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Expression of S100A6 in cardiac myocytes limits apoptosis induced by tumor necrosis factor-alpha.  


S100A6 is induced in myocardium post-infarction in vivo and in response to growth factors and inflammatory cytokines in vitro. Forced expression of S100A6 in cardiomyocytes inhibits regulation of cardiac specific gene expression in response to trophic stimulation. To define regulation and function of S100A6, we characterized the human S100A6 promoter and mapped upstream regulatory elements in rat neonatal cardiac myocytes, fibroblasts, and vascular smooth muscle cells and defined a functional role for S100A6 in tumor necrosis factor-alpha-induced myocyte apoptosis. The functional S100A6 promoter was localized to region -167/+134 containing 167 upstream base pairs. The S100A6 promoter is regulated by positive (-361/-167 and -588/-361) and negative (-1371/-1194) elements. Tumor necrosis factor-alpha induced the maximal S100A6 promoter and transcription factor NF-kappaB (p65 subunit). Electrophoretic mobility shift showed that tumor necrosis factor-alpha induced p65 binding to a potential NF-kappaB-binding site at -460/-451. Chromatin immunoprecipitation analysis revealed p65 is recruited to the S100A6 promoter upon tumor necrosis factor-alpha stimulation. The NF-kappaB inhibitor caffeic acid phenethyl ester and mutation of the NF-kappaB-binding site inhibited S100A6 promoter activation by tumor necrosis factor-alpha. Tumor necrosis factor-alpha induced cardiac myocyte apoptosis. Specific inhibition of S100A6 using a small interfering RNA directed against S100A6 potentiated tumor necrosis factor-alpha-induced myocyte apoptosis, whereas overexpression of S100A6 by gene transfer prevented tumor necrosis factor-alpha-induced myocyte apoptosis by interfering with p53 phosphorylation. These results demonstrate that S100A6 is induced by tumor necrosis factor-alpha via an NF-kappaB-dependent mechanism, serving a role in homeostasis to limit tumor necrosis factor-alpha-induced apoptosis by regulating p53 phosphorylation. PMID:18753141

Tsoporis, James N; Izhar, Shehla; Parker, Thomas G



Arsenite enhances tumor necrosis factor-{alpha}-induced expression of vascular cell adhesion molecule-1  

SciTech Connect

Epidemiological studies demonstrated a high association of vascular diseases with arsenite exposure. We hypothesize that arsenite potentiates the effect of proinflammatory cytokines on vascular endothelial cells, and hence contributes to atherosclerosis. In this study, we investigated the effect of arsenite and its induction of glutathione (GSH) on vascular cell adhesion molecule-1 (VCAM-1) protein expression in human umbilical vein endothelial cells (HUVECs) in response to tumor necrosis factor-{alpha} (TNF-{alpha}), a typical proinflammatory cytokine. Our study demonstrated that arsenite pretreatment potentiated the TNF-{alpha}-induced VCAM-1 expression with up-regulations of both activator protein-1 (AP-1) and nuclear factor-{kappa}B (NF-{kappa}B). To elucidate the role of GSH in regulation of AP-1, NF-{kappa}B, and VCAM-1 expression, we employed L-buthionine (S,R)-sulfoximine (BSO), a specific {gamma}-glutamylcysteine synthetase ({gamma}-GCS) inhibitor, to block intracellular GSH synthesis. Our investigation revealed that, by depleting GSH, arsenite attenuated the TNF-{alpha}-induced VCAM-1 expression as well as a potentiation of AP-1 and an attenuation of NF-{kappa}B activations by TNF-{alpha}. Moreover, we found that depletion of GSH would also attenuate the TNF-{alpha}-induced VCAM-1 expression with a down-regulation of the TNF-{alpha}-induced NF-{kappa}B activation and without significant effect on AP-1. On the other hand, the TNF-{alpha}-induced VCAM-1 expression could be completely abolished by inhibition of AP-1 or NF-{kappa}B activity, suggesting that activation of both AP-1 and NF-{kappa}B was necessary for VCAM-1 expression. In summary, we demonstrate that arsenite enhances the TNF-{alpha}-induced VCAM-1 expression in HUVECs via regulation of AP-1 and NF-{kappa}B activities in a GSH-sensitive manner. Our present study suggested a potential mechanism for arsenite in the induction of vascular inflammation and vascular diseases via modulating the actions of proinflammatory cytokines.

Tsou, T.-C. [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, 100 Shih-Chuan 1st Road, Kaohsiung 807, Taiwan (China)]. E-mail:; Yeh, Szu Ching [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, 100 Shih-Chuan 1st Road, Kaohsiung 807, Taiwan (China); Tsai, E.-M. [Department of Obstetrics and Gynecology, College of Medicine, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Department of Obstetrics and Gynecology, Kaohsiung Medical University Chung-Ho Memorial Hospital, Kaohsiung, Taiwan (China); Tsai, F.-Y. [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, 100 Shih-Chuan 1st Road, Kaohsiung 807, Taiwan (China); Chao, H.-R. [Department of Environmental and Safety Health Engineering, Chung Hwa College of Medical Technology, Tainan, Taiwan (China); Chang, Louis W. [Division of Environmental Health and Occupational Medicine, National Health Research Institutes, 100 Shih-Chuan 1st Road, Kaohsiung 807, Taiwan (China)



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


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-?) 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-? expression was evaluated. Both TNF-? 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-? promoter. In the presence of NEP the activity of TNF-? 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-? 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-? 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-? 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-? expression. PMID:24657783

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



Tumor necrosis factor alpha (TNF-?) autoregulates its expression and induces adhesion molecule expression in asthma.  


Subjects with mild asthma underwent repeated low-dose allergen exposure and bronchial biopsies were examined for the expression of TNF-? and adhesion molecules. Bronchial biopsies from moderately severe asthmatics were then tested in an explant culture system to assess the effect of Der p and CDP-870, a TNF-? blocking pegylated-antibody Fab, on expression of TNF-? and adhesion molecules. Low-dose allergen challenge significantly upregulated sub-mucosal mast cells, TNF-?(+) cells, and VCAM. When bronchial explants were exposed to Der p and CDP 870 for 24h, CDP 870 caused a significant reduction in TNF-? release both at baseline and following stimulation with Der p allergen. The bronchial biopsies showed significant upregulation of TNF-? positive cells and ICAM-1 following exposure to Der p (p=0.03) and this was reduced in the presence of CDP-870. So, allergen exposure up-regulates TNF-? expression in asthma and down-stream targets, including adhesion molecules that contribute to airway inflammation. PMID:21459047

Babu, Suresh K; Puddicombe, Sarah M; Arshad, Hasan H; Wilson, Susan J; Ward, John; Gozzard, Neil; Higgs, Gerry; Holgate, Stephen T; Davies, Donna E



Expression of transforming growth factor alpha and epidermal growth factor receptor in rat lung neoplasms induced by plutonium-239  

SciTech Connect

Ninety-two rat lung proliferative lesions and neoplasms induced by inhaled {sup 239}PuO{sub 2} were evaluated for aberrant expression of transforming growth factor alpha (TGF-{alpha}) and epidermal growth factor receptor (EGFR). Expression of TGF-{alpha} protein, measured by immunohistochemistry, was higher in 94% of the squamous cell carcinomas and 87% of the foci of alveolar epithelial squamous metaplasia than that exhibited by the normal-appearing, adjacent lung parenchyma. In contrast, only 20% of adenocarcinomas and foci of epithelial hyperplasia expressed elevated levels of TGF-{alpha}. Many neoplasms expressing TGF-{alpha} also expressed excessive levels of EGFR mRNA. Southern and DNA slot blot analyses showed that the elevated EGFR expression was not due to amplification of the EGFR gene. These data suggest that increased amounts of TGF-{alpha} were early alterations in the progression of plutonium-induced squamous cell carcinoma, and these increases may occur in parallel with overexpression of the receptor for this growth factor. Together, these alterations create a potential autocrine loop for sustaining clonal expansion of cells initiated by high-LET radiation. 44 refs., 4 figs., 1 tab.

Stegelmeier, B.L.; Gillett, N.A.; Hahn, F.F.; Kelly, G. [Inhalation Toxicology Research Institute, Albuquerque, NM (United States); Rebar, A.H. [Purdue Univ., West Lafayette, IN (United States)



5-Azacytidine treatment of HA-A melanoma cells induces Sp1 activity and concomitant transforming growth factor alpha expression.  

PubMed Central

Evidence indicates DNA methylation as a part of the regulatory machinery controlling mammalian gene expression. The human melanoma cell line HA-A expresses low levels of transforming growth factor alpha (TGF-alpha). TGF-alpha mRNA accumulated, however, in response to DNA demethylation induced by a nucleoside analog, 5-azacytidine (5-azaC). The importance of DNA methylation in the TGF-alpha promoter region was examined by a transient transfection assay with luciferase reporter plasmids containing a portion of the TGF-alpha promoter. 5-azaC treatment of HA-A cells before the transfection caused a significant increase in the luciferase activity. Since input plasmids were confirmed to remain unmethylated, DNA demethylation of the TGF-alpha promoter itself does not account for the observed increase in TGF-alpha mRNA. Using an electrophoretic mobility shift assay, enhanced formation of protein-TGF-alpha promoter complex was detected in response to 5-azaC treatment. This 5-azaC-induced complex was shown to contain the transcription factor Sp1 by the following criteria: the protein-DNA complex formed on the TGF-alpha promoter contained immunoreactive Sp1; the mobility of the complex in an electrophoretic mobility shift assay was similar to that formed by recombinant Sp1; and DNase I footprinting analysis demonstrated that the 5-azaC-induced complex produced a footprint on the TGF-alpha promoter identical to that of authentic Sp1. These observations suggest that 5-azaC induces TGF-alpha expression by augmenting the Sp1 activity. However, neither the Sp1 mRNA nor its protein was induced by 5-azaC. These results suggest that in HA-A cells, TGF-alpha expression is down-modulated by DNA methylation. In addition, this process may involve the specific regulation of Sp1 activity without altering the amount of the transcription factor. Images

Shin, T H; Paterson, A J; Grant, J H; Meluch, A A; Kudlow, J E



Prohibitin Inhibits Tumor Necrosis Factor alpha-induced Nuclear Factor-kappa B Nuclear Translocation via the Novel Mechanism of Decreasing Importin  3 Expression  

Microsoft Academic Search

Expression of prohibitin 1 (PHB), a multifunctional protein in the cell, is decreased during inflammatory bowel disease (IBD). Little is known regarding the regulation and role of PHB during intestinal inflammation. We examined the effect of tumor necrosis factor alpha (TNF-), a cytokine that plays a central role in the pathogenesis of IBD, on PHB expression and the effect of

Arianne L. Theiss; Aaron K. Jenkins; Ngozi I. Okoro; Jan-Michael A. Klapproth; Didier Merlin; Shanthi V. Sitaraman



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

PubMed Central

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

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



Expression of Tumor Necrosis Factor Alpha-Induced Protein 3 mRNA in Peripheral Blood Mononuclear Cells Negatively Correlates with Disease Severity in Psoriasis Vulgaris  

PubMed Central

Psoriasis vulgaris is considered a chronic inflammatory disease, but its immunopathogenesis has not been well understood. The tumor necrosis factor alpha-induced protein 3 (TNFAIP3) gene functions in negative-feedback regulation of inflammation, and its single nucleotide polymorphism is associated with psoriasis. However, the relationship between the expression level of the TNFAIP3 gene in immune cells and psoriasis is not known so far. In the present study, TNFAIP3 mRNA expression levels in peripheral blood mononuclear cells from 44 patients with psoriasis vulgaris and 30 healthy controls were determined using real-time reverse transcription-PCR analysis. We found that expression of TNFAIP3 mRNA in all patients negatively correlated with the psoriatic area and severity index (PASI) (r = ?0.5126; P = 0.0004) as well as with the percentage of body surface area affected by psoriasis (r = ?0.5013; P = 0.0005). Patients were divided into mild and severe groups based on the mean PASI score. Expression of TNFAIP3 mRNA in the mild group was higher than that in the severe group (P = 0.0064). Moreover, compared with that in healthy controls, the expression of TNFAIP3 mRNA in the mild group was significantly upregulated (P = 0.0004), but the expression of TNFAIP3 mRNA in the severe group was not. These results suggest that the expression level of TNFAIP3 plays an important role in the pathology of psoriasis vulgaris and that the loss of upregulation of TNFAIP3 expression may contribute to the severity of psoriasis vulgaris.

Jiang, Xuebing; Tian, Hongqing; Fan, Yuchen; Chen, Jie; Song, Yonghong; Wang, Shurong; Zhu, Faliang; Guo, Chun; Zhang, Lining



Greater expression of transforming growth factor alpha and proliferating cell nuclear antigen staining in mouse hepatoblastomas than hepatocellular carcinomas induced by a diethylnitrosamine-sodium phenobarbital regimen.  


Transforming growth factor alpha (TGF-alpha) is a potent stimulator of normal hepatocyte proliferation, considered to have relationship to the liver regeneration or carcinogenesis. In this study, we investigated immunohistochemically the association between expression of TGF-alpha and cell proliferation activity in mouse hepatoblastomas (HBs) and hepatocellular carcinomas (HCCs) induced in B6C3F1 mice by diethylnitrosamine and sodium phenobarbital. The TGF-alpha-positive rate in HBs (29.2%) was significantly higher than that in HCCs (12.7%). Likewise, the proliferating cell nuclear antigen-positive rate (22.2%) was higher than the HCC value (14.5%). On the individual data for both TGF-alpha and PCNA, most of the HBs showed higher positive rates than HCCs. In HBs, TGF-alpha was localized only in the nuclei, whereas some HCC cells stained positive both in their nuclei and cytoplasm (0.6%). These results suggest expression of TGF-alpha and its localization might be linked to cell proliferation and play a role in malignant progression of mouse HBs. PMID:11560253

Sakairi, T; Kobayashi, K; Goto, K; Okada, M; Kusakabe, M; Tsuchiya, T; Sugimoto, J; Sano, F; Mutai, M



A p75 tumor necrosis factor receptor-specific mutant of murine tumor necrosis factor alpha expressed from an adenovirus vector induces an antitumor response with reduced toxicity.  


The toxic effects of tumor necrosis factor alpha (TNFalpha) have greatly limited its use in tumor therapy. Recently, clear evidence has been obtained linking the p55 TNF receptor (TNFR) to the induction of systemic toxicity. We have generated a p75 murine TNFR (mTNFR)-specific mutant of mTNFalpha (D142N-A144R), cloned this gene into a recombinant adenovirus vector (Ad-75), and studied its efficacy for tumor immunotherapy of a murine transgenic breast cancer model. Cell culture supernatants from Ad-75-transduced cells showed no cytotoxic activity on L929 cells, but retained the ability to induce proliferation of a murine T-cell line (CT6); this activity was not blocked by soluble p55 mTNFR. Furthermore, it was shown that the mutant form of mTNFalpha was able to coimmunoprecipitate only with the p75 mTNFR and not with the p55 mTNFR. Tumors injected with Ad-75 became necrotic, and mice injected with < or =1 x 10(9) plaque-forming units showed no mortality, whereas both wild-type murine and human TNF vectors induced lethality at doses of 1 and 5 x 10(8) plaque-forming units. All Ad-TNF vectors induced partial or permanent tumor regressions, with cured mice showing immune memory against the tumor. These results demonstrate that a p75 mTNFR agonist expressed from a recombinant adenovirus vector does not induce mortality at doses that cause tumor regression. PMID:10505857

Marr, R A; Hitt, M; Gauldie, J; Muller, W J; Graham, F L



Tumor Necrosis Factor Alpha Enhances Influenza A Virus-Induced Expression of Antiviral Cytokines by Activating RIG-I Gene Expression  

Microsoft Academic Search

Epithelial cells of the lung are the primary targets for respiratory viruses. Virus-carried single-stranded RNA (ssRNA) can activate Toll-like receptors (TLRs) 7 and 8, whereas dsRNA is bound by TLR3 and a cytoplasmic RNA helicase, retinoic acid-inducible protein I (RIG-I). This recognition leads to the activation of host cell cytokine gene expression. Here we have studied the regulation of influenza

Sampsa Matikainen; Jukka Siren; Jorma Tissari; Ville Veckman; Jaana Pirhonen; Martina Severa; Qiang Sun; Rongtuan Lin; Seppo Meri; Gilles Uze; John Hiscott; Ilkka Julkunen



Flanking sequences for the human intercellular adhesion molecule-1 NF-kappaB response element are necessary for tumor necrosis factor alpha-induced gene expression.  


The regulated expression of intercellular adhesion molecule-1 (ICAM-1) by cytokines such as tumor necrosis factor alpha (TNF-alpha) plays an important role in inflammation and immune responses. Induction of ICAM-1 gene transcription by TNF-alpha has previously been shown to be dependent upon a region of the ICAM-1 5'-flanking sequences that contains a modified kappaB site. We demonstrate here that this modified kappaB site alone is insufficient for induction of transcription by TNF-alpha. Site-directed mutagenesis of both the kappaB site and specific flanking nucleotides demonstrates that both the specific 5'- and 3'-flanking sequences and the modified kappaB site are necessary for TNF-alpha induction. Further, site-directed mutagenesis of this modified kappaB site to a consensus kappaB site allows it to mediate transcriptional activation in response to TNF-alpha, even in the absence of specific flanking sequences. Transcription through this minimal ICAM-1 TNF-alpha-responsive region can be driven by co-expression of p65, and the minimal response element interacts with p65 and p50 in supershift mobility shift assays. However, when in vitro transcription/translation products for the Rel proteins are used in an electrophoretic mobility shift assay, only p65 is capable of binding the minimal response element while both p50 and p65 bind a consensus kappaB oligonucleotide. Additionally, in the absence of the specific flanking nucleotides, the ICAM-1 kappaB site is incapable of DNA-protein complex formation in both electrophoretic mobility shift assay and UV cross-linking/SDS-polyacrylamide gel electrophoresis analysis. These results demonstrate the requirement for specific flanking sequences surrounding a kappaB binding site for functional transcription factor binding and transactivation and TNF-alpha-mediated induction of ICAM-1. PMID:9188493

Paxton, L L; Li, L J; Secor, V; Duff, J L; Naik, S M; Shibagaki, N; Caughman, S W



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


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. PMID:22631659

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



An inhibitory effect of tumor necrosis factor-alpha antagonist to gene expression in monocrotaline-induced pulmonary hypertensive rats model  

PubMed Central

Purpose Tumor necrosis factor (TNF)-? is thought to contribute to pulmonary hypertension. We aimed to investigate the effect of infliximab (TNF-? antagonist) treatment on pathologic findings and gene expression in a monocrotaline-induced pulmonary hypertension rat model. Methods Six-week-old male Sprague-Dawley rats were allocated to 3 groups: control (C), single subcutaneous injection of normal saline (0.1 mL/kg); monocrotaline (M), single subcutaneous injection of monocrotaline (60 mg/kg); and monocrotaline + infliximab (M+I), single subcutaneous injection of monocrotaline plus single subcutaneous injection of infliximab (5 mg/kg). The rats were sacrificed after 1, 5, 7, 14, or 28 days. We examined changes in pathology and gene expression levels of TNF-?, endothelin-1 (ET-1), endothelin receptor A (ERA), endothelial nitric oxide synthase (eNOS), matrix metalloproteinase (MMP)2, and tissue inhibitor of matrix metalloproteinase (TIMP). Results The increase in medial wall thickness of the pulmonary arteriole in the M+I group was significantly lower than that in the M group on day 7 after infliximab treatment (P<0.05). The number of intra-acinar muscular arteries in the M+I group was lower than that in the M group on days 14 and 28 (P<0.05). Expression levels of TNF-?, ET-1, ERA, and MMP2 were significantly lower in the M+I group than in the M group on day 5, whereas eNOS and TIMP expressions were late in the M group (day 28). Conclusion Infliximab administration induced early changes in pathological findings and expression levels of TNF-?, and MMP2 in a monocrotaline-induced pulmonary hypertension rat model.

Kwon, Jung Hyun; Kim, Kwan Chang; Cho, Min-Sun; Kim, Hae Soon; Sohn, Sejung



Inhibition of tumor necrosis factor-{alpha}-induced expression of adhesion molecules in human endothelial cells by the saponins derived from roots of Platycodon grandiflorum  

SciTech Connect

Adhesion molecules play an important role in the development of atherogenesis and are produced by endothelial cells after being stimulated with various inflammatory cytokines. This study examined the effect of saponins that were isolated from the roots of Platycodon grandiflorum A. DC (Campanulaceae), Changkil saponins (CKS), on the cytokine-induced monocyte/human endothelial cell interaction, which is a crucial early event in atherogenesis. CKS significantly inhibited the TNF{alpha}-induced increase in monocyte adhesion to endothelial cells as well as decreased the protein and mRNA expression levels of vascular adhesion molecule-1 and intercellular cell adhesion molecule-1 on endothelial cells. Furthermore, CKS significantly inhibited the TNF{alpha}-induced production of intracellular reactive oxygen species (ROS) and activation of NF-{kappa}B by preventing I{kappa}B degradation and inhibiting I{kappa}B kinase activity. Overall, CKS has anti-atherosclerotic and anti-inflammatory activity, which is least in part the result of it reducing the cytokine-induced endothelial adhesion to monocytes by inhibiting intracellular ROS production, NF-{kappa}B activation, and cell adhesion molecule expression in endothelial cells.

Kim, Ji Young [Department of Pharmacy, College of Pharmacy, Research Center for Proteineous Materials, Chosun University, Kwangju (Korea, Republic of); Kim, Dong Hee [Department of Pathology, College of Oriental Medicine, Daejeon University, Daejeon (Korea, Republic of); Kim, Hyung Gyun [Department of Pharmacy, College of Pharmacy, Research Center for Proteineous Materials, Chosun University, Kwangju (Korea, Republic of); Song, Gyu-Yong [Department of Pharmacy, College of Pharmacy, Chungnam National University, Taejon (Korea, Republic of); Chung, Young Chul [Division of Food Science, Chinju International University, Chinju (Korea, Republic of); Roh, Seong Hwan [Jangsaeng Doraji Research Institute of Biotechnology, Jangsaeng Doraji Co., Ltd., Chinju (Korea, Republic of); Jeong, Hye Gwang [Department of Pharmacy, College of Pharmacy, Research Center for Proteineous Materials, Chosun University, Kwangju (Korea, Republic of)]. E-mail:



Tumor necrosis factor alpha-induced pulmonary vascular endothelial injury.  

PubMed Central

Tumor necrosis factor alpha (TNF-alpha) mediates components of the acute-phase response, stimulates granulocyte metabolism, and induces endothelial cell surface changes. We studied whether human recombinant TNF-alpha (rTNF-alpha) could increase pulmonary edema formation and pulmonary vascular permeability. Rabbits preinfused with 125I-albumin were administered rTNF-alpha or saline. Animals were sacrificed, and lung wet/dry weight ratios as well as bronchoalveolar lavage fluid and plasma 125I activities were determined. rTNF-alpha increased lung wet/dry weight ratios by 151% (P less than 0.02) and bronchoalveolar lavage fluid/plasma 125I activity ratios by 376% (P less than 0.01) compared with values for saline controls. Electron microscopy of lung sections demonstrated endothelial injury, perivascular edema, and extravasation of an ultrastructural permeability tracer. To demonstrate that rTNF-alpha could directly increase pulmonary vascular endothelial permeability in vitro, we studied albumin transfer across cultured porcine pulmonary artery endothelial cell monolayers. rTNF-alpha induced time-dependent dose-response increments in transendothelial albumin flux in the absence of granulocyte effector cells. These observations suggest that rTNF-alpha can provoke acute pulmonary vascular endothelial injury in vivo as well as in vitro. Images

Goldblum, S E; Hennig, B; Jay, M; Yoneda, K; McClain, C J



Molecular characterization and expression analysis of three hypoxia-inducible factor alpha subunits, HIF-1?/2?/3? of the hypoxia-sensitive freshwater species, Chinese sucker.  


Hypoxia-inducible factors (HIFs) are transcription factors that respond to changes in oxygen tension in the cellular environment. In this study, we identified full-length cDNAs of HIF-1?, HIF-2? and HIF-3? in an endangered hypoxia-sensitive fish species, Chinese sucker. The HIF-1?/2?/3? cDNAs are 3890, 3230 and 3374 bp in length, encoding 780, 782 and 632 amino acid residues, respectively. The real-time PCR results suggested that HIF-1? and HIF-3? mRNAs were highly expressed in liver and gonads, followed by spleen and muscle. Moreover, HIF-1? and HIF-3? transcription factors revealed similar developmental expression patterns, with the lowest expression at 48h post-fertilization, and reaching the highest expression level at 360 h post-fertilization. Short-term hypoxia exposure (2.2, 2.8 and 3.2mg/L dissolved oxygen for 24h) increased mRNA levels of HIF-1? and HIF-3?. HIF-2? mRNA showed similar expression patterns with that of HIF-1? and HIF-3?, however, its expression was extremely low in the spatio-temporal expression patterns and hypoxia treatment. This is the first report describing the potential to identify hypoxia-sensitive/tolerant fishes according to the number of the serine residues of fish oxygen-dependent degradation (ODD) domain. It was suggested that Cyprinomorpha fishes, with less than 40 serine residues in fish ODD domain were hypoxia-sensitive fishes and more than 40 serine residues in this domain were hypoxia-tolerant fishes. PMID:22342256

Chen, Nan; Chen, Li Ping; Zhang, Jie; Chen, Cheng; Wei, Xin Lan; Gul, Yasmeen; Wang, Wei Min; Wang, Huan Ling



Down-regulatory effect of usnic acid on nuclear factor-kappaB-dependent tumor necrosis factor-alpha and inducible nitric oxide synthase expression in lipopolysaccharide-stimulated macrophages RAW 264.7.  


The purpose of this study was to investigate the molecular mechanisms that are responsible for the antiinflammatory effect of usnic acid (UA). UA is one of the most common and abundant lichen metabolites. The present study examined the effects of UA on the tumor necrosis factor-alpha (TNF-alpha) and nitric oxide (NO) production induced by lipopolysaccharide (LPS) in RAW264.7 macrophages and the underlying molecular mechanisms. UA decreased the TNF-alpha level in LPS-stimulated RAW264.7 macrophages in dose-dependent manner, the IC(50) value was 12.8 microM. RT-PCR analysis indicated that it inhibited TNF-alpha mRNA expression. Furthermore, it inhibited NO production in LPS-activated RAW264.7 macrophages, the IC(50) value was 4.7 microM. Western blot analysis showed that UA attenuated LPS-induced synthesis of iNOS protein and nuclear translocation of NF-kappaB p65 in the macrophages, in parallel. UA also inhibited LPS-mediated I-kappaBalpha degradation. Taken together, this suggests that UA has an antiinflammatory effect by inhibiting TNF-alpha and iNOS expression, possibly through suppression of nuclear translocation of NF-kappaB p65 and I-kappaBalpha degradation. PMID:19003951

Jin, Ju-qing; Li, Cui-qin; He, Lang-chong



The MC160 Protein Expressed by the Dermatotropic Poxvirus Molluscum Contagiosum Virus Prevents Tumor Necrosis Factor Alpha-Induced NF-?B Activation via Inhibition of I Kappa Kinase Complex Formation  

PubMed Central

The pluripotent cytokine tumor necrosis factor alpha (TNF-?) binds to its cognate TNF receptor I (TNF-RI) to stimulate inflammation via activation of the NF-?B transcription factor. To prevent the detrimental effects of TNF-? in keratinocytes infected with the molluscum contagiosum virus (MCV), this poxvirus is expected to produce proteins that block at least one step of the TNF-RI signal transduction pathway. One such product, the MC160 protein, is predicted to interfere with this cellular response because of its homology to other proteins that regulate TNF-RI-mediated signaling. We report here that expression of MC160 molecules did significantly reduce TNF-?-mediated NF-?B activation in 293T cells, as measured by gene reporter and gel mobility shift assays. Since we observed that MC160 decreased other NF-?B activation pathways, namely those activated by receptor-interacting protein, TNF receptor-associated factor 2, NF-?B-inducing kinase, or MyD88, we hypothesized that the MC160 product interfered with I kappa kinase (IKK) activation, an event common to multiple signal transduction pathways. Indeed, MC160 protein expression was associated with a reduction in in vitro IKK kinase activity and IKK subunit phosphorylation. Further, IKK1-IKK2 interactions were not detected in MC160-expressing cells, under conditions demonstrated to induce IKK complex formation, but interactions between the MC160 protein and the major IKK subunits were undetectable. Surprisingly, MC160 expression correlated with a decrease in IKK1, but not IKK2 levels, suggesting a mechanism for MC160 disruption of IKK1-IKK2 interactions. MCV has probably retained its MC160 gene to inhibit NF-?B activation by interfering with signaling via multiple biological mediators. In the context of an MCV infection in vivo, MC160 protein expression may dampen the cellular production of proinflammatory molecules and enhance persistent infections in host keratinocytes.

Nichols, Daniel Brian; Shisler, Joanna L.



Relation of tissue Doppler-derived myocardial velocities to serum levels and myocardial gene expression of tumor necrosis factor-alpha and inducible nitric oxide synthase in patients with ischemic cardiomyopathy having coronary artery bypass grafting.  


We evaluated the relation of segmental and annular tissue Doppler (TD) velocities to serum levels and myocardial gene expression of tumor necrosis factor-alpha (TNF-alpha) and inducible nitric oxide (NOS2) in humans. Seven patients with coronary artery disease underwent echocardiographic examination including TD imaging, along with transmural endomyocardial biopsies (2 biopsies per patient for a total of 14 specimens) at the time of bypass surgery. The specimens were analyzed for the number of mRNA copies of TNF-alpha and NOS2. In addition, serum levels of TNF-alpha and nitrite were determined before and after surgery. Normal segments (n = 7) had fewer numbers of mRNA copies of both TNF-alpha (54.6 vs 6.5, p = 0.002) and NOS2 (3,093 +/- 486 vs 661 +/- 259, p <0.001) than dysfunctional segments (n = 7). Peak systolic velocity (Sm) (13.3 +/- 1.4 vs 4.9 +/- 1.6 cm/s, p = 0.002) and early diastolic velocity (Em) (16.5 +/- 2.7 vs 8.8 +/- 1.3 cm/s, p = 0.02) were significantly higher in normal segments. A significant correlation was present among Em, Sm, and the number of mRNA copies of TNF-alpha and NOS2 at baseline and during infusion of dobutamine (r range -0.72 to -0.92, p <0.01 for all). Likewise, significant relations were present between the changes in serum cytokine levels and changes in the mitral annulus diastolic velocity, E/Ea ratio, and end-diastolic wall stress (r range 0.75 to 0.88, p expression, which are known to have negative inotropic and lusitropic effects when overexpressed. PMID:12356382

Kalra, Dinesh K; Ramchandani, Mahesh; Zhu, Xi; Lawrie, Gerald; Reardon, Michael J; Mann, Douglas L; Zoghbi, William A; Nagueh, Sherif F



Expression of transforming growth factor alpha in human tissues: Immunohistochemical study and Northern blot analysis  

Microsoft Academic Search

Summary The expression of transforming growth factor alpha (TGF-a) was examined in various human tissues and the fetus, using immunohistochemistry and Northern blot analysis. TGF-a immunoreactivity was detected mainly in the epithelial cells of the digestive tract, liver, pancreas, kidney, thyroid, adrenal, skin, mammary gland and genital organs. In the digestive tract, epithelial cells with regenerative change or hyperplastic change

Wataru Yasui; Zhong-Qiang Ji; Hiroki Kuniyasu; Ayse Ayhan; Hiroshi Yokozaki; Hisao Ito; Eiichi Tahara



Role of tumor necrosis factor alpha-induced protein 1 in paclitaxel resistance.  


Paclitaxel has been extensively used as an antitumor drug to treat a broad range of epithelial cancers, including breast and cervical cancers. However, the efficacy of this drug is greatly limited by the development of acquired resistance. Identification of the underlying resistance mechanisms may inform the development of new therapies that elicit long-term response of tumors to paclitaxel treatment. Here we report that increased expression of TNFAIP1 (tumor necrosis factor alpha-induced protein 1) confers acquired resistance to paclitaxel. TNFAIP1 is shown to compete with paclitaxel for binding to ?-tubulin, thereby preventing paclitaxel-induced tubulin polymerization, cell cycle arrest and ultimate cell death. We also show that expression of TNFAIP1 is regulated by the transcriptional factor Sp1. In a xenograft mouse model, increased expression of TNFAIP1 decreases, whereas knockdown of TNFAIP1 increases tumor response to paclitaxel. Therefore, these results reveal tnfaip1 as a novel paclitaxel-resistance associated gene and suggest that TNFAIP1 may represent a valuable therapeutic target for the treatment of cancer. PMID:23912453

Zhu, Y; Yao, Z; Wu, Z; Mei, Y; Wu, M



The Role of Tumor Necrosis Factor Alpha in Lipopolysaccharide\\/Ranitidine-Induced Inflammatory Liver Injury  

Microsoft Academic Search

Exposure to a nontoxic dose of bacterial lipopolysaccharide (LPS) increases the hepatotoxicity of the histamine-2 (H2) re- ceptor antagonist, ranitidine (RAN). Because some of the pathophysiologic effects associated with LPS are mediated through the expression and release of inflammatory mediators such as tumor necrosis factor alpha (TNF), this study was designed to gain insights into the role of TNF in

Francis F. Tukov; James P. Luyendyk; Patricia E. Ganey; R. A. Roth



Salmonella Flagellin Induces Tumor Necrosis Factor Alpha in a Human Promonocytic Cell Line  

Microsoft Academic Search

During infection of the gastrointestinal tract, salmonellae induce cytokine production and inflammatory responses which are believed to mediate tissue damage in the host. In a previous study, we reported that salmonellae possess the ability to stimulate tumor necrosis factor alpha (TNF-a) accumulation in primary human monocytes, as well as in the human promonocytic cell line U38. In this model system,




Tumour necrosis factor alpha and interleukin-1 beta induce specific subunits of NFKB to bind the HIV-1 enhancer: characterisation of transcription factors controlling human immunodeficiency virus type 1 gene expression in neural cells.  


In human astrocytoma and neuroblastoma cell lines tumour necrosis factor alpha (TNF alpha) and interleukin 1 beta (IL-1 beta) induced NFKB and an additional KB-specific protein of approximately 80 K to bind the HIV-1 enhancer. One nucleoprotein complex contained prototypical NFKB comprising of p50 and p65 subunits and a second contained the p65 subunit and an 80 K factor, p80. Over a 24 hr period of cytokine stimulation the p50/p65 complex of NFKB was present in the nucleus whilst the second complex declined in abundance after two hours due to the decline of p80. In unstimulated cells, DNAse I footprinting revealed a previously unidentified octamer-like binding site in the negative regulatory element (NRE) of the HIV-1 long terminal repeat (LTR) which specifically bound protein factors present in human astrocytoma, neuroblastoma and murine oligodendroglioma cell lines. A second unique motif, also in the NRE, was observed with extracts from one human neuroblastoma cell line and a murine oligodendroglioma. Footprinting analysis also confirmed that Sp1, TATA, Site A and Site B motifs of the LTR were occupied by nuclear factors present in neural cells. PMID:8074713

Swingler, S; Morris, A; Easton, A



Expression of tumour necrosis factor alpha and its receptors in carcinoma of the breast.  

PubMed Central

The expression of tumour necrosis factor alpha (TNF-alpha) and its two distinct receptors, TNF-R p55 and TNF-R p75, was assessed by immunocytochemistry in 28 primary breast cancer and three reduction mammoplasty specimens ('normal' breast tissue). Expression of TNF-alpha or TNF-R p75 was not detectable in normal breast tissue or in non-malignant breast tissue adjacent to the tumours. By contrast, TNF-R p55 was expressed by occasional stromal cells in normal tissue. TNF-alpha was expressed focally in 50% of the tumours studied, being largely localised to macrophage-like cells in the stroma. TNF-R p55 was expressed by a population of stromal cells in all the tumours examined, and a varying proportion of neoplastic cells in 75% of these tissues. TNF-R p75 was detected in about 70% of the tumours, immunoreactivity being confined mainly to cells in the stroma. In this preliminary study there was no association between the above cytokine parameters and such measures of tumour biology as lymph node status, tumour grade, proliferative activity or degree of angiogenesis. However, there was a correlation between the expression of TNF-R p55 by blood vessels and the number of leucocytes present. Images Figure 1

Pusztai, L.; Clover, L. M.; Cooper, K.; Starkey, P. M.; Lewis, C. E.; McGee, J. O.



Tumor necrosis factor alpha and adalimumab differentially regulate CD36 expression in human monocytes  

PubMed Central

In chronic inflammatory diseases, such as rheumatoid arthritis, inflammation acts as an independent cardiovascular risk factor and the use of anti-inflammatory drugs, such as anti-tumor necrosis factor alpha (anti-TNF?), may decrease this risk. The phagocytosis of oxidized low density lipoproteins (LDLs) accumulated in the subendothelium by mononuclear cells influences atherosclerosis and depends on CD36 expression. We investigated the role of TNF? and adalimumab, a human anti-TNF? monoclonal antibody widely used in human pathology, in CD36 expression in human monocytes. Human monocytes were prepared by adherence from whole-blood buffy-coat fractions from healthy donors. CD36 expression was assessed by RT-PCR and flow cytometry, with various TNF? or adalimumab concentrations. Implication of peroxisome proliferator-activated receptor (PPAR)? in the regulation of CD36 expression was assessed using specific inhibitor or gel shift assays. The impact of redox signaling was investigated using quantification of reactive oxygen species, antioxidant and a NADPH oxidase inhibitor. The F(ab')2 fragment of adalimumab was isolated and its effect was analyzed. TNF? inhibits both CD36 membrane expression and mRNA expression. This inhibition involves a reduction in PPAR? activation. In contrast, adalimumab increases both CD36 membrane expression and mRNA expression. This induction is independent of the Fc portion of adalimumab and involves redox signaling via NADPH oxidase activation. CD36 expression on human monocytes is inhibited by TNF? and independently increased by adalimumab. These data highlight that pro-inflammatory cytokines and their specific neutralization influence the expression of cellular receptors implicated in atherosclerosis. Further studies are needed to investigate the clinical implications of these results in accelerated atherosclerosis observed in rheumatoid arthritis.

Boyer, Jean Frederic; Balard, Patricia; Authier, Helene; Faucon, Bruno; Bernad, Jose; Mazieres, Bernard; Davignon, Jean-Luc; Cantagrel, Alain; Pipy, Bernard; Constantin, Arnaud



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...


Spontaneous Inflammatory Demyelinating Disease in Transgenic Mice Showing Central Nervous System-Specific Expression of Tumor Necrosis Factor alpha  

Microsoft Academic Search

Cytokines are now recognized to play important roles in the physiology of the central nervous system (CNS) during health and disease. Tumor necrosis factor alpha (TNF-alpha) has been implicated in the pathogenesis of several human CNS disorders including multiple sclerosis, AIDS dementia, and cerebral malaria. We have generated transgenic mice that constitutively express a murine TNF-alpha transgene, under the control

Lesley Probert; Katerina Akassoglou; Manolis Pasparakis; George Kontogeorgos; George Kollias




EPA Science Inventory



Critical Role of the Complement System in Group B Streptococcus-Induced Tumor Necrosis Factor Alpha Release  

Microsoft Academic Search

Group B Streptococcus (GBS) is a major cause of newborn sepsis and meningitis and induces systemic release of tumor necrosis factor alpha (TNF-), believed to play a role in morbidity and mortality. While previous studies have shown that GBS can induce TNF- release from monocytes and macrophages, little is known about the potential modulating effect of plasma or serum on

Ofer Levy; Rochelle M. Jean-Jacques; Colette Cywes; Richard B. Sisson; Kol A. Zarember; Paul J. Godowski; Jennifer L. Christianson; Hilde-Kari Guttormsen; Michael C. Carroll; Anne Nicholson-Weller; Michael R. Wessels



Production of human tumor necrosis factor alpha, interleukin-6, and interleukin-10 is induced by lactic acid bacteria.  

PubMed Central

To investigate the role of cytokines in interactions between lactic acid bacteria and the immune system, we measured production of tumor necrosis factor alpha, interleukin-6 (IL-6), and IL-10 from human peripheral blood mononuclear cells after stimulation with live or glutaraldehyde-fixed bacteria. Production of tumor necrosis factor alpha, IL-6, and, in some cases, IL-10 was induced in amounts even greater than those obtained with lipopolysaccharide as a stimulant. Our results suggest that lactic acid bacteria can stimulate nonspecific immunity.

Miettinen, M; Vuopio-Varkila, J; Varkila, K



Production of human tumor necrosis factor alpha, interleukin-6, and interleukin-10 is induced by lactic acid bacteria.  


To investigate the role of cytokines in interactions between lactic acid bacteria and the immune system, we measured production of tumor necrosis factor alpha, interleukin-6 (IL-6), and IL-10 from human peripheral blood mononuclear cells after stimulation with live or glutaraldehyde-fixed bacteria. Production of tumor necrosis factor alpha, IL-6, and, in some cases, IL-10 was induced in amounts even greater than those obtained with lipopolysaccharide as a stimulant. Our results suggest that lactic acid bacteria can stimulate nonspecific immunity. PMID:8945595

Miettinen, M; Vuopio-Varkila, J; Varkila, K



Molecular mechanisms of hepatocellular apoptosis induced by trovafloxacin-tumor necrosis factor-alpha interaction.  


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

Beggs, Kevin M; Fullerton, Aaron M; Miyakawa, Kazuhisa; Ganey, Patricia E; Roth, Robert A



Tumor necrosis factor alpha maintains denervation-induced homeostatic synaptic plasticity of mouse dentate granule cells  

PubMed Central

Neurons which lose part of their input respond with a compensatory increase in excitatory synaptic strength. This observation is of particular interest in the context of neurological diseases, which are accompanied by the loss of neurons and subsequent denervation of connected brain regions. However, while the cellular and molecular mechanisms of pharmacologically induced homeostatic synaptic plasticity have been identified to a certain degree, denervation-induced homeostatic synaptic plasticity remains not well understood. Here, we employed the entorhinal denervation in vitro model to study the role of tumor necrosis factor alpha (TNF?) on changes in excitatory synaptic strength of mouse dentate granule cells following partial deafferentation. Our experiments disclose that TNF? is required for the maintenance of a compensatory increase in excitatory synaptic strength at 3–4 days post lesion (dpl), but not for the induction of synaptic scaling at 1–2 dpl. Furthermore, laser capture microdissection combined with quantitative PCR demonstrates an increase in TNF?-mRNA levels in the denervated zone, which is consistent with our previous finding on a local, i.e., layer-specific increase in excitatory synaptic strength at 3–4 dpl. Immunostainings for the glial fibrillary acidic protein and TNF? suggest that astrocytes are a source of TNF? in our experimental setting. We conclude that TNF?-signaling is a major regulatory system that aims at maintaining the homeostatic synaptic response of denervated neurons.

Becker, Denise; Zahn, Nadine; Deller, Thomas; Vlachos, Andreas



Bacterial evasion of host immune defense: Yersinia enterocolitica encodes a suppressor for tumor necrosis factor alpha expression.  

PubMed Central

The ability of the enteropathogenic Yersinia enterocolitica to survive and proliferate in host tissue depends on a 70-kb plasmid known to encode a number of released Yersinia outer proteins that act as virulence factors by inducing cytotoxicity and inhibiting phagocytosis. This study demonstrates that one of the Yersinia outer proteins, the 41-kDa YopB, suppresses the production of tumor necrosis factor alpha (TNF-alpha), a macrophage-derived cytokine with central roles in the regulation of immune and inflammatory responses to infection. This conclusion is based on several lines of evidence. First, in macrophage cultures, suppression of TNF-alpha mRNA expression was induced by culture supernatant (CS+) of plasmid-bearing yersiniae, the effect which was blocked by anti-YopB antiserum. Second, suppression of TNF-alpha production, but not of interleukin-1 (IL-1) and IL-6, was induced by purified YopB. Third, in Yersinia-infected mice, no increase in TNF-alpha mRNA expression was observed in Peyer's patches, the primary site of bacterial invasion, compared with IL-1 (alpha and beta) mRNA. Finally, administration of anti-YopB antiserum to mice prior to infection with Y. enterocolitica increased TNF activity levels in Peyer's patches and coincided with a reduction in bacterial growth. The results thus provide direct evidence for a secreted eubacterial virulence factor that mediates selective suppression of TNF-alpha production. Although suppression of this single cytokine response is probably not sufficient to facilitate survival of the infecting organisms, the results suggest that suppression of TNF-alpha production by YopB significantly contributes to the evasion of Y. enterocolitica from antibacterial host defense.

Beuscher, H U; Rodel, F; Forsberg, A; Rollinghoff, M



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

PubMed Central

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

Turksen, K; Choi, Y; Fuchs, E



Tumor necrosis factor-alpha induces the phosphorylation of 28kDa stress proteins in endothelial cells: possible role in protection against cytotoxicity?  


Tumor necrosis factor-alpha has been shown to rapidly increase the phosphorylation of three 28 kDa proteins in bovine aortic endothelial cells but not in L929 cells. Tumor necrosis factor-alpha induces the necrosis of the latter cells but not of the former. Arsenite enhanced the phosphorylation of the same 28kDa proteins as tumor necrosis factor-alpha in the endothelial cells. As stress proteins often play a protective role, we suggest that the phosphorylation of these proteins in endothelial cells may be responsible for the resistance of these cells to tumor necrosis factor-alpha. PMID:2775267

Robaye, B; Hepburn, A; Lecocq, R; Fiers, W; Boeynaems, J M; Dumont, J E



Increased tumor necrosis factor alpha-converting enzyme activity induces insulin resistance and hepatosteatosis in mice.  


Tumor necrosis factor alpha-converting enzyme (TACE, also known as ADAM17) was recently involved in the pathogenesis of insulin resistance. We observed that TACE activity was significantly higher in livers of mice fed a high-fat diet (HFD) for 1 month, and this activity was increased in liver > white adipose tissue > muscle after 5 months compared with chow control. In mouse hepatocytes, C(2)C(12) myocytes, and 3T3F442A adipocytes, TACE activity was triggered by palmitic acid, lipolysaccharide, high glucose, and high insulin. TACE overexpression significantly impaired insulin-dependent phosphorylation of AKT, GSK3, and FoxO1 in mouse hepatocytes. To test the role of TACE activation in vivo, we used tissue inhibitor of metalloproteinase 3 (Timp3) null mice, because Timp3 is the specific inhibitor of TACE and Timp3(-/-) mice have higher TACE activity compared with wild-type (WT) mice. Timp3(-/-) mice fed a HFD for 5 months are glucose-intolerant and insulin-resistant; they showed macrovesicular steatosis and ballooning degeneration compared with WT mice, which presented only microvesicular steatosis. Shotgun proteomics analysis revealed that Timp3(-/-) liver showed a significant differential expression of 38 proteins, including lower levels of adenosine kinase, methionine adenosysltransferase I/III, and glycine N-methyltransferase and higher levels of liver fatty acid-binding protein 1. These changes in protein levels were also observed in hepatocytes infected with adenovirus encoding TACE. All these proteins play a role in fatty acid uptake, triglyceride synthesis, and methionine metabolism, providing a molecular explanation for the increased hepatosteatosis observed in Timp3(-/-) compared with WT mice. Conclusion: We have identified novel mechanisms, governed by the TACE-Timp3 interaction, involved in the determination of insulin resistance and liver steatosis during overfeeding in mice. PMID:19877183

Fiorentino, Loredana; Vivanti, Alessia; Cavalera, Michele; Marzano, Valeria; Ronci, Maurizio; Fabrizi, Marta; Menini, Stefano; Pugliese, Giuseppe; Menghini, Rossella; Khokha, Rama; Lauro, Renato; Urbani, Andrea; Federici, Massimo



Extracellular inosine participates in tumor necrosis factor-alpha induced nitric oxide production in cultured Sertoli cells.  


Recent reports have described purinergic modulation of tumor necrosis factor-alpha (TNF-alpha) signaling in neutrophils and astrocytes. In Sertoli cells, both TNF-R1 and TNF-R2 TNF-alpha receptors are present and this cytokine modulates many functions of these cells related to the maintenance of spermatogenesis. Sertoli cells express distinct purinoreceptors and previous work has shown that these cells secrete extracellular nucleotides and their metabolites. In this work, we studied the possible role of extracellular purines in TNF-alpha signaling in cultured Sertoli cells. This cytokine increased inosine concentration from 30 min to 6 h, with no effect at 24 h. Both TNF-alpha and inosine increased nitrite accumulation and nitric oxide synthase activity. Erythro-9-(2-hydroxy-3-nonyl)adenine (EHNA), an adenosine deaminase inhibitor, abolished the TNF-alpha induced inosine increase, nitrite accumulation and nitric oxide synthase activity. These results suggest that extracellular inosine acts as intermediary in TNF-alpha stimulated nitric oxide production in cultured Sertoli cells. PMID:16328964

de Souza, Luiz Fernando; Gelain, Daniel Pens; Jardim, Fernanda Rafaela; Ribeiro, Gisele Roncheti; Zim, Marcelo; Bernard, Elena Aida



Two Coordinated Mechanisms Underlie Tumor Necrosis Factor Alpha-Induced Immediate and Delayed I?B Kinase Activation  

PubMed Central

Tumor necrosis factor alpha (TNF-?)-induced NF-?B activation has been believed to depend on TRAF2- and cIAP1-mediated RIP1 ubiquitination. However, recent findings have challenged the notion that these proteins play essential roles in NF-?B activation. Here, by assessing the kinetics and amplitude of I?B kinase (IKK) activation, we report that TNF-?-induced immediate and robust activation of IKK requires K63-linked and linearly linked ubiquitination of RIP1 and that in the absence of RIP1 expression, TRAF2 and cIAP1 cooperatively induce delayed IKK activation by recruiting LUBAC to TNFR1. Knockdown of HOIP (a component of LUBAC) in RIP1-deficient cells completely impairs the recruitment and activation of IKK but does not affect K63-linked ubiquitination of TRAF2 and recruitment of TAK1 to TNFR1, suggesting that the K63-linked ubiquitin chain is not capable of recruiting IKK in vivo. We also demonstrate that TRAF2 and cIAP1 together, but not either one alone, directly catalyze linearly linked ubiquitination of RIP1. Importantly, in embryonic hepatocytes, TNF-? activates NF-?B through a RIP1-independent pathway. Thus, our findings clarify molecular details of this important signaling mechanism by providing evidence for the existence of two phases of IKK activation: the immediate phase, induced by TRAF2/cIAP1-mediated ubiquitination of RIP1, and the delayed phase, activated by TRAF2/cIAP1-dependent recruitment of LUBAC.

Blackwell, Ken; Zhang, Laiqun; Workman, Lauren M.; Ting, Adrian T.; Iwai, Kazuhiro



Tumor Necrosis Factor alpha (TNF{alpha}) regulates CD40 expression through SMAR1 phosphorylation  

SciTech Connect

CD40 plays an important role in mediating inflammatory response and is mainly induced by JAK/STAT phosphorylation cascade. TNF{alpha} is the key cytokine that activates CD40 during inflammation and tumorigenesis. We have earlier shown that SMAR1 can repress the transcription of Cyclin D1 promoter by forming a HDAC1 dependent repressor complex. In this study, we show that SMAR1 regulates the transcription of NF-{kappa}B target gene CD40. SMAR1 recruits HDAC1 and forms a repressor complex on CD40 promoter and keeps its basal transcription in check. Further, we show that TNF{alpha} stimulation induces SMAR1 phosphorylation at Ser-347 and promotes its cytoplasmic translocation, thus releasing its negative effect. Concomitantly, TNF{alpha} induced phosphorylation of STAT1 at Tyr-701 by JAK1 facilitates its nuclear translocation and activation of CD40 through p300 recruitment and core Histone-3 acetylation. Thus, TNF{alpha} mediated regulation of CD40 expression occurs by dual phosphorylation of SMAR1 and STAT1.

Singh, Kamini; Sinha, Surajit; Malonia, Sunil Kumar; Chattopadhyay, Samit, E-mail:



Ubiquitination-Deubiquitination by the TRIM27-USP7 Complex Regulates Tumor Necrosis Factor Alpha-Induced Apoptosis  

PubMed Central

Tumor necrosis factor alpha (TNF-?) plays a role in apoptosis and proliferation in multiple types of cells, and defects in TNF-?-induced apoptosis are associated with various autoimmune diseases. Here, we show that TRIM27, a tripartite motif (TRIM) protein containing RING finger, B-box, and coiled-coil domains, positively regulates TNF-?-induced apoptosis. Trim27-deficient mice are resistant to TNF-?–d-galactosamine-induced hepatocyte apoptosis. Trim27-deficient mouse embryonic fibroblasts (MEFs) are also resistant to TNF-?–cycloheximide-induced apoptosis. TRIM27 forms a complex with and ubiquitinates the ubiquitin-specific protease USP7, which deubiquitinates receptor-interacting protein 1 (RIP1), resulting in the positive regulation of TNF-?-induced apoptosis. Our findings indicate that the ubiquitination-deubiquitination cascade mediated by the TRIM27-USP7 complex plays an important role in TNF-?-induced apoptosis.

Zaman, Mohammad Mahabub-Uz; Nomura, Teruaki; Takagi, Tsuyoshi; Okamura, Tomoo; Jin, Wanzhu; Shinagawa, Toshie; Tanaka, Yasunori



Tumor necrosis factor-alpha promotes tumor growth by inducing vascular endothelial growth factor.  


Tumor necrosis factor (TNF)-? has been proved as an adjuvant therapy for tumor by FDA. However, the effect of chronic TNF-? expression for tumor is still controversial. In this study, we investigated the effect of low-dose TNF-? on tumor growth. We confirmed that low-dose TNF-? promoted angiogenesis of tumor in vivo, vascular endothelial growth factor (VEGF) and hypoxia-inducible factor (HIF)-1?, the transcription factor of VEGF, were both upregulated. Our results suggested that low-dose TNF-? was a powerful activator of angiogenesis in tumor and HIF-1?-VEGF pathway seemed to be the most important molecular mechanism. PMID:21740086

Jing, Yingying; Ma, Nannan; Fan, Tingting; Wang, Chenyang; Bu, Xinxin; Jiang, Guocheng; Li, Rong; Gao, Lu; Li, Ding; Wu, Mengchao; Wei, Lixin



An endoplasmic reticulum transmembrane prolyl 4-hydroxylase is induced by hypoxia and acts on hypoxia-inducible factor alpha.  


Prolyl 4-hydroxylases (P4Hs) act on collagens (C-P4Hs) and the oxygen-dependent degradation domains (ODDDs) of hypoxia-inducible factor alpha subunits (HIF-P4Hs) leading to degradation of the latter. We report data on a human P4H possessing a transmembrane domain (P4H-TM). Its gene is also found in zebrafish but not in flies and nematodes. Its sequence more closely resembles those of the C-P4Hs than the HIF-P4Hs, but it lacks the peptide substrate-binding domain of the C-P4Hs. P4H-TM levels in cultured cells are increased by hypoxia, and P4H-TM is N-glycosylated and is located in endoplasmic reticulum membranes with its catalytic site inside the lumen, a location differing from those of the HIF-P4Hs. Despite this, P4H-TM overexpression in cultured neuroblastoma cells reduced HIF-alpha ODDD reporter construct levels, and its small interfering RNA increased HIF-1alpha protein level, in the same way as those of HIF-P4Hs. Furthermore, recombinant P4H-TM hydroxylated the two critical prolines in HIF-1alpha ODDD in vitro, with a preference for the C-terminal proline, whereas it did not hydroxylate any prolines in recombinant type I procollagen chains. PMID:17726031

Koivunen, Peppi; Tiainen, Päivi; Hyvärinen, Jaana; Williams, Kim E; Sormunen, Raija; Klaus, Stephen J; Kivirikko, Kari I; Myllyharju, Johanna



Endothelial nitric-oxide synthase enhances lipopolysaccharide-stimulated tumor necrosis factor-alpha expression via cAMP-mediated p38 MAPK pathway in cardiomyocytes.  


The purpose of this study was to investigate the role of endothelial nitric-oxide synthase (eNOS), cAMP, and p38 MAPK in tumor necrosis factor-alpha (TNF-alpha) expression induced by lipopolysaccharide (LPS). LPS dose- and time-dependently induced phosphorylation of p38 MAPK and TNF-alpha expression in neonatal mouse cardiomyocytes. TNF-alpha expression was preceded by p38 MAPK phosphorylation, and selective inhibition of p38 MAPK abrogated LPS-induced TNF-alpha expression. Deficiency in eNOS decreased basal and LPS-stimulated TNF-alpha expression in cardiomyocytes. NOS inhibitor l-NAME attenuated LPS-induced p38 MAPK phosphorylation and TNF-alpha production in wild-type cardiomyocytes, whereas NO donor 2,2'-(hydroxynitrosohydrazono)bis-ethanamine (DETA-NO) (2 microm) or overexpression of eNOS by adenoviral gene transfer restored the response of eNOS(-/-) cardiomyocytes to LPS. These effects of NO were mediated through cAMP-dependent pathway based on the following facts. First, deficiency in eNOS decreased basal levels of intracellular cAMP, and DETA-NO elevated intracellular cAMP levels in eNOS(-/-) cardiomyocytes. Second, a cAMP analogue 8-Br-cAMP mimicked the effect of NO in eNOS(-/-) cardiomyocytes. Third, either inhibition of cAMP or cAMP-dependent protein kinase attenuated LPS-stimulated p38 MAPK phosphorylation and TNF-alpha production in wild-type cardiomyocytes. In conclusion, eNOS enhances LPS-stimulated TNF-alpha expression in cardiomyocytes. Activation of p38 MAPK is essential in LPS-stimulated TNF-alpha expression. Moreover, the effects of NO on LPS-stimulated TNF-alpha expression are mediated through cAMP/cAMP-dependent protein kinase-dependent p38 MAPK pathway in neonatal cardiomyocytes. PMID:12506117

Peng, Tianqing; Lu, Xiangru; Lei, Ming; Feng, Qingping



Molecular Characterization and Expression Analysis of Equine Vascular Endothelial Growth Factor Alpha (VEGF?) Gene in Horse (Equus caballus)  

PubMed Central

The objective of this study was to determine the molecular characteristics of the horse vascular endothelial growth factor alpha gene (VEGF?) by constructing a phylogenetic tree, and to investigate gene expression profiles in tissues and blood leukocytes after exercise for development of suitable biomarkers. Using published amino acid sequences of other vertebrate species (human, chimpanzee, mouse, rat, cow, pig, chicken and dog), we constructed a phylogenetic tree which showed that equine VEGF? belonged to the same clade of the pig VEGF?. Analysis for synonymous (Ks) and non-synonymous substitution ratios (Ka) revealed that the horse VEGF? underwent positive selection. RNA was extracted from blood samples before and after exercise and different tissue samples of three horses. Expression analyses using reverse transcription-polymerase chain reaction (RT-PCR) and quantitative-polymerase chain reaction (qPCR) showed ubiquitous expression of VEGF? mRNA in skeletal muscle, kidney, thyroid, lung, appendix, colon, spinal cord, and heart tissues. Analysis of differential expression of VEGF? gene in blood leukocytes after exercise indicated a unimodal pattern. These results will be useful in developing biomarkers that can predict the recovery capacity of racing horses.

Song, Ki-Duk; Cho, Hyun-Woo; Lee, Hak-Kyo; Cho, Byung Wook



Tumor necrosis factor-alpha-induced reactive oxygen species formation is mediated by JNK1-dependent ferritin degradation and elevation of labile iron pool.  


Tumor necrosis factor alpha induces increased reactive oxygen species (ROS) generation in different experimental models. However, the nature of this phenomenon is still unknown. We hypothesized that TNF-induced ROS formation is due to JNK-regulated ferritin degradation and an increase in labile iron pool (LIP). We used as a model human prostate cancer cells, DU145. TNF treatment induced ROS formation, which was reduced to the control level in cells pretreated with desferrioxamine, an iron chelator. TNF induced a drop in light chain of the ferritin level, as judged by immunoblotting and an increase in LIP, evaluated by calcein fluorescence. Moreover, we observed that the JNK inhibitor SP600125 abolished TNF-induced changes in LIP, which suggests that JNK kinases are involved in this process. To explore which one of the JNK kinases is responsible for these effects, DU145 cells were transiently transfected with plasmids encoding inactive mutants of JNK1 or JNK2. The cells expressing inactive JNK1 mutant, but not cells expressing JNK2 mutant or possessing an empty vector, were completely resistant to TNF-induced ROS generation, ferritin degradation, and an increase in LIP. These data suggest that TNF-induced ROS formation is mediated by JNK1, which regulates ferritin degradation and thus the level of highly reactive iron. PMID:17603935

Antosiewicz, Jedrzej; Ziolkowski, Wieslaw; Kaczor, Jan Jacek; Herman-Antosiewicz, Anna



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

SciTech Connect

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.

Madonna, Rosalinda [The Center for Cardiovascular Biology and Atherosclerosis Research, The University of Texas Health Science Center at Houston, Texas (United States) [The Center for Cardiovascular Biology and Atherosclerosis Research, The University of Texas Health Science Center at Houston, Texas (United States); Institute of Cardiology, and Center of Excellence on Aging, 'G. d'Annunzio' University, Chieti (Italy); Shelat, Harnath; Xue, Qun; Willerson, James T. [The Center for Cardiovascular Biology and Atherosclerosis Research, The University of Texas Health Science Center at Houston, Texas (United States) [The Center for Cardiovascular Biology and Atherosclerosis Research, The University of Texas Health Science Center at Houston, Texas (United States); The Texas Heart Institute at St. Luke's Episcopal Hospital, Houston, Texas (United States); De Caterina, Raffaele [Institute of Cardiology, and Center of Excellence on Aging, 'G. d'Annunzio' University, Chieti (Italy)] [Institute of Cardiology, and Center of Excellence on Aging, 'G. d'Annunzio' University, Chieti (Italy); Geng, Yong-Jian, E-mail: [The Center for Cardiovascular Biology and Atherosclerosis Research, The University of Texas Health Science Center at Houston, Texas (United States) [The Center for Cardiovascular Biology and Atherosclerosis Research, The University of Texas Health Science Center at Houston, Texas (United States); The Texas Heart Institute at St. Luke's Episcopal Hospital, Houston, Texas (United States)



Fish nutrients decrease expression levels of tumor necrosis factor-alpha in cultured human macrophages.  


Numerous studies have demonstrated the beneficial effects of fish consumption on inflammatory markers. Until now, these beneficial effects of fish consumption have been mostly linked to the omega-3 fatty acids (FA). The objective of the present study was to examine, in vitro, whether expression levels of genes involved in the inflammatory response differ in human macrophages incubated with casein hydrolysates (CH) or fish protein hydrolysates (FPH) in the presence or absence of omega-3 FA compared with omega-3 FA alone. Peripheral blood monocytes differentiated into macrophages from 10 men were incubated in the presence of omega-3 FA (10 microM eicosapentaenoic acid and 5 microM docosahexaenoic acid) or CH or FPH (10, 100, 1,000 microg) with or without omega-3 FA for 48 h. Results demonstrate that expression levels of tumor necrosis factoralpha (TNFalpha) had a tendency to be lower after the addition of FPH alone or CH with omega-3 FA compared with omega-3 FA treatment. Furthermore, the combination of FPH and omega-3 FA synergistically decreased expression levels of TNFalpha compared to treatment with omega-3 FA or FPH alone. No difference on gene expression levels of interleukin-6 was observed between treatments. In conclusion, these preliminary results suggest that the anti-inflammatory effects of fish consumption can be explained by a synergistic effect of the omega-3 FA with the protein components of fish on TNFalpha expression and therefore contribute to the beneficial effects of fish consumption. Hence, follow-up studies should be performed to confirm the effects of a diet rich in FPH and omega-3 FA on serum proinflammatory cytokine concentrations. PMID:19952281

Rudkowska, Iwona; Marcotte, Bruno; Pilon, Genevičve; Lavigne, Charles; Marette, André; Vohl, Marie-Claude



Effect of Estrogen on Hypothalamic Transforming Growth Factor Alpha and Gonadotropin-Releasing Hormone Gene Expression in the Female Rhesus Monkey  

Microsoft Academic Search

In order to study whether hypothalamic transforming growth factor alpha (TGF?) gene expression in the monkey is estrogen-sensitive, long-term ovariectomized rhesus macaques were implanted subcutaneously with either estradiol-containing (n = 3) or blank (n = 3) Silastic capsules. Blood samples were collected every other day while the animals were lightly sedated with ketamine hydrochloride to monitor circulating LH and estradiol

Mohammed El Majdoubi; Abhiram Sahu; Tony M. Plant



Tumor necrosis factor-alpha modulates monocyte/macrophage apoprotein E gene expression.  

PubMed Central

apo E has been shown to modulate cholesterol balance in arterial wall cells. Production of apo E by macrophages in atherosclerotic plaques could thereby influence the development of the plaque lesion. Cytokines, including TNF alpha, have been identified in human lesions, therefore, we undertook a series of studies to evaluate the effect of TNF alpha on monocyte/macrophage apo E production. The addition of TNF alpha to freshly isolated human monocytes led to a four- to fivefold increase of apo E mRNA abundance. The addition of TNF alpha to fully differentiated macrophages either had no effect or modestly inhibited apo E mRNA expression. THP1 human monocytic cells also responded to TNF alpha in a phenotype-specific manner. Treatment of these cells with TNF alpha produced a dose- and time-dependent increase in apo E mRNA. This increase was reflected in apo E synthesis and was associated with inhibition of DNA synthesis, and with induction of c-fos and ICAM-1 gene expression. Cell-permanent analogues of ceramide did not reproduce TNF alpha effect on apo E, but antagonists of protein kinase C did inhibit its effect. TNF alpha induction of apo E mRNA abundance was associated with stimulation of apo E promoter-dependent gene transcription. In summary, TNF alpha stimulates apo E gene transcription, mRNA abundance, and protein synthesis in the monocyte/macrophage in a phenotype-specific manner. Such regulation could significantly modify the amount of apo E present in vessel wall lesions. Images

Duan, H; Li, Z; Mazzone, T



Tumor Necrosis Factor Alpha Transcription in Macrophages Is Attenuated by an Autocrine Factor That Preferentially Induces NF-?B p50  

PubMed Central

Macrophages are a major source of proinflammatory cytokines such as tumor necrosis factor alpha (TNF-?), which are expressed during conditions of inflammation, infection, or injury. We identified an activity secreted by a macrophage tumor cell line that negatively regulates bacterial lipopolysaccharide (LPS)-induced expression of TNF-?. This activity, termed TNF-?-inhibiting factor (TIF), suppressed the induction of TNF-? expression in macrophages, whereas induction of three other proinflammatory cytokines (interleukin-1? [IL-1?], IL-6, and monocyte chemoattractant protein 1) was accelerated or enhanced. A similar or identical inhibitory activity was secreted by IC-21 macrophages following LPS stimulation. Inhibition of TNF-? expression by macrophage conditioned medium was associated with selective induction of the NF-?B p50 subunit. Hyperinduction of p50 occurred with delayed kinetics in LPS-stimulated macrophages but not in fibroblasts. Overexpression of p50 blocked LPS-induced transcription from a TNF-? promoter reporter construct, showing that this transcription factor is an inhibitor of the TNF-? gene. Repression of the TNF-? promoter by TIF required a distal region that includes three NF-?B binding sites with preferential affinity for p50 homodimers. Thus, the selective repression of the TNF-? promoter by TIF may be explained by the specific binding of inhibitory p50 homodimers. We propose that TIF serves as a negative autocrine signal to attenuate TNF-? expression in activated macrophages. TIF is distinct from the known TNF-?-inhibiting factors IL-4, IL-10, and transforming growth factor ? and may represent a novel cytokine.

Baer, Mark; Dillner, Allan; Schwartz, Richard C.; Sedon, Constance; Nedospasov, Sergei; Johnson, Peter F.



Effects of tumor necrosis factor-alpha and interferon-gamma on expressions of matrix metalloproteinase-2 and -9 in human bladder cancer cells.  


We have investigated the effects of tumor necrosis factor-alpha (TNF-alpha) and interferon (INF-gamma), the potent Bacillus Calmette-Guerin (BCG)-induced cytokines on the production of MMP-2, MMP-9, TIMP-1, TIMP-2 and MT1-MMP in high grade human bladder cancer cell lines, T-24, J-82 and HT-1376 cell lines. MMP-2 expression and activity were decreased in T-24 cells treated with both cytokines in a dose dependent manner. However, J-82 cells treated with TNF-alpha and INF-gamma revealed dose dependent increases of MMP-9 expression and activity with similar baseline expression and activity of MMP-2. HT-1376 cells after exposure to TNF-alpha only enhanced the expression and activity of MMP-9. These results indicate that TNF-alpha and INF-gamma could regulate the production of MMP-2 or MMP-9 on bladder cancer cells and their patterns of regulation are cell specific. Furthermore, this diverse response of bladder cancer cells to TNF-alpha and INF-gamma suggests that BCG immunotherapy may enhance the invasiveness of bladder cancer in certain conditions with induction of MMPs. PMID:10996723

Shin, K Y; Moon, H S; Park, H Y; Lee, T Y; Woo, Y N; Kim, H J; Lee, S J; Kong, G



The Biphasic mRNA Expression Pattern of Bovine Interleukin-8 in Pasteurella haemolytica Lipopolysaccharide-Stimulated Alveolar Macrophages Is Primarily due to Tumor Necrosis Factor Alpha  

PubMed Central

Pasteurella haemolytica serotype 1 is the bacterial agent responsible for the pathophysiological events associated with bovine pneumonic pasteurellosis. Our previous studies support a role for the lipopolysaccharide (LPS) from P. haemolytica in the induction of proinflammatory cytokines. One of the pathological hallmarks of bovine pneumonic pasteurellosis is an influx of neutrophils into the alveolar spaces. This pronounced influx suggests the local production of a chemotactic factor(s) such as interleukin-8 (IL-8). In the context of the lung, the alveolar macrophage appears to be the major producer of IL-8, a proinflammatory cytokine with potent neutrophil chemotactic activity. By using Northern blot analysis, we have examined the kinetics of IL-8 mRNA expression in P. haemolytica LPS-stimulated bovine alveolar macrophages and found that 1 ng of LPS per ml induces maximal expression of IL-8 mRNA. The results also indicate a biphasic time course expression pattern in which IL-8 mRNA levels peak between 1 and 2 h in the first phase and between 16 and 24 h in the second phase (P < 0.01). In addition, monospecific polyclonal antibodies were used to demonstrate the role of tumor necrosis factor alpha (TNF-?) in the second phase of IL-8 mRNA expression. Our findings support a role for P. haemolytica LPS and TNF-? in the induction of IL-8 from bovine alveolar macrophages.

Lafleur, Rhonda L.; Abrahamsen, Mitchell S.; Maheswaran, Samuel K.



Dyscoordinate expression of tumor necrosis factor-alpha by human blood monocytes and alveolar macrophages.  


Previous studies have shown that human alveolar macrophages produce less interleukin-1 (IL-1) in response to lipopolysaccharide (LPS) than do their precursors, blood monocytes. The purpose of this study was to compare the capacities of alveolar macrophages and blood monocytes to synthesize tumor necrosis factor (TNF) in response to LPS. Alveolar macrophages were obtained by bronchoalveolar lavage of healthy nonsmoking subjects, and blood monocytes were obtained by adherence of mononuclear cells to plastic. TNF activity was measured in supernatants and cell lysates as cytotoxicity to L929 fibroblasts (uptake of neutral red at 570 nm). TNF activity of alveolar macrophages stimulated at 10(6) cells/ml with LPS (10 micrograms/ml) for 16 h was 596 +/- 367, and of blood monocytes it was 60 +/- 84 U/ml (mean +/- SD, p less than 0.005). At no concentration of LPS and at no period of stimulation did alveolar macrophages express less TNF activity than did blood monocytes. In concurrent experiments, supernatants of LPS-stimulated alveolar macrophages contained less IL-1 activity than did blood monocytes. Lysates of both cell types contained less than 20% of total TNF activity. The TNF activity of LPS-stimulated alveolar macrophages was neutralized greater than 99% by monoclonal antibody to TNF-alpha; control monoclonal antibody OKT3 had no effect. Next, alveolar macrophages and blood monocytes were biosynthetically labeled with [3H]leucine during incubation with LPS; supernatants were immunoprecipitated with anti-TNF, and precipitates were electrophoresed on polyacrylamide gels. Autoradiographs indicated that immunoreactive TNF was produced by both blood monocytes and alveolar macrophages and that the relative molecular weights were identical (17,000).(ABSTRACT TRUNCATED AT 250 WORDS) PMID:2784642

Rich, E A; Panuska, J R; Wallis, R S; Wolf, C B; Leonard, M L; Ellner, J J



Interleukin (IL)-6 mRNA expression is stimulated by insulin, isoproterenol, tumour necrosis factor alpha, growth hormone, and IL-6 in 3T3-L1 adipocytes.  


Interleukin (IL)-6 has recently been shown to be an adipocyte-expressed cytokine. Its serum concentrations are elevated in insulin resistance and obesity. For further evaluation of IL-6 gene expression regulation, fully differentiated 3T3-L1 adipocytes were treated with various hormones known to induce insulin resistance. IL-6 mRNA content was measured by quantitative real-time reverse transcription-polymerase chain reaction. Interestingly, treatment of adipocytes with 100 nM insulin, 10 micro M isoproterenol, 10 ng/ml tumour necrosis factor alpha (TNFalpha), and 500 ng/ml growth hormone (GH) for 16 h stimulated IL-6 mRNA expression 2.3-fold, 47-fold, 74-fold, and 1.4-fold, respectively (p < 0.01). In contrast, treatment with 100 nM dexamethasone significantly decreased IL-6 expression to 32 % of control levels (p < 0.01), whereas triiodothyronine and angiotensin 2 did not have any effect. Furthermore, stimulation of IL-6 expression was time-dependent with maximal stimulatory effects detectable after 1 h of insulin, isoproterenol, and GH addition and 12 h of TNFalpha, respectively. Moreover, isoproterenol's effect could be almost completely reversed by pretreatment of 3T3-L1 cells with the beta-adrenergic antagonist propranolol and mimicked by stimulation of G S -proteins with cholera toxin and adenylyl cyclase with forskolin and dibutyryl cAMP, respectively. Finally, IL-6 strongly induced its own expression in a time-dependent fashion. Taken together, our results demonstrate that IL-6 expression in adipocytes is governed by an autocrine positive feedback loop and upregulated by insulin, isoproterenol, TNFalpha, and GH. In concert with this adipocytokine's upregulation in states of decreased insulin sensitivity such as obesity and diabetes, the data support a possible role of IL-6 as a selectively regulated mediator of insulin resistance. PMID:12734774

Fasshauer, M; Klein, J; Lossner, U; Paschke, R



Ferulic acid enhances the vasorelaxant effect of epigallocatechin gallate in tumor necrosis factor-alpha-induced inflammatory rat aorta.  


Previously, we demonstrated synergistic enhancement of vasorelaxation by combination treatment with Trp-His and epigallocatechin gallate (EGCg) in intact rat aorta. The aim of the present study was to determine whether this vasorelaxant synergy could be recapitulated in tumor necrosis factor-alpha (TNF-?)-induced inflammatory rat aorta, and to determine the extent of its modulation by anti-inflammatory phenolic acids. Synergistic enhancement of vasorelaxation in rat aorta by Trp-His and EGCg was significantly attenuated in the presence of TNF-?, an effect that was reversed by the addition of ferulic acid (FA, 250 ?M). Moreover, FA markedly enhanced EGCg-induced vasorelaxation, but not Trp-His-induced vasorelaxation, in TNF-?-treated aorta. Structure-activity analysis showed that the unsaturated 2-propenoic moiety and the methoxy group of FA were important for the enhancement of vasorelaxation by EGCg. The stimulation of EGCg-induced vasorelaxation by FA was antagonized by the nitric oxide synthase inhibitor N(G)-monomethyl-l-arginine acetate, while FA enhanced vasorelaxant properties of the endothelial nitric oxide (NO) synthase activator acetylcholine in TNF-?-treated inflammatory aorta. Moreover, the EGCg-stimulated NO production was also enhanced by FA in TNF-?-treated aorta. These data indicate that stimulation of NO production by FA enhances the vasorelaxant properties of EGCg in TNF-?-induced inflammatory aorta. PMID:24794014

Zhao, Jian; Suyama, Aki; Tanaka, Mitsuru; Matsui, Toshiro



Adenosine inhibits platelet-activating factor, but not tumour necrosis factor-alpha-induced priming of human neutrophils.  

PubMed Central

Regulation of the respiratory burst and its priming by recombinant human tumour necrosis factor-alpha (rhTNF-alpha) and platelet-activating factor (PAF) were investigated in human polymorphonuclear leucocytes (PMN). Adenosine (0.1-10 microM) pretreatment of PMN concentration-dependently inhibited the superoxide anion generation (O2-) in response to formyl-methionyl-leucyl-phenylalanine (FMLP). The priming by PAF (1 microM) for an increased O2- generation by FMLP-stimulated PMN was completely blocked by adenosine pretreatment. In contrast, rhTNF-alpha-induced priming was unaffected by adenosine. In addition, the direct stimulation of PMN O2- by rhTNF-alpha was also unaffected by adenosine as was rhTNF-alpha-induced PAF synthesis. FMLP-induced PAF synthesis was reduced by adenosine to a similar extent as the inhibition of the respiratory burst. Adenosine also inhibited PAF-, but not FMLP-induced increases in intracellular calcium in PMN. These findings indicate that short-term, direct stimulants (FMLP) or priming agents (PAF) are subject to modulation by the endothelial product adenosine, whereas the priming and direct stimulation of the respiratory burst by the longer-acting agent, rhTNF-alpha is unaffected. Moreover, differential inhibition of PMN activation by adenosine reveals important functional differences in the signalling mechanisms initiated by PAF, FMLP and rhTNF-alpha.

Stewart, A G; Harris, T



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

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.

Miettinen, Johanna A., E-mail: [Institute of Clinical Medicine, Department of Internal Medicine, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland); Pietilae, Mika [Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland)] [Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland); Salonen, Riikka J. [Institute of Clinical Medicine, Department of Internal Medicine, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland) [Institute of Clinical Medicine, Department of Internal Medicine, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland); Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland); Ohlmeier, Steffen [Proteomics Core Facility, Biocenter Oulu, Department of Biochemistry, University of Oulu, P.O. Box 3000, FIN-90014 Oulu (Finland)] [Proteomics Core Facility, Biocenter Oulu, Department of Biochemistry, University of Oulu, P.O. Box 3000, FIN-90014 Oulu (Finland); Ylitalo, Kari; Huikuri, Heikki V. [Institute of Clinical Medicine, Department of Internal Medicine, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland)] [Institute of Clinical Medicine, Department of Internal Medicine, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland); Lehenkari, Petri [Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland)] [Institute of Biomedicine, Department of Anatomy and Cell Biology, University of Oulu, P.O. Box 5000, FIN-90014 Oulu (Finland)



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

PubMed Central

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-?). 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-? was a key contributor to apoptosis, since (i) culture supernatants from cells infected with gonococci (MOI = 1) induced apoptosis in naďve cultures, suggesting that a soluble factor was responsible; (ii) gonococcal infection-induced apoptosis was inhibited with anti-TNF-? antibodies; and (iii) the addition of exogenous TNF-? induced apoptosis, which was inhibited by the presence of increasing numbers of bacteria (MOI = 10 to 100). These data suggest that TNF-?-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-?-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.

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



Lipoteichoic Acids from Lactobacillus Strains Elicit Strong Tumor Necrosis Factor Alpha-Inducing Activities in Macrophages through Toll-Like Receptor 2  

Microsoft Academic Search

Lactobacilli are nonpathogenic gram-positive inhabitants of microflora. At least some Lactobacillus strains have been postulated to have health beneficial effects, such as the stimulation of the immune system. Here we examined the stimulatory effects of lactobacilli on mouse immune cells. All six heat-killed Lactobacillus strains examined induced the secretion of tumor necrosis factor alpha (TNF-) from mouse splenic mononuclear cells,

Tetsuya Matsuguchi; Akimitsu Takagi; Takeshi Matsuzaki; Masato Nagaoka; Kimika Ishikawa; Teruo Yokokura; Yasunobu Yoshikai



Tumor Necrosis Factor alpha induced hypoexcitability in rat muscle evidenced in a model of ion currents and action potential.  


Sepsis and Tumor Necrosis Factor alpha (TNF?), a major pro-inflammatory mediator, have previously been shown to induce a decrease in the conductance of voltage-dependent sodium channels (NaV). Moreover, TNF? increased resting membrane potential, leading to hyperpolarization. NaV and resting potential are the two major factors of membrane excitability. Then we hypothesis that TNF? can decrease muscle membrane excitability. To evidence that role of TNF?, we carried out a simulation of the sodium and potassium currents and action potential (AP) of isolated muscle fibre. We used a computer model based on Hodgkin and Huxley equations, but also taking into account the sodium-potassium pump current. Our first aim was to optimise this model in control conditions according to our measurements of currents. Then the model was modified to fit the values measured experimentally in TNF?-containing medium in order to determine the modifications induced in the currents and hence in AP triggering. Our model provides a very good fit with experimental data on the ion currents. Moreover, it clearly shows that the triggering level of AP is increased in TNF?-containing medium, thus corresponding to a decreased excitability. PMID:23911204

Guillouët, Maďté; Rannou, Fabrice; Giroux-Metges, Marie-Agnčs; Droguet, Mickael; Pennec, Jean-Pierre



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

PubMed Central

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 node status (i.e. metastasis). These results suggest that tumour cell responsiveness to TNF-alpha produced by neighbouring TAMs may play a part in the regulation of TP expression by tumour cells as well as their metastatic behaviour. This may explain, in part, the relationship between increased macrophage infiltration and angiogenesis in breast cancer, and further supports the contention that TAMs may represent an important target for future anti-angiogenic therapies. Images Figure 1

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



Plasmodium Falciparum Pigment Induces Monocytes to Release High Levels of Tumor Necrosis Factor-Alpha and Interleukin-1Beta.  

National Technical Information Service (NTIS)

We show that high levels of tumor necrosis factor-alpha (TNF-alpha) activity were consistently detected when monocytes were cocultured with Plasmodium falciparum schizont stage-parasitized erythrocytes that subsequently ruptured. Isolated pigment recovere...

S. Pichyangkul P. Saengrai H. K. Webster



Virulence factor p60 of Listeria monocytogenes modulates innate immunity by inducing tumor necrosis factor alpha.  


We investigated the effect of p60, a virulence factor of Listeria monocytogenes, on host immune response in vitro and in vivo. Administration of p60 before a sublethal infection with L. monocytogenes enhanced innate host resistance in naďve mice. Mouse macrophage RAW264.7 cells produced tumor necrosis factor (TNF)-alpha in response to stimulation with recombinant p60. Toll-like receptor 4 may be involved in TNF-alpha production from RAW264.7 cells and enhanced host resistance induced by p60 administration. Our findings demonstrated that p60 modulates innate immune responses against L. monocytogenes infection. PMID:20337701

Sashinami, Hiroshi; Hu, Dong-Liang; Li, Sheng-Jun; Mitsui, Toshihito; Hakamada, Ken-Ichi; Ishiguro, Yoh; Fukuda, Shinsaku; Nakane, Akio



Systemic lupus erythematosus induced by anti-tumour necrosis factor alpha therapy: a French national survey.  


The development of drug-induced lupus remains a matter of concern in patients treated with anti-tumour necrosis factor (TNF) alpha. The incidence of such adverse effects is unknown. We undertook a retrospective national study to analyse such patients. Between June and October 2003, 866 rheumatology and internal medicine practitioners from all French hospital centres prescribing anti-TNF in rheumatic diseases registered on the website of the 'Club Rhumatismes et Inflammation' were contacted by email to obtain the files of patients with TNF-induced systemic lupus erythematosus. Twenty-two cases were collected, revealing two aspects of these manifestations. Ten patients (six patients receiving infliximab, four patients receiving etanercept) only had anti-DNA antibodies and skin manifestations one could classify as 'limited skin lupus' or 'toxidermia' in a context of autoimmunity, whereas 12 patients (nine patients receiving infliximab, three patients receiving etanercept) had more complete drug-induced lupus with systemic manifestations and at least four American Congress of Rheumatology criteria. One patient had central nervous system manifestations. No patients had lupus nephritis. The signs of lupus occurred within a mean of 9 months (range 3-16 months) in patients treated with infliximab and within a mean of 4 months (range 2-5 months) in patients treated with etanercept. In all cases after diagnosis was determined, anti-TNF was stopped and specific treatment introduced in eight patients: two patients received intravenous methylprednisolone, four patients received oral steroids (15-35 mg/day), and two patients received topical steroids. Lupus manifestations abated within a few weeks (median 8 weeks, standard deviation 3-16) in all patients except one with longer-lasting evolution (6 months). At that time, cautious estimations (unpublished data from Schering Plough Inc. and Wyeth Inc.) indicated that about 7700 patients had been exposed to infliximab and 3000 to etanercept for inflammatory arthritides in France. It thus appears that no drug was more implicated than the other in lupus syndromes, whose incidence was 15/7700 = 0.19% with infliximab and 7/3800 = 0.18% with etanercept. Clinicians should be aware that lupus syndromes with systemic manifestations may occur in patients under anti-TNF alpha treatment. PMID:15899041

De Bandt, Michel; Sibilia, Jean; Le Loët, Xavier; Prouzeau, Sebastian; Fautrel, Bruno; Marcelli, Christian; Boucquillard, Eric; Siame, Jean Louis; Mariette, Xavier



[Investigation of interleukin-10, tumor necrosis factor-alpha and interferon-gamma expression in experimental model of pulmonary aspergillosis].  


Pulmonary aspergillosis which is an important opportunistic infection in neutropenic patients, is usually caused by Aspergillus fumigatus. Since the pathogenesis of disease is not well understood, the main proposed mechanism is thought to be cell-mediated immunity and cytokine response. The aim of this study was to investigate the local production of cytokines in the lung tissues of rats with experimentally developed aspergillosis, by reverse transcriptase-polymerase chain reaction (RT-PCR). A total of 33 Wistar albino type rats were included in the study with the consent of Experimental Animal Ethics Committee. Twenty-five of the rats were infected with A.fumigatus by intratracheal way, while 8 animals were used as controls. The presence of A.fumigatus in the lung tissues of infected rats was confirmed with the use of quantitative culture and histologic staining methods. RNA isolation from the lung tissue samples of both groups were performed by a commercial kit (Qiagen, Germany). After obtaining complementary DNAs from the genomic RNAs, in-house qualitative and quantitative (real-time) PCR methods were used to amplify the target regions for interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-?) and interferon-gamma (IFN-?) by using specific primers (T?b Molbiol, Germany). Mean mRNA levels achieved by real-time PCR for IL-10, TNF-? and IFN-? in aspergillosis group were 6.5 x 106 copies/ml, 7.9 x 105 copies/ml and 2.2 x 103 copies/ml, respectively, while those values in control group were 4.3 x 102 copies/ml, 5.6 x 103 copies/ml and 1.3 x 102 copies/ml, respectively. Our data indicated that rat model of aspergillosis was associated with significantly increased expression of mRNA encoding IL-10 and TNF-? than controls (p< 0.05), however there was no statistically significant difference between the groups with respect to IFN-? expression (p= 0.53). In conclusion, the production of proinflammatory cytokines which mediate the influx of phagocytic cells might account for the localization of Aspergillus infection to the upper respiratory tract. The up-regulation of the expression of the immunomodulatory cytokine TNF-? and IL-10 in lung tissue from infected rats might be important to limit the extent of local tissue destruction, but might also account for the fact that infected rats are generally unable to clear the infection spontaneously. PMID:21644078

Ca?lar, Kayhan; Kalkanc?, Ay?e; Fidan, I??l; Aydo?an, Sibel; H?zel, Kenan; Dizbay, Murat; Poyraz, Aylar; Ku?timur, Semra



Tumor Necrosis Factor-Alpha Induced by Hepatitis B Virus Core Mediating the Immune Response for Hepatitis B Viral Clearance in Mice Model  

PubMed Central

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-?/? 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-?) 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-? 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-? production induced by HBcAg. These results provide evidences for TNF-? mediated innate immune mechanisms in HBV clearance and explain the mechanism of HBV reactivation during therapy with TNF blockage agents.

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



Interferon-gamma potentiates interleukin (IL)-6 and tumor necrosis factor-alpha but not IL-1beta induced by endotoxin in the brain.  


Because interferon-gamma (IFN gamma) is present in the central nervous system during neurologic diseases associated with inflammation, its effect on endotoxin-induced cytokines was studied. Cerebrospinal fluid (CSF) and serum levels of interleukin (IL)-1beta, IL-6, and tumor necrosis factor-alpha (TNF alpha), their messenger RNA expression in brain areas (hypothalamus, hippocampus, and striatum) and in spleen were evaluated 2 and 8 h after endotoxin [lipopolysaccharide (LPS), 25 microg/rat i.c.v.], IFN gamma (2.5 microg/rat i.c.v.) or after their coadministration in rats. CSF and serum IL-1beta levels were increased by LPS alone and IFN gamma coadministration did not furtherly increase them. IFN gamma potentiated LPS effect on IL-6 and TNF alpha levels in both CSF and serum. LPS and IFN-gamma coadministration did not alter IL-1beta messenger RNA expression induced by LPS in brain areas and in spleen, but it potentiated that of IL-6 and TNF alpha. The present in vivo data show that i.c.v. coadministration of LPS and IFN gamma results in a potentiation of cytokine production (IL-6 and TNF alpha) which may trigger a cascade of events relevant to neurodegenerative processes. This action is independent of IL-1beta because the production of this cytokine is not altered by IFN gamma treatment. PMID:9389504

De Simoni, M G; Terreni, L; Chiesa, R; Mangiarotti, F; Forloni, G L



Amiodarone Exposure During Modest Inflammation Induces Idiosyncrasy-like Liver Injury in Rats: Role of Tumor Necrosis Factor-alpha  

PubMed Central

Amiodarone [2-butyl-3-(3?,5?-diiodo-4’?-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.

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



Mycobacterium marinum SecA2 Promotes Stable Granulomas and Induces Tumor Necrosis Factor Alpha In Vivo  

PubMed Central

SecA2 is an ATPase present in some pathogenic Gram-positive bacteria, is required for translocation of a limited set of proteins across the cytosolic membrane, and plays an important role in virulence in several bacteria, including mycobacteria that cause diseases such as tuberculosis and leprosy. However, the mechanisms by which SecA2 affects virulence are incompletely understood. To investigate whether SecA2 modulates host immune responses in vivo, we studied Mycobacterium marinum infection in two different hosts: an established zebrafish model and a recently described mouse model. Here we show that M. marinum ?secA2 was attenuated for virulence in both host species and SecA2 was needed for normal granuloma numbers and for optimal tumor necrosis factor alpha response in both zebrafish and mice. M. marinum ?secA2 was more sensitive to SDS and had unique protrusions from its cell envelope when examined by cryo-electron tomography, suggesting that SecA2 is important for bacterial cell wall integrity. These results provide evidence that SecA2 induces granulomas and is required for bacterial modulation of the host response because it affects the mycobacterial cell envelope.

Watkins, Brigitte Y.; Joshi, Shilpa A.; Ball, David A.; Leggett, Hadley; Park, Summer; Kim, Janice; Austin, Cary D.; Paler-Martinez, Andres; Xu, Min; Downing, Kenneth H.



Upregulation of tumor necrosis factor-alpha in nucleus accumbens attenuates morphine-induced rewarding in a neuropathic pain model.  


Treatment of neuropathic pain with opioid analgesics remains controversial and a major concern is the risk of addiction. Here, we investigated this issue with spared nerve injury (SNI) model of neuropathic pain in rats and mice. SNI prevented conditioned place preference (CPP) induced by low dose (3.5mg/kg) of morphine (MOR), which was effective for anti-allodynia, but not by high dose (?5.0mg/kg) of MOR. Tumor necrosis factor-alpha (TNF-?) was upregulated in nucleus accumbens (NAcc) following SNI. The inhibitory effect of SNI on MOR-induced CPP was blocked by either genetic deletion of TNF receptor 1 (TNFR1) or microinjection of anti-TNF-? into the NAcc and was mimicked by intra-NAcc injection of TNF-? in sham rats. Furthermore, SNI reduced dopamine (DA) level and upregulated dopamine transporter (DAT) in the NAcc, but did not affect total tyrosine hydroxylase (TH) or phospho-TH (p-TH), a rate-limiting enzyme of catecholamine biosynthesis, in ventral tegmental area (VTA). Accordingly, the increase in DA reuptake but not decrease in its synthesis may lead to the reduction of DA level. Finally, the upregulation of DAT in the NAcc of SNI animals was again blocked by either genetic deletion of TNFR1 or NAcc injection of anti-TNF-?, and was mimicked by NAcc injection of TNF-? in sham animals. Thus, our data provided novel evidence that upregulation of TNF-? in NAcc may attenuate MOR-induced rewarding by upregulation of DAT in NAcc under neuropathic pain condition. PMID:24845379

Wu, Ying; Na, Xiaodong; Zang, Ying; Cui, Yu; Xin, Wenjun; Pang, Ruiping; Zhou, Lijun; Wei, Xuhong; Li, Yongyong; Liu, Xianguo



Bovine endometrial stromal cells support tumor necrosis factor alpha-induced bovine herpesvirus type 4 enhanced replication.  


Bovine uterine infections are the most important cause of economic losses in the cattle industry. Although the etiology of uterine diseases is mainly ascribed to bacterial infection, they can also be associated with viral infection, such as bovine herpesvirus 4 (BoHV-4), which is often a secondary agent following bacteria. Besides microbial infection, many inflammatory molecules belonging to the innate immune response orchestrate the outcome of the infection. In the present study, the interaction between BoHV-4-infected bovine endometrial stromal cells and tumor necrosis factor alpha (TNF-alpha) was investigated. Bovine herpesvirus 4 possesses a special tropism toward endometrial stromal cells. For this reason, a simian virus 40 (SV40) immortalized endometrial stromal cell line (SV40BESC) was established; it was proven that it was stable, it expressed toll-like receptors (TLRs; from 1 to 10) and TNF-alpha receptors I and II, and it was responsive to exogenous TNF-alpha. Further, an increase of BoHV-4 replication and cytopathic effect was observed in BoHV-4-infected and TNF-alpha-treated SV40BESCs. This increase of viral replication was associated with BoHV-4 immediate early 2 (IE2) gene promoter trans-activation through the interaction of the nuclear factor KB (NFKB) with the putative NFKB-responsive elements found within BoHV-4 IE2 gene promoter, and this interaction was abolished when NFKB-responsive elements were deleted. These data shed light on two important and rather controversial issues: the role of TNF-alpha receptor, which is weakly expressed in the stromal layer of the bovine uterus, as well as the possible interactions between proinflammatory molecules, viral replication, and chronic uterine disease. PMID:23515672

Jacca, Sarah; Franceschi, Valentina; Colagiorgi, Angelo; Sheldon, Martin; Donofrio, Gaetano



Down-regulation of integrins by von Hippel-Lindau (VHL) tumor suppressor protein is independent of VHL directed hypoxia-inducible factor alpha degradation  

PubMed Central

Inactivation of the von Hippel-Lindau (VHL) tumor suppressor gene occurs in the majority of clear-cell renal cell carcinomas (RCC). It was previously shown that VHL decreased the abundance of integrin ?2, ?5 and ?1 – consistent with VHL-associated changes in cell-cell and cell-extracellular matrix adhesions. We investigated the mechanism by which VHL down-regulates integrins. Although VHL can target hypoxia-inducible factor alpha (HIF?) subunits for degradation, VHL-dependent reduction of integrins was independent of O2 concentration and HIF? levels. VHL reduced the half-lives of integrins and this activity was blocked by proteasomal inhibition. Ectopic expression of Flag-VHL, while retaining activity for HIF? degradation, neither down-regulated integrins nor promoted adherens and tight intercellular junctions, in contrast to expression of wild-type VHL. Moreover, integrins co-immunoprecipitaed with wild type VHL, but not Flag-VHL. These data indicate that down-regulation of integrins by VHL is distinct from VHL’s regulation of HIF ? subunits, and suggests that loss of this activity contributes to VHL-associated RCC development through disruption of adherens and tight junctions.

Ji, Qingzhou; Burk, Robert D.



Involvement of tumour necrosis factor-alpha-related apoptosis-inducing ligand in enhanced cytotoxicity of lipopolysaccharide-stimulated dendritic cells to activated T cells.  


Dendritic cells (DC) are potent antigen-presenting cells (APC) specialized in T-cell mediated immune responses, and also play critical roles in the homeostasis of T cells for controlling immune responses. In the present study, we demonstrated that during mouse bone-marrow-derived DC activation of ovalbumin (OVA)-specific Ia-kb-restricted T hybridoma cells, MF2.2D9 and OVA257-264-specific H-2kb-restricted RF33.70 T cells, respectively, both hybridomas undergo cell death, partially mediated via apoptotic ligand-tumour necrosis factor-alpha (TNF-alpha)-related apoptosis-inducing ligand (TRAIL). Lipopolysaccharide enhanced the cytotoxic effect on the two activated T hybridoma cells, which was correlated with up-regulation of TRAIL-expression on DC to some extent. The activation of caspase-3 in activated T hybridoma cells cocultured with DC contributed to the programmed cell death pathway T cells underwent. Therefore, our results show that activation-induced cell death of T hybridoma cells can be influenced by DC, suggesting that DC may be involved in elimination of activated T cells at the end of primary immune responses. PMID:12100718

Yu, Yizhi; Liu, Shuxun; Wang, Wenya; Song, Wengang; Zhang, Minghui; Zhang, Weiping; Qin, Zhihai; Cao, Xuetao



Enhancement by tumor necrosis factor alpha of dengue virus-induced endothelial cell production of reactive nitrogen and oxygen species is key to hemorrhage development.  


Hemorrhage is a severe manifestation of dengue disease. Virus strain and host immune response have been implicated as the risk factors for hemorrhage development. To delineate the complex interplay between the virus and the host, we established a dengue hemorrhage model in immune-competent mice. Mice inoculated intradermally with dengue virus develop hemorrhage within 3 days. In the present study, we showed by the presence of NS1 antigen and viral nuclei acid that dengue virus actively infects the endothelium at 12 h and 24 h after inoculation. Temporal studies showed that beginning at day 2, there was macrophage infiltration into the vicinity of the endothelium, increased tumor necrosis factor alpha (TNF-alpha) production, and endothelial cell apoptosis in the tissues. In the meantime, endothelial cells in the hemorrhage tissues expressed inducible nitric oxide synthase (iNOS) and nitrotyrosine. In vitro studies showed that primary mouse and human endothelial cells were productively infected by dengue virus. Infection by dengue virus induced endothelial cell production of reactive nitrogen and oxygen species and apoptotic cell death, which was greatly enhanced by TNF-alpha. N(G)-nitro-L-arginine methyl ester and N-acetyl cysteine reversed the effects of dengue virus and TNF-alpha on endothelial cells. Importantly, hemorrhage development and the severity of hemorrhage were greatly reduced in mice lacking iNOS or p47(phox) or treatment with oxidase inhibitor, pointing to the critical roles of reactive nitrogen and oxygen species in dengue hemorrhage. PMID:18842737

Yen, Yu-Ting; Chen, Hsuen-Chin; Chen, Hseun-Chin; Lin, Yang-Ding; Shieh, Chi-Chang; Wu-Hsieh, Betty A



Enhancement by Tumor Necrosis Factor Alpha of Dengue Virus-Induced Endothelial Cell Production of Reactive Nitrogen and Oxygen Species Is Key to Hemorrhage Development?  

PubMed Central

Hemorrhage is a severe manifestation of dengue disease. Virus strain and host immune response have been implicated as the risk factors for hemorrhage development. To delineate the complex interplay between the virus and the host, we established a dengue hemorrhage model in immune-competent mice. Mice inoculated intradermally with dengue virus develop hemorrhage within 3 days. In the present study, we showed by the presence of NS1 antigen and viral nuclei acid that dengue virus actively infects the endothelium at 12 h and 24 h after inoculation. Temporal studies showed that beginning at day 2, there was macrophage infiltration into the vicinity of the endothelium, increased tumor necrosis factor alpha (TNF-?) production, and endothelial cell apoptosis in the tissues. In the meantime, endothelial cells in the hemorrhage tissues expressed inducible nitric oxide synthase (iNOS) and nitrotyrosine. In vitro studies showed that primary mouse and human endothelial cells were productively infected by dengue virus. Infection by dengue virus induced endothelial cell production of reactive nitrogen and oxygen species and apoptotic cell death, which was greatly enhanced by TNF-?. NG-Nitro-l-arginine methyl ester and N-acetyl cysteine reversed the effects of dengue virus and TNF-? on endothelial cells. Importantly, hemorrhage development and the severity of hemorrhage were greatly reduced in mice lacking iNOS or p47phox or treatment with oxidase inhibitor, pointing to the critical roles of reactive nitrogen and oxygen species in dengue hemorrhage.

Yen, Yu-Ting; Chen, Hseun-Chin; Lin, Yang-Ding; Shieh, Chi-Chang; Wu-Hsieh, Betty A.



Cell-type-specific expression of the platelet-derived growth factor alpha receptor: a role for GATA-binding protein.  

PubMed Central

Platelet-derived growth factor alpha receptor (PDGF alpha R) is a transmembrane tyrosine kinase receptor for all three existing PDGF isoforms, AA, AB, and BB. Transcripts of PDGF alpha R are detected as early as in fertilized mouse eggs and throughout adulthood in a time- and space-specific manner, thereby suggesting an important role of PDGFs in mammalian development. In this study, we have investigated the mechanism involved in cell-type-specific PDGF alpha R gene expression during early embryonic development. Using F9 embryonic carcinoma cells as an in vitro study model, we identified a differentiation-dependent enhancer element within the PDGF alpha R promoter that controlled receptor expression during parietal endoderm cell differentiation induced by retinoic acid and dibutyryl cyclic AMP treatment. The differentiation-dependent enhancer element sequence bore no resemblance to consensus DNA-binding sites of either the retinoic acid receptor family or the cyclic AMP-responsive element-binding protein family. It was composed of two identical 12-bp direct repeats separated by a 17-bp insert sequence enriched in C and A nucleotides. Although only a single repeat was needed to form specific DNA-protein complexes with factors present in F9 parietal endoderm cell extracts, both repeats together were necessary to display cell-type-specific enhancing activity. Mutational analysis revealed that the protein-binding sites within the repeat sequences were identical to GATA-binding sites. In this study, we provided evidence to suggest that a member of the GATA transcription factor family (GATA-4) is responsible for parietal endoderm-specific PDGF alpha R expression.

Wang, C; Song, B



Modulation of adhesion molecule expression on endothelial cells during the late asthmatic reaction: role of macrophage-derived tumour necrosis factor-alpha.  

PubMed Central

In a previous work we have demonstrated that in patients exhibiting a late allergic reaction (LAR), alveolar macrophages (AM) collected 18 h after bronchial allergen challenge produced high levels of IL-6 and tumour necrosis factor-alpha (TNF) which is known to up-regulate the endothelial cell expression of adhesion molecules participating in the development of the inflammatory reaction in bronchial asthma. For these reasons, we evaluated the effect of AM supernatants from asthmatic patients developing an LAR on intercellular adhesion molecule-1 (ICAM-1) and endothelial leucocyte adhesion molecule-1 (ELAM-1) expression by human endothelial cells. The expression of adhesion molecules was assessed by an ELISA method and compared with the effect of an optimal dose of human recombinant (hr) TNF. Results showed that AM supernatants, from challenged asthmatics developing an LAR, increased significantly the ICAM-1 and ELAM-1 expression on endothelial cells to a level similar to that obtained in the presence of hrTNF (500 U/ml) (P < 0.001 in both cases, respectively 90.4% and 75.2% of the level obtained with hrTNF). In contrast, AM supernatants from asthmatics at baseline or exhibiting, after challenge, a single early reaction had no significant effect on these parameters (P = NS in both cases, respectively 23.5% and 24.7% of the ICAM-1 expression, 22.7% and 15.3% of the ELAM-1 expression obtained with hrTNF). AM-derived TNF present in these supernatants was thought to play a key role in endothelial cell stimulation, since: (i) TNF concentration in AM supernatants correlated with its effect on ICAM-1 (r = 0.80, P < 10(-4)) and ELAM-1 expression (r = 0.88, P < 10(-5)); and (ii) a neutralizing anti-TNF antibody decreased their effect (68% and 80% respectively on ICAM-1 and ELAM-1 expression). Moreover, the role of IL-6 was excluded on the basis both of the hrIL-6 inefficiency to induce ICAM-1 and ELAM-1 synthesis, even in costimulation with hrTNF, and of anti-IL-6 antibody to neutralize the effect of AM supernatants. Our results suggest that, beside mast cells and lymphocytes, macrophages might participate in the induction of the local inflammatory reaction observed in bronchial asthma. During the LAR, cytokines and especially TNF are able, through an enhanced adhesion molecule expression on endothelial cells, to facilitate the bronchial cellular influx.

Lassalle, P; Gosset, P; Delneste, Y; Tsicopoulos, A; Capron, A; Joseph, M; Tonnel, A B



Low-molecular-weight hyaluronic acid induces nuclear factor-kappaB-dependent resistance against tumor necrosis factor alpha-mediated liver injury in mice.  


Liver resident NK1.1+ T cells are supposed to play a pivotal role in the onset of inflammatory liver injury in experimental mouse models such as concanavalin A (Con A)-induced hepatitis. These cells, expressing the adhesion receptor, CD44, are largely depleted from the liver by a single intravenous injection of low-molecular-weight fragments of hyaluronic acid (LMW-HA). Here, we report that LMW-HA pretreatment protected mice from liver injury in several models of T-cell- and macrophage-dependent, tumor necrosis factor alpha (TNF-alpha)-mediated inflammatory liver injury, i.e., from liver injury induced by either Con A or Pseudomonas exotoxin A (PEA) or PEA/lipopolysaccharide (LPS). Interestingly, apart from inhibition of cellular adhesion, pretreatment of mice with LMW-HA was also capable of preventing hepatocellular apoptosis and activation of caspase-3 induced by direct administration of recombinant murine (rmu) TNF-alpha to D-galactosamine (GalN)-sensitized mice. LMW-HA-induced hepatoprotection could be neutralized by pretreatment with the nuclear factor-kappaB (NF-kappaB) inhibitor, pyrrolidine dithiocarbamate (PDTC), demonstrating the involvement of NF-kappaB in the observed protective mechanism. Indeed, injection of LMW-HA rapidly induced the production of TNF-alpha by Kupffer cells and the translocation of NF-kappaB into hepatocellular nuclei. Both LMW-HA-induced TNF-alpha production and NF-kappaB translocation were blocked by pretreatment with PDTC. Our findings provide evidence for an unknown mechanism of LMW-HA-dependent protection from inflammatory liver disease, i.e., induction of TNF-alpha- and NF-kappaB-dependent cytoprotective proteins within the target parenchymal liver cells. PMID:11526540

Wolf, D; Schümann, J; Koerber, K; Kiemer, A K; Vollmar, A M; Sass, G; Papadopoulos, T; Bang, R; Klein, S D; Brüne, B; Tiegs, G



Molecular cloning and expression analysis of tumour necrosis factor-alpha in amoebic gill disease (AGD)-affected Atlantic salmon (Salmo salar L.).  


Tumour necrosis factor-alpha (TNF-alpha) is a key mediator of inflammation during amoebiasis of humans and mice. Atlantic salmon (Salmo salar L.) are also susceptible to infection by amoebae (Neoparamoeba spp.), inflicting a condition known as amoebic gill disease (AGD). Here, the role of TNF-alpha in AGD-pathogenesis was examined. Two Atlantic salmon TNF-alpha transcripts designated TNF-alpha1 and TNF-alpha2 together with their respective genes were cloned and sequenced. TNF-alpha1 is 1379 bp and consists of a 738 bp open reading frame (ORF) translating into a predicted protein of 246 amino acids. TNF-alpha2 is 1412 bp containing an ORF and translated protein the same lengths as TNF-alpha1. An anti-rainbow trout TNF-alpha polyclonal antibody that bound recombinant Atlantic salmon TNF-alpha1 and TNF-alpha2 was used to detect constitutive and inducible expression of TNF-alpha in various tissues. The anti-TNF-alpha antibody bound to a TNF-like protein approximately 60 kDa that was constitutively expressed in a number of tissues in healthy Atlantic salmon. However, this protein was not detected in lysates from mitogen-stimulated head kidney leucocytes, despite up-regulation of TNF-alpha mRNAs under the same conditions. During the early onset of AGD in Atlantic salmon, there were no demonstrable differences in the gill tissue expression of TNF-alpha1, TNF-alpha2 nor the interleukin-1 beta (IL-1beta), inducible nitric oxide synthase (iNOS) and interferon gamma (IFN-gamma) mRNAs compared to tissue from healthy fish. In Atlantic salmon with advanced AGD, IL-1beta but not TNF-alpha1 or TNF-alpha2 mRNAs was up-regulated and was lesion-restricted. Given that Neoparamoeba spp. modulated both TNF-alpha2 and IL-1beta in head kidney leucocytes in vitro, it appears that rather than being recalcitrant to Neoparamoeba spp.-mediated TNF-alpha expression, either the parasite can influence the cytokine response during infection, there is ineffective signalling for TNF-alpha expression, or there are too few cells at the site of infection with the capacity to produce TNF-alpha. These data support our previous observation that IL-1beta mRNA expression is up-regulated in AGD-affected tissue and that TNF-alpha is not intrinsic in AGD-pathogenesis. PMID:17566761

Morrison, R N; Zou, J; Secombes, C J; Scapigliati, G; Adams, M B; Nowak, B F



Protective role of normothermic, hyperthermic and estrogen preconditioning and pretreatment on tumour necrosis factor-alpha-induced damage  

PubMed Central

BACKGROUND: Tumour necrosis factor-alpha (TNF-?) has been reported to play an important role in ischemia reperfusion injury and ischemic preconditioning (IPC). However, its role is not completely understood. Recently, normothermic IPC (NIPC), hyperthermic IPC (HIPC), preconditioning (PC) with 17-beta estradiol (estrogen, E2) and E2 pretreatment were proven to be effective in reducing ischemia reperfusion injury. OBJECTIVES: To investigate the detrimental effects of TNF-? on the heart, and the protective effects of NIPC, HIPC, E2 PC and pretreatment on TNF-?-induced injury. METHODS: A Langendorff-perfused rat heart model was used for the present study. Hearts isolated from male rats were studied under eight different conditions (n=5 each): negative control; control treated with TNF-? without any further treatment; NIPC (preconditioned at 37°C); HIPC (preconditioned at 42°C); E2 PC; E2 pretreatment; normal, untreated hearts plus E2; or pretreated hearts perfused for 60 min with TNF-? and an E2-containing buffer. RESULTS: TNF-? treatment resulted in deterioration of heart function. HIPC offered better protection by significantly increasing left ventricular developed pressure (Pmax) and coronary flow (P<0.01), and by decreasing left ventricular end-diastolic pressure (P<0.01). NIPC or pretreatment of the hearts with E2 normalized left ventricular end-diastolic pressure, coronary flow and coronary vascular resistance (P<0.001); however, it did not normalize Pmax. The combination of E2 and HIPC did not show any synergetic protection; however, the addition of HIPC normalized Pmax (P<0.001). CONCLUSIONS: TNF-? treatment resulted in deterioration of heart hemodynamics, which were reversed by HIPC, E2 PC and pretreatment. The combination of these treatments did not add to the previously observed protection compared with when they were used individually.

Juggi, Jasbir S; Hoteit, Lamia J; Babiker, Fawzi A; Joseph, Shaji; Mustafa, Abu Salim



Role of tumor necrosis factor-alpha and TRAIL in high-dose radiation-induced bystander signaling in lung adenocarcinoma.  


In the present study, ionizing radiation (IR)-induced bystander effects were investigated in two lung cancer cell lines. A549 cells were found to be more resistant to radiation-conditioned medium (RCM) obtained from A549 cells when compared with the H460 exposed to RCM procured from H460 cells. Significant release of tumor necrosis factor-alpha (TNF-alpha) was observed in A549 cells after IR/RCM exposure, and the survival was reversed with neutralizing antibody against TNF-alpha. In H460 cells, significant release of TNF-related apoptosis-inducing ligand (TRAIL), but not TNF-alpha, was observed in response to IR, RCM exposure, or RCM + 2Gy, and neutralizing antibody against TRAIL diminished clonogenic inhibition. Mechanistically, TNF-alpha present in RCM of A549 was found to mediate nuclear factor-kappaB (NF-kappaB) translocation to nucleus, whereas the soluble TRAIL present in RCM of H460 cells mobilized the nuclear translocation of PAR-4 (a proapoptotic protein). Analysis of IR-inducible early growth response-1 (EGR-1) function showed that EGR-1 was functional in A549 cells but not in H460 cells. A significant decrease in RCM-mediated apoptosis was observed in both A549 cells stably expressing small interfering RNA EGR-1 and EGR-1(-/-) mouse embryonic fibroblast cells. Thus, the high-dose IR-induced bystander responses in A549 may be dependent on the EGR-1 function and its target gene TNF-alpha. These findings show that the reduced bystander response in A549 cells is due to activation of NF-kappaB signaling by TNF-alpha, whereas enhanced response to IR-induced bystander signaling in H460 cells was due to release of TRAIL associated with nuclear translocation of PAR-4. PMID:18089811

Shareef, Mohammed M; Cui, Nuan; Burikhanov, Ravshan; Gupta, Seema; Satishkumar, Sabapathi; Shajahan, Shahin; Mohiuddin, Mohammed; Rangnekar, Vivek M; Ahmed, Mansoor M



The Receptor for Interleukin 3 is Selectively Induced in Human Endothelial Cells by Tumor Necrosis Factor alpha and Potentiates Interleukin 8 Secretion and Neutrophil Transmigration  

Microsoft Academic Search

Interleukin (IL)-3 stimulates hemopoiesis in vitro. However, IL-3 is not normally found in bone marrow, raising doubts as to the in vivo role of IL-3. We have found that human umbilical vein endothelial cells (HUVEC) express functional high-affinity receptors for IL-3 after stimulation with tumor necrosis factor alpha (TNF-alpha), IL-1beta, or lipopolysaccharide, and that this receptor is involved in inflammatory

Eija I. Korpelainen; Jennifer R. Gamble; William B. Smith; Gregory J. Goodall; Sun Qiyu; Joanna M. Woodcock; Mara Dottore; Mathew A. Vadas; Angel F. Lopez



Effect of salvia miltiorrhiza Bge on left ventricular hypertrophy and the expression of tumor necrosis factor-alpha in spontaneously hypertensive rats.  


The effects of salvia miltiorrhiza Bge (SMB) on left ventricular hypertrophy (LVH) and the expression of tumor necrosis factor-alpha (TNF-alpha) in the left ventricle of spontaneously hypertensive rats and the action mechanism were investigated. Normal Wistar-kyoto (WKY) rats were used as negative control, and spontaneously hypertensive rats (SHR) were randomly assigned to receive placebo or SMB. SMB (1 g/kg.d) was injected intraperitoneally for 12 weeks. Systolic blood pressure (SBP) and left ventricular mass index (LVMI) were measured. HE, VG and immunohistochemical staining combined with computed morphometry were employed to evaluate the cardiomyocyte size, diameter, the collagen volume fraction (CVF), perivascular circumferential area (PVCA), and tumor necrosis factor-alpha (TNF-alpha) expression in the left ventricular tissue. The results showed, as compared with WKY rats, the SBP, LVMI, cardiomyocyte size, diameter, CVF, PCVA, and TNF-alpha expression were increased markedly in the 20-week-old spontaneously hypertensive rats. SMB decreased LVMI (P<0.01), size of cardiomyocytes (P<0.01), collagen volume fraction (P<0.01), perivascular circumferential area (P<0.01), and TNF-alpha expression (P<0.01), but had no effect on SBP (P>0.05). It was suggested that chronic administration of SMB could inhibit and reverse the development of LVH in spontaneously hypertensive rats independent of BP. TNF-alpha may be involved in the reversal mechanism of LVH by SMB. PMID:17641833

Sun, Lianping; Zheng, Zhi



Tumor necrosis factor-alpha is expressed in donor heart and predicts right ventricular failure after human heart transplantation.  


BACKGROUND-Myocardial failure is an important problem after heart transplantation. Right ventricular (RV) failure is most common, although its mechanisms remain poorly understood. Inflammatory cytokines play an important role in heart failure. We studied the expression of tumor necrosis factor (TNF)-alpha and other cytokines in donor myocardium and their relationship to the subsequent development of RV failure early after transplantation. METHODS AND RESULTS-Clinical details were obtained, and ventricular function was assessed by transesophageal echocardiography in 26 donors before heart retrieval. A donor RV biopsy was obtained immediately before transplantation, and each recipient was followed for the development of RV failure. Reverse transcriptase-polymerase chain reaction was performed to detect TNF-alpha, interleukin-2, interferon-gamma, and inducible nitric oxide synthase expression. Eight of 26 recipients (30.8%) developed RV failure. Seven of these 8 (87.5%) expressed TNF-alpha, but only 4 of the 18 (22.2%) who did not develop RV failure expressed TNF-alpha (P<0.005). As a predictor of RV failure, TNF-alpha mRNA had a sensitivity of 87.5%, a specificity of 83.3%, a positive predictive value of 70%, and a negative predictive value of 93.7%. Western blotting demonstrated more TNF-alpha protein in the myocardium of donor hearts that developed RV failure (658+/-60 versus 470+/-57 optical density units, P<0.05). Immunocytochemistry localized TNF-alpha expression to cardiac myocytes. Reverse transcriptase-polymerase chain reaction detected interferon-gamma in 2 (7.7%), interleukin-2 in 1 (3.8%), and inducible nitric oxide synthase mRNA in 1 (3.8%) of the 26 donor hearts, none of which developed RV failure. CONCLUSIONS-TNF-alpha expression in donor heart cardiac myocytes seems to predict the development of RV failure in patients early after heart transplantation. PMID:10899097

Birks, E J; Owen, V J; Burton, P B; Bishop, A E; Banner, N R; Khaghani, A; Polak, J M; Yacoub, M H



The Skp1 prolyl hydroxylase from Dictyostelium is related to the hypoxia-inducible factor-alpha class of animal prolyl 4-hydroxylases.  


Skp1 is a cytoplasmic and nuclear protein of eukaryotes best known as an adaptor in SCF ubiquitin-protein isopeptide ligases. In Dictyostelium, Skp1 is subject to 4-hydroxylation at Pro(143) and subsequent O-glycosylation by alpha-linked GlcNAc and other sugars. Soluble cytosolic extracts have Skp1 prolyl 4-hydroxylase (P4H) activity, which can be measured based on hydroxylation-dependent transfer of [(3)H]GlcNAc to recombinant Skp1 by recombinant (Skp1-protein)-hydroxyproline alpha-N-acetyl-d-glucosaminyltransferase. The Dictyostelium Skp1 P4H gene (phyA) was predicted using a bioinformatics approach, and the expected enzyme activity was confirmed by expression of phyA cDNA in Escherichia coli. The purified recombinant enzyme (P4H1) was dependent on physiological concentrations of O(2), alpha-ketoglutarate, and ascorbate and was inhibited by CoCl(2), 3,4-dihydroxybenzoate, and 3,4-dihydroxyphenyl acetate, as observed for known animal cytoplasmic P4Hs of the hypoxia-inducible factor-alpha (HIFalpha) class. Overexpression of phyA cDNA in Dictyostelium yielded increased enzyme activity in a soluble cytosolic extract. Disruption of the phyA locus by homologous recombination resulted in loss of detectable activity in extracts and blocked hydroxylation-dependent glycosylation of Skp1 based on molecular weight analysis by SDS-PAGE, demonstrating a requirement for P4H1 in vivo. The sequence and functional similarities of P4H1 to animal HIFalpha-type P4Hs suggest that hydroxylation of Skp1 may, like that of animal HIFalpha, be regulated by availability of O(2), alpha-ketoglutarate, and ascorbate, which might exert novel control over Skp1 glycosylation. PMID:15705570

van der Wel, Hanke; Ercan, Altan; West, Christopher M



Dermal tumour necrosis factor-alpha induces dendritic cell migration to draining lymph nodes, and possibly provides one stimulus for Langerhans' cell migration.  

PubMed Central

Previous studies have shown that following skin sensitization there is an accumulation of dendritic cells (DC) in lymph nodes draining the site of exposure. A significant number of the DC which arrive in the lymph nodes bear high levels of antigen, and the available evidence indicates that they are derived from epidermal Langerhans' cells (LC). Although freshly isolated LC are relatively inefficient antigen-presenting cells, the antigen-bearing DC which are found within draining nodes following skin sensitization are highly immunostimulatory. Recent investigations indicate that the functional maturation of LC as they migrate from the skin is reflected by an enhanced capacity to form stable clusters with lymphocytes, and is associated with an increased expression of membrane major histocompatibility complex (MHC) class II (Ia) antigen. By analogy with in vitro studies of LC maturation, it is possible that such changes are effected by granulocyte/macrophage colony-stimulating factor (GM-CSF) and interleukin-1 (IL-1), both of which are products of epidermal cells. The question remains as to the nature of the stimulus that initiates LC migration. In the present study we have examined in mice the effects of intradermal injection of tumour necrosis factor-alpha (TNF-alpha), another epidermal cytokine, on the accumulation of DC in lymph nodes. Murine recombinant TNF-alpha was found to cause a concentration- and time-dependent increase in the number of DC within draining nodes. Under the same conditions of exposure murine recombinant GM-CSF was without effect. Heat treatment of mouse TNF-alpha resulted in an equivalent inhibition of both DC accumulation and cytotoxic activity measured by in vitro bioassay. An interesting observation was that equal concentrations of human TNF-alpha, of equivalent specific activity, failed to influence the frequency of lymph node DC. These data demonstrate that TNF-alpha induces DC accumulation in draining lymph nodes, and we propose that this cytokine may provide one stimulus for LC migration during cutaneous immune responses.

Cumberbatch, M; Kimber, I



Depletion of OLFM4 gene inhibits cell growth and increases sensitization to hydrogen peroxide and tumor necrosis factor-alpha induced-apoptosis in gastric cancer cells  

PubMed Central

Background Human olfactomedin 4 (OLFM4) gene is a secreted glycoprotein more commonly known as the anti-apoptotic molecule GW112. OLFM4 is found to be frequently up-regulated in many types of human tumors including gastric cancer and it was believed to play significant role in the progression of gastric cancer. Although the function of OLFM4 has been indicated in many studies, recent evidence strongly suggests a cell or tissue type-dependent role of OLFM4 in cell growth and apoptosis. The aim of this study is to examine the role of gastric cancer-specific expression of OLFM4 in cell growth and apoptosis resistance. Methods OLFM4 expression was eliminated by RNA interference in SGC-7901 and MKN45 cells. Cell proliferation, anchorage-independent growth, cell cycle and apoptosis were characterized in vitro. Tumorigenicity was analyzed in vivo. The apoptosis and caspase-3 activation in response to hydrogen peroxide (H2O2) or tumor necrosis factor-alpha (TNF ?) were assessed in the presence or absence of caspase inhibitor Z-VAD-fmk. Results The elimination of OLFM4 protein by RNA interference in SGC-7901 and MKN45 cells significantly inhibits tumorigenicity both in vitro and in vivo by induction of cell G1 arrest (all P < 0.01). OLFM4 knockdown did not trigger obvious cell apoptosis but increased H2O2 or TNF ?-induced apoptosis and caspase-3 activity (all P < 0.01). Treatment of Z-VAD-fmk attenuated caspase-3 activity and significantly reversed the H2O2 or TNF ?-induced apoptosis in OLFM4 knockdown cells (all P < 0.01). Conclusion Our study suggests that depletion of OLFM4 significantly inhibits tumorigenicity of the gastric cancer SGC-7901 and MKN45 cells. Blocking OLFM4 expression can sensitize gastric cancer cells to H2O2 or TNF ? treatment by increasing caspase-3 dependent apoptosis. A combination strategy based on OLFM4 inhibition and anticancer drugs treatment may provide therapeutic potential in gastric cancer intervention.



Effect of Gastrodia elata on tumor necrosis factor-alpha-induced matrix metalloproteinase activity in endothelial cells.  


The aim of the present study was to investigate whether an ethanol extract of Gastrodia elata (EGE) rhizome, a traditional Korean herbal medical food, suppresses the endothelial extracellular matrix degradation induced by tumor necrosis factor (TNF)-alpha. Gelatin zymography results showed that pretreatment with EGE to human umbilical vein endothelial cells (HUVEC) decreased TNF-alpha-induced increase of matrix metalloproteinase (MMP)-2/-9 activities in the range of 1-50 microg/ml. Real-time qRT-PCR results also revealed that TNF-alpha-induced MMP-2/-9 mRNA expression levels were attenuated by pretreatment with EGE. These results provide new insights into the pathophysiological mechanisms for the anti-atherosclerotic properties of EGE in vascular diseases. PMID:19672675

Lee, Yun Jung; Hwang, Sun Mi; Kang, Dae Gill; Kim, Jin Sook; Lee, Ho Sub



Tumour necrosis factor-alpha expression in tumour islets confers a survival advantage in non-small cell lung cancer  

Microsoft Academic Search

BACKGROUND: The role of TNF? in cancer is complex with both pro-tumourigenic and anti-tumourigenic roles proposed. We hypothesised that anatomical microlocalisation is critical for its function. METHODS: This study used immunohistochemistry to investigate the expression of TNF? in the tumour islets and stroma with respect to survival in 133 patients with surgically resected NSCLC. RESULTS: TNF? expression was increased in

Chandra M Ohri; Aarti Shikotra; Ruth H Green; David A Waller; Peter Bradding



Epigenetic Regulation of Tumor Necrosis Factor Alpha  

Microsoft Academic Search

Tumor necrosis factor alpha (TNF-) is a potent cytokine which regulates inflammation via the induction of adhesion molecules and chemokine expression. Its expression is known to be regulated in a complex manner with transcription, message turnover, message splicing, translation, and protein cleavage from the cell surface all being independently regulated. This study examined both cell lines and primary cells to

K. E. Sullivan; A. B. M. Reddy; K. Dietzmann; A. R. Suriano; V. P. Kocieda; M. Stewart; M. Bhatia



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

SciTech Connect

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.

Han, Hyo-Kyung [BK21 Project Team, College of Pharmacy, Chosun University, Gwangju 501-759, South Korea (Korea, Republic of); Han, Chang Yeob [BK21 Project Team, College of Pharmacy, Chosun University, Gwangju 501-759, South Korea (Korea, Republic of); Cheon, Eun-Pa [BK21 Project Team, College of Pharmacy, Chosun University, Gwangju 501-759, South Korea (Korea, Republic of); Lee, Jaewon [College of Pharmacy, Pusan National University, Busan 609-735, South Korea (Korea, Republic of); Kang, Keon Wook [BK21 Project Team, College of Pharmacy, Chosun University, Gwangju 501-759, South Korea (Korea, Republic of)]. E-mail:



Down-regulation of cyclic-nucleotide phosphodiesterase 3B in 3T3-L1 adipocytes induced by tumour necrosis factor alpha and cAMP.  

PubMed Central

We have used murine 3T3-L1 cells, which differentiate in culture and acquire morphological and biochemical features of mature adipocytes, as a model for studying the expression of cyclic-nucleotide phosphodiesterase (PDE) 3B activity, protein and mRNA during differentiation and during long-term treatment of the cells with tumour necrosis factor alpha (TNF-alpha), a cytokine associated with insulin resistance, and a cAMP analogue, N(6),2'-O-dibutyryl cAMP (dbcAMP). PDE3B activity, protein and mRNA could be detected 4 days after the initiation of differentiation of 3T3-L1 preadipocytes. Treatment of 3T3-L1 adipocytes with 10 ng/ml TNF-alpha for 24 h produced a maximal (50%) decrease in PDE3B activity, protein and mRNA, which was well correlated with both activation of protein kinase A (PKA) and stimulation of lipolysis, presumably reflecting an increase in intracellular cAMP concentration. To investigate the effect of cAMP on PDE3B we treated 3T3-L1 adipocytes with dbcAMP. After 4 h with 0.5 mM dbcAMP, PDE3B activity was decreased by 80%, which was also correlated with a decrease in PDE3B protein and mRNA. This effect was abolished in the presence of N-[2-(bromocinnamylamino)ethyl]-5-isoquinolinesulphonamide] (H-89), a specific PKA inhibitor. We conclude that the lipolytic effect of TNF-alpha involves the down-regulation of PDE3B, which is associated with increased activation of PKA, presumably owing to increased levels of cAMP. In addition, the PKA activation induced by dbcAMP resulted in the down-regulation of PDE3B. These results, which suggest that PDE3B is a novel target for long-term regulation by TNF-alpha and cAMP, could contribute to the understanding of the mechanisms of insulin resistance.

Rahn Landstrom, T; Mei, J; Karlsson, M; Manganiello, V; Degerman, E



Tumor necrosis factor alpha (TNF-alpha) levels in the brain and cerebrospinal fluid after meningitis induced by Streptococcus pneumoniae.  


Bacterial meningitis due to Streptococcus pneumoniae is associated with a significant mortality rate and persisting neurologic sequelae including sensory-motor deficits, seizures, and impairments of learning and memory. The presence of proliferating bacteria within the subarachnoid and ventricular space compartments triggers an intense inflammatory host response at killing the invading microorganism. Proinflammatory mediators released in the process include tumor necrosis factor alpha (TNF-alpha), interleukin (IL)-1beta, IL-6. TNF-alpha have several effects, including cytotoxicity, antiviral activity, transcription factor activation, and immune response regulation. Thus, the aim of this study was to verify the levels of the TNF-alpha after pneumococcal meningitis in male Wistar rats. The animals underwent a magna cistern tap receiving either 10 microL sterile saline as a placebo or an equivalent volume of a S. pneumoniae suspension at the concentration 5 x 10(9)cfu/mL. The animals were killed at 0, 6, 12, 24, 48 and 96 h after induction. The brain was removed and hippocampus, cortex, prefrontal and cerebrospinal fluid (CSF) were isolated and used for the determination of TNF-alpha levels. We found an increase in TNF-alpha levels at 6h after induction of the meningitis in the hippocampus (p<0.01), frontal cortex (p<0.05), and cerebrospinal fluid (p<0.001).There was no alteration in the cortex. Our data suggest that TNF-alpha is involved in the pathophysiology of the pneumococcal meningitis and could be investigated as a putative biomarker for brain damage in the first hours. PMID:19835931

Barichello, Tatiana; dos Santos, Ivonete; Savi, Geovana D; Florentino, Anelise F; Silvestre, Cintia; Comim, Clarissa M; Feier, Gustavo; Sachs, Daniela; Teixeira, Mauro M; Teixeira, Antonio L; Quevedo, Joăo



Tumour necrosis factor-alpha expression in tumour islets confers a survival advantage in non-small cell lung cancer  

PubMed Central

Background The role of TNF? in cancer is complex with both pro-tumourigenic and anti-tumourigenic roles proposed. We hypothesised that anatomical microlocalisation is critical for its function. Methods This study used immunohistochemistry to investigate the expression of TNF? in the tumour islets and stroma with respect to survival in 133 patients with surgically resected NSCLC. Results TNF? expression was increased in the tumour islets of patients with above median survival (AMS) compared to those with below median survival (BMS)(p = 0.006), but similar in the stroma of both groups. Increasing tumour islet TNF? density was a favorable independent prognostic indicator (p = 0.048) while stromal TNF? density was an independent predictor of reduced survival (p = 0.007). Patients with high TNF? expression (upper tertile) had a significantly higher 5-year survival compared to patients in the lower tertile (43% versus 22%, p = 0.01). In patients with AMS, 100% of TNF?+ cells were macrophages and mast cells, compared to only 28% in the islets and 50% in the stroma of BMS patients (p < 0.001). Conclusions The expression of TNF? in the tumour islets of patients with NSCLC is associated with improved survival suggesting a role in the host anti-tumour immunological response. The expression of TNF? by macrophages and mast cells is critical for this relationship.



Tumour Necrosis Factor Alpha, Interferon Gamma and Substance P Are Novel Modulators of Extrapituitary Prolactin Expression in Human Skin  

PubMed Central

Human scalp skin and hair follicles (HFs) are extra-pituitary sources of prolactin (PRL). However, the intracutaneous regulation of PRL remains poorly understood. Therefore we investigated whether well-recognized regulators of pituitary PRL expression, which also impact on human skin physiology and pathology, regulate expression of PRL and its receptor (PRLR) in situ. This was studied in serum-free organ cultures of microdissected human scalp HFs and skin, i.e. excluding pituitary, neural and vascular inputs. Prolactin expression was confirmed at the gene and protein level in human truncal skin, where its expression significantly increased (p?=?0.049) during organ culture. There was, however, no evidence of PRL secretion into the culture medium as measured by ELISA. PRL immunoreactivity (IR) in female human epidermis was decreased by substance P (p?=?0.009), while neither the classical pituitary PRL inhibitor, dopamine, nor corticotropin-releasing hormone significantly modulated PRL IR in HFs or skin respectively. Interferon (IFN) ? increased PRL IR in the epithelium of human HFs (p?=?0.044) while tumour necrosis factor (TNF) ? decreased both PRL and PRLR IR. This study identifies substance P, TNF? and IFN? as novel modulators of PRL and PRLR expression in human skin, and suggests that intracutaneous PRL expression is not under dopaminergic control. Given the importance of PRL in human hair growth regulation and its possible role in the pathogenesis of several common skin diseases, targeting intracutaneous PRL production via these newly identified regulatory pathways may point towards novel therapeutic options for inflammatory dermatoses.

Langan, Ewan A.; Vidali, Silvia; Pigat, Natascha; Funk, Wolfgang; Lisztes, Erika; Biro, Tamas; Goffin, Vincent; Griffiths, Christopher E. M.; Paus, Ralf



Recombinant tumor necrosis factor. alpha. and interleukin 1. alpha. increase expression of c-abl protooncogene mRNA in cultured human marrow stromal cells  

SciTech Connect

Analysis of protooncogene RNA expression in marrow stromal cells from long-term marrow culture demonstrated high levels of c-abl 5-, 6-, and 7-kilobase (kb) RNA transcripts. In experiments on three independently derived simian virus 40-transformed marrow stromal cell lines, the expression of these c-abl transcripts was further increased in response to recombinant tumor necrosis factor {alpha} (1,000 units/ml) and interleukin 1{alpha} (10 units/ml). Although lymphocyte-conditioned medium predominantly up-regulated the 5-kb transcript, interleukin 1{alpha} primarily affected the 6-kb transcript. The up-regulation of the 5-kb c-abl message correlated with up-regulation of the granulocyte/macrophage colony-stimulating factor transcript and down-regulation of procollagen I transcripts in transformed cells. These data suggest that c-abl plays roles in the regulation of extracellular matrix expression and in the regulation of hematopoietic growth factors by stromal cells.

Andrews, D.F. III; Nemunaitis, J.J.; Singer, J.W. (Univ. of Washington, Seattle, WA (USA))



Pulmonary expression of tumor necrosis factor alpha, interleukin-1 beta, and interleukin-8 in the acute phase of bovine pneumonic pasteurellosis.  


Inflammatory cytokines are suspected to contribute to the pathogenesis of bovine pneumonic pasteurellosis (BPP) through neutrophil recruitment, leukocyte activation, and the induction of a broad array of soluble inflammatory mediators. An in vivo experimental model of BPP was used to characterize the pulmonary expression kinetics of tumor necrosis factor alpha (TNFalpha), interleukin-1 beta (IL-1beta), and interleukin-8 (IL-8) genes and proteins during the acute phase of disease development. Cytokine expression in bronchoalveolar lavage (BAL) fluid, BAL cells, and pneumonic lung parenchyma was quantitated by northern blot analysis, enzyme-linked immunosorbent assay (ELISA), and in situ hybridization at 2, 4, 8, 16, and 24 hours after endobronchial inoculation of Pasteurella (Mannheimia) haemolytica. Expression of TNFalpha, IL-1beta, and IL-8 was significantly increased in the airways and lung lesions of infected calves as compared with mock-infected controls. Although kinetic patterns varied, peak levels of cytokine mRNA occured within 8 hours postinfection (PI), and peak cytokine concentrations occurred within 16 hours PI. In all samples, IL-8 was expressed to the greatest extent and TNFalpha was least expressed. Expression of TNFalpha was restricted to alveolar macrophages. Alveolar and interstitial macrophages produced IL-1beta and IL-8 in the first 4 hours; bronchial and bronchiolar epithelial cells were also significant sources of IL-8 during this period. By 8 hours PI, neutrophils were the dominant source of both IL-1beta and IL-8. These findings demonstrate a spatial and temporal association between pulmonary expression of inflammatory cytokines and acute lung pathology, supporting the hypothesis that cytokines contribute to inflammatory lung injury in BPP. PMID:11355660

Malazdrewich, C; Ames, T R; Abrahamsen, M S; Maheswaran, S K



Phorbol ester-induced activation of a membrane-bound candidate pro-transforming growth factor-alpha processing enzyme.  


Transforming growth factor-alpha (TGF-alpha) is synthesized and transported to the cell surface as a membrane-anchored precursor (proTGF-alpha) that is converted to the soluble form by proteolytic cleavage. ProTGF-alpha cleavage is activated in response to tumor-promoting phorbol ester or to calcium ionophore. Mechanism(s) controlling conversion of the membrane-anchored precursor to the soluble TGF-alpha are unknown, and the responsible protease has not been identified. Using the fluorogenic substrate succinyl-Ala-Ala-Ala-4-methylcoumaryl-7-amide (Suc-Ala-Ala-Ala-MCA), containing the sequence similar to the cleavage sites for proTGF-alpha processing, we identified a putative candidate proTGF-alpha-converting enzyme in the membrane fractions of Chinese hamster ovary (CHO) cells. The enzyme activity was about 5-6-fold increased by exposure of the cells with phorbol ester or with calcium ionophore. The enzyme activity was released from the cells by trypsin added exogenously; hence, the activity probably locates on the outer surface of the cell membrane. The enzyme was partially purified from membranes of phorbol ester-treated CHO cells. The molecular mass of the enzyme was estimated to be about 84 kDa by gel filtration analysis. Inhibitor profile of the enzyme, including inhibition by serine protease inhibitors and no inhibition by elastatinal, coincides with that previously reported for proTGF-alpha cleavage activity on CHO cells. These findings suggest that the enzyme is a processing protease involved in the cleavage of proTGF-alpha and that induction of proTGF-alpha conversion by phorbol ester or by calcium ionophore can be attributed to activation of the processing protease. In the cell-free reconstitution system, the membrane-bound Suc-Ala-Ala-Ala-MCA-hydrolyzing enzyme from CHO cells was also activated by phorbol ester in the presence of the soluble fraction, ATP, and vanadate. Requirement of vanadate, an inhibitor for protein phosphatases, for activation of the enzyme in this system suggests that phosphorylation-dephosphorylation probably plays a key role in the regulated cleavage of proTGF-alpha. PMID:8051125

Harano, T; Mizuno, K



A tumor necrosis factor alpha- and interleukin 6-inducible protein that interacts with the small subunit of DNA polymerase delta and proliferating cell nuclear antigen.  


A cDNA encoding a protein of 36 kDa, polymerase delta-interacting protein 1 (PDIP1), that interacts with the small subunit (p50) of DNA polymerase delta (pol delta) was identified in a two-hybrid screen of a HepG2 cDNA library by using p50 as bait. The interaction of PDIP1 with p50 was confirmed by pull-down assays, and a similar assay was used to demonstrate that PDIP1 interacts directly with the proliferating cell nuclear antigen (PCNA). PCNA and p50 bound to PDIP1 simultaneously, and PDIP1 stimulated pol delta activity in vitro in the presence, but not the absence, of PCNA, suggesting that PDIP1 also interacts functionally with both p50 and PCNA. Subcellular localization studies demonstrated that PDIP1 is a nuclear protein that colocalizes with PCNA at replication foci. A putative PCNA-binding motif was identified within the C terminus of PDIP1, and a synthetic peptide containing this PCNA-binding motif was shown to bind PCNA by far-Western analysis. Northern analysis demonstrated that PDIP1 mRNA is present in a wide variety of human tissues. PDIP1 was found to be highly homologous to a previously identified protein, B12 [Wolf, F. W., Marks, R. M., Sarma. V., Byers, M. G., Katz, R. W., Shows, T. B. & Dixit, V. M. (1992) J. Biol. Chem. 267, 1317-1326], one of the early response genes induced by tumor necrosis factor alpha. PDIP1 synthesis can also be induced by tumor necrosis factor alpha and by IL-6, cytokines essential for liver regeneration after loss of hepatic tissue. It is suggested that PDIP1 provides a link between cytokine activation and DNA replication in liver as well as in other tissues. PMID:11593007

He, H; Tan, C K; Downey, K M; So, A G



A food-born heterocyclic amine, 2-amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), suppresses tumor necrosis factor-alpha expression in lipoteichoic acid-stimulated RAW 264.7 cells.  


2-Amino-1-methyl-6-phenylimidazo[4,5-b]pyridine (PhIP), a heterocyclic amine with strong carcinogenic and mutagenic potential, is created abundantly in the overcooking of meat and fish. Carcinogenic toxicants are often implicated in immunosuppression, where cancer cells are not easily eliminated by the host immune system. Here, we investigated the effect of PhIP on tumor necrosis factor-alpha (TNF-alpha) expression by murine macrophage-like cells (RAW 264.7) stimulated with lipoteichoic acid (LTA), a major virulence factor of Gram-positive bacteria. Upon exposure to LTA purified from Staphylococcus aureus, TNF-alpha expression was substantially induced, whereas pretreatment with PhIP significantly inhibited LTA-induced TNF-alpha expression. LTA is known to activate Toll-like receptor 2 (TLR2) and NF-kappaB, resulting in TNF-alpha expression. Interestingly, PhIP did not interfere with LTA-binding to TLR2, its stimulation of TLR2, or the DNA-binding activity of NF-kappaB. However, treatment with actinomycin D facilitated the PhIP-induced attenuation of TNF-alpha mRNA expression, implying that PhIP might decrease TNF-alpha mRNA stability rather than its biosynthesis. Furthermore, Western blot analysis demonstrated that PhIP reduced the phosphorylation of ERK1/2 and JNK but not p38 kinase in LTA-stimulated cells. The addition of a protein kinase C (PKC) activator, phorbol 12-myristate 13-acetate, rescued PhIP-inhibited TNF-alpha expression in LTA-stimulated cells. These results suggest that PhIP downregulates TNF-alpha expression in LTA-stimulated macrophages by decreasing TNF-alpha mRNA stability and signaling pathways related to PKC, ERK1/2, and JNK activation. PMID:18845389

Im, Jintaek; Choi, Hyung Shim; Kim, Sun Kyung; Woo, Sang Su; Ryu, Young Hee; Kang, Seok-Seong; Yun, Cheol-Heui; Han, Seung Hyun



Enhancement of tumor necrosis factor-alpha gene expression by low doses of prostaglandin E2 and cyclic GMP.  


Macrophage-derived PGE2 is usually considered to be a down-regulator of TNF-alpha production. However, we recently demonstrated that PGE2 may display dual activities in that low concentrations stimulated whereas higher doses suppressed TNF-alpha synthesis in resident peritoneal macrophages. To examine the underlying molecular mechanisms, we studied TNF-alpha gene expression in rat peritoneal macrophages and the murine PU5-1.8 macrophage line. In both macrophage types, PGE2 enhanced TNF-alpha gene transcription and production at an optimal concentration of 1 ng/ml. Furthermore, evidence was obtained that PGE2 may stimulate TNF-alpha mRNA accumulation via a rise of the intracellular messenger cGMP. Both, exogenously added as well as endogenously, by sodium nitroprusside generated cGMP were found to enhance TNF-alpha gene expression and production. These findings lend further support to the concept that cGMP may represent one of the positive signals for TNF-alpha synthesis. PMID:1965895

Gong, J H; Renz, H; Sprenger, H; Nain, M; Gemsa, D



Role of isoprenylcysteine carboxyl methyltransferase in tumor necrosis factor-alpha stimulation of expression of vascular cell adhesion molecule-1 in endothelial cells.  


We have previously shown that cytokine stimulation of the expression of vascular cell adhesion molecule-1 (VCAM-1), but not that of intercellular adhesion molecule-1 (ICAM-1), is redox sensitive in endothelial cells. Here, we investigated the role of isoprenylcysteine carboxyl methyltransferase (ICMTase), which methylates isoprenylated CAAX (where C indicates cysteine; A, aliphatic amino acids; and X, almost any other amino acid) proteins, including Rac1, a component of superoxide-generating NAD(P)H oxidase, in the expression of VCAM-1. Pretreatment of endothelial cells with N-acetyl-S-farnesyl-L-cysteine (AFC) or N-acetyl-S-geranylgeranyl-L-cysteine (AGGC), specific inhibitors of ICMTase, inhibited the tumor necrosis factor-alpha (TNF-alpha) stimulation of mRNA expression of VCAM-1 but not that of ICAM-1. Endothelial cells expressed constitutively active ICMTase, as suggested by the presence of methylated Rac1 and the methylation of AFC by the cells. TNF-alpha stimulation of the cells significantly increased the methylation of AFC and Rac1 in endothelial cells. That ICMTase was a component of the redox-sensitive signaling pathway was also suggested by the AFC inhibition of the generation of reactive oxygen species by TNF-alpha. Interestingly, the dominant-negative isoform of Rac1 was not selective but inhibited the TNF-alpha stimulation of the mRNA expression of VCAM-1 and ICAM-1. Thus, ICMTase is a critical component of the redox-sensitive VCAM-1-selective signaling pathway, and it appears to activate a discrete inflammatory signaling pathway, at least in part, through the methylation of Rac1. PMID:12006387

Ahmad, Mushtaq; Zhang, Yan; Zhang, Yong; Papharalambus, Christopher; Alexander, R Wayne



Tumor necrosis factor alpha and endothelin-1 increase P-glycoprotein expression and transport activity at the blood-brain barrier.  


The ATP-driven drug efflux pump, P-glycoprotein, is a critical and selective element of the blood-brain barrier and a primary impediment to pharmacotherapy of central nervous system (CNS) disorders. Thus, an understanding of how P-glycoprotein function is regulated has the potential to improve CNS therapy. We recently demonstrated rapid (minutes) and reversible inactivation of P-glycoprotein in rat brain capillaries signaled through tumor necrosis factor-alpha (TNF-alpha) and endothelin-1 (ET-1), components of the brain's innate immune response. In this study, we examined the longer-term consequences of continuous exposure of rat brain capillaries to low levels of TNF-alpha and ET-1. Exposing brain capillaries to TNF-alpha or ET-1 caused a rapid decrease in P-glycoprotein transport activity with no change in transporter protein expression. This was followed by a 2- to 3-h plateau at the low activity level and then by a sharp increase in both transport activity and protein expression. After 6 h, transport activity and transporter protein expression was double that of control samples. TNF-alpha signaled through TNF-R1, which in turn caused ET release and action through ETA and ETB receptors, nitric-oxide synthase, protein kinase C and nuclear factor-kappaB (NF-kappaB) and finally increased P-glycoprotein expression and transport activity. Assuming similar effects occur in vivo, the present results imply a tightening of the selective blood-brain barrier with chronic inflammation and thus reduced efficacy of CNS-acting drugs that are P-glycoprotein substrates. Moreover, involvement of NF-kappaB raises the possibility that other effectors acting through this transcription factor may have similar effects on this key blood-brain barrier transporter. PMID:17132686

Bauer, Björn; Hartz, Anika M S; Miller, David S



Gene expression for interleukin-2 and tumor necrosis factor-alpha in the spleen of old rats under physiological condition and during septic shock. Possible pharmacological modulation.  


Older individuals are more susceptible to infectious agents than younger and this is related to the disrepair of the immune defence mechanisms associated with aging. In this study we evaluated the activity of a new biological response modifier (BRM), pidotimod ((R)-3-[(S)-(5-oxo-2-pyrrolidinyl)carbonyl]-thiazolidine-4-carboxylic acid, PGT/1A, CAS 121808-62-6) in relation to the expression of some cytokine genes. We utilized 24 month-old Sprague-Dawley rats (n = 24), randomly divided into 4 groups: controls (n = 6), pidotimod-treated (n = 6; 200 mg/kg i.p., for 10 days), infected (n = 6; i.p. infection of E. coli CH 198) and pidotimod-treated + infected (n = 6). Poly(A+)RNA purified from the spleens of the animals killed 48 h after the infection was probed with Interleukin-2 (IL-2) and Tumor Necrosis Factor-alpha (TNF-alpha) cDNA clones. Northern blot analysis showed a slight signal of the IL-2 steady state mRNA in the groups of control, pidotimod-treated and infected animals, with an increase (20%) evident only in pidotimod + infected rats, 48 h after E. coli injection. On the contrary, the TNF-alpha mRNA levels were easily detectable in controls and infected rats and lower (20%, 40%) following the drug treatment, independent of i.p. infection. These results account for the BRM activity of pidotimod. PMID:7531977

Annoni, G; Arosio, B; Santambrogio, D; Cullurŕ, D; Gagliano, N; Uslenghi, C



Enhancement of Mycobacterium tuberculosis-Induced Tumor Necrosis Factor Alpha Production from Primary Human Monocytes by an Activated T-Cell Membrane-Mediated Mechanism  

PubMed Central

Mycobacterium tuberculosis alone induces small, donor-variable amounts of tumor necrosis factor alpha (TNF-?) from primary human monocytes in vitro. However, TNF-? release is increased 5- to 500-fold when fixed activated T cells (FAT) or their isolated, unfixed membranes are added to this system. This FAT-induced synergy was at least as potent as that induced by gamma interferon (IFN-?) at 100 U/ml. FAT-enhanced TNF-? production is at least in part transcriptionally mediated, as reflected by quantitative changes in TNF-? mRNA between 2 and 6 h poststimulation. Unlike IFN-?-cocultured cells, FAT-treated monocytes appeared not to have enhanced TNF-? message stability, suggesting that de novo transcription may be involved in this effect. Furthermore, M. tuberculosis alone induced only minimal DNA binding of monocyte NF-?B, but cells treated with M. tuberculosis and FAT potentiated NF-?B activity more effectively. It is therefore possible that one mechanism by which FAT synergize with M. tuberculosis to stimulate TNF-? production is via NF-?B-enhanced transcription. These data strongly suggest that in the interaction of cells involved in the immune response to M. tuberculosis, T-cell stimulation of monocyte TNF-? production involves a surface membrane interaction(s) as well as soluble mediators.

Warwick-Davies, Jan; Watson, Amanda J.; Griffin, George E.; Krishna, Sanjeev; Shattock, Robin J.



Effect of Gastrodia elata on tumor necrosis factor-alpha-induced matrix metalloproteinase activity in endothelial cells  

Microsoft Academic Search

The aim of the present study was to investigate whether an ethanol extract of Gastrodia elata (EGE) rhizome, a traditional Korean herbal medical food, suppresses the endothelial extracellular matrix degradation induced\\u000a by tumor necrosis factor (TNF)-?. Gelatin zymography results showed that pretreatment with EGE to human umbilical vein endothelial\\u000a cells (HUVEC) decreased TNF-?-induced increase of matrix metalloproteinase (MMP)-2\\/-9 activities in

Yun Jung Lee; Sun Mi Hwang; Dae Gill Kang; Jin Sook Kim; Ho Sub Lee



Two types of tumor necrosis factor-alpha in bluefin tuna (Thunnus orientalis) genes: Molecular cloning and expression profile in response to several immunological stimulants.  


Tumor necrosis factor-alpha (TNF-alpha) is a key inflammatory mediator and has also the potential as a prominent biomarker of innate immunity. In this study, we identified and characterized TNF-alpha from bluefin tuna, which is an important cultured species. Two types of TNF-alpha were also cloned incidentally (TNF1 and TNF2). The open reading frame of TNF1 and TNF2 cDNA encoded 247 and 245 amino acids, respectively. The amino acid sequence identity among sea perch, red sea bream, and tiger puffer was 73, 70, 59% for TNF1 and 49, 51, 45% for TNF2, respectively. The identity between TNF1 and TNF2 amino acid sequences of the bluefin tuna was only 43%. The positions of cysteine residues, transmembrane sequence, and protease cleavage site in bluefin tuna TNFs were similar with other reported fish and mammalian TNF-alpha. In a phylogenetic analysis, TNF1 is grouped with other reported Perciformes TNF-alpha. On the other hand, TNF2 is grouped with ayu TNF and is quite distant from the fish TNF-alpha group and lymphotoxin-beta group. While TNF1 mRNA showed no significant difference in all tissues, TNF2 mRNA was expressed significantly higher in the blood than in the gill, intestine, head kidney, spleen, heart, and ovary. In peripheral blood leucocytes (PBL), expressions of TNF2 mRNA were significantly increased by stimulation with lipopolysaccharide, phytohemagglutinin, concanavalin A, pokeweed mitogen, phorbol myristate acetate in vitro, but those of TNF1 were not. Recombinant mature TNF1 and TNF2 proteins significantly enhanced phagocytic activity of PBL. Our results suggest that bluefin tuna possess two types of TNF-alpha homologue, and TNF2 is a potential biomarker for innate immunity. PMID:19146959

Kadowaki, Takeshi; Harada, Hideaki; Sawada, Yoshifumi; Kohchi, Chie; Soma, Gen-Ichiro; Takahashi, Yukinori; Inagawa, Hiroyuki



Recombinant Guinea Pig Tumor Necrosis Factor Alpha Stimulates the Expression of Interleukin-12 and the Inhibition of Mycobacterium tuberculosis Growth in Macrophages  

PubMed Central

Tumor necrosis factor alpha (TNF-?) plays an important role in the host immune response to infection with the intracellular pathogen Mycobacterium tuberculosis. It is essential for the formation of protective tuberculous granulomas and regulates the expression of other cytokines which contribute to a protective immune response. Interleukin-12 (IL-12) is known to promote a Th1 response, which is essential for antimycobacterial resistance. Recombinant guinea pig TNF-? (rgpTNF-?) protein (17 kDa) was purified, and its bioactivity was confirmed by its cytotoxicity for L929 fibroblasts. High titers of polyclonal anti-gpTNF-? antibody were obtained by immunization of rabbits. Resident alveolar and peritoneal macrophages were isolated from guinea pigs and infected with either the H37Ra or H37Rv strain of M. tuberculosis. The mRNA levels for TNF-? and IL-12 p40 were measured using real-time PCR. IL-12 p40 mRNA was up-regulated in a dose-dependent manner by rgpTNF-? alone. In infected macrophages, a lower dose of rgpTNF-? intensified the mRNA levels of TNF-? and IL-12 p40. However, higher doses of rgpTNF-? suppressed TNF-? and IL-12 p40 mRNA. The antimycobacterial activity of macrophages was assessed by metabolic labeling of M. tuberculosis with [3H]uracil. Resident alveolar and peritoneal macrophages treated with anti-gpTNF-? antibody to block endogenous TNF-? exhibited increased intracellular mycobacterial growth. These data suggest that the dose of TNF-? is crucial to the stimulation of optimal expression of protective cytokines and that TNF-? contributes to the control of mycobacterial replication to promote host resistance against M. tuberculosis.

Cho, Hyosun; Lasco, Todd M.; Allen, Shannon Sedberry; Yoshimura, Teizo; McMurray, David N.



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

PubMed Central

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

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



Shedding of Tumor Necrosis Factor Receptor 1 Induced by Protein A Decreases Tumor Necrosis Factor Alpha Availability and Inflammation during Systemic Staphylococcus aureus Infection  

PubMed Central

Staphylococcus aureus infections are an important public health concern due to their increasing incidence and high rates of mortality. The success of S. aureus as a pathogen is highly related to its enormous capacity to evade the host immune response. The critical role of tumor necrosis factor alpha (TNF-?) in the initial host defense against systemic staphylococcal infection has been demonstrated in experimental models and may partially explain the lack of significant benefits observed in clinical trials attempting to neutralize this cytokine in septic patients. S. aureus protein A plays a key role in regulating inflammation through its ability to bind and signal through the TNF-? receptor 1 (TNFR1). In this study, we demonstrate that S. aureus, via protein A-mediated signaling, induces early shedding of TNFR1, which precedes the secretion of TNF-? in vitro and in vivo. The results obtained using a protein A-deficient mutant and tnfr1?/? mice strongly suggest that the increased levels of soluble TNFR1 present during experimental S. aureus infection may neutralize circulating TNF-? and impair the host inflammatory response. Early shedding of TNFR1 induced by protein A may constitute a novel mechanism by which S. aureus subverts the host immune response.

Giai, Constanza; Gonzalez, Cintia; Ledo, Camila; Garofalo, Ailin; Di Genaro, Maria Silvia; Sordelli, Daniel O.



Nimesulide and diclofenac inhibit lipopolysaccharide-induced hypothermia and tumour necrosis factor-alpha elevation in rats.  


The effects of nimesulide and diclofenac on lipopolysaccharide (LPS)-induced rectal temperature changes and serum tumour necrosis factor (TNF)-alpha elevation were investigated in rats. LPS (Escherichia coli O111:B4; 50 microg/kg, intraperitoneally) produces a dual body temperature response, in which initial hypothermia precedes fever. Serum TNF-alpha levels rise during the initial phase of the induced hypothermia. Nimesulide, a preferential inhibitor of cyclooxygenase-2 (0.05, 0.5 or 1 mg/kg, subcutaneously) completely abolished the hypothermia, resulting in an acceleration of the fever phase. However, the peak and plateau phases of fever were not changed by nimesulide treatment. Nimesulide (0.5 mg/kg) partially prevented serum TNF-alpha elevation. The non-selective cyclooxygenase inhibitor diclofenac inhibited hypothermia at all doses tested (0.03, 0.3 or 3 mg/kg, subcutaneously) although fever was completely abolished at the 3 mg/kg dose only. Diclofenac also partially abolished the elevation in serum TNF-alpha levels, but at the highest dose only (3 mg/kg). These data suggest that nimesulide and diclofenac can preferentially inhibit LPS-induced hypothermia at doses that do not abolish fever in rats. Both these drugs also reduced elevated TNF-alpha levels, a fact which may, at least partly, explain the antihypothermic effect of nimesulide. PMID:12570019

Dogan, Muberra Devrim; Ataoglu, Haluk; Akarsu, Eyup S



Effect of retinoic acid and vitamin D on the expression of interleukin-1 beta, tumour necrosis factor-alpha and interleukin-6 in the human monocytic cell line U937.  

PubMed Central

We have previously described a synergism between the two physiological hormones, retinoic acid (RA) and 1 alpha,25-dihydroxyvitamin D3 (VD) in the induction of U937 cell differentiation towards a more mature state. Herein, we investigated the regulation of cytokine production during RA and/or VD treatment of U937 cells. Cell differentiation was followed by measurement of their capacity to give oxidative responses, and interleukin-1 beta (IL-1 beta), tumour necrosis factor-alpha (TNF-alpha) and IL-6 gene and protein expression were determined in RA/VD-treated cells, activated or not with lipopolysaccharide (LPS). The undifferentiated and RA-treated U937 cells were unable to produce monokines even when they were stimulated by LPS. VD induced the monokine mRNA expression in U937 cells but failed to induce protein release. However, unlike RA, it primed the cells to secrete monokines upon endotoxin stimulation. A large enhancement of the production of the monokines both at mRNA and protein levels was observed in the U937 cells exposed to the combination of RA + VD. Nevertheless, protein release required a further step of activation of the RA + VD-primed cells. The co-inducer effect of RA and VD was not observed in HL-60 or THP-1 cells and seems to be restricted to U937 cells. These results on cytokine expression support our previous finding that a combination of RA and VD brings the U937 cells to a high stage of myeloid differentiation with major characteristics of monocytes/macrophages. Images Figure 1

Taimi, M; Defacque, H; Commes, T; Favero, J; Caron, E; Marti, J; Dornand, J



Tumor necrosis factor-alpha-induced reduction of glomerular filtration rate in rats with fulminant hepatic failure.  


The mechanism of renal failure during fulminant hepatic failure (FHF) or end-stage of liver disease is not fully understood. The present study aims to delineate the mechanisms of decreased glomerular filtration rate (GFR) in acute hepatic failure. A rat model of renal insufficiency in severe liver injury was established by lipopolysaccharide (LPS) plus D-galactosamine (GalN) exposure. GFR was evaluated by continuous infusion of fluorescein isothiocyanate-inulin with implanted micro-osmotic pumps. GalN/LPS intoxication resulted in severe hepatocyte toxicity as evidenced by liver histology and biochemical tests, whereas renal morphology remained normal. GFR was reduced by 33% of the controls 12?h after GalN/LPS exposure, accompanied with a decreased serum sodium levels, a marked increase in serum TNF-? and ET-1 levels as well as significantly upregulated renal type 1 inositol 1,4,5-trisphosphate receptor (IP3R1) expression. The upregulated IP3R1 expression was abrogated by the treatment of anti-TNF-? antibodies, but not by 2-aminoethoxydiphenylborate (2-APB), which blocks the inositol 1,4,5-trisphosphate signaling pathway. Treatments with either TNF-? antibodies or 2-APB also significantly improved the compromised GFR, elevated serum urea nitrogen and creatinine levels, and reversed the decrease in glomerular inulin space and the increase in glomerular calcium content in GalN/LPS-exposed rats. The extent of acute liver injury as reflected by serum ALT levels was much more attenuated by anti-TNF-? antibodies than by 2-APB. Liver histology further confirmed that anti-TNF-? antibodies conferred better protection than 2-APB in GalN/LPS-exposed rats. LPS-elicited TNF-? over-production is responsible for decreased GFR through IP3R1 overexpression, and the compromised GFR resulted in the development of acute renal failure in rats with FHF. PMID:24887412

Wang, Jing-Bo; Wang, Dong-Lei; Wang, Hai-Tao; Wang, Zhao-Han; Wen, Ying; Sun, Cui-Ming; Zhao, Yi-Tong; Wu, Jian; Liu, Pei



Mycobacterial 65-kilodalton heat shock protein induces tumor necrosis factor alpha and interleukin 6, reactive nitrogen intermediates, and toxoplasmastatic activity in murine peritoneal macrophages.  

PubMed Central

The 65-kDa heat shock protein (Hsp65) is supposed to play a role in host defense against infections with various microbial pathogens and in autoimmune inflammatory disorders. These effects are thought to result mainly from an Hsp65-specific T-lymphocyte-mediated immune response that recognizes conserved epitopes. The aim of the present study was to assess whether mycobacterial Hsp65 has a direct effect on resident murine peritoneal macrophages, independent of Hsp65-sensitized T lymphocytes. Exposure of peritoneal macrophages from naive C57BL/6 mice to the mycobacterial Hsp65 in vitro induced an enhanced release of tumor necrosis factor alpha (TNF-alpha) and interleukin 6. These cells also produced large amounts of reactive nitrogen intermediates (RNI) and inhibited the intracellular proliferation of Toxoplasma gondii. Small amounts of gamma interferon acted synergistically with Hsp65. Thus, exposure of murine macrophages to Hsp65 results in activation of these cells. The acquisition of these characteristics by peritoneal macrophages occurred in the absence of sensitized T lymphocytes. Addition of anti-TNF-alpha antiserum resulted in an attenuation of the Hsp65-induced release of RNI and toxoplasmastatic activity, indicating that endogenous TNF-alpha is involved in the Hsp65-induced macrophage activation. The conclusion of this study is that in vitro exposure of peritoneal macrophages to the mycobacterial Hsp65 induces the release of proinflammatory cytokines and RNI and results in inhibition of the intracellular proliferation of T. gondii. These effects on murine macrophages occur independently of Hsp65-specific T lymphocytes. The proinflammatory effect of Hsp65 demonstrated in this study suggests that this heat shock protein may play a role in the initiation of inflammation that adds to a non-species-specific resistance in the early stages of infections.

Peetermans, W E; Raats, C J; van Furth, R; Langermans, J A



Mechanistic links between oxidative/nitrosative stress and tumor necrosis factor alpha in letrozole-induced murine polycystic ovary: biochemical and pathological evidences for beneficial effect of pioglitazone.  


This study aimed to investigate the possible relationship between ovarian functionality and the oxidative response during cystogenesis induced by hyperandrogenization with letrozole and examine protective effect of the peroxisome proliferator-activated receptor gamma (PPAR-?) agonist, pioglitazone (PIO), in polycystic ovary (PCO). Ovarian cysts were induced by oral administration of letrozol (1 mg/kg/day) for 21 consecutive days in the female rats. Effective dose of PIO (20 mg/kg/day) was administrated orally for 21 days. Serum estradiol (E), progesterone (P), testosterone (T), and the ovarian immunomodulator prostaglandin E (PGE) were analyzed as biomarkers of ovarian function. To determine the role of oxidative stress in PCO, the level of cellular lipid peroxidation (LPO), superoxide dismutase (SOD), catalase (CAT), glutathione peroxidase (GPx), and peroxynitrite (ONOO), and tumor necrosis factor alpha (TNF-?) as a marker of inflammation and apoptosis were measured in serum and the ovaries. Letrozole-induced PCO in rats exhibited a significant increase in LPO and ONOO in serum and ovary while significantly decreased serum and ovarian SOD, CAT, and GPx. Serum T and TNF-?, and ovarian PGE were increased in animals with cysts compared with healthy controls, while E and P diminished. When compared to control group, letrozole-treated group showed irregular sexual cycles, polycystic ovaries characterized by high incidence of sub-capsular ovarian cyst with diminished or scant granulosa cell layer, increased number of atretic pre-antral and antral follicles and absence of corpus luteum. There were almost no primary, secondary, and tertiary follicles observed in PCO rats. All measured parameters were improved by PIO and reached close to normal levels. The present study further supports the role of oxidative/nitrosative stress and infiammatory responses in the pathogenesis of letrozole-induced hyperandrogenic PCO rats. Results indicate that PIO is able to exert direct antioxidative and anti-inflammatory effects on the endocrine, biochemical, and pathological alterations independent of its possible effects mediated via increased insulin sensitivity in hyperandrogenized PCO. PMID:22076494

Rezvanfar, M A; Rezvanfar, M A; Ahmadi, A; Saadi, H A Shojaei; Baeeri, M; Abdollahi, M



Proteomic analysis of tumor necrosis factor-alpha (TNF-?)-induced L6 myotube secretome reveals novel TNF-?-dependent myokines in diabetic skeletal muscle.  


There is a strong possibility that skeletal muscle can respond to irregular metabolic states by secreting specific cytokines. Obesity-related chronic inflammation, mediated by pro-inflammatory cytokines, is believed to be one of the causes of insulin resistance that results in type 2 diabetes. Here, we attempted to identify and characterize the members of the skeletal muscle secretome in response to tumor necrosis factor-alpha (TNF-?)-induced insulin resistance. To conduct this study, we comparatively analyzed the media levels of proteins released from L6 skeletal muscle cells. We found 28 TNF-? modulated secretory proteins by using separate filtering methods: Gene Ontology, SignalP, and SecretomeP, as well as the normalized Spectral Index for label-free quantification. Ten of these secretory proteins were increased and 18 secretory proteins were decreased by TNF-? treatment. Using microarray analysis of Zuker diabetic rat skeletal muscle combined with bioinformatics and Q-PCR, we found a correlation between TNF-?-mediated insulin resistance and type 2 diabetes. This novel approach combining analysis of the conditioned secretome and transcriptome has identified several previously unknown, TNF-?-dependent secretory proteins, which establish a foothold for research on the different causes of insulin resistance and their relationships with each other. PMID:22023146

Yoon, Jong Hyuk; Song, Parkyong; Jang, Jin-Hyeok; Kim, Dae-Kyum; Choi, Sunkyu; Kim, Jaeyoon; Ghim, Jaewang; Kim, Dayea; Park, Sehoon; Lee, Hyeongji; Kwak, Dongoh; Yea, Kyungmoo; Hwang, Daehee; Suh, Pann-Ghill; Ryu, Sung Ho



Mannose-capped Lipoarabinomannan from Mycobacterium tuberculosis induces soluble tumor necrosis factor receptor production through tumor necrosis factor alpha-converting enzyme activation.  


Primary Mycobacterium tuberculosis infection results in granuloma formation in lung tissue. A granuloma encapsulates mycobacterium-containing cells, thereby preventing dissemination and further infection. Tumor necrosis factor alpha (TNF-?) is a host-protective cytokine during M. tuberculosis infection due to its role in promoting and sustaining granuloma formation. TNF activity is regulated through the production of soluble TNF receptors (sTNFRI and sTNFRII). Therefore, we examined the potential production of endogenous sTNFRs during M. tuberculosis infection. Using the murine model of aerosol M. tuberculosis infection, we determined that levels of sTNFR production were elevated in bronchoalveolar lavage fluid 1 month following infection. An investigation of M. tuberculosis cell wall components identified that the known virulence factor mannose-capped lipoarabinomannan (ManLAM) was sufficient to induce sTNFR production, with sTNFRII being produced preferentially compared with sTNFRI. ManLAM stimulated the release of sTNFRs without TNF production, which corresponded to an increase in TNF-?-converting enzyme (TACE) activity. To determine the relevance of these findings, serum samples from M. tuberculosis-infected patients were tested and found to have an increase in the sTNFRII/sTNFRI ratio. These data identify a mechanism by which M. tuberculosis infection can promote the neutralization of TNF and furthermore suggest the potential use of the sTNFRII/sTNFRI ratio as an indicator of tuberculosis disease. PMID:22927046

Richmond, Jillian M; Duffy, Elizabeth R; Lee, Jinhee; Kaboli, Kavon; Kim, Yun Seong; Remick, Daniel G; Kornfeld, Hardy; Cruikshank, William W



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  

Microsoft Academic Search

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-) 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

Ursula Deiters; Marina Gumenscheimer; Chris Galanos; Peter F. Muhlradt



Tumor necrosis factor alpha- and inducible nitric oxide synthase-producing dendritic cells are rapidly recruited to the bladder in urinary tract infection but are dispensable for bacterial clearance.  


The role of dendritic cells (DC) in urinary tract infections (UTI) is unknown. These cells contribute directly to the innate defense against various viral and bacterial infections. Here, we studied their role in UTI using an experimental model induced by transurethral instillation of the uropathogenic Escherichia coli (UPEC) strain 536 into C57BL/6 mice. While few DC were found in the uninfected bladder, many had been recruited after 24 h, mostly to the submucosa and uroepithelium. They expressed markers of activation and maturation and exhibited the CD11b+ F4/80+ CD8- Gr-1- myeloid subtype. Also, tumor necrosis factor alpha (TNF-alpha)- and inducible nitric oxide synthase (iNOS)-producing CD11bINT DC (Tip-DC) were detected, which recently were proposed to be critical in the defense against bacterial infections. However, Tip-DC-deficient CCR2-/- mice did not show reduced clearance of UPEC from the infected bladder. Moreover, clearance was also unimpaired in CD11c-DTR mice depleted of all DC by injection of diphtheria toxin. This may be explained by the abundance of granulocytes and of iNOS- and TNF-alpha-producing non-DC that were able to replace Tip-DC functionality. These findings demonstrate that some of the abundant DC recruited in UTI contributed innate immune effector functions, which were, however, dispensable in the microenvironment of the bladder. PMID:16966414

Engel, Daniel; Dobrindt, Ulrich; Tittel, André; Peters, Petra; Maurer, Juliane; Gütgemann, Ines; Kaissling, Brigitte; Kuziel, William; Jung, Steffen; Kurts, Christian



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

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.

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



Blocking the janus-activated kinase pathway reduces tumor necrosis factor alpha-induced interleukin-18 bioactivity by caspase-1 inhibition  

PubMed Central

Introduction Our objective was to examine the role of the janus-activated kinase (JAK) pathway in the modulation of tumor necrosis factor-? (TNF)-induced-IL-18 bioactivity by reduction of caspase-1 function. Methods Caspase-1 expression in rheumatoid arthritis (RA) synovial fibroblasts treated with TNF was assessed by qRT-PCR and Western blot. Interleukin (IL)-18 was assessed by enzyme-linked immunosorbent assay (ELISA) in cell lysates and conditioned media and detected by immunofluorescence (IF) staining in RA synovial fibroblasts. The critical pathways for TNF-induced caspase-1 expression were determined by using chemical inhibitors of signaling followed by TNF stimulation. IL-18 bioactivity was assessed using human myelomonocytic KG-1 cells. Results TNF induced RA synovial fibroblast caspase-1 expression at the protein level in a time-dependant manner (P?induced-caspase-1 expression at the transcriptional and protein levels by approximately 60% and 40%, respectively (P?induced-caspase-1 expression at the transcriptional, protein, and activity levels by approximately 60%, 40%, and 53%, respectively (P?induced IL-18 and investigated roles of the ERK1/2 and JAK pathways. Blocking the JAK pathway, TNF induced intracytoplasmic granular IL-18 expression suggesting a defect of caspase-1. Finally, blocking the JAK pathway, we observed a reduction of IL-18 bioactivity by 52% in RA synovial fibroblasts (P?induced-IL-18 bioactivity by blocking capase-1. These data present a novel role for JAK inhibition in RA patients and emphasize JAK inhibition use as a new therapeutic option in RA management.



Adalimumab (tumor necrosis factor-blocker) reduces the expression of glial fibrillary acidic protein immunoreactivity increased by exogenous tumor necrosis factor alpha in an organotypic culture of porcine neuroretina  

PubMed Central

Purpose To determine if exogenous addition of tumor necrosis factor alpha (TNF?) exacerbates retinal reactive gliosis in an organotypic culture of porcine neuroretina and to evaluate if concomitant adalimumab, a TNF-blocker, diminishes it. Methods Porcine retinal explants from 20 eyeballs were cultured. Cultures with 100 pg/ml TNF?, 10 µg/ml adalimumab, 100 pg/ml TNF? plus 10 µg/ml adalimumab, or controls without additives were maintained for 9 days. Freshly detached retinas were processed in parallel. TNF? levels in control culture supernatants were quantified with enzyme-linked immunosorbent assay. Cryostat sections were doubly immunostained for glial fibrillary acidic protein (GFAP), a marker for reactive gliosis, and cellular retinaldehyde-binding protein (CRALBP), a marker for Müller cells. Sections were also labeled with the isolectin IB4, a label for microglia/macrophages. Results TNF? in control culture supernatants was detected only at day 1. Compared to the fresh neuroretinal samples, upregulation of GFAP and downregulation of CRALBP occurred during the 9 days of culture. Exogenous TNF? stimulated glial cells to upregulate GFAP and downregulate CRALBP immunoreactivity. TNF?-treated cultures also initiated the growth of gliotic membranes and underwent retinal disorganization. Adalimumab inhibited the spontaneous increases in GFAP and maintained CRALBP. In combination with TNF?, adalimumab reduced GFAP expression and conserved CRALBP, with only slight retinal disorganization. No appreciable changes in IB4 labeling were observed under the different culture conditions. Conclusions In cultured porcine neuroretina, spontaneous reactive gliosis and retinal disorganization were exacerbated by exogenous TNF?. Adalimumab reduced spontaneous changes and those induced by TNF?. Therefore, inhibiting TNF? may represent a novel approach to controlling retinal fibrosis observed in some human diseases.

Garcia-Gutierrez, M.T.; Srivastava, G.K.; Gayoso, M.J.; Gonzalo-Orden, J.M.; Pastor, J.C.



Promotion of mouse fibroblast collagen gene expression by mast cells stimulated via the Fc epsilon RI. Role for mast cell-derived transforming growth factor beta and tumor necrosis factor alpha  

PubMed Central

Chronic allergic diseases and other disorders associated with mast cell activation can also be associated with tissue fibrosis, but a direct link between mast cell mediator release and fibroblast collagen gene expression has not been established. Using in situ hybridization, we show that the elicitation of an IgE-dependent passive cutaneous anaphylaxis (PCA) reaction in mice results in a transient, but marked augmentation of steady state levels of type alpha-1 (I) collagen mRNA in the dermis. While peak levels of collagen mRNA expression in the skin are observed 16-24 h after mast cell activation, substantial numbers of dermal cells are strongly positive for collagen mRNA at 1 and 2 h after antigen challenge, before circulating inflammatory cells are recruited into the tissues. Furthermore, experiments in mast cell- reconstituted or genetically mast cell-deficient WBB6F1-W/Wv mice demonstrate that the increased expression of collagen mRNA at sites of PCA reactions is entirely mast cell dependent. In vitro studies show that the supernatants of mouse serosal mast cells activated via the Fc epsilon RI markedly increase type alpha-1 (I) collagen mRNA levels in mouse embryonic skin fibroblasts, and also upregulate collagen secretion by these cells. The ability of mast cell supernatants to induce increased steady state levels of collagen mRNA in mouse skin fibroblasts is markedly diminished by absorption with antibodies specific for either of two mast cell-derived cytokines, transforming growth factor beta (TGF-beta 1) or tumor necrosis factor alpha (TNF- alpha), and is eliminated entirely by absorption with antibodies against both cytokines. Taken together, these findings demonstrate that IgE-dependent mouse mast cell activation can induce a transient and marked increase in steady state levels of type alpha-1 (I) collagen mRNA in dermal fibroblasts and that mast cell-derived TGF-beta 1 and TNF-alpha importantly contribute to this effect.



Up-regulation of tumor necrosis factor-alpha and interferon-gamma expression in the spleen and lungs of mice infected with the human Babesia isolate WA1  

Microsoft Academic Search

We analyzed cytokine expression in mice infected with the intraerythrocytic parasites Babesia microti and WA1. In C3H\\/HeN mice, WA1 infections were fatal, whereas B. microti infections were resolved. We propose that the proinflammatory cytokines tumor necrosis factor-alpha (TNF?) and interferon-gamma\\u000a (IFN?) contribute to the WA1-associated disease. WA1 infection was characterized by up-regulation of TNF? and IFN? mRNA in\\u000a the spleen.

Ruth M. Hemmer; David A. Ferrick; Patricia A. Conrad



A Role for Protein Phosphatase 2A in Regulating p38 Mitogen Activated Protein Kinase Activation and Tumor Necrosis Factor-Alpha Expression during Influenza Virus Infection.  


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. PMID:23549267

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



Interaction of an Adenovirus E3 14.7-Kilodalton Protein with a Novel Tumor Necrosis Factor Alpha-Inducible Cellular Protein Containing Leucine Zipper Domains  

Microsoft Academic Search

Early region 3 (E3) of group C human adenoviruses (Ad) encodes several inhibitors of tumor necrosis factor alpha (TNF-a) cytolysis, including an E3 14.7-kDa protein (E3-14.7K) and a heterodimer containing two polypeptides of 10.4 and 14.5 kDa. To understand the mechanism by which the viral proteins inhibit TNF-a functions, the E3-14.7K protein was used to screen a HeLa cell cDNA



Activation of human endothelial cells by tumor necrosis factor-alpha results in profound changes in the expression of glycosylation-related genes.  


The endothelium plays a central role in the logistics of the immune system by allowing the selective transmigration of leukocytes, as well as the maintenance of the circulation and coagulation homeostasis. Evidence is increasing that the carbohydrate composition of the endothelial cell surface is critical for the cells to exert their physiological function. The major aim of this study is to unravel the mechanisms underlying the expression of carbohydrate structures by endothelial cells, which are involved in leukocyte adhesion and migration. Using quantitative real-time PCR, the expression profile of a selected group of 74 glycosylation-related genes has been determined in human umbilical vein endothelial cells (HUVEC) and human foreskin microvascular endothelial cells (FMVEC) under control and TNFalpha-induced conditions. The set of genes comprised 59 glycosyltransferases, 6 mannosidases and 9 sulfotransferases. In parallel, the overall cell surface glycan profile has been assessed by the use of glycan-specific lectins and monoclonal antibodies. The results demonstrate that HUVEC and FMVEC differ substantially in the expression of glycosylation-related genes and, accordingly, also in the presence of different glycan epitopes on the cell membrane. Induction of an inflamed phenotype of the cells by treatment with TNFalpha differentially modulates a set of these genes in HUVEC and FMVEC resulting in a change in the cell membrane associated glycans that are of importance in inflammation-related endothelial cell-surface processes. PMID:16080149

García-Vallejo, Juan J; Van Dijk, Willem; Van Het Hof, Bert; Van Die, Irma; Engelse, Marten A; Van Hinsbergh, Victor W M; Gringhuis, Sonja I



Expression of NADPH oxidases and enhanced H(2)O(2)-generating activity in human coronary artery endothelial cells upon induction with tumor necrosis factor-alpha.  


Tumor necrosis factor (TNF)-alpha, which potentiates reactive oxygen species (ROS) generation, is crucial for the development of coronary arteritis and aneurysm in Kawasaki disease. We hypothesized that vascular NADPH oxidase (Nox) enzymes participate in the TNF-alpha-triggered endothelial damage through elevating ROS generation. Thus, we herein examine the expression of Nox enzymes in human coronary artery endothelial cells (HCAEC) and the effects of TNF-alpha on Nox-mediated ROS generation. We show that HCAEC in culture spontaneously generate H(2)O(2) at basal level (0.53 nmol/min/mg protein). In searching for Nox components responsible for the H(2)O(2) generation, two distinct isoforms of Nox4 are found expressed in HCAEC: the prototype Nox4A and the shorter Nox4B, respectively in the postnuclear supernatant and the nuclear fractions. Other expressed Nox family components are: as mRNAs, Nox4C, Nox4D, Nox1, p51(nox), and Racs; as mRNAs and proteins, Nox2, p22(phox), p47(phox), and p67(phox). The H(2)O(2)-generating activity increases up to three-fold upon inclusion of TNF-alpha in culture, concomitantly with augmented expressions of Nox4A, p22(phox), p47(phox) and p67(phox) proteins. Together, these results suggest that Nox2 and Nox4A enzymes are induced by TNF-alpha endowing HCAEC with enhanced ROS-generating activity, which may play a role in the initial endothelial dysfunction through oxidative stress. PMID:18687299

Yoshida, Lucia S; Tsunawaki, Shohko



The role of 12/15-lipoxygenase in the expression of interleukin-6 and tumor necrosis factor-alpha in macrophages.  


12/15-lipoxygenase (12/15-LO) enzyme and products have been associated with inflammation and atherosclerosis. However, the mechanism of effects of the 12/15-LO products has not been fully clarified. To study the role of 12/15-LO in cytokine expression, experiments with direct additions of the12/15-LO products, 12(S)-hydroxyeicosa tetraenoic acid or 12(S)-hydroperoxyeicosa-5Z, 8Z, 10E, or 14Z-tetraenoic acid to macrophages were first carried out, and results showed that the 12/15-LO products stimulated mRNA and protein expression of IL-6 and TNF-alpha in a dose-dependent manner. In contrast, an inactive analogue of 12(S)-hydroxyeicosa tetraenoic acid had no effect. To further explore the role of endogenous 12/15-LO in cytokine expression, we used an in vitro and in vivo model to test the effect of 12/15-LO overexpression. The models included Plox-86 cells, a J774A.1 cell line that stably overexpresses leukocyte-type 12/15-LO and primary mouse peritoneal macrophages (MPMs) from 12/15-LO transgenic mice. The results showed a clear increase in IL-6 and TNF-alpha expression in Plox-86 cells and MPMs from 12/15-LO transgenic mice, compared with mock-transfected J774A.1 cells and MPMs from control C57BL6 mice. IL-1beta, IL-12, and monocyte chemoattractant protein (MCP)-1 mRNA were also increased in Plox-86 cells. These data clearly suggest a clear role of 12/15-LO pathway in cytokine production. We also demonstrated that signaling pathways including protein kinase C, p38 MAPK (p38), c-jun NH(2)-terminal kinase as well as nicotinamide adenine dinucleotide phosphate oxidase are important for 12-(S)-hydroxyeicosatetraenoic acid-induced increases in IL-6 and TNF-alpha gene expression. These results suggest a potentially important mechanism linking 12/15-LO activation to chronic inflammation and atherosclerosis. PMID:17170102

Wen, Yeshao; Gu, Jiali; Chakrabarti, Swarup K; Aylor, Kevin; Marshall, John; Takahashi, Yoshitaka; Yoshimoto, Tanihiro; Nadler, Jerry L



Tumor necrosis factor-alpha-induced cell killing and activation of transcription factor NF-kappaB are uncoupled in L929 cells.  


The induction of transcription factor NF-kappaB has been shown to counteract tumor necrosis factor (TNF)-alpha-induced cell death in various cell types. In this study, we investigated the role of NF-kappaB for TNF-alpha-triggered cell death in the widely used mouse cell line L929 by various approaches. Inhibition of the mitochondrial permeability transition by bongkrekic acid impaired TNF-alpha-induced cell death without affecting the activity of NF-kappaB. The reduction of NF-kappaB-mediated gene expression by the synthetic steroid dexamethasone was associated with a decrease in TNF-alpha-mediated cell killing, suggesting that NF-kappaB does not protect L929 cells from TNF-alpha-induced cell death. This concept was reinforced by experiments employing L929 cell lines stably overexpressing a transdominant negative form of IkappaB-alpha. These cell lines were unable to activate NF-kappaB and to inducibly express the IL-6 gene, but they showed the same susceptibility toward TNF-alpha-mediated cell death as L929 wild-type cells. PMID:9660769

Hehner, S P; Hofmann, T G; Ratter, F; Dumont, A; Dröge, W; Schmitz, M L



Eukaryotic translation initiation factor 5A induces apoptosis in colon cancer cells and associates with the nucleus in response to tumour necrosis factor {alpha} signalling  

SciTech Connect

Eukaryotic translation initiation factor 5A (eIF5A) is thought to function as a nucleocytoplasmic shuttle protein. There are reports of its involvement in cell proliferation, and more recently it has also been implicated in the regulation of apoptosis. In the present study, we examined the effects of eIF5A over-expression on apoptosis and of siRNA-mediated suppression of eIF5A on expression of the tumour suppressor protein, p53. Over-expression of either eIF5A or a mutant of eIF5A incapable of being hypusinated was found to induce apoptosis in colon carcinoma cells. Our results also indicate that eIF5A is required for expression of p53 following the induction of apoptosis by treatment with Actinomycin D. Depiction of eIF5A localization by indirect immunofluorescence has indicated, for the first time, that the protein is rapidly translocated from the cytoplasm to the nucleus by death receptor activation or following treatment with Actinomycin D. These findings collectively indicate that unhypusinated eIF5A may have pro-apoptotic functions and that eIF5A is rapidly translocated to the nucleus following the induction of apoptotic cell death.

Taylor, Catherine A. [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Sun Zhong [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Cliche, Dominic O. [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Ming, Hong [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Eshaque, Bithi [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Jin Songmu [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Hopkins, Marianne T. [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Thai, Boun [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada); Thompson, John E. [Department of Biology, University of Waterloo, 200 University Ave. W., Waterloo, Ontario, N2L 3G1 (Canada)]. E-mail:



Protective effect of photodegradation product of nifedipine against tumor necrosis factor alpha-induced oxidative stress in human glomerular endothelial cells.  


Recently, increasing evidence suggests that the antihypertensive drug nifedipine acts as a protective agent for endothelial cells, and that the activity is unrelated to its calcium channel blocking. Nitrosonifedipine (NO-NIF) is metabolically and photochemically produced from nifedipine, and NO-NIF has been recognized as a contaminant of nifedipine because it has no antihypertensive effect. Treatment of tumor necrosis factor-? (TNF-?) suppressed the cell viability and facilitated the expression of Inter-Cellular Adhesion Molecule 1(ICAM-1) in human glomerular endothelial cells (HGECs) though, pretreatment of NO-NIF significantly recovered the TNF-?-induced cell damage to the same extent as Trolox-C did, and suppressed the ICAM-1 expression in a concentration dependent manner. In addition, NO-NIF inhibited the cell toxicity induced by cumene hydroperoxide, which hampers the integrity of cell membrane through oxidative stress, as effective as Trolox-c. These data suggest that NO-NIF is a candidate for a new class of antioxidative drug that protect cells against oxidative stress in glomerular endothelial cells. PMID:21372496

Fukuhara, Yayoi; Tsuchiya, Koichiro; Horinouchi, Yuya; Tajima, Soichiro; Kihira, Yoshitaka; Hamano, Shuichi; Kawazoe, Kazuyoshi; Ikeda, Yasumasa; Ishizawa, Keisuke; Tomita, Shuhei; Tamaki, Toshiaki



Tumor necrosis factor alpha induces a serotonin dependent early increase in ciliary beat frequency and epithelial transport velocity in murine tracheae.  


The tracheal epithelium prevents via its highly effective clearance mechanism the contamination of the lower airways by pathogens. This mechanism is driven by ciliary bearing cells which are not only in contact with the gas phase; in addition they are also influenced by inflammatory mediators. These mediators can alter the protective function of the epithelium. Since the pro-inflammatoric cytokine tumor necrosis factor-? (TNF-?) plays a pivotal role within the inflammatory cascade, we investigated its effect onto the tracheal epithelium measured by its ciliary beat frequency and the particle transport velocity. In organ explant experiments the ciliary beat frequency and the particle transport velocity were measured under the application of TNF-? using tracheae from male C57BL6J mice. We observed a dose dependent TNF-? induced increase of both particle transport velocity and ciliary beat frequency. Knock out mice experiments made evident that the increase was depended on the expression of tumor necrosis factor receptor 1 (TNF-R1). The increases in ciliary beat frequency as well as the accelerated particle transport velocity were either inhibited by the unspecific serotonin antagonist methysergide or by cyproheptadine a specific 5-HT2 receptor antagonist. Thus, acetylcholine antagonists or nitric oxide synthase (NOS) inhibitors failed to inhibit the TNF-? induced activation. In conclusion, TNF-? may play a pivotal role in the protection of lower airways by inducing ciliary activity and increase in particle transport velocity via TNF-R1 and 5-HT2 receptor. PMID:24626175

Weiterer, Sebastian; Schulte, Dagmar; Müller, Sabrina; Kohlen, Thomas; Uhle, Florian; Weigand, Markus A; Henrich, Michael



Comparative in vitro toll-like receptor ligand induced cytokine profiles of Toda and Murrah buffaloes-Identification of tumour necrosis factor alpha promoter polymorphism.  


The objective of this study was to assess cytokine production upon activation of pattern recognition receptors responsible for sensing bacterial and viral pathogen associated molecular patterns in two genetically diverse buffalo breeds, Toda and Murrah. A very limited molecular-epidemiological analysis showed a higher prevalence of Anaplasma and Theileria in Murrah than Toda buffaloes. Toda buffalo peripheral blood mononuclear cells (PBMC) produced significantly higher levels of IFN ? and/or TNF ? mRNAs in response to peptidoglycan, poly I:C, lipopolysaccharide, imiquimod and CpG. Flagellin stimulation did not result in any significant differences in the expression levels of the cytokines tested between these breeds. The levels of ligand induced IFN ? and TNF ? mRNA and proteins also correlated except when induced with CpG. The proximal promoter region of TNF ? across these two breeds were also sequenced to detect SNPs and promoter assay performed to determine their role in altering the transcriptional activity. Two polymorphisms were identified at -737 (T/A) and -1092 (G/T) positions in Toda buffalo TNF ? promoter and promoter assay revealed higher transcription activity in Toda buffalos than in Murrah. This suggests that disease tolerance of these buffalo breeds could be due to the differences in their cytokine transcription levels in response to the respective PAMPs that may be at least in part determined by polymorphisms in the cytokine promoter regions. PMID:23084344

Vignesh, A R; Dhinakar Raj, G; Dhanasekaran, S; Tirumurugaan, K G; Raja, A



Up-regulation of tumor necrosis factor-alpha and interferon-gamma expression in the spleen and lungs of mice infected with the human Babesia isolate WA1.  


We analyzed cytokine expression in mice infected with the intraerythrocytic parasites Babesia microti and WA1. In C3H/HeN mice, WA1 infections were fatal, whereas B. microti infections were resolved. We propose that the proinflammatory cytokines tumor necrosis factor-alpha (TNFalpha) and interferon-gamma (IFNgamma) contribute to the WA1-associated disease. WA1 infection was characterized by up-regulation of TNFalpha and IFNgamma mRNA in the spleen. Previous studies in WA1-infected mice showed that pathologic lesions occurred primarily in the lungs, including pulmonary edema and intravascular margination of leukocytes. Analysis of cytokine expression in the lungs is important for an understanding of the disease process in WA1-infected mice. Expression of both TNFalpha and IFNgamma mRNA was increased in the lungs of WA1-infected mice. Immunohistochemical staining confirmed the upregulation of these proinflammatory cytokines in the lungs. Expression of TNFalpha and IFNgamma was not up-regulated in the lungs of B. microti-infected mice. The results implicate TNFalpha and IFNgamma in the pathogenesis of WA1-associated disease. PMID:10685843

Hemmer, R M; Ferrick, D A; Conrad, P A



Tumor necrosis factor alpha mediates resistance to Trypanosoma cruzi infection in mice by inducing nitric oxide production in infected gamma interferon-activated macrophages.  

PubMed Central

Cell invasion by Trypanosoma cruzi and its intracellular replication are essential for continuation of the parasite life cycle and for production of Chagas' disease. T. cruzi is able to replicate in nucleated cells and can be killed by activated macrophages. Gamma interferon (IFN-gamma) is one of the major stimuli for the activation of macrophages and has been shown to be a key activation factor for the killing of intracellular parasites through a mechanism dependent upon nitric oxide (NO) biosynthesis. We show that although the addition of exogenous tumor necrosis factor alpha (TNF-alpha) does not potentiate the trypanocidal activity of IFN-gamma in vitro, treatment of resistant C57BI/6 mice with an anti-TNF-alpha monoclonal antibody increased parasitemia and mortality. In addition, the anti-TNF-alpha-treated animals had decreased NO production, both in vivo and in vitro, suggesting an important role for TNF-alpha in controlling infection. In order to better understand the role of TNF-alpha in the macrophage-mediating killing of parasites, cultures of T. cruzi-infected macrophages were treated with an anti-TNF-alpha monoclonal antibody. IFN-gamma-activated macrophages failed to kill intracellular parasites following treatment with 100 micrograms of anti-TNF-alpha. In these cultures, the number of parasites released at various time points after infection was significantly increased while NO production was significantly reduced. We conclude that IFN-gamma-activated macrophages produce TNF-alpha after infection by T. cruzi and suggest that this cytokine plays a role in amplifying NO production and parasite killing.

Silva, J S; Vespa, G N; Cardoso, M A; Aliberti, J C; Cunha, F Q



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


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-1? (Hif-1?), the master regulatory protein of hypoxia, in MSCs remain unclear, due to the limited span of Hif-1? stabilization and the complex network of hypoxic responses. In this study, to further define the significance of Hif-1? 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-1? during such long-term biological processes. Using this model, we show that the stabilization of Hif-1? 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-1? 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-1? proteins in maintaining the primitive state of primary MSCs and on the cellular heterogeneities in hypoxic responses among MSC populations. PMID:24071737

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




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...


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

PubMed Central

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

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



Peptidoglycan and Lipoteichoic Acid from Staphylococcus aureus Induce Tumor Necrosis Factor Alpha, Interleukin 6 (IL-6), and IL-10 Production in Both T Cells and Monocytes in a Human Whole Blood Model  

PubMed Central

We have examined the ability of peptidoglycan (PepG) and lipoteichoic acid (LTA) isolated from Staphylococcus aureus to induce the release of tumor necrosis factor alpha (TNF-?), interleukin-6 (IL-6), and IL-10 in whole human blood and identified the cellular origins of these cytokines. Both PepG and LTA induced transient increases in TNF-? and IL-10 in plasma, with peak values at 6 and 12 h, respectively. IL-6 values increased throughout the experimental period (24 h). The TNF-?, IL-6, and IL-10 release induced by PepG and LTA was dose dependent. Only PepG was a potent inducer of TNF-? secretion. After stimulation of whole blood with PepG or LTA, very pure populations of monocytes (CD14 positive), T cells (CD2 positive), B cells (CD19 positive), and granulocytes (CD15 positive) were isolated by immunomagnetic separation and analyzed by reverse transcription-PCR for mRNA transcripts encoding TNF-?, IL-6, and IL-10. The TNF-? mRNA results were inconclusive. In contrast, PepG induced IL-6 and IL-10 mRNA accumulation in both T cells and monocytes. LTA, as well as lipopolysaccharide, induced IL-6 and IL-10 mRNA production in monocytes and possibly in T cells. Whether granulocytes and B cells produce cytokines in response to bacterial stimuli remains obscure. Blockade of the CD14 receptors with monoclonal antibodies (18D11) had no influence on the PepG-induced release of TNF-? but attenuated the LTA-induced release of the same cytokine. In conclusion, our data indicate that circulating T cells and monocytes contribute to cytokine production in sepsis caused by gram-positive bacteria.

Wang, J. E.; J?rgensen, P. F.; Almlof, M.; Thiemermann, C.; Foster, S. J.; Aasen, A. O.; Solberg, R.



Overexpression of Transforming Growth Factor alpha in Psoriatic Epidermis  

Microsoft Academic Search

Transforming growth factor alpha (TGF-alpha ) is produced by and required for the growth of epithelial cells and is angiogenic in vivo. Since epidermal hyperplasia and angiogenesis are hallmarks of psoriasis TGF-alpha gene expression was analyzed in epidermal biopsies of normal and psoriatic skin. TGF-alpha messenger RNA and protein are much more abundant in lesional psoriatic epidermis than in normal-appearing

James T. Elder; Gary J. Fisher; Patricia B. Lindquist; Gregory L. Bennett; Mark R. Pittelkow; Robert J. Coffey; Larry Ellingsworth; Rik Derynck; John J. Voorhees



Live Lactobacillus reuteri Is Essential for the Inhibitory Effect on Tumor Necrosis Factor Alpha-Induced Interleukin8 Expression  

Microsoft Academic Search

The mechanism of the apparent anti-inflammatory action of probiotic organisms is unclear. Lactobacillus reuteri is effective in inhibiting colitis in interleukin-10 (IL-10)-deficient mice. Nerve growth factor (NGF), in addition to its activity on neuronal cell growth, has significant anti-inflammatory effects in several experimen- tal systems in vitro and in vivo, including a model of colitis. Our experiments were designed to

Donglai Ma; Paul Forsythe; John Bienenstock



Recombinant human granulocyte macrophage colony-stimulating factor (rhGM-CSF) induces human macrophage production of transforming growth factor-alpha.  


Granulocyte-Macrophage Colony Stimulating Factor (GM-CSF) is known as an inducer of proliferation and functional activation of myeloid cells. This study was carried out to characterize the effect of purified recombinant human GM-CSF (rhGM-CSF) on induction of TGF-alpha in macrophages. Using Northern blot analysis and immunoassays, we show here that rhGM-CSF markedly stimulates production of TGF-alpha messenger RNA and protein in normal tonsil macrophages. The findings are consistent with macrophages being a normal inducible source of TGF-alpha which may be an important mediator of various activities of GM-CSF both in hematopoietic and non-hematopoietic cells. PMID:1934015

Zhu, J Q; Wu, J; Zhu, D X; Scharfman, A; Lamblin, G; Han, K K



Effects of estradiol and progesterone on tumor necrosis factor alpha-induced apoptosis in human hepatoma HuH7 cells  

Microsoft Academic Search

Oxidative stress, including the generation of reactive oxygen species (ROS), is known to be involved in apoptosis. Preventing apoptosis may thereby induce a malignant transformation of liver tumor cells. Estradiol (E2) is a potent endogenous antioxidant. We examined the proapoptotic role of progesterone as well as the antiapoptotic role of E2 in human hepatoma HuH-7 cells in a state of

Xinliang Cheng; Ichiro Shimizu; Ying Yuan; Mei Wei; Mi Shen; Huiwei Huang; Mari Urata; Katsutaka Sannomiya; Hiroshi Fukuno; Tomoko Hashimoto-Tamaoki; Susumu Ito



Adiponectin suppresses IkappaB kinase activation induced by tumor necrosis factor-alpha or high glucose in endothelial cells: role of cAMP and AMP kinase signaling.  


Adiponectin is a protein secreted from adipocytes that exhibits salutary effects in the vascular endothelium by signaling mechanisms that are not well understood. In obesity-related disease states and type 2 diabetes, circulating substances, including tumor necrosis factor-alpha (TNFalpha) and high glucose, activate IkappaB kinase (IKK)beta and reduce the abundance of its substrate, inhibitor of kappaB (IkappaB)alpha, leading to nuclear translocation of the transcription factor NF-kappaB and stimulation of an inflammatory signaling cascade closely associated with endothelial dysfunction. The present study demonstrates that the globular domain of adiponectin (gAd) potently suppresses the activation of IKKbeta by either TNFalpha or high glucose in human umbilical vein endothelial cells and ameliorates the associated loss of IkappaBalpha protein. Interestingly, activation of AMP kinase was substantially more effective than cAMP signaling in suppressing high glucose-induced IKKbeta activity, whereas both pathways were comparably active in suppressing the TNFalpha-induced increase in IKKbeta. Both cAMP/protein kinase A signaling and activation of the AMP kinase pathway played a role in the suppression by gAd of TNFalpha- and high glucose-mediated IKKbeta activation. These findings support an important role for adiponectin in anti-inflammatory signaling in the endothelium and also imply that multiple pathways are involved in the cellular effects of adiponectin. PMID:17940218

Wu, Xiangdong; Mahadev, Kalyankar; Fuchsel, Lauren; Ouedraogo, Raogo; Xu, Shi-Qiong; Goldstein, Barry J



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 Central

The proinflammatory cytokine tumor necrosis factor alpha (TNF-?) 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 SAP0/0 mice generated by gene targeting. We studied the lethal response of SAP0/0 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-1? (IL-1?) for desensitization and clearly involving a liver protein, the other by tolerizing mice for 5 days using small doses of human TNF-?. Although after IL-1? or after tolerization the SAP levels in the serum had risen fourfold in the control mice and not in the SAP0/0 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-? was not altered in SAP0/0 mice. We conclude that the presence or absence of SAP has no influence on the sensitivity of mice to TNF-?-induced hepatitis, SIRS, and hemorrhagic necrosis or on the endogenous protective mechanisms of desensitization or tolerization.

Van Molle, Wim; Hochepied, Tino; Brouckaert, Peter; Libert, Claude



Tumor necrosis factor-alpha- and hyperglycemia-induced insulin resistance. Evidence for different mechanisms and different effects on insulin signaling.  

PubMed Central

Inhibition of insulin receptor signaling by high glucose levels and by TNF-alpha was recently observed in different cell systems. The aim of the present study was to characterize the mechanism of TNF-alpha-induced insulin receptor inhibition and to compare the consequences of TNF-alpha- and hyperglycemia-induced insulin receptor inhibition for signal transduction downstream from the IR. TNF-alpha (0.5-10 nM) and high glucose (25 mM) showed similar rapid kinetics of inhibition (5-10 min, > 50%) of insulin receptor autophosphorylation in NIH3T3 cells overexpressing the human insulin receptor. TNF-alpha effects were completely prevented by the phosphotyrosine phosphatase (PTPase) inhibitors orthovanadate (40 microM) and phenylarsenoxide (35 microM), but they were unaffected by the protein kinase C (PKC) inhibitor H7 (0.1 mM), the phosphatidylinositol-3 kinase inhibitor wortmannin (5 microM), and the thiazolidindione troglitazone (CS045) (2 microgram/ml). In contrast, glucose effects were prevented by PKC inhibitors and CS045 but unaffected by PTPase inhibitors and wortmannin. To assess effects on downstream signaling, tyrosine phosphorylation of the following substrate proteins of the insulin receptor was determined: insulin receptor substrate-1, the coupling protein Shc, focal adhesion kinase (FAK125), and unidentified proteins of 130 kD, 60 kD. Hyperglycemia (25 mM glucose) and TNF-alpha showed analogous (> 50% inhibition) effects on tyrosine phosphorylation of insulin receptor substrate-1, Shc, p60, and p44, whereas opposite effects were observed for tyrosine phosphorylation of FAK125, which is dephosphorylated after insulin stimulation. Whereas TNF-alpha did not prevent insulin-induced dephosphorylation of FAK125, 25 mM glucose blocked this insulin effect completely. In summary, the data suggest that TNF-alpha and high glucose modulate insulin receptor-signaling through different mechanisms: (a) TNF-alpha modulates insulin receptor signals by PTPase activation, whereas glucose acts through activation of PKC. (b) Differences in modulation of the insulin receptor signaling cascade are found with TNF-alpha and high glucose: Hyperglycemia-induced insulin receptor inhibition blocks both insulin receptor-dependent tyrosine phosphorylation and dephosphorylation of insulin receptor substrate proteins. In contrast, TNF-alpha blocks only substrate phosphorylation, and it does not block insulin-induced substrate dephosphorylation. The different effects on FAK125 regulation allow the speculation that long-term cell effects related to FAK125 activity might develop in a different way in hyperglycemia- and TNF-alpha-dependent insulin resistance.

Kroder, G; Bossenmaier, B; Kellerer, M; Capp, E; Stoyanov, B; Muhlhofer, A; Berti, L; Horikoshi, H; Ullrich, A; Haring, H



Low levels of tumor necrosis factor alpha increase tumor growth by inducing an endothelial phenotype of monocytes recruited to the tumor site.  


Microenvironmental cues instruct infiltrating tumor-associated myeloid cells to drive malignant progression. A subpopulation of tumor-associated myeloid cells coexpressing endothelial and myeloid markers, although rare in peripheral blood, are primarily associated with tumors where they enhance tumor growth and angiogenesis. These biphenotypic vascular leukocytes result from the endothelial differentiation of myeloid progenitors, a process regulated by tumor necrosis factor (TNF)alpha in vitro. An in vivo increase in tumor-derived TNFalpha expression promoted tumor growth and vascularity of mouse melanoma, lung cancer, and mammary tumors. Notably, tumor growth was accompanied by a significant increase in myeloid/endothelial biphenotypic populations. TNFalpha-associated tumor growth, vascularity, and generation of tumor vascular leukocytes in mouse melanoma tumors were dependent on intact host TNFalpha receptors. Importantly, TNFalpha-expressing tumors did not exhibit increased inflammation over control tumors, suggesting a unique action related to myeloid to endothelial differentiation. Our studies suggest that TNFalpha constitutes a tumor microenvironment signal that biases recruited monocytes toward a proangiogenic/provasculogenic myeloid/endothelial phenotype. PMID:19118019

Li, Bin; Vincent, Alicia; Cates, Justin; Brantley-Sieders, Dana M; Polk, D Brent; Young, Pampee P



Serum-induced potentiation of tumor necrosis factor alpha production by human monocytes in response to staphylococcal peptidoglycan: involvement of different serum factors.  

PubMed Central

Peptidoglycan from a Staphylococcus epidermidis strain, isolated from a patient with septicemia, was preincubated with human serum. This mixture was then investigated for its potency to induce tumor necrosis factor (TNF) secretion by human blood monocytes. TNF was measured in the supernatants by using a bioassay and/or an enzyme-linked immunosorbent assay specific for TNF alpha (TNF-alpha). Although earlier studies indicated that staphylococcal peptidoglycan alone is a relatively poor stimulator of TNF-alpha production, the present study shows that human serum highly potentiates peptidoglycan-induced TNF-alpha release by human monocytes. In the presence of serum and in the low-dose range, peptidoglycan was almost as potent as endotoxin. At high peptidoglycan concentrations, monocytes showed an extremely high TNF-alpha response, but again only in the presence of serum. At low peptidoglycan doses, the stimulatory effect of serum was abrogated by heat treatment or depleting serum of complement components C1 and C3/C4, which suggests a role for the classical complement pathway. At high doses of peptidoglycan, the serum stimulatory effect depended mainly on immunoglobulin G.

Mattsson, E; Rollof, J; Verhoef, J; Van Dijk, H; Fleer, A



Tumor necrosis factor alpha and interleukin 1beta are responsible for in vitro myocardial cell depression induced by human septic shock serum.  


Previous studies have demonstrated the presence of myocardial depression in clinical and experimental septic shock. This depression is associated with the presence of a circulating myocardial depressant substance with physical characteristics consistent with cytokines. The present study utilized an in vitro myocardial cell assay to examine the role of various human recombinant cytokines, including tumor necrosis factor (TNF)alpha and interleukin (IL)1beta, in depression of cardiac myocyte contractile function induced by serum from humans with septic shock. The extent and velocity of electrically paced rat cardiac myocytes in tissue culture was quantified by a closed loop video tracking system. Individually, TNF-alpha and IL-1beta each caused significant concentration-dependent depression of maximum extent and peak velocity of myocyte shortening in vitro. In combination, TNF-alpha and IL-1beta induced depression of myocardial cell contractility at substantially lower concentrations consistent with a synergistic effect. Using immunoabsorption, removal of both TNF-alpha and IL-1beta (but not either alone) from the serum of five patients with acute septic shock and marked reversible myocardial depression resulted in elimination of serum myocardial depressant activity. IL-2, -4, -6, -8, -10, and interferon gamma failed to cause significant cardiac myocyte depression over a wide range of concentrations. These data demonstrate that TNF-alpha and IL-1beta cause depression of myocardial cell contraction in vitro and suggest that these two cytokines act synergistically to cause sepsis-associated myocardial depression in humans. PMID:8642298

Kumar, A; Thota, V; Dee, L; Olson, J; Uretz, E; Parrillo, J E



Tumor necrosis factor alpha and interleukin 1beta are responsible for in vitro myocardial cell depression induced by human septic shock serum  

PubMed Central

Previous studies have demonstrated the presence of myocardial depression in clinical and experimental septic shock. This depression is associated with the presence of a circulating myocardial depressant substance with physical characteristics consistent with cytokines. The present study utilized an in vitro myocardial cell assay to examine the role of various human recombinant cytokines, including tumor necrosis factor (TNF)alpha and interleukin (IL)1beta, in depression of cardiac myocyte contractile function induced by serum from humans with septic shock. The extent and velocity of electrically paced rat cardiac myocytes in tissue culture was quantified by a closed loop video tracking system. Individually, TNF-alpha and IL-1beta each caused significant concentration-dependent depression of maximum extent and peak velocity of myocyte shortening in vitro. In combination, TNF-alpha and IL-1beta induced depression of myocardial cell contractility at substantially lower concentrations consistent with a synergistic effect. Using immunoabsorption, removal of both TNF-alpha and IL-1beta (but not either alone) from the serum of five patients with acute septic shock and marked reversible myocardial depression resulted in elimination of serum myocardial depressant activity. IL-2, -4, -6, -8, - 10, and interferon gamma failed to cause significant cardiac myocyte depression over a wide range of concentrations. These data demonstrate that TNF-alpha and IL-1beta cause depression of myocardial cell contraction in vitro and suggest that these two cytokines act synergistically to cause sepsis-associated myocardial depression in humans.



Induced autocrine signaling through the epidermal growth factor receptor contributes to the response of mammary epithelial cells to tumor necrosis factor alpha  

SciTech Connect

In contrast to the well-known cytotoxic effects of tumor necrosis factor a (TNF) in many mammary cancer cells, we have found that TNF stimulates the proliferation and motility of human mammary epithelial cells (HMEC). Since the response of HMEC to TNF is similar to effects mediated by epidermal growth factor receptor (EGFR) activation, we explored the potential role of cross-talk through the EGFR signaling pathways in mediating cellular responses to TNF. Using a microarray enzyme-linked immunoassay, we found that exposure to TNF stimulated the dose-dependent shedding of the EGFR ligand transforming growth factor a (TGFa). Both proliferation and motility of HMEC induced by TNF was prevented either by inhibiting membrane protein shedding with a metalloprotease inhibitor, by blocking EGFR kinase activity, or by limiting ligand-receptor interactions with an antagonistic anti-EGFR antibody. EGFR activity was also necessary for TNF-induced release of MMP-9, a matrix metalloprotease thought to be an essential regulator of mammary cell migration. The cellular response to TNF was associated with a biphasic temporal pattern of extracellular signal-regulated kinase (ERK) phosphorylation, which was EGFR-dependent and modulated by inhibition of metalloprotease-mediated shedding. Significantly, the late phase of ERK phosphorylation, detectable within 4 hours after exposure, was blocked by the metalloprotease inhibitor batimastat, indicating that autocrine signaling through ligand shedding was responsible for this secondary wave of ERK activity. Our results indicate a novel and important role for metalloprotease activation and EGFR transmodulation in mediating the cellular response to TNF.

Chen, Wan-Nan U.; Woodbury, Ronald L.; Kathmann, Loel E.; Opresko, Lee; Zangar, Richard C.; Wiley, H S.; Thrall, Brian D.



Herpes Simplex Virus 1 E3 Ubiquitin Ligase ICP0 Protein Inhibits Tumor Necrosis Factor Alpha-Induced NF-?B Activation by Interacting with p65/RelA and p50/NF-?B1  

PubMed Central

NF-?B plays central roles in regulation of diverse biological processes, including innate and adaptive immunity and inflammation. HSV-1 is the archetypal member of the alphaherpesviruses, with a large genome encoding over 80 viral proteins, many of which are involved in virus-host interactions and show immune modulatory capabilities. In this study, we demonstrated that the HSV-1 ICP0 protein, a viral E3 ubiquitin ligase, was shown to significantly suppress tumor necrosis factor alpha (TNF-?)-mediated NF-?B activation. ICP0 was demonstrated to bind to the NF-?B subunits p65 and p50 by coimmunoprecipitation analysis. ICP0 bound to the Rel homology domain (RHD) of p65. Fluorescence microscopy demonstrated that ICP0 abolished nuclear translocation of p65 upon TNF-? stimulation. Also, ICP0 degraded p50 via its E3 ubiquitin ligase activity. The RING finger (RF) domain mutant ICP0 (ICP0-RF) lost its ability to inhibit TNF-?-mediated NF-?B activation and p65 nuclear translocation and degrade p50. Notably, the RF domain of ICP0 was sufficient to interact with p50 and abolish NF-?B reporter gene activity. Here, it is for the first time shown that HSV-1 ICP0 interacts with p65 and p50, degrades p50 through the ubiquitin-proteasome pathway, and prevents NF-?B-dependent gene expression, which may contribute to immune evasion and pathogenesis of HSV-1.

Zhang, Jie; Wang, Kezhen



Absence of tumor necrosis factor alpha, interleukin-6 (IL-6), and granulocyte-macrophage colony-stimulating factor expression but presence of IL-1beta, IL-8, and IL-10 expression in human monocytes exposed to viable or killed Ehrlichia chaffeensis.  

PubMed Central

Ehrlichia chaffeensis is a recently isolated minute gram-negative obligatory intracellular bacterium of monocytes/macrophages and is the etiologic agent of human monocytic ehrlichiosis. It is not known how macrophages respond when they encounter ehrlichiae in terms of cytokine production. In this study, we examined cytokine mRNA expression by incubating E. chaffeensis with THP-1 cells and performing competitive reverse transcription-PCR (RT-PCR). At 2 h postinfection, the levels of interleukin-1beta (IL-1beta), IL-8, and IL-10 mRNAs were significant but lower than those following Escherichia coli lipopolysaccharide (LPS) stimulation. Unlike the situation with E. coli LPS stimulation, however, IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), and tumor necrosis factor alpha (TNF-alpha) mRNAs were not induced. Time course and dose-response studies confirmed the absence of IL-6, GM-CSF, and TNF-alpha mRNA induction with E. chaffeensis. Viable E. chaffeensis organisms were not required for IL-1beta IO, IL-8, and IL-10 mRNA induction, since heat-killed E. chaffeensis induced identical time course responses. IL-1beta, IL-8, and IL-10 mRNAs were detected for up to 21, 21, and 24 h postexposure with E. chaffeensis, respectively, which were shut off more rapidly than with LPS stimulation. Although heat treatment of E. chaffeensis had no effect, periodate treatment completely abolished the ability of E. chaffeensis to induce IL-1beta, IL-8, and IL-10 mRNAs. The capture enzyme-linked immunosorbent assay result corresponded with the RT-PCR results, showing that viable and heat-killed E. chaffeensis produced and secreted the same levels of IL-1beta and IL-8. IL-10 production was significantly reduced by heat treatment. Periodate-treated ehrlichiae did not induce production of any of the cytokines tested. Anti-CD14 monoclonal antibody and polymyxin B did not inhibit IL-1beta mRNA expression upon exposure to E. chaffeensis. The absence of TNF-alpha, IL-6, and GM-CSF mRNA induction may delay the development of a protective immune response, thereby allowing E. chaffeensis to set up residence in macrophages.

Lee, E H; Rikihisa, Y



Herbal medicine 'Sho-saiko-to' induces tumour necrosis factor-alpha and granulocyte colony-stimulating factor in vitro in peripheral blood mononuclear cells of patients with hepatocellular carcinoma.  


'Sho-saiko-to' (TJ-9) is a Japanese herbal medicine that is commonly administered to patients with chronic viral liver disease in order to improve their overall physical condition and to prevent the development of liver cancer, The present in vitro study demonstrated that, by adding TJ-9 to cell cultures, there were dose-dependent increases in production levels of tumour necrosis factor-alpha (TNF-alpha) and granulocyte colony-stimulating factor (G-CSF) in peripheral mononuclear cells of patients with hepatocellular carcinoma accompanied by liver cirrhosis. Increases in the production of TNF-alpha and G-CSF in control cell cultures exposed to different herbal medicines were low, and this indicates the specificity of the response increases in production of these cytokines to TJ-9. TNF-alpha and G-CSF are known to play important roles in the biological defence mechanism. Administration of TJ-9 may, therefore, be beneficial for patients afflicted with intractable liver diseases because it could mildly induce these cytokines. PMID:8672758

Yamashiki, M; Nishimura, A; Nomoto, M; Suzuki, H; Kosaka, Y



Transforming Growth Factor ?1 Genotypes in Relation to TGF?1, Interleukin-8, and Tumor Necrosis Factor Alpha in Induced Sputum and Blood in Cystic Fibrosis  

PubMed Central

Background. High-producer TGF?1 genotypes are associated with severe lung disease in cystic fibrosis (CF), but studies combining IL-8, TNF?-, and TGF?1(+genotype) levels and their impact on CF lung disease are scarce. Aim. Assessing the relationship between TGF?1, IL-8, and TNF-? and lung disease in CF in an exacerbation-free interval. Methods. Twenty four patients delta F508 homozygous (median age 20.5?y, Shwachman score 75, FEV1(%) 83) and 8 controls (median age 27.5?y) were examined. TGF?1 was assessed in serum and induced sputum (IS) by ELISA, for IL-8 and TNF-? by chemiluminescence in IS and whole blood. Genotyping was performed for TGF?1 C?509T and T+869C utilizing RFLP. Results. TGF?1 in IS (CF/controls median 76.5/59.1?pg/mL, P < 0.074) was higher in CF. There was a negative correlation between TGF?1 in serum and lung function (LF) (FEV1 (r = ?0.488, P = 0.025), MEF 25 (r = ?0.425, P = 0.055), and VC (r = ?0.572, P = 0.007)). Genotypes had no impact on TGF?1 in IS, serum, and LF. In IS TGF?1 correlated with IL-8 (r = 0.593, P < 0.007) and TNF-? (r = 0.536, P < 0.018) in patients colonized by bacteria with flagellin. Conclusion. TGF?1 in serum not in IS correlates with LF. In patients colonized by bacteria with flagellin, TGF?1 correlates with IL-8 and TNF-? in IS.

Eickmeier, O.; Boom, L. v. d.; Schreiner, F.; Lentze, M. J.; NGampolo, D.; Schubert, R.; Zielen, S.; Schmitt-Grohe, S.



Activated extracellular signal-regulated kinases: association with epidermal growth factor receptor/transforming growth factor alpha expression in head and neck squamous carcinoma and inhibition by anti-epidermal growth factor receptor treatments.  


The expression of the activated mitogen-activated kinases/extracellular signal-regulated kinases (ERKs) ERK1 and ERK2 was characterized in 101 humanhead and neck squamous carcinoma specimens. Activated ERK1/2were detected at different levels in the majority of these tumors, as assayed by immunostaining with an antibody specific for the dually phosphorylated and activated ERK1 and ERK2. ERK1/2 activation levels were higher in tumors with advanced regional lymph node metastasis (P = 0.048) and in relapsed tumors (P = 0.021). The expression of epidermal growth factor (EGF) receptor (P = 0.037), transforming growth factor alpha (TGF-alpha; P < 0.001), and HER2 (P = 0.066; positive trend) correlated with activation of ERK1/2. In a multivariate analysis, both TGF-alpha (P < 0.0001) and HER2 (P = 0.045) were independently correlated with ERK1/2 activation. In turn, activation of ERK1/2 was associated with a higher Ki-67 proliferative index (P = 0.002). In EGF receptor-dependent model cells (A431 and DiFi), a specific EGF receptor tyrosine kinase inhibitor ("Iressa"; ZD1839) and a chimeric anti-EGF receptor antibody ("Cetuximab"; C225) inhibited ERK 1/2 activation at concentrations that inhibited autocrine cell proliferation. In patients on treatment with C225, the activation of ERK1/2 in skin, an EGF receptor-dependent tissue, was lower compared with control skin. Parallel changes were seen in keratinocyte Ki67 proliferation indexes in skin from C225-treated patients. Taken together, these studies provide support for a role of activation of ERK1/2 in head and neck squamous carcinoma and a correlation with EGF receptor/TGF-alpha expression. The inhibition of ERK1/2 activation in vitro and in vivo by compounds targeting the EGF receptor points to the interest of ERK1/2 as potential surrogate markers of EGF-receptor signaling in clinical therapeutic studies. PMID:11522647

Albanell, J; Codony-Servat, J; Rojo, F; Del Campo, J M; Sauleda, S; Anido, J; Raspall, G; Giralt, J; Roselló, J; Nicholson, R I; Mendelsohn, J; Baselga, J



Modulation of in vitro monocyte cytokine responses to Leishmania donovani. Interferon-gamma prevents parasite-induced inhibition of interleukin 1 production and primes monocytes to respond to Leishmania by producing both tumor necrosis factor-alpha and interleukin 1.  

PubMed Central

Cytokines produced by mononuclear cells are important regulatory and effector molecules and evidence has been presented to support a role at least for tumor necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) in host defense against Leishmania. In the present study, we examined the production of TNF-alpha and interleukin 1 (IL-1) by resting and IFN-gamma-primed peripheral blood monocytes infected in vitro with Leishmania donovani. Monocytes produced neither IL-1 nor TNF-alpha during challenge with Leishmania. Cells preinfected with Leishmania synthesized normal amounts of TNF-alpha, but had diminished production of IL-1 in response to stimulation with either S. aureus or lipopolysaccharide (LPS). The induction by S. aureus or LPS of IL-1 beta mRNA accumulation in infected cells was normal despite diminished intracellular or supernatant IL-1 protein and bioactivity. Thus, inhibition of IL-1 production by Leishmania most probably reflected diminished translation of IL-1 beta mRNA. Pretreatment of cells with IFN-gamma abrogated infection-induced inhibition of IL-1 production and primed cells for the production of both IL-1 and TNF-alpha upon subsequent exposure to Leishmania. These results indicate that L. donovani has evolved the capacity to infect mononuclear phagocytes, without stimulating the production of two potentially host-protective monokines. The ability of IFN-gamma to prime monocytes to produce TNF-alpha and IL-1 in response to infection with Leishmania and to prevent inhibition of IL-1 production may have implications for immunotherapy with this lymphokine. Images

Reiner, N E; Ng, W; Wilson, C B; McMaster, W R; Burchett, S K



Uniocular Anterior Chamber Inoculation of a Tumor Necrosis Factor Alpha-Expressing Recombinant of Herpes Simplex Virus Type 1 Results in More Rapid Destruction and Increased Viral Replication in the Retina of the Uninoculated Eye  

Microsoft Academic Search

Tumor necrosis factor alpha (TNF-) has been shown to have a protective role in the eyes and brains of herpes simplex virus type 1 (HSV-1)-infected mice. To determine whether overexpression of TNF- affected the course of virus infection following uniocular anterior chamber inoculation, a recombinant of HSV-1 that produces TNF- constitutively (KOSTNF) was constructed. BALB\\/c mice were injected with the

Mark A. Fields; Mei Zheng; Pam Wall; Scott Oberg; Sally S. Atherton



Tumor necrosis factor-alpha/nuclear transcription factor-kappaB signaling in periprosthetic osteolysis.  


Due to irreversible joint destruction caused by the various arthritides, more than 400,000 total joint arthroplasties are performed each year in the United States. As many as 20% of these require revision surgery because of aseptic loosening. The current paradigm to explain aseptic loosening is that wear debris generated from the prosthesis stimulates the release of proinflammatory cytokines (i.e., tumor necrosis factor-alpha and interleukins 1 and 6) following phagocytosis by resident macrophages. These cytokines, in turn, initiate an inflammatory response, with the development of an erosive pannus that stimulates bone resorption by osteoclasts. In support of this model, we have previously shown that human monocytes produce large quantities of tumor necrosis factor-alpha in response to titanium particles in vitro. In the current study, we characterized the role of tumor necrosis factor-alpha/nuclear transcription factor-kappaB signaling in the proinflammatory response to titanium particles in vitro and in vivo. Using the mouse macrophage cell line J774, we showed that these cells produce an amount of tumor necrosis factor-alpha in response to titanium particles similar to that produced by human peripheral blood monocytes. The production of tumor necrosis factor-alpha was preceded by a drop in cellular levels of inhibitory factor-kappaBalpha protein and translocation of p50/p65 nuclear transcription factor-KB to the nucleus 30 minutes after stimulation. Levels of tumor necrosis factor-alpha and inhibitory factor-kappaBalpha mRNA increased 30 minutes after stimulation, consistent with the activation of nuclear transcription factor-kappaB. Interleukin-6 mRNA was first seen 4 hours after the addition of the titanium particles, indicating that the production of this cytokine is secondary to the immediate nuclear transcription factor-kappaB response. To test the relevance of tumor necrosis factor-alpha/nuclear transcription factor-kappaB signaling in response to titanium particles in vivo, we adopted an animal model in which the particles were surgically implanted on the calvaria of mice. The animals displayed a dramatic histological response to the debris, with the formation of fibrous tissue and extensive bone resorption after only 1 week. With use of immunohistochemistry and tartrate-resistant acid phosphatase staining, tumor necrosis factor-alpha and osteoclasts were readily detected at the site of inflammation and bone resorption in the calvaria of the treated mice. By testing mice that genetically over-produce tumor necrosis factor-alpha (hTNFalpha-Tg), those defective in tumor necrosis factor-alpha signaling (TNF-RI-/-), and those that are nuclear transcription factor-kappaB1-deficient (NFkappaB1-/-), we evaluated the importance of tumor necrosis factor-alpha/nuclear transcription factor-kappaB signaling in the biological processes responsible for aseptic loosening. The hTNFalpha-Tg mice had a grossly exaggerated response, the TNF-RI(-/-) mice showed little evidence of inflammation or bone resorption, and the nuclear transcription factor-kappaB1(-/-) mice had an inflammatory response without bone resorption. On the basis of these results, we propose a model for periprosthetic osteolysis in which wear debris particles are phagocytosed by macrophages, resulting in the activation of nuclear transcription factor-kappaB and the production of tumor necrosis factor-alpha. Tumor necrosis factor-alpha directly induces fibroblast proliferation and tissue fibrosis and recruits or activates, or both, osteoclasts to resorb adjacent bone. PMID:10937636

Schwarz, E M; Lu, A P; Goater, J J; Benz, E B; Kollias, G; Rosier, R N; Puzas, J E; O'Keefe, R J



ETS-1 oncogenic activity mediated by transforming growth factor alpha.  


Inappropriate expression of Ets-1 is observed in a variety of human cancers, and its forced expression in cultured cells results in transformation, autonomous proliferation, and tumor formation. The basis by which Ets-1 confers autonomous growth, one of the primary hallmarks of cancer cells and a critical component of persistent proliferation, has yet to be fully explained. Using a variety of cancer cell lines, we show that inhibition of Ets-1 blocks tumor formation and cell proliferation in vivo and autonomous growth in culture. A screen of multiple diffusible growth factors revealed that inhibition of Ets-1 results in the specific downregulation of transforming growth factor alpha (TGFalpha), the proximal promoter region of which contains multiple ETS family DNA binding sites that can be directly bound and regulated by Ets-1. Notably, rescuing TGFalpha expression in Ets-1-silenced cells was sufficient to restore tumor cell proliferation in vivo and autonomous growth in culture. These results reveal a previously unrecognized mechanism by which Ets-1 oncogenic activity can be explained in human cancer through its ability to regulate the important cellular mitogen TGFalpha. PMID:20068146

Holterman, Chet E; Franovic, Aleksandra; Payette, Josianne; Lee, Stephen



Tumour necrosis factor-alpha inhibitor-induced hepatic injury in patients with rheumatoid arthritis: two case reports and an analysis of the laboratory data from the Slovenian national biologicals registry.  


Tumour necrosis factor-alpha (TNF-?) inhibitors are widely used in the management of patients with rheumatoid arthritis (RA) and spondylarthritides. However, TNF-? inhibition may lead to adverse events, including liver injury. The RA patients are frequently treated with several potentially hepatotoxic drugs concomitantly; hence, a causative link between TNF-? inhibitors and liver injury is usually difficult to establish. We report two cases of RA patients who developed histologically manifest liver injury shortly after the introduction of treatment with two different TNF-? inhibitors. Furthermore, we present the analysis of the laboratory data from the registry (the Slovenian national registry of rheumatologic patients treated with biologicals) and provide evidence that elevated levels of serum aminotransferase can be observed in patients treated with TNF-? inhibitors. Additionally, our analysis suggests no significant differences between the impact of adalimumab and etanercept on aminotransferase levels. Although the use of TNF-alpha inhibitors is safe and efficient, we suggest that continuous careful monitoring of aminotransferase levels in patients treated with these agents is probably warranted. PMID:22955878

Perdan-Pirkmajer, Katja; Ho?evar, Alojzija; Rotar, Žiga; Žibert, Janez; Marolt, Vera Ferlan; Gu?ev, Filip; Tomši?, Matija



Tumor Necrosis Factor Alpha: A Link between Neuroinflammation and Excitotoxicity  

PubMed Central

Tumor necrosis factor alpha (TNF-?) is a proinflammatory cytokine that exerts both homeostatic and pathophysiological roles in the central nervous system. In pathological conditions, microglia release large amounts of TNF-?; this de novo production of TNF-? is an important component of the so-called neuroinflammatory response that is associated with several neurological disorders. In addition, TNF-? can potentiate glutamate-mediated cytotoxicity by two complementary mechanisms: indirectly, by inhibiting glutamate transport on astrocytes, and directly, by rapidly triggering the surface expression of Ca+2 permeable-AMPA receptors and NMDA receptors, while decreasing inhibitory GABAA receptors on neurons. Thus, the net effect of TNF-? is to alter the balance of excitation and inhibition resulting in a higher synaptic excitatory/inhibitory ratio. This review summarizes the current knowledge of the cellular and molecular mechanisms by which TNF-? links the neuroinflammatory and excitotoxic processes that occur in several neurodegenerative diseases, but with a special emphasis on amyotrophic lateral sclerosis (ALS). As microglial activation and upregulation of TNF-? expression is a common feature of several CNS diseases, as well as chronic opioid exposure and neuropathic pain, modulating TNF-? signaling may represent a valuable target for intervention.

Olmos, Gabriel; Llado, Jeronia



Fusobacterium nucleatum Infection of Colonic Cells Stimulates MUC2 Mucin and Tumor Necrosis Factor Alpha?  

PubMed Central

The etiology of inflammatory bowel disease is not completely known, but it is influenced by the presence of normal gut microflora as well as yet-unrecognized pathogens. The anaerobic, Gram-negative bacterial species Fusobacterium nucleatum is a common resident of the human mouth and gut and varies in its pathogenic potential. In this study, we demonstrate that highly invasive F. nucleatum isolates derived from the inflamed guts of Crohn's disease patients evoked significantly greater MUC2 and tumor necrosis factor alpha (TNF-?) gene expression than minimally invasive strains isolated from the noninflamed gut in human colonic epithelial cells and in a rat ligated colonic loop model of infection. Only live F. nucleatum induced mucin secretion and TNF-? expression in direct contact with and/or during invasion of colonic cells. In rat colons, mucin secretion was augmented in response to a highly invasive F. nucleatum isolate but was unaffected by treatment with a minimally invasive strain. Taken together, these studies reveal that F. nucleatum may represent a challenging pathogen in the etiology of gut inflammatory diseases and highlight the importance of different pathotypes of candidate bacterial species in disease pathogenesis.

Dharmani, Poonam; Strauss, Jaclyn; Ambrose, Christian; Allen-Vercoe, Emma; Chadee, Kris



Development of orally bioavailable bicyclic pyrazolones as inhibitors of tumor necrosis factor-alpha production.  


2-Aryl-3-pyrimidinyl based tumor necrosis factor-alpha (TNF-alpha) inhibitors, which contain a novel bicyclic pyrazolone core, are described. Many showed low-nanomolar activity against lipopolysaccharide-induced TNF-alpha production in monocytic cells. Secondary screening data are presented for the pyrimidinyl bicyclic pyrazolones. Several of these analogues showed good oral bioavailability in rat and efficacy in the rat iodoacetate in vivo model. PMID:15139749

Clark, Michael P; Laughlin, Steven K; Laufersweiler, Matthew J; Bookland, Roger G; Brugel, Todd A; Golebiowski, Adam; Sabat, Mark P; Townes, Jennifer A; VanRens, John C; Djung, Jane F; Natchus, Michael G; De, Biswanath; Hsieh, Lily C; Xu, Susan C; Walter, Rick L; Mekel, Marlene J; Heitmeyer, Sandra A; Brown, Kimberly K; Juergens, Karen; Taiwo, Yetunde O; Janusz, Michael J



Tumor necrosis factor alpha is an autocrine growth factor for normal human B cells.  

PubMed Central

Transcription of the human tumor necrosis factor alpha (TNF-alpha) gene is one of the earliest events that occurs after stimulation of B or T cells via their antigen receptors. Antibody directed at surface immunoglobulin (anti-Ig) on B cells has previously been shown to induce a rapid burst of TNF-alpha gene transcription, which can be blocked by the immunosuppressants cyclosporin A (CsA) and FK506. Here, TNF-alpha gene transcription is shown also to be highly and rapidly induced in human B cells after stimulation via the CD40 and interleukin 4 pathways, which similarly is inhibited by CsA and a panel of CsA or FK506 analogues that block calcineurin phosphatase activity. Endogenous TNF-alpha produced after stimulation was involved in B-cell proliferation since anti-TNF-alpha monoclonal antibody inhibited both anti-Ig- and anti-CD40-induced B-cell proliferative responses. Moreover, addition of TNF-alpha during stimulation resulted in augmentation of B-cell proliferation, which was also inhibited by anti-TNF-alpha monoclonal antibody. Although lymphotoxin alpha (LT-alpha) mRNA is induced by both pathways, it is not blocked by CsA, whereas LT-beta mRNA is constitutively expressed in B cells. Thus, TNF-alpha is a necessary autocrine growth factor for human B cells stimulated via two independent CsA-sensitive pathways and plays a role similar to that of interleukin 2 in T-cell proliferation. The autocrine nature of TNF-alpha in activated B cells implies a potential role for this cytokine in infection-related polyclonal B-cell expansion and in B-cell malignancies. Images

Boussiotis, V A; Nadler, L M; Strominger, J L; Goldfeld, A E



Tumor necrosis factor. alpha. and interleukin 1 stimulate the human immunodeficiency virus enhancer by activation of the nuclear factor. kappa. B  

SciTech Connect

Binding of peptide hormones to surface membrane receptors leads to the transcription of specific genes within relevant target cells. How these signals are transduced to alter gene expression is largely unknown, but this mechanism probably involves a sequence of enzymatic steps that activate factors in the nucleus that modulate transcription. The authors demonstrate that two different peptide hormones, or cytokines, stimulate the human immunodeficiency virus enhancer, and this effect is mediated by nuclear factor (NF) {kappa}B. These cytokines, tumor necrosis factor {alpha} and interleukin 1, act on multiple cell types and represent the only naturally occurring activators of this transcription factor among eight cytokines examined. Although NF-{kappa}B binding can be stimulated by phorbol 12-myristate 13-acetate, tumor necrosis factor {alpha} acts through an independent mechanism, inducing NF-{kappa}B binding in HT-2 cells, which did not show increased binding in response to phorbol 12-myristate 13-acetate, and causing superinduction in Jurkat T-lymphoma cells. These findings suggest that human immunodeficiency virus gene expression can be induced in T cells without activating lymphokine secretion and that the role of these cytokines in the activation of latent human immunodeficiency virus infection deserves further clinical evaluation. Finally, this link between binding at the surface membrane and stimulation of a specific transcription factor should help define intermediates for these cytokine activation pathways.

Osborn, L.; Kunkel, S.; Nabel, G.J. (Howard Hughes Medical Institute, Ann Arbor, MI (USA))



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

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

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



Tumor Necrosis Factor Alpha and Inducible Nitric Oxide Synthase-Producing Dendritic Cells Are Rapidly Recruited to the Bladder in Urinary Tract Infection but Are Dispensable for Bacterial Clearance  

Microsoft Academic Search

The role of dendritic cells (DC) in urinary tract infections (UTI) is unknown. These cells contribute directly to the innate defense against various viral and bacterial infections. Here, we studied their role in UTI using an experimental model induced by transurethral instillation of the uropathogenic Escherichia coli (UPEC) strain 536 into C57BL\\/6 mice. While few DC were found in the

Daniel Engel; Ulrich Dobrindt; A. Tittel; P. Peters; J. Maurer; I. Gutgemann; B. Kaissling; W. Kuziel; S. Jung; C. Kurts



Elevated serum levels of tumor necrosis factor alpha in normal-weight women with polycystic ovary syndrome  

Microsoft Academic Search

Since an increase in tumor necrosis factor alpha (TNF?) expression has been associated with insulin resistance, this study was undertaken to determine the status of circulating TNF? and the relationship of TNF? with insulin levels, body weight, or both in women with polycystic ovary syndrome (PCOS). Fasting serum samples were analyzed in 34 subjects with PCOS, of whom 22 were

Frank Gonzalez; Kuldip Thusu; Ehad Abdel-Rahman; Anu Prabhala; Madonna Tomani; Paresh Dandona



Transforming growth factor alpha in arterioles: cell surface processing of its precursor by elastases.  

PubMed Central

Analysis of the transforming growth factor alpha (TGF alpha) cDNA predicts that the mature TGF alpha polypeptide is cleaved from the extracellular domain of its precursor, which is an integral membrane protein. Furthermore, the cleavage sites for the release of this mitogen are compatible with the participation of an elastaselike protease. We have immunohistochemically localized TGF alpha to the vascular smooth muscle cells in the arterioles. To investigate whether polymorphonuclear (PMN) leukocytic elastase, a blood-borne protease, could process the cell surface TGF alpha, NR6 cells were transfected with the rat TGF alpha cDNA. The cDNA encoded the entire open reading frame, and its expression was under the control of the mouse metallothionein I promoter. A cloned transfectant, termed 1B2, synthesized the TGF alpha precursor in a zinc-inducible manner, and the precursor was localized to the cell surface. Western blot (immunoblot) analysis indicated that treatment of the zinc-induced 1B2 cells with either PMN leukocytic or pancreatic elastase resulted in the release of the mature TGF alpha polypeptide. The released TGF alpha was bioactive, as it was capable of both competing with epidermal growth factor for binding to its receptor and stimulating [3H]thymidine incorporation in the mitogenic assay. Formaldehyde fixation of the 1B2 cells eliminated basal release of TGF alpha but allowed normal processing by both PMN leukocytic and pancreatic elastase to occur. However, human cathepsin G, bovine pancreatic alpha 1-chymotrypsin, collagenase, trypsin, subtilisin, and plasmin failed to release any detectable fragments of the TGF alpha precursor from the fixed cells. The location of TGF alpha in the arterioles and ability of PMN leukocytic elastase to process the membrane-bound TGF alpha precursor suggests a novel role for this elastase at the wound site. Images

Mueller, S G; Paterson, A J; Kudlow, J E



Down-regulation of tumor necrosis factor alpha activity by acute ethanol treatment in human peripheral blood monocytes  

Microsoft Academic Search

As the most commonly used drug that can modulate both metabolic and immune pathways, ethanol is evaluated in this report as a regulator of tumor necrosis factor alpha (TNF alpha) production in human peripheral blood monocytes (M phi) in combination with a variety of stimuli. While acute ethanol treatment did not induce TNF alpha in M phi, it was a

Bikash K. Verma; Miklos C. Fogarasi; Gyongyi Szabo



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


Osteosarcoma is the most common primary malignancy of bone and is characterized by a high malignant and metastatic potential. Transforming growth factor alpha (TGF-?) 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-? on human osteosarcoma is largely unknown. We found that TGF-? 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-?-mediated cell migration. We also found that the phosphatidylinositol 3'-kinase (PI3K)/Akt/NF-?B pathway was activated after TGF-? treatment, and TGF-?-induced expression of ICAM-1 and cell migration was inhibited by the specific inhibitors and siRNAs of PI3K, Akt, and NF-?B cascades. In addition, knockdown of TGF-? expression markedly decreased cell metastasis in vitro and in vivo. Our results indicate that TGF-?/EGFR interaction elicits PI3K and Akt activation, which in turn activates NF-?B, resulting in the expression of ICAM-1 and contributing the migration of human osteosarcoma cells. PMID:24685520

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



Tumor necrosis factor-alpha inhibits endothelium-dependent relaxation.  


Tumor necrosis factor-alpha (TNF-alpha) stimulates nitric oxide (NO) in vascular endothelium by induction of the enzyme NO synthase II (NOS II). We examined the effects of TNF-alpha on 1) endothelium-dependent (EDR) and endothelium-independent (EIR) relaxation and 2) contraction of bovine intralobar pulmonary arteries (BPA) and veins (BPV) in vitro. Acetylcholine (ACh), bradykinin (BK), histamine, and A23187 produced EDR of BPA contracted with a 50% effective concentration of U-46619 (15 nM), because relaxation was abolished by endothelium-rubbing and attenuated by L-NG-mono-methylarginine (L-NMMA; 300 microM). TNF-alpha (0.00417, 0.0417, 0.417, and 1.25 micrograms/ml) incubated with BPA for 60 min inhibited EDR of the BPA to ACh, BK, and histamine. The effects of TNF required 30 min for onset. Recovery of EDR occurred 3-4 h after washout of TNF-alpha. Pentoxifylline (1 microM) did not affect ACh-induced EDR but selectively reversed TNF-alpha-mediated inhibition of ACh-induced EDR. TNF-alpha-mediated inhibition of EDR was not reversible by L-NMMA, an inhibitor of NOS I and NOS II, the cyclooxygenase inhibitor ibuprofen, or CV-3908 (1 microM), a platelet-activating factor antagonist. The inhibitory effect of TNF-alpha on EDR was not mediated by nonspecific sensitization of the endothelium to human protein because recombinant human granulocyte colony-stimulating factor (10, 50, and 500 x 10(3) U/ml) did not affect EDR of BPA. The effect of TNF-alpha was specific for release of NO from the endothelium of BPA because TNF-alpha did not affect 1) EDR of BPV to ACh, BK, or ATP; 2) EIR of BPA or BPV to nitroprusside; and 3) contraction of either BPA or BPV to KCl, U-46619, histamine, norepinephrine, or serotonin. Thus TNF-alpha appears to selectively inhibit receptor-mediated EDR and NO release in BPA. TNF-alpha-mediated inhibition of EDR differs from that of L-arginine-based inhibitors and may represent an endogenous physiological mechanism of regulation of NO in the endothelium. PMID:8335573

Greenberg, S; Xie, J; Wang, Y; Cai, B; Kolls, J; Nelson, S; Hyman, A; Summer, W R; Lippton, H



Tumor necrosis factor alpha (TNF-?) gene polymorphism in alopecia areata  

Microsoft Academic Search

Tumor necrosis factor alpha (TNF-a) phenotypes of two polymorphic systems were determined in 50 patients with alopecia areata, a common inflammatory disease of the skin. The distribution of TNF-a T1, T2 phenotypes differed between patients with the patchy form of disease and patients with totalis\\/universalis disease. There was no significant difference in the distribution of TNF-a G,A phenotypes between patient

Gillian M. P. Galbraith; Janardan P. Pandey



Molecular Mechanism of Suppression of Testicular Steroidogenesis by Proinflammatory Cytokine Tumor Necrosis Factor Alpha  

PubMed Central

Tumor necrosis factor alpha (TNF-?) has been demonstrated to inhibit steroidogenesis in Leydig cells at the transcriptional level of steroidogenic enzymes. However, the molecular mechanism of this observed gene repression is not well understood. We now demonstrate that nuclear factor ?B (NF-?B) activated by TNF-? inhibits the transactivation of orphan nuclear receptors, which regulate the expression of steroidogenic-enzyme genes. TNF-? treatment suppressed the luteinizing-hormone-induced or Nur77/SF-1-stimulated promoter activity of steroidogenic-enzyme genes in Leydig cells. The TNF-?-mediated gene suppression was blocked by treatment with an inhibitor of NF-?B. In addition, overexpression of the p65 (RelA) subunit of NF-?B showed the same effect as TNF-? and inhibited Nur77 transactivation, suggesting the involvement of NF-?B activation in the observed gene repression. Physical association of Nur77 with p65 was revealed by mammalian two-hybrid, GST pull-down, and coimmunoprecipitation analyses. The NF-?B inhibition of Nur77 transactivation was likely due to the competition of p65 for Nur77 binding with coactivators. Finally, chromatin immunoprecipitation assays revealed that TNF-? treatment caused the recruitment of NF-?B to the promoter of the steroidogenic-enzyme P450c17 gene, supporting the hypothesis that the TNF-?-mediated gene repression involves NF-?B inhibition of the transcriptional activity of Nur77 and other orphan nuclear receptors. These findings provide a molecular mechanism underlying the inhibition of testicular steroidogenesis by proinflammatory cytokines.

Hong, Cheol Yi; Park, Jin Hee; Ahn, Ryun Seop; Im, Suhn Young; Choi, Hueng-Sik; Soh, Jaemog; Mellon, Synthia H.; Lee, Keesook



Inflammatory cytokines induce a reduction in E-cadherin expression and morphological changes in MDCK cells.  


Epithelial-mesenchymal transition (EMT) is a fundamental phenomenon in organisms that occurs during gastrulation, wound healing, and cancer metastasis. Various cytokines induce EMT processes through complex mechanisms. Inflammatory cytokines, such as tumor necrosis factor alpha (TNF-?) and interleukin 6 (IL-6), induce EMT in human cell lines. However, whether inflammatory cytokines can affect EMT processes in canine cell lines remains unclear. In this study, we investigated the role of transforming growth factor beta (TGF-?), TNF-?, and IL-6 in Madin-Darby canine kidney (MDCK) cells. We found that the localization of E-cadherin, a cell adhesion molecule, was shifted and that its expression was decreased. We also observed morphological changes in MDCK cells under persistent stimulation of inflammatory cytokines. Morphological changes in cells may occur during late stages of EMT processes; inflammatory cytokines may be important in these changes. PMID:24565002

Saito, Tomochika; Yoshida, Kota; Matsumoto, Kaori; Saeki, Kohei; Tanaka, Yuiko; Ong, Siew-Mei; Sasaki, Nobuo; Nishimura, Ryohei; Nakagawa, Takayuki



Tumor necrosis factor-alpha (TNF-?) enhances functional thermal and chemical responses of TRP cation channels in human synoviocytes  

PubMed Central

Background We have shown functional expression of several TRP channels on human synovial cells, proposing significance in known calcium dependent proliferative and secretory responses in joint inflammation. The present study further characterizes synoviocyte TRP expression and activation responses to thermal and osmotic stimuli after pre-treatment with proinflammatory mediator tumor necrosis factor alpha (TNF-?, EC50 1.3221 × 10-10g/L). Results Fluorescent imaging of Fura-2 loaded human SW982 synoviocytes reveals immediate and delayed cytosolic calcium oscillations elicited by (1) TRPV1 agonists capsaicin and resiniferatoxin (20 – 40% of cells), (2) moderate and noxious temperature change, and (3) osmotic stress TRPV4 activation (11.5% of cells). TNF-alpha pre-treatment (1 ng/ml, 8 – 16 hr) significantly increases (doubles) capsaicin responsive cell numbers and [Ca2+]i spike frequency, as well as enhances average amplitude of temperature induced [Ca2+]i responses. With TNF-alpha pre-treatment for 8, 12, and 16 hr, activation with 36 or 45 degree bath solution induces bimodal [Ca2+]i increase (temperature controlled chamber). Initial temperature induced rapid transient spikes and subsequent slower rise reflect TRPV1 and TRPV4 channel activation, respectively. Only after prolonged TNF-alpha exposure (12 and 16 hr) is recruitment of synoviocytes observed with sensitized TRPV4 responses to hypoosmolarity (3–4 fold increase). TNF-alpha increases TRPV1 (8 hr peak) and TRPV4 (12 hr peak) immunostaining, mRNA and protein expression, with a TRPV1 shift to membrane fractions. Conclusion TNF-? provides differentially enhanced synoviocyte TRPV1 and TRPV4 expression and [Ca2+]i response dependent on the TRP stimulus and time after exposure. Augmented relevance of TRPV1 and TRPV4 as inflammatory conditions persist would provide calcium mediated cell signaling required for pathophysiological responses of synoviocytes in inflammatory pain states.



Tumor necrosis factor-alpha induced enhancement of cryosurgery  

NASA Astrophysics Data System (ADS)

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-? 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-? 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-?. Both cryosurgery alone or TNF-? treatment alone caused only a minimal damage to the tumor tissue at the conditions used in the study. The combination of TNF-? 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-?.

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



Inducible expression systems for mycobacteria.  


A wide variety of inducible expression systems have been designed for Gram-negative bacteria, but adapting these systems to phylogenetically distinct species, such as mycobacteria, has proved notoriously difficult. Mycobacteria belong to a class of high G+C Gram-positive bacteria known as actinomycetes. Although comparatively few genetic tools are available for these organisms, those that do exist are more likely to be adaptable for use in mycobacteria. A compelling example of this rationale is the recent description of a tetracycline-responsive element from corynebacteria that functions in mycobacteria. Here we describe the use of two additional mycobacterial expression systems that are derived from endogenous regulons of Streptomyces and Rhodococcus spp. Each of the currently available systems has specific advantages and limitations, and the conditions that recommend the use of each will be discussed. PMID:20560072

Sassetti, Christopher M



Reduction of tumor necrosis factor-alpha (TNF-?) related nuclear factor-kappaB (NF-?B) translocation but not inhibitor kappa-B (I?-B)-degradation by Rho protein inhibition in human endothelial cells  

Microsoft Academic Search

Degradation of inhibitor kappa-B (I?-B) followed by translocation of nuclear factor-kappaB (NF-?B) into the nucleus and activation of gene expression is essential in tumor necrosis factor-alpha (TNF-?)-signaling. In order to analyze the role of Rho proteins in TNF-?-induced NF-?B-activation in human umbilical cord vein endothelial cells (HUVEC) we used Clostridium difficile toxin B-10463 (TcdB-10463) which inactivates RhoA\\/Rac1\\/Cdc42 by glucosylation and

Stefan Hippenstiel; Bernd Schmeck; Joachim Seybold; Matthias Krüll; Christoph v Eichel-Streiber; Norbert Suttorp



Bacterium-Generated Nitric Oxide Hijacks Host Tumor Necrosis Factor Alpha Signaling and Modulates the Host Cell Cycle In Vitro  

PubMed Central

In mammalian cells, nitric oxide (NO·) is an important signal molecule with concentration-dependent and often controversial functions of promoting cell survival and inducing cell death. An inducible nitric oxide synthase (iNOS) in various mammalian cells produces higher levels of NO· from l-arginine upon infections to eliminate pathogens. In this study, we reveal novel pathogenic roles of NO· generated by bacteria in bacterium-host cell cocultures using Moraxella catarrhalis, a respiratory tract disease-causing bacterium, as a biological producer of NO·. We recently demonstrated that M. catarrhalis cells that express the nitrite reductase (AniA protein) can produce NO· by reducing nitrite. Our study suggests that, in the presence of pathophysiological levels of nitrite, this opportunistic pathogen hijacks host cell signaling and modulates host gene expression through its ability to produce NO· from nitrite. Bacterium-generated NO· significantly increases the secretion of tumor necrosis factor alpha (TNF-?) and modulates the expression of apoptotic proteins, therefore triggering host cell programmed death partially through TNF-? signaling. Furthermore, our study reveals that bacterium-generated NO· stalls host cell division and directly results in the death of dividing cells by reducing the levels of an essential regulator of cell division. This study provides unique insight into why NO· may exert more severe cytotoxic effects on fast growing cells, providing an important molecular basis for NO·-mediated pathogenesis in infections and possible therapeutic applications of NO·-releasing molecules in tumorigenesis. This study strongly suggests that bacterium-generated NO· can play important pathogenic roles during infections.

Mocca, Brian



Production of Transforming Growth Factor alpha in Human Pancreatic Cancer Cells: Evidence for a Superagonist Autocrine Cycle  

Microsoft Academic Search

Previous work showed that cultured human pancreatic cancer cells overexpress the epidermal growth factor (EGF) receptor. In the present study, we sought to determine whether some of these cell lines produce transforming growth factor alpha (TGF-alpha ). Utilizing a radiolabeled TGF-alpha cDNA in hybridization experiments, we determined that ASPC-1, T3M4, PANC-1, COLO-357, and MIA PaCa-2 cell lines expressed TGF-alpha mRNA.

Jeffrey J. Smith; Rik Derynck; Murray Korc



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

SciTech Connect

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.

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 (PG)



Induced interleukin-8 expression in gliomas by tumor-associated macrophages.  


The tumor microenvironment affects tumor initiation, progression, and metastasis. However, it is still not clear how stromal cells interact with the tumor cells. By using a cytokine array immunoblot assay, we showed that interleukin (IL)-8, IL-6, and RANTES (regulated upon activation normal T-cell expressed and secreted) proteins were up-regulated in GBM8401 glioma cells after coculture with human THP-1-derived macrophages. IL-8 is a chemokine with leukocyte chemotactic, tumorigenic, and proangiogenic properties. To evaluate the correlation of IL-8 expression with tumor-associated macrophages and angiogenesis, 43 glioma specimens were studied. The results showed that the IL-8 mRNA expression and microvessel count in glioma surgical specimens correlated positively with the density of tumor-associated macrophages. We further showed that IL-8 mRNA expression in GBM8401 cells increased dramatically, by 2(8)-2(10)-fold, after being cocultured with macrophages. This increase could also be induced by macrophage-conditioned medium, tumor necrosis factor-alpha, IL-1alpha, and IL-1beta, and could be suppressed by anti-inflammatory agents including pyrrolidine dithiocarbamate, pentoxifylline, or dexamethasone. These findings imply that macrophage infiltration may be the common feature shared by cancer and inflammation, and macrophages could play a role in promoting glioma growth and angiogenesis by inducing IL-8 expression in glioma cells via inflammatory stimuli or the nuclear factor kappa B pathway. PMID:19156359

Hong, Tse-Ming; Teng, Lee-Jene; Shun, Chia-Tung; Peng, Mei-Chen; Tsai, Jui-Chang



Flavonoids inhibit cytokine-induced endothelial cell adhesion protein gene expression.  

PubMed Central

Treatment of human endothelial cells with cytokines such as interleukin-1, tumor necrosis factor-alpha (TNF-alpha) or interferon-gamma induces the expression of specific leukocyte adhesion molecules on the endothelial cell surface. Interfering with either leukocyte adhesion or adhesion protein upregulation is an important therapeutic target as evidenced by the potent anti-inflammatory actions of neutralizing antibodies to these ligands in various animal models and in patients. In the present study we report that cotreatment of human endothelial cells with certain hydroxyflavones and flavanols blocks cytokine-induced ICAM-1, VCAM-1, and E-selectin expression on human endothelial cells. One of the most potent flavones, apigenin, exhibited a dose- and time-dependent, reversible effect on adhesion protein expression as well as inhibiting adhesion protein upregulation at the transcriptional level. Apigenin also inhibited IL-1 alpha-induced prostaglandin synthesis and TNF-alpha-induced IL-6 and IL-8 production, suggesting that the hydroxyflavones may act as general inhibitors of cytokine-induced gene expression. Although apigenin did not inhibit TNF-alpha-induced nuclear translocation of NF-kappa B(p50(NFKB1)/p65(RelA)) we found this flavonoid did inhibit TNF-alpha induced beta-galactosidase activity in SW480 cells stably transfected with a beta-galactosidase reporter construct driven by four NF-kappa B elements, suggesting an action on NF-kappa B transcriptional activation. Adhesion of leukocytes to cytokine-treated endothelial cells was blocked in endothelial cells cotreated with apigenin. Finally, apigenin demonstrated potent anti-inflammatory activity in carrageenan induced rat paw edema and delayed type hypersensitivity in the mouse. We conclude that flavonoids offer important therapeutic potential for the treatment of a variety of inflammatory diseases involving an increase in leukocyte adhesion and trafficking. Images Figure 7 Figure 8 Figure 11

Gerritsen, M. E.; Carley, W. W.; Ranges, G. E.; Shen, C. P.; Phan, S. A.; Ligon, G. F.; Perry, C. A.



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

PubMed Central

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 due to exposure to cytokines and collagen. Thus, it is likely that MMP-13 expression is instrumental in the subepithelial collagenolysis and local invasion of the activated mucosal epithelium into the connective tissue. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6

Uitto, V. J.; Airola, K.; Vaalamo, M.; Johansson, N.; Putnins, E. E.; Firth, J. D.; Salonen, J.; Lopez-Otin, C.; Saarialho-Kere, U.; Kahari, V. M.



Protective role of ascorbic acid isolated from Cissus quadrangularis on NSAID induced toxicity through immunomodulating response and growth factors expression.  


The present study investigate the effect of ascorbic acid, the major bioactive component isolated from Cissus quadrangularis extract (CAA) on inflammatory cytokines and growth factors in non-steroidal anti-inflammatory drug (NSAID) induced gastric ulcer. Analysis of serum cytokine profile using enzymelinked immunosorbent assay (ELISA) showed a drastic increase in interleukin (IL)-1beta, IL-6, tumour necrosis factor-alpha (TNF)-alpha, interferon-gamma (IFN-gamma) and decrease in IL-10, Il-4 and prostaglandin E2 (PGE2) levels in NSAID (aspirin) treated rats. The reduction of growth factors such as transforming growth factor-alpha (TGF)-alpha and vascular endothelial cell growth factor (VEGF) by aspirin was determined by immunohistochemistry method. Administration of CAA produced significant protection against aspirin induced gastric toxicity by showing significant increase in PGE2, TGF-alpha, VEGF expression and accompanied by a significant inhibition of nitric oxide and regulating the levels of cytokines in rats. These findings suggest that CAA prevents gastric ulcer formation due to its immunomodulatory effect, antioxidant activity along with the ability to modulate PG synthesis and up-regulation of the growth factors. PMID:18773975

Jainu, Mallika; Mohan, Kunju Vijai



Thalidomide selectively inhibits tumor necrosis factor alpha production by stimulated human monocytes  

PubMed Central

Thalidomide selectively inhibits the production of human monocyte tumor necrosis factor alpha (TNF-alpha) when these cells are triggered with lipopolysaccharide and other agonists in culture. 40% inhibition occurs at the clinically achievable dose of the drug of 1 micrograms/ml. In contrast, the amount of total protein and individual proteins labeled with [35S]methionine and expressed on SDS-PAGE are not influenced. The amounts of interleukin 1 beta (IL-1 beta), IL-6, and granulocyte/macrophage colony-stimulating factor produced by monocytes remain unaltered. The selectivity of this drug may be useful in determining the role of TNF-alpha in vivo and modulating its toxic effects in a clinical setting.



Increased hypoxia-inducible factor 1? expression in lung cells of horses with recurrent airway obstruction  

PubMed Central

Background Recurrent airway obstruction (RAO, also known as equine heaves) is an inflammatory condition caused by exposure of susceptible horses to organic dusts in hay. The immunological processes responsible for the development and the persistence of airway inflammation are still largely unknown. Hypoxia-inducible factor (Hif) is mainly known as a major regulator of energy homeostasis and cellular adaptation to hypoxia. More recently however, Hif also emerged as an essential regulator of innate immune responses. Here, we aimed at investigating the potential involvement of Hif1-? in myeloid cells in horse with recurrent airway obstruction. Results In vitro, we observed that Hif is expressed in equine myeloid cells after hay dust stimulation and regulates genes such as tumor necrosis factor alpha (TNF-?), interleukin-8 (IL-8) and vascular endothelial growth factor A (VEGF-A). We further showed in vivo that airway challenge with hay dust upregulated Hif1-? mRNA expression in myeloid cells from the bronchoalveolar lavage fluid (BALF) of healthy and RAO-affected horses, with a more pronounced effect in cells from RAO-affected horses. Finally, Hif1-? mRNA expression in BALF cells from challenged horses correlated positively with lung dysfunction. Conclusion Taken together, our results suggest an important role for Hif1-? in myeloid cells during hay dust-induced inflammation in horses with RAO. We therefore propose that future research aiming at functional inactivation of Hif1 in lung myeloid cells could open new therapeutic perspectives for RAO.



Platelet-activating factor induces goblet cell hyperplasia and mucin gene expression in airways.  


In patients dying from asthma, extensive mucous plugging occurs in the airways, associated with goblet cell hyperplasia. In this study, we examined the hypothesis that platelet-activating factor (PAF) induces goblet cell hyperplasia and mucin gene expression. After instilling PAF into the airways of guinea pigs and rats, we stained airway goblet cells with Alcian blue/periodic acid-Schiff and determined the number of goblet cells and percentage of stained area within the epithelium. In guinea pigs, one instillation of PAF (10(-)5 M, 100 microl) increased the goblet cell-stained area time-dependently, beginning at 24 h, maximum at 72 h. PAF also caused tracheal recruitment of eosinophils by 24 h, maximum at 48 h. In rats, which have few goblet cells in airways, PAF (3 instillations, 10(-)5 M, 200 microl) caused striking goblet cell hyperplasia, greatest in peripheral airways. Tumor necrosis factor alpha (TNFalpha) alone had no significant effect on goblet cells, but together with PAF, it caused exaggerated goblet cell hyperplasia. In rat tracheas studied by in situ hybridization, PAF induced mucin MUC5 gene expression in epithelial cells that stained for mucosubstances. In summary, PAF induces goblet cell hyperplasia and TNFalpha potentiates this effect. PMID:9620929

Lou, Y P; Takeyama, K; Grattan, K M; Lausier, J A; Ueki, I F; Agustí, C; Nadel, J A




PubMed Central

Background: Alopecia areata (AA) is a common form of localized, nonscarring hair loss. It is characterized by the loss of hair in patches, total loss of scalp hair (alopecia totalis, AT), or total loss of body hair (alopecia universalis, AU). The cause of AA is unknown, although most evidence supports the hypothesis that AA is a T-cell-mediated autoimmune disease of the hair follicle and that cytokines play an important role. Aims: The aim of the study was to compare the serum levels of tumor necrosis factor-alpha (TNF-?) in patients with AA and the healthy subjects and also to investigate the difference between the localized form of the disease with the extensive forms like AT and AU. Materials and Methods: Sixty patients with AA and 20 healthy controls were enrolled in the study. Forty-six patients had localized AA (LAA), and 14 patients had AT, AU, or AT/AU. The serum levels of TNF-? were measured using enzyme-linked immunoassay techniques. Results: Serum levels of TNF-? were significantly higher in AA patients than in controls (10.31 ± 1.20 pg ml vs 9.59 ± 0.75 pg/ml, respectively). There was no significant difference in serum levels of TNF-? between patients with LAA and those with extensive forms of the disease. Conclusion: Our findings support the evidence that elevation of serum TNF-? is associated with AA. The exact role of serum TNF-? in AA should be additionally investigated in future studies.

Kasumagic-Halilovic, Emina; Prohic, Asja; Cavaljuga, Semra



Serum tumor necrosis factor-alpha profile in trauma patients.  


Tumor necrosis factor-alpha (TNF-alpha) has been implicated in several late consequences of trauma such as sepsis, multiple organ failure, and ischemia-reperfusion injury. However, no data are available to indicate whether TNF-alpha is involved in the initial pathophysiologic response to trauma. To address this issue, serum TNF-alpha was determined (by ELISA) longitudinally (first blood sample on admission) in 100 randomly selected trauma patients admitted to the emergency department and trauma division at Jefferson Medical Center, Philadelphia. The TNF-alpha levels were detectable at one or more time points in 35 patients. Mean values tended to be elevated (50.3 +/- 11.5 pg/mL) during the first 5 days, but this trend did not differ statistically from that in healthy controls (n = 12) and did not correlate with the severity of injury (Injury Severity Score and Glasgow Coma Scale score). The TNF-alpha response was not dependent on the mechanism and site of injury, the presence of shock (systolic blood pressure < 90 mm Hg), and the need for emergent surgery. Also, serum TNF-alpha levels were not significantly elevated in patients who subsequently developed multiple organ failure (n = 4), septic shock (n = 5), or both (n = 3). Taken together, these data do not support a role for circulating TNF-alpha in the initial acute inflammatory response to trauma. PMID:8230332

Rabinovici, R; John, R; Esser, K M; Vernick, J; Feuerstein, G



Purified Shiga-like toxins induce expression of proinflammatory cytokines from murine peritoneal macrophages.  

PubMed Central

Infections with Shiga toxin-producing Shigella dysenteriae type 1 and Shiga-like toxin (SLT)-producing Escherichia coli cause outbreaks of bloody diarrhea in which patients are at risk for developing life-threatening complications involving the renal and central nervous systems. Histopathology studies and in vitro experiments suggested that the toxins damage toxin receptor-expressing endothelial cells (EC) lining glomerular and central nervous system capillaries. In the presence of inducible host factors (cytokines), EC sensitivity to SLT toxicity was increased approximately 1 million-fold. We hypothesized that to manifest the vascular lesions characteristic of infection with toxin-producing bacteria, two signals were needed: systemic toxins and elevated proinflammatory cytokines (tumor necrosis factor alpha [TNF-alpha], interleukin 1 [IL-1], and IL-6). Human EC do not secrete these cytokines when stimulated with SLTs in vitro, suggesting that additional cells may be involved in pathogenesis. Therefore, we carried out comparative analyses of the capacity of purified (endotoxin-free) SLTs and lipopolysaccharides (LPS) to induce cytokine mRNA and proteins from murine macrophages. The cells were essentially refractory to SLT cytotoxicity, expressing low to undetectable levels of toxin receptor. SLTs and LPS induced TNF activity and IL-6 expression from macrophages, although dose response and kinetics of cytokine induction differed. LPS was a more effective inducing agent than SLTs. SLT-I-induced TNF activity and IL-6 expression were delayed compared with induction mediated by LPS. IL-1 alpha production required approximately 24 h of exposure to SLTs or LPS. Macrophages from LPS-hyporesponsive C3H/HeJ mice produced low levels of TNF activity when treated with SLT-I, suggesting that LPS and SLTs may utilize separate signaling pathways for cytokine induction. Images

Tesh, V L; Ramegowda, B; Samuel, J E



Transgenic expression of the human amphiregulin gene induces a psoriasis-like phenotype.  

PubMed Central

Amphiregulin (AR) is a heparin-binding, heparin-inhibited member of the epidermal growth factor (EGF) family and an autocrine growth factor for human keratinocytes. Previous studies have shown that AR expression is increased in psoriatic epidermis. To test the hypothesis that aberrant AR expression is central to the development of psoriatic lesions, we constructed a transgene (K14-ARGE) encoding a human keratin 14 promoter-driven AR gene. Our results indicate that transgene integration and subsequent expression of AR in basal keratinocytes correlated with a psoriasis-like skin phenotype. Afflicted mice demonstrated shortened life spans, prominent scaling and erythematous skin with alopecia, and occasional papillomatous epidermal growths. Histologic examination revealed extensive areas of marked hyperkeratosis with focal parakeratosis, acanthosis, dermal and epidermal lymphocytic and neutrophilic infiltration, and dilated blood vessels within the papillary dermis. Our results reveal that AR exerts activity in the skin that is distinct from that of transgenic transforming growth factor-alpha or other cytokines, and induces skin pathology with striking similarities to psoriasis. Our observations also link the keratinocyte EGF receptor-ligand system to psoriatic inflammation, and suggest that aberrant expression of AR in the epidermis may represent a critical step in the development or propagation of psoriatic lesions.

Cook, P W; Piepkorn, M; Clegg, C H; Plowman, G D; DeMay, J M; Brown, J R; Pittelkow, M R



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

SciTech Connect

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.

Zhong, Xia, E-mail: [Department of Emergency, Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China)] [Department of Emergency, Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Li, Xiaonan; Liu, Fuli; Tan, Hui [Department of Emergency, Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China)] [Department of Emergency, Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China); Shang, Deya, E-mail: [Department of Emergency, Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China)] [Department of Emergency, Provincial Hospital Affiliated to Shandong University, Jinan 250021 (China)



Heat-stressed CD4+ T lymphocytes: differential modulations of adhesiveness to extracellular matrix glycoproteins, proliferative responses and tumour necrosis factor-alpha secretion.  

PubMed Central

Although cells of the immune system often function under feverish conditions, the effects of elevated temperatures on T cells have not been fully elucidated. Herein, the effects of a 1-hr exposure to 41 degrees of CD4+ human T cell were studied. Heat-shock treatment of activated CD4+ T cells reduced their adhesion to fibronectin and laminin, the major adhesive glycoproteins of the extracellular matrix (ECM) by 25-40%. This decrease was partially due to a minor decrease in the surface expression of beta 1 integrins, which specifically interact with fibronectin and laminin. In contrast, the capacities of heat-stressed T cells to proliferate and to secrete tumour necrosis factor-alpha (TNF-alpha) were increased upon cell activation. In vivo, heat-treated antigen-primed murine T cells, injected directly into the antigen challenging site, induced a more severe delayed-type hypersensitivity (DTH) response than those not exposed to elevated temperatures. In contrast, the same heat-treated cells inoculated intravenously did not induce DTH, suggesting that these cells were impaired with respect to penetration of blood vessel walls. Thus, the effects of heat shock on key cellular functions are expressed in different manners: T-cell-ECM adhesiveness and subsequent extravasation are impaired, whereas their abilities to proliferate and to secrete TNF-alpha are augmented.

Hershkoviz, R; Alon, R; Mekori, Y A; Gilat, D; Cahalon, L; Miller, A; Lider, O



Induction of NKG2D ligands by gamma radiation and tumor necrosis factor-alpha may participate in the tissue damage during acute graft-versus-host disease.  


Immunopathology of acute graft-versus-host disease (aGVHD) involves secretion of proinflammatory cytokines with subsequent expression of danger signals by injured host tissues. This explanation, however, does not explain the cluster of aGVHD target organs (skin, gut, and liver). NKG2D ligands (MICA/B and ULBP1-3 proteins) are stress-induced molecules that act as danger signals to alert NK and alphabeta or gammadelta CD8 T cells through engagement of the activating NKG2D receptor. We observed a strong and reversible induction of MICA/B expression in skin and liver sections during aGVHD. Tumor necrosis factor-alpha and gamma-radiation up-regulated expression of MICA/B and ULBP proteins in vitro on skin and intestine epithelial cell lines and ex vivo in normal skin explants. This NKG2D-ligand induction was regulated by a complex interplay between NFkB and JNK activation pathways. Our data suggest that NKG2D ligand induction might participate in the amplification loop that leads to tissue damage during aGVHD. PMID:18360276

Gannagé, Monique; Buzyn, Agnčs; Bogiatzi, Sofia I; Lambert, Marion; Soumelis, Vassili; Dal Cortivo, Liliane; Cavazzana-Calvo, Marina; Brousse, Nicole; Caillat-Zucman, Sophie



The protective effects of ambroxol on radiation lung injury and influence on production of transforming growth factor beta1 and tumor necrosis factor alpha.  


The aim of this article was to investigate the effect of ambroxol on radiation lung injury and the expression of transforming growth factor beta(1) (TGF-beta(1)), as well as tumor necrosis factor alpha (TNF-alpha) in plasma. Totally, 120 patients with locally advanced lung cancer in radiotherapy were randomized into treatment and control groups. Patients in the treatment group took ambroxol orally at a dosage of 90 mg, three times per day for 3 months from the beginning of radiotherapy. The expression of TGF-beta(1) and TNF-alpha in plasma was analyzed. The clinical symptoms and lung diffusing capacity were monitored using high resolving power computed tomography. The level of TGF-beta(1) in the control group was increased (11.8 +/- 5.5 ng/ml), whereas in ambroxol-treated patients, the increase was not significant (5.6 +/- 2.6 ng/ml, P < 0.001). Radiotherapy-induced elevation of TNF-alpha levels, seen in control patients, was also abolished after treatment with ambroxol (5.1 +/- 1.0 vs. 2.4 +/- 0.8 ng/ml, P < 0.001). In the treatment group, carbon monoxide diffusion capacity was not significantly decreased at 6, 12, and 18 months post-radiotherapy, compared with the control group (P < 0.05). Ambroxol decreased the expression of TGF-beta(1) and TNF-alpha, and minimized the diminishment of lung diffusion capacity after radiotherapy. PMID:19636975

Xia, De-Hong; Xi, Lei; Xv, Chen; Mao, Wei-Dong; Shen, Wei-Sheng; Shu, Zhong-Qin; Yang, Hong-Zhi; Dai, Min



Drug-induced expression of intercellular adhesion molecule-1 on lesional keratinocytes in fixed drug eruption.  

PubMed Central

The mechanism(s) and the factor(s) that contribute to preferential localization of fixed drug eruption (FDE) lesions to certain skin sites remain speculative. Previous studies suggested that populations of T cells residing in the lesional epidermis may be involved in selective destruction of the epidermis in FDE. In this study, to define the earliest cellular and molecular events with potential relevance to activation of the epidermal T cells, expression of adhesion molecules on keratinocytes (KC) and vascular endothelium was examined sequentially in the lesional skin of FDE patients after challenge with the causative drug. Rapid and intense intercellular adhesion molecule-1 (ICAM-1) expression was induced on the vascular endothelium and KC as early as 1.5 hours after challenge, at which time E-selectin and vascular cell adhesion molecule-1 (VCAM-1) were not up-regulated. In vitro studies using skin organ culture showed that the lesional KC and endothelium responded more rapidly and intensely to express ICAM-1 to tumor necrosis factor-alpha or interferon-gamma compared with those in the nonlesional skin. Surprisingly, such selective induction of KC ICAM-1 restricted to the lesional skin was also observed after exposure to the causative drug alone in skin organ culture. Pretreatment of the lesional skin with anti-tumor necrosis factor completely abrogated in vitro induction of KC ICAM-1 expression by the drug. Drug-induced, TNF-alpha-dependent KC ICAM-1 expression in the lesional skin suggests that induction of ICAM-1 expression by the lesional KC after ingestion of the drug would probably provide a localized initiating stimulus for activation of the disease-associated epidermal T cells. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5

Teraki, Y.; Moriya, N.; Shiohara, T.



Physiological role of tumor necrosis factor alpha in traumatic muscle injury.  


degenerative and regenerative roles of tumor necrosis factor alpha (TNF-alpha), a pro-inflammatory cytokine with pleiotropic functions, were investigated by using TNF receptor 1 and 2 double knockout (TNFR-DKO) and TNF-alpha antibody neutralized mice following traumatic freeze injury to the tibialis anterior muscle. In wild-type control mice, TNF-alpha mRNA transcripts and protein increased following injury and gradually returned to control (uninjured) levels by 13 days. A reduction in MyoD mRNA expression occurred in TNF-alpha-deficient mice, although there were no visible differences in MyoD immunostaining or histological characteristics in regenerating muscles. At 5 days post-injury, the reductions in isometric strength in TNFR-DKO and TNF-alpha-depleted mice did not differ from that of wild-type mice but by 13 days after injury, the TNFR-DKO and TNF-alpha-depleted mice exhibited strength deficits twice that of wild-type mice (i.e., 27-31% vs 13%). Muscle injury was also accompanied by increased expression of interleukin-6 (IL-6), but IL-6-deficient mice demonstrated MyoD expression and recovery of isometric strength similar to that of wild-type mice. These data indicate that TNF-alpha is involved in the recovery of muscle function after traumatic muscle injury, and this effect might be associated with modulation of muscle regulatory genes, including MyoD. PMID:12207010

Warren, Gordon L; Hulderman, Tracy; Jensen, Nancy; McKinstry, Michael; Mishra, Michael; Luster, Michael I; Simeonova, Petia P



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

PubMed Central

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. Images

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



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

PubMed Central

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

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



NFkappaB-p65 dependent transcriptional regulation of glycosyltransferases in human colon adenocarcinoma HT-29 by stimulation with tumor necrosis factor alpha.  


Regulation of fucosyltransferases (FUTs) and sialyltransferases (STs) in a human colon adenocarcinoma cell line HT-29 and nuclear factor kappaB (NFkappaB)-p65 knockdown HT-29 cells was investigated after stimulation with tumor necrosis factor alpha (TNFalpha) using real time PCR. TNFalpha stimulation induced the biphasic increases in expression of NFkappaB-p65, ST3Gal I, FUT IV, ST3Gal IV and ST6GalNAc III mRNAs and the transient increase in expression of ST6Gal I mRNA and the decrease in ST3GalNAc IV mRNA. In NFkappaB-p65 knockdown HT-29 cells, the biphasic and transient increases in all of these mRNA expression induced with TNFalpha were diminished. On the other hand, NFkappaB-p65 siRNA enhanced the constitutive expression levels of ST3GalNAc IV mRNA which was suppressed by TNFalpha. Transcription activities of ST3Gal I reporter gene from nt -1050 5'-flanking region to translation initiation site which has consensus NFkappaB binding sites were up-regulated by stimulation with TNFalpha in HT-29 cells. The promoter activities for deletion constructs of each NFkappaB binding sites were determined using dual luciferase assay. The results indicated that constitutive promoter activities were detected at nt -120 5'-flanking translation initiation site and TNFalpha enhanced ST3Gal I gene expression through NFkappaB binding sites in HT-29 cells. Combination of stimulation with TNFalpha and NFkappaB knockdown with siRNA is useful for determination of NFkappaB dependent transcriptional regulation. PMID:17142966

Higai, Koji; Ishihara, Saki; Matsumoto, Kojiro



Molecular cloning and tissue distribution of pig transforming growth factor alpha.  

PubMed Central

Transforming growth factor alpha (TGF alpha) was originally identified as a product of tumour tissues and transformed cells in culture. Although it is now clear that expression of this factor is not restricted to neoplastic cells, there remains relatively little information about the sites of expression of TGF alpha in normal tissues. Therefore, an amplified DNA fragment encoding the pig TGF alpha precursor was cloned by reverse transcription-PCR (RT-PCR) using RNA isolated from normal skin tissue as the template. Nucleotide sequence analysis predicts a 160-residue transmembrane polypeptide that differs from the rat, mouse and human TGF alpha precursors at 14, 15 and six sites respectively. The distribution of TGF alpha mRNA in a wide variety of pig tissues was analysed by RT-PCR, using oligonucleotide primers based on the pig TGF alpha cDNA sequence. TGF alpha transcripts were detected in RNA isolated from 17 of the 22 tissues analysed, including four previously unreported sites. Using an antibody raised against a synthetic TGF alpha peptide, we have immunolocalized TGF alpha protein to cells within the red pulp of the spleen and to the distal convoluted tubules of the kidney. Images Figure 1 Figure 4 Figure 5

Vaughan, T J; James, P S; Pascall, J C; Brown, K D



Tumour necrosis factor-alpha regulates expression of the CCAAT-enhancer-binding proteins (C/EBPs) alpha and beta and determines the occupation of the C/EBP site in the promoter of the insulin-responsive glucose-transporter gene in 3T3-L1 adipocytes.  

PubMed Central

We have demonstrated previously that treatment of 3T3-L1 adipocytes with tumour necrosis factor-alpha (TNF) results in a rapid (4 h) and significant (75-80%) reduction in the rate of transcription of the GLUT4 gene. Control of GLUT4 gene transcription has been suggested at least in part to reside with the CCAAT-enhancer-binding protein (C/EBP) family (alpha, beta and delta isoforms) of transcription factors. Using electrophoretic mobility shift assays, we have examined the ability of TNF to alter the occupation of the C/EBP site in the GLUT4 promoter. The data suggest that in fully differentiated adipocytes the C/EBP site is a ligand for predominantly alpha/alpha homodimers; however, after exposure to TNF, a shift in occupancy of the site occurs and the ligands become alpha/beta heterodimers and beta/beta homodimers. Partner selection in dimer formation appears to be controlled by selective translocation of the beta-isoform from the cytosol to the nucleus after exposure of the cells to TNF.

Jain, R; Police, S; Phelps, K; Pekala, P H



Glial Tumor Necrosis Factor Alpha (TNF?) Generates Metaplastic Inhibition of Spinal Learning  

PubMed Central

Injury-induced overexpression of tumor necrosis factor alpha (TNF?) in the spinal cord can induce chronic neuroinflammation and excitotoxicity that ultimately undermines functional recovery. Here we investigate how TNF? might also act to upset spinal function by modulating spinal plasticity. Using a model of instrumental learning in the injured spinal cord, we have previously shown that peripheral intermittent stimulation can produce a plastic change in spinal plasticity (metaplasticity), resulting in the prolonged inhibition of spinal learning. We hypothesized that spinal metaplasticity may be mediated by TNF?. We found that intermittent stimulation increased protein levels in the spinal cord. Using intrathecal pharmacological manipulations, we showed TNF? to be both necessary and sufficient for the long-term inhibition of a spinal instrumental learning task. These effects were found to be dependent on glial production of TNF? and involved downstream alterations in calcium-permeable AMPA receptors. These findings suggest a crucial role for glial TNF? in undermining spinal learning, and demonstrate the therapeutic potential of inhibiting TNF? activity to rescue and restore adaptive spinal plasticity to the injured spinal cord. TNF? modulation represents a novel therapeutic target for improving rehabilitation after spinal cord injury.

Huie, J. Russell; Baumbauer, Kyle M.; Lee, Kuan H.; Bresnahan, Jacqueline C.; Beattie, Michael S.; Ferguson, Adam R.; Grau, James W.



Controlled release by Ca(2+)-sensitive recombinant human tumor necrosis factor-alpha liposomes.  


Recombinant human tumor necrosis factor-alpha (rHuTNF) was entrapped in liposomes consisting of Egg phosphatidylcholine (EggPC) alone, EggPC-egg phosphatidic acid (EggPA) or EggPC-egg phosphatidylglycerol (EggPG). These liposomes, stored in vials, were stable for a month at 4 degrees C. The rHuTNF release from the liposomes were examined in rat plasma and phosphate buffered saline (PBS). rHuTNF was released from the liposomes containing EggPA in rat plasma. The release of rHuTNF was inhibited by EDTA and was induced in PBS containing CaCl2, indicating that this release is induced by Ca2+ ion in the plasma. The release of rHuTNF was promoted by an increase of the EggPA content. In conclusion, we could obtain stable liposomes in a vial and this liposome could immediately release rHuTNF in the rat plasma. Furthermore we were able to control the release rate of rHuTNF from these liposomes. PMID:7774035

Yasui, K; Fujioka, H; Nakamura, Y



Pro-inflammatory cytokines induce c-fos expression followed by IL-6 release in human airway smooth muscle cells.  

PubMed Central

BACKGROUND: Airway smooth muscle (ASM) is considered to be a target for mediators released during airway inflammation. AIMS: To investigate the expression of c-fos, a constituent of the transcription factor activator protein-1, in human ASM cells. In addition, to measure the release of interleukin (IL)-6 into the conditioned medium of stimulated ASM cells, as well as DNA biosynthesis and changes in cell number. METHODS: Serum-deprived human ASM cells in the G0/G1 phase were stimulated with the pro-inflammatory cytokines; tumour necrosis factor-alpha, IL-1beta, IL-5 and IL-6. The expression of mRNA encoding the proto-oncogene c-fos was measured by Northern blot analysis. Cell proliferation was assessed by [3H]-thymidine incorporation assays and cell counting, and IL-6 levels in cell-conditioned medium were measured by enzyme-linked immunosorbent assay. RESULTS: All of the cytokines investigated induced a rapid (within 1 h) and transient increase in the expression of mRNA encoding c-fos, followed by the expression and enhanced release of IL-6. Cell proliferation remained unchanged in cytokine-stimulated cells. CONCLUSIONS: Cytokine-induced c-fos expression in human ASM cells could be described as a marker of cell 'activation'. The possible association of these results with airway inflammation, through secondary intracellular mechanisms such as cytokine production, is discussed.

McKay, S; Bromhaar, M M; de Jongste, J C; Hoogsteden, H C; Saxena, P R; Sharma, H S



Inhibition of cytokine-induced vascular cell adhesion molecule-1 expression; possible mechanism for anti-atherogenic effect of Agastache rugosa.  


Adhesion molecules such as vascular cell adhesion molecule-1 (VCAM-1) play an important role during the early stages of atherogenesis. Agastache rugosa has an anti-atherogenic effect in low density lipoprotein receptor -/- mice. Moreover, A. rugosa reduced macrophage infiltration and VCAM-1 expression has been localized in aortic endothelium that overlies early foam cell lesions. This study ascertained that tilianin (100 microM), a major component of A. rugosa, inhibits the tumor necrotic factor-alpha (TNF-alpha)-induced expression of VCAM-1 by 74% in cultured human umbilical vein endothelial cells (HUVECs). Also, tilianin (100 microM) reduced TNF-alpha-induced activation of nuclear factor-kappaB in HUVECs. PMID:11334881

Hong, J H; Choi, J H; Oh, S R; Lee, H K; Park, J H; Lee, K Y; Kim, J J; Jeong, T S; Oh, G T



Hypoxia Attenuates Purinergic P2X Receptor-Induced Inflammatory Gene Expression in Brainstem Microglia  

PubMed Central

Hypoxia and increased extracellular nucleotides are frequently coincident in the brainstem. Extracellular nucleotides are potent modulators of microglial inflammatory gene expression via P2X purinergic receptor activation. Although hypoxia is also known to modulate inflammatory gene expression, little is known about how hypoxia or P2X receptor activation alone affect inflammatory molecule production in brainstem microglia, nor how hypoxia and P2X receptor signaling interact when they occur together. In this study, we investigated the ability of a brief episode of hypoxia (2hrs) in the presence and absence of the non-selective P2X receptor agonist 2?(3?)-O-(4-benzoylbenzoyl)adenosine-5?-triphosphate (BzATP) to promote inflammatory gene expression in brainstem microglia in adult rats. We evaluated inducible nitric oxide synthase (iNOS), tumor necrosis factor alpha (TNF?) and interleukin-6 (IL-6) mRNA levels in immunomagnetically-isolated brainstem microglia. Whereas iNOS and IL-6 gene expression increased with hypoxia and BzATP alone, TNF? expression was unaffected. Surprisingly, BzATP-induced inflammatory effects are lost after hypoxia, suggesting that hypoxia impairs pro-inflammatory P2X receptor signaling. We also evaluated the expression of key P2X receptors activated by BzATP, namely P2X1, P2X4 and P2X7 receptors. Whereas hypoxia did not alter their expression, BzATP upregulated P2X4 and P2X7 mRNAs; these effects were ablated in hypoxia. Although both P2X4 and P2X7 receptor expression correlated with increased microglial iNOS and IL-6 levels in microglia from normoxic rats, in hypoxia, P2X7 only correlated with IL-6, and P2X4 correlated only with iNOS. In addition, correlations between P2X7 and P2X4 were lost following hypoxia, suggesting that P2X4 and P2X7 receptor signaling differs in normoxia and hypoxia. Together, these data suggest that hypoxia suppresses P2X receptor-induced inflammatory gene expression, indicating a potentially immunosuppressive role of extracellular nucleotides in brainstem microglia following exposure to hypoxia.

Smith, Stephanie M. C.; Mitchell, Gordon S.; Friedle, Scott A.; Sibigtroth, Christine M.; Vinit, Stephane; Watters, Jyoti J.



Behavioral Effects of Systemic Transforming Growth Factor-alpha in Syrian Hamsters  

PubMed Central

The growth factor, transforming growth factor-alpha (TGF-?) is strongly expressed in the hypothalamic circadian pacemaker, the suprachiasmatic nucleus (SCN). TGF-? is one of several SCN peptides recently suggested to function as a circadian output signal for the regulation of locomotor activity rhythms in nocturnal rodents. When infused in the brain, TGF-? suppresses activity. TGF-? suppresses other behaviors as well including feeding, resulting in weight loss. Elevated TGF-? is correlated with some cancers, and it is possible the TGF-? and its receptor, the epidermal growth factor receptor (EGFR), mediate fatigue and weight loss associated with cancer. If true for cancers outside of the brain, then systemic TGF-? should also affect behavior. We tested this hypothesis in hamsters with intraperitoneal injections or week-long subcutaneous infusions of TGF-?. Both treatments suppressed activity and infusions caused reduced food consumption and weight loss. To identify areas of the brain that might mediate these effects of systemic TGF-?, we used immunohistochemistry to localize cells with an activated MAP kinase signaling pathway (phosphorylated ERK1). Cells were activated in two hypothalamic areas, the paraventricular nucleus and a narrow region surrounding the third ventricle. These sites could be targets of TGF-? produced in the SCN but could also mediate effects of elevated TGF-? from tumors both within and outside the central nervous system.

Gilbert, Jenifer; Davis, Fred C.



Etanercept decreases tumor necrosis factor-alpha activity in chronic wound fluid.  


High levels of tumor necrosis factor-alpha (TNF-alpha), a pro-inflammatory cytokine, are present in the wound fluid of chronic nonhealing wounds. This leads to increased inflammation, cytokine expression, and ultimately results in impaired wound healing and tissue destruction. Etanercept is a recombinant fusion protein that consists of the soluble TNF receptor (p75) linked to the Fc portion of human IgG1. It is an effective inhibitor of TNF-alpha and has been shown to provide rapid and sustained improvement in rheumatoid arthritis by acting as a soluble receptor binding TNF-alpha and preventing its proinflammatory activities. Therefore, the aim of this study was to determine whether Etanercept could inhibit TNF-alpha activity in chronic wound fluid. Wound fluid was collected from the venous leg ulcers of 16 different patients. The effect of Etanercept on TNF-alpha activity was evaluated using both a TNF-alpha bioassay and an enzyme-linked immunoassay. Etanercept was found to reduce the cytotoxic effect of chronic wound fluid on L929 fibroblasts by approximately 30% and neutralized TNF-alpha binding in the enzyme-linked immunoassay by up to 80%. Direct application of Etanercept to chronic wounds may therefore reduce the inflammatory activity of TNF-alpha, which could reduce the chronicity of venous leg ulcers and thus aid in the healing of these wounds. PMID:16939569

Cowin, Allison J; Hatzirodos, Nicholas; Rigden, Justin; Fitridge, Robert; Belford, David A



Association of transforming growth-factor alpha gene polymorphisms with nonsyndromic cleft palate only (CPO)  

SciTech Connect

Genetic analysis and tissue-specific expression studies support a role for transforming growth-factor alpha (TGFA) in craniofacial development. Previous studies have confirmed an association of alleles for TGFA with nonsyndromic cleft lip with or without cleft palate (CL/P) in humans. The authors carried out a retrospective association study to determine whether specific allelic variants of the TGFA gene are also associated with cleft palate only (CPO). The PCR products from 12 overlapping sets of primers to the TGFA cDNA were examined by using single-strand conformational polymorphism analysis. Four DNA polymorphic sites for TGFA were identified in the 3[prime] untranslated region of the TGFA gene. These variants, as well as previously identified RFLPs for TGFA, were characterized in case and control populations for CPO by using X[sup 2] analysis. A significant association between alleles of TGFA and CPO was identified which further supports a role for this gene as one of the genetic determinants of craniofacial development. Sequence analysis of the variants disclosed a cluster of three variable sites within 30 bp of each other in the 3[prime] untranslated region previously associated with an antisense transcript. These studies extend the role for TGFA in craniofacial morphogenesis and support an interrelated mechanism underlying nonsyndromic forms of CL/P. 46 refs., 3 figs., 3 tabs.

Shiang, R. (Univ. of California, Irvine, CA (United States)); Lidral, A.C.; Ardinger, H.H.; Murray, J.C.; Romitti, P.A.; Munger, R.G.; Buetow, K.H.



Tumor necrosis factor alpha antagonism improves neurological recovery in murine intracerebral hemorrhage  

PubMed Central

Background Intracerebral hemorrhage (ICH) is a devastating stroke subtype characterized by a prominent neuroinflammatory response. Antagonism of pro-inflammatory cytokines by specific antibodies represents a compelling therapeutic strategy to improve neurological outcome in patients after ICH. To test this hypothesis, the tumor necrosis factor alpha (TNF-?) antibody CNTO5048 was administered to mice after ICH induction, and histological and functional endpoints were assessed. Methods Using 10 to 12-week-old C57BL/6J male mice, ICH was induced by collagenase injection into the left basal ganglia. Brain TNF-? concentration, microglia activation/macrophage recruitment, hematoma volume, cerebral edema, and rotorod latency were assessed in mice treated with the TNF-? antibody, CNTO5048, or vehicle. Results After ICH induction, mice treated with CNTO5048 demonstrated reduction in microglial activation/macrophage recruitment compared to vehicle-treated animals, as assessed by unbiased stereology (P = 0.049). This reduction in F4/80-positive cells was associated with a reduction in cleaved caspase-3 (P = 0.046) and cerebral edema (P = 0.026) despite similar hematoma volumes, when compared to mice treated with vehicle control. Treatment with CNTO5048 after ICH induction was associated with a reduction in functional deficit when compared to mice treated with vehicle control, as assessed by rotorod latencies (P = 0.024). Conclusions Post-injury treatment with the TNF-? antibody CNTO5048 results in less neuroinflammation and improved functional outcomes in a murine model of ICH.



Interleukin-6, but not tumour necrosis factor-alpha, increases lipogenesis in rat hepatocyte primary cultures.  

PubMed Central

The Kupffer-cell products interleukin-6 (IL-6) and tumour necrosis factor-alpha (TNF-alpha) have been shown to stimulate hepatic lipogenesis in vivo. Studies were performed to define the direct effects of these cytokines on lipogenesis in primary-culture rat hepatocytes. Hepatocytes were cultured in the presence of IL-6 or TNF-alpha for periods of 24-72 h. IL-6 increased hepatocyte protein content per microgram of DNA. IL-6 also caused a dose- and time-dependent induction of hepatocyte capacity for incorporation of [2-14C]pyruvate into fatty acids (56% increase by 12.5 ng/ml IL-6 after 72 h of cytokine exposure). This increase in cellular lipogenic capacity was confirmed by using 3H2O incorporation into fatty acids as tracer. TNF-alpha did not increase hepatocyte lipogenesis. In contrast with studies in vivo, neither IL-6 nor TNF-alpha had any acute (2 h of exposure) effects on rates of lipogenesis. Both IL-6 and TNF-alpha are known to increase macrophage prostaglandin synthesis acutely. The prostaglandin E agonist misoprostol free acid (0.1 microM) acutely increased hepatocyte lipogenic rates by 14%. Thus, IL-6 can directly induce hepatocyte lipogenic capacity, and E-series prostaglandins can antagonize the acute inhibition of lipogenesis by glucagon. The observations provide further evidence for the role of Kupffer-cell products in the regulation of hepatocyte metabolism.

Brass, E P; Vetter, W H



Rubus croceacanthus Leveille inhibits mast cell-mediated anaphylactic-like reaction and tumor necrosis factor-alpha secretion.  


This work aims at examining the effect of the concentrated methanol extract of Rubus croceacanthus Leveille (RCL) on mast cell-mediated anaphylactic-like reaction in a murine model. RCL inhibited compound 48/80-induced systemic anaphylactic-like reaction. When RCL was given as pre-treatment at concentrations ranging from 0.01 to 1 mg/ml, the histamine release from rat peritoneal mast cells induced by compound 48/80 or anti-dinitrophenyl (DNP) immunoglobulin E (IgE) was reduced in a dose-dependent manner. RCL also inhibited passive cutaneous anaphylaxis activated by anti-DNP IgE. In addition, RCL inhibited phorbol 12-myristate 13-acetate and A23187-induced tumor necrosis factor-alpha secretion from human mast cell line HMC-1 cells. These results indicate that RCL may possess a strong anti-anaphylactic activity. PMID:15340219

Moon, Phil-Dong; Choi, In-Young; Na, Ho-Jeong; Jeong, Hyun-Ja; Kim, Cheorl-Ho; Kim, Hyung-Ryong; Kim, Yun-Kyung; Park, Seong Kyu; Hong, Seung-Heon; Kim, Hyung-Min



Regulation of collagen deposition and lysyl oxidase by tumor necrosis factor-alpha in osteoblasts.  


Tumor necrosis factor-alpha (TNF-alpha) inhibits osteoblast function in vitro by inhibiting collagen deposition. Studies generally support that TNF-alpha does not inhibit collagen biosynthesis by osteoblasts but that collagen deposition is in some way diminished. The study investigated TNF-alpha regulation of biosynthetic enzymes and proteins crucial for posttranslational extracellular collagen maturation in osteoblasts including procollagen C-proteinases, procollagen C-proteinase enhancer, and lysyl oxidase. The working hypothesis is that such regulation could inhibit collagen deposition by osteoblasts. We report that in phenotypically normal MC3T3-E1 osteoblasts, TNF-alpha decreases collagen deposition without decreasing collagen mRNA levels or procollagen protein synthesis. Analyses of the cell layers revealed that TNF-alpha diminished the levels of mature collagen cross-links, pyridinoline and deoxypyridinoline. Further analyses revealed that the mRNA expression for lysyl oxidase, the determining enzyme required for collagen cross-linking, is down-regulated by TNF-alpha in a concentration- and time-dependent manner by up to 50%. The decrease was accompanied by a significant reduction of lysyl oxidase protein levels and enzyme activity. By contrast, Northern and Western blotting studies revealed that procollagen C-proteinases bone morphogenic protein-1 and mammalians Tolloid and procollagen C-proteinase enhancer were expressed in MC3T3-E1 cells and not down-regulated. The data together demonstrate that TNF-alpha does not inhibit collagen synthesis but does inhibit the expression and activity of lysyl oxidase in osteoblasts, thereby contributing to perturbed collagen cross-linking and accumulation. These studies identify a novel mechanism in which proinflammatory cytokine modulation of an extracellular biosynthetic enzyme plays a determining role in the control of collagen accumulation by osteoblasts. PMID:15138266

Pischon, Nicole; Darbois, Laurent M; Palamakumbura, Amitha H; Kessler, Efrat; Trackman, Philip C



Inhibition of the inflammatory cytokine tumor necrosis factor-alpha with etanercept provides protection against lethal H1N1 influenza infection in mice  

PubMed Central

Introduction Factors implicated in influenza-mediated morbidity and mortality include robust cytokine production (cytokine storm), excessive inflammatory infiltrates, and virus-induced tissue destruction. Tumor necrosis factor-alpha (TNF-?) is an important pro-inflammatory cytokine present during influenza infection, but it is unclear whether direct inhibition of TNF-? can elicit protection against influenza infection. Methods In this study, the commercially available TNF-? inhibitor etanercept was used to inhibit TNF-? induced by lethal A/FM/1/47 (H1N1) influenza virus infection of mice. The effects of TNF-? inhibition on mouse survival, pathologic changes, immune cell infiltration, inflammatory cytokine secretion, Toll-like receptor expression, and activation of the NF-?B (nuclear factor kappa B) signaling pathway were evaluated. Results The intranasal delivery of etanercept provided significant protection against mortality (30% of mice survived up to 14 days after infection) in mice treated with etanercept. In contrast, no survivors were found beyond 6 days in mice treated with saline after lethal challenge with H1N1 influenza virus. It was observed that etanercept significantly reduced inflammatory cell infiltration (for example, macrophages and neutrophils), inflammatory cytokine secretion (for example, interleukin-6, TNF-?, and interferon gamma), and expression of Toll-like receptors (TLR-3, TLR-4, and TLR-7). Etanercept also downregulated and inhibited the cascade proteins of the NF-?B signaling pathway (for example, MyD88, TRIF, NF-?B, and p65), as well as enhanced host control of virus replication. Conclusions These findings indicate that etanercept, by blocking TNF-?, can significantly downregulate excessive inflammatory immune responses and provide protection against lethal influenza infection, making its use a novel strategy for controlling severe influenza-induced viral pneumonia.



Tumor necrosis factor alpha-gene therapy for an established murine melanoma using RGD (Arg-Gly-Asp) fiber-mutant adenovirus vectors.  


Although adenovirus vectors (Ad) provide high-level transduction efficacy to many cell types, extremely high doses of Ad are required for sufficient gene transduction into several tumors, including melanoma. Here, we demonstrated that the expression of coxsackie-adenovirus receptor, a primitive Ad-receptor, was very low in murine and human melanoma cells. We also found that fiber-mutant Ad containing the Arg-Gly-Asp (RGD) sequence in the fiber knob remarkably augmented gene transduction efficacy in melanoma cells by targeting alpha(v)-integrins. In addition, intratumoral injection of RGD fiber-mutant Ad containing the tumor necrosis factor alpha gene (Ad-RGD-TNFalpha) revealed dramatic anti-tumor efficacy through hemolytic necrosis in an established murine B16 BL6 melanoma model. Ad-RGD-TNFalpha required one-tenth the dosage of Ad-TNFalpha to induce an equal therapeutic effect. These results suggest that alpha(v)-integrin-targeted Ad will be a very powerful tool for the advancement of melanoma gene therapy. PMID:11985794

Okada, Yuka; Okada, Naoki; Nakagawa, Shinsaku; Mizuguchi, Hiroyuki; Takahashi, Koichi; Mizuno, Nobuyasu; Fujita, Takuya; Yamamoto, Akira; Hayakawa, Takao; Mayumi, Tadanori



Microarray analysis revealing common and distinct functions of promyelocytic leukemia protein (PML) and tumor necrosis factor alpha (TNF?) signaling in endothelial cells  

PubMed Central

Background Promyelocytic leukemia protein (PML) is a tumor suppressor that is highly expressed in endothelial cells nonetheless its role in endothelial cell biology remains elusive. Tumor necrosis factor alpha (TNF?) is an important cytokine associated with many inflammation-related diseases. We have previously demonstrated that TNF? induces PML protein accumulation. We hypothesized that PML may play a role in TNF? signaling pathway. To identify potential PML target genes and investigate the putative crosstalk between PML’s function and TNF? signaling in endothelial cells, we carried out a microarray analysis in human primary umbilical endothelial cells (HUVECs). Results We found that PML and TNF? regulate common and distinct genes involved in a similar spectrum of biological processes, pathways and human diseases. More importantly, we found that PML is required for fine-tuning of TNF?-mediated immune and inflammatory responses. Furthermore, our data suggest that PML and TNF? synergistically regulate cell adhesion by engaging multiple molecular mechanisms. Our biological functional assays exemplified that adhesion of U937 human leukocytes to HUVECs is co-regulated by PML and TNF? signaling. Conclusions Together, our study identified PML as an essential regulator of TNF? signaling by revealing the crosstalk between PML knockdown-mediated effects and TNF?-elicited signaling, thereby providing novel insights into TNF? signaling in endothelial cells.



IkappaBalpha kinase inhibitor IKI-1 conferred tumor necrosis factor alpha sensitivity to pancreatic cancer cells and a xenograft tumor model.  


Tumor necrosis factor alpha (TNFalpha) has been used to treat patients with certain tumor types. However, its antitumor activity has been undermined by the activation of IkappaBalpha kinase (IKK), which in turn activates nuclear factor-kappaB (NF-kappaB) to help cancer cells survive. Therefore, inhibition of TNFalpha-induced IKK activity with specific IKK inhibitor represents an attractive strategy to treat cancer patients. This study reveals IKI-1 as a potent small molecule inhibitor of IKKalpha and IKKbeta, which effectively blocked TNFalpha-mediated IKK activation and subsequent NF-kappaB activity. Using gene profiling analysis, we show that IKI-1 blocked most of the TNFalpha-mediated mRNA expression, including many genes that play important roles in cell survival. We further show that in vitro and in vivo combination of TNFalpha with IKI-1 had superior potency than either agent alone. This increased potency was due primarily to the increased apoptosis in the presence of both TNFalpha and IKI-1. Additionally, IKKbeta small interfering RNA transfected cells were more sensitive to the treatment of TNFalpha. The study suggests that the limited efficacy of TNFalpha in cancer treatment was due in part to the activation of NF-kappaB, allowing tumor cells to escape apoptosis. Therefore, the combination of IKI-1 with TNFalpha may improve the efficacy of TNFalpha for certain tumor types. PMID:19010928

Zhang, Yixian; Gavriil, Maria; Lucas, Judy; Mandiyan, Sreekala; Follettie, Max; Diesl, Veronica; Sum, Fuk-Wah; Powell, Dennis; Haney, Steve; Abraham, Robert; Arndt, Kim



Tumor necrosis factor alpha, interleukin-1 beta, interleukin-6 and major histocompatibility complex molecules in the normal brain and after peripheral immune challenge.  


The capacity of the brain to activate an inflammatory reaction involving the production of cytokines in response to an immune challenge in the periphery has been well established. Interleukin-1 beta is a cytokine that responds with the most widespread pattern of expression followed by tumor necrosis factor alpha and interleukin-6. In addition, our laboratory has shown that class I major histocompatibility complex molecules are upregulated in the brain in response to peripheral administration of bacterial products. Remarkably, during recent years, all these immune genes have been shown to participate in activity-dependent structural synaptic changes in specific neurochemical circuitries in the normal brain. These processes range from the refinement of synaptic connections in sensory systems to learning and memory storage functions of the hippocampus. Therefore, the mechanisms of defense against pathogens can dramatically affect brain structure and function-inducing changes in cognition, mood and behavior. The immune reactions initiated by viruses, bacteria and parasites may result in latent vulnerabilities which could become manifest with future stressors or challenges. Other inflammatory challenges may function as triggers for uncovering pre-existing vulnerabilities or exacerbation of previous functional deficits, or clinical symptoms of neurological or psychiatric conditions. This review will discuss the importance of infections on basic neuronal processes owing to the alteration in the brain of the balance of cytokines involved in higher brain functions. PMID:16197804

Tonelli, Leonardo H; Postolache, Teodor T



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

PubMed Central

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.

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



Amurubicinol-induced eotaxin-3 expression in human NCI-H69 small cell lung carcinoma cells.  


We previously demonstrated the doxorubicin-induced expression of urokinase-type plasminogen activator (uPA), interleukin-8 (IL-8), monocyte chemoattractant protein-1 (MCP-1) and tumor necrosis factor-alpha in human RC-K8 lymphoma cells and NCI-H69 small cell lung carcinoma cells in which reactive oxygen species might be involved. Amurubicin hydrochloride (AMR), a novel derivative drug of doxorubicin, was recently introduced to clinical practice for treatment of lung cancer in Japan. Therefore, we investigated the effects of AMR on the expression of uPA and chemokines in NCI-H69 cells. AMR and its active form, amurubicinol hydrochloride (AMROH), both induced the expression of uPA, IL-8 and MCP-1 in H69 cells in a dose-dependent manner. When the cultured supernatant obtained from AMR-treated H69 cells was subcutaneously injected into rabbits, migration of a significant number of eosinophils was observed around the injected site. Antigen levels of eotaxin-3, a major migration-factor of eosinophils, were increased in AMROH-treated cells in parallel with the mRNA levels. The induction was observed below the clinically achievable concentration of AMR or AMROH. Thus, the simultaneous induction of uPA, IL-8, MCP-1 and eotaxin-3 may play a role in the pharmacological action of AMR through induction of the interaction between proinflammatory cells and lung carcinoma cells. PMID:16465414

Niiya, Masami; Niiya, Kenji; Shibakura, Misako; Asaumi, Noboru; Yoshida, Chikamasa; Tanimoto, Mitsune



A short-term treatment with tumor necrosis factor-alpha enhances stem cell phenotype of human dental pulp cells  

PubMed Central

Introduction During normal pulp tissue healing, inflammatory cytokines, such as tumor necrosis factor-alpha (TNF-?) or interleukins, act in the initial 48 hours (inflammatory phase) and play important roles not only as chemo-attractants of inflammatory cells and stem/progenitor cells but also in inducing a cascade of reactions toward tissue regeneration or reparative dentin formation or both. Previous reports have shown that inflammatory cytokines regulate the differentiation capacity of dental pulp stem/progenitor cells (DPCs), but none has interrogated the impact of these cytokines on the stem cell phenotype of stem/progenitor cells. This study investigated the effects of a short-term treatment with TNF-? on the stem cell phenotype and differentiation ability of human DPCs. Methods An in vivo mouse model of pulp exposure was performed for analysis of expression of the mesenchymal stem cell marker CD146 in DPCs during the initial stage of inflammatory response. For in vitro studies, human DPCs were isolated and incubated with TNF-? for 2 days and passaged to eliminate TNF-? completely. Analysis of stem cell phenotype was performed by quantification of cells positive for mesenchymal stem cell markers SSEA-4 (stage-specific embryonic antigen 4) and CD146 by flow cytometry as well as by quantitative analysis of telomerase activity and mRNA levels of OCT-4 and NANOG. Cell migration, colony-forming ability, and differentiation toward odontogenesis and adipogenesis were also investigated. Results The pulp exposure model revealed a strong staining for CD146 during the initial inflammatory response, at 2 days after pulp exposure. In vitro experiments demonstrated that a short-term (2-day) treatment of TNF-? increased by twofold the percentage of SSEA-4+ cells. Accordingly, STRO-1, CD146, and SSEA-4 protein levels as well as OCT-4 and NANOG mRNA levels were also significantly upregulated upon TNF-? treatment. A short-term TNF-? treatment also enhanced DPC function, including the ability to form cell colonies, to migrate, and to differentiate into odontogenic and adipogenic lineages. Conclusions A short-term treatment with TNF-? enhanced the stem cell phenotype, migration, and differentiation ability of DPCs.



Hepcidin expression in colon during trinitrobenzene sulfonic acid-induced colitis in rats  

PubMed Central

AIM: To investigate hepcidin expression, interleukin-6 (IL-6) production and iron levels in the rat colon in the presence of trinitrobenzene sulfonic acid (TNBS)-induced colitis. METHODS: In rats, we evaluated the severity of colitis induced by repeated TNBS administration using macroscopic and microscopic scoring systems and myeloperoxidase activity measurements. The colonic levels of hepcidin, tumor necrosis factor alpha (TNF-?), IL-10 and IL-6 were measured by Enzyme-Linked Immunosorbent Assay, and hepcidin-25 expression and iron deposition were analyzed by immunohistochemistry and the Prussian blue reaction, respectively. Stat-3 phosphorylation was assessed by Western blot analysis. Hematological parameters, iron and transferrin levels, and transferrin saturation were also measured. Additionally, the ability of iron, pathogen-derived molecules and IL-6 to induce hepcidin expression in HT-29 cells was evaluated. RESULTS: Repeated TNBS administration to rats resulted in macroscopically and microscopically detectable colon lesions and elevated colonic myeloperoxidase activity. Hepcidin-25 protein levels were increased in colonic surface epithelia in colitic rats (10.2 ± 4.0 pg/mg protein vs 71.0 ± 8.4 pg/mg protein, P < 0.01). Elevated IL-6 levels (8.2 ± 1.7 pg/mg protein vs 14.7 ± 0.7 pg/mg protein, P < 0.05), TNF-? levels (1.8 ± 1.2 pg/mg protein vs 7.4 ± 2.1 pg/mg protein, P < 0.05) and Stat-3 phosphorylation were also observed. Systemic alterations in iron homeostasis, hepcidin levels and anemia were not detected in colitic rats. Iron deposition in the colon was only observed during colitis. Hepcidin gene expression was increased in HT-29 cells after IL-6 and lipopolysaccharide [a toll-like receptor 4 (TLR-4) ligand] treatment. Deferoxamine, ferric citrate and peptidoglycan (a TLR-2 ligand) were unable to alter the in vitro expression of hepcidin in HT-29 cells. CONCLUSION: Colitis increased local hepcidin-25 expression, which was associated with the IL-6/Stat-3 signaling pathway. An increase in local iron sequestration was also observed, but additional studies are needed to determine whether this sequestration is a defensive or pathological response to intestinal inflammation.

Gotardo, Erica Martins Ferreira; Ribeiro, Gilberto de Almeida; Clemente, Thayane Rodrigues Leite; Moscato, Camila Henrique; Tome, Renata Bortolin Guerra; Rocha, Thalita; Pedrazzoli Jr, Jose; Ribeiro, Marcelo Lima; Gambero, Alessandra



Dissecting Cellulitis of the Scalp Responding to Intravenous Tumor Necrosis Factor-alpha Antagonist  

PubMed Central

The authors present the case of a 30-year-old male patient with a severe and long-standing dissecting cellulitis of the scalp. The disease did not respond to conventional treatment, including oral antibiotics, isotretinoin, and prednisolone. Quality of life was significantly impaired. After introduction of anti-tumor necrosis factor-alpha treatment (infliximab), the malodorous discharge stopped, inflammation was reduced significantly, nodules became flat, and pain decreased. The treatment was well tolerated although he developed a temporary psoriasiform rash after the second intravenous infusion. In conclusion, anti-tumor necrosis factor-alpha treatment is a new therapeutic option in this severe and recalcitrant disorder.

Wollina, Uwe; Gemmeke, Astrid; Koch, Andre



[Tumor necrosis factor-alpha in a children population with overweight].  


The child overweight is associated with overweight/obesity at the adult age. The obese adipose tissue produces an increase of proinflammatory cytokines as the tumor necrosis factor alpha (TNF-?), causing a deleterious effect on vascular functions. The aim of this work was to evaluate TNF-? levels in a children's population with overweight and its relationship with clinical and laboratory variables. Thirty overweight children were studied, with ages between 8-13 years old, and twenty control children. In both groups waist circumference was measured and body mass index (BMI) calculated. The inclusion criterium was a >85th < 95 th BMI percentile for age and sex. In both groups were determined: fasting blood glucose (glucose-oxidase method); plasma insulin (ECLIA); plasma fibrinogen (Fg, Clauss method); high sensitivity C reactive protein (hsCRP, immunoturbidimetric method); plasma myeloperoxidase (ELISA); TNF-? (ELISA); lipid profile (enzymatic methods); erythrosedimentation rate (ESR) and homeostasis model assessment index (HOMA). Data were expressed as the median and interquartile range. Correlations between variables were investigated with the Spearman's coefficient. A p < 0.05 was considered significant. The TNF-? levels were higher in overweight children [15.4 (13.2-24.0) vs. 12.7 (11.2-14.8) pg/ml; p = 0.028]. Levels of Fg, plasma insulin, HOMA index, uCRP and triglycerides were also statistically significant higher than the control group. The TNF-? was positively correlated with the waist circumference (r = 0.654; p = 0.021). The high TNF-? levels found, with the CRP and Fg levels, confirm a low grade proinflammatory state associated to abdominal obesity in the studied population. PMID:23924528

Carrizo, Teresita Del R; Díaz, Elba I; Velarde, María S; Prado, María M; Bazán, María C; Abregú, Adela V



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


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 iNOS expression. PMID:12839867

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



A Role for Tumor Necrosis Factor-Alpha (TNF?) in Experimental Bacillus cereus Endophthalmitis Pathogenesis  

PubMed Central

Purpose To determine the contribution of tumor necrosis factor-alpha (TNF?)e pathogenesis of experimental Bacillus cereus endophthalmitis. Methods Experimental B. cereus endophthalmitis was induced in wild-type control (B6.129F1) or age-matched homozygous TNF? knockout mice (TNF??/?, B6.129S6-Tnftm1Gk1/J). At various times postinfection, eyes were analyzed by electroretinography, and harvested for quantitation of bacteria, myeloperoxidase, proinflammatory cytokines and chemokines, and histological analysis. Results B. cereus replicated more rapidly in eyes of TNF??/? mice compared to that of B6.129F1 eyes. Retinal function decreased more rapidly in eyes of TNF??/? mice compared to that of B6.129F1 eyes. Retinal layers were not as structurally intact at 6 and 12 h postinfection in TNF??/? eyes compared to that of B6.129F1 eyes. Histological analysis suggested less PMN infiltration in the vitreous of TNF??/? mice than in B6.129F1 mice. B6.129F1 eyes also had greater myeloperoxidase concentrations than did eyes of TNF??/? mice. In general, concentrations of proinflammatory cytokines and chemokines (IL-1? IL-6, and MIP-1?) were greater in eyes of TNF??/? mice than in B6.129F1 eyes. Conclusions TNF? is important to intraocular pathogen containment by PMN during experimental B. cereus endophthalmitis. In the absence of TNF ?, less PMN migrated into the eye, facilitating faster bacterial replication and retinal function loss. Although greater concentrations of proinflammatory cytokines were synthesized in the absence of TNF?, the resulting inflammation was less, and an equally devastating course of infection occurred.

Ramadan, R.T.; Moyer, A.L.; Callegan, M.C.



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

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.

Wang, Cheng-hu; Cao, Guo-Fan [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China)] [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China); Jiang, Qin, E-mail: [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China)] [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China); Yao, Jin, E-mail: [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China)] [The Affiliated Eye Hospital of Nanjing Medical University, Nanjing 210029 (China)



Effect of leaves of Eriobotrya japonica on anaphylactic allergic reaction and production of tumor necrosis factor-alpha.  


Leaves of Eriobotrya japonica Lindl. (Rosaceae) (LEJL) have been used as traditional medicines for inflammatory diseases and chronic bronchitis. However, its effect on mast cell-mediated anaphylactic reaction is not known. The anaphylactic allergic reaction is involved in many allergic diseases such as asthma and allergic rhinitis. In this report, we investigate the effect of LEJL on the anaphylactic allergic reaction and studied its possible mechanisms of action. LEJL inhibited compound 48/80-induced systemic anaphylactic reactions and serum histamine release in mice. LEJL dose-dependently decreased the IgE-mediated passive cutaneous anaphylaxis and histamine release from mast cells. Furthermore, LEJL decreased the production of tumor necrosis factor-alpha in phorbol 12-myristate 13-acetate and A23187-stimulated human mast cells. These findings provide evidence that LEJL could be a candidate as an anti-allergic agent. PMID:19514997

Kim, Sang-Hyun; Kwon, Young-Ee; Park, Won-Hwan; Jeon, Hoon; Shin, Tae-Yong



Cell-specific effects of pentoxifylline on nitric oxide production and inducible nitric oxide synthase mRNA expression.  

PubMed Central

Cytokine-stimulated astrocytes and macrophages are potent producers of nitric oxide (NO), a free radical proposed to play an important role in organ-specific autoimmunity, including demyelinating diseases of the central nervous system. The aim of this study was to investigate effects of pentoxifylline (PTX), a phosphodiesterase inhibitor with immunomodulatory properties, on NO production and inducible NO synthase (iNOS) mRNA expression in rat astrocytes and macrophages. We have shown that PTX affects cytokine (interferon-gamma, IFN-gamma; interleukin-1, IL-1; tumour-necrosis factor-alpha, TNF-alpha)-induced NO production in both cell types, but in the opposite manner--enhancing in astrocytes and suppressive in macrophages. While PTX did not have any effect on enzymatic activity of iNOS in activated cells, expression of iNOS mRNA was elevated in astrocytes and decreased in macrophages treated with cytokines and PTX. Treatment with PTX alone affected neither NO production nor iNOS mRNA levels in astrocytes or macrophages. This study indicates involvement of different signalling pathways associated with iNOS induction in astrocytes and macrophages, thus emphasizing complexity of regulation of NO synthesis in different cell types. Images Figure 3 Figure 4

Trajkovic, V; Badovinac, V; Popadic, D; Hadzic, O; Stojkovic, M M



LXR antagonists induce ABCD2 expression.  


X-linked adrenoleukodystrophy (X-ALD) is a rare neurodegenerative disorder characterized by the accumulation of very-long-chain fatty acids resulting from a beta-oxidation defect. Oxidative stress and inflammation are also key components of the pathogenesis. X-ALD is caused by mutations in the ABCDI gene, which encodes for a peroxisomal half ABC transporter predicted to participate in the entry of VLCFA-CoA into the peroxisome, the unique site of their beta-oxidation. Two homologous peroxisomal ABC transporters, ABCD2 and ABCD3 have been proven to compensate for ABCD1 deficiency when overexpressed. Pharmacological induction of these target genes could therefore represent an alternative therapy for X-ALD patients. Since LXR activation was shown to repress ABCD2 expression, we investigated the effects of LXR antagonists in different cell lines. Cells were treated with GSK(17) (a LXR antagonist recently discovered from the GlaxoSmithKline compound collection), 22(S)-hydroxycholesterol (22S-HC, another LXR antagonist) and 22R-HC (an endogenous LXR agonist). We observed up-regulation of ABCD2,ABCD3 and CTNNB1 (the gene encoding for beta-catenin, which was recently demonstrated to induce ABCD2 expression) in human HepG2 hepatoma cells and in X-ALD skin fibroblasts treated with LXR antagonists. Interestingly, induction in X-ALD fibroblasts was concomitant with a decrease in oxidative stress. Rats treated with 22S-HC showed hepatic induction of the 3 genes of interest. In human, we show by multiple tissue expression array that expression of ABCD2 appears to be inversely correlated with NR1H3 (LXRalpha) expression. Altogether, antagonists of LXR that are currently developed in the context of dyslipidemia may find another indication with X-ALD. PMID:24239766

Gondcaille, Catherine; Genin, Emmanuelle C; Lopez, Tatiana E; Dias, Alexandre M M; Geillon, Flore; Andreoletti, Pierre; Cherkaoui-Malki, Mustapha; Nury, Thomas; Lizard, Gérard; Weinhofer, Isabelle; Berger, Johannes; Kase, Eili T; Trompier, Doriane; Savary, Stéphane



High-sensitivity C-reactive Protein and Tumor Necrosis Factor Alpha in Pseudoexfoliation Syndrome  

PubMed Central

Objectives The purpose of the present study was to determine the alterations in high-sensitivity C-reactive protein and Tumor Necrosis factor alpha levels in the blood serum of pseudoexfoliation syndrome cases (a disease with similar risk factors as systemic endothelial dysfunction diseases) and to compare the results with healthy individuals. Methods High-sensitivity C-reactive protein and Tumor Necrosis factor alpha levels were determined in 30 cases with pseudoexfoliation syndrome and in 30 control patients of the same age and sex, by enzyme-linked immunosorbent assay. Results The levels of high- sensitivity C-reactive protein and Tumor Necrosis factor alpha in the blood serum of patients with pseudoexfoliation syndrome (3.95±0.88 mg/l, 3.32±0.99 pg/ml, respectively) were significantly higher than in the control group (2.51±0.79mg/l, 0.43±0.15 pg/ml, respectively) p=0.001, p=0.002. Conclusion The results suggest that increased levels of high- sensitivity C-reactive protein and Tumor Necrosis factor alpha, as markers of inflammation and peripheral endothelial dysfunction in pseudoexfoliation syndrome, may be risk factors for systemic and ocular manifestations of pseudoexfoliation syndrome.

Sorkhabi, Rana; Ghorbanihaghjo, Amir; Ahoor, Mohamadhossein; Nahaei, Mehriar; Rashtchizadeh, Nadereh



Gemella morbillorum Bacteremia after Anti-Tumor Necrosis Factor Alpha as Acne Inversa Therapy  

PubMed Central

We present a case of fever, brain abscesses, and Gemella morbillorum bacteremia after anti-tumor necrosis factor alpha (TNF-?) therapy in a 21-year-old acne inversa patient currently taking long-term dapsone. To the best of our knowledge, this is the first report describing such a case. During antimicrobial therapy, the patient developed systemic varicella infection with severe thrombocytopenia.

Vossen, Matthias G.; Gattringer, Klaus B.; Khalifeh, Neda; Koreny, Maria; Spertini, Verena; Mallouhi, Ammar; Willeit, Markus; Volc-Platzer, Beatrix; Asboth, Friederike; Graninger, Wolfgang; Thalhammer, Florian



Mucosal tumour necrosis factor alpha and interleukin-6 in patients with Helicobacter pylori associated gastritis  

Microsoft Academic Search

The production of tumour necrosis factor alpha (TNF alpha) and interleukin-6 by human antral mucosa during short term culture in vitro has been measured by enzyme linked immunosorbent assay. TNF alpha and interleukin-6 concentrations in culture supernatants were significantly greater (p less than 0.001) in patients infected with Helicobacter pylori, all of whom had chronic gastritis, than in patients who

J E Crabtree; T M Shallcross; R V Heatley; J I Wyatt



Interleukin-1 beta and tumor necrosis factor alpha inhibit migration activity of chondrogenic progenitor cells from non-fibrillated osteoarthritic cartilage  

PubMed Central

Introduction The repair capability of traumatized articular cartilage is highly limited so that joint injuries often lead to osteoarthritis. Migratory chondrogenic progenitor cells (CPC) might represent a target cell population for in situ regeneration. This study aims to clarify, whether 1) CPC are present in regions of macroscopically intact cartilage from human osteoarthritic joints, 2) CPC migration is stimulated by single growth factors and the cocktail of factors released from traumatized cartilage and 3) CPC migration is influenced by cytokines present in traumatized joints. Methods We characterized the cells growing out from macroscopically intact human osteoarthritic cartilage using a panel of positive and negative surface markers and analyzed their differentiation capacity. The migratory response to platelet-derived growth factor (PDGF)-BB, insulin-like growth factor 1 (IGF-1), supernatants obtained from in vitro traumatized cartilage and interleukin-1 beta (IL-1?) as well as tumor necrosis factor alpha (TNF-?) were tested with a modified Boyden chamber assay. The influence of IL-1? and TNF-? was additionally examined by scratch assays and outgrowth experiments. Results A comparison of 25 quadruplicate marker combinations in CPC and bone-marrow derived mesenchymal stromal cells showed a similar expression profile. CPC cultures had the potential for adipogenic, osteogenic and chondrogenic differentiation. PDGF-BB and IGF-1, such as the supernatant from traumatized cartilage, induced a significant site-directed migratory response. IL-1? and TNF-? significantly reduced basal cell migration and abrogated the stimulative effect of the growth factors and the trauma supernatant. Both cytokines also inhibited cell migration in the scratch assay and primary outgrowth of CPC from cartilage tissue. In contrast, the cytokine IL-6, which is present in trauma supernatant, did not affect growth factor induced migration of CPC. Conclusion These results indicate that traumatized cartilage releases chemoattractive factors for CPC but IL-1? and TNF-? inhibit their migratory activity which might contribute to the low regenerative potential of cartilage in vivo.



Protease-activated receptor-2 induces proinflammatory cytokine and chemokine gene expression in canine keratinocytes.  


Although the molecular basis of the allergenicity remains to be fully elucidated, the ability of allergens to elicit allergic responses is at least partly attributed to their proteolytic activity. Protease-activated receptor-2 (PAR-2) is a G protein-coupled receptor that is activated by site-specific proteolysis by serine proteases and is known to mediate inflammatory processes in various tissues. In this study, we investigated the effects of trypsin, a major serine protease, and a human PAR-2 agonist peptide (SLIGKV-NH2) on proinflammatory cytokine and chemokine gene expression in the canine keratinocyte cell line CPEK. The expression of PAR-2 mRNA and protein in CPEK cells was detected by RT-PCR and Western blotting, respectively. The localization of PAR-2 in CPEK was examined by immunofluorescence. The mRNA expression levels of proinflammatory cytokines and chemokines were quantified by real-time RT-PCR. The free intracellular Ca(2+) concentration was measured using the Ca(2+)-sensitive fluorescent dye. CPEK cells constitutively expressed PAR-2 mRNA and protein. Stimulation of CPEK cells with trypsin induced significant upregulation of the mRNA expression levels of tumor necrosis factor alpha (TNF-?, P<0.05), granulocyte-macrophage colony-stimulating factor (GM-CSF, P<0.01), thymus and activation regulated chemokine (TARC/CCL17, P<0.01), and interleukin 8 (IL-8/CXCL8, P<0.01). Similarly, the PAR-2 agonist peptide increased the mRNA expression levels of TNF-? (P<0.05), GM-CSF (P<0.05), TARC/CCL17 (P<0.05), and IL-8/CXCL8 (P<0.05) in CPEK cells. Both trypsin and the PAR-2 agonist peptide increased the intracellular Ca(2+) concentration and PAR-2 internalization. These results suggest that PAR-2 activation can augment inflammatory cytokine and chemokine expression in canine keratinocytes, and it may initiate allergic inflammation through the proteolytic activity of allergens in canine atopic dermatitis. PMID:23465358

Maeda, Shingo; Maeda, Sadatoshi; Ohno, Koichi; Kaji, Noriyuki; Hori, Masatoshi; Fujino, Yasuhito; Tsujimoto, Hajime



Gravity-Induced Gene Expression in Plants.  

NASA Astrophysics Data System (ADS)

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 sequence identity as well as a conserved pattern of transcript abundance changes after gravity stimulation between corn pulvinus tissue and Arabidopsis root apices. The functions of these genes in gravitropic responses are currently being analyzed and should give us important information about evolutionary conserved elements in plant gravity signal transduction. (This research was funded by NASA). Kimbrough, J. M., R. Salinas-Mondragon, et al. (2004). "The Fast and Transient Transcriptional Network of Gravity and Mechanical Stimulation in the Arabidopsis Root Apex." Plant Physiol. 136(1): 2790-2805. Moseyko, N., T. Zhu, et al. (2002). "Transcription profiling of the early gravitropic response in Arabidopsis using high-density oligonucleotide probe microarrays." Plant Physiol 130(2): 720-8. Salinas-Mondragon, R., A. Brogan, et al. (2005). "Gravity and light: integrating transcriptional regulation in roots." Gravit Space Biol Bull 18(2): 121-2.

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


Role of p38 MAPK in LPS induced pro-inflammatory cytokine and chemokine gene expression in equine leukocytes.  


Endotoxemia occurs when bacterial lipopolysaccharide (LPS) in the blood induces a dysregulated inflammatory response, resulting in circulatory shock and multi-organ failure. Laminitis is a common complication in endotoxemic horses and is frequently the reason for humane euthanasia of these cases. Blood leukocytes are a principal target of LPS in endotoxemia leading to activation of multiple signal transduction pathways involved in the induction of a number of pro-inflammatory genes. In other animal models, the p38 mitogen activated protein kinase (MAPK) pathway has been associated with induced expression of tumor necrosis factor-alpha (TNFalpha), interleukin (IL)-1beta, IL-6 and IL-8. The goal of this study was to determine the role of the p38 MAPK pathway in the induction of these pro-inflammatory cytokine and chemokine genes in LPS-stimulated equine leukocytes. Stimulation of equine peripheral blood leukocytes resulted in an increase in TNFalpha, IL-1beta, IL-6 and IL-8 mRNA levels. Pharmacological inhibition of p38 MAPK activity with SB203580 or SB202190 reduced the ability of LPS stimulation to increase mRNA concentrations for all four genes. However, only SB203580 pretreatment significantly reduced LPS-stimulated IL-1beta and IL-8 mRNA expression and only pretreatment with SB202190 significantly reduced LPS-stimulated TNFalpha and IL-6 mRNA expression. From this study we conclude TNFalpha, IL-1beta, IL-6 and IL-8 are induced upon LPS stimulation of equine leukocytes and that this induction of gene expression is dependent on the p38 MAPK pathway. However, there are differences in the efficacy of the p38 inhibitors tested here that may be explained by differences in specificity or potency. This study provides evidence for the use of selective p38 MAPK inhibitors as potential therapeutics for the treatment of equine endotoxemia. PMID:19070370

Neuder, Laura E; Keener, Jamie M; Eckert, Rachael E; Trujillo, Jennifer C; Jones, Samuel L



S-Adenosylmethionine and methylthioadenosine inhibit cellular FLICE inhibitory protein expression and induce apoptosis in colon cancer cells.  


S-Adenosylmethionine (SAMe) and its metabolite 5'-methylthioadenosine (MTA) inhibit mitogen-induced proliferative response in liver and colon cancer cells. SAMe and MTA are also proapoptotic in liver cancer cells by selectively inducing Bcl-x(S) expression. The aims of this work were to assess whether these agents are proapoptotic in colon cancer cells, and if so, to elucidate the molecular mechanisms. We found that both SAMe and MTA are proapoptotic in HT-29 and RKO cells in a dose- and time-dependent manner. Gene microarray uncovered down-regulation of cellular FLICE inhibitory protein (cFLIP). SAMe and MTA treatment led to a decrease in the mRNA and protein levels of both the long and short cFLIP isoforms. This required de novo RNA synthesis and was associated with activation of procaspase-8, Bid cleavage, and release of cytochrome c from the mitochondria. Inhibiting caspase 8 activity or overexpression of cFLIP protected against apoptosis, whereas supplementing with polyamines did not. SAMe and MTA treatment sensitized RKO cells to tumor necrosis factor alpha-related apoptosis-inducing ligand-induced apoptosis. Although SAMe and MTA are proapoptotic in colon cancer cells, they have no toxic effects in NCM460 cells, a normal colon epithelial cell line. In contrast to liver cancer cells, SAMe and MTA had no effect on Bcl-x(S) expression in colon cancer cells. In conclusion, SAMe and MTA are proapoptotic in colon cancer cells but not normal colon epithelial cells. One molecular mechanism identified is the inhibition of cFLIP expression. SAMe and MTA may be attractive agents in the chemoprevention and treatment of colon cancer. PMID:19372210

Li, Tony W H; Zhang, Qingsong; Oh, Pilsoo; Xia, Meng; Chen, Hui; Bemanian, Sean; Lastra, Natalie; Circ, Magda; Moyer, Mary Pat; Mato, José M; Aw, Tak Yee; Lu, Shelly C



Wnt/beta-catenin signaling regulates cytokine-induced human inducible nitric oxide synthase expression by inhibiting nuclear factor-kappaB activation in cancer cells.  


The human inducible nitric oxide synthase (hiNOS) gene is regulated by nuclear factor kappaB (NF-kappaB) and has recently been shown to be a target of the Wnt/beta-catenin pathway. In this study, we tested the hypothesis that Wnt/beta-catenin signaling might regulate cytokine- or tumor necrosis factor alpha (TNFalpha)-induced hiNOS expression through interaction with NF-kappaB. A cytokine mixture of TNFalpha + interleukin (IL)-1beta + IFNgamma induced a 2- to 3-fold increase in hiNOS promoter activity in HCT116 and DLD1 colon cells, but produced a 2-fold decrease in SW480 colon cancer cells. A similar differential activity was seen in liver cancer cells (HepG2, Huh7, and Hep3B). Overexpression of beta-catenin produced a dose-dependent decrease in NF-kappaB reporter activity and decreased cytokine mixture-induced hiNOS promoter activity. Gel shift for TNFalpha-induced hiNOS NF-kappaB activation showed decreased p50 binding and decreased NF-kappaB reporter activity in the beta-catenin-mutant HAbeta18 cells. Conversely, enhanced p50 binding and increased NF-kappaB reporter activity were seen in HAbeta85 cells, which lack beta-catenin signaling. Coimmunoprecipitation confirmed that beta-catenin complexed with both p65 and p50 NF-kappaB proteins. NF-kappaB-dependent Traf1 protein expression also inversely correlated with the level of beta-catenin. Furthermore, SW480 cells stably transformed with wild-type adenomatous polyposis coli showed decreased beta-catenin protein and increased TNFalpha-induced p65 NF-kappaB binding as well as iNOS and Traf1 expression. Finally, beta-catenin inversely correlated with iNOS and Fas expression in vivo in hepatocellular carcinoma tumor samples. Our in vitro and in vivo data show that beta-catenin signaling inversely correlates with cytokine-induced hiNOS and other NF-kappaB-dependent gene expression. These findings underscore the complex role of Wnt/beta-catenin, NF-kappaB, and iNOS signaling in the pathophysiology of inflammation-associated carcinogenesis. PMID:19383900

Du, Qiang; Zhang, Xinglu; Cardinal, Jon; Cao, Zongxian; Guo, Zhong; Shao, Lifang; Geller, David A



Involvement of H-Ras and reactive oxygen species in proinflammatory cytokine-induced matrix metalloproteinase-13 expression in human articular chondrocytes.  


Proinflammatory cytokines such as interleukin-1 beta (IL-1?) and tumor necrosis factor alpha (TNF-?) enhance degradation of cartilage-specific, type II collagen by matrix metalloproteinase-13 (MMP-13). We investigated the previously unknown role of H-Ras and reactive oxygen species (ROS) in the cytokine induction of MMP-13 gene expression in human articular chondrocytes by using pharmacological inhibitors, RNA interference (RNAi) and antioxidants. Manumycin A, an inhibitor of H-Ras farnesylation by farnesyltransferase, suppressed IL-1?- and TNF-?-induced MMP-13 mRNA and protein expression. Small interfering RNA (siRNA)-mediated H-Ras silencing down-regulated MMP-13 mRNA and protein induction by IL-1? and TNF-?. Nicotinamide adenine dinucleotide phosphate oxidase (NADPH oxidase/NOX) inhibitor, diphenyleneiodonium (DPI) suppressed cytokine-induced MMP-13 expression and superoxide production. Apocynin, another NOX inhibitor, also diminished MMP-13 induction. Deoxyglucose an antimetabolite of glucose metabolism reduced MMP-13 increase. Role of NOX-mediated ROS production was reaffirmed by the observation that the antioxidants, trolox, nordihydroguaiaretic acid (NDGA), quercetin and resveratrol downregulated cytokine-induced MMP-13 mRNA and protein expression. These results provide strong pharmacological and genetic evidence for the implication of H-Ras and NADPH oxidase-generated superoxide production in MMP-13 gene regulation by IL-1? and TNF-?. These proteins could be potentially targeted for therapeutic inhibition of MMP-13-driven cartilage erosion by using H-Ras and NOX inhibitors and antioxidants. PMID:21211511

Ahmad, Rasheed; Sylvester, Judith; Ahmad, Mushtaq; Zafarullah, Muhammad



Asiatic Acid Alleviates Hemodynamic and Metabolic Alterations via Restoring eNOS/iNOS Expression, Oxidative Stress, and Inflammation in Diet-Induced Metabolic Syndrome Rats  

PubMed Central

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-?) 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-?, 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.

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



Preincubation of Candida albicans strains with amphotericin B reduces tumor necrosis factor alpha and interleukin-6 release by human monocytes.  

PubMed Central

The release of tumor necrosis factor alpha (TNF-alpha) and interleukin-6 (IL-6) by human monocytes stimulated with whole heat-killed Candida albicans CA3 (a clinical isolate) and CA2 (a germ tube-negative mutant) either treated or not treated with amphotericin B was investigated. The optimal release of the cytokines was observed at 24 h of incubation of the yeasts with the monocytes for both TNF-alpha and IL-6. The levels ranged from 10,500 to 19,000 U/ml for TNF-alpha and from 350 to 460 pg/ml for IL-6. Germ tube-negative mutant CA2 induced the release of TNF-alpha at levels significantly (P < 0.05) lower than those induced by clinical isolate CA3, while no major differences were observed between the two strains with regard to their capacity to induce the release of IL-6. In all instances, preincubation of the yeasts with a sublethal concentration of amphotericin significantly reduced cytokine production. These results suggest that drug-induced alterations of fungal outer structures may affect the interactions between the yeasts and the monocytes, resulting in a reduced level of secretion of cytokines.

Raponi, G; Ghezzi, M C; Mancini, C; Filadoro, F



The locus of tumor necrosis factor-alpha action in lung inflammation.  


The pulmonary host response to infection and inflammation appears, at least in part, to be compartmentalized from the systemic host response. Tumor necrosis factor-alpha (TNF-alpha) has been implicated in lung inflammation and injury, but its site(s) of action has not been clearly defined. To investigate this, transgenic mice (surfactant apoprotein C promotor/soluble TNF receptor type II-Fc fusion protein ([SPCTNFRIIFc] mice) were generated in which TNF-alpha was selectively antagonized in the distal lung through tissue-specific expression of sTNFRIIFc, a soluble TNF inhibitor. The lung inflammatory response in these mice to pulmonary challenge with Micropolyspora faeni antigen or lipopolysaccharide (LPS) was compared with the response of wild-type mice, wild-type mice treated with recombinant sTNFRIIFc intravenously, and type I TNF-receptor knockout mice. Recruitment of polymorphonuclear leukocytes (PMN) to the lung after challenge with M. faeni antigen was essentially abolished in the TNFRI knockout mice and markedly reduced in the SPCTNFRIIFc mice. Wild-type mice given sTNFRIIFc intravenously in amounts resulting in lung concentrations similar to those in SPCTNFRIIFc mice also showed significantly reduced lung PMN recruitment, whereas those given doses that achieved such concentrations in the blood but low levels in the lung did not. In contrast, PMN recruitment to the lung following aerosol challenge with LPS was reduced significantly in the TNFRI knockout mice and in mice given high-dose sTNFRIIFc intravenously, but was not reduced significantly in SPCTNFRIIFc mice. Thus, inhibition of PMN recruitment in response to M. faeni antigen correlated largely with the extent of intrapulmonary inhibition of TNF-alpha, whereas the response to LPS correlated best with the extent of extrapulmonary inhibition of TNF-alpha. These studies indicate that TNF-alpha may act at different loci to mediate lung inflammation, with the site of action depending in part on the nature of the inflammatory stimulus, and that SPCTNFRIIFc mice provide a tool by which the locus of TNF action can be addressed. PMID:9843922

Smith, S; Skerrett, S J; Chi, E Y; Jonas, M; Mohler, K; Wilson, C B



Upregulation of Tumor Necrosis Factor Alpha and Interleukin1bin Q Fever Endocarditis  

Microsoft Academic Search

The occurrence of Q fever endocarditis likely involves some alterations in the responses of monocytes, the in vivo targets ofCoxiella burnetii. To test this hypothesis, the production of the inflammatory cytokines tumor necrosis factor alpha, interleukin-1b, and interleukin-6 was assessed in monocytes from patients with Q fever endocarditis. Spontaneous transcription and secretion of tumor necrosis factor and interleukin-1 were signif-




Anti-Tumour Necrosis Factor-alpha Therapy in Crohn's Disease: Clinical and Health Economic Aspects  

Microsoft Academic Search

Objectives: Crohn's disease is a chronic, relapsing inflammatory disease of the gastro- intestinal tract. Tumour necrosis factor-alpha (TNF-?) is a pro-inflammatory protein that is believed to play a major role in the pathogenesis of Crohn's disease (CD). Infliximab, a chimeric anti-TNF-? proclonal antibody, which inhibits the bioactivity of TNF-?, is a recent and exciting strategy in the treatment of CD.

Fiona McGuire


Tumour necrosis factor-alpha and other cytokines in Guillain-Barré syndrome  

Microsoft Academic Search

The efficacy of plasma exchange implicates myelinotoxic humoral factors in the pathogenesis of Guillain-Barré syndrome. Candidate factors include autoantibodies to peripheral nerve myelin, which are not unique to Guillain-Barré syndrome; and cytokines such as tumour necrosis factor-alpha (TNF-alpha) which are T cell\\/macrophage products. Plasma cytokine concentrations were determined in 26 patients with Guillain-Barré syndrome undergoing plasma exchange, 25 with other

A R Exley; N Smith; J B Winer



Adalimumab (Humira ™ ): a promising monoclonal anti-tumor necrosis factor alpha in ophthalmology  

Microsoft Academic Search

Tumor necrosis factor alpha (TNF-?) is a key soluble mediator involved in the inflammatory cascade of many disorders including\\u000a uveitis. Among the anti-TNF-? agents, one of the most used in immune-mediated diseases, such as inflammatory arthropathies,\\u000a is adalimumab (Humira™, Abbott Pharmaceutical Inc.), a fully humanized antibody. The purpose of this review is to analyze the main pharmacological\\u000a and clinical aspects

Piergiorgio Neri; Manuela Zucchi; Pia Allegri; Marta Lettieri; Cesare Mariotti; Alfonso Giovannini



Tumor necrosis factor-alpha receptor ablation in a chronic MPTP mouse model of Parkinson's disease  

Microsoft Academic Search

Recently, we demonstrated that mice deficient of the pro-inflammatory cytokine tumor necrosis factor-alpha (TNF-alpha) were partly protected against 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) neurotoxicity. Here we extended the study and investigated TNF-alpha receptor 1 (?\\/?) (TNFR1) and TNF-alpha receptor 2 (?\\/?) (TNFR2) mice using a chronic MPTP dosing regimen (15mg\\/kg MPTP on 8 consecutive days). One week after the last MPTP treatment, HPLC

Andreas Leng; Anna Mura; Joram Feldon; Boris Ferger



Early administration of tumor necrosis factor-alpha antagonist promotes survival of transplanted neural stem cells and axon myelination after spinal cord injury in rats.  


Neural stem cell (NSC) transplantation has been reported to be a leading strategy to stimulate neuroplasticity, repair neuronal loss and promote the morphologic and functional recovery of spinal cord injury (SCI). However, massive death of transplanted NSCs is still a problem, which is considered to be related to a series of pro-inflammatory cytokines that induce apoptosis, extensive demyelination and axonal destruction. Tumor necrosis factor alpha (TNF-?), as one of the major inflammation initiators, contributes to secondary neural cell death. We previously found that the administration of the TNF-? antagonist etanercept during the acute phase of SCI can reduce the apoptosis of neurons and oligodendrocytes. To investigate whether etanercept can suppress transplanted NSC apoptosis and promote NSC survival, axon myelination and functional recovery, we tested the combination strategy of the early administration of etanercept and NSC transplantation. First we observed that etanercept suppressed the TNF-? expression and apoptosis of transplanted NSCs by Western blot, TUNEL and immunofluorescence staining. The Basso, Beattle and Bresnahan scale and motor-evoked potential were used to evaluate functional recovery. The results suggest significantly better recovery after combination therapy. Further, histopathological alterations were evaluated by hematoxylin and eosin staining and Nissl staining. These procedures showed that the early administration of etanercept improved survival of neurons in the ventral horn, restored neural morphology and produced a smaller cavity area. We observed most abundant NF-positive fibers after the combination treatment, indicating that combination therapy retained and promoted neural regeneration. Finally, the early suppression of TNF-? reduced the occurrence of demyelination, and the combination therapy led to more myelinated axons, as shown by electron microscopy. These data suggest that this strategy significantly protected transplanted NSCs via the anti-inflammation and anti-apoptosis effects of etanercept, promoting re-myelination, neural regeneration and locomotor function. PMID:24887643

Wang, Le; Wei, Fu-Xin; Cen, Jing-Sheng; Ping, Su-Ning; Li, Zi-Qing; Chen, Ning-Ning; Cui, Shang-Bin; Wan, Yong; Liu, Shao-Yu



Molecular Determinants of NF-kB-Inducing Kinase Action  

Microsoft Academic Search

NF-kB corresponds to an inducible eukaryotic transcription factor complex that is negatively regulated in resting cells by its physical assembly with a family of cytoplasmic ankyrin-rich inhibitors termed IkB. Stimulation of cells with various proinflammatory cytokines, including tumor necrosis factor alpha (TNF-a), induces nuclear NF-kB expression. TNF-a signaling involves the recruitment of at least three proteins (TRADD, RIP, and TRAF2)




Establishment and characterization of differentiated, nontransformed hepatocyte cell lines derived from mice transgenic for transforming growth factor alpha.  


Hepatocytes are extensively used in studies of gene regulation but cannot be maintained in long-term culture as replicating, differentiated cells while remaining nontumorigenic. We have derived two hepatocyte lines from livers of transgenic mice overexpressing transforming growth factor alpha, a potent hepatocyte mitogen, which overcome these limitations. The transgenic hepatocytes were maintained for > or = 2 months in serum-supplemented primary culture and gave rise to cell lines, of which two (AML12 and AML14) have been cultured for > 1.5 years (> 80 passages). Both lines have typical hepatocyte features such as peroxisomes and bile canalicular-like structures, do not grow in soft agar, and are nontumorigenic in nude mice. Like normal hepatocytes, AML cells express high levels of mRNA for serum (albumin, alpha 1-antitrypsin, and transferrin) and gap junction (connexins 26 and 32) proteins, secrete albumin, and contain solely isozyme 5 of lactate dehydrogenase. After extensive passaging, AML12 cells continue to strongly coexpress hepatocyte connexin mRNAs but do not display nonparenchymal cell markers. Although mRNA levels for some serum proteins progressively fall, high expression in late AML12 cultures may be regained by passage in serum-free medium. The AML14 line loses expression of both differentiated markers and transgene mRNA with extended passaging, and hepatocytic traits are only partially restored by passage in serum-free medium. These differentiated, nontumorigenic cell lines should serve as models in which to study hepatocyte growth and differentiation. PMID:7904757

Wu, J C; Merlino, G; Fausto, N



Minocycline reduces intracerebral hemorrhage-induced brain injury  

PubMed Central

Objectives Microglial activation and thrombin formation contribute to brain injury after intracerebral hemorrhage. Tumor necrosis factor-alpha and interleukin-1beta are two major pro-inflammatory cytokines. The present study investigated if thrombin stimulates tumor necrosis factor-alpha and interleukin-1beta secretion in vitro and if microglial inhibition reduces intracerebral hemorrhage -induced brain injury in vivo. Methods There were two parts in this study. In the first part, cultured rat microglial cells were treated with vehicle, thrombin (10 U/ml), or thrombin plus minocycline (1 or 10 ?M), an inhibitor of microglia activation. Levels of tumor necrosis factor-alpha and interleukin-1beta in culture medium were measured by Enzyme-Linked ImmunoSorbent Assay at 24 hours after thrombin treatment. In the second part, rats had an intracerebral injection of 100-?l autologous whole blood. Rats received minocycline or vehicle treatment. Brain edema was measured at day 3 and brain atrophy was determined at day 28 after intracerebral hemorrhage. Results Thrombin receptors were expressed in cultured microglia cells, and tumor necrosis factor-alpha and interleukin-1beta levels in the culture medium were increased after thrombin treatment. Minocycline reduced thrombin-induced upregulation of tumor necrosis factor-alpha and interleukin-1beta. In vivo, minocycline reduced perihematomal brain edema, neurological deficits and brain atrophy. Discussion Thrombin stimulates microglia to release the pro-inflammatory cytokines, tumor necrosis factor-alpha and interleukin-1beta, and microglial inhibition with minocycline reduces brain injury after intracerebral hemorrhage suggesting a critical role of microglia activation in intracerebral hemorrhage -related brain injury.

Wu, Jimin; Yang, Shuxu; Xi, Guohua; Fu, Guosheng; Keep, Richard F; Hua, Ya



Similar mechanisms of action of defined polysaccharides and lipopolysaccharides: characterization of binding and tumor necrosis factor alpha induction.  

PubMed Central

Little has been reported about the effects of different polysaccharides on cytokine production from human monocytes. In this study, we show that several well-defined polysaccharides, including polymers with different sizes of beta 1-4-linked D-mannuronic acid (poly-M, high-M alginate, and M-blocks) and cellulose oxidized in the C-6 position, induced human monocytes to produce tumor necrosis factor alpha (TNF-alpha). Poly-M was the most efficient polysaccharide tested and, on a weight basis, was approximately as efficient as lipopolysaccharide (LPS) from Escherichia coli. TNF-alpha production was shown to depend strongly on the molecular weights of poly-M and high-M alginate, with maximal TNF-alpha production occurring at molecular weights above 50,000 and 200,000, respectively. G-blocks, alpha 1-4-linked L-guluronic acid polymers that did not induce cytokine production from monocytes, reduced the cytokine production induced by the beta 1-4-linked polyuronic acids and LPS. Furthermore, both G-blocks and LPS were found to inhibit the binding of poly-M to monocytes, as measured by flow cytometry. In addition, we found that the binding of LPS to monocytes was inhibited by G-blocks, M-blocks, and poly-M. Our results indicate that beta 1-4-linked polyuronic acids and LPS may stimulate monocytes to produce TNF-alpha by similar mechanisms and may bind to a common receptor.

Otterlei, M; Sundan, A; Skjak-Braek, G; Ryan, L; Smidsr?d, O; Espevik, T



Lactobacilli and Streptococci Induce Interleukin12 (IL12), IL18, and Gamma Interferon Production in Human Peripheral Blood Mononuclear Cells  

Microsoft Academic Search

Human peripheral blood mononuclear cells (PBMC) were stimulated with three nonpathogenic Lactobacillus strains and with one pathogenic Streptococcus pyogenes strain, and cytokine gene expression and protein pro- duction were analyzed. All bacteria strongly induced interleukin-1b (IL-1b), IL-6, and tumor necrosis factor alpha mRNA expression and protein production. S. pyogenes was the most potent inducer of secretion of IL-12 and gamma




Production of tumor necrosis factors alpha and beta by human mononuclear leukocytes stimulated with mitogens, bacteria, and malarial parasites.  

PubMed Central

Tumor necrosis factors alpha and beta (TNF-alpha and TNF-beta) are multifaceted polypeptide cytokines which may mediate some of the significant changes in cellular homeostasis which accompany the invasion of the mammalian host by viruses, bacteria, and parasites. Although it is well established that bacterial lipopolysaccharide is a potent inducer of TNF-alpha, there is still very little known of the types of agents which can trigger the production of TNFs in mononuclear leukocytes. Using an enzyme-linked immunosorbent assay for measuring TNF-alpha and TNF-beta, we examined the capacity of various T-lymphocyte and beta-lymphocyte mitogens as well as microbial components to stimulate production of these cytokines in culture. The mitogens phytohemagglutinin, concanavalin A, and pokeweed mitogen induced production of both TNF-alpha and TNF-beta, while whole-killed Staphylococcus aureus and Bordetella pertussis, like lipopolysaccharide, were potent inducers of TNF-alpha but failed to stimulate TNF-beta production. TNF-alpha production was detectable within 1 h after stimulation, while TNF-beta production was not detected until after 8 h of culture. The bacterial products tetanus toxoid, purified protein derivative, pertussis filamentous hemagglutinin, and pertussis toxin were all able to induce TNF-alpha and TNF-beta production. Disrupted (frozen-thawed) Plasmodium falciparum-infected erythrocytes were also potent inducers of TNF-alpha and TNF-beta. The results demonstrated that a wide variety of microbial components are inducers of TNF-alpha. Some may not only be more effective than lipopolysaccharide but can also induce TNF-beta production. Furthermore, evidence is presented showing that TNF-beta but not TNF-alpha production correlates with lymphoproliferation.

Ferrante, A; Staugas, R E; Rowan-Kelly, B; Bresatz, S; Kumaratilake, L M; Rzepczyk, C M; Adolf, G R



Hypoxic-induced stress protein expression in rat cardiac myocytes  

Microsoft Academic Search

Mammalian stress proteins can be induced in cells and tissues exposed to a variety of conditions including hyperthermia and diminished Oâ supply. The authors have previously shown that the expression of three stress proteins (71, 85, and 95 kDa) was induced in cardiac tissue from mice exposed to hypoxic conditions. The expression of mRNAs coding for the 85 and 95

G. Howard; T. E. Geoghegan



Synthetic Riboswitches That Induce Gene Expression in Diverse Bacterial Species? †  

PubMed Central

We developed a series of ligand-inducible riboswitches that control gene expression in diverse species of Gram-negative and Gram-positive bacteria, including human pathogens that have few or no previously reported inducible expression systems. We anticipate that these riboswitches will be useful tools for genetic studies in a wide range of bacteria.

Topp, Shana; Reynoso, Colleen M. K.; Seeliger, Jessica C.; Goldlust, Ian S.; Desai, Shawn K.; Murat, Dorothee; Shen, Aimee; Puri, Aaron W.; Komeili, Arash; Bertozzi, Carolyn R.; Scott, June R.; Gallivan, Justin P.



Alpha-quartz-induced chemokine expression by rat lung epithelial cells: effects of in vivo and in vitro particle exposure.  

PubMed Central

Chemokines are chemotactic cytokines that can play a key role in leukocyte recruitment to sites of tissue injury or infection. Previous studies have demonstrated that exposure to alpha-quartz as well as other noxious particles increases chemokine gene expression in rat lung, although the cells responsible for chemokine expression and the mechanisms underlying this response have remained unclear. The present studies demonstrate that exposure of rats to alpha-quartz induced expression of mRNA for the chemokine macrophage-inflammatory protein (MIP)-2 in epithelial cells lining the terminal bronchioles and alveolar ducts as well as macrophages and alveolar type II cells in the more distal lung. Treatment of rats with an anti-MIP-2 antiserum before alpha-quartz exposure markedly attenuated neutrophilic infiltration of the lungs demonstrating an important role for MIP-2 in alpha-quartz-induced pulmonary inflammation. In vitro exposure of primary cultures of rat alveolar type II cells or the rat alveolar type II cell line RLE-6TN to tumor necrosis factor-alpha, endotoxin, or alpha-quartz increased mRNA for MIP-2 as well as the structurally and functionally similar chemokine cytokine-induced neutrophil chemoattractant but not the chemokine MIP-1 alpha. The alpha-quartz-induced increase in epithelial MIP-2 mRNA resulted, at least in part, from increased gene transcription and was associated with the release of active MIP-2 protein. Induction of RLE-6TN MIP-2 and cytokine-induced neutrophil chemoattractant mRNA expression was not unique to alpha-quartz, being also increased by crocidolite asbestus fibers but not by titanium dioxide or MMVF-10 glass fibers. These findings indicate that epithelial cells contribute to chemokine expression in rat lung after exposure to alpha-quartz and potentially other noxious particles and suggest that alpha-quartz-activated MIP-2 expression in vivo results, at least in part, from a direct action of the particles on the lung epithelium. Images Figure 1 Figure 3 Figure 5 Figure 6 Figure 7 Figure 8

Driscoll, K. E.; Howard, B. W.; Carter, J. M.; Asquith, T.; Johnston, C.; Detilleux, P.; Kunkel, S. L.; Isfort, R. J.



A novel sesquiterpenoid dimer parviflorene F induces apoptosis by up-regulating the expression of TRAIL-R2 and a caspase-dependent mechanism.  


Parviflorene F (1), a novel sesquiterpenoid dimer isolated from Curcuma parviflora Wall, is a cytotoxic compound. In this study, we examined the mechanism of its cytotoxic effect in HeLa cells. Treatment with 1 enhanced the mRNA and protein expression of TRAIL-R2 (tumor necrosis factor alpha-related apoptosis inducing ligand receptor 2). Apoptosis was induced by 1 as revealed by the distribution of DNA and Annexin V/PI staining using flow cytometry. In addition, 1-induced apoptosis was inhibited by human recombinant TRAIL-R2/Fc chimera protein, TRAIL-neutralizing fusion protein. Also, we found that 1 induced the activation of caspase-8, caspase-9, and caspase-3, indicating that the cytotoxic effect of 1 is correlated with apoptosis by a caspase-dependent mechanism through TRAIL-R2. In addition, 1 enhanced TRAIL-induced cell death against HeLa and TRAIL-resistant DLD1 cells. Taken together, up-regulation of TRAIL-R2 by 1 may contribute to sensitization of TRAIL-induced cell death. PMID:18036820

Ohtsuki, Takashi; Tamaki, Mayu; Toume, Kazuhumi; Ishibashi, Masami



Inflammatory stimuli reduce survival of serotonergic neurons and induce neuronal expression of indoleamine 2,3-dioxygenase in rat dorsal raphe nucleus organotypic brain slices.  


Neuroinflammation results in dysregulation of serotonergic neurons in the dorsal raphe nucleus (doR) and is considered to play an important role in the pathophysiology of depression. The aim of the present study was to induce neuroinflammation in a simple doR brain slice model using lipopolysaccharide (LPS), interferon-gamma (IFN?), beta-amyloid???? or tumor necrosis factor-alpha and to explore the survival of serotonergic neurons and the expression of the tryptophan degrading enzyme indoleamine 2,3-dioxygenase (IDO). Administration of pro-inflammatory stimuli reduced survival of serotonergic neurons in doR slices and increased IDO expression. IFN? most potently induced IDO expression, which co-localized with neurons, including serotonergic neurons, but not with microglia or astrocytes. IFN? did not induce PI-positive staining in slices, but increased the average nuclei size of IDO-positive cells. The inflammation-induced decline did not return to control levels, when slices were withdrawn from inflammation, pointing to neurodegeneration. The growth factors BDNF or GDNF did not counteract the inflammation-induced decrease in serotonergic neurons, except for LPS-induced neuronal decline. The inflammation-induced effect was not blocked by the NMDA-receptor antagonist MK-801. Further LPS, but not IFN? increased inflammatory markers and microglia activity. In conclusion, our data show that a range of inflammatory stimuli decline serotonergic neurons in doR slices and upregulate IDO expression. The data suggest that IDO does not contribute to serotonergic decline, but may serve as a marker of neurodegeneration. Neuroinflammation may contribute to the development of depression. PMID:21501664

Hochstrasser, T; Ullrich, C; Sperner-Unterweger, B; Humpel, C



Molecular cloning and characterization of beluga whale (Delphinapterus leucas) interleukin-1beta and tumor necrosis factor-alpha.  

PubMed Central

Interleukin-1beta (IL-1beta) and tumor necrosis factor-alpha (TNF-alpha) are cytokines produced primarily by monocytes and macrophages with regulatory effects in inflammation and multiple aspects of the immune response. As yet, no molecular data have been reported for IL-1beta and TNF-alpha of the beluga whale. In this study, we cloned and determined the entire cDNA sequence encoding beluga whale IL-1beta and TNF-alpha. The genetic relationship of the cytokine sequences was then analyzed with those from several mammalian species, including the human and the pig. The homology of beluga whale IL-1beta nucleic acid and deduced amino acid sequences with those from these mammalian species ranged from 74.6 to 86.0% and 62.7 to 77.1%, respectively, whereas that of TNF-alpha varied from 79.3 to 90.8% and 75.3 to 87.7%, respectively. Phylogenetic analyses based on deduced amino acid sequences showed that the beluga whale IL-1beta and TNF-alpha were most closely related to those of the ruminant species (cattle, sheep, and deer). The beluga whale IL-1beta- and TNF-alpha-encoding sequences were thereafter successfully expressed in Escherichia coli as fusion proteins by using procaryotic expression vectors. The fusion proteins were used to produce beluga whale IL-1beta- and TNF-alpha-specific rabbit antisera. Images Figure 3. Figure 4. Figure 5.

Denis, F; Archambault, D



Transforming growth factor-alpha, epidermal growth factor receptor, and proliferating potential in benign and malignant gliomas.  


Surgical specimens from six benign and 16 malignant human gliomas were investigated immunohistochemically to correlate the degree of malignancy, the distribution of transforming growth factor-alpha (TGF-alpha) and epidermal growth factor (EGF) receptor, and the potential for cell proliferation using monoclonal antibodies to TGF-alpha, EGF receptor, and Ki-67. Fourteen (88%) of the malignant gliomas and one (17%) of the benign gliomas were found to be positive for TGF-alpha, and 14 (88%) of the malignant gliomas and two (33%) of the benign gliomas expressed EGF receptor. The proliferation index with Ki-67 was 18.8% +/- 8.1% (mean +/- standard deviation) in malignant gliomas and 1.9% +/- 1.8% in benign gliomas. In general, cells positive for EGF receptor and Ki-67 were randomly distributed throughout the tumor tissue, and cells positive for TGF-alpha tended to be clustered without obvious relationship to areas of necrosis or blood vessels. In some tumors, cells positive for TGF-alpha, EGF receptor, and Ki-67 were associated in a focal distribution. The more frequent expression of TGF-alpha and EGF receptor in the highly proliferative malignant gliomas is compatible with a role for TGF-alpha and EGF receptor in the induction or stimulation of malignant gliomas. PMID:2045927

Maruno, M; Kovach, J S; Kelly, P J; Yanagihara, T



RelB Modulation of I?B? Stability as a Mechanism of Transcription Suppression of Interleukin-1? (IL-1?), IL-1?, and Tumor Necrosis Factor Alpha in Fibroblasts†  

PubMed Central

Members of the NF-?B/RelB family of transcription factors play important roles in the regulation of inflammatory and immune responses. RelB, a member of this family, has been characterized as a transcription activator and is involved in the constitutive NF-?B activity in lymphoid tissues. However, in a previous study we observed an overexpression of chemokines in RelB-deficient fibroblasts. Here we show that RelB is an important transcription suppressor in fibroblasts which limits the expression of proinflammatory mediators and may exert its function by modulating the stability of I?B? protein. Fibroblasts from relb?/? mice overexpress interleukin-1? (IL-1?), IL-1?, and tumor necrosis factor alpha in response to lipopolysaccharide (LPS) stimulation. These cells have an augmented and prolonged LPS-inducible IKK activity and an accelerated degradation which results in a diminished level of I?B? protein, despite an upregulated I?B? mRNA expression. Consequently, NF-?B activity was augmented and postinduction repression of NF-?B activity was impaired in these cells. The increased ?B-binding activity and cytokine overexpression was suppressed by introducing RelB cDNA or a dominant negative I?B? into relb?/? fibroblasts. Our findings suggest a novel transcription suppression function of RelB in fibroblasts.

Xia, Yiyang; Chen, Shizhong; Wang, Yibin; Mackman, Nigel; Ku, George; Lo, David; Feng, Lili



Tumor necrosis factor-alpha promotes survival in methotrexate-exposed macrophages by an NF-kappaB-dependent pathway  

PubMed Central

Introduction Methotrexate (MTX) induces macrophage apoptosis in vitro, but there is not much evidence for increased synovial macrophage apoptosis in MTX-treated patients. Macrophage apoptosis is reported, however, during clinical response to anti-tumor necrosis factor-alpha (TNF-?) treatments. This implies that TNF-? promotes macrophage survival and suggests that TNF-? may protect against MTX-induced apoptosis. We, therefore, investigated this proposal and the macrophage signaling pathways underlying it. Methods Caspase-3 activity, annexin-V binding/7-aminoactinomycin D (7-AAD) exclusion and cell-cycle analysis were used to measure steps in apoptosis of primary murine macrophages and cells of the RAW264.7 macrophage cell line that had been exposed to clinically-relevant concentrations of MTX and TNF-?. Results MTX induces apoptosis in primary murine macrophages at concentrations as low as 100 nM in vitro. TNF-?, which has a context-dependent ability to increase or to suppress apoptosis, efficiently suppresses MTX-induced macrophage apoptosis. This depends on NF-?B signaling, initiated through TNF Receptor Type 1 ligation. Macrophage colony stimulating factor, the primary macrophage survival and differentiation factor, does not activate NF-?B or protect macrophages from MTX-induced apoptosis. A weak NF-?B activator, Receptor Activator of NF-?B Ligand (RANKL) is likewise ineffective. Blocking NF-?B in TNF-?-exposed macrophages allowed pro-apoptotic actions of TNF-? to dominate, even in the absence of MTX. MTX itself does not promote apoptosis through interference with NF-?B signaling. Conclusions These findings provide another mechanism by which TNF-? sustains macrophage numbers in inflamed tissue and identify a further point of clinical complementarity between MTX and anti-TNF-? treatments for rheumatoid arthritis.



Bacterial endotoxin induces IL20 expression in the glial cells  

Microsoft Academic Search

The regulatory mechanisms leading to IL-20 expression during infection have not been elucidated. In the present study, we found that bacterial lipopolysaccharide (LPS) induced IL-20 expression in the primary cultured glial cells and RAW264.7 macrophage cell line. Pretreatment with protein synthesis inhibitor puromycin or cycloheximide failed to inhibit the expression of IL-20, suggesting that the expression was not dependent on

Toru Hosoi; Sachiyo Wada; Sawako Suzuki; Yasunobu Okuma; Shizuo Akira; Tadashi Matsuda; Yasuyuki Nomura



Tumour necrosis factor-alpha and the failing heart--pathophysiology and therapeutic implications.  


Immune activation plays a significant role in the development and progression of chronic heart failure (CHF). Indeed, pro-inflammatory cytokines, especially tumour necrosis factor-alpha (TNFalpha) are activated in this condition and exert direct detrimental actions on the myocardium. Physiological dampeners of TNFalpha production, such as interleukin-10, catecholamines, cortisol, and others fail in the course of the disease. However, the outcomes of two large-scale clinical trials with etanercept and infliximab, which directly antagonise TNFalpha have been rather disappointing. Nevertheless, TNFalpha antagonism remains a major target of CHF therapy, although counterbalancing this cytokine alone may not be sufficient. PMID:14685702

von Haehling, Stephan; Jankowska, Ewa A; Anker, Stefan D



Inhibition of Secretion of Interleukin-1? and Tumor Necrosis Factor Alpha by the Ketolide Antibiotic Telithromycin  

PubMed Central

The antibiotic telithromycin was examined for its effect on secretion of interleukin-1? (IL-1?), IL-1?, IL-6, IL-10, and tumor necrosis factor alpha (TNF-?) by lipopolysaccharide (LPS)-stimulated monocytes of eight human donors. Secretion of each cytokine was significantly increased by LPS alone, whereas treatment with telithromycin significantly inhibited secretion of IL-1? and TNF-? but not secretion of IL-1?, IL-6, and IL-10. Telithromycin had immunomodulatory effects as a result of alteration of secretion of IL-1? and TNF-? by monocytes.

Araujo, Fausto G.; Slifer, Teri L.; Remington, Jack S.



Inducible Gene Expression in Mammals: Plants Add to the Menu  

NSDL National Science Digital Library

Achieving inducible gene expression in mammalian cells with inexpensive and nontoxic inducers is an ongoing quest. The plant hormone abscisic acid has now been added to the list of compounds that can be used for regulating transcription and controlling protein function by induced proximity. These advances may enable new clinical applications of proximity-induced systems, and they highlight the value of fundamental research in plant biology.

Sean R. Cutler (Department of Botany and Plant Sciences;Center for Plant Cell Biology REV)



Survival and proliferation of cells expressing caspase-uncleavable Poly(ADP-ribose) polymerase in response to death-inducing DNA damage by an alkylating agent.  


To determine whether caspase-3-induced cleavage of poly(ADP-ribose) polymerase (PARP), a DNA damage-sensitive enzyme, alters the balance between survival and death of the cells following DNA damage, we created stable cell lines that express either caspase-uncleavable mutant or wild type PARP in the background of PARP (-/-) fibroblasts. The survival and apoptotic responses of these cells were compared after exposure to N-methyl-N'-nitro-N-nitrosoguanidine (MNNG), a DNA-damaging agent that activates PARP, or to tumor necrosis factor-alpha, which causes apoptosis without initial DNA damage. In response to MNNG, the cells with caspase-uncleavable PARP were very resistant to loss of viability or induction of apoptosis. Most significantly, approximately 25% of these cells survived and retained clonogenicity at a level of DNA damage that eliminated the cells with wild type PARP or PARP (-/-) cells. Expression of caspase-uncleavable PARP could not protect the cells from death induced by tumor necrosis factor, although there was a slower progression of apoptotic events in these cells. Therefore, one of the functions for cleavage of PARP during apoptosis induced by alkylating agents is to prevent survival of the extensively damaged cells. PMID:10601269

Halappanavar, S S; Rhun, Y L; Mounir, S; Martins, L M; Huot, J; Earnshaw, W C; Shah, G M



Efficient, Repeated Adenovirus-Mediated Gene Transfer in Mice Lacking both Tumor Necrosis Factor Alpha and Lymphotoxin ?  

PubMed Central

The efficiency of adenovirus-mediated gene transfer is now well established. However, the cellular and the humoral immune responses triggered by vector injection lead to the rapid elimination of the transduced cells and preclude any efficient readministration. The present investigation focuses on the role of tumor necrosis factor alpha (TNF-?), a proinflammatory cytokine, and the related cytokine lymphotoxin ? (LT?), in mounting an immune reaction against recombinant adenovirus vectors. After gene transfer in the liver, mice genetically deficient for both cytokines (TNF-?/LT??/?), in comparison with normal mice, presented a weak acute-phase inflammatory reaction, a reduction in cellular infiltrates in the liver, and a severely impaired T-cell proliferative response to both Adenoviral and transgene product antigens. Moreover, we observed a strong reduction in the humoral response to the vector and the transgene product, with a drastic reduction of anti-adenovirus immunoglobulin A and G antibody isotypes. In addition, the reduction in antibody response observed in TNF-?/LT??/? and TNF-?/LT?+/? mice versus TNF-?/LT?+/+ mice links antibody levels to TNF-?/LT? gene dosage. Due to the absence of neutralizing antibodies, the TNF-?/LT? knockout mice successfully express a second gene transduced by a second vector injection. The discovery of the pivotal role played by TNF-? in controlling the antibody response against adenovirus will allow more efficient adenovirus-based strategies for gene therapy to be proposed.

Benihoud, Karim; Saggio, Isabella; Opolon, Paule; Salone, Barbara; Amiot, Franck; Connault, Elisabeth; Chianale, Colette; Dautry, Francois; Yeh, Patrice; Perricaudet, Michel



Promoter haplotype combinations of the platelet-derived growth factor alpha-receptor gene predispose to human neural tube defects.  


Neural tube defects (NTDs), including anencephaly and spina bifida, are multifactorial diseases that occur with an incidence of 1 in 300 births in the United Kingdom. Mouse models have indicated that deregulated expression of the gene encoding the platelet-derived growth factor alpha-receptor (Pdgfra) causes congenital NTDs (refs. 2-4), whereas mutant forms of Pax-1 that have been associated with NTDs cause deregulated activation of the human PDGFRA promoter. There is an increasing awareness that genetic polymorphisms may have an important role in the susceptibility for NTDs (ref. 6). Here we identify five different haplotypes in the human PDGFRA promoter, of which the two most abundant ones, designated H1 and H2 alpha, differ in at least six polymorphic sites. In a transient transfection assay in human bone cells, the five haplotypes differ strongly in their ability to enhance reporter gene activity. In a group of patients with sporadic spina bifida, haplotypes with low transcriptional activity, including H1, were under-represented, whereas those with high transcriptional activity, including H2 alpha, were over-represented. When testing for haplotype combinations, H1 homozygotes were fully absent from the group of sporadic patients, whereas H1/H2 alpha heterozygotes were over-represented in the groups of both sporadic and familial spina bifida patients, but strongly under-represented in unrelated controls. Our data indicate that specific combinations of naturally occurring PDGFRA promoter haplotypes strongly affect NTD genesis. PMID:11175793

Joosten, P H; Toepoel, M; Mariman, E C; Van Zoelen, E J



Linkage between obesity and a marker near the tumor necrosis factor-alpha locus in Pima Indians.  

PubMed Central

Because tumor necrosis factor-alpha (TNF-alpha) expression is increased in adipose tissue of both rodent models of obesity and obese humans, it has been considered as a candidate gene for obesity. Pima Indians were scored for genotypes at three polymorphic dinucleotide repeat loci (markers) near the gene TNF-alpha at 6p21.3. In a sib-pair linkage analysis, percent body fat, as measured by hydrostatic weighing, was linked (304 sib-pairs, P = 0.002) to the marker closest (10 kb) to TNF-alpha. The same marker was associated (P = 0.01) by analysis of variance with BMI. To search for possible DNA variants in TNF-alpha that contribute to obesity, single stranded conformational polymorphism analysis was performed from 20 obese and 20 lean subjects. Primer pairs were designed for the entire TNF-alpha protein coding region and part of the promoter. Only a single polymorphism located in the promoter region was detected. No association could be demonstrated between alleles at this polymorphism and percent body fat. We conclude that the linkage of TNF-alpha to obesity might be due to a sequence variant undetected in TNF-alpha or due to a variant in some other closely linked gene.

Norman, R A; Bogardus, C; Ravussin, E



Transforming growth factor alpha treatment alters intracellular calcium levels in hair cells and protects them from ototoxic damage in vitro.  


To determine if transforming growth factor alpha (TGF alpha) pretreatment protects hair cells from aminoglycoside induced injury by modifying their intracellular calcium concentration, we assayed hair cell calcium levels in organ of Corti explants both before and after aminoglycoside (i.e. neomycin, 10(-3) M) exposure either with or without growth factor pretreatment. After TGF alpha (500 ng/ml) treatment, the intracellular calcium level of hair cells showed a five-fold increase as compared to the levels observed in the hair cells of control cultures. After ototoxin exposure, calcium levels in hair cells of control explants showed an increase relative to their baseline levels, while in the presence of growth factors pretreatment, hair cells showed a relative reduction in calcium levels. Pretreatment of organ of Corti explants afforded significant protection of hair cell stereocilia bundle morphology from ototoxic damage when compared to explants exposed to ototoxin alone. This study correlates a rise in hair cell calcium levels with the otoprotection of hair cells by TGF alpha in organ of Corti explants. PMID:9263032

Staecker, H; Dazert, S; Malgrange, B; Lefebvre, P P; Ryan, A F; Van de Water, T R



Respiratory syncytial virus replication in human lung epithelial cells: inhibition by tumor necrosis factor alpha and interferon beta.  


Respiratory syncytial virus (RSV) is the major pathogen causing severe lung disease in children. RSV initially replicates efficiently in the respiratory tract but becomes undetectable by 7 to 21 d after infection in normal children, suggesting that intrinsic cellular mechanisms, as yet undefined, may restrict virus replication. To provide an in vitro model to examine mechanisms that restrict RSV replication, three human lung epithelial cell lines were exposed to RSV in vitro and virus replication proceeded in a dose- and time-dependent manner, although less efficiently than the highly permissive CV-1 cell line (monkey kidney epithelial cell). Tumor necrosis factor alpha (TNF alpha) and/or interferon beta (IFN beta) markedly inhibited RSV replication in a dose- and time-dependent manner. TNF alpha combined with IFN beta essentially aborted RSV replication in A549 epithelial cells. TNF alpha and/or IFN beta did not induce cell membrane damage, cause cell lysis, or inhibit cellular protein synthesis. RSV-infected human alveolar macrophages, which produce TNF alpha, failed to productively infect lung epithelial cells in co-culture. Together these studies suggest that endogenous TNF alpha coupled with exogenous IFN beta could restrict RSV replication in lung epithelium. PMID:7551395

Merolla, R; Rebert, N A; Tsiviste, P T; Hoffmann, S P; Panuska, J R



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

PubMed Central

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.

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



Sensitive Immunoassay of a Biomarker Tumor Necrosis Factor-[alpha] Based on Poly(guanine)-Functionalized Silica Nanoparticle Label  

SciTech Connect

A novel electrochemical immunosensor for the detection of tumor necrosis factor-alpha (TNF-a) based on poly(guanine)-functionalized silica nanoparticles (NPs) label is presented. The detection of mouse TNF-a via immunological reaction is based on a dual amplification: 1) a large amount of guanine residues is introduced on the electrode surface through the silica nanoparticle and immunoreaction, 2) mediator-induced catalytic oxidation of guanine, which results in great enhancement of anodic current. The synthesized silica NP conjugates were characterized with atomic force microscopy, X-ray photoelectron spectroscopy, and electrochemistry. These experiments confirmed that poly[G] and avidin were immobilized on the surface of silica NPs. The performance of the electrochemical immunosensor was evaluated and some experiment parameters (e.g., concentration of Ru(bpy)32+, incubation time of TNF-a, etc.) were optimized. The detection of limit for TNF-a is found to be 5.0x10-11 g mL-1 (2.0 pM), which corresponds to 60 attomoles TNF-a in 30 uL. This immunosensor based on the poly[G] functionalized silica NP label offers great promise for rapid, simple, cost-effective analysis of biological samples.

Wang, Jun; Liu, Guodong; Engelhard, Mark H.; Lin, Yuehe



Differential induction of phosphatidylcholine hydrolysis, diacylglycerol formation and protein kinase C activation by epidermal growth factor and transforming growth factor-alpha in normal human skin fibroblasts and keratinocytes.  

PubMed Central

We have investigated coupling between the epidermal growth factor (EGF) receptor and the phospholipase C (PLC)/protein kinase C (PKC) signal-transduction system in normal skin fibroblasts and keratinocytes, for which EGF and transforming growth factor alpha (TGF-alpha) are mitogenic. EGF and TGF-alpha induced a rapid increase in tyrosine phosphorylation of the EGF receptor, in both fibroblasts and keratinocytes, but failed to induce tyrosine phosphorylation of PLC-gamma 1 or detectable phosphoinositide hydrolysis, as measured by two sensitive assays. In fibroblasts, EGF induced phosphatidylcholine (PC) hydrolysis, resulting in increased diacylglycerol (DAG). In contrast, in keratinocytes, there was no detectable PC hydrolysis or elevation of DAG in response to EGF or TGF-alpha. EGF and TGF-alpha activated PKC in fibroblasts, as evidenced by increased phosphorylation of a specific cellular PKC substrate (myristoylated alanine-rich C-kinase substrate, 'MARCKS'). In keratinocytes, TGF-alpha and EGF induced only a modest increase in MARCKS protein phosphorylation. This apparent modest activation of PKC, in the absence of detectable DAG formation, may have been mediated by arachidonic acid, which was released from keratinocytes in response to TGF-alpha, and has been shown to stimulate PKC activity in vitro. These data demonstrate that (1) in dermal fibroblasts and keratinocytes, which express normal levels of EGF receptors, EGF receptor activation is not coupled to tyrosine phosphorylation of PLC-gamma 1 or PtdIns hydrolysis, suggesting that these events are not required for the mitogenic activity of EGF or TGF-alpha in these cells, (2) coupling of EGF receptor to PC hydrolysis is cell-type specific, and (3) in skin fibroblasts, DAG, formed through EGF-induced PC hydrolysis, is capable of activating PKC. Images Figure 3 Figure 4 Figure 6

Reynolds, N J; Talwar, H S; Baldassare, J J; Henderson, P A; Elder, J T; Voorhees, J J; Fisher, G J



Inactivated Parapoxvirus ovis induces a transient increase in the expression of proinflammatory, Th1-related, and autoregulatory cytokines in mice  

PubMed Central

The immunostimulatory properties of inactivated Parapoxvirus ovis (iPPVO) have long been investigated in different animal species and experimental settings. In this study, we investigated the effects of iPPVO on cytokine expression in mice after intraperitoneal inoculation. Spleen and sera collected from iPPVO-treated mice at intervals after inoculation were submitted to cytokine mRNA determination by real-time PCR (qPCR), serum protein concentration by ELISA, and interferon (IFN)-?/? activity by bioassay. The spleen of iPPVO-treated animals showed a significant increase in mRNA expression of all cytokines assayed, with different kinetics and magnitude. Proinflammatory cytokines interleukin (IL)-1?, tumor necrosis factor-alpha (TNF-?), and IL-8 mRNA peaked at 24 hours postinoculation (hpi; 5.4-fold increase) and 48 hpi (3- and 10-fold increases), respectively. A 15-fold increase in IFN-? and 6-fold IL-12 mRNA increase were detected at 48 and 24 hpi, respectively. Increased expression of autoregulatory cytokines (Th2), mainly IL-10 and IL-4, could be detected at later times (72 and 96 hpi) with peaks of 4.7- and 4.9-fold increases, respectively. IFN-I antiviral activity against encephalomyocarditis virus was demonstrated in sera of treated animals between 6 and 12 hpi, with a >90% reduction in the number of plaques. Measurement of serum proteins by ELISA revealed increased levels of IL-1, TNF-?, IL-12, IFN-?, and IL-10, with kinetics similar to those observed by qPCR, especially for IL-12 and IFN-?. These data demonstrate that iPPVO induced a transient and complex cytokine response, initially represented by Th1-related cytokines followed by autoregulatory and Th2 cytokines.

Anziliero, D.; Weiblen, R.; Kreutz, L.C.; Spilki, F.; Flores, E.F.



Induction of macrophage-mediated production of tumor necrosis factor alpha by an L-form derived from Staphylococcus aureus.  

PubMed Central

We investigated the capability of an L-form derived from Staphylococcus aureus to induce tumor necrosis factor alpha (TNF-alpha) production in murine peritoneal macrophages. The activity for TNF-alpha induction was found in the membrane fraction of the L-form but not in the cytoplasmal fraction purified by the sucrose step gradient centrifugation. TNF-alpha mRNA was also detected in macrophages stimulated with L-form membranes. L-form induced TNF-alpha production in macrophages from both lipopolysaccharide-responsive and -unresponsive mouse strains. Regardless of the presence of polymyxin B, the activity of TNF-alpha induction of L-form was mostly found in the phenol layer, but not in the aqueous layer, both of which were prepared by phenol extraction method. Fractions of L-form membranes representing molecular masses of approximately between 29 and 36 kDa were primarily responsible for inducing the production of TNF-alpha consistently. Moreover, this stimulatory effect was abolished by digestion with Streptomyces griseus protease. In Western blot (immunoblot) analysis with anti-lipoteichoic acid antibody, two bands (65 and 45 kDa) were observed in the sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the phenol layer, whereas one band (14 kDa) was observed in either the aqueous layer or lipoteichoic acid of S. aureus. These results suggest that the component in the membrane of the L-form, distinct from cell wall components such as teichoic acid or lipopolysaccharide, possesses the capability to stimulate TNF-alpha production by macrophages. Images

Kuwano, K; Akashi, A; Matsu-ura, I; Nishimoto, M; Arai, S



Hantavirus Infection Induces the Expression of RANTES and IP-10 without Causing Increased Permeability in Human Lung Microvascular Endothelial Cells  

PubMed Central

Sin Nombre virus (SNV) and Hantaan virus (HTN) infect endothelial cells and are associated with different patterns of increased vascular permeability during human disease. It is thought that such patterns of increased vascular permeability are a consequence of endothelial activation and subsequent dysfunction mediated by differential immune responses to hantavirus infection. In this study, the ability of hantavirus to directly induce activation of human lung microvascular endothelial cells (HMVEC-Ls) was examined. No virus-specific modulation in the constitutive or cytokine-induced expression of cellular adhesion molecules (CD40, CD54, CD61, CD62E, CD62P, CD106, and major histocompatibility complex classes I and II) or in cytokines and chemokines (eotaxin, tumor necrosis factor alpha, interleukin 1? [IL-1?], IL-6, IL-8, MCP-1, MIP-1?, and MIP-1?) was detected at either the protein or message level in hantavirus-infected HMVEC-Ls. Furthermore, no virus-specific enhancement of paracellular or transcellular permeability or changes in the organization and distribution of endothelial intercellular junctional proteins was observed. However, infection with either HTN or SNV resulted in detectable levels of the chemokines RANTES and IP-10 (the 10-kDa interferon-inducible protein) in HMVEC-Ls within 72 h and was associated with nuclear translocation of interferon regulatory factor 3 (IRF-3) and IRF-7. Gamma interferon (IFN-?)-induced expression of RANTES and IP-10 could also be detected in uninfected HMVEC-Ls and was associated with nuclear translocation of IRF-1 and IRF-3. Treatment of hantavirus-infected HMVEC-Ls with IFN-? for 24 h resulted in a synergistic enhancement in the expression of both RANTES and IP-10 and was associated with nuclear translocation of IRF-1, IRF-3, IRF-7, and NF-?B p65. These results reveal a possible mechanism by which hantavirus infection and a TH1 immune response can cooperate to synergistically enhance chemokine expression by HMVEC-Ls and trigger immune-mediated increases in vascular permeability.

Sundstrom, J. Bruce; McMullan, Laura K.; Spiropoulou, Christina F.; Hooper, W. Craig; Ansari, Aftab A.; Peters, Clarence J.; Rollin, Pierre E.



Salmonella induces prominent gene expression in the rat colon  

Microsoft Academic Search

BACKGROUND: Salmonella enteritidis is suggested to translocate in the small intestine. In vivo it induces gene expression changes in the ileal mucosa and Peyer's patches. Stimulation of Salmonella translocation by dietary prebiotics fermented in colon suggests involvement of the colon as well. However, effects of Salmonella on colonic gene expression in vivo are largely unknown. We aimed to characterize time

Wendy Rodenburg; Jaap Keijer; Evelien Kramer; Susanne Roosing; Carolien Vink; Martijn B Katan; Roelof van der Meer; Ingeborg MJ Bovee-Oudenhoven



Vocalization Induced CFos Expression in Marmoset Cortex  

PubMed Central

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.

Miller, Cory T.; DiMauro, Audrey; Pistorio, Ashley; Hendry, Stewart; Wang, Xiaoqin



Induction of interleukin (IL)-1beta and IL-8 mRNA expression in porcine macrophages by lipopolysaccharide from Serpulina hyodysenteriae.  

PubMed Central

Lipopolysaccharide (LPS) is a classic inducer of inflammatory cytokines and is a key virulence factor for most gram-negative pathogens. The effect of phenol-water (LPS) and butanol-water (endotoxin) extracts from Serpulina hyodysenteriae on inflammatory cytokine mRNA expression from porcine alveolar macrophages was investigated. The LPS and endotoxin extracts from S. hyodysenteriae induced a dose-dependent expression of interleukin 1beta (IL-1beta) and IL-8 which was weak compared with the responses induced by Escherichia coli LPS. In addition, the spirochetal extracts induced no detectable upregulation of mRNA expression for either IL-6 or tumor necrosis factor alpha.

Sacco, R E; Nibbelink, S K; Baarsch, M J; Murtaugh, M P; Wannemuehler, M J



Amlodipine inhibits pro-inflammatory cytokines and free radical production and inducible nitric oxide synthase expression in lipopolysaccharide/interferon-gamma-stimulated cultured vascular smooth muscle cells.  


Overproduction of nitric oxide (NO) from inducible nitric oxide synthase (iNOS) is importantly involved in the pathogenesis of endotoxemia and atherosclerosis. Calcium antagonists are commonly used as cardiovascular drugs and have a beneficial effect on prolonging survival in various models of endotoxin shock. The present study was to investigate the effect of a calcium antagonist amlodipine on nitrite, tumor necrosis factor-alpha (TNF-alpha) and interleukin-1beta (IL-1beta) formation and iNOS induction both in lipopolysaccharide (LPS) and interferon-gamma (IFN-gamma)-treated rat aortic smooth muscle cells (RASMC) and in a rat model of endotoxemia. Incubation with amlodipine (0.1 - 10 microM) for 24 h resulted in a significant and dose-dependent attenuation in medium nitrite, TNF-alpha and IL-1beta formation as well as iNOS protein expression in LPS/IFN-gamma-treated RASMC. In addition, amlodipine inhibited leucigenin-induced superoxide formation in RASMC. In the rat endotoxic model, the serum nitrite/nitrate, TNF-alpha and IL-1beta levels as well as iNOS protein expression of lungs were also suppressed by administration of amlodipine (50 microg/kg, i.v.). These results suggest that amlodipine may exert vascular beneficial effects by suppressing pro-inflammatory cytokines and free radical generation as well as iNOS induction in smooth muscle cells during activation of inflammatory mechanism. PMID:12120758

Chou, Tz-Chong; Yang, Shih-Ping; Pei, Dee



Production of Rantes/CCL5 in human gingival fibroblasts challenged with tumor necrosis factor alpha.  


Chemokines are small-secreted proteins that stimulate the directional migration of leukocytes and thereby mediate the inflammatory process. The present study investigates the capacity of human gingival fibroblasts to produce the beta chemokine Rantes/CCL5. In situ hybridization, immunohistochemistry and ELISA were used to measure the induction of Rantes/CCL5 at the mRNA and protein levels, both in unstimulated gingival fibroblasts as well as in fibroblasts treated with the proinflammatory cytokines tumor necrosis factor (TNF)alpha or interleukin (IL)-1beta. TNFalpha in different concentrations (0.1-10 ng/ml) induced Rantes/CCL5 mRNA expression and protein production in 24-h cultures of human gingival fibroblasts. The expression of Rantes/CCL5-mRNA and protein production, induced by TNFalpha, was evident at 6 h and thereafter increased continuously during the study period (24 h). IL-1beta (3-300 pg/ml) also enhanced the production of Rantes/CCL5 in gingival fibroblasts. The amount of Rantes/CCL5 induced by IL-1beta (300 pg/ml), however, was less than that induced by TNFalpha (10 ng/ml). The study suggests that human gingival fibroblasts, by producing the chemokine Rantes/CCL5, participate in the regulation of the host response during the inflammatory process in the periodontal tissue. PMID:11330934

Mustafa, M; Wondimu, B; Bakhiet, M; Modéer, T



CYP3A induction by N-hydroxyformamide tumor necrosis factor-alpha converting enzyme/matrix metalloproteinase inhibitors use of a pregname X receptor activation assay and primary hepatocyte culture for assessing induction potential in humans.  


A series of N-hydroxyformamide tumor necrosis factor-alpha converting enzyme (TACE)/matrix metalloprotease (MMP) inhibitors were evaluated for their potential to induce human cytochrome P450 3A (CYP3A). Two in vitro assays were used: 1) a cell-based reporter gene assay for activation of the pregnane X receptor (PXR), and 2) a primary "sandwich" culture of human hepatocytes. Approximately 50 TACE/MMP inhibitors were evaluated in the human PXR assay. A range of PXR activation was observed, 0 to 150% of the activation of the known human CYP3A inducer rifampicin. Three TACE/MMP inhibitors were evaluated in rat and human hepatocytes. Significantly higher PXR activation/CYP3A induction was observed in PXR/hepatocyte models, respectively, for (2R,3S) 3-(formyl-hydroxyamino)-2-(2-methyl-1-propyl)-4-methylpentanoic acid [(1S,2S)-2-methyl-1-(2-pyridylcarbamoyl)-1-butyl]amide (GW3333) compared with (2R,3S)-6,6,6-trifluoro-3-[formyl(hydroxy)amino]-2-isobutyl-N-[(1S,2R)-2-methoxy-1-[(1,3-thiazol-2-ylamino)carbonyl]propyl]hexanamide (GW6495) and (2R)-N-[(1S)-2,2-dimethyl-1-[(methylamino)carbonyl]-propyl]-2-[(1S)-1-[formyl(hydroxy)amino]ethyl]-5-phenylpentanamide (GI4023). The CYP3A induction level achieved with GW3333 at a concentration of approximately 10 microM in human hepatocytes was comparable to that achieved with rifampicin at a concentration of 10 microM. The extent of rodent CYP3A induction caused by GW3333 was confirmed in vivo after daily oral administration for 14 days to rats. In conclusion, GW3333 is a potential inducer of CYP3A expression in vivo in humans, but other N-hydroxyformamides are less likely to induce CYP3A. PMID:12814963

Tippin, Timothy K; Hamilton, Geraldine; Moore, Linda; Beaudet, Elizabeth J; Jolley, Summer; Brodie, Thomas A; Andrews, Robert C; Becherer, J David; McDougald, Darryl L; Gaul, Michael D; Hoivik, Debie J; Mellon-Kusibab, Kathy; Lehmann, Jurgen; Kliewer, Steven; Novick, Steven; Laethem, Ron; Zhao, Zhiyang; LeCluyse, Edward L



Medicinal flowers. XXVII. New flavanone and chalcone glycosides, arenariumosides I, II, III, and IV, and tumor necrosis factor-alpha inhibitors from everlasting, flowers of Helichrysum arenarium.  


The methanolic extract from the flowers of Helichrysum arenarium L. MOENCH was found to show inhibitory effect on tumor necrosis factor-alpha (TNF-alpha, 1 ng/ml)-induced cytotoxicity in L929 cells. From the methanolic extract, 50 constituents including four new flavanone and chalcone glycosides named arenariumosides I (1), II (2), III (3), and IV (4) were isolated. The stereostructures of 1-4 were elucidated on the basis of chemical and physicochemical evidence. Among the constituents, naringenin 7-O-beta-D-glucopyranoside (7), apigenin 7-O-beta-D-glucopyranoside (14), apigenin 7-O-gentiobioside (16), and apigenin 7,4'-di-O-beta-D-glucopyranoside (17) significantly inhibited TNF-alpha-induced cytotoxicity in L929 cells at 30 microM. PMID:19336929

Morikawa, Toshio; Wang, Li-Bo; Nakamura, Seikou; Ninomiya, Kiyofumi; Yokoyama, Eri; Matsuda, Hisashi; Muraoka, Osamu; Wu, Li-Jun; Yoshikawa, Masayuki



Gomisin A decreases the LPS-induced expression of iNOS and COX-2 and activation of RIP2/NF-?B in mouse peritoneal macrophages.  


Abstract Gomisin A (GA), a lignan component contained in the fruit of Schisandra chinensis Baillon, improves hepatic cell degeneration, vasodilatory activity and insulin sensitivity. These effects also impact the immune system, including various inflammatory mediators and cytokines. In this study, the anti-inflammatory effect of GA on lipopolysaccharide-stimulated mouse peritoneal macrophages was studied. Pretreatment with GA attenuated the expression of receptor-interacting protein 2 (RIP2) and I?B kinase-? (IKK-?) as well as IKK-? phosphorylation. The activation of nuclear factor-kappa B (NF-?B) in the nucleus, the phosphorylation of I?B? and degradation of I?B? in the cytosol were suppressed by GA. GA decreased the production and mRNA expression of the inflammatory cytokines tumor necrosis factor-alpha (TNF-?) and interleukin (IL)-6. In addition, expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) and production of nitric oxide were decreased by pretreatment with GA. In conclusion, these results show that the anti-inflammatory properties of GA potentially result from the inhibition of COX-2, iNOS, IL-6, TNF-? and NO through the down-regulation of RIP2 and NF-?B activation. These results impact the development of potential health products for preventing and treating inflammatory diseases. PMID:24749675

Jeong, Hyun-Ja; Han, Na-Ra; Kim, Kyu-Yeob; Choi, Il-Sook; Kim, Hyung-Min



Express yourself: bold individuals induce enhanced morphological defences.  


Organisms display an impressive array of defence strategies in nature. Inducible defences (changes in morphology and/or behaviour within a prey's lifetime) allow prey to decrease vulnerability to predators and avoid unnecessary costs of expression. Many studies report considerable interindividual variation in the degree to which inducible defences are expressed, yet what underlies this variation is poorly understood. Here, we show that individuals differing in a key personality trait also differ in the magnitude of morphological defence expression. Crucian carp showing risky behaviours (bold individuals) expressed a significantly greater morphological defence response when exposed to a natural enemy when compared with shy individuals. Furthermore, we show that fish of different personality types differ in their behavioural plasticity, with shy fish exhibiting greater absolute plasticity than bold fish. Our data suggest that individuals with bold personalities may be able to compensate for their risk-prone behavioural type by expressing enhanced morphological defences. PMID:24335987

Hulthén, Kaj; Chapman, Ben B; Nilsson, P Anders; Hollander, Johan; Brönmark, Christer



Sequential Analysis of Gene Expression after an Osteogenic Stimulus - c- fos Expression Is Induced in Osteocytes  

Microsoft Academic Search

We have recently developed an experimental model whereby mechanical stimulation induces osteogenesis in the caudal vertebrae of rats. We used this model to assess expression of genes induced by mechanical loading. Bulk preparations of mRNA extracted after loading did not-show >2-fold increases in expression of mRNA for matrix proteins or growth factors in Northern blotting analysis. c-jun was undetectable. However,

T. Inaoka; J. M. Lean; T. Bessho; J. W. M. Chow; A. Mackay; T. Kokubo; T. J. Chambers



The expression of TIPE1 in murine tissues and human cell lines  

Microsoft Academic Search

Members of the tumor necrosis factor-alpha-induced protein-8 (TNFAIP8 or TIPE) family play important roles in immune homeostasis and cancer. TIPE1 (TNFAIP8-like 1) is a new member of the TIPE family that may regulate cell death. However, due to the lack of a suitable antibody, the nature of cells and tissues that express TIPE1 protein has not been determined. In this

Jian Cui; Guizhong Zhang; Chunyan Hao; Yan Wang; Yunwei Lou; Wenqian Zhang; Juan Wang; Suxia Liu



Bilateral diaphragmatic paralysis associated with the use of the tumor necrosis factor-alpha inhibitor adalimumab  

PubMed Central

A 51-year-old woman was referred for evaluation of progressive dyspnea of 3 months— duration. She had received 3 doses of adalimumab for treatment of rheumatoid arthritis prior to the onset of her dyspnea. Her chest examination revealed absent diaphragmatic movement with inspiration. Spirometry showed a severe restrictive defect. Radiologic studies confirmed the diagnosis of bilateral diaphragmatic paralysis. Laboratory and radiologic workup excluded other possible causes of the diagnosis. Adalimumab was discontinued, and she was treated with bilevel positive airway pressure ventilation and intravenous immunoglobulin. Three months later, the diaphragmatic paralysis persisted. This is the second reported case of bilateral diaphragmatic paralysis occurring in a patient who had received adalimumab. Acute neuropathies are rare side effects of tumor necrosis factor-alpha inhibitors.

Martin, Alan William; Rosenblatt, Randall Lee



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

PubMed Central

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-?) 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-? monoclonal antibody, infliximab (Remicade®).

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



Immunoelectron microscopic localisation of transforming growth factor alpha in rat colon.  

PubMed Central

Transforming growth factor alpha (TGF alpha) is a polypeptide, which binds to the epidermal growth factor receptor to carry out its function related to cell proliferation and differentiation. The ultrastructural localisation of TGF alpha was studied in both the proximal and the distal colon. The columnar cells, lining the surface epithelium of the proximal colon, showed a strong immunoreactivity in the polyribosomes and in the interdigitations of the lateral membrane. The columnar cells of the crypts and the goblet cells in both the proximal and the distal colon showed the immunostaining in the cis and trans cisternae of the Golgi apparatus. TGF alpha seems to be processed differently in the surface columnar cells and in the crypt columnar cells and goblet cells. Moreover, it probably has different roles in proliferation and differentiation. Images Figure 1 Figure 2 Figure 3 Figure 4 Figure 5 Figure 6 Figure 7

Perez-Tomas, R; Cullere, X; Asbert, M; Diaz-Ruiz, C



Role of tumor necrosis factor alpha in pathogenesis of pneumococcal pneumonia in mice.  

PubMed Central

The production and role of tumor necrosis factor alpha (TNF-alpha) in pneumococcal pneumonia were investigated in a mouse pneumonia model. When approximately 10(6) CFU of Streptococcus pneumoniae TUM19 were used to inoculate CBA/J mice intranasally, TNF-alpha levels in the lungs and serum began to increase from 1 and 3 days after infection, respectively, concomitantly with the increase in bacterial counts in the lungs. Anti-TNF-alpha antibody accelerated bacterial proliferation in the blood and the death of the mice. Although serum levels of immunoglobulin G antibody against the infecting bacteria were not affected by the anti-TNF-alpha antibody treatment, neutrophil counts in the blood were decreased by the treatment. These results suggest that TNF-alpha produced in the course of pneumococcal pneumonia prevents bacteremia by increasing the number of neutrophils in the blood.

Takashima, K; Tateda, K; Matsumoto, T; Iizawa, Y; Nakao, M; Yamaguchi, K



Detection of in vivo production of tumour necrosis factor-alpha by human thyroid epithelial cells.  

PubMed Central

We have established previously that human thyroid epithelial cells (TEC) from patients with autoimmune thyroiditis are able to synthesize cytokines, such as interleukin-1 (IL-1) and interleukin-6 (IL-6). This paper examines TEC in sections from autoimmune thyroiditis for the in vivo production of tumour necrosis factor-alpha (TNF-alpha) and interferon-gamma (IFN-gamma) using the combined techniques of in situ hybridization and immunohistochemistry. Thyroid tissue from patients with Graves' disease, Hashimoto's disease and non-toxic goitre was examined and both mRNA and the protein of TNF-alpha were detected in TEC on frozen sections. Representative figures of only Graves' samples are illustrated in this paper. In contrast, using the same methods, IFN-gamma was detected only in the infiltrating cells and not in TEC of thyroid tissue from the patients. Images Figure 1 Figure 2

Zheng, R Q; Abney, E R; Chu, C Q; Field, M; Maini, R N; Lamb, J R; Feldmann, M



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

SciTech Connect

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.

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



Suppression of Lefty expression in induced pluripotent cancer cells.  


Cancer and stem cells share the ability to silence tumor suppressors. We focused on Lefty, which encodes one of the most abundant tumor suppressors in embryonic stem (ES) cells and is not expressed in somatic cancer cells. We found that transforming growth factor ? (TGF-?) induced demethylation of the Lefty B cytosine-phosphate-guanine (CpG) island and increased Lefty expression (10-200 times) in human pancreatic cancer cells and human liver cancer cells (PLC/PRF/5 and HLF). Expression of Cripto, another important factor in Nodal-Lefty signaling, was not increased after adding TGF-?. We generated reprogrammed cancer cells that revealed high expression of immature marker proteins, high proliferation, and the potential to express morphological patterns of ectoderm, mesoderm, and endoderm, suggesting that these cells may have cancer stem cell-like phenotypes. We investigated Lefty and found that reprogrammed human liver cancer cells (induced pluripotent cancer cells) displayed a much lower ability to express Lefty, although less Lefty B CpG methylation was also observed. We also found that a MEK inhibitor dramatically enhanced Lefty expression in human pancreatic cancers with mutated ras, whereas Lefty B CpG methylation was not decreased. These observations indicate that despite the demethylation of DNA strands in promoter regions of pluripotency-associated genes, including Lefty gene, Lefty expression was not induced well in reprogrammed cells. Of note was the fact that Lefty is abundantly expressed in human ES cells but not in induced pluripotent stem (iPS) cells. We thus think that reprogrammed cancer cells share the mechanism for expression of Lefty with iPS cells. This shared mechanism may contribute to the cancerous transformation of iPS cells. PMID:23407711

Saito, Akiko; Ochiai, Hiromi; Okada, Shoko; Miyata, Naoteru; Azuma, Toshifumi



Adenosine inhibits tumor necrosis factor-alpha release from mouse peritoneal macrophages via A2A and A2B but not the A3 adenosine receptor.  


Adenosine is elaborated in injured tissues where it suppresses inflammatory responses of essentially all immune cells, including production of proinflammatory cytokines such as tumor necrosis factor-alpha (TNF-alpha). Most of the anti-inflammatory actions of adenosine have been attributed to signaling through the A(2A) adenosine receptor (A(2A)AR). Previously, however, it has been shown that the A(3)AR agonist N(6)-(3-iodobenzyl)adenosine-5'-N-methylcarboxamide (IB-MECA) potently inhibited TNF-alpha release from macrophages obtained from A(2A)AR "knockout" (A(2A)KO) mice, suggesting that the A(3)AR may also regulate cytokine expression. Here, we confirmed that the A(2A)AR is the primary AR subtype that suppresses TNF-alpha release from thioglycollate-elicited mouse peritoneal macrophages induced by both Toll-like receptor-dependent (TLR) and TLR-independent stimuli, but we determined that the A(2B)AR rather than the A(3)AR mediates the non-A(2A)AR actions of adenosine since 1) the ability of IB-MECA to inhibit TNF-alpha release was not altered in macrophages isolated from A(3)KO mice, and 2) the A(2B)AR antagonist 1,3-dipropyl-8-[4-[((4-cyanophenyl)carbamoylmethyl)oxy]phenyl]xanthine (MRS 1754) blocked the ability of the nonselective AR agonist adenosine-5'-N-ethylcarboxamide (NECA) to inhibit TNF-alpha release from macrophages isolated from A(2A)KO mice. Although A(2B)ARs seem capable of inhibiting TNF-alpha release, the A(2A)AR plays a dominant suppressive role since MRS 1754 did not block the ability of NECA to inhibit TNF-alpha release from macrophages isolated from wild-type (WT) mice. Furthermore, the potency and efficacy of adenosine to inhibit TNF-alpha release from WT macrophages were not influenced by blocking A(2B)ARs with MRS 1754. The data indicate that adenosine suppresses TNF-alpha release from macrophages primarily via A(2A)ARs, although the A(2B)AR seems to play an underlying inhibitory role that may contribute to the anti-inflammatory actions of adenosine under select circumstances. PMID:16339914

Kreckler, Laura M; Wan, Tina C; Ge, Zhi-Dong; Auchampach, John A



Tumour necrosis factor alpha activates nuclear factor kappaB signalling to reduce N-type voltage-gated Ca2+ current in postganglionic sympathetic neurons.  


Inflammation has profound effects on the innervation of affected tissues, including altered neuronal excitability and neurotransmitter release. As Ca(2+) influx through voltage-gated Ca(2+) channels (VGCCs) is a critical determinant of excitation-secretion coupling in nerve terminals, the aim of this study was to characterize the effect of overnight incubation in the inflammatory mediator tumour necrosis factor alpha (TNFalpha; 1 nM) on VGCCs in dissociated neurons from mouse superior mesenteric ganglia (SMG). Voltage-gated Ca(2+) currents (I(Ca)) were measured using the perforated patch clamp technique and the VGCC subtypes present in SMG neurons were estimated based on inhibition by selective VGCC blockers: omega-conotoxin GVIA (300 nM; N-type), nifedipine (10 microM; L-type), and omega-conotoxin MVIIC (300 nM; N-, P/Q-type). We used intracellular Ca(2+) imaging with Fura-2 AM to compare Ca(2+) influx during depolarizations in control and TNFalpha-treated neurons. TNF receptor and VGCC mRNA expression were measured using PCR, and channel alpha subunit (CaV2.2) was localized with immunohistochemistry. Incubation in TNFalpha significantly decreased I(Ca) amplitude and depolarization-induced Ca(2+) influx. The reduction in I(Ca) was limited to omega-conotoxin GVIA-sensitive N-type Ca(2+) channels. Depletion of glial cells by incubation in cytosine arabinoside (5 microM) did not affect I(Ca) inhibition by TNFalpha. Preincubation of neurons with SC-514 (20 microM) or BAY 11-7082 (1 microM), which both inhibit nuclear factor kappaB signalling, prevented the reduction in I(Ca) by TNFalpha. Inhibition of N-type VGCCs following TNFalpha incubation was associated with a decrease in CaV2.2 mRNA and reduced membrane localization of CaV2.2 immunoreactivity. These data suggest that TNFalpha inhibits I(Ca) in SMG neurons and identify a novel role for NF-kappaB in the regulation of neurotransmitter release during inflammatory conditions with elevated circulating TNFalpha, such as Crohn's disease and Guillain-Barré syndrome. PMID:19403618

Motagally, Mohamed A; Lukewich, Mark K; Chisholm, Susan P; Neshat, Shadia; Lomax, Alan E



Epidermal growth factor (EGF), transforming growth factor-alpha (TGF-alpha), and basic fibroblast growth factor (bFGF) differentially influence neural precursor cells of mouse embryonic mesencephalon.  


Growth factors are key elements in the process of neural cell differentiation. We examined the effects of classical mitogens on neural precursor cells, by culturing mouse cells of the embryonic (13.5 days postcoitum) mesencephalon and treating them with epidermal growth factor (EGF), transforming growth factor-alpha (TGF-alpha), basic fibroblast growth factor (bFGF), nerve growth factor (NGF), and transforming growth factor-beta (TGF-beta). Our initial results show that EGF, TGF-alpha, or bFGF, but not NGF or TGF-beta, induced general proliferation of the cultured cells, followed by formation of colonies. Combinations of these three growth factors suggest that most cells with the capacity to form colonies responded to EGF, TGF-alpha, or bFGF. The number of colonies increased significantly when EGF, but not TGF-alpha, was used in combination with bFGF. Furthermore, a population responding only to EGF + bFGF was detected in the dorsal mesencephalon. The colony-forming activity of bFGF was dependent on insulin, but bFGF and insulin cooperation was indirect since we could not observe colony formation in subcultures of cells derived from colonies, even in the presence of insulin. Cells obtained from our colonies displayed neuronal and glial morphology and expressed markers of both neurons and astrocytes; nestin, a marker of neural precursor cells, was also expressed in the majority of colonies. Growth factors also influenced neuronal maturation; the best neurite outgrowth was obtained from cells derived from bFGF-induced colonies cultured in the presence of EGF + bFGF. These data indicate the existence of neural precursor cells in the embryonic mesencephalon that respond differentially to growth factors. PMID:8568917

Santa-Olalla, J; Covarrubias, L



The anti-inflammatory activity of the polyphenol resveratrol may be partially related to inhibition of tumour necrosis factor-alpha (TNF-alpha) pre-mRNA splicing.  


The present study shows for the first time that the polyphenol resveratrol (RESV) blocks processing of tumour necrosis factor-alpha (TNF-alpha) pre-mRNA in mature mRNA. This study was carried out in turbot (Psetta maxima (L.)), a fish species that we are using to evaluate the effects of RESV on the inflammatory response in vertebrates. Treatment of turbot head kidney leucocytes with polysaccharides from the seaweed Ulva rigida (ulvan) resulted in an increase in TNF-alpha expression. RESV did not inhibit transcription but almost completely inhibited the production of mRNA in ulvan-induced cells and caused a notable increase in the level of unspliced TNF-alpha pre-mRNA. RESV also induced accumulation of IL-1beta pre-mRNA at the expense of mature mRNA, although the effects on IL-1beta were less evident than those on TNF-alpha. However, the housekeeping gene was not affected by RESV. We also evaluated the effects of RESV in vivo under an inflammatory stimulus and found an inhibitory effect on TNF-alpha and IL-1beta pre-mRNA splicing in turbot head kidney at 24 and 48h post-injection. In addition, RESV also reduced migration of cells to the peritoneal cavity under the same inflammatory stimulus. The results show that this fish species may be a useful model for analysing the effects of RESV on TNF-alpha and IL-1beta expression, and suggest that RESV could be used to decrease the levels of pro-inflammatory cytokines in vivo and to reduce inflammatory reactions in certain inflammatory diseases. PMID:19945165

Leiro, José M; Varela, Monica; Piazzon, M Carla; Arranz, Juan A; Noya, Manuel; Lamas, Jesus



Influence of ?S-globin haplotypes and hydroxyurea on tumor necrosis factor-alpha levels in sickle cell anemia  

PubMed Central

Background: Sickle cell anemia is a chronic inflammatory disease characterized by an increased production of proinflammatory cytokines including tumor necrosis factor-alpha. Hydroxyurea, by decreasing the polymerization of hemoglobin, reduces inflammatory states. The effect of the genetic polymorphisms of sickle cell patients on tumor necrosis factor-alpha levels remains unknown. Objective: The aim of this study was to investigate the association of tumor necrosis factor-alpha levels with ?-globin haplotypes and the use of hydroxyurea. Methods: A cross-sectional study was performed of 67 patients with sickle cell anemia diagnosed at steady-state in a referral hospital in Fortaleza, Ceará, Brazil. A group of 26 healthy individuals was used as control. ?S-haplotype analysis was performed by restriction fragment length polymorphism-polymerase chain reaction. The tumor necrosis factor-alpha levels were measured by the enzyme-linked immunosorbent assay test. Laboratory data (complete blood count and fetal hemoglobin) and information regarding the use of hydroxyurea were obtained from medical records. Statistical analysis was performed using R software with the Kruskal-Wallis and Mann-Whitney tests. Statistical significance was established for p-values < 0.05 for all analyses. Results: The mean age of the participants was 35.48 years. Patients with sickle cell anemia had significantly higher tumor necrosis factor-alpha levels than controls (p-values < 0.0001). Tumor necrosis factor-alpha levels were lower in sickle cell anemia patients who were receiving hydroxyurea treatment than those who were not (p-value = 0.1249). Sickle cell anemia patients with Bantu/n genotype had significantly higher levels than patients with the Bantu/Benin genotype (p-value = 0.0021). Conclusion: In summary, ?S-globin haplotypes, but not hydroxyurea therapy, have a role in modulating tumor necrosis factor-alpha levels in sickle cell anemia adults at steady-state. Many previous studies have investigated prognosis and inflammatory states in sickle cell anemia patients, but the discovery that tumor necrosis factor-alpha levels vary according to the genetic polymorphism of the patient is a new finding.

Laurentino, Marilia Rocha; Maia, Pedro Aurio; Barbosa, Maritza Cavalcante; Bandeira, Izabel Cristina Justino; Rocha, Lilianne Brito da Silva; Goncalves, Romelia Pinheiro



Construction of a doxycycline inducible adipogenic lentiviral expression system  

PubMed Central

To provide a tool for research on regulating adipocyte differentiation, tetracycline inducible (Tet on) lentiviral expression vectors under the control of an adipose-specific promoter were constructed. The lowest basal expression in the absence of doxycycline and most efficient dose-dependent, doxycycline-induced transient overexpression was observed using vectors constructed with a combination of Tetracycline Responsive Element (TRE) and reverse tetracycline-controlled TransActivator advanced (rtTAadv), transfected in white (3T3-L1) and brown (HIB-1B) preadipocytes cell lines. The results demonstrate that doxycycline adipogenic inducible expression can be achieved using a pLenti TRE / rtTA adv under the control of the truncated aP2 promoter in HIB-1B preadipocytes.

Liu, Q.; Hill, P.J.; Karamitri, A.; Ryan, K.J.P.; Chen, H.Y.; Lomax, M.A.



Prothrombotic effects of tumor necrosis factor alpha in vivo are amplified by the absence of TNF-alpha receptor subtype 1 and require TNF-alpha receptor subtype 2  

PubMed Central

Introduction Elevated serum levels of the proinflammatory cytokine tumor necrosis factor alpha (TNF?) correlate with an increased risk for atherothrombotic events and TNF? is known to induce prothrombotic molecules in endothelial cells. Based on the preexisting evidence for the impact of TNF? in the pathogenesis of autoimmune disorders and their known association with an acquired hypercoagulability, we investigated the effects of TNF? and the role of the TNF receptor subtypes TNFR1 and TNFR2 for arteriolar thrombosis in vivo. Methods Arteriolar thrombosis and platelet-rolling in vivo were investigated in wildtype, TNFR1-/-, TNFR2-/- and TNFR1-/R2-/- C57BL/6 mice using intravital microscopy in the dorsal skinfold chamber microcirculation model. In vitro, expression of prothrombotic molecules was assessed in human endothelial cells by real-time PCR and flow cytometry. Results In wildtype mice, stimulation with TNF? significantly accelerated thrombotic vessel occlusion in vivo upon ferric chloride injury. Arteriolar thrombosis was much more pronounced in TNFR1-/- animals, where TNF? additionally led to increased platelet-endothelium-interaction. TNF? dependent prothrombotic effects were not observed in TNFR2-/- and TNFR1-/R2- mice. In vitro, stimulation of human platelet rich plasma with TNF? did not influence aggregation properties. In human endothelial cells, TNF? induced superoxide production, p-selectin, tissue factor and PAI-1, and suppressed thrombomodulin, resulting in an accelerated endothelial dependent blood clotting in vitro. Additionally, TNF? caused the release of soluble mediators by endothelial cells which induced prothrombotic and suppressed anticoagulant genes comparable to direct TNF? effects. Conclusions TNF? accelerates thrombus formation in an in vivo model of arteriolar thrombosis. Its prothrombotic effects in vivo require TNFR2 and are partly compensated by TNFR1. In vitro studies indicate endothelial mechanisms to be responsible for prothrombotic TNF? effects. Our results support a more selective therapeutic approach in anticytokine therapy favouring TNFR2 specific antagonists.



Transient, Inducible, Placenta-Specific Gene Expression in Mice  

PubMed Central

Molecular understanding of placental functions and pregnancy disorders is limited by the absence of methods for placenta-specific gene manipulation. Although persistent placenta-specific gene expression has been achieved by lentivirus-based gene delivery methods, developmentally and physiologically important placental genes have highly stage-specific functions, requiring controllable, transient expression systems for functional analysis. Here, we describe an inducible, placenta-specific gene expression system that enables high-level, transient transgene expression and monitoring of gene expression by live bioluminescence imaging in mouse placenta at different stages of pregnancy. We used the third generation tetracycline-responsive tranactivator protein Tet-On 3G, with 10- to 100-fold increased sensitivity to doxycycline (Dox) compared with previous versions, enabling unusually sensitive on-off control of gene expression in vivo. Transgenic mice expressing Tet-On 3G were created using a new integrase-based, site-specific approach, yielding high-level transgene expression driven by a ubiquitous promoter. Blastocysts from these mice were transduced with the Tet-On 3G-response element promoter-driving firefly luciferase using lentivirus-mediated placenta-specific gene delivery and transferred into wild-type pseudopregnant recipients for placenta-specific, Dox-inducible gene expression. Systemic Dox administration at various time points during pregnancy led to transient, placenta-specific firefly luciferase expression as early as d 5 of pregnancy in a Dox dose-dependent manner. This system enables, for the first time, reliable pregnancy stage-specific induction of gene expression in the placenta and live monitoring of gene expression during pregnancy. It will be widely applicable to studies of both placental development and pregnancy, and the site-specific Tet-On G3 mouse will be valuable for studies in a broad range of tissues.

Fan, Xiujun; Petitt, Matthew; Gamboa, Matthew; Huang, Mei; Dhal, Sabita; Druzin, Maurice L.; Wu, Joseph C.



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

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

Guerder, S; Picarella, D E; Linsley, P S; Flavell, R A



Asbestos fibres and man made mineral fibres: induction and release of tumour necrosis factor-alpha from rat alveolar macrophages.  

PubMed Central

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

Ljungman, A G; Lindahl, M; Tagesson, C



Tumor necrosis factor alpha inhibits contractions to sympathetic nerve stimulation by a nitric oxide-dependent mechanism.  


Gram-negative sepsis and administration of tumor necrosis factor alpha (TNF alpha) are associated with hypotension and peripheral neuropathies suggestive of impaired sympathetic neurotransmission. We examined the effect of TNF alpha on the responses of the bovine pulmonary artery (BPA) to transmural sympathetic nerve stimulation (SNS). BPA contracted to SNS (0.5-32 Hz, 5-10 V, 2-msec duration, 2-msec delay) in a frequency-dependent manner. The contractions of the BPA to SNS were mediated by norepinephrine and activation of postsynaptic alpha 1-adrenoceptors, since they were attenuated by prazosin. Maximum contraction of the BPA to SNS was significantly enhanced (148 +/- 37% increase, n = 6) after inhibition of nitric oxide synthase with L-NG-monomethylarginine (LNMMA, 500 microM), an effect abrogated by L-arginine (1 mM). TNF alpha (0.0042, 0.042, and 0.42 micrograms/ml) selectively inhibited contractions of the BPA to SNS without affecting the contraction of the BPA to exogenous norepinephrine. In BPA incubated with LNMMA (5-500 microM), TNF alpha facilitated rather than inhibited SNS. TNF alpha increased the formation of amperiometrically measured free nitric oxide in bovine adrenal chromaffin cells in primary culture. The data show that in the absence of LNMMA, TNF alpha releases free nitric oxide from a sympathetic neuron and selectively inhibits the contractions of the BPA to SNS. In BPA in which nitric oxide synthase I is inhibited by LNMMA, TNF alpha amplifies the contractions to SNS, even in the absence of endothelium. Thus, TNF alpha can modify vascular smooth muscle tone by affecting SNS. TNF alpha inhibits SNS at the level of the neuron by a mechanism involving the L-arginine-nitric oxide pathway. TNF alpha-induced suppression of SNS and neurotransmission may contribute to the hypotension and peripheral neuropathy of sepsis. PMID:7688901

Xie, J; Wang, Y; Kolls, J; Malinski, T; Nelson, S; Summer, W; Greenberg, S S



Tumor Necrosis Factor Alpha-Mediated Toxic Shock in Trypanosoma cruzi-Infected Interleukin 10-Deficient Mice  

PubMed Central

Using interleukin-10 (IL-10)-deficient (IL-10?/?) mice, previous studies revealed a pathological immune response after infection with Trypanosoma cruzi that is associated with CD4+ T cells and overproduction of proinflammatory cytokines. In this study we further investigate the pathology and potential mediators for the mortality in infected animals. T. cruzi-infected IL-10?/? mice showed reduced parasitemia accompanied by increased systemic release of gamma interferon (IFN-?), IL-12, and reactive nitrogen intermediates and overproduction of tumor necrosis factor alpha (TNF-?). Despite this early resistance, IL-10?/? mice died within the third week of infection, whereas all control mice survived acute infection. The clinical manifestation with weight loss, hypothermia, hypoglycemia, hyperkalemia, and increased liver-derived enzymes in the blood together with hepatic necrosis and intravascular coagulation in moribund mice indicated a toxic shock-like syndrome, possibly mediated by the systemic TNF-? overproduction. Indeed, high production of systemic TNF-? significantly correlated with mortality, and moribund mice died with critically high TNF-? concentrations in the blood. Consequent treatment with anti-TNF-? antiserum attenuated pathological changes in T. cruzi-infected IL-10?/? mice and significantly prolonged survival; the mice died during the fourth week postinfection, again with a striking correlation between regaining high systemic TNF-? concentrations and the time of death. Since elevated serum IL-12 and IFN-? concentrations were not affected by the administration of antiserum, these studies suggest that TNF-? is the direct mediator of this toxic shock syndrome. In conclusion, induction of endogenous IL-10 during experimentally induced Chagas' disease seems to be crucial for counterregulating an overshooting proinflammatory cytokine response resulting in TNF-?-mediated toxic shock.

Holscher, Christoph; Mohrs, Markus; Dai, Wen Juan; Kohler, Gabriele; Ryffel, Bernhard; Schaub, Gunter A.; Mossmann, Horst; Brombacher, Frank



Gigantol isolated from the whole plants of Cymbidium goeringii inhibits the LPS-induced iNOS and COX-2 expression via NF-kappaB inactivation in RAW 264.7 macrophages cells.  


During our efforts to find bioactive natural products with anti-inflammatory activity, we isolated gigantol from the whole plants of Cymbidium goeringii (Orchidaceae) by activity-guided chromatographic fractionation. Gigantol was found to have potent inhibitory effects on LPS-induced nitric oxide (NO) and prostaglandin E (2) (PGE (2)) production in RAW 264.7 cells. Consistent with these findings, gigantol suppressed the expression of inducible nitric oxide synthase (iNOS) and cyclooxygenase-2 (COX-2) at the protein and mRNA levels in RAW 264.7 cells in a concentration-dependent manner. Our data also indicate that gigantol is a potent inhibitor of tumor necrosis factor-alpha (TNF-alpha), interleukin-1beta (IL-1beta) and interleukin-6 (IL-6) release and influenced the mRNA expression levels of these cytokines in a dose-dependent manner. Furthermore, a reporter gene assay for nuclear factor kappa B (NF-kappaB) and an electromobility shift assay (EMSA) demonstrated that gigantol effectively inhibited the activation of NF-kappaB, which is necessary for the expression of iNOS, COX-2, TNF-alpha, IL-1beta and IL-6. Thus, our studies suggest that gigantol inhibits LPS-induced iNOS and COX-2 expression by blocking NF- kappaB activation. PMID:16924582

Won, Jong-Heon; Kim, Ji-Yeon; Yun, Kyung-Jin; Lee, Jin-Hee; Back, Nam-In; Chung, Hae-Gon; Chung, Sun A; Jeong, Tae-Sook; Choi, Myung-Sook; Lee, Kyung-Tae



Osteopontin expression in angiotensin II-induced tubulointerstitial nephritis  

Microsoft Academic Search

Osteopontin expression in angiotensin II-induced tubulointerstitial nephritis. Osteopontin is an arginine-glycine-aspartate (RGD) containing secreted phosphoprotein recently shown to stimulate a local macrophage influx when injected subcutaneously in mice. We examined the effect of angiotensin II infusion on renal injury and osteopontin expression in the rat kidney by in situ hybridization and immunohistochemistry. Preceding pathologic changes in tubular and interstitial cells,

Cecilia M Giachelli; Raimund Pichler; Donna Lombardi; David T Denhardt; Charles E Alpers; Stephen M Schwartz; Richard J Johnson



Expression of hypoxia-inducible factor 1  in thyroid carcinomas  

Microsoft Academic Search

Hypoxia-inducible factor 1a (HIF-1a) is upregulated by hypoxia and oncogenic signalling in many solid tumours. Its regulation and function in thyroid carcinomas are unknown. We evaluated the regulation of HIF-1a and target gene expression in primary thyroid carcinomas and thyroid carcinoma cell lines (BcPAP, WRO, FTC-133 and 8505c). HIF-1a was not detectable in normal tissue but was expressed in thyroid

N Burrows; J Resch; R L Cowen; R von Wasielewski; C Hoang-Vu; C M West; K J Williams; G Brabant



Human mesangial cells express inducible macrophage scavenger receptor  

Microsoft Academic Search

Human mesangial cells express inducible macrophage scavenger receptor.BackgroundType A scavenger receptors (Scr) mediate the uptake of modified low-density lipoproteins by macrophages. The accumulation of lipids via this process is thought to lead to foam cell formation in atherosclerotic plaques. Human mesangial cells (HMCs) have not been previously shown to express Scr in normal culture. We therefore investigated whether there is

Xiong Z. Ruan; Zac Varghese; Stephen H. Powis; John F. Moorhead



Ganoderma lucidum inhibits inducible nitric oxide synthase expression in macrophages  

Microsoft Academic Search

Nitric oxide (NO) is a principal mediator in many physiological and pathological processes. Overproduction of NO via the inducible nitric oxide synthase (iNOS) has cytotoxic effect through the formation of peroxynitrite with superoxide anion. The iNOS is mainly expressed in macrophages and is able to produce large amount of NO. The expression of iNOS is mainly regulated at the transcriptional

Connie W. H. Woo; Ricky Y. K. Man; Yaw L. Siow; Patrick C. Choy; Eric W. Y. Wan; Chak S. Lau; Karmin O



Cyanate Is a Novel Inducer of Endothelial ICAM-1 Expression  

PubMed Central

Abstract Aim Recent work has shown that humans are significantly exposed to isocyanic acid/cyanate, which is generated when coal, biomass, or tobacco is burned. In vivo, cyanate is formed by the phagocyte protein myeloperoxidase and by breakdown of urea. Carbamylation of proteins through cyanate has been demonstrated to predict cardiovascular risk and is thought to promote vascular dysfunction; however, the underlying mechanisms remain unclear. Results: Here, we show that cyanate induces intercellular cell adhesion molecule-1 (ICAM-1) expression with subsequently enhanced neutrophil adhesion in human coronary artery endothelial cells. Cyanate triggers ICAM-1 expression through a mechanism depending on activation of the mitogen-activated protein kinase p38 and nuclear factor-kappaB. Endothelial ICAM-1 expression was not induced when low-molecular-weight substances were removed from cell culture medium, thus ruling out a role of carbamylated (lipo)proteins in ICAM-1 induction. In mice, oral administration of cyanate induced marked endothelial ICAM-1 expression in the aorta. Moreover, in patients with end-stage renal disease, the extent of plasma protein carbamylation (a marker for cyanate exposure) significantly correlated with plasma levels of soluble ICAM-1. Innovation: Here, we demonstrate for the first time that cyanate, rather than carbamylated lipoproteins, induces vascular ICAM-1 expression in vivo. Conclusion: Collectively, our data raise the possibility that cyanate amplifies vascular inflammation, linking inflammation, smoking, and uremia. Antioxid. Redox Signal. 16, 129–137.

El-Gamal, Dalia; Holzer, Michael; Gauster, Martin; Schicho, Rudolf; Binder, Veronika; Konya, Viktoria; Wadsack, Christian; Schuligoi, Rufina; Heinemann, Akos



Leptin deficiency-induced obesity exacerbates ultraviolet B radiation-induced cyclooxygenase-2 expression and cell survival signals in ultraviolet B-irradiated mouse skin  

SciTech Connect

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.

Sharma, Som D. [Department of Dermatology, University of Alabama at Birmingham, Birmingham, AL 35294 (United States); Katiyar, Santosh K., E-mail: skatiyar@uab.ed [Birmingham Veterans Administration Medical Center, Birmingham, AL 35294 (United States); Department of Dermatology, University of Alabama at Birmingham, Birmingham, AL 35294 (United States); Clinical Nutrition Research Center, University of Alabama at Birmingham, Birmingham, AL 35294 (United States); Comprehensive Cancer Center, University of Alabama at Birmingham, Birmingham, AL 35294 (United States)



Short-Lived Effector CD8 T Cells Induced by Genetically Attenuated Malaria Parasite Vaccination Express CD11c  

PubMed Central

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-?), 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-?, tumor necrosis factor alpha (TNF-?), 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.

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.



Inducible nitric oxide synthase expression in human cerebral infarcts  

Microsoft Academic Search

The inducible or “immunological” isoform of nitric oxide synthase (iNOS) is induced in many cell types by inflammatory stimuli\\u000a and synthesizes toxic amounts of NO. In rodent models of focal cerebral ischemia, iNOS is expressed in neutrophils invading\\u000a the injured brain and in local blood vessels. Studies with iNOS inhibitors and iNOS null mice indicate that NO produced by\\u000a iNOS

Colleen Forster; H. Brent Clark; M. Elizabeth Ross; Constantino Iadecola



Thiostrepton-induced gene expression in Streptomyces lividans.  


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. PMID:2537819

Murakami, T; Holt, T G; Thompson, C J



Icariin attenuates LPS-induced acute inflammatory responses: Involvement of PI3K\\/Akt and NF-?B signaling pathway  

Microsoft Academic Search

This study aimed to investigate the mechanism underlying the attenuation of LPS-induced lung inflammation by icariin in vivo and in vitro. The anti-inflammatory effects of icariin on LPS-induced acute inflammatory and the molecular mechanism were investigated. Pretreatment with icarrin (20mg\\/kg) could attenuate acute lung inflammation by inhibiting mRNA expressions of tumor necrosis factor alpha (TNF-?), interleukin-6 (IL-6), metalloproteinase cycloxygenase-2 (COX-2),

Chang-Qing Xu; Bao-Jun Liu; Jin-Feng Wu; Yan-Chun Xu; Xiao-Hong Duan; Yu-Xue Cao; Jing-Cheng Dong



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

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

Poli, G; Kinter, A L; Fauci, A S



Expression of interleukin-15 and inflammatory cytokines in skeletal muscles of STZ-induced diabetic rats: effect of resistance exercise training.  


Skeletal muscle atrophy is associated with type-1 diabetes. Skeletal muscle is the source of pro- and anti-inflammatory cytokines that can mediate muscle hypertrophy and atrophy, while resistance exercise can modulate both muscle mass and muscle cytokine expression. This study determined the effects of a 5-week resistance exercise training regimen on the expression of muscle cytokines in healthy and streptozotocin-induced diabetic rats, with special emphasis on interleukin-15 (IL-15), a muscle-derived cytokine proposed to be involved in muscle hypertrophy or responses to stress. Induction of diabetes reduced muscle weight in both the fast flexor hallucis longus (FHL) and slow soleus muscles, while resistance training preserved FHL muscle weight in diabetic rats. IL-15 protein content was increased by training in both FHL and soleus muscles, as well as serum, in normal and diabetic rats. With regard to proinflammatory cytokines, muscle IL-6 levels were increased in diabetic rats, while training decreased muscle IL-6 levels in diabetic rats; training had no effect on FHL muscle IL-6 levels in healthy rats. Also, tumor necrosis factor-alpha (TNF-?) and IL-1? levels were increased by diabetes, but not changed by training. In conclusion, we found that in diabetic rats, resistance training increased muscle and serum IL-15 levels, decreased muscle IL-6 levels, and preserved FHL muscle mass. PMID:24006180

Molanouri Shamsi, M; Hassan, Z H; Gharakhanlou, R; Quinn, L S; Azadmanesh, K; Baghersad, L; Isanejad, A; Mahdavi, M



Expression of Glucocorticoid Induced Leucine Zipper (GILZ) in Cardiomyocytes  

PubMed Central

Glucocorticoids (GCs) are frequently prescribed pharmacological agents most notably for their immunosuppressive effects. Endogenous GCs mediate biological processes such as energy metabolism and tissue development. At the cellular level, GCs bind to the Glucocorticoid Receptor (GR), a cytosolic protein that translocates to the nuclei and functions to alter transcription upon ligand binding. Amongst a long list of genes activated by GCs is the Glucocorticoid Induced Leucine Zipper (GILZ). GC induced GILZ expression has been well established in lymphocytes and mediates GC induced apoptosis. Unlike lymphocytes, cardiomyocytes respond to GCs by gaining resistance against apoptosis. We determined GILZ expression in cardiomyocytes in vivo and in vitro. Expression of GILZ in mouse hearts as a result of GC administration was confirmed by Western blot analyses. GCs induced dose and time dependent elevation of GILZ expression in primary cultured rat cardiomyocytes, with dexamethasone (Dex) as low as 0.1 M being effective. Time course analysis indicated that GILZ protein levels increased at 8 hr and peaked at 48 hr after exposure to 1 M Dex. H9c2(2-1) cell line showed a similar response of GILZ induction by Dex as primary cultured rat cardiomyocytes, providing a convenient model for studying the biological significance of GILZ expression. With corticosterone (CT), an endogenous form of corticosteroids in rodents, 0.1–2.5 M was found to induce GILZ in H9c2(2-1) cells. Time course analysis with 1 M CT indicated induction of GILZ at 6 hr with peak expression at 18 hr. Inhibition of the GR by mifepristone led to blunting of GILZ induction by GCs. Our data demonstrate GILZ induction in cardiomyocytes both in vivo and in vitro by GCs, pointing to H9c2(2-1) cells as a valid model for studying the biological function of GILZ in cardiomyocytes.

Aguilar, David C.; Strom, Josh; Xu, Beibei; Kappeler, Kyle; Chen, Qin M.



Lutzomyia longipalpis Salivary Gland Homogenate Impairs Cytokine Production and Costimulatory Molecule Expression on Human Monocytes and Dendritic Cells  

Microsoft Academic Search

In this report, we describe an investigation of the effects of Lutzomyia longipalpis sand fly salivary gland homogenates (SGH) on cytokine production and expression of costimulatory molecules on human monocytes, macrophages (Ms), and dendritic cells (DCs). SGH of L. longipalpis induced an increase in interleukin-6 (IL-6), IL-8 and IL-12p40 production but a decrease in tumor necrosis factor alpha and IL-10

Dirceu J. Costa; Cecő ´ lia Favali; Jorge Clarencio; Lő ´ lian Afonso; Viviane Conceicao; JoseCarlos Miranda; Richard G. Titus; Jesus Valenzuela; Manoel Barral-Netto; Aldina Barral; Claudia Ida Brodskyn



Increased gene expression of scavenger receptors and proinflammatory markers in peripheral blood mononuclear cells of hyperlipidemic males  

Microsoft Academic Search

Interactions between peripheral blood mononuclear cells (PBMCs) and those within plaques are suggested to be pathophysiologically\\u000a relevant to lipid-induced arteriosclerosis. In this study, gene expressions of scavenger receptors (CD36, CD68), LPS receptor\\u000a (CD14), proinflammatory (tumor necrosis factor alpha [TNF?], CD40, interleukin-1 beta [IL-1?]) and oxidative stress-related\\u000a (manganese superoxide dismutase [MnSOD]) markers were analyzed in PBMCs of clinically asymptomatic males with

Gabriel A. Bonaterra; Wulf Hildebrandt; Anne Bodens; Roland Sauer; Klaus A. Dugi; Hans-Peter Deigner; Dan Turcanu; Helmut Heinle; Wulf Dröge; Jürgen Metz; Ralf Kinscherf



Human Cytomegalovirus Attenuates Interleukin1  and Tumor Necrosis Factor Alpha Proinflammatory Signaling by Inhibition of NF B Activation  

Microsoft Academic Search

Viral infection is associated with a vigorous inflammatory response characterized by cellular infiltration and release of the proinflammatory cytokines interleukin-1 (IL-1) and tumor necrosis factor alpha (TNF-). In the present study, we identified a novel function of human cytomegalovirus (HCMV) that results in inhibition of IL-1 and TNF- signaling pathways. The effect on these pathways was limited to cells infected

Michael A. Jarvis; Jamie A. Borton; Amy M. Keech; John Wong; William J. Britt; Bruce E. Magun; Jay A. Nelson



Salmonellae Activate Tumor Necrosis Factor Alpha Production in a Human Promonocytic Cell Line via a Released Polypeptide  

Microsoft Academic Search

Invasive strains of Salmonella spp. cause both systemic and localized infections in humans. The ability to resist infection and some aspects of the tissue pathology associated with the presence of Salmonella in the gastrointestinal tract have been shown to be mediated in part by the induction of tumor necrosis factor alpha (TNF-a), a proinflammatory cytokine produced by activated macrophages and




Further evidence that acanthosis nigricans maligna is linked to enhanced secretion by the tumour of transforming growth factor alpha  

Microsoft Academic Search

The pathogenesis of cutaneous paraneoplastic syndromes is still under discussion. Since many of these syndromes, including acanthosis nigricans, are proliferative skin disorders it is believed that products secreted by the tumour stimulate the keratinocytes to proliferate. Growth factors like transforming growth factor alpha (TGF-alpha) are known to be highly mitogenic for keratinocytes in vitro. Here we report on a patient

K. Wilgenbus; A. Lentner; R. Kuckelkorn; St. Handt; Ch. Mittermayer



Association of tumour necrosis factor alpha ?308 gene polymorphism with primary open-angle glaucoma in Chinese  

Microsoft Academic Search

Purpose Genetic factors are known to play a role in the aetiology of glaucoma, and in particular the role of the immune system is highly suspected. In this study, we evaluated the association between tumour necrosis factor alpha ?308 (TNF ??308) and primary open-angle glaucoma (POAG).Methods A total of sixty POAG patients and 103 healthy volunteers as control group were

H-J Lin; F-J Tsai; W-C Chen; Y-R Shi; Y Hsu; S-W Tsai



Impaired Intramembranous Bone Formation during Bone Repair in the Absence of Tumor Necrosis Factor-Alpha Signaling  

Microsoft Academic Search

Tumor necrosis factor-alpha (TNF-?) is known to mediate bone resorption; however, its role in osteogenesis has not been fully elucidated. In order to investigate the direct role of TNF-? signaling in the recruitment and differentiation of osteoblasts, two separate models of bone repair were used, marrow ablation and simple transverse fractures. These models were carried out in the tibiae of

L. C. Gerstenfeld; T.-J. Cho; T. Kon; T. Aizawa; J. Cruceta; B. D. Graves; T. A. Einhorn



TNF? induces HIF-1? expression through activation of IKK?  

PubMed Central

The transcription factor hypoxia-inducible factor 1? (HIF-1?) is regulated by oxygen availability as well as various inflammatory mediators, including tumor necrosis factor ? (TNF?). Early work suggested that the phosphatidylinositol-3-kinase (PI3K) and mitogen-activated protein kinase (MAPK) signaling pathways are involved in TNF?-mediated HIF-1? accumulation and activation under normoxic conditions. Here, we provide evidence showing that I?Bkinase ? (IKK?) is required for HIF-1? regulation by TNF?. We found that TNF? enhances HIF-1? protein expression in various breast cancer cell lines under either normoxic or hypoxia-mimicking conditions, but has little effect on the HIF-1? mRNA level. Increased HIF-1? expression was found in IKK? stable clones and transient transfectants, and depletion of IKK? consistently reduced the amount of HIF-1? protein. Treatment of cells with the IKK? inhibitor Bay 11-7082 reduced the TNF?-induced HIF-1? expression, suggesting that IKK? is required in this signaling pathway. Decreased expression of vascular endothelial growth factor (VEGF), a direct target of HIF-1?, was shown in IKK?-knockout mouse embryonic fibroblast cells. We further demonstrated a positive correlation between IKK? and VEGF expression in primary human breast cancer specimens. Our findings indicate that TNF?-induced HIF-1? accumulation is IKK? dependent, and may enable further understanding of the HIF-1? regulation by inflammatory signals.

Kuo, Hsu-Ping; Lee, Dung-Fang; Xia, Weiya; Wei, Yongkun; Hung, Mien-Chie



An Essential Role for NF-kappaB in Preventing TNF-alpha-Induced Cell Death  

Microsoft Academic Search

Studies on mice deficient in nuclear factor kappa B (NF-kappaB) subunits have shown that this transcription factor is important for lymphocyte responses to antigens and cytokine-inducible gene expression. In particular, the RelA (p65) subunit is required for induction of tumor necrosis factor-alpha (TNF-alpha)-dependent genes. Treatment of RelA-deficient (RelA-\\/-) mouse fibroblasts and macrophages with TNF-alpha resulted in a significant reduction in

Amer A. Beg; David Baltimore



Wound healing potential of Ocimum sanctum Linn. with induction of tumor necrosis factor-alpha.  


Ocimum sanctum, a well known herb in Indian medicine, possesses various therapeutic properties including healing properties and cytokine induction. Wound healing activity of cold aqueous extract of O. sanctum leaves along with its effect on tumor necrosis factor-alpha (TNF-alpha) was assessed using excision model of wound repair in Wistar albino rats. After application of the O. sanctum extract, rate of epithelization with an increase in wound contraction was observed. In animals, treated with 10% O. sanctum extract in petroleum jelly, wound healing was faster as compared to control group which were treated with petroleum jelly alone but significant accelerated healing was noticed in animals which in addition to the topical application of 10% extract of O. sanctum, were prefed with 250 mg/kg body weight of aqueous O. sanctum extract daily for 20 consecutive days. During wound healing phase TNF-alpha level was found to be up regulated by O. sanctum treatment. Early wound healing may be pronounced due to O. sanctum extract, by elevating TNF-alpha production. PMID:20726339

Goel, Anjana; Kumar, Sandeep; Singh, Dilip Kumar; Bhatia, Ashok Kumar



Dissociative symptoms reflect levels of tumor necrosis factor alpha in patients with unipolar depression  

PubMed Central

Recent evidence indicates that the nature of interactions between the nervous system and immune system is important in the pathogenesis of depression. Specifically, alterations in pro-inflammatory cytokines have been related to the development of several psychological and neurobiological manifestations of depressive disorder, as well as to stress exposure. A number of findings point to tumor necrosis factor alpha (TNF-?) as one of the central factors in these processes. Accordingly, in the present study, we test the hypothesis that specific influences of chronic stressors related to traumatic stress and dissociation are related to alterations in TNF-? levels. We performed psychometric measurement of depression (Beck Depression Inventory [BDI]-II), traumatic stress symptoms (Trauma Symptom Checklist [TSC]-40), and psychological and somatoform dissociation (Dissociative Experiences Scale [DES] and Somatoform Dissociation Questionnaire [SDQ]-20, respectively), and immunochemical measure of serum TNF-? in 66 inpatients with unipolar depression (mean age 43.1 ± 7.3 years). The results show that TNF-? is significantly related to DES (Spearman R=?0.42, P<0.01), SDQ-20 (Spearman R=?0.38, P<0.01), and TSC-40 (Spearman R=?0.41, P<0.01), but not to BDI-II. Results of the present study suggest that TNF-? levels are related to dissociative symptoms and stress exposure in depressed patients.

Bizik, Gustav; Bob, Petr; Raboch, Jiri; Pavlat, Josef; Uhrova, Jana; Benakova, Hana; Zima, Tomas



Faecal interleukin-8 and tumour necrosis factor-alpha concentrations in cystic fibrosis.  

PubMed Central

Interleukin-8 (IL-8) and tumour necrosis factor-alpha (TNF-alpha) concentrations were measured in faecal samples from nine patients with cystic fibrosis and nine healthy age matched controls. The patients were assessed with Shwachman score, apparent energy absorption, pancreatic enzyme dosage, simple spirometry, and presence of pseudomonal colonisation. Median (range) wet stool IL-8 and TNF-alpha concentrations in patients were 32,113 pg/g (21,656-178,128) and 3187 pg/g (368-17,611) respectively, compared with < 43.5 pg (IL-8)/g (< 22-4079) and 99 pg (TNF-alpha)/g (< 0.26-231) in controls. IL-8 concentration was negatively correlated with Shwachman score (r = -0.79) and pancreatic enzyme dosage (r = -0.77), but not with energy absorption. Seven patients were mature enough to cooperate with spirometry. Their IL-8 concentrations correlated with percentage predicted forced expiratory volume in one second (r = -0.78). IL-8 concentration was greater in four patients with, than five without, established pseudomonal colonisation: median difference 134,583 pg/g. TNF-alpha concentration was not correlated with measures of disease severity. Faecal IL-8 concentration might reflect the severity of pulmonary inflammation in cystic fibrosis and could provide an easily obtainable marker of disease activity.

Briars, G L; Dean, T P; Murphy, J L; Rolles, C J; Warner, J O



Topical application of the tumour necrosis factor-alpha antibody infliximab improves healing of chronic wounds.  


The role of tumour necrosis factor-alpha (TNF-alpha) in wound healing is not clear. Elevated levels of TNF-alpha have been observed in fluids from chronic wounds and have been shown to decrease over time during the healing process. Therapeutic antibodies such as infliximab can inhibit TNF-alpha activity. In this case series, we applied infliximab topically to eight patients with chronic ulcers of more than 4-month durations. The ulcers had multifactorial aetiology, with chronic venous insufficiency being the most prominent factor. All the ulcers had failed to respond to any previous conventional treatment. Infliximab was applied repeatedly to ulcers either as a 10 mg/ml solution and covered with an adhesive sheet or as a gel formulation (0.45, 1, or 4.5 mg/g) under a hydrofiber dressing/adhesive sheet. Improvement was assessed by measuring the percentage of change in the ulcer surface area. Seven of the eight patients (12 of 14 ulcers) responded to treatment with infliximab. After 4 weeks of treatment, surface area was reduced by more than 50% in 6 of the 14 treated ulcers. Within 8 weeks, five ulcers completely healed, while another four were reduced by more than 75% in size. Chronic, therapy-resistant leg ulcers responded well to repeated topical administration of a solution or a gel containing the TNF-alpha antibody, infliximab. Randomised controlled studies should be conducted to further evaluate the effect of topical infliximab on chronic wound healing. PMID:16984574

Streit, Markus; Beleznay, Zsuzsanna; Braathen, Lasse R



Estrogen Induces Vav1 Expression in Human Breast Cancer Cells  

PubMed Central

Vav1, a guanine nucleotide exchange factor (GEF) for Rho family GTPases, is a hematopoietic protein involved in a variety of cellular events. In recent years, aberrant expression of Vav1 has been reported in non-hematopoietic cancers including human breast cancer. It remains to be answered how Vav1 is expressed and what Vav1 does in its non-resident tissues. In this study, we aimed to explore the mechanism for Vav1 expression in breast cancer cells in correlation with estrogen-ER pathway. We not only verified the ectopic expression of Vav1 in human breast cancer cell lines, but also observed that Vav1 expression was induced by 17?-estradiol (E2), a typical estrogen receptor (ER) ligand, in ER-positive cell lines. On the other hand, Tamoxifen, a selective estrogen receptor modulator (SERM), and ICI 182,780, an ER antagonist, suppressed the expression of Vav1. The estrogen receptor modulating Vav1 expression was identified to be ? form, not ?. Furthermore, treatment of E2 increased the transcription of vav1 gene by enhancing the promoter activity, though there was no recognizable estrogen response element (ERE). Nevertheless, two regions at the vav1 gene promoter were defined to be responsible for E2-induced activation of vav1 promoter. Chromatin immunoprecipitation (ChIP) and co-immunoprecipitation (Co-IP) analyses suggested that ER? might access to the vav1 promoter via interacting with transcription factors, c-Myb and ELF-1. Consequently, the enhanced expression of Vav1 led to the elevation of Cyclin D1 and the progression of cell cycle. The present study implies that estrogen-ER modulates the transcription and expression of Vav1, which may contribute to the proliferation of cancerous cells.

Du, Ming-juan; Chen, Xiang-dong; Zhou, Xiao-li; Wan, Ya-juan; Lan, Bei; Zhang, Cui-zhu; Cao, Youjia



Role and new perspectives of transforming growth factor-alpha (TGF-alpha) in adenocarcinoma of the gastro-oesophageal junction.  


The incidence of gastro-oesophageal junction (GEJ) adenocarcinoma is increasing in Western countries and prognosis is poor since metastasis is most often present at diagnosis. We examined samples from 87 resected type II GEJ adenocarcinomas, 30 of these with endoscopic diagnostic biopsy material, to evaluate transforming growth factor alpha (TGF-a) expression and p53 overexpression by immunohistochemistry and in situ hybridization (for TGF-alpha), in relation to biological and clinical behaviour. TGF-alpha messenger RNA (mRNA) and protein were detectable in neoplastic cells in 56% and 64% cases respectively. TGF-alpha mRNA was detected in intra- and peritumoral lymphocytes and those of metastatic lymph nodes. TGF-alpha protein expression was significantly associated with tumour progression (P= 0.025) and lymph node metastasis (P < 0.05). The strong TGF-alpha expression found in neoplastic cells inside blood and lymphatic vessels and in metastatic localizations suggests that TGF-a-positive GEJ adenocarcinomas could have a more aggressive biological phenotype. The expression of TGF-alpha mRNA and protein in both inflammatory and neoplastic cells indicates that TGF-alpha is directly synthesized by both cell compartments. Finally, since TGF-alpha expression was associated with lymph node metastasis, its detection in preoperative perendoscopic biopsies might identify patients with more aggressive tumours who may need additional therapy, including neo-adjuvant treatment. PMID:10732760

D'Errico, A; Barozzi, C; Fiorentino, M; Carella, R; Di Simone, M; Ferruzzi, L; Mattioli, S; Grigioni, W F



Radiocurability by Targeting Tumor Necrosis Factor-{alpha} Using a Bispecific Antibody in Carcinoembryonic Antigen Transgenic Mice  

SciTech Connect

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.

Larbouret, Christel; Robert, Bruno [INSERM, Centre de Recherche en Cancerologie de Montpellier, Universite de Montpellier, and CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Linard, Christine [IRSN, Fontenay-aux-Roses (France); Teulon, Isabelle [INSERM, Centre de Recherche en Cancerologie de Montpellier, Universite de Montpellier, and CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Gourgou, Sophie M.Sc. [Unite de Biostatistiques en Oncologie, CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Bibeau, Frederic [Departement d'Anatomie Pathologique, CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Martineau, Pierre [Institut de Biotechnologie et Pharmacologie, CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Santoro, Lore [INSERM, Centre de Recherche en Cancerologie de Montpellier, Universite de Montpellier, and CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Departement d'Oncologie Radiotherapie, CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Pouget, Jean-Pierre; Pelegrin, Andre [INSERM, Centre de Recherche en Cancerologie de Montpellier, Universite de Montpellier, and CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Azria, David [INSERM, Centre de Recherche en Cancerologie de Montpellier, Universite de Montpellier, and CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France); Departement d'Oncologie Radiotherapie, CRLC Val d'Aurelle-Paul Lamarque, Montpellier (France)], E-mail:



Purinergic signaling induces thrombospondin-1 expression in astrocytes  

PubMed Central

Thrombospondin (TSP)-1, a multidomain glycoprotein, is secreted from astrocytes and promotes synaptogenesis. However, little is known about the mechanisms regulating its expression and release. In this article, we report that purinergic signaling participates in the production and secretion of TSP-1. Treatment of primary cultures of rat cortical astrocytes with extracellular ATP caused an increase in TSP-1 expression in a time- and concentration-dependent manner and was inhibited by antagonists of P2 and P1 purinergic receptors. Agonist studies revealed that UTP, but not 2?,3?-O-(4-benzoyl)benzoyl-ATP, 2-methylthio-ADP, adenosine, or 5?-N-ethyl-carboxamidoadenosine, caused a significant increase in TSP-1 expression. In addition, release of TSP-1 was stimulated by ATP and UTP but not by 2-methylthio-ADP or adenosine. Additional studies indicated that P2Y4 receptors stimulate both TSP-1 expression and release. P2Y receptors are coupled to protein kinase cascades, and signaling studies demonstrated that blockade of mitogen-activated protein kinases or Akt inhibited ATP- and UTP-induced TSP-1 expression. Using an in vitro model of CNS trauma that stimulates release of ATP, we found that TSP-1 expression increased after mechanical strain and was completely blocked by a P2 receptor antagonist and by inhibition of p38/mitogen-activated protein kinase and Akt, thereby indicating a major role for P2 receptor/protein kinase signaling in TSP-1 expression induced by trauma. We conclude that TSP-1 expression can be regulated by activation of P2Y receptors, particularly P2Y4, coupled to protein kinase signaling pathways and suggest that purinergic signaling may be an important factor in TSP-1-mediated cell-matrix and cell–cell interactions such as those occurring during development and repair.

Tran, Minh D.; Neary, Joseph T.



Hypoxia-inducible regulation of placental BOK expression.  


BOK (BCL-2-related ovarian killer) is a member of the pro-apoptotic BCL-2 family that is highly expressed in the human placenta. BOK excess causes increased trophoblast autophagy and apoptosis in pre-eclampsia, a pathological condition of hypoxia and oxidative stress. In the present study, we identified an HRE (hypoxia-response element) at the junction of exon-1 and intron-1 (+229 to +279) in the human BOK gene, as well as an antisense transcript driven by a promoter located in intron-2. The isolated BOK-HRE bound hypoxia-inducible HIF (hypoxia-inducible factor) proteins in vitro as well as in trophoblastic JEG3 cells and was functional in its natural position as well as in front of a heterologous promoter. Being a reverted repeat, the BOK-HRE functioned in both orientations. This directionless feature of the BOK-HRE facilitates hypoxia regulation via HIF of both BOK and its antisense transcript as demonstrated by RNAi knockdown of the HIF system. Although the antisense transcript was expressed in several human carcinoma cell lines, including choriocarcinoma-derived JEG3 cells, no antisense-regulated mechanism for BOK expression was noted. Taken together, these findings indicate that hypoxia-induced expression of BOK in placental cells is regulated via HIF and is not affected by its antisense transcript. PMID:24806027

Luo, Daochun; Caniggia, Isabella; Post, Martin



Cytokine expression and signaling in drug-induced cellular senescence.  


Cellular senescence guards against cancer and modulates aging; however, the underlying mechanisms remain poorly understood. Here, we show that genotoxic drugs capable of inducing premature senescence in normal and cancer cells, such as 5-bromo-2'-deoxyuridine (BrdU), distamycin A (DMA), aphidicolin and hydroxyurea, persistently activate Janus kinase-signal transducer and activator of transcription (JAK/STAT) signaling and expression of interferon-stimulated genes (ISGs), such as MX1, OAS, ISG15, STAT1, PML, IRF1 and IRF7, in several human cancer cell lines. JAK1/STAT-activating ligands, interleukin 10 (IL10), IL20, IL24, interferon gamma (IFNgamma), IFNbeta and IL6, were also expressed by senescent cells, supporting autocrine/paracrine activation of JAK1/STAT. Furthermore, cytokine genes, including proinflammatory IL1, tumor necrosis factor and transforming growth factor families, were highly expressed. The strongest inducer of JAK/STAT signaling, cytokine production and senescence was BrdU combined with DMA. RNA interference-mediated knockdown of JAK1 abolished expression of ISGs, but not DNA damage signaling or senescence. Thus, although DNA damage signaling, p53 and RB activation, and the cytokine/chemokine secretory phenotype are apparently shared by all types of senescence, our data reveal so far unprecedented activation of the IFNbeta-STAT1-ISGs axis, and indicate a less prominent causative role of IL6-JAK/STAT signaling in genotoxic drug-induced senescence compared with reports on oncogene-induced or replicative senescence. These results highlight shared and unique features of drug-induced cellular senescence, and implicate induction of cancer secretory phenotype in chemotherapy. PMID:19802007

Novakova, Z; Hubackova, S; Kosar, M; Janderova-Rossmeislova, L; Dobrovolna, J; Vasicova, P; Vancurova, M; Horejsi, Z; Hozak, P; Bartek, J; Hodny, Z



Thrombin Induces VEGF Expression in Human Retinal Pigment Epithelial Cells  

PubMed Central

PURPOSE The aim of the present study was to investigate effects of thrombin and thrombin in combination with other proangiogenic factors on VEGF expression in hRPE cells. METHODS hRPE cells were stimulated with thrombin TNF-?, monocytes and TGF-?2. Following stimulation, conditioned medium and lysed cells were subjected to ELISA, Western blotting, immunocytochemistry and RT-PCR analyses. Inhibitors specific for various signal transduction pathways were employed to determine the signal pathways involved. RESULTS Treatment of RPE cells with thrombin resulted in dose- and time-dependent increases in VEGF mRNA levels and protein production. hRPE VEGF expression is predominantly PAR-1 dependent. About 80% of thrombin-induced VEGF secretion was abrogated by inhibitors of MAPK/ERK kinase (MEK), p38, c-Jun NH2-terminal kinase (JNK), protein tyrosine kinase (PTK), phosphatidylinositol 3-kinase (PI3K), protein kinase C (PKC), nuclear factor-?B (NF-?B), and reactive oxygen species (ROS). Analyses of VEGF protein production and mRNA synthesis revealed that VEGF induction by thrombin plus TNF-? or co-culture with monocytes was additive, while that by co-incubation with TGF-?2 was synergistic. The co-stimulated VEGF production by TGF-?2 plus thrombin averaged threefold higher than the sum of that induced by each agent alone. Furthermore, BAPTA, a calcium chelator, blocked the VEGF secretion induced by thrombin and thrombin plus TGF-?2 by 65% and 20%, respectively, but had no effect on that by TGF-?2 alone. CONCLUSIONS Thrombin alone and in combination with TNF-?, monocytes and TGF-?2 potently stimulated VEGF expression in hRPE cells via multiple signal pathways. The thrombin-induced calcium mobilization may play an important permissive role in maximizing TGF-?2–induced VEGF expression in RPE cells.

Bian, Zong-Mei; Elner, Susan G.; Elner, Victor M.



A novel function of transcription factor alpha-Pal/NRF-1: increasing neurite outgrowth.  


Alpha-Pal/NRF-1 is a critical regulator of the promoter of human IAP/CD47 gene, a gene related to memory formation in rodents. However, its function in neurons was unknown. We found that stable or transient expression of full-length alpha-Pal/NRF-1 in human neuroblastoma IMR-32 cells significantly induced neurite outgrowth and increased the length of neurites both in medium containing 10% fetal bovine serum and in serum-free medium. In contrast, the dominant-negative mutant of alpha-Pal/NRF-1 inhibited the induction and extension of neurites. Ectopic expression of full-length alpha-Pal/NRF-1 also increased the induction of neurite outgrowth in primary mouse cortical neurons. The IAP antisense cDNA significantly inhibited the increase of neurite outgrowth by alpha-Pal/NRF-1. These findings indicate that a novel function of alpha-Pal/NRF-1 is to regulate neuronal differentiation, and that this function is mediated partly via its downstream IAP gene. PMID:15992771

Chang, Wen-Teng; Chen, Hsiun-ing; Chiou, Rong-Jing; Chen, Chen-Yun; Huang, A-Min



Prostaglandin E2 inhibits advanced glycation end product-induced adhesion molecule expression on monocytes, cytokine production, and lymphocyte proliferation during human mixed lymphocyte reaction.  


Posttransplant diabetes mellitus is a frequent complication among transplant recipients. Ligation of advanced glycation end products (AGEs) with their receptor on monocytes/macrophages plays a role in diabetes complications. The enhancement of adhesion molecule expression on monocytes/macrophages activates T cells, reducing allograft survival. In previous work, we found that toxic AGEs, AGE-2 and AGE-3, induced the expression of intracellular adhesion molecule-1, B7.1, B7.2, and CD40 on monocytes, production of interferon-gamma and tumor necrosis factor alpha, and lymphocyte proliferation during human