Influence of the A and B subunits of cholera toxin (CT) and Escherichia coli toxin (LT) on TNF-α release from macrophages (original) (raw)
Related papers
Cellular Immunology, 1990
Clostridium dz@cile toxin A causes severe intestinal inflammation and fluid secretion in rabbit ileum and is chemotactic for neutrophils in vitro. The mechanism of intestinal injury produced by toxin A appears to involve direct epithelial cell damage as well as recruitment of an inflammatory cell response. The current study was undertaken to determine if toxin A can directly stimulate a proliferative response in lymphocytes. Highly purified toxin A, in the presence ofthe calcium ionophore, ionomycin, stimulated substantial ['Hlthymidine incorporation by murine splenic lymphocytes, which was maximal at 1 Om9 Mtoxin A and 800 rig/ml ionomycin. Removal of T cells with anti-Thy-l.2 antibody plus complement had no effect on the proliferative response induced by toxin A. However, [3H]thymidine incorporation in response to toxin A was significantly inhibited (P < 0.00 1) by the removal of macrophages from splenocyte suspensions and was restored by the addition of peritoneal macrophages or cell-free supernatant from toxin A-treated macrophage cultures. Analysis of the toxin A-treated macrophage supernatants showed high levels of IL-I, but not IL-2 or IL-4. The combination of recombinant IL-1 plus ionomycin was found to stimulate ['Hlthymidine incorporation by T cell-depleted splenic lymphocytes. These results suggest that toxin A stimulates the release of IL-1, and possibly other factors, from macrophages which can costimulate murine B lymphocytes. o 1990 Academic Press. Inc.
Previously study showed that LT-R192G, a less toxic mutant of the thermolabile enterotoxin of E. coli (LT), decreased murine regulatory CD4+CD25+Foxp3+ T cells in vitro. To go further, we tested other molecules of the cholera toxin family, i.e. native LT, the cholera toxin (CT), and their B subunits devoid of enzymatic activity, LTB and CTB. Cells from mesenteric lymph nodes of BALB/c mice were incubated with these molecules, and CD4+CD25+Foxp3+ T cells were analyzed by cytometry after two days of culture. All molecules, except CTB, decreased CD4+CD25+Foxp3+ T cells. T cell apoptosis has already been reported with these molecules, preferential effect on CD8+ T cells. So, we analyzed CD3+CD8+ and CD4+CD25+/- T cell apoptosis in the presence of LT, LT-R192G, LTB, CT and CTB in our in vitro model to detect possible preferential apoptosis of Tregs. Unexpectedly, CTB did not affect CD8+ and CD4+ T cells. Whereas LT, LT-R192G and LTB induced rapid apoptosis of CD8+ T cells with a decrease in CD3 expression, CT induced less rapid apoptosis with no decrease in CD3 expression. We observed CD4 T cell apoptosis but did not find any preferential apoptosis of CD4+CD25+ T cells (containing Foxp3+ T cells) compared with CD4+CD25- T cells. In conclusion, the enzymatic activity seems to play a significant role in apoptosis induction by CT, as CTB has no effect. In contrast, receptor binding may be essential in the observed effects for LT. Moreover, these results show the complexity and heterogeneity in the impact of these molecules.
The effects of tumor necrosis factor (TNF) derivatives on TNF receptors
Cytokine, 1993
The pleiotropic cytokine TNF has been implicated in the regulation of many immune and inflammatory responses in vivo, and in addition exerts a wide range of effects on target cells in vitro. However, although two cell surface receptors for TNF have been identified, and their cDNAs cloned, the amino acid residues necessary for the biological activity of TNF have not been characterized. We have therefore constructed derivatives of TNF termed 'muteins', in which the first 3 to 7 amino acids of native TNF-a have been replaced, using synthetic cDNA expressed in E. co/i. In the present study we compare the effects of native TNF-a and muteins III, IV, V and VI in several different in vitro systems and in one in vivo model. We observed binding to the ~75 TNF receptor on Jijoye Burkitt lymphoma cells with native TNF-cw and mutein III alone, whereas the ~55 TNF receptor on the human epithelioid carcinoma cell line HeLa bound TNF-a, mutein III and mutein V. Muteins IV and VI failed to recognize either TNF receptor. WEHI 164 fibrosarcoma cells were killed by muteins III, V and VI. Human umbilical vein endothelial cells responded to native TNF-(U and to muteins III, IV and V, but not to mutein VI, by increasing the surface expression of ICAM-antigen and secretion of the cytokines GM-CSF and IL-6. All four compounds were pro-inflammatory in a mouse in vivo model. The results presented in this report confirm that N-terminal amino acids are critical for both receptor binding and biological activity of TNF-cw. Amino acid substitution in the N-terminal region of the TNF-a molecule may provide an insight into the mechanism of TNF-a receptor binding, and further elucidate the structural requirements for its biological activity.
Murine macrophage activation by staphylococcal exotoxins
Infection and immunity, 1991
We investigated the ability of staphylococcal enterotoxins A and B, exfoliative toxins A and B, and toxic shock syndrome toxin 1 to activate macrophages. All of the toxins tested had the potential to stimulate tumoricidal activity in peritoneal macrophages from lipopolysaccharide-responsive C3HeB/FeJ mice. In contrast, none of the toxins activated cytotoxicity in lipopolysaccharide-unresponsive macrophages from C3H/HeJ mice. We also studied toxin stimulation of monokine secretion. Staphylococcal enterotoxin A, toxic shock syndrome toxin 1, and both exfoliative toxins triggered C3HeB/FeJ macrophages to secrete tumor necrosis factor alpha, but enterotoxin B induced only marginal amounts of tumor necrosis factor. All of the toxins used stimulated interleukin-6 production by macrophages from both strains of mice. Nitric oxide is produced in response to the exfoliative toxins only by the lipopolysaccharide-responsive macrophages. These results suggest that macrophages respond differently...
1993
Thekinetics oftumornecrosis factor alpha (TNF-ca) production, thecorrelation between cytokine levels and mortality ratesafter lethal challenge, andtheproduction ofa native inhibitor ofTNF-aactivity was investigated inmiceimmunized withformalin-kilied Escherichia coli. Groups ofmicewereinjected for8weeks witheither untreated bacteria orbacteria treated with 0.5 MICofaztreonam andsubsequently challenged with 10050%lethal doses ofviable E.coli. Micereceiving saline only(controls) diedwithin 24h.Themortality of miceimmunized withaztreonam-treated E.coli was significantly lowerthanthatofmiceimmunized with untreated E.coli. There were no measurable levels ofTNF-ceinseraobtained fromcontrol miceduring the entire period ofimmunization. TNF-ca levels ranging from90to306U/mlwere measured 90minafter each vaccination inseraobtained frommiceimmunized withuntreated E.coli. Serafrommiceimmunized with antibiotic-treated E.coli showedlowerTNF-alevels, ranging from40to128U/ml. TNF-alevels measured 90...