TNF-alpha Protein Research Articles

Pioglitazone suppresses the lipopolysaccharide‐induced production of inflammatory factors in mouse macrophages by inactivating NF‐κB

TZDs (thiazolidinediones) are prescribed as anti-Type II diabetes drugs, but little is known regarding whether TZDs regulate the expression of sPLA2 (secretory phospholipase A2) in macrophages. We have investigated the effects of pioglitazone on LPS (lipopolysaccharide)-induced production of TNF-alpha (tumour necrosis factor alpha), sPLA2-V and -X (groups V and X sPLA2) in RAW 264.7 macrophages. TNF-alpha, sPLA2-V and -X mRNA and protein expression were determined by RT-PCR (reverse transcriptase-PCR) and Western blot analysis, respectively. The activity of NF-kappaB (nuclear factor kappaB) was determined by Western blot and confocal microscopy. LPS induced TNF-alpha, sPLA2-V and sPLA2-X mRNA and protein expression. Pretreatment with 10 mumol/l pioglitazone significantly suppressed LPS-induced TNF-alpha, sPLA2-V and sPLA2-X mRNA and protein expression. LPS induced NF-kappaB expression and translocation in the nucleus, but the inductive effects were inhibited by pioglitazone. Our findings indicate that pioglitazone inhibits production of inflammatory factors induced by LPS in murine macrophage cells by inactivating NF-kappaB. Pioglitazone appears to play an anti-inflammatory role in the atherosclerotic process.

Expression of 8-oxoguanine DNA glycosylase 1 (OGG1) and the level of p53 and TNF-alpha proteins in peripheral lymphocytes in patients with Alzheimer's disease

Expression of 8-oxoguanine DNA glycosylase 1 (OGG1) and the level of p53 and TNF-alpha proteins in peripheral lymphocytes in patients with Alzheimer's disease

Fragile X‐related protein FXR1 controls post‐transcriptional suppression of lipopolysaccharide‐induced tumour necrosis factor‐α production by transforming growth factor‐β1

Tumour necrosis factor-alpha (TNF-alpha) is a key mediator of inflammation in host defence against infection and in autoimmune disease. Its production is controlled post-transcriptionally by multiple RNA-binding proteins that interact with the TNF-alpha AU-rich element and regulate its expression; one of these is Fragile X mental retardation-related protein 1 (FXR1). The anti-inflammatory cytokine transforming growth factor-beta1 (TGF-beta1), which is involved in the homeostatic regulation of TNF-alpha, causes post-transcriptional suppression of lipopolysaccharide (LPS)-induced TNF-alpha production. We report here that this depends on FXR1. Using RAW 264.7 cells and bone marrow-derived macrophages (BMDMphi) stimulated with LPS and TGF-beta1, we show that TGF-beta1 inhibits TNF-alpha protein secretion, whereas TNF-alpha mRNA expression remains unchanged. This response is recapitulated by the 3'-UTR of TNF-alpha, which is known to bind FXR1. TGF-beta1 induces FXR1 with a pattern of expression distinct from that of tristetraprolin, T-cell intracellular antigen 1, or human antigen R. When FXR1 is knocked down, TGF-beta1 is no longer able to inhibit LPS-induced TNF-alpha protein production, and overexpression of FXR1 suppresses LPS-induced TNF-alpha protein production. Targeting the p38 mitogen-activated protein kinase pathway of LPS-treated cells with small molecule inhibitors can induce FXR1 protein and mRNA expression. In summary, TGF-beta1 opposes LPS-induced stabilization of TNF-alpha mRNA and reduces the amount of TNF-alpha protein, through induction of expression of the mRNA-binding protein FXR1.

Pharmacological pre‐ and post‐conditioning with the sphingosine‐1‐phosphate receptor modulator FTY720 after myocardial ischaemia–reperfusion

Our recent experiments demonstrated that the Sphingosine-1-phosphate (S1P) receptor agonist FTY720 (2-amino-2-[2-(4-octylphenyl)ethyl]-1,3-propanediol hydrochloride) improves recovery of function after myocardial ischaemia-reperfusion ex vivo. Therefore, we tested the hypothesis that pharmacological post-conditioning with FTY720 reduces infarct size after myocardial ischaemia-reperfusion in vivo. Myocardial ischaemia was induced in Wistar rats by ligation of the left coronary artery for 45 min. FTY720 (0.5 mg kg(-1)) was applied i.p. either once, before reperfusion, or twice, 24 h before myocardial ischaemia and before reperfusion. After 24 h reperfusion, we determined infarct size by triphenyltetrazolium chloride staining and granulocyte infiltration by immunohistochemistry. Tumour necrosis factor-alpha (TNF)-alpha concentration was determined by elisa. S1P receptor expression was studied by Western blot. Calcium transients were evaluated in Indo-1-loaded cardiomyocytes. In both groups, FTY720 significantly reduced lymphocyte count in peripheral blood. FTY720 treatment attenuated granulocyte infiltration and TNF-alpha protein expression in reperfused myocardium. However, both treatment regimens were not able to reduce infarct size. FTY720 increased mortality due to induction of fatal ventricular tachyarrhythmias when administered once before reperfusion, but protected against reperfusion arrhythmias when given 24 h prior to ischaemia. Pretreatment selectively down-regulated S1P(1) receptor expression within the myocardium. S1P receptor agonists did not induce calcium deregulation in cardiomyocytes. FTY720 applied during reperfusion did not reduce infarct size but increased mortality during myocardial ischaemia-reperfusion due to induction of arrhythmias. Pretreatment with FTY720 before ischaemia abrogated the deleterious pro-arrhythmic effects without reducing infarct size.

Methylglyoxal modulates immune responses: relevance to diabetes

Increased methylglyoxal (MG) concentrations and formation of advanced glycation end-products (AGEs) are major pathways of glycaemic damage in diabetes, leading to vascular and neuronal complications. Diabetes patients also suffer increased susceptibility to many common infections, the underlying causes of which remain elusive. We hypothesized that immune glycation damage may account for this increased susceptibility. We previously showed that the reaction mixture (RM) for MG glycation of peptide blocks up regulation of CD83 in myeloid cells and inhibits primary stimulation of T cells. Here, we continue to investigate immune glycation damage, assessing surface and intracellular cytokine protein expression by flow cytometry, T-cell proliferation using a carboxyfluorescein succinimidyl ester assay, and mRNA levels by RT-PCR. We show that the immunomodulatory component of this RM was MG itself, with MG alone causing equivalent block of CD83 and loss of primary stimulation. Block of CD83 expression could be reversed by MG scavenger N-acetyl cysteine. Further, MG within RM inhibited stimulated production of interleukin (IL)-10 protein from myeloid cells plus interferon (IFN)-γ and tumour necrosis factor (TNF)-α from T cells. Loss of IL-10 and IFN-γ was confirmed by RT-PCR analysis of mRNA, while TNF-α message was raised. Loss of TNF-α protein was also shown by ELISA of culture supernatants. In addition, MG reduced major histocompatibility complex (MHC) class I expression on the surface of myeloid cells and increased their propensity to apoptose. We conclude that MG is a potent suppressor of myeloid and T-cell immune function and may be a major player in diabetes-associated susceptibility to infection.

Cigu Xiaozhi pills's influence on lipid peroxidation and TNF-α expressionin liver tissues of rats with nonalcoholic steatohepatitis

To investigate the effect of Cigu Xiaozhi pills on expression of tumor necrosis factor alpha (TNF-alpha) in rat with nonalcoholic steatoheptatitis (NASH). Fifty male SD rats were divided randomly into five groups: the normal group (n = 10), the model group (n = 10), the high dosage group (n = 10) and the little dosage group of the treatment (n = 10), Dongbao Gantai group (n = 10). Rats in normal control group were fed with standard diet. Rats in other four groups were established models of nonalcoholic steatohepatitis and were treated simultaneously with traditional Chinese medicine and positive medicine. Histopathological changes in the liver were observed. The serum ALT, AST, TG, TC and hepatic MDA, SOD, GPX were detected histologically. The expression of TNF-alpha in the liver was determined using the immunohistochemical technique and RT-PCR. In model group, extensive adipose degeneration and inflammatory cell infiltration were found in the liver. In treatment groups steatohepatitis were markedly alleviated. Compared with model group, TG, TC, ALT, AST of the serum and MDA of liver tissue were reduced significantly, and SOD, GPX were increased significantly in treatment groups and positive control group (P < 0.05). TNF-alpha was not expressed almost in normal rat liver and was expressed highly in model rat liver. Compared with model group, the TNF-alpha mRNA and protein expression were significantly lower in liver of treatment groups and positive control group (P < 0.05). And the effects of high dosage group surpass than those of Dongbao Gantai group (P < 0.05). TNF-alpha plays an important role in NASH pathogenesis. Cigu Xiaozhi pills can effectively treat experimental nonalcoholic steatohepatitis in rats, and its mechanism may be associated with ameliorating hepatocellular steatosis, removing the free radicals and enhancing the capability of anti-oxidation and anti-inflammatory.

Abstract 4378: Aflatoxin B1 inhibits LPS-induced TNF-alpha production in RAW 264.7 cells

Abstract Aflatoxin B1 (AFB1), a potent hepatocarcinogen, is known to impair specific and nonspecific immune responses. Recent studies suggested that AFB1 affects macrophage functions. Macrophages play an important role in a host defense against tumors and bacteria. In addition, some macrophage products, including tumor necrosis alpha (TNF-alpha), can be cytotoxic. In the present study, we investigated the effect of aflatoxin B1 (AFB1) on LPS-induced TNF-alpha production by a mouse macrophage cell line, RAW 264.7 cells. Cells were pretreated with increasing concentrations of AFB1 for 2 hours followed by stimulation with lipopolysaccharide (LPS) for 6 h. The concentration of TNF-alpha released in the culture medium was assessed by enzyme-linked immunosorbent assay (ELISA). AFB1 at 1, 10, 50 or 100 µM reduced the production of TNF-alpha induced by LPS. Compared to vehicle control, there was a greater reduction of TNF-alpha production with increased concentration of AFB1 pretreatment and LPS stimulation. AFB1 at 100 µM decreased the synthesis of both TNF-alpha mRNA and protein in RAW 264.7 cells. To explore the mechanism of AFB1 activation by RAW 264.7 cells, we examined the expression of CYP3A4 and CYP3A11, two isoforms of cytochrome p450 known to activate AFB1. Our data indicated that CYP3A11 but not CYP3A4 was significantly expressed in RAW 264.7 cells. These results suggest that AFB1 pretreatment suppresses LPS-induced TNF production in RAW 264.7 cells. (Supported by NIH grant R01ES04139.) Citation Format: {Authors}. {Abstract title} [abstract]. In: Proceedings of the 101st Annual Meeting of the American Association for Cancer Research; 2010 Apr 17-21; Washington, DC. Philadelphia (PA): AACR; Cancer Res 2010;70(8 Suppl):Abstract nr 4378.

Chronic low frequency/low volume resistance training reduces pro-inflammatory cytokine protein levels and TLR4 mRNA in rat skeletal muscle

Skeletal muscle is the source of pro- and anti-inflammatory cytokines, and recently, it has been recognized as an important source of interleukin 6 (IL-6), a cytokine that exerts inhibitory effects on several pro-inflammatory cytokines. Although dynamic chronic resistance training has been shown to produce the known "repeated bout effect", which abolishes the acute muscle damage, performing of high-intensity resistance training has been regarded highly advisable, at least from the hypertrophy perspective. On the other hand, a more therapeutic, "non-damaging" resistance training program, mainly composed of concentric forces, low frequency/low volume of training, and the same exercise, could theoretically benefit the muscle when the main issue is to avoid muscle inflammation (as in the treatment of several "low-grade" inflammatory diseases) because the acute effect of each resistance exercise session could be diminished/avoided, at the same time that the muscle is still being overloaded in a concentric manner. However, the benefits of such "less demanding" resistance training schedule on the muscle inflammatory profile have never been investigated. Therefore, we assessed the protein expression of IL-6, TNF-alpha, IL-10, IL-10/TNF-alpha ratio, and HSP70 levels and mRNA expression of SCF(beta-TrCP), IL-15, and TLR-4 in the skeletal muscle of rats submitted to resistance training. Briefly, animals were randomly assigned to either a control group (S, n = 8) or a resistance-trained group (T, n = 7). Trained rats were exercised over a duration of 12 weeks (two times per day, two times per week). Detection of IL-6, TNF-alpha, IL-10, and HSP70 protein expression was carried out by western blotting and SCF(beta-TrCP) (SKP Cullin F-Box Protein Ligases), a class of enzymes involved in the ubiquitination of protein substrates to proteasomal degradation, IL-15, and TLR-4 by RT-PCR. Our results show a decreased expression of TNF-alpha and TLR4 mRNA (40 and 60%, respectively; p < 0.05) in the plantar muscle from trained, when compared with control rats. In conclusion, exercise training induced decreased TNF-alpha and TLR-4 expressions, resulting in a modified IL-10/TNF-alpha ratio in the skeletal muscle. These data show that, in healthy rats, 12-week resistance training, predominantly composed of concentric stimuli and low frequency/low volume schedule, down regulates skeletal muscle production of cytokines involved in the onset, maintenance, and regulation of inflammation.

Association between TNF-α and Entamoeba histolytica Diarrhea

An association between tumor necrosis factor alpha (TNF-alpha) and Entamoeba histolytica diarrhea was assessed in a cohort of 138 non-related Bangladeshi children who have been prospectively followed since 2001. Peripheral blood mononuclear cells (PBMCs) obtained at study entry were purified, cultured, and stimulated with soluble amebic antigen before cytokine measurement from supernatant. Higher levels of TNF-alpha were associated with increased risk of first (P = 0.01) and recurrent E. histolytica-related diarrheal episodes (P = 0.005). Children who developed E. histolytica diarrhea had significantly higher TNF-alpha protein levels than those who experienced asymptomatic E. histolytica infection (P value = 0.027) or no infection (P value = 0.017). Microarray studies performed using RNA isolated from acute and convalescent whole blood and colon biopsy samples revealed higher but non-significant TNF-alpha messenger RNA (mRNA) levels in subjects with acute E. histolytica diarrhea compared with convalescence. We conclude that there is an association between higher TNF-alpha production and E. histolytica diarrhea.

Contraction-induced changes in TNFα and Akt-mediated signalling are associated with increased myofibrillar protein in rat skeletal muscle

Resistance training results in skeletal muscle hypertrophy, but the molecular signalling mechanisms responsible for this altered phenotype are incompletely understood. We used a resistance training (RT) protocol consisting of three sessions [day 1 (d1), day 3 (d3), day 5 (d5)] separated by 48 h recovery (squat exercise, 4 sets x 10 repetitions, 3 min recovery) to determine early signalling responses to RT in rodent skeletal muscle. Six animals per group were killed 3 h after each resistance training session and 24 and 48 h after the last training session (d5). There was a robust increase in TNFalpha protein expression, and IKK(Ser180/181) and p38MAPK(Thr180/Tyr182) phosphorylation on d1 (P < 0.05), which abated with subsequent RT, returning to control levels by d5 for TNFalpha and IKK(Ser180/181). There was a trend for a decrease in MuRF-1 protein expression, 48 h following d5 of training (P = 0.08). Notably, muscle myofibrillar protein concentration was elevated compared to control 24 and 48 h following RT (P < 0.05). Akt(Ser473) and mTOR(Ser2448) phosphorylation were unchanged throughout RT. Phosphorylation of p70S6k(Thr389) increased 3 h post-exercise on d1, d3 and d5 (P < 0.05), whilst phosphorylation of S6(Ser235/236) increased on d1 and d3 (P < 0.05). Our results show a rapid attenuation of inflammatory signalling with repeated bouts of resistance exercise, concomitant with summation in translation initiation signalling in skeletal muscle. Indeed, the cumulative effect of these signalling events was associated with myofibrillar protein accretion, which likely contributes to the early adaptations in response to resistance training overload in the skeletal muscle.

Asthma control, adiposity, and adipokines among inner-city adolescents

There is an association between adiposity and asthma prevalence, but the relationship to asthma control is unclear. We sought to understand the relationships among adiposity, sex, and asthma control in inner-city adolescents with asthma. We prospectively followed 368 adolescents with moderate-to-severe asthma (ages 12-20 years) living in 10 urban areas for 1 year. Asthma symptoms and exacerbations were recorded, and pulmonary function and exhaled nitric oxide levels were measured every 6 weeks. Adiposity measures (body mass index [BMI] and dual-energy X-ray absorptiometric scans) were made, and blood was collected for measurement of allergy markers, adiponectin, leptin, TNF-alpha, IL-6, and C-reactive protein levels. More than 60% of female subjects and 50% of male subjects were above the 85th percentile of BMI for age. Higher BMI was associated with more symptom days (R = 0.18, P = .02) and exacerbations (R = 0.18, P = .06) among female subjects only. Adiponectin was inversely related to asthma symptoms (R = -0.18, P < .05) and exacerbations (R = -0.20, P < .05) and positively with FEV(1)/forced vital capacity ratio (R = 0.15, P < .05) in male subjects only independent of body size. There was no relationship between adiposity or adipokines and total IgE levels, blood eosinophil counts, and exhaled nitric oxide levels. Dual-energy X-ray absorptiometry provided little additional value in relating adiposity to asthma outcome in this population of adolescents. Adiposity is associated with poorer asthma control in female subjects. Adiponectin is associated with improved asthma control in male subjects.

Preconditioning with Inhalative Carbon Monoxide Protects Rat Retinal Ganglion Cells from Ischemia/Reperfusion Injury

PURPOSE. Retinal ischemia/reperfusion (I/R) injury damages retinal neurons. Carbon monoxide (CO) recently attracted attention as cytoprotective because of its anti-inflammatory and antiapoptotic effects. Rapid preconditioning of retinal neurons by inhaled CO before I/R injury may reduce inflammation and apoptosis in retinal ganglion cells (RGCs). METHODS. I/R injury was performed on the left eyes of rats (n = 8) with or without inhaled CO preconditioning (250 ppm) for 1 hour before ischemia. Densities of fluorogold-prelabeled RGCs were analyzed 7 days after injury in whole-mounts. Retinal tissue was further harvested to analyze protein expression of TNF-alpha, HSP-70, and mitogen-activated protein kinases (MAPKs) pERK1/2 and p-p38. DNA-binding activities of the transcription factors NF-kappaB, AP-1, CREB, and HSF-1 were determined to elucidate a possible pathway of neuroprotection. RESULTS. Seven days after I/R injury, RGC death decreased by 52% in the CO preconditioning group compared with controls receiving room air (P < 0.001). Similarly, CO inhalation resulted in attenuated caspase-3 activity and TNF-alpha protein expression. In contrast, HSP-70 protein expression was elevated in the retina after CO. CREB and HSF-1 showed CO-dependent regulation and p-p38 MAPK. CONCLUSIONS. Rapid preconditioning with CO mediates anti-inflammatory and antiapoptotic effects in retinal I/R injury, thus making it neuroprotective. Further studies are needed to evaluate whether CO posttreatment may represent a therapeutic option counteracting ischemic neuronal injury.

Peritoneal Inflammation in Pigs is Associated with Early Mitochondrial Dysfunction in Liver and Kidney

The objective of this study was to investigate early effects of peritoneal inflammation on the mitochondrial function in the vital organs, liver and kidney, and their relation to inflammatory and oxidative stress mediators. The study was performed on 14 domestic pigs. Peritoneal inflammation was induced in anesthetized pigs after a midline laparotomy by autologous feces. Fluid resuscitation maintained a MAP above 60 mmHg. Animals were sacrificed 12 h later, and tissue samples were obtained to determine mitochondrial function, mRNA levels of relevant genes [inducible NO synthase (iNOS), inducible HO (HO-1), tumor necrosis factor-alpha (TNF-alpha)], generation of reactive oxygen species (ROS), and HO-1 activity. We found impaired mitochondrial function in both liver and kidney, based on decreased state 3 respiration in the liver and increased states 2 and 4 respiration in the kidney at 12 h. This was accompanied by increased TNF-alpha protein in the blood and up-regulation of TNF-alpha mRNA in the liver. Free iron was elevated in the liver but not in the kidney. In the kidney, mitochondrial ROS production was increased. Nitric oxide levels in blood remained unchanged, corresponding to unchanged levels of iNOS mRNA expression in liver and kidney. Similarly, HO-1 mRNA and heme oxygenase (HO)-activity were unchanged. The inflammatory response in the absence of characteristic septic symptoms was not associated with morphological organ damage at this early time point. Peritoneal inflammation in pigs caused mitochondrial dysfunction in liver and kidney, preceding signs of organ damage. We did not find proof that mitochondrial dysfunction was due to increased levels of either nitric oxide (NO) or products of HO, but it was accompanied by increased levels of oxidative stress markers.

Direct Activation of TACE-Mediated Ectodomain Shedding by p38 MAP Kinase Regulates EGF Receptor-Dependent Cell Proliferation

Inflammatory stimuli activate ectodomain shedding of TNF-alpha, L-selectin, and other transmembrane proteins. We show that p38 MAP kinase, which is activated in response to inflammatory or stress signals, directly activates TACE, a membrane-associated metalloprotease that is also known as ADAM17 and effects shedding in response to growth factors and Erk MAP kinase activation. p38alpha MAP kinase interacts with the cytoplasmic domain of TACE and phosphorylates it on Thr(735), which is required for TACE-mediated ectodomain shedding. Activation of TACE by p38 MAP kinase results in the release of TGF-alpha family ligands, which activate EGF receptor signaling, leading to enhanced cell proliferation. Conversely, depletion of p38alpha MAP kinase activity suppresses EGF receptor signaling and downstream Erk MAP kinase signaling, as well as autocrine EGF receptor-dependent proliferation. Autocrine EGF receptor activation through TACE-mediated ectodomain shedding intimately links inflammation and cancer progression and may play a role in stress and conditions that relate to p38 MAP kinase activation.

Remote Ischemic Preconditioning by Hindlimb Occlusion Prevents Liver Ischemic/Reperfusion Injury

Hepatocellular injury caused by ischemia-reperfusion of the liver occurs in a number of clinical situations including major trauma, elective surgery of the liver, and liver transplantation. Several strategies have been used to prevent liver injury following ischemia-reperfusion (I/R). Among these, ischemic preconditioning has shown promise as a preventative approach. In this manuscript, we hypothesized that use of remote ischemic preconditioning by brief hindlimb ischemia might prevent liver dysfunction in a mouse model of liver I/R. C57/B mice were subjected to 60 minutes of partial liver ischemia with or without antecedent hindlimb vascular I/R. Blood was drawn for serum alanine aminotransferase levels at times following liver reperfusion. Liver inflammation was assessed by measuring serum and liver tumor necrosis factor (TNF)-alpha protein and mRNA. The role of toll-like receptor 4 (TLR4) in mediating protection was determined using the mouse strain HeJ, which has a mutated TLR4. Antecedent hindlimb ischemia (10 minutes) lessened I/R-induced elevation of serum alanine aminotransferase compared with untreated I/R animals. This protection correlated with a reduction in serum TNF-alpha protein levels as well as liver TNF-alpha mRNA and apoptosis. High Mobility Group-Box 1 (HMG-B1) levels in the blood were elevated after hindlimb ischemia and injection of HMG-B1 prior to liver recapitulated the protective effect of hindlimb occlusion. TLR4-mutant HeJ mice did not demonstrate protection with hindlimb preconditioning. Brief hindlimb occlusion prevents liver I/R injury. This effect appears to be related to release of HMG-B1 and is dependent on the presence of a functional TLR4. Remote ischemia preconditioning represents a novel approach to preventing distant organ injury.

Signaling pathways involved in LPS induced TNFalpha production in human adipocytes

BackgroundThe development of obesity has been linked to an inflammatory process, and the role of adipose tissue in the secretion of pro-inflammatory molecules such as IL-6 or TNFalpha has now been largely confirmed. Although TNFalpha secretion by adipose cells is probably induced, most notably by TLR ligands, the activation and secretion pathways of this cytokine are not yet entirely understood. Moreover, given that macrophagic infiltration is a characteristic of obesity, it is difficult to clearly establish the level of involvement of the different cellular types present within the adipose tissue during inflammation.MethodsPrimary cultures of human adipocytes and human peripheral blood mononuclear cells were used. Cells were treated with a pathogen-associated molecular pattern: LPS, with and without several kinase inhibitors. Western blot for p38 MAP Kinase was performed on cell lysates. TNFalpha mRNA was detected in cells by RT-PCR and TNFalpha protein was detected in supernatants by ELISA assays.ResultsWe show for the first time that the production of TNFalpha in mature human adipocytes is mainly dependent upon two pathways: NFkappaB and p38 MAP Kinase. Moreover, we demonstrate that the PI3Kinase pathway is clearly involved in the first step of the LPS-pathway. Lastly, we show that adipocytes are able to secrete a large amount of TNFalpha compared to macrophages.ConclusionThis study clearly demonstrates that the LPS induced activation pathway is an integral part of the inflammatory process linked to obesity, and that adipocytes are responsible for most of the secreted TNFalpha in inflamed adipose tissue, through TLR4 activation.

Tumor Necrosis Factor-alpha as a possible auto-/paracrine factor affecting estrous cycle in the cat uterus

Tumor Necrosis Factor-alpha (TNFalpha) is a pleiotrophic cytokine, affects either normal or tumor cells, and influences cellular differentiation. TNFalpha role in female reproduction has been proven to be mediated through an influence on prostaglandin (PGs) synthesis and output. To evaluate the possible role of TNFalpha in an auto-/paracrine regulation in the cat uterus, mRNA expression coding for TNFalpha and its receptors (TNFR1 and TNFR2), and TNFalpha protein content at different stages of the estrous cycle were investigated. Additionally, TNFalpha involvement in PG secretion at different stages of the estrous cycle was investigated by in vitro tissue culture. Gene expressions coding for TNFalpha and TNFR1 were the highest at diestrus (P < 0.05). TNFalpha protein expression was the lowest at interestrus (P < 0.05). Nevertheless, TNFR2 was not affected by the estrous stage. TNFalpha at a dose of 1 ng/ml significantly increased PGF2alpha secretion at estrus (P < 0.01) and PGE2 secretion at diestrus (P < 0.001) after 12h incubation. Overall findings indicate that TNFalpha locally produced in the cat's uterus, stimulates PG secretion in an estrous cycle-related manner.

Anti-Inflammatory Effect of Guan-Xin-Er-Hao via the Nuclear Factor-Kappa B Signaling Pathway in Rats with Acute Myocardial Infarction

A traditional Chinese medicine, Guan-Xin-Er-Hao (GXEH), is a famous multiple target therapeutic polypharmaceutical. Our aim was to evaluate whether or not oral administration of GXEH has an anti-inflammatory effect associated with blockade of nuclear factor-kappa B (NF-kappaB), and to investigate the NF-kappaB-mediated pro-inflammatory cytokines expression pathway during acute myocardial infarction (AMI) in rats. Sprague-Dawley rats were randomly assigned to four groups: oral GXEH administered at 15 or 5 g/kg, the vehicle control and sham-operated groups. Thirty minutes after giving GXEH or 0.9% NaCl (p.o.) once, coronary arteries were occluded except for the sham-operated rats. We measured 24-h infarct size, 3-h expression of NF-kappaB protein in the myocardial left ventricular tissues and serum levels of tumor necrosis factor-alpha (TNF-alpha), interleukin-6 (IL-6), and C-reactive protein (CRP). Compared with rats receiving vehicle, rats administered 15 g/kg GXEH had significantly reduced 24-h infarct size, expression of NF-kappaB protein and serum concentrations of TNF-alpha, IL-6, and CRP. GXEH at 5 g/kg did not have a significant effect on these parameters. In conclusion, GXEH exhibited an anti-inflammatory effect through inhibition of the NF-kappaB-mediated signaling pathway leading to downregulation of pro-inflammatory cytokine expression. These findings provide new evidence of the cardioprotective effect of GXEH through reduction of infarct size by mediating lots of endogenous materials via multiple pathways to act on myocardial cells in the treatment of cardiovascular disease.

Effect of probiotics on pro-inflammatory cytokines and NF-κB activation in ulcerative colitis

To demonstrate the therapeutic effect of probiotics in patients with ulcerative colitis (UC), and their effect on inflammatory mediators and nuclear factor (NF)-kappaB activation in these patients. Thirty patients with mild to moderate UC were randomly classified into two groups: sulfasalazine group, who received sulfasalazine 2400 mg/d; and probiotic group, who received sulfasalazine 2400 mg/d with probiotic. The patients were investigated before and after 8 wk of treatment with probiotic (Lactobacillus delbruekii and Lactobacillus fermentum). Colonic activity of myeloperoxidase (MPO) was assayed with UV spectrophotometry, the colonic content of interleukin (IL)-6 was determined by enzyme-linked immunosorbent assay (ELISA), fecal calprotectin was determined by ELISA, and expression of NF-kappaB p65 and tumor necrosis factor (TNF)-alpha proteins in colonic tissue was identified by immunohistochemistry and reverse transcription polymerase chain reaction, respectively. At the start of the study, colonic mucosal injury and inflammation were demonstrated in UC patients by hematoxylin and eosin staining, and an increase in colonic MPO activity, fecal calprotectin, and expression of colonic TNF-alpha and NF-kappaB p65 proteins. The use of probiotic for 8 wk significantly ameliorated the inflammation by decreasing the colonic concentration of IL-6, expression of TNF-alpha and NF-kappaB p65, leukocyte recruitment, as demonstrated by a decrease in colonic MPO activity, and the level of fecal calprotectin compared to sulfasalazine group and the control group (P < 0.05). Oral supplementation with probiotics could be helpful in maintaining remission and preventing relapse of UC.

The effect of non-steroidal anti-inflammatory agents on behavioural changes and cytokine production following systemic inflammation: Implications for a role of COX-1

Systemic inflammation gives rise to metabolic and behavioural changes, largely mediated by pro-inflammatory cytokines and prostaglandin production (PGE2) at the blood–brain barrier. Despite numerous studies, the exact biological pathways that give rise to these changes remains elusive. This study investigated the mechanisms underlying immune-to-brain communication following systemic inflammation using various anti-inflammatory agents.Mice were pre-treated with selective cyclo-oxygenase (COX) inhibitors, thromboxane synthase inhibitors or dexamethasone, followed by intra-peritoneal injection of lipopolysaccharide (LPS). Changes in body temperature, open-field activity, and burrowing were assessed and mRNA and/or protein levels of inflammatory mediators measured in serum and brain.LPS-induced systemic inflammation resulted in behavioural changes and increased production of IL-6, IL-1β and TNF-α, as well as PGE2 in serum and brain. Indomethacin and ibuprofen reversed the effect of LPS on behaviour without changing peripheral or central IL-6, IL-1β and TNF-α mRNA levels. In contrast, dexamethasone did not alter LPS-induced behavioural changes, despite complete inhibition of cytokine production. A selective COX-1 inhibitor, piroxicam, but not the selective COX-2 inhibitor, nimesulide, reversed the LPS-induced behavioural changes without affecting IL-6, IL-1β and TNF-α protein expression levels in the periphery or mRNA levels in the hippocampus.Our results suggest that the acute LPS-induced changes in burrowing and open-field activity depend on COX-1. We further show that COX-1 is not responsible for the induction of brain IL-6, IL-1β and TNF-α synthesis or LPS-induced hypothermia. Our results may have implications for novel therapeutic strategies to treat or prevent neurological diseases with an inflammatory component.