Cardiodepression by tumor necrosis factor-alpha

A 75-year-old man with carcinoma of the prostate presented with a pruritic, erythematous plaque involving the scrotal skin. Histological examination revealed extramammary Paget's disease. The intraepidermal tumour cells expressed prostate-specific antigen in keeping with a prostatic origin.     

Exogenous bovine surfactant suppresses tumor necrosis factor-alpha release by murine macrophages stimulated by genital mycoplasmas

Tumor necrosis factor-alpha (TNF-alpha) is a proinflammatory cytokine that appears to play a significant role in the development of neonatal chronic lung disease (CLD). Inflammation and CLD are also associated with respiratory tract colonization with genital mycoplasmas. The possible protective roles of surfactant in mitigating the inflammatory response to these microbes were investigated. Murine RAW 264.7 macrophages were preincubated with an exogenous surfactant and exposed overnight to sterile media, lipopolysaccharide (LPS), Mycoplasma hominis, or Ureaplasma urealyticum. Macrophages released TNF-alpha in response to challenge with LPS, U. urealyticum, and M. hominis in a concentration-dependent fashion. Surfactant suppressed LPS and M. hominis induced TNF-alpha production in a dose-dependent manner but suppressed U. urealyticum-mediated TNF-alpha production only at the higher dose tested. Similar effects were seen in hyperoxia (95% O2). Thus, exogenous bovine surfactant significantly inhibits the production of TNF-alpha by murine macrophages stimulated with genital mycoplasmas and bacterial LPS.     

Glia increase degeneration of hippocampal neurons through release of tumor necrosis factor-alpha

A multicenter, randomized, double-blind, crossover, placebo-controlled study was conducted in 90 isosorbide dinitrate responders showing stable angina to compare the efficacy of molsidomine retard, 8 mg b.i.d., with that of molsidomine, 4 mg t.i.d., for 6 weeks. Total work performance (workload x min) was significantly improved, compared with baseline and placebo until 8 and 12 h after molsidomine and molsidomine retard administration, respectively. ST-segment depression decreased significantly under the two treatments at 60 W as well as at maximal exercise. The rate-pressure product (heart rate x systolic blood pressure) decreased and increased significantly at submaximal and maximal exercise level, respectively. All these effects remained significant after 6-week treatment, with only the ST segment showing a nonsignificant tendency to improvement at maximal work. The frequency of anginal attacks and of sublingual nitroderivative-tablets consumption decreased significantly with molsidomine, 4 mg, and molsidomine retard, 8 mg. However, overall results showed that the latter form reduces myocardial ischemia more efficiently at submaximal exercise level, has a more prolonged effect on exercise tolerance, and maintains it at a somewhat higher level after 6-week treatment.     

Alveolar macrophage release of tumor necrosis factor-alpha in chronic alcoholics without liver disease

Alcohol is an immunosuppressive drug, and chronic abuse has been associated with increased susceptibility to a variety of infections, including bacterial pneumonia and tuberculosis. Alveolar macrophages are the resident phagocytes of the lung and play a central role in lung host defenses against infection ranging from direct antibacterial activity to the release of proinflammatory cytokines such as tumor necrosis factor-alpha (TNFalpha). TNFalpha, in particular, plays a key role in the development of the early inflammatory response. In this study, we investigated the effects of chronic alcohol consumption on alveolar macrophage release of TNFalpha in vitro. We prospectively studied lipopolysaccharide (LPS)-stimulated release of TNFalpha from alveolar macrophages obtained from bronchoalveolar lavage fluid (BALF) in 22 alcoholic (18 smokers, 4 nonsmokers) and 7 nondrinking healthy volunteers (3 smokers, 4 nonsmokers). The total number of cells recovered by bronchoalveolar lavage (BAL) and their differential distribution were not significantly different in alcoholics versus controls (43 +/- 8 x 10(6) and 39 +/- 13 x 10(6), respectively). However, the total number of cells recovered from BALF was significantly higher in smokers (51 +/- 8 x 10(6)) than in nonsmokers (19 +/- 5 x 10(6)). Spontaneous (basal) release of TNFalpha by alveolar macrophages was the same in alcoholics and controls. In contrast, LPS-stimulated release of TNFalpha was significantly suppressed in alcoholics compared with that of controls (1343 +/- 271 vs. 3806 +/- 926 U TNF/ml/10(6) cells, respectively, p < 0.015). When controlled for smoking, LPS-stimulated TNFalpha production was suppressed in alcoholic nonsmokers (563 +/- 413 U TNF/ml/10(6)) compared with control nonsmokers (5113 +/- 1264 U TNF/ml/10(6)). LPS-stimulated TNFalpha production was also less in control smokers (2063 +/- 386 U TNF/ml/10(6) cells) than in control nonsmokers (5113 +/- 1264 U TNF/ml/10(6) cells). There was no difference in TNFalpha production between smoking alcoholics and smoking control subjects. We conclude that chronic alcohol consumption significantly suppresses LPS-stimulated alveolar macrophage production of TNFalpha. This effect is obscured if the subject also smokes. Because TNFalpha production is an important element in host defense, this may explain, in part, the susceptibility of chronic alcohol abusers to a variety of infections.     

Expression and cleavage of tumor necrosis factor-alpha and tumor necrosis factor receptors by human monocytic cell lines upon direct contact with stimulated T cells

Tumor necrosis factor-alpha (TNF-alpha) is a potent cytokine in inflammatory processes. A variety of mechanisms that modulate its activity have been described, one being its binding to soluble receptors (sTNFR). In this study, we demonstrate that human monocytic cells such as THP-1 respond to direct contact with a membrane preparation of stimulated HUT-78 cells by producing TNF-alpha and by releasing sTNFR-p75, but not sTNFR-p55, with different kinetics. TNF-alpha concentration peaked after 12 h of contact and then decreased, whereas sTNFR-p75 production increased progressively upon cell/cell contact. The decrease in TNF-alpha concentration is not due to trapping of TNF-alpha by its soluble receptors or other soluble or cell-associated molecules, but rather to a proteolytic activity associated to THP-1 cells. On the other hand, the increase in sTNFR-p75 release does not result from an increase in the cleavage of pre-existing cell-associated sTNFR-p75 but from an increase in TNFR-p75 expression, immediately followed by the cleavage of its extracellular domain. Phenylmethylsulfonylfluoride, a serine protease inhibitor, has a negative effect on both TNF-alpha degradation and sTNFR-p75 release by THP-1 cells. Thus, there may be an enzymatic activity associated to THP-1 cells that plays an important role in the neutralization of TNF-alpha activity both by degrading the molecule and by cleaving its receptors at the cell surface.     

Feedback inhibition of macrophage tumor necrosis factor-alpha production by tristetraprolin

Tumor necrosis factor-alpha (TNF-alpha) is a major mediator of both acute and chronic inflammatory responses in many diseases. Tristetraprolin (TTP), the prototype of a class of Cys-Cys-Cys-His (CCCH) zinc finger proteins, inhibited TNF-alpha production from macrophages by destabilizing its messenger RNA. This effect appeared to result from direct TTP binding to the AU-rich element of the TNF-alpha messenger RNA. TTP is a cytosolic protein in these cells, and its biosynthesis was induced by the same agents that stimulate TNF-alpha production, including TNF-alpha itself. These findings identify TTP as a component of a negative feedback loop that interferes with TNF-alpha production by destabilizing its messenger RNA. This pathway represents a potential target for anti-TNF-alpha therapies.     

Induction and release of manganese superoxide dismutase caused by tumor necrosis factor-alpha from mitochondria in human umbilical vein endothelial cells

The effects of tumor necrosis factor-alpha (TNF-alpha) on cultured human umbilical vein endothelial cells (EC) and five cancer cell lines, A549, ME180, A2780, KURAMOCHI, and Hela, were compared. While A549, A2780, KURAMOCHI, and Hela cells were fairly resistant to the cytolytic effects of TNF-alpha, ME180 cells were sensitive. EC were also less sensitive to TNF-alpha than ME180 cells as judged by the viability of individual cells and by the release of lactate dehydrogenase (LDH) into the medium. Manganese superoxide dismutase (Mn-SOD) was markedly induced by these cytokines in EC and in A549 cells but not in ME180 cells. The levels of Mn-SOD in the conditioned medium of EC were dramatically increased after stimulation with cytokines, whereas those in ME180 and A549 cells were relatively low. The amount of Mn-SOD released appears to be comparable to that from cells lysed by other means. Immunoblot analysis of Mn-SOD in the medium showed that the molecular mass of the immunoreactive protein was the same as mitochondrial Mn-SOD, indicating that no proteolysis had occurred. These data suggest that in vivo the TNF-alpha produced by cancer cells may induce Mn-SOD in vascular endothelial cells, resulting in release of a relatively large amount of this protein into the serum.     

Obstructive jaundice in rats results in exaggerated hepatic production of tumor necrosis factor-alpha and systemic and tissue tumor necrosis factor-alpha levels after endotoxin

BACKGROUND: Obstructive jaundice (OJ) predisposes patients to postoperative sepsis. We determined whether OJ led to an increased endotoxin stimulated tumor necrosis factor-alpha (TNF-alpha) production by macrophage-rich organs and whether a lack of intraluminal gut bile contributed to this increased sensitivity. METHODS: Rats underwent laparotomy and common bile duct ligation and division (CBDL) or sham operation after they were given low-dose endotoxin or saline solution (NS). TNF-alpha levels in plasma, perfusate from the isolated perfused rat liver, and tissue from lung, spleen, and liver were measured 90 minutes later. An additional group underwent creation of a choledochal-vesical fistula and endotoxin administration. RESULTS: The plasma TNF-alpha, liver perfusate TNF-alpha, and the tissue TNF-alpha levels in liver, lung, and spleen were significantly elevated in the CBDL + endotoxin (CBDL + ETX) group compared with the SHAM + ETX and CBDL + NS groups (p < 0.05). The choledochal-vesical fistula group after endotoxin had plasma TNF-alpha levels only 27% that of the CBDL + ETX group (p < 0.05). CONCLUSIONS: OJ sensitizes macrophage-rich organs to produce larger amounts of TNF-alpha in response to endotoxin. This sensitization is not solely due to decreased intraluminal gut bile.     

Interleukin-4 and -13 inhibit tumor necrosis factor-alpha mRNA translational activation in lipopolysaccharide-induced mouse macrophages

The production of tumor necrosis factor-alpha (TNF-alpha) by lipopolysaccharide (LPS)-stimulated macrophages can be markedly inhibited by the two closely related cytokines, interleukin (IL)-4 and IL-13. To investigate the molecular mechanism of this inhibition, we analyzed the effect of the two cytokines on TNF-alpha production and TNF-alpha mRNA accumulation in the mouse macrophage cell lines RAW 264.7 and J774 stimulated by LPS. Whereas LPS-induced TNF-alpha production is strongly suppressed by both cytokines, TNF-alpha mRNA accumulation is not significantly affected, indicating that IL-4 and IL-13 induce a translational repression of TNF-alpha mRNA. Transfection of reporter gene constructs containing different regions of the TNF-alpha gene revealed that the inhibitory action of IL-4 and IL-13 is mediated by the UA-rich sequence present in the TNF-alpha mRNA 3'-untranslated region.     

Study of antiinfectious potential of recombinant tumor necrosis factor-alpha

46 patients with coronary heart disease with hypercholesterolemia were exposed to therapeutic plasmapheresis (TP) in combination with alpha-tocopherol treatment (AT). The results of 3-month follow-up with assessment of the clinical status, lipid spectrum, lipid peroxidation, concentration of ceruloplasmin indicated high hypolipidemic effectiveness of TP 2-3 weeks after the treatment as shown by inhibition of lipid peroxidation and antioxidant system. The addition of AT prolonged the hypolipidemic effect of TP, promoted optimization of plasma antioxidant potential (a rise in HDL, stabilization of ceruloplasmin levels).     

Selective induction of tumor necrosis factor receptor type II gene expression by tumor necrosis factor-alpha in C6 glioma cells

The aim of this study, in rabbit tibia, was an evaluation of the early reactions of the tissues to the insertion of polylactic membranes, used in connection with titanium implants. The specimens were retrieved after 1-4 weeks, and a histological analysis was performed. It was possible to see that, in the early implantation phases, no degradation of the macrostructure of the membrane was present. On the outer portion of the membrane many multinucleated giant cells (MGC) were present and membrane fragments were present inside the cytoplasm of these cells. These cells could explain the inflammatory processes reported, in some reports, with the use of materials made by polylactic and polyglycolic acid. We did not observe detrimental effects in the bone tissue around the membrane, and the membrane appeared to have a mechanical stability for the time necessary for bone regeneration.     

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

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

Acetazolamide treatment prevents in vitro endotoxin-stimulated tumor necrosis factor release in mouse macrophages

A 22-year-old woman was admitted to our hospital with fever, generalized lymphadenopathy, and pancytopenia in February 1995. She was diagnosed as having systemic lupus erythematosus (SLE) based on positivity for anti-nuclear antibody, and polyarthritis among other findings. A diagnosis of disseminated intravascular coagulation (DIC) was made based on the increase of FDP and other data (DIC score: 7). We also detected an anti-fibrinogen antibody. Lymph node biopsy revealed subacute necrotizing inflammation and there were on signs of the hemophagocytic phenomenon in bone marrow. The DIC score improved and the anti-fibrinogen antibody disappeared in association with the response of SLE to prednisolone therapy. The onset of SLE associated with DIC has never been reported before, as far as we could determine. The mechanism of DIC associated with SLE may be related to endothelial damage caused by immune complexes.     

Effect of esculentoside H on release of tumor necrosis factor from mouse peritoneal macrophages

Effect of esculentoside H (EH) on release of tumor necrosis factor (TNF) from murine peritoneal macrophage (Mphi) in vitro was studied. The results showed that EH (12.5-200 micrograms.ml-1) induced the thioglycolate-broth elicited peritoneal Mphi to release TNF into supernatants in a dose-dependent manner, and higher levels of TNF activity were detected in the supernatants from EH-stimulated calcimycin-primed M? culture. EH-induced TNF release had a different type of kinetics compared with that of lipopolysaccharides (LPS). LPS-induced release of TNF increased rapidly until 6 h after LPS stimulation, then declined gradually, while EH-induced TNF release increased gradually after EH stimulation and reached its peak at approximately 24 h later. These results suggested that the anti-tumor mechanisms of Phytolaccaceae may be related to the capacity of EH for TNF release.     

Molecular design of hybrid tumor necrosis factor-alpha III: polyethylene glycol-modified tumor necrosis factor-alpha has markedly enhanced antitumor potency due to longer plasma half-life and higher tumor accumulation

We have reported that chemical modification of tumor necrosis factor-alpha (TNF-alpha) with polyethylene glycol (PEG) markedly increases its antitumor potency without any adverse side effects. MPEG-TNF-alpha, especially, in which 56% of the lysine amino groups of TNF-alpha are coupled with PEG, exhibits 100-fold more antitumor activity in vivo than native TNF-alpha in the Meth-A murine sarcoma model. In this study, we investigated the pharmacokinetics of PEG-modified TNF-alpha with various molecular sizes to clarify the mechanisms of the enhanced antitumor potency of MPEG-TNF-alpha. The plasma half-lives of modified TNF-alpha increased with increasing molecular size. The decreased plasma clearance of modified TNF-alpha was partially caused by the shielding effect of the proteolytic sites in TNF-alpha by the attached PEG and the decreased transport from blood to various tissues. Almost all native TNF-alpha was uniformly distributed to the kidney and reticuloendothelial system within 1 hr of an intravenous administration, and rapidly disappeared from these tissues at 3 hr. However, very little native TNF-alpha was transported into the tumor. The absolute distributed amount and distribution profile of modified TNF-alpha to tissues other than the tumor were the same as those of native TNF-alpha, whereas the plasma levels of the modified TNF-alpha were higher than plasma levels of the native TNF-alpha. The tumor distribution of modified TNF-alpha was markedly enhanced compared with native TNF-alpha and gradually increased over time. About 9-fold more MPEG-TNF-alpha was distributed to the tumor than native TNF-alpha. Thus, we found that the marked increase in the antitumor potency of PEG-modified TNF-alpha resulted from the enhanced blood residency and tumor accumulation. The antitumor effect of MPEG-TNF-alpha against sarcoma-180 other than Meth-A fibrosarcoma was also about 100 times greater than that of native TNF-alpha when systemically administered. The optimal PEGylation of TNF-alpha facilitated its antitumor potency and MPEG-TNF-alpha may be useful systemic antitumor therapeutic drug.     

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.     

In vitro mechanism(s) of ultraviolet-induced tumor necrosis factor-alpha release in a human keratinocyte cell line

It has been demonstrated that ultraviolet (UV) irradiation is able to induce both in vivo and in vitro, tumor necrosis factor-alpha (TNF) release. The purpose of the present study was to evaluate, using a human keratinocyte cell line NCTC 2544, the mechanism(s) of UV-induced TNF release and the ability of commonly used sunscreens to modulate UV-induced TNF release. TNF release can be partially prevented both by adding an anti-human IL-1 alpha antibody after irradiation, suggesting an autocrine effect of IL-1 alpha in inducing TNF release, and by adding antioxidants indicating also a role of oxidant species. TPCK, a I kappa-B alpha protease inhibitor, was able to virtually abolish UV-induced TNF release, indicating that UV-induced TNF release requires NF-kappa B activation. Anti-human IL-1 beta antibody was ineffective as expected, considering that keratinocytes are unable to process pre-IL-1 beta to its active form. To evaluate the sunscreen's modulation on UV-induced TNF release, confluent cells were irradiated, in the presence or absence of the tested sunscreens (Uvinul MS40, Uvinul P25 and Uvinul DS49). Different IC50 values could be calculated, which may be related to different UV absorption spectrums: Uvinul MS40 offers great protection by virtue of its broader absorption spectrum, closely followed by Uvinul P25 and finally by Uvinul DS49.     

Differential induction of metallothionein synthesis by interleukin-6 and tumor necrosis factor-alpha in rat tissues

Metallothionein (MT) synthesis induced by the inflammatory cytokines, interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF), was studied in vivo. Administration of recombinant human IL-6 or TNF to rats caused the acute phase responses including rapid decreases in plasma zinc (Zn), and increases in plasma copper (Cu) and ceruloplasmin. Hepatic concentration of MT-I, one of MT isoforms, began to increase within 3 h after the injection of IL-6 or TNF. In IL-6-treated rats, MT-I concentration in liver reached a maximum level at 12 h and decreased with a transient rebound, whereas, in TNF-treated rats, a high level of MT-I lasted for about 48 h. MT-II, the other MT isoform, was induced more than MT-I in liver by both cytokines. MT-I was also induced in lung and heart by TNF, but little by IL-6. The data suggest that IL-6 may be responsible for MT synthesis in liver, whereas TNF may be responsible not only in liver but also in lung and heart. Furthermore plasma concentration of MT did not always reflect the enhanced concentration of MT by TNF and IL-6 in liver, suggesting involvement of many factors influencing plasma MT levels. The interrelation between IL-6 and TNF for MT synthesis has also been discussed.