedly enhances the muscarinic response and that these currents have a non-linear dependence on membrane voltage, increasing substantially with depolarization. 3. Using the optimal conditions for evoking the largest ACh responses, (28 mM [Ca2+]o, 0 mV, omission of serum and Hepes from the media), the light-evoked currents obtained in RNA-injected oocytes were remarkably enhanced, and responses to multiple light stimuli could be obtained. 4. The light response appeared to desensitize, even after long periods of recovery and pigment regeneration. By contrast, the ACh responses continued to appear normal. These results suggest that desensitization of photoresponses expressed in Xenopus oocytes involve changes at early stages of the pathway, resulting in a reduced ability of rhodopsin to couple to the endogenous signalling system. Images Fig. 3 PMID:7692039

  3. Tomato leaf spatial expression of stress-induced Asr genes.

    PubMed

    Maskin, Laura; Maldonado, Sara; Iusem, Norberto D

    2008-12-01

    Asr1 and Asr2 are water stress-inducible genes belonging to the Asr gene family, which transcriptionally regulate a sugar transporter gene, at least in grape. Using an in situ RNA hybridization methodology, we determined that, in basal conditions, expression of Asr2 in tomato leaves is detected in the phloem tissue, particularly in companion phloem cells. When plants are exposed to water stress, Asr2 expression is contained in companion cells but expands occasionally to mesophyll cells. In contrast, Asr1 transcript localization seems to be sparse in leaf vascular tissue under both non-stress and stress conditions. The occurrence of Asr transcripts precisely in companion cells is in accordance with the cell type specificity reported for hexose-transporter protein molecules in grape encoded by the only Asr-target gene known to date. The results are discussed in light of the reported scarcity of plasmodesmata between companion cells and the rest of leaf tissue in the family Solanaceae.

  4. Anxious behavior induces elevated hippocampal Cb2 receptor gene expression.

    PubMed

    Robertson, James M; Achua, Justin K; Smith, Justin P; Prince, Melissa A; Staton, Clarissa D; Ronan, Patrick J; Summers, Tangi R; Summers, Cliff H

    2017-04-07

    Anxiety is differentially expressed across a continuum of stressful/fearful intensity, influenced endocannabinoid systems and receptors. The hippocampus plays important roles in the regulation of affective behavior, emotion, and anxiety, as well as memory. Location of Cb1/Cb2 receptor action could be important in determining emotional valence, because while the dorsal hippocampus is involved in spatial memory and cognition, the ventral hippocampus has projections to the PFC, BNST, amygdala, and HPA axis, and is important for emotional responses to stress. During repeated social defeat in a Stress-Alternatives Model arena (SAM; an oval open field with escape portals only large enough for smaller mice), smaller C57BL6/N mice are subject to fear conditioning (tone=CS), and attacked by novel larger aggressive CD1 mice (US) over four daily (5min) trials. Each SAM trial presents an opportunity for escape or submission, with stable behavioral responses established by the second day of interaction. Additional groups had access to a running wheel. Social aggression plus fear conditioning stimulates enhanced Cb2 receptor gene expression in the dorsal CA1, dorsal and ventral dentate gyrus subregions in animals displaying a submissive behavioral phenotype. Escape behavior is associated with reduced Cb2 expression in the dorsal CA1 region, with freezing and escape latency correlated with mRNA levels. Escaping and submitting animals with access to running wheels had increased Cb2 mRNA in dorsal DG/CA1. These results suggest that the Cb2 receptor system is rapidly induced during anxiogenic social interactions plus fear conditioning or exercise; with responses potentially adaptive for coping mechanisms.

  5. Epigenetic regulation of inducible gene expression in the immune system.

    PubMed

    Lim, Pek Siew; Li, Jasmine; Holloway, Adele F; Rao, Sudha

    2013-07-01

    T cells are exquisitely poised to respond rapidly to pathogens and have proved an instructive model for exploring the regulation of inducible genes. Individual genes respond to antigenic stimulation in different ways, and it has become clear that the interplay between transcription factors and the chromatin platform of individual genes governs these responses. Our understanding of the complexity of the chromatin platform and the epigenetic mechanisms that contribute to transcriptional control has expanded dramatically in recent years. These mechanisms include the presence/absence of histone modification marks, which form an epigenetic signature to mark active or inactive genes. These signatures are dynamically added or removed by epigenetic enzymes, comprising an array of histone-modifying enzymes, including the more recently recognized chromatin-associated signalling kinases. In addition, chromatin-remodelling complexes physically alter the chromatin structure to regulate chromatin accessibility to transcriptional regulatory factors. The advent of genome-wide technologies has enabled characterization of the chromatin landscape of T cells in terms of histone occupancy, histone modification patterns and transcription factor association with specific genomic regulatory regions, generating a picture of the T-cell epigenome. Here, we discuss the multi-layered regulation of inducible gene expression in the immune system, focusing on the interplay between transcription factors, and the T-cell epigenome, including the role played by chromatin remodellers and epigenetic enzymes. We will also use IL2, a key inducible cytokine gene in T cells, as an example of how the different layers of epigenetic mechanisms regulate immune responsive genes during T-cell activation.

  6. A high affinity HSF-1 binding site in the 5'-untranslated region of the murine tumor necrosis factor-alpha gene is a transcriptional repressor.

    PubMed

    Singh, Ishwar S; He, Ju-Ren; Calderwood, Stuart; Hasday, Jeffrey D

    2002-02-15

    Tumor necrosis factor-alpha (TNFalpha) is a pivotal early mediator of host defenses that is essential for survival in infections. We previously reported that exposing macrophages to febrile range temperatures (FRT) (38.5-40 degrees C) markedly attenuates TNFalpha expression by causing abrupt and premature cessation of transcription. We showed that this inhibitory effect of FRT is mediated by an alternatively activated repressor form of heat shock factor 1 (HSF-1) and that a fragment of the TNFalpha gene comprising a minimal 85-nucleotide (nt) proximal promoter and the 138-nt 5'-untranslated region (UTR) was sufficient for mediating this effect. In the present study we have used an electrophoretic mobility shift assay (EMSA) to identify a high affinity binding site for HSF-1 in the 5'-UTR of the TNFalpha gene and have used a chromosome immunoprecipitation assay to show that HSF-1 binds to this region of the endogenous TNFalpha gene. Mutational inactivation of this site blocks the inhibitory effect of overexpressed HSF-1 on activity of the minimal TNFalpha promoter (-85/+138) in Raw 264.7 murine macrophages, identifying this site as an HSF-1-dependent repressor. However, the same mutation fails to block repression of a full-length (-1080/+138) TNFalpha promoter construct by HSF-1 overexpression, and HSF-1 binds to upstream sequences in the regions -1080/-845, -533/-196, and -326/-39 nt in EMSA, suggesting that additional HSF-1-dependent repressor elements are present upstream of the minimal -85-nt promoter. Furthermore, although mutation of the HSF-1 binding site in the minimal TNFalpha promoter construct abrogates HSF-1-mediated repression, the same mutation fails to abrogate repression of this construct by high levels of HSF-1 overexpression or exposure to 39.5 degrees C. This suggests that HSF-1 might repress TNFalpha transcription through redundant mechanisms, some of which might not require high affinity binding of HSF-1.

  7. Genetic polymorphisms of tumour necrosis factor alpha (TNF-α) promoter gene and response to TNF-α inhibitors in Spanish patients with inflammatory bowel disease.

    PubMed

    López-Hernández, R; Valdés, M; Campillo, J A; Martínez-Garcia, P; Salama, H; Salgado, G; Boix, F; Moya-Quiles, M R; Minguela, A; Sánchez-Torres, A; Miras, M; Garcia, A; Carballo, F; Álvarez-López, M R; Muro, M

    2014-02-01

    Tumour necrosis factor alpha (TNF-α) has an important role in inflammatory response. Alterations in the regulation of TNF-α have been implicated in a variety of inflammatory disorders, including Inflammatory bowel disease (IBD). Indeed, a common treatment for IBD is the use of TNF-α inhibitors. Polymorphisms in the TNF-α promoter region are known to affect the level of gene expression. Our aim was to investigate the influence of these single nucleotide polymorphisms (SNPs) in TNF-α promoter gene play in the risk of IBD in a Spanish population and their individual response to anti-TNF-α treatment. DNA samples from patients with IBD and controls were screened for TNF-α -238G/A (rs361525) and -308G/A (rs1800629) SNPs by PCR-SSOP using a microbeads luminex assay and compared with response to TNF-α inhibitors. There were not statistical differences in -238G/A and -308G/A allele and genotype frequencies between patients. However, we found an increased frequency of -308A allele and -308GA genotype in these nonresponders patients to TNF-α inhibitors with respect to responders patients (Pc < 0.05). This -308GA genotype has been classified as high producer of this cytokine. This fact could actually be interesting to explain the different response of patients with IBD with respect to TNF-α inhibitors. TNF-α promoter gene polymorphism does not seem to play a role in IBD susceptibility, but particular TNF-α genotypes may be involved in the different responses to TNF-α inhibitor treatment in Spanish patients with IBD.

  8. Tumor necrosis factor alpha/cachectin stimulates eosinophil oxidant production and toxicity towards human endothelium

    PubMed Central

    1990-01-01

    Eosinophils (EOs) participate in a variety of inflammatory states characterized by endothelial cell damage, such as vasculitis, pneumonitis, and endocarditis. We find that 100 U/ml TNF- alpha/cachectin (TNF), a concentration attainable in the blood of humans with parasitic infestations, stimulates highly purified populations of EOs to damage human umbilical vein endothelial cells (HUVEC), a model of human endothelium. This TNF-dependent EO cytotoxicity is strongly inhibited by heparin and methyprednisolone but unaffected by the platelet-activating factor antagonist BN52012 or scavengers of superoxide anion and H2O2, superoxide dismutase and catalase. However, addition of a physiologically relevant concentration of Br- (100 microM) enhances EO/TNF damage to HUVEC, implicating the possible participation of EO peroxidase (EPO) in the killing mechanism. EOs adherent to FCS-coated plastic wells more than double their production of superoxide anion and the cytotoxic EPO-derived oxidant HOBr when exposed to TNF, showing that TNF activates the respiratory burst of EOs attached to a "physiologic" surface. Unlike PMNs, EOs were not irreversibly activated to kill unopsonized endothelium by previous exposure to TNF, and did not degranulate or upregulate CR3 expression as detected by Mo1 in the presence of 100 U/ml TNF. HUVEC exposed 18 h to TNF were considerably more susceptible to lysis by PMA-activated EOs and reagent H2O2, demonstrating a direct effect of TNF upon endothelium, perhaps through inhibition of antioxidant defenses. These findings suggest that abnormally elevated serum levels of TNF may provoke EOs to damage endothelial cells and thereby play a role in the pathogenesis of tissue damage in hypereosinophilic states. PMID:1972179

  9. Tumor necrosis factor alpha/cachectin stimulates eosinophil oxidant production and toxicity towards human endothelium.

    PubMed

    Slungaard, A; Vercellotti, G M; Walker, G; Nelson, R D; Jacob, H S

    1990-06-01

    Eosinophils (EOs) participate in a variety of inflammatory states characterized by endothelial cell damage, such as vasculitis, pneumonitis, and endocarditis. We find that 100 U/ml TNF-alpha/cachectin (TNF), a concentration attainable in the blood of humans with parasitic infestations, stimulates highly purified populations of EOs to damage human umbilical vein endothelial cells (HUVEC), a model of human endothelium. This TNF-dependent EO cytotoxicity is strongly inhibited by heparin and methyprednisolone but unaffected by the platelet-activating factor antagonist BN52012 or scavengers of superoxide anion and H2O2, superoxide dismutase and catalase. However, addition of a physiologically relevant concentration of Br- (100 microM) enhances EO/TNF damage to HUVEC, implicating the possible participation of EO peroxidase (EPO) in the killing mechanism. EOs adherent to FCS-coated plastic wells more than double their production of superoxide anion and the cytotoxic EPO-derived oxidant HOBr when exposed to TNF, showing that TNF activates the respiratory burst of EOs attached to a "physiologic" surface. Unlike PMNs, EOs were not irreversibly activated to kill unopsonized endothelium by previous exposure to TNF, and did not degranulate or upregulate CR3 expression as detected by Mo1 in the presence of 100 U/ml TNF. HUVEC exposed 18 h to TNF were considerably more susceptible to lysis by PMA-activated EOs and reagent H2O2, demonstrating a direct effect of TNF upon endothelium, perhaps through inhibition of antioxidant defenses. These findings suggest that abnormally elevated serum levels of TNF may provoke EOs to damage endothelial cells and thereby play a role in the pathogenesis of tissue damage in hypereosinophilic states.

  10. Prognostic and Therapeutic Values of Tumor Necrosis Factor-Alpha in Hepatocellular Carcinoma

    PubMed Central

    Wang, Hongmei; Liu, Jianmin; Hu, Xuemei; Liu, Shanshan; He, Baojun

    2016-01-01

    Background Hepatocellular carcinoma (HCC) causes many deaths worldwide every year, especially in Asia. It is characterized by high malignancy, recurrence, and short survival time. Inflammation is closely related to the initiation and development of HCC. Tumor necrosis factor-α (TNF-α), an essential inflammatory mediator, has been studied as a potential therapy target in many cancers. However, its potential role in HCC diagnosis and therapy is still unclear. Material/Methods In our study, we detected the TNF-α expression in both human HCC tumor tissue and HCC cell lines HepG2 and HuH7. Then, we detected the effect of anti-TNF-α treatment and it synergistic function with 5-FU in an HCC xenograft mouse model and in HCC cell lines. Results Survival analysis and Cox regression analysis based on 97 HCC patients indicated that a high level of TNF-α is an independent predictor of poor survival in HCC patients. Anti-TNF-α treatment by infliximab synergizes with Fluorouracil (5-FU) by promoting apoptosis of HCC tumor cells through complement-dependent cytotoxicity (CDC) and antibody-dependent cellular cytotoxicity (ADCC) effects. Conclusions Based on these data, we conclude that anti-TNF-α treatment could be a good way to increase the effect of classic chemotherapy of HCC patients, especially for the patients who have modest response to classic chemotherapy, such as 5-FU. TNF-α could also be used as a biomarker to help in early diagnosis of HCC. PMID:27739418

  11. Apolipoprotein A-I inhibits the production of interleukin-1beta and tumor necrosis factor-alpha by blocking contact-mediated activation of monocytes by T lymphocytes.

    PubMed

    Hyka, N; Dayer, J M; Modoux, C; Kohno, T; Edwards, C K; Roux-Lombard, P; Burger, D

    2001-04-15

    Tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta), essential components in the pathogenesis of immunoinflammatory diseases, are strongly induced in monocytes by direct contact with stimulated T lymphocytes. This study demonstrates that adult human serum (HS) but not fetal calf or cord blood serum displays inhibitory activity toward the contact-mediated activation of monocytes by stimulated T cells, decreasing the production of both TNF-alpha and IL-1beta. Fractionation of HS and N-terminal microsequencing as well as electroelution of material subjected to preparative electrophoresis revealed that apolipoprotein A-I (apo A-I), a "negative" acute-phase protein, was the inhibitory factor. Functional assays and flow cytometry analyses show that high-density lipoprotein (HDL)-associated apo A-I inhibits contact-mediated activation of monocytes by binding to stimulated T cells, thus inhibiting TNF-alpha and IL-1beta production at both protein and messenger RNA levels. Furthermore, apo A-I inhibits monocyte inflammatory functions in peripheral blood mononuclear cells activated by either specific antigens or lectins without affecting cell proliferation. These results demonstrate a new anti-inflammatory activity of HDL-associated apo A-I that might have modulating functions in nonseptic conditions. Therefore, because HDL has been shown to bind and neutralize lipopolysaccharide, HDL appears to play an important part in modulating both acute and chronic inflammation. The novel anti-inflammatory function of apo A-I reported here might lead to new therapeutic approaches in inflammatory diseases such as rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, and atherosclerosis.

  12. Differential effects of pentoxifylline on the production of tumour necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) by monocytes and T cells.

    PubMed Central

    Schandené, L; Vandenbussche, P; Crusiaux, A; Alègre, M L; Abramowicz, D; Dupont, E; Content, J; Goldman, M

    1992-01-01

    Pentoxifylline (PTX) is a methylxanthine compound known to inhibit the production of tumour necrosis factor-alpha (TNF-alpha) by monocytic cells. In this study, we found that PTX differentially regulates the production of TNF-alpha and interleukin-6 (IL-6). Indeed, PTX at high concentrations triggers the production of IL-6 but not of TNF-alpha by peripheral blood mononuclear cells (PBMC). Further experiments indicated that monocytes are responsible for this PTX-induced IL-6 production. When PBMC were stimulated with LPS, PTX was found to inhibit the secretion of TNF-alpha as well as the accumulation of TNF-alpha messenger RNA (mRNA). In contrast, no inhibitory effect was observed on the induction of IL-6. Similar results were obtained when PBMC were stimulated with OKT3 monoclonal antibody (mAb). In addition, the in vivo administration of PTX in transplant patients receiving the first dose of OKT3 allowed to decrease the systemic release of TNF-alpha but not of IL-6. Since monocytes represent a major source of TNF-alpha and IL-6 in these settings, additional experiments were performed in vitro on purified T cells stimulated with the CLB-T3/3, an anti-CD3 mAb which does not require the presence of accessory cells to activate T cells. In this system, PTX was found to inhibit the secretion of both TNF-alpha and IL-6 by T cells. We suggest that cAMP could be involved in these differential effects of PTX on production of TNF-alpha and of IL-6. Images Figure 3 PMID:1385797

  13. Inhibition of sup 125 I organification and thyroid hormone release by interleukin-1, tumor necrosis factor-alpha, and interferon-gamma in human thyrocytes in suspension culture

    SciTech Connect

    Sato, K.; Satoh, T.; Shizume, K.; Ozawa, M.; Han, D.C.; Imamura, H.; Tsushima, T.; Demura, H.; Kanaji, Y.; Ito, Y. )

    1990-06-01

    To elucidate the mechanism of decreased 131I uptake by the thyroid gland in patients with subacute thyroiditis and painless thyroiditis, human thyroid follicles were cultured with interleukin-1 (IL-1), tumor necrosis factor-alpha (TNF alpha), and/or interferon-gamma (IFN gamma), and the effects of these cytokines on thyroid function were studied in vitro. When human thyrocytes were cultured in RPMI-1640 medium containing 0.5% fetal calf serum and TSH for 5-8 days, the cells incorporated 125I, synthesized de novo (125I)iodotyrosines and (125I)iodothyronines, and secreted (125I)T4 and (125I)T3 into the medium. IL-1 alpha and IL-1 beta inhibited 125I incorporation and (125I)iodothyronine release in a concentration-dependent manner. The minimal inhibitory effect was detected at 10 pg/ml. Electron microscopic examination revealed a marked decrease in lysosome formation in IL-1-treated thyrocytes. TNF alpha and IFN gamma also inhibited thyroid function in a concentration-dependent manner. Furthermore, when thyrocytes were cultured with IL-1, TNF alpha and IFN gamma, these cytokines more than additively inhibited thyroid function. Although the main mechanism of 131I uptake suppression in the thyroid gland in subacute thyroiditis is due to cellular damage and suppression of TSH release, our present findings suggest that IL-1, TNF alpha, and IFN gamma produced in the inflammatory process within the thyroid gland further inhibit iodine incorporation and at least partly account for the decreased 131I uptake by the thyroid gland in destruction-induced hyperthyroidism.

  14. p38 Mitogen-Activated Protein Kinase Mediates Lipopolysaccharide and Tumor Necrosis Factor Alpha Induction of Shiga Toxin 2 Sensitivity in Human Umbilical Vein Endothelial Cells▿ †

    PubMed Central

    Stone, Matthew K.; Kolling, Glynis L.; Lindner, Matthew H.; Obrig, Tom G.

    2008-01-01

    Escherichia coli O157:H7 Shiga toxin 2 (Stx2), one of the causative agents of hemolytic-uremic syndrome, is toxic to endothelial cells, including primary cultured human umbilical vein endothelial cells (HUVEC). This sensitivity of cells to Stx2 can be increased with either lipopolysaccharide (LPS) or tumor necrosis factor alpha (TNF-α). The goal of the present study was to identify the intracellular signaling pathway(s) by which LPS and TNF-α sensitize HUVEC to the cytotoxic effects of Stx2. To identify these pathways, specific pharmacological inhibitors and small interfering RNAs were tested with cell viability endpoints. A time course and dose response experiment for HUVEC exposure to LPS and TNF-α showed that a relatively short exposure to either agonist was sufficient to sensitize the cells to Stx2 and that both agonists stimulated intracellular signaling pathways within a short time. Cell viability assays indicated that the p38 mitogen-activated protein kinase (MAPK) inhibitors SB202190 and SB203580 and the general protein synthesis inhibitor cycloheximide inhibited both the LPS and TNF-α sensitization of HUVEC to Stx2, while all other inhibitors tested did not inhibit this sensitization. Additionally, SB202190 reduced the cellular globotriaosylceramide content under LPS- and TNF-α-induced conditions. In conclusion, our results show that LPS and TNF-α induction of Stx2 sensitivity in HUVEC is mediated through a pathway that includes p38 MAPK. These results indicate that inhibition of p38 MAPK in endothelial cells may protect a host from the deleterious effects of Stx2. PMID:18086809

  15. Porphyromonas gingivalis Infection during Pregnancy Increases Maternal Tumor Necrosis Factor Alpha, Suppresses Maternal Interleukin-10, and Enhances Fetal Growth Restriction and Resorption in Mice

    PubMed Central

    Lin, Dongming; Smith, Mary Alice; Champagne, Catherine; Elter, John; Beck, James; Offenbacher, Steven

    2003-01-01

    Epidemiological studies have shown a potential association between maternal periodontitis and pregnancy complications. We used a pregnant murine model to study the effect of infection with the periodontal pathogen Porphyromonas gingivalis on pregnancy outcomes. Female BALB/c mice were inoculated with heat-killed P. gingivalis (109 CFU) in a subcutaneous chamber and mated 2 weeks later. At gestation day (GD) 7.5, mice were challenged with live P. gingivalis (107 CFU) (n = 20) or broth (control; n = 8) and sacrificed at GD 16.5. Fetal growth restriction (FGR, <0.46 g) was defined as fetuses with weights 2 standard deviations (SD) smaller than controls (0.56 ± 0.05 g [mean ± SD]). Among the 20 challenged mice, 8 had both normal-weight (0.51 ± 0.11 g) and FGR (0.34 ± 0.1 g) fetuses within the same litter. All other challenged dams had normal-weight fetuses (0.57 ± 0.04 g). Maternal liver, uterus, and spleen samples were examined for P. gingivalis DNA using a PCR technique. Of the eight challenged mice with FGR fetuses, three had PCR signals for P. gingivalis in liver and uterus, but not in the spleen. Liver, uterus, and spleen were negative for P. gingivalis DNA among all other challenged and control mice. In serum of dams with FGR fetuses, tumor necrosis factor alpha levels were elevated significantly, while interluekin-10 levels were significantly reduced compared to levels in dams with normal fetuses. P. gingivalis-specific serum immunoglobulin G levels were significantly elevated in dams with FGR fetuses compared to dams without any FGR fetuses. These data demonstrate that P. gingivalis-induced murine FGR is associated with systemic dissemination of the organism and activated maternal immune and inflammatory responses. PMID:12933859

  16. Combined Intraperitoneal and Intrathecal Etanercept Reduce Increased Brain Tumor Necrosis Factor-Alpha and Asymmetric Dimethylarginine Levels and Rescues Spatial Deficits in Young Rats after Bile Duct Ligation

    PubMed Central

    Sheen, Jiunn-Ming; Chen, Yu-Chieh; Hsu, Mei-Hsin; Tain, You-Lin; Yu, Hong-Ren; Huang, Li-Tung

    2016-01-01

    Background: Rats subjected to bile duct ligation (BDL) exhibit increased systemic oxidative stress and brain dysfunction characteristic of hepatic encephalopathy (HE), including fatigue, neurotransmitter alterations, cognitive and motor impairment, and brain inflammation. The levels of tumor necrosis factor-alpha (TNF-α) and asymmetric dimethylarginine (ADMA) are both increased in plasma and brain in encephalopathy induced by chronic liver failure. This study first determined the temporal profiles of TNF-α and ADMA in the plasma, brain cortex, and hippocampus in young BDL rats. Next, we examined whether etanercept was beneficial in preventing brain damage. Methods: Young rats underwent sham ligation or BDL at day 17 ± 1 for 4 weeks. Treatment group rats were administered etanercept (10 mg/kg) intraperitoneally (IP) three times per week with or without etanercept (100 μg) intrathecally (IT) three times in total. Results: We found increased plasma TNF-α, soluble tumor necrosis factor receptor 1 (sTNFR1), soluble tumor necrosis factor receptor 2 (sTNFR2), and ADMA levels, increased cortical TNF-α mRNA and protein and ADMA, and hippocampal TNF-α mRNA and protein, and spatial defects in young BDL rats. The increase in cortex TNF-α mRNA and ADMA were reduced by IP etanercept or combined IP and IT etanercept. Dually IP/IT etanercept administration reduced the increased cortical and hippocampal TNF-α mRNA and protein level as well as spatial deficits. Conclusions: We conclude that combined intraperitoneal and intrathecal etanercept reduce increased brain TNF-α and ADMA levels and rescues spatial deficits in young rats after BDL. PMID:27445694

  17. Antitumor activity of tumor necrosis factor-alpha conjugated with polyvinylpyrrolidone on solid tumors in mice.

    PubMed

    Kamada, H; Tsutsumi, Y; Yamamoto, Y; Kihira, T; Kaneda, Y; Mu, Y; Kodaira, H; Tsunoda, S I; Nakagawa, S; Mayumi, T

    2000-11-15

    We attempted the development of a novel polymer conjugation to further improve the therapeutic potency of antitumor cytokines compared with PEGylation for clinical application. Compared with native tumor necrosis factor (TNF)-alpha in vitro, specific bioactivities of polyvinyl-pyrrolidone (PVP)-modified TNF-alphas (PVP-TNF-alphas) were decreased by increasing the degree of PVP attachment. PVP-TNF-alpha fraction 3, Mr 101,000, had the most effective antitumor activity of the various PVP-TNF-alphas in vivo. PVP-TNF-alpha fraction 3 had >200-fold higher antitumor effect than native TNF-alpha, and the antitumor activity of PVP-TNF-alpha fraction 3 was >2-fold higher than that of MPEG-TNF-alpha (Mr 108,000), which had the highest antitumor activity among the polyethylene glycol (PEG)-conjugated TNF-alphas. Additionally, a high dose of native TNF-alpha induced toxic side effects such as body weight reduction, piloerection. and tissue inflammation, whereas no side effects were observed after i.v. administration of PVP-TNF-alpha fraction 3. The plasma half-life of PVP-TNF-alpha fraction 3 (360 min) was about 80- and 3-fold longer than those of native TNF-alpha (4.6 mm) and MPEG-TNF-alpha (122 min), respectively. The mechanism of increased antitumor effect in vivo caused the prolongation of plasma half-life and increase in stability. These results suggested that PVP is a useful polymeric modifier for bioconjugation of TNF-alpha to increase its antitumor potency, and multifunctionally bioconjugated TNF-alpha may be a potentiated antitumor agent for clinical use.

  18. Pituitary gene expression differs in D-galactose-induced cell senescence and steroid-induced prolactinomas.

    PubMed

    Zhang, Tiehui; Zhao, Binhai; Li, Jia; Zhang, Chunlei; Li, Hongzhi; Wu, Jiang; Zhang, Shiming; Hui, Guozhen

    2015-04-01

    In general, pituitary tumors are benign with low mitotic activity. Premature senescence has been considered to be a significant mechanism underlying this uniquely benign pituitary tumor. The present study aims to compare the expression of the associated proteins involved in premature senescence pathways among normal, aging and pituitary adenoma cells. We successfully induced the aging pituitary using continuous D‑galactose (D‑gal) injection as well as a prolactin‑secreting pituitary tumor via diethylstilbestrol implants. Compared with normal pituitary cells, the aging pituitary tissues rev