Role Of Tumor Necrosis Factor-alpha Research Articles

Tumor necrosis factor‐α inhibits seizures in mice via p75 receptors

Brain inflammatory reactions have been described in various neurological disorders, including epilepsy. Although there is clear evidence that cytokines affect neuroglial functions and blood-brain barrier permeability, scarce information is available on the functional consequences of brain inflammation on seizures. We studied the role of tumor necrosis factor-alpha (TNF)-alpha and its p55 and p75 receptors in seizure modulation. We found that intrahippocampal injection of murine recombinant TNF-alpha potently inhibits seizure in mice while human recombinant TNF-alpha, which shows strong specificity for mouse p55 receptors, was ineffective. p75 receptors were detected in mouse hippocampal neurons, whereas p55 receptors were absent. Transgenic mice with a perturbed TNF-alpha system showed profound alterations in seizure susceptibility: astrocytic overexpression of TNF-alpha was associated with reduced seizures, whereas mice lacking TNF-alpha p75 or both p55 and p75, receptors showed prolonged seizures. Mice deficient in p55 receptor only showed reduced seizures; and both p75 and TNF receptor-associated factor 2 protein levels were upregulated in their hippocampi. Our findings show that increased brain levels of TNF-alpha result in significant inhibition of seizures in mice, and this action is mediated by neuronal p75 receptors. This evidence highlights a novel function of TNF-alpha in brain and indicates a new system for anticonvulsive intervention.

Contribution of Transmembrane Tumor Necrosis Factor to Host Defense against Mycobacterium bovis Bacillus Calmette-Guerin and Mycobacterium tuberculosis Infections

To study the specific role of transmembrane tumor necrosis factor (TmTNF) in host defense mechanisms against bacillus Calmette-Guerin (BCG) and Mycobacterium tuberculosis infections, we compared the immune responses of TNF/lymphotoxin (LT)-alpha(-/-) mice expressing a noncleavable transgenic TmTNF (TmTNF tg) to those of TNF/LT-alpha(-/-) and wild-type mice. Susceptibility of TNF/LT-alpha(-/-) mice to BCG infection was associated with impaired induction of systemic RANTES but not of monocyte chemoattractant protein 1 (MCP-1), the development of excessive local and systemic Th1-type immune responses, and a substantially reduced inducible nitric oxide synthase (iNOS) activity. Resistance of TmTNF tg mice to BCG infection was associated with efficient activation of iNOS in granulomas and with the regulated release of local and systemic chemokines and Th1-type cytokines. However, M. tuberculosis infection of TmTNF tg mice resulted in longer survival and enhanced resistance compared to TNF/LT-alpha(-/-) mice but higher sensitivity than wild-type mice. TmTNF tg mice exhibited reduced pulmonary iNOS expression and showed an exacerbated cellular infiltration in the lungs despite a modest bacillary content. Our data thus indicate a role for TmTNF in host defense against mycobacteria by contributing to induction and regulation of Th1-type cytokine and chemokine expression leading to development of bactericidal granulomas expressing iNOS, which critically determines susceptibility versus resistance of the host to mycobacterial infections.

Evaluating the role of tumor necrosis factor-alpha in experimental pulmonary tuberculosis in the guinea pig

Tumor necrosis factor-alpha (TNF-alpha) is suggested to play multiple roles in immune and pathologic responses in tuberculosis. In this study, we have developed a system for the expression of recombinant guinea pig TNF-alpha (rgpTNF-alpha). Using rgpTNF-alpha along with neutralizing anti-rgpTNF-alpha antiserum, we tested the effect of modulating the levels of TNF-alpha on antigen-specific T cell proliferation in splenocytes. By neutralizing TNF-alpha in the supernatant of PPD-pulsed splenocytes with anti-rgpTNF-alpha, we observed hyperproliferation. Conversely, the addition of rgpTNF-alpha resulted in a significant suppression of PPD-induced lymphoproliferation. In addition, when unvaccinated and BCG-vaccinated guinea pigs were treated with polyclonal rgpTNF-alpha antiserum throughout the first 3 weeks following low-dose, pulmonary infection with Mycobacterium tuberculosis H37Rv, we observed splenomegaly in BCG-vaccinated guinea pigs. We also detected higher levels of splenic granuloma organization in the non-vaccinated group as well as a significant number of plasma cells associated with granulomata from the BCG-vaccinated group. These results suggest that modulating the availability of TNF-alpha in BCG-vaccinated guinea pigs can lead to immuno-dysregulation and, perhaps, the inappropriate enhancement of humoral immunity. Conversely, abrogating TNF-alpha activity in the context of a hyperinflammatory response in non-vaccinated guinea pigs may, in fact, rescue them from immunopathological consequences of overproducing TNF-alpha.

Essential role of tumor necrosis factor-alpha in the induction of hepatitis B virus (HBV) specific cytotoxic T lymphocyte: Senji Kasahara, Kazuki Ando, Kuniaki Saito, Gifu University School of Medicine, Gifu Japan; Kenji Sekikawa, National Institute of Animal Health, Tsukuba Japan; Mitsuru Seishima, Hisataka Moriwaki, Gifu University School of Medicine, Gifu Japan

Essential role of tumor necrosis factor-alpha in the induction of hepatitis B virus (HBV) specific cytotoxic T lymphocyte: Senji Kasahara, Kazuki Ando, Kuniaki Saito, Gifu University School of Medicine, Gifu Japan; Kenji Sekikawa, National Institute of Animal Health, Tsukuba Japan; Mitsuru Seishima, Hisataka Moriwaki, Gifu University School of Medicine, Gifu Japan

Immunohistochemical analysis of TNF-alpha and HSP-60 in women with tubal factor infertility associated with Chlamydia trachomatis.

To explore the roles of tumor necrosis factor-alpha (TNF-alpha) and heat shock protein 60 (HSP-60) in women with tubal factor infertility (TFI) associated with Chlamydia trachomatis, and to determine the mechanisms of fallopian adhesions in Chlamydia trachomatis (CT) infections, the expressions of TNF-alpha and HSP-60 were quantitatively determined in 60 cases of TFI and 30 controls by immunohistochemical technique. The patients with TFI were further divided into group A and group B according to the CT-DNA of cervical specimens of PCR. The quantitative analysis was conducted by employing computerized image analysis system. It is found that the expressions of TNF-alpha and HSP-60 were much higher in TFI patients than those of controls. Among CT-HSP responders, a stronger expression was correlated with more severe salpingeal pathology. It is concluded that TNF-alpha and HSP-60 play very important roles in fallopian tube adhesion and occlusion in TFI due to CT infection.

The role of tumour necrosis factor‐α in renal dysfunction following mild haemorrhage in rats

Mild haemorrhage occasionally causes delayed death following failure of kidney or multiple organs, but the precise mechanisms have not yet been identified. We investigated the role of tumour necrosis factor-alpha (TNF-alpha), known as a major pro-inflammatory cytokine that leads to multiple organ failure, on the renal damage induced by mild haemorrhage. A mild haemorrhagic state was induced in male anaesthetized rats by bleeding via a common carotid catheter for 20 min at 16.7% of total body blood, 1.09 ml/100 g body weight, without fluid resuscitation. Mean arterial pressure and heart rate decreased soon after haemorrhaging but returned to baseline level up to 5 h after bleeding. TNF-alpha mRNA expression in the kidney and serum TNF-alpha levels were highest at 1 h after bleeding. Intraperitoneal pretreatment with FR167653, an inhibitory compound of TNF-alpha production, as well as of interleukin (IL)-1beta, significantly inhibited the increase in TNF-alpha. The inflammatory cell infiltration and tubular cell injury induced by haemorrhage were suppressed, and the renal dysfunction was dramatically improved by the FR167653 treatment. The morphological changes were also less in the treated group than in those that had not been treated. TNF-alpha has been reported to have striking effects on IL-1beta release and activation of neutrophils, and to play a pivotal role in the expression of the other pro-inflammatory cytokines. Our data show that endogenously-derived TNF-alpha does play a key role in the renal dysfunction during mild haemorrhage. These results should be useful to forensic pathologists to explain the pathogenesis of renal dysfunction induced by a mild haemorrhage and to identify the cause of death where there are no significant morphological changes after mild haemorrhage.

Inflammatory cytokines in acute renal failure

A growing body of evidence indicates that inflammatory mechanisms contribute to toxin-induced acute renal failure as well as ischemia/reperfusion injury. A role for tumor necrosis factor-alpha (TNF-alpha) in mediating the inflammatory injury in cisplatin-induced acute renal failure has recently been established. Cisplatin induces the expression of TNF-alpha and TNF receptor subtype 2 (TNFR2) within the kidney. Genetic deletion of either TNF-alpha or TNFR2 substantially reduces cisplatin-induced renal failure and also necrosis and apoptosis within the kidney. Studies will be required to determine if pharmacologic inhibition of TNF-alpha might reduce cisplatin-induced renal failure in humans.

Obliterative airway disease in tracheal allografts requires tumor necrosis factor alpha

Abstract Introduction: Obliterative Bronchiolitis (OB), characterized by epithelial denudation, inflammation, and fibrosis of small airways, is the major complication affecting long-term lung and heart-lung transplant recipients. Tumor necrosis factor-alpha (TNF-a) is known to mediate and promote inflammation and fibrosis in chronic lung injury. This study clarified the role of TNF-a in an established obliterative bronchiolitis model. Methods: Rat tracheas and mainstem bronchi were transplanted from MHC-disparate Brown-Norway donors into Lewis recipients, and explanted on days 7 & 14. Treated groups received a TNF-a translational inhibitor, RDP-58, beginning immediately post-transplant or on day-7 once luminal occlusion had partially developed. Computerized morphometry assessed epithelial denudation and luminal obliteration. Separate tracheal specimens were processed for TNF-a mRNA expression by RQRT-PCR, and protein localization and expression by immunohistochemistry. EMSAs evaluated NFkB activation. Results: 14-day control allografts averaged 58% occlusion and 98% epithelial denudation. These parameters were significantly improved with TNF-a inhibition, averaging 34% luminal obliteration (p Conclusions: TNF-a promotes obliterative airway disease in tracheal allografts via an NFkB-dependent mechanism. TNF inhibition may even be beneficial in ameliorating disease progression after patients present with established lesions.

Tumour necrosis factor-alpha receptor 1 and 2 polymorphisms in inflammatory bowel disease and their association with response to infliximab.

The role of tumour necrosis factor-alpha in the pathogenesis of inflammatory bowel disorders is well-known and is underscored by the effectiveness of antitumour necrosis factor-alpha treatment. Tumour necrosis factor-alpha exerts its effect by binding TNFR1 and TNFR2, which genes map to inflammatory bowel disorders susceptibility loci. Since TNFR1 and TNFR2 are good candidate genes for inflammatory bowel disorders, we studied the functional TNFR2T587G and the TNFR1A36G mutation in 344 Crohn's disease and 152 ulcerative colitis patients and investigated the relation with disease phenotypes. An association with response to infliximab was evaluated in 166 Crohn's disease patients. The TNFR2 587G allele was more frequent in ulcerative colitis compared with controls (P = 0.03). Both single nucleotide polymorphisms were negatively associated with smoking at diagnosis in Crohn's disease (TNFR1A36G odds ratio: 0.614, 95% confidence interval: 0.452, 0.99 and TNFR2T587G odds ratio: 0.572, 95% confidence interval: 0.820, 0.875). There was a positive association between pancolitis and the TNFR1A36G polymorphism in ulcerative colitis (odds ratio: 5.341, 95% confidence interval: 1.484, 19.39). The biological response to infliximab was lower in patients carrying TNFR1 36G (odds ratio: 0.47, 95% confidence interval: 0.234, 0.946). The TNFR2 587G allele was more frequent in ulcerative colitis. Both single nucleotide polymorphisms were negatively associated with smoking in Crohn's disease. A relation between TNFR1A36G and pancolitis was found in ulcerative colitis. There was no clear effect of the polymorphisms on infliximab response although, the TNFR1 minor was associated with a lower response to infliximab.

Norepinephrine induces apoptosis in neonatal rat cardiomyocytes through a reactive oxygen species-TNF alpha-caspase signaling pathway.

Norepinephrine (NE)-induced apoptosis in cardiomyocytes is an important cause of heart failure. Previous studies revealed that reactive oxygen species (ROS) are involved in apoptosis. Tumor necrosis factor-alpha (TNF), a well-known mediator that stimulates apoptosis, is not only produced by macrophages but also by cardiomyocytes. Until now, the role of TNF and its relationship to ROS in NE-induced apoptosis of cardiomyocytes have never been investigated. Neonatal rat cardiomyocytes were treated with various concentrations of NE. Apoptosis of cardiomyocytes was determined using the TUNEL assay. The level of secreted TNF was measured by ELISA and TNF mRNA expression was determined by semiquantitative reverse transcriptional polymerase chain reaction. Caspase activity was measured by a fluorogenic protease assay kit. Anti-TNF antibodies, caspase inhibitors and antioxidants (N-acetyl-L-cysteine or vitamin C) were added to determine if they could inhibit the apoptotic effect of NE. NE induced apoptosis of cardiomyocytes in a dose- and time-dependent manner. NE up-regulated TNF mRNA expression and increased TNF secretion and caspase-2,-3,-6, and -9 activities. A neutralizing anti-TNF antibody and caspase-2 and -3 inhibitors significantly attenuated NE-induced apoptosis. Antioxidants completely abrogated NE-induced TNF secretion, caspase activation, and apoptotic death. NE induced apoptosis in neonatal rat cardiomyocytes through a ROS-TNF-caspase signaling pathway.

Tumor necrosis factor alpha as an endogenous stimulator for circulating coupling factor 6.

We recently showed that mitochondrial coupling factor 6 (CF6) is present as a pressor substance and a prostacyclin inhibitor in systemic circulation. However, the regulation mechanism for circulating CF6 is unknown. We investigated the role of tumor necrosis factor-alpha (TNF-alpha) in the generation and release of CF6. We used two kinds of cells, human umbilical vein endothelial cells (HUVEC) and ECV-304. The concentration of CF6 in the medium increased with time in both ECV-304 and HUVEC. Treatment of ECV-304 and HUVEC with TNF-alpha enhanced the release of CF6 in a dose-dependent manner concomitantly with the decrease in CF6 content in the mitochondria at 24 h. The released CF6 was characterized to be an active full-length peptide by Western blot. The ratio of CF6 to GAPDH mRNA, measured by real-time polymerase chain reaction, was 1.7 fold increased at 1 h after exposure to TNF-alpha in ECV-304 and HUVEC. This enhanced gene expression and release was blocked or suppressed by 70% by stable transfection of dominant negative mutant I kappa B kinase alpha whose efficacy was confirmed by blockade of translocation of NF-kappa B p65 protein and of degradation of I kappa B alpha protein. Flow cytometry analysis revealed that the cell surface-associated CF6 was significantly increased at 24 h after TNF-alpha in a dose-dependent manner. TNF-alpha stimulates the gene expression of CF6 via activation of NF-kappa B signaling pathway, and promotes the release of CF6 from ECV-304 and HUVEC.

Prevention of hepatopulmonary syndrome and hyperdynamic state by pentoxifylline in cirrhotic rats.

Inhibition of tumour necrosis factor-alpha (TNF-alpha), levels of which are increased in the blood of cirrhotic rats, prevents hyperdynamic circulatory state, mainly by decreasing the vascular overproduction of nitric oxide. Hepatopulmonary syndrome, which is characterised by intrapulmonary vascular dilatation and increased alveolar to arterial oxygen tension difference (PA-a,O2), is mainly related to pulmonary over-production of NO by macrophages accumulated in lung vessels. Since TNF-alpha is a potent activator of macrophagic inducible nitric oxide synthase (NOS), the aim of this study was to investigate whether TNF-alpha inhibition prevented hepatopulmonary syndrome and hyperdynamic circulatory state in rats with cirrhosis. TNF-alpha was inhibited by 5 weeks of pentoxifylline (10 mg x kg body weigh(-1) x day(-1)) in rats with cirrhosis induced by common bile duct ligation. Cardiac output, pulmonary and systemic vascular resistance, PA-a,O2 and cerebral uptake of intravenous technetium-99m-labelled albumin macroaggregates (which reflects intrapulmonary vascular dilatation) were similar in sham- and pentoxifylline-treated cirrhotic rats. Blood TNF-alpha concentrations and pulmonary intravascular macrophage sequestration, as assessed by morphometric analysis and radioactive colloid uptake, were decreased with pentoxifylline. Pentoxifylline also prevented increases in aorta and lung NOS activities and inducible NOS expression. Thus pentoxifylline prevents development of hyperdynamic circulatory state and hepatopulmonary syndrome, probably by inhibiting the effects of tumour necrosis factor-alpha on vascular nitric oxide synthase and intravascular macrophages. These results support an important role for tumour necrosis factor-alpha in the genesis of hepatopulmonary syndrome.

Role of tumor necrosis factor-alpha in methamphetamine-induced drug dependence and neurotoxicity.

Tumor necrosis factor-alpha (TNF-alpha), a proinflammatory cytokine, is now emerging as an important modulator of the function of the CNS. Methamphetamine (METH) is a widely abused psychostimulant that causes euphoria, hyperactivity, and drug dependence. High doses of METH cause long-term neurotoxicity in dopaminergic neurons. In this study, we investigated a role of TNF-alpha in METH-induced dependence and neurotoxicity. Repeated treatment with METH (2 mg/kg for 5 d) in rats induced a significant increase in TNF-alpha mRNA and protein expression in the brain. Exogenous TNF-alpha (1-4 microg) blocked locomotor-stimulating and rewarding effects of METH, as well as METH (4 mg/kg; four times at 2 hr intervals)-induced dopaminergic neurotoxicity in mice. To examine a role of endogenous TNF-alpha in behavioral and neurochemical effects of METH, we used mice with targeted deletions of the TNF-alpha gene. TNF-alpha-(-/-) mice showed enhanced responses to the locomotor-sensitizing, rewarding, and neurotoxic effects of METH compared with wild-type mice. We also examined the role of TNF-alpha in METH-induced dopamine (DA) release and uptake in vitro and in vivo in C57BL/6 mice. Exogenous TNF-alpha (4 microg) attenuated the METH-induced increase in extracellular striatal DA in vivo and potentiated striatal DA uptake into synaptosomes in vitro and in vivo. Furthermore, TNF-alpha activated vesicular DA uptake by itself and diminished the METH-induced decrease in vesicular DA uptake. Our findings suggest that TNF-alpha plays a neuroprotective role in METH-induced drug dependence and neurotoxicity by activating plasmalemmal and vesicular DA transporter as well as inhibiting METH-induced increase in extracellular DA levels.

Involvement of tumor necrosis factor-alpha and interleukin-8 in antigen-induced arthritis of the rabbit temporomandibular joint.

In temporomandibular joint (TMJ) arthritis, knowledge is limited about the source of the inflammatory mediators. The aim of this study was to investigate the immunohistochemical role of tumor necrosis factor-alpha (TNF-alpha) and interleukin-8 (IL-8) in the development of the antigen-induced arthritis of the rabbit TMJ. Unilateral TMJ arthritis was induced in 28 adult rabbits. From 6 h to 6 weeks after induction of arthritis, the topology of TNF-alpha and IL-8 was observed. Positive reaction for TNF-alpha of synovial cells was observed within 3 days after induction and at 3 weeks after induction. TNF-alpha positive vascular endothelial cells and chondrocytes were identified throughout the observation period. IL-8 was detected only during the acute stage. The cytokines TNF-alpha and IL-8 were observed in specific cells depending on the stage. TNF-alpha was particularly related with angiogenesis and cartilage destruction and IL-8 was involved in the acute stage of inflammation.

The role of tumor necrosis factor-alpha and interleukin-1 in the mammalian testis and their involvement in testicular torsion and autoimmune orchitis

This review will focus the roles of TNF-alpha, IL-1 alpha, and IL-1 beta in the mammalian testis and in two testicular pathologies, testicular torsion and orchitis. TNF alpha in the testis is produced by round spermatids, pachytene spermatocytes, and testicular macrophages. The type 1 TNF receptor has been found on Sertoli and Leydig cells and numerous studies suggest a paracrine mode of action for TNF alpha in the normal testis. IL-1 alpha has been reported to be produced by Sertoli cells, testicular macrophages, and possibly postmeiotic germ cells. IL-1 receptors have been reported on Sertoli cells, Leydig cells, testicular macrophages, and germ cells suggesting both autocrine and paracrine functions. While these proinflammatory cytokines have important roles in normal testicular homeostasis, an elevation of their expression can lead to testicular dysfunctions. Testicular torsion is a clinical pathology with results in testicular ischemia and surgical intervention is often required for reperfusion. A pivotal role for IL-1beta in the pathology of testicular torsion has been recently described whereby an increase in IL-1beta production after reperfusion of the testis is correlated with the activation of the stress-related kinase, c-jun N-terminal kinase, and ultimately resulting in neutrophil recruitment to the testis and germ cell apoptosis. In autoimmune orchitis, on the other hand, TNF alpha produced by T-lymphocytes and macrophages of the testis has been implicated in the development and progression of the disease. Thus, both proinflammatory cytokines, TNF alpha and IL-1, have significant roles in normal testicular functions as well as in certain testicular pathologies.

Expression of leucocyte-endothelial adhesion molecules is limited to intercellular adhesion molecule-1 (ICAM-1) in the lung of pneumoconiotic patients: role of tumour necrosis factor-alpha (TNF-alpha).

Coal workers' pneumoconiosis (CWP) is characterized by a chronic inflammatory lung reaction associated with macrophage accumulation in alveolar spaces. In this study, we investigated in CWP the implication of adhesion molecules such as E-selectin, ICAM-1 and vascular cell adhesion molecule-1 (VCAM-1) and the role of TNF-alpha which is one of the cytokines inducing their expression. Adhesion molecule expression was analysed by immunohistochemistry on lung biopsies from patients with CWP and from healthy subjects. In parallel, soluble adhesion molecules were detected in bronchoalveolar lavage fluids (BALF) from patients by specific ELISA. The involvement of TNF in the induction of these adhesion molecules was measured (i) by immunohistochemistry on sections from lung fragments, and (ii) by evaluating in vitro the expression of adhesion molecules on endothelial cells and on alveolar epithelial cells in the presence of alveolar macrophage supernatants. In control subjects, a weak staining of ICAM-1 was detected only in alveolar walls, while E-selectin and VCAM-1 were undetectable. In pneumoconiotic patients, ICAM-1 was expressed at a high level by endothelium, by alveolar and bronchial epithelial cells and by alveolar macrophages. E-selectin and VCAM-1 expression remained undetectable. Measurement of soluble adhesion molecule showed that only the concentration of sICAM-1 was significantly increased in BALF from patients with CWP compared with controls. The involvement of TNF in this ICAM-1 expression was shown by the in vitro effect of alveolar macrophage supernatants on adhesion molecule expression by endothelial cells and epithelial cells (this effect was neutralized by anti-TNF antibodies) and by the increased production of TNF in the lung of pneumoconiotic patients. These data provide evidence for the involvement of ICAM-1, induced at least in part by alveolar macrophage-derived TNF, in the development of the inflammatory reaction in CWP.

Role of tumour necrosis factor-alpha (TNF-alpha) in the induction of HIV-1 gp120-mediated CD4+ T cell anergy.

The HIV-1 envelope glycoprotein (gp 120) is known to induce antigen-specific and non-specific CD4+ T cell anergy. We found that early T cell activation, as indicated by HLA-DP expression in the early G1 (G1A) phase of the cell cycle, and the inhibition of mitogen-mediated IL-2 production induced by gp120, required TNF-alpha produced by gp120-stimulated macrophages. In the presence of an antibody to TNF-alpha, these changes induced by gp120 were inhibited, while recombinant TNF-alpha induced similar abnormalities of CD4+ T cells, even in the absence of gp120. On the other hand, inhibition of the mixed lymphocyte reaction (MLR) in CD4+ T cells by gp120, which may be related to gp120-mediated down-regulation of CD4 expression on T cells and activation of protein tyrosine kinase p56(lck) in CD4+ T cells, was observed even in the absence of macrophage-derived TNF-alpha induced by gp120. These observations indicate that both TNF-alpha-dependent and independent events contribute to gp120-mediated CD4+ T cell anergy, and TNF-alpha appears to play an important role in inducing CD4+ T cell anergy in HIV-1 infection.

Polymorphisms in the promoter of the TNFa gene are not associated with recurrent pregnancy loss and TNFa serum levels in a Caucasian population of 380 women

Objective: The tumor necrosis factor alpha (TNFa) gene plays an important role in immunology and inflammation. Conflicting results have been obtained regarding the role of TNFa during pregnancy and recurrent pregnancy loss (RPL). This study investigated the relationship between RPL and two polymorphisms in the promoter of the TNF-a gene (TNFa-308 and TNFa-863).Materials and Methods: Genotyping was performed in 168 RPL women and 212 ethnically matched healthy individuals. In addition, we performed analysis of TNFa serum protein levels.Results: TNFa-308 allele frequencies among women with IRM and controls were 88.9% and 88.4%, respectively, for allele-308G (wild type), and 11.1% and 11.6%, respectively, for allele-308A (mutant). No association between allele-308A and the occurrence of IRM was found (p = 0.9, odds ratio [OR] 0.9; confidence interval [CI] 0.38-2.30). Genotype frequencies were also not significantly different between the study and the control groups. The mean TNFa serum levels in women with IRM and controls were 12.84 (SD±3.1) pg/mL and 14.97 (SD±5.47) pg/mL (p = 0.2).Conclusion: We conclude that genetic factors are not a major determinant of early pregnancy failure and TNF-a cytokine production. Objective: The tumor necrosis factor alpha (TNFa) gene plays an important role in immunology and inflammation. Conflicting results have been obtained regarding the role of TNFa during pregnancy and recurrent pregnancy loss (RPL). This study investigated the relationship between RPL and two polymorphisms in the promoter of the TNF-a gene (TNFa-308 and TNFa-863). Materials and Methods: Genotyping was performed in 168 RPL women and 212 ethnically matched healthy individuals. In addition, we performed analysis of TNFa serum protein levels. Results: TNFa-308 allele frequencies among women with IRM and controls were 88.9% and 88.4%, respectively, for allele-308G (wild type), and 11.1% and 11.6%, respectively, for allele-308A (mutant). No association between allele-308A and the occurrence of IRM was found (p = 0.9, odds ratio [OR] 0.9; confidence interval [CI] 0.38-2.30). Genotype frequencies were also not significantly different between the study and the control groups. The mean TNFa serum levels in women with IRM and controls were 12.84 (SD±3.1) pg/mL and 14.97 (SD±5.47) pg/mL (p = 0.2). Conclusion: We conclude that genetic factors are not a major determinant of early pregnancy failure and TNF-a cytokine production.

Tumor Necrosis Factor-α Induces Nuclear Factor-κB-dependent TRPC1 Expression in Endothelial Cells

We investigated the role of tumor necrosis factor-alpha (TNF-alpha) in activating the store-operated Ca2+ channels in endothelial cells via the expression of transient receptor potential channel (TRPC) isoforms. We observed that TNF-alpha exposure of human umbilical vein endothelial cells resulted in TRPC1 mRNA and protein expression, whereas it had no effect on TRPC3, TRPC4, or TRPC5 expression. The TRPC1 expression was associated with increased Ca2+ influx after intracellular Ca2+ store depletion with either thrombin or thapsigargin. We cloned the 5'-regulatory region of the human TRPC1 (hTRPC1) gene which contained a TATA box and CCAAT sequence close to the transcription initiation site. We also identified four nuclear factor-kappaB (NF-kappaB)-binding sites in the 5'-regulatory region. To address the contribution of NF-kappaB in the mechanism of TRPC1 expression, we determined the effects of TNF-alpha on expression of the reporter luciferase after transfection of hTRPC1 promoter-luciferase (hTRPC1-Pro-Luc) construct in the human dermal microvascular endothelial cell line. Reporter activity increased >4-fold at 4 h after TNF-alpha challenge. TNF-alpha-induced increase in reporter activity was markedly reduced by co-expression of either kinase-defective IKKbeta kinase mutant or non-phosphorylatable IkappaB mutant. Treatment with NEMO-binding domain peptide, which prevents NF-kappaB activation by selectively inhibiting IKKgamma interaction with IKK complex, also blocked the TNF-alpha-induced TRPC1 expression. Thus, TNF-alpha induces TRPC1 expression through an NF-kappaB-dependent pathway in endothelial cells, which can trigger augmented Ca2+ entry following Ca2+ store depletion. The augmented Ca2+ entry secondary to TRPC1 expression may be an important mechanism of endothelial injury induced by TNF-alpha.

L-carnitine alleviates alcohol-induced liver damage in rats: role of tumour necrosis factor-alpha.

Excessive alcohol intake induces hepatic fatty infiltration, which has been suggested to sensitize the liver to further damage. To test this hypothesis, L-carnitine, a constitutional lipotropic compound, was administered to rats chronically treated with ethanol by liquid diet feeding for 10 weeks. Ethanol administration caused marked steatosis, mild inflammation and elevated plasma alanine aminotransferase and tumour necrosis factor alpha (TNF-alpha) concentrations. Dietary supplementation with L-carnitine significantly reduced all these parameters as well as the hepatic concentration of thiobarbituric acid reactive substances, an indicator of lipid peroxidation products. Pretreatment with L-carnitine also significantly blunted ethanol-induced stimulation of TNF-alpha release by isolated Kupffer cells. This study provides direct support for the notion that steatosis sensitizes the liver to further damage and suggests an involvement of TNF-alpha in this process.